Issue 4 - August 2010 - Pacini Editore

Periodico bimestrale – POSTE ITALIANE SPA - Spedizione in Abbonamento Postale - D.L. 353/2003 conv. in L. 27/02/2004 n° 46 art. 1, comma 1, DCB PISA 

Aut. Trib. di Genova n. 75 del 22/06/1949 

Journal of the Italian Society of Anatomic Pathology 

and Diagnostic Cytopathology, 

Italian Division of the International Academy of Pathology 

Società Italiana di Anatomia Patologica e Citopatologia Diagnostica, 

Divisione Italiana della International Academy of Pathology 

Cited in Index Medicus/MEDLINE, BIOSIS Previews, SCOPUS 

Vol. 102 August 2010

IL BOARD DI PPPFAD 

Bruno Murer, Mattia Barbareschi, a nome del GIPP 

Un servizio di aggiornamento scientifico realizzato con il contributo di

✄ 

Care Colleghe e Cari Colleghi, 

PPPFAD, il programma formazione a distanza web-based 

che il Grruppo Italiano di Patologia PleuroPolmonare 

(GIPP) ha organizzato lo scorso anno, grazie ad un grant 

educazionale non condizionante di Lilly Italia, ha avuto un 

grande successo, con quasi 400 colleghi che hanno visitato 

il nostro sito e affrontato i 24 casi proposti fra il 2009 e 

i primi mesi del 2010! 

Abbiamo così deciso di proseguire l’attività formativa anche 

nel corso del 2010, adottando però una nuova formula 

che speriamo possa essere gradita a tutti. Si tratta di un percorso che da un lato 

propone nuovamente la formula della visione di casi problematici prevalentemente 

di natura neoplastica con un format più snello e vivace, dall’altro ci offre la 

straordinaria possibilità di partecipare a un grande studio di concordanza diagnostica 

nella applicazione della classificazione degli istotipi dei tumori del polmone, 

con la possibilità anche di fare una fotografia reale delle possibilità/difficoltà 

diagnostiche sia sulle piccole biopsie che sui pezzi operatori. 

Da tale studio di concordanza potremo trarre un lavoro scientifico di sicuro interesse 

che cercheremo di pubblicare a nome di tutti i partecipanti. Lo studio di concordanza 

inizierà con una serie di casi su piccole biopsie per poi proseguire nella 

seconda metà del progetto con i casi operatori, sui quali cercheremo di applicare 

anche i nuovi schemi classificativi che sono attualmente in corso di pubblicazione. 

PPPFAD risponde appieno alle caratteristiche educazionali richieste dal nuovo 

Sistema nazionale ECM, presso il quale è stato accreditato appunto quale programma 

di formazione a distanza. 

Sul website di www.pppfad.it troverete dunque il doppio percorso formativo col 

quale poter conseguire i crediti ECM-FAD. 

Con la certezza che possiate gradire e condividere lo sforzo del GIIPP per offrire 

ai patologi italiani le migliori occasioni di aggiornamento e di confronto, a Voi tutti 

l’augurio di un buon lavoro! 

Bruno Murer 

Coordinatore GIPP 

Si prega di scrivere in stampatello leggibile e restituire a: Infomedica - Via P. Giannone, 10 - 10121 Torino - Fax 011.859890 - info@infomedica.com 

Desidero ricevere l’accesso a PPPFAD online per il 2010 

Nome Cognome E-mail 

Ospedale/Istituto di cura Qualifica 

Indirizzo di lavoro 

Help Desk: per qualsiasi necessità l’utente può contattare Infomedica, il provider che cura l’iniziativa: 

tel. 011 859990 dal lunedì al venerdì, ore 9.30 - 12.30 e 14.30-17.30 - E-mail: info@pppfad.it 

CAP Città Prov. Telefono 

Ai sensi dell’art. 13 del DL 196/03 e succ. modifiche e integrazioni La informiamo che i Suoi dati personali verranno trattati dal Titolare con strumenti informatici nel pieno rispetto della normativa applicabile al 

fine dell’invio della pubblicazione richiesta e dell’accesso al sito di PPPFAD online. Il conferimento dei Suoi dati è facoltativo; tuttavia la mancata compilazione del presente modulo non consentirà l’accesso al sito. 

Titolare del trattamento è Infomedica Srl e Lei potrà esercitare i diritti riconosciuti ex art. 7 DL 196/03 con richiesta rivolta a Infomedica Srl., via P. Giannone 10, 10121 Torino, tel. 011.859990. Pienamente informato 

delle finalità e modalità del trattamento dei dati, con la compilazione del presente modulo do il mio consenso al trattamento dei dati ivi indicati e all’invio di materiale pubblicitario, promozionale e commerciale. 

Data Firma 

1. PPPFAD 2010 

È il nuovo programma FAD realizzato dal GIPP, 

Gruppo Italiano di Studio di Patologia Pleuropolmonare 

2. COSA PUBBLICA PPPFAD 

Un duplice percorso formativo che prevede: 

- 24 casi clinici (2 al mese) 

- 100 casi di concordanza diagnostica articolati 

in 20 moduli da 10 casi ciascuno 

- A completamento, la possibilità di partecipare 

ad una survey finalizzata all’elaborazione 

dello studio scientifico promosso dal GIPP 

3. COME ACCEDERE A PPPFAD 

Accedere a PPPFAD è facile: 

- digitare l’indirizzo Internet: www.pppfad.it 

- inserire il codice di attivazione fornito 

- registrarsi scegliendo le proprie username e 

password 

4. CREDITI ECM-FAD 

- PPPFAD è accreditato quale servizio di Formazione 

a Distanza presso il Sistema nazionale 

ECM 

- Eroga dunque crediti ECM-FAD: 32 crediti 

annui 

- Per acquisirli è necessario registrarsi e completare 

i casi clinici proposti da www.pppfad.it

Journal of the Italian Society of Anatomic Pathology 

and Diagnostic Cytopathology, 

Italian Division of the International Academy of Pathology 

Editor-in-Chief 

Marco Chilosi, Verona 

Associate Editor 

Roberto Fiocca, Genova 

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Società Italiana di Anatomia Patologica e Citopatologia Diagnostica, 

Divisione Italiana della International Academy of Pathology 

Cited in Index Medicus/MEDLINE, BIOSIS Previews, SCOPUS 

Governing Board 

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Vol. 102 August 2010

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PRESIDENTE COMITATO 

SCIENTIFICO 

Vincenzo Eusebi 

COMITATO SCIENTIFICO 

Arrigo Bondi 

Cesare Bordi 

Marco Chilosi 

Maria Pia Foschini 

Giorgio Gardini 

Felice Giangaspero 

Walter F. Grigioni 

Giovanni Lanza 

Antonio Maiorana 

Giuseppe Martinelli 

COMITATI 

Roberto Nannini 

Stefano Pileri 

Anna Sapino 

Gian Luigi Taddei 

Giovanni Tallini 

PRESIDENTE COMITATO 

ORGANIZZATORE 

Arrigo Bondi 

COMITATO 

ORGANIZZATORE 

Andrea Ambrosini Spaltro 

Gian Piero Casadei 

Giovanna Cenacchi 

Guido Collina 

Antonia D’Errico 

Giuseppina Ferro 

Michelangelo Fiorentino 

Luisa Losi 

Luca Morandi 

Annalisa Pession 

Pier Paolo Piccaluga 

Teresa Ragazzini 

Donatella Santini

Pathologica 2010;102:127-235 leCTureS 

The role of magnetic resonance in the 

surveillance of women at high risk of 

hereditary breast cancer 

Wednesday, September 22 nd , 2010 

Symposium Susan G. Komen: Breast pathology 

F. Podo, F. Santoro, F. Sardanelli * 

Department of Cell Biology and Neurosciences, Istituto Superiore di 

Sanità, Rome; * Università di Milano, IRCCS Policlinico San Donato, 

Milano, Italy 

Background. Breast cancer (BC) affects up to 1:7 to 1:11 

women in Western Countries. Although BC is mainly a sporadic 

disease, about 15% of cases are clustered in families 

with highly or moderately elevated BC incidence. Pathogenic 

mutations in high-risk genes at autosomic dominant inheritance 

are held responsible for about 5% of BC cases, in which 

the disease may have early onset with a estimated cumulative 

lifetime risk as high as 50% to 85%. About 50% of hereditary 

BC cases can be explained by mutations in BRCA1 and 

BRCA2 genes. BRCA1 mutations are frequently associated 

with triple negative BC (TNBC), defined by lack of estrogen 

and progesterone receptor expression and absence of ErbB2 

(or HER2) amplification (reviewed in Bosch A. et al. Cancer 

Res and Treatment Reviews 2010; Podo F. et al., Mol Oncol 

2010). In women at high risk of breast cancer, screening mammography 

has shown a lower sensitivity (29-50%) compared 

with that of the screening addressed to the general female 

population (80%), with higher percentages of interval cancers 

(35-50% vs 20-25%) and higher nodal involvement (20-56% 

vs 22%). In the last decade a number of prospective, non-randomized 

studies have been conducted in Europe and North 

America to assess the value of dynamic contrast-enhanced 

magnetic resonance imaging (MRI) as a screening tool to be 

used as an adjunct to ×-ray mammography (XM), or to XM 

and ultrasonography (US) for the surveillance of women at 

high genetic-familiar risk of BC 1-10 . We will summarize the 

results of two consecutive multicenter, prospective, non-randomized 

studies coordinated in Italy by the Istituto Superiore 

di Sanità, the ISS-HIBCRIT Study 6 9 11 , carried out from June 

2000 to March 2008 in 18 Centers, and the four-year ISSIN- 

HBCR study, whose screening activities started in 2008 with 

the collaboration of a Network of 23 Centers located in 13 

regions. 

Methods. Both studies prospectively compared clinical breast 

examination (CBE), XM, US, and MRI for screening women 

at genetic-familial high risk of BC. CBE, XM, US, and MRI 

were used for repeated, yearly screening of women either 

proven to be BRCA1 or BRCA2 mutation carriers (BRCA + ), 

or untested first degree relatives of BRCA + , or enrolled only 

on the basis of strong family history of breast and/or ovarian 

cancer. Histopathology or at least one-year negative follow-up 

were used as the reference standard. 

Results. The HIBCRIT study enrolled 501 asymptomatic 

women (mean age 46.0 ± 11.8 years; median age 45 years); 

69% proven carriers of BRCA1 or BRCA2 mutation (or first 

degree relatives of proven carriers) with a BRCA1:BRCA2 

ratio of 1.3; 43.5% women had previous BC and/or ovarian 

cancer. A total of 1592 annual rounds (3.2 rounds/woman) 

Moderators: R. Masetti (Roma), V. Eusebi (Bologna) 

were performed. A total of 52 breast cancers were detected: 

49 screen-detected and 3 interval cancers (all three TNBC); 44 

invasive, 8 in situ; only 4 at stage ≥ pT2; 32 G3 grade; 72% 

(28 out of 39 patients explored for nodal status) were nodenegative. 

Of 43 invasive BC for which histopathological findings 

were reported, 18 (43%) were TNBC (12 associated with 

BRCA1 mutation, 3 with BRCA2 mutation and 3 detected 

in untested women with a strong family history of BC). The 

detection rate per year was 3.0% (95% CI 1.9%-4.0%) for 

BRCA1 or BRCA2 mutation carriers and 3.3% overall (95% 

CI 2.4%-4.1%). Cancer was detected only with MRI in 15 

out of 40 patients examined with all modalities. MRI showed 

the highest sensitivity (91%) compared with CBE (18%), 

XM (50%), ultrasonography (52%) and to the combination 

of mammography plus US (63%) (p < 0.001). Specificity for 

CBE, mammography, ultrasonography, mammography plus 

ultrasonography and MRI was 99%, 99%, 98%, 98% and 

97%; positive predictive value (PPV) was 56%, 71%, 62%, 

56% and 56%; negative predictive value 96%, 97%, 98%, 

98% and 100%, without significant differences. The area 

under the curve at ROC analyses was significantly higher for 

MRI (0.97) than for mammography (0.83) and ultrasonography 

(0.82) (p < 0.001) and was not significantly increased in 

the combination of MRI with either mammography or ultrasonography 

or both modalities. 

The ISSIN-HBCR study enrolled 662 women in two years 

(1.4 rounds/woman). The population characteristics are not 

significantly different from those of the HIBCRIT study in 

terms of mean and median age, percentage of BRCA mutation 

carriers (70%) and percentage of women with previous breast 

cancer. A total of 29 breast cancers have already been detected: 

24 screen-detected and 5 interval cancers (four TNBC, 

one not yet reported); 28 invasive, 1 in situ. The distribution 

of pT stages, G grade and percentage of nodal involvement 

are very close to those of the eight-year HIBCRIT study. The 

sensitivity of the MRI similarly outperformed that of the other 

imaging modalities or their combination; 33% of cases were 

detected by MRI only. 

Conclusions. In conclusion, the consolidated results of the 

HIBCRIT study showed that: a) MRI largely outperformed 

XM, US and their combination for screening high-risk women 

under and over 50 years of age; b) over 30% of tumors were 

detected by MRI only; c) the PPV of MRI reached 56%; 

d) over 40% of detected tumors were either in situ or smaller 

than 1 cm; e) over 70% of invasive tumors were lymph-node 

negative; f) women at high genetic-familial risk with personal 

history of BC should be included in a multimodality surveillance 

including annual MRI; g) the increasing incidence with 

age suggests not to reduce intensive surveillance in menopausal 

women. Preliminary results of the ISSIN-HBCR study 

confirm these trends. Further studies are needed: to better define 

risk-reduction strategies for BRCA1 mutation carriers in 

relation to the high risk of TNBC; to identify MRI parameters 

related to TNBC diagnosis, prognosis and prediction of therapy 

response; to evaluate the benefits of surveillance of high 

risk women compared with other strategies of risk reduction,

128 

according to gene mutation and age; to possibly optimize and 

simplify the protocols of multimodality surveillance; to evaluate 

the cost/benefit ratio of extending secondary prevention 

programs to women at intermediate risk of breast cancer. 

references 

1 Kriege M, Brekelmans CT et al. N Engl J Med. 2004;351:427-37. 

2 Warner E, Plewes DB, et al. JAMA. 2004;292:1317-25. 

3 Leach MO, Boggis CR, et al. Lancet. 2005;365(9473):1769-78. 

4 Kuhl CK, Schrading S, et al. J Clin Oncol 2005;20;23:8469-76. 

5 Lehman CD, Blume JD, et al. Cancer 2005;103:1898-905. 

6 Sardanelli F, Podo F, et al. Radiology 2007;242:698-715. 

7 Hagen AI, Kvistad KA, et al. Breast 2007;16:367-74. 

8 Riedl CC, Ponhold L, et al. Clin Cancer Res. 2007;13:6144-52. 

9 Sardanelli F, Podo F. Eur Radiol 2007;17(4):873-87. 

10 Kuhl C, Weigel S et al. J Clin Oncol 2010;28(9):1450-7. 

11 Podo F, Sardanelli F et al., submitted. 

Preoperative breast mri: which evidence? 

F. Sardanelli 

Dipartimento di Scienze Medico-Chirurgiche, Università di Milano, 

Unità di Radiologia, IRCCS Policlinico San Donato, Milan, Italy 

(francesco.sardanelli@unimi.it) 

Background. Breast conserving treatment (BCT), comprising 

breast conserving surgery (BCS) plus radiation therapy, 

is equally effective to mastectomy, in terms of survival, for 

early-stage cancers as demonstrated in randomized controlled 

trials (RCTs) and confirmed in a meta-analysis 1 . Of 

importance, four of the six RCTs of BCS included in this 

meta-analysis show a significantly lower risk of locoregional 

recurrence in favor of mastectomy (odds ratio 1.561) 1 . Thus, 

BCS should always aim to completely remove tumoral tissue 

and obtain clear margins. 

Evidence on MRI’s detection capability. MRI has a superior 

sensitivity compared with mammography in assessing index 

tumor size and in detecting ipsilateral multifocal or multicenter 

cancers, as demonstrated also in a multicenter study 2 . 

However, MRI may fail to detect all cancers when the whole 

breast is used as a pathological reference standard 3 , especially 

when ductal carcinoma in situ (DCIS) is considered 4 . The 

advantage of MRI has been shown to be non-significant in 

fatty breasts, while significant in scattered fibroglandular, 

heterogeneously, or extremely dense breasts 3 . MRI has also 

been shown to detect extensive intraductal component, but 

may overestimate or underestimate this finding in 11-28% 

and 17-28% of cases, respectively 5-7 . In a meta-analysis of 

19 studies 8 , the impact of pre-operative MRI on ipsilateral 

surgical planning was evaluated reporting surgical outcomes 

as follows 8 : 

– 8.1% conversion from wide local excision to mastectomy 

due to true positive findings; 

– 1.1% conversion from wide local excision to mastectomy 

due to false positive findings; 

– 3.0% conversion from wide local excision to wider/additional 

excision due to true positive findings; 

– 4.4% conversion from wide local excision to wider/additional 

excision due to false positive findings. 

Furthermore, several studies have shown that MRI can detect 

otherwise occult contralateral malignancy in women newly 

diagnosed with invasive cancer for about 3% of patients 9 . 

A meta-analysis of 22 studies 10 showed that MRI yields an 

incremental cancer detection rate equal to 4.1% with a positive 

predictive value of 47.9% due to a false positive detection 

rate of 5.2% (true positives/false positives = 0.92). In this 

analysis 10 35% of contralateral cancers were DCIS with a 

5 th triennial congress of the italian society of anatomic Pathology and diagnostic cytoPathology 

mean diameter of 7 mm, 65% invasive with a mean diameter 

of 9.3 mm, the majority of the latter were node negative 10 . A 

higher probability of an added diagnostic value of MRI for 

local staging has been shown for particular patient subgroups. 

In a recent systematic review of patients with invasive lobular 

cancer, additional ipsilateral lesions were found to be detected 

with MRI in 32% of cases, contralateral lesions in 7% 

while surgical management was changed in 28%.11 In these 

patients, MRI showed a 93% pooled sensitivity and a high 

correlation with pathologic tumor extent. 11 Women with an 

inherited high risk for breast cancer have a high probability of 

a more accurate local staging with MRI. The rate of multifocal 

and multicenter cancers in these women was reported as 

high as 45-50% 12 13 . In one study, the percentage of breasts 

with exact detection of the number of malignant lesions was 

reported to be 0% for mammography, 33% for sonography, 

and 71% for MRI 13 . Regarding the assessment of tumor extent, 

a retrospective analysis by Deurloo et al. 14 . reported that 

patients younger than 58 years of age with irregular lesion 

margins at mammography and discrepancy in tumor extent 

(including spiculated lesions and suspicious microcalcifications) 

by > 10 mm between mammography and sonography, 

had a 50% probability of complementary value of MRI over 

conventional imaging vs 16% in the remaining patients. Last 

but not least, MRI identifies a fraction of candidates for partial 

breast irradiation (PBI) who are affected with multifocal, 

multicentric, or contralateral cancer and may therefore not be 

suitable for this approach in treatment, about 5-10% according 

three recent studies 15-17 . This should be considered in the 

light that the American Society for Radiation Oncology has 

recently established the possibility of using PBI “outside a 

clinical trial” at least for patient subgroups 18 . Up to recent 

times, we have lacked evidence on patient outcomes in favor 

of, or against, pre-operative MRI. Conflicting retrospective 

studies on outcomes have been reported 19-22 , intrinsically 

limited by non-randomization. Unfortunately, the results of 

the COMICE 23 study, indicating the absence of benefit from 

MRI (about 19% of re-excision rate in both the MRI and non- 

MRI arms), are flawed by relevant limitations mainly due to 

very few experience with breast MRI by the large number of 

centers involved in that trial 24 . 

The potential and the drawbacks of MRI. Using tissue 

needle sampling of MRI-detected additional findings (through 

second-look sonography or MR-guidance), we will potentially 

drastically reduce overtreatment due to MRI false positives. 

As a consequence, using the estimates of Houssami et al. 8 , we 

would have only the 11.1% rate of MRI-induced potentially 

correct changes of surgical planning for the breast harboring 

the index lesion. To place this into context, we should consider 

the routine rate of positive margins after BCS, ranging from 

20% to 40% or more,25 and that of local recurrences after 

BCT, usually considered from 5% to 10% at ten years26 and 

reported about 9% at 20 years 27 . A similar reasoning can be 

proposed for the detection of contralateral cancers. Consistent 

use of MR-guided biopsy could strongly reduce the surgical 

treatment of false positives (about 5%) 10 , offering the chance 

to treat the synchronous contralateral cancers in about 4% of 

the women10 with simultaneous surgery. This rate should be 

compared with the 0.5-1% annual risk of contralateral breast 

cancer in women with a previous history of breast cancer 28 29 . 

We could speculate that only ipsilateral recurrences or contralateral 

cancers which would have appeared in the first years 

after BCT might be avoided by pre-operative MRI 26 . Thus, 

this comparison gives a relatively balanced result for contralateral 

cancers: with a 0.75% annual rate of contralateral

lectures 

cancers and an anticipated MRI diagnosis up to 3-4 years, we 

have a 2-3% cumulative rate of contralateral cancers in the 

first few years to be compared with a rate of MRI-detected 

contralateral cancers of 3-4% 9 10 . A larger discrepancy is obtained 

if we hypothesize a similar cumulative rate (2-3%) for 

local recurrences in the first years, to be compared with the 

11.1% rate 8 of MRI-induced correct changes of surgical planning 

for the breast harboring the index lesion. However, the 

rate of MRI-detected ipsilateral and contralateral cancers is 

probably overestimated due to the fact that pre-operative MRI 

has been performed in non-consecutive (selected) series 26 , i.e. 

through selection of patients with a probable higher likelihood 

of ipsilateral and contralateral cancers (for example dense 

breasts, or high-risk patients) to MRI. A publication bias is 

also hypothesized. Moreover, it is hard to evaluate the combination 

of the two aspects from a patient-based perspective: 

pre-operative MRI could determine an unnecessary wider/additional 

ipsilateral excision but also anticipate the diagnosis 

of contralateral cancer (or vice versa), thus avoiding the second 

cancer event in future, and receiving treatment for both 

breasts upfront; it may be argued that a bilateral advantage or 

a bilateral overtreatment could happen as a consequence. This 

interpretation considers the fact that systemic therapy may 

prevent some of the contralateral cancers 26 detected upfront 

by MRI only. At present, potential outcome benefits of preoperative 

MRI may include a possible reduction in the rate of 

the following events: surgical intervention needed to achieve 

free margins; ipsilateral recurrences; secondary mastectomies; 

and contralateral malignancy. On the other hand, we should 

consider that the use of MRI has been reported to be associated 

with an increased higher rate of mastectomy 22 26 30 31 and 

with a treatment delay of 22.4 days 24 . 

Perspectives on indications for pre-operative MRI. Acceptable 

indications for pre-operative MRI can be presently 

defined for subgroups of patients inwhom a larger potential 

benefit in term of local staging might be expected. This approach 

should be considered also for future RCTs evaluating 

pre-operative MRI. In fact, if the advantages of MRI would 

be relevant only for particular subgroups, RCTs on the average 

population of women newly diagnosed with a breast 

cancer may dilute the benefit and probably reduce power for 

achieving significance in subgroup analysis. Patients with a 

potential higher anticipated benefit from preoperative MRI 

can be identified as those: 1. with mammographically (heterogeneously 

or extremely) dense breasts; 2. with a unilateral 

multifocal/multicentric cancer or a synchronous bilateral cancer; 

3. with a lobular invasive cancer; 4. at high-risk for breast 

cancer; 5. with a cancer which shows a discrepancy in size 

of > 1 cm between mammography and sonography; 6. under 

consideration for PBI. 

More limited evidence exists in favor of MRI for evaluating 

candidates for total skin sparing mastectomy in order to 

decide saving or not the nipple32 or for patients with Paget’s 

disease 33-35 . Further research is needed in particular on these 

indications. Irrespective of whether the clinical team routinely 

uses preoperative MRI or not, the following issues are 

paramount: A. women newly diagnosed with breast cancer 

should always be informed of the potential risks and benefits 

of pre-operative MRI; B. results of pre-operative MRI should 

be interpreted taking into account clinical breast examination, 

mammography, sonography and verified by percutaneous biopsy; 

C. MRI-only detected lesions require MR-guidance for 

needle biopsy and pre-surgical localization, and these should 

be available or potentially accessible if pre-operative MRI is 

to be implemented; D. total therapy delay due to pre-operative 

129 

MRI (including MRI induced work-up) should not exceed one 

month; E. changes in therapy planning resulting from pre-operative 

MRI should be decided by a multidisciplinary team. 

Conclusions. In reality, and considering the detection capability 

of MRI, we cannot wait for conclusive evidence in favor 

of or against preoperative MRI. To deny this examination to 

all women newly diagnosed with breast cancer is a questionable 

decision because the evidence is ‘uncertain’ rather than 

against a benefit from preoperative MRI. In this context, to 

define general rules to be shared by breast cancer specialists is 

the first goal to avoid inappropriate use of this diagnostic step. 

To propose pre-operative MRI for subgroups of women as 

here defined can be a practical strategy for the present. Finally, 

the woman’s preference should be also carefully considered in 

order to decide whether to perform or not to perform preoperative 

MRI, according to evidence-based medicine basic 

principles 36 . From this standpoint we should also consider that 

mastectomy in 2010 is no longer the same surgical approach 

performed thirty or forty years ago. Immediate reconstruction, 

skin- and nipple-sparing mastectomy changed the scenario at 

least in terms of cosmetic results. Part of the reported increase 

in mastectomy rate may be due to the availability of these 

options. The large meta-analysis of Clarke et al. on the effect 

of radiation therapy concludes that “differences in local treatment 

that substantially affect local recurrence rates would, in 

the hypothetical absence of any other causes of death, avoid 

about one breast cancer death over the next 15 years for every 

four local recurrences avoided, and should reduce 15-year 

overall mortality” 37 . MRI is not radiation therapy but guiding 

a more effective surgery might potentially provide a similar 

effect. High-quality clinical research on pre-operative MRI is 

needed, especially RCTs. 

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6 Van Goethem M, et al. Eur J Radiol 2007;62:273-82. 

7 Kim do Y, et al. Korean J Radiol 2007;8:32-9. 

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9 Lehman CD, et al. N Engl J Med 2007;356:1295-303. 

10 Brennan ME, et al. J Clin Oncol 2009;27:5640-9. 

11 Mann RM, et al. Breast Cancer Res Treat 2008;107:1-14. 

12 Kuhl CK, et al. J Clin Oncol 2005;23:8469-76. 

13 Sardanelli F, et al. Radiology 2007;242:698-715. 

14 Deurloo EE, et al. Eur Radiol 2006;16:692-701. 

15 Al-Hallaq HA, et al. Cancer 2008;113:2408-14. 

16 Godinez J, et al. AJR Am J Roentgenol 2008;191:272-7. 

17 Tendulkar RD, et al. Cancer 2009;15(115):1621-30. 

18 Smith BD, et al. Int J Radiat Oncol Biol Phys 2009;74:987-1001. 

19 Fischer U, et al. Eur Radiol 2004;14:1725-31. 

20 Solin LJ, et al. J Clin Oncol 2008;26:386-91. 

21 Pengel KE, et al. Breast Cancer Res Treat 2009;116:161-9. 

22 Bleicher RJ, et al. ASCO Breast 2008 [abstract 227]. 

23 Turnbull L, et al. Lancet 2010;375:563-71. 

24 Morris EA. Lancet 2010;375:528-30. 

25 Pleijhuis RG, et al. Ann Surg Oncol. 2009;16:2717-30. 

26 Houssami N, Hayes DF. CA Cancer J Clin 2009;59:290-302. 

27 Veronesi U, et al. N Engl J Med 2002;347:1227-32. 

28 Adami HO, et al. Cancer 1985;55:643-7. 

29 Rutqvist LE, et al. J Natl Cancer Inst 1991;83:1299-306. 

30 Foote RL, et al. Breast 2008;17:555-62. 

31 Katipamula R, et al. J Clin Oncol 2008;26(Suppl):a509. 

32 Wijayanayagam A, et al. Arch Surg 2008;143:38-45. 

33 Frei KA, et al. Invest Radiol 2005;40:363-7. 

34 Haddad N, et al. J Radiol 2007;88:579-84. 

35 Morrogh M, et al. J Am Coll Surg 2008;206:316-21. 

36 Sackett DL, et al. BMJ 1996;312:71-2. 

37 Clarke M, et al. Lancet2005;366:2087-106.

130 

New and old entities in breast pathology 

V. Eusebi 

Sezione di Anatomia Istologia e Citologia Patologica “M. Malpighi” 

Università di Bologna 

To use Goethe’s words “we see what we know”. As we know 

very little new entities usually emerge as the result of better 

technology as well as more accurate methods of analysis. In 

addition some lesions tend to become obsolete and periodically 

are rediscovered and rejuvenated. 

In recent years very powerful molecular techniques have appeared 

which has lead to the statement by some molecular 

pathologists that by the year 2020, histopathology is going 

to be history and all diagnostic work up is going to be in the 

hands of scientists or machines. 

This might be the case although, to use the words of Lorenzo 

il Magnifico, “del diman non c’è certezza”. For the time being 

it appears that in spite of great expectations in molecular 

techniques, no very consistent achievements have been obtained. 

One example for all. Perou et al. 1 , at the beginning of 

this century, using a DNA array technique, reclassified breast 

cancer among groups different from those classically used. 

After nearly 10 years since the publication of Perou’s article, 

it appears that the new classification is not very useful in 

routine practice. One for example is the basal like carcinoma. 

In spite of myriads of papers published on it, there is still no 

a consensus on the definition of this type of tumor. Basal like 

molecular profile appears to be in common with a heterogeneous 

group of tumors which include very aggressive lesions 

that are G. 3 and triple negative carcinomas together with 

lesions that are quasi benign as well differentiated adenoid 

cystic carcinomas. 

Therefore we discuss here cases whose definition is mainly 

based on morphology. 

Breast carcinomas are simulated by a number of inflammatory 

conditions. 

Nodules either single, multiple or even bilateral are shown 

by IgG4-related sclerosing mastitis 2 which is a new entry of 

the syndrome of the IgG4-related sclerosing disease. This is a 

recently recognized syndrome characterized clinically by tumour-like 

enlargement of one or more exocrine glands as well 

as extra-glandular tissue. There is raised serum IgG4 level and 

histologically there is lymphoplasmacytic infiltration together 

with sclerosis. There are an increased number of IgG4-secreting 

plasma cells. The syndrome is believed to be autoimmune 

in origin, it was originally observed in autoimmune sclerosing-pancreatitis 

but a number of different sites have been 

reported such as hepatobiliary tree, lachrymal glands, salivary 

glands, lymph node, prostate, lung, kidney, retro-peritoneum 

and mesentery, mediastinum, meninges and breast 2-5 . In this 

latter site, of the 5 cases reported 2 were unicentric, 3 multifocal 

and 1 bilateral. Breast lesions are characterized by dense 

masses of lymphocytic infiltrate associated to intense sclerosis 

and loss of lobules. Reactive lymphoid follicles can be seen 

but granulomas as well as lympho- epithelial lesions are lacking. 

IgG4+ ought to be no less than 50% of IgG+ elements. 

This “inflammatory” lesion has to be distinguished from 

low grade B cell lymphoma and Castleman’s disease. In addition 

an inflammatory quasi neoplastic condition is Rosai 

Dorfman’s disease that can affect the breast. Of the 7 cases 

reported by Green et al. 6 , 3 patients had disease confined to 

the breast, one had involvement of the breast and ipsilateral 

auxiliary lymph nodes and two had bilateral breast involvement. 

A xantomatous infiltrate with scattered Touton’s giant 

cells and patchy lymphocytic infiltrate are the features of 


Erdheim-Chester (E-C) disease that may involve the breast 

presenting as bilateral clinically malignant breast masses. 

E-C disease is a rare non Langerhans cell histiocytosis of 

unknown aetiology. The commonest sites of involvement are 

long bones, skin, orbit pituitary and retro peritoneum. A number 

of granulomatous mastitis can clinically simulate a breast 

carcinoma among which idiopathic granulomatous mastititis 7 , 

sarcoid 8 and cat scratch disease. 

Among the lesions that are rejuvenated, the most obsolete 

entity that only recently has been brought up again is infiltrating 

epitheliosis 9 . 

Infiltrating epitheliosis (IE) was described by Azzopardi in 

1979 in Chapter 9 “Overdiagnosis of malignancy” of his book 

“Problems in breast pathology” 10 as “a lesion which is not 

uncommon but which has not received adequate recognition 

in the literature”, a statement very pertinent 30 years later. 

Infiltrating epitheliosis (IE) is usually a microscopic lesion, 

observed incidentally in cystic disease but which may infrequently 

present as a palpable lump. The lesion is generally located 

far from the nipple as epitheliosis (also known as usual 

duct hyperplasia-UDH), which is the main component, affects 

the acinar, terminal duct lobular unit (TDLU) and small duct 

portions of the mammary lobes 10 . 

The lesion is a complex epithelial-stromal interaction composed 

of epitheliosis (UDH) which constitutes the bulk of the 

lesion, and sclero-elastotic stromal changes. 

At low power IE appears as an asymmetrical lesion, with 

scleroelastotic areas located randomly either in the centre 

or at the periphery. The borders vary from irregular to circumscribed. 

In palpable lesions the scleroelastotic areas can 

be multiple. 

Morphologically the lesion is composed of Epitheliosis (synonym 

usual duct hyperplasia), and Scleroelastotic areas the 

latter characterized by a stromal reaction which is not only 

desmoplastic, but may also contain dense sclerotic and hyaline 

collagenous bands…” not unlike the appearances seen in 

a keloid” 10 . Finally abundant elastic tissue (elastosis) is seen 

intermingled with the desmoplastic reaction or around small 

ducts forming nodular foci. Infiltrating epitheliosis has to be 

distinguished from Radial Scar (benign scleroelastotic lesion 

simulating invasive duct carcinoma) which Hamperl in 1975 

described as a microscopic lesion that he named in German 

“strahligen narben”. In the summary this was translated as 

“radial scar” 11 . A few months later, Eusebi et al. 12 independently 

reported on the mammographic, macroscopic and 

microscopic features 4 cases showing a lesion they named in 

Italian “lesioni focali scleroelastotiche mammarie simulanti il 

carcinoma infiltrante”. In the summary this was translated as 

“mammary focal scleroelastotic lesions simulating an infiltrating 

carcinoma”. 

Both papers dealt with the same identical lesion 10 , the only 

difference being the size. The lesions described by Hamperl 11 

were selected on microscopic grounds and therefore were 

minute. Those reported by Eusebi et al. 12 were selected at 

mammography and all were palpable nodules, the largest 

measuring 2.5 cm in greatest axis. 

Both reports were in languages (German and Italian) that, 

especially 30 years ago, were not readily available to the 

scientific community. As a result the histological features of 

radial scar (scleroelastotic lesion) were not fully appreciated 

and the terms radial scar and infiltrating epitheliosis (and its 

synonyms) that describe two different lesions (see later) are 

used interchangeably by many authors. 

RS has a central zone of sclero-hyaline fibrous tissue mixed 

with abundant elastic tissue (elastosis). The sclero-elastotic

lectures 

nidus is surrounded by proliferating benign epithelium usually 

consisting of sclerosing adenosis and less frequently of epitheliosis, 

together with dilated small ducts that radiate towards 

the periphery. The zoning phenomenon, most evident at low 

power, is distinctive and very useful in frozen sections when 

distinguishing between invasive duct carcinoma and RS. The 

sclero-elastotic center on close analysis is seen to consist of 

round to elongated plaques of sclerohyaline tissue surrounded 

by elastosis a feature called obliterating mastopathy a phenomenon 

of involution of ducts as seen in the late stages of 

duct ectasia 13 . 

Finally both RS and IE must be distinguished from tubular 

carcinoma (TC) (Tab. I) 14 . 

In conclusion it seems that IE and RS are two morphologically 

and histogenetically distinct lesions. Whilst it may be a controversial 

view, careful histological evaluation reveals distinct 

differences between the two entities allowing for their separation. 

This is important as although both are clinically benign 

they appear to have different relationships with carcinoma. IE 

by definition has florid epitheliosis which is a proliferative 

process that is so extensive that it has been considered a “low 

risk duct intraepithelial neoplasia” 14 and accordingly should 

be regarded a probable risk “marker” for carcinoma 15 . RS on 

the other hand is the result of involutionary processes 16 . If a 

carcinoma arises in it, it is an infrequent phenomenon comparable 

to cases of fibroadenomas that harbour CIS but it should 

not be included in the list of precancerous lesions. 

references 

1. Sorlie T, Perou CM, Tibshirani R, et al. Gene expression patterns of 

breast carcinomas distinguish tumor subclasses with clinical implications. 

Medical Science 2001;98:10869-74. 

2 Cheuk W, Chan AC, Lam WL, et al. IgG4-related sclerosing mastitis: 

description of a new member of the IgG4-related sclerosing diseases. 

Am J Surg. Pathol 2009;33:1058-64. 

3 Chan SK, Cheuk W, Chan KT, Chan JK. IgG4-related sclerosing 

pachymeningitis: a previously unrecognized form of central nervous 

system involvement in IgG4-related sclerosing disease. Am. J. Surg. 

Pathol 2009;33:1249-52. 

4 Cheuk W, Yuen HKL, Chu SYY, et al. Lymphadenopathy of IgG4related 

sclerosing disease. Am J Surg Pathol 2008;32:671-81. 

5 Cheuk W, Lee KC, Chong LY, et al. IgG4-related Sclerosing disease: 

a potential new etiology of cutaneous pseudolymphoma. Am J Surg. 

Pathol 2009;33:1713-9. 

6 Green I, Dorfman RF, Rosai J. Breast involvement by extranodal 

Rosai-Dorfman disease: report of seven cases. Am J Surg Pathol 

1997;21(6):664-8. 

7 Donn W, Rebbeck P, Wilson C, et al. Idiopatic granulomatous mastitis. 

Arch Pathol Lab Med 1994;18:822-5. 

8 Shapiro JL, Goldblum JR, Petras RE. A clinicopathologic study 

of 42 patients with granulomatous gastritis. Am J Surg Pathol 

1996;20(4):462-70. 

9 Eusebi V, Millis RR. Epitheliosis, infiltrating epitheliosis, and radial 

scar. Semin Diagn Pathol 2010;27:5-12. 

10 Azzopardi JG, Ahmed A, Millis RR. Problems in breast pathology. 

London: W.B. Saunders Company 1979. 

11 Hamperl H. Strahlige narben und obliterierende mastopathie. Virchows 

Arch A Pathol Anat 1975;369:55-68. 

12 Eusebi V, Grassigli A, Grosso F. Lesioni focali sclero-elastotiche mammarie 

simulanti il carcinoma infiltrante. Pathologica 1976;68:507-18. 

13 Davies JD. Hyperelastosis, obliteration and fibrous plaques in major 

ducts of the human breast. J Pathol 1973;110:13-26. 

14 Tavassoli FA, Eusebi V. Tumors of the breast. 4 edn. Washington, DC: 

American Registry of Pathology/AFIP 2009. 

15 Wellings SR, Alpers CE. Subgross pathologic features and incidence 

of radial scars in the breast. Hum Pathol 1984;15:475-9. 

16 Jacobs TW, Byrne C, Colditz G, et al. Radial scars in benign breastbiopsy 

specimens and the risk of breast cancer. N. Engl. J Med 

1999;340:430-6. 

131 

Tab. I. infiltrating epitheliosis (ie), sclero-elastotic lesion (rs) and 

tubular carcinoma (tc). 

IE RS TC 

Epitheliosis Bulk of the 

lesion 

may be 

present at 

periphery 

not a feature 

Streaming of 

proliferating 

epithelial 

cells 

Present not a feature not a feature 

Zoning not a feature Present not a feature 

Obliterative 

mastopathy 

not a feature Present not a feature 

Sclerotic 

tissue 

Desmoplastic 

stroma 

Periductal 

elastosis 

Perivenous 

elastosis 

Glandular 

structures 

not a feature Present may be 

present 

Present not a feature Present 

not a feature Present may be 

present 

rare rare frequent 

solid 

“fingers” 

sometimes 

squamoid 

features 

entrapped angulated 

“tear drops” 

In situ 

carcinoma 

not a feature not a feature frequent 

Myoepithelial 

cells 

Present Present absent 

Basal lamina Present Present absent 

Molecularly targeted therapies in breast cancer: 

a rapidly evolving scenario 

F. Montemurro, E. Geuna, A. Milani, M. Aglietta 

Divisione Universitaria di Oncologia Medica I; Fondazione del 

Piemonte per l’Oncologia/IRCC Candiolo; Strada Provinciale 142, 

Km 3.95, 10060 Candiolo 

Over the last two decades, a greater understanding of the 

heterogeneous biology of breast cancer has resulted in the 

development of newer, molecularly targeted therapeutic approaches. 

Currently, the presence or absence of two main therapeutic 

targets, hormone receptors and HER2, recapitulates 

more complex biological definitions of breast cancer subtypes 

based on gene expression profiling 1 . Although endocrine 

therapy is historically considered the first form of biologically 

targeted therapy, the monoclonal antibody trastuzumab was 

the first compound that was rationally designed to target a 

biologically relevant alteration 2 . The evolution of HER2-targeting 

is a good example on how the concept of “biologically 

targeted therapy” has evolved over the years. 

HER2 is a tyrosine kinase receptor belonging to the epidermal 

growth factor receptor (EGFR) family 3 . Other members of 

this family include HER2, HER3 and HER4. Overexpression 

of HER2, which results from HER2 gene-amplification (from 

now on defined HER2-positivity), occurs in some 15-20% 

of all breast cancers and characterizes a biologically distinct 

subset of this disease 4 . HER2-positivity is associated with 

adverse histopathological features, propensity to metastasize 

to viscera and to the central nervous system, poor prognosis 

and resistance to endocrine therapy 4 . Compared with chemo-

132 

therapy alone, the addition of trastuzumab to chemotherapy 

boosts response rate, progression-free survival and overall 

survival in patients with metastatic disease 5 6 . In patients 

with operable, HER2-positive breast cancer, the inclusion 

of trastuzumab in adjuvant chemotherapy programs reduces 

the risk of relapse and prolongs survival 7-9 . More recently, 

several other HER2-targeting agents have shown clinical efficacy 

both in trastuzumab-naïve and in trastuzumab-resistant 

patients and several others are expected in the near future 10 . 

This tremendous research effort has become necessary because 

resistance to HER2-inhibition is a major challenge. In 

fact, as a single agent or in combination with chemotherapy, 

trastuzumab induces tumor regression in about 20-30% and 

60-70% of HER2-positive metastatic breast cancer patients, 

respectively 11 . Unfortunately, the vast majority of patients, 

including those with impressive initial responses, will ultimately 

show disease progression. 

Overcoming primary and acquired resistance to trastuzumab 

has been the focus of several preclinical and clinical investigations 

to increase the efficiency of HER2-targeting. 

These studies have clarified several aspects of the high level 

of interaction between signal transduction pathways, which 

account for the ability of cancer cells to circumvent inhibition. 

For example, tyrosine-kinase receptors can be seen as one 

layer of a complex, multilayered network 12 . Other layers are 

represented by extracellular ligands and downstream signalling 

pathways. By virtue of this architecture, a “core function” 

like for example proliferation or survival may be sustained by 

different effectors, in a bow-tie structure. This high level of 

integration is the result of an evolutionary process that started 

with a single ancestral tyrosine-kinase receptor, activated by 

one ligand and transmitting signals through a single cascade 

of intracellular mediators. 

The four EGFR family members have probably originated 

from a single receptor through gene duplication. Inactive 

monomers form homo- and heterodimeric structures with 

other members of the family, resulting in receptor activation 

and phosphorilation of downstream signalling effectors. 

HER2 has an “always-on” structure and lacks the capacity to 

interact with growth-factors ligands. HER3 has no tyrosine 

kinase activity. Despite this loss of functions, both HER and 

HER3 form hetherodimers with other EGFR members that 

are capable of generating potent cellular signals 3 . Apart from 

this “family-specific” cooperation, HER receptors can engage 

“external cooperation” with members of other families of 

tyrosine-kinase receptors, like for example the Insulin-like 

growth 1 receptor or with the estrogen receptor pathway 13 14 . 

Multiple ligands and intracellular cross-talk between signal 

transduction pathways complete this complex evolutionary 

network. This architecture has properties that are critical for 

both normal and cancer cells 12 . Robustness, which is the ability 

of the system to function despite external (environmental) 

and internal (genetic) perturbations, is ensured by modularity 

and redundancy. Furthermore, the system is able to learn how 

to circumvent common, single-hit perturbations (network 

training). It appears more and more evident that simultaneous 

targeting at several different levels in this multi-layered 

biological network is required for maximum clinical efficacy. 

Multiple targeting can be accomplished by using single agents 


with the ability to inhibit different substrates or by cocktails 

of selective or non-selective inhibitors. Furthermore, it can 

involve other members of the HER2 family or also connected 

“external” pathways. Examples of multiple targeting are 

already available in the clinic: pan-HER inhibitors, combinations 

of HER inhibitors with endocrine agents, antiangiogenic 

compounds and heat shock protein inhibitors 15 . HER2- negative 

tumors can be targeted successfully with antiangiogenetic 

agents 15 . Even “triple negative tumors” (hormone-receptors 

and HER2 negative) are no-longer a “targetless” subgroup 

since the therapeutic success achieved by PARP-inhibitors 16 . 

Due to several unanswered questions on the optimal use of 

these agents, this rapidly evolving scenario requires rigorously 

conducted clinical studies to select patients who are 

most likely to benefit from treatments. 

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breast tumours. Nature 2000;406:747-52. 

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antibody in patients with HER2/neu-overexpressing metastatic breast 

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3 Yarden Y, Sliwkowski MX. Untangling the ErbB signalling network. 

Nat Rev Mol Cell Biol 2001;2:127-37. 

4 Slamon DJ, Clark GM, Wong SG, et al. Human breast cancer: correlation 

of relapse and survival with amplification of the HER-2/neu 

oncogene. Science 1987;235:177-82. 

5 Slamon DJ, Leyland-Jones B, Shak S, et al. Use of chemotherapy plus 

a monoclonal antibody against HER2 for metastatic breast cancer that 

overexpresses HER2. N Engl J Med 2001;344:783-92. 

6 Marty M, Cognetti F, Maraninchi D, et al. Randomized phase II trial 

of the efficacy and safety of trastuzumab combined with docetaxel 

in patients with human epidermal growth factor receptor 2-positive 

metastatic breast cancer administered as first-line treatment: the 

M77001 study group. J Clin Oncol 2005;23:4265-74. 

7 Smith I, Procter M, Gelber RD, et al. 2-year follow-up of trastuzumab 

after adjuvant chemotherapy in HER2-positive breast cancer: a randomised 

controlled trial. Lancet 2007;369:29-36. 

8 Romond EH, Perez EA, Bryant J, et al. Trastuzumab plus adjuvant 

chemotherapy for operable HER2-positive breast cancer. N Engl J 

Med 2005;353:1673-84. 

9 Joensuu H, Bono P, Kataja V, et al. Fluorouracil, epirubicin, and cyclophosphamide 

with either docetaxel or vinorelbine, with or without 

trastuzumab, as adjuvant treatments of breast cancer: final results of 

the FinHer Trial. J Clin Oncol 2009;27:5685-92. 

10 Metzger-Filho O, Vora T, Awada A. Management of metastatic 

HER2-positive breast cancer progression after adjuvant trastuzumab 

therapy ΓÇô current evidence and future trends. Expert Opin Invest 

Drugs 2010;19:S31-9. 

11 Montemurro F, Valabrega G, Aglietta M. Trastuzumab-based combination 

therapy for breast cancer. Expert Opin Pharmacother 

2004;5:81-96. 

12 Citri A, Yarden Y. EGF-ERBB signalling: towards the systems level. 

Nat Rev Mol Cell Biol 2006;7:505-16. 

13 Nahta R, Yuan LX, Zhang B, et al. Insulin-like growth factor-I receptor/human 

epidermal growth factor receptor 2 heterodimerization 

contributes to trastuzumab resistance of breast cancer cells. Cancer 

Res 2005;65:11118-28. 

14 Bender LM, Nahta R. Her2 cross talk and therapeutic resistance in 

breast cancer. Front Biosci 2008;13:3906-12. 

15 Rosen LS, Ashurst HL, Chap L. Targeting signal transduction pathways 

in metastatic breast cancer: a comprehensive review. Oncologist 

2010;15:216-35. 

16 Fong PC, Boss DS, Yap TA, et al. Inhibition of poly(ADP-ribose) 

polymerase in tumors from BRCA mutation carriers. N Engl J Med 

2009;361:123-34.

lectures 

Standardization of techniques in anatomic pathology 

Nucleic acids extraction from ffPe tissues 

S. Bonin, G. Stanta 

ACADEM Department-Cattinara Hospital-University of Trieste, Italy 

Background. Archival formalin-fixed and paraffin embedded 

tissues are the most abundant supply of clinical material. 

Formalin fixation and paraffin embedding represents the standard 

methods used in any hospital to process clinical tissues, 

allowing permanent preservation of tissues, with easy storage 

and optimal histologic quality. However, nucleic acids extractions 

and analysis from formalin-fixed and paraffin embedded 

tissues still remains a hang-up issue, as formalin induces crosslinkage 

of nucleic acids to proteins and covalently modifies 

the bases of nucleic acids, impairing extraction of macromolecules 

1 . Nevertheless, it is possible to extract and analyse 

nucleic acids from formalin fixed paraffin embedded tissues 

(FFPE). Mostly, PCR based assays are performed in FFPE 

samples. At the present time most of the methods applied in 

molecular pathology are laboratory-based assays and commercial 

kits not directly intended for diagnostic purposes. Therefore 

there is a need to establish guidelines with standardised 

procedures for molecular methods, starting from an analysis 

of the currently used methods of nucleic acids extraction. A 

collaborative study including 13 European laboratories of the 

IMPACTS group (www.impactsnetwork.eu) has been carried 

out to evaluate the performance of nucleic acids extractions 

using the same formalin-fixed paraffin-embedded specimens 

to assess if the different methodologies used for nucleic acids 

extraction in different laboratories might affect the results 2 . 

By the use different protocols, but the same tests for quality 

controls, the authors demonstrated that most of the homemade 

protocols and commercial kits allow the extraction of nucleic 

acids, but for data comparison quality tests are needed 2 . 

Materials and Methods. The extractions procedures for 

DNA and RNA differ for some main aspects related to the 

different characteristics of the two macromolecules. The 

extraction protocols we analysed for DNA could be mainly 

divided in three groups: 

1. DNA extraction with alcohol precipitation of the DNA. 

2. DNA extraction without further precipitation or purification 

(crude extract). 

3. DNA extraction using commercial kits following silica 

based adsorption columns for DNA purification. 

For RNA the methods could be roughly divided into: 

a. RNA extraction with phenol extraction and isopropanol 

precipitation (homemade protocols and commercial solutions). 

b. RNA extraction using silica based columns for purification. 

Nucleic acids quantity was detected by means of spectrophotometer 

measurements, while nucleic acids’ quality was 

assessed by means of PCR and RT-PCR analysis with increasing 

lengths amplicons. 

Results. As a general consideration for both DNA and RNA, 

the purity and quantity assessment by spectral photometry did 

not correlate with the maximum amplifiable length. 

DNA extraction protocols that used purification of the extracted 

DNA, gave comparable results in terms of yield and 

purity of the DNA. DNA quality, assessed by PCR-amplifi- 

Moderators: G. Bussolati (Torino), G. Stanta (Trieste) 

133 

ability was not drastically affected by the use of different 

DNA-extraction protocols, without a single DNA-extraction 

protocol prevailing on others 2 . 

For RNA, the isolation methods affected the yield of the 

extracted RNA, even when extraction conditions are similar. 

Regarding the amplifiability, all extraction methods resulted 

in RNA of useful quality for expression analyses in archival 

and diagnostic tissues, since shorter amplicons are sufficient 

for quantitative PCR 2 . However, column based methods provided 

best RNA quality assessed by RT-PCR. 

In any case to overcome the inter-laboratory variability, further 

standardization of the techniques, especially quality control 

procedure, is recommended for research and diagnostic 

applications in molecular pathology. 

references 

1 Dotti I., Bonin S., Basili G., et al. Effects of formalin, methacarn, 

and fineFIX fixatives on RNA preservation. Diagn Mol Pathol 

2010;19:112-22. 

2 Bonin S., Hlubeck F., Benhattar J., et al. Multicentre Validation Study 

of Nucleic Acids Extraction from FFPE Tissues. Virchow Archiv (in 

press). 

role of microrNAs in defining tissue of origin of 

metastatic cancer 

A. Vecchione 

Roma 

MicroRNAs (miRNAs) are evolutionarily conserved, endogenous, 

small non-coding RNA molecules processed from 

precursors and in their mature forms, serve as important 

gene regulators that have the capacity to down-regulate gene 

expression through translation inhibition and promotion of 

miRNA degradation mediated by specific target site binding 

to the 3’-untranslated region of target genes. According to 

the most recent version of miRBase (v.15.0), 4000 different 

mature miRNA sequences have been identified in humans 

to date. A unique attribute of miRNAs that renders them potentially 

useful for the molecular diagnosis of tumors is their 

tissue and cell lineage specificity. Many of the miRNAs are 

highly specific in their expression in specific tissues and cell 

types, and this specificity is often retained in the corresponding 

tumor tissues. Identification of cell origin by profiling of 

miRNAs is more efficient compared with global analysis of 

mRNAs, because the former is not confounded by such a large 

pool of irrelevant genes because of the relatively small number 

of miRNA species. Therefore, miRNAs could facilitate the 

accurate diagnosis of tumors that are difficult to classify with 

respect to the tissue origin by conventional means, for example, 

metastatic cancer of unknown primary origin, a highly 

aggressive malignancy that poses diagnostic and management 

difficulties. Deregulation of miRNAs occurs frequently during 

tumorigenesis, making them attractive candidates for molecular 

detection of malignancy. A subset of miRNAs can often 

be found up- or down-regulated in tumors compared with the 

normal tissues in a specific tumor type or more globally in a 

number of different tumor types. Not only is the identification 

of these miRNAs critical in the understanding of the pathogenetic 

role of miRNAs in cancer development, it can also 

provide a diagnostic methodology for distinguishing tumors

134 

from normal tissues of different cell origins. In the current 

setting of clinical diagnostic practice, where morphological 

and antigenic evaluation appears to be adequate for accurate 

diagnostic separation between tumor and normal tissues in 

the vast majority of biopsies, the utility of miRNA analysis in 

this arena may not be too apparent. However, the differential 

expression of miRNAs between tumors and normal tissues 

may be exploited in the diagnosis of samples where cells are 

scant or poorly preserved, which may render diagnosis of a 

malignancy by traditional methods difficult. 

Despite the many exciting developments that demonstrate the 

potential capacity of miRNAs for tumor classification and 

prognosis, their practical use as biomarkers in a routine clinical 

setting is still in its infancy. To ascertain and accelerate its 

advancement beyond the developmental phase, efforts should 

be made to perform retrospective and prospective studies in 

global and gene-specific analysis of miRNAs on multiple 

independent cohorts of patient samples using standardized 

methodologies. 

fish 

C. Marchiò, P. Gugliotta, C. Botta, L. Verdun di Cantogno, 

A. Sapino 

Department of Biomedical Sciences and Human Oncology, Turin, 

Italy 

In Situ hybridization (ISH), is a molecular technique that has 

been available since 1969 and over time it has become part 

of the diagnostic armamentarium of pathologists in different 

fields (Lambros et al, 2007). Unlike other molecular biology 

techniques ISH is unique as it is based on a visual assessment 

of probe copy numbers directly at the microscope and can 

be performed on interphase nuclei, i.e. on tissue sections of 

archival samples (Lambros et al, 2007). Two modalities of 

in situ hybridisation have already been introduced in pathology 

laboratories: fluorescent in situ hybridisation (FISH) 

and chromogenic in situ hybridisation (CISH). FISH is most 

widely used and has already made prime time in terms of 

diagnosis, prognosis and prediction in several diseases (sarcomas, 

lymphomas and leukaemias, breast cancer, oligodendrogliomas, 

gastric cancer, melanomas). 

The success of this techniques stems from the fact that they 

allow for semi-quantitative assessment of gains, losses and 

amplifications directly on tissue sections, combining molecular 

genetics with traditional pathology (Marchiò and Reis 

Filho, 2008). FISH can be performed on both histological 

and cytological specimens, according to distinct protocols. In 

particular, for histological samples the pre-analytic phase of 

tissue collection, preservation and preparation is of paramount 

importance in order to obtain a good performance of the assay. 

Ideally tissue specimens should be fixed immediately after 

gross sampling (5mm thick tissue sample slices); neutralbuffered 

formalin is recommended, alcohol-based fixatives 


can be used as well, however they often give a fluorescent 

background which can be troublesome for interpretation of the 

results; the time of fixation should range from 6 to 24 hours. 

A second and crucial step in represented by section cutting: in 

order to be able to appreciate the signals and not to underestimate 

chromosome abnormalities, sections are recommended 

to be 2-5 µ thick. It has to be mentioned that FISH analysis is 

also feasible on Tissue Micro Array (TMA) sections, following 

a specific protocol (Sapino et al, 2006): once again, the 

pre-analytical phase, with proper sampling of tumour area and 

high quality tissue cutting, plays a central role. 

The analytic phase includes different variables, such as assay 

validation, equipment calibration, use of standardized laboratory 

procedures, training and competency assessment of staff, 

type of pre-treatments, test reagents, use of standardized 

control materials and automated laboratory methods (Wolff 

et al, 2007). These variables are closely related with laboratory 

competency and efficiency, thus suggesting such type 

of analyses should be delegated to specific laboratories with 

long-standing experience and high work-load of the assay. 

Automated machines for FISH analysis have been recently 

introduced and are of assistance to the pathologist in scanning 

and scoring areas of interest. 

Post-analytic phase issues are represented by interpretation 

criteria, use of image analysis, reporting elements, laboratory 

accreditation, pathologist competency assessment and 

quality assurance procedures. Undoubtedly, quality controls 

(QCs) represent a very important mechanism, even though 

a consensus has not been reached on the way QCs should be 

organized. In Italy, we have recently set up a “ring study” 

for QC of FISH analysis 11 institutions are have adhered to: 

single Institutions make cases with different amplification 

status circulate among the others to perform FISH in their 

own laboratories. Afterwards data are centrally collected and 

analysed by a referral centre, with particular attention to consensus 

on reported results. 

Taken together, all variables discussed in different phases of 

FISH analysis play a role in the definition of a good FISH experiment, 

therefore standardization, competency of operators 

(technicians, biologists, pathologists) and quality check are 

eagerly warranted, as also pointed out in different guidelines 

by experts of single diseases (see for example (Wolff et al, 

2007)). 

references 

Lambros MB, Natrajan R, Reis-Filho JS. Chromogenic and fluorescent in 

situ hybridization in breast cancer. Hum Pathol 2007;38:1105-22. 

Marchiò C, Reis Filho JS. Molecular diagnosis in breast cancer. Diagnostic 

Molecular Pathology 2008;14. 

Sapino A, Marchiò C, Senetta R, et al. Routine assessment of prognostic 

factors in breast cancer using a multicore tissue microarray procedure. 

Virchows Arch 2006;449:288-96. 

Wolff AC, Hammond ME, Schwartz JN, et al. American Society of Clinical 

Oncology/College of American Pathologists guideline recommendations 

for human epidermal growth factor receptor 2 testing in breast 

cancer. J Clin Oncol 2007;25:118-45.

lectures 

Thyroid pathologies: 

non-conventional thyroid lesions and emerging diagnostic problems 

Clinical-pathological and molecular features of 

papillary thyroid carcinomas smaller than 2 cm 

F. Basolo, L. Torregrossa, R. Giannini, M. Miccoli * , 

C. Lupi, E. Sensi, P. Berti, R. Elisei ** , P. Vitti ** , A. Baggiani * , 

P. Miccoli 

Department of Surgery, University of Pisa, Italy; * Department of 

Experimental Pathology B.M.I.E., Biostatistics Research Unit, University 

of Pisa, Italy; ** Department of Endocrinology, University of 

Pisa, Italy 

Background. The actual incidence of thyroid cancer is predominantly 

characterized by the increased detection of small 

PTCs: about 87% of cases consist of cancers measuring 2 cm 

or smaller 1 . According to TNM staging system, evaluation of 

the degree of neoplastic infiltration beyond the thyroid capsule 

remains a unique parameter that can be evaluated by histopathologic 

examination to label a Papillary Thyroid Carcinoma 

(PTC) measuring ≤ 2 cm in size as a pT1 or pT3 tumor 2 . PTCs 

smaller than 2 cm in size that are limited to the thyroid gland 

and without lymph node metastasis are considered low-risk tumors 

and can be successfully treated with total thyroidectomy 

alone, whereas PTCs of the same size but with extrathyroidal 

extension justify a more aggressive treatment 3 . 

In recent years, BRAF V600E has emerged as a promising 

prognostic factor in the risk stratification of PTC 4 . Many studies 

have demonstrated significant correlations between BRAF 

mutation and high-risk clinical-pathological features of PTCs 

in overall analyses of tumors of all sizes 4 . 

In the current study, we correlated the BRAF V600E mutation 

with both clinical-pathological features and the degree of 

neoplastic infiltration to redefine the reliability of the actual 

system of risk stratification. 

Methods. The presence of BRAF mutations was examined 

in a large group of PTCs smaller than 2 cm (overall 1,060 

patients: 254 males and 806 females, mean age 44.6 ± 13.3 

years) divided into four degrees of neoplastic infiltration: 

a) “totally encapsulated”; b) “not encapsulated without thyroid 

capsule invasion”; c) “thyroid capsule invasion”; and 

d) “extrathyroidal extension.” 

Results. The overall frequency of the BRAF V600E mutation 

was 44.6%. In both univariate and multivariate analyses, 

BRAF mutations showed a strong association with PTC variants 

(classical and tall cell), tumor size (1,1-2 cm), multifocality, 

absence of tumor capsule, extrathyroidal extension, lymph 

node metastasis, and higher AJCC stage. In PTCs staged as 

pT1 with thyroid capsule invasion, the frequency of BRAF 

mutations was significantly higher than in pT1 tumors that did 

not invade the thyroid capsule (67.3% vs. 31.8%, respectively, 

p < 0.0001). No statistically significant difference in BRAF alterations 

was found between pT1 tumors with thyroid capsule 

invasion and pT3 tumors (67.3% and 67.5%, respectively). 

In conclusion, we suggest that evaluation of BRAF status 

even in pT1 tumors would be useful to improve risk stratification 

and patient management, although follow-up data 

are necessary. 

references 

1 Davies L, Welch HG. Increasing incidence of thyroid cancer in the 

United States, 1973-2002. JAMA 2006;295:2164-7. 

Moderators: M. Papotti (Torino), G. Gardini (Reggio Emilia) 

135 

2 Edge SB, Byrd DR, Carducci MA, et al. American Joint Committee on 

Cancer (AJCC). Cancer staging manual. Seventh edition. New York: 

Springer: 2009. 

3 Pacini F, Schlumberger M, Dralle H, et al. European consensus for the 

management of patients with differentiated thyroid carcinoma of the 

follicular epithelium. Eur J Endocrinol 2006;154:787-803. 

4 Xing M. BRAF mutation in papillary thyroid cancer: pathogenic role, 

molecular bases, and clinical implications. Endocr Rev 2007;28:742- 

62. 

Mitochondrial DNA changes and oncogenic 

mutation of nuclear genes in thyroid oncocytic 

nodules: BrAf v600e and reT/PTC are associated 

with papillary carcinomas showing oncocytic 

features, but rAS mutations are uncommon in 

oncocytic follicular adenomas and carcinomas 

D. De Biase, E. Bonora * , L. Morandi, G. Gasparre * , G. Romeo 

* , G. Tallini 

Sezione di Anatomia, Istologia e Citologia Patologica “M. Malpighi”, 

Univerisità di Bologna, Ospedale Bellaria, Via Altura 3, 40139 Bologna, 

Italy; * U.O. Genetica Medica, Dipartimento di Scienze Ginecologiche, 

Ostetriche, Pediatriche Policlinico “S. Orsola-Malpighi” 

Background. Oncocytic neoplasms are tumors composed of 

cells characterized by an aberrant amount of mitochondria that 

is responsible for their ‘swollen’ (i.e. ‘oncocytic’, from the 

greek onkoustai, to swell) appearance 1 . These neoplasms may 

occur at various sites but are particularly common in the thyroid 

gland. Thyroid oncocytic tumors (with the exception of 

the rare oncocytic variant of medullary carcinoma) originate 

from follicular cells 1 . They can be benign (oncocytic adenomas) 

or malignant (oncocytic carcinomas) and have been the 

subject of both fascination and controversy for pathologists. 

One area of disagreement has concerned the definition of 

thyroid oncocytic tumors as a separate tumor category 1 . It is 

now accepted that oncocytic tumors in the thyroid should be 

viewed as special subtypes or variants, since their features 

are distinct enough to set them apart from corresponding neoplasm 

lacking accumulation of mitochondria 2 . Accordingly, 

oncocytic thyroid carcinomas are now classified as variants 

of follicular carcinomas (commonly) or of papillary carcinomas 

(less commonly) 2 . The obvious cellular derangement of 

oncocytic cells, with complete dysregulation of the mitochondrial 

mass and metabolism, have spurred some investigators 

to study the molecular mechanisms underlying the genesis 

of this peculiar phenotype 3 . Disruptive mitochondrial DNA 

(mtDNA) mutations in complex I subunits of the respiratory 

chain have recently been shown to be very common in thyroid 

oncocytic lesions, after the entire mtDNA of many cases has 

been sequenced, while in vitro experiments have demonstrated 

that complex I mtDNA mutations actually cause the 

decreased production of ATP associated with the oncocytic 

phenotype 3-5 . 

Somatic alterations of various nuclear genes are known 

to occur in thyroid tumors, including point mutations and 

rearrangements 6 . The most frequent oncogenic alterations 

involve the RET, RAS and BRAF genes 6 . These genes code 

for proteins involved in the linear signalling that goes from 

the tyrosine kinase receptors at the plasma membrane and

136 

RAS to cytoplasmic Ser/Thr kinases like BRAF, MEK, ERK 

- the MAPK pathway - whose activity is pivotal in controlling 

cell proliferation 6 . While these oncogenic events in thyroid 

tumors that are not oncocytic have been well studied, their 

prevalence in oncocytic thyroid lesions is unclear. Unclear is 

also their relationship with mtDNA mutations. 

Methods. H-, K-and N-Ras mutations as well as BRAF exon 

15 mutation and RET/PTC rearrangements were analyzed in 

45 thyroid oncocytic tumors (15 hyperplastic adenomatous 

nodules, HANonc; 8 follicular adenomas, FAonc; 14 follicular 

carcinomas, FConc; 8 papillary carcinomas with oncocytic 

features, PConc) that had been previously characterized for 

their mtDNA abnormalities 4 . Nucleic acids were extracted 

from paraffin-embedded neoplastic tissue after examination 

of the corresponding Hematoxylin and Eosins stained sections 

using a commercial kit (RecoverAll Total Nucleic Acid 

Isolation, Applied Biosystems/Ambion – Austin, TX). Direct 

sequencing was used to analyze H-, K- and N-Ras and BRAF 

exon 15, qRT-PCR to identify RET/PTC1 and RET/PTC3 rearrangement. 

The entire mtDNA was sequenced and mtDNA 

mutations were classified as disruptive, possibly/probably 

damaging and absent. 

Results. We found three cases with RAS mutations, two 

cases with a BRAF V600E activating mutation, one case with a 

RET/PTC1 rearrangement. All the above nuclear DNA alterations 

did not overlap in any given tumor. Of the RAS mutated 

cases, one had a NRAS Q61R mutation and was diagnosed as a 

minimally invasive FConc; the second case had a KRAS Q61R , 

and was diagnosed as FAonc; a third case had HRAS Q61R mutation, 

was diagnosed as a follicular variant PConc; mtDNA 

mutations were identified in the last two cases, and in both 

the mtDNA mutations were classified as possibly/probably 

damaging. Both cases with the BRAF V600E activating mutation 

were diagnosed as PConc, tall cell variant and did not have 

mtDNA mutations. The single RET/PTC1 mutated case was 

a Warthin-like PConc, with no mtDNA alterations. mtDNA 

mutations classified as disruptive were identified in 5/14 

(35.7%) FConc, 2/8 (25.0%) FAonc, 4/15 (26.7%) HANonc, 

1/8 (12.5%) PConc. mtDNA mutations classified as possibly/ 

probably damaging were identified in 3/14 (21.4%) FConc, 

4/8 (50.0%) FAonc, 3/15 (20.0%) HANonc, 3/8 (37.5%) 

PConc. 

Conclusion. RAS, BRAF V600E mutations and RET/PTC rearrangement 

have been identified in malignant oncocytic tumors 

and in one follicular adenoma. RAS mutations are uncommon 

in oncocytic follicular neoplasms (both carcinomas and 

adenoma), suggesting that other tumorigenic events may play 

a role in their development. BRAF V600E mutations are associated 

with tall cell variant papillary carcinomas with oncocytic 

features. Pathogenic mtDNA alterations may overlap with the 

oncogenic mutations commonly found in non-oncocytic thyroid 

tumors. Disruptive mtDNA mutations are more common 

in oncocytic follicular carcinomas than in papillary oncocytic 

carcinomas. They are also more common in oncocytic follicular 

neoplasms – both carcinomas and adenomas – and 

hyperplastic adenomatous nodules with oncocytic features, 

when compared with papillary oncocytic carcinomas. 

references 

1 Tallini G. Oncocytic tumors. Virchows Archives A (Anat Pathol) 

1998;433:5-12. 

2 World Health Organization Classification of Tumors-Pathology and 

Genetics, Tumors of Endocrine Organs. Lyon (France): IARC Press, 

2004. 

3 Gasparre G, Bonora E, Tallini G, et al. Molecular features of thyroid 

oncocytic tumors. Mol Cell Endocrinol. 2010;321:67-76. 


4 Gasparre G, Porcelli AM, Bonora E, et al. Disruptive mitochondrial 

DNA mutations in complex I subunits are markers of oncocytic phenotype 

in thyroid tumors. Proceedings of the National Academy of 

Sciences USA 2007;104:9001-6. 

5 Bonora E, Porcelli AM, Gasparre G, et al. Defective Oxidative Phosphorylation 

in Thyroid Oncocytic Carcinoma Is Associated with 

Pathogenic Mitochondrial DNA Mutations Affecting Complexes I and 

III. Cancer Research 2006;66:6087-96. 

6 Knauf JA, Fagin JA. Role of MAPK pathway oncoproteins pathogenesis 

and as drug targets. Current Opinion in Cell Biology 2009;21:296- 

303. 

Trabecular neoplasms of the thyroid 

M. Volante, I. Rapa, M. Papotti. 

Department of Clinical and Biological Sciences at San Luigi Hospital, 

University of Turin, Orbassano, Turin, Italy 

Follicular and papillary growth patterns represent the most 

common architectural features within thyroid nodules, in both 

benign and malignant settings. Alternative to these, nodules 

having a non follicular-non papillary structure may be encountered, 

being solid/trabecular arrangement the most common 

feature. In general, irrespective of the biological nature of 

the lesion under analysis, trabecular growth is represented by 

sheets of cells regularly arranged in one or few rows or more 

irregularly anastomosing, separated by usually scarce connective 

tissue and a thin vascular network. The solid growth is an 

extreme of the trabecular architecture, being represented by a 

more nodular arrangement with a wider thickness of cellular 

islands and a more irregular and dispersed vascular network. 

However, the border between compact trabecular growth and 

solid pattern is poorly defined and since this latter is usually 

mixed with and represents an architectural arrangement parallel 

to the trabecular one, they will be considered together. 

When dealing with a trabecular lesion in the thyroid, a wide 

range of differential diagnoses exists, representing one of the 

major diagnostic problems in the routine thyroid practice 1 , 

and include the following, among others: 

a) Lesions derived from the follicular epithelium with 

papillary carcinoma-type nuclear features. When follicular 

cell derivation is morphologically or immunophenotypically 

evident, the nuclear features – as conventionally considered 

for follicular/papillary lesions – should be carefully examined 

to check the presence of the diagnostic features of papillary 

carcinoma. If clear-cut papillary-type nuclei are recognized, 

the following two entities have to be considered. The 

solid variant of papillary carcinoma is a rare and still poorly 

characterized variant of papillary thyroid carcinoma, most 

commonly found in children and young adults especially in 

radiation-exposed individuals; the presence of irregular clear 

nuclei with grooving and pseudo-inclusions is the cytological 

hallmark whereas the solid growth pattern is accompanied 

by vascular invasion and extra-thyroidal extension in about 

one-third of cases. The clinical behaviour of the solid variant 

of papillary carcinoma is characterized by a slightly higher 

frequency of distant metastases and less favourable prognosis 

than classical papillary carcinoma 2 . Hyalinizing trabecular 

tumor (HTT) is a trabecular neoplasm of follicular derivation 

with peculiar nuclear, architectural and histochemical 

features, with a benign behaviour in the vast majority of 

cases reported so far 3 . HTT is a well circumscribed lesion, 

lacking morphological signs of capsular or vascular invasion. 

Two principal features are diagnostic of HTT: a uniform 

and diffuse solid and trabecular architecture, with markedly 

hyalinized deposits containing basal membrane-type material, 

typically located within the trabeculae rather than in the inter-

lectures 

posed stroma together with nuclear features, including clear 

nuclei with grooves and pseudoinclusions, resembling those 

of papillary carcinoma. 

b) Lesions derived from the follicular epithelium with 

dark round/convoluted nuclei. Trabecular (embryonal) 

adenoma is a variant of follicular adenoma, both of conventional 

and oncocytic types, with predominant/pure trabecular 

growth pattern associated to high cellularity and oedematous 

modifications of the stroma. The name embryonal adenoma is 

related to the morphologic resemblance of this tumor to the 

early stages of the developing thyroid. Follicular carcinoma 

with solid/trabecular growth pattern: focal or predominant 

trabecular growth may be observed in follicular carcinoma, 

similarly to follicular adenoma but with a higher frequency, 

especially in oncocytic forms. The differential diagnosis with 

adenomas relies on the identification on capsular and/or vascular 

invasion, whereas its distinction from poorly differentiated 

carcinoma is outlined below. Poorly differentiated carcinoma 

is characterized by a predominant trabecular, insular 

and/or solid growth together with the presence of unequivocal 

high grade histology, high mitotic count and necrosis 4 . 

c) Primary thyroid tumors not derived from the follicular 

epithelium. Medullary carcinoma. As stated in the WHO 

classification, the diagnosis of medullary carcinoma should 

be considered in any thyroid nodule showing unusual features. 

Its histological appearance is widely variable, being 

tumor cells most commonly arranged in nests or trabeculae 

with a variable amount of fibrovascular stroma. The presence 

of amyloid deposition is characteristic but not constant. Cytological 

features suggestive of medullary carcinoma are the 

presence of round to oval nuclei, without prominent nucleoli 

and with coarse chromatin. Primary thyroid paraganglioma 

is a very rare tumor showing a striking female predominance, 

it is usually confined to the thyroid gland and composed of 

neoplastic cells arranged in the typical lobular growth with 

fine connective tissue interposed. Architecture and cytology 

are similar to those of medullary carcinoma which shares a 

common neuroendocrine origin and represents the major differential 

diagnosis, but the absence of calcitonin immunoreactivity 

rules out the diagnosis of the latter. 

d) Extra-thyroidal lesions. Parathyroid lesions: intra-thyroidal 

parathyroid tissue is not uncommon and should be 

searched for during surgical interventions for primary or secondary 

hyperparathyroidism. When clinical hyperparathyroidism 

is not evident, hyperplastic or adenomatous parathyroid 

tissue showing typical trabecular arrangement might be confused 

with follicular cell-derived nodules or even medullary 

carcinoma. In the presence of a follicular patterned associated 

component, the recognition of bi-refringent crystals is useful 

in distinguishing thyroid from parathyroid gland tissues. More 

complicated is the differential diagnosis between malignant 

thyroid nodules and parathyroid carcinoma involving the 

thyroid gland, which present vascular and capsular invasion, 

trabecular growth with sometimes oncocytic changes, and in 

a fraction of cases necrosis and mitotic activity. Metastases: 

despite its high vascularisation, the thyroid is not a frequent 

site of tumor spread, according to clinical evidence. Metastases 

have as most common primary sites the kidney, lung, 

breast and gastrointestinal tract. In most instances, thyroid 

metastases present as solitary masses thus entering in the 

differential diagnosis with primary thyroid nodules. With special 

reference to kidney primaries, the clear cell/oncocytoid 

features and frequent trabecular growth might be source of 

misdiagnosis with other benign and malignant trabecular lesions 

primary of the thyroid. 

137 

references 

1 Volante M, Papotti M. A practical diagnostic approach to solid/trabecular 

nodules in the thyroid. Endocr Pathol 2008;19:75-81. 

2 Nikiforov YE, Erickson LA, Nikiforova MN, et al. Solid variant of 

papillary thyroid carcinoma: incidence, clinical-pathologic characteristics, 

molecular analysis, and biologic behavior. Am J Surg Pathol 

2001;25:1478-84. 

3 Carney JA, Hirokawa M, Lloyd RV, et al. Hyalinizing trabecular 

tumors of the thyroid gland are almost all benign. Am J Surg Pathol 

2008;32:1877-89. 

4 Volante M, Collini P, Nikiforov YE, et al. Poorly differentiated thyroid 

carcinoma: the Turin proposal for the use of uniform diagnostic 

criteria and an algorithmic diagnostic approach. Am J Surg Path 

2007;31:1256-64. 

Clinicopathological and prognostic features 

of well-differentiated capsulated carcinomas 

S. Piana, G. Gardini 

Department of Pathology, Arcispedale “Santa Maria Nuova”, Reggio 

Emilia, Italy 

Background. There is a conspicuous number of well-differentiated 

thyroid neoplasms of follicular cells characterized 

by encapsulation and a follicular pattern of growth (“follicular-patterned 

tumors”) which are currently designated as 

malignant if they show evidence of capsular/vascular invasion 

and/or exhibit the nuclear features of the papillary family of 

neoplasms 1 2 . 

The vast majority of these tumors have an excellent prognosis, 

but an aggressive therapy is often unnecessarily carried out. 

This group comprises firstly the minimally invasive follicular 

carcinoma (MIFCa) and the encapsulated follicular variant of 

papillary carcinoma (FV-PTC). The category of MIFCa continues 

to be controversial and a consensus regarding the minimal 

criteria for its diagnosis is still missing 3 . However, some 

studies have indicated that capsular invasion in the absence of 

vascular invasion does not appear to significantly affect the 

outcome of these tumors 4 . 

The FV-PTC is currently defined as a thyroid malignancy 

with a predominant or exclusive follicular growth pattern displaying 

the characteristic nuclear features of papillary thyroid 

carcinoma 5 . Multiple studies have demonstrated great interobserver 

variability in the diagnosis of these tumors, even 

among experts in thyroid pathology, thus emphasizing the difficulties 

in properly defining the criteria for the diagnosis of 

this particular type of papillary thyroid carcinoma 6 . The most 

difficult circumstance for diagnosis arises when these tumors 

are well-circumscribed an encapsulated. 

To these categories, the Chernobyl Pathologists Group 7 has 

proposed the addition of the well-differentiated carcinomas, 

not otherwise specified (WDC) for tumors with features 

intermediate between FCa and FV-PTC, and well-differentiated/follicular 

tumor of uncertain malignant behaviour 

(WDT-UMP and FT-UMP) for tumors with “incomplete” 

nuclear changes and/or “incomplete” capsular invasion, respectively. 

This alternate terminology reflects our current 

incomplete knowledge and it offers the advantage of avoiding 

unnecessary aggressive treatment for a tumor that shows an 

overwhelmingly innocuous behaviour following conservative 

surgery. 

Methods. If the above mentioned types of well-differentiated 

encapsulated carcinomas have an excellent prognosis, they 

should not be represented in a series of fatal thyroid carcinomas 

comprising all histologic types. Therefore, the files of 

the Department of Pathology of the Arcispedale Santa Maria 

Nuova in Reggio Emilia, Italy, were searched for all cases di-

138 

agnosed as thyroid carcinoma of any type from 1979 to March 

2004. A total of 1039 cases was found. Representative slides 

from each case were selected and re-classified according to 

the criteria recommended by standard texts 1 2 with the addition 

of the categories suggested by the Chernobyl pathology 

group 7 . Follow-up and clinical information were obtained 

from the Reggio Emilia Cancer Registry and from the files of 

the Pathology and Endocrinology Department. Follow-up was 

available in 1009 cases and ranged from 4.5 to 29 years (median, 

9.8 years; mean, 11.9 years) or until death. Among the 

1009 cases, 159 patients had died; thyroid carcinoma was the 

cause of death for 67 of the 159 patients, and these 67 cases 

are the focus of the study. 

Results. Among the 67 patients deceased as a consequence of 

thyroid carcinoma, there were none of the tumors belonging to 

any of the categories above mentioned, that is MIFCa, FV-PTC, 

WDT-UMP, and FT-UMP. In fact, the vast majority of these 

tumors shows a shows an exceedingly innocuous behaviour 

following conservative surgery. The overdiagnosis of this condition 

may lead to excessive treatment, including total thyroidectomy 

followed by radioactive iodide therapy. This acquires a 

particular importance with the encapsulated variant of FV-PTC, 

which is associated with an excellent prognosis and for which 

distant blood-borne metastasis has been rarely documented 8 . 

The results of this study and a critical review of the pertinent 

literature indicate that tumors with these features are associated 

with an extremely favourable outcome and that they 

do not play a significant role in the fatality rate of thyroid 

carcinoma 9 10 . 

references 

1 DeLellis RA, Lloyd RV, Heitz PU, et al. Tumours of Endocrine Organs, 

World Health Organization Classification of Tumours; Pathology 

and Genetics. Lyon: IARC Press 2004. 

2 Rosai J, Carcangiu ML, DeLellis RA. Tumors of the thyroid gland, 

Atlas of Tumor Pathology, Third Series, Fascicle 5, Washington, D.C: 

Armed Forces Institute of Pathology, AFIP 1992. 

3 Franc B, De La Salmoniere P, Lange F, et al. Interobserver and intraobserver 

reproducibility in the histopathology of follicular thyroid 

carcinoma. Hum Pathol 2003;34:1092-100. 

4 Van Heerden JA, Ray ID, Goellner JR, et al. Follicular thyroid carcinoma 

with capsular invasion alone: a non-threatening malignancy. 

Surgery 1992;112:1130-8. 

5 Carcangiu ML, Zampi G, Pupi A, et al. Papillary carcinoma of the thyroid. 

A clinicopathologic study of 241 cases treated at the University 

of Florence, Italy. Cancer 1985;55:805-28. 

6 Lloyd RV, Erickson LA, Casey MB, et al. Observer variation in the 

diagnosis of follicular variant of papillary thyroid carcinoma. Am J 

Surg Pathol 2004;28:1336-40. 

7 Williams ED (on behalf of the Chernobyl Pathologists Group). Two 

proposals regarding the terminology of thyroid tumors. Intern J Surg 

Pathol 2000;8:181-4. 

8 Chan JKC. Strict criteria should be applied in the diagnosis of encapsulated 

follicular variant of papillary thyroid carcinoma. Am J Clin 

Pathol 2002;117:16-18. 

9 Piana S, Frasoldati A, Di Felice E et al. Encapsulated well-differentiated 

follicular-patterned thyroid carcinomas do not play a significant 

role in the fatality rates from thyroid carcinoma. Am J Surg Pathol 

(E-pub ahead of print). 

10 Rosai J. The encapsulated follicular variant of papillary thyroid carcinoma; 

back to the drawing board. Endocr Pathol 2010;21:7-11. 


Vascular lesions of the thyroid 

M. Papotti, M. Volante. 

Department of Clinical and Biological Sciences, University of Turin 

at San Luigi Hospital, Orbassano (Torino), Italy 

Vascular lesions of the thyroid gland include benign endothelial 

proliferations of reactive (Masson’s “hemangioma”), 

benign neoplasms (cavernous hemangioma) and the rare 

malignant angiosarcomas. These latter occur in pure form 

or combined with anaplastic carcinoma (angio-sarcomatoid 

carcinoma). 

Reactive endothelial proliferations. In long standing nodular 

goiter, regressive changes are common, including oedema, 

fibrosis and calcification. Haemorrhage is an additional 

event, which can be associated to complete nodule infarction, 

followed by reparative processes such as granulation tissue 

and reactive endothelial hyperplasia, closely resembling 

intravascular papillary endothelial proliferations (Masson’s 

phenomenon). In these cases, intraluminal papillary projections 

in vascular spaces are lined by plump endothelial cells 

with occasional atypias possibly leading to a suspicion of 

malignancy. This condition may be an uncommon consequence 

of fine needle aspiration biopsy or the result of spontaneous 

intranodular haemorrhage/infarction. Completely 

infarcted goiter nodules are a challenge for clinicians, radiologists 

and pathologists: at ultrasound investigation, such 

nodules having prominent vascular endothelial hyperplasia 

are typically hyporeflecting and unhomogeneous and/or 

calcified, all features simulating malignancy. At light microscopy, 

the diagnosis of goiter may be missed (especially 

in fine needle aspiration cytological material) in the absence 

of residual micro- or macro-follicles due to haemorrhage and 

endothelial hyperplasia. 

WHAFFT. Another condition associated to vascular proliferation 

in the thyroid gland was described under the acronym 

WHAFFT, which stands for “Worrisome Histologic Alterations 

Following Fine needle aspiration of the Thyroid”. The 

alterations caused by the fine needle aspiration passages 

included haemorrhage,, fibrosis, calcification and worrisome 

lesions, like nuclear atypia, squamous metaplasia, capsular 

pseudoinvasion, and plump endothelial hyperplasia in vascular 

spaces, mimicking vascular tumors. 

Thyroid hemangioma. It is very rare and generally results 

from subsequent organization of intranodular hemorrhagic 

events in goiter, thus suggesting their reactive rather than 

neoplastic nature. 

Angiosarcoma and Sarcomatoid carcinoma. Thyroid angiosarcoma 

(or malignant hemangioendothelioma) was originally 

described in mountain areas and associated to endemic 

goiter. Grossly, a large extensively hemorrhagic mass is 

recognized in the presence of multinodular goiter in the 

surrounding parenchyma. Microscopically, elongated cells 

either lining vascular spaces and protruding into them, or 

arranged in small solid sheets are identified. Eosinophilic cytoplasm, 

polygonal shape, large and hyperchromatic nucleus, 

prominent nucleoli and numerous mitoses are typically present, 

with occasional intracytoplasmic lumina. Tumor cells are 

reactive for FVIII-related antigen, CD31, CD34 and vimentin, 

as well as for cytokeratin (focally). The histogenesis of 

thyroid angiosarcomas is controversial being the hypothesis 

that all such tumors are indeed (angio)sarcomatoid anaplastic 

carcinomas contrasted by the alternative evidence on the 

existence of rare true angiosarcoma cases of the thyroid. 

Whether reactive endothelial hyperplasia in long standing

lectures 

goiter nodules represents an intermediate step in the development 

of malignant vascular growths remains to be defined. 

From a microscopic point of view, the distinction between a 

Breast hamartoma with apocrine glandular 

structure without myoepithelium 

I. Castellano, L. Macrì, G. Canavese, A. Sapino 

Torino 

We describe a case of a 46-years old woman with painless 

mass of the left breast slowly enlarging in the last year. There 

was no family history of breast cancer and the patient was not 

under any pharmacological or hormonal treatments. Clinical 

examination reveals in the upper external quadrant a well-circumscribed, 

mobile, round nodule similar to a fibroadenoma. 

Ultrasound examination demonstrated a heterogeneous lesion 

with lobulated contour and a thin capsule, measuring 5 cm in 

diameter. The patient underwent fine needle aspiration with a 

diagnosis of hypercellular lesion, categorized as C3. Core biopsy 

of the mass, performed in another institution, revealed a 

fibroadenomatous epithelial lesion with usual ductal epithelial 

hyperplasia classified as B3. Quadrantectomy was performed. 

Grossly the mass was bilobated firm and rather circumscribed, 

grey-white on cut surface. Histologically, the nodule showed 

a stromal proliferation of interlobular elongated fibroblasts 

with lobular structures surrounded by sclero-hyaline stroma or 

stroma with a pseudo-angiomatous appearance. Lobular structures 

showed a histological pattern similar to the so called 

“gynecomastia-like hyperplasia” with columnar cells. In 

distinct foci ductal structures disposed in an organoid pattern 

or small cysts were formed by cytologically normal apocrine 

cells. No myoepithelial cells were seen at the periphery of 

these apocrine glands. Staining for p63 confirmed the absence 

of myoepithelial cells. The final diagnosis was of Hamartoma 

of the breast. The peculiar appearance of the apocrine gland 

without myoepithelila cells was described as a possible event 

mimicking pseudoinvasion. 

undifferentiated and poorly differentiated 

sinonasal malignancies 

A. Franchi 

Division of Anatomic Pathology, Department of Critical Care Medicine 

and Surgery, University of Florence Medical School, Florence, 

Italy 

Malignant tumours of the nasal cavities and paranasal sinuses 

represent about 3.6% of all malignancies arising in the head 

and neck area. In this complex anatomic region a significant 

number of neoplasms may present with “undifferentiated” 

Slide seminar: Breast 

Moderators: A. Sapino (Torino), M.P. Foschini (Bologna) 

Head and neck pathologies 

Moderators: M.P. Foschini (Bologna), E. Maiorano (Bari) 

139 

benign (pseudoangiomatous) lesion and a malignant vascular 

tumor is extremely difficult on both surgical and fine needle 

aspiration materials. 

Hamartoma is generically defines as “a malformation that 

resembles a neoplasm, grossly and even microscopically, but 

results from faulty development in an organ; it is composed of 

an abnormal mixture of tissue elements, or an abnormal proportion 

of a single element normally present at that site…”. 

Breast Hamartoma is uncommon, with incidence of 0.7% of 

benign breast tumors. It presents as a painless slow growing 

breast mass, not attached to the underlying structures, in patients 

predominantly in 5 th or 6 th decade of life. The mammographic 

appearance corresponds to a “breast in breast” mass, 

generally without microcacifications. Although the lesion is 

almost always benign, rare case reports describe cancer inside 

hamartoma, so complete excision is the only way to rule out 

malignancy. However, recurrences have been described in 

almost 10% of patients, mainly due to multifocal disease. The 

most interesting feature of the present case was the presence 

of several distinct foci of apocrine glands devoid of myoepithelial 

cells often arranged in an organoid (lobular) pattern. 

Cserni G. (Histopathology 52:239-262) described a similar 

pattern in apocrine glands of a low-grade phyllodes tumour. 

With the exception of microglandular adenosis, lack of myoepithelium 

is generally considered a hallmark of malignancy 

and invasion in breast pathology. However, as discussed by 

Cserni “the absence of myoepithelial cells around apocrine 

glandular structures of the breast does not necessarily imply 

malignancy, and may also be seen in some benign lesions”. 

To rule out malignancy the following criteria were taken into 

account: the presence of an organoid, lobular growth pattern 

which is generally associated with benign changes, lack of 

cellular atypia, monomorphic nuclei and absence of mitotic 

activity in apocrine cells, which are instead typical features 

of apocrine carcinomas. The patient is free of disease at one 

year follow-up. 

light microscopic morphology. In general, they represent a 

group of clinically aggressive neoplasms, although the knowledge 

has progressively evolved towards the need for careful 

differential diagnosis, because some of these entities present 

distinct clinico-pathologic features and biologic behaviour, 

warranting individualized treatment strategies. In addition, 

a number of studies have recently defined the use of novel 

diagnostic markers for sinonasal carcinomas, and there is 

increasing evidence of the importance of the role of immunophenotyping 

and genotyping for differentiating among these 

neoplasms. This presentation will focus on recent acquisitions

140 

concerning the diagnosis of sinonasal undifferentiated and 

poorly differentiated carcinomas, neuroendocrine carcinoma 

and olfactory neuroblastoma. 

Sinonasal poorly differentiated and undifferentiated 

carcinomas 

The group of high grade poorly differentiated and undifferentiated 

sinonasal carcinomas include nasopharyngeal-type 

undifferentiated carcinoma (lymphoepithelioma), sinonasal 

undifferentiated carcinoma (SNUC), NUT midline carcinoma, 

and poorly differentiated keratinizing and non-keratinizing 

variants of squamous cell carcinoma. 

Nasopharyngeal-type undifferentiated carcinoma is typically 

associated with EBV infection, and this is a useful feature 

to separate this entity from other sinonasal undifferentiated 

carcinomas, which are typically EBV negative 1 . 

SNUC is a rare highly aggressive tumour of uncertain histogenesis. 

The term “undifferentiated” has been applied inconsistently 

in the past, but should now be applied more selectively 

with better methods of cell study. By definition SNUC 

does not show any overt squamous or glandular differentiation, 

whereas neuroendocrine features have been frequently 

noted, both histologically and immunohistochemically 2 . 

SNUC can be distinguished from poorly differentiated 

squamous cell carcinoma variants for the different pattern 

of cytokeratins subtypes expression, since SNUC is positive 

for simple epithelia cytokeratins and lacks the expression 

of cytokeratins 5/6 and 13, which instead are expressed by 

squamous cell carcinoma variants 3 . In addition, SNUC has a 

limited expression of p63, which is present in squamous cell 

carcinoma variants 4 . 

NUT midline carcinoma (NMC) is a rare, clinically aggressive 

carcinoma, which is defined by a translocation involving 

the NUT (nuclear protein in testis) gene on chromosome 

15q14 and, in most cases, the BRD4 gene on chromosome 

19p13.1. Initial cases were reported in young patients affected 

by intrathoracic carcinomas, but it is now well established 

that these tumours may occur in adults and involve 

other anatomic sites, including the sinonasal tract 5 . So far 

less than ten cases have been described in the nasal cavity 

and paranasal sinuses. These tumours affected young adults 

of both sexes and showed an aggressive clinical behaviour. 

However, there is certainly an underestimation of their occurrence 

due to the lack of specific diagnostic features. 

Histologically, these carcinomas are composed of undifferentiated 

basaloid cells with focal, often abrupt, squamous 

differentiation. Therefore, the diagnosis of NMC requires 

the demonstration of the NUT translocation, which can be 

achieved by karyotyping, reverse transcription polymerase 

chain reaction (RT-PCR), and FISH. Recently, a monoclonal 

antibody to NUT has been developed, which showed a sensitivity 

of 87%, a specificity of 100%, a negative predictive 

value of 99%, and a positive predictive value of 100% when 

tested in a large panel of carcinoma tissues 6 . Moreover, the 

expression of normal NUT protein is limited to the germ cells 

of the testis and ovary, thus increasing the reliability of the 

use of immunohistochemistry in the diagnosis of NMC. The 

use of this antibody may help to separate NMC from other 

poorly differentiated sinonasal carcinomas, thus contributing 

to their clinico-pathologic characterisation. In addition, 

it appears that the distinction of NMC from other sinonasal 

carcinomas is of clinical relevance, in view of the favourable 

response to certain treatment regimes, including chemotherapy 

according to Ewing’s sarcoma protocols 7 or docetaxel 

and radiotherapy 8 . 


Small cell carcinoma, neuroendocrine type (SCC- 

NET) 

Currently, WHO classification of head & neck tumours, 

places SCCNET in the category of neuroendocrine tumours 

together with carcinoid tumour, which can be further sub-classified 

into typical and atypical 9 . SCCNET of the nasal cavities 

and paranasal sinuses is a very uncommon neoplasm of which 

only small series and isolate case reports have been reported 

in the English literature 10 . A critical review of these reports 

reveals that in some cases the clinico-pathological features 

of the lesions described were more consistent with other diagnoses, 

including olfactory neuroblastoma and SNUC. This 

underlines the current lack of criteria, including a definition 

of a panel of immunohistochemical markers, to make the diagnosis 

of SCCNET. Small cell neuroendocrine carcinoma of 

the sinonasal tract is histologically indistinguishable from its 

pulmonary counterpart. Immunohistochemically, it is positive 

for cytokeratins and neuroendocrine markers such as NSE 

(neuron specific enolase), synaptophysin, and chromogranin, 

although with variable intensity 10 . As small cell neuroendocrine 

carcinomas of other sites, sinonasal tumours express 

CD57 11 . These features allow the distinction from SNUC, 

malignant melanoma, olfactory neuroblastoma, lymphoma, 

Ewing’s sarcoma/PNET and rhabdomyosarcoma. 

Olfactory Neuroblastoma (ON) 

ON is a rare neoplasm occurring in a broad age range, which 

most commonly originates in the region of the cribriform plate 

from the olfactory mucosa 12 . More frequently, the tumour 

grows in nests separated by fibrovascular septa, or sometimes 

it may show a diffuse growth pattern. The neoplastic cells 

typically have small and round nuclei with stippled chromatin, 

absent or small nucleoli, and scanty cytoplasm. They are 

embedded in a fibrillary background formed by cell processes. 

Homer-Wright type of rosettes, or more rarely Flexner rosettes 

can be found. Immunohistochemically, ON shows diffuse 

positivity for NSE and synaptophysin, while chromogranin, 

GFAP and leu-7 are less often positive. S-100 protein stains 

sustentacular cells around neoplastic nests, but in less differentiated 

tumours there may be few scattered S-100 protein 

positive cells. Neurofilament protein and other markers of neural 

differentiation are more often expressed in tumours with 

diffuse, sheet-like pattern. Cytokeratins are generally negative, 

although in ON with nesting pattern a few tumours cells may 

exhibit staining for low molecular weight cytokeratins. A subgroup 

of ON with gland-like formations and more widespread 

cytokeratin positivity has been designated “olfactory neuroepithelioma” 

13 . EMA is consistently negative, as they are CD99, 

CD45, HMB-45 and muscle markers. Ultrastructural analysis 

shows evidence of neuroblastic differentiation, including the 

presence of dendritic processes containing dense core granules 

and neurotubules, and occasional synaptic junctions. ON lacks 

the t(11; 22) translocation of Ewing’s sarcoma/PNET. 

references 

1 Cerilli LA, Holst VA, Brandwein MS, et al. Sinonasal undifferentiated 

carcinoma: immunohistochemical profile and lack of EBV association. 

Am J Surg Pathol 2001;25:156-63. 

2 Mills SE. Neuroectodermal neoplasms of the head and neck with emphasis 

on neuroendocrine carcinomas. Mod Pathol 2002;15:264-78. 

3 Franchi A, Moroni M, Massi D, et al. Sinonasal undifferentiated carcinoma, 

nasopharyngeal-type undifferentiated carcinoma, and keratinizing 

and nonkeratinizing squamous cell carcinoma express different 

cytokeratin patterns. Am J Surg Pathol 2002;26:1597-604. 

4 Bourne TD, Bellizzi AM, Stelow EB, et al. p63 expression in olfactory 

neuroblastoma and other small cell tumors of the sinonasal tract. Am 

J Clin Pathol 2008;130:213-8.

lectures 

5 French CA, Kutok JL, Faquin WC, et al. Midline carcinoma of 

children and young adults with NUT rearrangement. J Clin Oncol 

2004;22:4135-9. 

6 Haack H, Johnson LA, Fry CJ, et al. Diagnosis of NUT midline carcinoma 

using a NUT-specific monoclonal antibody. Am J Surg Pathol 

2009;33:984-91 

7 Mertens F, Wiebe T, Adlercreutz C, Mandahl N, French CA. Successful 

treatment of a child with t(15;19)-positive tumor. Pediatr Blood 

Cancer. 2007;49:1015-7. 

8 Engleson J, Soller M, Panagopoulos I, et al. Midline carcinoma with 

t(15;19) and BRD4-NUT fusion oncogene in a 30-year-old female with 

response to docetaxel and radiotherapy. BMC Cancer 2006;6:69. 

9 Perez-Ordonez B. Neuroendocrine tumours. In: Barnes L, Eveson JW, 

Reichart P, Sidransky D (eds.). WHO Classification of Tumours. Pathology 

and Genetics of Head and Neck Tumours. Lyon: IARC Press 

2005, pp. 26-7. 

10 Perez-Ordonez B, Caruana SM, Huvos AG, et al. Small cell neuroendocrine 

carcinoma of the nasal cavity and paranasal sinuses. Hum 

Pathol 1998;29:826-32. 

11 Morice WG, Ferreiro JA. Distinction of basaloid squamous cell carcinoma 

from adenoid cystic and small cell undifferentiated carcinoma 

by immunohistochemistry. Hum Pathol 1998;29:609-12. 

12 Dulguerov P, Allal AS, Calcaterra TC. Esthesioneuroblastoma: a meta 

analysis and review. Lancet Oncol 2001;2:683-90. 

13 Sugita Y, Kusano K, Tokunaga O, et al. Olfactory neuroepithelioma: 

an immunohistochemical and ultrastructural study. Neuropathol 

2006;26:400-8. 

Minor salivary gland tumors 

G. De Rosa 

Department of Biomorphological and Functional Sciences, Pathology 

Section, University of Naples Federico II, School of Medicine, 

Naples, Italy 

Background. Minor salivary gland tumors (MSGT) are uncommon, 

representing approximately 10-15% of all salivary 

neoplasms and 3-5% of all head and neck tumors. Despite this 

relatively low frequency, these neoplasms show a high variety 

in clinical behavior and morphology, constituting a heteroge- 

141 

neous group with a broad range of histological types. All the 

histotypes of WHO classification of salivary gland tumors 

may arise in minor salivary gland, but some tumors, as canalicular 

adenoma, polymorphous low grade adenocarcinoma, 

and sialadenoma papilliferum are exclusive or predominant 

in these sites. Unlike parotid and submandibular tumors, most 

MSGT are malignant, with different rates among the various 

anatomical locations. 

Results. In our Department we identified 91 cases of 

MSGT from 1993 to 2009; in agreement with the data 

of the literature, the most frequent benign type was the 

pleomophic adenoma, while among the malignant tumors, 

polymorphous low grade adenocarcinoma was the most 

common, followed by adenoid-cystic and mucoepidermoid 

carcinoma. 

The diagnosis of MSGT may be difficult, because many histotypes 

show overlapping morphological features; cribriform 

areas, clear cells, bilayered and papillary pattern are present in 

many different benign and low grade neoplasms complicating 

the differential diagnosis. In this setting, the search of stromal 

and perineural invasion is one of the most important feature 

of malignancy; however, near always the specimen are small 

incisional biopsy and is not possible to assessing the tumor 

borders and the tumor interface with adjacent tissues, and then 

differentiate between benign tumor and malignancies. In these 

cases, a definitive diagnosis should be deferred to complete 

excision or a larger-size biopsy. 

At the present, the role of immunohistochemistry in diagnosis 

of MSGT is limitated; in fact the immunohistochemical 

staining may demonstrate the coexistence of glandular and 

myoepithelial components, and the presence of dual luminalabluminal 

cell differentiation, but most benign and low grade 

tumors exhibit these same features. On the contrary, Ki67 

proliferative index may be useful in distinguishing a benign 

lesion from a malignant tumors, being 10% the reliable cut-off 

value for malignancy. 

Gynaecological pathologies in lynch syndrome 

Clinicopathologic features of gynecologic 

tumors in lynch syndrome 

M.L. Carcangiu 

Dipartimento di Patologia e Laboratorio, Fondazione IRCSS Istituto 

Nazionale Tumori, Milano, Italia 

Background. Women with hereditary non-polyposis colorectal 

cancer (HNPCC)/Lynch Syndrome have a high risk for gynaecological 

cancer and in particular for endometrial cancer (EC). 

Methods. The pertinent literature on the field from 2000 to 

present was retrieved trough a med-line search and critically 

reviewed. 

Results. Lynch syndrome (LS), also known as hereditary polyposis 

colorectal cancer syndrome (HNPCC), is an autosomal 

dominant inherited cancer susceptibility syndrome characterized 

by a high risk of colorectal, endometrial, and other 

tumors, including ovarian, gastric, urinary, and biliary tract 

cancer. The genetic basis of this syndrome is a mutation in one 

of the known DNA mismatch-repair genes: MLH1, MSH2, 

and MSH6. Women with LS have a 20%-60% lifetime risk of 

Moderators: M.L. Carcangiu (Milano), C. Riva (Varese) 

endometrial cancer and approximately 50% of them present 

first with endometrial cancer. 

Clinical-pathologic data on LS-related EC are scanty. Most 

authors have stated that they occur in a younger age group 

and that most of them show low grade endometrioid histology. 

A different picture emerges from the series of Broaddus 

et al., in which the non-endometrioid types made up 14% of 

the LS-related EC, as opposed to 2.4% in the control cases; no 

details were given on the histologic subtypes comprising the 

non-endometrioid group. In our series of endometrial carcinomas 

from 23 patients (mean age 46.2 years) with MSH2 (16), 

MLH1 (6) and MLH1/MSH2 (1) constitutional mutations, 

there were 13 (56.5%) endometrioid endometrial carcinomas 

(EECA) and 10 (43.4%) non-endometrioid endometrial 

carcinomas (N-EECA) with a predominance of the clear cell 

type frequently combined with an endometrioid carcinoma 

as opposed to the control group made by 66 sporadic tumors 

in same age patients where 44 (95.6%) were of endometrioid 

type and 2 (4.3%) non-endometrioid (OR 0.59, 0.013-0.27). 

Furthermore the endometrioid cancers in women with a germ-

142 

line LS mutation had a higher FIGO grade and more frequent 

vascular invasion than their sporadic counterparts. In our series 

we also identified 2 cases of Malignant Mixed Mullerian 

Tumor in 2 MSH2-mutated sisters, a finding that has also been 

recently reported in a MLH1-mutated patient 

Regarding the FIGO grade, 46.1% of the LS women with 

EECA in our study had a grade III tumor, a frequency appreciably 

higher than that seen in the control group. 

As far as the FIGO Stage is concerned, Boks et al. and Broaddus 

et al. had commented on the relatively high number of 

Stage III tumors they found in their series of LS-related EC, 

especially in view of the relatively young age of the patients; 

regrettably, no information was provided on the stage distribution 

according to histologic type. In our group of patients, 

the tumor stage distribution at presentation of both EECA and 

N-EECA paralleled those observed in their sporadic counterparts, 

with the majority of EECA presenting at stage I and 

most N-EECA (with or without an endometrioid component) 

presenting at higher stages. 

The overall survival of our patients at the end of the followup 

was lower than that reported in previous studies and was 

strongly influenced by histologic type and tumor stage, as 

shown by the fact that 4 of the 5 patients who died of EC had 

a N-EECA. By contrast, the EECA were characterized by 

a uniformly favourable outcome, with only a tumor-related 

death in a 67-year-old woman who had a grade III, stage IIB 

tumor. 

LS associated ovarian carcinomas account for 10-15% of 

hereditary ovarian carcinomas. Histologically, they tend to be 

more frequently of endometrioid and clear cell types. Findings 

from a study by Crijnen et al. on 26 patients with LS associated 

ovarian cancer showed that the survival rate for ovarian 

cancer was not significantly different between these patients 

and controls with sporadic ovarian cancer. 

references 

Boks DE, Trujillo AP, Voogd AC, et al. Survival analysis of endometrial 

carcinoma associated with hereditary nonpolyposis colorectal cancer. 

Int J Cancer 2002;102:198-200. 

Broaddus RR, Lynch HT, Chen LM, et al. Pathologic features of endometrial 

carcinoma associated with HNPCC: a comparison with sporadic 

endometrial carcinoma. Cancer 2006;106:87-94. 

Carcangiu ML, Radice P, Casalini P, et al. Lynch syndrome--related 

endometrial carcinomas show a high frequency of non-endometrioid 

types and of high FIGO grade endometrioid types. Int J Surg Pathol 

2010;18:21-6. 

Crijnen ThEM, Janssen-Heijnen MLG, Gelderblom H, et al. Survival of 

patients with ovarian cancer due to a mismatch repair defect. Fam 

Cancer 2005;4:301-305. 

Lynch HT, Casey MJ, Snyder CL, et al. Hereditary ovarian carcinoma: 

heterogeneity, molecular genetics, pathology, and management. Mol 

Oncol 2009;3:97-137. 

Molecular and immunohistochemical features 

of endometrial carcinoma in HNPCC/lS 

C. Riva 

Dept of Human Morphology, University of Insubria, Varese 

Endometrial carcinoma (EC) is the most common extracolonic 

neoplasia in HNPCC/Lynch Syndrome (LS) patients. 

Men with LS have a 74% lifetime risk of colorectal cancer 

(CRC), in women the risk is over 30%, but more than 40% to 

50% will develop EC. 

LS is characterized by germline mutations of DNA MMR 

genes. The MMR system repairs DNA replications errors 

that are not immediately corrected by DNA polymerase and, 

therefore, it plays a crucial role in DNA replication accuracy. 


Human (h) MMR genes and corresponding proteins include 

hMSH2, hMSH6, hMLH1, and hPMS2. 

Functional inactivation of MMR genes by mutations or epigenetic 

changes leads to the accumulation of insertions/deletions. 

These are easily identified in short DNA tandem repeat 

sequences (microsatellites) and this phenotype is known 

as microsatellites instability (MSI). MSI can be present in 

tumors from LS patients but it is also observed in a fraction 

(15-25%) of various sporadic neoplasms, including EC. 

Among germline mutations are recognized 1) truncating mutations 

leading either to a loss or to a easily degradable protein 

and allowing to a negative immunohistochemical (IHC) staining 

of MMR protein and 2) missense point mutations leading 

to an amino acid substitution, affecting chemical stability or 

functionality; in point mutations often MMR protein is still 

immunoreactive and atypical clinical scenarios are observed. 

Analysis for germline mutations in MMR genes is confirmatory 

for LS diagnosis. Mutational analysis is very expensive 

and time consuming, therefore pre-selection of high risk 

patients for mutation search is very important. This purpose 

can be achieved by employing MSI tumor DNA analysis (by 

PCR), IHC for 4 MMR proteins (hMLH1, hMSH2, hMSH6 

and PMS2) and methylation assays. 

An epigenetic (sporadic) cause of MSI is more common than 

LS. Most of sporadic MSI CRCs and ECs arise via hMLH1 

promoter methylation, therefore methylation analysis seems 

useful to distinguish between sporadic and heritable cases. 

MSI analysis is performed using the NCI panel of 5 PCR 

microsatellite markers (BAT-25, BAT-26, D2S123, D5S346 

and D17S250). Tumors are classified as MSI-H (two or more 

markers show MSI), MSI-L (one marker shows MSI) and 

MSS (none marker shows MSI). Not all LS associated ECs are 

MSI-H, while most MSI-H ECs are sporadic tumors, therefore 

MSI analysis may fail to detect a number of ECs arising in a 

hereditary setting. 

DNA-MMR protein IHC is an effective method to detect MSI 

and it is useful as a screening of LS. Infact MLH1, MSH2, 

MSH6 and PMS2 are lacking in tumor cell nuclei by IHC in 

up 1/3 of ECs; this derives in most cases from MLH1 promoter 

ipermethylation, while mutations accounts for the rest. IHC 

interpretation can be problematic: in general only a complete 

loss of expression in the setting af a positive internal control 

(endometrial stroma, normal glands, lymphocytes, etc) seems 

to be reliable. In rare cases with inconclusive IHC results and a 

clinical LS suspicion, an alternative test should be performed. 

IHC with MLH1, MSH2 and MSH6 antibodies shows an high 

sensitivity (91%) and a specificity of only 83%, due to lack of 

correlation between loss of MSH6 and MSI-H. In fact LS-associated 

EC is fivefold more frequently associated with MSH6 

mutations compared to CRC and these mutations usually do 

not leave to MSI-H. The positive predictive value of IHC for 

detect a germline mutation, especially with absence of MSH2 

and MSH6 is very high.Therefore it has been suggested that 

IHC loss of these proteins may be a sufficient evidence of LS. 

On the other hand, IHC can be easy performed in the majority 

of pathology laboratories, is a convenient test and, in addition, 

it can address specific genes sequencing. 

Since LS detection methods cannot be applied routinely to every 

EC, various screening algorithms have been proposed. In 

particular some data suggest the application of IHC for DNA 

MMR proteins in ECs patients with strong personal or family 

history, age of onset less than 50 years, lower uterine segment 

localisation, a synchronous ovarian clear cell carcinoma and, 

finally, morphological features characterized by peritumoral 

or tumor infiltrating lymphocytes and tumor heterogeneity 

including dedifferentiated areas.

lectures 

The EC patients with abnormal IHC results are referred for a 

genetic evaluation including MSI analysis, if indicated a methylation 

test and mutational analysis of the screened genes. 

references 

Garg K, Leitao M, Kauff N, et al. Selection of endometrial carcinomas 

for DNA mismatch repair protein immunohistochemistry using patient 

age ansd tumor morphology enhances detection of mismatch repair 

abnormalities. Am J Surg Pathol 2009;33:925-33. 

Garg K, Soslow RA. Lynch Syndrome (hereditary non-polyposis colorectal 

cancer) and endometrial carcinoma. J Clin Pathol 2009;62 679- 

84. 

Karamurzin Y, Rutgers KL. DNA Mismatch Repair Deficiency in endometrial 

carcinoma. Int J Gynecol Pathol 2009;28,3;239-52. 

Resnick K, Straughn JM, Backes F, et al. Lynch Syndrome screening 

strategies among newly diagnosed endometrial cancer patients. Obstet 

Gynecol 2009;114:530-6. 

lynch syndrome and new model of 

individualized gynaecological cancer prevention 

M.G. Tibiletti 

U.O. Anatomia Patologica Ospedale di Circolo, Università 

dell’Insubria Varese 

Lynch syndrome (LS), or hereditary non-polyposis colorectal 

cancer (HNPCC), is an autosomal dominant syndrome (MIM) 

that predisposes its carriers to multiple malignancies including 

colorectal cancer (CRC), endometrial cancer (EC), ovarian 

cancer (OC), and cancer of the renal pelvis and ureter, stomach, 

pancreas, small bowel and brain. 

Traditionally Lynch syndrome has been perceived as a CRC 

dominated syndrome. However, in women with Lynch syndrome, 

the incidence of EC equals or exceeds that of CRC, 

and in more than 50% of cases, these women present a gynaecological 

cancer as their first malignancy. 

LS is caused by a germline mutation in one of the DNA 

mismatch repair genes: MLH1, MSH2, MSH6, and PMS2. 

Deficient mismatch repair protein activity leads to DNA microsatellite 

instability (MSI) and absent immunoistochemical 

protein expression in tumour tissue. This pattern of abnormal 

staining provides guidance as to which of the MMR genes is 

likely to harbour a germline mutation. 

143 

The initial (1991) and revised (1998) Amsterdam criteria 

were developed to identify families at high risk for LS. 

These criteria required colorectal or other LS-associated 

cancers in three first- degree relatives, occurring in at least 

two successive generations, and in one individual under the 

age of 50 years. These criteria were recognized to have poor 

sensitivity in identifying individuals carrying an LS gene 

mutation. Therefore, the Bethesda guidelines were introduced 

to broaden testing recommendations and to identify a greater 

proportion of affected individuals. The Bethesda guidelines 

recommended molecular testing for LS in different groups 

of patients including two gynaecologic cancer populations: 

patients with endometrial cancer diagnosed before 45 years of 

age and those with two LS-related cancers. 

In 2006 an European group of experts in LS established guidelines 

for the clinical management of LS (Mallorca guidelines 

J Med Genet 2007) in order to improve the identification and 

the care of these families. 

Identification of LS in affected individuals has important 

implications for screening in individuals as well as family 

members, as close screening and surveillance has been shown 

to reduce the mortality of colorectal cancer by over 60%. 

The frequency of germline DNA mismatch repair gene mutations 

among unselected patients with EC has been found to 

be 1.8% to 2.1% which is similar to the frequency of LS in 

colorectal carcinoma. In patients younger than 50 years, the 

incidence is increased up to 9%. The identification of these 

patients is important for several reasons. Affected patients are 

at risk for multiple synchronous and metachronous tumors. 

These individuals would therefore benefit from surveillance 

measures to detect other LS associated tumours; their family 

members may benefit from genetic testing to determine carrier 

status and to have adequate surveillance measures. 

references 

Meyer LA, et al. Endometrial cancer and Lynch Syndrome: Clinical and 

pathological considerations. Cancer Control 2009;16:14-22. 

Vasen H, et al. Guidelines for the clinical management of Lynch syndrome 

(HNPCC). J Med Genet 2007;44;353-62. 

Walsh CS, et al. Lynch syndrome among gynecologic oncology patients 

meeting Bethesda guidelines for screening. Gynecon Oncol 

2010;116:516-21. 

Pathologica symposium: new frontiers in immunohistochemistry 

Immunoprofile of renal tumors 

D. Segala, M. Brunelli, G. Martignoni 

Department of diagnostic pathology, University of Verona, Verona, 

Italy 

The WHO 2004 classification of renal cell neoplasms includes 

numerous entities characterized by different prognosis and the 

correct subtyping is an important procedures to predict the 

behavior of these tumors. 

Among major renal histotypes, oncocytoma has the best prognosis 

followed by chromophobe, papillary, clear cell and collecting 

duct renal cell carcinomas (RCCs) 1-4 . The overlapping 

morphological features and the increasing description of novel 

potential entities determine the difficulties of some histologic 

distinctions. That is why traditional histology needs today the 

support of more specific markers that should be consistently 

detected also on small biopsies. In fact, new minimal invasive 

forms of therapies, such as criotherapy and diathermocoagulation 

will require a pre-operative diagnosis 5 6 . 

Moreover, reliable predictive factors are essential for the 

stratification of patients into clinically meaningful categories. 

Staging has recently improved with the development of 

integrated systems 7 8 , however the information obtained by 

molecular tumor markers are expected to revolutionize the 

staging of RCC. 

Immunohistochemistry is the most easily available and not 

expensive ancillary technique used by pathologists, therefore 

it is the most important field to be improved. 

Diagnostic immunohistochemical markers 

Clear cell renal cell carcinoma. Clear cell RCC is immmunohistochemically 

characterized by a high positive rate for 

CD10 (82%) 9-11 . In our experience also CD13 is a good immunohistochemical 

marker of clear cell renal cell carcinoma 

being positive in 81% 12 . Most clear cell RCCs typically show

144 

a restricted expression pattern of cytokeratins (CK) with limited 

cases expressing CK7, CK8, CK19, high weight CKs and 

a large portion of cases positive for CK18 13 14 . Parvalbumin 

was found to be constantly absent 10 . Alpha-methylacyl-CoA 

racemase (AMACR) positivity has been detected in 25% 

whereas S100A1 immmunostaining has been observed in 75% 

of the cases 15 16 . Around more than a half of the clear cell 

RCCs reveals immunoreactivities for vimentin, RCC Marker 

and Epithelial Membrane Antigen (EMA) 9 14 17 . 

Papillary renal cell carcinoma. Papillary RCCs typically 

express CK7 (87%), 8, 18 and 19, Vimentin (90%) and they 

constantly show AMACR immunostain 14 15 18 . CK7 expression 

is more frequently observed in type 1 (87-100%) than 

type 2 (20-50%) 18 such as EMA (type 1 ranging from 72 to 

100% and type 2 from 13 to 17%) 17 19 whereas E-cadherin 

is reported predominantly in type 2 17 . An high incidence 

of positivity for CD10 (59-90%), BerEP4, EMA and RCC 

Marker is reported 10 20 . S100A1 is reported in 92% of papillary 

RCCs 16 which occasionally express high molecular 

weight CKs (26%) 11 . 

Chromophobe renal cell carcinoma. Chromophobe RCCs 

is strongly positive for parvalbumin in all primary and 

metastatic tumors 21-23 . Chromophobe RCCs are also positive 

for CK7 in 73-100% of the samples 13 14 . CD10 expression 

has been found in 26% of chromophobe RCCs, five of 

the seven (71%) showing aggressive features 10 . CKs 8, 18, 

EMA and E-cadherin are frequently positive whereas chromophobe 

RCCs do not usually express vimentin 13 14 17 24 . 

Immunohistochemical membrane expression of c-KIT 

is frequently found in chromophobe RCC however c-kit 

mutation has not been found 25 . AMACR is usually not 

expressed and only 4% of chromophobe RCCs are positive 

for S100A1 15 16 . 

Collecting duct carcinoma. The immunohistochemical profile 

of these carcinomas shows high molecular weight CKs, 

EMA, vimentin, lectin Ulex europaeus agglutinin and peanut 

lectin agglutinin (Arachis hypogaea) immunostain 26 . 

Oncocytoma 

Most of oncocytomas are immunoreactive for CKs (86%), 

EMA (86%), E-cadherin (71%), parvalbumin (70%) and c- 

KIT (100%) 13 14 17 21 27 28 . Vimentin and RCC marker are usually 

not expressed 14 . Althought contrasting results have been 

reported for CK7 in renal oncocytoma, it actually seems that 

only a focal immunoreactivity of a few cells can be found 29 30 . 

S100A1 is expressed in 92% of this neoplasm 16 . 

Renal mucinous tubular and spindle cell carcinoma. This 

histotype immunostains for CKs (CK5/6, 7, 8, 13, 14, 17, 18, 

19, 20), high molecular weight CKs 1, 5, 10, 14, E-cadherin 

and vimentin, but CD10 and RCC marker are usually not 

expressed 31-33 . Immunoreactivity for AMACR (93%), CK7 

(81%) and EMA (95%) have also been reported 33 . 

TFE-family translocation renal cell carcinomas. TFE-family 

translocation renal cell carcinomas bear specific translocations 

that results in overexpression of TFE3 or TFEB, 

genes that are strictly related to microphtalmia transctiption 

factor (MiTF). Different translocations involving chromosome 

Xp11.2 bring TFE3 fusion gene product overexpression, 

whereas TFEB overexpression is the result of the specific 

translocation t(6;11)(p21;q12). 

Immunohistochemistry for TFE3 and TFEB is the most reliable 

test able to distinguish TFE-family translocation renal 

cell carcinomas from formalin-fixed and paraffin-embedded 

archive tissue, but sometimes troubles using these antibodies 

have been reported. These tumors were also consistently im- 


munoreactive for the RCC antigen and CD10 and negative or 

focally positive for citokeratins 34-37 . 

Our group have recently described the immunohistochemical 

expression of Cathepsin-K, a protein described in osteoclasts 

to be modulated by the expression of MiTF, in 17 cytogenetically 

demonstrated TFE3 and TFEB renal cell carcinomas and 

in a large group of renal tumors 38 . Cathepsin-K was positive 

in all TFEB renal cell carcinoma and in 60% of TFE3 renal 

cell carcinomas, whereas all other renal tumors were negative. 

Therefore cathepsin-K could be a useful marker alternative to 

TFE3 and TFEB. 

End-stage renal disease associated tumors. Tumors arising 

in kidneys with end-stage renal disease include those 

resembling sporadic renal tumors such and tumors distinct 

from them that Tickoo at al named “acquired cystic diseaseassociated 

renal cell carcinoma” and “clear cell papillary renal 

cell carcinoma of the end-stage renal kidneys” 39 . This last 

neoplasms seem to display distinctive histologic features not 

easily referable to the histotypes described in the WHO 2004 

classification system. 

Clear-cell papillary renal cell carcinoma of the end-stage 

kidney, unlike papillary RCC, were costantly negative for 

AMACR, but unlike clear-cell RCC all tumors tested showed 

strong immunoexpression for CK7 39 . Gobbo et al. found similar 

tumors in normal kidneys 40 . They also observed the lack of 

immunoexpression of CD10. 

Tumors with a strict related immunohistochemical pattern and 

similar morphological features have also been recently described 

and called RCC with prominent angioleiomyomatous 

proliferation 41 . This tumors are characterized by a various 

grade of stromal proliferation beside the epithelial structures. 

To date the correlation between these two entities is not already 

demonstrated. 

Prognostic molecular markers 

Nomograms assigning numerical scores to various clinical 

and pathological prognostic indicators, excluding molecular 

markers, has been proposed, however a wide variety of molecular 

markers have been examined and some seem promising 

to legitimize further research to prove their value as 

prognostic tools. 

Among tumour suppressor genes p53 overexpression has been 

described as a significant molecular predictor of tumor recurrence, 

especially in clear cell RCC and the loss of p27/kip1 

expression is described as a possible prognostic and diagnostic 

marker of tumor development and/or progression 42-44 . 

Ki-67 proliferation index has been shown to be a prognostic 

factor in both univariate and multivariate analysis, although 

conflicting evidence has challenged these findings 45 46 . 

COX-2 expression in patients with renal cell carcinoma is 

associated with several clinicopathological factors, and appeared 

to play an important role in tumor cell proliferation, 

but is not a significant prognostic factor 47 48 . 

The adipose differentiation-related protein (ADFP) is a lipid 

storage droplet-associated protein and its transcription is considered 

to be regulated by the von Hippel-Lindau/hypoxia-inducible 

factor pathway. ADFP expression status may provide 

useful prognostic information as a biomolecular marker in 

patients with clear cell RCC 49 . 

Decreased carbonic anhydrase IX (CAIX) levels are independently 

associated with poor survival in advanced RCC. CAIX 

reflects significant changes in tumor biology, which should 

be used to predict clinical outcome and identify high-risk patients 

in need for adjuvant immunotherapy and CAIX-targeted 

therapies 50 .

lectures 

Epithelial growth factor receptor (EGF-R) positivity is more 

common in clear cell (81%) than in papillary tumours (40%). 

Membranous location of EGF-R immunostaining is associated 

with good prognosis in renal cell carcinoma 51 52 . 

Vascular endothelial growth factor (VEGF) protein expression 

is a significant independent predictor of outcome and suggests 

that VEGF is involved in angiogenesis in clear RCCs 53 . 

Patients with EpCam expressing clear cell RCC showed a 

trend toward a better prognosis in a Cox regression analysis 

including stage, grade, and necrosis 54 . 

Conclusions 

Among the large number of molecular markers proposed 

in recent years, CD10, parvalbumin, AMACR, CK7 and 

S100A1 seem the more promising immunostains for an accurate 

diagnostic panel. Immunohistochemical prognostic 

markers useful in the daily routine work are lacking to date 

and TNM staging, grading sec. Fuhrman and necrosis appear 

as prognostic information to include in the pathological 

report. 

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Molecular diagnosis of solid tumours. 

A practical approach for organ pathologies 

Molecular diagnosis in solid tumor: the breast 

A. Sapino, C. Marchiò 

Dipartimento di Scienze Biomediche e Oncologia Umana. Torino. 

Italy 

Molecular techniques are, nowadays, in common use in pathology 

laboratories, especially in the field of cancer diagnosis. 

In breast pathology, molecular testing continues to expand 

as requests by the oncologists of more precise prediction on 

response to treatment and risk of recurrence increase. 

In situ hybridization techniques, such ad FISH/CISH, SISH 

to test HER2 gene status, are the basic and most widely used 

molecular tests applied to breast cancer diagnosis. However, 

following the results of the first studies of microarrays used 

as prognostic/ predictive tools, countless prognostic and/or 

predictive signatures have been developed. Two of these 

signatures, the MammaPrint ® (Agendia BV, Amsterdam, 

Netherlands) and the Oncotype DX ® (Genomic Health Inc., 

Redwood City, CA, USA) have achieved the FDA approval. 

In Italy, both of them are commercially available only for 

patients’ use. In particular, the first assay is based upon a 

multi-gene prognostic predictive score comprising 70 genes 

and works on mRNA extracted form fresh cancer tissues. 

This signature segregates patients in two categories: one of 

good prognosis (“low-risk” group), and one of poor prognosis 

(“high-risk” group). The Oncotype DX ® is an RT-PCR based 

test that is based on the mRNA expression levels of only 21 

genes (16 cancer related genes and 5 reference genes) and is 

presented as single Recurrence Score, which is a continuous 

variable ranging between 0 and 100 divided into three risk 

Moderators: G. Tallini (Bologna), G. Stanta (Trieste) 

groups: low (< 18), intermediate (18-31) and high (RS ≥31), 

for clinical decision-making. The main goal of both signatures 

is to safely spare patients at “low molecular risk” with border 

line biological risk from chemotherapy. However extensive 

validation of MammaPrint ® and of Oncotype DX ® represents 

the main challenge in integrating them in the standard of 

breast patients care. Combining molecular assay results with 

the pathological and clinical features will pave the way to a 

new era in breast oncology. 

Molecular diagnosis of lung cancer 

A. Marchetti 

Sezione di Diagnostica Molecolare, Dipartimento di Oncologia e 

Medicina Sperimentale, Università “G. D’Annunzio”, Chieti, Italia 

Lung cancer is the most frequent cause of cancer-related 

morbidity and mortality in industrialised countries, and about 

80% of primary lung cancers are non-small cell lung carcinomas 

(NSCLCs). The two most common subtypes of NSCLC, 

squamous cell carcinoma (SCC) and adenocarcinoma (AC) 

derive from different compartments in the lung. The main 

molecular pathways involved in the pathogenesis of NSCLC 

include: a) growth promoting pathways (EGFR, KRAS PI3K, 

ALK), b) growth inhibitory pathways (p53, Rb, P14ARF, 

STK11), c) apoptotic pathways (Bcl-2, Bax, Fas/FasL), d) 

pathways involved in DNA repair and immortalisation processes. 

A number of epigenetic changes, including DNA 

methylation, histone/chromatin protein modification, and 

micro-RNA expression can also contribute to tumour develop-

lectures 

ment. Cumulative information suggests that the SCC and AC 

subtypes progress through different carcinogenic pathways, 

but the genetic aberrations promoting such differences are 

poorly understood. 

The recent advent of targeted therapies in the management of 

NSCLC patients have greatly enhanced the interest for predictive 

molecular markers that could allow to select patients 

maximising efficacy and avoiding toxic effects of treatments. 

The identification of predictive biomarkers that can guide 

treatment decisions is an important step for individualized 

therapy and in ultimately improving patient outcomes. 

Monoclonal antibodies and small-molecule tyrosine kinase 

inhibitors (TKIs) targeting the Epidermal Growth Factor 

Receptor (EGFR) and the Vascular Endothelial Growth Factor 

(VEGF) have recently emerged as effective agents for 

the treatment of patients with advanced NSCLC. In addition, 

several novel agents have been developed which may 

overcome acquired resistance to these treatments or target 

other deregulated cell pathways. Potential biomarkers for 

the selection of patients with NSCLC most likely to benefit 

from tyrosine kinase inhibitors include mutations, gene copy 

number increase and single-nucleotide polymorphisms of 

the EGFR gene, EGFR protein expression and oncogenic 

mutation on the KRAS gene. Additional biomarkers that may 

predict response to other recently developed targeted therapies 

are under investigation. 

A number of different techniques including fluorescence in 

situ hybridization (FISH), PCR amplification followed by 

sequencing or other mutation detection assays, and reversetranscription 

PCR, have been used to rapidly and efficiently 

characterize these biomarkers. The current weight of evidence 

for using these methods to analyse biomarkers for personalized 

therapy for a rapid characterization of NSCLCs to be 

treated with targeted agents will be presented. 

Integration of molecular diagnostics into thyroid 

cytological practice 

G. Troncone 

Dipartimento di Scienze Biomorfologiche e Funzionali, Università di 

Napoli “Federico II” 

Background. Although thyroid Fine-needle aspiration (FNA) 

is much more accurate than the clinical, biochemical or radiological 

assessments, the method is highly dependent on 

the operator experience 1 . Conventional wisdom dictates that 

FNA is more efficient when an experienced cytopathologist 

ensures the proper smearing technique and the rapid interpretation 

of air-dried Diff-Quick-stained smears 2 . Molecular 

testing of thyroid nodules for a panel of mutations refines the 

cytological diagnosis of a thyroid cell malignancy 3 . In particular 

the V600E BRAF mutation, highly specific for papillary 

carcinoma, is also emerging as an independent marker of 

clinical aggressiveness 4 5 . Preoperative knowledge of BRAF 

mutation may be helpful to tailor the surgical treatment for 

any individual patient 5 . However, the full application of this 

test from dedicated research labs to cytopathology outpatient 

settings has not completely been accomplished 6 . A number 

of pratical issues have not been investigated, as most of the 

studies were retrospective. To widespread the use of this test, 

sample collection procedures have to be standardized step by 

step. In particular, the way in which the aspirated samples is 

aliquoted into routine smears and the buffer for DNA extraction 

is crucial; in this step the “informativeness” of the material 

both for cytopathological and molecular diagnosis needs 

147 

to be carefully preserved. Here we present a study recently 

undertaken to assess whether our method of FNA preparation 

is suitable to implement BRAF testing without interfering 

with routine cytology. 

Methods. One-hundred and twenty-eight cases were picked 

up consecutively without any selection among the FNAs 

routinely performed in the outpatient clinic, at the University 

“Federico II” of Naples. Totally, three needle passes were 

taken in each case. As usual Diff-Quick smears were prepared 

from the first two passages by the nodule. When the adequacy 

criteria were fulfilled, the whole tissue material from the 

third pass was collected into a tube containing 500 µl of the 

nucleic acids preservative solution (RNA later. Ambion). In 

the case that the first two needle passes failed to provide a 

fully satisfactory sample, the third needle pass was used for 

direct smears and the remainder material was collected for 

molecular testing. Cases were classified according to scheme 

suggested by the NCI Thyroid FNA State of the Science Conference1. 

Regardless of the collection method, all samples 

were similarly processed and exon 15 BRAF mutational 

analysis was performed as previously described 6 . 

Results. Basing on a satisfactory on-site evaluation, a BRAF 

dedicated third pass was performed in 44 (34%) cases; concordance 

between preliminary impressions and the final diagnosis 

was found in 42/44 (96%) cases. Conversely, in 84 (66%) 

cases additional smears were prepared from the third pass. 

This latter group included two cases (2,3%) in which the final 

diagnosis could not be rendered due to scant cellularity. Final 

cytological diagnosis of most nodules 110/128 (86%) cases 

was benign; this category included nodular goiter (n = 61), 

colloid nodules (n = 39), goiter with Hurtle changes (n = 4), 

goiter with associated chronic lymphocytic thyroiditis (n = 4) 

and hyperplastic/adenomatoid nodule in goiter (n = 2). In six 

cases (4,6%) mixed features of both hyperplastic/adenomatoid 

nodules and follicular neoplasm were observed; in these case 

a diagnosis of follicular lesion of undetermined significance 

(FLUS) was issued. In two cases (1,5%) follicular neoplasms 

features were observed. In three cases (2,3%), there was a suspicion 

of papillary thyroid cancer. Five cases (3,9%) showing 

clear-cut PTC nuclear changes were diagnosed as malignant. 

DNA was isolated from 128 consecutive samples collected 

during thyroid FNA. In 44 cases the whole tissue material 

obtained from a dedicated pass was extracted, whereas in the 

remaining 84 cases only the remainder material was employed 

for DNA extraction. The quantity of isolated nucleic acids 

ranged from 500 pg/µl to 309 ng/µl in the first group and from 

500 pg/µl to 16,5 ng/µl to in the second group. Higher average 

of extracted DNA concentration was observed in the dedicated 

pass group (25,9 ng/µl vs 7,9 ng/µl). Benign FNA had 

a BRAF dedicated pass in 32%, whereas more often (68%) 

the third pass was dedicated to the preparation of additional 

smears. Conversely, the dedicated dedicated pass was often 

performed in the FLUS (50%), follicular lesions (100%), 

suspect (33%) and malignant (60%) groups. The vast majority 

of samples (95.3%) showed successful exon 15 BRAF amplification. 

Only 6 samples (4.6%), nearly all from the needle 

rinsing group (5/6), had insufficient and/or poor quality DNA 

and were excluded from the analysis. Two of these cases 

were inadequate for cytological diagnosis too. BRAFV600E 

mutation was found in three cases. One case had a cytological 

diagnosis of suspect for PTC, whereas other two cases had a 

diagnosis of PTC. All other diagnostic group (benign, FLUS, 

follicular neoplasms) showed wild type exon 15 BRAF in all 

examined cases. We conclude that the FNA collection protocol 

here shown proved to be highly efficient. Cytological

148 

and molecular tests gave adequate results respectively in the 

98,4% and in 95,3%. In particular our method of aliquoting 

the aspirated samples into routine smears and the buffer for 

DNA extraction did not affects the “informativeness” of the 

cytopathological diagnosis. Recently, Xing suggested that, 

for prognostic purpose, perhaps all patients with cytologically 

diagnosed PTC should be preoperatively tested for BRAF 

mutation. In this respect this test provides information that 

are additional and non redundant to those provided by a well 

taken and correctly interpreted thyroid FNA. Our data showed 

that in a routine clinical setting FNA specimens can properly 

be handled to provide both morphological and molecular information. 

Although a large number of studies have reported 

BRAF analysis of thyroid nodules aspirates only recently a 

prospective study, on a large of number and with a complete 

molecular analysis, was published. However on site-evaluation 

was performed only in a minority of cases and the issues 

of sample collection was not taken into account 7 . Our study 

focusing on the single steps required to aliquot the aspirated 

material into routine smears and DNA extraction buffer may 

help to implement BRAF testing in the prognostic evaluation 

of PTC diagnosed by FNA. Our proposed method ensures 

that this test does not interfere with conventional cytology 

diagnostic accuracy. 

references 

1 Baloch ZW, et al. Cytojournal 2008;5:6. 

2 Alexander EK, et al. J Clin Endocrinol Metab 2002;87:4924-7. 

3 Cohen Y, Xet al. J Natl Cancer Inst 2003;95:625-7. 

4 Xing M, et al. J Clin Endocrinol Metab 2005;90:6373-9. 

5 Xing M. Endocr Rev 2007;28:742-62. 

6 Troncone G, et al. Cytojournal 2008;5:2. 

7 Nikiforov YE, et al. J Clin Endocrinol Metab 2009;94:2092-8 

Molecular diagnostic of solid tumors: a practical 

approach for systematic pathology. urinary 

system 

D. Segala, M. Brunelli, G. Martignoni 

Department of diagnostic pathology, University of Verona, Verona, 

Italy 

Clinically robust molecular tests are necessary in current 

clinical management of urological malignancies in order to 

improve the faculties of choosing the right therapy and screening 

patients for target therapies. 

Several promising biomarkers for diagnosis, prognosis and 

target therapy are now under evaluation. 

Urothelial carcinoma of the bladder. The five-years survival 

rate for localized bladder cancers and distant metastasis are 

94% and 6%, respectively. This fact highlights the importance 

of detection and appropriate therapeutic intervention at early 

stages of disease. 

Two forms of noninvasive bladder cancer are well known 

to have a distinct histology and clinical behaviour: papillary 

urothelial carcinoma rarely invades and metastasized, whereas 

flat carcinoma in situ (CIS) is known to have a high rate of 

invasion and metastasis. 

Molecular evidences of the presence of distinct pathogenesis 

for this two phenotype of urothelial carcinoma are now increasing. 

Both tyrosine kinase receptor FGFR-3 and H-RAS 

oncogene are primarily involved in the pathogenetic pathway 

of low-grade papillary urothelial carcinoma 1 . Flat carcinoma 

in situ and invasive urothelial carcinoma predominantly involve 

tumour suppressor genes p53, p16 and Rb 2-4 . Tumor 

angiogenetic factors are also involved the tumor-promoting 


extracellular environment 5 6 . 

Chromosomal aberrations are also described in the pathogenesis 

of bladder cancer. Chromosome 9 alterations are established 

to be the earliest events in both pathways of urothelial 

carcinoma 7 8 . Moreover, gains of chromosome 3p, 7p, 17q, 

and 9p21 deletions (p16 locus) are of special interest given 

their potential diagnostic value. 

The diagnostic process is usually supported by cistoscopy, 

while urine cytology is the most widely used non-invasive 

test to detect urothelial tumors. However, the letter is limited 

by its low sensitivity. Recently the FDA approved a new 

technique which seems to show better specificity ranges, the 

multitarget multicolor fluorescence in situ hybridization assay 

(UroVysion TM ) 9-12 , that is based on frequent numerical 

chromosomal alterations detection and it consists of fluorescently 

labelled DNA probes to the pericentromeric regions 

of chromosome 3 (red), 7 (green), 17 (aqua) and to the locus 

9p21 (gold). With the exception of one study 13 , UroVysion TM 

appears to enhance the sensitivity of routine cytology analysis 

and it can be used in combination with routine cytology in 

case with atypical cytology. 

Diagnostic applications of UroVysion TM on histology has also 

been suggested, such as the distinction of inverted urothelial 

papilloma and bladder carcinoma with endophytic growth pattern. 

In fact, a study described chromosomal abnormalities in 

72% of bladder carcinomas, in contrast to the absence of gains 

and deletions in inverted papilloma 14 . 

Since an increasing number of data suggests the presence of 

the same typical urothelial carcinoma chromosomal aberration 

also in rare histologic subtypes 15 16 (e.g. clear cell adenocarcinoma 

and small cell carcinoma of the bladder), the possible 

use of UroVysion TM could be a valid tool in the diagnosis of 

these rare entities. 

Adenocarcinoma of the prostate. The challenge in the years 

to come will be to introduce new gene-based diagnostic and 

prognostic tests in algorithms integrating the other known 

clinical and pathological factors to better manage diagnostic 

and therapeutic strategies. 

In seek of this purpose, in the last decade an extensive list of 

molecular biomarkers has been evaluated. One of the most 

notable discoveries is presence of recurrent chromosomal rearrangements, 

which lead to a fusion of the androgen-responsive 

promoter elements of the TMPRSS2 gene (21q22) to one of 

the three of the ETS transcription factors family members ERG 

(21q22), ETV (7p21) and ETV4 (17q21) 17 . The prognostic role 

of these rearrangements remains controversial, but this discovery 

has a great implication in terms of providing new markers 

for molecular diagnostic and target therapy 18 19 . 

A large number of prognostic molecular markers still waits 

for additional studies before eventually undergoing clinical 

trials. p27 and p53 tumour suppressor genes expression has 

been demonstrated to have a correlation with progression after 

prostatectomy 20-23 . Furthermore, several recent studies have 

established the importance of PTEN/PI3K/mTOR (mammalian 

target of rapamycin) in cell growth, proliferation and oncogenesis 

of prostate cancer 24-29 . Finally, some papers suggest the potential 

usefulness of proliferation index (ki-67) 30 , microvessel 

density 31 and nuclear morphometry 32 , while some others lack 

to confirm their prognostic validity 23 33 34 . 

Gene expression profiling studies using cDNA microarrays 

identified three genetic-differentiated subclasses of prostate 

tumours 35 . High grade, advanced stage and recurrent tumours 

where much more represented among two of the three subtypes, 

one of which also included most lymph node metastases. 

Another study, using array-based comparative genomic

lectures 

hybridization (array CGH), identified a series of recurrent 

DNA aberrations 36 . Deletions at 5q21 and 6q15 were associated 

with favourable outcome group; 8p21 (NKX3-1) and 

21q22 (TMPRSS2-ERG fusion) deletion group, 8q24 (MYC) 

and 16p13 gains and loss at 10q23 (PTEN) and 16q23 groups 

correlating with metastatic disease. 

Germ cell tumors of the testis. The etiopathogenesis of germ 

cell tumors of the testis depend on both environmental and 

genetic factors acting on the primordial germ cells/gonocyte 

that led to the precursor lesion called intratubular germ cell 

neoplasia. This precursor can progress to invasive components 

divide into seminomas and nonseminomas, with different 

histology and therapeutic response. 

Several immunohistochemical markers are described as useful 

in differential diagnosis of TGCTs. Briefly, seminoma is 

characterized by the expression of OCT3/4 37 38 , PALP 39 40 , 

c-KIT 41 and the variable expression of citokeratins 42 , embryonal 

carcinoma stains for CD30 41 , OCT3/4 37 38 , PLAP, SOX2 43 

and citokeratins, yolk sac tumor shows positivity for AFP 44 

and PALP but it is negative for OCT3/4 38 . Choriocarcinoma 

is lighted by the immunoexpression of β-hCG 40 . 

Studies of familial cases and genome-wide analysis do not 

reveal a constant genetic origin, suggesting that multiple 

foci must contribute to the development and progression of 

TGCTs. 

In line with the origin of TGCTs, individuals with disorders 

of sex development show an increased risk for this kind of 

cancer, in which they Y chromosome genetic material is crucial 

45 46 . In fact, the recently described microdeletion of the 

long arm of the Y chromosome, involving the AZoospermia 

Factor (AZF)c region 47 , seems to be a significant risk factor 

for impaired spermatogenesis 48 . 

Among somatic chromosomal changes in TGCTs the only 

recurrent structural imbalance appears to be the gain of 

chromosome 12p 49 50 , mostly as isochromosomes, that is described 

as related to tumor progression. Possible diagnostic 

applications of Interphase Fluorescence In Situ Hybridization 

(FISH) analysis of chromosome 12p abnormalities have been 

proposed, such as the distinction of Epidermoid Cysts of the 

testis, a benign lesion that lack the gain of 12p, to pure mature 

Teratomas 51 . 

Rarely primitive and metastatic germ cell malignancies present 

with histologic features of somatic-type origin. A FISH 

study demonstrated the presence of 12p abnormalities in 6 out 

of 10 metastatic somatic-type malignancies in patients with 

history of testicular or mediastinal germ cell tumors, suggesting 

the utility of this genetic marker in differential diagnosis 

of metastatic tumors 52 . 

Renal Cell Carcinoma (RCC). Histological subtyping of 

renal cell tumors is now considered an important prognostic 

factor in order to plan appropriate follow-up strategies. 

Moreover, numerous targeted agents have been developed 

for treatment of patients with renal cell carcinoma. For these 

reasons cytogenetical analyses are becoming an essential improvement 

in the routine diagnostic practice. 

Clear cell RCC is characterized by the mutation of the Von 

Hippel-Lindau syndrome (VHL) gene mapping in the chromosomal 

region 3p25 and the deletion of chromosome 3p, 

easy detectable by FISH analysis 53 . Papillary RCC shows trisomy 

of chromosomes 7, 17 and loss of the Y chromosome 54 . 

The distinction of clear cell papillary RCC 55 56 , a recently 

described rare entity, from Clear cell RCC and papillary RCC 

can be difficult; FISH can resolve this differential diagnosis, 

in fact this lesion don’t show gains of chromosomes 7 and 

17 and 3p deletion 56 . Metanephric adenoma and mucinous 

149 

tubular spindle cell carcinoma, two others rare entities present 

in WHO 2004 classification, could be misdiagnosed as 

papillary RCC. FISH analysis for chromosome 7 and 17 is 

helpful since there are no numerical alterations of these chromosomes 

in these tumors 57-60 . Among oncocytic neoplasms, 

chromophobe RCC is characterized by a combination of loss 

of chromosomes Y, 1, 2, 6, 10, 17 and 21, both classic and eosinophilic 

variants and renal oncocytoma, a benign lesion that 

sometimes enters in differential diagnosis with eosinophilic 

variant of chromophobe RCC, showed normal karyotype in 

the majority of cases 61 . 

On the other hand, the information obtained by molecular 

tumor markers are expected to revolutionize the staging of 

RCC. A large set of immunohistochemical markers such as 

Ki-67 62 63 , p53 64-66 , CAIX 67 , ADPF 68 , EGFR 69 and VEGR 70 

have been investigated. A negative prognostic role different 

mTOR pathway members was recently underlined 71 72 . Moreover, 

some prognostic chromosomic aberrations are recently 

observed in clear cell RCCs: gains or losses of 5q21 73 and 

loss of chromosome 14q 74 have been shown a correlation with 

progression (5p21), higher stage, higher histologic grade and 

poorer outcome (14q). Loss of chromosome 9p, observed in 

18% of clear cell RCC, is described as an independent negative 

prognostic factor 75 . 

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broader perspective. Nat Rev Cancer 2005;5:210-22. 

51 Cheng L, Zhang S, MacLennan GT, et al. Interphase fluorescence in 

situ hybridization analysis of chromosome 12p abnormalities is useful 

for distinguishing epidermoid cysts of the testis from pure mature 

teratoma. Clin Cancer Res 2006;12:5668-72. 

52 Kernek KM, Brunelli M, Ulbright TM, et al. Fluorescence in situ hybridization 

analysis of chromosome 12p in paraffin-embedded tissue 

is useful for establishing germ cell origin of metastatic tumors. Mod 

Pathol 2004;17:1309-13. 

53 Yamaguchi S, Yoshihiro S, Matsuyama H, et al. The allelic loss of 

chromosome 3p25 with c-myc gain is related to the development of 

clear-cell renal cell carcinoma. Clin Genet 2003;63:184-91. 

54 Brunelli M, Eble JN, Zhang S, et al. Gains of chromosomes 7, 17, 12, 

16, and 20 and loss of Y occur early in the evolution of papillary renal 

cell neoplasia: a fluorescent in situ hybridization study. Mod Pathol 

2003;16:1053-9. 

55 Tickoo SK, dePeralta-Venturina MN, Harik LR, et al. Spectrum of 

epithelial neoplasms in end-stage renal disease: an experience from 

66 tumor-bearing kidneys with emphasis on histologic patterns distinct 

from those in sporadic adult renal neoplasia. Am J Surg Pathol 

2006;30:141-53. 

56 Gobbo S, Eble JN, Grignon DJ, et al. Clear Cell Papillary Renal Cell 

Carcinoma: A Distinct Histopathologic and Molecular Genetic Entity. 

Am J Surg Pathol 2008 in press. 

57 Cossu-Rocca P, Eble JN, Delahunt B, et al. Renal mucinous tubular 

and spindle carcinoma lacks the gains of chromosomes 7 and 17 and 

losses of chromosome Y that are prevalent in papillary renal cell carcinoma. 

Mod Pathol 2006;19:488-93. 

58 Srigley J. Mucinous tubular and spindle cell carcinoma. In: Eble JN, 

Sauter G, Epstein JI, Sesterhenn IA (eds). World Health Organization 

Classification of Tumours: Pathology and Genetics of Tumours of the 

Urinary System and Male Genital Organs. Lyon: IARC Press 2004, 

p. 40. 

59 Renshaw AA, Maurici D, Fletcher JA. Cytologic and fluorescence 

in situ hybridization (FISH) examination of metanephric adenoma. 

Diagn Cytopathol 1997;16:107-11. 

60 Brunelli M, Eble JN, Zhang S, et al. Metanephric adenoma lacks 

the gains of chromosomes 7 and 17 and loss of Y that are typical of 

papillary renal cell carcinoma and papillary adenoma. Mod Pathol 

2003;16:1060-3.

lectures 

61 Brunelli M, Eble JN, Zhang S, et al Eosinophilic and classic chromophobe 

renal cell carcinomas have similar frequent losses of multiple 

chromosomes from among chromosomes 1, 2, 6, 10, and 17, and this 

pattern of genetic abnormality is not present in renal oncocytoma. 

Mod Pathol 2005;18:161-9. 

62 Visapaa H, Bui M, Huang Y, et al. Correlation of Ki-67 and gelsolin 

expression to clinical outcome in renal clear cell carcinoma. Urology 

2003;61:845-50. 

63 Dudderidge TJ, Stoeber K, Loddo M, et al. Mcm2, Geminin, and KI67 

define proliferative state and are prognostic markers in renal cell 

carcinoma. Clin Cancer Res 2005;11:2510-7. 

64 Zigeuner R, Ratschek M, Rehak P, et al. Value of p53 as a prognostic 

marker in histologic subtypes of renal cell carcinoma: a systematic 

analysis of primary and metastatic tumor tissue. Urology 2004;63:651- 

5. 

65 Shvarts O, Seligson D, Lam J, et al. p53 is an independent predictor of 

tumor recurrence and progression after nephrectomy in patients with 

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66 Rioux-Leclercq N, Turlin B, Bansard J, et al. Value of immunohistochemical 

Ki-67 and p53 determinations as predictive factors of 

outcome in renal cell carcinoma. Urology 2000;55:501-5. 

67 Bui MH, Seligson D, Han KR, et al. Carbonic anhydrase IX is an 

independent predictor of survival in advanced renal clear cell carcinoma: 

implications for prognosis and therapy. Clin Cancer Res 

2003;9:802-11. 

The new test of the screening for the prevention 

of cervical carcinoma: experience in Abruzzo 

region 

C. Angeloni 

Coordinator of Screening Project on Cervical Carcinoma in Abruzzo 

Background. The scientific evidence that the infection of 

Human Papilloma Virus (HPV) is the main cause of the cervical 

carcinoma has opened a new scenery in terms of primary 

prevention with vaccination and secondary prevention with 

the introduction of new screening technologies. The test of 

HPV DNA as the test of the main screening demonstrated 

indeed a sensibility that is absolutely higher than the Pap test 

both for women aged between 25 and 34 and for women of superior 

years without showing any significant over-diagnosis. 

These recent data and the consideration of the raised predictive 

negative value of HPV DNA test can make one consider 

the possibility to use this test as main test for the screening, 

reserving to the Pap the triage in secondary level for positive 

HPV DNA test cases of high risk. Another element in favour 

of the introduction of HPV DNA test is the higher reproducibility 

comparing to Pap test. 

Therefore the National Centre for Disease Prevention and 

Control (CCM) by the Health Ministry is considering to 

modify further the Guidelines. It is fundamental that the pilot 

projects co-ordinate activities among themselves and share 

data, results and protocols to produce a series of conclusive 

data for the applicability of this strategy. 

The GISCi (Italian Study Group on Cervical Carcinoma) 

shares this position that foresees the introduction of HPV test 

in the main screening with controlled applications to test it in 

practice and it has arranged a proper document by consensus. 

Methods. The main objective of the study is to evaluate the 

applicability of the screening programme based on HPV test 

SIAPeC-IAP meets SICI 

Moderators: P. Maioli (Ravenna), A. Bondi (Bologna) 

151 

68 Yao M, Tabuchi H, Nagashima Y, et al. Gene expression analysis of 

renal carcinoma: adipose differentiation-related protein as a potential 

diagnostic and prognostic biomarker for clear-cell renal carcinoma. J 

Pathol 2005;205:377-87. 

69 Moch H, Sauter G, Buchholz N, et al. Epidermal growth factor receptor 

expression is associated with rapid tumor cell proliferation in 

renal cell carcinoma. Hum Pathol 1997;28:1255-9. 

70 Jacobsen J, Grankvist K, Rasmuson T, et al. Expression of vascular 

endothelial growth factor protein in human renal cell carcinoma. BJU 

Int 2004;93:297-302. 

71 Pantuck AJ, Seligson DB, Klatte T, et al. Prognostic relevance of the 

mTOR pathway in renal cell carcinoma: implications for molecular 

patient selection for targeted therapy. Cancer 2007;109:2257-67. 

72 Pantuck AJ, Thomas G, Belldegrun AS, et al. Mammalian target of rapamycin 

inhibitors in renal cell carcinoma: current status and future 

applications. Semin Oncol 2006;33:607-13. 

73 Nagao K, Yoshihiro S, Matsuyama H, et al. Clinical significance of allelic 

loss of chromosome region 5q22.3 approximately q23.2 in nonpapillary 

renal cell carcinoma. Cancer Genet Cytogenet 2002;136:23-30. 

74 Herbers J, Schullerus D, Muller H, et al. Significance of chromosome 

arm 14q loss in nonpapillary renal cell carcinomas. Genes Chromosomes 

Cancer 1997;19:29-35. 

75 Brunelli M, Eccher A, Gobbo S, et al. Loss of chromosome 9p is an 

independent prognostic factor in patients with clear cell renal cell 

carcinoma. Mod Pathol 2008;21:1-6. 

in a regional territory, already under coverage with a traditional 

methodology and particularly characterized by centralized 

management of the programme and deep experience of new 

technologies and computer based systems. The study represents 

an important diagnostic instrument of a new protocol of 

screening in a scenery with different complexity. The results 

obtained from this study might represent a preliminary and 

necessary element for the introduction of HPV test as routine 

test within screening programmes. 

Women between 25 and 64 years old resident in Abruzzo 

region and suitable to the screening, will be asked to have 

another HPV test done, after three years from the previous 

screening. 

In our Regional Project we have decided to adopt the strategy 

HC2 as main screening followed by the triage with cytology 

for all ages organized by the screening (25-64 years old) for a 

major adaptability and simplicity of using the programme and 

also for having a lowest cost, as a double strategy of screening 

is not forseen, so the number of professional staff can be 

limited and the centralization of the cytological triage can be 

facilitated. We are expecting to modify the actual strategy of 

the screening presupposing a long period of the rescreening 

(5 years?) with a consequent reduction of costs and a more 

favourable model of costs/benefits. 

The area of l’Aquila and its surroundings, devastated by last 

year earthquake, will be reached in different ways, such as 

mobile medical vehicles. 

The study will involve around 60.000 women in 2010. 

The samples will be taken in decentralized sampling centres 

in Abruzzo, using the vial for the ThinPrep (Hologic). Women 

will be shown the procedures of HPV test, along with its clinical 

and preventive importance, however, they will be asked 

to fill in an agreement form. In the agreement form signed by 

the women there will be expected the creation of a biological 

bank to preserve the residual rates of the material taken from

152 

HPV test that, as it introduced by the protocol, could be used 

for successive cytological tests of triage and for further elaborations 

of studies with a particular attention for the markers 

of the specificity of the infection caused by HPV and of the 

progression of the neoplastic pathology. 

The samples will be sent to the medical centres of Atri and 

Sulmona, chosen to perform the test for the research of high 

risk DNA HPV (Hybrid Capture 2, cut-off 1pg/ml), using 

an automated system which will make it possible to process 

a high number of samples a day, with very fast turnaround, 

costs saving and quality improvement. Women with negative 

results will receive a letter with the test results along with an 

invitation to a new screening test after 3 years. At the moment, 

the 3 years time lag is prudentially recommended, but 

it is more likely that it will be extended to a 5-6 years, when 

all the evidences on the duration of protective effects will 

be proved and when the information will be updated by the 

Ministry of Health. 

The typing of positive results of high risk is centralized in 

laboratories of molecular biology in Atri and Sulmona; in fact 

we have forseen a triage with a specified type typing to value 

better HPV positive women considering that also women 

HPV 16 and 18 positive have a high risk to develop a Cin3+. 

HPV HG test with positive results of high risk will be reported 

to the Centre of Lanciano, entitled to cytological readings 

of second level which will take care of slides preparation, 

colouring and reading slides. Women with positive cytology 

will be asked to take a colposcopy examination. Women with 

negative cytology will receive an invitation letter for a further 

examination in one year. 

To guarantee the necessary sharing of results in order to create 

conclusive data on the application of this strategy, we have 

adopted a protocol analogous of other studies ongoing. 

The software used for the screening is the only one in Abruzzo: 

it’s based on Web servers and LAN networks connected 

with the colon rectum screening. The software managed by 

Winsap on Web, adopted by Abruzzo region for the screening 

that uses the base of traced records of the regional vital 

statistics and is continuously updated by our operators, has 

been adjusted by the creation of a HPV module. A particular 

element of quality has been presented by the traced record 

produced individually, transmissible through the New Sanitary 

Information Service (NSIS) to the national Data Ware 

House that allows to equalize, to centralize and to simplify 

statistical analysis. 

A new screening methodology, which involves a first level 

test different from the traditional one, and which detects a 

viral infection sexually transmitted, needs of course a different 

approach in terms of communication strategies. To 

avoid useless over-treatments, it’s necessary to introduce a 

new way of communication, which has to be scientific, but 

at the same time easy to understand without creating anxiety 

in women. 

For these reasons the project introduces, for the new screening 

type, the use of information material, scientifically correct 

and easy to understand by population, people involved in the 

medical centers and doctors of general medicine. Each invitation 

letter for the test comes with a brochure, written with 

simple and clear terms. 

The given information illustrates the concept of the cervical 

oncogenic risk underlining how the virus test results negative 

for 90% of women over 30 and therefore allows to include 

the tested subject among those of extremely reduced at risk 

to develop a significant cervical pathology whereas the persistent 

positivity in the virus test represents a simple indicator 


of a probable development of cervical pathology in years 

analogously of any other test of screening usually made in the 

medical prevention (weight, nutrition, etc.). 

For all the levels of our regional project of screening there will 

be settled a Quality Assurance programme. The creation of 

a biological bank will allow to study molecular mechanisms 

especially with regard to the determinants of progression and 

regression of the infection itself and of the intraepithelial 

cervical lesions. 

We have already known that only persistent infections of HPV 

are associated with a high risk of precancerous lesions. At this 

moment persistent infections can only be valued by repeating 

the test after 12 months whereas a clinical validation is necessary 

for the study and the characterization of markers of the 

integration HPV-DNA and DNA cells that could signal the 

latency state, the persistence of the infection and the progression 

to cancer. 

At the moment there are not biomarkers of specificity in the 

algorithms such as p16 and p16 Plus dual kit or mRNA, which 

are extremely encouraging, but still under specific experimental 

studies: it will be the person in charge of reading the 

cytological triage to decide whether to use it or not. 

In case of a CIN diagnosis, it is up to the pathologist, to guarantee 

a more accurate diagnosis, to search for a confirmation 

with the p16 histological test. 

Results. The cost of the strategy of the screening with HPV 

test as first level will be established regarding to the costs 

met in the last decade with the use of a traditional strategy 

of screening (Pap test I level) and with the adoption of new 

technologies and computerized systems of cytological reading 

(see attached: study ARINT of Abruzzo region and the project 

of the research applied for programmes of screening by law 

138 approved and financed by the CCM of the Health Ministry 

for the year 2009) considering also the possible saving 

derive from the eventuality of the expected extension of the 

interval of the screening. 

The heterogeneity of accounting systems and even more the 

lacking criterions of analytic accounting stand in the way of 

an activity based costing system that would be essential for 

estimating the financial requirements. 

On the other hand, the necessary overcoming of the criterion 

obsoleted by the historic cost requires analysis and applications 

of alternative systems. A recent decree Calderoli (known 

as decree on ‘Federalism’ converted in law recently) has 

moved in this direction establishing that costs having reference 

according to the letter m) of the second paragraph of the 

article 117 of the Constitution (that concerns Essential Levels 

of Assistance including screenings) ‘should determine with 

respect the standard costs associated on essential levels of 

services established by the state law, to be distributed in terms 

of efficiency and appropriateness in all the national territory’ 

(articles 6, paragraph 1, letter b). 

It is about a sector of studies not having been yet explored and 

not lacking of difficulties also because the decree does not 

make clear what ‘standard costs’ means and therefore how it 

should be calculated. 

Moreover, for some economists it seems to be an unrealistic 

idea that the efficient specific cost could be calculated for 

every singular service of the National Health Service (SSN) 

and then have by a simple summation the costs of services 

of the Essential Level of Assistance (LEA) in the decision of 

the total requirements. They think: ‘it appears substantially 

out of reach for services of prevention and for those of territorial 

medicine’ to prevent a tariff system analogous of that 

of hospitals (for regional decree DRG), also because of the

lectures 

heterogeneity of accounting systems and the lacking valuation 

systems for cost centres. 

Even believing that the determination of an ‘efficient cost’ 

is hardly an achievable desire, sometimes it is possible to 

prevent a charging of non-hospital services and programmes 

of screenings, just like our regional one that arranges an 

analytical accounting system for cost centres, represent a possible 

application of innovative systems in the accounting and 

organizational system. 

Therefore coherently of what has been arranged recently, using 

the study of the valuation of costs by law 138, we are going 

to perform also the innovative valuation of standard costs 

in our regional project of screening. 

Our preliminary data, based on about 12.000 HPV DNA test, 

show 9% positive rate with a positive PAPtest triage of 30%. 

High-throughput type-specific detection of HPV 

using massarray technology 

V. Mantovani, E. Marasco, P. Garagnani, M. Cricca * , 

M. Zerbini * , P. Chieco 

Centro Ricerca Biomedica Applicata (CRBA); * U.O. Microbiologia e 

Virologia, Policlinico “S. Orsola-Malpighi”, Bologna 

Background. The persistent infection of oncogenic high 

risk Human Papillomavirus (HPV) is one of the major risk 

factors for cervical cancer and the presence of HPV DNA is 

detected in nearly 100% of invasive cervical cancers. Several 

studies showed that the HPV DNA can be included in the 

screening for the prevention of cervical cancer in addition 

to, or in substitution of the current cytological analysis (PAP 

test), increasing the sensitivity and discovering earlier the 

carcinogenesis process. However, this diagnostic approach is 

not yet extensively applied because of the high costs of the 

molecular tests. 

Methods. Aim of our project is the development and the 

evaluation of a reliable, low-cost method based on mass spectrometry 

(MALDI-TOF) for type-specific detection of HPV 

DNA. The protocol has been optimized on the MassARRAY 

platform (Sequenom) located in the CRBA laboratories. 

The mass-spectrometry is a precise technology, often taken 

as gold standard for biochemical analyses. The highthroughput 

MassARRAY platform can simultaneously test 

384 samples and it is well suitable for large screening. The 

methodology doesn’t need expensive immunometric reagents, 

it can simultaneously perform several tests and it is 

easily automatable. 

Results. The test here proposed identifies the twelve high risk 

HPV types (16, 18, 31, 33, 35, 39, 45, 51, 52,56, 58 and 59), 

in addition to the six probably high risk (26, 53, 66, 68, 73 

and 82). Sensitivity and specificity are very high for the major 

types. In addition, our method identified some HPV infections 

missed by standard hybridization test. 

A low cost detection of the most relevant HPV types may be 

an important advance in assessing the impact of HPV vaccines, 

allowing the monitoring of the future changes in typespecific 

prevalence in infection and cancer. Our approach is 

very innovative in comparison with the existing methods, 

combining high efficiency, high sensitivity and low costs, 

suitable for routinely diagnostic activity, as well as for current 

screening programs. 

Streamlining the urinary oncology cytological 

service in the metropolitan area 

153 

R. Rapezzi, A. Bondi 

Oncological Science Department, U.O. Anatomy, pathological histology 

and cytodiagnostic departmentOspedale Maggiore, Bologna Local 

Health Unit (AUSL) 

Background. Bladder carcinoma in men ranks fourth in 

terms of frequency after prostate, lung and colorectal cancer, 

accounting for 5.5% of all cancer cases; among women it 

ranks eighth in terms of frequency, accounting for 2.3% of 

all female neoplasias. The higher incidence among men than 

women (4:1 ratio) is presumably associated with greater exposure 

to risk factors in the workplace such as chemical agents 

(2-naftilamine, benzidine, 4-aminobiphenile), as well as with 

a more widespread cigarette smoking habit. 

Cigarette smoking increases by two to five times the risk associated 

with bladder carcinoma; those who quit reduce such 

risk by 30-60%. 

Other risk factors associated with this type of carcinoma are 

recurring infections; the genetic-hereditary risk causes, on 

the contrary, play only a marginal role. From a histology 

viewpoint, in Europe and North America, bladder carcinomas 

are transitional in 90-95% of cases, in 3% of cases squamous, 

while 2% are adenocarcinomas and other histotypes are less 

than 1%. Out of the total transitional carcinoma cases, 70% 

are superficial and about 30% invasive. Transitional carcinomas 

present a high risk of relapse; five-year survival rates 

are closely correlated to staging; they range from 85-65% 

for stage 0-I to 14% in the case of metastatic carcinomas. In 

superficial bladder carcinomas, the histological grading provides 

important clues as to how the disease is progressing: low 

grades show a 4-5% progression, while in high-grade cases 

the figure reaches 39%. Good survival rates and the subsequent 

follow-up mean that the citological urine examination 

– in spite of its limitations – is important both for the initial 

diagnosis of a urinary system pathology, and for the follow-up 

of oncological patients. 

Bladder carcinomas may be asymptomatic, but in 80% of cases 

they are associated with haematuria:persistant macrohaematuria 

or microhaematuria, therefore, are the most frequent 

indicators of the need for a cytological test. In Europe the 

highest incidence of bladder carcinoma (Clinical guidelines 

2005 Boccardo and Silvestrini CNR-MIUR) was reported in 

Italy, with 14,000 new cases among men and 3,000 among 

women, followed by Spain and Switzerland; the highest mortality 

rates, on the other hand, are reported in Denmark,Spain, 

Poland and Malta. According to the Mortality Register of the 

Emilia Romagna Region (1998-2004) this type of tumour 

causes about five hundreds deaths a year, of which just over 

one hundred are women and almost four hundred men. The 

relative risk, in the Municipality of Bologna only, is > 1.3 

and the distribution pattern is extremely uniform, especially 

among males. This tumour seldom appears before the age of 

forty; this is why the reorganisation proposed for the province 

of Bologna mainly involves citizens over sixty, who account 

for more than 30% of the resident population. 

Methods. Health care for citizens in the Bologna provincial 

area is guaranteed by the Bologna and Imola local health 

units. The provincial user base on 31/12/2008 consisted of 

1,105,764 people; nine hundred thousand of them refer to the 

Bologna Local Health Unit which includes fifty municipalities 

(Imola covers ten) and is one of the largest health trusts 

in Italy.

154 

In total there are twelve hospitals, one health hub, nine accredited 

clinics and seven districts; this means that any action or 

change regarding the system requires substantial organisation 

work. The privatization of health trusts and the complexity 

of the established facilities mean that it is becoming increasing 

urgent to plan, streamline and control the health services 

rendered. Within this framework and in order to meet these 

requirements, a reorganisation process has been started as regards 

urinary cytology in the whole province of Bologna, involving 

all stakeholders in the process, first and foremost the 

Pathological Anatomy and Unified Booking (CUP) services. 

The project included all four Pathological Anatomy departments 

dealing with this type of test: in Bologna these are the 

Ospedale Maggiore, the Ospedale Bellaria and the S. Orsola 

Hospital; in Imola it is the Pathological Anatomy department 

of the Imola Hospital. The project started with an assessment 

of the various process phases. 

The analysis revealed differences both regarding the sampling 

indications and the way the test was booked through the CUP; 

on the other hand, consistent features emerged both as regards 

the test’s setting-up, a single filtering layer with polycarbonate 

membranes, and the waiting times which could be as long 

as 60 days. The old method required delivery of a fresh urine 

sample for three days; this means that the patients had to go 

through five “steps” before getting the result (one booking 

from the CUP, three deliveries to the sample delivery Point, 

one report collection from the office in charge), and that the 

Pathological Anatomy departments needed to receive and 

process the three samples separately and then draft the reports 

on different days. The new method was introduced in April 

2009; it involves the use of an alcohol-based fixative liquid to 

correctly preserve the cellular elements in the sample which 

can be stored in a cool place for up to a week. The appropriate 

amount of fixative is stored in the three jars contained in 

the sponge housings which are part of the Kit provided by the 

manufacturer. The Kits are delivered both to CUP offices and 

to the chemists’ authorised to book these tests (about three 

hundred booking points). The Kit is delivered to the patient 

when the test is booked, together with a fact sheet explaining 

both the precautions and sampling method to be followed, 

plus the simplified questionnaire on the patient’s medical history 

which he/she has to fill in. This information is necessary 

in order to reconstruct a clinical record of patients who, in the 

case of the Bologna area, can choose between three different 

Pathological Anatomy departments which are not part of a 

network. 

Results. The patient returns the Kit, all of it at the same time, 

after having collected the urine at home for three days, following 

the method illustrated; an efficient transport system connects 

the delivery point to the Pathological Anatomy service 

in charge of the area, making sure that the material is promptly 

delivered. Here the material is all received together, which 

means that the Service secretariat has less work to do in the 

process; the setting-up involves completely filtering the three 

samples, all together, for each patient, then collecting the cells 

onto a single slide. As a consequence the process is less timeconsuming 

for the professionals involved, and it also allows 

for the production of a single report. The patients collect their 

reports on the set date from the booking structure; as a whole 

their number of visits to public facilities when a urine cytology 

examination is required have been reduced to three (one 

booking from the CUP, one journey to the delivery point, and 

one to collect the report). The delivery point staff has reduced 

the amount of time necessary for each user because there is 

only one delivery to be received, as opposed to three for the 


same number of days. The new process is proving advantageous 

for everybody, with the exception of the laboratory staff 

in charge of setting up the material because the filtration time 

has increased. In any case, the implementation of the system, 

after the necessary trial period, has led to easier accessibility to 

the test, and at the same time to a reduction of the waiting time 

required. The waiting times for a urine cytology exam now 

range between 3 and 15 days. Moreover, an agreement with 

the CUP2000 company, allows the Pathological Anatomy services 

to directly manage the booking schedules. A statistical 

report, provided on a regular basis by the CUP booking office 

management, makes it possible for the Pathological Anatomy 

departments to monitor the relevant trends, thus extending or 

reducing the booking schedules in order to meet the demand. 

The new method has been extended to hospital wards and 

homogeneously covers the whole provincial area. 

emerging prognostic and predictive factors 

in breast cancer: where are we? 

M. Mottolese, A. Di Benedetto, E. Melucci, S. Buglioni, 

L. Perracchio. 

Pathology Department, Regina Elena National Cancer Institute, 

Rome, Italy 

Background. Breast cancer (BC) is the most commonly 

occurring malignancy in women and is responsible for approximately 

500 000 deaths per year worldwide. Due to the 

remarkable heterogeneity of BC, mostly driven by genetic 

variability, a number of clinical and bio-pathological factors 

are routinely used to determine prognostic predictions of 

clinical relevance. Furthermore, decisions about the adjuvant 

chemotherapy (CT), mainly in early stage of the disease, are 

affected by a complex interplay of factors and guidelines 

stratify BC patients into prognostic subsets suggesting treatment 

protocols on the basis of the reported estimates of efficacy 

1 2 . Classical prognostic parameters include patient age, 

axillary lymph node status, tumor size, histological features 

(especially histological grade and lymphovascular invasion), 

estrogen receptor (ER)-progesterone receptor (PgR), and 

HER2 status. In addition, a recent report from the St Gallen 

International Expert Consensus recommends the use of proliferation 

markers (eg, Ki-67 and mitosis) and multigene assays 

when choosing appropriate systemic CT 3 . Although these factors 

may be of great clinical value, their role in determining 

prognosis and evaluating risk in an individual patient with BC 

is more limited, since patients with similar combinations of 

features may have very different clinical outcomes. In recent 

years gene expression profiling have been increasingly used 

aimed to improve BC classification and to assess prognosis 

and response to therapy 4 . Although the precise role of these 

novel molecular techniques in the routine management of BC 

patients is yet under investigation, certainly they may provide 

prognostic and predictive information often more useful than 

those provided by the traditional clinical and pathological 

factors 5 . In addition, thanks to this extended biological knowledge, 

we have the opportunity of identifying genes involved 

in responsiveness to therapy acquiring relevant information 

on drug resistance mechanisms. This may lead to the characterization 

of new therapeutic targets and the subsequent availability 

of more treatment options for patients with resistant 

disease 6 . 

HER2 Expression and Response toTrastuzumab and 

Chemotherapy. It is well established that expression of 

HER-2 is predictive of response to trastuzumab 7 . Retrospec-

lectures 

tive analysis of several adjuvant randomized studies indicated 

that HER-2 overexpression is associated with greater benefit 

from adjuvant anthracycline (AC)-containing regimens than 

from cyclophosphamide, methotrexate, and 5-fluorouracil 

(CMF)-type regimens 8 . Furthermore, the addition of paclitaxel 

to AC-based CT with AC-based chemotherapy alone 

also showed that the benefit from inclusion of paclitaxel was 

largely restricted to HER-2–overexpressing tumors 9 . Based 

on the available evidence, the American Society for Clinical 

Oncology Expert Panel on Tumor Markers in BC concluded 

that high levels of HER-2 expression may identify patients 

that will particularly benefit from AC-based adjuvant therapy, 

although HER2 negativity should not be used alone to exclude 

patients from this treatment 10 . 

Topoisomerase (TOP2A) Expression and Response to 

Anthracyclines. TOP2A, an essential component of the cell 

division machinery, is the molecular target of AC and high 

levels of the enzyme cause the formation of large amounts of 

AC:TOP2A complexes within the nucleus. In a retrospective 

analysis of two randomized studies, TOP2A amplification 

was a significant predictive factor for greater benefit from cyclophosphamide, 

epirubicin, and 5-fluorouracil therapy than 

from CMF 11 . These results were confirmed in a population of 

patients with metastatic BC who participated in a randomized 

clinical trial of AC-based CT with or without trastuzumab 12 . 

The FDA recently approved a TOP2A fluorescence in situ 

hybridization (FISH) assay (TOP2A FISH pharmDx; Dako, 

Glostrup, Denmark) to measure amplification of this gene in 

BC specimens. 

Molecular Classification. Microarray studies, using an intrinsic 

gene set, have now shown that differences in gene 

expression can account for much of the diversity in BC 4 13 14 . 

The largest difference in overall gene expression profile is 

observed between tumors that were ER positive or negative. 

ER negative tumors are further sub-divided into HER2 

positive and negative 4 . Therefore, hierarchical clustering of 

microarray data classified BC into four main groups: Luminal 

(LA, LB), HER2 and Basal-like/Triple negative subtypes. 

Luminal-type cancers are mostly ER positive, and patients 

with LA BC have the most favorable long-term survival (with 

endocrine therapy) compared with the other types, whereas 

HER-2–positive tumors are more sensitive to CT associated 

to anti HER2 trastuzumab therapy 15 . 

Gene expression profiling and prognosis. Gene-expression 

profiling has shown promise to distinguish between patients at 

low and high risk for developing distant metastases and identify 

those who are likely to benefit from adjuvant therapy 16 . 

The Rotterdam gene set, one of the prognostic genetic tests 

now commercially available, is a single 76-gene prognostic 

signature, able to predict distant metastatic recurrence with a 

sensitivity of 93% and a specificity of 48% 17 . The gene signature 

known as wound response indicator (WRI) identifies 

BC patients with a significant shorter overall and disease free 

survival than patients whose tumors did not express this gene 

signature 18 . The oncotype DX is a 21-gene indicator which, 

through an algorithm based on the expression levels of these 

genes, allows a Recurrence Score (RS) to be computed for 

each specimen correlated with the rate of distant recurrence at 

10 years. The assay uses fixed tumor specimens, rather than 

frozen tissue 19 . The MammaPrint-70-gene profile was developed 

from patients with lymph-node negative £55 years of 

155 

age BC. The assay uses frozen tumor specimens and separates 

patients developing distant metastases from those disease free 

within 5 years. The internal and external validation of this 

gene set led to the clearance of this test by the U.S. Food and 

Drug Administration (FDA), allowing the test to be marketed 

as a prognostic marker to be used with other clinicopathologic 

factors 20 . 

Conclusions. Current BC treatment guidelines are based on 

clinical trial evidence obtained in defined patient populations, 

and treatment algorithms are developed by relating clinical 

trial findings to specific patient subgroups. Nevertheless, better 

prognostic and, in particular, predictive factors are needed 

to assist in treatment decision-making on an individual patient 

basis. Considering prognostic markers, gene profiling seems 

promising, although further validation, particularly with 

respect to potential ‘predictive interactions’, is mandatory. 

For predictive testing, emerging evidence suggests TOP2A 

amplification or deletions may be appropriate factors for 

selecting patients for AC dosing. Gene expression profiling 

may become important as a tool to define predictive factors; 

however, more accurate statistical approaches able to analyze 

gene expression profiles, based on functional hypotheses 

about gene networks, will be essential. Finally, the recent 

discovery of a class of small noncoding endogenous RNA 

molecules, namely microRNA, which are frequently dysregulated 

in cancer has uncovered an entirely new repertoire of 

molecular factors upstream of gene expression. The potential 

role of miRNAs in BC management, particularly in improving 

current prognostic tools and achieving the goal of individualized 

cancer treatment is under investigation 21 . 

references 

1 Early Breast Cancer Trialists’ Collaborative Group. Lancet 

2005;365:1687-717. 

2 Lonning PE, Knappskog S, Staalesen V, et al. Ann Oncol 2007;18:1293- 

306. 

3 Goldhirsch A, Ingle JN, Gelber RD, et al. Ann Oncol 2009;20:1319- 

29. 

4 Sorlie T, Perou CM, Tibshirani R, et al. PNAS 2001;98:10869-74. 

5 Parker JS, Mullins M, Cheang MC, et al. J Clin Oncol 2009;27:1160- 

7. 

6 Geyer FC, Reis-Filho JS. Int J Surg Pathol 2009;17:285-302. 

7 Ménard S, Balsari A, Tagliabue E, et al. Ann Oncol 2008;19:1706- 

12. 

8 Pritchard KI, Shepherd LE, O’Malley FP et al. N Engl J Med 

2006;354:2103-11. 

9 Hayes DF, Thor AD, Dressler LG, et al. N Engl J Med 2007;357:1496- 

506. 

10 Harris L, Fritsche H, Mennel R, et al. J Clin Oncol 2007;25:5287- 

312. 

11 Knoop AS, Knudsen H, Balslev E, et al. J Clin Oncol 2005;23:7483- 

90. 

12 Press MF, Sauter G, Buyse M, et al. J Clin Oncol 2007;25(suppl 

18S):524. 

13 Lonning PE, Sorlie T, Borresen-Dale AL. Nat Clin Pract Oncol 

2005;2:26-33. 

14 Sotiriou C, Neo SY, McShane LM, et al. PNAS 2003;100:10393-8. 

15 Schnitt SJ. Mod Pathol 2010;23:S60-4. 

16 van de Vijver M. The Oncologist 2005;10(Suppl 2):30-4. 

17 Wang Y, Klijn JG, Zhang Y, et al. Lancet 2005;365:671-9. 

18 Chang HY, Sneddon JB, Alizadeh AA, et al. PLoS Biol 2004;2:E7. 

19 Paik S, Tang G, Shak S, et al. J Clin Oncol 2006;24:3726-34. 

20 van’t Veer LJ, Dai H, van de Vijver MJ, et al. Nature 2002;415:530- 

6. 

21 Heneghan HM, Miller N, Lowery AJ, et al. J Oncol 2009;doi:10.1155 

/2010/950201

156 

Back to histological subtyping of nsclc in the 

era of personalized treatments 

M. Papotti, L. Righi, M Volante 

Department of Clinical and Biological Sciences, University of Turin 

at San Luigi Hospital, Orbassano (Torino), Italy 

Background. Lung cancer classification was devised to 

work especially on surgical specimens and recognizes four 

major histological subtypes, namely squamous cell carcinoma 

(SQC), adenocarcinoma (ADC), large cell (LCC) and small 

cell carcinomas (SCLC). Such classification is more difficult 

to apply on cytological samples or small biopsies, especially 

in the presence of poorly differentiated tumors or of limiting 

factors, including tumor heterogeneity, extent of necrosis, 

limited number of viable cells, marked artifacts. Since in past 

years all non-small cell lung cancers (NSCLC), were generally 

treated with similar chemotherapy regimens, irrespective 

of the histotype, as opposed to SCLC, accurate lung cancer 

subtyping became less relevant for clinical purposes, a fact 

that lead pathologists to concentrate their efforts on the correct 

recognition of SCLC, only. As a matter of fact, on cytological 

or small biopsy samples, most pathologists are able 

to correctly differentiate SCLC from NSCLC, and within the 

NSCLC group to identify well- or moderately-differentiated 

SQC or ADC. However, a high percentage of cases are still 

simply diagnosed as NSCLC, as they are poorly differentiated 

tumors, lacking clear-cut morphologic signs of differentiation. 

In recent years, the advent of targeted therapies and novel 

chemotherapeutic agents showing differential efficacy or toxicity 

on specific NSCLC subtypes required a sudden call back 

to histological subtyping, which is becoming the milestone 

for personalized therapy (especially for unoperable patients, 

whose treatment will eventually be based on the biopsy diagnosis, 

only). 

A useful, rapid and cheap tool to identify squamous or glandular 

differentiation and characterize poorly differentiated 

NSCLC could be immunohistochemistry. Although according 

to the WHO classification “…classification is largely based 

on standard hematoxylin & eosin sections…”, the immunophenotypic 

profile may provide information in terms of probability 

level that a given neoplasm has squamous or glandular 

differentiation. This latter information may be of predictive 

value, assisting the clinician in selecting the most appropriate 

treatment for advanced NSCLC affected patients. 

Lung cancer histological subtypes that are morphologically recognizable 

on small biopsy fragments or cytological samples are 

basically three, i.e. ADC, SQC and SCLC. Other tumor types 

of lung carcinomas such as large-cell carcinoma (LCC) and 

its variants (eg large-cell neuroendocrine carcinoma, etc), or 

sarcomatoid carcinomas can be definitely diagnosed on surgical 

specimens, only. However, ADC variants cannot always be easily 

identified, with special reference to non invasive subtypes 

(former bronchiolo-alveolar carcinoma, now Adenocarcinoma 

in situ/minimally invasive adenocarcinoma), in the absence of 

the whole tumor specimen available for thorough examination. 

Molecular tests may be applied also in biopsy material and may 

help to refine the diagnosis, since some correlations were observed 

between molecular profile and histological subtype (eg 


SIAPeC-IAP meets the Adriatic Society of Pathology. 25 th year 

Moderators: M. Del Vecchio (Ascoli Piceno), V. Pisac Presutic (Spalato) 

EGFR mutations in BAC or mixed or papillary ADC, or K-RAS 

mutations in mucinous ADC). 

As stated, immunohistochemistry is also very helpful to identify 

the three most frequent lung tumor phenotypes: glandular, 

squamous and neuroendocrine. The group of large cell carcinomas 

frequently has an heterogeneous immunohistochemical 

profile, probably reflecting divergent differentiation mainly 

along squamous and glandular lineages. Adenocarcinomas are 

generally positive for TTF-1, surfactant apo-protein A (SPA), 

napsin-A, and cytokeratin 7 (CK7) and negative for CK5/6, 

CK20 and p63 (an exception is mucinous adenocarcinoma, 

which expresses CK20 rather than TTF-1 or SPA). Squamous 

cell carcinoma consistently and strongly react with p63, CK5 

and desmocollin-3, and virtually never for TTF-1. Neuroendocrine 

large and small cell carcinomas are known to express 

chromogranin A, synaptophysin and CD56, among others. 

In the daily practice a panel of immunohistochemical markers 

is generally employed, based on availability of reagents 

and the pathologist’s personal experience. The most widely 

applied panel for NSCLC sub-classification includes TTF- 

1, p63, CK7 and CK5. The former two are nuclear markers 

and seem more reliable in poorly cellular samples. In such 

cases, cocktails of antibodies can also be used, to reduce the 

number of necessary glass slides (for example p63+CK5 vs 

TTF1+CK7). With regard to the interpretation of results, 

TTF-1 is virtually never expressed in SQC, but stains only 

70-80% of ADC (depending on tumor grade and the presence 

of mucinous features). By contrast, p63 expression in SQC is 

robust and not influenced by tumor grade, although p63 immunoreactivity 

has been observed in a small subset of ADC 

(p40 seems a more squamous carcinoma specific marker, in 

this respect). Finally, in the presence of ambiguous phenotypes 

or discrepant marker reactivity, additional antibodies 

may be used. Promising results were obtained with napsin- 

A, MUC5AC or desmocollin-3 in discriminating pulmonary 

ADC from SQC. 

Once a morphological and/or immunohistochemistry-assisted 

accurate lung cancer subtyping has been obtained, the 

final step of pathological characterisation of lung tumors 

is their molecular profile. This can be optimally defined in 

surgical specimens, but can be assessed in small cytological 

or biopsy samples, too. EGFR or K-ras mutational status is 

the most common requirement for personalizing treatments, 

but new targets are emerging for specific drugs including 

c-met mutations, ALK fusion products and the levels of 

specific enzymes, such as ERCC1, thymidilate synthase or 

topoisomerase II. 

Conclusions. 1) An accurate histological subtyping of so 

called “NSCLC” may further improve the efficacy or reduce 

the toxicity associated to novel therapeutic options; 2) although 

lung cancer diagnosis is generally based on haematoxylin/eosin-stain, 

immunohistochemistry can be helpful, if not 

mandatory, in defining the histotype (or the most likely differentiation 

lineage) of poorly differentiated tumors; 3) TTF-1 & 

CK7 and p63 & CK5 seem to date the most valuable markers 

for ADC and SQC, respectively; 4) novel diagnostic markers 

may allow to abandon the “NSCLC” category, thus reducing 

as much as possible the number of unclassified cases.

lectures 

Mutation analyses of KRAS in colorectal cancer 

and egfr in non-small cell lung cancer: a task 

for pathologists? 

M. Dietel 

Institute of Pathology, Charité Universitätsmedizin Berlin, Germany 

Background. Colorectal cancer (CRC) and non-small cell 

lung carcinoma (NSCLC) are the two most common malignancy 

in the western world with estimated 350,000 new cases 

reported for the United States in 2008 1 . Since both tumors 

are being predominantly diagnosed at advanced stage, the option 

of a curative therapy then does no longer exist in many 

instances and chemotherapy is often the treatment of choice. 

However, conventional anti-cancer drugs have been shown 

to be of limited value accompanied by strong side effects. 

This situation was the driving force to search for new more 

specific drugs. 

The membrane bound epidermal growth factor receptor 

(EGFR1) molecule was found to be of major importance in 

growth stimulation und thus was identified as a possible target 

which inhibition may lead to reduced tumor growth. Meanwhile 

there exist two types of EGFR-inhibitors which are 

approved for clinical application, i.e. the therapeutical anti- 

EGFR antibodies cetuximab (Erbitux ®) ) and Panitumumab 

(Vectibix ® ) as well as the tyrosine kinase inhibitors (TKI) 

erlotinib (Tarceva ® )) and gefitinib (Iressa ® ). In several clinical 

studies it became obvious that not all tumors respond equally 

but that certain genetic characteristics are the prerequisite to 

clinical benefit. This was the background to link the application 

of the drugs to pre-therapeutic eligibility tests and that 

in Europe KRAS mutation testing as well as EGFR mutation 

testing became a prerequisite for anti-epidermal growth factor 

receptor therapy of metastatic colorectal cancer since the end 

of 2007 2 3 and metastatic non-small cell lung cancer (NSCLC) 

in 2009, respectively. 

Since the analyses have to be performed using carefully selected 

tumor tissue the tests should be done only in Institutes 

of Pathology where pathologists are able to check istology, 

select the tumor tissue adequate for molecular testing and sign 

out a combined morphological/molecular report. In addition 

each institute should participate in interdisciplinary ring trials 

(round robin tests) which should be lead by an independent 

organisation. For that purpose the German Society of Pathology 

(DGP) and the German Association of Pathologist have 

(BDP) created the QuiP (Quality in Pathology) – initiative 

which organizes interlaboratory tests and confers the respected 

certification. Only if a reliable morphological diagnosis 

is combined with a solid genetic analysis a robust basis for 

targeted therapy is given. 

Methods. A multitude of procedures and methods are available 

for detecting KRAS/EGFR mutations in tumor samples 

(Weichert 4-11). In the Berlin Institute the following selection 

of techniques has been found useful, reliable and applicable 

for routine molecular pathology. 

Tissue selection. As shown in several studies 12 13 a precise selection 

of the tissue to be analysed is mandatory. This should 

be done by an experienced pathologist indicating the area on 

H&E-stained slides estimating the percentage of tumor in relation 

to the whole tissue section. Subsequently a manual microdissection 

has to be done by the technician. Tissue samples 

can be stored for years prior to molecular analyses. 

DNA preparation. For DNA preparation the three unstained 

slides were used. The putative tumor areas corresponding to 

157 

the marks on the H&E slide were microdissected. DNA preparation 

was done as described previously 14 . In brief, microdisseceted 

tissue was transferred to 180 µl ATL-buffer (Qiagen, 

Hilden, Germany) and kept for 10 min at 95°C. After cooling 

down to room temperature, 20 µl of proteinase K solution 

were added. After gentle mixing, the sample was incubated at 

55°C until complete lysis (after about 2 h). The further steps 

of isolation of DNA follow the protocol “Tissue Protocol- 

QIAamp DNA Mini Kit” (Qiagen). The nucleic acids were 

eluted at a volume of 60-100 µl and DNA content was estimated 

with a Nanodrop 1000 (PeqLab, Erlangen Germany). 

For sequence analyses the techniques of Sanger sequencing, 

pyro-sequencing, chip analyses and melting curve analyses 

were applied (technical details see Weichert et al. 15 ). 

Results. Institute of Pathology, Berlin. For the analysis of 

the somatic KRAS genotype of 263 patients we used Sangersequencing, 

pyrosequencing, melting curve analysis and chip 

hybridization in parallel. We used Sanger sequencing as the 

reference method. For all cases DNA of sufficient quality was 

prepared. Overall mean (± SD) DNA yield was 196.8 ng/µl 

(± 142.5 ng/µl). Array analysis, melting curve analysis and 

pyrosequencing, using DNA from the same preparation, was 

performed in 233, 188 and 136 cases, respectively. 

Using Sanger sequencing 108 out of 260 cases (41.5%) were 

found to have a mutation in either codon 12 (31.9%) or codon 

13 (9.6%) of the KRAS gene. The most frequent mutations 

were p.G12D (12.7%), p.G12V (10.8%) and p.G13D (9.2%). 

Similar distributions were seen with the other three methods. 

The array analysis identified 104 out of 223 cases (44.6%) to 

harbor somatic mutations, melting curve analysis found 77 

out of 188 cases (41%) to be mutated, and by pyrosequencing 

51 out of 136 cases (37.5%) were reported to carry mutations. 

A crossover comparison of the four methods yielded k values 

exceeding 0.9 (for more details see 15 ). 

Analogue test comparisons were done for EGFR mutation 

analyses using tissue from NSCLC after surgical tumor resection 

revealing similar results. 

QuiP-Initiative. Detailed descriptions of the results regarding 

round robin tests for KRAS- and EGFR-testing are published 

elsewhere. In summary, around Germany currently there exist 

over 60 Institutes of Pathology certified for KRAS testing and 

53 certified for EGFR testing (for details visit the home page 

of the DGP 17 ). 

Discussion. Somatic gain-of-function mutations in the KRAS 

gene of CRCs predict the lack of response to anti-EGFR therapy 

with cetuximab and panitumumab, and KRAS mutational 

screening prior to treatment with either drug has become mandatory 

in Europe since the end of 2007. A similar situation 

become relevant in 2009 when clinical approval of gefitinib 

was limited to “tumours that have tested positive for EGFR 

with an activating mutation” (EMEA 2009;2,3). This resulted 

in a fast growing completely new branch of routine predictive 

diagnostic molecular pathology. 

The pre-therapeutical analyses which guide patients’ therapy 

are chance and challenge for pathologists as a new personal 

responsibility is given. To fulfil this following points have to 

be considered: 

1. The test has to be done only in certified Institutes of Pathology, 

e.g. with a QuiP-certicate, for details see ref. 17 . 

2. The responsible pathologists should be experienced in morphology 

and molecular testing. 

3. The responsible pathologists should: 

• re-confirm the histological diagnosis on an H&E slide 

and

158 

• he should identify and mark the tumor area for enrichment 

of tumor cells. 

4. This is followed by manual microdissection to assure that 

at least 40% of the material for the molecular analysis is 

indeed tumor tissue. 

5. The selected tumor tissue then should be analyzed following 

the procedure and recommendations described above 

and by others. 

6. Finally the responsible pathologists should prepare a combined 

report giving details on the histology and the molecular 

result. 

If these criteria are met molecular pathology is facing an excellent 

future coming closer to clinical decisions and thus to 

the patients. 

references 

1 Jemal A, Siegel R, Ward E, et al. Cancer statistics, 2008. CA Cancer J 

Clin. 2008;58:71-96. 

2 http://www.emea.europa.eu/humandocs/Humans/EPAR/erbitux/erbitux.htm 

3 http://www.emea.europa.eu/humandocs/Humans/EPAR/vectibix/ 

vectibix.htm 

4 Simi L, Pratesi N, Vignoli M, et al. High-resolution melting analysis 

for rapid detection of KRAS, BRAF, and PIK3CA gene mutations in 

colorectal cancer. Am J Clin Pathol 2008;130(2):247-53. 

5 Ogino S, Kawasaki T, Brahmandam M, et al. Sensitive sequencing 

method for KRAS mutation detection by Pyrosequencing. J Mol Diagn. 

2005;7:413-21. 

6 Clayton SJ, Scott FM, Walker J, et al. K-ras point mutation detection 

in lung cancer: comparison of two approaches to somatic mutation 

detection using ARMS allele-specific amplification. Clin Chem 

2000;46:1929-38. 

The role of the pathologist in the assessment of 

kidney adequacy 

G. Monga, G. Mazzucco * 

Dipartimento di Scienze Mediche. Facoltà di Medicina e Chirurgia. 

Università del Piemonte Orientale, Amedeo Avogadro. Novara; * Dipartimento 

di Scienze Biomediche e Oncologia Umana. Facoltà di 

Medicina e Chirurgia. Università di Torino 

Every year, no more than 1/3-1/4 of patients awaiting kidney 

transplant can receive the graft. This shortage of grafts has 

led to an ever increasing use of kidneys from marginal deceased 

donors (subjects aged ≥ 55 years or < 55 years with 

a history of hypertension and/or diabetes, or deceased after 

a cerebrovascular incident). At present, pretransplant renal 

biopsy (PTRB) is the most reliable method available to assess 

the kidney state. 

However, there are several problems connected to this diagnostic 

procedure: 

1. Morphologic evaluation of fixed and paraffin embedded 

samples vs frozen tissue. The former procedure is greatly 

favoured. Indeed, the frozen sections technique allows for a 

faster evaluation, offering briefer diagnostic times. However, 

the price is paid by the quality of the material, which is less 

satisfactory than that available after fixation and paraffin 

embedding. 

2. Semiquantitative vs morphometric evaluation of the morphologic 

changes. The latter procedure is more accurate, but 


Transplantation pathology 

Moderators: F.W. Grigioni (Bologna), M. Rugge (Padova) 

7 Lilleberg SL, Durocher J, Sanders C, et al. High sensitivity scanning 

of colorectal tumors and matched plasma DNA for mutations in APC, 

TP53, K-RAS, and BRAF genes with a novel DHPLC fluorescence 

detection platform. Ann NY Acad Sci 2004;1022:250-6. 

8 Rothschild CB, Brewer CS, Loggie B, et al. Detection of colorectal 

cancer K-ras mutations using a simplified oligonucleotide ligation 

assay. J Immunol Methods 1997;206:11-9. 

9 Emanuel JR, Damico C, Ahn S, et al. Highly sensitive nonradioactive 

single-strand conformational polymorphism: detection of Ki-ras mutations. 

Diagn Mol Pathol 1996;5:260-4. 

10 Keohavong P, Zhu D, Whiteside TL, et al. Detection of infrequent and 

multiple K-ras mutations in human tumors and tumor-adjacent tissues. 

Anal Biochem 1997;247:394-403. 

11 Ward R, Hawkins N, O’Grady R, et al. Restriction endonucleasemediated 

selective polymerase chain reaction: a novel assay for the 

detection of K-ras mutations in clinical samples. Am J Pathol 1998, 

153:373-9. 

12 Amado RG, Wolf M, Peeters M, et al. Wild-type KRAS is required for 

panitumumab efficacy in patients with metastatic colorectal cancer. J 

Clin Oncol 2008;26(10):1626-34. 

13 Lièvre A, Bachet JB, Boige V, et al. KRAS mutations as an independent 

prognostic factor in patients with advanced colorectal cancer 

treated with cetuximab. J Clin Oncol 2008;26(3):374-9. 

14 Petersen I, Schewe C, Schlüns K, et al. Inter-laboratory validation of 

PCR-based HPV detection in pathology specimens. Virchows Arch 

2007;451:701-16. 

15 Weichert W, Schewe C, Lehmann A, et al. KRAS Genotyping of 

Paraffin-Embedded Colorectal Cancer Tissue in Routine: Diagnostics 

Comparison of Methods and Impact of Histology. J Mol Diagn 

2010;12:35-42. 

16 Neumann J, Zeindl-Eberhart E, Kirchner T, et al. Frequency and 

type of KRAS mutations in routine diagnostic analysis of metastatic 

colorectal cancer. Pathol Res Pract. 2009;205(12):858-62. 

17 http://www.dgp-berlin.de 

time consuming and, therefore, unsuitable in an emergency 

diagnostic setting. It follows that semiquantitative evaluation 

must be used. 

3. Bioptical procedure: needle (NB) vs wedge (WB) biopsy. 

Opinions as to the primacy are conflicting among both surgeons 

and pathologists. WB offers larger amounts of tissue, 

but, according to several authors, increases the risk of an overestimation 

of glomerular sclerosis (GS) and interstitial fibrosis 

and allows for only limited sampling of the deeper renal tissue 

where larger arteries are present. 

4. The choice of the morphological parameters for the grading 

of the kidney damage. Global GS alone has been used, but 

opinions on this procedure are conflicting. At present, besides 

GS, three other parameters (interstitial fibrosis, tubular atrophy 

and vascular arterio-arteriolosclerotic damage) are usually 

included in different scoring systems. 

PTRB was considered useful in this setting in the prediction of 

short- and long-term graft outcome, in supplying a reference 

frame in the interpretation of subsequent graft biopsies and 

mandatory in the assessment of kidney adequacy for single 

and/or double transplant or its being discarded 1 . 

Since PTRB is justified on the assumption that it represents 

the real state of the whole kidney, it follows that the critical 

issue is its reliability, i.e. how accurately it represents the true 

histology of the whole kidney. This question prompted a study 

which has already been published 2 dealing with a comparative 

evaluation, according to the Karpinski et al. scoring system 3

lectures 

of 154 PTRB (118 NB and 36 WB) and the matched 154 kidneys, 

subsequently untransplanted for different reasons and 

used as gold standard. 

The concordance between the total score at biopsy and 

that on the kidney was fairly good, with higher values for 

NB (k 0.73) than for WB (k 0.57). When the individual 

parameters (global GS, interstitial fibrosis, tubular atrophy 

and vascular damage) were considered, agreement between 

biopsy and matched kidneys was found to be low for GS, 

mainly in the NB (k 0.18), intermediate for tubular atrophy 

and interstitial fibrosis and high for vascular changes (k 0.75 

and 0.74 for NB and WB respectively). It is worth stressing 

that agreement for each parameter (including GS) consistently 

increased when the biopsy contained more tissue. If 

the biopsies were subdivided into three categories, according 

to the number of the glomeruli (A: 6-10 glomeruli; B: 11-24 

glomeruli; C ≤ 25 glomeruli) considered as the index of the 

tissue amount, reliable results were achieved in those with 

more than 11 glomeruli. 

As to the judgement of the fitness for the transplant of the kid- 

159 

neys, biopsy and whole kidney evaluation agreed in 100/118 

cases of the NB group and in 29/36 cases of the WB group. 

Conclusive remarks 

1. PTRB supplies reliable information as to the actual kidney 

state, with slightly better results with NB than with WB. 

2. Although the biopsy size does play a role, samples with 

over 10 glomeruli suffice for a satisfactory evaluation. 

3. Vascular damage is the most faithful single parameter, 

whereas it is not the case of global GS, the estimation of 

which requires some caution. A multiparametric evaluation is 

strongly recommended. 

references 

1 Remuzzi G, Ruggenenti P. Renal transplantation: single or dual for 

donors aging = 60 years? Analyses & Commentaries. Transplantation 

2000;69:2000-4. 

2 Mazzucco G, Magnani C, Fortunato M, et al. Nephrology, Dialysis 

and Transplantation (in press). 

3 Karpinski J, Lajoie G, Cattran D, et al. Outcome of kidney transplantation 

from high-risk donors is determined by both structure and fuction. 

Transplantation 1999;67:1162-7. 

Pharmacogenetics in cancer: transfer of translational research 

into clinical practice 

Pharmacogenetics in cancer care: transferring 

translational research into clinical practice 

G. Toffoli 

Director Experimental and Clinical Pharmacology Unit, CRO National 

Cancer Institute (Aviano) 

Antitumoral drugs are characterized by a low therapeutic 

index, with a modest difference between toxic and efficient 

dose: this could determine in some individuals a number of 

toxic effects, to which a real benefit in terms of antitumoral 

activity does not always correspond. These compounds are 

also characterized by a sometimes very evident interindividual 

variability, and the same drug can determine in different individuals 

a different toxic effect or a different antitumoral 

activity. It is then important to personalize the antitumoral 

activity, that is to give each patient the right drug at the right 

dose. Therapy personalization represents one of the most 

important future challenges in the therapy of oncological 

patients. On this ground it is crucial to identify the biomolecular 

specificities of the neoplastic cells towards which 

the antitumoral drugs must be vehiculated, or to identify the 

genetic specificities of the patient that can explain the reason 

for the differential effect of antitumoral drugs. Pharmacogenetics, 

the study of the genetic basis of the interindividual 

differences in response to drugs results from the intersection 

between genetics and pharmacology is a notably interesting 

field for the possible implications in the clinical practice. 

Genetic polymorphisms, that is the structural alterations of 

genes involved in the metabolism, transport or drug interaction 

with the cellular target, have been described both for the 

drugs traditionally employed in the oncological patient, and 

for the so-called targeted molecules recently employed in 

clinic. Pharmacogenetics investigates the individual genetic 

specificities which are relatively common among the population 

and that are involved in antitumoral drugs action. The 

Moderators: A. Scarpa (Verona), P.P. Piccaluga (Bologna) 

most common form of genetic polymorphism is represented 

by the SNPs (single nucleotide polymorphisms). Other forms 

of polymorphisms are represented by abnormal nucleotide sequence 

repetitions (microsatellites or midisatellites), genomic 

duplications, pseudogenes etc. At present, more than 100 million 

SNPs of human genome have been described: it is then 

important to individuate those which have an effective impact 

on drug action, modulating their pharmacogenetics and pharmacogenomics 

(toxicity and response) and that can influence 

the ultimate result of the antitumoral therapy, influencing the 

patients survival. Sometimes the pharmacologic effect cannot 

be explained by the action of a single polymorphism. The 

complex metabolic or intercellular transport ways that often 

characterize antitumoral drugs require the involvement of 

more genes and only the precise definition of polymorphisms 

of all these genes results of some utility for prognosis. In the 

study of interactions between genome and drug, we can aim 

at a single polymorphism, applying in this case a pharmacogenetic 

approach or we can analyze the combined effect of 

more polymorphisms, employing in that case a pharmacogenomic 

approach. Basically, the strategies adopted at present 

in pharmacogenetics/omic studies are three: “candidate gene 

approach”, “gene pathway approach” and “genome wide 

analyses”. In the so-called “candidate gene approach” the 

polymorphisms of a single gene considered critical for the 

action of the drug are analyzed. This strategy presupposes 

that the action of the gene in question is actually important 

for the pharmacologic action and that the polymorphism or 

the polymorphisms of interest have an effective prognostic 

power in the clinical practice. It is also important to establish 

what is the combined effect of the polymorphisms present 

in a single gene and the modality of their distribution, that 

is if the contemporary presence of more polymorphisms in a 

single gene is casual or not (linkage disequilibrium). At present, 

the kits recommended by Food and Drug Administration

160 

(FDA) in the oncological field for single polymorphisms are 

relatively few (UGT1A1*28 (irinotecan) (invader assay) and 

TPMT (6-MP, azatioprine) (pro-Predict Py), nevertheless, the 

data recently produced in literature are encouraging for further 

clinical development. 

In the so-called “gene pathway approach”, the polymorphism 

of the genes involved in the complex metabolic ways of antitumoral 

drugs are analyzed. The rationale of this approach is 

that the pharmacologic effect is a multifactorial event and that 

the pleiotropic effect could depend on polymorphisms located 

in different genes. In this context, it is important to define 

for each antitumoral drug all polymorphisms with prognostic 

potentiality, and the attention of pharmaceutical companies is 

to design pharmacogenetic kits including all polymorphisms 

that are relevant for the action of a specific drug. 

The “genome wide analysis” represents an innovative strategy 

in the field of pharmacogenomics obtained by the recent 

advances in the technological field. With this approach, the 

entire human genome is analyzed, in an attempt to define 

clusters of polymorphisms predictive of the pharmacological 

effect. At present, this strategy is employed principally in the 

research field, to individuate those polymorphisms whose real 

meaning will be successively validated, but the possibility to 

adopt this strategy in the clinical practice represents a stimulating 

perspective, analogous to what is being done with the 

microarrays of gene expression in human tumors. 

Gene polymorphisms that could influence the biochemistry of 

the most common tumoral drugs have been described. Among 

the polymorphisms of enzymes involved in the metabolism 

of phase I of antitumoral compounds, the attention is pointed 

at the polymorphic variants of cytochrome isoforms, in particular 

3A4 and 3A5 (irinotecan, taxanes, alkilant agents). 

For what concerns phase II enzymes, we particularly focus 

on the polymorphisms in uridindifosfoglucoronosiltransferasi 

and in particular on UGT1A1*28 polymorphism in the gene 

that inactivates SN38, the active principle of irinotecan. We 

have recently published (Toffoli et al. J. Clin. Oncol. 2010) 

a phase I pharmacogenetic study to define the maximum 

tolerated dose of irinotecan associated with FOLFIRI regimen 

based on patients genetic characteristics. The study has 

demonstrated that the patients carrying the wild type genotype 

(UGT1A1) can tolerate higher drug dose with respect to patients 

carrying UGT1A1*28 polymorphism. We observed an 

increased response rate in patients receiving the higher dose 

of irinotecan. 

Other polymorphic variants of phase II enzymes concern the 

isoforms of glutathione S-transferase gene (GST). Numerous 

polymorphic variants of these genes that include allelic deletion 

or point mutations have been described. Recently, it was 

found how polymorphic variants of isoforms of GST gene can 

influence toxicity, and in particular neurotoxicity to chemotherapeutic 

regimens with oxaliplatin. 

Numerous polymorphisms have been described in genes 

involved in the action of fluoropyrimidines (thymidylate 

sintetasi, methylenetetrahydrofolate reductase and dihydropyrimidine 

dehydrogenase). In particular, IVS 14 + 1G > A 

variant dihydropyrimidine dehydrogenase can be associated 

with severe toxicity, even lethal, by fluorouracil. This polymorphism 

is extremely rare in our population (< 1%) but it 

should be attentively evaluated before starting a therapy with 

fluoropyrimidines. 

Finally, polymorphic variants have been described for the 

genes involved in the transmembrane transport of antitumoral 

drugs and in particular in MDR1 gene that codes for Pgp and 

for other “multidrug resistant proteins” MRPs. The real im- 


pact of these polymorphisms on toxicity and tumor response 

as well as on drug pharmacokinetics of these transporters 

needs further investigation. 

In conclusion, pharmacogenetic/omics represents a potential 

instrument for personalizing cancer therapy. This approach 

seems extremely interesting to reduce toxic effects of antitumoral 

drugs and for increasing their efficacy. Nevertheless, it 

is still necessary to validate this innovative perspective, both 

in analytical terms (sensibility, specificity and reproducibility 

of tests) and for what concerns the precise relations among 

genotype, pharmacokinetics and pharmacodynamics (toxicity 

and response). Finally, it is crucial to define the clinical utility 

of this approach. In particular, if based on a pharmacogenetic 

test the therapeutic regimen could be modified, what pharmacologic 

alternatives can be used and especially what will 

be the effect of these modifications in terms of toxicity and 

response to therapy. 

Pharmacogenomics of brain tumors 

L. Morandi, D. de Biase, G. Marucci, A Pession * , G Tallini 

Dipartimento di Ematologia e Scienze Oncologiche, Sezione di Anatomia 

Istologia e Citologia Patologica “M. Malpighi” Università- 

ASL Ospedale Bellaria; * Dipartimento di Patologia Sperimentale 

Università di Bologna 

Background. Epigenetic silencing of the MGMT gene by 

promoter methylation is associated with loss of MGMT expression, 

diminished DNA-repair activity and longer overall 

survival in patients with glioblastoma who, in addition to 

radiotherapy, received alkylating chemotherapy with carmustine 

or temozolomide (TMZ) 1-4 . The MGMT gene is located 

on chromosome 10q26 and encodes a DNA-repair protein 

that removes alkyl groups from the O 6 position of guanine, an 

important site of DNA alkylation. The restoration of the DNA 

consumes the MGMT protein, which the cell must replenish. 

Left unrepaired, chemotherapy-induced lesions, especially 

O6-methylguanine, trigger cytotoxicity and apoptosis 4 5 . High 

levels of MGMT activity in cancer cells create a resistant phenotype 

by blunting the therapeutic effect of alkylating agents 

and may be an important determinant of treatment failure 4-6 . 

Patients with glioblastoma containing a methylated MGMT 

promoter showed a major benefit from temozolomide 3 . Given 

the key roles of cytosine methylation, there has been a wide 

interest in the development of procedures for DNA methylation 

analyses 7-10 . Here we present a novel methylation sensitive 

quantitative real time PCR assay (MS-qLNAPCR) which 

permits high throughput quantification of the methylation 

status of the MGMT promoter in an accurate, very sensitive 

and cost-effective manner. High specificity was achieved 

recognizing methylated and unmethylated CpGs by 3’-locked 

nucleic acid (LNA) primers and molecular beacon probes. In 

order to calculate the ratio between methylated and unmethylated 

MGMT allele, the CpG islands of SNURF were selected 

as a reference gene. SNURF belongs to the 15q imprinted 

center mapped on 15q12. The maternal allele is usually methylated, 

while the paternal one is unmethylated 11 . In theory in 

a homogeneous population of cells of the same individual, 

the methylated maternal alleles should be balanced with the 

unmethylated paternal alleles if the tumor cells did not acquire 

any deletion for this locus or aberrant methylation of the paternal 

allele (loss of imprinting). This feature allows an easy 

and precise calculation of the ratio between the methylated 

and unmethylated alleles of MGMT following the method 

described by Ginzinger et al. 12 .

lectures 

Methods. 159 GBM patients followed prospectively between 

April 2004 and October 2008 were included in this study. 

After bisulfite treatment, methylated and unmethylated CpGs 

were detected by previously described MS-PCR 2 3 and by 

MS-qLNAPCR using LNA primers and molecular beacon 

probes 23 . SNURF was used as a reference for normalization 

allowing calculation of the percentage of MGMT methylation 

using the ∆∆Ct method 11 12 . Two SNPs adjacent to MGMT 

(rs8473; rs3740427) were used for loss of heterozygosity 

(LOH) analysis. They were interrogated by allele specific 

quantitative PCR using LNA at 3’- end of the discriminating 

primer as previously described 13 14 . 

Results. Concordance between already described nested MS- 

PCR and MS-qLNAPCR was found in 158 of 159 samples 

(99.4%). MGMT promoter was found to be methylated using 

MS-qLNAPCR in 70 patients (44.02%), and completely unmethylated 

in 89 samples (55.97%). Median overall survival 

was of 24 months, being 20 months and 36 months, in patients 

with MGMT unmethylated and methylated, respectively. Considering 

MGMT methylation data provided by MSqLNAPCR 

as a binary variable, overall survival was different between 

patients with GBM samples harboring MGMT promoter unmethylated 

and other patients with any percentage of MGMT 

methylation (p = 0.003). This difference was retained using 

other cut off values for MGMT methylation rate (i.e. 10% 

and 20% of methylated allele), while the difference was lost 

when 50% of MGMT methylated allele was used as cut-off. 

LNA modified primers for mMGMT showed higher PCR 

efficiency (slope: -3.271; efficiency: 102%,) than unmodified 

primers (slope: -4.339; efficiency: 69%). The analytical 

detection limit of 0.01% was reached only for LNA modified 

primers, while conventional primers showed a detection limit 

of 0.1%. To the best of our knowledge, we are the first to 

describe and validate a method to quantitate DNA methylation 

using LNA modified primers and an imprinted gene as 

a reference, instead of a methylation independent calibrator 

such as ACTB 15 . In our opinion ACTB does not represent the 

best reference gene for normalization because it is located at 

7p15-p12, a chromosomal site subject to copy number variations 

in gliomas 16 17 , and because it is close EGFR (located at 

7p12) that is amplified in about 40% of GBMs 18 . We use of 

an imprinted gene (SNURF) as an internal control, because it 

may check the efficiency of the assay from DNA purification, 

through bisulfite treatment to PCR. Additionally it should be 

used as a reference because mSNURF and uSNURF mimic the 

biallelic MGMT status. In fact, in normal cells the maternal allele 

of SNURF is methylated at the promoter locus, while the 

paternal allele of the same gene is usually unmethylated and 

expressed 11 . This condition is thus similar to a tumor population 

of cells in which the MGMT is methylated at one of the 

two alleles. In order to consider SNURF as an ideal reference, 

the ratio between methylated and unmethylated SNURF alleles 

might not be disturbed by copy number changes, or by 

loss of imprinting, both of which are common in cancer. However, 

several references demonstrate that SNURF, which has 

been mapped at 15q12, is hardly ever altered in gliomas 19 20 . 

These data were confirmed by our study because the methylated 

and unmethylated SNURF ∆Ct of the most part of cases 

(91.2%) was nearly always very close to 0. The SNURF methylation 

values outside the normality range in 14 of the 159 

GBMs (8.8%) may be due to a distinct CpG methylation pattern 

among tumor cell population, to partial loss of one allele 

(LOI: loss of imprinting), or to a methylation machinery disorder 

that methylates the paternal allele (GOI: gain of imprinting). 

The requirement for using SNURF as a reference is that 

161 

methylated and unmethylated SNURF alleles are at a ratio of 

1:1, and in our series in only one of these 14 cases did the ∆Ct 

of SNURF have a negative impact on the final calculation for 

the mMGMT/uMGMT ratio. In these circumstances we avoid 

using SNURF as a reference loosing relative quantification data. 

Additionally in cases with balanced ratio, the ∆Ct between 

uMGMT and uSNURF or mSNURF contributed to check 

for allelic imbalance status, as the PCR efficiencies of each 

marker are very closed to each other. These data were verified 

interrogating by ASqPCR the following SNPs adjacent to 

MGMT gene: rs8473 and rs3740427. Concordant results were 

obtained and 28.2% of cases showed AI. Among them 75.6% 

revealed MGMT methylation. AI alone and AI associated with 

MGMT methylation were not correlated with longer overall 

survival after TMZ treatment respect to cases with balanced 

copy number at this locus. Quantitative analysis showed a 

bimodal distribution of ratio values between methylated and 

unmethylated MGMT alleles, with two prevalent groups with 

ratios between 0.001-0.33 and 0.67-1, respectively. This bimodal 

distribution is similar to that found by Vlassenbroeck 

et al. 21 In summary, we report and clinically validate an accurate, 

robust, and cost effective MS-qLNAPCR protocol for 

the detection and quantification of methylated MGMT alleles. 

Using MS-qLNAPCR we demonstrate that even low levels of 

MGMT promoter methylation have to be taken into account to 

predict response to temozolomide-chemotherapy 22-24 . 

references 

1 Gerson SL. MGMT: its role in cancer aetiology and cancer therapeutics. 

Nat Rev Cancer 2004;4(4):296-307. 

2 Esteller M, Garcia-Foncillas J, Andion E, et al. Inactivation of the 

DNA-repair gene MGMT and the clinical response of gliomas to alkylating 

agents. N Engl J Med 2000;343:1350-4. 

3 Hegi ME, Diserens AC, Gorlia T, et al. MGMT gene silencing 

and benefit from temozolomide in glioblastoma. N Engl J Med 

2005;352(10):997-1003. 

4 Liu L, Markowitz S, Gerson SL. Mismatch repair mutations override 

alkyltransferase in conferring resistance to temozolomide but not to 

1,3-bis(2-chloroethyl)nitrosourea. Cancer Res 1996;56(23):5375-9. 

5 Gerson SL. MGMT: its role in cancer aetiology and cancer therapeutics. 

Nat Rev Cancer 2004;4(4):296-307. 

6 Komine C, Watanabe T, Katayama Y, et al. Promoter hypermethylation 

of the DNA repair gene O6-methylguanine-DNA methyltransferase 

is an independent predictor of shortened progression free 

survival in patients with low-grade diffuse astrocytomas. Brain Pathol 

2003;13(2):176-84. 

7 Liu ZJ, Maekawa M. Polymerase chain reaction-based methods of 

DNA methylation analysis. Anal Biochem 2003;317(2):259-65. 

8 Cottrell SE, Distler J, Goodman NS, et al. A real-time PCR assay for 

DNA-methylation using methylation-specific blockers. Nucleic Acids 

Res 2004;32(1):e10. 

9 Esteller M, Garcia-Foncillas J, Andion E, et al. Inactivation of the 

DNA-repair gene MGMT and the clinical response of gliomas to alkylating 

agents. N Engl J Med 2000;343:1350-4. 

10 Jeuken JW, Cornelissen SJ, Vriezen M, et al. MS-MLPA: an attractive 

alternative laboratory assay for robust, reliable, and semiquantitative 

detection of MGMT promoter hypermethylation in gliomas. Lab Invest 

2007;87(10):1055-65. 

11 El-Maarri O, Buiting K, Peery EG, et al. Maternal methylation 

imprints on human chromosome 15 are established during or after 

fertilization. Nat Genet 2001;27(3):341-4. 

12 Ginzinger DG, Godfrey TE, Nigro J, et al. Measurement of DNA copy 

number at microsatellite loci using quantitative PCR analysis. Cancer 

Res 2000;60(19):5405-9. 

13 Latorra D, Campbell K, Wolter A, et al. Enhanced allele-specific PCR 

discrimination in SNP genotyping using 3’ locked nucleic acid (LNA) 

primers. Hum Mutat 2003;22(1):79-85. 

14 Marucci G, Morandi L, Bartolomei I, et al. Amyotrophic lateral sclerosis 

with mutation of the Cu/Zn superoxide dismutase gene (SOD1) 

in a patient with Down syndrome. Neuromuscul Disord 2007;17(9- 

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162 

15 Vlassenbroeck I, Califice S, Diserens AC, et al. Validation of realtime 

methylation-specific PCR to determine O6-methylguanine-DNA 

methyltransferase gene promoter methylation in glioma. J Mol Diagn 

2008;10(4):332-7. 

16 Roversi G, Pfundt R, Moroni RF, et al. Identification of novel genomic 

markers related to progression to glioblastoma through genomic profiling 

of 25 primary glioma cell lines. Oncogene 2006;25(10):1571- 

83. 

17 Yin D, Ogawa S, Kawamata N, et al. High-resolution genomic copy 

number profiling of glioblastoma multiforme by single nucleotide 

polymorphism DNA microarray. Mol Cancer Res 2009;7(5):665-77. 

18 von Deimling A, Louis DN, von Ammon K, et al. Association of epidermal 

growth factor receptor gene amplification with loss of chromosome 

10 in human glioblastoma multiforme. J Neurosurg 1992;77:295- 

301. 

19 Freire P, Vilela M, Deus H, et al. Exploratory analysis of the copy 

number alterations in glioblastoma multiforme. PLoS One 2008;3(12): 

e4076. 

20 Lo KC, Bailey D, Burkhardt T, et al. Comprehensive analysis of loss 

of heterozygosity events in glioblastoma using the 100K SNP mapping 

arrays and comparison with copy number abnormalities defined by 

BAC array comparative genomic hybridization. Genes Chromosomes 

Cancer 2008;47(3):221-37. 

21 Vlassenbroeck I, Califice S, Diserens AC, et al. Validation of realtime 

methylation-specific PCR to determine O6-methylguanine-DNA 

methyltransferase gene promoter methylation in glioma. J Mol Diagn 

2008;10(4):332-7. 

22 Brandes AA, Tosoni A, Franceschi E, et al. Recurrence pattern after 

temozolomide concomitant with and adjuvant to radiotherapy in newly 

diagnosed patients with glioblastoma: correlation With MGMT promoter 

methylation status. J Clin Oncol 2009;27(8):1275-9. 

23 Morandi L, Franceschi E, De Biase D, et al. Promoter methylation 

analysis of O6-methylguanine-DNA methyltransferase in glioblastoma: 

detection by locked nucleic acid based quantitative PCR 

using an imprinted gene (SNURF) as a reference. BMC Cancer 

2010;10:48 

24 Brandes AA, Franceschi E, Tosoni A, et al. O(6)-methylguanine DNAmethyltransferase 

methylation status can change between first surgery 

for newly diagnosed glioblastoma and second surgery for recurrence: 

clinical implications. Neuro Oncol 2010;12(3):283-8. 

Pharmacogenetics in hematologic neoplasia 

S. Rasi, A. Bruscaggin, S. Franceschetti, R. Bruna, D. Rossi, 

G. Gaidano 

Division of Hematology, Department of Clinical and Experimental 

Medicine, Amedeo avogadro University of Eastern Piedmont, 28100 

Novara 

The field of pharmacogenetics investigates how genetic inheritance 

might influence the patients’ individual response to 

drugs. In fact, the majority of drugs exhibit large interindividual 

variability in their efficacy and toxicity, and this individual 

host response is influenced by genetic polymorphisms, 

in particular single nucleotide polymorphisms (SNPs). 

The main aim of pharmacogenetics is to optimize therapy 

based on the patient’s genotype, in order to maximize the 

therapeutic index of a given drug or regimen. Indeed, several 

pharmacogenetic studies have documented that host SNPs 

affecting genes involved in drug metabolism, detoxification, 

transport and targeting are in fact responsible, at least in part, 

for the interindividual variability in efficacy and toxicity of 

a given pharmacological treatment. Moreover, several drugs 

utilized in therapeutic treatment of hematologic neoplasms 

rely on DNA damage as part of their mechanisms of tumor 

cell killing. On these bases, treatment benefit and/or toxicities 

in patients may be modulated by the host DNA repair capacity. 

Also in this context, pharmacogenetic studies have shown 

that SNPs within genes of DNA repair pathways alter the host 

DNA repair capacity, thus affecting the individual response to 

drugs and the prognosis of the patients. 


One of the most consolidated pharmacogenetic model in 

hematological-oncology is represented by the case of acute 

lymphoblastic leukemia (ALL) of childhood. Among hematological 

neoplasms, ALL of childhood is the sole disease 

whose pharmacogenetics has been characterized in detail, 

and has entered clinical practice. The enzyme thiopurine Smethyltransferase 

(TPMT), classified as a phase II enzyme, 

metabolizes chemotherapeutic agents, and in particular is 

responsible for the metabolism of 6-mercaptopurine. The 

activity of the TPMT enzyme displays marked variability in 

the population, being influenced by SNPs of the TPMT gene 

that determine a reduction of the cellular content of TPMT. 

In particular, three allelic variants (TPMT*2, TPMT*3A, 

and TPMT*3C) are responsible for more than 90% of cases 

with an intermediate- or low-enzyme activity. ALL patients 

with a wild-type TPMT allele (TPMT*1 or TPMT*1S) and 

a non functional variant allele (TPMT*2, TPMT*3A, and 

TPMT*3C) have an intermediate activity of TPMT, while 

patients with two non functional variant alleles are TPMT 

deficient. In the context of ALL of childhood, the TPMT 

genotype identifies patients who are at risk of hematopoietic 

toxicity after thiopurine therapy. In fact, patients who are 

homozygous carriers of these SNPs and are therefore devoid 

of TPMT activity, are at higher risk of hematological toxicity 

when treated with 6-mercaptopurine. Instead, patients who 

are heterozygous carriers of these SNPs have an intermediate 

risk of dose-limiting toxicity. A 6-mercaptopurine dose reduction 

of 90% is generally required for homozygous-TPMT 

deficiency patients, whereas the TPMT heterozygous carriers 

require a mean dose reduction only of 35%. 

Instead, scant information is available about the impact of 

pharmacogenetics in predicting outcome and toxicity in the 

context of non-Hodgkin lymphoma (NHL) and available information 

is restricted to a limited number of studies. Studies 

in follicular lymphoma and diffuse large B cell lymphoma 

(DLBCL) aimed at assessing the association between antilymphoma 

efficacy of rituximab and SNPs of Fcγ receptors 

have reported conflicting results. Fcγ receptors are are involved 

in rituximab antibody-dependent cellular toxicity. A 

few studies have identified SNPs located in the Fcγ receptors 

(FcγRIIIa 158V/F and FcγRIIa 131H/R) as being associated 

with tumor response in patients treated with rituximab as 

first-line therapy. On the other hand, both in the context of 

follicular lymphoma and in DLBCL, other studies failed to 

identify an association between prognosis and SNPs of Fcγ 

receptors. 

In the context of DLBCL, the host pharmacogenetic background 

predicts efficacy of R-CHOP21 (R=rituximab, 

C=cyclophosphamide, H=doxorubicin, O=vincristine and 

P=prednisone) chemotherapy program, that is considered the 

standard treatment for this disease. In fact, SNPs modulating 

gene expression and/or function of enzymes involved 

in R-CHOP pharmacogenetics may contribute to prognostic 

stratification and prediction of toxicity in DLBCL patients 

treated with R-CHOP21. In particular, host SNPs affecting 

alkylating agent detoxification (GSTA1 rs3957357) and 

doxorubicin pharmacodynamics (CYBA rs4673) may predict 

survival in DLBCL treated with R-CHOP21. GSTA1 encodes 

a glutathione S-transferase that catalyses the conjugation of 

cyclophosphamide with glutathione to facilitate excretion. 

In particular, the GSTA1 rs3957357T minor allele associates 

with reduced levels of GSTA1 enzyme in healthy individuals, 

and predicts for reduced detoxification of alkylating agents, 

thus increasing tumor cell exposure to drug. In fact, DL- 

BCL patients carrying GSTA1 rs3957357 CT/TT genotypes

lectures 

displayed a better outcome compared to patients who carry 

the GSTA1 rs3957357 CC genotype. CYBA is a gene that 

encodes the p22phox subunit of the NAD(P)H oxidase complex. 

Individuals carrying the CYBA rs4673T minor allele 

have a substantial reduction in ROS (reactive oxygen species) 

generation by NAD(P)H oxidase. Because ROS generation is 

one of the antitumor mechanisms of doxorubicin, the CYBA 

rs4673T minor allele is expected to reduce the tumor cytotoxicity 

of doxorubicin based regimens. According to this 

model, DLBCL patients carrying CYBA rs4673 TT genotype 

displayed a poorer outcome compared to patients who carried 

the CYBA rs4673 CT/TT genotypes. 

Only few studies have shown that SNPs affecting genes involved 

in R-CHOP pharmacogenetics, in particular NAD(P)H 

oxidase subunit genes and ATP binding transporter genes, 

may predict toxicities of the CHOP regimen in DLBCL. In 

fact, in addition to outcome, the pharmacogenetic background 

of the host appears to be relevant for predicting R-CHOP21 

toxicity in DLBCL patients. In the context of DLBCL treated 

with R-CHOP21, a SNP of the NCF4 gene (rs1883112), that 

encodes the p40phox subunit of the NAD(P)H oxidase, recurs 

as a protective genotype against both hematologic and nonhematologic 

toxicities of R-CHOP21. In fact, carriers of the 

NCF4 rs1883112 G minor allele experience less frequently 

hematologic, infective and cardiac toxicity. NCF4 rs1883112 

affects the gene promoter with potential consequences on 

NCF4 expression and ROS generation, that may have a relevant 

function in several R-CHOP21 toxicities. Increased 

exposure to anthracycline-derived ROS is a widely accepted 

mechanisms of doxorubicin cardiotoxicity, and moreover 

ROS are involved in neutrophil death upon exposure to 

chemotherapy. In an other study, CHOP-induced cardiotoxicity 

has been shown to associate with selected SNPs of the 

NAD(P)H oxidase complex and with SNPs of ATP-binding 

cassette genes. 

In the context of DLBCL, SNPs modulating gene expression 

and/or function of enzymes involved in DNA repair pathways 

may contribute to the prognostic stratification in DLBCL 

patients treated with R-CHOP21. DNA damage is one of the 

163 

mainstay of cancer treatment. Cells can repair DNA damage 

induced by chemotherapeutic agents through several major 

repair pathways. Several drugs utilized in DLBCL treatment, 

including drugs of R-CHOP and second line regimens, rely on 

DNA damage for tumor killing. On these bases, DNA repair 

capacity may modulate treatment benefit and/or toxicities 

in DLBCL patients. In particular, a SNP of the MLH1 gene 

(rs1799977) is a predictor of survival in DLBCL treated with 

R-CHOP21. MLH1 rs1799977 is a nonsynonymous SNP 

causing the I219V amino acidic substitution on MLH1, a gene 

that encodes the mutL homolog 1 protein of the mismatch 

repair (MMR) pathway. In silico, MLH1 rs1799977 is predicted 

as a pathological SNP. In vitro, the G variant allele of 

MLH1 rs1799977 is known to associate with a reduction of 

MLH1 protein expression and function, and loss of MLH1 in 

tumor cells is known to induce refractoriness to doxorubicin 

and platinum compounds. According to these observations, 

DLBCL patients who carried the MLH1 rs1799977 AG/GG 

genotype displayed an increased risk of death compared to 

patients who carried the MLH1 rs1799977 AA genotypes. 

The poor prognosis heralded by MLH1 rs1799977 AG/GG 

genotype in DLBCL is due to an increased risk of failing both 

first and second line treatment. Consistently, DLBCL carriers 

of the MLH1 rs1799977 AG/GG genotypes displayed poor 

outcome possibly due to lack of MLH1 function. 

These studies of pharmacogenetics may contribute to increase 

knowledge in the field of advanced genomics applied to 

hematologic tumors. In particular, the expected results concern 

the possibility of identifying a priori, e.g. at the time of 

diagnosis, specific host pharmacogenetic profiles that may 

predict efficacy and toxicity of a given treatment. Overall, 

studies of host pharmacogenetics can therefore enable: i) the 

identification of novel markers for prognostic stratification 

and for toxicity prediction of pharmacological treatments in 

the context of hematological neoplasms; ii) the construction 

of a model for dose adjustement of drugs in order to maximize 

therapeutic benefit and minimize treatment toxicity; iii) the 

design of a personalized drug treatment for specific types of 

hematological neoplasms.

164 

role of endoscopy for polypoid 

and non-polypoid colonic lesions 

L.H. Eusebi, F. Bazzoli 

Department of Internal Medicine and Gastroenterology, University 

of Bologna, Italy 

Colorectal cancer (CRC) is the third most common cancer 

diagnosed in men and women and a leading cause cancer-related 

death worldwide. Declining rates in CRC incidence and 

mortality have been revealed by recent trends, due to reduced 

exposure to risk factors, screening’s effect on early detection 

and prevention of neoplastic and pre-neoplastic lesions, and 

improved treatment. 

Most colorectal cancers are believed to evolve through adenoma-carcinoma 

sequence; therefore, the goal of CRC screening 

is to reduce mortality through a reduction in incidence of advanced 

disease. Indeed, CRC screening can achieve this goal 

through the detection of adenomatous polyps and of earlystage 

adenocarcinomas, followed by endoscopic resection of 

such lesions. 

Screening programs are based on patients risk stratification, 

evaluating their personal, familial and clinical history. Indeed, 

identifying high risk patients, such as those with a family or 

personal history of CRC or adenomatous polyps, inflammatory 

bowel disease or of genetic syndromes such as Hereditary 

Non Polyposic Colorectal Cancer (HNPCC) and Familial 

Adenomatous Polyposis (FAP), allows to determine the most 

adequate screening strategy. 

Among the available screening techniques, several aspects 

underline the advantage of endoscopy; indeed, colonoscopy 

is the only single-stage strategy not requiring pretesting and 

polyps can be removed immediately during the screening 

procedure; besides, all other forms of screening, if positive, 

require colonoscopy as a second procedure 1 . 

The long term colorectal cancer incidence and mortality reduction 

provided by endoscopic polypectomy has been confirmed 

by the findings of the National Polyps Study. Indeed, in this 

study, patients with adenomas who had undergone endoscopic 

resection experienced a 76-90% reduction in CRC incidence 

compared with the expected general population incidence 2 . 

Other case-control and cohort studies have reported lower risk 

reduction rates of CRC after therapeutic colonoscopy than the 

National Polyp Study. Indeed, although endoscopy has a major 

protective role against colorectal cancer, colonoscopy is not an 

infallible “gold standard” and colonic lesions might still develop 

despite surveillance screening; detection miss-rates vary 

from 27% for adenomas < 5 mm to about 2% for CRC 3 4 . 

Several reasons might explain the imperfect colonoscopy 

protection such as the presence of rapidly growing tumours 

(HNPCC, increased risk of microsatellite instability in “interval” 

cancers), ineffective polypectomy, incomplete bowel 

preparation or ineffective application of current colonoscopic 

detection technologies 5 . 

Therefore, quality indicators and targets for colonoscopy, 

such as bowel preparation quality, cecal intubation rate, mean 


Thursday, September 23 rd , 2010 

Colon neoplasms 

Moderators: G. Lanza (Ferrara), M. Risio (Torino) 

colonoscopic withdrawal time, polyp detection rate and adverse 

or unplanned events occurring within 24 hours of colonoscopy, 

have been suggested to improve the effectiveness of 

the endoscopic inspection 6 . 

Furthermore, an underlying principle of quality improvement 

in colonoscopy is that such quality indicators must be 

recorded and monitored during examination. 

In Emilia-Romagna, CRC screening started in 2005 and since 

then more than 1000000 people have been involved, about one 

third of the adult population. Among people that were positive 

at the Faecal Occult Blood Test, 79% underwent colonoscopy. 

During endoscopic examination, 16% of patients were 

diagnosed with non-advanced adenomas, 32% with advanced 

adenomas and in 6% of cases CRC was found. 

Finally, although recent trend confirm increasing rates of patients 

applying to the screening programs and distribution of 

colorectal cancer stages has shifted towards earlier stages, in 

the near term, even greater incidence and mortality reductions 

could be achieved if a greater proportion of adults received 

regular CRC screening. 

references 

1 Levin B, Lieberman DA, McFarland B, et al. Screening and surveillance 

for the early detection of colorectal cancer and adenomatous 

polyps, 2008: a joint guideline from the American Cancer Society, the 

US Multi-Society Task Force on Colorectal Cancer, and the American 

College of Radiology. CA Cancer J Clin 2008;58:130-60. 

2 Winawer SJ, Zauber AG, Ho MN, et al. Prevention of colorectal 

cancer by colonoscopic polypectomy. The National Polyp Study Workgroup. 

N Engl J Med 1993;329:1977-81. 

3 Rex DK, Cutler CS, Lemmel GT, et al. Colonoscopic miss rates of adenomas 

determined by back-to-back colonoscopies. Gastroenterology 

1997;112:24-8. 

4 Bressler B, Paszat LF, Vinden C, et al. Colonoscopic miss rates for 

right-sided colon cancer: a population-based analysis. Gastroenterology 

2004;127:452-6. 

5 Rex DK, Eid E. Considerations regarding the present and future roles 

of colonoscopy in colorectal cancer prevention. Clin Gastroenterol 

Hepatol 2008;6:506-14. 

6 Lieberman D, Nadel M, Smith RA, et al. Standardized colonoscopy reporting 

and data system: report of the Quality Assurance Task Group 

of the National Colorectal Cancer Roundtable. Gastrointest Endosc 

2007;65:757-66. 

Histology quality assurance of neoplastic 

colorectal lesions 

P. Cassoni, A. Cassenti, P. Cardone, I. Castellano, M. Risio * 

Department of Biological Sciences and Human Oncology, University 

of Turin; * IRCC, Candiolo, Italy 

The diagnostic agreement on the neoplastic lesions of the colorectum 

may be affected by several factors, including lack of 

standardization in terminology, and objective diagnostic “grey 

areas”. The recent publication of the European guidelines for 

pathology quality assurance in colorectal cancer screening 

represented a robust attempt in limiting diagnostic discordance, 

and outlined the major criteria which should be adopted 

in order to achieve a good diagnostic agreement in key

lectures 

features crucial for screening patient management, such as, 

among others, grading of dysplasia, villousness, recognition 

of invasion. In fact, these are controversial histology features 

not only in the field of screen-detected adenomatous polyps, 

but in general account for a limited inter and intra-observer 

reproducibility within the routinary histopathology diagnoses 

of colorectal lesions. 

Moreover, in colorectal carcinomas, some recently introduced 

additional diagnostic criteria (i.e. evaluation of tumour budding, 

or definition of the entity of regression in chemo-radiotreated 

advanced rectum cancers) requires to pathologists 

a continuous resetting of previously acquired diagnostic 

categories. When dealing with either controversial or newly 

introduced entities, a satisfactory diagnostic standard can be 

achieved by verifying the diagnostic concordance to a second 

opinion. We recently verified, in a large series of screendetected 

polyps, that telepathology can be a reliable tool to 

diffuse microscopic images in the context of quality control 

efforts in screening, given its cost-effectiveness advantage 

in regard to preparation, distribution, and circulation of glass 

slides. In addition, improvement in image digitization technology 

have led to significant progress in manageability, and actually 

pathologists can interact with the slide image acquired 

on the pc through digital virtual microscopy, which organizes 

the acquisition process scanning the entire histologic slide at 

the selected resolutions, and provides the observer, through 

real-time image compression, with either the actual overview 

image and series of microscopic images derived from zooming 

of well defined histologic areas. The concordance between 

optic and virtual microscopy on the screen-detected cases was 

good (k-statistics = 0,8), and therefore the digital approach 

could be considerate an accurate tool for histology quality 

assurance in this context and deserves consideration in the 

validation of controversial and/or new diagnostic topics in the 

field of colorectal tumors. 

Morphological and immunohistochemical 

factors with prognostic and predictive roles 

in colorectal carcinoma 

L. Terracciano 

Department of Patology, University Hospital, Basel, Switzerland 

Colorectal cancer (CRC) is one of the most common malignancies 

in the Western world. Despite improvements in 

surgical techniques, dosing and scheduling of adjuvant and 

neoadjuvant therapy, the 5 year survival rate for patients with 

CRC decreases significantly from 93.2% to8.1% with tumour 

progression. The TNM stage remains the “gold standard” 

endomyocardial biopsy 

O. Leone 

Azienda Ospedaliero-Universitaria “S. Orsola-Malpighi”, Anatomia 

ed Istologia Patologica, Bologna, Italy 

The endomyocardial biopsy (EMB) is now routinely used in 

diagnostic strategies for cardiac diseases in the main Centres 

Cardiac pathology 

Moderators: G. Thiene (Padova), P. Gallo (Roma) 

165 

prognostic factor although patients within the same stage can 

demonstrate considerable variation in terms of prognosis. 

TNM stage is also used to help guide the clinical management 

of CRC. Patients with stage III disease may be candidates for 

postoperative chemotherapy, while those with stage II typically 

undergo surgery only. In addition to TNM stage, several 

other tumour related features have been identified as essential 

or important prognostic factors. Venous and lymphatic invasion 

represent a crucial step in the formation of micrometastases 

and eventually macroscopic tumour growth at a secondary 

site. Tumour budding is associated with an infiltrative tumour 

border, and shown to be an independent risk factor of local 

spread, lymph node and distant metastasis, recurrence and 

worse survival following curative surgery. Molecular characterisation 

is expected to improve the identification of patients 

with more aggressive tumours there by leading to individualised 

treatment protocols. 

Despite promising results using genotyping, mutation analysis 

and allelic imbalance, the applicability of these methods to 

routine practice is likely to have limited impact. By contrast, 

immunohistochemistry is frequently employed as a routine 

diagnostic test and is relatively inexpensive. Nevertheless, 

immuno-histochemical markers have yet to find routine application 

as prognostic factors in CRC in which TNM stage and 

other morphologically defined features continue to provide 

the clinical gold standard. 

Promising studies on potential prognostic markers are often 

followed up by subsequent reports contradicting these initial 

results. Several sources of discrepancy between different 

reports have been acknowledged including methodological 

differences, poor study design, non-standardised assays 

which lack reproducibility and inappropriate or misleading 

statistical analyses often performed on underpowered patient 

samples that are too small to enable meaningful conclusions 

to be drawn. 

The wide range of scoring methods employed to evaluate immunoreactivity 

as well as the use of pre-determined and often 

unvalidated or unjustified cut-off scores for tumour marker 

“positivity” is a major contributor to the conflicting results 

reported in the literature on the same tumour marker. We have 

recently proposed the use of receiver operating characteristic 

(ROC) curve analysis as an alternative method for determining 

the threshold values for tumour marker “positivity” and 

have applied this method to several well-established and novel 

tumour markers in CRC for a range of clinical endpoints. 

Recent data, produced by our group as by other Authors, 

regarding immunohistochemical biomarkers, as RHAMM, 

TOP-K, PTEN, RKIP, seems to indicate them as new promising 

prognostic markers in CRC. 

specializing in diagnosis and treatment of cardiac failure, 

although there is a considerable and debatable diversity in 

recourse to this technique, for two main reasons: 

• the shortage of specifically trained cardiovascular pathologists 

1 , able to deal appropriately with cardiac disease diagnostics 

and to apply suitable protocols and diagnostic criteria to 

obtain all necessary information in these complex diseases;

166 

• the absence in many Centers of a team approach integrating 

the competences of pathologists and clinicians. 

Important prerequisited in using appropriately EMB 2 are: 

• perfoming EMB in Centers with a high volume of cases and 

expert operators, in order to minimize procedural risks; 

• improving pathological diagnostic reproducibility and reducing 

the percentage of nonspecific pathological diagnoses, 

referring patients to Centers with an expert cardiovascular 

pathologist and making use of adequate protocols and 

standardized diagnostic criteria; 

• optimizing EMB diagnostic sensitivity limits in multi-microfocal 

diseases, in common with all other non-targeted 

bioptic techniques, applying when warranted imaging techniques 

able to focus on the sampling site. 

Since the 1970’s, when EMB was beginning to come into 

diagnostic use for heart transplanted patients, the indications 

for diagnostic EMB have become increasingly more targeted 

and the protocols more elaborate resulting in an increased 

potential for information. 

Recently, in evidence of the renewed interest in EMB, major 

Guidelines have been published: 

• the joint clinical Guidelines of the American Heart Association, 

the American College of Cardiology and the Europen 

Society of Cardiology 3 ; 

• the Position Paper on Endomyocardial biopsy promoted 

by the “Association for Italian Cardiovascular Pathology”, 

a jointly document produced by Italian cardiovascular 

pathologists and the representatives of the main Italian 

cardiologic scientific societies 2 . Part II of the document 

specifically addresses everyday diagnostic practice, dealing 

with the diagnostic role, particular technical notes, 

protocols and diagnostic criteria for each cardiac disease 

requiring EMB. 

The principal clinical conditions that require EMB are cardiac 

failure 4 5 , rhythm disorders 6 , cardiac masses 7 and heart 

transplantation 8 . 

Here let us confine ourselves to cardiomyopathies 9 10 , the specific 

topic of the Symposium, whose complexity is very much 

stresses by the most recent classifications. 

The diagnostic iter in cardiomyopathies starts when a cardiologist 

identifies some clinical, functional and morphological 

phenotypes, frequently aspecific and potentially caused by 

numerous different diseases, whose course and therapies are 

very different. 

Here the role of pathologist 2 is: 

• to give a definite diagnosis, when possible; 

• to exclude some diseases, guiding forwards a diagnostic 

program; 

• to provide useful information for therapeutic choice and 

prognosis; 

• to contribute to monitoring the clinical evolution of the 

disease and therapeutic program efficacy; 

• to help decrease diagnostic errors 11 ; 

• to guide genetic tests, when appropriate. 

It is noteworthy that, even if the main target of a diagnostic 

test is to identify a specific disease, excluding certain diseases 

is equally important, especially when the clinical picture is 

aspecific. 

The diagnostic potential of EMB in various cardiac diseases 

may be very different, so the level of its diagnostic contribution 

in a specific disease may vary from a definite diagnosis 

to a probable diagnosi, to a possible diagnosis, or even an 

aspecific picture 2 . 

The most significant contribution of EMB is in the diagnosis 

of secondary myocardial diseases 10 , either involving only 


or mainly the heart or as a part of a multi-organ systemic 

disease. 

It is possible to optimize EMB diagnostic accuracy by taking 

certain precautions, which may be considered general rules: 

• careful selection of EMB candidates 4 and the evaluation 

of EMB effects on the overall clinical management of 

the patient. EMB is performed only after the other basic 

clinical-instrumental tests have already excluded various 

diseases and focused more closely on a possible diagnosis. 

An appropriate indication for EMB is the first step towards 

decreasing nonspecific diagnoses. 

• Appropriate EMB timing and adequate bioptic sampling 12 

with multiple specimens, from different sites 13 (possibly 

guided by imaging techniques) in various cardiac diseases. 

• The knowledge of diagnostic potential in various cardiac 

diseases. 

• The use of protocols in which the traditional histological 

examination should be supported by other tissue investigation 

techniques (histochemical, histoenzymatic, immunohistochemical, 

molecular, ultrastructural), opportunely 

selected on the basis of the histological picture or of clinical 

suspicion. 

By way of example, I will describe the EMB diagnostic role 

and the tissue investigations required to increase information 

in some cardiomyopathies. 

Inflammatory cardiomyopathies 2 . EMB, integrating the information 

from the histological picture, immunohistochemical 

tests, molecular tests checking of possible viral genomes 14 , 

may rapidly provide: 

• a definite diagnosis of myocardial involvement; 

• the etiology ot the disease in many cases; 

• the degree of activity of the disease; 

• monitoring of the disease course and the efficacy of therapy; 

• cardiac localization in inflammatory systemic autoimmune 

diseases. 

Appropriate EMB timing and adequate sampling are essential. 

Amyloidotic Cardiomyopathy 2 . EMB is the only test able to 

reach a definite diagnosis of myocardial involvement. 

Moreover, when it is included in a complete clinical-instrumental 

program, it may: 

• contribute to etiological diagnosis using immunohistochemical 

tests on both histological and ultrastructural specimens, 

to identify the main fibrillar component; 

• provide further morphological data as to location, amount 

and type of distribution of deposits, myocardial damage and 

any associated inflammatory reactions; 

• guide genetic analysis in familial forms. 

Definite diagnosis of cardiac involvement and identification 

of type of amyloidosis is essential for therapy. 

Arrythmogenic right ventricle cardiomyopathy 2 . EMB is a 

major diagnostic standard in the score system for the diagnosis 

of ARVC and it may provide: 

• probable diagnosis of cardiac involvement showing myocardial 

atrophy with fibrosis or fibro-fatty replacement and 

differential diagnosis with myocarditis, sarcoidosis, dilated 

cardiomyopathy and idiopathic forms; 

• evaluation of the extent of myocite morphologic compromise. 

Diagnostic accuracy increases if the site of bioptic samples is 

selected using imaging- or electroanatomic voltage mappingguided 

techniques 

Cardiomyopathies in storage diseases 2 . (Glycogenoses, 

Anderson-Fabry disease, Desmin related cardiomyopathy).

lectures 

EMB with the help of ultrastructural and immunohistochemical 

tests may: 

• provide a definite diagnosis of myocardial involvement; 

• grade the extent of cardiac disease and, in Fabry’s disease, 

check the effects of enzymatic substitutive therapy on intramyocite 

accumulation; 

• raise diagnostic suspicions in absence of a clinical suspicion 

or in cases with generic clinic diagnosis of hypertrophic 

cardiomyopathy, suggesting subsequent molecular and 

biochemical tests; 

• guide genetic tests. 

Hemochromatosis/hemosiderosis 2 . EMB using only Perls’ 

staining may: 

• provide a definite diagnosis of myocardial involvement; 

• grade myocardial involvement; 

• evaluate the extent of morphologic myocyte involvement. 

Dystrophin-related cardiomyopathies, Lamin-related cardiomyopathies, 

Emery-Dreifus disease 2 . In these diseases, 

the role of immunohistochemistry has recently been gaining 

ground in: 

• providing a definite diagnosis of cardiac involvement in 

Duchenne MD (a disease whose diagnosis does not normally 

require a EMB) and a definite/probable diagnosis in 

Becker MD and in X-linked cardiomyopathy X21.2 MIM 

302045. In these latter forms EMB may be the sole diagnostic 

tool able to raise the question of a dystrophin gene linked 

disease; 

• raising diagnostic suspicion in laminopathies; 

• guiding genetic analysis; 

In all cases, EMB can exclude other diseases. 

references 

1 Thiene G, Veinot JP, Angelini A, et al. AECVP and SCVP 2009 recommendations 

for training in cardiovascular pathology. Association 

for European Cardiovascular Pathology and Society for cardiovascular 

Pathology Task Force on Training in Cardiovascular Pathology. 

Cardiovasc Pathol 2010;19:129-35. 

2 Leone O, Rapezzi C, Sinagra G, et al. Documento di consenso sulla 

biopsia endomiocardica promosso dall’Associazione per la Patologia 

Cardiovascolare Italiana. G Ital Cardiol 2009;10(Suppl 1-9):1S-50. 

3 Cooper LT, Baughman KL, Feldman AM, et al. The Role of Endomyocardial 

Biopsy in the Management of Cardiovascular Disease. A 

Scientific Statement from the American Heart Association, the American 

College of Cardiology, and the European Society of Cardiology. 

Circulation 2007;116:2216-33. 

4 Perkan A, Di Lenarda A, Sinagra G. Dilated cardiomyopathy: indication 

and role of endomyocardial biopsy. Ital Heart J Suppl 2002;3:419- 

25. 

5 Ardehali H, Qasim A, Cappola T, et al. Endomyocardial biopsy plays 

a role in diagnosing patients with unexplained cardiomyopathy. Am 

Heart J 2004;147:919-23. 

6 Basso C, Corrado D, Marcus FL, et al. Arrhythmogenic right ventricular 

cardiomyopathy. Lancet 2009;373:1289-300. 

7 Flipse TR, Tazelaar HD, Holmes DR Jr. Diagnosis of malignant cardiac 

disease by endomyocardial biopsy. Mayo Clin Proc 1990;65:1415- 

22. 

8 Stewart S, Winters GL, Fishbein MC, et al. Revision of the 1990 

Working Formulation for the standardization of nomenclature in the 

diagnosis of heart rejection. J Heart Lung Transplant 2005;24:1710- 

20. 

9 Veinot JP. Diagnostic endomyocardial biopsy pathology–general biopsy 

considerations, and its use for myocarditis and cardiomyopathy: 

a review. Can J Cardiol 2002;18:55-65. 

10 Veinot JP. Diagnostic endomyocardial biopsy pathology: secondary 

myocardial diseases and other clinical indications–a review. Can J 

Cardiol 2002;18:287-96. 

11 Luk A, Metawee M, Ahn E, et al. Do clinical diagnoses correlate with 

pathological diagnoses in cardiac transplant patients? The importance 

of endomyocardial biopsy. Can J Cardiol 2009;25:e48-54. 

12 Billingham ME. Acute myocarditis: is sampling error a controindication 

for diagnostic biopsies? J Am Coll Cardiol 1989;14:921-2. 

167 

13 Burke AP, Farb A, Robinowitz M, et al. Serial sectioning and multiple 

level examination of endomyocardial biopsies for the diagnosis of 

myocarditis. Mod Pathol 1991;4:690-3. 

14 Calabrese F, Thiene G. Myocarditis and inflammatory cardiomyopathy: 

microbiological and molecular biological aspects. Cardiovasc 

Res 2003;60:11-25. 

Molecular diagnosis of myocarditis 

A. Angelini 

Dept of Medical-Diagnostic Sciences and Special Therapies, University 

of Padua, Padua, Italy 

Myocarditis is a non-ischemic inflammatory disease of the 

myocardium associated with cardiac dysfunction 1 . It most often 

results from infectious agents, hypersensitivity responses, 

or immune related injury. In spite of the development of 

various diagnostic modalities, early and definite diagnosis of 

myocarditis still depends on the detection of inflammatory 

infiltrates in endomyocardial biopsy specimens according to 

Dallas criteria 2 . Routine application of immunohistochemestry 

(for characterization of inflammatory cell infiltration) and 

molecular analysis (PCR for identification of infective agents) 

have become an essential part of diagnostic armamentarium 

for a more precise biopsy report 3-6 . 

Three different forms of the diseases can be recognized: 

a) non-infectious disease due to allergic/immune causes (giant 

cell-myocarditis; rheumatic and eosinophilic myocardities; 

forms associated with immune systemic diseases; virusnegative 

myocardities with or without circulating anti-heart 

antibodies), b) infectious (bacterial, protozoan and viral) 

and c) myocardities in which the immune mechanism starts 

or is supported by an infection. Unfortunately, the clinical 

diagnosis of myocarditis still remains a challenge owing to 

the non-specific pattern of the clinical presentation and to 

the lack of universally accepted and standardized diagnostic 

criteria. Since the spectrum of clinical presentation is broad, 

including asymptomatic electrocardiographic abnormalities 

reported during enterovirus epidemic, vague symptoms of 

flu-like syndrome, congestive heart failure of recent onset, 

cardiogenic shock and sudden death, many false-positive 

and false-negative clinical diagnoses may be expected 7 . 

EMB,despite the low sensitivity due to the frequent sampling 

errors and to the lack of quantitative diagnostic criteria still 

represents the main diagnostic tool for myocarditis. Viral 

myocarditis usually lacks specific cytopathic effects and can 

cause different magnitude in the inflammatory response both 

as consequence of the virulence of the causative agents as 

well as the host status. As suggested in the consensus document 

8 on EMB published by the Associazione per la Patologia 

Cardiovascolare Italiana endomyocardial biopsy (EMB) 

still represent the gold standard for the diagnosis even though 

myocardial scintigraphy and MRI may help in diagnosis. 

EMB is mandatory: 

• to reach a definite diagnosis of myocardial involvement; 

• to contribute to etiological diagnosis (infectious microorganism, 

immune aetiology); 

• to indicate the degree of activity of the disease. 

Pathological diagnosis. In diagnosing myocarditis important 

points include: 

• timing of EMB in relation to the onset of symptoms; 

• number and size of bioptic fragments; 

• contemporary checks for heart auto-antibodies in serum 7 . 

Histological examination: In hematoxylin-eosin stained sections 

1) inflammatory infiltrate, 2) myocellular damage and 

3) fibrosis.

168 

Further elements to look for are: 

a) inflammatory infiltrate types (lymphocytic, polymorphous, 

granulomatous, eosinophilic) and extention using semiquantitative 

or quantitative evaluation; 

b) myocellular damage (not only necrosis or myocytolysis but 

also other alterations, such as cytoplasmic vacuolization, 

apoptosis, and atrophy); 

c) pattern of fibrosis (interstitial, perivascular, subendocardial) 

and its extent using semi-quantitative or quantitative evaluation. 

Histo-morphological stains: 

• Azan-Mallory trichrome is useful to highlight and quantify 

fibrosis. 

• Weigert-Van Gieson, which highlights elastic fibres in 

brown/black, and allows for evaluation of vessel wall structure 

and endocardial fibroelastosis. 

Immunohistochemical tests: immunohistochemical analysis 

represents a fundamental corollary to traditional histology 

enabling the characterization of inflammatory infiltrate and, 

when this is present in very small quantities, its identification. 

Antibody panel to use: CD45, CD68/PGM1, CD3, CD4, CD8, 

CD20, HLA-ABC, HLA-DR. 

Morphometric quantification: it is desirable to quantify 

inflammatory infiltrate (currently a lymphocytic infiltrate 

> 7 T lymphocytes/mm 2 is considered as pathologic) using 

computerized morphometry on immunohistochemical 

sections stained with anti-CD3 antibody. Morphometry on 

Azan-Mallory trichrome stained sections may allow precise 

quantification of fibrosis. 

Molecular tests and in particular techniques of gene amplification, 

such as polymerase chain reaction (PCR) or nested- 

PCR, because of their high sensitivity, allow the amplification 

of viral DNA or RNA, thus detecting any viral genome 

present in the small samples of EMB tissue. Nowadays, 

sequence analysis and the identification of replicating virus 

forms are increasingly utilized to characterize infective agents 

precisely 5-9 . If myocarditis is clinically suspected, at least the 

following most frequent cardiotropic virus genomes must be 

checked for in myocardial tissue: enterovirus, adenovirus, 

cytomegalovirus, Epstein Barr virus, herpes simplex virus, Influenza 

A and B viruses; B19 parvovirus and C hepatitis virus. 

In the setting of positive PCR results blood samples collected 

at the time of the biopsy should be tested: if positivity for the 

same virus is present in both myocardial tissue and blood, it is 

necessary to quantify its load with quantitative PCR analysis 

to exclude any haematic contamination of the myocardial 

specimen. Gene sequencing is a more sophisticated technique 

allowing the characterization of the infective agents as well as 

its virulence and cardiotropism. Different serotypes can bear 

a different virulence and cardiotropism and guide prognosis 

and therapeutic interventions. However the detection of viral 

genome does not necessarily imply a direct pathogenetic role, 

since it could be an innocent by stander. Also a negative PCR 

does not exclude viral disease. Final diagnosis of myocarditis 

must be the results of an integrated clinical, instrumental, 

morphological and molecular approach. 

Key points 

• The accurate diagnosis of myocarditis requires a representative 

number of specimens to be subjected to complete 

traditional histopathological, and immunohistochemical 

(lymphocyte types, HLA, C3-C4) and molecular virological 

(a study of the presence/persistence of RNA and DNA virus 

genome) analysis. 


• The application of immunohistochemical methodologies 

(which allows the identification of inflammatory infiltrates, 

their more adequate quantification and the evaluation of 

myocardial expression of immunological activation markers) 

increases interpretative capacity, especially in cases 

of prevalently autoimmune mechanism responsible for 

“chronic myocarditis”. 

• It is mandatory to apply molecular tests, especially gene 

amplification techniques such as the Polymerase Chain 

Reaction (PCR), quantitative (real time-PCR) or qualitative 

(nested-PCR), which, nowadays, because of their high 

sensitivity, allow the identification even of a small number 

of copies of any viral genome present in the EMB. 

• The exclusion of a viral aetiology is an essential requirement 

in considering a myocarditis as immuno-mediated 

(both antibody- and cell- mediated) and choosing the most 

appropriate therapeutic strategy 8 9 . 

• Etiological characterization is important also in assessing 

prognosis. 

references 

1 Maron BJ, Towbin JA, Thiene G, et al. Contemporary definitions and 

classification of the cardiomyopathies. An American Heart Association 

Scientific statement from the Council on clinical cardiology, heart 

failure and transplantation Committee; quality of care and outcomes 

research and functional genomics and translational biology interdisciplinary 

working Groups; and Council on epidemiology and prevention. 

Circulation 2006;113:1807-16. 

2 Aretz HT, Billingham ME, Edwards WD, et al. Myocarditis: a histopathological 

definition and classification. Am J Cardiovasc Pathol 

1987;1:3-14. 

3 Angelini A, Crosato M, Boffa GM, et al. Active vs borderline myocarditis: 

clinicopathological correlates and prognostic implications. 

Heart 2002;87:210-5. 

4 Calabrese F, Rigo E, Milanesi O, et al. Molecular diagnosis of myocarditis 

and dilated cardiomyopathy in children. Clinico-pathologic 

features and prognostic implications. Diagn Mol Pathol 2002;11:212- 

21. 

5 Calabrese F, Thiene G. Myocarditis and inflammatory cardiomyopathy: 

microbiological and molecular biological aspects. Cardiovasc 

Res 2003;60:11-25. 

6 Calabrese F, Carturan E, Thiene G. Cardiac infections: focus on molecular 

diagnosis. Cardiovascular Pathology 2010;19:171-182. 

7 Caforio AL, Calabrese F, Angelini A et al. A prospective study of 

biopsy-proven myocarditis: prognostic relevance of clinical and aetiopathogenetic 

features at diagnosis. Eur Heart J 2007;28:1326-33. 

8 Documento di consenso sulla biopsia endomiocardica promosso 

dall’Associazione per la Patologia Cardiovascolare Italiana. G. Ital 

Cardiol 2009;10(Suppl):1-9. 

9 Kuhl U, Pauschinger M, Noutsias M, et al. High prevalence of 

viral genomes and multiple viral infections in the myocardium of 

adults with “idiopathic” left ventricular dysfunction. Circulation 

2005;111(7):887-93. 

Diagnostic and terapeuetic work-up for 

cardiovascular diseases: the role of pathologists 

E. Arbustini, M. Grasso, M. Diegoli, A. Agozzino, M. Concardi 

Centre for Inherited Cardiovascular Diseases, Transplant Research 

Area, IRCCS Foundation Policlinico San Matteo, Pavia, Italy 

Background. The progression of knowledge on the pathologic 

basis of cardiovascular diseases and the development of biotechnological 

tools for pathological and molecular studies are 

progressively increasing the number of specific vs. descriptive 

diagnoses. Disease-specific diagnostic work-up tailored on 

phenotypes (percorsi diagnostico-terapeutici assistanziali: 

PDTA) constitute the tool; in this scenario pathology may play 

a fundamental role for diagnosis, prognostic stratification,

lectures 

treatment guide, and evaluation of the benefits of treatments. 

This short review concentrates on cardiovascular diseases 

and discusses a few examples, but similar considerations can 

extend to other disease settings. 

Methods. To evaluate the clinical impact of disease-specific 

work-up in cardiovascular diseases, we generated an 

outpatient centre where cardiovascular pathology, clinics and 

genetics are integrated and other multidisciplinary contributions 

are herein organised according to the type of disease. 

The centre policy is to offer patients and families tailored care 

strategies, starting from the specific diagnosis, contrapposed 

to the concept of descriptive diagnosis. The activity started 

in early 80ies with cardiovascular pathology; then it progressively 

incorporated genetics and clinics and finally evolved to 

the organisation of multidisciplinary teams of specialists that 

are now active on precise disease-specific protocols (PDTA). 

The centre participates to national and international quality 

controls and is equipped with instruments that can provide 

accurate, precise, and high-quality data for patient care and 

for research; it consists of three integrated units: 1) the cardiovascular 

pathology lab that is equipped with automated 

instruments for tissue and blood processing, light, electron 

and confocal microscopes, cell culture and cell sorter facilities 

and slide scanner; 2) the molecular genetic lab that is fully 

equipped for genomics and transcriptomics/gene expression 

profiling, as well as for very basic protein studies (Western 

Blot) for immunohostichemistry/immunoblotting comparative 

studies; 3) the clinical out-patient centre that permanently 

hosts cardiologists and geneticists, as well as nurses and administrative 

personnel to organise and run the disease specific 

diagnostic work-up for patients and families; on scheduled 

programs, specialists of disciplines that are included in the 

different diagnostic work-up have regular access to the centre. 

Personnel active in the three units includes 24 between 

pathologists, cardiologists, geneticists, engineers, biologists, 

technicians, nurses and a person responsible for international 

liaisons. 

The pathology provided the basic knowledge and tools for 

the entire organisation: from pathology to clinics and genetics 

first (1984-2000) and now from clinics to pathology and genetics 

(2001-2010). Each disease-specific diagnostic work-up 

is organised as follows: 1. The centre is contacted by medical 

doctors, specialists of different disciplines, according to 

the disease, or directly by patients or voluntary associations. 

2. All clinical/pathological reports are evaluated before the 

access of patients to the centre and tailored work-up is then 

planned, including clinical multidisciplinary evaluations, 

biopsy procedures, genetic testing and counselling (when 

necessary). 

Results. The Centre now offers multidisciplinary diagnostic 

work-up for acquired (inflammatory and autoimmune) and 

heritable cardiovascular diseases manifesting with vascular 

life-threatening events, heart failure and arrhythmias. Major 

groups of diseases are: genetic aneurismal syndromes [Marfan 

Syndrome, Loeys-Dietz Syndromes, Elhers Danlos Syndrome 

type IV, Thoracic Aortic Aneurysm and Dissections (TAAD), 

familial and non-familial Bicuspid Aorta (BAV)], heritable 

an acquired cardiomyopathies [hypertrophic sarcomeric and 

non-sarcomeric], dilated, restrictive and arrhythmogenic right 

ventricular cardiomyopathies and phenocopies), complex syndromes 

with cardiovascular involvement [such as Noonan, Alstrom, 

Holt-Oram and Barth syndromes, MELAS, MERFF], 

lysosomal diseases such as Danon Disease, Pompe Disease, 

Anderson Fabry Disease] as well as other more rare conditions. 

The centre currently follows more than 2500 families. 

169 

Based on the prevalence of each of the above diseases, more 

than 180.000 individuals are affected in our country. 

Acute myocardial diseases: examples of acute non-ischemic 

myocardial illnesses. Excluding the most prevalent ischemic 

heart disease, a setting in which novel diagnostic and treatments 

dramatically decreased mortality and improved survival, 

acute myocardial diseases also include myocarditis of 

viral origin (PVB19 or EV), idiopathic giant cell myocarditis 

(possible phenotype: ”fulminant myocarditis”), acute rheumatic 

carditis, pheochromocytoma (contraction band necrosis 

without inflammation), acute hypercalcaemia (myocardial 

microcalcifications), neoplastic infiltration [such as thymoma), 

septic emboli, drug toxicity (eosinophilic infiltrates in 

the absence of hypereosinophilc syndromes associated with 

FIP1L1/PDGFR alpha fusion transcript), as well as other 

more rare conditions. In the PDTA for acute heart failure, the 

correct diagnosis of the above rare causes may be life saving. 

The probability of specific pathologic diagnosis depends on 

the participation of the pathologists to the overall multidisciplinary 

evaluation/discussion. Most descriptive diagnoses can 

be done on H&E stain of endomyocardial biopsies (EMB). 

The myocardium however has a limited spectrum of morphologic 

responses to different types of insults; a clinically 

oriented hypothesis may guide the choice of appropriate immunohistochemical 

and molecular studies. 

Chronic myocardial diseases: the example of cardiomyopathies. 

The modern diagnostic strategies for cardiomyopathies 

require that pathologists know the clinical phenotypes, 

the imaging features and the genetic bases of these diseases. 

The intergation of this knowledge is the basis for providing a 

useful contribution of tissue studies in cardiomyopathies. 

Dilated cardiomyopathy (1:2500) (DCM) is a clinical descriptive 

diagnosis; pathology and genetics may provide a specific 

diagnosis. Up to 50% DCM are heritable diseases with genetic 

heterogeneity (up to 35 disease genes). A DCM phenotype 

may result from defects of genes that code for nuclear envelope 

proteins (Lamins and Emerin), desomosome proteins, 

myocyte membrane proteins (Dystrophin and Dystrophin-associated 

glycoproteins), mitochondrial proteins (Tafazzin). 

The appropriate immunostain may provide diagnostic information. 

Genetic test is the gold standard, when available. Both 

pathology and genetics integrate to document the functional 

impact of the mutation on the myocardial tissue. 

Hypertrophic phenotypes (1:500, or more). As DCM, Hypertrophic 

Cardiomyopathy (HCM) is a descriptive diagnosis: it 

simply indicates idiopathic increased left ventricular thickness. 

HCMs are grouped in sarcomeric (13 disease-genes that 

code for sarcomeric proteins) and non-sarcomeric, mostly 

lysosomal diseases such as glycogenosis (ex. Pompe disease), 

sphingolipidoses (ex. Anderson Fabry Disease), Danon 

Disease) or Protein receptor Kinase AG2 (PRKAG2)-related 

cardiomyopathy. The non-sarcomeric group also includes mitochondrial 

DNA-related cardiomyopathes, as well as pathologic 

phenocopies of lysosomal diseases such as cloroquine 

myocardial toxicity. Depending of the available facilities, the 

diagnosis can be obtained by genetic testing; when genetic 

tests are not available, the EMB uniquely contributes to the 

diagnosis. A novel role for EMB is therefore emerging in the 

scenario of non-sarcomeric HCM phenotypes. 

Restrictive cardiomyopathy (RCM) is characterised by functional 

restrictive pattern without hypertrophy (prevalence 

< 1:100,000). There are several genetic, systemic autoimmune, 

neuromuscular and inflammatory diseases that may 

cause a RCM phenotype. The starting point for establishing a 

specific diagnosis is the evaluation of cardiac and non-cardiac

170 

markers associated with RCM and the family phenotype. In 

this context EMB may play a fundamental role for the diagnosis 

of Desminopathies in patients with pure RCM associated 

with AVB. When cataract is also present, CRYAB gene should 

be considered as more likely candidate gene. Troponinopathies 

(defects of troponin T and I) may manifest as pure RCM 

w/o hypertrophy with high risk of sudden death: in this case, 

they are not associated with AVB and/or myopathy: a potentially 

useful maker is the mismatch between hypertrophy 

(minimal, if any) and disarray (present). 

Cardiac Amyloidoses: thickened hearts with restrictive pattern. 

With more than 20 different amyloidogenic proteins 

potentially causing systemic deposits, tissue studies document 

the presence of amyloid deposits in myocardium, valves, 

vessels, epicardial fat and provides the characterisation of 

the amyloidogenic protein by immunoelectron microscopy. 

In systemic amyloidoses, this result can be easily achieved 

by sampling periumbelical fat, but in non-systemic cardiac 

amyloidoses EMB is the only diagnostic option. Pathologists 

are also involved in monitoring treatment effects as well as 

in detecting possible recurrence of amyloid in transplanted 

organs; no imaging tool is equally informative. 

Arrhythmogenic right ventricular cardiomyopathy (ARVC) 

(1:5000). Pathology played a key role in the disease characteri- 

eosinophilic esophagitis:clinical aspects 

M. Marini 

Pediatric Gastrointestinal Endoscopy, University Hospital Santa Maria 

alle Scotte, Siena (Italy) 

Introduction. Eosinophilic esophagitis (EE) is an increasingly 

recognized disorder previously described in children, 

and emerged as a clinical disorder that afflicts adults, presents 

with dysphagia and food impaction and characterized by a 

dense eosinophilic infiltration of the surface lining of the 

esophagus 1-3 . 

EE is a primary esophageal disorder without associated eosinophilic 

infiltration of the stomach or intestine. 

There are many other diseases that can cause eosinophils 

in the tissue of the esophagus, including gastroesophageal 

reflux disease(GERD), parasitic infections, fungal infections, 

inflammatory bowel diseases 

(Crohn), certain cancers, recurrent vomiting,collagen vascular 

disorders, eosinophilic gastroenteritis. and others. These 

diseases need to be ruled out before primary EE can be diagnosed. 

Symptoms. EE has many different presentations. In the 

majority of studies to date, individuals affected by EE have 

been predominantly male children and adolescents; patients 

commonly have difficulty eating, failure to thrive, vomiting, 

epigastric or chest pain, dysphagia, and food impaction 4 5 . 

Adult patients typically have recurrent dysphagia and food 

impactions that are refractory to anti-GERD therapy; in fact, 

recent studies indicate that 10%–50% of adult male patients 

with these symptoms have EE 6 7 . 

Although a fixed stricture could account for the esophageal dysphagia 

and food impaction observed in some patients with EE 8 , 


Gastrointestinal inflammatory diseases 

Moderators: C. Capella (Varese), E. Tavani (Rho) 

sation. ARVC/D typically affects the right ventricle; it is clinically 

characterised by life-threatening ventricular arrhythmias. 

The pathologic markers are fibro-fatty replacement of RV 

myocardium with myocyte degeneration. ARVC/D is a desmosomal 

cardiomyopathy: defects of one of the 5 major proteins 

of the desmosome, plakophillin, plakoglobin, desmoglein, 

desmocollin and desmoplakin cause loss of continuity between 

myocytes caused by failing desmosomes, progressive degeneration 

and death of the myocytes leading to fibro-fatty repair. 

Although genetic testing is now available in tertiary centres 

that perform specific cardiogenetic programs, new directions 

for pathologic diagnosis include immuno-staining with antiplakoglobin 

and gap-junction protein Cx 43 antibodies, that 

should be markedly decreased. Cardiac sarcoidosis is included 

in the list of phenocopies that may mimic ARVC/D, as does 

myocarditis: EMB is the gold standard fro the diagnosis. 

Conclusion. As owners of a systematic approach to the pathologic 

bases of the diseases in general, and cardiovascular in 

particular, pathologists can either coordinate or deeply integrate 

in disease-specific clinical work-ups that represent the 

frontiers of sustainable health programs (the right diagnosis 

and care‡ right patients vs. descriptive diagnoses and care‡ 

all patients). The model is expanding in our country as well as 

throughout Europe. 

evidence is mounting that the esophagus displays impaired 

smooth muscle function, likely from asynchrony of circular and 

longitudinal muscle contraction during swallowing 9 . 

A variety of motor disturbances that are reversible with 

therapy have been reported in patients with EE 10 11 . 

Patients often have personal and family histories of asthma, allergic 

rhinitis,atopic dermatitis, food and drug allergies, eosinophilia, 

elevated serum levels of immunoglobulin E (IgE), and 

positive allergic skin and radio allergo sorbent tests (RAST). 

Although EE was originally recognized in pediatric patients, it 

has similar characteristics(including atopic sensitization) and 

occurrence rates in adults. The disease has also been recognized 

in patients older than 90 years. EE is a chronic disorder 

that has no significant evidence of spontaneous remission, 

even over a 14-year period,but some patients have seasonal 

variations in symptoms, consistent with an etiology related to 

airborne allergen exposure. 

Diagnosis. A number of endoscopic features have been described, 

including strictures (frequently proximal), mucosal 

rings (often multiple), mucosal ulceration, linear furrows, 

small-caliber esophagus, and multiple white papules (eosinophilic 

microabscesses) 12-18 . 

Vasilopoulos have proposed a classification system for EE 

that includes three types. Type 1 is called the early small 

caliber esophagus, type 2 is the advanced small caliber 

esophagus and type 3 is the ringed esophagus. It is not clear 

whether these types represent isolated variants of this disorder 

or whether they are sequential stages in its evolution 19 . 

The whitish exudates seen in EE can be fine, pinpoint and 

scattered and are often mistaken for Candida or debris. The 

exudates actually represent collections of eosinophils, which 

can also appear as mucosal nodules or plaques.

lectures 

The most dramatic endoscopic finding of EE is the appearance 

of a long and deep mucosal tear (with visualization of 

the submucosa and muscle fibers) following passage of a 

dilator. These tears or mucosal rents can also occur simply 

with insertion of the endoscope, particularly when there is an 

unrecognized diffusely narrowed esophagus. 

Endoscopic ultrasound findings in EE include thickening of 

the mucosa, submucosa and muscularis propria 20 . 

Endoscopy is used to obtain biopsy samples from patients 

with proton pump inhibitor (PPI)-refractory upper gastrointestinal 

symptoms. These tissue samples are analyzed by 

microscopy;a minimum of 15 eosinophils/hpf indicates that 

a patient has EE. 

However, it is now recognized that there can be significant 

overlap between GERD and allergic EE. Eosinophil counts 

greater than 100 per hpf can be seen in some patients with 

GERD, even at multiple levels of the esophagus 21 . 

In facts a diagnosis of allergic EE cannot be made until GERD 

is ruled out either by ambulatory pH testing or by an eight 

week trial of a PPI taken twice daily, followed by repeat endoscopy 

with biopsies. 

Recently analysis of transcription profiles has indicate dysregulated 

expression of 1% of the human genome, including overexpression 

of eotaxin-3, is found in samples from patients with 

EE. This gene expression profile can be used to distinguish between 

biopsy specimens from patients with EE, patients with 

reflux esophagitis (RE), and normal individuals (NL). 

In patients successfully treated with dietary modification and/ 

or glucocorticoids, eosinophil numbers in biopsy samples are 

reduced to 1/hpf and the transcriptome more closely resembles 

that of NL, although there are residual gene expression differences 

between patients treated for EE and RE and NL 22 . 

Managment. Optimal treatment for EE has not been defined. 

Management has been better evaluated in children where 

dietary restrictions and treatment with corticosteroids have 

proven effective, but compliance and toxicity, respectively, 

have limited usefulness. Fluticasone propionate (FP) applied 

topically appears to be equally effective and better tolerated 

23 24 . 

Strictures in adults have been managed by dilation. Approaches 

include use of antihistamines, sodium cromoglycate,and 

systemic and topical corticosteroids; andleukotriene receptor 

antagonists. 

Mepolizumab, an anti-interleukin-5 monoclonal antibody, recently 

has been reported to have histologic and clinical benefit 

in an adult case of EE. 

Esophageal dilation has been associated with deep mucosal 

tears, severe pain, and perforation 25 . 

Experience with (FP), an inhaled corticosteroid routinely used 

in the management of asthma, has shown benefit in a pediatric 

population with EE and, more recently, in adults 26-28 . 

FP has not always been efficacious, particularly in the allergic 

EE subgroup. 

The likelihood of response to anti-GER therapy decreases with 

increased severity of eosinophilic inflammation of the esophageal 

mucosa. The eosinophil density within the esophageal 

mucosa required for the diagnosis of AEE is often defined by 

R15 eosinophils per high power field (eos/hpf). 

An eosinophil density of %5 eos/hpf is thought to represent 

gastric-acid–mediated injury. 

Consequently, the diagnosis and treatment of patients with 

moderate tissue eosinophilia, ie, 6 to 14 eos/hpf, is unclear. 

Because AEE may take months to years to evolve, moderate 

esophageal eosinophilia may represent a mild or evolving 

form of EE 29 . 

171 

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172 

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Clin GastroenterolHepatol 2004;2:568-75. 

le esofagiti eosinofile: aspetti istopatologici 

C. Vindigni 

Division of Pathological Anatomy, AOUS, Siena, Italy 

Eosinophilic esophagitis (EE) is a primary disease of the 

esophagus characterized by esophageal and/or upper gastrointestinal 

tract symptoms and by dense esophageal eosinophilia 

associated with a normal gastric and duodenal mucosa and absence 

of pathologic gastroesophageal reflux disease (GERD) 

as evidenced by a normal pH monitoring study or lack of 

response to high dose PPI medication 1 . 

EE is more predominant in males, young adults and children, 

and is often associated with a history of allergic disease 2 3 . 

Many reports suggest familial clustering of the disease but it 

is difficult to determine whether this represents genetic predisposition 

or similar environmental exposure 4 . 

Recent studies suggest an increase in the prevalence of EE, 

but it is unclear whether this is due to a true escalation in 

incidence or to a better awareness of the disease by both gastroenterologists 

and pathologists 5 . 

The diagnosis of EE is based on the clinical presentation, 

endoscopic features and histopathological findings. Failure to 

respond to anti-reflux therapy and dense eosinophilic infiltration 

in oesophageal biopsies are essential. 

At endoscopy, several mucosal abnormalities have been identified, 

including friability, white specks, whitish exudates, 

“crepe paper mucosa”, narrow caliber esophagus, longitudinal 

furrows, and transient or fixed rings; some studies have also 

reported a normal mucosa 6 . It has been reported that multiple 

biopsy specimens from different areas, including the distal, 

mid and proximal oesophagus, improve the diagnostic ability 

because of the heterogeneous distribution of eosinophilic infiltration. 

Biopsies should also be obtained from the stomach 

and duodenum to rule out eosinophilic gastroenteritis 7 . 

The diagnostic criterion for the diagnosis of EE is an intense 

eosinophil infiltration in oesophageal squamous epithelium 

but the number and the method used varies among studies. It 

has been recommended that intraepithelial eosinophils should 

be counted in the most intensely inflamed HPF of the biopsy 

at x400 magnification 7 . A consensus opinion as to the histological 

diagnostic criteria is still lacking but most of the pathologists 

believe that the presence of > 20 eosinophils in one 

HPF or > 15 eosinophils in multiple HPFs is diagnostic for 

EE 5 . This is based on studies that showed that GERD is often 

associated with less than seven eosinophils per HPF 8 . Other 

associated histopathologic features have been observed in EE 

as eosinophils degranulation, eosinophilic microabscesses, 

preferential superficial distribution of eosinophilic inflammation, 

basal zone hyperplasia and papillary lengthening, lamina 

propria fibrosis. These features may be helpful to the pathologist 

for the diagnosis of EE and should be included in the pathology 

report in addition to the number of eosinophils 7 . 

The relationship between GERD and EE is not clear, and it 

must be kept in mind that these entities may sometimes coexis 

9 . Absolute eosinophil counts cannot be used to establish 

a definitive diagnosis of EE. Consensus recommendations 

state that the diagnosis of EE should only be made in the 

proper clinical context and therefore close communication 


between the pathologist and gastroenterologist is necessary 

for the diagnosis 10 . 

Recognition of EE and the differential diagnosis from GERD 

is critical for appropiate patient care. 

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Diagnosis of infectiuos enterocolitis 

S. Antinori, C. Parravicini, A.L. Ridolfo, L. Fociani, 

G.L.Vago 

Department of Clinical Sciences “L. Sacco”, Section of Infectious Diseases 

and Immunopathology, University of Milano, Italy 

Background. Infectious eneterocolitis (IE) is frequently a 

short course and self-limiting disease in otherwise healthy 

adults; however, severe and potential life threatening course 

may be observed in particular in infants, elderly and immunocompromised 

individuals. IE are most often caused by 

ingestion of contaminated food or water. In addiction, it may 

arise from reactivation of quiescent infectious agents (i.e., cytomegalovirus, 

CMV), in particular in immunocompromised 

hosts. 

The characteristics of the illness and the epidemiologic setting 

are essential for differential diagnosis and evaluation 1 . Moreover, 

there is general agreement that a distinct diagnostic approach 

is required in three different epidemiologic setting of 

IE, i.e., community-acquired (including traveller’s diarrhea), 

nosocomial and in immunocompromised hosts 2 . 

Methods. Four case vignettes have been used to addressed 

the challenges in diagnosing severe IE in three different epidemiological 

scenarios. 

Results. 

Case 1. A 23 years old women presented with fever and chills 

lasting two weeks after returning from a vacation in Bali. On 

admission she was febrile (39°C) and physical examination 

revealed mild hepatomegaly. Blood analysis showed anaemia 

(Hb 9.6 g/dl, MCV 69 fl), leukocytosis (WBC 12.000/µl), 

and elevated liver enzymes (ALT 104 U/l, AST 91 U/l). 

After 3 days she complained of diffuse abdominal pain and 

bloody diarrhea. Abdominal radiography and ultrasound

lectures 

were negative. Cultures of blood and stool for Salmonella, 

Shigella, Campylobacter and Clostridium difficile toxins were 

negative. She underwent a colonoscopy which showed several 

discrete purulent ulcers along the sigmoid colon and rectum, 

and disseminated haemorrhagic suffusions with an intense 

hyperaemic and oedematous mucosa in the transversal and 

descending colon. Multiple colonic biopsies showed marked 

chronic inflammation of the lamina propria extending into 

the submucosa with scattered crypt abscesses, erosions, and 

several CMV inclusions in endothelial cells. CMV pp65antigenemia 

(11 positive cells/slide) and CMV serology (IgM 

11.9, IgG 0.9) were compatible with acute CMV infection. 

Intravenous therapy with ganciclovir was started with a rapid 

clinical response. 

Case 2. A 71-year-old man with underlying diabetes mellitus, 

hypertension and hypertriglyceridemia was admitted to our 

hospital with bloody diarrhea and a three-day history of severe 

watery diarrhea, vomiting and cramping abdominal pain. The 

patient had just returned from a trip in Madagascar where he 

had eaten raw meat of zebu and had drunk tap water. 

On physical examination he was alert, dehydrated with sick 

appearance, and showed abdominal distention with tenderness 

and guarding to deep palpation in the lower quadrants. Laboratory 

studies revealed leukocytosis (10,720/µl), hyperglicemia 

(284 mg/dl), and acute renal failure (creatinine 6.5 mg/ 

dl) with hyperkaliemia (7.3 mmol/l) and severe metabolic 

acidosis (pH 7.16, HCO3 - 9.3 mmol/l, lactate 14.2 mmol/l). 

The patient underwent hemodyalisis, parenteral hydratation 

and correction of acidosis and was put on empirical antibiotic 

therapy (levofloxacin and metronidazole). Stool culture 

for Salmonella, Shigella, Campylobatcer, Yersinia as well 

as C. difficile toxin resulted negative; examination of fresh 

and stained samples of stool for ova and parasite was negative. 

An abdominal CT demonstrated ascites and colon-wall 

thickening. Due to the development of toxic megacolon the 

patient underwent explorative laparotomy which showed diffuse 

ascitic fluid and thickening of intestinal loops. A biopsy 

of rectal mucosa showed an acute proctosigmoiditis without 

demonstrable microorganisms. A blood culture obtained at 

admission grew Shigella sonnei that was sensitive to the ongoing 

antimicrobial therapy. The patient was discharged after 

one month hospital stay. 

Case 3. A 83-year-old man with multiple comorbidities 

(gastroresection, COPD, coronary artery disease, lower limb 

Kaposi’s sarcoma, psoriasis with psoriatic arthritis and hypertension) 

was admitted to our hospital with fever and vomiting. 

During the previous two months he had repeated hospitalizations 

and antibiotic treatments for urinary tract infection. On 

admission he had anaemia (Hb 8.3 g/dl; HT 28%), normal 

leukocytes (5380/µl), increased creatinine (1.7 mg/dl); a chest 

X-ray showed reticulo-nodular infiltrates. He was started on 

piperacillin-tazobactam after blood and urine cultures had 

been taken. Three days later he developed watery diarrhea 

with 4 to 5 loose stools daily along with poor appetite and 

vomiting. Stool culture for Salmonella, Shigella and C. difficile 

were negative whereas two stool samples were positive 

for C. difficile toxins. He subsequently developed stypsis, 

worsening abdominal pain and marked leukocytosis (27,320/ 

µl). A plain abdominal X-ray and abdominal CT demonstrated 

signs of ileus with marked thickening of the wall of rectum 

and sigma. Despite treatment with vancomycin plus intravenous 

metronidazole the patient died 15 days later. 

Case 4. A 39-year-old HIV-infected woman not taking antiretroviral 

therapy presented with a 4-month history of watery 

and bloody diarrhea, cramping abdominal pain and weight 

173 

loss. Her last CD4 cell count was 400/µl and plasma HIV load 

11,726 copies/ml. Previous stool examinations for bacteria, 

mycobacteria and protozoa gave negative results; antigliadin 

and transglutaminase antibodies were absent, while fecal occult 

blood tests were persistently positive. On admission physical 

examination was remarkable for fever (39°C), tenderness 

in left lower abdomen and external haemorrhoid. Laboratory 

tests showed anemia (9.5 g/dl), high CRP (211 mg/l), hypokaliemia 

(2.5 mmol/l), hypoalbuminemia (2.4 g/dl). Abdominal 

CT-scan showed moderate hepato-splenomegaly, without 

other relevant findings. Colonoscopy revealed the presence 

of multiple ulcerations in the left colon that were biopsied. 

The ulcers where characterized by a complete loss of the 

lamina propria, of the muscolaris mucosae and of part of the 

submucosal tissues, with a dense lymphoplasmocytic infiltrate. 

By immunohistochemistry, most of the lymphoid cells 

in the ulcerative lesions were a mixture of CD3/CD4+ and 

CD3/CD8+ T lymphocytes, intermingled with CD20+ B cells, 

CD138+ plasma cells and CD30+/CD15- blastic cells. Immunohistochemistry 

for CMV and in-situ hybridization for EBV/ 

EBER were negative. Extensive microbiologic and serologic 

investigations in faeces and peripheral blood were uniformly 

negative. A complete autoantibody panel was also negative. 

Empiric antimicrobial therapy with gancyclovir, ciprofloxacin 

and metronidazole was started but profuse diarrhoea persisted 

unchanged. After a massive rectal bleeding a new colonoscopy 

demonstrated multiple ulcers and a recto-vaginal fistula. 

Methylprednisolone (20.mg bid) was empirically added to the 

antibiotic regimen and antiretroviral treatment was started. 

After 15 days a follow-up colonoscopy was perfomed but the 

procedure was complicated by perforation of the sigma and 

subsequent emergency laparotomy with left hemicolectomy 

and ileostomy. The patient was then treated with a prolonged 

course of tapering steroids along with combined antiretroviral 

therapy. She is now without diarrhoea or rectal bleeding for 

6 months. 

Discussion. The first two vignettes, which respectively 

describe a case of CMV colitis and a case of shigellosis, 

highlight the diagnostic work-up of acute bloody diarrhoea 

(ABD) with special emphasis on travellers. Initial microbiological 

work-up should include bacteria (Salmonella, Shigella, 

Campylobacter, Yersinia, E. coli O157:H7) and parasites 

(Entamoeba histolytica; Schistosoma), which are the most 

frequent agents involved in ABD. Routine microbiologic 

cultures usually target the first three microorganisms whereas 

special requests are needed for Yersinia and E. coli O157: 

H7. A test for C. difficile should be included as it may cause 

infections running a severe course also in non-traditional risk 

groups including healthy persons in the community without 

antimicrobial exposure 3 . Freshly passed stool examination is 

required to search for trophozoites of E. histolytica and for 

ova of Schistosoma; moreover staining of fixed faecal smears 

with iron-hematoxylin or Ziehl-Neelsen can determine the 

presence of the E. histolytica/E. dispar complex. Imaging 

studies, in particular abdominal CT scan with oral or intravenous 

contrast, are useful to assess anatomic localization and 

extent of bowel involvement and complications such as perforation. 

Colonoscopy permits to identify and characterize mucosal 

lesions, and obtain biopsies that would allow differential 

diagnosis with non infectious diseases (e.g. inflammatory 

bowel disease, colon cancer or ischemic colitis) and detect 

unexpected infections (e.g. schistosomiasis) and infections 

that cannot be diagnosed otherwise (e.g. CMV colitis). Bowel 

infections with CMV is typical of immunocompromised patients; 

it has been also increasingly observed in IBD with most

174 

studies supporting a role for active CMV infection in causing 

exacerbations of the disease. Although rare, CMV colitis may 

occur in the immunocompetent-host during primary infection 

and may be potentially severe erosive disease with significant 

morbidity. 

The third vignette is exemplificative of nosocomial IE that is 

commonly defined as a diarrhoeic illness that occurs after 3 

days of hospitalization. C. difficile is the most important cause 

of nosocomial diarrhea in adults; the infection causes a toxin 

mediated intestinal disease that may range from mild watery 

diarrhea to life-threatening colitis. Its diagnosis is based primarily 

on the detection of C. difficile toxin A or toxin B. 

The last vignette underscore the diagnostic challenge of diarrhoeic 

syndrome in the setting of HIV/AIDS; the diagnostic 

work-up is largely influenced by the stage of the disease with 

classic intestinal opportunistic infections (e.g. microsporidiosis; 

cryptosporidiosis; isosporiasis; CMV colitis; intestinal 


COelIAC DISeASe 

mycobacterioses) occuring in patient with less than 100 CD4+ 

lymphocytes/µl. If after exhaustive stool studies no pathogen 

is isolated there is clearly a role for invasive endoscopic evaluations, 

particularly in patients with severe refractory diarrhoea 

4 . Active idiopathic ulcerative colitis has been described 

in HIV-positive patients independently of the depression of 

peripheral CD4 cells count. 

references 

1 Thielman NM, Guerrant RL. Acute infectious diarrhea. N Engl J Med 

2004;350:38-47. 

2 Pawlowski SW, Warren CA, Guerrant R. Diagnosis and treatment of 

acute or persistent diarrhea. Gastroenterology 2009;136:1874-86. 

3 Rupnik M, Wilcox MH, Gerding DN. Clostridium difficile infection: 

new developments in epidemiology and pathogenesis. Nature Rev 

Microbiol 2009;7:526-36. 

4 Cello JP, Day LW. Idiopathic AIDS enteropathy and treatment 

of gastrointestinal opportunistic pathogens. Gastroenterology 

2009;136:1952-65. 

Initial lesions – Differential diagnosis – refractory forms 

Gluten- and non-gluten-dependent 

non-atrophic lesions: the clinician 

and the pahologist viewpoint 

U. Volta, G. Caio, E. Tavani * , A. Andorno * 

Dipartimento di Malattie dell’Apparato Digerente e Medicina Interna, 

Policlinico “S. Orsola-Malpighi”, Bologna, Italia, * U.O Anatomia 

Patologica, A.O. “G. Salvini” – Ospedale di Rho, Italia 

Non-atrophic lesions of the small bowel mucosa are characterised 

by minimal intestinal lesions with an increased 

number of intraepithelial lymphocytes (IEL), with or without 

crypt hyperplasia and in presence of a normal architecture 

of intestinal villi. Minimal intestinal lesions, though 

non-specific for coeliac disease (CD), are included in the 

wide spectrum of gluten-dependent histological damage and 

they can be an expression of potential CD. However, they 

can be a sign of many disorders other than gluten-sensitive 

enteropathy. Therefore, it is important to differentiate patients 

with gluten-dependent intestinal damage from those 

with non gluten- dependent intestinal lesions in order to 

plan the correct treatment and follow-up. On the basis of 

the up-to-date histological classifications small intestinal 

non-atrophic lesions correspond to type 1 (IEL increase) 

and type 2 lesions (IEL increase with crypt hyperplasia), 

according to Marsh classification, modified by Oberhuber 1 

or to grade A, according to the most recent Corazza-Villanacci 

classification, which gathers Marsh-Oberhuber type 

1 and 2 lesions 2 . 

IEL are normally present in the intestinal mucosa of healthy 

people where they play a pivotal role in the surveillance and 

activation of the immune system. The majority of these lymphocytes 

is represented by T cells expressing α/β receptor. In 

the normal mucosa only 3% of IEL express γ/δ T-cell receptor. 

The upper normal limit of IEL in the duodenum, once 

fixed in 40/100 epithelial cells, is generally acknowledged 

to be 25/100, evaluating the mean value of the lymphocyte 

counts in five different points. 

Moderators: V. Villanacci (Brescia), A. D’Errico (Bologna) 

Non-atrophic lesions of the intestinal mucosa have been observed 

in 2.2%-24% of patients undergoing duodenal biopsy 

due to the clinical suspect of small bowel disease 3-5 . As well 

known, the typical histological picture of CD is based on more 

o less severe villous atrophy with a significant decrease of 

villous/crypt ratio, an increased number of IEL and crypts hyperplasia. 

When the villous architecture and the villous/crypt 

ratio are normal, the increased number of IEL and crypt hyperplasia 

show a very low predictive value for CD, since only 

in a small percentage of these patients (about 10%) the aetiology 

of the intestinal damage is attributable to a developing 

gluten-sensitive enteropathy 6 . In the remaining 90% of cases 

the finding of non-atrophic lesions of small intestinal mucosa 

is an expression of a wide spectrum of non-gluten-dependent 

disorders such as food allergy, Crohn disease, lymphocytic 

colitis, bacterial and parasitic infections (giardiasis is one of 

the most frequent one), common variable immunodeficiency 

(CVID), autoimmune disorders (Hashimoto thyroiditis, diabetes 

mellitus type 1, rheumatoid arthritis, systemic lupus 

erythematosus), small bowel bacterial overgrowth, non-steroidal 

anti-inflammatory drug treatment and helicobacter pylori 

infection. Moreover, it must be remembered that an increased 

number of IEL is present in 10-38% of 1 st degree relatives 

of coeliac patients without a pathological significance in the 

majority of cases. 

Although the diagnostic criteria for gluten-sensitive enteropathy 

clearly establish that non-atrophic lesions of small bowel 

mucosa are compatible, but not specific for CD, one of the 

emerging problems encountered in the clinical practice is the 

over-diagnosis of CD, improperly performed on the basis of 

these minimal changes in the intestinal mucosa, without reference 

to the other predictive factors for the identification of the 

gluten-sensitivity 7 . The importance of these wrong diagnoses 

of gluten-sensitive enteropathy is still much more relevant, 

if we consider that, following these diagnostic mistakes, a 

lifelong, expensive and socially limiting gluten-free diet and a 

periodical follow-up are recommended for an inexistent disor-

lectures 

der. On the other hand, the identification of the small percentage 

of patients, in whom non-atrophic intestinal lesions can 

predict the development of CD, is mandatory since this allows 

to confine the follow-up for confirming the gluten-dependent 

origin of intestinal damage to this small subgroup, avoiding 

to monitor uselessly the majority of other patients. To achieve 

this goal, it is relevant that: 

– small intestinal mucosal lesions are evaluated in the clinical, 

serological and genetic context; 

– histological evaluation of intestinal damage must be performed 

in the respect of well-defined technical rules. 

Since the clinical-serological context is extremely variable, 

the interpretation of the morphological changes performed by 

the pathologist should be flexible, above all at the beginning 

of the diagnostic work-up, drawing up the conclusion only 

when the whole “scenario” will be assessed together with the 

clinician. 

As for the technical rules concerning small intestinal histology, 

first of all it is relevant to underline that mucosal lesions 

in CD are not always continuous, but they can be patchy and 

irregular. Therefore, at least four biopsy samples from the 

second-third portion of duodenum should be picked up. 

It is essential that biopsy samples are correctly oriented. This 

is important not only for the evaluation of atrophic lesions, 

but also for a correct interpretation of minimal changes of 

small intestinal mucosa. A well-oriented intestinal biopsy allows 

a good evaluation of villous/crypt ratio (≥ 3:1 in a normal 

mucosa) and above all an accurate count of IEL, which is very 

difficult to obtain with transversal sections and/or convoluted 

villi. 

When an increase of IEL is suspected as the sole marker of 

intestinal mucosa damage, the use of immunohistochemistry 

represents a mandatory adjunctive technique to their counting, 

allowing to stain CD3+ and CD8+ T lymphocytes. An associated 

evaluation of CD4+ T lymphocytes, though suggested 

in the past, seems to be useless in the histological work-up 

of intestinal mucosa. IEL counts should be done taking into 

consideration five villi; moving from villous tips, lymphocytes 

present in 20 enterocytes (10 on the right and 10 on 

the left part of the villi) must be enumerated, establishing the 

mean value. The mean value is 9.2 lymphocytes/20 epithelial 

cells (range 5.8-21.8) in subjects with an abnormal mucosa 

(gluten-sensitivity or other pathological conditions) vs 4.6 

lymphocytes/20 epithelial cells (range 1.4-7.8) in subjects 

with a normal mucosa. The lymphocyte distribution along 

villi is substantially homogeneous; a higher lymphocytic 

concentration in the villous tip helps to identify patients with 

gluten-sensitivity 8 . Immunohistochemical characterization of 

lymphocyte populations in the intraepithelial compartment 

by using duodenal biopsy frozen sections may be useful in 

identifying gluten-sensitive patients. It is well established 

that a high density of T-cells with γ/δ receptors in he surface 

epithelium is a characteristic feature of gluten sensitivity. 

The mean proportion of γ/δ T cells in gluten-dependent nonatrophic 

lesions varies from 20% to 30%, whereas, when 

non-atrophic lesions are non-gluten-dependent, γ/δ T cells are 

about 2%-3%. However, this modality has limited diagnostic 

utility due to the non-availability of an assay for identifying 

γ/δ T cells in formalin-fixed, paraffin-embedded tissue. 

The characterization of crypt mitotic index by measuring the 

number of K67+ cells by immunohistochemistry can help to 

differentiate between gluten-dependent and non gluten-dependent 

intestinal damage. A value of K67+ cells higher than 

60-65% is suggestive for a condition of gluten sensitivity. 

On the contrary, many attempts to evaluate variations of the 

175 

immunohistochemical expression of tissue transglutaminase 

activity did not produce interesting results. 

Non-atrophic intestinal lesions should be evaluated in the 

context of clinical, serological and genetic data 9 . Although 

the diagnosis of CD on the basis of clinical data is an utopia, 

among symptoms, that can rise the suspect of gluten-sensitve 

enteropathy, there are bowel abnormalities, including both severe 

diarrhoea and marked constipation, weight loss, recurrent 

abdominal pain, iron-deficiency anaemia, hypertransaminasaemia 

of unknown origin, unexpected osteoporosis, recurrent 

miscarriages and CD-related autoimmune disorders. 

In order to establish if non-atrophic lesions are or not 

are gluten-dependent the detection of CD-related serological 

markers is much more relevant 10 . Many subjects with 

minimal changes of small bowel mucosa are classified as 

potential coeliacs on the basis of an antibody pattern that is 

not specific for CD. It is well-known that anti endomysial 

antibodies (EmA) of IgA class are the immunologic marker 

with a nearly always absolute specificity for CD. Anti tissue 

transglutaminase antibodies (anti-tTG) of IgA class display a 

very high predictive value for identifying CD when positive 

at a very high titer (> 5x the cut-off), whereas they show at 

least 10% of false positives when their antibody titer is very 

low (< 2x the cut-off). Therefore, their positivity, particularly 

at a low titer, should be always confirmed by EmA finding. 

Antibodies to deamidated gliadin peptides (DGP-AGA) of 

IgG class are another immunological marker which proved to 

be particularly useful in differentiating between gluten- and 

non-gluten-dependent intestinal lesions. Their specificity for 

gluten sensitivity is far higher than that of IgA anti-tTG and 

very close to that of IgA EmA. Moreover, DGP-AGA display 

a higher diagnostic accuracy than the traditional and obsolete 

AGA test. After the introduction of DGP-AGA in the workup 

of CD, the traditional AGA lost their last indication for 

CD screening, that remained the identification of CD in the 

infancy (children aged less than 2 years). 

Another immunological sign predictive of gluten-sensitive intestinal 

damage is the finding of IgA anti-tTG in small bowel 

biopsies. Indeed, these antibodies can appear at intestinal level 

when they are still negative in patients’ sera and when the intestinal 

barrier is quite normal or shows only mild abnormalities 

11 . Most of these patients, left on a gluten containing diet, 

display after a less or more lasting follow-up the development 

of villous atrophy associated with the appearance of serum 

antibodies. 

The puzzle of gluten sensitivity comprises another relevant 

element represented by genetic testing. As generally acknowledged, 

CD is closely related to a well-defined HLA pattern, 

characterized by positivity for HLA-DQ2 and -DQ8. The 

positivity of the test (finding of DQ2 or DQ8 or DQB1*02) is 

never diagnostic for CD by itself since about 30% of the general 

population displays the same HLA pattern of CD patients. 

The most important clinical message of the test comes from 

its negativity since the absence of DQ2, DQ8 and DQB1*02 

allows to exclude CD (negative predictive value 100%) 9 . In 

patients with a suspect of potential CD (positive serology 

with mild or absent histological lesions) HLA genotyping 

for DQ2 and DQ8 is useful to reinforce (when positive) or 

exclude (when negative) the suspect of a gluten-dependent 

intestinal damage. In patients with non atrophic intestinal lesions 

and negative serology the HLA negativity should alert 

us to search for another cause of small bowel abnormalities 

(CVID, giardiasis, helicobacter pylori infection, etc.). In patients 

with positivity at low titer for IgA anti-tTG, with IgA 

EmA and IgG DGP-AGA negativity, the absence of HLA-

176 

DQ2 and -DQ8 gives evidence that IgA anti-tTG are likely 

“false positives”. 

The erroneous interpretation of clinical, serological and genetic 

data significantly contributes to the plethora of false 

CD diagnoses performed in patients with minimal intestinal 

changes. The most frequent pitfalls leading to an over-diagnosis 

of CD in patients with non-atrophic intestinal lesions 

are the positivity for IgG anti-tTG in absence of selective IgA 

deficiency, the isolated finding of HLA-DQ2 or -DQ8 (that 

are only expression of a genetic predisposition for CD), the 

low titer positivity for IgA anti-tTG associated with negativity 

for IgA EmA, the isolated positivty for IgA AGA in children 

older than 2 years and in adults, and a gluten hypersensitivity 

on a clinical ground, often caused by irritable bowel syndrome 

or wheat allergy 

To sum up, the finding of non-atrophic intestinal lesions is 

a non-specific immunopathological phenomenon, that has a 

large number of possible causes, and by itself is never synonymous 

of gluten-sensitive enteropathy. A correct histological 

evaluation is recommended in order to avoid false CD diagnoses, 

caused by artifacts due to a not well-oriented biopsy. The 

evaluation of IEL count in villous tip as well as of γ/δ T-cell 

receptor lymphocytes can be of help in distinguishing between 

non-celiac patients and patients at risk of developing CD. 

A careful evaluation of the clinical, serological and genetic 

aspects must be carried out in all patients with non-atrophic 

lesions in order to identify the minority of cases affected by 

potential CD or who must undergo a periodic follow-up for 

the possible development of gluten-sensitive enteropathy, 

and to eliminate this suspect in the majority of cases, who 

should be studied for pathological conditions other than gluten-sensitivity. 

Awareness of the wide spectrum of disorders 

Immunohistochemical markers in cytology 

of neoplastic thyroid disease 

M. Volante, L. Daniele, M. Papotti. 

Department of Clinical and Biological Sciences, University of Turin, 

Turin, Italy 

Thyroid nodules represent a common clinical problem. 

The prevalence of palpable thyroid proliferations in adults 

increases with age, with an average of 4-7% for the United 

States population but higher in iodine-deficient areas where 

sub-clinical nodules are frequently incidentally discovered 

following thyroid ultrasound-scan. More than 90% of these 

thyroid proliferations are benign and for this reason a reliable 

and systematic approach to their evaluation represents 

an important task to be pursued for avoiding a surgical overtreatment. 

Ultrasound-guided fine-needle aspiration biopsy 

(FNAB) is the gold standard for thyroid nodule evaluation, 

but it is widely known that this method has some intrinsic 

limitations and immunohistochemistry represents the most 

reliable technique to assess the cytomorphological diagnosis 

in difficult cases. The application of immunohistochemistry 

in thyroid cytology may have as a first aim the identification 

the cell type of origin of the lesion, including thyroglobulin 

or TTF-1 for follicular cell-derived lesions, calcitonin, 


Thyroid cytology 

Moderators: A. Fassina (Padova), M. Papotti (Torino) 

associated with non-atrophic lesions of small bowel mucosa 

is important to guide the clinician and the pathologist toward 

a correct diagnosis. 

references 

1 Oberhuber G, Granditsch G, Vogelsang H. The histopathology of celiac 

disease: time for a standardized report scheme for pathologists. 

Eur J Gastroenterol Hepatol 1999;11:1185-94. 

2 Corazza GR, Villanacci V. Coeliac Disease. J Clin Pathol 2005;58;573- 

4. 

3 Brown I, Mino-Kenudson M, Deshpande V, et al. Intraepithelial 

lymphocytosis in architecturally preserved proximal small intestinal 

mucosa. Arch Pathol Lab Med 2006;130:1020-5. 

4 Biagi F, Bianchi PI, Campanella J, et al. The prevalence and the 

causes of minimal intestinal lesions in patients complaining of symptoms 

suggestive of enteropathy. A follow-up study. J Clin Pathol 

2008;61:1116-8. 

5 Lahdeaho ML, Kaukinen K, Collin P, et al. Celiac disease: from 

inflammation to atrophy, a long-term follow-up study. J Pediatr Gastroenterol 

Nutr 2005;41:44-8. 

6 Kakar S, Nehra V, Murray JA, et al. Significance of intraepithelial 

lymphocytosis in small bowel biopsy samples with normal mucosa 

architecture. Am J Gastroenterol 2003;98:2027-33. 

7 Upton MP. “Give us this day our daily bread”. Evolving concepts in 

celiac sprue. Arch Pathol Lab Med 2008;132:1594-9. 

8 Biagi F, Luinetti O, Campanella J, et al. Intraepithelial lymphocytes in 

the villous tip do they indicate potential coeliac disease? J Clin Pathol 

2004;57:835-9. 

9 Volta U, Villanacci V, Tavani E, et al. La diagnosi di malattia celiaca. 

Pathologica 2007;99:412-4. 

10 Volta U, Granito A, Fiorini E, et al. Usefulness of antibodies to deamidated 

gliadin peptides in celiac disease diagnosis and follow-up. Dig 

Dis Sci 2008;53:1582-8. 

11 Salmi TT, Collin P, Järvinen O, et al. Immunoglobulin A autoantibodies 

against transglutaminase 2 in the small intestinal mucosa predict 

forthcoming coeliac disease. Aliment Pharmacol Ther. 2006;24:541- 

52. 

chromogranin A or CEA for medullary carcinoma, parathyroid 

hormone for parathyroid lesions, pan-cytokeratin for 

anaplastic carcinoma, lymphoid markers for lymphomas, 

among others. However, the major applicative field is represented 

by the distinction between benign (i.e. microfollicular 

nodular hyperplasia and follicular adenoma) and malignant 

follicular lesions (i.e. follicular thyroid carcinoma and follicular 

variant of papillary carcinoma). In fact, these follicular 

thyroid nodules, that remain indeterminate at thyroid FNAB 

cytology (classified TIR3 according to recently proposed 

SIAPEC guidelines), are referred to surgery more for diagnosis 

than for therapeutic purposes, and less that 10-20% 

of such cases will prove to be malignant at final histology. 

To reduce the number of follicular proliferations referred 

to surgery, and therefore to reduce costs for public health, 

several immunocytochemical markers have been proposed 

to distinguish malignant from benign follicular proliferations. 

The use of immunocytochemical markers on FNAB 

material may be generally employed on smears, although 

cell block preparations seem to be more reliable in this 

specific setting. The markers proposed are mainly related to 

tumor-associated abnormal expression of cellular antigens, 

such as cell surface mesothelial antigen HBME-1 (HBME1), 

cytokeratin-19 (CK19), thyreoperoxidase (TPO), keratan-

lectures 

sulfate (KS), or to the specific expression of cell-cycle or 

apoptosis related molecules, such as galectin-3 (GAL-3), or 

oncogenes, such as RET. As a general comment, it is generally 

advisable to rely not on a single marker but rather on a 

combination, to achieve the best specificity and sensitivity 1- 

3 . The role of one of the most employed markers, GAL-3, has 

been recently validated in a large prospective multicentric 

study from an Italian population 4 and confirmed an overall 

sensitivity and specificity of this immunocytochemical test 

of 85% and 93%, respectively, with estimated positive and 

negative predictive values of 83% and 94% respectively. 

More than 91% of indeterminate (TIR3) follicular thyroid 

nodules enrolled in this study were considered correctly 

classified preoperatively. 

Breast cytology: reporting 

Breast cytology 

177 

references 

1 Maruta J, Hashimoto H, Yamashita H, et al. Immunostaining of galectin-3 

and CD44v6 using fine-needle aspiration for distinguishing follicular 

carcinoma from adenoma. Diagn Cytopathol 2004;31:392-6. 

2 Rossi ED, Raffaelli M, Minimo C, et al. Immunocytochemical evaluation 

of thyroid neoplasms on thin-layer smears from fine-needle 

aspiration biopsies. Cancer 2005;105:87-95. 

3 Saggiorato E, De Pompa R, Volante M, et al. Characterization of 

thyroid ‘follicular neoplasms’ in fine-needle aspiration cytological 

specimens using a panel of immunohistochemical markers: a proposal 

for clinical application. Endocr Relat Cancer 2005;12:305-317. 

4 Bartolazzi A, Orlandi F, Saggiorato E, et al., Italian Thyroid Cancer 

Study Group (ITCSG). Galectin-3-expression analysis in the surgical 

selection of follicular thyroid nodules with indeterminate fine-needle 

aspiration cytology: a prospective multicentre study. Lancet Oncol 

2008;9:543-9. 

Moderators: A. Bellomi (Mantova), A. Leotta (Lamezia Terme) 

L Di Bonito, F Martellani, D Bonifacio, S Dudine, M Di Napoli, 

E Isidoro, E Ober, E Leonardo, A Romano, A Zacchi,, 

T Al Omoush, D Bonazza, A De Pellegrin, O Haxhijmeri, M. 

Petris, L Zandonà, V Bandiera * , F Giudici * , L Torelli * , M 

Bortul *** , M Tonutti ** , F Zanconati 

U.C.O. Anatomia e Istologia Patologica Azienda Ospedaliero Universitaria 

Ospedali Riuniti Università di Trieste; * Dipartimento di 

Matematica e Informatica Università di Trieste; ** U.C.O. Radiologia 

Universitaria Azienda Ospedaliero Universitaria Ospedali Riuniti 

Trieste; *** U.C.O. Clinica Chirurgica Azienda Ospedaliero Universitaria 

Ospedali Riuniti Università di Trieste 

Fine needle aspiration cytology (FNAC) is widely used as 

first choice approach for the definition of the radiologically 

dubious or suspicious cases or to confirm their benign origin. 

In experienced hands, FNAC represents a reliable technique 

that has many advantages: it is a simple and fast exam with 

minimal invasiveness and low costs. 

Regarding FNAC reporting we refer to the guidelines proposed 

by the English Screening Program 1 subsequently adopted by 

the European guidelines 2 . The use of these categories by the 

Breast Unit of Trieste since 2002 provides a standardization 

of the diagnostic report. According to this classification’s system 

each lesion is placed in one of the five categories (C1-5) 

shortly described below. 

C1 = INADEQUATE; includes all those cases which do not 

provide the possibility to solve a specific diagnostic problem 

(poor cellularity, bad technical preparation, excessive inflammatory 

or blood’s elements,); 

C2 = BENIGN; there’s no evidence of malignancy. It includes 

all cases characterized by absence of nuclear or morphological 

alterations. 

C3 = PROBABLY BENIGN; 

includes all cases in which 

the smear’s cells are not 

certainly interpretable as benign. 

Management of such 

cases requires correlation of 

cytology with clinical and / 

or radiological aspect. 

C4 = SUSPICIOUS FOR 

MALIGNANCY; the cellular 

Tab. I 

Data 

2008-2009 

Histology 

malignant 

Histology 

benign 

c5 cytology 

malignant 

appearance, although highly suggestive for malignancy, is not 

conclusive. This category includes the cases with few highly 

atypical cells and some very well-differentiated tumors. These 

lesions must undergo biopsy to obtain a conclusive diagnosis 

or, in cases with low cellularity, FNAC can be repeated. 

C5 = MALIGNANT; cytological features are diagnostic for 

malignancy. Sometimes, through FNAC the histotype of malignancy 

can be determined. 

Trieste’s Breast Unit has been using FNAC as first morphological 

investigation for many years, in particular, in 2008- 

2009, it was used as first diagnostic approach for 1835 cases 

(88.6%) out of 2091. Thanks to FNAC, 742 lesions were 

diagnosed as benign (C2) (with clinical follow-up confirmation) 

avoiding more invasive histological investigations (i.e. 

microbiopsy and surgical biopsies). Besides that, thanks to 

the FNAC it was possible, for cases with surgical indication, 

to plan a targeted intervention: excisional nodulectomy for 

large benign (C2) or likely benign (C3) lesions, conservative 

treatment for malignant monofocal or small lesions (quadrantectomy) 

or mastectomy for malignant multifocal or locally 

advanced tumors. For suspicious lesions (C4) surgical approach 

(nodulectomy vs. diagnostic quadrantectomy with or 

without Sentinel Lymph Node) was decided considering the 

type of radiological suspicion. 

The advantage of using of the diagnostic categories is the possibility 

to correlate with the final outcome of the histology or 

follow-up. In Table I cyto-histological correlations of FNAC 

of breast nodules only (excluding the sampling of lymph 

nodes and chest wall’s nodules). 

The table shows a prevalence of C5 and C2 lesions: C5 were 

626 (29.9%) and C2 were 790 (37.8%) accounting for 67,7% 

of the total lesions investigated. 

c4 cytology 

suspicious 

c3 cytology 

atypical 

c2 cytology 

benign 

c1 cytology 

inadequate Total 

591 72 14 1 10 688 

1 19 76 47 13 156 

No histology 34 4 75 742 57 912 

Total C 626 95 165 790 80 1756

178 

Cyto-histological correlation allows continuous monitoring 

of quality’s indicators of breast diagnostic cytology provided 

by the laboratory, with the possibility to compare them with 

the standard suggested by the guidelines; the results are summarized 

in Table II. 

Tab. II 

Quality 

Indicators 

Two-years period 

2008-2009: 1756 


Standard (%) 

As 86.6% > 60% 

Cs 98.5% > 80% 

Sbx 30.1% / 

Spe 76.6% > 60% 

C5 Ppv 99.8% > 95% 

C4 Ppv 79.1% 70-80% 

C3 Ppv 8.5% < 20% 

Fis- 0.14% < 5% 

Fis+ 0.14% < 1% 

Ina 4.5% < 25% 

Inca 1.4% < 10% 

Sus 14.7% < 20% 

as: absolute sensitivity; cs: full sensitivity; 

sBX: specificity (only biopsied cases); 

spe: full specificity; 

c5 Ppv: positive predictive value of c5; c4 Ppv: positive predictive 

value of c4; c3 Ppv: positive predictive value of c3; fis-: rate of false 

negatives; fis+: rate of false positives; ina: inadequate’s rate; inca: 

inadequate rate with final diagnosis of cancer; sus: rate of suspects. 

The direct participation of the cytopathologist in all sampling 

sessions has allowed to get optimal smears without artifacts 

with an immediate adequacy assessment. This has allowed 

the immediate repetition – in the same session – of the cases 

with suboptimal material, helping to minimize the number of 

inadequates (4.5%), well below the maximum allowed value 

(25%). This activity also allows the pathologist a constant 

dialogue with the radiologist, providing a chance to choose 

collectively the most appropriate method for solving the 

single diagnostic doubt. 

The positive predictive value of C5 has been constantly 

maintaining at high levels: 99.8% (standard required: > 95%): 

respect of this parameter is necessary to omit frozen sections 

in all C5 cases, allowing to plan conservative surgery without 

further confirmation. 

The positive predictive value of C4 (95 lesions, 5.4%) was 

79.1% (standard value 70-80%). The positive predictive value 

of C3 (165 lesions, 9.3%), that according to the guidelines 

must remain below 20%, was 8.5%. Overall inconclusive lesions 

(C3 and C4) were 14.7% with a rate of suspicious cases 

well below 20%. 

The use of diagnostic categories in cytology reporting has 

found wide acceptance among radiologists and surgeons 

because it allows to apply to each lesion, a precise diagnostic/ 

therapeutic pathway and it represents, in our experience, an 

essential element of the report itself. 

references 

1 Guidelines for Cytology Procedures and Reporting in Breast Cancer 

Screening - Cytology Sub-Group of the National Coordinating Commitee 

for Breast Screening Pathology. NHS-BSP 1993;22. 

2 European guidelines for quality assurance in breast cancer screening 

and diagnosis, fourth edition, 2006. 

Management and standardization in anatomic pathology 

Analysis of the SIAPeC-IAP study and future perspectives 

Analysis of the results of the research/study 

SIAPeC-IAP 

E. Trinchero 

Public Management and Policy Department, SDA Bocconi School of 

Management, Milan, Italy 

Background. The possible aims related to the definition 

of standard times for the execution of the proper and typical 

activities of a Pathology Unit basically consists of the 

staff definition/identification, the personnel planning and 

management, the eventual restructuring and reorganisation 

of the Unit, the rationalisation in the employment of human 

resources, the definition of a rational and quantitative base for 

the budgeting negotiation. The methodologies for the determination 

of the standard times of health services are several. 

The possible alternatives, on which the research done for (and 

in cooperation with) Italian Society of Anatomic Pathology 

and Cytopathology (SIAPEC) is based, can be schematised 

as follows. 

1. Methodologies based on a TOP-DOWN or “synthetic” 

APPROACH. Following this approach, the calculation of 

the workloads is made on the final output. The standard 

work time per unit needed for the production of each typol- 

Moderators: C. Angeli (Vercelli), F. Crivelli (Gallarate) 

ogy of output is calculated by dividing the total time which 

each professional profile actually works by the output which 

is actually produced. The TOP-DOWN approach typically 

allows for the definition of time standards through synthetic 

surveys, which are simple and quick and allow for the determination 

of a good overview of the situation. This approach 

is focused on the service: it produces standards which can be 

compared between Organisation Units or hospitals applying 

the same method, although it consider neither the health organization 

processes nor the quality of the services offered. 

The validity of the result is strongly affected by the way in 

which estimations are introduced, by the method of data 

collection and by the survey sources used, although, being a 

synthetic approach, it implies a diffused and non-answerable 

involvement. 

2. Methodologies which follow a BOTTOM-UP or “analytical” 

APPROACH. Following this approach, the workload 

calculation is made on the procedures subdivided into microphases. 

The standard unit work time needed for the production 

of each typology of output is calculated by the analytical 

measurement of the time necessary for the execution of each 

procedural micro-phase in terms of man-time. The determination 

of standard loads requires analytical surveys, which

lectures 

absorbs time to the business applying it: for this reason this 

methodology involves a high motivation and participation, but 

once implemented, it can surely be a useful tool for the management 

of Organisation or Department Units. It is focused on 

the business processes and produces results that are very little 

comparable between different Units or hospitals. 

Actually, the most diffused approach among the existing 

methodologies for the determination of workloads, which is 

applied by different businesses to comply with law obligations, 

is the top-down approach. The reason for this choice 

can be mainly identified in the fact that the complexity of 

the health system does not allow for analytical surveys about 

the procedures (which are very often not explicit) within the 

deadline set by the legislation. Furthermore, this approach 

makes it possible to obtain a global overview on the use of 

human resources, thus on the efficiency levels both at a business 

and inter-business level (regional or national), by comparing 

similar structures, and it allows for the determination 

of standard loads which are common to different businesses. 

It overcomes the problem of the determination of standard 

loads by comparing production times of similar realities. 

Last but not least, we can affirm that this method respects 

more the professionalism of the operator, who is not considered 

as a mere executor of tasks, but is made responsible for 

a determined set of objectives. 

Methods. The project on standard load measurement for 

Pathology Unit has been developed and implemented into the 

following phases: i) creation of the work team and definition 

of the hospitals sample; ii) choice of the methodology; iii) creation 

of the reference activities list; iv) attribution of activity 

to the different professional profiles involved; v) creation of 

the informatics support for the collection and analysis of the 

information; vi) data collection; vii) data elaboration and 

simulation. 

The analysis of the national and regional legislation carried 

out by the hospital reference persons has not evidenced anything 

particular that would have driven the choice towards a 

particular methodological approach. The choice of the methodology 

has been influenced by the cost of the information 

collection, on one side, and by the necessity to obtain a standard 

that would have made possible the comparison among 

different hospitals realities (limitation of the standard variability), 

on the other. Therefore, the work team has decided 

for the application of the methodology with TOP DOWN 

APPROACH for the calculation of the standard execution 

time of the activities, together with observations based on the 

BOTTOM UP APPROACH to determine the standard load 

179 

of some specific and particularly critical activities (Exfoliative 

cervico-vaginal cytology; exfoliative cervico-vaginal 

cytology on thin-layer preparation; cervical-vaginal drawing; 

autopsy with histology). 

The work team has decided to test such method on data concerning 

human resources and activities in a limited number of 

Italian Pathology Units (8) over the three-year period 2003- 

2005 and to extend the observation to a larger number of 

Italian Pathology Units (27) over the period 2005-2007. The 

27 Italian Pathology Units are distributed as follows: 20 in the 

North Area; 6 in the Central Area and 1 in the South Area. 

Concerning the typology of health organizations to which the 

experimenting Pathology Units belongs, 5 Units belong to 

Teaching Hospitals, 1 Unit belong to a Cancer Institute, the 

others belong to General Hospitals. 

As far as the observation of the activities is concerned, the 

work team has decided to adopt the 2002 SIAPEC activities 

list (“Nomenclatore tariffario” – second revision) which 

already included a weight system defined by SIAPEC itself, 

and which is also used to attribute the activities to the different 

professional profiles. 

Results. Pathologist, Biologist and Pathology Technician 

are the key professional profiles on which the analysis 

is focused. The average amount of hours over the period 

2005-2007 of the whole sample varies very much both in 

case of the same professional profile and in case of different 

professional profiles. This can be explained by a different 

labour organisation. Also the average amount of activities 

of the whole sample over the three years 2005-2007 varies 

concerning both the total amount produced, and the weight 

of the activities observed with the BOTTOM UP approach 

(Pap test and autopsies) on the total of the activity produced. 

In order to limit the variability phenomenon, the work team 

has decided to exclude the following from the data analysis: 

i) the centres with values strongly above average and ii) the 

centres with values well below average. The work team has 

decided not to consider outlier the centres with values below 

average for some professional profiles and above average 

for others. Therefore the analysis has been carried out on 

the data of 22 centres. The Average Time per weight unit 

(total hours/total points) of the sample excluded the outlier 

centres, both per year and aggregated for the three-years 

period (Tab. I) shows anyway a great variety among the 

centres, probably due to a different distribution of the activities 

among the professional profiles, thus due to a different 

labour organisation. 

Tab. I 

Nurses Path/Bio OTA Administrative PathTechn. 

min/point min/ point min/ point min/ point min/ point 

avarage time 1.37 9.76 3.21 3.45 14.54 

avarage time 

liguria 

0.00 11.21 4.23 1.69 17.03 

avarage time 

Piemonte+Vda 

0.46 10.86 2.76 3.64 17.33 

avarage time 

lombardia 

0.73 6.92 3.78 3.31 14.42 

avarage time 

north 

0.43 8.59 3.37 3.06 15.29 

avarage time 

centre 

2.53 12.36 2.30 1.68 11.45

180 

The appropriateness of pathological reports 

M. Pavesi 

S.O.C. Anatomia e Istologia Patologica, ASLAL Casale Monferrato 

Italia 

Pathologists are faced with two different kinds of relationships: 

one with the patient (from whom the specimen was 

taken) and one with the patient’s doctor, either a specialist or 

a general practitioner. 

Therefore, a pathologist’s report should be intelligible to 

addressees differing significantly from a cultural and a linguistic 

point of view but having the same needs: discovering 

pathologies quickly and accurately in order to programme a 

well-timed therapy. 

In the light of these facts, pathologists’ reports should be 

concise (stating the final diagnosis briefly), clear (easily interpretable, 

though not giving up the usually necessary scientific 

terminology) and have a univocal interpretation (understood 

by both specialist and family doctors despite their different 

training). 

In any case, the appropriateness of pathological reports 

cannot do without the evolution of etiological and pathogenetical 

findings of illnesses, together with a complete and 

complex evaluation of the prognostic factors of cancers. 

These aspects are of paramount importance to oncologists, 

who are to stadiate the illness and may use new drugs in the 

target therapy, which has also been legislatively approved 

for certain neoplastic diseases (breast, colonic and lung 

carcinoma). 

Therefore, an appropriate report should be complete and include, 

where possible, information about the biological and 

Dermoscopy and histopathology of nevi and 

melanomas: pitfalls and diagnostic correlations 

G. Ferrara 

Anatomic Pathology Unit, Gaetano Rummo General Hospital, Benevento, 

Italy, (E-mail gerardo.ferrara@libero.it) 

The increasing use of dermoscopy in preoperative diagnosis 

of melanocytic skin neoplasms (MSN) is impacting on routine 

histopathology to a relevant extent. We herein present 

the dermoscopic-pathologic features of some cases of histopathologically 

controversial MSN. By illustrating these cases, 

we would like to emphasize at least three different fields of 

interest for a combined (clinico-)dermoscopic-pathologic 

diagnostic approach, namely: information about the evolution 

of lesions; detection of gross sampling errors; definition of 

peculiar clinicopathologic entities. The theoretical and practical 

aspects of a close interaction among dermoscopists and 

histopathologists are itemized in detail. 

references 

1 Ferrara G, Argenziano G, Soyer HP, et al. Dermoscopic and histopathologic 

diagnosis of equivocal melanocytic skin lesions. An interdisciplinary 

study on 107 cases. Cancer 2002;95:1094-100. 

2 Ferrara G, Argenyi Z, Argenziano G, et al. The influence of the clinical 

information in the histopathologic diagnosis of melanocytic skin neoplasms. 

PLoS ONE 4(4):e5375. doi:10.1371/journal.pone.0005375. 


Pigmented skin lesions 

Moderators: G. Massi (Roma), G. Collina (Bologna) 

clinical course of the illness and all the factors that define its 

pathological stadiation. 

Moreover, pathologists’ reports should be precise and unequivocally 

specify the received and examined tissue, any 

technical and special stains integration into normal procedures 

for diagnostic reasons, and the clinical and anamnestic data 

which are believed useful to pathological diagnoses. 

In the light of this, pathologists organise their final report in a 

complex way, again with the aim of giving more complete and 

clearer information about the pathology in question. 

Sometimes an attitude as such may turn out to be counterproductive, 

insofar as pathologists are unconsciously led to 

make deductions on a purely unscientific basis: the need for a 

complete consultation on pathologies should not confuse the 

objectivity of the observation. 

Observations should never be considered of minor importance 

as pathology is mainly a morphological study. As such, it 

cannot do without a descriptive observation of the extracted 

tissue. Ancillary colouring techniques are an essential aid to 

the final diagnosis, but should not lead to conclusions based 

on their interpretation only. 

The final diagnosis of reports should result from an integration 

of morphological data and biological information obtained 

from the tissue in question (immunophenotype, molecular biology) 

together with clinical, serological and anamnestic data 

of the patient from whom the specimen for the pathological 

diagnosis was taken. 

Pathologists’ proper behaviour while writing the final report 

safeguards themselves and their whole staff from legal measures 

in case of patients’ complaints against inappropriate 

medical treatments. 

3 Bauer J, Metzler G, Rassner G, et al. Dermatoscopy turns histopathologists’s 

attention to the suspicious area in melanocytic lesions. Arch 

Dermatol 2001;137:1338-40. 

Spitz nevi, atypical spitz tumors and spitzoid melanomas: diagnostic 

application of fluorescence in situ hybridization and 

p16 immunohistochemical stain 

C. Clemente, F. Cetti Serbelloni, S. Pagliarini, L. Scopsi * 

Pathology and * Cancer Genetics Services, Casa di Cura S. Pio 

X, Milan, Italy 

Background. Recently, Abbott Molecular commercialized a 

multi-color FISH probe mixture to assist pathologists in the 

differential diagnosis of difficult melanocytic lesions. The 

probe mixture includes a centromeric probe for chromosome 6 

and unique sequence probes for the RREB1 gene (6p25), MYB 

gene (6q23-q23), and CCND1 gene (11q13). The centromeric 

probe (CEP6) was included as a control for the ploidy level 

of chromosome 6, while the other three were chosen because 

their respective chromosomal regions have most frequently 

shown amplifications or deletions in melanoma. After a preliminary 

study aimed at evaluating the technical application of 

the kit 1 , we wanted to test this new tool on an array of spitzoid 

lesions including: Spitz/Reed nevi, Spitz/Reed tumors with 

atypical features, spitzoid melanomas, and other more complex 

lesions with features simulating the previous ones. The

lectures 

p16 immunohistochemical stain developed by mtm Laboratories 

AG was also tested. The spitzoid lesions represent the 

most difficult area in the differential diagnosis of melanocytic 

tumors and in our second opinion experience Reed nevus is 

the most frequent entity misdiagnosed for melanoma (Clemente, 

unpublished). 

Methods. Sections from 112 archival paraffin blocks corresponding 

to 109 patients were obtained (four cases had 

two different specimens each). Eighty of the 112 specimens 

were from consultation files and came from a wide array 

of Italian health institutions. The selection criteria adopted 

included those listed in the background section. H&E slides 

were used to accurately identify the area(s) of interest, which 

were marked with a glass pen on the back of the slides to be 

used in the FISH procedure. The samples for FISH analysis 

were treated strictly following the manufacturer’s protocol as 

specified in the kit’s instructions (SP). The evaluation was 

carried out using an Olympus BX 51 fluorescent microscope 

equipped with a filter set including DAPI, spectrum aqua, 

spectrum green, spectrum yellow and spectrum red. Scoring 

was restricted to cells from the areas previously identified on 

the matched H&E sections and was carried out by two observers 

separately, without prior knowledge of the diagnosis 

(LS and FCS). Whenever feasible, scoring was done on 150 

(75 + 75) non-overlapping intact nuclei. A specimen was considered 

positive if at least one of the following criteria was 

met: CCND1 % gain > 38, RREB1 % gain > 29, percent loss 

of MYB against CEP6 > 40%, percent gain of RREB1 against 

CEP6 > 55%. A specimen was labeled as FISH-negative if 

none of the above criteria were met. 

P16 immunostaining was tested on a large series of 342 cases: 

68 Spitz/Reed nevi, 22 atypical Spitz/Reed tumors, 56 dysplastic 

nevi, 47 common nevi and 130 melanomas. 

Results. Seven FISH samples were not assessable because 

of technical reasons. For practical purposes, specimens were 

divided into four main categories: benign nevi, atypical 

melanocytic tumors, dysplastic nevi, malignant melanomas. 

Positivity ensued mainly from loss of MYB, followed by gain 

in RREB1 and gain in CCND1. Forty out of 46 histologically 

benign nevi scored negative: among these were all Reed nevi 

and 90% of Spitz nevi. The six positive nevi included two 

Spitz nevi, three Spitz-like compound nevi and one epithelioid 

blue nevus. Of the 21 atypical tumors, 16 scored negative; 

the five scoring positive included three atypical Spitz 

tumors, one atypical cellular blue nevus and one atypical 

epithelioid melanocytic (Spitz-like) tumor. Only one out of 

seven dysplastic nevi scored positive. 10 out of 31 melanoma 

specimens scored positive, the remaining resulting negative. 

Among these latter: one is a vulvar lesion of a 9-year girl; 

two others have features of nevoid and one of desmoplastic 

melanoma, two types of melanoma we found negative also 

in a previous study performed with this FISH tool 1 . One is 

a melanoma arising in (and mixed with) a nevus and it cannot 

be excluded that a portion of the nuclei evaluated during 

the scoring procedure belonged to normal melanocytes. This 

drawback, which is also present in specimens where the lesion 

is represented by small nests or even single cells in close 

contact with the epithelial cells or lymphocytes, calls for caution 

in interpreting the results. Interestingly, a lymph node 

metastasis from this same patient was FISH-processed too 

and resulted strongly positive. Five other negative melanomas 

belonged to the superficial spreading category and five to the 

spitzoid type. A preliminary look at possible links between 

FISH results and prognostic factors in melanomas showed 

181 

that positivity was mainly associated with the worst presenting 

signs and that the strongest positivity was restricted to the 

three metastasis. 

A p16 positive moderate to intense stain was present in 78% 

of Spitz/Reed nevi, 41% of melanomas, 36% of atypical 

Spitz/Reed tumor, 68% of dysplastic nevi and 64% of common 

nevi. 

reference 

1 Clemente C, Bettio D, Venci A, et al. A fluorescence in situ hybridization 

(FISH) procedure to assist in differentiating benign from malignant 

melanocytic lesions. Pathologica 2009;101(5):169-74. 

Nevoid Melanoma 

G. Collina 

Bologna 

Nevoid melanoma is one of the most deceptive lesions in 

dermatopathology. Probably it is the 

hottest issue in the area of pigmented lesions at the moment, 

given that atypical Spitz/nevi tumors 

are well studied and more often approached in practical work 

with a defensive attitude. 

The term nevoid melanoma was used by Schmoeckel, Castro 

and Braun-Falco in 1985 to describe primary cutaneous malignant 

melanomas with histological features suggestive of 

benign nevocytic nevi 1 . They stated that some of the following 

histological characteristics were always observed: cellular 

atypia, mitoses, adnexa infiltration in the deeper dermis, infiltrative 

growth, pigmented tumor cells, sharply demarcated 

tumor nests, and the absence of maturation. 

The clinical behavior of nevoid melanoma does not differ significantly 

from ordinary melanoma, and tumor thickness was 

the most important prognostic criterion. Lesions which share 

similar histological findings were called borderline melanoma 

by Reed, Clark and Mimh in 1975 2 . In 1985 the Mhim group 3 

initially described this subset of lesions with the term minimal 

deviation melanoma, but in 1995 they decided to use the more 

committal and popular term of nevoid melanoma, suggesting 

that proper attention to cytological detail and subtle architectural 

features will aid in recognizing this unusual variant of 

malignant melanoma 4 5 . Similar observations were made by 

Zembowicz et al. 5 

Three paradigmatic cases are discussed: 

1) a nevoid melanoma which turned out to be a benign nevus; 

2) a previous benign nevus which behaved as a melanoma; 

3) I don’t know-case. 

references 

1 Schmoeckel C, Castro CE, Braun-Falco O. Nevoid malignant melanoma. 

Arch Dermatol Res 1985;9:362-9. 

2 Reed RJ, Ichinose H, Clark WH Jr, et al. Common and uncommon melanocytic 

nevi and borderline melanomas. Sem Oncol 1975;2:119-47. 

3 Mérot Y, Mihm MC Jr. Unusual and unknown aspect of cutaneous 

malignant melanoma: minimal deviation malignant melanoma. Retrospective 

study of 4 cases. Ann Dermatol Venereol 1985;112:325- 

6. 

4 Wong TY, Duncan LM, Mihm MC Jr. Melanoma mimicking dermal 

Spitz’s nevus (“nevoid” melanoma). Semin Surg Oncol 1993;9:188- 

93. 

5 Wong TY, Suster S, Duncan LM, et al. Nevoid melanoma: a clinicopathological 

study of seven cases of malignant melanoma mimicking 

spindle and epithelioid cell nevus and verrucous dermal nevus. Hum 

Pathol 1995;26:171-9. 

6 Zembowicz A, McCusker M, Chiarelli C, et al. Morphological 

analysis of nevoid melanoma: a study of 20 cases with a review of the 

literature. Am J Dermatopathol 2001;23:167-75.

182 

The granulomatous pattern in skin diseases 

A.M. Cesinaro 

Department of Anatomic Pathology, Azienda Ospedaliero-Universitaria, 

Policlinico di Modena, Italia 

Background. The granulomatous reaction pattern is characterized 

by the presence of granulomata, i.e. collections of 

histiocytes or epithelioid histiocytes, in the dermis, with or 

without admixed multinucleated giant cells and other types 

of inflammatory cells. The granulomata can show peculiar 

arrangements, accessory features such as necrosis, suppuration, 

or necrobiosis, and the presence of organisms or foreign 

material. Based on the histological features, granulomata can 

be sub-classified in the following types: sarcoidal, with the 

classic “naked” appearance; tuberculoid, characterized by 

central “caseation” necrosis; necrobiotic, showing more loose 

arrangement and necrobiosis (collagenolysis); suppurative, 

featuring central collections of neutrophils; foreign body-type, 

in which foreign material, either exogenous or endogenous, 

is identifiable; xanthogranulomatous, characterized by histiocytes 

with foamy or pale cytoplasm and admixture of other 

inflammatory cells; a miscellanea of other conditions 1 . 

Case report nr. 1. A 28-years old woman, born in Philippines, 

presented a solitary, annular plaque on the lower leg, 

asymptomatic and slowly enlarging in the last few months. A 

4-mm punch biopsy was performed on the border of the lesion 

and sent for histological examination with a clinical diagnosis 

of granuloma annulare. The haematoxylin-eosin stained slide 

showed a granulomatous inflammatory infiltrate in the superficial 

and deep dermis, coupled to a moderate lymphocytic 

component admixed with few plasma cells. The granulomata 

were arranged mostly around vessels and encased a nerve, 

as highlighted by immunostaining for S-100 protein. Special 

stains (PAS, Grocott, Ziehl-Neelsen, Fite) failed to show microorganisms. 

PCR studies were not performed. A diagnosis 

suspicious for leprosy, tuberculoid type, was rendered. The 

patient was referred to a national centre for infectious diseases 

(Genoa) for further investigations. The diagnosis of leprosy, 

tuberculoid type, was confirmed. 

The presence of granulomata should always suggest to look 

for an infectious agent. It is also recommended to perform 

special stains on multiple sections. Despite exhaustive search, 

these stains can fail to demonstrate the presence of microorganisms. 

The presence of perineural granulomatous inflammation 

in this case strongly addressed toward the diagnosis 

of leprosy. 

Case report nr. 2. A 55-years old woman, living in Sicily, 

complained of erythematous plaques on face and trunk for 

2 years. Histological examination of a punch biopsy from 

the dorsum showed a granulomatous inflammatory infiltrate 

throughout the dermis, around vessels and also surrounding 

nerves. Special stains were negative. The pattern of distribution 

suggested an infectious disease, i.e. leprosy, but this possibility 

was excluded by further investigations. The clinical 

history of the patient allowed to achieve the right diagnosis: 

the woman suffered from hypogammaglobulinemia and biliary 

cirrhosis and had had a diagnosis of common variable immunodeficiency. 

Patients with immunodeficiency disorders 

can develop cutaneous lesions with a granulomatous reaction 


Slide seminar: Non-neoplastic skin diseases 

Moderators: C. Angeli (Vercelli), D. Massi (Firenze) 

pattern 2 . Moreover, a patient with congenital combined immunodeficiency 

has been reported, whose cutaneous lesions 

featured granulomata with perineural distribution 3 , analogously 

to the present case. 

Besides the two stereotypical cutaneous granulomatous diseases, 

i.e. granuloma annulare and necrobiosis lipoidica, the 

granulomatous reaction pattern in the skin can have several 

causes and associations. It can be due to the deposition of 

foreign material, or to prolonged sun-light exposure leading to 

actinic changes, such as the group of so-called elastolytic granulomata. 

It can be observed in a large number of infectious 

diseases (TBC and non tuberculous mycobacteriosis, leprosy, 

leishmaniasis, fungal infections). It can be related to systemic 

conditions, such as sarcoidosis, Crohn’s disease, Rosai-Dorfman 

disease, haematological disorders, immunologic disorders, 

and also to the use of certain drugs. On the other hand, 

it is known also that a skin disease characterized by a peculiar 

granulomatous pattern at histology, such as granuloma annulare, 

can show protean clinical manifestations 4 . Infrequently, 

mycosis fungoides features a granulomatous pattern that 

overlaps the histological characters of granuloma annulare. 

Only few subtle clues allow to make the differential diagnosis 

between the two diseases, and sometimes the differentiation is 

almost impossible and can only rely on molecular biology 5 . 

Moreover, granuloma annulare-like features can be observed 

in certain drug reactions, again with only subtle histological 

differences 6 . Finally, pathologists should be aware of the possibility 

that an apparently innocent granulomatous reaction can 

hide a life-threatening condition, such as lymphoma 7 . 

All these observations underline the importance of the clinicopathological 

correlations when one is dealing with a granulomatous 

reaction in the skin, since histology alone could not 

be sufficient and sometimes can also lead toward the wrong 

diagnosis. 

references 

1 Weedon D. Skin Pathology. 3 rd Ed. 

2 Mitra A, Pollock B, Gooi J, et al. Cutaneous granulomas associated 

with primary immunodeficiency disorders. Br J Dermatol 

2005;153:194-9. 

3 Krupnick AI, Shim H, Phelps RG, et al. Cutaneous granulomas 

masquerading as tuberculoid leprosy in a patient with congenital 

combined immunodeficiency. Mt Sinai J Med 2001;68:326-30. 

4 McKee PH, Calonje E, Granter SR. Pathology of the skin with clinical 

correlations. Vol. 1. 3 rd Ed. 

5 Su LD, Kim YH, LeBoit PE, et al. Interstitial mycosis fungoides, a 

variant of mycosis fungoides resembling granuloma annulare and 

inflammatory morphea. J Cutan Pathol 2002;29:135-41. 

6 Magro CM, Crowson AN, Shapiro BL. The interstitial granulomatous 

drug reaction: a distinctive clinical and pathological entity. J Cutan 

Pathol 1998;25:72-8. 

7 Scarabello A, Leinweber B, Ardigò M, et al. Cutaneous lymphomas 

with prominent granulomatous reaction: a potential pitfall in the histopathologic 

diagnosis of cutaneous T- and B-cell lymphomas. Am J 

Surg Pathol 2002;26:1259-68. 

Non-neoplastic skin diseases: Case n. 2 

G. Collina 

Bologna 

Clinical History. 30-year-old man showed coppery-red papules 

localized in the trunk and limbs. Previous clinical history

lectures 

was unremarkable and the patients was in good health. A 

papule present in the leg was biopsied. 

Histopathology. A sparse, mostly superficial, perivascular 

infiltrate made up overwhelmingly of lymphocytes arranged 

also in patchy lichenoid fashion that obscures focally the base 

of unevenly hyperplastic epidermis topped by parakeratosis in 

mounds staggered in the lower half of a stratum corneum is 

characteristic of secondary syphilis. Among the lymphocytes, 

especially in the immediate vicinity of venules of the superficial 

plexus, are numerous plasma cells. Vacuolar alteration in 

company with a sprinkling of lymphocytes along the dermoepidermal 

junction and a tad of spongiosis in loci within surface 

epidermis are also present. 

Diagnosis. Secondary syphilis 

Discussion. The case presented is an example of secondary 

syphilis occurring in a 30-year-old patient. Clinically, this 

could, conceivably, be a drug reaction, but the possibility can 

be excluded by the assessment of the rest of the integument, 

the results of the histology and of studies serologically. The 

lesion are papules mostly because of somewhat lichenoid arrangement 

of the infiltrate of lymphocyte and plasma cells 

and the peculiar orange hue is consequence, in part, of the 

combination of widely dilated venules which in vivo housed 

countless erythrocytes and the peculiar distribution of inflammatory 

cells, those two findings are present in the upper part 

of the dermis. 

The histological findings were those of a lichenoid-psoriasiform 

dermatitis in which plasma cells predominate. This was 

strongly suggestive of secondary syphilis. The diagnosis was 

confirmed by serology. We could not demonstrate Treponema 

pallidum (TP) on histological sections using Warthin Starry 

stain. 

The correlation between clinical and histopathological findings 

were crucial for achieving the correct diagnosis. 

Acquired syphilis caused by TP has affected humanity since at 

least the fifteen century, but the advent of penicillin reduced 

the incidence of the disease in the rich world so that many clinicians 

are nowadays unfamiliar with its signs and symptoms. 

Recently the incidence of syphilis is rising because is linked 

to the immunodeficiency virus infection. 

TP is generally spread to contact between infectious lesions 

ad disrupted epithelium at the site of minor trauma during the 

intercourse. The transmission rate is between 10% and 60%. 

The disease shows four clinical detectable phase. Primary 

syphilis is defined as the typical chancre that appear clinically 

as a regular edge, regular based, hard and bottom-like 

ulceration measuring up to one centimetre in diameter. Unless 

secondary infected, chancre is not painful. Multiple lesion 

may be present and 25% of patients (predominately woman) 

diagnosed at second-stage syphilis had no history of primary 

infection. As a rule, the chancre heals spontaneously in 3-8 

weeks and rarely persist for more than three months. Histologically, 

fully developed lesions are constituted by dense 

inflammatory infiltrate composed of lymphocytes, histiocytes 

and plasma cells. The blood vessels are increased in number 

and are bordered by plump endothelial cells. Oedema is a 

features in the upper dermis along with the ulcerations of the 

epidermis; this latter covered by fibrin and crust. 

Secondary syphilis results from the haematogenous dissemination 

of the TP, resulting in more widespread clinical signs 

accompanied by fever, malaise and generalized lymphoadenopathy. 

A generalized eruption can occur comprising orange 

maculae, papules and papulosquamous lesions resembling 

guttate psoriasis. Rarely pustules are present. The three most 

common histopatological patterns of secondary syphilis are 

183 

psoriasiform, lichenoid and psoriasiform-lichenoid and they 

occur in conjunction with superficial and deep perivascular 

infiltrate in which plasma cells predominate. Exocytosis of 

lymphocytes spongiform pustulations (which harbour TP) 

and parakeratosis may be seen. Parakeratosis may be broad 

or paltry. Granulomatous inflammation may be seen in older 

lesions. 

Primary and secondary lesions may be unnoticed by the patient 

who then passes in the latent phase of the disease. 

Tertiary syphilis is categorized into nodular tertiary syphilis 

confined to the skin; benign gummatous syphilis principally 

affecting skin, bone and liver; syphilitic hepatic cirrhosis, 

cardiovascular syphilis and neurosyphilis. 

The histopathological findings of tertiary syphilis are those of 

necrotizing granolumatous reaction. 

Secondary syphilis should be differentiated from other inflammatory 

or neoplastic disease of the skin. When the inflammatory 

infiltrate is particular heavy and lymphocytes show atypical 

features the possibility of mycosis fungoides may be suggested. 

In this latter condition lymphocytes predominate and 

plasma cells are usually absent. Mucha- Haberman disease 

is composed almost entirely by lymphocytes. Psoriasis lacks 

histiocytes and plasma cells and usually the inflammatory 

infiltrate is more superficial and does not involve the blood 

vessels of mid dermis. In syphilis the present of spongiform 

pustules may be observed, but nary attenuation of suprapapillary 

plate is a feature. 

Secondary syphilis should also be differentiated from all 

lichenoid dermatitis in which lymphocytes predominate such 

as lichenoid drug eruption, lichenoid photodermatitis and 

lichenoid discoid lupus erithematosus. In syphilis, except for 

the early second phase in which plasma cells may be paltry, 

even absent, the infiltrate being made up nearly exclusively of 

lymphocytes, the presence of plasma cells may be considered 

a signal of this venereal disease. 

Pityriasis lichenoides and cutaneous 

vasculitides 

C. Miracco 

Section of Pathological Anatomy- Department of Human Pathology 

and Oncology- Siena- Italy 

Background. Pityriasis lichenoides (PL) is an uncommon inflammatory 

skin disorder of unknown histogenesis, that may 

occur either in acute (pityriasis lichenoides et varioliformis 

acuta, PLEVA; and febrile ulceronecrotic Mucha-Habermann 

disease) or chronic (pityriasis lichenoides chronica, PLC) 

form. Acute PL is usually a self-limiting polymorphous eruption 

of macules, papules, and pustulae, which may evolve 

into hemorrhagic and necrotic lesions; recurrences over the 

years, as well as lethal febrile cases are not infrequent. PLC, 

the prolonged form of the disease, has a more indolent clinical 

course, with recurrent erythematous, scaling papules, tending 

to regress within some weeks. By some authors PL is classified 

among the cutaneous lymphocytic vasculitides, although 

classical signs of blood vessel damage are usually missing. PL 

by most is instead included among the interface-dermatitides, 

due to the heavy inflammatory infiltrate obscuring the dermalepidermal 

junction. No invasion of vessel walls by inflammatory 

cells is in fact observed in PLC; and non-vasculitic vessel 

changes are the rule in acute PL, in some lesions, however, a 

damage of vessel walls, with fibrinoid necrosis and leukocytoclasis 

may occur; in these cases, other histological findings 

are relevant to exclude a true skin vasculitic process.

184 

Cases and Methods. Two cases of PLEVA occurred in an 

8-year-old boy and in a 24-year-old man will be shown. In 

the first case, the diagnosis was supported by clinical data and 

diagnosis. In the second case, there was no clinical diagnosis, 

however a description of an eruption of papules and hemorrhagic 

lesions was given. The histological diagnosis was of 

PLEVA in both cases, based on the observation of a wedgeshaped 

inflammatory infiltrate, obscuring the dermal-epidermal 

junction, with variable degrees of keratinocyte damage, 

up to necrosis and epidermal ulceration, accompanied by 

dermal vessel alterations, with erythrocyte extravasation. 

Results and Discussion. Histological findings of PLEVA, 

as well as of other true skin vasculitides and other common 

forms of cutaneous pseudovasculitides will be shown and 

discussed for the differential diagnosis. On the one hand, vas- 


cular changes, including engorgement and oscuring of dilated 

dermal vessels by lymphocytes, and endothelial proliferation, 

as well as extravasation of erythrocytes in the dermis and epidermis, 

usually seen in acute PL, are helpful diagnostic clues 

in the differential diagnosis with other diseases characterized 

by an interface dermatitis. On the other hand, the wedgeshaped 

pattern of the lesion, with a dense, predominantly 

lymphocytic infiltrate at the dermal-epidermal junction, with 

lymphocyte exocytosis, and vacuolar alteration of the basal 

layer, are useful diagnostic criteria for the differential diagnosis 

with true vasculitides and pseudovasculitides. Clinics will 

also be shown and compared with histological findings. An 

haematoxylin and eosin stain, supported by clinical findings, 

is usually sufficient for a correct diagnosis, which is mandatory 

for an adequate treatment of PLEVA patients.

lectures 

WHO classification of tumours of 

haematopoietic and lymphatic tissues: 

methods, current issues, future perspectives 

S. Pileri 

Bologna 

As indicated above, there is no one “gold standard”, by which 

all diseases are defined in the WHO classification. Morphology 

is always important, and many diseases have characteristic 

or even diagnostic morphologic features. Immunophenotype 

and genetic features are an important part of the definition of 

these diseases, and the availability of this information makes 

arriving at consensus definitions much easier than when only 

subjective morphologic criteria were available. Immunophenotyping 

studies are used in routine diagnosis in the vast 

majority of haematologic malignancies, both to determine 

lineage in malignant processes and to distinguish benign from 

malignant processes. Many diseases have a characteristic 

immunophenotype, such that one would hesitate to make the 

diagnosis in the absence of the immunophenotype, while in 

others the immunophenotype is only part of the diagnosis. In 

some lymphoid and in many myeloid neoplasms a specific 

genetic abnormality is the key defining criterion, while others 

lack specific known genetic abnormalities. Some genetic 

abnormalities, while characteristic of one disease, are not specific 

(such as,, or rearrangements or mutations in), and others 

are prognostic factors in several diseases (such as mutations 

or). The inclusion of immunophenotypic features and genetic 

abnormalities to define entities not only provides an objective 

criterion for disease recognition but has identified antigens, 

genes or pathways that can be targeted for therapy; the success 

of rituximab, an anti-CD20 molecule, in the treatment of Bcell 

neoplasms, and of imatinib in the treatment of leukemias 

associated with ABL1 and other rearrangements involving 

tryosine kinase genes are testament to this approach. Finally, 

some diseases require knowledge of clinical features – age, 

nodal vs extranodal presentation, specific anatomic site, and 

history of cytotoxic and other therapies – to make the diagnosis. 

Most of the diseases described in the WHO classification 

are considered to be distinct entities; however, some are not 

as clearly defined, and these are listed as provisional entities. 

In addition, borderline categories have been created in this 

edition for cases that do not clearly fit into one category, so 

that well-defined categories can be kept homogeneous, and 

the borderline cases can be studied further. 

Hepatitis C virus (HCV) related lymphomas: a new 

model for lymphomagenesis 

M. Paulli, M. Lucioni, G. Fiandrino, M. Nicola, L. Arcaini * 

Pathology Section, Department of Human Pathology and * Division of 

Hemathology, University of Pavia, Foundation IRCCS Policlinico San 

Matteo, Pavia, Italy 

Immunodeficiency, autoimmunity and sustained activation of 

the lymphoid system, which frequently accompanies chronic 

friday, September 24 th , 2010 

Symposium on haemolymph pathology: the experience of the WHO 

Moderator: S. Pileri (Bologna) 

185 

infections, represent a risk factor for subsequent lymphoma 

development. Similarly, congenital and acquired immunodeficiencies 

associated with HIV infection and solid organ 

transplantation increase the risk of developing B-cell NHLs. 

On the other hand, autoimmune diseases such as Sjögren 

syndrome are also associated with an increased risk of lymphomas. 

The geographic heterogeneity in the incidence of 

B-cell non Hodgkin lymphomas (NHLs) suggests that also 

environmental factors such as infections might have a role in 

lymphomagenesis. 

In fact, particular lymphoma subtypes have been associated 

with specific microbial infections, which may promote 

lymphomagenesis by creating a favourable environment for 

transformation, with increased proliferation and decreased 

apoptosis of lymphoid cells, via direct or indirect mechanisms. 

Lymphotropic transforming viruses such as Epstein 

Barr virus (EBV), human herpes virus 8 (HHV8) and human 

T-lymphotropic virus 1 (HTLV-1) directly infect a subset of 

lymphoid cells in which they express viral oncogenes, with 

transforming abilities. An alternative scenario has emerged 

for microbial species that do not directly infect or transform 

lymphoid cells, but may persist chronically in host tissues and 

trigger a sustained lymphoid proliferation, giving a selective 

advantage to lymphoid clones that initially are still dependent 

upon antigenic stimulation. In this setting, additional oncogenic 

events may occur, leading the lymphoid proliferation 

to become independent of antigenic stimulation. The best 

documented model for indirect lymphomagenesis mediated 

by infectious agents is represented by Helicobacter pylori 

in gastric marginal zone lymphoma, but similar mechanisms 

have been more recently proposed for Borrelia burgdoferi in 

cutaneous B-cell lymphomas, Chlamydia psittaci in ocular 

adnexal lymphoma of mucosa-associated lymphoid tissue 

(MALT) and Hepatitis C virus (HCV) in several B-cell 

NHLs. 

HCV, a positive single-stranded RNA virus, belongs to the 

family of Flaviviruses; it affects millions of patients worldwide 

and represents a major burden in term of morbidity, mortality 

and social costs. Estimated prevalence in Italy is up to 10%, 

representing one of the highest rates among western countries. 

It is now well recognized that, in addition to hepatic manifestations, 

HCV infection is linked to a spectrum of cryoglobulinemic 

and non-cryoglobulinemic B-cell lymphoproliferative 

disorders. A meta-analysis showed that prevalence of HCV 

infection in both nodal or extranodal B-cell non-Hodgkin’s 

lymphoma (NHL) patients is 15% in comparison with 1.5% 

in the general population; this association is more evident in 

geographic areas with high HCV seroprevalence. A study in 

2004 estimated that for Italy, due to high seroprevalence rates, 

the attributable risk for HCV infection in lymphoma development 

would be up to 10% of all new cases. 

Reports in the literature dealing with lymphoproliferative 

disorders associated with HCV infection indicate variable incidences 

for the various lymphoma histotypes. This variability 

may reflect alternative classification approaches, differences 

in HCV infection rates among geographic areas and variable

186 

oncogenic potential and/or lymphoid tropism for different 

HCV serotypes, even if the latter hypothesis is most doubtful. 

In spite of some discrepancies, HCV-associated B-cell NHL 

seem to have distinctive clinicopathological features including 

a predilection for extranodal localizations and an overrepresentation 

of the diffuse large B-cell (DLBCL) and marginal 

zone (MZL) histological subtypes; some authors also reported 

an increased incidence in the female sex. 

The association of HCV infection with lymphoma of liver, 

salivary glands, and spleen was investigated by many Authors. 

Among MZL subtypes, we reported a stronger association 

with HCV infection in the splenic and nodal form as well 

as in non-gastric MALT lymphomas. Notably, the association 

of HCV with splenic MZL with villous lymphocytes has been 

proposed as a paradigmatic example of a HCV-driven lymphoproliferative 

disorder. 

Recently, presence of HCV infection has been also detected in 

cases of primary cutaneous B-cell lymphomas, irrespectively 

of clinicopathological subtype. Our group has also recently 

described a series of HCV-positive patients who developed 

a MZL, characterized by unusual lipoma-like subcutaneous 

presentation, and a relatively indolent clinical course. Extensive 

molecular and genetic investigations seem to indicate 

that HCV infection may play a causative role in this peculiar 

lymphoma subset. 

In addition to epidemiological studies, the possible pathogenetic 

role for HCV in mixed cryoglobulinemia and lymphoproliferative 

disorders have been confirmed by lymphoma 

regression following antiviral therapy, that has been observed 

in some patients. Antiviral therapy with interferon-α (IFNα) 

is known to be highly efficacious in the treatment of 

type II mixed cryoglobulinemia. As for NHL developing in 

HCV-positive patients, anti-viral therapy with IFN-α with or 

without ribavirin was reported to induce complete remission 

of the disease in up to 75% of cases irrespective of their histological 

subtype even if, altogether, the number of analyzed 

cases appears disproportionately low. High rates of molecular 

remission with disappearance of previously documented of 

IgH rearrangements and/or t(14;18) translocation have also 

been reported. 

From the biological point of view, it is known that HCV can 

infect B-cells in vitro; until now, however, there is a lack of 

evidence for a direct infection of lymphoid cells by HCV as 

a trigger for lymphoma development. In fact, only a fairly 

small subset of LNHs arising in HCV positive patients actually 

host viral genome; on the contrary, the virus itself has 

been detected in accompanying stromal cells of affected 

lymph nodes. In addition, HCV is a RNA virus lacking DNA 

intermediates in its replicative cycle and it seems unlikely that 

integration of HCV genome into the host might take place. 

Therefore, it appears more plausible that lymphoid antigendriven 

proliferation represents an early and facilitating event 

leading to lymphoma through an indirect pathway. A clue for 

this comes from the analysis of B-cell clones obtained from 

HCV-positive controls. Among the latter, IgH rearrangements 

(monoclonal or oligoclonal) are demonstrable in up to 

100% of patients with type II mixed cryoglobulinemia; approximately 

8 to 10% of all type II mixed cryoglobulinemia 

patients actually will develop NHL after a long follow-up. 

Additionally, recombinant E2 viral protein exposed on HCV 

virion surface was demonstrated as sufficient to induce hypermutations 

at the immunoglobulin locus when binding to 

CD81 coreceptor of B lymphocytes. Finally, a biased usage 

of IGH genes was documented in HCV-associate lymphomas 


and a certain degree of homology was detected between B-cell 

receptors (BCR) and anti-viral antibodies in HCV-positive 

NHL patients. 

In conclusion epidemiologic data, molecular findings and 

clinical evidence of lymphoma regression after antiviral therapy 

seem to indicate that HCV may play a causative role in 

some B-cell NHLs. Some recent clinico-pathological reports 

confirm that lymphomas associated with HCV infection may 

have peculiar clinical features (such as subcutaneous presentation 

mimicking lipoma), that may deserve particular attention 

in order to be properly recognized. 

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lectures 

Burkitt lymphoma, fifty years plus: the 

beginning and the future 

E. Rogena * ** , S. Lazzi * , G. De Falco * , M.R. Ambrosio * , 

M. Onorati * , B.J. Rocca * , C. Bellan * , PP. Piccaluga *** , P. 

Pileri *** , L. Leoncini * 

* Department of Human Pathology and Oncology, Anatomic Pathology 

Section, University of Siena, Italy; ** UON/KNH, Nairoby, Kenya; 

*** Department of Haeematopathology and Oncology “L. & A. Seràgnoli”, 

University of Bologna, Italy 

Introduction. The first clinical records of Burkitt lymphoma 

(BL) are found in the Mengo Hospital case-notes of Sir Albert 

Cook at the beginning of the 20 th century. Sporadic reports 

of tumours resembling BL, but often reported as atypical 

neuroblastomas, appeared in publications from Africa in the 

first half of the century. Denis Burkitt’s seminar publication 

in 1957 not only described the clinical features of this tumour 

but showed that the jaw tumours and the visceral tumours 

were part of the same disease entity. 

Burkitt lymphoma (BL) is now listed in the World Health 

Organization (WHO) classification of lymphoid tumors as a 

B-cell lymphoma with an extremely short doubling time that 

often presents in extranodal sites or as an acute leukaemia. 

It is composed of monomorphic medium-sized transformed 

cells. Translocation involving MYC is the most common 

genetic alteration. A combination of several diagnostic techniques 

(morphology, genetic analysis or immunophenotyping) 

is necessary for the diagnosis of BL. 

Some clinical aspects of BL. Three clinical variants are recognized: 

endemic BL (eBL), sporadic BL (sBL), and immunodeficiency-associated 

BL (ID-BL). Each clinical subset affects 

different populations and can present in different forms. 

Endemic BL occurs in equatorial Africa, representing the 

most common childhood malignancy with an incidence peak 

at 4 to 7 years and a male:female ratio of 2:1. EBV is present 

in 90-95% of the neoplastic cells in most of the patients; the 

jaw is the more frequent site of presentation. Sporadic BL 

is seen throughout the world, mainly in children and young 

adults. The incidence is low, representing only 1-2% of all 

lymphomas in Western Europe and in USA. The male: female 

ratio is 2 or 3:1. EBV may be detected in 30-40% of cases and 

the classical presentation is with a bulky disease involving the 

distal ileum/proximal cecum. Immunodeficiency-associated 

BL is primarily seen in association with the human immunodeficiency 

virus (HIV) infection, often occurring as the initial 

manifestation of the acquired immunodeficiency syndrome 

(AIDS). EBV is identified only in 25-40% of cases. ID-BL 

often presents with bulky lymph node involvement. 

However, cases with clinical features and presentation typical 

of sporadic forms, occurring in adults and being HIV and 

EBV negative, have been reported also in endemic areas. 

BL is an highly aggressive tumour and most patients present 

with advanced clinical stage (bulky disease with a high 

tumour burden, infiltration of the bone marrow, extracerebral 

nervous system and liver) but it is potentially curable and 

intensive chemotherapy leads to up to 90% cure rates in low 

stage disease and 60-80% in patients with advanced disease. 

The prognosis is better in children than in adults. Relapse 

usually occurs in the first year after diagnosis. Although the 

tumour is curable, many patients still die of the disease mainly 

in Africa. 

Role of infectious agents and environmental factors in 

the pathogenesis of BL. In Africa Burkitt’s lymphoma is 

endemic in wet regions with mean minimum temperatures 

> 15,5°C and annual rainfall. The question as to why Burkitt’s 

187 

lymphoma (BL) is endemic in equatorial Africa has intrigued 

scientists since the first clinical description of this neoplasm 

by Denis Burkitt. Investigation of the geographic restriction 

demonstrated that BL occurred primarily where there was 

sustained and intense exposure to holoendemic malaria. This 

gave rise to the hypothesis that the tumour might be caused by 

a mosquito borne virus. Following an intensive search for this 

virus Antony Epstein and his colleagues later discovered the 

Epstein Barr virus in biopsy samples of BL sent from Uganda. 

Recent advances in virology, parasitology and immunology 

have helped to elucidate the important contributions of malarial 

infection and Epstein-Barr virus to lymphomagenesis 

in African endemic Burkitt’s lymphoma. P. falciparum infection 

suppresses cytotoxic T cells immunity against EBV. The 

importance of cytotoxic T cells in controlling EBV infections 

has been well established and it has long been hypothesized 

that EBV immunity is suppressed as a consequence of repeated 

malaria infections. However, the means by which malaria 

orchestrate deficiencies in anti-EBV immunity that result in 

tumorigenesis have not been so well explained. Recent studies 

have demonstrated that malaria-induced T cell disfunction 

is age-dependent, transient, EBV-specific, and differentially 

affects EBV-specific T cell memory subsets. In addition, 

through interaction with the Toll like receptor (TLR)9, P. falciparum 

infection may lead to perturbation of peripheral B 

cell subsets and reactivation and expansion of latently infected 

memory B cells, where the virus persist in healthy carriers. 

In fact, according to recent studies, EBV first infect naïve B 

cells and activates a growth program in these cells so they 

can differentiate into resting memory B cells or plasma cells 

through the process of the germinal center (GC) reaction. The 

virus thus gains access to the memory B cell compartment, its 

main reservoir during persistence, where no latent viral genes 

are expressed. However, when these latently infected memory 

cells divide, they express the EBNA-1 protein (latency I), the 

same viral latency program as found in BL primary tumours. 

Thus, it is then reasonable to argue that the cell of origin of 

endemic BL may be the memory B cells. This is in accordance 

also with a recent investigation suggesting that EBV-negative 

tumours derive from early centroblasts, whereas EBV 

positive cases derive from memory or late germinal center B. 

However, this is in contrast with the GC phenotype shared by 

all of the BL variants. This discrepancy may be explained by 

the assumption that in EBV-positive BL B cells follow the 

normal differentiation process of the GC as they reach the 

differentiation stage of memory cells in terms of VH gene 

mutation but do not follow the normal differentiation process 

in terms of morphology, immunophenotype and gene expression. 

This process is primed by BCL6 and requires the activation 

of BLIMP-1, which in turn represses c-MYC and BCL6 

genes. Recent observations have indicated that BCL6 and 

BLIMP-1 are targeted by different microRNAs (miRNAs), a 

class of small non-coding RNAs acting as post-transcriptional 

regulators of gene expression. Interestingly, recent data give 

evidence that hsa-miR-127, which is the master regulator of 

B cell fate through the interaction with BLIMP-1 and XBP1, 

is strongly up-regulated only in EBV-positive BL, whereas 

EBV-negative cases show levels of expression similar to GC 

cells and reactive lymph nodes. It is not currently known how 

EBV can regulate miRNAs expression but preliminary results 

confirm the existence of an active interplay between EBV 

gene product and cellular miRNAs. 

However, epidemiological studies suggest that malaria and 

EBV alone cannot account for the distribution of endemic BL 

in high risk regions. Selenium, arboviruses and plant extracts

188 

are additional environmental factors that appear to be important 

in the pathogenesis of eBL. It has been recently postulated 

a possible role of Euphobia tirucalis, a plant commonly used 

in the traditional medicine, as a cofactor in the development 

of eBL by its modulation of the expression of the latency 

genes of EBV, and the up-regulation of anti-apoptotic factor 

as BCL-2. 

The incidence of non-Hodgkin’s lymphoma (NHL) is greatly 

increased in HIV-infected individuals, being the second most 

common neoplasm (after Kaposi’s sarcoma). BL occurs in 

less immunodeficient patients and develops when the CD4 

count is relatively high, being the immunosuppression per se 

not sufficient to explain the relatively high prevalence of BL 

in this setting. Tat protein of HIV is a likely candidate to contribute 

to tumour pathogenesis in HIV-infected patients. Tat is 

an early non-structural protein necessary for virus replication 

which is secreted by infected cells and taken up by uninfected 

cells. Deregulation of cellular genes and functions by Tat can 

cause abnormalities that may contribute to AIDS pathogenesis 

and to the development of AIDS-associated disorders. The 

molecular mechanism underlying Tat’s pleiotropic activity 

may include the generation of functional heterodimers of Tat 

with cell cycle proteins, resulting in uncontrolled cell proliferation. 

Another mechanism, through which Tat may influence 

HIV-mediated transformation, is by hyper-activation 

of transcription by interacting with chromatin remodelling 

complexes. In addition, HIV and EBV through viral-encoding 

miRNAs may interfere with the physiological regulation of 

cell functions maintained by cellular miRNAs. 

Morphology and molecular signature of BL. BL classically 

shows a monomorphic, medium-sized cells diffusely infiltrating 

into the tissue, with a starry sky pattern. The stars representing 

tingible body histiocytes and the sky representing the 

neoplastic lymphoid cells. The lymphoid cells show a regular 

cytoplasmic border (non-cleaved), a non-vesicular nucleus 

with two or four basophilic nucleoli. Mitoses are frequent, up 

to 4%. The proliferation index as shown by Ki-67 is almost 


always greater than 95%. The characteristic immunophenotype 

of BL is CD 19, CD 20, CD22 (B cell associated antigens) 

positive; CD10, BCL6, CD38, CD 77, CD 43 positive. 

The cells are usually negative or weekly positive for BCL2 

and Tdt-negative. Most tumours show MYC translocation at 

band 8q24 to the Ig heavy chain region 14q32 or less commonly 

22q11 or kappa, 2p12 light chain loci. Up to 10% of 

cases may lack a demonstrable MYC translocation by FISH. 

These tumors show a similar expression of MYC as the cases 

carrying the typical translocation. There are several explanations 

for this, such as the fact that the break point can occur 

in different regions that cannot be recognized by commercial 

probes. Whatever the genetic mechanism may be, these cases 

have a different molecular pattern due to miRNA deregulation 

involved in MYC pathway. This supports the hypothesis that 

is the over-expression of MYC independently by the genetic 

mechanism that is important for the neoplastic transformation. 

Furthermore, it should be considered that MYC translocation 

in not specific for BL. Additional alterations may be then responsible 

for the typical BL signature. Other genetic and epigenetic 

aberrations, occurring in a subset of BL, can involve 

p16, TP53, P73, BAX, P130/RB2, and BCL6. 

Gene expression profile (GEP) studies have demonstrated a 

consistent gene expression signature for BL, which is clearly 

distinct from that of diffuse large B cell lymphoma (DLBCL), 

but intermediate cases were also found. In addition, preliminary 

studies have shown also difference in GEP among 

the clinical variants of sBL, eBL and ID-BL, having similar 

GEP different from sBL. Of note, these differences regard 

significant cellular pathways probably reflecting the different 

pathogenetic mechanisms. 

A look into the future. The different findings, point out at 

the importance of a more discriminative and accurate GEP, as 

that of miRNAs, to clarify differences between BL subtypes, 

to better explain the pathogenetic mechanisms involved in 

virus-associated diseases, and eventually to help designing 

more tailored treatments. 

Neoplastic pathologies of the bladder and prostate 

2009 International Society of urological 

Pathology consensus conference 

on standardization of pathology reporting 

of radical prostatectomy specimens 

R. Montironi, A. Lopez-Beltran * , L. Cheng ** 

Section of Pathological Anatomy, Polytechnic University of the Marche 

Region, School of Medicine, United Hospitals, Ancona, Italy; 

* Department of Pathology, Reina Sofia University Hospital and Faculty 

of Medicine, Cordoba, Spain; ** Department of Pathology and 

Laboratory Medicine, Indiana University School of Medicine, Indianapolis, 

IN, USA 

The 2009 International Society of Urological Pathology 

consensus conference in Boston, made recommendations regarding 

the standardization of pathology reporting of radical 

prostatectomy specimens. 

Issues relating to the handling and processing of radical prostatectomy 

specimens were coordinated by working group 1. 

Most uropathologists followed similar procedures for fixation 

Moderators: G. Mikuz (Innsbruck), R. Montironi (Ancona) 

of radical and prostatectomy specimens, with 51% of respondents 

transporting tissue in formalin. There was also consensus 

that the prostate weight without the seminal vesicles should be 

recorded. There was consensus that the surface of the prostate 

should be painted. It was agreed that both the prostate apex 

and base should be examined by the core method with sagittal 

sectioning of the tissue sample. There was consensus that the 

gland should be fully fixed before sectioning Both partial or 

complete embedding of prostates was considered to be acceptable 

as long as the method of partial embedding is stated. No 

consensus was determined regarding the necessity of weighing 

and measuring the length of the seminal vesicles, the 

preparation of whole mounts rather than standardized blocks 

and the methodology for sampling of fresh tissue for research 

purposes, and it was agreed that these should be left to the 

discretion of the working pathologist. 

Issues relating to the substaging of pT2 prostate cancers according 

to the TNM 2002/2009 system, reporting of tumor 

size/volume and zonal location of prostate cancers were coor-

lectures 

dinated by working group 2. A survey circulated prior to the 

consensus conference demonstrated that 74% of the 157 participants 

considered pT2 substaging of prostate cancer to be of 

clinical and/or academic relevance. The survey also revealed 

a considerable variation in the frequency of reporting of pT2b 

substage prostate cancer which was likely a consequence of 

the variable methodologies used to distinguish pT2a from 

pT2b tumors. Overview of the literature indicates that current 

pT2 substaging criteria lack clinical relevance and the majority 

of conference attendees expressed dissatisfaction with the 

current pT2 substaging system. Despite this, no consensus 

was achieved relating to standardization of the method used 

to distinguish the various pT2 substages. For these reasons it 

was agreed that reporting of pT2 substages should, at present, 

be optional. Several studies have shown that prostate cancer 

volume is significantly correlated with other clinico-pathological 

features including Gleason score and extraprostatic 

extension of tumor; however, most studies fail to demonstrate 

this to have prognostic significance on multivariate analysis. 

Consensus was reached with regard to the reporting of some 

quantitative measure of the volume of tumor in a prostatectomy 

specimen, without prescribing a specific methodology. 

Incorporation of the zonal and/or anterior location of the 

dominant/index tumor in the pathology report was accepted 

by most participants, but a formal definition of the identifying 

features of the dominant/index tumor remained undecided. 

Issues relating to extraprostatic extension (pT3a disease), bladder 

neck invasion, microvascular invasion, and the definition 

of pT4 were coordinated by working group 3. It was agreed 

that prostate cancer can be categorized as pT3a in the absence 

of adipose tissue involvement when cancer bulges beyond the 

contour of the gland or beyond the condensed smooth muscle 

of the prostate at posterior and posterolateral sites. Extraprostatic 

extension can also be identified anteriorly. It was 

agreed that the location of extraprostatic extension should be 

reported. While there was consensus that the amount of extraprostatic 

extension should be quantitated, there was no agreement 

as to which method of quantitation should be employed. 

There was overwhelming consensus that microscopic urinary 

bladder neck involvement by carcinoma should be reported as 

stage pT3a and that lymphovascular invasion by carcinoma 

should be reported. It is recommended that these elements be 

considered in the development of practice guidelines and in 

the daily practice of urologic surgical pathology. 

Issues relating to the infiltration of tumor into the seminal 

vesicles and regional lymph nodes were coordinated by working 

group 4. There was a consensus that complete blocking of 

the seminal vesicles was not necessary, though sampling of 

the junction of the seminal vesicles and prostate was considered 

to be mandatory. Sampling of the vas deferens margins 

was not considered as obligatory. There was consensus that 

muscular wall invasion of the extraprostatic seminal vesicle 

only should be considered as seminal vesicle invasion. Categorization 

into types of seminal vesicle spread was considered 

not to be necessary. For examination of lymph nodes, 

special techniques such as frozen sectioning were considered 

to be of use only in high risk cases. There was no consensus 

on the optimal sampling method, though it was agreed that all 

lymph nodes should be completely blocked as a minimum. It 

was also agreed that a count of the number of lymph nodes 

harvested should be attempted. It was agreed that in view of 

recent evidence, the diameter of the largest lymph node metastasis 

should be measured. 

Issues relating to surgical margin assessment were coordinated 

by working group 5. Pathologists agreed that tumor 

189 

extending close to the “capsular” margin, yet not to it, should 

be reported as a negative margin, and that locations of positive 

margins should be indicated as either posterior, posterolateral, 

lateral, anterior at the prostatic apex, mid-prostate or base. 

Other items of consensus included specifying the extent of 

any positive margin as milimeters of involvement; tumor in 

skeletal muscle at the apical perpendicular margin section, in 

the absence of accompanying benign glands, to be considered 

organ confined; and that proximal and distal margins be uniformly 

referred to as bladder neck and prostatic apex respectively. 

Grading of tumor at positive margins was to be left 

to the discretion of the reporting pathologists. There was no 

consensus as to how the surgical margin should be regarded 

when tumor is present at the inked edge of the tissue, in the 

absence of transected glands at the apical margin. Pathologists 

also did not achieve agreement on the reporting approach to 

benign prostatic glands at an inked surgical margin where no 

carcinoma is present. 

Variants and variations of bladder cancer 

A. Lopez-Beltran 

Cordoba University Medical School, Cordoba, Spain 

Urothelial carcinoma has a propensity for divergent differentiation 

with the most common being squamous, followed 

by glandular. Virtually the whole spectrum of bladder cancer 

variants may be seen in variable proportions accompanying 

otherwise typical urothelial carcinoma. The clinical outcome 

of some of these variants differs from typical urothelial carcinoma; 

therefore, recognition of these variants is important 

Urothelial carcinoma with mixed differentiation. About 

20% of urothelial carcinomas contain areas of glandular or 

squamous differentiation. Squamous differentiation, defined 

by the presence of intercellular bridges or keratinization, 

occurs in 21% of urothelial carcinomas of the bladder. Its 

frequency increases with grade and stage. Detailed histologic 

maps of urothelial carcinoma with squamous differentiation 

have shown that the proportion of the squamous component 

may vary considerably, with some cases having urothelial 

carcinoma in situ as the only urothelial component. These 

cases may have a less favorable response to therapy than 

pure urothelial carcinoma. Of 91 patients with metastatic 

carcinoma, 83% with mixed adenocarcinoma and 46% with 

mixed squamous cell carcinoma experienced disease progression 

despite intense chemotherapy, whereas it progressed in 

< 30% of patients with pure urothelial carcinoma. Low-grade 

urothelial carcinoma with focal squamous differentiation has a 

higher recurrence rate. Tumors with any identifiable urothelial 

element are classified as urothelial carcinoma with squamous 

differentiation, and an estimate of the percentage of squamous 

component should be provided. Cytokeratin 14, L1 antigen 

and Caveolin-1 have been reported as immunohistochemical 

markers of squamous differentiation. 

Glandular differentiation is less common than squamous 

differentiation and may be present in about 6% of urothelial 

carcinomas of the bladder. Glandular differentiation is defined 

as the presence of true glandular spaces within the tumor. 

These may be tubular or enteric glands with mucin secretion. 

A colloid-mucinous pattern characterized by nests of cells 

“floating” in extracellular mucin, occasionally with signet 

ring cells, may be present. Cytoplasmic mucin-containing 

cells are, present in 14-63% of typical urothelial carcinoma 

and are not considered to represent glandular differentiation. 

The diagnosis of adenocarcinoma is reserved for pure tumors.

190 

A tumor with mixed glandular and urothelial differentiation 

is classified as urothelial carcinoma with glandular differentiation, 

and an estimate of the percentage of glandular 

component should be provided. The expression of MUC5ACapomucin 

may be useful as immunohistochemical marker of 

glandular differentiation in urothelial tumors. When small 

cell carcinoma is present in association with urothelial carcinoma, 

even focally, it portends a poor prognosis. Small cell 

carcinoma is an important finding and usually dictates more 

aggressive therapy (see following section). 

Small cell carcinoma. Small cell carcinoma is a malignant 

neuroendocrine neoplasm derived from the urothelium which 

histologically mimics its pulmonary counterpart. Patients with 

small cell carcinoma of the urinary bladder have a dismal prognosis. 

In a recent series of 64 cases of small cell carcinoma of 

the urinary bladder, the mean age at diagnosis was 66 years 

and the male to female ratio was 3.3:1; 88% presented with 

hematuria. All the patients except one had muscle-invasive 

disease at presentation. Thirty-eight patients (59%) underwent 

cystectomy and 66% of patients had lymph node metastasis at 

the time of cystectomy with regional lymph nodes, bone, liver 

and lung being the most common locations. Twenty cases 

(32%) were pure small cell carcinoma; 44 cases (68%) cases 

consisted of small cell carcinoma with other histological types 

(urothelial carcinoma, 35 cases; adenocarcinoma, 4 cases; 

sarcomatoid urothelial carcinoma, 2 cases; and 3 cases with 

both adenocarcinoma and urothelial carcinoma). Patients with 

organ-confined cancers had marginally better survival than 

those with non organ-confined cancer (p = 0.06). Overall, 1year, 

18-month, 3-year, and 5-year cancer-specific survivals 

were 56%, 41%, 23%, and 16%, respectively. 

At histology, they consist of small, rather uniform cells, with 

nuclear molding, scant cytoplasm and nuclei containing finely 

stippled chromatin and inconspicuous nucleoli. Mitoses are 

present and may be frequent. Necrosis is common and there 

may be DNA encrustation of blood vessels walls (Azzopardi 

phenomenon). Most cases have areas of urothelial carcinoma 

sometimes in the form of flat urothelial carcinoma in situ and 

exceptionally, squamous cell carcinoma, adenocarcinoma or 

sarcomatoid carcinoma. This is important, because the presence 

of these differentiated areas does not contradict the diagnosis 

of small cell carcinoma. Neuroendocrine granules are 

found with electron microscopy, but the immunohistochemical 

profile reveals neuronal-specific enolase in 87% of cases, and 

chromogranin A only in a third of cases. Some cases are also 

reactive against synaptophysin, PGP 9.5, thyroid transcription 

factor-1 (TTF-1), p53 (DO7), and Ki67 (MIB-1). The diagnosis 

of small cell carcinoma can be made on morphologic 

grounds alone, even if neuroendocrine differentiation cannot 

be demonstrated. Frequently small cell carcinoma expresses 

cytokeratin which supports the hypothesis of urothelial origin. 

The recent finding of c-kit and c-erbB2 expression by immunohistochemistry 

opens new possibilities for therapy in small 

cell carcinoma of the bladder. 

Nested variant. The nested variant of urothelial carcinoma 

is an aggressive neoplasm with fewer than 50 reported cases. 

There is a marked male predominance, and 70% of patients 

died 4-40 months after diagnosis, in spite of therapy. This rare 

pattern of urothelial carcinoma was first described as a tumor 

with a “deceptively benign” appearance that closely resembles 

Brunn’s nests infiltrating the lamina propria. Some nests have 

small tubular lumens that eventually can predominate. Nuclei 

generally show little or no atypia, but invariably the tumor 

contains foci or unequivocal cancer with cells exhibiting enlarged 

nucleoli and coarse nuclear chromatin. This feature is 


most apparent in the deeper aspects of the cancer. The differential 

diagnosis of the nested variant of urothelial carcinoma 

includes prominent Brunn’s nests, cystitis cystica and glandularis, 

inverted papilloma, nephrogenic metaplasia, carcinoid 

tumor, paraganglionic tissue, and paraganglioma. 

Micropapillary carcinoma. Micropapillary carcinoma is a 

distinct variant of urothelial carcinoma that resembles papillary 

serous carcinoma of the ovary, and approximately 60 

cases were reported in the literature. There is a male predominance 

and the patients ages range from fifth to the ninth decade 

with a mean age of 66 years. The most common presenting 

symptom is hematuria. The first description of micropapillary 

carcinoma consisted of 18 patients whose ages ranged from 

47 to 81 years (mean 67) with a male-to-female ratio of 5:1. 

Seven patients died of carcinoma. The micropapillary component 

is found in association with noninvasive papillary or 

invasive urothelial carcinoma in 80% of reported cases, consisting 

of slender delicate filiform processes or small papillary 

clusters of tumor cells; when present in invasive carcinoma, 

it is composed of infiltrating tight clusters of micropapillary 

aggregates that are often within lacunae that are negative for 

endothelial markers. Twenty-five percent of cases show glandular 

differentiation, and some authors consider it as a variant 

of adenocarcinoma. 99 Psammoma bodies are infrequent. 

Vascular and lymphatic invasion is common, and most cases 

show invasion of the muscularis propria or deeper, often with 

metastases. Immunohistochemical studies in one large series 

disclosed immunoreactivity of the micropapillary carcinoma 

in 20 of 20 cases for MUC1 (EMA), Cytokeratin 7 and 20, 

and Leu M-1. The presence of a surface micropapillary component 

in bladder biopsy specimens with cancer is an unfavorable 

prognostic feature, and deeper biopsies may be useful to 

determine the level of muscle invasion. The main differential 

consideration is serous micropapillary ovarian carcinoma in 

women or mesothelioma in both genders. 

Microcystic carcinoma. The microcystic variant of invasive 

urothelial carcinoma is characterized by the formation of microcysts, 

macrocysts, or tubular structures with cysts ranging 

from microscopic up to 1-2 cm in diameter. The cysts and 

tubules may be empty or contain necrotic debris or mucin that 

stains with periodic acid-Schiff stain with diastase predigestion. 

This variant of cancer may be confused with benign 

proliferations such as florid polypoid cystitis cystica and 

glandularis and nephrogenic metaplasia. 

Lymphoepithelioma-like carcinoma. Carcinoma that histologically 

resembles lymphoepithelioma of the nasopharynx 

has recently been described in the urinary bladder, with fewer 

than 40 cases reported. Disease in the urinary bladder is more 

common in men than in women (3:1 ratio) and occurs in 

late adulthood (range: 52-81 years; mean: 69 years). Most 

patients present with hematuria. The tumor is solitary and 

usually involves the dome, posterior wall, or trigone, often 

with a sessile growth pattern. Histologically, it may be pure or 

mixed with typical urothelial carcinoma, the later being focal 

and inconspicuous in some instances. Glandular and squamous 

differentiation may be seen. The tumor is composed of 

nests, sheets, and cords of undifferentiated cells with large 

pleomorphic nuclei and prominent nucleoli. The cytoplasmic 

borders are poorly defined, imparting a syncytial appearance. 

The background consists of a prominent lymphoid stroma 

that includes T and B lymphocytes, plasma cells, histiocytes, 

and occasional neutrophils or eosinophils. Epstein-Barr virus 

infection has not been identified in lymphoepithelioma-like 

carcinoma of the bladder. This tumor, thus far has been found 

to be responsive to chemotherapy when it is encountered in

lectures 

its pure form. The epithelial cells of this tumor stain with several 

cytokeratin markers as follows: AE1/AE3, CK8, CK 7, 

and they are rarely positive for CK20. The major differential 

diagnostic considerations are poorly differentiated urothelial 

carcinoma with lymphoid stroma, poorly differentiated squamous 

cell carcinoma, and lymphoma. Immunohistochemistry 

reveals cytokeratin immunoreactivivity in the malignant cells, 

confirming their epithelial nature. Most reported cases of the 

urinary bladder had a relatively favorable prognosis when pure 

or predominant, but when lymphoepithelioma-like carcinoma 

is focally present in an otherwise typical urothelial carcinoma, 

the disease behaves as it does in patients with conventional 

urothelial carcinoma of the same grade and stage. 

Plasmacytoid carcinoma. Zukerberg et al. described bladder 

carcinoma in two patients that diffusely permeated the 

bladder wall and was composed of cells with a monotonous 

appearance mimicking lymphoma. The tumor cells were medium-sized, 

with eosinophilic cytoplasm and eccentric nuclei 

producing a plasmacytoid appearance. The epithelial nature 

of the malignancy was confirmed by immunohistochemistry. 

Differential diagnostic considerations include lymphoma 

(plasmacytoid type) and multiple myeloma. Identification 

of an epithelial component confirms the diagnosis. In a series 

report of 6 cases, the male-to female ratio was 2:1, and 

the age range was 54-73 years. All cases stained positively 

for cytokeratin cocktail, cytokeratin 20 and 7, and all were 

negative for leukocyte common antigen. Five of six patients 

died of disease (mean survival, 23 months). Some case may 

express CD138. 

Inverted papilloma-like carcinoma. The potential for misinterpretation 

of urothelial carcinoma with endophytic growth 

as inverted papilloma is high. By definition, this variant of 

urothelial carcinoma has significant nuclear pleomorphism, 

mitotic figures, and architectural abnormalities consistent 

with low- or high-grade urothelial carcinoma. In most cases, 

the overlying epithelium has similar abnormalities and often 

contains typical urothelial carcinoma. Inverted papilloma-type 

carcinoma with minimal cytologic and architectural abnormalities 

have high mitotic activity. An exophytic papillary 

or invasive component is often associated with the inverted 

element. However, in cases of inverted papilloma fragmented 

during transurethral resection, a pseudoexophytic pattern 

may result. In some instances, both inverted papilloma and 

inverted papilloma-type carcinoma are intimately mixed. 

Large papillary tumors with prominent endophytic growth 

“invade” the lamina propria with a pushing border. Unless this 

pattern is accompanied by true destructive stromal invasion 

the likelihood of metastasis is minimal, because the basement 

membrane is not truly breached. 

Urothelial carcinoma with syncytiotrophoblastic giant 

Cells. Syncytiotrophoblastic giant cells are present in up to 

12% of cases of urothelial carcinoma, producing substantial 

amounts of immunoreactive beta-human chorionic gonadotropin 

(HCG) indicative of syncytiotrophoblastic differentiation. 

The number of HCG-immunoreactive cells is inversely associated 

with cancer grade. Secretion of HCG into the serum 

may be associated with a poor response to radiation therapy. 

The most important differential diagnostic consideration is 

choriocarcinoma; most but not all cases previously reported 

as primary choriocarcinoma of the bladder represent urothelial 

carcinoma with syncytiotrophoblasts. 

Pleomorphic Giant cell carcinoma. High grade urothelial 

carcinoma may contain epithelial tumor giant cells or the 

tumor may appear undifferentiated, resembling giant cell 

carcinoma of the lung. This variant is very infrequent. Malig- 

191 

nant giant cells in urothelial carcinoma, when present in great 

numbers, portend a poor prognosis, similar to that associated 

with giant cell carcinoma in the lung. The giant cells display 

cytokeratin and vimentin immunoreactivity. 

Clear cell (glycogen-rich) carcinoma. Up to two-thirds of 

cases of urothelial carcinoma have foci of clear cell change resulting 

from abundant glycogen. The glycogen-rich clear cell 

“variant” of urothelial carcinoma, recently described, appears 

to represent the extreme end of the morphologic spectrum, 

consisting predominantly or exclusively of cells with abundant 

clear cytoplasm that stains for cytokeratin 7. 

Sarcomatoid carcinoma with/without heterologous elements 

(carcinosarcoma, metaplastic carcinoma). The term 

sarcomatoid variant of urothelial carcinoma should be used 

for all biphasic malignant neoplasms exhibiting morphologic 

and/or immunohistochemical evidence of epithelial and mesenchymal 

differentiation (with the presence or absence of 

heterologous elements acknowledged in the report). There 

is considerable confusion and disagreement in the literature 

regarding nomenclature and histogenesis of these tumors. In 

some series, both carcinosarcoma and sarcomatoid carcinoma 

are included as “sarcomatoid carcinoma”. In others they are 

regarded as separate entities. 

The gross appearance is characteristically “sarcoma-like,” dull 

gray with infiltrative margins. The tumors are often polypoid 

with large intraluminal masses. Microscopically, sarcomatoid 

carcinoma is composed of urothelial, glandular or small cell 

component showing variable degrees of differentiation. Carcinoma 

in situ is present in 30% of cases and occasionally 

is the only apparent epithelial component. A small subset of 

sarcomatoid carcinoma may have a prominent myxoid stroma. 

The mesenchymal component most frequently observed is a 

undifferentiated high grade spindle cell neoplasm. The most 

common heterologous element is osteosarcoma followed 

by chondrosarcoma, rhabdomyosarcoma, leiomyosarcoma, 

liposarcoma, angiosarcoma or multiple types of heterologous 

differentiation may be present. By immunohistochemistry, 

epithelial elements react with cytokeratins, whereas stromal 

elements react with vimentin or specific markers corresponding 

to the mesenchymal differentiation. The sarcomatoid phenotype 

retains the epithelial nature of the cells by immunohistochemistry 

or electronmicroscopy. Recent molecular studies 

strongly argue for a monoclonal origin of both components in 

sarcomatoid carcinoma and carcinosarcoma. The mean age is 

66 years (range, 50-77 years). Pathological stage is the best 

predictor of survival in sarcomatoid carcinoma. The major 

differential diagnostic consideration is urothelial carcinoma 

with pseudo-sarcomatous stroma, a rare entity with reactive 

stroma. In cases with exclusively spindle cells, the main 

differential diagnostic consideration is sarcoma, particularly 

leiomyosarcoma. Immunostaining with cytokeratin is helpful 

in this setting. Sarcomatoid carcinoma with prominent myxoid 

and sclerosing stroma may be mistaken for inflammatory 

pseudotumor. 

Lipoid-cell variant. Lipoid cell variant, is a rare neoplasm 

defined by the WHO (1999, 2004) as an urothelial carcinoma 

which exhibits transition to a cell type resembling signet-ring 

lipoblasts. It is currently considered to be an ill-defined tumor 

variant, and whether it should be classified as carcinosarcoma 

remains to be established. Clinicopathologic features and 

the immunohistochemical findings in seven reported cases 

showed gross hematuria as the initial symptom. All patients 

were elderly men (mean age, 74 years; range, 63-94 years). 

On microscopic examination, the extension of the lipid cell 

pattern varied from 10%-30% of the tumor specimen, with

192 

associated micropapillary (n = 1), plasmocytoid (n = 2), and 

grade 3 conventional urothelial carcinomas (n = 4). The immunohistochemical 

results showed an epithelial phenotype 

of the lipoid cell component characterized by diffuse staining 

with cytokeratins AE1/AE3. 

Undifferentiated carcinoma. This category contains tumors 

that cannot be otherwise classified. To our knowledge, they 

are extremely rare. 

References 

Lopez-Beltran A, Cheng L. Histologic variants of urothelial 

carcinoma: differential diagnosis and clinical implications. 

Hum Pathol 2006;37:1371-88. 

Inverted bladder neoplasms: inverted papilloma 

M. Colecchia, B. Paolini 

Dipartimento di Patologia, Fondazione IRCCS IstitutoTumori di Milano, 

Italy 

Inverted papilloma comprises less than 1% of urothelial 

neoplasms and is generally considered a benign neoplasm. 

The mean age at diagnosis is 64 years and a peak frequency 

is in the 6 th and 7 th decades. It is more common in men than 

women. The etiology is uncertain 1 . Recent molecular data 

supports its benign nature based in the low amount of genetic 

anomalies found in most cases 2 . Most cases of inverted papilloma 

are located in the bladder trigone, but inverted papilloma 

can also be found in the uretere, renal pelvis, urethra 1 . 

Macroscopically it is characteristically sessile or pedunculated, 

smooth surfaced, small and single but large multifocal 

lesions may occur 3 . Microscopically inverted papillomas had 

a relatively smooth surface covered by histologically and 

cytologically normal urothelium and consists of intramucosal 

and submucosal anastomising islands and trabeculae of 

urothelium. There are two main patterns: the trabecular and 

glandular patterns 4 . In both patterns of inverted papilloma, 

the epithelial elements are surrounded by an intact membrane 

and delicate fibrovascular stroma. Unusual growth patterns 

of inverted papilloma include basaloid, hyperplastic, spindle 

cell and neuroendocrine patterns. Mild cytologic atypia could 

be observed in inverted papilloma, and the precise demarcation 

with carcinoma is unresolved; in rare cases nuclear 

Clinical pathways: is there a role for 

pathologists? 

E. Bonoldi, A. Parafioriti * , G. Coggi 

Unit of Pathology, IRCCS Ospedale Maggiore Policlinico, Milan, 

Italy; * Unit of Pathology Gaetano Pini Institute, Milan, Italy 

A critical or clinical pathway (CP) is the coordinate, accurately 

planned sequence of interventions by health care professionals 

for a single patient or a group of patients requiring 

treatment for a particular diagnosis or problem. CPs are tools 

used as references for Evidence Based health-care. They have 

been implemented internationally since the 1980s. by different 

health care systems all over the world. 

CPs are characterized, according to the European Pathways 


Clinical governance 

Moderators: G. Coggi (Milano), G. Barresi (Messina) 

atypia may be prominent but these atypical nuclear features 

are considered degenerative in nature. Immunohistochemical 

expression of a number of biomarkers, including Ki 67, p 

53 and CK 20has been shown to be of diagnostic value. In 

a study by Jones 0/15 inverted papillomas stains positively 

for Ki67 and CK 20 and only 1/15 stained positively for 

p 53. Urovysion FISH produced normal results for all cases 

of inverted papilloma 5 . Mitotic figures are rare or absent in 

inverted papilloma, unlike carcinoma and generally are less 

than 1/10 hpf, while in inverted carcinoma mean number 

was 8/10 HPF in a large series study 5 6 . The number of cases 

with coexistent urothelial carcinoma in situ or carcinoma has 

increased recently. Inverted papilloma are usually diploid 7 . 

Sung and collagues examined a series of 39 inverted papillomas 

using LOH analysis and showed a very low incidence 

of LOH at genetic loci that are frequently lost in both urothelial 

carcinomas and papillary urothelial neoplasms of low 

malignant potential 2 . Recurrent lesions have been observed in 

< 1% of the reported cases 8 . 

references 

1 Sauter G. Inverted papilloma. In: Eble JN, Sauter G, Epstein JI, et al. 

(eds). Pathology and genetics of Tumors of the Urinary System and 

Male Genital Organs. Lyon: Iarcc Press 2004. 

2 Sung MT, Eble JN, Wang M, et al. Inverted papilloma of the urinary 

bladder: a molecular genetic appraisal. Mod Pathol 2006;19(10):1289- 

94. 

3 Rozanski TA. Inverted papilloma: an unusual recurrent, multiple and 

multifocal lesion. J Urol 1996155(4):1391. 

4 Kunze E, Schauer A, Schmitt M. Histology and histogenesis of two different 

types of inverted urothelial papillomas. Cancer 1983;51(2):348- 

58. 

5 Jones TD, Zhang S, Lopez-Beltran A, et al. Urothelial carcinoma with 

an inverted growth pattern can be distinguished from inverted papilloma 

by fluorescence in situ hybridization, immunohistochemistry, and 

morphologic analysis. Am J Surg Pathol 2007;31(12):1861-7. 

6 Amin MB, Gómez JA, Young RH. Urothelial transitional cell carcinoma 

with endophytic growth patterns: a discussion of patterns of 

invasion and problems associated with assessment of invasion in 18 

cases. Am J Surg Pathol 1997;21(9):1057-68. 

7 Cheville JC, Wu K, Sebo TJ, et al. Inverted urothelial papilloma: is 

ploidy, MIB-1 proliferative activity, or p53protein accumulation predictive 

of urothelial carcinoma? Cancer 2000;88(3):632-6. 

8 Cheng CW, Chan LW, Chan CK, et al. Is surveillance necessary for 

inverted papilloma in the urinary bladder and urethra? NZ J Surg 

2005;75(4):213-7. 

Association by five parameters: 1) multidisciplinary approach 

to plan care 2) translation of guide lines or evidence 

into local structures 3) detailed description of the steps 

in a course of diagnosis and treatment, with regard to the 

algorithms, guide-lines, protocols and procedures adopted 

4) evaluation in progress of the timeframes of intervention 

5) standardization of care for a specific clinical problem in a 

specific population. 

CPs are developed through collaborative efforts of clinicians, 

case managers, nurses and other allied health care professionals 

with the aim of improving the quality of patient care and 

minimizing costs. 

Indeed CP aims to improve, in particular, the continuity and 

coordination of care, across different medical specialties.

lectures 

Although the choice of a standardized CP is mainly a clinician’s 

task, still an important role is left to other health care 

professional in the field of Laboratory Medicine, Radiology, 

Radiotherapy etc. 

With regard to Laboratory Medicine, a significant role is 

played by Pathologists. 

The latter, in fact, can deeply contribute to the development of 

CPs, for both neoplastic and non-neoplastic diseases, in prevention, 

diagnosis, control of response to therapy and follow-up. 

The histopathological diagnosis itself is in fact of paramount 

relevance in addressing a patient’s health process towards the 

most appropriate CP or shifting it, from one CP to another. 

For example, in case of superficial lymphoadenopathy, the 

choice of CP will be considerably different with a diagnosis of 

metastatic disease, rather than of lymphoproliferative disease, 

or of a reactive process. Even more different will be the CP 

should a fine needle biopsy approach or a surgical excision be 

chosen, on the basis of specific diagnostic clinical questions. 

The CP followed by patients affected by breast cancer, for 

example, is going to become more and more tailored on the 

basis of pathological and biological features of the single 

case, just in virtue of the increasingly appropriate pathological 

diagnosis, which must fulfill the criteria of Evidence Based 

Pathology, reporting prognostic and predictive factors. 

The multidisciplinary team of health professional involved 

in planning and construction of a CP will refer to guide lines 

(systematically developed statements to assist practitioners 

for making decision in diagnostic process and/or health 

care), protocols and procedures (diagnostic and/or treatment 

recommendations based on guide lines), in order to decrease 

individual variability. Development of optimal CP is gained 

through the following steps: 

– Select topic: topic selection generally concentrates on high 

frequency, high cost diagnoses and procedures. 

– Select a team: crucial to success is to develop a sound 

multidisciplinary team, in order to guarantee coverage of 

the different fields and professionals involved in patient 

care. Lack of active commitment and participation plays a 

dramatic role in failure of a CP. 

– Evaluate the current process of care in order to understand 

current variation. 

– Evaluate medical evidence and gold-standard practice. 

– Determine the format of CP: a simple check list could be 

an optimal method for both attending and implementing the 

pathway. 

– Document and analyze variance using performance indicators 

corresponding to key features in process improvement 

– Pathway implementation by educational projects addressed 

to the staff and accurate selection of individual roles. 

Among the benefits to health care organization in introducing 

CP, special interest gains the reduction of unnecessary variation 

in patient care, of delay in discharge and the improvement 

of cost-effectiveness of clinical services. 

Notwithstanding the potential barriers to the introduction of 

CPs, integrated processes running the whole course from prevention 

to diagnosis, treatment and rehabilitation can really 

help to provide explicit and well-defined standards of care, 

while supporting clinical effectiveness, risk management and 

clinical audit. 

references 

Coffey RJ, et al. Qual Manag Health Care 1992;1(1):45-54. 

Campbell SS, et al. British Medical Journal 1998;316(7125):133-7. 

Coffey RJ, et al. Qual Manag Health Care 2005;14(1):46-55. 

Talmor D, et al. Crit Care Med 2006;34(11):2738-47. 

Patkar V, Fox J. Stud. Health Technol Inform 2008;139:233-42. 

The control process in pathology 

G. Angeli 

Pathology Unit, Vercelli Hospital, Italy 

193 

The control process consists of sequential actions taken by 

management to establish performance standards, measure 

and evaluate performance and take corrective actions where 

indicated. 

The control process is practiced by all areas and levels of 

an organization. The basic process remains the same: 1) setting 

performance standards; 2) measuring the performance; 

3) evaluating the performance; 4) making effective use of 

feedback and taking corrective actions when necessary. 

Feedback information can be used either to confirm or to correct 

organizational performance. Effective use of feedback is 

a powerful tool for the control of work performance. Using 

feedback is a crucial point, because if the actual performance 

does not meet the performance standard, management will 

take corrective actions. This presupposes that the standards 

are fair and can be met. This is particularly true when we are 

dealing with the objectives assigned to the Pathology Unit, 

which have to be by definitions realistic and related to the 

given resources. 

Control is a dynamic and ongoing process. Such a process assumes 

as given the aims and strategies of the organization and 

takes place in the context of the strategic planning. 

The first step is the definition of performance standards, that 

are organizational goals stated in concrete and measurable 

performance terms. If the standards are unrealistically high, 

the organization will not obtain the desired results and will be 

judged a failure. If the performance standards are set too low, 

an organization may easily get the desired standards and be 

considered a success when a much more productive use of the 

assigned resources was actually possible. The aim is to create 

fair and equitable standards. This can be done by examining 

past performance as well as the performance of other institutions 

with similar characteristics in a benchmarking manner. 

In the context of control process it is necessary to decide 

what performance to measure, when to measure, and how to 

measure. Most important point is to define suitable and easily 

measurable indicators of the different phases of the process. 

Such indicators may be of quantitative or qualitative types, as 

for example those of the balance score card. The indicators so 

identified become part of reporting documents. 

Once the performance standard has been defined and the measures 

taken, the next step of the control process is to evaluate 

the performance. This is the process in which the measured 

performance is compared with the performance standard. The 

information derived from the performance evaluation is in 

each case used in a constructive manner, either confirmative 

or corrective, on the job itself. This is the feedback mechanism. 

The control process can operate at three different levels: input, 

process and output. When the control process operates before 

the actual activity is called input control. It allows the organization 

to correct defective performance before making the 

final commitment of resources. Example of input control is 

budget. Process control takes place as the work is performed, 

so it can assures that the actual performance meets the desired 

standards. Examples of this kind of control are quality 

controls. The output control operates at the end of the process 

and involves the final product of the process itself. It includes 

quality controls of the final product and audits. 

No organization has unlimited resources, so controls are 

necessary. Among financial controls budget has a primary

194 

importance, because it can be used as financial performance 

standard. Since budgets are created for organizations with 

limited resources, they give the financial framework and 

define the boundaries within which the allocation of those 

resources must take place. Budgets aid management in the 

performance of the basic managerial functions: planning, 

organizing and control. 

Budget is a plan expressed in quantitative, usually monetary, 

terms referred to a given period of time, generally one year, 

with a cost center or a responsibility center approach. So responsibility 

centers commit themselves to reach the objectives 

with assigned resource, then superior levels assure a positive 

evaluation of this behavior. 

There are two different kinds of approach to budgeting process: 

top-down, when the budget is prepared by top management; 

bottom-up, when the budget is prepared by lower levels 

managers and then submitted to top management for approval. 

The second approach has the advantage that the budgeting 

process is carry out by the same that have created it. 

The budgeting process needs regular reports which allow to 

compare expected inputs and outputs with the actual ones and 

to start confirmative or corrective feedbacks. Such a process 

should be flexible and subject to periodic review. Flexible 

or probabilistic budgets take into account potential environmental 

changes, and may be changed in front of a modified 

context. 

Audits are another kind of process control, either external or 

internal to the organization. Audits are formal evaluations 

of the performance of an organization in term of financial 

situation or quality of the final product (clinical audits, for 

example). 

Most important function of control process is the behavioral 

control, in term of evaluation and motivation of the people, 

who represent the most valuable non material resource of an 

organization. 

references 

Anthony RN, Young DW. Management control in nonprofit organization. 

The McGraw – Hill Co. 1999. 

Montana PJ, Charnow BH. Management. Barron’s Educational Series 

2008. 

evidence based pathology 

A. Parafioriti, E. Bonoldi * , E. Armiraglio * , G. Coggi 

Unit of Pathology Gaetano Pini Institute, Milan, Italy; * Unit of Pathology, 

IRCCS Ospedale Maggiore Policlinico, Milan, Italy 

Evidence Based Pathology (EBP) is an integral part and an 

off-shoot of Evidence Based Medicine (EBM), specialized 

in diagnosis and prognosis of human diseases. Pathologists 

play a vital role in clinical diagnosis: decisions based on the 

pathological contribution to diagnosis are made on a daily 

basis and have far-reaching consequences for patients and the 

health service. The same diagnostic role also plays a key part 

in screening, in classifying disease and as a reference standard 

for clinical disease. EBM has mostly focused on therapy and 

has provided a solid basis for some treatment, emphasizing the 

importance of the randomized controlled trial to determine, 

through perfectly designed experiments, the best treatments. 

Diagnosis is more complex than therapy and less attention 

has been paid to developing a rigorous, systematic approach 


to this field of medicine. Well-tried methods of EBM can be 

used for diagnosis. The process starts with identifying a real 

clinical problem and then, following rules of EBM, translating 

this into an answerable question. Possible solutions are 

obtained through a search for evidence and critically evaluated. 

When solutions are appropriate they can be applied in 

practice, otherwise we need for further research. 

In anatomic pathology, the pathway of diagnostic inference 

is usually leaded by clinical instinct, supported by experience 

according the so-called “skill full eye”. 

Such a trajectory could look like an “artistic process” rather 

than a scientific procedure. 

Incorporation of EBP in everyday practice can really help to 

plan a diagnostic method informed by objective evidences 

derived from well-designed studies, rather than based on 

problem-solving experience. 

In placing a lesion in a defined category, pathologists should 

always link the observation of morphological elements to the 

information obtained by ancillary tests, clinical data and to the 

knowledge derived from the literature. This requires to rely 

upon a rigorous algorithm defining the hierarchy of morphological 

and biotechnological features, so that, the final report 

can be reproducible. 

Actually, many pathology practices are neither objective, nor 

precise because guidelines for standardization are lacking. 

EBP helps to unify the current methodology primarily based 

on “pattern recognition” to newer types of tissue-based testing, 

including but not limited, to analyses of scores, cut-off, 

genomic or proteomic features. 

A final diagnosis, EBP-directed, allows a real customized 

therapy, in observance of the unique biology of an individual 

patient’s disease and more accurately can predict outcome and 

effectiveness of treatment. 

Moreover, we must not forget that everything that is done in 

healthcare system has an “opportunity cost”. Reproducibility 

of a diagnosis, obtained according to EBP criteria, can help to 

minimize both error possibility and potential costs, including 

medico-legal consequences. 

In conclusion, we believe that EBP is concerned with ensuring 

that all the aspects of generating a test result (sampling, 

morphological assessment and final report) are based both on 

judgment ability stemming from experience and on reproducible 

and clinically relevant criteria. 

Evidence based diagnosis needs more systematic reviews and 

appropriate tools, with special regards to electronic databases 

and meta-analysis (Pub-Med, Cochrane Library, Clinical Evidence), 

and more high quality research information in order to 

translate the data obtained to real decision making. 

The challenge to clinicians, pathologists, educators, researchers, 

funders, journal editors, and publishers is to work together 

to make this happen. 

references 

Annual Meeting of the Association of directors of Anatomical and Surgical 

Pathology; March 24, 2007; San Diego, CA. 

Booth A. Mapping the evidence base of pathology. J Pathol 1999;188:344- 

50. 

Crawford J.M. Original research in pathology: judgment, or evidencebased 

medicine. Lab. Invest 2007;87:104-14. 

Marchevsky AM, Wick MR. Evidence-Based Medicine, Medical Decision 

Analysis, and Pathology. Hum Pathol 2004;35:1179-88. 

Wick MR, Marchevsky AM, Foucar E. Evidence-Based Medicine. Semin 

Diagn Pathol 2005;22(2):105-77.

lectures 

Procedures and guidelines 

R. Giardini, E. Tavani, D. Ientile 

Istituti Ospitalieri, Cremona, Italy, Rho Hospital, Rho, Italy, Bucheri 

La Ferla Fatebenefratelli Hospital, Palermo Italy 

Procedures manuals and guidelines are born as tools with 

the aims to permit to the professionals to make “informed 

choices” based on the analysis of scientific tests and on the 

evaluation of risk and benefit of every action. Moreover, procedure 

manuals and guidelines proved to be a tool of updating 

for professionals, of education and information for patients 

and an external reference to verify what the pathologist is able 

to produce. 

Given the high level of information interchange among pathologists 

of the same hospital and among different structures 

(counselling activity, data centralization, regional quality 

controls, screening campaigns), the operative practice in a 

Division of Anatomical Phatology, is usually and, to say, 

physiologically, based on standard protocols, well defined 

and accepted among professionals, even if not very diffused 

trough divisions and known only in very specialized (by organs 

or pathology) divisions. 

Nevertheless there is often an implied information interchange, 

not formalized and not standardized by methodologically 

rigorous procedures clearly declared. On the other hand 

there is an implicit need of validated references proved by 

those almost always high level procedures, optimized over the 

time, by study groups of the National Scientific Society or into 

regional branches of the same society. 

Therefore it seems to us appropriate to start again a “new-old” 

discussion in our National Scientific Society about the proper 

and practical use of work tools such us the procedures manual 

and the diagnostic guidelines. We think that such tools are 

more and more relevant in the diagnostic activity and express 

a specific value both for crediting and certification of the Anatomical 

Pathology divisions and also for the role that their use 

assume in the risk management related to our work. 

Procedures and guidelines 

Moderator: R. Giardini (Cremona) 

195 

A relevant remark concerning an appropriate consideration 

about the real meaning of the terms “Procedures manual” and 

“Guidelines”, often not properly utilized as synonyms. 

In the Anglo-Saxon literature, due to the prevalence of health 

insurance systems imposing more rigorous rules, a better defined 

meaning of those words is used, whereas in our language 

those rules are vaguer. 

If from one side we can agree about the value of “procedures” 

in order to standardize as much as possible the “technicalmethodological” 

course in different extent of a pathology 

field, on the other side we have to ask ourselves if there is a 

meaning in talking about our “guidelines”, out of an indispensable 

multidisciplinary context, where we are working in 

the every day life. 

As suggested by the “manual of direction” published by the 

Italian Superior Institute of Health, perhaps it would better not 

to talk about anatomopathological guidelines but about “diagnostic 

algorithms”, intended as supports to the diagnostic 

iter, based on the literature data (EBM) and/or on necessaries 

consensus conferences. 

In summary our terms “procedures” and our “interpretative 

algorithms” should find their natural context in real guidelines, 

established by the contribution of multi-specialist teams. 

In this sense, the FONCAM manual looks as a very good 

example to us. 

Based on this not only semantic, but also substantial, argueing, 

stay the medicolegal aspect of the use of procedures and 

guidelines, more and more relevant in our professional field. 

There is a very open question about that and at this point an 

indispensable discussion among expert pathologist should be 

open. 

The aim of this session is to make possible a workgroup that 

can guarantee, with the collaboration of corporate specialists 

and single interested pathologists, the necessary homogeneity 

and coverage of as much diagnostic fields as possible, constituting 

a valid support to our professional activity, in every 

context could be expressed. 

Slide seminar: lymphoma surgical pathology 

Primary large B cell lymphoma presenting as 

soft tissues mass: a case report and review of 

the literature 

U. Gianelli, E. Bonoldi, E. Armiraglio * , A. Di Bernardo, A. 

Moro ** , A. Parafioriti * 

Dipartimento Interospedaliero di Anatomia Patologica: U. O. C. 

Anatomia Patologica, Università di Milano, Fondazione IRCCS “Ca’ 

Granda”, Ospedale Maggiore Policlinico, Milano, Italia; * U.O. Anatomia 

Patologica A.O. Istituto Ortopedico Gaetano Pini, Milano, Italia; 

** U.O. Anatomia Patologica A.O. San Paolo, Milano, Italia 

We describe a case of a 73 years old man, who was admitted 

at the Istituto Ortopedico “G. Pini” of Milan, because of a 

continuous pain in the pelvis, for 7 months. 

Moderators: V. Franco (Palermo), S. Pileri (Bologna) 

The patient had a previous history of a car accident occurring 

four years before, from which he had reported multiple 

fractures of the ribs and of the left pelvis, together with lung 

contusions. 

At the time of the first observation the patient suffered of 

cruralgia and the physical examination revealed a swelling of 

the left buttock. 

Peripheral blood examination showed the following parameters: 

WBC: 11,1 × 10 9 /l, RBC: 4,72 × 10 12 /l, Hb: 12,3 g/dl, 

PTL: 422 × 10 9 /l (LF: N = 81, Eo = 0, Ba = 0,3; l = 11, 

Mo = 6,7), ESR 100 mm. 

The CT scan revealed a mass 4 cm. thick, covering the foramen 

ovale and extending to the pelvis, located posterior to the 

iliac muscle and anterior to the muscle of the left buttock.

196 

Bone scintigraphy was negative and didn’t show any pathological 

uptake. 

Ecothomograpy displayed a grossly hypoecogenic mass growing 

between the iliac muscle and the bone. 

In the suspect of soft tissue malignancy an open biopsy of the 

mass was performed. 

Histological examination revealed a solid proliferation of 

large cells with abundant clear to granular and eosinophilic 

cytoplasm, with a nested and diffuse growth pattern, focally 

alveolar, infiltrating the skeletal muscle and the adipose tissues. 

The nuclei of tumoral cells were irregular with prominent 

single or multiple nucleoli. Differential diagnosis included: 

clear cells sarcoma, amelanotic melanoma, metastatic 

large cells carcinoma or diffuse large cells lymphoma. 

Neoplastic cells presented the following immunophenotype: 

CD45 (+), CD79a (+), CD20 (+), CD10 (-), bcl-6 (+), MUM- 

1 (+), NKIC3 (+/-), CD3 (-), CD5 (-), CD30 (-), EMA (-), 

S-100 (-), HMB45 (-), Melan-A (-), CD68R (-), Actin (-), 

Desmin (-), CK (MNF116, CAM 5.2, AE1-AE3) (-). The 

proliferation index was ki-67 (MIB1): 60-70%. 

The morphology and the immunophenotype supported a diagnosis 

of diffuse large B-cell lymphomas (DLBC), NOS, of 

non-germinal centre-like immunophenotype. Because of all 

other staging procedures did not reveal any localization of 

the neoplasm, a final diagnosis of primary DLBC of the soft 

tissues was performed. 

Primary DLBC of the soft tissue are extremely rare and the 

two most important series of these types of NHLs were described 

about 20 years ago. 

The Armed Forces Institute of Pathology collected 75 cases of 

primary lymphomas of the soft tissues in a period spanning 20 

years. The Mayo Clinic was able to find 8 of these cases out 

of 7000 NHLs collected in period of 10 years. Other sporadic 

cases have been described in the literature as case report. 

All histological types of NHL have been found in soft tissues, 

including low- and high-grade tumours. Most of the tumours 

occur in the soft tissues of the thigh, followed by abdominal 

wall, arm and leg. Intermuscolar tissue rather than the muscle 

itself appears to be the tissue most frequently involved. 

This case emphasizes the importance to include Non Hodgkin 

large B cell lymphoma in the differential diagnosis of poorly 

differentiated neoplasms arising in soft tissues and skeletal 

muscle. 

Detection of primary lymphoma, by means of immunohistochemical 

investigation may spare the patient invasive surgical 

treatment and provide properly clinical management. 

A case of CD5 negative, diffuse large B-cell 

lymphoma with unusual expression of cyclin D1 

M. Lucioni1 , R. Riboni1 , F. Novara2 , G. Fiandrino1 , S. Kindl3 , 

O. Zuffardi2 , M. Paulli1 1 Anatomic Pathology, Foundation IRCCS Policlinico San Matteo, 

University of Pavia, Pavia Italy; 2 Human Genetics, University of 

Pavia, Italy; 3 Anatomic Pathology, Ospedale Guglielmo da Saliceto, 

Piacenza, Italy 

Background. The nuclear protein cyclin D1 plays a major 

role in cell cycle control since it promotes transition from G1 

to S-phase and, therefore, cell proliferation. Based on its well 

defined molecular function, cyclin D1 deregulation may contribute 

to the pathogenesis of certain lymphoma subtypes 1 . 

Overexpression of cyclin D1 is, with few exceptions, restricted 

to three lymphoma subtypes: mantle mantle cell lymphoma 

(MCL), plasma cell myeloma and hairy cell leukemia 2 . 


Cyclin D1 is aberrantly expressed in 90% of MCL as a result 

of t(11;14)(q13;q32), between the immunoglobulin heavy 

chain (IGH) and cyclin D1 (CCDN1) genes. Expression of 

CD5 is also highly characteristic, being present in at least 90% 

of mantle cell lymphomas; that notwithstanding, existence of 

CD5- cases is well recognized. Plasma cell myelomas express 

cyclin D1 in about 40% of cases, some of which show t(11;14). 

In hairy cell leukemia, deregulated expression of cyclin D1 is 

very common, apparently in the absence of the t(11;14) 2 . 

In these settings, immunostain for cyclin D1 may be diagnostically 

useful, also providing prognostic information in the 

case of multiple myeloma. Cyclin D1 detection is crucial to 

identificate the pleomorphic and blastoid variants of MCL, 

two aggressive forms of MCL characterized by large cells 

(different from those of classical MCL), high label index and 

complex kariotypes. Cyclin D1 expression is of paramount 

importance in the differential diagnosis between these MCL 

variants and diffuse large B-cell lymphoma (DLBCL), a heterogeneous 

group of aggressive lymphomas with respect to 

morphology, phenotype, genetic features and clinical behaviour, 

that is usually not associated with t(11;14). Herein we 

present an unusual case of DLBCL expressing cyclin D1 in 

the absence of CCND1 translocation, addressing the dilemma 

of the differential diagnosis for B-cell lymphomas consisting 

of large cells and its possible biological significance. 

Case history. A 69-year-old male patient presented with 

superficial laterocervical and submandibular lymphadenopathies, 

since 2 months. He was well and reported no systemic 

symptoms. Laboratory investigations showed normal white 

cell count and a raised lactate dehydrogenase level. Because 

of clinical suspicion of a lymphoproliferative disorder, the 

patient underwent surgical excision of a right laterocervical 

lymph node. 

Histological examination revealed complete nodal architecture 

effacement by a lymphoid proliferation consisting of medium 

to large atypical cells, with round, elongated or pleomorphic 

vescicular nuclei, one or more prominent nucleoli and relatively 

abundant cytoplasm. Numerous mitoses were observed. 

The lymphoid proliferation showed a predominant diffuse 

growth pattern, with residual vaguely nodular appearance still 

detectable at low magnification. Accompanying interstitial 

collagenous fibrosis was found, sometimes in broad bands. 

By immunohistochemistry, the neoplastic cells strongly expressed 

CD20, CD79a and PAX5 but were negative for CD3 

and CD5. Immunostains for cyclin D1, using both mouse 

monoclonal antibody P2D11F11 and rabbit monoclonal antibody 

SP4, showed moderate nuclear expression in about 60% 

of tumor cells. Additional staining for T-cell leukaemia 1 

(TCL1) oncogene and the nuclear transcription factor SOX11 

were negative. Histogenetic profile (CD10-, bcl-6+, MUM1+) 

was consistent with a post-germinal center (GC) derivation. 

Lymphoma cells were also strongly positive for bcl-2 and 

p53, whereas CD23, IgD and IgM immunostains were negative. 

Variable expression of CD30 was also found in 30-40% 

of lymphoma cells. Mib1/Ki-67 label index was about 40%. 

Molecular biology analysis documented monoclonal IGH 

gene rearrangement, with use of IGHV 1-69 gene segment and 

high load of somatic mutations (8,59%) at direct sequencing. 

Search for BCL-1/CCND1 a BCL-2 gene rearrangements by 

PCR was negative. 

Interphase DNA fluorescence in situ hybridization (FISH) 

with IGH/cyclin D1 fusion probes (Vysis ® ) and cyclin D1 

split probes (DakoCytomation ® ), failed to detect any translocation 

involving CCND1 gene. Further FISH studies revealed 

no translocations involving BCL-2 or c-MYC genes. We also

lectures 

performed whole genome array-CGH analysis using the 60K 

platform (41.5 KB overall median probe spacing) according 

to the manufacturer’s protocol (Agilent Technologies ® ). Array 

CGH experiment revealed complex kariotype, characterized 

by the following copy number alterations (CNAs): losses of 

1p36.33-p12 (120 Mb), 1p35.3-p35.2 (1,6 Mb), 3, 10, 15, 

16q12.12-q12.2 (2,9 Mb), 18, Y; gains of 1q21.1-q44 (102 

Mb), Xp22.11-q11.1, Xq11.1-q28. The observed log 2ratios of 

all CNAs suggested that the rearrangements were in mosaic. 

Based on the integration of morphologic findings, tomor cell 

phenotype, molecular and genetic data, our final diagnosis 

was of DLBCL, centroblastic with aberrant cyclin D1 expression. 

Following this diagnosis the patient began immunochemotherapy 

with rituximab, cyclophosphamide, doxorubicin, 

vincristine and prednisolone (R-CHOP). 

Discussion. The present case is interesting because of unusual 

expression of cyclin D1 in a DLBCL centroblastic subtype. 

Immunohistochemical detection of cyclin D1 in a B cell lymphoma 

consisting of medium to large cells usually suggests a 

diagnosis of pleomorphic or blastoid MCL; however in our 

case several features argued against this hypothesis. Actually, 

t(11;14)(q13;q32) or variants, that are found in virtually all 

cases of cyclin D1+ MCLs, were lacking. Aberrant phenotypes 

have been described in MCL and also in blastoid and 

pleomorphic variants; rare cases seem to express bcl-6 (1,6% 

in a series) 3 CD10, or MUM1, others lack CD5 or IgM/IgD 4 . 

In spite of this, our case showed a CD5-, bcl-6+, MUM1+ 

phenotype, which is far more in keeping with DLBCL. In 

addition to bcl-6 and MUM1 co-expression, the presence of a 

high degree of somatic hypermutation in IGHV genes would 

be exceptional in MCL and suggests post-GC histogenesis 5 . 

Array CGH revealed a highly complex kariotype, but the most 

frequent chromosomal aberrations accompanying CCND1 

translocation in MCL were lacking 2 . 

TCL1 and SOX11 are two immunohistochemical markers recently 

employed in the characterization of B-cell lymphomas, 

being positive in the vast majority of MCLs. In particular, 

strong TCL1 expression has been detected in the majority of 

lymphomas of pre-GC derivation, including MCL, whereas 

lymphomas deriving from GC and post-GC B-cells are usually 

negative, with the exception of Burkitt lymphoma 6 . 

SOX11 is a neural transcription factor that is expressed in 

lymphoblastic lymphoma (almost always), MCLs (up to 

93% of cases), and in a subset of Burkitt lymphoma (33%). 7 

Negativity for both TCL1 and SOX11 in the present case also 

seems to exclude MCL, thus confirming the usefulness and 

specificity of these novel markers in the differential diagnosis 

of mature B-cell lymphomas. 

Cyclin D1 positivity is exceptional in DLBCL and mainly 

restricted to single case reports 8 9 . Few studies have systematically 

examined the immunohistochemical expression of cyclin 

D1 by DLBCL. Most authors found no or only occasional 

cyclin D1 expression in DLBCL, but we can not exclude that 

the recent introduction of more sensitive and reliable antibodies 

(such as SP4), may result in the detection of an increased 

number of cyclin D1+ cases 10 . Indeed, a more recent study by 

Ehringer et al reported the immunohistochemical expression 

of cyclin D1 in 10 (4,3%) of 231 DLBCL, some of which 

showing structural aberrations at CCND1 locus, but only one 

carrying t(11;14) 11 . 

These and our findings confirm the existence of CD5-, cyclin 

D1+ DLBCL, in the absence of t(11;14), even if these cases 

are very rare. Therefore, in current haematopathology practice 

cyclin D1 immunopositivity alone may be not sufficient 

in distinguishing pleomorphic/blastoid MCL from DLBCL. 

197 

FISH detection of a t(11;14)(q13;q32) appears preferable for a 

definitive diagnosis of MCL, at least in equivocal cases. 

The mechanism of aberrant expression of Cyclin D1 in the 

present case is unclear. The normal pattern of FISH analysis 

using CCND1 probes and the absence of gains at the 11q13 

locus suggest that the overexpression of cyclin D1 is related 

to posttranslational mechanisms. DLBCL represents a biologically 

and genetically heterogeneous group of aggressive 

lymphomas. Cyclin D1 is one of the key regulators of the 

cell cycle and elevated levels of cyclin D1 expression may 

accelerate G1/S-phase transition and therefore tumor cell proliferation. 

Our and other findings seem to suggest that cyclin 

D1 overexpression may play a role in the pathogenesis of a 

subset of DLBCL. 

references 

1 Baldin V, Lukas J, Marcote MJ, et al. Cyclin D1 is a nuclear protein 

required for cell cycle progression in G1. Genes Dev 1993;7:812- 

821. 

2 WHO Classification of Tumours of Haematopoietic and Lymphoid 

Tissues. Eds Swerdlow SH, Campo E, Harris NL, Jaffe ES, Pileri SA, 

Stein H, Thiele J, Vardiman JW. Lyon: IARC Press 2008. 

3 Camacho FI, Garcia JF, Cigudosa JC, et al. Aberrant Bcl6 protein 

expression in mantle cell lymphoma. Am J Surg Pathol 2004;28:1051- 

6. 

4 Gualco G, Weiss LM, Harrington WJ Jr, et al. BCL-6, MUM1 and 

CD10 expression in mantle cell lymphoma. Appl Immunohistochem 

Mol Morphol 2010;18:103-8. 

5 Kienle D, Krober A, Katzemberger T, et al. VH mutation status and 

VDJ rearrangement structure in mantle cell lymphoma: correlation 

with genomic aberrations, clinical characteristics, and outcome. 

Blood 2003;102:3003-9. 

6 Herling M, Patel KA, His ED, et al. TCL1 in B-cell tumors retains its 

normal B-cell pattern of regulation and is a marker of differentiation 

stage. Am J Surg Pathol 2007;31:1123-9. 

7 Mozos A, Royo C, Hartmann E, et al. SOX11 expression is highly specific 

for mantle cell lymphoma and identifies the cyclin D1-negative 

subtype. Haematologica 2009;94:1555-62. 

8 Rodriguez-Justo M, Huang Y, Ye H, et al Cyclin D1-positive diffuse 

large B-cell lymphoma. Histopathology 2008;52:889-904. 

9 Teruya-Feldstein J, Gopalan A, Moskowitz CH. CD5 negative, Cyclin 

D1-positive diffuse large B-cell lymphoma (DLBCL) presenting as 

ruptured spleen. Appl Immunohistochem Mol Morphol 2009;17:255- 

8. 

10 Cheuk W, Wong KOY, Wong CSC, et al. Consistent immunostaining 

for cyclin D1 can be achieved on a routine basis using a newly available 

rabbit monoclonal antibody. Am J Surg Pathol 2004;28:801-7. 

11 Ehringer M, Linderoth J, Christensson B, et al. A subset of CD5- diffuse 

large B-cell lymphomas expresses nuclear cyclin D1 with aberrations 

at the CCND1 locus. Am J Clin Pathol 2008;129:630-8. 

Diagnosing splenic marginal zone lymphoma in 

the bone marrow 

C. Tripodo, E. Iannitto * 

Dipartimento di Patologia Umana, Università di Palermo; * Unità di 

Ematologia con Trapianto di Midollo Osseo, Università di Palermo 

Splenic marginal zone lymphoma (SMZL) is an uncommon 

B-cell neoplasm listed as a distinct pathological entity in 

the WHO classification of tumours of haematopoietic and 

lymphoid tissues. SMZL is characterized by a commonly 

asymptomatic presentation, indolent clinical course, and 

overall survival usually exceeding ten years. SMZL diagnosis 

has been classically based on spleen histology, following the 

evidence that the spleen harbours most of the disease burden. 

Nevertheless, in most cases, the diagnosis of SMZL can be 

achieved by the combination of clinical and laboratory data, 

peripheral blood examination, and bone marrow histopathology, 

thus avoiding splenectomy.

198 

Here we discuss the approach to SMZL diagnosis on BM histopathology, 

by describing the prototypical case of a 63 years 

old female patient. The patient presented to the Haematology 

Unit of our University Hospital following the occasional finding 

of splenomegaly during a routine clinical examination. 

The spleen was palpable 3cm below costal margin and displayed 

a maximum diameter of 15cm on ultrasound imaging. 

No superficial or deep-sited lymphadenopaty was identified 

by physical examination and CT scan, respectively. 

Peripheral blood counts were the following: Hb 11 g/dl, 

HCT 37, WBC 5.8 × 10 9 /l, Neut. 1.5 × 10 9 /l, Lym. 3.8 × 10 9 /l, 

Mono 0.4 × 10 9 /l, PLT 270 × 10 9 /l. LDH was within normal 

range and β2-microglobulin was 3.3 mg/ml. Total serum protein 

levels were normal. However, electrophoresis revealed 

an inconspicuous monoclonal band in the gamma region. On 

immuno-electrophoresis, the monoclonal component proved 

to be of the IgM-kappa type. 

Peripheral blood smear analysis revealed the presence of a 

fraction of circulating lymphocytes (about 8%) with characteristic 

“villous” morphology (i.e. the presence of large polar 

villi). On flow cytometry, circulating lymphocytes showed the 

following phenotype: CD19+, CD20+, CD3-, CD5-, CD4-, 

CD8-, CD10-, CD23-, CD25-, CD43-, kappa-light-chain 

restricted. 

Bone marrow histopathology highlighted a slightly hypercellular 

marrow (55% overall cellularity) with preserved haematopoiesis 

and showing the presence of a mixed nodular, 

interstitial, and intra-sinusoidal infiltration by medium-sized 

lymphocytes, accounting for nearly 40% of the haematopoietic 

parenchyma. Immunohistochemistry confirmed the 

CD20+CD79a+CD5-CD2-CD23-CD10-IgM+ phenotype of 

the neoplastic lymphoid cells. 

On the bases of these data, a diagnosis of SMZL was performed 

and the patient was followed-up adopting a watchful 

waiting policy. 

Bone marrow examination is a crucial step in the diagnosis of 

splenic lymphomas. The presence of an intrasinusoidal pattern 

of infiltration (either alone or in combination with other patterns) 

can be frequently observed in B- and T-cell lymphomas 

with preferential splenic localization other than SMZL, such 

as hairy cell leukemia (HCL), HCL-variant, hepatosplenic Tcell 

lymphoma, all sharing a tropism for sinusoidal vascular 

niches. 

A pediatric natural killer lymphoma/leukemia 

with indolent course 

M. Ungari 

I Servizio di Anatomia Patologica, Spedali Civili di Brescia, Italia 

Case report. A 3 years old girl showed several erythematous 

skin lesions as well as scabby nasal lesion characterized by 

spontaneous regression. During the following 5 years, these 

lesions occurred every two months. At the age of 8, lesions 

recurred weekly. Physical examination showed enlarged right 

cervical and inguinal lymph nodes, hepatomegaly and normal 

spleen. A computed tomography (CT) scan revealed multiple 

adenopaties in the laterocervical, submandibular, subclavian 

and axillary regions. 

Haematological findings showed a slight anemia (10g/dl) and 

leucocyte counts (4,800 µl) with lymphocytosis (70%). The 

flow cytometric analysis of peripheral blood showed a natural 

killer lymphocyte population (CD2+, CD3-, CD7+, CD16+, 

CD56+). Morphological and immunophenotypical analysis of 

the bone marrow were normal. 


Antibodies vs EBV were tested and the IgG titer was1/128, 

whilst IgM titer was negative. The CT scan and a lumbar 

puncture were negative. 

A skin and a cervical lymph node biopsies were performed. 

The skin showed extensive necrosis, infiltration of atypical 

lymphocytes and histiocytes with angioaggressive and 

angiodestructive behavior. The lymph node parenchyma 

revealed an effaced architecture, with a diffuse infiltration 

of small-to-medium atypical lymphoid cells, that frequently 

enchroached upon the wall of large vessels; subcapsular areas 

of necrosis were also evident. On imprints, numerous cells 

with azurophilic granules were identified. Immunophenotypical 

examination performed on frozen and paraffin sections 

revealed a dominant infiltration of cells displaying a NK 

phenotype, with expression of CD2, CD56, TIA1, Perforin, 

and CD94, and negativity for CD3, CD4, CD5, CD8, CD30, 

CD57, the B-cell associated antigens CD19 and CD20, and 

those associated with dendritic plasmacytoid cells CD123 and 

BDCa2. Finally on both skin and lymph node sections high 

number of cells were positive for EBV, detected with EBER 

in situ hybridation technique. TCR-gamma rearrangement 

study did not show clonal bands. 

A chemotherapy according NHL protocol for anaplastic 

lymphoma was administered. A good clinical response in 

both skin and lymph nodes was abtained since the first cycle 

of treatment; flow cytometric analysis of PB cells showed 

a decrease (< 20%) of the peripheral NK-lymphoma cells. 

Following the second cycle of chemotherapy, full remission 

was obtained. Forty months after interruption of treatment, 

complete recovery was still enduring. 

Discussion. Cytotoxic lymphomas are tumors derived from 

T or NK lymphocytes with a cytotoxic phenotype. Neoplastic 

cells typically express at least one cytotoxic protein such as 

T-cell intracellular antigen (TIA)-1, granzyme B, or perforin 

1 2 . The World Health Organization (WHO) 3 lists them 

as distinct (extranodal NK/T-cell lymphoma nasal type and 

cutaneous γ/δ-cell lymphoma) or provisional entities (primary 

cuteneous aggressive epidermotropic CD8+ cytotoxic T-cell 

lymphoma). A closely related entity seen mainly in children 

is hydroa vacciniforme-like lymphoma. This disease shows 

overlap with NK/T-cell lymphoma, nasal type, of which it can 

be considered a variant 4 . However the expression of cytotoxic 

proteins is not restricted to a specific group of lymphomas as 

they can be observed in micosis fungoides, cutaneous CD30+ 

lymphoproliferative disorders and subcutaneous “panniculitis-like” 

T-cell lymphoma. Cytotoxic proteins do not have 

any diagnostic or prognostic value per se, and their expression 

should be evaluated in the context of the clinico-pathologic 

and molecular features of the lesions 4 . 

Negativity for T-cell markers and germline rearrangement of 

T lymphocytes, together with positivity for EBV in neoplastic 

cells, should be interpreted as a strong hint towards a diagnosis 

of extranodal NK/T cell lymphoma, nasal type 5 . This lymphoma 

is commonly located in the nasal cavity, but involvement 

of the skin can be observed. The prognosis in adults is usually 

unsuccessfull. Fifteen pediatric cases showed a better outcome 

than adults 6 . Complete remission was obtained and remained 

steady in two-thirds of the patients with localized disease. 

Among the 6 cases in stage IV, two had a successful outcome 

after treatment with high dose chemotherapy and hematopoietic 

stem cell transplantation. Moreover, it has been suggested 

that NK/T-cell lymphomas that express CD94, a lectin that 

inhibits NK function, may have a better prognosis 7 . 

This case showed unexpected long interval between the first 

signs of the disease and the diagnosis. At the age of 3, the

lectures 

first cutaneous lesions were evident and characterized by 

spontaneous regression. Unfortunately, no skin biopsy was 

performed at the age of 3. 

references 

1 Massone C, Chott A, Metze D, et al. Subcutaneous, blastic natural 

killer (NK) NK/T-cell and other cytotoxic lymphomas of the skin: a 

morphologic, immunophenotypic and molecular study of 50 patients. 


2 Kluin PM, Feller A, Gaulard P, et al. Peripheral T/NK-cell lymphoma: 

a report of the IXth Workshop of the European association for Haematopathology. 

Histopathology 2001;38:250-70. 

Bioethics in research on Human Tissue 

C. Faralli 

Bologna 

199 

3 Swerdlow SH, Campo E, Harris NL, et al. (eds). WHO Classification 

of Tumours of Haematopoietic and Lymphoid Tissues. Lyon: IARC 

Press 2008. 

4 Nava VE, Jaffe ES, The Pathology of NK-Cell Lymphomas and Leukemias. 

Adv Anat Pathol 2005;12:27-34. 

5 Cerroni L, Gatter K, Kerl H. Skin Lymphoma – The Illustrated Guide; 

3 rd Edition, 2009. 

6 Shaw PH, Cohn SL, Morgan ER, et al. Natural killer cell lymphoma: 

report of two pediatric cases, therapeutic options, and review of the 

literature. Cancer 2001;91:642-6. 

7 Lin CW, Chen YH, Chuang YC, et al. CD94 transcripts imply a better 

prognosis in nasal-type extranodal NK/T-cell lymphoma. Blood. 

2003;102:2623-31. 

Network of biobanks of archived tissues in Italy 

Research on human tissue, and in particular the collection and 

storage of biological samples to that end, raises bioethical 

problems relative to the tissue donor’s consent and privacy as 

well as to the commerciability of the human body. 

Consent. Informed consent is one of the keystones on which 

bioethics is built, because through informed consent the basic 

human liberties are exercised. 

Indeed, the principle of consent had been codified, and the 

occasion for it was the aberrant human experimentation carried 

out in the first half of the twentieth century: this led to 

the Nuremberg Code. And the principle has also been made 

part of other international documents, such as the Declaration 

of Helsinki and the Oviedo Convention, as well as in all European 

recommendations and directives directly or indirectly 

concerned with health. 

Under the principle of informed consent, such as it has 

evolved through the aforementioned documents and through 

the opinion of the Supreme Corte di Cassazione in Italy, no 

consent is valid unless accompanied by an adequate notice 

providing the following information, especially where the 

collection of human tissue is concerned: 1) the purpose for 

drawing and storing samples of tissue; 2) the techniques 

used to this end; 3) the location of the facilities that will be 

analyzing and storing the samples; 4) the lenght of storing 

samples; 5) the means used to guarantee the donor’s privacy; 

and, not least; 6) a requirement expressly stated by the Italian 

Data Protection Commissioner for genetic data, the methods 

enabling donors to withdraw their consent and destroy their 

samples if they so choose. 

Since biobanks cannot be created without processing biological 

samples, the issue that has emerged in this connection is 

that of the “right not to know:” this right, stating that a human 

being should be able to decide not to know his or her health or 

life prospects, was codified into law in Italy in 2007 through 

an authorization by the country’s Data Protection Commissioner. 

The hypothesis of an open consent or a trust consent has 

emerged in Northern European countries. 

A fundamental twofold requisite needs to be met in compliance 

with ethical, scientific, and legal standards: on the one 

hand, someone must be appointed who will be responsible for 

Moderators: V. Eusebi (Bologna), G. Stanta (Trieste) 

the tissue samples; on the other hand, dedicated ethical committees 

must be set up to which to turn in seeking an opinion 

on the use of a biobank and the single biological samples in 

it. 

Privacy and Discrimination. Clearly, as is the case with 

DNA sequences, biological samples can be used in ways that 

seriously infringe a person’s privacy, the risk being that of 

genetic discrimination, all the more so that the advance of 

knowledge is making for greater and greater possibilities, 

correspondingly increasing the potential for research to be 

conducted with such aims as bring greater harm. 

Indeed, the use of databanks containing tissue samples constitutes 

“processing of sensitive data” and can prove enticing 

to insurers, employers, and, not least, drug companies, 

among others. Under European law – and under Italian law 

in particular (especially the rules stated in the aforementioned 

authorization by the country’s Data Protection Commissioner 

for genetic data, an authorization applying as well to the 

processing of biological samples)—much attention is paid to 

the risk of unauthorized access to tissue samples, and for this 

reason strong security measures are provided for to restrict access 

to data as far as possible and to subject the tissue samples 

themselves to the most exacting formal, physical, and technological 

controls, while also limiting as far as possiple the 

maximum period over which such samples may be stored. 

Commercialization. Biobanks containing human tissues are 

additionally exposed to the risk of being improperly or illegally 

used for commerical purposes. The Oviedo Convention 

on the human genome and biomedicine accordingly expressly 

provides that the human body and its parts may not be used for 

profit, a provision based in part on an altruistic and solidarity 

principle. 

Network di biobanche europeo 

G Stanta 

A.C.A.D.E.M. Department, University of Trieste, Italy. 

Translational research on prognostic and therapy predictive 

biomarkers is strictly connected to the availability of normal 

and pathological tissues. In the last ten years the traditional 

biobanking system has been collecting many thousands of 

frozen tissues, but these repositories are being rapidly used. 

In the meantime the need for tissues has been increasing 

and their unavailability could slow down research. For these

200 

reasons a European project “Archive Tissues: Improving Molecular 

Medicine Research and Clinical Practice (IMPACTS)” 

(www.impactsnetwork.eu) was implemented in the past years 

to validate molecular methods in formalin-fixed and paraffinembedded 

tissues, which are usually preserved for a very long 

time in any pathology department of any hospital (Archive 

Tissues – AT). The project was also devoted to start biobanking 

procedures on this kind of tissues and on bioethical implications 

1 . Recently the usefulness of these tissues has been 

evaluated by the Biobanking and Biomolecular Resources 

Research Infrastructure (BBMRI) 2 . 

The IMPACTS group, together with the European Society 

of Pathology (ESP) and BBMRI is trying to develop a pan- 

European network of archive tissue biobanking that could 

be a very important instrument for accelerating the clinical 

application of molecular medicine. The possibility to carry 

out multicentric projects on a voluntary and collaborative 

basis with the collection of a very high number of human 

pathological tissues correlated with clinical information or 

the possibility to collect a sufficient number of rare lesions 

is an important issue that must be pursued. The reason for 

this is that AT are those tissues available for any patient in 

any European hospital and in most of the cases the only tissues 

available. That’s why any clinical procedure must take 

AT into consideration, and pathologists must take in charge 

molecular analysis in this kind of tissues. The role of pathologists 

starts from their ability to identify a huge number 

of pathological entities and to recognize the heterogenity of 

pathological tissues. In this way effective microdissections 

can be performed before any molecular examination. The 

unrecognized tissue variabilities could be the basis of many 

mistaken results reported in literature. Such mistakes could 

lead not only to wrong, but also to confusing research with 

retardation of a correct development. 

After performing multicentric studies to give validated methods 

for this kind of analysis, the IMPACTS group prepared 

the final version of “Guidelines for molecular analysis in archive 

tissues”. Pre-analytical conditions and molecular procedures 

were carefully evaluated to try to standardize the results 

of research. The network itself can represent a reference point 

for this kind of molecular procedures. 

The pan-European network can be organized as a virtual network 

that can be activated on a voluntary and collaborative 

basis. The existing biorepositories in the pathology departments 

are the basis for the development of the network. When 

a pathology department decides to participate in a multicentric 

project it can very easily modify its function of biorepository 

to the function of biobank just through the anonymization 

of tissue samples. The reason for this is that pathologists are 

already privacy guarantors, since they are involved in the diagnostic 

process and therefore subject to professional secrecy. 

They can explore the already existing computerized clinical 

files to connect tissues with clinical data and follow-up information. 

They are also in charge of choosing the cases and 

defining microdissection criteria. The best choice of cases, 

tissues and the validity of methods can guarantee the results 

of the studies in the best way ever. 

references 

1 Stanta G, Cescato A, Bonin S, et al. Bioethics considerations for 

medical research in human archive tissues: the point of view of the 

researcher” Virchows Arch 2008;453:117-9. 

2 Hainaut P., Bevilacqua G, Bosman F, et al. The role of the pathologist 

in tissue banking: European Consensus Expert Group Report. 

Virchows Arch. 2010 Feb 16. [Epub ahead of print] PubMed PMID: 

20157825. 


Quality control in biobanking 

G. Bevilacqua 

Division of Surgical, Molecular and Ultrastructural Pathology, University 

of Pisa and Pisa University Hospital, Italy 

Introduction. Over the past 20 years, biobanking of human 

specimens has become a central activity underpinning all 

aspects of biomedical research as well as the development 

of personalized medicine. Biobanking encompasses a wide 

range of specimen types and sample collection designs, ranging 

from population-based biobanking of specimens from 

healthy subjects in large, epidemiological cohorts to specific 

biobanking of diseased tissues obtained in the course of clinical 

interventions. Human tissue biobanking is of particular 

importance for implementation of novel biomarkers into 

clinical trials, as well as for the application of a wide range 

of new technologies (-“omics”) to the discovery and validation 

of new, molecular patterns of disease. Heterogeneity and 

variability of pre-analytical practices is a major source of error 

in analyzing biobanked specimens. In recent years, large international 

efforts have converged towards the harmonization 

of standard operating procedures for biobanking, providing a 

basis for improving reproducibility and comparability of molecular 

data as well as for designing large, multicentric studies 

involving specimen exchanges among different centers. The 

most critical steps in the workflow of biospecimen acquisition 

and annotation for biobanking involve hospital pathologists. 

Pathology is the cornerstone of tissue biobanking. The 

most basic minimal standard for any biobanking operation 

is to identify and define the nature and origin of the tissues 

to be kept in the biobank. This requires specialized pathology 

expertise. Furthermore, pathologists also make decisions 

on what should be biobanked, making sure that the timing 

of all operations is consistent with both the requirements of 

clinical diagnosis and the optimal preservation of biological 

products. Pathologists also play a central role in the design 

of studies involving banked biospecimens and in the dialogue 

between clinicians and researchers. The rapid development 

of biobanking as an essential process in translational research 

and personalized medicine places strong demands on the work 

of the pathologist. This document summarizes the conclusions 

of a Pathology Expert Group Meeting that took place 

in Munich in December 2008 within the European Biological 

and Biomolecular Research Infrastructure (BBMRI) Program. 

The experts have considered all aspects of the involvement of 

the pathologist in the biobanking process. They also discussed 

the impact of biobanking on pathology practice. The recommendations 

developed in the document are aimed at providing 

guidance for pathologists as well as for institutions hosting 

biobanks on how to better integrate and support pathological 

activities within the framework of biobanks that fulfill international 

standards. 

Scope and definition. 1. The focus of the working group is 

the banking for research of human tissues in a clinical context. 

This activity is hereby defined as “tissue banking”. It includes, 

but is not limited to, the banking of residual specimens obtained 

in the course of clinical procedures as well as of “postmortem 

material.” 2. Tissue banking is a chain of operations 

that includes informing patients and obtaining the proper 

consent (depending on local requirements), data acquisition, 

tissue procurement, annotation, preservation, storage, quality 

control, cataloguing, managing of access, processing and 

distribution. Pathology expertise is required at several steps. 

Tissue banking also requires expertise in cryobiology, quality 

management, legal/ ethical aspects, project management, staff

lectures 

management, administration and networking. 3. “Pathology 

archives” represent a special type of tissue repository that may 

support tissue banking, provided that they fulfill required standards 

with respect to a. documentation of variations; b. cataloguing; 

c. rules of access; d. fulfillment of legal requirements 

for use as research resource. The primary role of these archives 

is to document diagnosis and to support later/metachronous 

diagnostic analyses but they should be developed in a way that 

allows them to fulfill roles in research as well. 

Tissue banking: critical role in articulating translational 

research and personalized medicine. 1. Tissue banking in 

a clinical context is essential for the procurement of high 

quality samples for translational research aimed at biomarker 

discovery and validation as well as identification of new 

targets for therapy. It is therefore a strategic activity for research 

and innovation in biomedicine. 2. Tissue banking is 

critical for implementing and applying biomarkers in clinical 

practice. It lays the foundations for the discovery of new 

targets for therapy and for drug discovery. It sets conditions 

and procedures allowing patients to benefit from new developments 

in biomarkers as well as personalized medicine and 

is therefore beneficial for future diagnosis and treatment and 

for public health. In this vision, each patient contributes to 

the care that will be provided to the future patients. 3. Translational 

research on biomarkers encompasses three overlapping 

phases: discovery, validation, and implementation. Each 

phase has different requirements in terms of tissue banking. 

4. Discovery phase is aimed at identifying biomarkers and 

molecular targets for therapy, establishing their prevalence 

and formulating hypotheses on their biological and medical 

significance in ex vivo analyses. This requires access to well 

annotated and pathologically reviewed case series, either 

based on specimens collected and processed in the course 

of clinical diagnostic activities or in specific tissue collection 

protocols. 5. Validation phase is aimed at demonstrating 

the effect and significance of a potential biomarker. This 

requires applying ex vivo analyses within study designs with 

adequate epidemiological and statistical power. Such designs 

may be comparable to those of clinical trials except that they 

do not necessarily imply de novo specimen collection using 

invasive procedures. In a number of cases, these studies can 

be constructed using retrospective or prospective collections. 

6. Implementation phase is aimed at translating biomarkers 

into clinical practice in affordable, cost-effective conditions 

and at integrating new biomarkers into diagnostic practice. 

This requires applying biomarkers to a large series of specimens 

collected using standard operating clinical protocols. 

Role of the pathologist. 1. The pathologist has an essential 

role in tissue banking. His medical and scientific expertise is 

required at two distinct phases in the process of tissue banking: 

a. in making diagnostic decisions, providing specific annotations 

and overseeing specimen procurement and preservation, 

and b. in reviewing specimens and providing information 

prior to specimen processing and distribution to research laboratories. 

2. Through his role in tissue banking, the pathologist 

is a key actor in the continuity between research and medical 

care. 3. The pathologist adds value and expertise to the definition 

of the banked tissue and is a critical scientific contributor 

to research carried out on the specimen. 4. The pathologist 

validates the appropriateness of the banked tissue specimen 

and its use for a particular research purpose, excluding conflicts 

with diagnostic purposes. 5. The pathologist has a key 

role as custodian of the banked specimens. Tissue collections 

are best developed in the context of a pathology department 

or pathology service. 

201 

Role of institutions. 1. Tissue banking is not the exclusive responsibility 

of pathology departments. It should be run in the 

context of institutions (mainly hospitals or universities) that 

are responsible for providing the whole chain of expertise and 

the organizational frame required for tissue banking. 2. Institutions 

are responsible for the maintenance, sustainability, and 

accessibility of tissue banks, adequate level of training of the 

staff and the protection of patient rights. Full cost calculation 

is an essential step in guaranteeing the sustainability of the 

tissue bank. 

Tissue banking in clinical trials. 1. Clinical trials offer a 

wide range of designs with added value for the discovery, 

validation and implementation of potential new biomarkers. 

2. Using biomarkers is critical for the interpretation of many 

therapeutic trials in particular for defining the characteristics 

of responders vs. non-responders. 3. In future medical care, 

biomarkers will become mandatory for allocating patients 

to appropriate therapeutic protocols. 4. The participation of 

a biobank into a clinical trial should obey to the same strict 

technical, legal, and ethical standards independently of the 

type of promoter, academic, or industrial. 

Improving standards for tissue banking within clinical 

practice. 1. There are technical differences in current standards 

for tissue processing in pathology practice and in tissue 

banking. 2. Many protocols used in tissue banking, e.g., for 

duration of fixation, optimal time for preservation and duration 

of storage, are mainly based on experience rather than 

evidence. 3. There is a need for more adequate markers of 

quality for the tissue-banking process for the qualification of 

banked tissue specimens for specific research applications. 

4. Discovery, validation, and implementation of biomarkers 

and therapeutic targets in the clinics require a very large series 

of specimens with inter-laboratory comparison. Such studies 

need strong networking between dedicated platforms using 

harmonized, comparable protocols. 

Incentives for increasing the participation of pathologists. 

1. Tissue banking is an important mechanism by which pathologists 

participate in generating and increasing knowledge 

in biomedicine. 2. In many instances, the involvement of the 

pathologist adds scientific value to the banked specimens beyond 

the requirements of routine diagnosis. This added value 

corresponds to an intellectual property. 3. Tissue-banking 

activities entail considerable costs and demands on pathology 

staff time. 

Conclusions and perspectives: a strategic vision for tissue 

banking in Europe. Today, tissue banks have a key role in 

the process of biomarker and drug target discovery through the 

procurement of annotated specimens to innovative research 

programs. In addition to this research role, the use of cellular 

and molecular biomarkers is rapidly becoming a standard part 

of hospital pathology practice and of therapeutic decision 

schemes. Tissue banking is the key mechanism for pathologists 

to get involved in translating newly discovered biomarkers 

into clinical practice. Furthermore, tissue banking will rapidly 

become an intrinsic part of pathology requirements in the 

context of standard clinical care. Given its strong linkage with 

clinical activities, tissue banking is best performed at the local 

level, and its sustainability requires investment in infrastructure 

at the local and/or regional and national levels, to avoid 

duplication of effort and achieve critical mass necessary to address 

major academic research programs, as well as to secure 

a strong position in addressing the needs of industry. Therefore, 

tissue banks must be organized in operational networks. 

Implementation of biomarkers will require large networks interconnecting 

tissue banks, analysis and distribution platforms

202 

and several other data resources such as databases of clinical 

information and population-based disease registries. Biobank 

networks should have fully documented standard operating 

procedures, share tissue bank catalogues, and clear rules for 

access. They should also be able to run research projects based 

on collections developed in several tissue banks. Such projects 

may be retrospective (using previously banked specimens) or 

prospective. Running the same, hypothesis-driven collection 

protocol through a large network of tissue banks that adhere 

to the same standards will allow assembling large case series 

addressing a wide range of clinical conditions. In developing 

such protocols, the diversity of European populations and 

ecological contexts is an asset for the design of sophisticated 

case–case comparison studies. To achieve this vision, it is essential 

to perform innovative research on improving all aspects 

of specimen processing, including the development of quality 

controls applicable to retrospective collections. This requires a 

dedicated effort from funding agencies and from the scientific 

and medical publication community. Training of highly qualified 

tissue-banking professionals will increase the standards 

of tissue banking as well as the recognition of tissue banking 

as an integral part of biomedicine. This will also facilitate the 

development and dissemination of a corpus of harmonized, 

evidence-based tissue-banking procedures. 

Biobanking and SIAPeC-IAP 

O. Nappi 

Past President SIAPEC-IAP; Unit of Anatomic Pathology, “Antonio 

Cardarelli” Hospital, Naples, Italy 

In 2005, the Executive Committee of Italian Society of Pathology 

and Cytopathology, IAP Italian Division (SIAPEC- 

IAP) decided to institute a “Project tissue biobanking task 

force Group”. At that time, the Biobanking was becoming 

an emerging issue with several problems and much confusion. 

The main problem was that,,although cells and tissues 


are daily managed by pathologists, these latter looked to be 

excluded from any kind of initiatives concerning the topic, 

being confined to a marginal role. 

The first goal of the Group has been that to involve “the pathologist” 

and possibly put him at the center of any discussion, 

guidelines extension or tissue banking management project, 

on the basis of the well recognized competence in selecting, 

preserving, storing and studyng human cells and tissues. 

Following these ideas, the Group was invited to contribute to 

write the “Guidelines for the Institution and certification of 

Biobanks”, a document commissioned by Italian Government 

in 2006 1 . 

In 2008 the Group published a thematic issue of Pathologica 2 

where several papers on national and international experiences, 

legal and ethic aspects, sample conservation and quality 

control, informatics,cost analysis and other related topics 

have been presented by most of the Group’s members. All the 

published papers have been presented in Italian and English 

languages. 

Moreover, Congress Sessions, Specific educational Courses, 

National and International meetings have been organized 

under the patronage of the SIAPEC-IAP and his Biobanking 

Group. 

Next step is the constitution of a permanent “SIAPEC-IAP 

Working Group on Biobanking” connected also to European 

Society of Pathology, that should continue to produce documents 

and organize educational events, having the perspective 

of building a progressive Italian biobanking network within a 

wider European one, that can involve the majority, if not all, 

of national anatomic pathology services. 

references 

1 Linee guida per l’istutuzione e l’accreditamento delle biobanche. 

Presidenza del Consiglio dei Ministri. Comitato Nazionale per la biosicurezza 

e le biotecnologie. Rapporto del Gruppo di lavoro. 19 Aprile 

2006. 

2 AA.VV. Tissue banking: a tool in Anatomic Pathology. Pathologica 

2008;100:43-148. 

The cancer crisis in Africa: diagnostic anatomo-pathology 

using a multidisciplinary approach 

ultrasound and fine needle aspiration: 

a low-costs multidiciplinary approach 

S. Guzzetti 

Department of Histopathology, Ospedale Evangelico Valdese – ASL 

TO1, Turin, Italy; Member of “Patologi Oltre Frontiera”, NGO 

Recently, the role of pathology in developing countries has 

grown and diversified: the increasing demand for projects 

not only dedicated to the improvement of the diagnostic level 

in low-resource settings but also the mandatory involvement 

of pathology in establishing specific programs of preventive 

medicine is making our specialization even more essential in 

these contexts. 

Regardless the project type, the major problems are due to the 

shortage of skilled personnel and to the rational use of available 

economical resources. 

Since 1999, the Association “Patologi Oltre Frontiera, NGO” 

(APOF) has developed projects dedicated to the improvement 

Moderators: F. Bonetti (Verona), C. Clemente (Milano) 

of pathology in developing countries in cooperation either 

with local institutions or with other Italian specialists in order 

to provide sustainable and multidisciplinary diagnostic tools. 

In some of these projects, the combined use of ultrasound with 

fine needle aspiration (FNA) made possible a quick, safety and 

cheap diagnose for a large group of detectable pathologies. 

In Mwanza, Tanzania, APOF restructured and reorganized 

the Department of Pathology of the local referral hospital, the 

Bugando Medical Center, also improving the cytology and 

applying it first for a specifically set outpatient clinic for FNA 

on palpable masses, then for ultrasound-guided FNA even for 

inpatients. 

Similarly, at the Mtendere Mission Hospital in Chirundu, 

Zambia, APOF not only provides for the building of a new 

Pathology Department, but also organized several missions of 

pathologist and radiologists in order to introduce ultrasound 

and cytology, together with a program of remote diagnostics 

through a system of telepathology.

lectures 

The use of ultrasound has proven essential in another project 

in Bethlehem, Palestine, where, together with the local Ministry 

of Health and the Italian Cooperation, APOF is developing 

a pilot program for breast cancer screening. 

In conclusion, multidisciplinarity and low-cost technologies 

can play a key role in the improving of diagnostics in developing 

countries. 

An Italian-Palestinian cooperation project. 

The role of pathology dept. in the prevention 

and treatment of cancer: the experience of Beit 

Jala Government Hospital 

R. Shriam, S. Guzzetti * , D. Fenocchio ** , P. Giovenali ** 

Dept. Of Pathology, Beit Jala Government Hospital, Palestine (West 

Bank); * Associazione Patologi oltre Frontiera, Serv. Anatomia Patologica, 

Ospedale Evangelico Valdese, Torino; ** Associazione Patologi 

oltre Frontiera, Serv. Citologia e Istologia Diagnostica, Ospedale 

“S. Maria della Misericordia”, Perugia 

Introduction. The aims of “Associazione Patologi oltre Frontiera” 

(APOF) a nonprofit organization (NGO), established 

since 1999 are to implement and improve diagnostic oncology, 

prevention and treatment of cancer in developing and 

emerging countries. 

Background. Under agreement between Bethlehem Municipality, 

the Provincial Authority of Venice, United Nation Developing 

Program (UNDP), Italian Cooperation: Unità Tecnica 

Locale (UTL) Of Jerusalem and Palestinian Ministry of 

Health (MOH), pathologists of APOF assessed the feasibility 

of a project to advance pathology service for the West Bank 

area in Beit Jala Government Hospital (BJGH) and submitted 

a proposal with a 3-years plan to strengthen and stabilize cytological 

and histopathological diagnostics, which are essential 

for the development of oncology services. 

The proposal was accepted by the Ministry of Health and by 

UNDP in February 2006. 

The project started in collaboration with UTL, including the 

construction of the pathology laboratory on 4th floor (160 m 2 ) 

of the Beit Jala Hospital and the technical assistance in the 

program for prevention and early detection of breast cancer 

in West Bank. 

At the starting of the project, despite major efforts by UNDP 

to recruit at least one physician specialized in pathology from 

the Palestinian Territory, the position for pathologist to run 

and manage the pathology service at BJGH was vacant, and 

the laboratory was not yet able to prepare adequate specimens 

1) Dept of Pathology 

histo-cytological cases in BJgh (2006-2010) 

n. of cases malignancy 

n % 

Breast 747 191 25.6 

gastro-intestinal 419 144 34.4 

urinary tract + Prostate 249 70 28.1 

lymph nodes 205 49 23.9 

thyroid 356 29 8.1 

lung 129 18 14.0 

gynecopathology 2860 75 2.6 

other 9,414 261 2.8 

total 14,379 837 5.8 

203 

for histopathology; for this reason, a pathologist from Nablus 

Rafidia Hospital was recruited on a part-time contract, as 

consultant, for three years and a Palestinian physician, Dr 

Riad Shriam, was identified to complete his studies in surgical 

pathology in Italy (University of Pisa, Scuola di Specializzazione 

in Anatomia Patologica) and since 2009 he is the head 

of Pathology Dept. of BJGH 

The following staff was recruited and trained during the years 

2006-2009: 

• Two technicians; one of them was trained in Italy for a 3months 

stage in immunohistochemistry and tumor markers. 

• One medical secretary. 

• Three specialist pathologists. 

Another physician has nearly completed the specialty fellowship 

in pathology in Jordan. 

Nowadays the lab is fully equipped with all of the instruments 

needed. 

The Palestinian Ministry of Health, the main partner in this 

project, whose input has been essential for the successful 

maintenance of this project, will provide reagents and disposables 

needed for the pathology lab at BJGH. 

APOF, in collaboration with UTL and MOH, organized in 

April 2009 a training course in Bethlehem with the following 

aims: 

• staff training; 

• development of guidelines ad protocols and procedures of 

breast cancer management. 

The main aim was to raise the capacity of the staff in identifying 

the cytological patterns of breast cancer and precursor 

lesions and to gain knowledge of pathological classification, 

grading, staging and prognostic markers of breast cancer. 

The course consisted in training on: 

• pre-operative diagnosis by means of FNAC and micro-biopsy 

(needle core biopsy); 

• breast surgical pathology; 

• evaluation of prognostic/therapeutic markers and 

• developing local guidelines and protocols on breast cancer 

management, according to European Guidelines. 

In the year 2009, MOH, in collaboration with Italian Cooperation, 

started a national Palestinian program for early detection 

of breast cancer. Bethlehem Governorate was identified as 

pilot area and breast screening by mammography started in 

BJGH with fully involvement of Pathology Dept. 

Results. The reported data are relative to admitted patients, 

outpatients and specimens referred to Pathology Lab of BJGH 

from other 6 hospital in South West Bank, between January 

2006 and May 2010

204 

fnac’s in BJgh (2006-2010). 

2) Screening 

The Breast Unit of BJGH opened in January 2009: since then 

and until April 2010, were performed 1.919 mammograms 

and/or ultrasounds, 103 out of those were diagnosed as suspicious 

or positive (5.4%) and FNA was suggested. FNA was 

actually performed in 57 women; 20 were positive or highly 

suspicious (1.0%), 6 were diagnosed as benign but with uncertain 

malignant potential (0.3%), 23 were negative and 8 

unsatisfactory. 

Conclusion. The APOF cooperation project, in collaboration 

with Palestinian MOH, leaded to the establishment and organization 

of a modern and effective Pathology Dept. A specific 


n. of cases malignancy 

n % 

Breast 232 45 19.4 

thyroid 102 5 4.9 

other 68 34 50.0 

total 402 84 20.9 

Breast pathology cases in BJgh (2006-2010). 

n. of cases malignancy controlled false positive false negative 

fnac 232 45 53 1 5 

core biopsies 23 9 5 0 1 

surgical specimens 492 137 = = = 

total 747 191 

program was developed to train and bring up-to-date physicians 

and technicians of the pathology laboratory at BJGH. 

The main beneficiaries of the project are the Palestinians, 

resident in the West Bank who can benefit from the improved 

health services, the medical staff who received training to better 

fulfill their specialist tasks, and the Ministry of Health who has 

improved medical facilities to serve the Palestinian people 

The general objective to improve the level of advanced medical 

services in the West Bank, has obtained a good support by 

the establishment of this new pathology service, needful in a 

general hospital with surgical and oncological depts. and for 

screening programs. 

Men and women in pathology, the strength of the foundation 

Camillo Golgi: a genius of observation 

P. Mazzarello 

Museo per la Storia dell’Università di Pavia, Dipartimento di Medicina 

Sperimentale, Sezione di Patologia Generale, Università di Pavia, 

Pavia Italy 

Camillo Golgi was born at Corteno (today Corteno Golgi), 

a small mountain village in the North of Italy on the 7 th July 

1843. He studied medicine at the University of Pavia where 

he graduated in 1865. After his graduation, Golgi started his 

clinical activity at the San Matteo Hospital in various medical, 

surgical and dermatological wards. However he soon 

became assistant at the Psychiatric Clinic headed by Cesare 

Lombroso who sparked his vocation to study the brain. Following 

the tenets of the positivist scientific philosophy, advocated 

by Lombroso, anatomical and anthropological data 

became, at that time, the tools by which biology could explore 

neuropsychiatric diseases. Thus Golgi, in collaboration 

with Lombroso, began to investigate the etiology of mental 

and neurological illness from an experimental and antimetaphysical 

point of view. Meanwhile in the free time that 

his hospital duties allowed, Golgi attended the Institute of 

General Pathology under the direction of Giulio Bizzozero, 

the rising exponent of the new experimental medicine which 

had as its emblem the microscope. From Bizzozero, Golgi 

acquired a passion for histological investigation, the direct 

means of penetrating the formidable unknown of the archi- 

Moderators: C. Bondi (Parma), S. Uccini (Roma) 

tecture of the nervous system. Even if three years younger 

than Golgi, Bizzozero thus became his master, patron and 

the “catalyst” of his mind. Under his direction, Golgi began 

to publish works between 1870 and 1872, the most important 

of which were dedicated to the study of the neuroglia and 

which were flatteringly quoted in international literature. By 

1872 Golgi had acquired a solid reputation as a clinician and 

histologist but this was not considered enough to earn him a 

satisfactory position at the University. On 1872 pressured by 

his father, Golgi took part in and won the competition for the 

post of Chief Physician at the Pio Luogo degli Incurabili, a 

hospital for chronic diseases, at Abbiategrasso near Milan. 

Everything suggested that, with Golgi’s arrival in a small 

town hospital, his research activity were about to end for 

good. However after some initial difficulties, Golgi set up a 

rudimentary laboratory consisting of a microscope and a few 

instruments in the kitchen of Golgi’s small quarters. On 16 

February 1873, Golgi in a rush wrote the following words 

to his friend Nicolò Manfredi: “I spend long hours at the 

microscope. I am delighted that I have found a new reaction 

to demonstrate, even to the blind, the structure of the interstitial 

stroma of the cerebral cortex. I let the silver nitrate 

react with pieces of brain hardened in potassium dichromate. 

I have already obtained magnificent results and hope 

to do even better”. This is the first record of the invention 

of the black reaction known nowadays as “Golgi staining” 

or “Golgi impregnation” that was a breakthrough for brain

lectures 

structure research. The black reaction consists of a first phase 

of hardening the tissue in potassium dichromate followed by 

the impregnation of the nervous elements by silver nitrate. 

The final result is a preparation in which the silhouette of the 

nerve cell appear in all its morphological complexity with all 

its ramifications, which could be followed and analysed even 

at a great distance from the cell body. The great advantage 

of this technique is that, for reasons that are still unknown, 

a precipitate of silver chromate randomly stains only a few 

cells in black (usually from 1 to 5%), and completely spares 

other surrounding cells, allowing the individual elements to 

emerge from the nervous puzzle. The discovery of the black 

reaction provided the spark to a truly scientific revolution 

which allowed the morphology and the basic architecture of 

the cerebral tissue to be displayed in all its complexity, thus 

contributing to the foundations of modern neurosciences. 

Golgi remained in Abbiategrasso until January 1876; there he 

discovered the constant presence of the axon in nerve cells, 

the branching of the axon, the presence of striatal and cortical 

lesions in a case of chorea and performed studies on the structure 

of the cerebellum (describing the so-called Golgi cells 

of the cerebellar cortex), and of olfactory bulbs. Meanwhile 

he began to elaborate on a general theory of brain organization, 

the so-called “diffuse nervous net” according to which 

the axons are connected (through direct fusion or intimate 

contact) in a diffuse network along which the nervous impulse 

is propagated. Although this concept was in polemical 

opposition to the “neuron theory”, ironically the indefatigable 

paladin of this theory, the Spaniard Santiago Ramón y Cajal, 

became such by using the Golgi stain. 

After the discovery of the black reaction Golgi became Professor 

of Histology at the University of Pavia in 1876 and 

from 1879 onward, he also became Professor of General 

Pathology and honorary chief with direct clinical responsibilities 

of a medical ward at the San Matteo Hospital. In 1878 

he described two kinds of tendinous sensory corpuscles: the 

Golgi tendon organ (proprioceptors) and the Golgi-Mazzoni 

corpuscles (transductor of pressure stimuli). Then he invented 

the staining method with potassium dichromate and mercuric 

chloride (1878-79), discovered the myelin annular apparatus 

(horny funnel of Golgi-Rezzonico, 1879) and analysed several 

regions of the nervous system in detail providing beautiful 

illustrations of them (Golgi, 1885). Between 1885 and 1892 

he concentrated on studying human malaria. He was soon 

able not only to determine the entire intraerythrocytic cycle of 

development of the malaria parasites for tertian and quartan 

(Golgi cycle), but he also discovered the temporal relation 

between the recurrent febrile bout and the segmentation of 

the parasite (Golgi law). Meanwhile he concentrated on the 

study of kidney histology, histopathology and histogenesis 

(1884-1889) and discovered the important relationship between 

the vascular pole of the Malpighian glomerulus and the 

distal tubule, which plays an important role in the regulation 

of blood pressure. 

A skilled physician who always refused private activity, he 

also published important clinical studies on peritoneal blood 

transfusion, intestinal worm infection, regeneration and pathological 

changes of the kidney. He also observed independently 

from the Swedish histologist Erik Müller, the canaliculi of the 

parietal cells of the gastric glands, often called Müller-Golgi 

205 

tubules. At the end of 1893 he was elected, for the first time, 

Rector of the University of Pavia, and held the position until 

1896. Thereafter Golgi returned to the study of the nervous 

system and using a variant of his black reaction he was able 

to observe, in 1897, a “reticulum” in the cytoplasm of cells of 

spinal ganglia, the so-call internal reticular apparatus, subsequently 

christened the Golgi apparatus or Golgi complex. 

Meanwhile Golgi observed the perineuronal net which constitutes 

a reticular structure enveloping many neurons. 

On the twentieth century, Golgi’s scientific creativity faded. 

His time was divided between new responsibilities in the 

direction of Pavia University (of which he was again made 

Rector from 1901 to 1909) and the Senate of the Italian Kingdom, 

of which he was elected member from 1900. In 1906 

he reached the pinnacle of his international fame, when he 

received the Nobel prize for Physiology or Medicine, which, 

ironically, was also won by his eternal scientific rival Ramón 

y Cajal. 

During the First World War, Golgi directed the Military 

Hospital Collegio Borromeo of Pavia, and promoted the rehabilitatory 

treatment for the war-wounded. In 1918 he retired 

from the University of Pavia at the age of 75, but continued 

to teach Histology as Professor Emeritus until the beginning 

of the 1920’s. 

During his life he was elected honorary doctor of the Universities 

of Cambridge, Geneva, Kristiania (Oslo), Athens and 

Paris (Université de la Sorbonne). He had been Dean of the 

Medical Faculty of the University of Pavia and member of a 

number of international academies and scientific societies. 

He died in Pavia on 21 January 1926. 

In Golgi’s laboratory Carlo Martinotti identified the cell 

named after him in the cerebral cortex, Aldo Perroncito described 

the phases of regeneration in the peripheral nerves, 

Emilio Veratti observed the T system linked to the sarcoplasmic 

reticulum and Adelchi Negri discovered the intraneuronal 

inclusions (the Negri bodies) in animals and humans infected 

with the rabies virus. Many other scientists spent periods 

of study and specialization in Golgi’s laboratory such as 

Giovanni Battista Grassi, the discoverer of the Anopheles 

which transmit human malaria, Antonio Carini who discovered 

the Pneumocystis carinii (recently renamed P. Jiroveci) 

and Fritjof Nansen a Norwegian zoologist and a Nobel Prize 

winner for Peace in 1922. 

references 

Golgi G. Sulla struttura della sostanza grigia del cervello. Gazzetta 

Medica Italiana – Lombardia 1873;33:244-6. 

Golgi G. I recenti studi sull’istologia del sistema nervoso centrale. 

Rivista critica. Rivista Sperimentale di Freniatria e Medicina Legale, 

1875;1:121-30 (first part), 260-74 (second part). 

Golgi C. Sulla fina anatomia degli organi centrali del sistema nervoso. 

Reggio Emilia: Tipografia di Stefano Calderini e Figlio 1885. 

Golgi C. Opera Omnia. Vol. I-III. Fusari R, Marenghi G, Sala L (eds). 

Milano: Hoepli Editore 1903. 

Golgi C. Opera Omnia. Vol. IV. Sala L, Veratti E, Sala G. (eds). Milano: 

Hoepli Editore 1929. 

Mazzarello P, Garbarino C, Calligaro A. How Camillo Golgi became “the 

Golgi”. Febs Letters 2009;583:3732-7. 

Mazzarello P. Golgi. A biography of the founder of modern neuroscience. 

Transl. by and A. Badiani and H. Buchtel. New York: Oxford 

University Press 2010. 

Mazzarello P. The rise and fall of Golgi’s school. Brain Research Reviews 

(in press).

206 

In the reconstruction and internationalization 

process in post-unification Italy 

S. Tugnoli Pàttaro 

Department of Philosophy, University of Bologna, Italy 

Background. “We regret to announce the death of Dr. Giuseppina 

Cattani [1859-1914], lecturer on general pathology, 

first in the university of Turin, later in that of Bologna. Her 

name is associated with that of Tizzoni in the investigation 

of tetanus. She was also the author of several memories embodying 

the results of independent research. The state of her 

health made it impossible for her to continue her labours as a 

university teacher, but she continued to direct the laboratories 

of the civil hospitals and the observation asylum of her native 

town, Imola” 1 . 

After Cattani died, her name seemed to have been consigned 

to oblivion, as evidenced by the fact that it does not appear in 

any general historico-scientific bibliographical dictionary, not 

only abroad but also in Italy. 

Today, however, it is much easier to talk about Giuseppina 

Cattani than it was even only a few years ago. 

Indeed, in gender studies – an area of investigation which 

originated in 1968-70 in parallel with the developing feminist 

movement, first in the United States and then in Europe, and 

which has been intensifying since the 1980s – a broad and 

intense historiographical effort has been undertaken leading to 

unprecedented historiographical findings of great interest, no 

doubt heralding further developments down the line. 

These findings have really lead to the “rediscovery” of documents 

bearing witness to the role that female scientists have 

played in the history of scientific thought. In this process, 

involving an effort to recover forgotten documentary sources, 

Giuseppina Cattani has herself begun to regain in the history 

of science the sort of visibility she certainly deserves. 

For which reason she now receives special mention even in 

foreign dictionaries of female scientists. Suffice it to mention 

in this regard that in a listing of female scientists, she figured 

among the “leading contributors,” along with the French- 

American neurologist Augusta Dejerine Klurapke; more to the 

point, the vast research Cattani has done on tetanus has earned 

for Italy a position as a “leading Western country for pre-1901 

bacteriology research by women” 2 . 

Methods. The method to be used will be as follows. We will 

first provide an outline pointing out key moments in the life 

and training of Giuseppina Cattani as a woman and as a scientist, 

to this end relying on recently discovered documents 

to which we will refer the reader for further study. We will 


Giuseppina Cattani 

Woman and scientist 

then proceed to an interpretation of the primary and secondary 

sources collected, offering a new, contextualized reading of 

Cattani at the historical moment in which she lived. 

Results. We will have two main objectives. 

1) The first of these is to illustrate three closely interconnected 

perspectives from which Cattani as a historical figure can be 

rendered. One such perspective will be that of her scientific 

contribution, recognizing a primary role for her discovery of 

a tetanus vaccine with Guido Tizzoni, a discovery that immediately 

gave her prominence in the international effort in 

search for the best cure against tetanus, and owing to which 

Guido Tizzoni was awarded a Nobel Prize in 1895; the second 

perspective is that of her civil-political commitment, militating 

in the ranks of internationalist and socialist movements 

that university students and scholars across all areas of study 

(from literature to medicine) took part in at that time; lastly, 

we will discuss her role in the women’s liberation movement, 

where she fought for equal rights with men – political rights, 

and especially the right to education and the right to practice a 

profession (the medical profession where she was concerned, 

including research and medical teaching at university) – in a 

society that denied women both types of rights. 

2) Our second objective will be to illustrate the interest that 

Cattani has for us today once we take the three aforementioned 

aspects of her activity and view them in the context of 

the social, political, economic, and scientific ferment that was 

stirring in her time in Europe and especially in Italy. She was 

born on the eve of Italian Unification (1861) and died five 

months after the outbreak of World War I (1914). She thus 

lived at a time when the construction of the Italian state was 

in full swing, a construction that even in the face of enormous 

difficulties, and sometimes of contradictions, was pursued 

according to a specific design, on a political level as well as 

on a cultural and a scientific level, in which last respect the 

effort was to lift the country from its provincialism and make 

it an international player, singling out strategic points around 

which to develop academic research and teaching. Cattani 

partook in full of the élan and civil and scientific commitment 

that distinguished many of her contemporaries, from humanists 

to scientists—but with a huge additional hurdle, that of 

being a woman in a “world without women,” in the words of 

David F. Noble (1992). 

references 

1 Obituary. BMJ 1915;1:577-8. 

2 Creese MRS, Creese TM. Ladies in the Laboratory II: West European 

Women in Science, 1800-1900; A Survey of Their Contributions to 

Research. Lanham, MD: Scarecrow Press 2004, p. 287.

lectures 

Management of Gestational Trophoblastic Disease 

M.J. Seckl 

Gestational Trophoblastic Disease Centre, Charing Cross Hospital 

campus of Imperial College London, UK 

Introduction. Gestational trophoblastic disease (GTD) is a 

spectrum of pregnancy related disorders comprising the premalignant 

conditions of complete (CHM) and partial (PHM) 

hydatidiform moles (HM) through to the malignant conditions 

of invasive mole, choriocarcinoma (CC) and the rare placental 

site trophoblastic tumour (PSTT). The latter three conditions 

are also collectively known as gestational trophoblastic 

tumours or neoplasia (GTN). Sixty years ago, most women 

with this group of diseases could expect to die. However, with 

modern management and careful follow-up protocols, overall 

cure rates can exceed 98% with retention of fertility. This 

success can be explained by 3 factors: 1) the development of 

effective therapies, 2) the use of human chorionic gonadotrophin 

(hCG) as a biomarker and 3) centralization of care. 

What is Hydatidiform Mole (HM) and who gets it? HM 

affect 1-3 per 1000 pregnancies. About 10% of hydatidiform 

moles subsequently transform into one of the malignant forms 

of GTD. HM are abnormal conceptions resulting in excessive 

placental, and little or no fetal, development. HM can affect 

women throughout the reproductive-age range but are more 

common at the extremes of childbearing age. Thus, women 

< 16 years old have a six-fold higher risk of developing the 

disease compared to women aged 16-40, whilst those conceiving 

aged > 50 have a 1 in 3 chance of having a molar pregnancy. 

Interestingly, the previously documented higher incidence 

in women of far-eastern origin, although still greater than for 

Caucasions, is now falling to more closely match the rates 

seen in the UK and other western countries. The reasons for 

this are not clear but might reflect dietary changes. HM can 

also rarely develop (1:100,000 pregnancies) as part of twin or 

multiple gestations. 

How does GTD present clinically? In the United Kingdom, 

most women with HM present with vaginal bleeding and/or 

suspected miscarriage in early pregnancy, prompting a pelvic 

ultrasound examination, although in one observational study 

of 41 women with confirmed CHM, 40% were entirely asymptomatic, 

being detected following routine early pregnancy sonographic 

examinations. The remaining majority presented with 

vaginal bleeding; only 2% reported symptoms of hyperemesis 

and none had any other systemic manifestations. Vaginal bleeding 

in early pregnancy is of course common and is often innocent, 

but such symptoms should precipitate an early ultrasound 

examination. The presence of material in the uterus in the absence 

of a viable pregnancy will lead to uterine evacuation with 

examination of products for identification of pathology. 

How is a diagnosis of GTD made? The grape-like or hydropic 

change most commonly found with CHM occurs mainly in 

the second-trimester and an ultrasound performed at this stage 

shows a classical snow-storm like appearance. However, most 

women develop vaginal bleeding in the first trimester and now 

undergo uterine evacuation around 8-9 weeks of gestation in 

the UK. At this time, there is minimal hydropic change which 

makes early sonographic diagnosis of hydatidiform moles less 

reliable. Two large, recent retrospective studies from centres 

Trophoblast pathologies 

Moderators: E. Fulcheri (Genova), A. Salerno (Bologna) 

207 

in London have reported that correct pre-evacuation identification 

of molar pregnancy by ultrasound in the first and early 

second trimester is achieved in around only 40-60% of cases 

in routine clinical practice. In the largest study, of > 1,000 

patients referred to a regional trophoblastic disease centre, 

only 40% had a pre-evacuation ultrasound diagnosis suggesting 

molar pregnancy, including 80% of complete and 30% of 

partial moles. The sonographic diagnosis in the majority of 

cases was simple miscarriage, with the diagnosis of HM being 

dependent on subsequent routine histological examination of 

the products of conception. The implications of not sending 

evacuated uterine contents for histology are clear, since if the 

diagnosis is not made, subsequent monitoring for malignant 

change is not instituted and such women have a significantly 

increased risk of life threatening complications such as uterine 

perforation and severe haemorrhage; in a retrospective study 

of 51 women with HM following pregnancy termination, 

women without a known histological diagnosis were significantly 

more likely to have subsequent life-threatening complications, 

or require surgical intervention and chemotherapy 

compared to those in whom a histological diagnosis of HM 

made been made. 

Pathology of GTD. It follows from the above that pathological 

diagnosis of HM is essential. All GTD is derived from 

components of the normal human placenta, hydatidiform 

moles (HM) plus CC, and PSTT, representing villous and 

interstitial trophoblast, respectively. Most CHM and PHM 

have distinctive morphological characteristics, although these 

features have changed in recent years with earlier gestational 

ages at evacuation (median 8-9 weeks in the UK). First-trimester 

CHM show characteristic abnormal ‘budding’ villous architecture 

with trophoblast hyperplasia, stromal karyorrhectic 

debris and collapsed villous blood vessels. In contrast, early 

PHM show patchy villous hydrops with scattered abnormally 

shaped irregular villi with trophoblastic pseudoinclusions and 

patchy trophoblast hyperplasia. The morphological distinction 

between non-molar miscarriage (NMM) and PHM can sometimes 

be difficult, since villous dysmorphism may be present 

but NMM do not show trophoblast hyperplasia characteristic 

of PHM. Ancillary techniques may rarely be required including 

immunostaining with p57 KIP2 , the product of cyclin 

dependent kinase inhibitor CDKN1C. This is expressed from 

the maternal allele as nuclear staining of cytotrophoblast and 

villous mesenchyme in placenta of all gestations other than 

androgenetic CHM. In addition, ploidy analysis by in-situ 

hybridisation or flow cytometry can distinguish diploid from 

triploid conceptions, so facilitating the diagnosis of PHM but 

not distinguishing CHM vs diploid non-molar, or molar vs 

non-molar triploidy, which require molecular investigations. 

CC are malignant hCG-producing epithelial tumours demonstrating 

central necrosis and characteristic biphasic architecture 

recapitulating cytotrophoblast-like cells and multinucleate, 

pleomorphic syncytiotrophoblast-like areas; mononuclear 

cells may predominate in some cases, especially post-chemotherapy. 

Intraplacental CC may occur and probably represent 

the source of metastatic CC following term pregnancies. 

Neonatal choriocarcinoma is well-described, with most cases 

now thought to represent metastatic spread from an intraplacental 

choriocarcinoma. PSTT is the malignant equivalent of

208 

extravillous interstitial implantation site-like trophoblast and 

forms uterine lesions with less haemorrhage and necrosis, 

and lower hCG levels, than CC. A specific variant of PSST 

with distinctive hyalinization has been reported, Epithelioid 

Trophoblastic Tumour (ETT) which is clinically thought to 

behave like PSTT, but data are still relatively sparse. 

How are HM initially managed? Since in many cases the diagnosis 

of HM is unsuspected until histological examination, 

it is important that all products of conception from non-viable 

pregnancies and those suspected of molar disease are submitted 

for routine pathological evaluation. In clinically suspected 

cases, initial management is suction uterine evacuation, (sharp 

curettage should be avoided to minimise the risk of uterine 

perforation). Usually, initial evacuation is sufficient to remove 

most molar material and any residual tissue subsequently involutes. 

A second uterine evacuation when there is evidence 

of persisting disease with further vaginal bleeding, regrowth 

of molar material and a plateaued or rising hCG is not usually 

recommended because 70% of patients undergoing a second 

evacuation will still need chemotherapy which is safe and curative, 

and each procedure carries a risk of uterine perforation, 

infection and massive haemorrhage. 

What’s the risk of malignant disease and should women be 

screened? In benign disease, patient hCG levels spontaneously 

return to normal, but in those who develop GTN, the hCG 

concentration plateaus or rises. The risk of malignant sequelae 

following a complete or partial HM is 15% and 0.5% respectively. 

This is detected in almost all cases by regular hCG 

monitoring using an hCG assay capable of detecting all the 

different forms of the hormone that can be produced in cancer, 

with sensitivity and specificity of almost 100%. In general, in 

cancer medicine one would perform a tissue biopsy to prove 

malignancy. However, malignant GTD is highly vascular and 

re-biopsy is contra-indicated, since it may be associated with 

life-threatening haemorrhage. To ensure reliable monitoring 

of hCG levels after a molar pregnancy, all patients in the UK 

are registered with one of three centres: Ninewells Hospital 

(Dundee), Weston-Park Hospital (Sheffield) and Charing 

Cross Hospital (London). Although most other countries do 

not have a centralized screening program, the majority will 

have designated regional centres to manage GTN. 

Any patient with a diagnosis of HM, either clinically or following 

routine histological diagnosis, should be registered 

for surveillance with a specialist centre. In the UK, this is 

usually done by the Gynaecologist either using paper or webbased 

registration. The patient and managing doctors are then 

posted an information pack and the referring hospital asked 

to provide histological material for central pathology review 

where available. Patients then submit regular samples for hCG 

monitoring. The protocols for monitoring hCG differ slightly 

between centres regarding the frequency and type of samples 

required but the principle is to monitor the patient at least until 

hCG levels have returned to normal. At Charing Cross Hospital 

hCG is measured in serum and urine for several months of 

normal values and following this protocol the risk of missing 

treatable disease is about 1:1400. Intriguingly, reactivation 

of molar disease may occasionally occur after a subsequent 

pregnancy, even several years later, therefore repeat hCG 

monitoring at 6 and 10 weeks after any further pregnancy is 

recommended. Following a molar pregnancy, the risk of a 

subsequent mole rises to 1:80, but most women have normal 

pregnancies after their first hydatidiform mole. Other centres 

in the world have advocated using abbreviated hCG followup 

protocols, particularly for partial mole, where the risk of 

malignant progression is lower. However, this increases the 


risk of undetected malignant disease and since hCG testing is 

cheap and prevents life-threatening complications, we do not 

advocate such shortened follow-up. 

Factors that increase the risk of malignant progression of 

HM. Logically, one might expect that hydatidiform moles that 

progress to later gestations before evacuation should acquire 

more genetic changes with increased malignant transformation. 

However, two studies, one of twin pregnancies comprising 

a mole and healthy co-twin and the other in singleton 

molar pregnancies, indicate that gestational timing of molar 

evacuation does not affect the risk of developing malignant 

disease. Conversely, the method of evacuation does appear 

important since procedures that induce uterine contractions 

could theoretically increase the risk of persistent disease and 

systemic spread. Finally, evidence based on retrospective series 

of patients from the UK suggest that the hormones in the 

combined oral contraceptive pill (OCP) may increase the risk 

of malignant sequelae in a subset of women in whom hCG 

levels remains raised, therefore UK centres currently recommend 

avoidance of the OCP until hCG levels have returned to 

normal. This area remains controversial, however, with data 

from other countries suggesting that OCP use may be safe. 

Who requires chemotherapy after HM? Most patients will 

exhibit plateaued or rising hCG levels indicative of GTN 

(usually invasive mole or choriocarcinoma) with or without 

vaginal bleeding. If bleeding is severe this is, in itself, an indication 

for chemotherapy to reduce haemorrhage even if the 

hCG level is falling. Women with hCG levels > 20,000 IU/l 

one month after molar evacuation are at risk of uterine 

perforation and chemotherapy is required to help preserve 

fertility, and histological diagnosis of choriocarcinoma or placental 

site trophoblastic tumour, or the presence of metastases 

should prompt urgent referral for treatment. The commonest 

metastatic disease site is lung, which may be associated with 

dyspnoea, cough, haemoptysis, and/or chest pain, but any site 

can become involved. Consequently, any woman of childbearing 

age presenting with possible metastatic disease should 

have GTN included in the differential diagnosis. A positive 

serum or urine hCG test will suggest the diagnosis and should 

prompt referral to a GTD centre. 

What happens to patients referred for specialist treatment? 

In the UK treatment is provided at two specialist centres 

(Sheffield and London), with similar treatments offered 

in many countries. In order to determine the chemotherapy 

regimen required, women are assessed to estimate their risk 

of having disease which might become resistant to single 

drug therapy with methotrexate. Risk scoring will usually be 

determined based on history, examination, serum hCG concentration, 

Doppler pelvic ultrasound and chest radiograph. 

About 2/3 of women with low risk (score 0-6) disease will 

be cured with methotrexate alone, whilst women at high risk 

(score > 7) require combination drug chemotherapy, usually 

involving etoposide, methotrexate and Dactinomycin alternating 

weekly with cyclophosphamide and oncovine (EMA/CO). 

Analysis of UK data reveals that the vast majority of women 

following molar pregnancy have low-risk disease, since they 

are on hCG surveillance and the onset of malignant disease 

is detected early. They receive methotrexate injections intramuscularly 

alternating daily, with folinic acid tablets, for 

one week, repeated every two weeks. Therapy is continued 

until the hCG has been normal (< 5 IU/l on the Charing Cross 

hCG assay) for 6 weeks. This regimen is well tolerated, with 

only 2% suffering troublesome side-effects such as mouth 

ulcers and sore eyes, which are managed with mouthwashes 

and hypromellose eyedrops respectively, and sometimes by

lectures 

increasing the folinic acid dose. Patients are admitted for their 

first cycle of methotrexate due to the potential risk of bleeding. 

Additional treatment courses are usually administered by 

a practice nurse, GP or local hospital. 

Response to therapy is assessed by a falling hCG serum concentration 

monitored twice weekly. In one third of women, 

treatment is changed either because of drug resistance or, very 

occasionally, severe toxicity (mouth ulcers or methotrexateinduced 

serositis). Those developing methotrexate resistance 

at relatively low hCG concentrations (hCG < 100 IU/l) are 

usually cured with Dactinomycin, which is slightly more 

toxic, causing hair loss, myelosuppression, mouth ulcers and 

nausea. The remaining resistant patients, and the occasional 

patients not cured by Dactinomycin, are effectively salvaged 

with EMA/CO chemotherapy. This requires an overnight hospital 

stay every two weeks, and is more toxic, inducing alopecia, 

myelosupression, lethargy, nausea and other short-term 

problems. Moreover, in contrast to methotrexate which has 

no long-term toxicity, EMA/CO hastens the menopause by 

about 3 years, and increases the risk of a second malignancy 

by about 1.5 fold compared to the general population. None 

of the therapies affects fertility and the overall outlook is excellent 

with an almost 100% cure rate for women developing 

GTN after an HM based on a study of 485 patients with GTN 

following HM. 

Presentation and Management of High Risk GTN. Most 

high risk GTN patients present with multiple metastases 

months or years after the causative pregnancy which could 

have been of any type. Symptoms and signs will vary depending 

on the location of disease. Patients with brain metastasis 

may present with seizures, headaches or hemiparesis whilst 

those with lung metastasis or disease in the pulmonary vasculature 

might have haemoptysis, shortness of breath and/or 

pleuritic chest pains. Menstrual irregularity may be present but 

is not universal, so unless clinicians consider GTN in the differential 

of metastatic disease and measure the serum or urine 

hCG the diagnosis can be overlooked. If the hCG is raised, the 

patient should be immediately discussed with the nearest GTD 

centre regarding further management. Imaging investigations 

should include CT body, MRI brain, Doppler ultrasound and 

MRI pelvis. If the brain scan is normal then a lumbar puncture 

to assess the CSF:serum hCG ratio (normal less than 1:60) can 

help to exclude occult CNS disease. Biopsy of these highly 

vascular tumours should be avoided to prevent life threatening 

haemorrhage. However, where a lesion is easily accessible and 

bleeding can be controlled then excision biopsy may be helpful. 

This is particularly important if a PSTT or non-gestational 

tumour might be present, since their management differs from 

gestational choriocarcinoma. Fortunately, PSTT has a distinct 

histological appearance and comparison of microsatellite polymorphisms 

in the tumour with DNA from the patient and her 

partner can determine whether the tumour is gestational. The 

phenotypic appearance of the tumour is not always reliable 

and rarely non-gestational carcinomas may appear morphologically 

very similar to gestational choriocarcinomas, and 

conversely, the latter can occasionally mimic other epithelial 

tumours. Chemotherapy is effective at curing the gestational 

tumours whilst the chance of survival from a non-gestational 

tumour reflects the primary site of origin. 

The patients scoring over 7 are at high risk of developing drug 

resistance and so are very unlikely to be cured with single 

agent chemotherapy. Consequently, several different multiagent 

therapies have been developed. At Charing Cross, after 

many years of progressive experience, a regimen was developed 

consisting of etoposide, methotrexate and actinomycin 

209 

D (EMA) alternating weekly with cyclophosphamide and 

vincristine (CO). This has been widely adopted worldwide 

because it appears to be effective with predictable and easily 

managed short-term toxicity. Indeed, a retrospective comparison 

from the Korean GTD centre’s experience of MFA, MAC, 

CHAMOCA with EMA-CO demonstrated a remission rate 

of 63.3% (31/49), 67.5% (27/40), 76.2% (32/45) and 90.6% 

(87/96), respectively. The EMA/CO regimen requires one 

overnight stay every 2 weeks and causes reversible alopecia. 

It is myelosuppressive but G-CSF support helps to maintain 

neutrophil count, treatment intensity and avoid neutropaenic 

febrile episodes. 

The cumulative 5-year survival of patients treated with this 

schedule varies between 75-90%, and of 272 cases at Charing 

Cross was 86.2% (95% CI 81.9% to 90.5%). While these results 

were good, the presence of liver or brain metastases correlated 

with only 27% or 70% long-term survival, respectively 

and was just 10% with both liver and brain metastases. The 

reasons why these patients have adverse outcomes is unclear 

but most did not have a prior HM, were not registered for 

follow-up and consequently presented with extensive disease. 

Furthermore, many deaths occurred soon after admission from 

haemorrhage or metabolic complications of overwhelming 

disease. Indeed, if deaths within 4 weeks (before adequate 

chemotherapy can be given) are excluded, survival of patients 

with brain metastasis is similar to other patients. The situation 

with liver metastasis may be similar; of 37 patients with liver 

metastasis treated between 1977-2005 at Charing Cross, overall 

survival had increased to approximately 50% at 5 years 

but if early deaths were excluded, survival was nearly 70% 

(ISSTD Conference 2009). In addition to disease extent, other 

factors associated with poor outcome include the type of, and 

duration from, the antecedent pregnancy and the prior use of 

chemotherapy. 

To reduce early deaths in patients with very advanced disease, 

we have found that commencing chemotherapy gently with 

low dose etoposide and cisplatin (100 mg/m 2 and 20 mg/m 2 , 

respectively for two days) combined with dexamethasone 

24 mgs in 24 hours to diminish tumour oedema has been 

helpful. Further details on the management and modifications 

of treatment required for these and other challenging clinical 

situations such as brain metastasis and pulmonary failure 

are beyond the scope of the present review but are contained 

within the following references. 

Similar to low-risk disease, therapy is continued for 6 weeks 

of normal hCG values or 8 weeks if poor prognostic features 

such as liver or brain metastases are present (19). Patients are 

then re-imaged to document the post-treatment appearance for 

future comparison. Removal of residual masses is unnecessary 

as it does not reduce the risk of recurrence which is less 

than about 3%. 

Follow-up post-chemotherapy. Post-treatment, patients are 

followed-up with hCG measurements weekly for 6 weeks, 

two-weekly for 3 months and then with diminishing frequency 

until just 6-monthly urine samples are requested according to 

the Charing Cross protocol In the UK, the follow-up continues 

indefinitely since insufficient data is available to determine a 

safe time to stop, but is variable in other countries. Of 1708 

GTN patients at Charing Cross Hospital, including women 

presenting after non-molar pregnancies, the overall relapse 

rate was 3.5%, most of which occurred within the first year 

post-treatment. Therefore, women are advised not to become 

pregnant for 12 months since this may mask early detection 

of relapsed disease. Fertility is unaffected by either low risk 

methotrexate or high risk combination agent chemotherapy

210 

with EMA/CO. However, the latter brings forwards the date 

of the menopause by about 3 years. Second cancer are also increased 

by combination agent chemotherapy by about 1.5 fold 

compared to the general population but single agent therapy 

has no measurable effect. 

Survey on the incidence of gestational 

trophoblastic disease in histopathology: 

rare or underdiagnosed? 

A. Salerno 

Bologna 

Background. The epidemiology of gestational trophoblastic 

disease (GTD) is not well understood. Despite extensive epidemiological 

data spanning more than 50 years, the extent to 

which genetic and environmental factors including race, age 

and geographic location influence the variably in reported 

differences in incidence rates for GTD throughout the world 

is uncertain. Three obstacles limit the interpretation of most 

published studies: case definition, case detection and identification 

of the population at risk. 

Case definition: many reports lack a precise and reproducible 

case definition of the disease entities encompassed by the 

term “gestational trophoblastic disease”. Until recently there 

was no universally accepted classification for GTD. Several 

systems continue to be used for staging, including the World 

Health Organization(WHO) Scoring Index, the FIGO system 

and others. WHO currently divides GTD variants into hydatiform 

mole, choriocarcinoma, placental site thophoblastic 

tumour, miscellaneous trophoblastic tumour (exaggerated 

placental site, placental site nodule, or plaque), and unclassified 

trophoblastic lesions. Gestational trophoblastic neoplasia 

includes invasive mole, choriocarcinoma, and placental 

site trophoblastic tumor. Low risk patients according to the 

FIGO system will follow the same terapies regardless of the 

histologic features. Likewise, treatment for a trophoblastic 

pulmunary nodule would be the same regardless the initial 

histological feature of the uterine contents. An accurate nosologic 

definition of GTD is important to understanding its 

epidemiology, but histologic classification schemes may have 

little clinical usefulness. 

Case detection: published reports are subject to errors in ascertaining 

cases of GTD disease. Over-reporting of pregnancies 

involving GTD relative to other pregnancies can occur in 

hospital-based studies, especially in less developed countries 

because patients with problem pregnancies or cancer are more 

likely to receive hospital care than are those with uncomplicated 

deliveries, which may occur routinely at home. Underreporting 

of cases may also be common: hydatiform moles 

may be spontaneously expelled in many women who never 

receive medical attention and choriocarcinoma may not be diagnosed 

in women who died without medical care or without 

pathological diagnosis. 

Identification of the population at risk: reported rates of GTD 

are difficult to interpret because different denominators are 

used in published studies since only pregnant women are at 

risk of GTD, traditional census statistics, which do not take 

fertility levels into account, are inappropriate for use in calculating 

incidence rates. The preferred denominator for women 

at risk of GTD disease is all women who have conceived. 

This number is not known for populations and several approximations 

have been used as denominators. The number 

of pregnancies usually includes live births, stillbirths, and 

abortions (and ectopic pregnancies) and this represents the 


closest approximation to the population at risk. The number of 

deliveries is not so good as a denominator because it excludes 

spontaneous abortions and induced abortions. The number of 

live births is the poorest approximation of the population at 

risk because it excludes still more conceptions at risk, but it 

can be the only or the most complete information available. 

Since different denominators have been used in calculating 

the incidence rates, comparison of rates may be misleading. 

To the extent that denominator underestimate the size of the 

population at risk, estimates of the incidence of GTD will 

be too high. Use of births or live births in hospitals as the 

denominator, especially in regions where childbirth at home 

is customary, may account in part for high rates of gestational 

trophoblastic disease reported from less-developed countries. 

Populations-based studies relying on centralized pathology 

institute or comprehensive hospital surveillance should provide 

the most accurate estimates of the incidence of GTD. 

Several epidemiological studies using cases recorded by regional 

and national registries have been published. They are 

clearly superior to hospital-based studies, but in some cases 

only malignant variants were recorded and other cases without 

histologic confirmation were excluded. Few have identified 

all GTD cases by histological sub-type, and all potential risk 

factors including dietary, environmental gravidity, parity, age 

at diagnosis and ethnicity have not always been recorded. 

Nevertheless, because population-based registries permit the 

calculation of incidence rates using women residents, live 

births and pregnancies within a well-defined geographic region, 

comparison can be made across registries through the 

world. The reported incidence of gestational trophoblastic 

disease varies dramatically among different regions of the 

world. The published incidences, wich express the rates per 

1000 live births, range from 0.7 in Australia to 4.6 in Hawaii. 

Some of the highest rates were reported from hospital-based 

studies in Indonesia, the Philippines and Mexico. There is 

little epidemiological data from the Indian sub-continent, with 

the exception of a reported rate of choriocarcinoma of 19.2 per 

1000 pregnancies. In North America and in Europe the rates 

of hydatiform mole and GTN are approximately 0.5-1/1000 

pregnancies and 0.2-0.7/1000 pregnancies, respectively. Italian 

studies reported rates of 0,8 hydatiform mole/1000 deliveries 

and 1GTD/1500 pregnancies. 

The aim of this study is to recognize the difficulties in collecting 

data from different type of Hospital in different part 

of Italy and to compare the rates obtained with reported data, 

using different denominators as pregnancies, deliveries and 

live births. 

Methods. Pathologists (mostly belonging to APEFA-Gruppo 

Italiano di Anatomia Patologica dell’Embrione, del Feto e dei 

loro Annessi) from hospitals located in many regions of Italy 

were asked to retrieve from their hospital records the number 

of histopathological diagnosis of Complete Hydatiform Mole, 

Partial Hydatiform Mole, Gestational Choriocarcinoma and 

Placental site Trophoblastic Tumour, excluding referral cases. 

They were asked also to provide the number of total pregnancies 

(live births, still births, spontaneous abortions, induced 

abortions) registered in their hospital. Public official data 

from Regions or National statistic registries were also used 

when available. 

Pathologists and Hospitals. Giovanni Botta, Ospedale 

Sant’Anna, Torino; Francesca Garbini Ospedale Careggi, 

Firenze; Giovanni Angeli Ospedale S. Andrea, Vercelli; 

Mario Abrate Ospedale di Savigliano (CN); Gaetano Pietro 

Bulfamante Università di Milano, Polo S. Paolo Milano; Valeria 

Lucchini Ospedale San Gerardo Monza; Gertrud Fichtel

lectures 

Azienda sanitaria dell’Alto Adige, Bolzano; Yuri Musizzano 

AOU S. Martino, Università di Genova; Luigi Caliendo ASL 

2 savonese Ospedale S. Paolo, Savona; Cristina Vignale 

Ospedale Città di Imperia, Imperia; Massimo Palladino EO 

Spedali Galliera Genova; Francesca Saro ASL 4 Chiavarese 

Ospedale di Sestri Levante (GE); Eugenio Merlo Ospedale Civile 

Padre Antero Miconi, Genova; Filippo Licausi Ospedale 

S. Corona Pietra Ligure (SV); Gianfranco Carfagna Ospedali 

Civili di Sanremo; Marina Gualco Ist. Nazionale Ricerca sul 

Cancro-IST, Genova; Maria Paola Bonasoni IRCCS Istituo 

Gianna Gaslini, Genova; Tommaso Ragusa A.O. Villa Scassi, 

Genova; Paolo Dessanti Ospedale S. Andrea, La Spezia; 

Daniela Danieli Ospedale ASL n. 6, Vicenza; Tiziana Salviato 

Ospedale S. Maria Degli Angeli, Pordenone; Adriano 

Zangrandi Ospedale Civile, Piacenza; Giovanna Giordano, 

Università-Azienda Ospedaliera, Parma; Maria Carolina Gelli 

Arcispedale S. Maria Nuova, Reggio Emilia; Francesco 

Rivasi Università di Modena; Angela Salerno Ospedale Maggiore 

AUSL Bologna; Luigi Serra Ospedale di Forlì; Evandro 

Nigrisoli Ospedale Bufalini, Cesena; Silvia Zago Ospedale 

Civile S. M. delle Croci, Ravenna; Monica Ricci Ospedale Infermi 

Rimini; Vincenzo Nardini Azienda Ospedale-Università 

di Pisa; Fiovo Marziani Ospedale di Terni; Evelina Silvestri 

Ospedale Camillo Forlanini, Roma; Gianfranco Zannoni Università 

Cattolica, Roma; Maria Teresa Ramieri Ospedale di 

Frosinone; Ugo Buonocore AORN Cardarelli, Napoli; Maria 

D’Armiento Università di Napoli Federico II; Leonardo Resta 

Anatomia Patologica Policlinico Universitario, Bari; Gabriella 

Ottoveggio Presidio Ospedaliero G. F. Ingrassia, Palermo 

Results. The data retrieval has resulted in various difficulties. 

The most frequent causes of incompleteness or lack of 

data on the numbers at numerator were: partial computerization; 

the computer system has changed over time and the old 

archives are not readily available; encoding specific disease 

has changed over time and may be ambiguous or incorrect; 

diagnoses (especially partial mole) not confirmed clinically or 

by other means can not be counted or could have been coded 

differently. 

The most frequent causes of incompleteness or lack of denominator 

data were: data (most often those of recent years) have 

not yet been developed or are not available; the aggregated 

data are available but in a different way from hospital to hospital 

(live births and stillbirths combined) and not comparable; 

the data from different hospitals referring to the same institute 

of pathology are not comparable. 

In preliminary data the rates of complete hydatiform mole 

and choriocarcinoma are approximately 0.6/1000 pregnancies 

(0.9 /1000 deliveries) and 0.02/1000 pregnancies (0.04/1000 

deliveries) respectively. 

references 

Fasoli M., Ratti E, Franceschi S, et al. Management of gestational trophoblastic 

disease: results of a Cooperative study. Obstet Gynecol 

1982;60:205. 

Golfier F, Raudrant D, Frappart L, et al. First epidemiological data from 

the French Trophoblastic Disease Reference Center. Am J Obstet 

Gynecol 2007;196:172e1-e5,. 

Grimes DA. Epidemiology of gestational trophoblastic diseases. Am J 

Obstet Gynecol 1984;150:309-18. 

Ngan S, Seckl MJ. Gestational trophoblastic neoplasia management: an 

update. Curr Opin Oncol 2007;19:486-91. 

Smith HO. Gestational trophoblastic disease epidemiology and trends. 

Clin Obstet Gynecol 2003;46:541. 

Steigrad SJ. Epidemiology of gestational trophoblastic diseases. Best 

Pract Res Clin Obstet Gynaecol 2003;17(6):837-47. 

Thama BWL, Everardb JE, Tidyc JA, et al. Gestational trophoblastic 

disease in the Asian population of Northern England and North Wales. 

BJOG 2003;110(6):555-9. 

Diagnostics of early Spontaneous Abortion 

E. Fulcheri, Y. Musizzano 

Anatomic Pathology, DISC, University of Genoa 

211 

Classically, early spontaneous abortion (ESA) can be defined 

as occasional, repeated, or recurrent; we discussed the diagnostic 

problems of these three groups several years ago, in a 

leading article considering every aspect of embryo, fetal and 

neonatal pathology 1 . The role of the pathologist has acquired 

more and more importance in the diagnostics of miscarriages 

and in the interpretation of the related infertility. Actually, 

a different socio-anthropological attitude has changed the 

features of female population over the last decades and is still 

inducing significant mutations in the composition of rural 

and urban ethnic groups. The most striking aspects of this 

phenomenon can be resumed in: 1) an increasing number of 

pregnancies in advanced maternal age; 2) the wish of only 

one pregnancy, well planned and programmed with a definite 

timing; 3) the characteristics of immigrant populations, 

whose needs about maternity and fecundity are very different 

from those of the past and present aboriginal population. It is 

evident that, nowadays, a generic diagnosis formulated on the 

abortion specimen cannot be considered satisfactory. At the 

same time, it is clear that accurate and adherent diagnoses can 

be made only in optimal conditions, and when clinical data 

and cytogenetics are available. Anyway, given the abovementioned 

conditions and the fact that occasional abortion 

does not permit accurate collection of clinical informations, 

the problem should be considered from different viewpoints; 

in brief, four different types of specimen exist, representing 

the following situations. 1) First occasional ESA in a woman 

with negative clinical history; only the date of last menses is 

known, and a few more data can be referred. 2) First ESA in 

a patient treated for infertility or other disease; in this case, 

clinical data should be very exhaustive, with particular regard 

to the patient’s history. 3) Repeated or recurrent ESA; even in 

this case thorough clinical data should be provided. 4) Review 

of chorio-decidual specimens from previous ESAs in a recurrent 

aborter. 

These specimens can be available in the same laboratory or 

elsewhere; unfortunately, sometimes they are not available 

owing to the lack of previous histological examination. This 

item will be discussed later, in the forthcoming debate. 

While facing any of these situations, we need to define the 

level of diagnostic accuracy needed and the appropriate type 

of histological report. Undoubtedly, the perspective of an 

indisputable diagnosis is unrealistic and this occurrence is feasible 

in a very few situations. Hence, in some cases we shall 

formulate our diagnosis in terms of highest likelihood; due to 

the above-discussed problems, even this approach is possible 

only in selected cases. Thus, in the majority of cases we’ll 

make orientating diagnoses that, particularly in the setting of 

infertility and repeated ESAs, take on great importance. Nevertheless, 

some situations remain in which it is only possible 

to rule out a given condition; even this approach, can play 

an important role in the evaluation of repeated or recurrent 

ESAs. 

In order to follow this scale based on diagnostic complexity, 

the ability of the pathologist to explain chorionic and decidual 

findings is essential. First, some fundamental differences in 

the cause-and-effect mechanism underlying the generic definition 

of detachment of gestational sac should be defined: 

actually, in some cases the detachment following any cause 

of abortion can be itself the cause of the pregnancy loss (e.g. 

abruptio placentae), while in other instances it represents the

212 

common evolution of other noxae From this point of view, 

a diagnosis such as “abortion owing to detachment of gestational 

sac” appears inconclusive. 

Before outlining the diagnostic procedure, it seems necessary 

to focus on the sampling methods. As a rule, the whole specimen 

should be submitted to histology, requiring 4 to 6 biocassettes. 

Preliminarily, the fragments should be examined on a 

large Petri dish. The latter allows to recognize the presence 

of a gestational sac and its macroscopic features (complete 

and sealed, complete and open, presence of parts of embryo 

and adnexa, according to Fujikura classification) The embryo 

examination should be discussed as a distinct issue. Microautopsy 

of the embryo, as we defined it in a previous meeting 

in Pisa 2 , requires the use of a dissecting steromicroscope, and 

inclusion in epoxy resins is unavoidable; in our institution, 

this method is routinely used since 1994. 

Diagnostic flow chart. When approaching an abortion specimen 

(Fig. 1), adequacy should be first considered; the latter 

relates not only to the amount, but also to the representativeness 

of the received fragments. A specimen should be 

regarded as adequate when it includes parts of basal decidua 

(marked by the presence of Nitabuch’s stria), parietal decidua 

and chorionic villi. Other structures can be seldom identifed, 

such as the cord, amniotic sac, or yolk sac 3 . 

The identification of the basal decidua represents the first step 

in the evaluation of the specimen, since it allows to recognize 

superficial maternal vessels and to evaluate the modifications 

induced by extravillous (intermediate) trophoblast. Conversely, 

it is virtually impossible to investigate deep decidual 

vessels, which are usually not included in the specimen. Investigation 

of the parietal decidua, apart from pregnancy 

Fig.1. checklist for the microscopic examination of abortion 

specimens according to uni en iso 9001-2000 norm, no. 9122. 

ao10 (modified from: musizzano et al. 6 ). 


stromal modifications and Arias-Stella reaction, permits the 

evaluation of decidual vessels that were not modified by extravillous 

trophoblast and so very similar to deep vessels in 

the implantation site; the features of intima and vascular wall 

should be described. 

The most important feature of the chorionic plate is the 

branching of chorionic villi. Trophoblast layer is bilaminar 

all over the first trimester, featuring a cap of cytotrophoblasts 

only in terminal villi. These normal features should be accurately 

researched and documented in the histopathological 

report. All modifications, in terms of uneven branching, abnormal 

trophoblast proliferation and degeneration, as well as 

every modification of the villous outline (invagination, inclusion, 

angular or slender shape) are unmistakable findings that 

suggest karyotype abnormalities. 

Other important features of the villi are represented by the 

type and distribution of capillaries and by the presence of 

red blood cells and erythroblasts in the lumen, according to 

well established proportions. Similarly, every abnormality 

in vascular distribution, as well as the presence of incomplete 

vascularization in the villi, shall suggest karyotypic 

abnormality. 

Finally, stromal degeneration and fibrinoid deposition, though 

common findings in many conditions and hence not peculiar 

to a given etiology, are useful to correlate and sharpen the 

observations in the setting of the above discussed protocol. 

While approaching ESA pathology, it is useful the knowledge 

of the features, histopathological modifications and diagnostic 

findings peculiar to some large categories. 

The first condition is represented by abnormal karyotype, that 

is responsible for a huge amount of cases, at present estimated 

around 70%. 

Morphological and structural findings in the villi peculiar to 

karyotype abnormalities: abnormal branching; avascular villi; 

Irregular outline of the villi; trophoblast invagination; trophoblast 

inclusion; stromal hydrops (edema). 

Morphological and structural findings in the villi suggestive 

of karyotype abnormalities: irregular branching; uneven, 

sometimes lacking blood vessels; Uneven, discontinuous 

trophoblast double layer (cyto- and syncytiotrophoblast); 

vacuolated syncytiotrophoblast; hystiocyte-like stromal cells; 

fibrinoid degeneration of villous stroma. 

Secondly, acute or chronic infections should be considered, 

the term chronic indicating long standing, eventually remittent 

or steady and latent, conditions. Nowadays, the majority 

of infections and subsequent inflammatory states can be reliably 

identified. Unfortunately, the isolation of microorganisms 

(viruses, bacteria) is today more difficult despite the 

large amount of antibodies available, hence in many cases the 

villitis or chorioamniositis remains “aspecific”. 

Morphological and structural findings in the villi peculiar of 

acute infections: stromal edema of the villi; dilated villous 

vessels; granulocytic infiltrate (villitis/intervillitis) 

Morphological and structural findings in the villi suggestive 

of chronic infections: mild stromal fibrosis of the villi; collapsed 

vessels; mineralization of trophoblast basement membrane; 

syncytiotrophoblast hyperplasia (sprouts). 

Another diagnostic category is represented by maternal vasculopathies 

and related conditions. The latter include, foremost, 

maternal autoimmunity (even subclinical), hypertension, and 

decidual vasculopathy as defined by the AFIP Atlas of Placental 

Pathology 4 . Frequently, these pictures partially overlap 

thus further complicating diagnostics. In the last years, our efforts 

were aimed at the definition of an examination protocol 

mainly based on vascular decidual findings both in routine

lectures 

H&E slides and after some cost effective and easy-to-do histochemical 

and immunohistochemical stainings. 

Although in many cases only orientating diagnoses are feasible, 

some histopathological findings, eventually confirmed 

by histochemistry and immunohistochemistry, tend to recur 

and sometimes to cluster, so that at least three situations can 

be defined 5 : unconverted decidual vessels strongly suggest 

maternal luteal defect; diminished or disrupted smooth muscle 

cells, intimal thickening, and inflammation are more probably 

related to autoimmune maternal diseases; finally, fibrinoid 

necrosis (staining blue with Weigert-fibrin) and/or hypertrophy 

of vascular walls (confirmed by the increase of smooth 

A review on pathology report coding practices 

V. Della Mea 

Medical Informatics, Telemedicine & Health Lab; Dept. of Mathematics 

and Computer Science, University of Udine Italy 

Background. Text included in clinical documents, including 

anatomic pathology reports is often complemented by a 

concise version of the content, obtained by coding them using 

terms (and corresponding alphanumeric codes) coming from 

one or more terminologies or classifications. 

Coding may be aimed at different applications, where the 

most traditional are administration and finance aspects (e.g. 

healthcare intervention reimbursements) and epidemiology 

(e.g., disease statistics provided by national statistics 

institutes and WHO). The practical advantage provided by 

coding to the coder (e.g., the reporting pathologist), when 

applied into some computerized information system, is the 

possibility to retrospectively retrieve reports according to 

coded content. Coding also provides the opportunity of 

comparing and collecting anatomic pathology data independently 

from document language, and also automated 

decision support. 

In the anatomic pathology report, coding may involve various 

aspects, not always and not all present and coded, including: 

– site of sampling, for which a terminology describing human 

anatomy is needed; 

– macroscopic and microscopic description of the sample, for 

which a terminology is needed to describe morphological 

aspects; 

– the diagnosis, or which a terminology of diseases is needed 

that provides the adequate amount of detail to describe anatomic 

pathology diagnoses; 

– and finally, procedures applied to the sample (e.g., stainings), 

for which a terminology collecting all possible procedures 

is needed. 

SNOMED is historically a very large terminology that provides 

all above mentioned components, plus others aimed at 

describing other content of a general clinical record including 

other reports. 

In the world, SNOMED is likely the most used terminology 

for anatomic pathology report coding, but is not the only one 

available, nor is used in just one, i.e., the last available, version. 

In fact, in some countries national terminologies have 

been developed, like ADICAP in France and READ codes in 

SINOMeD-NAP 

Moderators: F. Crivelli (Gallarate), C. Francescutti (Udine) 

213 

muscle actin-positive cells) are the usual findings in classical 

decidual vasculopathy according to the AFIP. 

references 

1 Fulcheri En et al. Pathologica. 2006;98(1):1-36. 

2 Fulcheri E. Pathologica 1997;89(6):624. 

3 Fulcheri E, Mariuzzi GM. Patologia della gravidanza. In: Mariuzzi 

GM (ed). Anatomia Patologica e correlazioni anatomo-cliniche. Padova: 

Piccin Nuova Libraria 2007, pp. 1946-8. 

4 Kraus FT, et al. AFIP Atlas of nontumor pathology. N. 3. Placental 

pathology 2004. 

5 Musizzano Y, Fulcheri E. Virchows Arch 2010;456:543-60. 

6 Musizzano Y, Fulcheri E. Decidual vascular patterns in first-trimester 

abortions. Virchows Arch 2010;456:543-60. 

UK, while in other countries WHO’s diagnostic classification 

have been used (ICD9-CM, ICD10, ICD-O). 

SNOMED was initially developed by the College of American 

Pathologists, but since few years is maintained by the 

International Health Terminology Standards Development 

Organisation (IHTSDO), based in Denmark and currently involving 

15 countries 1 . While for long time it was used only in 

Pathology, application is broadening towards other areas 2 . 

The present paper reviews the current practices of pathology 

report countries in various countries. 

Methods. A questionnaire was developed to survey pathology 

report coding practices in the different countries, starting from 

those participating into the COST action IC0604 “Telepathology 

Network in Europe: EURO-TELEPATH” 3 4 . Survey was 

implemented using GoogleApps in order to provide an online 

fillable version, but was also collected by interviewing participants 

to the European Congress of Telepathology and Virtual 

Microscopy, held in Vilnius in 2010. 

Questions in the survey were means at collecting information 

on the presence and application of a national policy for pathology 

report coding, on who established the policy, on which 

codings are used for the various sections of the report, on 

IHSTDO involvement and on SNOMED translation plans. 

Results. Representatives of 12 countries answered the questionnaire, 

of which 10 from EU countries. All countries present 

at least some coding practice, but is applied by the totality 

of pathology institutes in 25% of countries. In two cases, this 

is made without a national policy, although when present (8 

countries) it is provided by professional associations in all 

but one case. This also means that usually coding decisions 

are left at the Pathology Institute level, so that heterogeneous 

behaviour can be found in a country for both the coding and 

the terminology used. 

SNOMED-CT is formally adopted in two countries, although 

this does not mean that is also practically used by a majority 

of their Pathology institutes. Older versions of SNOMED, 

often nationally or even locally adapted to purpose, are most 

often used, together with national terminologies. When coding 

is done, site of disease and diagnosis are always coded. 

Procedures are much less subject to coding, and description 

almost never, although sometimes morphological aspects are 

enclosed in the diagnostic coding. 

Conclusions. Pathology report coding seems a common practice 

in most countries, although the terminologies used for

214 

that are actually heterogeneous. This makes sharing of data 

more difficult, although trans-coding (i.e., converting one 

coding into another) is already used for other terminologies 

and classifications. 

In a previous assessment of SNOMED implementations, Giannangelo 

and Fenton 5 found out that a considerable amount 

of software vendors need a business case for why SNOMED 

CT should be deployed in their systems. In addition to that, 

some vendors indicated that if institutions were to require 

it, then they would proceed with including SNOMED CT in 

their products. Unfortunately, report coding policies are not 

always available or mandatory in countries involved in the 

present survey. 

Translation in national languages has been started and done by 

few countries 6 , but it can be a problem due to the large size 

of the whole SNOMED-CT terminology. Pathologists use just 

a part of the whole vocabulary, which could be more easily 

translated than the whole SNOMED-CT. 

Although respondents were among people involved in telepathology 

and digital pathology, awareness about policies and 

plans about coding, IHSTDO and SNOMED was not much 

diffused, perhaps because perceived as a secondary aspect of 

digitalization. 

Uniform implementation of a single coding systems seems 

still more easily rechaed where a common information system 

is adopted throughout the country, like in Netherlands. 

However, pathologists produce a considerable amount of 

information that would be better exploited if made available 

also in a coded, sharable form. 

Acknowledgements. The present review has been carried 

out inside the activities of the COST Action IC0604 “Telepathology 

Network in Europe: EURO-TELEPATH”, Working 

Group 2 “Informatics Standards in Pathology”. 

references 

1 International Health Terminology Standards Development Organisation 

(IHTSDO): http://www.ihtsdo.org/ 

2 Cornet R, de Keizer N. Forty years of SNOMED: a literature review. 

BMC Med Inform Decis Mak. 2008;8(Suppl 1):S2. 

3 COST Action IC0604 “Telepathology Network in Europe: EURO- 

TELEPATH”: http://www.conganat.org/eurotelepath/ 

4 Garcia Rojo M, Punys V, Slodkowska J, et al. Digital pathology in 

Europe: coordinating patient care and research efforts. Stud Health 

Technol Inform 2009;150:997-1001. 

5 Giannangelo K, Fenton SH. SNOMED CT survey: an assessment of 

implementation in EMR/EHR applications. Perspect Health Inf Manag 

2008;5:7. 

6 Klein GO, Chen R. Translation of SNOMED CT - strategies 

and description of a pilot project. Stud Health Technol Inform 

2009;146:673-7. 

Personal data protection in italian pathology 

services 

G. Negrini 

Ospedali Area Ovest AUSL Bologna, Italy 

Background. In Italy, since 2003 there is a regulatory body of 

personal data, so called Privacy Code (Dlgs 196/2003) * . 

According to this legal framework, special rules were established 

in order to protect all sensitive data, especially those 

relating to health. 

Moreover, within these ones, genetic data have a far greater 

protection. 

Pathologist’s activities usually come across a lot of privacy 

matters, for instance: 


– informed consent to personal data treatment, related to patient 

care; 

– use of personal data for clinical studies, epidemiology, research; 

– biological samples collections; 

– electronic documentation. 

Discussion. Firstly, we have to answer the question if the 

patient consent must be related only to clinical needs or even 

further occurrences. 

Nowadays most physicians deem data as essential for purposes 

other than each patient’s treatment. 

Nevertheless, it is not enough to give generic information to 

patients about foreseeable purposes of study or research, consequently 

we have to clear for which study or research data 

could be used. 

All that may seem too restrictive. 

When we ask for the patient consent we couldn’t know some 

needs that arise only later. 

Our legislation ** relieves us from the patient consent only if 

a research or study was provided by the National Research 

Program, 45 days after the notice to Privacy Authority. 

Otherwise, when seeking consent is too expensive, because of 

a great deal of patients or actual difficulties, we have to acquire 

a favorable opinion of a local ethic committee, followed 

by the approval of the Privacy Authority. 

About genetic data, some Authors speculated about what they 

are really and confuted genetic exceptionalism 1 , but our legal 

system requires a separate, written consent *** . 

About that one, a previous information should explain: detailed 

list of all specific purposes to be achieved, possible 

findings, the right to object to data processing, whether the 

data subject is allowed to limit the scope of their communication 

and the transfer of biological samples, retention period of 

genetic data and biological samples. 

When the consent to search is withdrawn, the related biological 

samples must be destroyed, if they are still identifiable. 

Many expedients should be arranged to prevent the risks of 

undue accesses, for instance: every room where genetic data 

are stored needs special controls. 

People, who enter it, after the closing time, must be identified 

and recorded. 

Storage, use and transport of biological samples must be 

carried out to ensure their quality, integrity, availability and 

traceability. 

Genetic data should be transmitted electronically by certified 

electronic mail after encrypting and digital signature. 

If genetic data or biological samples are acquired for clinical 

purposes, a different use is allowed only for a purpose related 

to the former, unless a new consent or the anonymization of 

data or samples. 

If seeking new consent is too expensive, they can get positive 

evaluation of local ethic committee, followed by approval of 

Privacy Authority. 

However, we consider that, except for rare hereditary cancers, 

genetic characteristics of tumor tissues can not be qualified as 

genetic data, because they are limited to some body parts and 

don’t affect germ cells. 

With regard to biological samples, we should ask ourselves: 

‘Who owns? Who can decide what to do with them? Is there 

a real property right of biological materials 2-5 ? 

The answers determine the resolution of several issues: their 

use for additional purposes, responsibility of their retention, 

storage time, right of access (for example: could patients get 

their samples back?).

lectures 

The Convention on Human Rights and Biomedicine **** 

– Oviedo, 1997 – article 22 provides: 

“Disposal of a removed part of the human body. 

When in the course of an intervention any part of a human 

body is removed, it may be stored and used for a purpose 

other than that for which it was removed, only if this is done 

in conformity with appropriate information and consent procedures.” 

Well, what happens if the patient dies? 

May the relatives – the heirs – succeed to the rights of the 

deceased? 

All these issues have been discussed for a long time in many 

countries, but we are still waiting for a definite answer. 

Recently, some Authors have suggested an interesting innovation 

with regard the patients’ consent and their rights 6 . 

They argued the need of rethinking the relationship between 

patients - or donors not patients- and researchers and proposed 

to replace the current informed consent with trusted consent, 

at least with reference to research biobanks. 

New themes and questions are now related to the development 

of electronic health records. 

In our country, regional governments are building health information 

systems, designed to create files with the medical 

history of each person, potentially from birth to death 7 8 . 

Our Privacy Authority has recently launched guidelines on 

reports on line as well as on electronic record and electronic 

file ***** . 

Patients have the right to give or withhold their consent to the 

implementation of such a file. 

It should be possible excluding some items of medical information, 

so that patients can decide, after being duly informed, 

whether or not they want to disclose certain events (blanking). 

However, blanking can become a threat to clinical decisions 

when these are based on partial information. 

Despite of the easy electronic consultation, we must ensure 

only professionals who provide care to the patients are entitled 

to consult their records. 

Another issue is the patients right of access to their data: may 

the documents containing severe diagnosis be directly known 

by the patient, without previous discussion with a doctor, as 

it is now possible (e.g.: on line reports, access to electronic 

health record)? 

To avoid such an impact, some health organizations block 

critical reports until an interview with a doctor. 

What’s above is a limited summary: how can we extricate 

ourselves from all these entanglements? 

Pending regulatory precise answers, we should make an effort 

to go beyond the rule of thumb, to seek balanced solutions, 

in compliance with existing legal rules and prevalent ethical 

principles. 

references 

1 Rothstein MA. Genetic Exceptionalism and Legislative Pragmatism. 

Hastings Center Report 2005;35:27-33. 

2 Negrini G. Materiali biologici donati: incertezze di inquadramento 

giuridico. Rischio Sanità 2009;34:16-21. 

3 Negrini G. Raccolte di materiali biologici: interrogativi. De Qualitate 

2008;2:8-25. 

4 Furness PN, Nicholson ML. Obtaining explicit consent for the use of 

archival tissue samples: practical issues. J Med Ethics 2004;30:561- 

4. 

5 Negrini G, La Pietra L. Campioni biologici conservati nelle strutture 

sanitarie: interrogativi e problemi aperti. Professione. Cultura e 

pratica del medico d’oggi 2005;1:34-41. 

6 Boniolo G, Di Fiore P, Pece S. Trusted Consent and Research Biobanks. 

Towards a new alliance between researchers and donors. 

215 

Bioethics 2010 JUN doi:10.1111/j.1467-8519.2010.01823.x 

7 Negrini G, la Pietra L. Opportunità e criticità del fascicolo sanitario 

elettronico. Professione & Clinical Governance 2009;7:30-7. 

8 Moruzzi M. Health e Fascicolo Sanitario Elettronico. Il Sole 24 Ore. 

Milano 2009 

Notes 

* www.garanteprivacy.it 

** Art. 110 Dlgs 196/2003 

*** General Authorisation for the processing of genetic data, 27/2/2007 

http://www.garanteprivacy.it/garante/doc.jsp?ID=1389918 

**** http://www.coe.int/t/dg3/healthbioethic/Activities/01_ 

Oviedo%20Convention/ 

***** http://www.garanteprivacy.it/garante/doc.jsp?ID=1681147 

http://www.garanteprivacy.it/garante/doc.jsp?ID=1634116 

NAP Italia – The new nomenclature for the 

anatomic pathology 

P. Crucitti, A. Bondi 

U.O. Anatomia Patologica, Maggiore Hospital - AUSL Bologna, 

Italy 

Background. Classification is a recognised method to organize 

in a systematic way all scientific informations. In medical 

field, the broad diagnosis terminology has produced the internationally 

utilized SNOMED code, with the subset “Microglossary 

for Pathology”, translated in Italian. SNOMED has 

an international copyright and is registered by the College of 

American Pathologists (CAP): use of the code is under payment 

of the rights for workstation. 

Adverse events during SNOMED distribution in Italy caused 

a consequent autonomous development of “self-made” codes, 

often not in line with international version and that not allow 

exchanges of data between different Anatomic Pathology. 

All this situation became an obstacle for data extraction from 

different Institutions and, as a consequence, for Tumour Registries 

end official epidemiological archives, raising the need 

of one national language. From these assumptions, role of 

SIAPEC has been to join different codes from many Anatomic 

Pathology in Italy and re-direct Italian Pathology to the international 

scenario, favouring and coordinating cultural and 

scientific collaboration between varies Associations involved 

in medical informatics, diagnostic coding, epidemiology and 

tumour registry. Actors of this project are Scientific Corporations 

of the field (SIAPEC-IAP and AIRTum), the Italian 

Contributor Centre of OMS for Sanitary Codes (CC-OMS) 

and few Regional Sanitary Agencies (Friuli-Venezia Giulia, 

Liguria and the cooperation of Emilia-Romagna and Lombardia). 

Aim of the Nomenclature for Anatomic Pathology 

(NAP) is to become the national reference, for improvements 

and updating, shared from the Pathologist community. 

Methods. NAP development is on-going, but we can briefly 

schematize the general proceeding: 

1. Identification of a work group, constituted by Pathologies 

from different Italian Anatomic Pathology, an operator editing 

different tables (see below), sometimes in collaboration 

with other centre; a group coordinator. 

2. Census of main code versions utilised between Italian 

pathologists. Requirement to software houses (SH) of the 

area, to get different nomenclature distributed. Eventually 

other sources can be Institutions, that have enriched autonomously 

the Nomenclature. 

3. Definition of coding rules and preparation of NAP, mainly 

for section related to non-tumours definitions: extra-tumour 

morphologies, topography, procedures, other sections. 

4. ICD-O 3 acquisition to create the core of NAP.

216 

5. Realization of other axes, starting from Microglossary from 

Pathology of 1995, lately integrated with varies versions 

collected from produced from the work group promoted 

from the executive in 2005. 

6. Definition of a first NAP edition and official SIAPEC approval. 

7. Creation of transcoding tables for new terms (point 2) and 

distribution from SH, in application to each administration 

management. 

8. Realization of a transmission protocol among archives, 

eventually according to HL7. 

Through the construction of intermediate “tables of comparison” 

we found “alignment rules”, containing all indications 

for records to be modified, deleted or added to uniform any 

table according to these rules. 

Results. At present NAP sections already completed are: 

a table containing morphology of tumours (M-8 and M-9), 

based on ICD-O 3 perfectly in line with SNOMED CT 2002, 

a table containing topography codes and - to be released - a 

table containing all morphology (extra-tumour). All tables are 

organised according to a record track, including alphanumeric 

code, Italian and English description, an index code to trace 

source of each record, a link to SNOMED / ICD-O topography 

and finally a hierarchy code to structure each table (see 

below). The NAP code for tumours is completely compatible 

with SNOMED International and it’s composed by: Axis definition 

(1 capital letter) / dash / tumour definition (4 Digits) / 

Stem cells and genetic skeletal diseases – role 

in pathogenesis and therapy 

M. Riminucci 

Department of Experimental Medicine, University La Sapienza, 

Rome, Italy 

Fibrous dysplasia (FD) (polyostotic fibrous dysplasia, Mc- 

Cune/Albright syndrome, OMIM#174800), is a non inherited 

skeletal dysplasia caused by mutations of GNAS, the gene 

encoding the alpha subunit of the stimulatory G protein (Gs). 

Mono and polyostotic forms of the disease are recognized, 

the latter frequently associated with extraskeletal disorders 

in complex clinical settings such as the McCune-Albright 

syndrome (MAS, polyostotic FD, café-au-lait spots and endocrine 

lesions) and the Mazabraud’s syndrome (MS, FD and 

muscular myxomas). 

FD lesions develop during the post-natal growth and replace 

normal skeletal tissues (bone, adipose and hematopoietic marrow) 

with woven bone and fibrotic marrow. Affected skeletal 

segments often become mechanically insufficient and in most 

patients, severe invalidity ensues from recurrent fractures and 

deformities. 

Long thought of as an undefined fibro-osseous disease, FD has 

become over the last few years a unique model of human disease 

affecting both embryonic and post-natal stem cells. The 

somatic nature of the mutation and the distribution of mutated 

cells in derivatives of all germ layers, point to a pluripotent 

embryonic cell as the initial (although silent) target in which 

the molecular lesion occurs. The profound derangement of the 


Germ cells in tumoural pathology 

Moderator: C.A. Beltrami (Udine) 

behaviour (1 digit) / Differentiation (1 digit) / 1 Extra digit / 

Synonyms (1 letter). 

Id link allows to trace each code/definition for correction, 

modification. 

The reference table is also modified and enriched with new 

terms deriving from different Anatomic Pathology: new 

tables and codes will be available through the Italian Portal of 

Classifications, tool of the Italian Collaborating Centre, for a 

continuous upgrade of NAP. 

NAP code is also related to the creation of a digital Atlas 

for rare cases promoted from Emilia-Romagna region and 

presented in current SIAPEC congress (see presentation from 

S. LEGA et al) 

Record track: 

Field name Length Data type Function 

cod 10 text naP code - icd-o 

/snomed 

txt 120 text italian description 

txt_en 120 text english description 

id 8 number unique identification 

of the term 

icd 15 text reference to icd-o 

topography 

ref 80 text link to snomed 

topography 

super 10 text hierarchy code 

bone/bone marrow microenvironment observed at affected 

sites and reproduced upon ectopic transplantation of FD osteprogenitors, 

indicate post-natal skeletal stem cells (originating 

from the mutated embryonic clone) as the late effectors in 

which the mutational event displays its adverse effects. 

Reinterpretation of FD as a stem cell disease has opened new 

avenues to investigation of its pathogenetic mechanisms, generation 

of suitable experimental models and development of 

specific therapeutic approaches. 

Many complex changes occurring at affected skeletal sites, 

such as the inappropriate bone resorption and bone matrix 

hypomineralization, which cannot be explained based on the 

activity of mutated osteoblasts solely, have been reinterpreted 

in light of the abnormal expansion and function of mutated 

osteoprogenitors. 

Lentivector-mediated trasduction of normal ES and post-natal 

skeletal stem cells with the disease gene has provided appropriate 

experimental models to investigate the molecular 

responses induced by the GNAS mutation at different developmental 

stages and to seek new potential pharmacological 

targets. 

Finally, the recognition of FD as a stem cell disease has emphasized 

the need for innovative, stem cells based therapeutical 

approaches as the only possibility to cure the disease radically. 

However, the develoment of new strategies based on 

either the replacement or the genetic manipulation of mutated 

FD skeletal stem cells requires the availability of suitable in 

vivo models. To this aim, we have recently generated the first 

transgenic murine models of the diseases.

lectures 

Case n. 1 

Aggressive psammomatoid cemento-ossifying 

fibroma of the sinonasal region 

L. Roncati, A. Maiorana 

Department of Laboratory Services, Pathologic Anatomy and Forensic 

Medicine, Section of Pathologic Anatomy, University of Modena 

and Reggio Emilia, Modena, Italy 

Background. An 18 years-old woman with a clinical history 

of chronic sinusitis and headache was evaluated for 

nasal airway obstruction associated with recent, painful and 

widespread left hemifacial swelling. No history of trauma 

was elicited. Computed tomography of the maxillo-facial area 

showed an expansive-erosive neoformation, predominantly 

isodense to surrounding soft tissues, that occupied the left 

maxillary sinus, eroding the left hemipalate and the left jawbone. 

The left orbital floor was thinned and slightly raised. 

The right maxillary sinus, together with both sphenoidal and 

frontal sinuses and ethmoid cells were regularly pneumatized. 

A biopsy of the left maxillary sinus was performed. Following 

the diagnosis, the patient underwent a first surgical intervention 

under the intraoperative direction of the pathologist. 

A left hemimaxillectomy with reconstruction of the orbital 

floor, postero-lateral wall of the maxillary sinus (maxillary 

alveolar process) and zygomatic process using revascularized 

fibula was performed. Nine months later a recurrence in the 

left orbital floor required surgical revision of the orbital area. 

After a recurrence-free follow-up period of two years, the 

patient underwent another surgical intervention for removal 

of fixation devices in the orbital floor and remodelling of the 

homologous bone graft with alloplastic tissue. 

Methods. All specimens were routinely processed for histopathological 

examination (fixation in 4% formaldehyde, 

paraffin embedding, staining of sections with hematoxylineosin). 

Immunohistochemistry for CD34 (Ventana), EMA 

(Ventana) and MIB-1 (Dako) was performed. 

Results. The bioptic sample was a small fragment measuring 

cm 1 × 0.5 × 0.5. The surgical material consisted of numerous 

white, hard tissue fragments that, when put together, measured 

approximately cm 3.8 × 3.5 × 3.2. All fragments showed a 

fibro-osseous lesion composed of a fibrous stromal component 

of monomorphic spindle or polygonal cells that embedded bony 

trabeculae of varying shapes, mostly curvilinear, and numerous 

round-to-oval calcific elements similar to psammoma bodies 

(so-called “psammomatoid bodies” or “ossicles”). Some bony 

trabeculae were rimmed by osteoblasts accompanied by isolated 

osteoclasts. A cystic component with hypocellular fibrous 

septa, reminiscent of aneurysmal bone cyst, was also present. 

Nuclear atypias and mitotic activity were not detected. Immunohistochemical 

reactions for CD34 and EMA were negative. 

The histological findings led to the diagnosis of “aggressive 

psammomatoid cemento-ossifying fibroma” of the sinonasal 

region, also known as “extragnathic cemento-ossifying fibroma”. 

Saturday, September 25 th , 2010 

Slide seminar: Histopathology 

Moderators: V. Eusebi (Bologna), G. Pelosi (Milano) 

217 

Discussion. The definition of “aggressive (or juvenile) psammomatoid 

cemento-ossifying fibroma” identifies a complex 

extragnathic fibro-osseous mesenchymal proliferation that 

may arise in every site of the sinonasal tract (nasal cavity, 

paranasal sinuses, turbinates, nasolacrimal duct). The tumor 

is similar to cemento-ossifying fibromas that occur in the 

gnathic region and was postulated to arise from mesenchymal 

elements of the periodontal ligament 1 . In the common definition, 

the use of descriptive adjectives such as “aggressive”, 

“active” or “juvenile” refers to the tumor capability to exhibit, 

especially in young patients (first and second decades) of 

both sexes, a locally aggressive behaviour, mainly characterized 

by invasion and destruction of surrounding anatomic 

structures (cranial cavity, orbit, palate, nasopharynx), with 

tendency to recur after surgical resection, in particular in cases 

of simultaneous involvement of multiple sites. Some authors 

have suggested the adjective “juvenile” to be dropped, since 

the lesion is not limited to the young age and can even affect 

people in the fifth or sixth decades of life. Although isolated 

cases were able to cause death of the patient, due to local involvement 

of vital intracranial areas, the aggressive behaviour 

does not imply an evolution into metastatic disease. The most 

common symptoms are sinusitis, headache, nasal obstruction, 

facial swelling, visual disturbances and progressive blindness, 

exophthalmos and proptosis. Epileptic seizures, smell disturbances 

and facial deformities are rarely observed. A case of 

juvenile aggressive ossifying fibroma presenting as mucocele 

of the ethmoid sinus has also been reported 2 . Clinicopathological 

integration, with detailed analysis of the radiographic 

projections, is essential to reach the correct diagnosis. At radiological 

imaging, the lesion is round-shaped and initially appears 

hyperdense suggesting the presence of a calcified matrix. 

In advanced stages, it can either show a multiloculated internal 

appearance with variable density, usually surrounded by a 

sclerotic bone rim, or can exhibit lytic growth in the adjacent 

bones or soft tissues. Aggressive growth is evidenced by invasion 

of anatomic compartments and bulging or displacement 

of surrounding bone structures. Grossly, the tumor can assume 

a compact tight aspect or a multicystic appearance with accumulations 

of coagulated blood material, resulting in a color 

change from greysh to brownish. Histologically, it shows a 

cellular stroma stuffed by numerous, spherical, concentric 

mineralized elements (psammomatoid ossicles) with variable 

foci of stromal myxoid degeneration and occasional multinucleated 

giant cells, in particular around vascular spaces. The 

differential diagnosis 3 4 of sinonasal psammomatoid cementoossifying 

fibroma includes a group of pseudoneoplastic proliferations 

(fibrous dysplasia, aneurysmal bone cyst, giant cell 

reparative granuloma) and benign or malignant neoplastic conditions 

such as osteoma, ossifying fibroma, giant cell tumor, 

myxoma/fibromyxoma, chondromyxoid fibroma, psammomatous 

meningioma 5 , osteoblastoma and osteosarcoma. The support 

of radiological findings associated with the detection of a 

large number of pathognomonic psammomatoid ossicles in the 

histological section does usually allow the correct diagnosis to

218 

be made. Surgical treatment may be conservative (endoscopic 

tumor resection) or highly demolitive (craniofacial resection, 

enucleation), with cosmetic deformities and increased risks for 

secondary blindness, meningitis and brain abscesses. Adjuvant 

therapeutic strategies (radiation therapy) have proved ineffective 

and can, at times, promote tumor dedifferentiation. The sinonasal 

psammomatoid cemento-ossifying fibroma is a prime 

example of how a radiosurgical multidisciplinary approach, 

directed by the intra-operative diagnosis of the pathologist, is 

imperative in order to define the extent of the disease and allow 

its total excision. 

references 

1 Wenig BM, Pilch BZ. Tumors of the upper respiratory tract. In: 

Fletcher CDM (ed.) Diagnostic histopathology of tumors, II ed, vol. 1 

Churchill-Livingstone 2007. 

2 Vaidya AM, Chow JM, et al. Juvenile aggressive ossifying fibroma 

presenting as an ethmoid sinus mucocele. Otolaryngol Head Neck 

Surg 1998;119:665-8. 

3 Wenig BM, Vinh TN, et al. Aggressive psammomatoid ossifying fibroma 

of the sinonasal region. A clinicopathologic study of a distinct 

group of fibro-osseous lesions. Cancer 1995;76:1155-1165. 

4 Slootweg PJ, Panders AK, et al. Psammomatoid ossifying fibroma of 

the paranasal sinuses. An extragnathic variant of cemento-ossifying 

fibroma. J Cran Max Fac Surg 1993;21:294-7. 

5 Granados R, Carrillo R, et al. Psammomatoid ossifying fibromas: 

immunohistochemical analysis and differential diagnosis with psammomatous 

meningiomas of craniofacial bones. Oral Surg Oral Med 

Oral Pathol Oral Radiol Endod 2006;101:614-9. 

Case n. 2 

Myxoid liposarcoma of the foot 

L. Roncati, A. Maiorana 

Integrated Department of Laboratory Services, Pathologic Anatomy 

and Forensic Medicine, Section of Pathologic Anatomy, University of 

Modena and Reggio Emilia, Modena, Italy 

Background. A 52 years-old woman underwent surgical 

excision of a nodular, mobile, fatty neoformation of the right 

big toe, which had been present for 2 years and had gradually 

grown in size. The lesion caused pain on walking. Grossly, 

the nodule was soft and well defined, measuring 3.8 × 2.7 × 

2.5 cm. Its cut surface appeared uniformly greyish and translucent. 

Methods. Immunohistochemistry was performed on paraffin-embedded 

sections using a broad panel of antibodies, such 

as α-smooth muscle actin (Ventana), desmin (Ventana), pankeratins 

(MoAb MNF 116 – Dako), S100 protein (Ventana), 

CD34 (Ventana). Interphase F.I.S.H. was performed on formalin-fixed 

paraffin-embedded tissue using the CHOP Dual 

Color Break Apart Rearrangement Probe (Abbott). 

Results. Histologically, the neoplasia was composed of 

spindle cells, immersed in abundant myxoid stroma, with focal 

marked mucinous accumulation. A vascular crow ‘s feet 

pattern of thin-walled branching vessels was present, together 

with a focal component of round cells. The tumor reached 

the margin of resection. Immunohistochemical reactions for 

α-smooth muscle actin, desmin, pankeratins, S100 protein, 

CD34 were negative. Interphase F.I.S.H. revealed the presence 

of the t(12;16) (q13;p11) translocation with FUS-CHOP 

fusion gene. The diagnosis of “myxoid liposarcoma with focal 

round cell component” (5-10% of tumor cell population) was 

rendered. Since tumor removal had been incomplete, re-excision 

was necessary followed by radiotherapy (10 Gy focused 

on the surgical site). The patient is alive and well after a 3 year 

follow-up period. 


Discussion. Approximately 75% of myxoid liposarcomas 

arise in the deep soft tissue of the buttocks and lower limbs, in 

particular thigh, groin and popliteal region, giving rise to neurovascular 

and muscular compressive symptoms. Rare cases 

localize in the feet. Isolated reports were described in unusual 

locations 1 , such as breast, ovary, scrotum, spermatic cord, 

vulva and thyroid. The tumour may be seldom accompanied 

by a paraneoplastic neurological syndrome (subacute complete 

ophtalmoplegia), asssociated with anti-Hu antibody 2 . Grossly, 

pure myxoid liposarcomas are multinodular gelatinous masses 

with a variable yellow tint, whereas predominantly round cell 

liposarcomas show a white fleshy appearance. The small size 

of the tumor (< 5 cm) may make the clinical diagnosis of 

malignancy difficult, since the findings of large size (> 5 cm) 

and rapid enlargement are the major clinical indicators for 

malignancy. Presurgical superficial (U/S-Scan) and deep 

(MRI-Scan) radiological investigation are useful to determine 

the size, shape, outline and, in particular, presence of typical 

cystic zones. Histologically, the differentiated myxoid component 

shows a low cellularity (paucicellular myxoid liposarcoma), 

composed of spindle/round cells scattered in a myxoid 

matrix of hyaluronic acid (Alcian blue-positive), sometimes 

with a microcystic pattern or lace-like configuration (pooling 

phenomenon). The occurrence of a plexiform capillary vascular 

network and signet ring lipoblasts in different stages of 

maturation are important diagnostic clues. Mitotic figures are 

tipically rare or absent. As myxoid liposarcoma loses its differentiation, 

it assumes a round cell appearance, characterized 

by the progressive accumulation of primitive round cells with 

high nuclear/cytoplasmic ratio and prominent nucleoli, growing 

in sheets and accompanied by vascular texture attenuation 

(cellular myxoid liposarcoma). The amount of round cells correlates 

with the development of distant metastases and should 

be always estimated in a well-sampled specimen (one section 

per centimetre tumor diameter). In more than 95% of myxoid/ 

round cell liposarcomas, a classical t (12;16) (q13;p11) or t 

(12;22) (q13;q12) translocation can be found, giving rise, respectively, 

to chimeric FUS-CHOP or EWSR1-CHOP genes. 

The identification of such translocations in lipomatous tumors 

is a powerful tool that aides in the correct identification of 

myxoid/round cell liposarcomas. 

A comparative study 3 of 16 cases of tumors previously diagnosed 

as primary myxoid/round cell liposarcoma of the retroperitoneum 

and 18 cases of myxoid/round cell liposarcoma of 

the extremities disclosed that FUS-CHOP or EWSR1-CHOP 

fusion genes were present only in tumors of the limbs, being 

absent in retroperitoneal tumors. The findings suggested 

that apparent primary myxoid/round cell liposarcomas of the 

retroperitoneum are actually well-differentiated (or dedifferentiated) 

liposarcomas with morphological features mimicking 

myxoid/round cell liposarcomas. As a consequence, the 

detection of FUS-CHOP or EWSR1-CHOP fusion genes in an 

apparent myxoid/round cell liposarcoma of the retroperitoneal 

area should lead to the suggestion of metastasis and prompt 

search for a primary localization outside the retroperitoneum. 

Similarly, a further study carried-out in 15 cases of presumed 

“multifocal” myxoid/round cell liposarcoma 4 was able to 

evidence (using LOH and RT-PCR for determination of the 

FUS-CHOP and EWSR1-CHOP breakpoints) the existence 

of a clonal relationship in the different tumors arisen in the 

same patient, supporting the “metastatic” nature of the apparent 

“multifocal” myxoid/round cell liposarcoma, with 

obvious consequences on therapeutic strategies. Cytogenetics 

may also be helpful in the routine differential diagnosis of 

myxoid liposarcoma. Potential mimics 5 6 include a wide range

lectures 

of benign or malignant subcutaneous (myxolipoma, myxoid 

nodular fasciitis, superficial angiomyxoma, aggressive angiomyxoma, 

myxoid dermatofibrosarcoma protuberans, myxoid 

neurofibroma, dermal nerve sheath myxoma) and deep-seated 

myxoid soft tissue tumors (low grade myxoid malignant fibrous 

histiocytoma, myxofibrosarcoma, extraskeletal myxoid 

chondrosarcoma, myxoid synovial sarcoma, myxoid leiomyosarcoma, 

myxoid malignant peripheral nerve sheath tumour). 

Intratumoral hemorrhage is a common finding in myxoid 

liposarcoma and, if present, can simulate a vascular neoplasia. 

The main histological parameters to follow in the differential 

diagnosis of myxoid soft tissue tumors are the architectural 

pattern, vascular pattern, cellularity and cytological aspects. 

Immunohistochemistry and histochemical reactions for mucosubstances 

may provide additional useful information. When 

pure myxoid liposarcoma loses its differentiation and assumes 

a round cell appearance, the differential diagnosis moves 

towards other round cell neoplasias, such as rhabdomyosarcoma, 

poorly differentiated synovial sarcoma, Ewing’s sarcoma/ 

primitive neuroectodermal tumor, lymphoma, melanoma and 

carcinoma, making the correct identification of myxoid soft 

tissue tumors a continuous diagnostic challenge. 

Pure myxoid liposarcoma exhibits high risk of local recurrence 

and a 20% rate of distant metastases. On the other side, 

round cell liposarcoma gives rise to metastases in approximately 

70% of cases. Both tumors show an unusual pattern of 

spread, since metastases arise mainly in extrapulmonary sites 

(2/3 of cases), such as soft tissues (mediastinum, retroperitoneum, 

thorax, distant extremity), bone (spinal cord, ribs), 

liver and serous membranes, lung metastases being observed 

in approximately 30% of cases. The average survival rate is 

80% at 5 years and 50% at 10 years, being mainly related to 

the quality of local excision. The time interval of appearance 

of the first metastasis is on average 68 months 7 . According to 

standard surgical procedures, wide excision with safety histopathological 

margins of at least 1 cm, should be performed 

each time the presence of a near-by neurovascular axis can 

allow it. Surgery is usually followed by radiotherapy and, 

sometimes, in biologically aggressive tumors, by adjuvant 

chemotherapy (Doxorubicine, Ifosfamide, Dacarbazine), following 

the indications of histopathological examination (size 

> 7.5 cm, number of round cells > 25%, p53 overexpression 

at immunostaining, R1 or R2 initial resection margins) 8 . The 

tumor is higly chemo- and radiosensitive, when compared 

with other soft tissue sarcomas, and its sensitivity to radiation 

treatment appears mainly to be related to the occurrence of 

the typical vascular crow’s feet pattern, since ionizing radiations 

can induce vascular damage with consequent hypoxic 

death of tumor cells. The chemo-radiosensitivity of myxoid 

liposarcoma and the possibility of using new drugs, such as 

trabectedin (ET-743; Yondelis), the first marine-derived anticancer 

product, able to induce the detachment of FUS-CHOP 

protein from the promoters of target genes and reduce neoplastic 

growth 9 , allow a more conservative surgical approach 

(limb-sparing surgery) in cases of large tumors affecting the 

limbs. 

references 

1 Weiss SW, Goldblum JR. Liposarcoma. In: Enziger & Weiss’s soft 

tissue tumors, V ed. Mosby Elsevier 2008;16:477-516. 

2 Chan JW. Subacute complete ophthalmoplegia: an anti-Hu paraneoplastic 

manifestation of myxoid liposarcoma. Clin Experiment Ophthalmol 

2007;35(5):491-2. 

3 De Vreeze RS, de Jong D, Tielen IH, et al. Primary retroperitoneal 

myxoid/round cell liposarcoma is a nonexisting disease: an 

immunohistochemical and molecular biological analysis. Mod Pathol 

2009;22(2):223-31. 

219 

4 De Vreeze R, de Jong D, Nederlof P, et al. Multifocal myxoid liposarcoma--metastasis 

or second primary tumor?: a molecular biological 

analysis. J Mol Diagn 2010;12(2):238-43. 

5 Ninfo VV, Montesco MC. Myxoid tumors of soft tissues: a challenging 

pathological diagnosis. Adv Clin Path 1998;2(2):101-15. 

6 Graadt van Roggen JF, Hogendoorn PC, et al. Myxoid tumours of soft 

tissue. Histopathology 1999;35(4):291-312. 

7 Kempson RL, Fletcher CDM, Evans HL, et al. Lipomatous tumors. In: 

Tumors of the Soft Tissues, III ed. AFIP 2001;4:187-238. 

8 Loubignac F, Bourtoul C, Chapel F. Myxoid liposarcoma: a rare 

soft-tissue tumor with a misleading benign appearance. World J Surg 

Oncol 2009;22;7:42. 

9 Germano G, Frapolli R, Simone M, et al. Antitumor and anti-inflammatory 

effects of trabectedin on human myxoid liposarcoma cells. 

Cancer Res 2010;70(6):2235-44. 

Case n. 3 

Peripheral primitive neuroectodermal tumor 

(pPNeT) of the small bowel 

M. Milione, A. Testi, F. Perrone, F. Melotti, S. Pilotti, 

G. Pelosi 

Dipartimento di Patologia Diagnostica e Laboratorio, Fondazione 

IRCCS Istituto Nazionale dei Tumori, Milano 

Clinical History. A 43-year-old man was admitted, on March 

2010, to Tivoli City Hospital with acute abdomen and intermittent 

abdominal pain of 5 months’ duration. His family history 

was non-contributory. Routine laboratory data on admission 

were within normal limits. Tumor markers, such as carcinoembryonic 

antigen (CEA) and carbohydrate antigen 19-9 (CA 

19-9) showed normal values; Computed tomographic (CT) 

scan of the abdomen showed ascitic fluid and an 80-mm diameter 

mass in ileo-cecal region. Laparotomy was conducted. 

The tumor consisted of polycystic components, and part of its 

surface adhered tightly to the initial portion of the jejunum 

whereas ascending colon, and greater omentum were normal. 

The tumor exhibited movability and could be easily dislocated 

from the abdominal cavity. An en-bloc resection of the tumor, 

including the ascending colon and proximal jejunum (each 

20 cm in length), and greater omentum, was performed. Endto-end 

jejunojejunostomy and colono-colonostomy were also 

carried out. Macroscopically, the resected tumor was 10 cm 

in major size, and 290 g in weight; it was elastic, and soft 

in consistency. The cut surface of the tumor showed mostly 

polycystic and, partially, solid features. There were bleeding 

and necrotizing parts inside the tumor. The tumour ulcerated 

the mucosa. Mucosal surfaces away from the ulcerated area 

were normal. 

After surgery no lesion were detected with Octreoscan study. 

Plasmatic chromogaranin levels were normal. 

Discussion. Microscopically, the specimens showed a sheetlike 

proliferation of spindle-to-polygonal cells with large 

vesicular nuclei with thin vascular stroma. Focally the tumor 

formed ribbon-like or rosette structures. Moderate mitosis 

was noted (10/50 high-power field) No invasive growth or 

metastasis to lymph nodes was noted. Immunohistochemestry 

showed the tumour cells to stain focal positively with: 

cytokeratin (CK) AE1/AE3, CAM 5.2, synaptophisin (SYN), 

chromogranin A (CgA) and CD 117, strongly diffuse positively 

with vimentin, CD 99, FlI-1and negatively for epithelial 

membrane antigen (EMA), S100, leucocyte common antigen 

(LCA), CD 34, smooth muscle actine (SMA), carcinoembryonic 

antigen (CEA), desmin an d WT-1. Proliferative index 

of neoplasia valued with MIB-1/Ki-67 immunostaining was 

about 30%. P53 was hyperexpressed. Based on histology and

220 

immunohistochemistry findings, the tumor was diagnosed as 

a extraskeletal ES/PNET, suspected of the small bowel, rather 

than an epitheliod GIST, or neuroendocrine poorly differentiated 

carcinoma (PDEC). Primitive neuroectodermal tumor 

(PNET) arises in soft tissue, and is thought to be of neural 

crest origin. This tumor, which usually develops in the chest 

wall and the extremities of children and adolescents 1-9 , is very 

rare, and is highly aggressive and malignant, and is characteristic 

of small round-cell tumours (SRCT). Pathology diagnosis 

in the present case was difficult because ES/PNET is 

extremely rare among tumors of the small intestine. Diagnosis 

required the exclusion of similar tumors showing undifferentiated 

small round cell morphology, including neuroblastoma, 

malignant lymphoma 9 , rhabdomyosarcoma, gastrointestinal 

stromal tumor (GIST), and desmoplastic small round cell 

tumor (DSRCT). Lymphoma could be excluded because the 

tumor cells did not express leukocyte common antigen (LCA). 

DSCRT could be excluded because the tumor lacked desmoplastic 

stromal reaction, WT-1 and EMA immunoreactivity, 

but expressed CD99 and CD117/c-kit. Neuroblastoma, rhabdomyosarcoma, 

could be excluded based on microscopy and 

immunohistochemical data CD99 is expressed in ES/PNET 

and other malignancies such as lymphoblastic lymphoma, 

rhabdomyosarcoma, DSRCT, synovial sarcoma, and solitary 

fibrous tumors. Small Bowel PDEC was favoured by clinical 

presentation (age, site, occlusive syndrome with peritonitis 

and ascitis) and was sustained by IHC positivity for cytokeratins, 

chromogranin A and synaptophysin, but morphology (no 

intratumoral necrosis, no perivascular glomeruloid pattern 

and abundant pseudorosette formations) and MIB 1/Ki-67 

value were against this diagnosis. CD117 was originally a 

marker for c-kit, a transmembrane tyrosine kinase normally 

expressed by Cajal’s interstitial intestine cells and now known 

to be a stem cell marker. CD117 is expressed in a variety of 

cancers including gastrointestinal stromal tumors, malignant 

melanoma, mastocytosis, acute myelocytic leukemia, anaplastic 

lymphoma, germinoma, and ES/PNET. The EWS gene 

rearrangement in 22q12 demonstration facilitates a prompt 

discrimination between ES/PNET and other morphologically 

similar round cell tumors as more than 90% of all ES/PNET. 

Molecular analysis for C-KIT gene were performed on 

formalin fixed paraffin embedded material, genomic DNA 

amplification has shown wild type exons 9, 11,13 and 17. 

According to the exclusive diagnosis, the present case was 

ultimately diagnosed an extraskeletal ES/PNET. Clinically, 

peripheral primitive neuroectodermal tumor (pPNET) may 

occur anywhere in the body 10 11 . Systematic revision of 54 

cases of extracranial pPNET encountered at Memorial Sloan- 

Kettering Cancer Center over a 20-year period and found that 

the primary sites were thoraco-pulmonary (25 cases), pelvis 

(12 cases), retroperitoneum or abdomen (10 cases), limb (five 

cases), neck (one case) and unknown (one case) 1 . Isolated 

cases of pPNET have been reported in various visceral sites, 

including the pancreas 12 , heart 13 , kidney 14 , ovary 15 , uterus, 

testis, urinary bladder and parotid gland 1 . Despite combined 

therapy with surgery, chemotherapy and irradiation, the prognosis 

of pPNET is poor. Kushner et al. indicated that only 

25% of patients with tumors greater than 5 cm were alive at 

24 months 1 . In conclusion, we have documented a rare case of 

pPNET arising from the small bowel with perforation at onset. 

The perforation was considered to be caused by massive invasion 

of the tumor cells with prominent tumor necrosis and/or 

local ischemic changes. It is important for both surgeons and 

pathologists to remember that intraabdominal pPNET may 

present with acute abdomen. 


references 

1 Kushner BH, Hajdu SI, Gulati SC, et al. Extracranial primitive neuroectodermal 

tumors. Cancer 1991;67:1825-9. 

2 Marina NM, Etcubanas E, Parham DM, et al. Peripheral primitive 

neuroectodermal tumor (peripheral neuroepithelioma) in children. 

Cancer 1989;64:1952-60. 

3 Harper PG, Pringle J, Souhami RL. Neuroepithelioma-a rare malignant 

peripheral nerve tumor of primitive origin. Cancer 1981;48:2282-7. 

4 Askin FB, Rosai J, Sibley RK, et al. Malignant small cell tumor of the 

thoracopulmonary region in childhood. Cancer 1979;43:2438-51. 

5 Dehner LP. Primitive neuroectodermal tumor and Ewing’s sarcoma. 


6 Mor Y, Nass D, Raviv G, Neumann Y, et al. Malignant peripheral 

primitive neuroectodermal tumor (PNET) of the kidney. Med Pediatr 

Oncol 1994;23:437-40. 

7 Furman J, Murphy WM, Jelsma PF, et al. Primary primitive neuroectodermal 

tumor of the kidney. Am J Clin Pathol 1996;106:339-44. 

8 Horn LC, Fischer U, Bilek K. Primitive neuroectodermal tumor of the 

cervix uteri: a case report. Gen Diagn Pathol 1996/97;142:227-30. 

9 Horie Y, Kato M. Peripheral primitive neuroectodermal tumor of the 

small bowel mesentery: a case showing perforation at onset. Pathol Int 

2000;50:398-403. 

10 Enzinger FM, Weiss SW. Primitive neuroectodermal tumors and related 

lesions. In: Soft Tissue Tumors, 3rd ed. New York: Mosby 1995, 

pp. 929-64. 

11 Deb RA, Desai SB, Amonkar PP, et al. Primar primitive neuroectodermal 

tumour of the parotid gland. Histopathology 1998;33:375-8. 

12 Danner DB, Hruban RH, Pitt HA, et al. Primitive neuroectodermal 

tumor arising in the pancreas. Mod Pathol 1994;7:200-4. 

13 Charney DA, Charney JM, Ghali VS, et al. Primitive neuroectodermal 

tumor of the myocardium: A case report, review of the literature, 

immunohistochemical, and ultrastructural study. Hum Pathol 

1996;27:1365-19. 

14 Marley EF, Liapis H, Humphrey PA, et al. Primitive neuroectodermal 

tumor of the kidney. Another enigma: A pathologic, immunohistochemical, 

and molecular diagnostic study. Am. J. Surg. Pathol. 

1997;21:354-9. 

15 Kawaguchi S, Fukuda T, Miyamoto S, et al. Peripheral primitive neuroectodermal 

tumor of the ovary confirmed by CD99 immunostaining, 

karyotypic analysis, and RT-PCR for EWS/FLI-1 chimeric mRNA. Am 

J Surg Pathol 1998;22:1417-22. 

Case n. 4 

Pleomorphic lobular carcinoma in situ of breast 

G. Falconieri, V. Angione, S. Pizzolitto 

Struttura Operativa Complessa di Anatomia Patologica, Azienda 

Ospedaliero Universitaria, Udine, Italy 

Clinical History. A quadrant biopsy is performed in a 55-yearold 

woman who has mammographic evidence of suspicious 

microcalcifications and a core needle biopsy suspicious for 

“ductal” carcinoma in situ with comedonecrosis. Specimen inspection 

reveals poorly demarcated white–gray consolidations 

that, on cut surface, express yellowish comedo-like material. 

Tissue material is fixed in formalin and routinely processed. 

A panel of antibodies was applied to paraffin sections directed 

against estrogen (ER) and progesterone (PR) receptors, p63, 

E-cadherin, low- and high-molecular weight keratins. Hematoxylin-eosin 

stained sections feature gland units irregularly 

distended by a population of medium-sized to large epithelial 

cells with highly atypical nuclei. Central necrosis and microcalcifications 

are noticed. Tumor cells are discohesive, with 

amphophilic to slightly eosinophilic cytoplasm. No infiltrative 

component is recognized. Tumor cells are positive for highmolecular-weight 

keratins as well as ER/PR; they are negative 

for E-cadherin. Cells positive for p63 regularly decorate the 

basal layers of the affected units. The combined features were 

consistent with high-grade pleomorphic lobular carcinoma in 

situ (PLCIS) with comedonecrosis.

lectures 

Discussion. In most cases, recognition of classic lobular 

carcinoma in situ (LCIS) is prompted by a number of histologic 

and cytologic features, including distention of terminal 

tubulolobular units by fairly uniform, small to medium-sized 

round epithelial cells involving more than 50% of the acini 

within a terminal duct lobular unit. Tumor cells are discohesive 

and show mild nuclear atypia with an increased nuclear 

to cytoplasmic ratio; mitoses are rare. LCIS is positive for 

both ER/PR and negative for e-cadherin; it tends to express 

a greater amount of high-molecular-weight keratins. The 

diagnosis of LCIS has several clinical implications: notably, 

it is considered a marker of increased risk for invasive 

breast cancer, either ipsi- or contralaterally, but its incidental 

recognition in core needle biopsy specimens is not generally 

considered an indication for further surgery. In fact, 

unlike ductal carcinoma in situ (DCIS), LCIS documented 

at resection margins is managed conservatively by means of 

tamoxifen and/or follow-up. Variants of LCIS have been reported, 

mirroring a number of architectural and/or cytologic 

changes. Tumour cells may be of larger size, show increased 

variation of cell shape and size, and may exhibit variable 

nuclear pleomorphism (e.g., two- to threefold variation in 

nuclear size), an increased nuclear/cytoplasmic ratio, and 

nucleoli. These lesions are also referred to as pleomorphic 

LCIS (PLCIS). In addition, tumor-cell necrosis (so-called 

comedonecrosis) is often encountered. Tumor cells may also 

feature a relatively abundant, stainable, or “apocrine” cytoplasm. 

Because of their different clinical implications, these 

variants of LCIS must be distinguished from high-grade (or 

grade III) DCIS, in particular when they are associated with 

comedonecrosis. On a morphologic ground, it should be 

pointed out that PLCIS still maintains the basic microscopic 

features of conventional lobular neoplasia and that several 

features militate against DCIS. Conventional areas of LCIS 

may be present next to the PLCIS foci suggesting that the 

two conditions are closely related. Although at a first glance 

necrosis, microcalcification, and high-grade nuclei suggest 

intraductal carcinoma, discohesion of tumor cells is a clue to 

lobular neoplasia. On the other hand, comedonecrosis is not 

a distinctive feature of DCIS, since it can be seen in several 

other proliferative conditions of the breast, such as classic 

LCIS or florid papillomatosis. Like common LCIS, the 

high-grade variant is also consistently positive for ER/PR, 

although in a lesser amount, and negative for E-cadherin, 

whereas a reverse immunostaining pattern is often seen in 

high-grade DCIS. The apocrine variant of PLCIS characteristically 

shows much less ER/PR content and several cytogenetic 

alteration including 16p gain and several losses (11q, 

13q, 17p). In the multistep pathway of lobular neoplasia, it is 

also postulated that apocrine PLCIS is the potential precursor 

of apocrine infiltrating lobular carcinoma. The proliferative 

ki67 index of PLCIS is significantly higher. At times, 

the distinction between DCIS and LCIS may be problematic 

due to “packing” of tumor cells in LCIS and heterogeneous 

e-cadherin staining in some DCIS. It is also possible that 

some such cases might represent true mixed tumor, thus 

indicating that the diagnoses of DCIS and LCIS are not 

mutually exclusive. It is suggested that in situ tumors with 

a mixed phenotype be treated as DCIS. On the other hand, 

the management of patients with PLCIS is still controversial, 

with informed opinions recommending either a conservative 

221 

approach (along to the lines commonly adopted for classic 

LCIS) or more effective surgical measures. A number of 

drawbacks (including the paucity of series, limited data, lack 

of prospective and clinically validated studies) preclude firm 

conclusions. In addition, it is not known whether the breast 

cancer risk (level and laterality) associated with this lesion 

is comparable with that of conventional LCIS. However, 

infiltrating lobular carcinoma may be associated with PLCIS 

in as much as 45% of cases, especially in post-menopausal 

women. Furthermore, an increased phenotypic aggressiveness 

characterizes PLCIS-related invasive lobular carcinoma 

as a high nuclear grade and the overexpression of Her2, p53 

and c-myc oncogenes indicate. For these reasons, accountable 

experts recommend that PLCIS treatment should be 

probably more “DCIS-tailored”, including surgical excision 

of the entire lesion or a quadrant biopsy in cases of PL- 

CIS diagnosed on core needle biopsy. Given the increased 

chance of an associated invasive component the opportunity 

of a sentinel lymph node biopsy should be also considered. 

references 

Barsky SH, Bose S. Should LCIS Be Regarded as a Heterogeneous Disease? 

Breast J 1999;5:407-12. 

Bentz JS, Yassa N, Clayton F. Pleomorphic lobular carcinoma of 

the breast: clinicopathologic features of 12 cases. Mod Pathol. 

1998;11:814-22. 

Cangiarella J, Guth A, Axelrod D, et al. Is surgical excision necessary 

for the management of atypical lobular hyperplasia and lobular carcinoma 

in situ diagnosed on core needle biopsy?: a report of 38 cases 

and review of the literature. Arch Pathol Lab Med 2008;132:979-83. 

Chen YY, Hwang ES, Roy R, et al. Genetic and phenotypic characteristics 

of pleomorphic lobular carcinoma in situ of the breast. Am J Surg 

Pathol 2009;33:1683-94. 

Chivukula M, Haynik DM, Brufsky A, et al. Pleomorphic lobular carcinoma 

in situ (PLCIS) on breast core needle biopsies: clinical significance 

and immunoprofile. Am J Surg Pathol. 2008;32:1721-6. 

Fadare O. Pleomorphic lobular carcinoma in situ of the breast composed 

almost entirely of signet ring cells. Pathol Int 2006;56:683-7. 

Fadare O, Dadmanesh F, Alvarado-Cabrero I, et al. Lobular intraepithelial 

neoplasia [lobular carcinoma in situ] with comedo-type 

necrosis: A clinicopathologic study of 18 cases. Am J Surg Pathol 

2006;30:1445-53. 

Hanby AM, Hughes TA. In situ and invasive lobular neoplasia of the 

breast. Histopathology 2008;52:58-66. 

Jacobs TW, Pliss N, Kouria G, et al. Carcinomas in situ of the breast with 

indeterminate features: role of E-cadherin staining in categorization. 


Koerner F, Maluf H. Uncommon morphologic patterns of lobular neoplasia. 

Ann Diagn Pathol 1999;3:249-59. 

Maluf HM. Differential diagnosis of solid carcinoma in situ. Semin Diagn 

Pathol 2004;21:25-31. 

Maluf HM, Swanson PE, Koerner FC. Solid low-grade in situ carcinoma 

of the breast: role of associated lesions and E-cadherin in differential 

diagnosis. Am J Surg Pathol 2001;25:237-44. 

Middleton LP, Palacios DM, Bryant BR, et al. Pleomorphic lobular carcinoma: 

morphology, immunohistochemistry, and molecular analysis. 


Reis-Filho JS, Simpson PT, Jones C, et al. Pleomorphic lobular carcinoma 

of the breast: role of comprehensive molecular pathology in 

characterization of an entity. J Pathol 2005;207:1-13. 

Schnitt SJ, Morrow M. Lobular carcinoma in situ: current concepts and 

controversies. Semin Diagn Pathol 1999;16:209-23. 

Sneige N, Wang J, Baker BA, et al. Clinical, histopathologic, and biologic 

features of pleomorphic lobular (ductal-lobular) carcinoma in 

situ of the breast: a report of 24 cases. Mod Pathol 2002;15:1044-50. 

Wahed A, Connelly J, Reese T. E-cadherin expression in pleomorphic 

lobular carcinoma: an aid to differentiation from ductal carcinoma. 

Ann Diagn Pathol 2002;6:349-51.

222 

Case n. 5 

Pulmonary metastasis of rhabdoid melanoma 

G. Falconieri, M. Rocco, S. Pizzolitto 

Struttura Operativa Complessa di Anatomia Patologica, Azienda 

Ospedaliero Universitaria, Udine, Italy 

Clinical History. A peripheral nodular opacity is documented 

in a 49 year-old, otherwise healthy woman during a routine 

plain chest-X ray film. About 24 years previously a diagnosis 

of invasive melanoma, no further specified, had been done on 

excisional skin biopsy. A wedge-shaped lung biopsy is carried 

out. The lung specimen reveals a 2 cm, sub-pleural grey 

nodule. A panel of antibodies is applied to paraffin sections 

directed against broad spectrum keratins, TTF1, leucocyte 

common antigen, S100 protein, HMB45, Melan A, tyrosinase 

and MITF-1. Hematoxylin-eosin stained sections feature a 

discohesive proliferation of epithelioid cells. Cytoplasm is 

relatively abundant, brightly eosinophilic to glassy, sometimes 

pushing the nucleus at the periphery and imparting a 

plasmacytoid appearance to tumor cells. Nuclei are vescicular, 

with finely dispersed chromatin and prominent nucleoli. 

Mitotic activity is brisk. Tumor cells are positive for s100 

protein, HMB45, melan A, MITF1 and tyrosinase and negative 

for the other markers. The overall features are consistent 

with metastatic melanoma, with rhabdoid cells. Follow-up at 

3 years is uneventful. 

Discussion. Traditionally, rhabdoid cells are defined as variably 

sized elements featuring abundant, eosinophilic and 

fibrillary cytoplasm recalling rudimentary skeletal muscle 

differentiation. A number of malignant tumors may exhibit a 

rhabdoid phenotype, including carcinoma, melanoma, large 

cell lymphoma, mesenchymal malignancies such as nerve 

sheath sarcoma or synovial sarcoma. Intrathoracic tumors featuring 

rhabdoid cells raise additional interpretive challenges. 

Rhabdoid carcinomas are rare in the lung. Current review of 

the literature shows that less than 40 cases of rhabdoid pulmonary 

carcinoma have been compiled in the American and 

English literature. This carcinoma is considered aggressive if 

compared with the more common non-small cell carcinoma. 

The rhabdoid component may be focal and associated with a 

conventional large cell carcinoma or adenocarcinoma. In the 

series by Tamboli et al., 11 cases of lung rhabdoid carcinomas 

were described: the patients were all adults and presented 

mostly in advanced stage (III or IV) carcinoma. History of 

cigarette smoking could be elicited in more than half of the 

patients. However, the rhabdoid cells were inconsistently 

observed. In addition, in all cases an epithelial differentiation 

could be recognized while in four cases, features of glandular 

differentiation were seen. No specific immunohistochemical 

profile was detected, although most tumors reacted for 

keratins and vimentin. Interestingly, no immunostaining was 

reported for TTF-1, an antibody that frequently reacts with 

conventional adenocarcinoma of lung. Metastatic deposits 

had a predominance of rhabdoid cells. The prognosis for 

pulmonary rhabdoid carcinoma is generally poor; however, 

long-term survivors are apparently not exceptional. In the case 

reported by Hiroshima the patient was a 70-year-old woman 

with a stage I tumor. Recurrence was detected 6 years after 

thoracic surgery and was comparable with the primary lesion. 

The survival time noted in this case, which is even longer for 

the more common non-small cell carcinoma, suggests that 

localized diseases may have a greater chance of curability. 

Similarly, more than 5-year survival was observed in the case 


reported by Kaneko et al., who described a 59-year-old man 

presenting with a lung-confined tumor. The patient developed 

an adrenal metastasis 3 years after lobectomy. On the other 

hand, in their study of 14 patients, Shimazaki et al. found 

that rhabdoid cell-rich tumors entailed a poorer prognosis 

regardless of tumor stage, and their observation was also 

validated by a statistical analysis suggesting than the number 

of rhabdoid cells may be a significant prognosticator. Interestingly, 

if the compiled cases of rhabdoid lung carcinoma are 

considered, lymph node metastases have been documented in 

13 of 33 cases, the proportion of lung tumors without local 

metastases at presentation being roughly two thirds. Also, we 

are not aware of cases in which the metastases were found 

prior to the lung tumor. Immunohistochemistry and electron 

microscopy suggest an epithelial lineage, given the consistent 

immunoreactivity for keratins and ultrastructural features 

such as paranuclear intermediate filaments. Tumor size and 

stage did not appear to have any predictive means. 

Notably, the differential diagnosis of extra-renal rhabdoid 

neoplasm may be a difficult task because of its numerous 

microscopic mimickers. In the lung, the matter is further 

compounded by the existence of site-specific simulators: for 

the sake of brevity, only some of such lesions will be briefly 

discussed. The differentiation may be difficult on pure morphologic 

grounds and must be clinically driven, first. Then, 

the judicious use of a limited antibody panel may suffice for 

reliably segregating these lesions and singling out those that 

may benefit from specific treatments. Large cell lymphoma 

may occur as a primary pulmonary and/or mediastinal lesion. 

Tumor cells are often immunopositive for LCA and lymphoid 

antigens such as CD3, CD20, or CD30 may be expressed. 

Specific pulmonary lymphoproliferative disorders such as 

lymphomatoid granulomatosis may be recognized by virtue 

of clinical and pathologic features, although resorting to immunohistochemistry 

is useful to rule out other conditions. 

Melanoma, as the case at issue indicates, can be virtually 

indistinguishable from rhabdoid tumors as well as from any 

other malignancy inasmuch as rhabdoid changes are often 

detected in recurrent or metastatic lesions; immunoreactivity 

for S100 protein, in absence of staining for other antigens, is, 

however, expected in most cases of melanoma. Epithelioid angiosarcoma 

may rarely but not exceptionally occur in the lung 

as a primary tumor, either alone or associated with extensive 

pleural involvement. Poorly differentiated forms may lack 

evidence of diagnostic clues such as a freely anastomosing 

vascular channel pattern or intracellular lumina. In addition, 

tumor cells may have a rich stainable cytoplasm, recalling that 

seen in rhabdoid tumors, and epithelioid angiosarcoma cells 

may react to antibodies against keratins, thus suggesting a 

tumor of true epithelial lineage. However, clues such as abortive 

vessels or intracellular lumina may be often recognized; 

notably, angiosarcoma is often positive for factor VIII-related 

antigen, CD31, CD34 and fli-1. 

references 

Attems JH, Lintner F. Pseudomesotheliomatous adenocarcinoma of the 

lung with rhabdoid features. Pathol Res Pract 2001;197:841-6. 

Cavazza A, Colby TV, Tsokos M, et al. Lung tumors with a rhabdoid 

phenotype. Am J Clin Pathol 1996;105:182-8. 

Chetty R. Combined large cell neuroendocrine, small cell and squamous 

carcinomas of the lung with rhabdoid cells. Pathology 2000;32:209- 

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Chetty R, Bhana B, Batitang S, et al. Lung carcinomas composed of rhabdoid 

cells. Eur J Surg Oncol 1997;23:432-4. 

Falconieri G, Moran CA, Pizzolitto S, et al. Intrathoracic rhabdoid carcinoma: 

a clinicopathological, immunohistochemical, and ultrastructural 

study of 6 cases. Ann Diagn Pathol 2005;9:279-83.

lectures 

Hiroshima K, Shibuya K, Shimamura F, et al. Pulmonary large cell carcinoma 

with rhabdoid phenotype. Ultrastruct Pathol 2003;27:55-9. 

Kaneko T, Honda T, Fukushima M, et al. Large cell carcinoma of the 

lung with a rhabdoid phenotype. Pathol In. 2002;52:643-7. 

Rubenchik I, Dardick I, Auger M. Cytopathology and ultrastructure of 

primary rhabdoid tumor of lung. Ultrastruct Pathol 1996;20:355-60. 

Shimazaki H, Aida S, Sato M, et al. Lung carcinoma with rhabdoid cells: 

a clinicopathological study and survival analysis of 14 cases. Histopathology 

2001;38:425-34. 

Tamboli P, Toprani TH, Amin MB, et al. Carcinoma of lung with rhabdoid 

features. Hum Pathol 2004;35:8-13. 

Wick MR, Ritter JH, Dehner LP. Malignant rhabdoid tumors: a clinicopathologic 

review and conceptual discussion. Semin Diagn Pathol 

1995;12:233-48. 

Case n. 6 

Small cell neuroendocrine carcinoma 

with myofibroblastic and skeletal muscle 

differentation 

G. Pelosi 


IRCCS Istituto Nazionale dei Tumori, Milano 

Clinical history. A 76-year-old Caucasian man, former smoker 

since 30 years (previously he smoked 30 cigarettes/day for 

at least 20 years), underwent left upper lobectomy for Aspergillus 

infection (fungus ball) in 1980. The patient experienced 

hemoptysis in 2003, when a granulomatous inflammation was 

found in the left main bronchus (likely related to the previous 

surgery) that was treated with LASER therapy. In May 

2007, the patient began to complain of hemoptysis again and 

a chest X ray examination showed a huge tumor mass in the 

right upper lobe, measuring 6-7 cm in diameter. A total body 

CT scan investigation confirmed the presence of this tumor 

mass in the right upper lobe, sized 67 × 48 × 50 mm, which 

compressed the lobar bronchus and apparently infiltrated the 

azygos vein but was not associated with pleural effusion or 

distant metastases. As PET scan examination did not reveal 

distant metastases, a right upper lobectomy with a complete 

hilar-mediastinal lymphadenectomy was performed at the end 

of May 2007. The postoperative clinical course was uneventful, 

and the patient was discharged ten days later in good 

general conditions. The tumor measured 7 cm in its greatest 

dimension, was located in the pulmonary upper lobe, attained 

the visceral pleura but with no azygos vein infiltration and 

showed necrosis and hemorrhage with friable tissue on cut 

section. Histopathologic examination revealed a biphasic tumor 

composed of 1) a high-grade neuroendocrine carcinoma 

component arranged in nests, trabeculae or solid aggregates 

with finely granular chromatin and inconspicuous nucleoli, 

and 2) a spindle to pleomorphic cell component with abundant 

collagen deposition resembling high-grade sarcoma. The two 

components were intimately intermingled with each other, but 

a slight prevalence of the sarcoma-like component (55-60%) 

was noted. The immunohistochemical study revealed a strong 

and diffuse positivity for cytokeratins (AE1-AE3) in the epithelial-like 

component and for CD56 in all tumor cells, and 

a more variable immunoreactivity for S-100 protein, GFAP, 

synaptophysin, sarcomeric actin, neurofilaments, TTF-1, 

smooth-muscle actin, calponin, caldesmon and CD10. Co-expression 

of cytokeratins, desmin and myogenin was localized 

in the same aggregates of tumors cells exhibiting epithelial 

features, suggesting dipartite differentiation. Ki-67 labeling 

index was higher in the epithelial-like component (80 to 90%) 

than in the sarcoma-like population (30 to 40%). Electron 

223 

microscopy study showed myofibroblastic differentiation 

in the spindle cell component, whereas neuroendocrine differentiation 

in the epithelial cell-like component shows and, 

less frequently, coexisting bundles of contractile filaments 

containing abortive Z bands reminiscent of skeletal muscle 

differentiation, suggesting co-localized rhabdomyoblastic and 

neuroendocrine differentiation. Tumor staging was pT3N1M0 

because of a single peribronchial lymph node metastasis. After 

adjuvant chemotherapy, the patient is alive and well with 

no sign of metastatic disease. 

Discussion. The case here reported is a combined small-cell 

carcinoma with skeletal muscle differentiation and spindle 

cell sarcoma component of myofibroblastic type. Combined 

small-cell carcinoma variant makes up about 10 to 30% of 

all small cell carcinomas of the lung. It refers to the variable 

admixture of small-cell and non-small cell carcinoma elements, 

the latter usually including squamous cell carcinoma, 

adenocarcinoma and/or large-cell carcinoma 1 , and much more 

uncommonly spindle cell 2 3 or giant cell carcinoma 4 5 . Other 

exceedingly rare combinations in the theme of neuroendocrine 

carcinomas of the lung include associations of atypical 

carcinoid and rhabdomyosarcoma 6 , small cell carcinoma 

plus adenocarcinoma and spindle-shaped cell tumor 7 , small 

cell carcinoma plus squamous cell carcinoma and spindle 

cell carcinoma 8 , small cell carcinoma plus sarcomatoid carcinoma 

with either spindle cell or giant cell carcinoma 9 , and 

carcinoid and adenocarcinoma 10 11 . Moreover, occurrence of 

small cell carcinoma with skeletal muscle differentiation has 

been described in the larynx 12 , as well as in the skin, nasal 

cavity and urinary bladder 13 , and combination of high-grade 

neuroendocrine carcinoma and alveolar rhabdomyosarcoma is 

also on record in the anorectal junction 14 . Although intimate 

intermingling of small cell carcinoma elements with scattered 

rhabdomyoblastic cells 13 and even tripartite differentation in 

individual cells with concurrent epidermoid, glandular and 

neuroendocrine features 15 or dipartite differentiation with 

rhabdomyogenous and cytokeratin expression within the same 

mesenchymal tumor cells of carcinosarcomas 16 have been described 

in the literature, the current case is worth of mention 

because of dipartite differentiation of small cell carcinoma 

and rhabdomyosarcoma coexisting with spindle cell sarcoma, 

probably derived from a common protoepithelial stem cell. It 

has been demonstrated that additional genetic alterations may 

be found in the mesenchymal component of sarcomatoid carcinomas 

of the lung, which were lacking in the epithelial one, 

suggesting mesenchymal transformation during epithelial carcinogenesis 

17 . In our case, the spindle cell component lacked 

any epithelial or rhabdomyogenous differentiation, probably 

because of a complete myofibroblastic/smooth muscle transdifferentiation 

of carcinomatous cells or early divergence during 

tumor progression of the same ancestor lesion. 

references 

1 Travis W, Brambilla E, Muller-Hermelink H, et al. Tumours of the 

lung, pleura, thymus and heart. Edited by Cancer IAfRo. Lyon: IARC 

Press 2004, p. 344. 

2 Tsubota Y, Kawaguchi T, Hoso T, et al. A combined small cell and 

spindle cell carcinoma of the lung. Report of a unique case with immunohistochemical 

and ultrastructural studies. Am J Surg Pathol 

1992;16:1108-15. 

3 Niho S, Yokose T, Nagai K, et al. A case of synchronous double 

primary lung cancer with neuroendocrine features. Jpn J Clin Oncol 

1999;29:219-25. 

4 Bégin P, Sahai S, Wang N. Giant cell formation in small cell carcinoma 

of the lung. Cancer 1983;52:1875-9. 

5 Müller K, Fisseler-Eckhoff A. Small cell bronchial cancer--pathologic 

anatomy. Langenbecks Arch Chir Suppl Kongressbd 1991:534-543.

224 

6 Rainosek D, Ro J, Ordonez N, et al. Sarcomatoid carcinoma of the 

lung. A case with atypical carcinoid and rhabdomyosarcomatous 

components. Am J Clin Pathol 1994;102:360-4. 

7 Hsiao H, Tsai H, Liu Y, et al. A rare case of combined small-cell 

lung cancer with unusual soft tissue metastasis. Kaohsiung J Med Sci 

2006;22:352-6. 

8 Gotoh M, Yamamoto Y, Huang C, et al. A combined small cell carcinoma 

of the lung containing three components: small cell, spindle cell 

and squamous cell carcinoma. Eur J Cardiothorac Surg 2004;26:1047- 

9. 

9 Fishback N, Travis W, Moran C, et al. Pleomorphic (spindle/giant 

cell) carcinoma of the lung. A clinicopathologic correlation of 78 

cases. Cancer 1994;73:2936-45. 

10 Sen F, Borczuk AC. Combined carcinoid tumor of the lung: a combination 

of carcinoid and adenocarcinoma. Lung Cancer 1998;21:53-8. 

11 Cavazza A, Toffanetti R, Ferrari G, et al. Combined neoplasia of the 

lung: description ofa acase of adenocarcinoma mixed with typical 

carcinoid. Pathologica 2001;93:216-220. 

12 Doglioni C, Ferlito A, Chiamenti C,et al. Laryngeal carcinoma showing 

multidirectional epithelial neuroendocrine and sarcomatous differentation. 

ORL J Otorhinolaryngol Relat Spec 1990;52:316-26. 

13 Eusebi V, Damiani S, Pasquinelli G, et al. Small cell neuroendocrine 

carcinoma with skeletal muscle differentation. Am J Surg pathol 

2000;24:223-30. 

14 Roncaroli F, Montironi R, Feliciotti F, et al. Sarcomatoid carcinoma 

of the anorectal junction with neuroendocrine and rhabdomyoblastic 

features. Am J Surg Pathol 1995;19:217-23. 

15 McDowell EM, Trump BF. Pulmonary smell cell carcinoma showing 

tripartite differentiation in individual cells. Hum Pathol. 1981;12:286- 

94. 

16 Wick MR, Ritter JH, Humphrey PA. Sarcomatoid carcinomas of the 

lung: a clinicopathologic review. Am J Clin Pathol 1997;108:40-53. 

17 Dacic S, Finkelstein SD, Sasatomi E, et al. Molecular pathogenesis of 

pulmonary carcinosarcoma as determined by microdissection-based 

allelotyping. Am J Surg Pathol 2002;26:510-6. 

Case n. 7 

Hepatocellular carcinoma with unusual 

endocrine features 

M. Milione, F. Melotti, A. Carbone * , G. Pelosi 


IRCCS Istituto Nazionale dei Tumori, Milano; * IRCCS - C.R.O. Centro 

di Riferimento Oncologico di Aviano, Udine 

Clinical History. During a follow-up visit for chronic hepatitis 

C in a 36-years old man, a nodular lesion measuring around 

1 cm in diameter was detected in the S5 liver segment using 

abdominal ultrasonography. Serum a-fetoprotein (AFP) was 

25.3 ng/ml. Two months later a new nodule measuring just 

about 3,3 mm in diameter was found in the S6 liver segment. 

Three months later the S6 nodule grew up rapidly and was approximately 

2.9 cm in diameter. The tumor in S5 grew slowly 

and also achieved approximately 2.3 cm. AFP levels were 

elevated to 3787.0 ng/ml. Hepatitis B surface antigen and antibody 

were negative. Carcinoembryonic antigen (CEA) was 

within normal limits. Using computed tomography (CT) the 

S6 tumor was enhanced in the early phase and the S5 tumor 

showed peripheral enhancement in the early phase. Lymph 

nodes were not distinguished. In abdominal ultrasonography 

(US) the S5 and S6 tumors demonstrated a mosaic pattern. 

At our institution, a liver core biopsy was performed on both 

nodules. 

Discussion. Histologically the S6 tumor was composed of 

round tumor cells with moderately represented eosinophilic 

cytoplasm and round nuclei showing a trabecular organization. 

The tumor was reliable with moderately differentiated 

HCC, but was intermingled with small round cells with 

scarce cytoplasm, which resembled those found in endocrine 


carcinoma (EC). This population shaped solid nests along 

with the trabecular arrangement intermitted with pseudoacinar 

areas and rosette formation. It stained positively for 

chromogranin-A, synaptophysin, CD56, strongly for CK19, 

alpha phetoprotein (AFP) and Glypican 3; and it resulted 

negative for Hep Par1, CK7, CK20, TTF1, serotonin, insulin, 

and glucagon. MIB-1 immunostain demonstrated high proliferative 

activity (70-80% of the cells were positive). p53 was 

positive in 60% of neoplastic cells. The diagnosis of HCC 

with Endocrine features was thus established. The S5 nodule 

showed morphologic and immunophenotipic features of classic 

well differentiated (G1) HCC. On rare occasions endocrine 

character appear within a Hapatocelluar Carcinoma (HCC) 

nodule. Origin of hepatic endocrine tumors is vague, the most 

interesting hypotheses planned are: A) derivation from the endocrine 

cells nearby in the intrahepatic bile duct epithelium 1 

and B) endocrine differentiation of a distinct malignant stem 

cell originator of additional hepatic malignant tumors 2 . The 

growth of a lot of hepatic carcinoids consisting of uniform tumor 

cells, in a noncirrhotic liver without necrosis or degeneration, 

wired the former notion. Carcinomas of the extrahepatic 

biliary system showed a high rate of endocrine differentiation, 

and had a poor prognosis 3 . Intrahepatic cholangiocarcinoma 

and HCC have also been reported to go through endocrine 

differentiation. Positivity on immunohistochemistry for neuroendocrine 

markers in some cases of HCC was described 4 5 . 

These findings support the second hypothesis. It is also well 

known that in HCC, a less well-differentiated tumor clone, 

arises within the original tumor and proliferates, eventually 

replacing the latter. As a result, a clone undergoing endocrine 

differentiation may proliferate, replacing the entire tumor, and 

form a complete EC. In the present case, the primary hepatic 

EC microscopically resembled an HCC. The occurrence of 

this EC on a background of hepatitis C cirrhosis and its coexistence 

with another HCC strongly suggest that this primary 

hepatic EC did not arise de novo from the endocrine system in 

the hepatic parenchyma, and leads us to speculate that one of 

the HCCs underwent endocrine differentiation and thereafter 

transformed into a EC. In general, endocrine carcinomas grow 

rapidly and have a poor prognosis. Endocrine differentiation 

in other organ cancers has more malignant behavior (3,5) In 

the present case the S6 tumor with neuroendocrine carcinoma 

showed more rapid growth than a typical HCC of S5. In addition, 

labeling index of p53 and Ki-67 of the small round 

endocrine component were significantly higher than those of 

the HCC component, and suggesting that the former has more 

abnormalities of p53 and higher proliferative activity. HCC 

with endocrine appearance has higher proliferative activity 

and malignant potential than ordinary HCC. Few authors have 

described primary endocrine tumors in the liver combined 

with hepatocellular carcinoma, but these represented differentiation 

of the malignant liver cells into a endocrine tumor 6 7 . 

Separately from the collision and combined types, endocrine 

tumors can also arise in an isolated manner primarily in the 

liver in the shape of carcinoids or highgrade tumors represented 

by small cell carcinomas 1 8 . The distinction between primary 

and metastatic endocrine tumors is important in making 

the diagnosis of primary hepatic EC, as the liver is the most 

common site of metastasis for these tumors. We investigated 

the full body, principally the lung, pancreas, and gastrointestinal 

tract, for additional primary lesions by CT, MRI, and 

endoscopy, but were incapable to find any primary lesions 

outside the liver. It is imperative to distinguish carcinoid 

tumors and EC clinic pathologically as EC is more malignant 

and has poorer prognosis 2 . Even though various hormones

lectures 

such as serotonin and gastrin have been frequently established 

in primary hepatic carcinoid, such positive staining is hard to 

identify in primary liver EC because of poor differentiation. 

On the other hand, some collective immunohistochemical 

features may origin misunderstanding, and care should hence 

be taken in the diagnosis of EC. 

references 

1 Pilichowska M, Kimura N, Ouchi A, et al. Primary hepatic carcinoid 

and neuroendocrine carcinoma clinicopathological and immunohistochemical 

study of five cases. Pathol Int 1999;49:318-24. 

2 Gould VE, Banner BF, Baerwaldt M. Neuroendocrine neoplasms in 

unusual primary sites. Diagn Histopathol 1981;4:263-77. 

3 Hsu W, Dezidel DJ, Gould VE, et al. Neuroendocrine differentiation 

and prognosis of extrahepatic biliary tract carcinomas. Surgery 

1991;110:604-10. 

4 Artopoulos JG, Destuni C. Primary mixed hepatocellular carcinoma 

with carcinoid characteristics: a case report. Hepato- Gastroenterology 

1994;41:442-4. 

5 Alpert LI, Zak FG, Werthamer S, et al. Cholangiocarcinoma: a 

clinicopathologic study of five cases with ultrastructural observations. 

Hum Pathol 1974;5:709-28. 

6 Barsky SH, Linnoila I, Triche, TJ, et al. Hepatocellular carcinoma 

with carcinoid features. Hum Pathol 1984;15(9):892-4. 

7 Yamaguchi R, Nakashima O, Ogata T, et al Hepatocellular carcinoma 

with an unusual neuroendocrine component. Pathol Int 

2004;54(11):861-5. 

8 Rückert RI, Rückert JC, Dörffel Y, et al. Primary hepatic neuroendocrine 

tumor: successful hepatectomy in two cases and review of the 

literature. Digestion 1999;60(2):110-6. 

Case n. 8 

Intrahepatic cholangiocarcinoma with thyroidlike 

features 

V. Eusebi 

Dipartimento di Ematologia e Scienze Oncologiche “L. e A. Seragnoli”, 

Osp. Bellaria, Anatomia Patologica, Bologna 

Clinical history. A 52-year-old male suffering from abdominal 

pain for several months was admitted to hospital. Family 

history was not relevant. A CT scan of the abdomen revealed 

a homogeneous enhancing lesion of the right hepatic lobe of 

18 cm in greatest axis. No other lesions were present in the 

chest, head and neck and urogenital apparatus. Laboratory 

data, including thyroid function tests, were within normal 

range. A core biopsy of the liver mass was obtained which led 

to the diagnosis of cholangiocarcinoma. This was followed by 

hepatic lobectomy. Eighteen months after surgery the patient 

is alive with no evidence of recurrence or metastatic disease. 

Histology from pre operative biopsy as well as from surgical 

specimen was identical. At low power the lesion was circumscribed 

by a thin fibrous capsule and showed a remarkable 

follicular architecture. Follicles were of various sizes, ranging 

from small to large. The content of follicles closely resembled 

colloid being pale eosinophilic with occasional vacuoles at 

the interface with the epithelium. The neoplastic cells were 

mostly cuboidal with granular eosinophilic cytoplasm. Nuclei 

were round to ovoid in shape, with scanty chromatin and 

inconspicuous nucleoli. Some nuclei were clear and showed 

neat membrane that was occasionally grooved. Mitoses were 

1/10 hpf (x400). The eosinophilic luminal content of follicles 

was rich in mucosubstances as evidenced by positivity for Alcian 

blue pH 2.5 and PAS after diastase digestion. Neoplastic 

cells were positive for CK7, CK19, CAM 5.2 and CK AE1 

225 

and consistently negative for CEA, CK20, CD 56, hepatic 

specific antigen, thyroglobulin, TTF-1, CD56, synaptophysin 

and chromogranin. 

Discussion. The case here reported was a large hepatic 

tumour showing spongy cut surface with occasional bloodfilled 

cystic spaces. Histologically a remarkable follicular 

architecture was evident. Neoplastic cells were mostly cuboidal 

with regular nuclei showing grooves and clearing of 

the chromatin. The macroscopic and histological features 

are similar to a follicular variant of papillary carcinoma of 

the thyroid. Metastasis to the liver from well-differentiated 

carcinomas of the thyroid is a well known, although rare phenomenon 

1 . An additional remote possibility would be that of 

ectopic normal thyroid tissue in the liver as the case reported 

by Strohschneider et al. 2 Nevertheless in the present patient 

the neoplastic cells were all negative for thyroglobulin and 

TTF-1 at the variance with well differentiated thyroid neoplasms 

and ectopic normal thyroid tissues wherever they are 

found. Finally no evidence of a thyroid lesion of any kind was 

found by clinical investigations, ultrasonography, CT scan 

and laboratory tests. As no other primary was found after 13 

months, including breast, kidney and pancreas, the cholangiocellular 

nature of the hepatic neoplasm was then favoured 

for the presence of mucosubstances and further supported by 

immunohistochemistry that evidenced CK7, CK19 and CAM 

5.2 positivity along with CK20 and hepatic specific antigen 

negativity. In differential diagnosis bile duct adenoma and 

biliary cystoadenomas of the serous variant have to be taken 

in consideration. It seems that a similar case had been reported 

by Foucar et al. 7 who described an unusual variant of peripheral 

well-differentiated cholangiocarcinoma in a 27 yr-old 

pregnant patient with histological features very similar to the 

present case. Neoplastic lesions having thyroid-like features 

have been reported in the breast and in the kidney 1 3-6 . In the 

breast the several cases described were superimposable to the 

tall cell variant of papillary carcinoma of thyroid 1 3 . The cases 

reported in kidney were very similar to follicular carcinoma 

of thyroid 4-6 . Therefore it seems that thyroidlike features can 

occur in tumours located in different organs and liver has to 

be added to the list. To be aware of the existence of tumours 

in the liver histologically mimicking follicular carcinomas 

of thyroid can avoid diagnostic erroneous interpretation and 

more cases are needed to establish the biological behaviour. 

references 

1 Eusebi V, Damiani S, Ellis IO, et al. Breast tumor resembling the 

tall cell variant of papillary thyroid carcinoma. Am J Surg Pathol 

2003;27:1114-8. 

2 Strohschneider T, Timm D, Worbes C. Ectopic thyroid gland tissue in 

the liver. Der Chirurg 1993;64:751-3. 

3 Tosi AL, Ragazzi M, Asioli S, et al. Breast tumor resembling the tall 

cell variant of papillary thyroid carcinoma: report of 4 cases with 

evidence of malignant potential. Int J Surg Pathol 2007;5:14-9. 

4 Jung SJ, Chung JI, Park SH, et al. Thyroid follicular carcinoma-like 

tumor of the kidney: a case report with morphologic, immunohistochemical 

and genetic analysis. Am J Surg Pathol 2006;30:411-5. 

5 Sterlacci W, Verdofer I, Gabriel M, et al. Thyroid follicular carcinoma-like 

renale tumor: a case report with morphologic, immunophenotypic, 

cytogenetic and scintigraphic studies. Virchows Arch 

2008;452:91-5. 

6 Amin MB, Gupta R, Ondrej H, et al. Primary thyroid like follicular 

carcinoma of the kidney: report of six cases of a histologically distinct 

adult renal epithelial neoplasm. Am J Surg Pathol 2009;33:393-400. 

7 Foucar E, Kaplan LR, Gold JH, et al. Well differentiated peripheral 

cholangiocarcinoma with unusual clinical course. Gastroenterlogy 

1979;77:347-53.

226 

Glioneuronal tumors with leptomeningeal 

dissemination 

Marina P. Gardiman, Matteo Fassan. 

Department of Diagnostic Medical Sciences and Special Therapies, 

Pathology Unit, University of Padova, Padova (PD), Italy 

Background. Glioneuronal tumors are a group of primary 

brain neoplasm of relatively recent acquisition in the World 

Health Organization (WHO) Classification of Central Nervous 

System (CNS) tumors which has been recently expanded with 

new recognized entities such as rosette-forming tumor of the 

fourth ventricle, the papillary glioneuronal tumor and rosetted 

glioneuronal tumor/glioneuronal tumor with neuropil-like 

islands 1 . Glioneuronal tumors are characterized by a biphasic 

neurocytic and glial population. The neuronal component consists 

of synaptophysin-positive neurocytes with round nuclei 

and clear cytoplasm occasionally intermingled with neurons 

and intermediated-size “ganglioid” cells, whereas the glial 

component exhibits features of glial fibrillary acidic protein 

(GFAP) positive astrocytes. The histogenesis of these tumors 

is unclear, but an origin from multipotent precursors capable 

of divergent differentiation has been suggested 2 . 

Leptomeningeal dissemination in glioneuronal tumors is very 

rare, but the incidence in low grade gliomas (a name for a 

wide variety of neoplasm of glial or mixed glial-neuronal 

origin) is estimated at 5% at diagnosis and 7-10% at progression. 

Among neoplasm of astrocytic origin, it is well known 

that pilocytic astrocitoma can disseminate 3 but also a new 

codified glial neoplasm in the 2007 WHO Classification of 

tumors of the SNC, such as pilomyxoid astrocitoma, shows a 

characteristic high tendency to disseminate. 

The spreading along the subarachnoidal spaces of cerebrospinal 

fluid (CSF)of glioneuronal neoplasms has been reported 

in the last few years more frequently than in the past probably 

related to the more diffuse use of magnetic resonance imaging 

(MRI) in tumor staging and follow-up. Well-established 

examples of glioneuronal tumors with leptomeningeal dissemination 

include ganglioglioma and pleomorphic xantoastrocitoma 

4 . 

In diagnostic practice is still possible to encounter glioneuronal 

tumors that cannot be placed into any of the well-defined 

WHO categories despite this growing list of entities. We have 

recently published four pediatric cases of diffuse leptomeningeal 

tumors which cannot easily be classified in the currently 

used CNS WHO classification, but have the histological and 

immunohistochemical criteria to be considered as glioneuronal 

tumors. 

Methods. Cases were retrieved from the Institutional files of 

the authors. One case had been previously reported as spinal 

low-grade neoplasm with diffuse leptomeningeal dissemination 

5 . Pathology reports and the histological slides were reviewed. 

Immunohistochemical analysis was performed using 

the standard avidin-biotin-peroxidase method. Fluorescence 

in situ hybridization (FISH) was performed on formalin-fixed, 

paraffin-embedded tissues of one of the considered cases. 

Clinical findings: the children were all admitted to the Padova 

University Hospital with the symptoms and sign of 

hydrocephalus (morning headache, vomiting and increased 

cranial circumference). Neuroradiological findings: tumors 


Problems in neuropathology 

Moderators: F. Giangaspero (Roma), G. Cenacchi (Bologna) 

were characterized by similar radiological appearance, i.e. 

contrast-enhanced head and spine magnetic resonance images 

revealed a tetraventricular communicating hydrocephalus, a 

diffuse cerebral and spinal leptomeningeal enhancement, a 

marked progressive cortical and subcortical cystic involvement 

of the cerebellum, basal temporal and frontal lobes, 

brainstem and spinal cord without evidence of a primary 

intraparenchymal mass. 

Results. In all cases a dural biopsy was performed. Minute 

samples characterized by a pearly opacified surface and 

an increased consistence were obtained. The histological 

samples showed thickened desmoplastic leptomeninges with 

sclerohyaline bands and enlarged capillary-sized blood vessels 

diffusely infiltrated by a monotonous population of cells 

arranged in straight lines or in small lobules within a compact 

to loosely fibrillary stroma. Cells were characterized by round 

to oval nuclei with finely granular dispersed chromatin, inconspicuous 

nucleoli with clear oligodendrocyte-like features 

with perinuclear haloes. No mitosis, necrosis, calcifications, 

lymphoid infiltration, myxoid changes or endothelial vascular 

proliferation were observed. No Rosenthal fibers, nor rosettes 

or pseudorosettes were detected. 

Tumor cells showed diffuse immunoreactivity for synaptophysin 

and S100, patchy reactivity for GFAP and negative for 

neurofilaments or epithelial membrane antigen. Proliferation 

index, as percentage of MIB1-positive cells [MIB1 labeling 

index (MIB1 L.I.)], was always less than 1%. 

Only in case #3, after a first dural biopsy (performed in 

2002), we obtained a significative sample of tissue from the 

lesion appeared on the inner surface of the lateral ventricle 

frontal horn (2007). The analyzed sample was composed by 

a biphasic architecture. The more differentiated part of the 

tumor was abutting in the ventricle lumen and composed by 

uniform small cuboidal cells with round nuclei and scant clear 

cytoplasm intermingled with “ganglioid” cells occasionally 

arranged in perivascular pseudorosettes or pseudopapillary 

structures. 

Additional features include fibrillary areas mimicking neuropil 

and rare foci of microvascular proliferation of the capillary-sized 

blood vessels. The inner part of the tumor showed 

anaplastic histological features with increased cellularity and 

a diffuse honeycomb pattern of growth. The neoplastic oligodendrocyte-like 

cells, diffusely infiltrating the brain parenchyma 

and showed mild polymorphism with hyperchromatic 

nuclei. Endothelial proliferation in the branching capillaries 

was evident. No tumoral necrosis was observed. An increased 

mitotic activity (three mitotic figures (10 high-power field) 

with an MIB1 L. I. higher than 5% was detected. FISH 

analysis revealed deletion of 1p, whereas 19q was intact. A 

significant number of nuclei were immunopositive for Neu-N. 

Tumor cells were also diffusely synaptophysin positive. Scattered 

cells were GFAP positive. 

Discussion. The main histological characteristics of these 

tumors affecting our four pediatric patients deserve special 

considerations. On microscopic examination, these tumors 

were composed by cells characterized by round to oval nuclei 

with inconspicuous nucleoli with clear oligodendrocyte-like 

cytoplasm. These histological findings might have favored the 

diagnosis of oligodendrogliomas and oligodendrogliomatosis 

in some of the similar cases presented in the Literature 6 7 .

lectures 

Moreover, the diagnosis of oligodendrogliomas was achieved 

only on cytologic criteria, that is, clear cytoplasm and round 

nuclei caused by the lack of specific markers for oligodendrogliomas. 

As previously described in a case of diffuse leptomeningeal 

oligodendroglioma 7 , we found the deletion of 1p 

in one of our cases. This deletion is neither pathognomonic 

of oligodendroglioma 8 nor particularly frequent in pediatric 

cases that usually do not show 1p/19q co-deletions 9 . In oligodendrogliomas, 

synaptophysin immunoreactivity is usually 

caused by residual parenchyma and is frequently seen at the 

infiltrating tumor borders. In our cases, the constant immunohistochemical 

profiles observed (i.e.: the positive reactivity 

for synaptophysin and Neu-N) strongly suggest a glioneuronal 

commitment of the neoplasm. 

Also, neurocytomas and dysembrioplastic neuroepithelial 

tumor (DNT) show similar histological/immunophenotypical 

profile 10 , but in our cases a common characteristic was 

the absence of a primary neoplastic mass in contrast with the 

pathological evidence of the other glioneuronal tumors (i.e.: 

DNT, extraventricular neurocytoma, papillary glioneuronal 

tumor and rosette-forming glioneuronal tumor) 1 . 

A possible explanation about the origin of these diffuse leptomeningeal 

tumors could be isolated groups of glioneuronal 

progenitor cells entrapped in the context of the leptomeninges 

during the primitive migration. These embryonal cells 

could be able of divergent differentiation with neuronal, 

oligodendroglial and astrocytic features 11 . In fact, cases of 

morphologically classic oligodendroglioma with neurocytic 

rosettes or neurocytomas arboring 1p/19q deletion have been 

described 11 12 . The description of these entities suggests a 

histogenetic overlap between oligodendroglioma and extraventricular 

neurocytoma 11 , and further supports the existence 

of a new “superfamily” of tumors with oligodendroglial and 

neurocytic potential in which our series of diffuse leptomeningeal 

glioneuronal tumors could be included. 

Interestingly, in the other similar cases presented in the literature, 

but considered as diffuse leptomeningeal oligodendrogliomas, 

the immunohistochemical profiles are quite variable 

and sometimes inconsistent which could be related to the 

glioneuronal nature of the described neoplasms, and further 

indicate the difficulty to classify these types of tumors. 

Three of four patients are alive up to 2 years of follow-up, following 

minimal to no clinical intervention, and these data suggest 

these tumors as neoplasm with a slow progressive and quite 

indolent course. However, in case #3, the subsequent appearance 

of a bulking neoplastic intraventricular lesion with anaplasia, 

high mitotic index and focal vascular endothelial proliferation 

suggests a potential aggressive biological transformation. 

In conclusion, we hypothesized that the tumors affecting the 

children we described represent a new nosological entity characterized 

by: i) intense enhancement of subarachnoidal space 

with cystic lesions; ii) diffuse leptomeningeal infiltration by 

glioneuronalcells without a primary mass; and iii) quite indolent 

course. 

For these reasons, this group of neoplasms could be descriptively 

named “diffuse leptomeningeal glioneuronal tumors.” 

Further studies and larger validation are needed to test our 

hypothesis and to consider “leptomeningeal glioneuronal 

tumors” as a distinct nosological entity in the SNC tumor 

classification. 

references 

1 Louis DN, Ohgaki H, Wiestler OD, et al. WHO Classification of Tumours 

of the Central Nervous System, 4th ed. Lyon: IARC:2007. 

2 Allende DS, Prayson RA. The Expanding Family of Glioneuronal 

Tumors. Adv Anat Pathol 2009;16:33-9. 

227 

3 Kreiger PA, Okada Y, Simon S, et al. Losses of chromosomes 1p 

and 19q are rare in pediatric oligodendrogliomas. Acta Neuropathol 

2005;109:387-92. 

4 Passone E, Pizzolitto S, D’Agostini S, et al. Non-anaplastic pleomorphic 

xantoastrocitoma with neuroradiological evidences of leptomeningeal 

dissemination. Childs Nerv Syst 2006;22:614-8. 

5 Perilongo G, Gardiman M, Bisaglia L, et al. Spinal low-grade neoplasms 

with estensive leptomeningeal dissemination in children. 

Childs Nerv Syst 2002;18:505-12. 

6 Armao DM, Stone J, Castillo M, et al. Diffuse leptomeningeal oligodendrogliomatosis: 

radiologic/pathologic correlation. Am J Neuroradiol 

2000;21:1122-6. 

7 Bourne TD, Mandell JW, Matsumoto JA, et al. Primary disseminated 

leptomeningeal oligodendroglioma with 1p deletion. J Neurosurg 

2006;105:465-9. 

8 Brandes AA, Tosoni A, Cavallo G, et al. Correlation between O6methylguanine 

DNA methyltransferase promoter methylation status, 

1p and 19q deletions and response to temozolomide in anaplastic 

and recurrent oligodendroglioma: a prospective GICNO study. J Clin 

Oncol 2006;24:4746-53. 

9 Kreiger PA, Okada Y, Simon S, et al. Losses of chromosomes 1p 

and 19q are rare in pediatric oligodendrogliomas. Acta Neuropathol 

2005;109:387-92. 

10 Yamamoto T, Komori T, Shibata N, et al. Multifocal neurocytoma/ 

gangliocytoma with extensive leptomeningeal dissemination in the 

brain and spinal cord. Am J Surg Pathol 1996;20:363-70. 

11 Perry A, Scheithauer BW, Macaulay RJB, et al. Oligodendrogliomas 

with neurocytic differentiation. A report of 4 cases with diagnostic and 

histogenetic implication. J Neuropathol Exp Neurol 2002;61:947-55. 

12 Perry A, Fuller CE, Banerjee R, et al. Ancillary FISH analysis for 1p 

and 19q status: preliminary observations in 287 gliomas and oligodendroglioma 

mimics. Front Biosci 2003;8:a1-9. 

Prognostic factors in meningiomas 

V. Barresi 

Department of Human Pathology, University of Messina, Italy 

Meningiomas account for approximately 24-30% of primary 

intracranial tumors; they occur most commonly in middleaged 

and elderly patients and show a female predominance 1 . 

According to the WHO classification system, meningiomas 

are classified into several histotypes, the most common of 

which are represented by meningothelial, fibrous and transitional 

meningiomas 1 . Moreover, three histological grades 

of increasing malignancy are recognized for these tumours 1 , 

with most of meningiomas falling into grade I and presenting 

as indolent neoplasias 1 . 

The main prognostic questions regarding meningiomas involve 

prediction of recurrence and, for malignant variants, 

prediction of survival. The most important clinical factor 

in recurrence risk is represented by the extent of surgical 

resection, which is influenced by tumor site, extent of invasion 

and by the attachment to vital intracranial structures. 

According to Simpson’s scale 2 , the degree of surgical resection 

is commonly classified into five grades (grade 1: macroscopically 

complete removal, including dura and bone; 

grade 2: macroscopically complete removal, with apparently 

reliable coagulation of dural attachments; grade 3: macroscopic 

complete excision of the solid tumor, but insufficient 

dural coagulation or bone excision; grade 4: partial removal 

of the tumor; grade 5: simple decompression), displaying 

increasing recurrence risk. A major issue relates to the recurrence 

of totally resected (Simpson’s grade 1) meningiomas 

which, in spite of total macroscopic removal including dura 

and bone, display a recurrence rate around 10% 2 . It has been 

hypothesized that the development of recurrences of these 

meningiomas may be related to the presence of microscopic 

clusters of neoplastic cells left in the dura mater or in the

228 

arachnoid membrane 3 4 in relationship to their biological 

activity. 

In the last years a number of studies has been carried out 

in order to evidence histo-prognostic factors able to predict 

the clinical course of meningiomas. At now, the histological 

grade and the proliferation index are considered to be the 

most powerful histological prognosticators for the outcome 

of these neoplasias 5 . Indeed, grade I meningiomas display 

recurrence rates of 7-25%, atypical meningiomas recur in 

29-52% of cases and anaplastic variant at rates of 50-94% 5 . 

Moreover, a mitotic index higher than 4 mitoses/10 HPF has 

been significantly associated with 8-fold higher recurrence 

rates of meningiomas 6 ; similarly, Ki-67 labeling index has 

been shown to significantly correlate with the prognosis of 

these tumors 5 7 . 

The absence of progesterone receptors expression is also regarded 

as a significant prognostic factor for the development 

of recurrences in meningiomas 8 , but only in association with 

high mitotic index and histological grade. High vascularity 

and peri-tumoral vasogenic oedema have been also proposed 

as significant prognostic factors correlated with adverse 

clinical course of meningiomas 5 . In recent years our group 

has evaluated the prognostic role of neo-angiogenesis and 

its regulators on the outcome of meningiomas, showing that 

a high quantity of tumour neo-angiogenesis, reflected by a 

high microvessel density (MVD) appears to be significantly 

associated with a shorter overall survival and with the development 

of recurrences in totally resected neoplasias. In 

addition, according to our findings, a high ratio between the 

concentrations of the pro-angiogenic vascular endothelial 

growth factor (VEGF) and the anti-angiogenic semaphorin3A 

(SEMA3A) in the microenvironment of the tumour behaves 

as a negative predictor of recurrences in meningiomas. We 

may hypothesize that neo-angiogenesis is blocked or stimulated 

depending on the prevalence of VEGF or SEMA3A and 

that microscopic not-removed neoplastic foci may grow and 

give rise to recurrent tumours for their higher capability to 

stimulate proliferation and neo-angiogenesis in relationship to 

VEGF/SEMA3A balance in favour of VEGF. 

Finally, in our Department the prognostic role of proteins involved 

in the regulation of neoplastic growth and progression 

has been also tested in meningiomas. Basing on our results, 

high expression of caveolin-1, a 22 KDa protein which stimulates 

the proliferation of neoplastic cells, or that of matrix 

metalloproteinase-9, an enzyme involved in the invasive potential 

of tumour cells, appear as negative prognostic factors 

for meningiomas. On the other hand, a low expression of the 

CAAT-enhancer binding protein δ (CEBP/δ) seems to be associated 

with lower recurrence risk of these neoplasias. 

references 

1 Perry A, Louis DN, Scheithauer BW, et al. Meningiomas. In: Louis 

DN, Ohgaki H, Wiestler OD, et al (eds). WHO Classification of 

Tumors of the Central Nervous System. Lyon: IARCC press 2007, 

pp. 164-72. 

2 Simpson D. The recurrence of intracranial meningiomas after surgical 

treatment. J Neurol Neurosurg Psychiatry 1957;20:22-39. 

3 Kamitani H, Masuzawa H, Kanazawa I, et al. Recurrence of convexity 

meningiomas: tumour cells in the arachnoid membrane. Surg Neurol 

2001;56:228-35. 

4 Kinjo T, al-Mefty O, Kanaan I. Grade zero removal of supratentorial 

convexity meningiomas. Neurosurgery 1993;33:394-9. 

5 Perry A, Stafford SL, Scheithauer BW, et al. “Malignancy” in meningiomas: 

a clinico-pathological study of 116 patients with grading 

implications. Cancer 1999;85:2046-56. 

6 Perry A, Stafford SL, Scheithauer BW, et al. Meningioma grading: an 

analysis of histological parameters. Am J Surg Pathol 1997;21:1455- 

65. 


7 Perry A, Stafford SL, Scheithauer BW, et al. The prognostic significance 

of MIB-1, p53, and DNA flow cytometry in completely resected 

primary meningiomas. Cancer 1998;82:2262-9. 

8 Perry A, Cai DX, Scheithauer BW, et al. Merlin, DAL-1 and progesterone 

receptor expression in clinico-pathologic subset of meningioma: 

a correlative immunohistochemical study of 175 cases. J Neuropathol 

Exp Neurol 2000;59:872-9. 

Molecular alterations in embryonal tumors 

of central nervous system 

M. Gessi 

Inst. of Neuropathology, University of Bonn Medical Center, Bonn, 

Germany 

Embryonal tumors of the CNS are described as malignant 

small, round cell tumors with possible divergent patterns of 

differentiation. According to the World Health Organization 

classification (2007), this group of tumors includes: medulloblastoma, 

the most common embryonal tumor subtype, 

the central nervous system primitive neuroectodermal tumor 

(CNS-PNET), and the atypical teratoid/rhabdoid tumor 

(ATRT). Although they could share common light microscopy 

features, embryonal tumors appear to evolve by alterations 

of a wide spectrum of genetic pathways. 

The largest part of molecular research on embryonal tumors 

has been focused on medulloblastoma biology, and over the 

years, many aspects of signalling pathways regulating the 

biology of this tumor, have been revealed. The most common 

genetic alteration in medulloblastoma is the loss of chromosome 

17p, often in association with isochromosome 17q: 

i(17)(q10), occurring in 30-50% of cases. Although the precise 

role in medulloblastoma oncogenesis and its prognostic 

significance are not known, the region 17p13.2-13.3 harbors 

many tumor suppressor genes, including TP53. However, 

while sporadic TP53 mutations are uncommon in medulloblastoma, 

germline mutations of TP53 gene, resulting in the 

Li–Fraumeni syndrome, have been related to an increased 

medulloblastoma incidence. 

Numerous investigations have also demonstrated the pivotal 

role of the sonic hedgehog (SHH) signaling pathway in medulloblastoma 

pathogenesis. Molecular alterations in components 

of the SHH pathway (i.e. the inactivating mutations of 

PTCH1 and SUFU and/or activating mutations of SMO), have 

been found in 15-20% of sporadic medulloblastomas. Moreover, 

patients with Gorlin’s syndrome (harboring germline 

alterations in PTCH1 gene) present also an high incidence of 

several tumor types, including medulloblastoma. 

Alterations of the WNT signaling pathway have also been 

implicated to the development of medulloblastoma: approximately 

15 to 20% of sporadic medulloblastoma present 

mutations in genes, including APC, AXIN-1, AXIN-2 or 

CTNNB1-β-catenin, all members of the WNT pathway. 

Genomic amplifications of n-Myc and c-Myc are commonly 

encountered in medulloblastoma and characterize a subset of 

clinically aggressive tumors, frequently with large cell/anaplastic 

histological features. Other molecular pathways, including 

the tyrosin-kinase family receptors Erbb, insulin-like 

growth factor 1 receptor (IGF1R) and PDGFR, have been also 

directly implicated in medulloblastoma pathogenesis. 

Molecular genetic investigations and transcriptional expression 

profile analyses have shown that atypical teratoid/rhabdoid 

tumors (ATRT) are distinct from other embryonal 

tumors. Inactivating deletions or mutations of the tumor suppressor 

gene hSNF5/INI-1, located in the chromosomal region 

22q11.2, encoding a subunit of the SWI/SNF family of chro-

lectures 

matin-remodelling complexes, have been found in more than 

75% of cases. Although its specific tumor suppressor function 

remains still unknown, the alteration of the hSNF5/INI-1 is 

now considered as a crucial step in the pathogenesis of most 

ATRT and today its recognition is a powerful diagnostic tool 

for the diagnosis of these tumors. 

In contrast to ATRTs and medulloblastomas, various molecular 

pathways have been hypothesized to play a role in 

the CNS-PNET pathogenesis but, due to the rarity and the 

heterogeneity of these tumors, only a limited number of studies 

on large series of cases have been made. Alterations affecting 

genes of various molecular pathways (i.e. WNT, Tp53, 

RASFF1A, n-Myc and c-Myc) have been also described in 

CNS-PNET cases. Recently, new findings demonstrating the 

implication of microRNAs (miR-517c and miR-520g) in the 

biology of CNS-PNET and ependymoblastomas have been 

reported. 

In conclusion, many progresses in the molecular characterization 

of medulloblastoma and other embryonal tumors have 

been made. However, these biological data are still the subject 

of intensive translational research in order to define new tools 

for the improvement of patients risk stratification procedures 

as well as their management. 

Surgical pathology of epilepsy 

G. Marucci 

Section of Pathology, Department of Haemathology and Oncological 

Sciences Section of Pathology, Bellaria Hospital, University of 

Bologna, Italy 

Background. Surgical approach has become a useful alternative 

to treat refractory epilepsy, with a postoperative favorable 

outcome that in temporal lobe epilepsy accounts for about 

70% of patients. Cases must be studied with a multidisciplinary 

approach that involves pathologists, neurosurgeons, 

neurologists and neuroradiologists. First of all pathologist 

should be present in operation room to better understand 

the adopted surgical strategy and the correct orientation of 

removed specimens. A large series of histopathological pictures 

may be found in such tissues: hippocampal sclerosis 

(HS), focal cortical dysplasia (FCD), mild cortical dysplasia, 

hamartomas, vascular lesions, low-grade glioneuronal tumors, 

scars or gliotic lesions. Although numerous histochemical and 

immunohistochemical stains have been proposed in literature, 

in everyday practice Nissl, Kluver and anti-NeuN antiserum 

could be considered the most easy and useful tools in histological 

diagnosis. 

Methods. The lesions that typically are encountered in epilepsy 

surgery are represented by HS, FCD and heterotopias. 

Neuropathological classification of HS proposed in 1992 by 

Wyler et al. was based on a semiquantitative evaluation of cell 

loss in the Ammon Horn subfields, resulting in a distinction 

of five grades, although in routine classic and severe Ammon 

Horn Sclerosis are the most frequent reported diagnosis. In 

2007 Blümcke et al. recognized five patterns of HS adopting 

a computerized cluster analysis, and applied the term of MTS 

(mesial temporal sclerosis) 1A to histological pictures similar 

to the classic Ammon Horn Sclerosis and the term of MTS 1B 

to the severe Ammon Horn Sclerosis. Recently it has emerged 

a growing interest in evaluating also the presence of alterations 

in Dentate Gyrus, in particular the so called granular cell 

dispersion. 

Presence of FCD are usually assessed following the scheme 

proposed by Palmini et al., who distinguished type IA (iso- 

229 

lated architectural abnormalities), type IB (architectural abnormalities 

plus giant or immature neurons), type IIA (architectural 

abnormalities with dysmorphic neurons but without 

balloon cells) and type IIB (architectural abnormalities with 

dysmorphic neurons and balloon cells). 

The term of heterotopia is applied to alterations of cortical 

development in which apparently normal brain tissue is mislocated 

in abnormal sites. The most common subtype is represented 

by nodular heterotopia characterized by the presence of 

heterotopic islands of grey matter into the white matter. 

The other lesions found in these patients are not specific of 

epilepsy surgery setting: regarding low-grade tumors a comparison 

between tumors operated with the so called tailored 

resection (characterized by anterior-mesial temporal resection 

along with amygdalohippocampectomy) and with simple lesionectomy 

was performed. 

Finally in some cases a fresh 0,5 cm 3 tissue sample from 

Dentate Gyrus of the hippocampus has been collected immediately 

after removing for tissue culture. 

Results. Between April 2001 and April 2010 110 patients 

(52 males and 58 females) with drug resistant temporal lobe 

epilepsy underwent epilepsy surgery in Bellaria Hospital, 

Bologna. Histological examination has evidenced 15 cases of 

hippocampal sclerosis, 20 cases of focal cortical dysplasia, 

41 cases of hippocampal sclerosis associated to focal cortical 

dysplasia (dual pathology), 27 cases of low grade tumor, 2 

cases of nodular heterotopia, 3 cases of vascular lesions and 

2 cases of encephalocele. A favorable post-surgical epilepsy 

outcome was achieved in 77% of cases: in particular in 61% 

of cases it was obtained a complete disappearance of seizures 

(Engel Class IA). Furthermore it has been demonstrated a better 

seizure outcome for temporo-mesial glioneuronal tumors 

associated with epilepsy in patients who underwent tailored 

resection rather than simple lesionectomy. Finally surgical 

approach makes available hippocampi not only for routine 

histological examination but also for further studies. Adult 

neural stem cells are undifferentiated cells that are present 

in the adult brain and are capable to divide and differentiate 

into astrocytes, oligodendrocytes and neurons. These cells are 

present in the subgranular zone (SGZ) of the Dentate Gyrus of 

the hippocampus and it has been demonstrated the possibility 

to generate neurosphere from the SGZ. 

Inflammatory myopathies 

G Cenacchi 

Dipartimento Clinico di Scienze Radiologiche e Istocitopatologiche, 

“Alma Mater Studiorum” Università di Bologna, Italy 

Background. The inflammatory myopathies (IM) are an 

heterogeneous group of acquired disorders of skeletal muscle 

with undefined etiology and pathogenesis. Inflammatory 

myopathies can be subdivided in two main groups: infectious 

myositis and immunogenic myositis. The autoimmune myopathies 

include polymyositis (PM), dermatomyositis (DM), 

overlap syndromes, and inclusion body myositis (IBM). Recent 

findings have confirmed that PM is an uncommon, but 

frequently misdiagnosed disorder: PM mimics many other 

myopathies and remains a diagnosis of exclusion. Muscle 

biopsy is the gold standard for the diagnosis; the histological 

cornerstone is the identification of cellular infiltrates in muscle 

tissue, however infiltrates are not always present. Induction of 

Major Histocompatibility Complex class I (MHC-I) antigen 

in muscle fibres precedes inflammatory infiltrates, persists 

in chronic phase, and is unaffected by immunosuppressive

230 

therapy so it is considered a good marker of IM. Many Authors 

consider only the sarcolemmal MHC-I staining even if 

evidences that a reticular pattern of internal MHC-I reactivity 

in fibres of myositis patients are reported. 

Methods and Results. We revised 64 adult muscle biopsies 

from a file of the Pathology Department of the Azienda Ospedaliera-Universitaria 

S. Orsola-Malpighi to evaluate the 

diagnostic role of both immunohistochemistry for MHC-I 

stain (samples were scored by two independent and blinded 

investigators and an average of 580 fibres were evaluated 

for each biopsy. The percentage of MHC-I internal labelled 

fibres was determined and interobserver reproducibility was 

evaluated) and transmission electron microscopy. The positive 

muscle fibres displayed MHC-I staining of the cytoplasm 

rather than of the sarcolemma. Positive fibres were observed 

in all samples (21 IM cases and 43 controls). Interobserver reproducibility 

was moderate (K = 0,568). The specificity of the 

test was of 100% when the percentage of the internal labelled 

fibres was higher than 50%, as mean of the two observers. Ultrastructural 

studies are necessary in many cases, especially in 

IBM, by screening other myopathies with inflammation, such 

as dystrophies, toxic and metabolic myopathies. IBM or dystrophies, 

especially without a positive family history, can be 

Colangite sclerosante e pancreatite 

autoimmune 

L. Terracciano 

Department of Patology, University Hospital, Basel, Switzerland 

Case history. A 51 year-old man was referred to University 

Hospiatl Basel, Switzerland,with abdominal pain, jaundice 

weight loss and diarrhea as presenting symptoms. Laboratory 

examinations showed an increase of transaminases and cholestatic 

parameters, A liver biopsy was performed and a diagnosis 

of sclerosing cholangitis was rendered. Two months later 

because of persisting abdominal pain and diffuse enlargement 

of the pancreas on imaging, pancreas carcinoma was suspected. 

The patient underwent a duodeno-pancreatic resection. 

Histology was consistent with autoimmune pancreatitis. 

4 months later a further liver biopsy showed the full-blown 

picture of autoimmnue sclerosing cholangitis. 

Primary sclerosing cholangitis is a cholestatic disease characterized 

by patchy inflammation, fibrosis, and stricturing of 

the intrahepatic and/or extrahepatic bile ducts.1 The diagnosis 

of primary sclerosing cholangitis is based on characteristic 

cholangio-graphic findings, in combination with clinical, biochemical, 

and histological features. 

The disease lacks a definitive etiological factor, although a 

strong association with inflammatory bowel disease is well 

recognized. 

Autoimmune pancreatitis (AIP) is a recently recognized clinicopathological 

entity, which was first described by Sarles in 

1961 as a “chronic inflammatory sclerosis of the pancreas” 

of possible autoimmune pathogenesis associated with hypergamma-globulinemia. 

The disease has been gaining new 

attention for the last two decades, and the term “autoimmune 

pancreatitis”, coined by Yoshida in 1995, has only recently 


Slide seminar: epatic pathology 

Moderators: G. Faa (Cagliari), L. Terracciano (Basilea) 

diagnosed as PM and can therefore be treated unsuccessfully 

and unnecessarily for many years, thereby exposing patients 

to the long-term side-effects of prednisolone and immunosuppressant. 

In sporadic IBM, in addition to autoimmune 

inflammation, there are degenerative features characterized 

by vacuolization and accumulation of stressor and amyloid-related 

molecules. The diagnosis of IBM rests on morphological 

criteria including inflammatory infiltrates with mononuclear 

cell invasion of non-necrotic muscle fibers, rimmed vacuoles 

and either intracellular amyloid or inclusions consisting of 15- 

18 nm filaments at the ultrastructural level. In PM, multifocal 

lymphocytic infiltrations invade healthy muscle fibers: in addition 

to primary inflammation there are vacuolated muscle 

fibers containing lamellar membranous residues and amorphous 

electron dense material. When the intramuscular blood 

vessels show endothelial hyperplasia with tubuloreticular 

profiles, fibrin thrombi and obliteration of capillary lumina, 

the diagnosis may be DM. 

Conclusions. This review outlines the fundamentally different 

pathology between different IM as evolved the past few 

years, provides a critical analysis of the diagnostic markers, 

and summarizes the most significant developments on their 

pathogenesis as relate to therapeutic strategies. 

been widely accepted in the scientific literature. Due to the 

possible involvement of the biliary tract, the term autoimmune 

pancreatocholangitis (AIPC) has been introduced. The main 

reasons for the rising interest in investigating AIPC reside in 

its increasing frequency, partly due to an increased awareness 

of the disease but also due to a potentially increased incidence 

in the last 20-30 years, its not yet clarified aetiology and 

pathogenesis and ist still undefined clinical spectrum. The 

coexistence of AIPC with other autoimmune-related diseases, 

such as Sjo¨gren’s syndrome, inflammatory bowel diseases 

(IBD) 

and rheumathoid arthritis, the presence of immunologic abnormalities 

in subsets of patients (hypergammaglobulinemia, 

elevated serum IgG4 levels, presence of auto-antibodies), 

and the association with a specific HLA-haplotype in the 

Japanese population, represent the main pieces of evidence 

of an autoimmune pathogenesis of the disease. All lesions 

incorporated into the spectrum of the disease, including the 

pancreatic manifestations are characterized by a plasma cellrich, 

often mass-forming inflammatory process with numerous 

IgG4-positive plasma cells. Altough very similar to primary 

sclerosding cholangitis, IgG4 sclerosing cholangitis not 

rarely show peculiar histological features. As in the pancreas, 

biliary involvement by IgG4-related autoimmune disease can 

be diffuse or localized, producing either a generalized but irregular 

thickening or a tumefactive lesion. The histological 

appearance is similar in both situations: lymphoplasmacytic 

inflammation, fibrosis (often with a swirling or storiform arrangement) 

and obliterative phlebitis. Despite the dense periluminal 

inflammation, the biliary epithelium is usually intact. 

This is in distinct contrast to PSC, which often produces mucosal 

erosion. In another contrast with PSC, the inflammatory 

process is often more dense at the periphery of the duct. This

lectures 

phenomenon is partly due to dense inflammation in the walls 

of periductal vein branches, but lymphoplasmacytic inflammation 

around nerve twigs and forming nodular infiltrates in 

periductal soft tissue are also characteristic features of IAC. 

While lymphocytes and plasma cells predominate, eosinophils 

can be numerous and are occasionally numerically dominant. 

Neutrophils, commonly seen in PSC, are not a feature of 

IgG4 cholangitis. Immunohistochemical staining for IgG4 is 

a useful tool for confirming the diagnosis of IgG4 cholangitis. 

Accumulating evidence suggests that the bile duct lesions and 

the concomitant pancreatitis in patients with IgG4 cholangitis 

improve with corticosteroid treatment which distinguishes 

IgG4 cholangitis from PSC. 

Post liver transplant complications (PlTC): 

recurrence of hepatitis (rH) or cellular rejection 

(Cr)? 

E. David 

Anatomia Patologica, II Az. Osp. Molinette Torino 

PLTC are constituted by a various group of diseases that are 

crucial for the clinical management of patients in liver transplantation 

(LT). 

Liver biopsy (LB) is the gold standard for diagnosis of rejection. 

The search of clinico-pathological correlations, advantaging 

of the sequential evaluation of follow-up biopsies, 

represent the most valuable working method. 

Frequently, different pathologic processes are present on the 

same LB. As a matter of fact, most chronic liver diseases can 

recur in the graft, and LBs may display features that require 

differential diagnosis between (acute or chronic) CR and de 

novo conditions such as de novo autoimmune hepatitis, drug 

toxicity and vascular lesions. 

We recommend the slides to be initially examined by the 

pathologist blindly to clinical data, a diagnosis or possible 

differential diagnoses being formulated, then histology to be 

compared with the available clinical data and the final diagnosis 

to be discussed with the physicians. Pathologists must be 

familiar with atypical presentation of PLTC and aware to recognize 

the primary process that has to be primarily treated. 

PLTC are traditionally distinguished as early and late events, 

but both RH and CR can occur early. 

We propose to distinguish three broad categories of PLTC: 

RH, CR and de novo diseases. 

Rejection can be distinguished in early acute CR, late CR 

with“atypical” features, chronic rejection and antibody-mediated 

rejection. Acute CR is the commonest cause of early graft 

dysfunction, its incidence ranging from 24 to 80%. A RAI 

(Rejection Activity Index) may be used, with an histological 

scoring system from 0 to 9. 

HCV related- cirrhosis represents a very common indication 

for LT and unfortunately the recurrence of HCV infection is 

universal and immediate. HCV RH occurs in up to 90% of 

patients at 5 years from transplantation, nevertheless some 

patients will present an indolent course, whereas others will 

rapidly progress to cirrhosis. Compared to non-transplanted 

HCV patients who develop cirrhosis at a rate of less than 5% 

over 5 years, the course of post-transplant recurrent HCV is 

accelerated with up to 20- 40% progressing to cirrhosis within 

5 years. Factors influencing the prevalence and severity of 

disease recurrence include: the virus genotype,the host immunogenetic 

background and the immunosoppressive treatment. 

231 

Both hepatitis B and D relapse, but prophylactic measures 

have significantly decreased their recurrence. 

Differential diagnosis between acute CR and RH is crucial on 

LB because immunosuppresive treatment is associated with 

an increased risk of allograft cirrhosis and mortality. 

Histologically, acute CR is characterized by a various combination 

of features of predominantly mononuclear (including 

blastic or activated lymphocytes, neutrophils and eosinophil) 

portal inflammation, of subendothelial inflammation in portal 

and/or terminal hepatic veins, and of bile duct inflammation 

and damage. 

Lobular necroinflammatory activity and interfacie inflammation 

with ductular reaction is usually more prominent in RH 

than in CR. But 

CR and RH may coexist, and in such cases it is mandatory to 

identify the predominant process to be treated. 

In protocol LB, (hepatitis-like) necroinflammatory lesions 

occur in 40% of adult LT recipients after 12 months from 

transplantion and in 60% of pediatric patients after 10 years, 

whit normal serological tests. The possible causes of this 

idiopathic post transplantation hepatitis include: atypical 

rejection, de novo autoimmune hepatitis and infection from 

unknown agents. A significant percentage of these patients 

may show progression to cirrhosis without significant liver 

test abnormalites. 

In conclusion, LB may represent a diagnostic challenge for 

the pathologists, nevertheless a systematic approach to morphological 

analysis of liver lesions can satisfactorily identify 

or give the clue for diagnosis of clinical syndromes such as 

immuno reactions (rejection), hepatitis, cholestasis, drug toxicity, 

and for prognostic staging of evolutive PLTC. 

A focal liver lesion in a young body-builder 

M. Roncalli, L. Di Tommaso, A. Destro * , E. David ** , 

L. Terracciano *** 

Department of Pathology University of Milan School of Medicine & 

IRCCS Humanitas Clinical Institute, Rozzano, Milan, Italy; * Molecular 

Genetic Laboratory, IRCCS Humanitas Clinical Institute, Rozzano, 

Milan, Italy; ** Anatomia Patologica II° Azienda Ospedaliera 

Molinette, Torino, Italy; *** Institute of Pathology, University Hospital 

Basel, Basel, Switzerland 

Clinical history. A 35 years old asymptomatic man with a 

history of anabolic steroid intake (body builder) underwent 

a surgical resection of a 6.5 cm. focal liver lesion located in 

II-III liver segments. The lesion was incidentally discovered 

after a routine US of the liver. Grossly the lesion appeared as a 

greenish, unencapsulated nodule of 6.5 cm, with well-defined 

margins and located in the context of an otherwise unremarkable 

parenchyma. The nodule margins were close to those of 

the surgical resection. Microscopical examination revealed a 

well differentiated hepatocellular proliferation composed by 

hepatocytes with focally increased N/C ratio and organized in 

trabecular and small acinar structures. 

The main diagnostic issue was hepatocellular adenoma vs 

well differentiated hepatocellular carcinoma. A number of 

histochemical, immunocytochemical and molecular studies 

were carried out to address the diagnostic issue. 

The patient is alive and well 6 years after the original diagnosis. 

The discussion will focus on the differential diagnosis and on 

the possible pathogenetic links between liver cell adenoma 

and carcinoma.

232 

Molecular diagnosis in colorectal cancer 

A. Scarpa 

Department of Pathology and ARC-Net Research Center, Verona University 

Hospital, Verona, Italy 

Colorectal cancer is a disease whose moleclar basis are clearer 

than those of many other cancers. Besides a precise histologic 

diagnosis and pathological staging, the pathologist can provide 

a molecular characterization of a colorectal neoplasia, 

thus permitting to 1) unveil the existence of a hederitary 

syndrome, 2) help assessing prognosis, 3) predict response 

to therapy. 

Hereditary cancer syndromes account for 1-5% of all colorectal 

neoplasms. The main forms are the familial adenomatous 

polyposis (FAP) in its classic variant, due to mutations in the 

APC gene, and its attenuated form with biallelic mutations of 

MYH gene (MAP) and the nonpoliposyc forms as HNPCC 

due to mutations in mismatch repair genes (MLH1, MSH2, 

MSH6, PMS2). The cost/benefit of sequencing of entire genes 

is too high to permit the analysis of patients with colorectal 

cancer. One of the main objectives is therefore to stratify 

colorectal cancer patients according to different levels of 

genetic risk using a stepwise procedure. The first approach to 

help identifying hereditary syndromes is clinical and resides 

in the study of the phenotype and the genealogic tree that is of 

great help for FAP and MAP. For HNPCC a major help also 

comes from the analysis of the tumor samples for the presence 

of the molecular phenomenon called microsatellite instablity 

(characterizing the vast majority of HNPCC cancers) and/or 

for the immunohistochemical expression of mismatch repair 

proteins. The lack of expression of one of the proteins suggests 

the presence of a somatic homozygous mutation in cancer 

cells and indicates which gene is to be sequencedd to find 

a germline mutation. Finding a germline mutation defines the 

the follow-up for the patient and the surveillance programme 

for the family. 

Microsatellite instabilty is also observed in 10-15% of sporadic 

colorectal cancers and is due to the somatic inactivation of 

a mismatch repair gene, more frequently MLH1. Patients with 

sporadic colorectal cancers of this molecular phenotype show 

a longer survival especialy in Stages II and III. Wheter or not 

these patients benefit from adjuvant therapies remains to be 

determined. An additional prognostic indiocator in colorectal 

cancer is the presence of P53 mutations that seems to be associated 

with a worse prognosis, higher risk of metastasis and 

resistance to chemio and radiotherapy. 

The recent development of targetted drugs specifically inhibiting 

the receptor of the epidermal growth factor (EGFR) as 

cetuximab or panitumab is giving new hopes for the treatment 

of metastatic colorectal cancer (mCRC) resistent to standard 

chemotherapy. Various studies have unequivocally shown 

that KRAS mutational status is able to predict response to 

anti-EGFR therapy in patients with mCRC. Recently the 

American Society of Clinical Oncology (ASCO) and the 

Agenzia Italiana del Farmaco (AIFA) have suggested that 


Wednesday, September 22 nd , 2010 

Molecular diagnosis of solid tumours. 

A practical approach for organ pathologies 

all patients with mCRC who are candidates for anti-EGFR 

therapy must have the KRAS status assessed and in the case 

a mutation in codons 12 and 13 should not be subjected to 

therapy. The sequencing analysis of DNA prepared from 

paraffin embedded tissues is highly efficient pending a cancer 

cell enrichment to more than 60%. In our experience on the 

latest 375 patients with mCRC, we observed that 163 (43.5%) 

had a KRAS mutation (162 in codons 12 and 13, 1 in codon 

22), 5 (1.5%) were not PCR amplifiable and 207 (55%) had a 

wild type KRAS sequence. 

Analysis of genes frequently mutated in CRC is also effective 

in selecting candidate patients for treatment with anti- 

EGFR drugs cetuximab and panitumumab. Approximately 

40% of CRC patients harbour a K-Ras mutation conferring 

resistance to anti-EGFR drugs. Of the remaining 60% with 

a wild type K-Ras tumor, 5-10% carry the B-Raf activating 

mutation V600E, which also negates response to these agents 

and dictates a very poor prognosis. This prompted the search 

and discovery of a novel selective B-Raf inhibitor targeting 

tumors with V600E mutation. An additional 20% of K-Ras wt 

CRC patients are resistant to anti-EGFR drugs, due to activating 

mutations in exons 9 and 20 of PIK3CA, thus providing 

the rationale to test novel agents targeting PI3K/Akt pathway. 

More recently, a signature of 6 genes among a 57 gene set 

was associated with response to cetuximab among Ki-Ras wt 

CRC patients. 

In the same fashion, in gastric cancer the identification of 

genetic lesions such as PIK3CA mutations or HER2 amplifications, 

reported in 15% of tumors, may allow treatment with 

targeted agents not otherwise indicated for this disease. 

In pancreatic cancer, a recent global genomic analysis revealed 

12 cell signalling pathways altered in 67-100% of 

tumors, including among the others, genes such as Hedgehog, 

TGFß and Wnt/Notch, for which novel targeted agents are 

now available. In addition, the identification of BRCA2 mutations, 

which hamper DNA repair efficiency, provides the 

opportunity of a synthetic lethality therapeutic approach by 

combining PARP inhibitors with DNA-damaging agents such 

as platinum derivatives. 

The large body of information emerging from cancer gene/ 

protein expression profiles is making a major contribution in 

the clinically efficient sub-classification of cancers. This will 

further help in the selection of patients, through the use of 

reliable biomarkers, who may benefit from specific targeted 

agents or chemotherapeutics. 

In several types of GI cancers, particularly colorectal cancer 

(CRC), analysis of a limited set of genes currently allows 

more tailored treatment. Along with the formerly reported Oncotype 

DX colon, a novel 38 gene signature named Coloprint, 

can predict prognosis in stage II and III CRC patients, identifying 

those more likely to benefit from adjuvant treatment. 

Finally, genotyping profiles are providing useful information, 

to more specifically predict activity and toxicity of several 

previously available chemotherapeutics currently used in GI 

cancer.

lectures 

Deficit of DNA mismatch repair: 

diagnostic algorithm and clinical implications 

G. Lanza, I. Maestri, L. Ulazzi, R. Gafà 

Sezione di Anatomia Patologica, Università di Ferrara, Ferrara, Italia 

Thursday, September 23 rd , 2010 

Nearly 15% of colorectal carcinomas (CRCs) display microsatellite 

instability (MSI or MSI-H, high frequency MSI) 

caused by impairment of the DNA Mismatch Repair (MMR) 

system. The distinction between MMR-proficient (MMRp) 

and MMR-deficient (MMRd) tumors represents a fundamental 

step in the molecular classification of CRC with important 

clinical implications. 

Most MSI-H CRCs (70%) are sporadic and in these tumors 

inactivation of the MMR system is determined by somatic promoter 

methylation of the MLH1 gene. The remaining MSI-H 

CRCs are hereditary (Lynch syndrome) and produced by germline 

mutations of a MMR gene (MLH1, MSH2, MSH6, and 

more rarely PMS2) with somatic inactivation of the second wild 

type allele. It has been consistently demonstrated that inactivation 

of the MMR genes is associated with immunohistochemical 

loss of expression of the corresponding protein. In addition, 

as MMR proteins work as heterodimers, abnormalities of the 

obligatory partners (MSH2 and MLH1) will result in degradation 

of their dimers and concurrent loss of expression of both 

the obligatory and secondary partner proteins (MSH2/MSH6 

and MLH1/PMS2). Conversely, abnormalities in genes of the 

secondary partner proteins (MSH6 and PMS2) will determine 

selective loss of MSH6 and PMS2 expression, respectively, as 

their function is compensated by other proteins. Therefore, immunohistochemical 

analysis of MMR proteins espression represents 

a rapid and reliable test for the identification of MMRd 

colorectal tumors, also indicating the gene that is most likely 

inactivated. Many studies demonstrated an excellent correlation 

of the results obtained by immunohistochemistry and MSI 

analysis. Only a small fraction of hereditary cases with missense 

mutations (generally of MLH1) leading to nonfunctional 

proteins with maintained antigenicity might result MSI-H with 

normal expression of the MMR proteins. 

Colon neoplasms 

233 

Lynch syndrome accounts for 2-3% of all CRCs. MSI testing 

and immunohistochemical analysis of MMR proteins 

expression are worldwide employed for the identification of 

colorectal cancer patients with presumptive Lynch syndrome, 

to be tested for MMR genes germline mutations. It is recommended 

that MSI or MMR protein expression analyses should 

be carried out on tumors from patients clinically at high risk or 

selected on the basis of the revised Bethesda guidelines. However, 

molecular screening investigations performed on large 

series of unselected surgically removed colorectal cancers 

indicated that a large fraction of Lynch syndrome cases should 

be unrecognized using common criteria of selection. These 

data suggest that screening of all CRCs for MSI or abnormal 

MMR protein expression should be a more effective approach 

for the identification of hereditary cases. 

Recent studies showed that sporadic MSI-H MLH1-negative 

tumors frequently harbour BRAF V600E gene mutations. 

Conversely, BRAF mutations have not been detected in 

MSI-H MLH1-negative tumors from patients with Lynch syndrome. 

Also in our experience BRAF gene mutation analysis 

could be employed as an aid for discriminating hereditary 

from sporadic MLH1-negative MSI-H carcinomas. 

MMR status has been clearly demonstrated to be an independent 

prognostic indicator in colorectal cancer. Patients with 

stage II and III MSI carcinomas display higher survival rates 

with respect to patients with non-MSI tumors. In addition, 

emerging data suggest that patients with MSI tumors don’t 

have significant benefit from adjuvant 5-fluorouracil-based 

chemotherapy. Although the use of MMR status assessment 

as a prognostic and predictive test has not yet been validated 

and incorporated into clinical practice, it is advisable to perform 

this analysis in stage II colon cancer patients. Owing to 

the favourable outcome and lack of benefit of current standard 

treatment, patients with stage II MSI-H colon cancers should 

not receive adjuvant chemotherapy. 

In conclusion, accumulated evidence indicates that MMR 

status evaluation is of great importance in the management 

of CRC patients. Pathologists have an essential role in MMR 

status testing.

234 

Grey zones of hodgkin lymphoma: report of 

a case with features intermediate between 

primary mediastinal B-cell lymphoma, classical 

hodgkin lymphoma and nodular lymphocytepredominant 

Hodgkin lymphoma 

A. Zamò 1 , G. Todeschini 2 , R. Zanotti 2 , F. Benedetti 2 , F. 

Menestrina 1 

1 Department of Pathology and Diagnostics, University of Verona; 

2 Department of Medicine, University of Verona 

Background. Hodgkin lymphoma (HL) was one of the first 

lymphomas to be defined as an entity on morphological and 

clinical grounds, and diagnostic criteria seem straightforward 1 . 

Yet, accurate morphological evaluation coupled to the use of 

extensive immunohistochemical panels have highlighted the 

presence of several “grey zones” (GZ). A GZ can be defined 

in several ways: as a morphological overlap, as a composite 

morphology with or without a transition area, as an aberrant 

immunophenotype, or as a mixture of these conditions. 

In brief, GZ of HL include only one WHO-defined entity, 

called “B-cell lymphoma, unclassifiable, with features intermediate 

between diffuse large B-cell lymphoma and classical 

Hodgkin lymphoma” 2 , and other non-WHO-defined GZ, 

namely between T-cell rich diffuse large B-cell lymphoma 

and nodular lymphocyte predominant HL, between classical 

HL and anaplastic large cell lymphoma and also other composite 

HL and non-Hodgkin lymphoma, not included in the 

previous categories. 

We report a case showing features intermediate between 

primary mediastinal B-cell lymphoma (PMBL), classical HL 

(cHL) and nodular lymphocyte-predominant Hodgkin lymphoma 

(N-LPHL). 

Case description. A 37-year old male presented in another 

hospital in January 2009 with dyspepsia, night fever and 

sweats. In February the left arm became swollen, and a chest 

X-ray was taken, showing a mediastinal enlargement. CT 

scan confirmed the presence of a 90 x 60 mm mass as well as 

multiple lymphadenopathies, including subcarinal, supraclavicular 

and axillary (bilateral). The patient was classified as 

stage IIB bulky mediastinal. 

Laboratory analyses showed the follwing values: Hemoglobin 

14.1 g/dl, Platelets 290x10 9 /L, Leukocytes 6.8x10 9 /L, Neutrophils 

5.4x 10 9 /L, Lymphocytes 0.53x10 9 /L, ESR 36 mm, 

normal beta2-microglobulin and LDH values. 

A first lymph node biopsy was taken, and the patient was 

diagnosed with nodular sclerosis cHL. 

After two cycles of ABVD chemotherapy, PET-scan showed 

an increased SUV in paratracheal region, and persistence of 

supraclavicular lymphadenopathies. CT scan also showed a 

decrease of the mediastinal mass (38 x 18 mm). A second 

biopsy (consisting of two supraclavicular lymph node fragments) 

was taken and sent to our Department. 

Histopathological examination showed two different pictures 

in the two fragments. 

In one of the fragments, the lymph node structure was partly 

preserved, with several reactive follicles present. Focally large 


friday, September 24 th , 2010 

Slide seminar: lymphoma surgical pathology 

Moderators: V. Franco (Palermo), S. Pileri (Bologna) 

atypical cell were present, showing a morphology reminiscent 

sometimes of LH cells, sometimes of HRS cells. These cells 

were mostly positive for CD20, PAX5, BOB1, OCT2, p63, 

MUM1/IRF4, partially for CD79a and CD30; BCL6 was expressed 

only focally and EBER was negative. The microenviroment 

was suggestive of N-LPHL, including B-cell nodules 

composed mostly by small cells, that embedded the large 

cells, although these were found also outside the nodules. 

The second fragment showed a diffuse infiltration of lymphoid 

cells, showing marked polymorphism, where the dominant 

cells were medium to large-sized, frequently with a clear 

cytoplasm; often neoplastic cells showed a “pop-corn” or 

more rarely “sternbergoid” appearance. Compartmentalizing 

sclerosis was focally present. Neoplastic cells were diffusely 

positive for CD20, PAX5, BOB1 and OCT2, variably positive 

for CD79a, CD30, MDC, MUM1/IRF4, BCL6, p63, focally 

positive for CD23, negative for EBER. Ki-67 marked around 

50% of cells. 

The final diagnosis was “B-cell lymphoma, unclassifiable, 

with features intermediate between diffuse large B-cell lymphoma 

and classical Hodgkin lymphoma” (grey zone lymphoma) 

with a comment explaining the peculiarity of the case. 

The patient underwent one R-CHOP cycle, followed by one 

R-DHAP, and then sequential high-dose therapy associated 

with four infusion of Rituximab and stem cell reinfusion after 

high dose of Cytarabine and after Mitoxantrone and high dose 

of Melphalan, (completed on the 29 th of January this year). 

After therapy, CT scan showed a further reduction of the 

mediastinal mass (4mm), while PET-scan was completely 

negative. At the last follow-up (19 th of July 2010) the patient 

was in complete remission. 

Discussion. B-cell lymphoma, unclassifiable, with features 

intermediate between diffuse large B-cell lymphoma and classical 

Hodgkin lymphoma has been recognized as an entity in 

the 2008 WHO classification 2 . However, the introduction of 

this category has stirred some discussion, mostly concerning 

the acceptance by clinicians, who might face a problem in 

deciding the most appropriate therapeutic regimen. 

This category is heterogeneous by definition, comprising a 

mixture of features (both morphological and immunophenotypical) 

of PMBL and HL. Adding even more complexity, 

composite PMBL and cHL lymphoma, are also mentioned 

in a very short paragraph of the WHO blue book, and were 

included in the “B-cell lymphoma, unclassifiable, with features 

intermediate between diffuse large B-cell lymphoma and 

classical Hodgkin lymphoma category” by EAHP panellists 

at the last EAHP lymphoma workshop (Bordeaux 2008). This 

approach is also reflected in the ICD-O code chosen for this 

entity, that is 9596/3, corresponding to “composite Hodgkin 

and non-Hodgkin lymphoma”. 

The clinical presentation of this lymphoma is usually similar 

to PMBL, although a male prevalence has been reported 3 4 . 

The most common morphology is similar to PMBL, but with 

greater cellular heterogeneity, including many HRS-like cells, 

sometimes with the presence of areas that closely resemble 

cHL.

lectures 

The immunophenotype is positive for B-cell markers (CD20, 

CD79a, PAX5), even in HRS-like cells (whereas in cHL 

CD20 is usually weak or negative, and CD79a usually negative) 

although some aberrant markers may be present, like 

CD15 or diffuse CD30 expression (focal CD30 expression is 

very common in PMBL). EBER is usually negative. 

Recently Hoeller et al. have published a work trying to spot 

the most significant immunophenotypic differences between 

PMBL and HL 5 . The authors proposed a diagnostic algorithm 

based on BOB1, CD79a and cyclin E. They also confirmed 

p63 (TP73L) as a useful and highly reproducible marker of 

PMBL as previously reported by our group 6 . 

To our knowledge the case we describe is the first mediastinal 

lymphoma with features intermediate between PMBL, cHL 

and N-LPHL. These intermediate features were mostly evident 

comparing the two different fragments, one showing intermediate 

features between cHL and N-LPHL, and one between 

cHL and PMBL. Intermediate features were both morphological 

and immunophenotypical. Classical HL features included 

a large nucleolus in many large cells, partial CD30 expression 

and very strong MDC expression. N-LPHL features included 

several cells resembling LH cells, diffuse positivity for CD20, 

partial positivity for CD79a as well as p63, BOB1, OCT2 and 

BCL6 expression, and a PTGC-like microenvironment in one 

of the fragments. PMBL features included the presence in one 

of the fragments of clusters of medium to large cells with clear 

cytoplasm, showing CD20, CD79a, p63, BOB1, OCT2 and 

BCL6 expression, as well as partial CD23 expression. 

Our personal interpretation is that the first fragment might 

represent an initial lesion of PMBL. Should only one biopsy 

have been taken, the patient might have been misdiagnosed. 

Several cases of synchronous or metachronous PMBL and HL 

have been reported; more commonly relapses show features of 

PMBL, raising the suspicion that this component might have 

been present ab initio and relapsed because of inadequate 

therapy. 

235 

Conclusion. The experience gained from this and other similar 

cases supports two conclusions: 

a) in case of a mediastinal mass, the largest possible biopsy 

should be taken; needle biopsies should be completely 

avoided; 

b) more studies are needed to understand the nature of GZ 

lymphomas, but the hypothesis that most grey zone cases 

should be considered inside the morphologic and phenotypic 

spectrum of PMBL seems sound (also for therapeutic 

purposes). 

references 

1 Stein H. Hodgkin lymphoma. In: Swerdlow SH, Campo E, Harris NL, 

Jaffe ES, Pileri SA, Stein H, et al., eds. WHO Classification of Tumours 

of Haematopoietic an Lymphoid Tissues. Lyon, France: IARC 

2008, pp. 322. [Bosman FT, Jaffe ES, Lakhani SR, Ohgaki H (Series 

Editor): World Health Organization Classification of Tumours]. 

2 Jaffe ES, Stein H, Swerdlow SH, Campo E, Pileri SA, Harris NL. 

B-cell lymphoma, unclassifiable, with features intermediate between 

diffuse large B-cell lymphoma and classical Hodgkin lymphoma. In: 

Swerdlow SH, Campo E, Harris NL, Jaffe ES, Pileri SA, Stein H, 

et al., eds. WHO Classification of Tumours of Haematopoietic an 

Lymphoid Tissues. Lyon, France: IARC 2008, pp. 267-268. [Bosman 

FT, Jaffe ES, Lakhani SR, Ohgaki H (Series Editor): World Health 

Organization Classification of Tumours]. 

3 Garcia JF, Mollejo M, Fraga M, Forteza J, Muniesa JA, Perez-Guillermo 

M, et al. Large B-cell lymphoma with Hodgkin’s features. Histopathology 

2005;47:101-10. 

4 Traverse-Glehen A, Pittaluga S, Gaulard P, Sorbara L, Alonso MA, 

Raffeld M, et al. Mediastinal gray zone lymphoma: the missing link 

between classic Hodgkin’s lymphoma and mediastinal large B-cell 

lymphoma. Am J Surg Pathol 2005;29:1411-21. 

5 Hoeller S, Zihler D, Zlobec I, Obermann EC, Pileri SA, Dirnhofer S, 

et al. BOB.1, CD79a and cyclin E are the most appropriate markers to 

discriminate classical Hodgkin’s lymphoma from primary mediastinal 

large B-cell lymphoma. Histopathology 2010:56:217-28. 

6 Zamo A, Malpeli G, Scarpa A, Doglioni C, Chilosi M, Menestrina F. 

Expression of TP73L is a helpful diagnostic marker of primary mediastinal 

large B-cell lymphomas. Mod Pathol 2005;18:1448-53.

Pathologica 2010;102:237-383 OrAl COMMuNICATIONS AND POSTerS 

P16 INK4a is a useful marker in uterine 

adenocarcinoma classification 

M.A. Caponio, T. Addati, S. Petroni, O. Popescu, G. Giannone, 

R. Di Girolamo, V. Rubini, A. Kardashi, G. Simone 

Pathological Anatomy Unit, NCI “Giovanni Paolo II”, Bari, Italy 

Background. Endocervical adenocarcinomas (ADCs) are increasing 

each year. Determining primary site of uterine ADC 

can be problematic due to the overlapping morphology of endocervical 

and endometrial ADCs. The same problem can regard 

metastatic ADCs of extra-uterine origin. P16 INK4a is a molecular 

biomarker potentially useful in discriminating endocervical ADC, 

diffuse positive (P), from endometrial negative (N) or focally 

positive (FP) and metastatic ADCs of extra-uterine origin and 

from reactive glandular cells. The aim of this study was to investigate 

p16 expression in endocervical, endometrial and metastatic 

ADCs of extra-uterine origin. 

Methods. We observed 43 cervical biopsies of uterine ADC. 

P16 INK4a (CINtec p16 Histology Kit) has been investigated in all 

histological samples. 

Results. On 43 cervical biopsies the following diagnosis were 

performed: 5 endocervical ADCs, 8 endometrioid-type endometrial 

ADCs, 5 endometrial serous-papillary ADCs, 12 extra-uterine 

ADCs, 11 NOS ADCs and 2 endocervical biopsies whitout 

atypia. Three out of 5 endocervical ADCs were p16 P, 1 FP and 

1 case was p16 N. One endometrioid ADCs resulted p16 P, 4 

FP and 3 N. Three endometrial serous-papillary ADC have been 

classified p16 P and 2 FP. On NOS ADCs, 3 resulted p16 P, 5 FP 

and 3 N. Of the 7 ADCs of ovarian origin, 4 resulted P and 3 N. 

In 2 out four cases from large intestine FP was detected, whereas 

2 resulted N. One case of breast origin was N such as 2 endocervical 

samples without atypia. 

P16 positive was prevalent in endocervical and serous papillary 

ADCs of endometrial or ovarian origin, whereas endometrioid 

ADCs, such as metastatic non ovarian lesions generally presented 

only focal or negative immunostaining. Some bias in diagnostic 

use of p16, leading to disagreement between bioptic and surgical 

sample could be due to sampling problems and neoplastic heterogeneity. 

P16 seems to be an useful marker in ADCs particularly 

in reclassifying NOS ADCs. 

Predictors of recurrence or progression in 

pituitary adenomas differ according to tumour 

subtypes: a classification-tree approach 

1,2)A. Righi, 3)P. Agati, 4)G. Frank, 5)M. Faustini-fustini, 6)R. 

Agati, 4)D. Mazzatenta, 1)A.Farnedi, 6)F. Menetti, 1)G. Marucci, 

1)M.P. Foschini 

1)Anatomia patologica, Dipartimento ematologia-oncologia, Università di 

Bologna, Ospedale Bellaria, Bologna, Italia; 2)Scienze biomediche e oncologia 

umana, Molinette, Torino, Italia; 3)Dipartimento di statistica “P. Fortunati”, 

Università di Bologna, Bologna, Italia; 4)Centro di chirurgia dei tumori 

ipofisari, Ospedale Bellaria, Bologna, Italia; 5)Unità di endocrinologia, Dipartimento 

di medicina, Ospedale Bellaria, Bologna,Italia; 6)Dipartimento 

di Neuroradiologia, Ospedale Bellaria, Bologna, Italia 

Background. It is difficult to evaluate the recurrence and progression 

potential of pituitary adenomas (PA) at presentation. 

The World Health Organization Classification of Endocrine 

Tumors suggests that invasion of the surrounding structures, 

size at presentation, Ki67 labelling index (LI) higher than 

3%, and extensive p53 expression are indicators of aggressive 

behaviour. Nevertheless, Ki67 and p53 LIs evaluation is 

subject to inter-observer variability and their cut-off value is 

controversial. 

Methods. Aim of the present study is to analyse the prognostic 

value of age, invasion, size, Ki67, and p53 protein LIs (evaluated 

using a digital image analysis) in a series of 166 pituitary adenomas 

with a minimum follow-up of 6 years using the receiver 

operator characteristic (ROC) curve and the classification and 

regression tree analysis (CART). 

Results. In the un-stratified dataset, the commonly used threshold 

index of 3% has a high specificity (93.2%) but a very low sensitivity 

(27.8%); p53 LI, even if slightly higher in PA with progression 

or recurrence, is not significant using ROC curve and CART 

analyses. On the contrary, the CART-derived tree evidences that 

each PA subtype has its specific prognostic factors. In cases of 

PRL and ACTH type PA, the Ki67 LI has the main prognostic 

value. Specifically, a cut-off of 4.40% shows the highest accuracy 

in the PRL type of PA, while the cut off in the group of ACTH 

is 1.70%. Invasion as evaluated by MRI emerges as the most 

important prognostic factor in cases of non-functioning PA since 

it identifies 5 of 6 (83.3%) cases with recurrence or progression. 

In the non-invasive subgroup, the Ki67 LI is useful in identifying 

patients at risk of recurrence/progression. On CART analysis, GH 

adenomas show different prognostic features since patient age 

and sex appear to be the most useful. 

Conclusions. In conclusion, the CART algorithm generates decision 

trees which appear to be useful to identify PA with high risk 

of recurrence. 

Pilomatrix carcinoma arising in a pilomatrixoma 

L. Alessandrini, R. Salmaso, R. Cappellesso, A. Fassina 

Dipartimento di Scienze Medicodiagnostiche e Terapeutiche, Università 

di Padova, Italia 

Background. Pilomatrix carcinoma is a rare malignant tumor of 

hair matrix differentiation, occurring most frequently in elderly 

patients on posterior neck, pre- and retro-auricolar area. It can be 

distinguished from its benign counterpart mainly for the presence 

of necrosis, islands of basaloid cells with high mitotic index and 

true capsular infiltration. 

Methods. 81-year-old patient presented an ulcerated lesion in 

the left lower eyelid which was excised and routinely processed 

for H&E, PAS, and for immunohistochemistry for cytokeratins 

(CKs), Epithelial Membrane Antigen, Carcinoembryonic antigen, 

S-100 and MIB-1. 

Results. The lesion was a symmetric dermal nodule with an overlying 

ulcerated epidermis, characterized by peripheral lobules of 

basaloid cells arranged in sheets and nests and a central area with 

“ghost” cells, and pale eosinophilic cytoplasm. In most basaloid 

nests, cells had hyperchromatic nuclei and prominent nucleoli, with 

high mitotic rate (4-8 mitoses/HPF), as confirmed by MIB-1 reaction 

(40%), with pushing margins with islands of infiltration of the 

surrounding capsule. Squamous differentiation was demonstrated 

by positive and strong staining for CKs, negative in “ghost” cells. 

Foci of necrosis and a patchy lymphocytic infiltrate were present, 

whereas vascular and perineural invasion was not detected. Besides 

the histological features of malignancy, benign aspects were identified: 

few basaloid lobules composed of uniformly sized, typical 

cells with low mitotic rate and “ghost” cells formation towards the 

centre of the tumor. Only 50 cases of pilomatrix carcinoma have so 

far been reported, which usually arises de novo, and only occasionally 

in a pilomatrixoma: neither molecular biology nor immunohistochemistry 

are helpful in distinguishing the two entities, and their 

distinction remains uniquely on the histological recognition. 

PIK3CA gene mutations in lung neuroendocrine 

tumors 

A. Capodanno, G. Alì, L. Boldrini, G. Riccardo, A. Servadio, 

M.I. Rotondo, G. Fontanini 

Surgery, Santa Chiara Hospital, Pisa, Italy 

Background. Lung neuroendocrine tumors represent about 20% 

of all lung carcinomas and comprise a large spectrum of tumors 

that share structural, morphologic, immunohistochemical, and

238 

ultrastructural features. Lung neuroendocrine tumors are classified 

into four main groups with different biologic aggressiveness: 

typical carcinoids (TCs), atypical carcinoids (ATs), large-cell 

neuroendocrine carcinomas (LCNCs), and small-cell lung carcinomas 

(SCLCs). Recently, somatic mutations in the phosphatidylinositol-3-kinase 

(PI3K) catalytic subunit (PIK3CA) gene 

have been reported in several human cancers. The cancer-associated 

PIK3CA mutations lead to an enhanced enzymatic activity, 

the upregulation of the downstream signalling cascade, and the 

oncogenic cell transformation. In this study, we investigated the 

PIK3CA gene status in lung neuroendocrine tumors. 

Methods. Mutations in the helical and kinase domains of the 

PIK3CA gene were determined by direct gene sequencing analysis 

in 189 lung neuroendocrine tumors, including 80 TCs, 17 

ACs, 17 LCNCs, and 75 SCLCs. 

Results. The frequency of PIK3CA gene mutation in lung neuroendocrine 

tumors was 52/189 (27.5%). The mutation distribution 

was approximately twice in the kinase domain (37/52) compared 

with the helical domain (15/52). The most prevalent PIK3CA 

gene anomalies were the H1047R and G1049S mutations in the 

kinase domain and the E542K and E547K mutations in the helical 

domain. No significant associations were observed between 

PIK3CA gene status and age, sex, or lymph node status of the 

patients. However, we found a significant association between 

PIK3CA gene status and lung neuroendocrine tumor histology 

(p=0.029) with PIK3CA mutations that were more frequent in 

more aggressive AC, LCNC, and SCLC histotypes. Our study is 

the first report of PIK3CA gene mutations in lung neuroendocrine 

tumors and the high frequency of mutations suggests an important 

role of PIK3 kinase in tumorigenesis of these tumors. 

role of glucocorticoides and matrix 

metalloproteinases in the pathogenesis of pelvic 

organ prolapse: a clinicopathological study of 34 

cases 

1)A.M. Altavilla, 2)D. Caliandro, 2)M. Politano, 1)L. Carluccio, 

2)L. Milano 

1)U.O. di Anatomia Patologica, Pia Fondazione “Card .G. Panico”, 

Azienda Ospedaliera, Tricase (Le), Italia; 2)U.O. di Ostetricia e Ginecologia, 

Pia Fondazione “Card. G. Panico”, Azienda Ospedaliera, Tricase 

(Le), Italia 

Background. Pelvic organ prolapse(POP) is a debilitating disorder 

for women. Risk factors are known but the pathogenesis is 

unclear. Imbalance between metalloproteinases(MMPs) and their 

inhibitors TIMPs plays an important role in connective remodelling 

process. Many data suggest that glucocorticoides(GC) excess 

damage matrix homeostasis. The aim of our study is to evaluate in 

incontinent women connective tissue alterations and the immunohistochemical 

expression of MMP2/TIMP2, in uterosacral ligament, 

a pelvic support, compatibly with changes of cortisol levels 

and with an Hypothalamic-Pituitary-Adrenal axis activation. 

Methods. We analyzed the uterosacral ligaments specimen of 

16 incontinent women and as controls from 18 women who 

underwent benign gynaecologic surgery. Histochemistry for trichrome 

and elastic stain and immunohistochemistry for MMP2 

and TIMP2 were performed on paraffin embedded sections. The 

slides were scored by the pathologist blinded to diagnoses. GCs 

profile was evaluated on the response of basal Cortisol-ACTH 

ratio before and after a dexamethasone-suppression test(0.5mg). 

Statistic analyses: mean±SD, Spearman’s rank, chi-squared test, 

p-values of < 0.05 significant. 

Results. There was no difference(ns) in parity, menopausal status 

and age between groups, the only significant datum was stress 

incontinence(p < 0.05). In POP group GCs levels were increased 

compared to controls, excluding effects of age and parity. Women 

with POP compared to those without revealed a decrease of 

collagen cellularity. The score dispersion rate of elastin did not 


show difference between groups. POP group revealed an important 

higher MMP2 expression than non-POP group(p < 0.01). 

The ratio of MMP2/TIMP2 was higher in the POP-group than 

in controls. These data are consistent with increased collagen 

break-down as a pathologic aetiology of prolapse with a laxity 

of collagen content due to connective degradation rather than a 

decrease of collagen synthesis and with GCs influence. 

Nasal seromucinous hamartoma (microglandular 

adenosis): a morphological and molecular study 

of five cases 

A. Ambrosini Spaltro, L. Morandi, D.V. Spagnolo * , A. Cavazza 

** , M. Brisigotti *** , S. Damiani, V. Eusebi 

Sezione di Anatomia Patologica, Dipartimento di Oncologia ed Ematologia, 

Università di Bologna, Ospedale Bellaria, Bologna, Italia; * Department 

of Anatomical Pathology, PathWest Laboratory Medicine, Queen 

Elizabeth II Medical Centre, Nedlands, Western Australia; ** Unità Operativa 

di Anatomia Patologica, Arcispedale S. Maria Nuova, Reggio Emilia, 

Italia; *** Unità Operativa di Anatomia Patologica, Ospedale Infermi, 

Rimini, Italia; 

Background. Seromucinous hamartoma (SH) is a rare glandular 

lesion of the sinonasal tract and nasopharynx. Cases reported in 

the literature are limited and there are few follow-up data. 

Methods. The clinicopathological features of five cases of nasal 

SH were analyzed. Immunoreactivity for alpha-smooth muscle 

actin (α-SMA), calponin, desmin, p63, CK 14, laminin, collagen 

IV, S100 protein, Ki-67 and EMA was assessed. Molecular 

analyses for clonality using mtDNA (mitochondrial DNA) were 

conducted. The mtDNA of five cases with normal nasal mucosa 

obtained by turbinate resection was used as the “normal” counterpart 

for genetic analysis. 

Results and comments. Patients (3 F and 2 M) ranged from 49 

to 66 yrs in age. All lesions were located in the nasal cavity. In 4 

cases with follow-up there was no recurrence. 

In all cases the lamina propria exhibited a proliferation of small 

seromucinous glands embedded in a dense, fibrotic stroma. Neither 

mitotic activity nor nuclear atypia were observed. 

Around the small proliferating seromucinous glands, no immunoreactivity 

for p63 and CK 14 was detected, but expression of laminin 

(2 of 5 cases) and collagen IV (all cases) was observed. Glandular 

epithelial cells were positive for S100 protein in all cases, for 

EMA in 4/5 cases and Ki-67 positivity ranged from 1% to 2%. The 

immediately periglandular stromal cells were reactive for calponin 

in all cases, for α-SMA in 4/5 cases and focally for desmin in 2/5 

cases, while the intervening stroma was completely negative. 

In the 5 cases with normal nasal mucosa the mean mutation rate 

was 0.83% (0.23% homoplasmy, 0.67% heteroplasmy), while the 

lesional cases showed a higher mutation rate, especially in heteroplasmy 

(0.52% homoplasmy, 2.02% heteroplasmy). 

These features indicate that this unusual glandular proliferation is a 

hyperplastic lesion both at morphological and molecular levels. It also 

shares some similarities with microglandular adenosis of the breast. 

endoscopic mucosal resection and endoscopic 

submucosal dissection as an alternative treatment 

for dysplastic lesions and early cancer of the 

stomach 

M.R. Ambrosio, M. Onorati, B.J. Rocca, V. Mourmouras, 

M. Mario * , L. Barbagli, F. De Luca, C. Vindigni 

Department of Human Pathology and Oncology-Anatomic Pathology Section, 

Santa Maria delle Scotte, Siena, Italy; * Gastroenteric Unit, Santa 

Maria Delle Scotte, Siena, Italy 

Background. Endoscopic mucosal resection (EMR) and endoscopic 

submucosal dissection (ESD) have been developed as 

treatment for gastric dysplastic lesions and early gastric cancer in 

Japan and in the Western world.

oral communications and Posters 

Methods. During the period 1998-2009, 34 EMR and 10 ESD for 

a total of 44 lesions, histologically diagnosed as dysplasia or early 

cancer, were performed in the Hospital of Siena. 

Results. 24 cases (55%) were piecemeal resections and 20 

(45%) were en bloc resections. The lesions were mostly located 

in the body (19 cases) and the size ranged from 0.8 to 3.5 cm. According 

to the endoscopic Paris Classification, 19 cases were type 

I (Is = 18, Ip = 1), 11 type II, prevalently IIa (10 cases) and 14 

mixed types, prevalently IIa+IIc (11 cases). Complications were 

represented by two cases of perforations and two cases of late 

onset of bleeding, readily treated and resolved. According to the 

histological Vienna classification, 16 cases were type 3, 8 were 

4.1, 14 were 5.1 and 6 were 5.2. Complete (lateral and deep margins 

free) and incomplete resection was confirmed histologically 

in 32 (76%) and 10 cases (24%) lesions respectively; in two cases 

margin involvement was not evaluable. Seven patients underwent 

surgical treatment because of the involvement of the cut ends. In 

one of these cases the diagnosis was that of high grade dysplasia, 

three cases were T1, in two cases the cancer involved the muscular 

layer (T2) and in one case there was no cancer. All cases were 

N0. The median follow-up period was 30 months (range 3-135). 

In three cases of lesion 3, one recurred after 9 months and two 

after 60 months; a case of 5.1 lesion recurred after 12 months. 

Conclusions. Our data suggest that EMR may provide an alternative 

treatment to surgery for selected cases of preneoplastic and 

superficial neoplastic gastric lesions. 

Cortical thymic epithelial tumors have an 

increased risk of developing additional 

malignancies: lack of immunologic surveillance? 

M.R. Ambrosio, B.J. Rocca, F. Granato * , D. Spina, S. Lazzi, 

L. Leoncini 

Human Pathology and Oncology-Anatomic Pathology Section, Santa Maria 

delle Scotte, Siena, Italy; * Cardiothoracic and Vascular Surgery, Thoracic 

Surgery Unit, Santa Maria delle 

Background. The increased risk of developing an additional malignancy 

(AM) before or after a thymic epithelial tumors (TET) 

has not yet been fully examined and no relations with histologic 

pattern of thymic neoplasma was found. 

Methods. 52 patients who underwent surgical excision for TET 

were studied. Based on the WHO classification, the tumors were 

classified as A, AB (B1 and B2-like) and B thymoma, and thymic 

carcinoma (C). A control population was provided by the creation 

of a further database comprising 114 patients with colorectal 

cancer (CC). 

Results. Patients with TET showed a statistically significant 

higher risk of developing AM compared to patients with CC 

(12/52 vs 11/114 patients, p = 0.0374). The association between 

TETs and AM was related to the TET histotype. B2, B3, AB 

(B2-like) and C were histotypes more correlated with the onset 

of an AM. Taking into consideration the histogenesis of these 

thymomas prevalently from cortical epithelial cells (cTECs), 

cases were divided into two sub-groups. Sub-group 1 included 

29 patients with A, AB (B1-like) and B1 thymomas, sub-group 2 

comprised 23 patients with AB (B2-like), B2, B3 thymomas and 

C. Sub-group 2 showing a statistically significant higher risk of 

developing an AM (p = 0.008). The time interval (TI) between 

the appearance of the first and second tumor in TET group was 

significantly shorter than those in CC group (p = 0.014), with 

TETs following AM in many cases (n = 10). 

Conclusions. Patients affected by TETs have a significantly 

higher risk of developing AM and this risk is considerably greater 

in tumors exhibiting a prevalent cortical origin. This may be 

related to the role of cTECs in presenting foreign antigen. The 

generally low TI values between TETs and other malignancies 

suggest the potential presence of an immunological impairment 

that often appears prior to evidence of TET. 

239 

Aberrant expression of Tfr1/CD71 in thyroid 

carcinomas identifies a novel potential diagnostic 

marker and therapeutic target 

1)A. Torrisi, 1)P. Amico, 2)I. Cataldo, 3)R. Parenti, 1)G.M. Vecchio, 

4)R. Perris, 1)G. Magro 

1)Dipartimento “G.F. Ingrassia” - Università di Catania, Azienda Ospedaliero-Universitaria 

–Policlinico Vittorio Emanuele, Catania, Italia; 

2)Dipartimento di Patologia, Università di Verona, Verona, Italia; 3)Dipartimento 

di Scienze Fisiologiche, Università di Catania, Catania, Italia; 

4)Comt, Università di Parma, Parma, Italia 

Background. Type I receptor for transferrin (TfR1/CD71) is 

overexpressed in several malignant tumors. We investigated the 

expression of TfR1/CD71 in benign and malignant thyroid tissues. 

Methods. Tissue samples, including benign lesions and follicular-derived 

carcinomas, from 241 patients and a total of 35 benign 

and malignant fresh specimens were assayed for TfR1/CD71 expression 

by RT-PRC, Western blot and immunohistochemistry. 

Results. We found that transcription of TfR1/CD71 gene is constitutive 

in thyroid epithelia, but the mRNA is differently translated 

in benign and malignant tissues. In benign tissues low levels 

of TfR1/CD71 were found, whereas most carcinomas exhibited 

overexpression of the receptor, predominantly in the cytoplasm 

of neoplastic cells. The highest expression level was detected in 

primary and metastatic papillary carcinomas and anaplastic carcinomas, 

with a positivity ranged from 86% to 100% of the cases. 

Our findings suggest that altered expression of TfR1/CD71 is a 

marker of malignancy in thyroid tissues where it is useful in distinguishing 

PTC from benign lesions with PTC-like cyto-architectural 

alterations and follicular variant PTC from benign follicularpatterned 

lesions. The present observations support the rationale 

for the use of radiolabeled transferrin/transferrin analogs and/or 

anti-TfR1/CD71 antibodies for diagnostic and/or radiotherapeutic 

purposes in TfR1/CD71-expressing thyroid tumors. 

Comparison of three different methods for the 

analysis of codon G12 and G13 of the KrAS gene 

1)Andreozzi MC. 2)Bihl MP. 3)Foesrster A. 4)Rufle A. 5)Tornillo 

L. 6)Terracciano LM. 

1)Institute of pathology, University of basel, Basel, Switzerland 2)Institute 

of pathology, University of basel, Basel, Switzerland 3)Institute of 

pathology, University of b, Basel, Switzerland 4)Institute of pathology, 

University of basel, Basel, Switzerland 5)Institute of pathology, University 

of basel, Basel, Switzerland 6)Institute of pathology, University of basel, 

Basel, Switzerland 

Aims and Methods. KRAS mutation screening has been achieved 

high importance in selecting the right therapy for patients with 

colorectal cancer and non-small-cell lung cancer especially in 

metastatic disease stage. Screening for KRAS mutations in these 

patients provide additional information on optimizing treatment 

options with targeted drugs. Paraffin embedded tissue from 100 

colon carcinomas were randomly selected to include a wide spectrum 

of KRAS mutations. A comparison of three different methods 

for the analysis of the KRAS gene of codon G12 and G13 

using the same DNA preparation for all methods was performed. 

For the molecular analysis the following methods were used: 

Pyrosequencing. First step: Amplification of the sequence for 

analyzation by PCR. Second step: Enzymatic reaction cascade 

during the synthesis of the previously amplified sequence that 

converts the specific nucleotide incorporation into light. 

INFINITI ® : The analyzer is designed to measure fluorescence 

signals of labelled DNA target hybridized to BioFilmChip ® microarrays. 

The analyzer automates the assays and integrates all 

the discrete processes of sample (PCR amplicons) handling, reagent 

management, hybridization, detection, and result analysis.

240 

Dideoxysequencing. The DNA to be sequenced is amplified by 

PCR. 

Results. The following mutations were detected in codon 12 and 

13 as follows: G12A 5x, G12C 2x, G12D 16x, G12S 3x, G12V 

15x, G13C 1x, G13D 17x, G13R 1x and 40 samples were wild 

type. 

Conclusions. This is the first study, which analyzed three different 

methods in a comparative matter. All these 3 methods are 

suitable for detecting hot spot mutations in the Kras oncogene. 

Infinity technology seems to be more sensitive in samples contaminated 

with high amounts of non mutated cells or in samples 

of low DNA quality. Wrong results were all a problem of sensitivity 

(1% false negative for Infinity and Dideoxysequencing 

technology, respectively). 

Vegfa amplification in different neoplastic 

entities: tissue microarray analysis on 2292 tissue 

samples 

1)Andreozzi MC. 2)Vlajnic T. 3)Zlobec I. 4)Bihl MP. 5)Tornillo 

L. 6)Schneider S. 7)Lugli A. 8)Terracciano LM. 


of pathology, University of basel, Basel, Switzerland 3)Institute of pathology, 


of basel, Basel, Switzerland 5)Institute of pathology, University of 

basel, Basel, Switzerland 6)Institute of pathology, University of basel, Basel, 

Switzerland 7)Institute of pathology, University of basel, Basel, Switzerland 

8)Institute of pathology, University of basel, Basel, Switzerland 

Aims. Angiogenesis plays an important role in progression of 

several tumor types. Evidence from preclinical and clinical studies 

indicates that vascular endothelial growth factor (VEGFA) is 

the predominant angiogenic factor. The aim of this study was a 

systematic investigation of VEGFA amplification in a large survey 

of solid human tumors in tissue microarray format. 

Methods. FISH analysis of the VEGFA gene was performed 

in a multi tumor array (n = 2292) including 132 different tumor 

categories and 31 normal tissue types. Additionally VEGFA gene 

amplification was evaluated in a further large series of sporadic 

CRC resections (n = 1280) and the obtained data were compared 

to relevant clinico-pathological features. 

Results. VEGFA amplification was detected in carcinoma of 

colon (n = 39; 3%), gall bladder (n = 5; 13.2%), pancreas (n = 3; 

6.5%), prostate (n = 6; 15.8%), stomach (n = 6; 14.3%), testis 

seminoma (n = 4; 8.5%) and colon adenoma (n = 7; 9.2%). VEG- 

FA amplification in CRC significantly correlated with higher 

T stage and higher tumor grade, presence of vascular invasion, 

right sided location and with worse survival in univariate and 

multivariable analysis. 

Conclusions. Albeit rare, VEGFA amplification can be detected 

in several different tumor entities. In CRC it highlights a small 

subset of CRCs with aggressive phenotype. Additional studies are 

needed to evaluate its significance in other neoplastic entities. 

extraskeletal ewing sarcoma in a 78-years-old 

woman: a case report 

U.F. Angelotti, R. Scamarcio, A. Scivetti, A. Colagrande, 

C. Traversi, A. Cimmino, L. Resta 

Anatomia patologica, Policlinico, Bari, Italia 

Background. Extraskeletal Ewing sarcoma (EES) is a rare soft tissue 

tumour morphologically indistinguishable from the more common 

Ewing Sarcoma of bone. It must be differentiated from other 

small, blue round cell tumours, including primitive neuroectodermal 

tumour and neuroblastoma. The age at the time of diagnosis, 

unlike its osseous counterpart, has a wide range, from infancy to 

the elderly, and has a slight predominance in male patient. 


Methods. We present a case of EES in the retroperitoneum of a 

78-year-old woman which in 1993 has been diagnosed of Ewing 

sarcoma of the upper third of right arm. The history clinical is 

silent until April 2000 when it occurs disease recurrence in the 

distal third of the right arm, followed in February 2003 by intervention 

of right axillary lymph nodes metastatic dissection and 

in November 2007 by secondary localization of Ewing sarcoma 

at level of the fifth hepatic segment. For about four months, the 

patient complains of pain in the epi-mesogastrial area, anorexia, 

fatigue and malaise; the computed tomographic scan shows solid 

expansive mass, suggestive of peritoneal metastasis, in the mesogastrial 

median area. 

Results. The tumour lacks of immunoreactivity for epithelial, 

lymphoid, vascular, neuroendocrine, neural and muscle markers. 

Immunohistochemically, the tumor was positive for vimentin, 

CD99, slightly positive for CD45LC, but negative for CKpool, 

CD117, CK-20, MPO. Extraskeletal Ewing sarcoma was 

confirmed by electron microscopy, which showed a prominent 

nucleus with marginated chromatin, few organelles and abundant 

glycogen. Primitive neuroectodermal tumour was excluded because 

of the lack of neural differentiation by histologic analysis, 

immunohistochemistry and electron microscopy. This case serves 

to remind the reader that EES is not a tumour that occurs exclusively 

in young patients. 

The question of reproducibility in cytology, 

histology and colposcopy 

1) D. Antonini 2) A. Marsico 3) A. Anastasio 4) M. I. Rostan 5) 

R. Navone 

1) Ospedale degli Infermi, UO di Anatomia Patologica, Biella, Italia 2) 

Ospedale Koelliker, UO di Anatomia Patologica, Torino, Italia 3)Dipartimento 

di Scienze Biomediche e Oncologia Umana dell’Università di Torino 

(Sez. di Anatomia Patologica), Italia 4) Dipartimento di Scienze Biomediche 

e Oncologia Umana dell’Università di Torino (Sez. di Anatomia 

Patologica), Italia 5) Dipartimento di Scienze Biomediche e Oncologia 

Umana dell’Università di Torino (Sez. di Anatomia Patologica), Italia 

Background. It is well known that colposcopy has a low specificity 

(Barrasso, 1998), above all if used as a 1st level test. Indeed, 

should a positive Pap test be followed by a colposcopic grade 1 

abnormal transformation zone (ATZ), then, 79% of cases will be 

histologically positive; without a previous positive cytology, the 

values of positivity of histology are very low i.e. in the range of 

20% for grade 1 colposcopical ATZ 

Methods. We evaluated the correlation amongst cytology, histology 

and colposcopy in a spontaneous screening group (21,451 

cases), where colposcopy was done at the same time as the Pap 

test. A biopsy was also carried out in the presence of grade I or 

higher ATZ. 

Results. A total of 21,451 Pap tests were done along with colposcopies. 

There were 2,175 abnormal colposcopy results (mostly 

grade 1 ANTZ). The colposcopic diagnosis were: white epithelium 

(1,002 cases), 603 keratosis, 279 punctate, 280 mosaic and 

11 carcinoma. The cyto-histological diagnosis of the abnormal 

colposcopy results included 210 L-SIL, 56 H-SIL and 11 carcinomas 

(277 cases). There were 170 abnormal cytology results 

(confirmed by histology) (113 L-SIL, 54 H-SIL, 2 endocervical 

adenocarcinoma in situ (AIS) and 1 carcinoma) in patients 

with a normal colposcopy result (G 0). Whilst there were 1.898 

abnormal colposcopy results associated to normal cytology and 

histology. 

Our data showed that 89.1% of the patients had both a negative 

colposcopy and Pap test and that 1.3% of the cases were both positive. 

However, there were 8.8% of patients in whom, although 

the colposcopy was positive, the cytology and histology were 

negative. Whilst, despite the fact that a 0.8% of the Pap tests and 

histology were positive, their colposcopies were negative. The 

discordance between the colposcopy and histo-cytology results


indicates that colposcopy alone, i.e. without the association of 

cytology and histology, is not able to offer a definitive diagnosis. 

This is particularly true for abnormal colposcopy results (grade I 

ANTZ or higher) that should always have an anatomopathological 

confirmation. 

Neonatal neuroblastoma mimicking 

sacrococcygeal teratoma: an autoptic case. 

Vincenzo Arena*, Ilaria Pennacchia*, Egidio Stigliano*, Fabio 

De Giorgio°, Arnaldo Carbone*, Fabio Maria Vecchio* 

*Institute of Pathology; °Institute of Legal Medicine; Catholic University 

of Sacred Heart, Roma 

Background. Neonatal masses occurring in the sacrococcygeal 

region are mostly teratomas. We report herein a case of neonatal 

neuroblastoma in a newborn male infant delivered after a normal 

pregnancy. 

Methods. The neonate was brought to our hospital after a normal 

vaginal delivery (38 weeks of gestation), because of an extremely 

large sacrococcygeal mass. A RMI showed an enormous neoplastic 

mass with an undifferentiated and uniform internal structure, 

which extended from the sacrococcygeal region to the celiac 

region and was pressing on the rectum and the bladder. Multiple 

metastatic lesions to the liver were seen too. The general condition 

of the child rapidly worsened and he died before a biopsy 

was performed on the mass. For this reason the autoptic examination 

was required and revealed the presence of a tumor mass of 

12 cm in diameter, which was extremely firm and immobile and 

adherent to the rectum. Multiple repetitive lesions were noted 

both in the liver and adrenal glands. A macroscopic diagnosis 

of “likely” malignant teratoma was made. Interestingly, histopathologic 

examination of the tumor showed rosette formation 

and neuroblastoma cells with small nuclei and fibroid cytoplasm. 

The tumor cells were strongly immunopositive for NSE and Sinaptophysin. 

The histopathological diagnosis was neuroblastoma, 

classified as stroma poor, undifferentiated in the Shimada pathological 

classification 

Discussion. The differential diagnosis of sacrococcygeal neoplasms 

includes several lesions like meningomyelocele, lipoma 

and lhymphangioma; however, in newborns they are most often 

teratomas. Our case suggests that neuroblastoma should be considered 

in differential diagnosis by pathologists who perform 

perinatal autopsies and confirms once again the role of autopsy 

in the correct nosographic definition of diseases. 

Chondroma of the hand with osteoid formation 

Vincenzo Arena*; Ilaria Pennacchia; Arnaldo Capelli; Arnaldo 

Carbone; Fabio Maria Vecchio 

Institute of Pathology Catholic University of Sacred Heart Roma (Italy) 

Background. Chondroma is the most common bone tumor arising 

in the hand. Histologically the majority of them consist of 

mature hyaline cartilage arranged in a lobular pattern. Frequently 

the cartilage has focal or diffuse calcification. 

Methods. A 36 year-old woman presented with a swelling of the 

proximal phalanx of the 3 th finger of the left hand appeared five 

months before, with no history of a previous traumatic event. At xray 

the lesion extended up to the ulnar cortex without evidence of a 

pathologic fracture. Histologically the lesion was composed of well 

differentiated chondrocytes and mature hyaline cartilage. Areas of 

myxoid stroma with scattered cells without any atypical features 

were also present. Neither double-nucleated cells nor clusters 

of chondrocytes were seen. No mitotic figures were seen [MIB- 

1 < 2%]. Interestingly focal deposits of osteoid within the lesion 

were also seen. The patient had no relapse with 1 year follow-up. 

Results. In chondroma of the hand. in case the myxoid component 

is predominant, a myxoid variant of chondroma should 

241 

be considered. In this context, the main differential diagnosis 

is myxoid chondrosarcoma. In our case, a diagnosis of malignancy 

was not considered due to the absence of cellular 

pleomorphism and because of the hand being a very unusual 

site for malignant chondroid neoplasms. As for the unusual 

presence of osteoid matrix formation, it is described in both 

chondroblastoma and in chondromyxoid fibroma. The lesion 

we described did not fullfill all the criteria for a diagnosis of 

the above mentioned entities. However, we believe the finding 

of a osteoid matrix is stricking in the setting of a chondroma of 

the hand. It is common for both benign and malignant cartilage 

tumors, to undergo pathologic fracture, making the histology 

of new bone formation associated with the cartilage somewhat 

complex but in the case presented, neither radiologic nor 

pathologic signs of fracture were seen. 

refined Immunohistochemistry scoring criteria for 

Her-2 “borderline” breast cancer: study on 230 cases. 

Arena Vincenzo, Pennacchia Ilaria, Fabio Maria Vecchio, Arnaldo 

Carbone 

Institute of Pathology, Catholic University of Sacred Heart, Rome 

Background. Two methods are used for measuring HER-2 in the 

clinical setting: immunohistochemical analysis (IHC) and fluorescence 

in situ hybridization (FISH). Convenience dictates that IHC 

remains the screening test for HER-2 status in patients with breast 

cancer, adopting FISH as second-line diagnostic tool in case of 

doubtful IHC results. Aim of this study is to investigate IHC criteria 

for scoring HER2 in order to refine the “2+” category. 

Methods. Two hundred thirty cases resulted IHC/2+ (DAKO 

scoring system) were subsequently evaluated for HER-2 gene 

amplification by FISH. Granularity of signal, linear, even though 

not complete, membrane decoration, signal intensity (1+ to 3+ 

score) and presence of linear paired definite membrane signal 

between cells (“track” feature) were blinded evaluated by us and 

discordant cases were discussed until an agreement was reached. 

Results. A granular staining pattern was seen in 73% of HER-2 

FISH negative cases (p = 0.0006). Seventy four percent of FISH 

positive cases showed linear membrane decoration of some extent 

(p = 0.011). An intense overall IHC signal (3+) was observed in 

45% of FISH positive cases (p = 0.0009). Fifty nine percent of 

FISH positive cases presented “track” images in > 25% of cell 

population (p = 0.054; ns), whereas 68% of FISH positive cases 

presented simultaneously “rimming” and strong IHC reactivity 

(p = 0.0002). 

Conclusions. Combined intensity and linearity of membrane 

signal, even though limited (intense partial membrane staining) 

resulted the best aid for the pathologist in making final scoring 

decision in borderline IHC HER-2 tests. In our opinion, the 

effort of the pathologist in adding IHC details for refining the 

worldwide validated criteria for IHC HER-2 assessment, could 

reduce the number of “borderline” cases undergoing FISH with a 

significant benefit to economy lab. 

Are we losing the value of autopsy? evidence 

from an epidemiological descriptive study 

Vincenzo Arena*; Luca Valerio#; Ilaria Pennacchia*; Fabio De 

Giorgio§;Bruno Federico°; Arnaldo Capelli*; Fabio Maria Vecchio* 

*Institute of Pathology; #Institute of Hygiene; §Institute of Legal Medicine; 

Catholic University of Sacred Heart, Roma; °Department of Health 

and Sport Sciences; University of Cassino 

Background. Substantial evidence shows the high accuracy of 

autopsy relative to clinical diagnosis in determining the cause of 

death. Traditionally, pathologists provide clinicians with a feedback 

for improvement, by identifying the diseases at greater risk

242 

of error. Nevertheless, the number of autopsies has been on the 

decrease in all countries over the last twenty years. 

Methods. We describe with a statistical analysis the variations in 

the characteristics of cases referred to pathologists in Our Hospital 

between two three-year periods over 20 years, to investigate 

the extent of the problem and the possible objective and cultural 

causes. Data were derived from the hospital’s Pathology registry, 

which includes data on all autopsies and is updated daily. 

Results. Autopsies have decreased in number and their composition 

has changed significantly: in terms of epidemiology, with a 

surge of neonatal autopsies; in terms of requesting wards and diseases 

diagnosed, with heart surgery, emergency and cardiovascular 

diseases and their respective diagnoses having increased in importance 

relative to requests from specialists in infectious diseases. 

Discussio. Our data show that such evolution is not consistent 

with that of the causes of hospital mortality in Italy over the same 

period: the cause of the phenomenon must be elsewhere. 

The decision of the clinician to ask for autopsy is mainly dictated 

by the latter’s accuracy relative to clinical diagnosis as well as by 

the increasing need for defensive medicine. 

Conclusion. There seems to be a paradox in the attitude of clinicians 

towards autopsy diagnosis: while demand for autopsies for 

the traditional purposes decreases, more autopsies are requested 

for what are likely to be medico-legal reasons. Therefore, accuracy 

of pathologic diagnosis in the clinicians’ opinion has not 

changed, but, for the same reason, what pathology can offer to 

clinics, education and research outside defensive medicine cannot 

have changed either. 

limits of TIr-3 reporting in Thyroid fine-Needle 

Cytology: 3-Year experience from A Single 

Academic Center 

1)Ascoli V. 2)Bosco D. 3)Taffon C. 4)Marinelli L. 5)De Mattia 

D. 6)Grillo L. 7)Nardi F. 

Anatomia Patologica, Dipartimento di Medicina Sperimentale, Università 

La Sapienza, Azienda Policlinico Umberto I°, Roma, Italia 

Introduction. The SIAPEC has proposed a 5-tier reporting system 

for thyroid fine-needle cytology (FNC), which include the 

“indeterminate/inconclusive” category (TIR-3). This category 

encompasses follicular-patterned lesions. In such cases, only histology 

(and no cytology alone) can provide a final diagnosis. 

Methods. In our laboratory, the 5-tier reporting system has been 

used for 7579 thyroid aspirates in the last 3 years (period 2007/ 

June 2009: 5680; period July-2009/April-2009: 1899). We assessed 

the distribution of aspirates by the 5 diagnostic categories 

in the two periods, and then we focused on TIR-3 cases by evaluating 

the proportion of surgically treated cases in our hospital and 

the histological diagnosis. 

Results. A total of 319 cases (4,2%) were interpreted as TIR-3. 

Of these 319 cases 125 had surgical follow-up in our hospital 

(39,2%). Overall, the surgical yield of malignancy was 22.4% (23 

papillary carcinoma, 1 follicular carcinoma, 3 Hürthle cell carcinoma, 

1 metastatic renal cell carcinoma); 24% were adenomas 

(20 follicular and 10 Hürthle cell adenomas) and the remaining 

53.6% were negative (53 nodular hyperplasia, 11 Hashimoto’s 

thyroiditis, 3 chronic thyroiditis). Six occult papillary carcinoma 

were identified as incidental finding (4 controlateral lobe; 2 omolateral 

lobe/additional nodule). 

Conclusions. Our survey is limited that a major fraction of TIR-3 

FNC (60%) had no surgical/histological follow-up in our hospital. 

Nevertheless, our results are in agreement with the literature 

concerning malignancy rate of TIR-3 (22%) and prevalence of 

occult thyroid carcinoma; the high fraction of benign nodules 

(> 50%) indicate that some TIR-3 cases could rather benefit of a 

repeat aspirate after an appropriate interval of observation instead 

of unnecessary surgery. For this is crucial the collaboration of pathologists 

with clinicians and radiologists (ultrasound findings). 


Cyclosporine-induced gingival overgrowth is 

associated to increased Transglutaminase -2 

expression 

1)Asioli S. 2)Cassoni P. 3)Righi A. 4)Cassenti A. 5)Maletta F. 

6)Carossa S. 7)Navone R. 

1)Scienze biomediche e oncologia umana, Molinette,Torino, Italia 2)Scienze 

biomediche e oncologia umana, Molinette, Torino, Italia 3)Scienze biomediche 

e oncologia umana, Molinette, Torino, Italia 4)Scienze biomediche 

e oncologia umana, Molinette, Torino, Italia 5)Scienze biomediche 

e oncologia umana, Molinette, Torino, Italia 6)Sezione di riabilitazione 

orale e maxillofaciale, Molinette, Torino, Italia 7)Scienze biomediche e 

oncologia umana, Molinette, Torino, Italia 

Background. Cyclosporine A induced gingival overgrowth, 

which is characterized by an extracellular matrix increase, is due 

to an altered balance between collagen synthesis and degradation. 

Cyclosporine A is a potent immunosuppressant used to prevent 

organ transplant rejection and to treat various autoimmune diseases. 

Methods. This study proposed to verify if transglutaminase 2, an 

enzyme that is thought to be responsible for the assembling and 

remodeling of extracellular matrix, played some kind of role in 

the pathogenesis of the cyclosporine A-induced gingival overgrowth, 

its expression in the gingival overgrowth was compared 

to normal tissue to evidence any differences. 

Cyclosporine A-induced gingival overgrowth tissues were collected 

from 21 liver transplanted patients and case-controlled 

with 20 non-hyperplastic gingival biopsies from healthy patients 

who had had previous periodontal treatment. Both the presence 

and tissue distribution of transglutaminase 2 was determined in 

the two groups by immunohistochemistry and analysed in comparison 

to the tissue morphology and expression of lymphocyte 

related antigens (CD3 and CD20) and a vessel related marker 

(CD34). 

Results. A significant increase in the transglutaminase 2 expression 

was observed within the stromal component in the cyclosporine 

A treated patients compared to controls (p < 0.001). An 

increased transglutaminase 2 expression in mesenchymal cells 

and/or extracellular matrix in gingival overgrowth suggests 

that this molecule has a role in the pathogenesis of the disease. 

Further studies will investigate the therapeutic effect of antitransglutaminase 

2 drugs (putrescine or 1,4-diaminobutane) in 

these patients. 

Nuclear membrane decoration by emerin staining 

improves cytological detection of papillary 

thyroid carcinomas 

Asioli S., Maletta F., Pacchioni D., Lupo R., Bussolati G. 

Biomedical sciences and human oncology, Molinette, Torino, Italia 

Background. The diagnosis of follicular lesions is a grey zone in 

thyroid fine-needle aspiration (FNA) cytology. Our study aims to 

verify if staining with Emerin is a helpful marker of the follicular 

variant of papillary thyroid carcinoma (FVPTC) in the differential 

diagnosis of follicular-patterned lesions. 

Methods. We designed both a prospective study on smears 

and Thin Prep specimens to prove the feasibility of the procedure 

and a retrospective study on 78 FNA cell-blocks from 

cases which, after surgery, turned out to be either benign (34 

cases) or malignant lesions (44, of which 31 PTC). From each 

sample, we obtained two slides, one stained with Hematoxylin 

and Eosin (H&E) and the other with immunohistochemistry 

(IHC) for Emerin. In Thy3 cases, HBME-1 and Gal 3 stains 

were also done. Two observers gave a judgement in Thy 

categories (British Thyroid Association) on H&E, Emerin, 

HBME-1 and Gal 3 stained slides. 

Results. The prospective study demonstrated that Emerin staining 

is an effective tool for nuclear membrane decoration and


amplification of nuclear irregularities. In the retrospective study, 

inter-observer agreement proved higher in Emerin-stained slides 

(K of Cohen-Fleiss = 0.6890) than H&E-stained slides (K of 

Cohen-Fleiss = 0.4878). Sensitivity and overall accuracy were 

higher for Emerin (respectively, 77.27% and 84.61%) than H&Estained 

slides (respectively, 36.36% and 62.82%). Emerin staining 

proved able to identify all cases of PTC, including all cases 

of FVPTC. In Thy3 cases, Emerin’s sensitivity and specificity 

(64% and 96%) proved higher than HBME-1’s (60% and 88%), 

and Gal3’s (61% and 68%). 

Conclusions. Emerin stain, is a useful tool in the cytological 

diagnosis of thyroid lesions. It enhances detection of nuclear 

irregularities typical of PTC, thus helping to solve inconclusive 

FNA cases, mainly in those cases of FVPTC with a reduced 

expression of nuclear irregularities in the traditional stains 

(H&E). 

Her2 overexpression in patients with small tumor 

size and node-negative breast cancer: a high risk 

group? 

1)Petroni S. 1)Asselti M. 2)Giotta F. 3)Quero C. 4)DAamico C. 

5)Marzano A.L. 6)Daprile R. 7)Palma F. 8)Simone G. 

1)Pathological Anatomy Unit, NCI “Giovanni Paolo II”, Bari, Italia 

2)Medical and Experimetal Oncology Department, NCI “Giovanni Paolo 

II”, Bari, Italia 3)Pathological Anatomy Unit, NCI “Giovanni Paolo II”, 

Bari, Italia 4)Senology Unit, NCI “Giovanni Paolo II”, Bari, Italia 5)Pathological 

Anatomy Unit, NCI “Giovanni Paolo II”, Bari, Italia 6)Pathological 



Anatomy Unit, NCI “Giovanni Paolo II”, Bari, Italia 

Background. Approximately 25% of breast cancer is HER2/ 

Neu overexpressed and/or amplified being both prognostic 

and predictive factors associated with worse disease-free and 

overall survivals. HER/Neu+ patients with metastatic tumor or 

in adjuvant systemic therapies are eligible for treatment with 

Trastuzumab according to the results of the major phase III clinical 

trials that do not include small (pT1a or pT1b), node negative 

breast cancer. Patients with node-negative breast carcinoma 

have a good prognosis but in ~ 20-30%, cases a recurrence of 

disease is present. The aim of this study is to evaluate the risk 

of recurrence in women with pT1a or pT1b, N0, M0, HER2+ 

breast cancer. 

Methods. We collected 279 women with small invasive breast 

cancer, pT1, N0, M0 from 2004 to 2009. 89 out of 279 

(32.3%) < 1cm sized (pT1a and b), node-negative (median age: 

56 ys) entered the study. In all cases ER, PgR, Ki-67 status 

and expression of Her2/Neu, using immunoistochemistry, were 

evaluated. Hormonal receptors and Ki-67 were scored according 

to St Gallen conference guidelines; expression of Her2/Neu was 

detected using HercepTest (DAKO) and scored according to FDA 

guidelines. 

Results. 18 out of 89 cases were pT1a (20.2%) and 71 pT1b. 68 

tumors (76.4%) were ER +, 65 (73.0%) were PgR + and 23 cases 

(26%) showed a high proliferative activity (Ki-67 index: > 20%). 

10 out of 89 (11.2%), 2 pT1a and 8 pT1b, evidenced Her2/Neu 

overexpression: only one case was G1, 2 were ER+/PgR+, 7 

showed high expression of Ki-67. 

Follow-up data (mean FU: 44.4 months; range 18-156 m.) of 

the 10 patients overexpressing HER/Neu were available and 

evidenced one relapse (Local and metacronous cancer) in a 

woman only treated with radio therapy, in 5 patients treated with 

herceptin no relapse occurred. Our results suggest that Her2/Neu 

expression could be a significant marker of risk to relapse of disease, 

being a prognostic and predictive factor also in small breast 

carcinoma with pT1a or pT1b, N0, M0. 

references 

Chavez-MacGregor M. HER2-neu positivity in patients with small and 

243 

node-negative breast cancer (pT1a,b,N0,M0): a high risk group? Clin 

Adv Hematol Oncol 2009;7(9):591-8. 

Colleoni M. Minimal and small size invasive breast cancer with no axillary 

lymph node involvement: the need for tailored adjuvant therapies. 

Ann Oncol 2004;15(11):1633-9. 

Curigliano G. Clinical relevance of HER2 overexpression/amplification 

in patients with small tumor size and node-negative breast cancer. J 

Clin Oncol 2009;27(34):5693-9. 

APC molecular alterations in ileal midgut carcinoid 

tumors 

1)Azzoni C. 2)Bottarelli L. 3)Pizzi S. 4)D’Adda T. 5)Tamburini 

E. 6)Rindi G. 7)Silini EM. 8)Bordi C. 

1)Dip patologia e medicina di laboratorio, sez anatomia patologica, Università 

di Parma, Parma, Italia 2)Dip patologia e medicina di laboratorio, 

sez anatomia patologica, Università di Parma, Parma, Italia 3)Dip 

patologia e medicina di laboratorio, sez anatomia patologica, Università 

di Parma, Parma, Italia 4)Dip patologia e medicina di laboratorio, sez 

anatomia patologica, Università di Parma, Parma, Italia 5)Dip patologia 

e medicina di laboratorio, sez anatomia patologica, Università di Parma, 

Parma, Italia 6)Istituto di anatomia patologica, Policlinico universitario 

a. gemelli, Roma, Italia 7)Dip patologia e medicina di laboratorio, sez 

anatomia patologica, Università di Parma, Parma, Italia 8)Dip patologia 

e medicina di laboratorio, sez anatomia patologica, Università di Parma, 

Parma, Italia 

Background. Classical midgut carcinoids are well-differentiated 

neuroendocrine tumors arising from lower jejunum, ileum, 

caecum and ascending colon. Despite recent advances in the diagnosis 

and treatment, no etiologic factors have been associated 

with these tumors, little is known about their molecular features 

and no molecular markers useful for their prognostication have 

been identified. A high frequency of cytoplasmic accumulation 

or nuclear translocation of β-catenin has been described in gastrointestinal 

carcinoid tumors but the role of Wnt pathway in the 

genesis of ileal carcinoid tumors remains unknown. 

Methods. We investigated 30 ileal carcinoid tumors from 14 

male and 16 female patients for loss of heterozigosity (LOH) of 

the APC gene using the microsatellite markers D5S346 (5q21- 

22) and D5S1965 (5q23) and by direct sequencing of the gene 

using four sets of primers amplifying three overlapping portions 

of exon 15. All tumors proved to be composed of EC cells by 

either serotonin immunostaining or Masson-Fontana argentaffin 

reaction. The ileal carcinoids were classified according to the 

WHO criteria (all WDEC class) and ENETS grading and staging 

(grades G1: 24 cases, G2: 6 cases; stages IIA: 6 tumors, IIIA: 1 

tumor, IIIB: 11 tumors and IV: 12 tumors). 

Results. LOH was found in 15% of ileal carcinoids not correlating 

with any clinicopathological feature. APC gene mutations 

were detected in 23% of tumors, in 5 of which mutations were 

also present in the associated metastatic tissues. Moreover, in 

15 (50%) carcinoids the mutational analysis identified a single 

nucleotide polymorphism (SNP) in 1493ACG > ACA (T1493T) 

as demonstrated also by Pizzi et al. in a series of 60 endocrine 

tumours of the gastroenteropancreatic tract. 

Our results indicate that the SNP in the codon 1493 of APC gene 

is commonly found in ileal carcinoids, a finding that requires 

further investigation. The data do not support a relevant role of 

the APC pathway in this type of endocrine tumors. 

ultrarapid immunoistochemistry in intraoperative 

evaluation of sentinel lymph nodes in breast 

cancer 

1)Baldin P. 2)Cucchi M.C. 3)Foschini M.P. 

1)Dipartimento di Ematologia e Oncologia “L e A Seragnoli” Università 

di Bologna Sezione di Anatomia Patologica, Ospedale Bellaria, Bologna, 

Italia 2)U.O. di Chirurgia Oncologica, Ospedale Bellaria, Bologna, Italia 

3)Dipartimento di Ematologia e Oncologia “L e A Seragnoli” Università 

di Bologna Sezione di Anatomia Patologica, Ospedale Bellaria, Italia

244 

Background. In cases of breast cancer, examination of sentinel 

lymph nodes (SNs) is essential to establish the status of regional 

lymph nodes. To avoid the need of two separate surgical resections, 

it is important to develop a reliable method of intraoperative 

examination (IO) of SNs during the excision of the tumour. 

To shorten the technical immunohistochemical procedure it has 

been suggested to employ an ultrarapid immunohistochemical 

method (UICH) for keratins. 

The aim of the present study is to assess whether the use of 

ultrarapid cytokeratin stain (UICH) enhances the intraoperative 

detection of lymph nodal metastases in breast cancer patients 

compared with routine frozen (RF) sections. 

Methods. A consecutive series of 70 cases of IO examination 

of SNs was evaluated with RF and UIHC at the same time. The 

protocol described in Cancer (2005;104:14-9) was followed. In 

addition 100 consecutive cases of SNs were evaluated with RF 

only. All RF sections were compared with the related paraffin 

embedded sections which were subsenquently obtained. Major 

discordance was considered when the difference between intraoperative 

and definitive examination led to a delayed axillary 

dissection. 

Results. In the case group (70 patients), SNs showed tumoral 

involvement in 18 cases (27.7%) (12 macrometastases, 2 micrometastases 

and 4 ITCs). In 65 cases (92,8%) the IO diagnoses 

were similar to those of paraffin sections. In 5 cases paraffin 

sections showed additional neoplastic cells, leading in 3 patients 

(4.2%) to a delayed axillary dissection. The consecutive group 

of 100 patients displayed neoplastic cell in SN in 27 cases (27%) 

(23 macrometastases and 4 micrometastases) with 9 major discordances 

(9%). No false positive cases were detected (100% 

specificity). 

In conclusion UHIC allows a more accurate IO evaluation of SNs 

leading to a lower number of delayed axillary dissections. 

Survey on the quality perceived by internal 

customers on pathology service 

1)Baldoni C. 2)Bondi A. 3)Muraro L. 

1) Anatomia Patologica, Dipartimento Oncologico, Ospedale Maggiore, 

Bologna, Italia 2) Dipartimento Oncologico, Ospedale Maggiore, Bologna, 

Italia 3)Staff aziendale, Ospedale Maggiore, Bologna, Italia 

Background. Evaluation of customer satisfaction in health care 

is a recommended tool in the path of accreditation of healthcare 

facilities. For this reason we develop a questionnaire that would 

allow to highlight the quality of services perceived by customers 

/ users who come to our service. 

Methods. The survey was conducted by administering a questionnaire 

published on the intranet, which could be completed 

online within a month. Respondents were contacted via email 

with a message containing instructions and a link to open the 

questionnaire. Were taken into consideration the following areas: 

access to services, waiting time for answers, information, quality 

and relational aspects. The results of the survey, which was attended 

by hospital and territory doctors and nurse coordinators, 

were collected and processed statistically by a member of OU 

Quality Company. 

Results. The most positive aspects from the perspective of internal 

customers were the quality of services provided and availability 

of pathologists to participate in study groups, clinical audits, 

surveys and general willingness to cooperate shown by all staff. 

The most critical were the response time for histological diagnosis, 

followed by the response time of cytological diagnosis and 

the timing and way of delivery of reports / references. 

Conclusion. This type of check was particularly significant as it 

provided useful information on the expectations, needs and evaluation 

of clinical services allowing to understand and identify critical 

points and to activate the process of continuous improvement 

in the delivery performance. 


Hepatocellular carcinoma induce abnormal 

vascular organization: study on the role of their 

tumor-infiltring stromal cells 

1)Balzarini P. 2)Benetti A. 3)Benerini gatta L. 4)Berenzi A. 

5)Dessy E. 6)Portolani N. 7)Giulini SM. 8)Grigolato P. 9)Alessandri 

G. 

1)2nd department of pathology, school of medicine, P.le spedali civili di 

brescia, Brescia, Italia 2)2nd department of pathology, school of medicine, 

P.le spedali civili di brescia, Brescia, Italia 3)2nd department of pathology, 

school of medicine, P.le spedali civili di brescia, Brescia, Italia 

4)2nd department of pathology, school of medicine, P.le spedali civili di 

brescia, Brescia, Italia 5)2nd department of pathology, school of medicine, 

P.le spedali civili di brescia, Brescia, Italia 6)Department of medical 

and surgical sciences, P.le spedali civili di brescia, Brescia, Italia 7)Department 

of medical and surgical sciences, P.le spedali civili di brescia, 

Brescia, Italia 8)2nd department of pathology, school of medicine, P.le 

spedali civili di brescia, Brescia, Italia 9)Cellular neurobiology laboratory, 

department of ce, Fondazione neurological institute “carlo besta”, 

Milano, Italia 

Backgruond. Hepatocellular Carcinoma (HCC) is one of the 

most common neoplasms worldwide. Unfortunately, conventional 

therapy is not effective and a possible explanation for this 

failure is the abnormal architectural organization of the HCC 

vasculature, which causes poor blood flow and blood stagnation, 

leading to inadequate delivery of chemotherapeutic drugs 

to cancer cells. The aim of this work was to study the phenotypic 

and functional features of stromal cells (StCs) infiltrating HCC 

(HCC-StCs) both in vivo and in vitro and their relationship in 

HCC-induced abnormal neovascularization. 

Methods. Neoplastic and normal liver tissue was obtained from 

20 patients who underwent curative resection of HCC. Histologic 

and immunostaining was performed on formalin fixed and 

embedded paraffin tissue. Cultures of StCs were obtained from 

fresh tissue, neoplastic and adjacent not neoplastic liver sample. 

Monoclonal antibodies (mAbs) used for this study were anti- 

CD105, CD44, CD54, NG2, TGFβ1, TGFβ2, TGFβ3, Smooth 

Muscle Actin (SMA) and PDGF. We analyzed, also, the presence 

of endothelial cells (ECs) markers such as CD31, CD34, Ve-Cad, 

Ang-1 and Ang-2 to evaluate vascular tumor infiltration. 

Results. We defined the immunopathological features of HCC 

biopsies, in particular the grade of malignancy and the rate of 

microvascualr density (MVD). By immunohistochemistry, we 

found that, compared to adjacent not neoplastic liver counterpart, 

HCC-StCs have a reduced expression of mural cell markers NG2 

and SMA both in vitro and in vivo. Moreover, HCC-StCs showed 

a lower expression of the adhesion molecule NCAM, resulting 

in a lower capacity of HCC-StCs to adhere on ECs by using an 

adhesion test in vitro. We conclude that HCC-StCs have lost the 

capacity to interact with ECs and this may concur to produce the 

vascular abnormalities observed in HCC-infiltrating vessels. 

Significance of egfr expression in de novo and 

progressed atypical and anaplastic meningiomas: 

an immunohistochemical and fluorescence in situ 

hybridization study 

1) Barbagallo G.M. 2) Albanese V. 3)Castaing M. 4) Lanzafame 

S. 

1)Dipartimento di Neurochirurgia, Azienda Ospedaliero-Universitaria 

Policlinico OVE, Catania, Italia 2)Dipartimento di Neurochirurgia, Azienda 

Ospedaliero-Universitaria Policlinico OVE, Catania, Italia 3) Dipartimento 

G.F. Ingrassia Istituto di Igiene, Catania, Italia 4) Dipartimento 

G.F. Ingrassia Anatomia Patologica, Azienda Ospedaliero-Universitaria 

Policlinico-OVE, Catania, Italia 

Background. The gene encoding EGFR is located on chromosome 

7. It encodes a 170 kD protein, which is a transmembrane 

receptor responsible for sensing its extracellular ligands, such 

as EGF and TGF-α and for transducting this proliferation sig-


nal. The purpose of this study is to assess the EGFR protein 

expression and the EGFR gene amplification in meningiomas 

of different grade. We investigated whether there is a difference 

in the EGFR protein expression and the EGFR gene amplification 

between the so called de novo malignant meningiomas and 

meningiomas with malignant progression. We also assessed the 

prognostic value of the EGFR expression on overall survival in 

different groups of meningiomas. 

Methods. All cases of meningiomas diagnosed from year 2000 

to 2009 at the Pathology Department of the University of Catania 

were reviewed. Five meningiomas with recurrences and progression 

were selected. They were compared with fifteen meningiomas 

without recurrences. 

Results. The group of G I-II meningiomas without progression 

showed a tendency to a better survival than the group of G I-II 

meningiomas with recurrences and progression. The group of 

G III meningiomas without progression showed a tendency to a 

better survival than the group of G III meningiomas with recurrences 

and progression. The comparison between EGFR expression 

at baseline and after progression have showed an increased 

expression of EGFR protein in the last group. The progression 

from benign to atypical or anaplastic meningiomas may be due to 

the increased expression of EGFR protein. However there was no 

difference in the EGFR expression between the group of G I-II 

de novo meningiomas and the group of G I-II progressed meningiomas. 

The comparison between the group of G III de novo and 

progressed meningiomas and EGFR expression was not statistically 

significant. Our FISH study demonstrated an increase in the 

number of EGFR gene copies in only 1 of the 20 meningiomas. 

eGfr molecular expression, evaluated by 

Immunohistochemistry (IHC) and In Situ 

fluorescent Hybridization (fISH), in lung 

carcinomas 

1)Baron L. 2)Postiglione M. 3)Trombetta C. 4)Maiello F.M. 

5)Quarto F. 

1)S.o.c. di anatomia ed istologia patologica e citop, P.o. san leonardo, 

Castellammare di stabia, Italia 2)S.o.c. di anatomia ed istologia patologica 

e citop, P.o. san leonardo, Castellammare di stabia, Italia 3)S.o.c. di 

anatomia ed istologia patologica e citop, P.o. san leonardo, Castellammare 

di stabia, Italia 4)S.o.c. anatomia patologica, Ospedale dei pellegrini, 

Napoli, Italia 5)S.o.c. di anatomia ed istologia patologica e citop, P.o. san 

leonardo, Castellammare di stabia, Italia 

Background. There is an increasing knowledge of underlying 

molecular mechanisms involving EGFR for targeted lung cancer 

therapies. 

Materials and methods. EGFR overexpression by IHC and 

EGFR gene copy number by FISH were performed on surgical 

histological samples from 49 primary not small cell lung carcinoma 

(NSCLC): 32 adenocarcinomas(ADC),17 squamous cell 

carcinomas(SCC), obtained by casistic of other institution (§). 

Comparisons of the proportions of variables within pathologic 

characteristics were assessed by using χ2 test. 

Results. We were able to detect EGFR overexpression in 18.3% 

of the analyzed tumors (9 cases) and it was more frequent in SCC 

(5 cases, 29.4%) than in ADC (4 cases, 12.5%). 

Non statistically significant differences in degree of differentiation, 

lymph-node status or pathologic stage were seen. 

EGFR amplification by FISH was found in 14.3%, according 

to criteria suggested by Varella-Garcia(Diagnostic Pathology,2006). 

The presence of amplification didn’t correlate with 

any of morphologic parameters analyzed. 

Instead some variables, such as number of EGFR gene copies per 

cell and ratio of EGFR gene to chromosome 7, determined by 

FISH, could be associated to tumor differentiation, lymph-node 

status and tumor stage. 

Out of 7 cases with gene amplification (4 ADC and 3 SCC), 4 

245 

cases (2 ADC and 2 SCC) showed EGFR overexpression by IHC 

too, and 3 cases (2ADC and 1SCC),negative by IHC, had gene 

amplification. 

The level of agreement for EGFR overexpression by IHC and 

EGFR gene amplification by FISH demonstrated a K of 0.74 

(considerable agreement sec. Landis and Koch criteria). 

Conclusions. EGFR protein expression could be couple with 

gene amplification in most cases of NSCLC, although these results 

are based on a single institution experience with a relatively 

small number of patients and so our data need to be verified in a 

larger cohort of patients. 

Primary non-Hodgkin’s lymphoma of ovaries: 

a case report 

1)Barresi E. 2)Schiavo N. 3)Paniccià bonifazi A. 4)Reghellin D. 

5)Rucco V. 6)Lestani M. 

1)U.o.c. anatomia patologica, Ulss 5 “ovest vicentino”, Arzignano (vi), 

Italia 2)U.o.c. anatomia patologica, Ulss 5 “ovest vicentino”, Arzignano 

(vi), Italia 3)U.o.c. anatomia patologica, Ulss 5 “ovest vicentino”, Arzignano 

(vi), Italia 4)U.o.c. anatomia patologica, Ulss 5 “ovest vicentino”, 

Arzignano (vi), Italia 5)U.o.c. anatomia patologica, Ulss 5 “ovest vicentino”, 

Arzignano (vi), Italia 6)U.o.c. anatomia patologica, Ulss 5 “ovest 

vicentino”, Arzignano (vi), Italia 

Background. Primary ovarian lymphoma is an extremely rare 

disease (0.5% of non-Hodgkin’s lymphomas and 1.5% of all 

ovarian neoplasm) and limited count reports about it have been 

recorded in the literature. Usually it is a secondary localization of 

a systemic disease. 

Methods. We describe a case of 36-year-old woman with a left 

ovary mass incidentally discovered (a CT scan performed for 

chronic pelvic pain revealed a tumour). Neoplastic haematic 

markers were negative. A left salpingo-ophorectomy, biopsies 

of right ovary, endometrium and peritoneum were performed in 

laparoscopy, preserving controlateral ovary and uterus. 

Results. On gross examination the ovary measured 

7,5 × 5 × 4,7 cm, with a smooth and thin surface. Neoplasm 

appeared homogeneously solid, white-pink, with areas of necrosis 

on surface of section. Microscopically ovarian tissue was 

replaced by diffuse sheets of mixed medium to large lymphoid 

centroblastic-like cells. These elements, frequently in apoptosis 

or mitoses, are growing in cords or in alveolar-like structures. 

Neither follicular structures nor “starry sky” pattern were observed. 

Immunohistochemically neoplastic cells expressed CD20, 

CD10 and BCL6 (weak); they were negative for CD3, CD5, 

BCL2 and MUM1, with a high proliferate rate (> 90%). In addition, 

neoplasm was negative for primary ovaric neoplastic 

markers (calretinin, inhibin, cytokeratin 7, cytokeratin 20 and placental 

alkaline phosphatase). Ovaric, endometrial and peritoneal 

biopsies were negative. 

Conclusions. Histological and immunohistochemical findings 

revealed a non-Hodgkin diffuse large B-cell lymphoma BCL2- 

and BCL6+. This histotype must be differentiated from B-cell 

lymphoma “unclassifiable”, with features intermediate between 

diffuse large B-cell lymphoma and Burkitt lymphoma. MYC rearrangement 

must be studied in order to better defined lymphoma 

subtype and appropriate therapy. 

Progression of disease in stage I colorectal 

carcinoma: are there histo-prognostic markers? 

1)V. Barresi, 1)R. Lucianò, 1)E. Vitarelli, 1)A. Ieni, 2)L. Reggiani 

Bonetti, 4)C. Di Gregorio, 1)C. Crisafulli, 3)M. Ponz de 

Leon, 1)G. Barresi 

1)Patologia umana, Università di Messina, Messina, Italia; 2)Dip. 

Integrato di Laboratorio, Anatomia Patologica, Az. Universitaria 

Policlinico,Modena, Italia; 3)Medicina e specialità mediche, Università 

di Modena e Reggio-Emilia , Modena, Italia; 4)U.O. di Anatomia patologica, 

Ospedale di carpi, Carpi, Italia

246 

Background. TNM stage I CRC is commonly characterized by 

a good prognosis, with 5-year survival around 80-90%. According 

to most recent protocols, affected patients are only submitted 

to surgical treatments although a percentage of them experience 

disease progression. As a consequence, the identification of prognostic 

markers able to predict the clinical course of stage I CRC 

may be useful in order to select and submit to adjuvant treatments 

those patients with higher progression risk. In view of this, in recent 

studies we tested the eventual prognostic role of the quantity 

of neo-angiogenesis, reflected by microvessel density (MVD), 

of the immunohistochemical expression of the pro-angiogenic 

vascular endothelial growth factor (VEGF) as well as of NGAL, 

an iron-binding protein which is involved in colorectal cancer 

progression, in stage I CRC. 

Methods. MVD as well as VEGF and NGAL immuno-expression 

were analyzed and compared in two subgroups of surgically 

resected stage I CRC: the first included cases obtained from patients 

deceased because of disease progression, whereby the second 

comprised cancers from patients still alive with no evidence 

of disease progression five years after the initial diagnosis. The 

prognostic value of MVD and of VEGF or NGAL expression on 

the overall survival to CRC was investigated. 

Results. NGAL positive cases as well as high MVD counts and 

VEGF expression were significantly more frequent in the CRCs 

from patients deceased of the disease in comparison to those 

from patients alive after five years from surgery. Furthermore, 

NGAL expression as well as high VEGF expression and MVD 

appeared as significant negative prognostic markers related to a 

shorter overall survival to stage I CRC, with VEGF and NGAL as 

independent variables at multivariate analysis. If our preliminary 

results will be further validated, assessment of these markers in 

CRC specimens might be used in order to select those patients 

with a higher progression risk and to submit them to adjuvant 

therapies useful to prevent adverse outcome. 

Performance of fine needle cytology (fNC) 

in Italian breast screening programme 

1)Rossi S. 2)Beccati MD. 3)Nenci I. 

1) Anatomia, Istologia e Citologia patologica, Azienda Ospedaliero-Universitaria 

di Ferrara, Italia” e slittare i numeri delle successive di conseguenza. 

2)Diagnostica citopatologica, Azienda Ospedaliero-Universitaria 

di Ferrara, Ferrara, Italia 3)Anatomia, Istologia e Citologia patologica, 

Università di Ferrara, Ferrara, Italia 

Introduction. The Italian Breast Screening Programme started in 

1996. All data of the target population and the screening-assessment 

process were annually recorded for quality assurance, since 

1997 by GISMa-ONS. Fine needle cytology (FNC) is the most 

diffuse technique for assessment of the breast abnormalities after 

mammography. Core biopsy (CB) were also used. 

Methods. We compared the accuracy for FNC and CB collected 

by the SQTM-Project (Computerized Report for Quality of Diagnosis 

and Therapy for Breast Tumour) by GISMa-ONS, in the 

2007 1 , according to the parameters defined in the European guidelines 

for quality assurance, 2006. We have used the five-point 

classification system for cytology (C1-5) and core biopsy (B1-5). 

We correlated FNC and CB with radiology (R1-5). We calculated 

also the Risk of Malignancy (%) on a series of 1688 FNC performed 

by the Breast-Screening-Programme Ferrara,1997-2006, 

to evaluate if cytology may assist in clinical management. 

Results. SQTM-Project assembled 3136 screen-detected operated 

tumours. The parameters for cytology were (%): AS 66,09; CS 

89,97; PPV C5 99,37, C4 92,33, C3 57,61; FN and FP rate 1,68 

and 0,42, inadequate rate 9,45. The parameters for histology were 

(%): AS 86,41; CS 94,42; PPV B5 99,40, B4 70,96, B3 33,01; 

FN and FP 1,72 and 0,51, miss rate from cancer 5,57. Screening 

Ferrara. The Risk of Malignancy was (%): C1 14,54; C2 2,18; 

C3 32,66; C4 88,84; C5 99,67. 


Conclusions. SQTM-Project. The sensitivity and PPV for cancer 

were high in both techniques. Since cytology is fast, non invasive 

and cost-effective it was the first choice in R4-5 categories. PPV 

for B3 were better than C3. Microhistology was preferable in R3. 

Screening Ferrara. Based on the Risk of Malignancy we propose 

the following clinical management: C1, C2/R4-5, C3/R3 microbiopsy; 

C2/R1-2, screening; C2/R3, microbiopsy or FU; C3/R1-2, 

FU; C3/R4-5 excisional biopsy; C4-5 therapeutic excision and 

sentinel lymphnode. 

references 

1 ONS, Ottavo Rapporto 2009, Ed. M. Zappa. 

ubch10 in cervical intraepithelial neoplasia (CIN) 

as a novel marker of cell proliferation 

1)Bellevicine C. 2)Desiderio D. 3)Varone V. 4)De luca C. 5)Malapelle 

U. 6)Troncone G. 

1)Scienze biomorfologiche e funzionali, Università di Napoli “Federico 

II”, Napoli, Italia 2)Scienze biomorfologiche e funzionali, Università di 

Napoli “Federico II”, Napoli, Italia 3)Scienze biomorfologiche e funzionali, 

Università di Napoli “Federico II”, Napoli, Italia 4)Scienze biomorfologiche 

e funzionali, Università di Napoli “Federico II”, Napoli, Italia 



Napoli “Federico II”, Napoli, Italia 

Backgrounds. Morphological diagnosis of CIN has intraobserver 

and interobserver variability. Ki67 may sometimes be aspecific. 

UbcH10 is a novel proliferation marker. Here we analyzed 

UbcH10 in CIN by RT-PCR and by immunohistochemistry 

(IHC) in relation to Ki-67. 

Methods. Cervical biopsies representative of cervicitis (n = 18), 

CIN I (n = 14), CIN II (n = 14) and CIN III (n = 6) were UbcH10 

and Ki-67 immunostained; a layer(s)-based approach (negative, 

1/3+, 2/3+ and 3/3+) was applied. In addition, UbcH10 RT-PCR 

was also performed on fresh biopsies. 

Results. Most cases of cervicitis were Ki67 (95%) and Ubch10 

(78%) negative. In CIN I, UbcH10 and Ki67 yielded the same 

staining pattern (negative: 42%; 1/3+: 28%; 2/3+: 14%; 3/3+: 

14%). Higher levels of expression were found in CIN II both for 

Ubch10 (negative: 21%; 1/3+: 14%; 2/3+: 64%; 3/3+: 7%) and 

Ki67 (negative: 14%; 1/3+:21%, 2/3+: 50%; 3/3+: 14%). Similarly, 

in CIN III UbcH10 (2/3+: 50%; 3/3+: 50%) and Ki-67 (2/3+: 

17%; 3/3+: 83%) were highly expressed. Consistently, UbcH10 

mRNA levels also increased according to the severity of CIN. 

Conclusion. UbcH10 a both qRT-PCR and IHC levels is useful 

to increase CIN diagnostic accuracy. Its role in cervical cytology 

is currently under investigation. 

IMP3 expression in phyllodes tumours of breast 

1)Bellezza G. 2)Ferri I. 3)Loreti E. 4)Del Sordo R. 5)Colella R. 

6)Sidoni A. 7)Cavaliere A. 

1)Institute of Pathological Anatomy, Perugia University, Perugia, Italy 





6)Institute of Pathological Anatomy, Terni, Perugia University, Italy 7)Institute 

of Pathological Anatomy, Perugia University, Perugia, Italy 

Background. Phyllodes tumours (PTs) of the breast are uncommon 

neoplasms with potential for local recurrence or metastatic 

spread. The WHO classification 1 divided PTs into benign, borderline 

and malignant. However, prognostic assessments based 

solely on histological parameters, can be problematic and many 

biological markers have been proposed. In this study, we investigated 

if IMP3, a member of the insulin-like growth factor II 

mRNA binding protein, was differently expressed in the three 

groups of PTs and could be predictive of behaviour.


Methods. Sixty-two PTs were classified by morphological criteria, 

proposed by WHO, in 40 benign, 13 borderline and 9 malignant. 

Immunohistochemical expression of IMP3 was evaluated 

in stromal neoplastic cells; cases with more than 10% of positive 

cells were considered as positive. Some other variables, including 

surgery, status margin and pathological features, were also 

compared among PTs subgroups. 

Results. Malignant PTs were more frequent in older patients 

(mean: 59 years) and larger in size (mean: 90 mm). Twelve 

patients, who experienced local recurrences (7 benign, 3 borderline 

and 2 malignant), were originally treated mainly by simple 

lumpectomy. In these cases surgical margins were positive more 

frequently (45% of cases) than in non recurrent tumours (26%). 

In 3 malignant cases lymph nodes and lung metastases were also 

seen. IMP3 expression was observed in 9 cases (15%). In benign 

and borderline PTs IMP3 was present respectively in 5% and 15% 

of cases, while in malignant PTs in 56% (p = .001). No differences 

were noted between PTs that did and did not recur, while, interestingly, 

IMP3 expression was higher (50% of cases) in recurrences. 

Conclusions. In conclusion, our study showed that IMP3 could 

be an helpful diagnostic tool to discriminate benign and borderline 

from malignant PTs and its expression in recurrences seems 

to be related with a more aggressive behaviour. 

references 

1 Tavassoli et al., WHO, 2003. 

eosinophlic dysplasia of the cervix uteri: 

morphology and immunostochemical features 

1)Bellisano G. 2)Peer I. 3)Faa G. 4)Ambu R. 5)Tolu G.A. 

6)Kasal A. 7)Antoniazzi S. 8)Vittadello F. 9)Egarter-Vigl E. 

10)Negri G. 

1)U.O. Anatomia Patologica, San Martino, Oristano, Italia / Anatomia 

Patologica - Università di Cagliari, San Giovanni di Dio, Cagliari, Italia 

2)Anatomia Patologica, Ospedale Centrale, Bolzano, Italia 3)Anatomia 

Patologica - Università di Cagliari, San Giovanni di Dio, Cagliari, 

Italia 4)Anatomia Patologica - Università di Cagliari, San Giovanni di 

Dio, Cagliari, Italia 5)U.O. Anatomia Patologica, San Martino, Oristano, 

Italia 6)Anatomia Patologica - Università di Cagliari, Ospedale Centrale, 

Bolzano, Italia 7)Anatomia Patologica, Ospedale Centrale, Bolzano, 

Italia 8)Explora, Ricerca ed analisi statistica, Padova, Italia 9)Anatomia 

Patologica, Ospedale Centrale, Bolzano, Italia 10)Anatomia Patologica, 

Ospedale Centrale, Bolzano, Italia 

Background. Eosinophilic dysplasia (ED) of the cervix uteri is a 

particular kind of dysplasia that retains metaplastic features. The 

aim of this study was to evaluate the biologic potentiality of ED 

using p16ink4a (p16) and HPV-L1 (L1) as markers of HPV-induced 

carcinogenesis 1 . 

Methods. Histological samples from 82 women with a previous 

diagnosis of ED were collected from the archive of the Department 

of Pathology of the Central Hospital of Bolzano, Italy. All 

cases were reviewed using the diagnostic criteria for ED described 

by Ma et al. 2 : 1) lack of normal maturation; 2) relatively 

abundant eosinophilic cytoplasm and distinct cell borders compared 

with conventional HSIL; 3) mildly to moderately increased 

nuclear/cytoplasmic ratio; and 4) dysplastic nuclei showing nuclear 

enlargement, hyperchromasia, variable nuclear membrane 

irregularities, and appreciable nucleoli. Immunohistochemical 

analysis for p16 and L1 was performed on all ED specimens and 

on 31 control specimens with a high-grade Cervical Intraepithelial 

Neoplasia (CIN 2-3) of usual type. 

Results. After revision of the histological samples, features of 

ED were confirmed in 66 out of 82 (81%) samples. The original 

diagnosis was CIN1 in 6 out of 66 cases, CIN 2 in 37 and CIN3 

in 23. In 58 out of 66 (88%) specimens, ED was associated with 

CIN of usual type. Diffuse p16 expression was detected in all 66 

ED, whereas L1 was expressed in 18 out of 66 (27%) cases. L1+ 

ED were most often (67%) associated with an original CIN1 di- 

247 

agnosis. In conclusion, Eosinophilic dysplasia of the cervix uteri 

is frequently associated with CIN of usual type and mostly shows 

a high-grade immunohistochemical pattern (p16+/L1-). However, 

HPV-L1 may be, expressed in some ED (p16+/L1+), with a pattern 

similar to that of still productive low-grade lesions, this could 

indicate a higher tendency to spontaneous regression. 

references 

1 Negri G, Bellisano G, Zannoni GF, et al. p16ink4a and HPV L1 

Immunohistochemistry is Helpful for Estimating the Behavior of 

Low-grade Dysplastic Lesions of the Cervix Uteri. Am J Surg Pathol 

2008;32:1715-20. 

2 Ma L, Fisk J, Zhang R, et al. Eosinophilic Dysplasia of the Cervix: 

A Newly Recognized Variant of Cervical Squamous Intraepithelial 

Neoplasia. Am J Surg Pathol 2004;11:1474-84. 

Haemaobium eggs detection in human bladder 

cancer and sporocysts in snail vectors 

1)Benerini Gatta L. 2)Balzarini P. 3)Cadei M. 4)Castelli F. 

5)Grigolato P. 

1)2nd department of pathology, Spedali civili di brescia, Brescia, Italia 



4)Institute of infectious and tropical desaese, Spedali civili di brescia, 

Brescia, Italia 5)2nd department of pathology, Spedali civili di brescia, 

Brescia, Italia 

Background. Schistosomiasis or bilharzia is a tropical parasitic 

disease caused by blood-dwelling fluke worms of the genus 

Schistosoma. In Burkina Faso the main schistosomes infecting 

human beings are S. haematobium, transmitted by Bulinus snails 

and causing urinary schistosomiasis. Schistosoma haematobium 

eggs play a central role in the development of bladder cancer. 

Investigation of eggs in the urine is the most sensitive and specific 

method for diagnosing active schistosomiasis. But the eggs 

may not be detected in urine during chronic parasitation stages 

and cancer. So, the final diagnosis is based on the presence of 

granulomas or dysplastic cells and schistosoma eggs in the 

submucosa of bladder biopsies. We were interested in set up a 

molecular method in which S. heamatobium eggs were detected 

by immunohistochemistry and polymerase chain reaction, in 

formalin-fixed and paraffin-embedded human bladder cancer or 

snail vectors. 

Methods. A total of four vesicals carcinoma were obtained from 

patients undergoing curative intent surgical resections at the S. 

Camille Medical Centre, Nanorò, Burkina Faso. Bulinus snails 

were collected from transmission site of Nanorò region of the 

Burkina Faso. Immunohistochemistry, polymerase chain reaction 

(PCR) and sequencing of S. haematobium eggs was led in formalin-fixed 

and paraffin-embedded tissues. 

Results. We report four cases of vesical cancer schistosomiasis-related. 

Our data showed that the immunoreaction and amplification 

detection led to correct diagnosis of the specific species of Schistosoma 

in the human cancer. Finally we suggest the use of molecular 

methods in the snail vectors for the detection of sporocysts. 

l1 AND p16 proteins and HPV DNA in the low-grade 

cervical intraepithelial neoplasia (CIN1) 

1)Benerini Gatta L. 2)Berenzi A. 3)Balzarini P. 4)Dessy E. 5)Angiero 

F. 6)Alessandri G. 7)Grigolato P. 8)Benetti A. 





5)Department of pathology, University of milano - bicocca, Milano, Italia 

6)Cellular neurobiology laboratory, Fondazione neurological institute 

“carlo besta”, Milano, Italia 7)2nd department of pathology, Spedali civili 

di brescia, Brescia, Italia 8)2nd department of pathology, Spedali civili 

di brescia, Brescia, Italia

248 

Background. We evaluated the expression of p16, Ki-67, L1 

proteins, and HPV DNA, as molecular markers for diagnosis and 

transforming potentiality of low cervical intraepithelial neoplasia 

(CIN1). 

Methods. Cervical specimens from 72 patients, including 32 

cases of CIN1, 10 of CIN2, 10 of CIN3/CIS, 10 of squamous 

cell carcinoma (SCC), and 10 cases of chronic cervicitis, were 

collected. The expression of p16, Ki-67, L1 antigens was evaluated 

by immunohistochemical methods. The HPV/nested PCR 

method was applied to amplify the HPV/L1 region and high risk 

E6/E7 genome 16-18-33-35-52-58. Catalyzed Signal-Amplified 

Colorimetric DNA In Situ Hybridization (CSAC/ISH) of high 

oncogenic viral risk was also applied. 

Results. Ki-67 and p16 increased linearly from control cases to 

CIN1, CIN2 and CIN3 cases, with a peak in the SCC cases. In 

contrast L1 expression was inversely correlated with malignant 

transformation. We divided CIN1 patients into four groups: 

L1-p16+, L1+p16-, L1-p16-, and L1+p16+ and we combined 

immunohistochemical results with HPV/PCR, L1/PCR and highrisk 

E6/E7 genome and CSAC/ISH data. We found that only the 

L1-p16+ group correlated with malignant transformation (100% 

of CIN2, CIN3 and SCC cases) and was present in the 23% of 

the CIN1. Moreover, 52% of CIN1 cases showed the presence of 

HPV/DNA+. In particular, within L1-p16+ group, in 4 out of 7 

cases there was high risk E6/E7 HPV genome and, in one case, it 

was integrated into host DNA, as confirmed by CSAC/ISH. We 

conclude that in CIN1 patients, the HPV DNA, in particular high 

risk E6/E7 genome, has to be investigated in order to distinguish 

high from low risk oncogenic patient groups. 

Human papillomavirus DNA and p16 protein 

expression in squamous cell carcinoma of the lung 

1)Benerini Gatta L. 2)Dessy E. 3)Berenzi A. 4)Benetti A. 5)Balzarini 

P. 6)Tironi A. 7)Angiero F. 8)Grigolato P. 

1)2nd department of pathology, P.le spedali civili di brescia, Brescia, 

Italia 2)2nd department of pathology, P.le spedali civili di brescia, Brescia, 

Italia 3)2nd department of pathology, P.le spedali civili di brescia, 

Brescia, Italia 4)2nd department of pathology, P.le spedali civili di brescia, 

Brescia, Italia 5)2nd department of pathology, P.le spedali civili di 

brescia, Brescia, Italia 6)2nd department of pathology, P.le spedali civili 

di brescia, Brescia, Italia 7)Pathological anatomy, Università di milano 

bicocca, Milano, Italia 8)2nd department of pathology, P.le spedali civili 

di brescia, Brescia, Italia 

Background. HPV is a small DNA virus that usually infects 

squamous epithelial cells. In malignant transformation of uterine 

cervix, the expression of the E6/E7 viral proteins is associated to 

the alterated expression of p16 protein (a key protein in cell cycle 

regulation). Data on human papilloma virus (HPV) involvement 

in preneoplastic and neoplastic lesions of the lung are limited 

and conflicting. To investigate the role of HPV infection in lung 

tumorigenesis, we studied the expression of p16 protein and the 

relation with the presence of HPV DNA in lung squamous cell 

carcinoma (SCC). 

Methods. 41 cases of formalin fixed and paraffin-embedded human 

lung specimens were obtained from the archives of the 2 nd 

Department of Pathology, Spedali Civili, University of Brescia, 

Italy. 31 cases of lung primary SCC and 10 control cases (non 

neoplastic non squamous specimens), in the study were included. 

Genomic and viral DNA of SCC samples were obtained from the 

paraffin block. DNA was extracted and HPV DNA was detected 

by nested polymerase chain reaction. The expression of p16 protein 

was evaluated by immunohistochemistry. 

Results. Two cases of SCC were positive for HPV PCR. The 

expression of the p16 protein was demonstrated immunohistochemically 

in the same specimens. The presence of HPV DNA 

was correlated to p16 protein expression. The results suggest that 

the HPV DNA might play a pivotal role in development and/or 

progression of a small group of lung SCC. 


Perspective evaluation of proteomic analysis in 

prostate cancer and benign prostatic hyperplasia 

1)Bergamini S. 2)Bellei E. 3)Monari E. 4)Reggiani Bonetti L. 

5)De Gaetani C. 6)Sighinolfi M.C. 7)Micali S. 8)De Stefani S. 

9)Bianchi G. 10)Tomasi A. 

1) Dipartimento Integrato di Laboratori, Anatomia Patologica e Medicina 

Legale, Università di Modena e Reggio Emilia, Modena, Italia 2) Dipartimento 

Integrato di Laboratori, Anatomia Patologica e Medicina Legale, 

Università di Modena e Reggio Emilia, Modena, Italia 3)Dipartimento Integrato 

di Laboratori, Anatomia Patologica e Medicina Legale, Università 

di Modena e Reggio Emilia, Modena, Italia 4) Dipartimento Integrato di 

Laboratori, Anatomia Patologica e Medicina Legale, Università di Modena 

e Reggio Emilia, Modena, Italia 5) Dipartimento Integrato di Laboratori, 

Anatomia Patologica e Medicina Legale, Università di Modena e 

Reggio Emilia, Modena, Italia 6) UO di Urologia, Università di Modena 

e Reggio Emilia, Modena, Italia 7)UO di Urologia, Università di Modena 



e Reggio Emilia, Mo 

Background. The limited sensitivity and specificity of PSA for 

diagnosis of prostate cancer (PCa) highlight the need of more 

predictive diagnostic markers: the proteomic analysis might represents 

a good approach for their discovery and identification. In 

this study, we described a proteomic investigation on serum of 

82 patients. 

Methods. We recruited 28 patients with PCa (Group 1) and 30 

subjects with benign prostatic hyperplasia (BPH) and without 

PCa histologically confirmed, as control (Group 2). The mean 

of the age was 67.0 years (SD ± 6.6) in Group 1 and 67.8 years 

(SD ± 7.0) in Group 2, respectively; the mean of PSA value was 

9.9 ng/ml in Group 1 and 6.2 ng/ml in Group 2. The serum was 

depleted of the 7 high-abundance proteins by Multiple Affinity 

Removal System (MARS HuPL7, Agilent) to permit the detection 

of the low-abundance proteins. The samples proteomic profile was 

obtained using the Surface Enhanced Laser Desorption/Ionization 

Time-of-Flight-Mass Spectrometry (SELDI-TOF-MS). Two different 

types of chromatographic surfaces (ProteinChip) were used: 

the IMAC30 (metal affinity) and the H50 (hydrophobic surface). 

Results. Proteomic analysis has revealed several cluster of peaks 

according to the ProteinChip used. In particolar, the IMAC30 

ProteinChip showed three protein peaks differentially expressed 

(p < 0.05) in BPH compared to PCa (2210, 2929 and 9082 kDa). 

Separating the Group 2 in two different subgroups (BPH with or 

without prostatitis) has emerged that these cluster peaks remained 

differentially expressed among the BPH/prostatitis patients and 

those with PCa. The three protein peaks could therefore selectively 

characterize the presence of PCa. These data are preliminary 

and require additional assessments to confirm the presence of 

differentially expressed proteins. However, the SELDI-TOF-MS 

technique might represents an innovative and promising approach 

for the discovery of potential predictive biomarkers of PCa. 

Her2 testing in gastric cancer. 

immunohistochemistry (IHC) and fluorescence 

in situ hybridization (fISH) comparison 

1)Bettelli S. 2)Fontana A. 3)Losi L. 4)Reggiani Bonetti L. 

5)Bertolini F. 6)Zironi S. 7)Scarabelli L. 8)Luppi G. 9)Conte PF. 

10)Maiorana A. 

1)Dip Integr di Lab, Anat Pat e Med Leg, Az. Ospedaliero Universitaria 

Policlinico, Modena, Italia 2) Dip. Oncologia, Ematol e Mal Respiratorie, 

Az. Ospedaliero Universitaria Policlinico, Modena, Italia 3)Dip Integr 

di Lab, Anat Pat e Med Leg, Az. Ospedaliero Universitaria Policlinico, 

Modena, Italia 4)Dip Integr di Lab, Anat Pat e Med Leg, Az. Ospedaliero 

Universitaria Policlinico, Modena, Italia 5)Dip. Oncologia, Ematol e Mal 

Respiratorie, Az. Ospedaliero Universitaria Policlinico, Modena, Italia 

6)Dip. Oncologia, Ematol e Mal Respiratorie, Az. Ospedaliero Universitaria 

Policlinico, Modena, Italia 7)Dip. Oncologia, Ematol e Mal Respiratorie, 

Az. Ospedaliero Universitaria Policlinico Modena, Italia 8)Dip.


Oncologia, Ematol e Mal Respiratorie, Az. Ospedaliero Universitaria 

Policlinico, Modena, Italia 9)Dip. Oncologia, Ematol e Mal Respiratorie, 

Az. Ospedaliero Universitaria Policlinico, Modena, Italia 10)Dip Integr di 

Lab, Anat Pat e Med Leg, Az. Ospedaliero Universitaria Policlinico 

Background. Overexpression of the HER2 protein in gastric cancer 

have been reported from 6% to 35% of cases. Tumor localization, 

histological type, tumor grading and tumor heterogeneity are 

mostly related to this wide range. 

Methods. From October 2009, we tested HER2 status in 26 

gastric cancers newly diagnosed in the Pathologic Anatomy of 

Modena. IHC was performed in 20 patients, while FISH analysis 

in 16. ICH and FISH were both carried out in 10 patients. 

Results. HER2 protein overexpression was observed in 3/20 

(15%) patients. HER2 amplification was detected in 5/16 (31%). 

Among cases evaluate through both IHC and FISH, 3/10 (30%) 

showed IHC score 0, but high amplification by FISH. Differently 

from what is generally observed in breast cancer, our data showed 

an unexpected discrepancy between FISH and ISH results in the 

assessment of HER 2 status in a single institution analysis of 

gastric cancer patients. 

Muscle spindle and pacinian corpuscle: 

conceptions, misconceptions, and the far-fetched 

hypothesis of an experienced surgical pathologist 

1)M. Bisceglia 2)S. Bisceglia 3)M.L. Bisceglia 

1)Department of Pathology, Division of Anatomic Pathology, IRCCS – 

Casa Sollievo della Sofferenza Hospital, San Giovanni Rotondo, Italy; 2) 

Nursing School, Faculty of Medicine, University of Foggia,Italy; 3) School 

of Pharmacy, University of Parma, Parma, Italy 

Background. The muscle or neuromuscular spindle is a proprioceptive 

microanatomic structure, which together with the Golgi tendon 

organ, is responsible for the reflex arc that determines the tonic state 

of a muscle. It is penetrated by both sensory and motor nerve fibres, 

gamma (mainly) and beta efferents, and therefore has sensory and 

motor functions 1 . The neuromuscular spindles are found in all skeletal 

muscles with facial muscles and (perhaps) diaphragm the only 

exceptions. The Pacinian corpuscle (or corpuscle of Vater-Pacini) is 

a pure mechanoreceptor, responsive to pressure. It is typically found 

in the skin, subcutis and superficial soft tissues, but uncommonly 

may also be seen in the soft tissue of body cavities and in the serosa 

and subserosa of visceral organs. 1 Very rarely the Pacinian corpuscle 

may be found in skeletal muscle, chiefly in relation to fascia 

and aponeuroses, and, when found in skeletal muscle, the Pacinian 

corpuscle is intimately related to the neuromuscular spindle. Usually 

both muscle spindle and Pacini corpuscle are too unremarkable 

microanatomical findings to focus on during the course of routine 

work in surgical pathology. 

Objectives. i. To report on the incidental finding of a curious 

muscle spindle, the “fibrous” capsule of which mimicked the “lamellar” 

body of Pacinian corpuscle. ii. To describe the sequence of 

events leading to the correct recognition of the muscle spindle. iii. 

To emphasize the fundamental anatomical notions ignored by the 

pathologist; iv. To list the pathological conditions, partly theoretical, 

which can affect the two aforesaid microanatomic structures. 

Materials. During the microscopic examination of a resection 

margin of a skeletal muscle surgical resection specimen harbouring 

a capillary haemangioma, removed from the thigh of a 

42-year old male, a structure which at first glance appeared to 

be a neuromuscular spindle was noted by the pathologist (MB), 

an unremarkable finding in that context. Two students, one (SB) 

who was prepared to take his anatomy exam at the completion 

of the first year of nursing school, and the other (MLB), in her 

last year of pharmacy school, were asked to identify that microscopic 

structure. Both students answered that the structure 

under the microscope was a Pacinian corpuscle. Their concordant 

answers provoked the testing pathologist to scrutinize the slide 

more closely and come to suspect that this muscle spindle with a 

249 

“lamellar” fibrous capsule was likely to be a “hybrid” structure, 

in other words a new finding, specifically a composite structure 

comprised of a peripherally located Pacinian corpuscle wrapped 

around a true central neuromuscular spindle. The suspicion became 

convincing since nowhere in any of the other sections from 

the entire surgical specimen was a similar structure found nor had 

the pathologist ever seen something similar, despite having some 

experience in musculoskeletal and neuromuscular pathology. 

Methods. A true Pacinian corpuscle from an archival skin resection 

specimen was examined and immunostained, with the 

following expected results: the “onion skin-like” lamellar body 

of the Pacinian corpuscle was EMA-positive; the sensory nerve 

fibre penetrating the lamellar body was neurofilament-positive; 

and the Schwann cells enveloping the nerve fibre axon were 

S-100 positive. The new finding (thought to be chimeric/composite 

muscle Pacinian spindle), was also immunostained: the 

intrafusal fibres of the spindle were obviously desmin positive; 

the “lamellar” sheath, namely what was supposed to be the 

wrapping Pacinian corpuscle, was successfully EMA positive 

and CD34 negative; neurofilament stained d axons in the center 

of the spindle. 

Discussion. In essence, in transverse section, the muscle spindle 

normally measures 200 µm and is made of skeletal muscle microfibres 

(variably 5 to 14 in number) surrounded by a “fibrous” 

capsule made of 9 to 15 concentric, usually tightly arranged, 

layers of flattened epithelial-like cells (also called “capsular 

sheet cells”). This “fibrous” capsule represents an extension of 

the perineurium enclosing the nerve fibres serving the intrafusal 

muscle fibres 1 2 . The imaginary hypothesis which was construed 

to support fusion of the Pacinian corpuscle and muscle spindle 

was based on the misconception that the “fibrous” capsule was 

made of EMA negative ordinary fibrous connective tissue. 

Actually this “fibrous” capsule of the muscle spindle can be 

confidently equated to the terminal perineurium ensheathing the 

peripheral nerve twigs, made of EMA positive cells. The Vater- 

Pacini corpuscle, normally measuring up 2 mm in length, is made 

of a lamellar body and a small central core, the former made of 30 

or more concentric loosely arranged lamellae, composed of flat 

perineurial cells, the latter containing the terminal non-myelinated 

sensory nerve fibre. The neuromuscular spindle may normally 

vary in size and number, but in some circumstances they 

also vary in arrangement, appearing in tandem, even sharing a 

common capsule, and in groups. The intrafusal fibres are affected 

in some neuromuscular diseases (myotonic dystrophy for splitting, 

poliomyelitis for rarifying), or appear pseudohypertrophic 

in others as in Werdnig-Hoffman disease (in comparison with the 

extrafusal fibres) 3 , and were supposed to be the tissue from which 

alveolar soft part sarcoma arises 4 . The “fibrous” capsule may 

become thickened as in ageing or cellular, fibrosed or edematous 

as in Duchenne dystrophy 3 . In our case the muscle spindle was 

oversized (350 to 400 µm in size after several measurements 

on transverse sections), and exhibited a thickened capsule with 

separated lamellae encroaching on the periaxial space, which is 

normally present between intrafusal fibres and the capsule: we 

could not ascertain whether this was only anatomical variation 

or due to some unknown cause (?trauma). The Vater-Pacini corpuscles 

may also normally vary in size and number according to 

specific anatomical locations, and may also appear hyperplastic 

(“Pacinian corpuscle hyperplasia” 5 ) or simulate neoplastic conditions 

(Pacinian neuroma, Pacinian neurofibroma o Pacinian 

perineurioma). Finally, at least in theory, one cannot exclude that 

perineuriomas might arise from EMA positive perineurial cells 

either of the muscle spindle capsule or the lamellar body of the 

Vater-Pacini corpuscle. 

references 

1 Standring S. Gray’s Anatomy. 40 th Edition, Churchill Livingstone 

2008, pp. 59-60 

2 Banks RW, Barker D. The Muscle Spindle. In: Engel AG, Franzini-

250 

Armstrong C (eds). Myology. Third Edition. New York: McGraw-Hill, 

2004, pp. 489-509. 

3 Swash M. Pathology of the muscle spindle. In: Mastaglia FL, Walton 

J (eds). Skeletal Muscle Pathology. Edinburgh: Churchill Livingstone 

1982, pp. 508-36. 

4 Christopherson WM, Foote FW Jr, Stewart FW. Alveolar soft-part sarcomas; 

structurally characteristic tumors of uncertain histogenesis. 

Cancer 1952;5:100-11. 

5 Reznik M, Thiry A, Fridman V. Painful hyperplasia and hypertrophy 

of Pacinian corpuscles in the hand: report of two cases with immunohistochemical 

and ultrastructural studies, and a review of the 

literature. Am J Dermatopathol 1998;20:203-7. 

Glomus tumor of stomach. report of 8 cases 

and review of the literature. 

1) Bisceglia M. 2) Bleiweiss I. 3)Ben Dor D. 4) Magro G. 5) 

Sickel J. 6) Carosi I. 7) Miettinen M. 

1)Department of Pathology, IRCCS Casa Sollievo della Sofferenza Hospital, 

San Giovanni Rotondo, Italy 2)Department of Pathology, Mount 

Sinai School of Medicine, New York, NY, USA 3)Department of Pathology, 

Barzilai Medical Center, Ashkelon, Israel 4)Department of Pathology, 

University of Catania, Catania, Italy 5)Department of Pathology, El Camino 

Hospital, Grant Road Mountain View, CA, USA 6) Department of 

Pathology, IRCCS Casa Sollievo della Sofferenza Hospital, San Giovanni 

Rotondo, Italy 7)Department of Soft Tissue Pathology, Armed Forces Institute 

of Pathology, Washington, DC, USA. 

Background. Glomus tumors (GT) are thought to originate from 

glomocytes. Glomocytes are round, distinct, epithelioid cells with 

ultrastructural and immunohistochemical features of modified 

smooth muscle cells, functioning as sphincters of Hoyer-Sucquet 

canals of glomera. Glomera represent normal arteriovenous shunts, 

abundantly supplied with nerve fibers that act as regulators of temperature 

in several locations throughout the body. The most common 

locations of glomera are the skin and peripheral soft tissue of 

the distal parts of extremities. However, they are also encountered 

in large cavities and visceral organs, including the alimentary tract, 

where their function is related to absorption. Paralleling the usual 

distribution of glomera, GT are most common in the skin and soft 

tissue of acral sites. GT of the stomach is a very rare neoplasm that 

was first recognized by Saul Kay et al. in 1951, who reported 3 

cases at that time 1 . Since then, most published cases have appeared 

as single case reports with very few series on record. 

Objectives. 1. To present our personal cases of GT of stomach. 

2. To comprehensively review the English literature on this 

subject and document the total number of gastric GT reported 

so far. 

Methods. 1 A systematic search of our combined databases was 

performed to identify cases of gastric GT. 2. A computerized 

literature search of PubMed/Medline was performed between 

1951 and April 2010 using [glomus tumor of stomach] as a search 

term. 

Results. Analysis of cases. 8 original (unpublished) cases were 

found in our institutional files: 6 cases were males and 2 cases 

were females, all were adults or elderly. The ages of the males 

ranged between 38 and 81 (with the others 54, 62, 67, 79 in 

between); the two females were 48 and 55, respectively. Gastric 

bleeding was the presenting symptom in 7 out of 8 cases 

(hematemesis in 5, melena 2). The tumors ranged in size from 

1.5 to 8 cm. The majority of tumors were located in the antrum 

(4); 1 tumor was in the body, 1 was in the fundus, and in 2 the 

site was not recorded. Mucosal ulceration was seen in 6 cases. 

Either leiomyoma or lipoma was the preoperative diagnosis in 

4 cases. All tumors were surgically treated: partial gastrectomy 

in 4, wedge resection in 3, lumpectomy in 1. All tumors were 

intramural, well circumscribed and confined to the muscularis 

propria of the organ, except for one, the largest, which was locally 

invasive. All tumors were histologically benign (very low mitotic 

index, absence of necrosis, no cytological atypia, no spindling), 


except for one, the largest, which exhibited high mitotic rate, 

coagulation necrosis, and nuclear atypia, and was diagnosed as 

malignant. The morphological pattern was predominantly classic 

solid glomus tumor, with associated glomangiomatous areas in 3. 

Immunohistochemically: vimentin and alpha-SMA were positive 

in all; cytokeratins, CD117, and neuroendocrine markers were 

always negative. Follow-up: 6 patients are alive with no evidence 

of disease (follow-up ranging from 3 to 18 years; median 8), 1 

patient (the eldest) shortly died after surgery, the ma1ignant case 

was lost to follow-up. Review of the Literature. To date 104 GT 

have been recorded in the English Literature since 1951. Only 

4 papers included more than 1 case 1-4 . The two largest series 

were compiled in 1969 by Appelman and Helwig 3 and in 2002 

by Miettinen et al. 4 , both from the Armed Forces Institutes of 

Pathology (Washington, D.C.), who reported 12 and 31 cases, 

respectively. In most cases the tumor was solitary, but in 4 cases 

multiple tumors were described. Most GTs are histologically 

benign, but 3 malignant cases have been published. 

Discussion. GTs most often occur in the gastric antrum of adults, 

without any sex predilection. The signs and symptoms can be 

variable: bleeding due to surface ulceration is a common finding, 

and the bleeding can be quite profuse due to the extensively vascularized 

nature of the lesion, leading in some instances to anemia 

or to emergency surgical gastrectomy; other frequent clinical 

manifestations are pain, nausea and vomiting. The majority of 

the lesions are solitary, although multiple gastric glomus tumors 

have been described 3 . The main differential diagnosis is with epithelioid 

GIST and carcinoid. Analogous to their soft tissue counterpart, 

large size, high mitotic index (≥ 5M:50HPF), cytologic 

atypia including spindling, necrosis, and local infiltrative growth 

are all possible histologic indicators of malignancy 5 in GT of the 

stomach. Additionally we note that, in the experience of one of us 

(MM), one gastric GT with limited atypia (spindling present) and 

mitotic rate < 5/50 metastasized and killed the patient 4 . 

Conclusions. 1. GT of stomach is rare (according to Miettinen 

et al. 4 < 1% compared to GIST). 2. Most gastric GT are benign. 

3. Malignant GT of stomach is extremely rare, but may occur. 

4. Smooth muscle differentiation is a constant immunohistochemical 

finding. 

references 

1 Kay S, Callahan WP Jr, Murray MR, et al. Glomus tumors of the stomach. 

Cancer 1951;4:726-36. 

2 Allen RA, Dahlin DC. Glomus tumor of the stomach: report of 2 cases. 

Proc Staff Meet Mayo Clin 1954;29:429-36. 

3 Appelman HD, Helwig EB. Glomus tumors of the stomach. Cancer 

1969;23:203-13. 

4 Miettinen M, Paal E, Lasota J, et al. Gastrointestinal glomus tumors: a 

clinicopathologic, immunohistochemical, and molecular genetic study 

of 32 cases. Am J Surg Pathol 2002;26:301-11. 

5 Folpe AL, Fanburg-Smith JC, Miettinen M, et al. Atypical and malignant 

glomus tumors: analysis of 52 cases, with a proposal for the 

reclassification of glomus tumors. Am J Surg Pathol 2001;25:1-12. 

lipofuscin-like granules of the juxtaglomerular 

apparatus of the kidney. The diagnostic 

significance of a quasi-normal subcellular 

structure only incidentally encountered in the 

course of routine ultrastructural evaluation of 

renal biopsies for diagnostic purposes 

1) Bisceglia M. 2) D’Errico M. 3) Carosi I. 4) Grasso M.A. 

5)Pasquinelli G. 

1)Unit of Anatomic Pathology, IRCCS Casa Sollievo della Sofferenza Hospital, 

San Giovanni Rotondo, Italy 2)Unit of Nephrology, IRCCS Casa 

Sollievo della Sofferenza Hospital, San Giovanni Rotondo, Italy 3) Unit of 

Anatomic Pathology, IRCCS Casa Sollievo della Sofferenza Hospital, San 

Giovanni Rotondo, Italy 4)Hospital Pharmacy Program, School of Pharmacy, 

“La Sapienza” University, Rome, Italy 5)Department of Clinical 

Pathology, University of Bologna, Italy


Background. In a systematic study of 114 human kidney tissue 

specimens, from patients ranging in age from 2 to 70, in 1965 Biava 

and West reported the light and electron microscopic features 

of a particular type of “lipofuscin-like granule” mainly in the 

cytoplasm of vascular smooth muscle cells, but also in juxtaglomerular 

cells, and (more rarely) the lacis cells of human kidneys 1 . 

The granules were found in all cases, including healthy kidneys, 

but their number correlated with age, arterial hypertension and 

diabetes mellitus, and were larger in diabetic than in non-diabetic 

patients. Neither the awareness of the existence of this finding 

nor its clinical significance is widespread among either pathologists 

or nephrologists. Standard nephropathology textbooks do 

not even mention these structures either in the normal anatomy 

section nor in any other specific pathology chapter. The afore 

mentioned lipofuscin-like granules were incidentally observed 

in 4 cases during the routine electron microscope examination of 

440 renal biopsies performed at Casa Sollievo della Sofferenza 

hospital between 1991 and 2009, for investigation of medical 

nephropathies. 

Objectives. 1. To review the world literature to find out how 

many articles dealing with this particular type of granule in the 

context of renal pathology were on record. 2. To report the ultrastructural 

features of this finding. 3. To see if systemic arterial 

hypertension and/or the status of diabetes mellitus were present 

also in our cases as originally demonstrated by Biava and West 

45 years earlier. 4. To correlate the significance, if any, of these 

granules with the specific renal disease for which kidney biopsies 

were performed. 

Materials and Methods. 1. A comprehensive PubMed-Medline 

search was performed between 1962 and May 2010 using widely 

several search terms, including lipofuscin, lipofuscin-like granules, 

and fingerprint in the context of the kidney. 2. The clinical 

histories and biochemical data as well as the electron microphotographs 

of these cases were retrospectively reviewed with regard 

to both the key status-symptoms of hypertension and diabetes and 

to the specific primary renal disease leading to biopsy. 

Results. 1. Only two articles (1 Russian, 1 Japanese), other than 

the original one by Biava and West 1 , mentioning lipofuscin-like 

granules, were found. 2. Our cases included 2 males and 2 females. 

All patients were adults and their ages were 67, 66, 55, 69, 

respectively. All renal biopsies were studied by immunofluorescence, 

light microscopy and electron microscopy. The pathological 

diagnoses were the following: minimal change disease, focal 

segmental glomerulosclerosis, idiopathic membrano-proliferative 

glomerulonephritis type I, and minimal change disease, respectively 

in the same order. The corresponding suspected clinical 

diagnoses were: immunologic glomerulonephritis-NOS vs renal 

amlyoidosis, chronic glomerulonephritis-NOS, lupus nephritis, 

and membranous glomerulonephritis vs amyloidosis, respectively. 

Two patients had moderate proteinuria, and two had nephrotic 

syndrome. In no case the clinical information of systemic hypertension 

was given to the pathologist prior to pathological biopsy 

examination. Though the retrospective review of the clinical 

charts revealed that all patients were affected by this condition, 

one in association with diabetes mellitus. All patients had other 

associated systemic diseases (rheumatoid arthritis, diabetes mellitus, 

Sjögren syndrome), except for one (the 69-year old female). 

Discussion. In the work by Biava and West the lipofuscin-like 

granules were found – in decreasing order of frequency – in 

the smooth muscle cells of the afferent glomerular arteriole, in 

myoepithelioid juxtaglomerular cells (along with different specific 

renin-containing granules), and in lacis cells 1 . They also 

noted that these granules are of 0.5 to 4.0 µ in size and visible at 

light microscopy, being argyrophilic, and PAS-positive diastase 

resistant 1 . We did not directly search for fingerprint profiles 

in juxtaglomerular apparatus elements either during electron 

microscopical examination, or in examination of standard histological 

sections. Although these are eye-catching findings, they 

251 

have always been incidental, and although we were aware of the 

existence of these structures as a nonspecific finding in renal arterioles 

from citation in a neuropathology paper on Kufs’ disease, 

due to the interest in this subject matter of one of us (GP), regrettably 

we admit to never having given them any specific clinical 

significance. Instead Biava and West attributed to them a relative 

clinical significance, due to their increased number or size in 

arterial hypertension and/or diabetes mellitus, respectively 1 . In 

electron microscopy in our cases, as in the keystone and seminal 

paper by Biava and West, these lipofuscin-like granules appear as 

dense bodies with a lipid component, a coarsely granular matrix, 

and a crystalloidal component which may appear in band or dot 

pattern, according to the plane of sectioning. The band pattern of 

these crystalloids is homologous to the fingerprint profiles seen 

in other diseases such as neuronal ceroid-lipofuscinosis or the 

semicircularly organized (fingerprint) linear immune deposits 

seen in the above mentioned glomerulopathies. Parenthetically, 

but noteworthy, fingerprint profiles have also been observed by 

one of us (GP) in the smooth muscle cells of the arterioles in a 

rectal mucosal biopsy in a case of neuronal ceroidolipofuscinosis. 

Although not in the scope of this report, another open question 

concerning these structures is the patho-physiologic mechanism 

of their formation. The answer cannot be other than hypothetical, 

and is likely due to either oxidative damage to cytosolic structures 

or mitochondrial oxidative stress, possibly related to the continual 

adrenergic nervous stimulation in connection with blood flow and 

to ageing-related impairment of their proteasome processing systems; 

however for this mechanism we refer to specialized papers 

addressing this matter. 

Conclusions. 1. Lipofuscin-like granules are subcellular, quasiphysiologic, 

finding mainly in smooth muscle cells of the walls of 

renal arterioles, which increases in number and/or size in subjects 

affected by arterial hypertension and diabetes. 2. They do not correlate 

with a specific primary renal disease. 3. The pathologist has 

to be aware of these lipofuscin-like granules in order not to confuse 

them with other similar findings having a specific diagnostic 

significance (such as the fingerprint organized immune deposits 

associated with specific glomerulopathies). 4. The nephrologist 

should always be alert for either treated or untreated clinical arterial 

hypertension in their patients and inform the pathologist as 

such. 5. Additional systematic observations are needed in order 

to further our understanding of these almost completely neglected 

subcellular structures. 

references 

1 Biava C, West M. Lipofuscin-like granules in vascular smooth 

muscle and juxtaglomerular cells of human kidneys. Am J Pathol. 

1965;47:287-313. 

Solitary fibrous tumor of the meninges. literature 

review with a report of 5 additional cases 

1)Bisceglia M. 2)Dimitri L. 3)Carotenuto V. 4)Bianco M. 5)Monte 

V. 6)Giannatempo G. 7)D’Angelo V. 

1)Unit of Anatomical Pathology, IRCCS “Casa Sollievo della Sofferenza” 

Hospital, San Giovanni Rotondo, Italy 2)Unit of Anatomical Pathology, 

IRCCS “Casa Sollievo della Sofferenza” Hospital, San Giovanni Rotondo, 

Italy 3)Unit of Neurosurgery, IRCCS “Casa Sollievo della Sofferenza” 

Hospital, San Giovanni Rotondo, Italy 4) Unit of Neurosurgery, IRCCS 

“Casa Sollievo della Sofferenza” Hospital, San Giovanni Rotondo, Italy 

5) Unit of Neurosurgery, IRCCS “Casa Sollievo della Sofferenza” Hospital, 

San Giovanni Rotondo, Italy 6)Unit of Radiology, IRCCS “Casa 

Sollievo della Sofferenza” Hospital, San Giovanni Rotondo, Italy 7) Unit 

of Neurosurgery, IRCCS “Casa Sollievo della Sofferenza” Hospital, San 

Giovanni Rotondo, Italy 

Background. Solitary fibrous tumour (SFT) is a spindle cell mesenchymal 

tumor first described in the pleura by Klemperer and 

Rabin in 1931 as a distinct pathologic entity (solitary fibrous mesothelioma, 

submesothelial fibroma). Suster et al. were the first

252 

to report the occurrence of these tumors in extrapleural location, 

specifically in the somatic soft tissues 1 . A number of cases in 

many different locations have been added to the literature since 2 , 

and in 1996 the first seven meningeal cases were reported, five of 

which were intracranial and two intraspinal, respectively 3 . SFT 

is thought to originate from the almost ubiquitous CD34 positive 

fibroblast (“dendritic interstitial cell”), which has also been identified 

in the dural tissue 4 . 

Objectives. 1. To comprehensively review the world literature 

concerning SFT involving the central nervous system (CNS). 

3. To report 5 personal additional cases. 2. To ascertain the frequency 

of this tumor as a proportion of all primary meningothelial 

and non-meningothelial meningeal-based mesenchymal tumors 

in our 17 years experience at the Casa Sollievo della Sofferenza 

hospital in S. Giovanni Rotondo. 4. To briefly discuss its distinction 

from hemangiopericytoma (HPC) of the meninges. 

Materials and Methods. 1. A comprehensive PubMed-Medline 

search was performed between 1996 and May 2010, using 

[central nervous system AND solitary fibrous tumor], [meninges 

AND solitary fibrous tumor], [brain AND solitary fibrous tumor], 

[intraventricular AND solitary fibrous tumor], [medullary cord 

AND solitary fibrous tumors], and [intramedullary AND solitary 

fibrous tumors] as search terms. 2. A systematic search of our 

database was performed to retrieve all meningeal-based primary 

mesenchymal tumors from our files. 3. To review the clinical 

charts and to follow-up the affected patients. 

Results. Literature Review. In 2004 Caroli E et al. found 56 previously 

reported cases of CNS SFT to which they added 4 cases 

of their own 5 , and in 2009 Mekni et al. independently reviewed 

the same subject and added 8 cases of their own, the number of 

cases reported to 2007 thus totalling 96 6 . By using all the afore 

mentioned search terms, around 150 cases have been found on 

record. Most CNS SFT were dural-based, but a significant proportion 

(≅ 15%) of other CNS locations not directly attached to 

the dura (intraventricular, intramedullary, cerebello-pontine, and 

subpial-intracerebral, in descending order of frequency) are also 

on record A few cases (≅ 10%) exhibited histological malignant 

features either at presentation or upon recurrence. 

Frequency of SFT in our files. Out of 806 meningeal-based 

primary mesenchymal tumors retrieved in total, 786 were meningiomas, 

15 hemangiopericytomas, 5 were SFTs, of which 4 

intracranial and 1 intraspinal, respectively, and 2 were meningeal 

sarcoma unspecified. 

Personal SFT Case Reports. All cases were surgically operated, 

with or without adjunctive radiation therapy. Case 1: male, aged 

47, with an intraspinal (T3-T4) tumor 2 cm in size; the tumor was 

totally excised grossly; the patient is alive with no evidence of 

disease (ANED) at 11 years. Case 2: male, aged 75, with a tumor 

in the posterior cranial fossa (PCF) 3.5 cm in size; the tumor was 

grossly incompletely excised and treated with adjunctive radiosurgery 

(gamma knife); the patient experienced 2 recurrences at 4 and 

7 years, which were treated with surgery and radiotherapy, respectively, 

and eventually – after partial response to radiotherapy – he 

died of disease 10 years after diagnosis at the age of 85. Case 3: 

male, aged 64, with a PCF tumor 4.5 cm in size; the tumor was 

surgically totally excised; the patient is ANED at 5½ years. Case 

4: male, aged 76, with a second tumor recurrence 6.5 in size cm in 

the PCF (the primary and first recurrence were surgically resected 

in an outside institution 15 and 7 years earlier and diagnosed as 

fibrous meningioma and SFT, respectively – the slides were not 

available for review); the recurrence was grossly totally resected; 

eventually the patient died 26 years after the first surgical operation. 

Case 5: female, aged 59, with a PCF tumor of 4 cm, which 

was grossly totally removed; the patient is ANED at 3 years. 

Intraoperative findings. All tumors were dural-based. Histological 

descriptions. All tumors were diagnosed as classical SFT, 

except case 4 (myxoid variant). Immunohistochemistry. All cases 

showed the classical immunoprofile: Vimentin, CD34, CD99, 


BCL2 were all diffusely positive in all cases, except for CD34 

in case 4 which was patchily positive; S-100, EMA, alpha SMA, 

and desmin all were negative; the mitotic index was very low 

(< 1M/10HPF) in all cases, except in case 4 (2M/10HPF); the 

MIB-1/Ki67 labeling index was very low (< 2%) in all, except 

in case 4 (10%). 

Discussion. CNS-SFT is a tumor of adulthood, though occasionally 

seen in the pediatric population. It is mostly dural-based but 

non-dural-based occurrences (intraparenchymal or intraventricular) 

have also been recorded. The majority of CNS-SFT are intracranial, 

but one fourth of the cases are intraspinal. Some intraspinal 

tumors may also arise from the spinal nerve roots, as in our case 

1. Preoperatively they are mostly often diagnosed as meningioma; 

intraspinal tumors may also be diagnosed as neurinoma, as in 

our case 1. Grossly they are usually well circumscribed, but may 

infiltrate into the brain, nerve roots and even skull base. Histologically 

SFT is comprised of short spindle cells mostly arranged in a 

“patternless pattern” (ordinary form), but occasionally organized 

in fascicles, with alternating bands of eosinophilic collagen. Similarly 

to its somatic soft tissue counterpart, morphological variants 

(epithelioid, cellular, and myxoid 7 ) of CNS SFT have also been 

observed. Our case 4 was characterized as SFT myxoid variant. 

The classic cell morphology is usually bland, but – analogous to 

soft tissue – anaplastic variants are also on record, with necrosis, 

nuclear atypicalities, high mitotic rate (> 4M:10HPF), and high Ki- 

67/MIB-1 index. In the third tumor recurrence of our case 4 a high 

Ki-67/MIB-1 index was seen along with a few mitotic figures, but 

necrosis or frank nuclear atypia were absent. SFT is a pathological 

entity which needs to be distinguished from other dural-based 

neoplasms, mostly (fibrous) meningioma, neurinomas, and HPC 8 9 . 

The correct diagnosis is usually made based on light microscopic 

features and the characteristic immunoprofile: CD34+/EMA- 

/S100- (adjunctive markers usually also positive are BCL2 and 

CD99), as distinguished from meningioma (usually EMA+/CD34- 

/S100-), and neurinoma (usually S100+/EMA-/CD34-). HPC 

exhibits an immunoprofile similar to SFT, but CD34 is expressed 

only in a minority of cases and in a weak and patchy pattern 8 . 

Recently some authors have suggested that CNS HPC should be included 

in the spectrum of SFT (cellular SFT) 10 , but this view is not 

shared by others 8 9 11 , since CNS HPC is a well-defined CNS entity, 

despite controversy regarding its histogenesis. CNS HPC shows a 

higher local recurrence rate, and more tumor-related deaths and 

extracranial metastases. In a very recent study 12 , comparing the 

biological behaviour of CNS HPC with that of soft tissue SFT (now 

in this setting by definition inclusive of soft tissue HPC) a noticeable 

clinical difference has been noted, with CNS HPC having a 

recurrence rate reaching > 92%. Thus although in soft tissues HPC 

has almost disappeared as category separate from SFT, in regards 

to CNS the SFT category is still kept separate from it 12 . However 

difficult cases to discriminate or even transitional (from primary to 

recurrent) cases between the two are acknowledged 8 10 . From the 

literature the biologic behaviour is usually benign, provided total 

tumor resection is accomplished, but follow-up data are limited 

and they need still be taken cautiously 5 . However, in comparison 

with its soft tissue counterpart, more aggressive examples (in terms 

of recurrence) have been seen in the CNS, but this is due to their 

frequent incomplete surgical excision. Extracranial metastases are 

extremely rare in CNS-SFT. Our case 2 recurred three times but the 

surgical excision of tumor had been grossly incomplete and surgery 

was not repeated. Our case 4, who sustained 2 recurrences and 

whose tumor eventually developed aggressive histological features 

(mitoses, and high Ki-67/MIB1 labeling index), died of tumor after 

26 years from primary tumor presentation. 

Conclusions. CNS SFT is a tumor distinct from fibrous meningioma 

and HPC. From the literature the behaviour appears to be 

generally benign, but recurrences have been recorded. Surgery is 

the treatment of choice, and tumor regrowth is to be anticipated 

when removal is not complete. The usefulness of radiotherapy


is not well documented. Long-term follow-up of the patients is 

always mandatory. 

references 

1 Suster S, Nascimento AG, Miettinen M, et al. Solitary fibrous tumors 

of soft tissue. A clinicopathologic and immunohistochemical study of 

12 cases. Am J Surg Pathol 1995;19:1257-66. 

2 Chan JK. Solitary fibrous tumour-everywhere, and a diagnosis in 

vogue. Histopathology 1997;31:568-76. Mod Pathol 1999;12:463-71. 

3 Carneiro SS, Scheithauer BW, Nascimento AG, et al. Solitary fibrous 

tumor of the meninges: a lesion distinct from fibrous meningioma. A 

clinicopathologic and immunohistochemical study. Am J Clin Pathol 

1996;106:217-24. 

4 Cummings TJ, Burchette JL, McLendon RE. CD34 and dural fibroblasts: 

the relationship to solitary fibrous tumor and meningioma. 

Acta Neuropathol 2001;102:349-54. 

5 Caroli E, Salvati M, Orlando ER, et al. Solitary fibrous tumors of the 

meninges: report of four cases and literature review. Neurosurg Rev 

2004;27:246-51. 

6 Mekni A, Kourda J, Hammouda KB, et al. Solitary fibrous tumour 

of the central nervous system: pathological study of eight cases and 

review of the literature. Pathology 2009;41:649-54. 

7 de Saint Aubain Somerhausen N, Rubin BP, Fletcher CD. Myxoid 

solitary fibrous tumor: a study of seven cases with emphasis on differential 

diagnosis. Mod Pathol 1999;12:463-71. 

8 Perry A, Scheithauer BW, Nascimento AG. The immunophenotypic 

spectrum of meningeal hemangiopericytoma: a comparison with fibrous 

meningioma and solitary fibrous tumor of meninges. Am J Surg 

Pathol 1997;21:1354-60. 

9 Tihan T, Viglione M, Rosenblum MK, et al. Solitary Fibrous Tumors 

in the Central Nervous System. A Clinicopathologic review of 18 cases 

and comparison to meningeal hemangiopericytomas. Arch Pathol Lab 

Med 2003;127:432-9. 

10 Gengler C, Guillou L. Solitary fibrous tumour and haemangiopericytoma: 

evolution of a concept. Histopathology 2006;48:63-74. 

11 a Paulus W, Scheithauer BW, Perry A, Hemangiopericytoma, in Louis 

DN, Ohgaki H, Wiestler OD, et al. (eds). Mesenchimal, non meningothelial 

tumors. Lyon: IARC Press 2007, pp. 173-7. b Giannini C, Rushing 

EJ, Hainfelier. In: Louis DN, Ohgaki H, Wiestler OD, et al. (ed). 

Hemangiopericytoma. WHO Classification of tumors of the central 

nervous system. Lyon: IARC Press 2007, pp. 178-80. 

12 Ambrosini-Spaltro A, Eusebi V. Meningeal hemangiopericytomas 

and emangiopericytoma/solitary fibrous tumors of extracranial soft 

tissues: a comparison. Virchows Arch 2010;456:343-54. 

TTf-1 and WT1 expression in embryonal soft tissue, 

visceral, and central nervous system tumors. An 

immunohistochemical study of 100 cases 

1)M. Bisceglia, 2)C. Galliani, 3)G. Lastilla, 4) J. Rosai 

1) Department of Pathology, IRCCS Casa Sollievo della Sofferenza Hospital, 

San Giovanni Rotondo, Italy; 2)Department of Pathology, Cook Children’s 

Medical Center, Fort Worth, TX, USA; 3)Department of Pathology, 

Polyclinic Hospital of Bari, Bari, Italy; and 4)Centro Diagnostico Italiano 

(CDI), Milan, Italy 

Background. TTF-1, a member of the NK-2 family of homeodomain 

transcription factors, is expressed in the early stages of thyroid, 

lung, and ventral forebrain development, and has been applied 

as a marker for epithelial-derived neoplasms of the lung and 

thyroid, both primary and secondary. In addition, a wide variety 

of cell and tissue types have been found to variably express TTF- 

1, including nephroblastoma. 1 WT1, encoded by Wilms’ tumor 

suppressor gene, controls the expression of growth factors that 

regulate glomerular capillary development, activates the bcl-2 

gene, and is normally expressed in the kidney (glomerular podocytes) 

and nephroblastomas. WT1 has also been seen expressed 

in other anatomical sites and neoplasms, including ovarian and 

endometrial cancer, mesothelioma, desmoplastic small round cell 

tumor, melanoma and acute leukemias. 

Objectives. To investigate TTF-1 and WT1 immunohistochemical 

nuclear expression of small round cell tumors of the soft tissues, 

viscera, and the central nervous system (CNS). 

253 

Materials and Methods. All cases were retrieved from the pathology 

files of the participating institutions, 9 were from outside 

institutions (6 consultation, 3 courtesy). Formalin-fixed, paraffin 

embedded tissues were obtained from 122 patients. Embryonal 

soft tissue and bone tumors (64 cases): 26 Ewing’s sarcoma/ 

primitive neuroectodermal tumors (EWS/pPNET), 13 peripheral 

(thoracoabdominal) neuroblastomas (pNB), 18 embryonal 

rhabdomyosarcomas (ERMS), and 5 desmoplastic small round 

cell tumors (DSRCT - 3 intraabdominal, and 2 extra-abdominal: 

1 dural-based intracranial, 1 pleural-based thoracic). Embryonal 

visceral tumors (12 cases): 5 hepatoblastomas (HB), 4 type I 

pleuropulmonary blastomas (PPB-I), 1 retinoblastoma (RB), 1 

pancreatoblastoma (1 PTB), 1 paraganglioblastoma (PGB), and 

1 embryonal liver sarcoma. Embryonal CNS tumors (24 cases): 

14 infratentorial PNET (medulloblastoma – MB), 6 central supratentorial 

PNET (cPNET), 3 central supratentorial neuroblastoma 

(cNB - including 1 olfactory neuroblastoma), and 1 pineoblastoma 

(PNB). We also studied 9 synovial sarcomas (SVS), a few 

differentiated CNS tumors (2 central neurocytomas, 3 pineocytomas, 

4 subependymomas), 2 ovarian small cell carcinoma of the 

hypercalcemic-type (SCC HC-type), 1 case of Merkel cell tumor, 

and one adult case of primitive-looking Merkel-like epithelialderived 

malignant tumor of the skin for contrast. To compare to 

our previous study of nephroblastoma 1 , we also tested WT1 reactivity 

in all these tumors. The medical records were abstracted 

for demographic information, specific anatomical sites, and 

diagnoses. Heat-induced antigen retrieval was used for detection 

of both markers. The following antibodies were used: monoclonal 

antibody TTF-1 (1:30 dilution; clone (8G7G3/1) and WT1 (1:50 

dilution; clone 6F-H2, directed against the amino terminal domain 

of WT1 protein). Appropriate positive and negative controls 

were used for each antibody. Immunohistochemical staining was 

performed using the labeled Envision system according to the 

manufacturer’s recommendations. 

Results. The series of embryonal tumors included 100 patients in 

total, 68 males and 32 females. 62 patients were in the pediatric 

age (≤ 21 years), 10 were young adults (> 21 and ≤ 30), and 28 

adults. Specifically, the patients’ age ranged between < 1 month 

and 42 months (mean 11.4 months), birth and 78 years (mean 

24.9 years), 1 and 18 (mean 5.12 years), and 5 and 62 (mean 

30.12 years), for embryonal tumors of peripheral nerve tissue 

(akin pNB), somatic soft tissue & bone, visceral organs, and 

CNS, respectively. The anatomical sites of pNB were: posterior 

mediastinum (3), adrenal (7), retroperitoneum (1), and 1 liver 

and 1 periaortic lymph node metastasis from adrenal. The rest 

of somatic and visceral tissues tumors involved the following 

anatomical locations: head & neck (7), genital organs (7), urinary 

bladder (4), retroperitoneum (2), liver (1), chest-wall (6), trunk (6 

- somatic superficial soft tissue), upper limb (1 - deep soft tissue), 

and serosal cavities (DSRCT 5 – intrabdominal 3, intracranial 1, 

thoracic 1); 3 tumors affected bone (skull 1, femur 2); 3 were 

pPNET metastases (1 each to hilar lymph node from lung, to skin 

from kidney, and to liver from unknown primary). There were 13 

visceral-based embryonal tumors (5 HB, 4 PPB-I, 1 PGB of the 

carotid body, 1 RB, 2 pPNET, 1 each of the kidney and colon). 

All CNS tumors were intraaxial (10 supratentorial; 14 infratentorial). 

Of 9 synovial sarcomas, 6 were female, 3 were male, with 

an age range between 10 and 72, and the anatomical sites were 

limbs (6 lower limbs, 1 upper limb), trunk (1 chestwall), and lung 

(1 metastasis). The 2 females with ovarian SCC HC-type were 

17 and 45 years of age, respectively. The immunohistochemical 

results are as follows: TTF-1 was negative in all the tumors 

tested, except for one case of suprasellar cPNET, which showed 

widespread immunopositivity in 40% of tumor cell nuclei; 

50% of PPB-I had alveolar-epithelial lining with nuclear TTF- 

1 immunostaining, serving as an internal control. No nuclear 

positivity for WT1 was found in any case of embryonal tumors. 

Of nonembryonal tumors, included in the study, the 2 cases of

254 

ovarian SCC HC-type showed nuclear moderate immunostaining 

in numerous tumor cells (diffuse in 1, and patchy in the other). 

However, strong WT1 cytoplasmic positivity was seen in all 18 

ERMS; moderate cytoplasmic staining was observed in 53.8% 

of pNB, 19.2% EWS/pPNET, 35.7% MB, 50% PPB-I, 60% 

DSRCT, 22.2% SVS, 50% cPNET, 40% of HB. WT1 was always 

and significantly positive in the endothelium of both normal and 

tumor vessels. 

Discussion. To the best of our knowledge, TTF-1 has not been 

previously investigated in a wide array of embryonal tumors. The 

impetus to perform such a study was stimulated by the discovery 

of a subset of nephroblastomas expressing nuclear TTF-1 immunopositivity 

1 . According to our results, negative TTF-1 may 

help in the differential diagnosis of primary PNET of the kidney 2 

versus nephroblastoma, which can express TTF-1. We could not 

confirm nuclear immunostaining in SVS as previously reported in 

one case metastatic to the lung 3 . The only TTF-1 positive cPNET 

is in agreement with reports of other positive peri-diencephalic 

neuroepithelial tumors 4 . Nuclear WT1 positivity in both ovarian 

SCC HC-type is in accordance with previous studies and in 

support of its müllerian origin 5 . Cytoplasmic WT1 positivity has 

been reported in ERMS 6 , and in the rhabdomyomatous component 

of nephroblastomas 1 . We exploited this property as an 

adjunctive marker in a case of spindle cell rhabdomyosarcoma 

of the heart 7 , supporting cytoplasmic WT1 as a marker for documenting 

skeletal muscle differentiation. Regarding the absence 

of WT1 nuclear immunoreactivity in DSRCT, it is in agreement 

with that of other investigators who used a similar monoclonal 

antibody 8 . The cytoplasmic reactivity for WT1 we observed in 

DSRCT might reflect its polyphenotypic nature, but needs to be 

elucidated. WT1 cytoplasmic immunopositivity in embryonal 

tumors of neural lineage and in the rest of soft tissue tumors is 

likely nonspecific, but cytoplasmic positivity in a case of MB is 

intriguing, since it was in fact an already known anaplastic MB, 

which we had previously diagnosed with immunohistochemical 

evidence for early rhabdomyoblastic differentiation (nuclear immunopositivity 

with myogenin). WT1 was consistently positive 

in the cytoplasm of endothelial cells mainly of tumoral vasculature, 

partly confirming previous experience 9 , and leading us 

to select WT1 as perhaps one of the most sensitive endothelial 

markers currently available (unpublished data of one of us [MB] ). 

Conclusion. Embryonal tumors of soft tissues or viscera, other 

than nephroblastoma, fail to express nuclear reaction with 

TTF-1. cPNET of the diencephalic region of the forebrain can 

express TTF-1 in tumor cell nuclei. ERMS consistently exhibits 

cytoplasmic WT1 immunopositivity. Other embryonal tumors, 

mainly of neural lineage and of the somatic soft tissues, may variably 

express cytoplasmic WT1 in a nonspecific fashion. WT1 is 

perhaps one of the most sensitive endothelial markers in surgical 

pathology. 

references 

1 Bisceglia M, Ragazzi M, Galliani CA, et al. TTF-1 expression in 

nephroblastoma. Am J Surg Pathol 2009;33:454-61. 

2 Parham DM, Roloson GJ, Feely M, et al. Primary malignant neuroepithelial 

tumors of the kidney: a clinicopathologic analysis of 146 adult 

and pediatric cases from the National Wilms’ Tumor Study Group 

Pathology Center. Am J Surg Pathol 2001;25:133-46. 

3 Lewis JS, Ritter JH, El-Mofty S. Alternative epithelial markers. In: 

Bridge JA, Beckwith JB (eds). Primary malignant neuroepithelial 

tumors of the kidney: a clinicopathologic sarcomatoid carcinomas of 

the head and neck, lung, and bladderp63, MOC-31, and TTF-1. Mod 

Pathol 2005;18:1471-81. 

4 Zamecnik J, Chanova M, Kodet R. Expression of thyroid transcription 

factor 1 in primary brain tumours. J Clin Pathol 2004;57:1111-3. 

5 Carlson JW, Nucci MR, Brodsky J, et al. Biomarker-assisted diagnosis 

of ovarian, cervical and pulmonary small cell carcinomas: the role 

of TTF-1, WT-1 and HPV analysis. Histopathology 2007;51:305-12. 

6 Carpentieri DF, Nichols K, Chou PM, et al. The expression of WT1 in 

the differentiation of rhabdomyosarcoma from other pediatric small 


round blue cell tumors. Mod Pathol 2002;15:1080-6. Comment in: 

Mod Pathol 2003;16:1178-; author reply 1179. 

7 Fraternali Orcioni G, Ravetti JL, Gaggero G, et al. Primary embryonal 

spindle cell cardiac rhabdomyosarcoma: case report. Pathol Res Pract 

2010;206:325-30. 

8 Barnoud R, Sabourin JC, Pasquier D, et al. Immunohistochemical 

expression of WT1 by desmoplastic small round cell tumor: a comparative 

study with other small round cell tumors. Am J Surg Pathol 

2000;24:830-6. 

9 Wagner N, Michiels JF, Schedl A, et al. The Wilms’ tumour suppressor 

WT1 is involved in endothelial cell proliferation and migration: 

expression in tumour vessels in vivo. Oncogene. 2008;27:3662-72. 

Primary embryonal rhabdomyosarcoma of prostate 

in adults. report of a case and review 

of the literature 

1)Bisceglia M. 2)Fiordelisi F. 3)Perrone G. 4)Dicandia L. 5)Cannazza 

V. 6)Ben Dor D. 

1)Unit of Anatomic Pathology, IRCCS “Casa Sollievo della Sofferenza”, 

San Giovanni Rotondo, Italy 2) Unit of Anatomic Pathology, IRCCS 

“Casa Sollievo della Sofferenza”, San Giovanni Rotondo, Italy 3) Unit of 

Anatomic Pathology, IRCCS “Casa Sollievo della Sofferenza”, San Giovanni 

Rotondo, Italy 4) Unit of Anatomic Pathology, IRCCS “Casa Sollievo 

della Sofferenza”, San Giovanni Rotondo, Italy 5)Unit of Radiology, 

P.O. di Scorrano, ASL - LE 2, Maglie, Italy 6)Department of Pathology, 

Barzilai Medical Center, Ashkelon, Israel 

Background. Embryonal rhadomyosarcoma (ERMS) is the most 

common tumor of the lower genitourinary tract occurring in the 

first 2 decades of life, both in males and females, mostly arising 

from the bladder, vagina, uterine cervix, prostate, and paratesticular 

region. ERMS of prostate in adults is extremely rare. In 

a systematic review, which was published by Waring et al. in 

1992, only 6 cases were found in the literature with adequate 

clinicopathological information, to which these authors added 3 

cases of their own. 1 Occasional case reports of ERMS in adults 

were published since. 

Objectives. To report one additional personal case of adult 

patient with ERMS and to comprehensively review the world 

literature on this subject. 

Case Report. In 2002, a 49-year-old male, who had undergone 

transurethral prostatic resection for benign nodular hyperplasia 

and left hydrocelectomy 4 and 2 years previously, respectively, 

was admitted for acute urinary retention. Previous pathological 

specimens were not available for review. At this time rectal 

digital examination revealed an enlarged firm prostate gland. 

Abdominal CT scan and US scan showed a prostatic tumor 9 cm 

in size with infiltrative margins, bulging into the urinary bladder 

and invading the perirectal adipose tissue. Bilateral hydronephrosis 

due to obstruction of both the ureters and enlarged iliac lymph 

nodes were also documented. At cystoscopy a polypoid tumor 

obstructing the prostatic urethra was seen and a transurethral 

tumor resection was performed. Light microscopic examination 

revealed a malignant tumor composed of an admixture of undifferentiated 

small round cells and scattered groups of spindleshaped 

cells with bipolar eosinophilic cytoplasmic extensions 

showing definite cross striations. Immunohistochemically the 

tumor cells were positive for vimentin, muscle specific actin, 

desmin, fast myosin, sarcomeric actin, and negative for CD34, 

EMA, S100 protein, PSA, PSAP; pan-cytokeratin (MNF116) 

was focally positive in a few cells. After several courses of 

neoadjuvant VAC-chemotherapy (vincristine, adriamycin, and 

cyclophosphomide), which reduced the tumor mass to 5 cm, the 

patient underwent radical cystoprostatectomy with bilateral seminal 

vesiculectomy and pelvic lymphadenectomy. Urinary diversion 

was accomplished with creation of bilateral ileal conduits. 

The original diagnosis was histologically confirmed on examination 

of the resection specimen. The urethral resection margin 

was positive for tumor. Both seminal vesicles, the iliac lymph


nodes, as well as the resection margins of both ureters, were all 

free of tumor. 6 months after surgery a huge pelvic recurrence 

of the ERMS, causing intestinal occlusion and bilateral ureteral 

obstruction which were relieved with percutaneous nephrostomy 

and transverse colostomy, was found and confirmed on needle 

biopsy. The patient became cachectic and severely debilitated 

and died 1 year after diagnosis. Distant metastases were not 

documented. Autopsy was not done. 

Literature Review. The literature was reviewed based on a computerized 

PubMed/Medline search, using [rhabdomyosarcoma 

AND prostate] as search terms, and the references lists of all the 

available publications on this subject, encompassing the interval 

between 1988 (the year when Waring’s et al. 1 review ended) and 

May 2010. 

Discussion. Sarcoma of prostate is rarely seen in adults, accounting 

for less than 5% of all malignant prostatic tumors. 

ERMS is the rarest type of sarcoma in this age group. Around 

40 cases of primary prostatic rhabdomyosarcoma have been reported 

so far in males ≥ 18 years of age from 1988 to May 2010. 

However, in compliance with Waring’s et al. inclusion/exclusion 

criteria 2 , less than 30 cases should be included, which, 

in addition to cases recorded in the afore-mentioned review, 

amount to a grand total of less than 40. ERMS mostly present 

with symptoms of progressive dysuria or urinary obstruction. 

Patients often present with locally advanced disease and at times 

with metastatic disease. A tumor mass is always discovered and 

the diagnosis is made on transrectal needle biopsy or transurethral 

resection or biopsy specimens. The differential diagnosis 

includes both stromal sarcomas arising from specific prostatic 

stroma, including STUMP (stromal tumors of uncertain malignant 

potential), and sarcomas of soft tissue-type, such as inflammatory 

myofibroblastic tumors, malignant peripheral nerve 

sheath tumors, leiomyosarcoma, and other types of rhabdomyosarcoma 

(alveolar and pleomorphic) 2 . Occasionally GIST from 

the rectum invading the prostate might also be a consideration. 

Immunohistochemistry is of utmost importance in ascertaining 

the correct diagnosis, which is based on immunopositivity for 

desmin and skeletal muscle markers (MyoD1, myogenin, fast 

myosin, sarcomeric actin, myoglobin, …). Predictive prognostic 

factors are stage-related. Adults with prostatic rhabdomyosarcomas 

do not respond to multimodal therapy and have a poor 

prognosis. Pediatric patients appear to respond much better than 

adults with combined modality treatment for sarcoma in general 

3-5 , and the rhabdomyosarcomatous group fares better than 

the nonrhabdomyosarcomatous one 3 . All adult patients with 

adequate follow-up died within 20 months after histological 

diagnosis with a mean survival of 8 to 10 months vs an overall 

5-year survival rate of 70-80% and a median survival of over 

10 years, respectively, in children 3 . Surgery is the mainstay of 

treatment. 

Conclusions. ERMS of prostate in adults is a very rare and aggressive 

disease. The long-term disease specific survival rate is 

poor. Stage influences the outcome. Early diagnosis and complete 

surgical resection offer the patients the best chance of improved 

survival. 

references 

1 Waring PM, Newland RC. Prostatic embryonal rhabdomyosarcoma in 

adults. A clinicopathologic review. Cancer 1992;69:755-62. 

2 Hansel DE, Herawi M, Montgomery E, et al. Spindle cell lesions of the 

adult prostate. Mod Pathol 2007;20:148-58. 

3 Janet NL, May AW, Akins RS. Sarcoma of the prostate: a single 

institutional review. Am J Clin Oncol 2009;32:27-9. 

4 Sexton WJ, Lance RE, Reyes AO, et al. Adult prostate sarcoma: the 

M.D. Anderson Cancer Center Experience. J Urol 2001;166:521-5. 

5 Mondaini N, Palli D, Saieva C, et al. Clinical characteristics and overall 

survival in genitourinary sarcomas treated with curative intent: a 

multicenter study. Eur Urol 2005;47:468-73. 

Aggressive angiomyxoma: a tumour with a wide 

morphological spectrum. A clinicopathological 

study of 27 cases including recurrent lesions 

255 

1)Gurrera A. 1)Amico P. 2)Bisceglia M. 1)Longo F. 3)Kazakov 

D. 3)Kacerovskà D. 3)Michal M. 1)Magro G. 

1)Dipartimento G.F. Ingrassia, Anatomia Patologica, Università di Catania, 

Azienda Ospedaliero-Universitaria Policlinico-Vittorio Emanuele, 

Catania, Italia; 2)Servizio Anatomia Patologica, IRCCS Ospedale Casa 

Sollievo della Sofferenza, San Giovanni Rotondo, Italia; 3)Sikl’s Department 

of Pathology, Charles University, Medical Faculty Hospital, Pilsen, 

Czech Republic 

Background. Aggressive angiomyxoma (AAM) is an uncommon 

fibro-myofibroblastic tumour, usually occurring in soft tissues of 

the vulvovaginal, pelvic and perineal regions of young females. 

Similar lesions have occasionally been reported in retroperitoneum 

and in perineum, para-anal and inguino-scrotal region. AAM 

is a locally infiltrative neoplasm with a significant risk of multiple 

local recurrences, with a low metastatic potential. 

Methods. The clinicopathological features of 27 cases of AAM 

are presented with emphasis on morphological heterogeneity of 

both primitive (22 cases) and recurrent tumours (5 cases). 

Results. Tumours usually presented as painless masses located in 

the vagina, vulva, and pelvi-perineum region of women ranging 

in age from 43 to 65 years. Grossly, most of tumours presented 

with ill-defined margins, ranging in size from 1.5 to 20 cm in 

greatest diameter and with a gelatinous to fibrous appearance at 

cut surface. Histologically, 59% of tumours were fibro-myxoid 

in appearance, while 26% and 15% were purely fibro-sclerotic or 

myxoid, respectively. Neoplastic cells were round to spindle or 

stellate in shape, with scanty cytoplasm and hyperchromatic nuclei. 

Cellularity was low in all but in 3 cases that were highly cellular. 

Mitoses were only rarely observed. Notably, smooth muscle 

cells, isolated or arranged in short fascicles, were found scattered 

throughout the stroma in 33% of cases. Vascular component was 

represented by small capillary-like to large blood vessels with 

perivascular hyalinization (48% of cases) and medial hypertrophy 

(37% of cases). Recurrent tumours were predominantly hypocellular 

fibro-sclerotic lesions (3 out 5 cases) that showed keloid-like 

collagen bands and a neurofibromatous-like pattern. In two of 

these cases, a complete sclerotic obliteration of blood vessels was 

seen resulting in confluent nodular structures closely reminiscent 

of corpora albicantia. 

The present study emphasizes that AAM is a tumour with a 

wide morphological spectrum ranging from a purely myxoid 

to hypocellular fibro-sclerotic lesion with a neurofibromatouslike 

appearance. This latter morphological feature, seen both 

in primitive and recurrent lesions, should be kept in mind by 

pathologist to avoid confusion with benign fibromatous lesions 

extracutaneous involvement of sporadic Kaposi’s 

sarcoma. A clinicopathologic study of a case series 

1)Bisceglia M. 2)Magro G.3) Panniello G. 4)G. Sanguedolce F. 

5)Ben Dor D. 

1)Department of Pathology, IRCCS Casa Sollievo della Sofferenza, San 

Giovanni Rotondo, Italy 2)Department of Pathology, University of Catania, 

Catania, Italy 3)Unit of Clinical Dermatology, Ospedali Riuniti, Foggia, 

Italy 4)Unit of Anatomic Pathology, Ospedali Riuniti, Foggia, Italy 5) 

Department of Pathology, Barzilai Medical Center, Ashkelon, Israel 

Background. Kaposi’s sarcoma (KS) is a peculiar tumor of vascular 

derivation and viral etiology (gammaherpesvirus HHV-8), 

occurring primarily in the skin 1 . KS is rare, comprising 0.1% 

of all malignancies worldwide. A variety of clinical forms have 

been identified: the sporadic, the endemic, the iatrogenic, and 

the epidemic 2 . The sporadic (or classical, European, Mediterranean) 

form primarily affects elderly Caucasian males with a

256 

predisposition for Eastern and Southern Europeans, and Jews of 

European (mainly Russian and Polish) and North African origin. 

KS mostly affects the skin of the acral sites, having a chronic 

and indolent clinical course and persisting for many years, with 

little propensity to spread to other organs 1 2 . In most cases the 

course is benign, but a fatal outcome after many years has also 

been observed. In sporadic KS soft tissue, bone, lymph nodes, 

and visceral organs (mainly the gastrointestinal tract) are rarely 

involved 1 2 . On occasion these unusual locations represent the 

only site of involvement. 

Objectives. To report on as well as to present a pictorial review 

of a series of extracutaneous KS involving different organs, either 

in isolation or in association with cutaneous manifestations. 

Materials and Methods. A systematic search of our combined 

databases based on institutional and personal consultation files 

was performed between 1985 and 2009 to identify extracutaneous 

cases of KS. All cases have been re-examined and immunohistochemically 

investigated with anti-LNA-1 (Latent Nuclear 

Antigen-1) HHV8 antibody (clone 13B10, dilution 1:20, Novocastra 

Laboratories, England, UK), if this had not already been 

performed at the time of the original examination. 

Results. 25 cases with extracutaneous involvement have been 

identified from about 750 cases of sporadic KS in our combined 

files. Patients’ ages ranged from 10 to 85 yrs. The male to female 

sex ratio was 11:1. Of these cases, 5 occurred in the soft tissues (4 

in the somatic soft tissue; 1 in the retroperitoneum, involving the 

right adrenal), 10 in the gastrointestinal tract (7 in the stomach, 

1 in the pharynx, 2 in the rectum), 2 in the bones (1 in the calcaneum 

and 1 in the lateral malleolus), 8 in the lymph node (3 of the 

neck, 4 inguinal, 1 axillary), and 1 in the parotid gland. All cases 

exhibited the classic predominantly spindle cell morphology, 

alternating with focal angiomatous-like foci. All cases exhibited 

diffuse and strong nuclear immunohistochemical reactivity with 

anti-LNA-1 HHV 8 antibody. 

Discussion. In the Mediterranean basin the frequency of sporadic 

KS is that of 1.5:100,000 people. The frequency of extracutaneous 

involvement for classical KS has been reported at 10%, 

which we think is an overestimate. Our rate is much lower, 

which may be because of the following: 1. KS patients are not 

systematically followed-up for visceral and mucosal involvement 

(most frequently in the gastrointestinal tract), which are usually 

asymptomatic; 2. we counted lesions per cases diagnosed and 

many patients had several skin lesions excised. All our cases 

of extracutaneous KS occurred in non-immunocompromised 

patients (non-AIDS associated KS, non-iatrogenic KS). However 

we included in this case series even those cases in which other 

conditions (usually multicentric Castleman’s disease, or non- 

Hodgkin’s lymphoma) were simultaneously found in association 

in the same organ (usually lymph node). In 10 cases we were 

aware of previous or concurrent skin lesions in the same patient. 

Although most cutaneous KS lesions are easily identified by an 

experienced pathologist, some lesions are not easy to diagnose if 

seen out of the usual anatomic context in which is KS expected 

to occur. Soft tissue KS may be misinterpreted as a different type 

of spindle cell sarcoma with an inflammatory component (mostly 

leiomyosarcoma), but HHV-8 immunohistochemcial testing is 

extremely useful for ruling this out 3 . On small biopsies KS in 

the gastrointestinal tract may be confused with granulation tissue 

or other reactive vasoproliferative lesions. KS in lymph nodes 

may be overlooked, since at times it is represented by small foci 

in association with other reactive or neoplastic lymphoproliferative 

processes, and may also be confused with foci of nodular 

spindle-cell vascular transformation. We did not follow-up these 

patients since this was not the scope of this report, however we 

are aware that in 2 of these cases the outcome was fatal following 

spread to visceral organs (lung, and brain), one of the two cases 

having in addition bone involvement, and the other had a huge 

local recurrence with primary retroperitoneal involvement (in the 


absence of skin changes), after a long disease course of 30 years 

and 13 years, respectively. Notably involvement of the adrenal in 

a non-HIV KS patient has been previously reported only once 4 . 

Also of interest is our youngest patient, currently in good health, 

who was diagnosed with KS of lymph node, the second pediatric 

case in the literature, and who was the subject of a separate report 

in 1988 5 . Parenthetically the first pediatric patient with classical 

KS involving the lymph node was also Italian. 

Conclusion. Extracutaneous KS does occur, but is rare. The pathologist 

should be aware of this occurrence. HHV-8 immunohistochemical 

testing is critical for KS diagnosis in these cases. 

references 

1 Bisceglia M, Bosman C, Carlesimo OA, et al. Kaposi’s sarcoma: a 

clinico-pathologic overview. Tumori 1991;77:291-310. 

2 Geraminejad P, Memar O, Aronson I, et al. Kaposi’s sarcoma and 

other manifestations of human herpesvirus 8. J Am Acad Dermatol 

2002;47:641-55. 

3 Cheuk W, Wong KO, Wong CS, et al. Immunostaining for human herpesvirus 

8 latent nuclear antigen-1 helps distinguish Kaposi sarcoma 

from its mimickers. Am J Clin Pathol 2004;121:335-42. 

4 Lazure T, Plantier F, Alsamad IA, et al. Bilateral adrenal Kaposi’s 

sarcoma in an HIV seronegative patient. J Urol 2001;166:1822-3. 

5 Bisceglia M, Amini M, Bosman C. Primary Kaposi’s sarcoma of the 

lymph node in children. Cancer. 1988;61:1715-8. 

Phosphaturic mesenchymal tumor and oncogenic 

osteomalacia. A clinicopathologic study of 14 

cases with emphasis on unusual features, and 

review of the literature 

1)Bisceglia M. 2)Parafioriti A. 3)Robbins P. 4)Elmberger G. 

5)Fusconi M. 6)Rendina D. 7)Alberghini M. 8)Viti R. 9)Armiraglio 

E. 10)Spagnolo D. 11)Pasquinelli G. 12)Varenna M. 

13)Guglielmi G. 14)Perrone E. 15)Scillitani A. 16)Mossetti G. 

1)Unit of Anatomic Pathology, IRCCS Casa Sollievo della Sofferenza 

Hospital, San Giovanni Rotondo, Italy 2)Unit of Anatomic Pathology, 

Gaetano Pini Institute, Milan, Italy 3)Department of Anatomic Pathology, 

PathWest Laboratory Medicine, Perth, Western Australia 4)Department of 

Pathology and Cytology, Karolinska University Hospital, Stockholm, Sweden 

5)Unit of Rheumatology, University of Bologna, Bologna, Italy 6)Department 

of Clinical and Experimental Medicine, Federico II University, 

Naples, Italy 7)Unit of Anatomic Pathology, Rizzoli Institute, Bologna, 

Italy 8)Unit of Endocrinology, IRCCS Casa Sollievo della Sofferenza Hospital, 

San Giovanni Rotondo, Italy 9)Unit of Anatomic Pathology, Gaetano 

Pini Institute, Milan, Italy 10)Department of Anatomic Pathology, 

PathWest Laboratory Medicine, Perth, Western Australia 11)Department 

of Clinical Pathology, University of Bologna, Bologna, Italy 12) Unit of 

Metabolic Bone Diseases, Gaetano Pini Institute, Milan, Italy 13) Unit of 

Radiology, IRCCS Casa Sollievo della Sofferenza Hospital, San Giovanni 

Rotondo, Italy 14) Unit of Nuclear Medicine, IRCCS Casa Sollievo della 

Sofferenza Hospital, San Giovanni Rotondo, Italy 15) Unit of Endocrinology, 

IRCCS Casa Sollievo della Sofferenza Hospital, San Giovanni 

Rotondo, Italy 16)Department of Clinical and Experimental Medicine, 

Federico II University, Naples, Italy 

Background. Most cases of oncogenic osteomalacia (OO) are 

caused by mesenchymal tumors (phosphaturic mesenchymal 

tumors - PMT), which overexpress fibroblast growth factor-23 

(FGF-23), a protein of the “phosphatonin” family, capable of inhibiting 

renal tubular phosphate transport. The typical laboratory 

findings secondary to phosphate loss are hypophosphatemia and 

hyperphosphaturia, which finally result in an inadequate mineralization 

of osteoid in mature bone, the metabolic disorder known 

as osteomalacia 1 . OO is vitamin-D resistant and dramatically 

cured by tumor excision. The 1 st case of PMT was described in 

1947, but the term PMT was coined in 1987 2 . PMT is rare and 

consistently located in soft tissue and bone of limbs and trunk; 

sinonasal cavities and acral parts are traditionally considered uncommon 

sites. In a review of the literature up to 2002, Folpe et al. 

found 109 cases on record, to which they added 29 new original 

cases of their own of 32 they studied in total. 3 Histologically PMT


corresponds to a polymorphous group of neoplastic entities, the 

vast majority of cases, particularly in soft tissue, are associated 

with a specific histopathologic entity, the mixed connective tissue 

variant (PMT-MCT) 3 , which is characterized by a distinctive 

admixture of bland spindled cells, osteoclast-like giant cells, microcysts, 

prominent and variously sized vasculature, smudgy to 

calcified cartilage-like matrix and metaplastic bone. Some cases 

have histological features of malignancy. PMT of craniofacial 

sinuses usually differs from PMT-MCT and closely resembles 

a sinonasal HPC-like tumor variant. Tumor discovery and histological 

recognition are frequently delayed in OO. Sometimes 

(< 10%) OO is not documented, but the diagnosis of PMT can 

be proposed reasonably on the basis of the histological features 

(aphosphaturic PMT) 3 . 

Objectives. 1. To describe the clinicopathologic features of our 

cases of OO with emphasis on unusual findings. 2. To comprehensively 

review the world literature between January 2002 and 

March 2010. 

Methods. 1. A systematic search of our combined databases 

was performed to identify cases of possible PMT. All clinicopathologic 

features, including serum biochemical determinations 

and imaging studies were reviewed. Electron microscopy was 

performed in one sinonasal case. 2. A computerized PubMed/ 

Medline search was performed, using 4oncogenic osteomalacia7, 

[tumor-induced osteomalacia], and 4phosphaturic mesenchymal 

tumor7 as search terms. 

Results. 21 cases were initially retrieved from the institutional 

files and personal consultation archives, of which 7 PMT with 

OO were excluded since they had been previously reported. Of 

these latter 7 cases, 4 had been surgically excised and histologically 

examined (1 intraosseous osteoblastoma of sacrum, 1 

sinonasal HPC-like tumor, 2 soft tissue PMT-MCT), and 3 were 

not operated on (1 vertebral hemangiomatosis, 2 tumors unidentified). 

The present study concerns 14 patients (age range 21-70, 

median 51) 2 of whom were included previously in Folpe’s et al. 

series, though without detailed clinical history or illustrations. 

5 were males, 9 were females: 13 with OO, and 1 asymptomatic. 

Imaging (standard X-ray, and/or CT, and/or MRI, and/or 

PET-CT, and/or bone scintigraphy) and appropriate biochemical 

studies were performed in all. Octreotide scan for somatostatin 

receptor imaging was positive in 3 of 6 cases so studied. FGF-23 

serum levels were elevated in all 6 cases assayed (FGF-23 failed 

to decrease in 2 cases with incompletely removed tumor, but 

normalised on re-excision). Longstanding symptoms and delayed 

diagnosis were frequent (6/14); there was failure to recognise the 

causal tumor in 2/14. PMT occurred as a soft tissue lesion of the 

foot in 3 cases, was intraosseous in 7 (2 in the femur, 1 each in the 

humerus, rib, ileum, C1-vertebra) and sinonasal in 2. Histologically 

PMT was MCT-type in 9 and HPC-like in 3. In sinonasal 

cases PMT was HPC-like in 1 and MCT-type in 1. All the excised 

tumors were histologically benign (12/12). 1 case examined ultrastructurally 

displayed suggestive neuroendocrine dense core 

granules. 1 case, which was immunostained for FGF-23, was 

positive. OO normalized after complete tumor removal in 10/12 

surgically treated cases with OO (repeat operations required in 

2). The 2 cases of OO with no evidence of PMT were diagnosed 

according to ASBMR criteria 4 and medically treated with phosphate 

and calcifediol supplementation with minimal benefit. The 

single case of histologically proven PMT without OO occurred 

as a soft tissue tumor of the hand in a 62-year old female. In our 

most recent literature review for the years 2002-2010, 107 cases 

of PMT were found (mostly adult patients; 2 in pediatric age). Of 

the 100 for which the site was known, 58 cases occurred in soft 

tissue, 24 in bone, 11 in nasal/paranasal cavities, 4 were adjacent 

to or involved the central nervous system coverings (2 intracranial; 

2 intraspinal) and 3 were visceral (1 each in tongue, liver and 

uterus). Acral location (bone and soft tissue) occurred in 13/100 

(foot in 11, hand in 2). 

257 

Conclusions. 1. PMT is a rare, poorly understood pathologic 

entity, often with delayed diagnosis. 2. Acral sites (especially 

foot) and sinonasal locations are not uncommon. 3. Aphosphaturic 

PMT is rare, but may occur. 4. PMT-MCT is the commonest 

histological variant, and may also occur in nasal/paranasal cavities. 

5. OO is cured by surgery, but fails to regress after incomplete 

tumor removal. 6. FGF-23 serum level is a sensitive tumor 

biomarker that allows clinical management. 7. “Orphan” OO is 

rarely established despite careful and repeat investigations. 

references 

1 Jan de Beur SM. Tumor-induced osteomalacia. JAMA. 2005;294:1260- 

7. 

2 Weidner N, Santa Cruz D. Phosphaturic mesenchymal tumors. 

A polymorphous group causing osteomalacia or rickets. Cancer 

1987;59:1442-54. 

3 Folpe AL, Fanburg-Smith JC, Billings SD, et al. Most osteomalaciaassociated 

mesenchymal tumors are a single histopathological entity. 

An analysis of 32 cases and a review of the literature. Am J Surg 

Pathol 2004;28:1-30. 

4 Jan de Beur SM. Tumor-induced osteomalacia. In: American Society 

for Bone and Mineral Research (ed). Primer on the metabolic bone 

diseases and disorders of mineral metabolism. American Society for 

Bone and Mineral Research 2006, pp 345-51. 

Primary malignant melanoma of the esophagus. 

A clinico-pathologic study of a case with literature 

review 

1)Bisceglia M. 2)Perri F. 3)Tardio M. 4)Vairo M. 5)Pasquinelli 

G. 

1)Unit of Anatomic Pathology, IRCCS Casa Sollievo della Sofferenza Hospital, 

San Giovanni Rotondo, Italy 2)Unit of Gastroenterology, IRCCS 

Casa Sollievo della Sofferenza Hospital, San Giovanni Rotondo, Italy 

3)Unit of Surgery, IRCCS Casa Sollievo della Sofferenza Hospital, San 

Giovanni Rotondo, Italy 4)Unit of Anatomic Pathology, IRCCS Casa Sollievo 

della Sofferenza Hospital, San Giovanni Rotondo, Italy 5)Department 

of Clinical Pathology, University of Bologna, Bologna, Italy 

Background. Primary malignant melanoma originating from 

internal organs is rare, but has been well documented in the 

literature 1 . Primary melanoma originating in the digestive tract 

is particularly rare, with the majority of cases involving the oral 

cavity and anorectum 1,2 . Primary malignant melanoma of the 

esophagus (PMME) has been the source mainly of case reports, 

and its frequency is estimated around 0.1-0.2% of all esophageal 

malignancies 3 . In a large epidemiologic study in USA the median 

age was 69 and the age-adjusted rate incidence that of 0.03 per 

million population 4 . Again in another large epidemiologic study 

in USA, from 1973 to 2004, 39 PMME were found on record 

of 659 total primary gastrointestinal malignant neoplasms 5 . In 

3 separate reviews, presented up to 1989, to 1999, and to (June) 

2005 a total of only 139, 154, and 262 cases, respectively, could 

be identified in the world literature 3 6 7 . 

Objectives. To describe a personal case of PMME, and to comprehensively 

review on a computerized search the world literature 

between 2005 and December 2009. 

Case Report. A 69-year-old man was admitted for complaints 

of abdominal distress and melena, who had never undergone 

surgery or been diagnosed previously with malignancy. An 

echoscan of the stomach and esophagus revealed an area of mural 

thickening at the level of the lower 3 rd of the esophagus and 

involving the cardia. On endoscopic examination, the tumor was 

exophytic polypoid. An endoscopic biopsy revealed a poor1y 

differentiated, malignant neoplasm. The patient underwent partial 

esophagectomy with total gastrectomy. The surgical specimen 

was sent for pathological examination. The resection specimen 

showed a large, ulcerated, partly fungating, tan red mural mass 

of 6 cm in greatest diameter. Histological examination revealed 

a malignant neoplasm composed of solid sheets or discrete nests 

of monotonous, highly malignant, tumor cells with large nuclei

258 

and prominent nucleoli. Areas of hemorrhage and necrosis were 

evident throughout all sections. Mitotic figures were numerous 

(> 50 per 10 HPF). The tumor was deeply infiltrating into the 

muscularis propria, and, in one area, a small focus of junctional 

melanocytic activity was found in the basal layer of the squamous 

mucosa at the level of the gastroesophageal junction. Histochemical 

reactions for mucin (PAS-D and mucicarmine) were negative 

in the tumor cells as negative was Fontana stain for argentaffin 

granules. Immunohistochemically the tumor was: strongly 

positive for vimentin, S-100 protein, HMB45, and Melan-A, and 

negative for cytokeratin AEl/AE3, CEA, MOC31, actin, desmin, 

EMA, CD10, CD20, CD45, CD99, CD117 and AFP. Electron 

microscopic examination was performed on tissue retrieved from 

the paraffin block, which in some of the cells displayed stage I 

premelanosomes and stage II melanosomes. All 15 perigastric 

and esophageal lymph nodes examined in total were free of tumor. 

(pT3, pN0, pMx). The final established diagnosis was that 

of amelanotic PMME. Follow-up: Careful physical examination 

in our patient following the postoperative diagnosis did not reveal 

any cutaneous lesion suspicious for melanoma or any tumor elsewhere 

in the body. Clinical follow-up demonstrated a recurrent 

lesion at the site of anastomosis 10 months after surgery. The 

patient died of disease 24 months after primary diagnosis. 

Discussion. Less than 60 additional PMMEs have been found 

up from July 2005 to May 2010, amounting to a grand total of 

320 since ever. PMME is thought to originate from foci of basal 

melanocytes present in the squamous epithelium 8 9 . The clinical 

features of PMME similar to those of carcinoma of the esophagus. 

Most patients are in their sixth and seventh decades, with a 

slight male predilection. Dysphagia, substernal pain, heartburn, 

and weight loss are the most common symptoms. Grossly the 

tumors are most often polypoid and ulcerated, and can vary in 

size from small lesions to large, bulky masses. The mucosa at the 

edges of the lesion can be pigmented, a phenomenon referred to 

as “melanosis” of the esophagus 8 9 . The majority of the tumors 

are grossly pigmented; however, rarely completely amelanotic 

lesions can occur, as in our case. The histologic differential diagnosis 

for this tumor, particular1y the amelanotic variant, is quite 

broad, since it may present either as a large epithelioid cell or 

spindle cell or small cell malignant neoplasm: poorly differentiated 

carcinoma, gastrointestinal stromal tumor (GIST) or other 

malignant mesenchymal neoplasms, such as leiomyosarcoma and 

malignant peripheral nerve sheath tumor, or non-Hodgkin malignant 

lymphoma. Immunohistochemistry should be able to clear1y 

define the melanocytic nature of the tumor cells. However, the 

main differential diagnosis of PMME is with a metastasis to 

this organ from melanoma of another site. The most important 

histological feature suggestive of PMME is the identification of 

junctional activity by atypical melanocytes within the basal layer 

of the squamous epithelium. In the present case, the combination 

of the absence of a tumor in any other location on thorough clinical 

examination, absence of development of other lesions in other 

organs after 10 months of follow-up, local recurrence at the site of 

surgery, and focal Pagetoid involvement in the squamous mucosa 

from the resected specimen all supported a diagnosis of primary 

melanoma of the esophagus in our patient. PMME is quite aggressive, 

likely more aggressive that its cutaneous counterparts, 

but this may be due to their larger size and depth of invasion at 

the time of diagnosis. Common sites of metastases are regional 

lymph nodes, liver, mediastinum, lung and brain. The average 

survival time following esophagectomy for primary melanoma 

is less than 1 year, with a 5 year survival of about 2%. Complete 

surgical excision is the standard treatment, followed by adjuvant 

radiation and chemotherapy. Local endoscopic laser treatment 

may play a role in palliation in locally advanced tumors that are 

unresectable. 

Conclusions. This thoroughly documented case is presented for 

its rarity and the differential diagnosis, especially for amelanotic 


PMME is emphasized. The world literature has been reviewed 

up to date. 

references 

1 Batsakis JG, Suarez P. Mucosal melanomas: a review. Adv Anat 

Pathol 2000;7:167-80. 

2 Mills SE, Cooper PH. Malignant melanoma of the digestive system. 

Pathol Annu 1983;18:1-26. 

3 Sabanathan S, Eng J, Pradhan GN. Primary malignant melanoma of 

the esophagus. Am J Gastroenterol 1989;84:1475-81. 

4 Coté TR, Sobin LH. Primary melanomas of the esophagus and anorectum: 

epidemiologic comparison with melanoma of the skin. Melanoma 

Res 2009;19:58-60. 

5 Cheung MC, Perez EA, Molina MA, et al. Defining the role of surgery 

for primary gastrointestinal tract melanoma. J Gastrointest Surg 

2008;12:731-8. 

6 Lam KY, Law S, Wong J. Malignant melanoma of the oesophagus: 

clinicopathological features, lack of p53 expression and steroid receptors 

and a review of the literature. Eur J Surg Oncol 1999;25:168-72. 

7 Vandewoude M, Cornelis A, Wyndaele D, et al. Acta Gastroenterol 

Belg 2006;69:12-4. (18) FDG-PET-scan in staging of primary malignant 

melanoma of the oesophagus: a case report. 

8 Sharma SS, Venkateswaran S, Chacko A, et al. Melanosis of the 

esophagus. An endoscopic, histochemical, and ultrastructural study. 

Gastroenterology 1991;100:13-6. 

9 Chang F, Deere H. Esophageal melanocytosis morphologic features 

and review of the literature. Arch Pathol Lab Med 2006;130:552-7. 

Immunosuppression-associated Kaposi’s sarcoma 

complicating chronic inflammatory bowel disease. 

A clinico-pathologic study of 2 cases with review 

of the literature 

1)Bisceglia M. 2)Piscitelli D. 3)Panniello G. 4)Sanguedolce F. 5) 

Serviddio G. 6) Bisceglia M.L. 7)Ben Dor D. 


Giovanni Rotondo, Italy 2)Department of Pathology, Polyclinic Hospital, 

Bari, Italy 3)Division of Clinical Dermatology, Ospedali Riuniti, Foggia, 

Italy 4)Division of Anatomic Pathology, Ospedali Riuniti, Foggia, Italy 

5) Institute of Internal Medicine, University of Foggia, Foggia, Italy 6) 

School of Pharmacy, University of Parma, Parma, Italy 7)Department of 

Pathology, Barzilai Medical Center, Ashkelon, Israel 


histogenesis and viral etiology (gammaherpesvirus HHV-8), 

occurring primarily in the skin 1 . A variety of clinical forms have 

been identified: the sporadic, the endemic, the iatrogenic, and the 

epidemic 2 . The iatrogenic form supervenes in immunocompromised 

hosts, i.e. transplanted patients with immunosuppression, patients 

receiving immunosuppressive therapies for haematological malignancies 

(mainly chronic lymphocytic leukemia, and non-Hodgkin’s 

and Hodgkin’s lymphomas), solid tumors (mainly carcinomas of the 

breast, followed by lung, colon, larynx, liver, pancreas, and kidney, 

in decreasing order of frequency), or inflammatory/autoimmune 

diseases, after long-term steroid treatment 1 2 . Also included in this 

rubric of KS are those rare cases of patients receiving blood transfusions, 

factor VIII containing plasma fractions, or platelets apheresis. 

Excluded from this iatrogenic category are those KS patients who 

either subsequently present with a second malignancy 3 or are simultaneously 

diagnosed with a second malignancy, in the absence 

of prior systemic anticancer therapy and/or any clinically detectable 

immunosuppression, though both conditions may have developed 

independently and coincidentally on the background of an altered 

immune system. Non-neoplastic medical conditions treated with 

immunosuppressive drugs (mainly corticosteroids), which are on 

record as (rarely) associated with, KS are: rheumatoid arthritis, giant 

cell arteritis (Horton arteritis), relapsing polychondritis, systemic 

lupus erythematosus (SLE), pemphigus vulgaris and inflammatory 

chronic intestinal diseases. 

Objectives. 1. To report on two cases of iatrogenic KS in patients 

receiving immunosuppressive therapy for chronic inflammatory 

bowel disease, one with chronic ulcerative colitis and the other


with Crohn’s disease, neither of whom had HIV infection, or had 

been receiving immunosuppressive treatment following transplantation. 

2. To review the world literature with regards to iatrogenic 

KS complicating chronic inflammatory intestinal diseases 

recorded between January 1980 and December 2009. 

Case reports. Case 1. A 50-year old man diagnosed six months 

previously with biopsy-proven severe ulcerative colitis and 

treated with corticosteroids and azathioprine, underwent emergency 

subtotal colectomy due to massive intestinal bleeding. 

Histological examination of the surgical specimen revealed 

widespread involvement of the colon by KS, in addition to 

ulcerative colitis. Following histological diagnosis, the patient 

underwent proctosigmoidectomy, and KS with ulcerative colitis 

was also seen in the rectum. The postoperative course was uneventful, 

the immunosuppressive treatment was withdrawn and 

the patient recovered. Case 2. A 65-year old man, diagnosed 

with biopsy-proven Crohn’s disease involving the duodenum 

and receiving near-continuous immunosuppressive treatment for 

5 years, underwent surgical resection of a 60 cm long segment of 

jejunum, due to repeat episodes of bowel occlusion. Histological 

examination of the specimen revealed KS in association with 

Crohn’s disease. 3 weeks after surgery the patient was severely 

debilitated, and died due to Candida Albicans sepsis. Autopsy 

was not performed. 

Discussion. Patients who receive immunosuppressive therapy 

are at increased risk for KS (immunosuppression-associated 

KS), the majority of whom are renal transplant patients. In addition 

to the role of immunologic impairment, other etiologic 

factors also play a role in this form of KS, and one of the recognized 

risk factors for this type of KS is ethnicity (Eastern and 

Mediterranean people as well as Jews of European and North 

African origin are at higher risk). The immunosuppression-associated 

form of KS occurs between a few months and a few 

years after starting therapy. Immunosuppression-associated KS 

may also affect the skin, but may be limited to the gastrointestinal 

tract. 9 cases of non-transplant associated iatrogenic KS 

afflicting patients with long-lasting inflammatory chronic bowel 

disease (chronic ulcerative colitis in 6 and Crohn’s disease in 

2) have been described so far in HIV-negative patients 4 5 . All 

cases were on long-standing immunsuppressive treatment. Two 

of these previously reported cases involved Italian patients, both 

affected by ulcerative colitis. One of our two patients (both Italian) 

was on immunosuppressive therapy for 5 months, while 

the other one was on it for 5 years. In both cases the initial 

diagnosis was morphological, but was subsequently confirmed 

with the anti-LAN-1 HHV-8 monoclonal antibody, when it 

became commercially available, with both cases showing strong 

and diffuse positivity. The differential diagnosis of KS of the 

intestine includes other types of spindle cell sarcomas such as 

angiosarcoma, GIST, leiomyosarcoma, and inflammatory myofibroblastic 

tumor (previously known as inflammatory fibrosarcomas): 

immunohistochemical testing with anti-HHV-8 antibody is 

critical, given its high sensitivity (almost 100%) and specificity 

(100%) 6 . After the skin the gastrointestinal tract is the most frequent 

anatomic site of involvement by KS, the stomach, being 

most frequently affected, followed by the colon. Colon may be 

affected by classic KS and may be the only site of involvement 

(exclusive of skin). Conversely, KS involvement limited to the 

skin may also occur secondarily to medical treatment for colonic 

inflammatory diseases. Nonetheless, immunosuppression-associated 

KS complicating chronic inflammatory diseases of the 

bowel is rare. Parenthetically we mention here, that AIDS related 

KS presenting as ulcerative colitis-like illness has also been 

observed, but this should not be confused with the subject in 

question. Genetic susceptibility is definitely part of the complex 

interplay in KS between the mechanism of cell proliferation, the 

apoptotic controlling machinery and an individual’s immune 

regulatory systems. 

259 

Conclusion. Immunosuppression-associated KS complicating 

ulcerative colitis and Crohn’s disease is rare, with 9 cases on 

record. Intestinal resection (especially proctocolectomy for ulcerative 

colitis) and the withdrawal of immunosuppressive drugs 

result in improvement of the patient’s general health. 

references 



2002;47:641-55. 


clinico-pathologic overview. Tumori 1991;77:291-310. 

3 Bisceglia M, Zenarola P, Melillo L, et al. Cutaneous presentation of 

acute myeloid leukemia in a “classical” Kaposi’s sarcoma patient. 

Tumori 1990;76:400-2. 

4 Girelli CM, Serio G, Rocca E, et al. Refractory ulcerative colitis and 

iatrogenic colorectal Kaposi’s sarcoma. Dig Liver Dis 2009;41:170- 

4. 

5 Tedesco M, Benevolo M, Frezza F, et al. Colorectal Kaposi’s sarcoma 

in an HIV-negative male in association with ulcerative rectocolitis: a 

case report. Anticancer Res 1999;19:3045-8. 




unclassified non-pleomorphic sarcoma versus 

de novo malignant solitary fibrous tumor versus 

monophasic fibrous synovial sarcoma – primary 

of the kidney. Pathologic and molecular study 

of a case with a long-term survivor 

1)Bisceglia M. 2)Trabucco S. 3)Albrizio M. 4)Palmiotti G. 5)Alberghini 

M. 6)Pasquinelli G. 7)Serio G. 


Giovanni Rotondo, Italy 2)Department of Pathology, University of Bari, 

Bari, Italy 3)Department of Pathology, Di Venere Hospital, Carbonara- 

Bari, Italy 4)Department of Clinical Oncology, Di Venere Hospital, Carbonara-Bari, 

Italy 5)Unit of Anatomic Pathology, Rizzoli Institute, Bologna, 

Italy 6)Department of Clinical Pathology, University of Bologna, 

Bologna, Italy 7) Department of Pathology, University of Bari, Bari, Italy 

Background. Solitary fibrous tumor of the kidney (SFTK) 

was first described in 1996 and can originate either in the renal 

capsule or parenchyma 1 2 . To date 38 cases of SFTK have been 

reported, most of them described by standard criteria as histologically 

benign, and carrying a favourable clinical prognosis 

(follow-up ranging 2 to 89 months) 1-3 . Only 2 cases of SFTK 

exhibiting malignant histological changes were reported in 2006 1 

and 2008 2 , respectively. The tumor in the first case was > 10cm 

in size and diffusely malignant, with the conventional (benign) 

features found only focally: the patient developed lung metastases 

4 months after surgery (malignant [secondary] SFTK arising in a 

preexisting tumor which had been followed clinically as a stable 

lesion for 4 years). 1 The tumor in the second case was 9 cm in 

diameter with a 3 cm nodular malignant area abruptly emerging 

from the surrounding typically bland SFT tissue (dedifferentiated 

SFTK or SFTK with sarcomatous overgrowth); this patient was 

free of disease 21 months after surgery. 3 Malignant SFTK in the 

absence of residual histologically benign SFT may be difficult if 

not impossible to assess as well as to differentiate from primary 

menophasic fibrous synovial sarcoma of the kidney (SSK). SSK, 

a rare neoplasm usually carrying a poor prognosis, was first described 

in 2000 4 5 . Primary SSK can exist in either a monophasic 

or a biphasic form, and may be misdiagnosed as another type of 

sarcomatous or sarcomatoid renal tumor, primary or metastatic. 

As its soft tissue counterpart, the diagnosis of primary SSK can 

be confirmed by molecular analysis, showing the characteristic 

t(X;18) (p11;q11) translocation. To date 54 cases of primary 

SSK have been reported, including 3 with rhabdoid features 6 

(rhabdoid variant of SSK). 

Objectives. To report a case of a primary nonpleomorphic renal

260 

sarcoma in a young lady which was difficult to categorize, in 

whom lung metastases developed 10 years after nephrectomy, 

and which on review was eventually classified as “unspecified 

nonpleomorphic sarcoma”. 

Methods - Case Report. In 2009 this patient was admitted for 

a huge left pulmonary pneumonia-like infiltrate, accompanied 

by an abundant pleural effusion. Total body 18 FGF-PET scan 

revealed high uptake in the corresponding left lung and the patient 

underwent minimally invasive open lung biopsy. A minute, 

2 mm in size, intrapulmonary, mesenchymal malignant tumor 

nodule was excised. Given a known history of left nephrectomy 

of 10 years earlier, the patient’s previous clinical chart and the 

original glass slides of that tumor were retrospectively reviewed 

and newly cut sections were either immunohistochemically analyzed 

or submitted for molecular investigation. In addition small 

fragments of paraffin-embedded tissue were deparaffinised and 

processed for ultrastructural investigation. 

Results. The past renal tumor was hypercellular and mitotically 

active, composed of atypical, monomorphic, round to oval closely 

apposed medium-sized tumor cells, and showed focal HPC-like 

growth pattern, hemorrhagic foci, necrosis and pseudocystic areas. 

The greatest tumor diameter was 12.5 cm. The tumor margins were 

infiltrative, entrapping the surrounding normal renal parenchyma. 

The renal pelvis was infiltrated, but hilar or perirenal adipose tissues 

as well as 5 paracaval/periaortic lymph nodes submitted for 

histological examination were not involved. Examination of the 

newly excised lung nodule showed that it was morphologically 

consistent with a metastasis from the renal tumor. The immunoprofile 

of both the renal primary and lung metastasis was: vimentin 

diffusely +ve; BCL-2 diffusely +ve; EMA focally +ve; CD34 

patchy +ve (strongly diffuse on restaining); CD99 focally +ve; CK 

(pankeratin, CK7, CK19) all negative; alpha-SMA, desmin, myogenin 

all negative; CD117 negative; Fli-1 negative; WT1 negative; 

TTF1 negative; S-100 negative; CD10, ER, and PGR all negative. 

Electron microscopy showed closely apposed oval-shaped cells 

lacking epithelial differentiation (no tonofibrils, no desmosomes), 

with absence of actin-like microfilaments. Basal lamina was not 

seen. Molecular analyses (RT-PCR and FISH analyses) did not 

demonstrate the t(X;18) (p11;q11) translocation of either SYT- 

SSX1 or SYT-SSX2 gene fusion. Taking all these findings into 

account the final diagnosis was “unclassified nonpleomorphic renal 

sarcoma” – probably de novo malignant SFTK. 

Discussion. The diagnosis of malignant SFTK was neither 

straightforward nor was it eventually completely accepted since 

no foci of benign SFT were found (areas of usual SFT had been 

seen so far in both the 2 afore-mentioned cases of malignant 

SFTK as well as in all cases of the recently recognized dedifferentiated 

SFT of soft tissue 7 . Furthermore, CD34 may also be 

positive in sarcomas other than malignant SFTK and is most often 

lost in the malignant and dedifferentiated areas of SFT in both 

renal and soft tissue cases 1 8 . Notwithstanding, de novo malignant 

SFT is a real possibility. SSK, which had not yet been described 

at the time of nephrectomy, was the main consideration on review, 

but its exclusion is based on both the absence of the specific 

translocation and presence of CD34 positivity (parenthetically 

CD34 positivity was also seen in 3 cases of intrathoracic SS 7 ); 

other renal primaries, excluded for more obvious reasons, were 

sarcomatoid renal cell carcinoma, primary renal fibrosarcoma, 

malignant nerve sheath tumor, monomorphic angiomyolipoma, 

extragonadal endometrial stromal sarcoma, inflammatory myofibroblastic 

tumor, congenital mesoblastic nephroma, malignant 

mixed epithelial stromal tumor, leiomyosarcoma, anaplastic 

sarcoma of the kidney with polyphenotypic features 9 , of recent 

identification, and (atypical) congenital mesoblastic nephroma. 

Follow-up: The patient was given several courses of chemotherapy, 

using Ifosfamide, Epirubicin and MESNA, and temporarily 

improved. Currently, 10 months following discovery of the lung 

metastases, she is alive with slight disease progression. 


references 

1 Fine SW, McCarthy DM, Chan TY, et al. Malignant solitary fibrous 

tumor of the kidney: report of a case and comprehensive review of the 

literature. Arch Pathol Lab Med 2006;130:857-61. 

2 Magro G, Emmanuele C, Lopes M, et al. Solitary fibrous tumour of the 

kidney with sarcomatous overgrowth. Case report and review of the 

literature. APMIS 2008;116:1020-5. 

3 Hirano D, Mashiko A, Murata Y, et al. A case of solitary fibrous tumor 

of the kidney: an immunohistochemical and ultrastructural study with 

a review of the literature. Med Mol Morphol 2009;42:239-44. 

4 Argani P, Faria PA, Epstein JI, et al. Primary renal synovial sarcoma: 

molecular and morphologic delineation of an entity previously included 

among embryonal sarcomas of the kidney. Am J Surg Pathol 

2000;24:1087-96. 

5 Kim DH, Sohn JH, Lee MC, et al. Primary synovial sarcoma of the 

kidney. Am J Surg Pathol 2000;24:1097-104. 

6 Jun SY, Choi J, Kang GH, et al. Synovial sarcoma of the kidney 

with rhabdoid features: report of three cases. Am J Surg Pathol 

2004;28:634-7. 

7 Bégueret H, Galateau-Salle F, Guillou L, et al. Primary intrathoracic 

synovial sarcoma: a clinicopathologic study of 40 t(X;18)-positive 

cases from the French Sarcoma Group and the Mesopath Group. Am 

J Surg Pathol 2005;29:339-46. 

8 Mosquera JM, Fletcher CD. Expanding the spectrum of malignant progression 

in solitary fibrous tumors: a study of 8 cases with a discrete 

anaplastic component – is this dedifferentiated SFT? Am J Surg Pathol 

2009;33:1314-21. 

9 Vujanić GM, Kelsey A, Perlman EJ, et al. Anaplastic sarcoma of the 

kidney: a clinicopathologic study of 20 cases of a new entity with 

polyphenotypic features. Am J Surg Pathol 2007;31:1459-68. 

Oncocytic adrenocortical neoplasms – a distinct 

entity. report of 13 additional cases with emphasis 

on new diagnostic criteria and clinicopathologic 

correlation 

1)Bisceglia M. 2)Wong D.D. 3)Havlat M.F. 4)McCallum D 

5)Platten M.A. 6)Spagnolo D.V. 


San Giovanni Rotondo, Italy 2)Department of Anatomic Pathology, 

PathWest Laboratory Medicine, Perth, Western Australia 3)Department of 

Histopathology, St John of God Pathology, Subiaco, Western Australia 4) 

Department of Anatomic Pathology, PathWest Laboratory Medicine, Perth, 

Western Australia 5) Department of Anatomic Pathology, PathWest Laboratory 

Medicine, Perth, Western Australia 6) Department of Anatomic 

Pathology, PathWest Laboratory Medicine, Perth, Western Australia 

Background. The first oncocytic adrenocortical neoplasm (OAN) 

was reported in 1986 1 . Over the ensuing 25 years OAN has been 

established as a distinct entity, and interest is now largely focused 

on diagnostic criteria and predicting their behaviour. The original 

Weiss system 2 for conventional (nononcocytic) adrenocortical 

neoplasms does not apply to OAN. The Lin-Weiss-Bisceglia 

(LWB) system (applicable for resectable neoplasms only) was 

proposed in 2004-2005 3 4 , and is gaining widespread acceptance 

5 6 . In the LWB system definitional criteria, major criteria, 

and minor criteria allow classification of OANs as benign, borderline 

or malignant. 

Objectives. 1. To describe the clinicopathologic features of 

13 new OANs; 2. to review retrospectively all OANs from the 

world literature in the context of the LWB criteria; 3. to analyse 

statistically and correlate outcome data according to LWB tumor 

categories; and 4. to assess if there are behavioural differences 

between malignant OAN and conventional adrenocortical carcinoma. 

Methods. A systematic search of our combined databases was 

performed to identify cases of “pure” OANs as previously 

defined 3 4 , and confirmed as oncocytic immunohistochemically 

and/or ultrastructurally. A comprehensive PubMed-Medline 

search was performed between January 1980 and August 2009. 

Follow-up data were collected for all reported cases and median 

and 5 year survivals were estimated using the Kaplan-Meier


method. Differences in survival curves between cases classified 

histologically as benign, borderline and malignant were analysed 

using the Log-Rank test. 

Results. We found 13 new OANs in 7 females and 6 males with 

a median age of 41 years (range 22-69). 6 patients showed either 

clinical or biochemical hormone hypersecretion. All tumors 

were encapsulated: median size 80mm (range 7-285), median 

weight 155g (range 15-5720). According to LWB criteria 3 

were benign, 2 borderline and 8 malignant. Of the latter, local 

recurrence occurred in 3, distant metastases in 1 and death in 3. 

1 case was associated with an ipsilateral adrenal myelolipoma 

and 1 (gigantic) malignant OAN is the largest on record. The 

occurrence of “small oncocytes” was a frequent focal finding. 

All tumors were strongly immunopositive with mES-13 and 

9 were immunoreactive for calretinin, a novel finding in this 

context. All 4 cases examined ultrastructurally showed typical 

oncocytic features with an abundance of mitochondria. Kaplan- 

Meier curves for recurrence/metastases and death (p < 0.001 for 

both, using Log-Rank test), were applied to 84 of 109 cases (our 

13 OAN and 96 from the literature) with sufficient data to allow 

analysis. This revealed the ability of the LWB system to reliably 

predict future risk in OAN. The overall median survival for 

malignant OAN was 58 months (95%CI 27.5 to 88.5), notably 

better than the reported 14-32 months for conventional adrenocortical 

carcinoma 7 . 

Conclusions. 1. We report 13 new cases of OANs; 2. report the 

value of mES-13 immunostaining in establishing oncocytic differentiation; 

3. show the value of the LWB criteria in categorising 

OAN as benign, borderline or malignant; and 4. provide 

preliminary evidence of a better prognosis for malignant OANs 

compared with conventional adrenocortical carcinomas. 

references 

1 Kakimoto S, Yushita Y, Sanefuji T, et al. Non-hormonal adrenocortical 

adenoma with oncocytoma-like appearances. Hinyokika Kiyo 

1986;32:757-63. 

2 Weiss LM. Comparative histologic study of 43 metastasizing and nonmetastasizing 

adrenocortical tumors. Am J Surg Pathol 1984;8:163- 

9. 

3 Bisceglia M, Ludovico O, Di Mattia A, et al. Adrenocortical oncocytic 

tumors: report of 10 cases and review of the literature. Int J Surg 

Pathol 2004;12:231-43. 

4 Bisceglia M, Ben-Dor D, Pasquinelli G. Oncocytic Adrenocortical 

Tumors. Pathol Case Rev 2005;10:228-42. 

5 Lack E. Adrenal Cortical Carcinoma. Tumours of the Adrenal Gland 

and Paraganglia. 4th ed. Washington DC: Armed Forces Institute of 

Pathology 2008. 

6 Lau SK, Weiss LM. The Weiss system for evaluating adrenocortical 

neoplasms: 25 years later. Hum Pathol 2009;40:757-68. 

7 Bilimoria KY, Shen WT, Elaraj D, et al. Adrenocortical carcinoma in 

the United States: treatment utilization and prognostic factors. Cancer. 

2008;113:3130-6. 

Na + /H + exchanger regulatory factor 1 (NHerf1) 

expression in colorectal cancerogenesis 

1)”Mangia A. 1)Bisceglie D. 2)Malfettone A. 3)Asselti M. 

4)Bellizzi A. 5)Daprile R. 6)Paradiso A. 7)Simone G. 

1)Clinical experimental oncology laboratory, Ncc “Giovanni Paolo II”, 

Bari, Italy 2)Clinical experimental oncology laboratory, Ncc “Giovanni 

Paolo II”, Bari, Italy 3)Department of pathology, Ncc “Giovanni Paolo 

II”, Bari, Italy 4)Clinical experimental oncology laboratory, Ncc “Giovanni 

Paolo II”, Bari, Italy 5)Department of pathology, Ncc “Giovanni 

Paolo II”, Bari, Italy 6)Clinical experimental oncology laboratory, Ncc 

“Giovanni Paolo II”, Bari, Italy 7)Department of pathology, Ncc “Giovanni 

Paolo II”, Bari, Italy 

Background. Na + /H + exchanger regulatory factor 1 (NHERF1) 

is a candidate tumor suppressor gene. NHERF1 protein expression 

has been demonstrated to be altered in several cancers. An 

increased cytoplasmic NHERF1 expression suggests a key role of 

261 

its localization/compartmentalization in defining cancerogenesis 

and progression, but its role in colorectal carcinoma remains still 

undefined. 

Methods. We examined immunohistochemically the expression 

pattern and sub-cellular localization of NHERF1 in 51 patients 

with advanced colorectal cancers matched with surrounding 

nontumoral epithelium, in metastatic lymph nodes and hepatic 

metastases from each patient. 

Results. NHERF1 showed a different localization and expression 

in the different compartments of colorectal cancer samples. 

In nontumoral epithelium tissues, NHERF1 immunoreactivity 

was present as cytoplasmic, membranous and nuclear staining, 

while in tumor and metastatic tissues NHERF1 was present as 

diffuse cytoplasmic and nuclear staining. The median of cytoplasmic-NHERF1 

positive cells was significantly higher in primary 

tumors (70%), metastatic lymph nodes (60%) and hepatic 

metastases (70%) than normal tissues (10%) (p < 0.0001). In 

contrast, we had observed a low membranous protein expression 

in all tumoral tissues examined respect to normal tissues (0% vs 

5% respectively; p < 0.0001). Nuclear-NHERF1 expression was 

higher in tumor (18%) and metastatic tissues (15%) than normal 

tissues (11%) (p = 0.006; p < 0.01 respectively). 

Colorectal cancerogenesis is characterized by increased cytoplasmic 

expression of NHERF1 as the tumour progresses, suggesting 

its role in this process. The switch from membranous to cytoplasmic 

expression is compatible with a dual role for NHERF1 as a tumour 

suppressor or tumour promoter dependent on its sub-cellular 

localization. Indeed, the increasing nuclear NHERF1 expression 

suggest that this protein can move to the nucleus and may induce 

expression of genes determining the malignant phenotype. 

references 

Cardone RA, et al. The NHERF1 PDZ2 domain regulates PKA-RhoAp38-mediated 

NHE1 activation and invasion in breast tumor cells. 

Mol Biol Cell 2007;18:1768-80. 

Mangia A, et al. Biological role of NHERF1 protein expression in breast 

cancer. Histopathology 2009;55:600-8. 

Song J, et al. Expression and clinicopathological significance of oestrogenresponsive 

ezrin-radixin-moesin-binding phosphoprotein 50 in 

breast cancer. Histopathology 2007;51:40-53. 

expression of p-AKT and p-mTOr in a large series 

of BP-NeTs 

1)Boldrini L. 2)Capodanno A. 3)Servadio A. 4)Rotondo M I. 

5)Pelliccioni S. 6)Fontanini G. 

1)Surgery, Santa Chiara Hospital pisana, Pisa, Italy 2)Surgery, Santa 

Chiara Hospital, Pisa, Italy 3)Surgery, Santa Chiara Hospital, Pisa, Italy 

4)Surgery, Santa Chiara Hospital, Pisa, Italy 5)Molecular Diagnostic, 

Santa Chiara Hospital, Pisa, Italy 6)Surgery, Santa Chiara Hospital, 

Pisa, Italy 

Background. Bronchopulmonary neuroendocrine tumors (BP- 

NETs) comprise about 20% of all lung cancers. They are separated 

into 4 subgroups: typical carcinoid tumor (TC), atypical carcinoid 

tumor (AC), large-cell neuroendocrine carcinoma (LCNEC), 

and small-cell lung carcinoma (SCLC), which exhibit different 

biological characteristics that have been extensively investigated 

to identify features for diagnosis, prognosis and therapy for this 

special lung tumor category. 

The signalling pathway involving AKT/mTOR (the mammalian 

target of rapamycin) is one of the main regulators of 

cell growth and proliferation and is located at the crossroad of 

several major signal transduction molecules (PTEN/Pi3-kinase, 

AMKP, Ras/Raf). The only available literature data on AKT/ 

mTOR in NE lung tumours are represented by experimental 

models in SCLC cells. The purpose of this study was to evaluate 

the expression of phosphorylated AKT/mTOR in a large 

series of BP-NETs and to investigate the correlations with 

clinicopathological parameters.

262 

Methods. p-AKT (Ser473) and p-mTOR (Ser2448) were 

determined by immunohistochemistry in a series of 210 BP- 

NETs, including 85 SCLC, 17 LCNEC, 26 AC, 75 TC, and 7 

tumorlets. 

Results. High p-AKT expression was found in the majority of 

tumorlets and carcinoids, whereas a lower expression was found 

in SCLC and LCLNC p = 0.0001). The expression of p-mTOR 

was also statistically different in tumorlets and carcinoids, that 

showed higher p-mTOR expression, comparing with SCLC and 

LCNEC (p = 0.0002). Furthermore, p-mTOR expression was 

higher in T1-T2 tumor stages compared to higher stages in all 

BP-NETs (p = 0.0008). 

Our results suggest a role for Akt/mTOR pathway in BP-NETs, 

particularly in carcinoids. Moreover, mTOR could represent a 

useful marker in this type of tumors with important applications 

in the clinico-therapeutic management of patients. 

Granulomatous reaction in gastric carcinomas: an 

immunohistochemical and ultrastructural study 

1)R. Caruso 1)Bonanno A. 2)Quattrocchi E. 3)Napoli P. 4)Fedele 

F. 

1)Patologia umana, Policlinico universitario, Messina, Italia 2)Patologia 

umana, Policlinico universitario, Messina, Italia 3)Servizio anatomia patologica, 

Ospedale papardo, Messina, Italia 4)Patologia umana, Policlinico 

universitario, Messina, Italia 

Granuloma is a focal, compact collection of inflammatory cells 

in which mononuclear phagocytes predominate. The authors 

report 9 cases of papillary-tubular gastric adenocarcinomas 

characterized by mature granulomas associated with recent microhemorrhages. 

Mature granulomas were composed of foamy, 

CD68-positive histiocytes with occasional giant cells. Hemosiderin-containing 

macrophages were present in the tumor stroma, 

suggesting phagocytosis of erythrocytes. Under electron microscopy, 

mature (nonepithelioid) granulomas and clusters containing 

1 macrophage and 1-3 eosinophils were found. This study 

provides morphological examples of skewed type II macrophage 

infiltration in gastric adenocarcinomas that is involved in scavenging 

activity, particularly erythrophagocytosis, formation of 

mature (nonepithelioid granulomas), and heterotypic aggregation 

with eosinophils. 

left ventricular hemangioma 

1) Bondi F. 2)Del Giglio 

1)Endocrinologia, Ospedale S. Maria delle croci, Ravenna, Italia 2)Department 

of cardiovascular surgery, Villa maria cecilia hospital, Cotignola, 

lugo (ra), Italia 

Background. Primary cardiac tumors are rare. The large majority 

of cardiac tumors are benign; hemangiomas account for < 10% 

of all primary cardiac tumors in children and they are usually asymptomatic 

when diagnosed after infancy. Cardiac hemangiomas 

are often found incidentally at autopsy or with imaging, usually 

hocardiography. 

Metohods. A 16-year-old previously healthy boy presented with 

a heart murmur and was found by transthoracic echocardiography 

to have a single mobile tumor in the left ventricular. A diagnosis 

of probable cardiac hemangioma was made on the basis of its 

MRI signal intensity characteristics indicating high vascularity. 

The polipoid mass appeared to be localized in the left ventricle 

and its implant base was in the lateral border of the posterior 

papillary muscle. The tumor was surgically excised. 

Results. At gross inspection, tumor consisted of exophytic 

polypoid mass. The size was 1.7 × 1.5 × 1 cm. On cut section, 

tumor had microcytic appearance with areas of hemorrhage. 

Histopathological features were consistent with an unusual type 

of hemangioma composed of large, endothelial-lined, thin-walled 

channels and intervening dense proliferation of capillary-sized 


vessels. Although hemangioma was predominantly exophytic, 

there was infiltration of superficial myocardium. No evidence of 

atypia, cellular pleiomorphism, high mitotic count, or necrosis 

were found. Immunohistochemical profile of tumor consistent 

with with strong staining for CD31 and factor VIII. The diagnosis 

of cardiac hemangioma, capillary type, was made. 

Conclusions. Cardiac hemangiomas are rare tumors therefore 

it is difficult to make a definitive preoperative diagnosis. Other 

cardiac tumors that may have strong gadolinium enhancement 

include pheochomocytoma, angiosarcoma, myxoma, and rhabdomyosarcoma. 

Cardiac angiosarcomas are exceptionally aggressive, 

are usually large, centrally necrotic, and frequently extend 

into the pericardium. 

The cell blocks: it could be a real -biopsy 

1)Bondi F. 2)Salerno V. 

1)Endocrinologia, Ospedae S. Maria delle croci, Ravenna, Italia 2)Oncologia, 

Ospedale Umberto I, Lugo, Italia 

Background. For pathologist, an essential step in the mastery of 

aspiration cytology is the ability to translate the cytologic patterns 

into histologic tissue patterns of diagnostic value. 

The fine needle aspiration cytology (FNAC) in nodular lesions 

has a limited diagnostic use for the impossibility to obtain multiple 

sections for an immunohistochemical analysis. 

Methods. Often from standard FNA is possible to obtain thin 

cores or multiple tissue fragments, especially in tissue rich of cell 

as lymph node and solid tumours. The FNA samples, previously 

centrifugation, are assembled with a drop of tromboplastina to 

produce a clot. The clot is fixed in 10% solution of buffered isotonic 

formalin and processed as for routine histology. Cell blocks 

may give some idea of tissue architecture and allow multiple section 

for immunohistochemistry. 

Results. We always prepare the cell blocks and a cytologic 

smearing from fresh material in FNA of neoplastic lesions from 

different organs and tissues. This gives us tissue fragments for 

value histologic pattern of the lesions and on which perform 

immunohistochemistry and/or the molecular pathology (FISH, 

EGFR, K-ras ecc). In the review of our series we have observed 

that the cell blocks is useful to differentiate tumoral histotypes 

(in particular of the parotid gland and of the lung), primary from 

metastatic tumours, lymphomas, undifferentiated carcinomas 

from sarcomas and melanomas, neuroendocrine tumours and it 

was essential to diagnose: parotid gland melanoma metastasis, 

lymph node alveolar rhabdomyosarcoma metastasis, lymph 

node gastric leiomyosarcoma (GIST) metastasis, thyroid gland 

colic ADK metastasis, adrenal gland leiomyosarcoma, giant cells 

MFH, pulmonary angiosarcoma. 

follow-up of borderline cervical cytology cases 

negative for atypia with indication to repeat pap 

test in a group of spontaneous screening 

1)Bonfadini M.G. 2)Magnani C. 3)Rostan I. 4)Marsico A. 

5)Navone R. 

1)Servizio di citologia, Clinica san gaudenzio, Novara, Italia 2)Servizio di 

citologia, Clinica san gaudenzio, Novara, Italia 3)Sc. biomediche e oncologia 

umana-sez. anatomia pat., Universita di torino, Torino, Italia 4)U.o. 

di anatomia patologica, Osp. koelliker, Torino, Italia 5)Sc. biomediche e 

oncologia umana-sez. anatomia pat., Universita di torino, Torino, Italia 

Background. The Bethesda 2001 System foresees a diagnosis of 

ASC-US, ASC-H and AGC for borderline lesions and a subdivision 

of the cervical slides into negative or positive, with reactive 

cell changes (RCC) placed into the negative category. Our 

research, using an adequate follow-up, i.e. at least 2 cytological 

tests and/or 1 negative histological result, in a case group of cervical 

cytology with a 3-9 year follow-up) aims to establish a final 

diagnosis, both for borderline atypia and RCC.


Materials. 146,020 cytological cervical samples showed 1,845 

ASC\AGC (1.3%), 604 (0.4%) L-SIL, 432 (0.3%) H-SIL, 56 

squamous carcinoma (0.04%) and 33 adenocarcinoma (0.02%). 

Amongst the 1,845 borderline cases, 455 of the ASC and 54 

of the AGC had a follow-up considered to be sufficient, as did 

806/4,577 with a negative diagnosis for atypia with indication for 

a further cytology test to assess RCC (mainly infections, above 

all vaginosis) and ASC-AGC with a reduced legibility (quality) 

of the samples. 

Results. 305/ 445 cases of ASC (68.6%) were benign, 97 (21.8%) 

SIL were low grade, 37 (8.3%) SIL were high grade and 6 (1.3%) 

were carcinoma. 35/54 cases of AGC (64.8%), were benign and 

19 (35.2%) were malignant (13 H-SIL, 1 squamous carcinoma 

and 5 adenocarcinoma). When the ASC were subdivided into 

ASC-US and ASC-H, in the 1 st group (393 cases), there were 

295 (77.0%) definitive benign diagnosis, 75 (17.8%) L-SIL, 20 

(4.7%) H-SIL and 5 carcinoma (0.8%). Whilst in the 2nd group, 

(37 cases) there were 11 (37.9%) negative cases, 7 (20.7%) L- 

SIL, 18 (37.9%) H-SIL and 2 (3.5%) squamous carcinoma. 

764/806 RCC (94.8%) were benign, 29 (3.6%) were low grade 

SIL, 11 were high grade SIL (1.4%), 1 (0.1%) was a squamous 

carcinoma and 1 (0.1%) adenocarcinoma. 

In conclusion, a high predictive capacity was confirmed for ASC- 

H as was the possibility of false negatives for the RCC, due to 

poor quality samples. This is in line with the Bethesda System 

management recommendations for ASC-US, where a repeated 

Pap test after adequate therapy, in the case of infection, is recommended. 

Ki67 and p53 immunohistochemical evaluation in 

malignant and potentially malignant oral lesions 

based on samples obtained by curette 

1)Bonfadini M.G. 2)Magnani C. 3)Rostan I. 4)Pentenero M. 

5)Gandolfo S. 6)Navone R. 

1)Servizio di citologia, Clinica san gaudenzio, Novara, Italia 2)Servizio di 

citologia, Clinica san gaudenzio, Novara, Italia 3)Sc. biomediche e oncologia 

umana-sez. anatomia pat., Universita di torino, Torino, Italia 4)Sc. 

cliniche e biologiche-sez. med. e oncologia orale, Universita di torino, 

Torino, Italia 5)Sc. cliniche e biologiche-sez. med. e oncologia orale, Universita 

di torino, Torino, Italia 6)Sc. biomediche e oncologia umana-sez. 

anatomia pat., Universita di torino, Torino, Italia 

Background. As oral squamous carcinoma is often diagnosed 

in the late stages, its survival rate is low. This may be due to 

the diagnostic difficulty and the fact that the lesion may develop 

without evident dysplastic morphology. It is well known that 

both Ki67 and p53, may be good markers for neoplastic and 

preneoplastic oral lesions, as is DNA content. Therefore, we 

compared the results of immunohistochemistry (IIC) to those of 

the DNA ploidy and the microhistological diagnosis, according 

to the method already described by our group [Navone R et al. 

Oral Potentially Malignant Lesions: First Level Microhistological 

Diagnosis from Tissue Fragments Sampled in Liquid-Based 

Diagnostic Cytology. J Oral Pathol Med 2008;37:358-363]. 

Methods. Curette sampling (AcuDispo Curette, Acuderm inc) 

was carried out in 111 patients with lesions suspicious for carcinoma 

or potentially malignant lesions (PMLs) of the oral cavity. 

Microhistology was done and the immunohistochemical reactions 

assessed (Ki 67 e p53) and the data of IIC were compared to the 

ploidy data already published by our group (Pentenero M et al.: 

DNA Aneuploidy and dysplasia in oral potentially malignant 

disorders. Oral Oncol 2009, 45:887-890). 

Results. 11/111 cases were squamous carcinoma, 23 high grade 

dysplasia, 22 low grade lesions and 55 keratosis without dysplasia. 

The Ki67 (p = 0.00006) and the dysplasia grade had a statistically 

positive correlation; suprabasal p53 had a lower correlation 

(p = 0.02) as it was positive also for some keratosis cases without 

evidence of dysplasia. 

263 

In conclusion, in the light of the strong correlation with preneoplastic 

and neoplastic lesions, above all the possibility of progression, 

Ki67 and p53 IIC seems to be useful in oral PMLs. Clinical 

follow-up is indicated for p53 positive cases without dysplasia. 

expression of stem cells markers CD133, CD117, 

and CD44 in prostatic adenocarcinomas is not 

associated with stage and grading 

1)Bosisio F.M. 2)Leone B.E. 

1)Scienze chirurgiche (università milano-bicocca), Desio, Desio, Italia 

2)Scienze chirurgiche (università di milano-bicocca), Desio, Desio, Italia 

Background. The stem cells phenotype, when present in tumours, 

may be able to explain some features of neoplastic progression, 

as invasiveness or metastatization, and is hypotetically related to 

a more aggressive behaviour. Stemness in prostatic cancer cells 

have been studied so far only in cancer cell lines or in restricted 

groups of cases. Aim of this study is to correlate the expression 

of stem cells markers CD133, CD44, and CD117 in 113 cases of 

acinar adenocarcinoma of the prostate primarily with tumor stage 

and grading of the neoplasia, then with other prognostic and differentiation 

variables, such as immunohistochemical staining for 

CXCR4, p53, cyclin D1, e-cadherin, vimentin, Ki-67 proliferation 

index, and basal, luminal or neuroendocrine phenotype. 

Methods. Immunohistochemistry for CD133, CD117, and CD44 

was performed on tissue microarrays of 113 prostate adenocarcinomas. 

Data were correlated to stage and grade, and with other 

immunohistochemical markers of prognostic or differentiation 

significance by statistical analysis. 

Results. CD133 resulted positive in 21 out of 113 cases (18.5%), 

CD44 in 86 out of 113 (76.1%), CD117 in 87 out of 113 (76.9%). 

A simultaneous expression of the three stem cell markers 

(CD133+, CD117+, CD44+) was found in 15 cases (13.3%). 

None of the stemness markers, individually or simultaneously 

considered, showed any kind of correlation with stage, grading, 

and prognostic and differentiation markers, with the exception of 

CXCR4, a putative mediator of invasiveness and itself related to 

staminal phenotype. 

Conclusion. The hypothetic stem cell phenotype of the prostate 

cancer cells, defined by CD133, CD44, and CD117, is not able to 

distinguish a subset of tumours with specific prognostic or differentiation 

features. Selection of other tumor-initiating cell markers 

or resolution of technical problems related to the choice and use 

of monoclonal antibodies is mandatory to better explore this field 

of knowledge of tumor biology. 

Cyclin D1 overexpression in ewing’s sarcoma/ 

PNeT: a potential marker helpful in the differential 

diagnosis of small round cell tumours 

1)F. Brancato, 2)R. Alaggio, 1)A. Gurrera, 1)E. Vasquez, 1)G. 

Magro 

1)Dipartimento G.F. Ingrassia, Anatomia Patologica - Università di Catania, 

Azienda Ospedaliero Universitaria Policlinico Vittorio Emanuele, 

Catania, Italia; 2)Anatomia Patologica, Università di Padova, Azienda 

Ospedaliera, Padova, Italia 

Background. Ewing’s sarcoma(ES)/neuro-ectodermal primitive 

tumour (PNET) is a small round cell sarcoma showing varying 

degrees of neuroectodermal differentiation and the characteristic 

t(11;22) (q24q12) chromosomal translocation. Although the diagnosis 

of ES/PNET is morphologically suspected, immunohistochemical 

analysis, including CD99, HNK1, FLI1 protein and 

caveolin-1, is mandatory in confirming it. Unfortunately, these 

markers are not highly specific, being also expressed in a wide 

variety of pediatric small round cell tumours. In vitro studies have 

shown that tumour cell lines of ES/PNET overexpress cyclin D1, 

suggesting its key role in the mechanisms that regulate normal

264 

cell cycle during G1-S. Additionally, in one immunohistochemical 

study the expression of cyclin D1 was found in approximately 

42% of ES/PNETs, but without any significant correlation with 

prognosis. 

Methods. The aim of this paper was to study the comparative 

immunohistochemical expression of cyclin D1 in 20 cases of pediatric 

ES/PNETs, 10 cases of embryonal rhabdomyosarcoma, 10 

cases of alveolar rhabdomyosarcoma, including the solid variant, 

and 5 cases of desmoplastic round cell tumours to assess its potential 

usefulness in the differential diagnosis of these tumours. 

Results. Notably 100% of ES/PNETs expressed cyclin D1, 

with a diffuse extension, ranging from 60 to 100% of neoplastic 

cells. In contrast, desmoplastic round cell tumours showed 

immunoreactivity restricted to 10-20% of cells, whereas both 

embryonal and alveolar rhabdomyosarcomas lacked cyclin D1 

immunoreactivity. Our preliminary results suggest that immunohistochemical 

cyclin D1 overexpression may be exploitable 

as an additional marker in the differential diagnosis of Ewing’s 

sarcoma/PNET, rhabdomyosarcoma and desmoplastic small 

round cell tumours. This finding, evaluated appropriately in 

the context of a large panel of antibodies, may be helpful especially 

when dealing with small incisional biopsies or ambiguous 

immunohistochemical results due to sub-optimal fixation, 

non-specific immunoreactivity or polyphenotypic profile by 

neoplastic cells. 

Abnormalities of chromosome 3 and 3q in 

squamous lung carcinoma: genotypic patterns 

with potential clinical impact 

Brunelli M., Eccher A., Martignoni G., Brunello E., Parolini C., 

Pedron S., Menestrina F., Chilosi M. * 

1)Department of pathology and diagnostic, Policlinico G.B. Rossi, Verona, 

Italy; *Department of pathology and diagnostic, Policlinico G.B. 

Rossi, Verona, Italy 

Background. Amplification of chromosome 3q is the most common 

genomic aberration in squamous pulmonary carcinoma, 

however few reports distinguish chromosomal amplification 

due to an increase of the locus specific region 3q or to the entire 

chromosome 3. Nowadays, the distinction of the primary event 

(amplification) vs the second (polysomy of a chromosome) is 

mandatory, due to potential impact at a diagnostic or prognostic 

levels. We sought to evaluate the subtypes of genotypic abnormalites 

of the entire chromosme 3 and the distal locus specific 3q 

in a serie of squamous lung adenocarcinoma. 

Methods. 18 squamous lung adenocarcinomas were recruited. 

Immunophenotyping was performed by using antibodies for CK5, 

p63, TTF-1 and CK7. Fluorescence in situ hybridization analysis 

(FISH) was used to assess the centromeric region of the chromosome 

3 (CEP3) and the locus specific gene (LSI) 3q (Olympus). 

Polysomy of chromosome 3 without 3q amplification (more than 

three CEP3 fluoresecent signals in 18% of the neoplastic nuclei), 

polysomy of chromosme 3 with 3q amplification (ratio LSI 3q/ 

CEP3 > 2.2 in polysomic cells) and amplification of LSI 3q without 

polysomy of chromosome 3 were differently scored. 

Results. All cases displayed CK5 and p63 positivity. TTF-1 and 

CK20 were negative. Focal CK7 was observed in 12/18 cases. 

4/18 (22%) squamous lung adenocarcinoma showed amplification 

of 3q without polysomy of chromosome 3, 9/18 (50%) polysomy 

of chromosome 3 with 3q amplification and 5/18 (28%) 

polysomy of chromosome 3 without 3q amplification. Overall, 

squamous pulmonary carcinoma usually show centromeric and 

locus specific abnormalites on chromosome 3/3q; however there 

are three distinctive patterns that may have potential value at 

the prognostic level or when evaluating different therapeutical 

strategies. The clinical impact of these multifaceted genotypic 

abnormalities need further investigation. 


expanding immunophenotypical and molecular 

features of tubulo-cystic renal cell carcinoma 

1)Brunelli M. 2)Segala D. 3)Gobbo S. 4)Bersani S. 5)Bragantini 

E. 6)Gardiman M. 7)Tardanico R. 8)Chilosi M. 9)Menestrina F. 

10)Martignoni G. 

1)Pathology, University of verona, policlinico “G.B. Rossi”, Verona, Italy 




5)Pathology, Ospedale “santa chiara”, Trento, Italy 6)Pathology, University 

of padua, Padova, Italy 7)Pathology, spedali civili, Brescia, Italy 



10)Pathology, University of verona, policlinico “G.B. Rossi”, Verona, 

Italy 

Background. Among new emerging description of renal neoplasms, 

the tubulo-cystic renal cell carcinoma may be considered 

a unique morphologic entity, with its distinctive gross and microscopic 

features. However, before it is accepted as a distinct renal 

cell carcinoma subtype, further studies are needed to document 

a characteristic molecular signature associated with this tumor. 

It has also been questioned its relationship to papillary renal cell 

carcinoma. We sought to evaluate the fluorescent molecular signature 

expanding the chromosomal in situ analysis. 

Methods. Ten tubulo-cystic renal cell carcinoma were recruited, 

5 of which from a single patient. Clinico-pathological analysis 

were recorded. Immunophenotypical analysis using monoclonal 

antibody against Cytokeratin 7 (CK7), S100A1, Parvalbumin 

(PV), AMACR, CD10 were performed. Chromosomes 7, 12, 16, 

17, 20 and Y and locus specific gene 7q31 (c-met), c-myc, EGFR, 

p53, Her-2 were tested. 

Results. Patients age ranged from 45 to 67, with a male preponderance 

(5:1). One patient showed 5 similar nodules. Another 

patient presented a synchronous papillary renal cell carcinoma. 

Tumours had a diameter ranged from 0,8 to 3,5 cm and all staged 

pT1a. Grading sec. Furhman was G1-G2 in 7 cases and G3 in 3 

cases. Cases stained immuno-positive for CD10 (10/10, 100%), 

S100A1 (10/10, 100%) and AMACR (9/10, 90%); PV was 

weakly and focally positive in 3 cases (3/10, 30%), while only 

one case immunoexpressed CK7 (1/10, 10%). Entrapped tubules 

into the neoplasms were positive for CK7. LSI-7q31 c-met was 

gained in all cases. Two out of 5 gained chromosome 7 and 17. 

Three out of 5 cases showed gains of p53, c-myc, EGFR. One 

case showed loss of Y. Chromosome 20 were wild. The LSI 

EGFR set wild. 

Tubulo-cystic renal cell carcinoma are low staged and graded 

tumours. Findings of c-met gains is similar to those reported in 

papillary renal cell carcinoma, however the other chromosomes 

do not show overlapping features. 

New emerging morphological subtypes of papillary 

renal cell carcinoma: gains of the 7q31 (C-MeT) 

as a molecular signature 

1)Brunelli M. 2)Segala D. 3)Gobbo S. 4)Bersani S. 5)Eccher A. 

6)Chilosi M. 7)Menestrina F. 8)Martignoni G. 









Background. Papillary renal cell carcinomas (RCCs) are morphologically 

divided into type 1 and type 2. Recently, other three 

subtypes of papillary RCCs have been described such as the on-


cocytic and the spindle cells variants and papillary RCCs showing 

clear cell changes. While gains of the entire chromosomes 7 and 

17 have been described in all aforementioned subtypes of papillary 

RCCs, the status of the 7q31 gene was not assessed in the 

new emerging subtypes. Gains of the locus specific region 7q31 

(C-MET) in considered an hallmark of the papillary subtype of 

RCCs. 

Methods. 35 papillary RCCs including 12 type 1, 12 type 2, 6 

oncocytic, 2 with spindle cells and 3 with clear cell changes were 

recruited. Immunohistochemical analysis included AMACRracemase 

and cytokeratin 7. Chromosome 7 and 17 have been 

assessed by fluorescence in situ hybridization analysis by using 

centromeric CEP7 and CEP17 (Olympus) probes. The locus specific 

probes mapping the 7q31 region (C-MET) was also used. 

Single, double and gains of fluorescent signals was scored per 

neoplastic nuclei per each case. 

Results. AMACR stained all cases. Cytokeratin 7 stained all 

cases except 4 type 2, 3 oncocytic and 2 spindle cells subtypes 

of papillary RCCs. Both chromosomes 7 and 17 gains were 

observed in 11/12 type 1, in 8/12 type 2, 4/6 oncocytic, in 2/2 

spindle cells and in 3/3 with clear cell changes tumours. Gains 

of the locus specific gene 7q31 was observed in all type 1, 9/12 

type 2, in 4/6 oncocytic, in 2/2 spindle cells and in 3/3 with clear 

cell changes tumours. The mean score for assessing chromosomal 

gains was 45% (range 15 to 76%). 

In conclusion, the new emerging morphological subtypes of papillary 

RCCs have the molecular signature, such as gains of the 

locus specific gene 7q31 (C-MET), that belongs to the genomic 

profile of the papillary neoplasms. These findings may have a 

primary potential value at a diagnostic level. 

lack of the tailored use of anthracycline 

in the lobular subtype of breast carcinoma: 

evidence on the Topoisomerase-IIA Amplicon 

1)Brunello E. 2)Brunelli M. 3)Manfrin E. 4)Nottegar A. 

5)Bersani S. 6)Vergine M. 7)Menestrina F. 8)Martignoni G. 

9)Bonetti F. 

1)Pathology and diagnostic, Policlinico G.B. Rossi, Verona, Italy 2)Pathology 

and diagnostic, Policlinico G.B. Rossi, Verona, Italy 3)Pathology and 

diagnostic, Policlinico G.B. Rossi, Verona, Italy 4)Pathology and diagnostic, 

Policlinico G.B. Rossi, Verona, Italy 5)Pathology an, Policlinico G.B. 

Rossi, Verona, Italy 6)Pathology and diagnostic, Policlinico G.B. Rossi, 

Verona, Italy 7)Pathology and diagnostic, Policlinico G.B. Rossi, Verona, 

Italy 8)Pathology and diagnostic, Policlinico G.B. Rossi, Verona, Italy 

9)Pathology and diagnostic, Policlinico G.B. Rossi, Verona, Italy 

Background. In breast carcinoma, topoisomerase-IIa gene amplification 

appears to be a marker predictor of response to 

anthracyclines therapy, with few contrasting data. Interestingly, 

the lobular subtype usually does not respond to chemotherapies 

such as those including doxorubicin/anthracycline. Few data area 

available when matching topoisomerase-IIa gene and lobular 

breast carcinoma, thus we sought to analyze the topoisomerase- 

IIa status in the lobular subtype. 

Methods. 46 infiltrative lobular carcinomas, 13 with matched 

loco-regional lymph-nodal metastases were recruited. Tissue microarrays 

have been built by punching three neoplastic cores per 

case. Whole tumorous tissue sections were simultaneously evaluated. 

Topoisomerase-IIa gene amplification was analyzed by 

both chromogenic (Zytolight) (CISH) and fluorescent (Olympus) 

(FISH) in situ analyses. We also assessed the Her-2/neu status by 

CISH, FISH and SISH (Ventana). Amplification was scored as 

recommanded criteria. HER-2 immunoexpression was assessed 

by using Hercept test. 

Results. 44/46 (95%) of the cases did not reveal topoisomerase-IIa 

amplification whereas two of the 46 (5%) cases were 

amplified. Eleven of the 13 metastatic sites did not reveal amplification 

neither in the primary nor in matched metastases (85%); 

265 

the two remaining were amplified (15%). All cases revealed an 

homogeneous status on all three neoplastic cores. The two cases 

showing Her-2/neu and topoisomerase-IIa amplification scored 

3+ the remaining not-amplified cases scored 0 or 1+ in 40 and 

2+ in 4 cases. 

In conclusion, the infiltrative lobular subtype of breast carcinoma 

does not usually show topoisomerase-IIa gene amplification 

neither in the primary nor lymph-nodal metastases. In the era 

of personalised and tailored therapies, patients affected by the 

lobular subtype of breast carcinoma lack in most of the cases the 

biological rationale for receiving the common chemotherapy that 

include anthracycline. 

Subcutaneous ewing sarcoma / PNeT as a second 

cancer in a previously irradiated young patient. 

An uncommon type of post-irradiation soft tissue 

sarcoma 

1)Bruno M. 2)D‚Antona G.I. 3)Vita G. 4) Dicandia L. 

5)Bisceglia M. 

1)Division of Anatomic Pathology, Madonna delle Grazie Hospital, Matera, 

Italy 2)Division of Anatomic Pathology, Madonna delle Grazie Hospital, 

Matera, Italy 3)Division of Anatomic Pathology, IRCCS Institute 

of Cancer, Rionero in Vulture, Italy 4)Department of Pathology, IRCCS 

Casa Sollievo della Sofferenza Hospital, San Giovanni Rotondo, Italy 


San Giovanni Rotondo, Italy 

Background. A second cancer is defined as a histologically 

distinct cancer that develops in survivors after the first cancer. 

The risk determinants of second cancer are multifactorial, and the 

younger age of patients at the time of diagnosis of the first cancer, 

the exposure to high-dose radiation therapy, the administration 

of certain chemotherapeutic agents, and known genetic predisposition 

to cancer each play a role. Second cancers account for 

6-10% of all malignant tumor diagnoses in USA 1 . The increased 

relative risk of developing a second cancer has been assessed in 

both adults and children, and is higher (> 2-fold) in the latter 1 . 

Soft tissue sarcomas account for a small proportion of second 

cancers, with an estimated frequency of < 10% 2 . The most common 

histologic type of soft tissue sarcomas as second cancers 

include mostly high-grade sarcomas, such as rhabdomyosarcoma, 

malignant peripheral nerve sheath tumor, fibrosarcoma, leiomyosarcoma, 

synovial sarcoma, alveolar soft part sarcoma, and Ewing 

sarcoma / primitive neuroectodermal tumor (PNET). 

Objectives. To report a case of superficial soft tissue Ewing 

sarcoma / PNET as a second cancer in a young patient previously 

treated for Hodgkin’s disease (HD). 

Case Report. A 18-year old boy developed a palpable soft tissue 

mass at the level of the lateral border of his breast region, which 

was surgically removed. This young patient had a known history 

of HD, nodular sclerosis type (BNLI-II), stage IIB, involving the 

neck and mediastinal lymph nodes, which had been diagnosed 

approximately 4 years earlier. The patient had been treated with 

chemotherapy (ABVD regimen therapeutic protocol) and conventional 

three-dimensional radiotherapy (Photon 6-10MW with 

a total dose delivered of 25,5 Gy with a daily fraction of 150 

cGy). The excised tumor was 3 cm in size, grossly circumscribed, 

but unencapsulated, and on cut section appeared as a solid, greyish, 

and fleshy nodule, surrounded by healthy adipose tissue. Microscopically 

the tumor was composed of undifferentiated small 

round cells, with scanty to moderate glycogen-rich cytoplasm 

and vesicular nuclei, and showing scattered mitoses. Focally the 

resection margins were very near to tumor. Immunohistochemically, 

the tumor cells were diffusely positive for vimentin, CD99/ 

O13, and FLI-1, and negative for skeletal muscle, hematolymphoid, 

neural and neuroendocrine, and epithelial markers (EMA, 

cytokeratins w.s.). The tumor was diagnosed as Ewing sarcoma/ 

peripheral PNET, and a second local excision performed. Follow-

266 

up: the patient was given courses of chemotherapy (IVADO regimen 

therapeutic protocol). Currently he is alive with no evidence 

of disease at 1 year after the second cancer diagnosis. 

Discussion. The cancer cure rate in children has greatly improved 

with modern multimodal treatment protocols and nowadays approximately 

70% of overall pediatric patients previously treated 

for their (childhood) malignancies are long-term survivors 3 . 

Patients with childhood or adolescence cancer are at a 3.6-fold 

increased risk for development of a second cancer relative to the 

general population, and the estimated cumulative incidence of 

a second cancer is 3.5% at 25 years 1 . The determinants of the 

increased relative risk of developing a second cancer are factors 

that are partly host-related and partly therapy-related: the former 

including the increased susceptibility of stem cells to mutagenic 

effects and the higher rate of cell proliferation during development 

and differentiation (the substrate for their enhanced radiosensitivity), 

and for the latter radiation therapy and the usage of 

certain chemotherapeutic agents (alkylating drugs and anthracyclines) 

3 . The most common cancers associated with the development 

of second cancers are hereditary retinoblastoma, soft-tissue 

sarcomas, including Ewing sarcoma 2 , and HD 1 3 . The increased 

relative risk for the development of a second cancer in HD, the 

highest rate for a cancer with no known genetic predisposition 3 

may be at least partly due to the disease itself as an independent 

risk factor, and most likely to the specific chemo- and radiationtherapy 

required for treatment. The therapy-related factor in HD 

are anthracyclines and radiotherapy, mostly the latter 1-5 . The case 

herein described may be qualified as a post-irradiation sarcoma, 

based on the following standard criteria: previous exposure to irradiation, 

tumor site within the radiation field, latency of several 

years, and histologic distinction from the primary tumor. Postirradiation 

soft tissue sarcomas are a well known but rare entity 

1 2 4 5 . As to their therapy all low-grade tumors and high-grade 

tumors 5 cm or smaller may be treated with a margin-negative 

surgical excision, and systemic chemotherapy can be considered 

when a negative margin is difficult or impossible to obtain 6 . 

However it is suggested to treat also with chemotherapy those 

known chemosensitive soft tissue sarcomas, as Ewing sarcoma 

and rhabdomyosarcoma, which proved to be as chemosensitive 

as second neoplasms as they are as primaries 7 . Ewing sarcoma 

/ PNET has already been documented as second malignant neoplasms 

appearing outside the irradiated area following treatment 

for HD (1 case in the Italian joint AIEOP-INT/Milan series of 

cases registered during the period of 1979-2004), angiosarcoma 

or fibrosarcoma (1 case each 7 ), and in the irradiated area in a patient 

who had been treated for chronic myeloid leukemia (1 case 

– same series). Ewing sarcoma / PNET often is mostly a sarcoma 

of bone and deep soft tissue, occasionally involving even visceral 

organs. Ewing sarcoma occurs very rarely as a primary superficial 

soft tissue tumor 8 9 . 

Conclusions. Post-irradiation soft tissue sarcomas are rare. They 

occur only in irradiated tissues. HD is the most common childhood 

first malignancy at risk of developing second cancer later 

both in childhood and in adulthood. Our case is presented for its 

rarity and the described association. 

references 

1 Bhatia S, Sklar C. Second cancers in survivors of childhood CANCER. 

Nat Rew Cancer 2002;2:124-32. 

2 Nguyen F, Rubino C, Guerin S, et al. Risk of a second malignant neoplasm 

after cancer in childhood treated with radiotherapy: correlation 

with the integral dose restricted to the irradiated fields. Int J Radiation 

Oncology Biol Phys 2008;70:908-15. 

3 Neglia JP, Friedman DL, Yasui Y, et al. Second malignant neoplasms 

in five-year survivors of childhood cancer: childhood ancer survivor 

study. J Natl Cancer Inst 2001;93:618-29. 

4 Schneider U, Lomax A, Timmermann B. Second cancers in children 

treated with modern radiotherapy techniques. Radiother Oncol 

2008;89:135-40. 


5 Hall EJ, Wuu C-S. Radiation-induced second cancers: the impact of 

3D-CRT and IMRT. Int J Radiation Oncology Biol Phys 2003;56:83- 

8. 

6 Patel SR. Radiation-induced sarcoma. Curr Treat Options Oncol 

2000;1:258-61. 

7 Bisogno G, Sotti G, Nowicki1 Y, et al. Soft tissue sarcoma as a 

second malignant neoplasm in the pediatric age group. Cancer 

2004;100:1758-65. 

8 Banerjee SS, Agbamu DA, Eyden BP, et al. Clinicopathological characteristics 

of peripheral primitive neuroectodermal tumour of skin and 

subcutaneous tissue. Histopathology 1997;31:355-66. 

9 Bisceglia M, Fisher C, Suster S, et al. Tumoral, quasitumoral and 

pseudotumoral lesions of the superficial and somatic soft tissue: new 

entities and new variants of old entities recorded during the last 25 

years. Part VII: excerpta V. Pathologica 2005;97:92-114. 

epigenetic silencing of wnt-inhibitors: activation 

of a constitutive wnt signalling in oral cancer 

1)Bufo P. 2)Pannone G. 3)Santoro A. 4)Sanguedolce F. 5)Franco 

R. 6)Losito S. 7)Botti G. 8)Lo muzio L. 

1)Department of surgical sciences, section of anatom, Riuniti, Foggia, 

Italy 2)Department of surgical sciences, section of anatom, Riuniti, Foggia, 

Italy 3)Department of surgical sciences, section of anatom, Riuniti, 

Foggia, Italy 4)Department of surgical sciences, section of anatom, Riuniti, 

Foggia, Italy 5)Istituto nazionale per lo studio e la cura dei tum, Fondazione 

“G. Pascale”, Napoli, Italy 6)Istituto nazionale per lo studio e la 

cura dei tum, Fondazione “G. Pascale”, Napoli, Italy 7)Istituto nazionale 

per lo studio e la cura dei tum, Fondazione “G. Pascale”, Napoli, Italy 

8)Department of surgical sciences, Irccs crob - centro di riferimento oncologico 

di b, Rionero in vulture, Italy 

Background. Epigenetic DNA methylations plays an important 

role in oral carcinogenesis. The soluble frizzled receptor protein 

(SFRP) family together with WIF-1 and DKK-3 encodes antagonists 

of the WNT pathway. Silencing of these genes leads 

to constitutive WNT signalling. Because aberrant expression of 

ß-catenin might be associated with the epigenetic inactivation of 

WNT inhibitors, we analyzed, in a collection of primary OSCC 

with matched normal oral mucosa, the methylation status of a 

complete panel of genes, SFRP-1, SFRP-2, SFRP-4, SFRP-5, 

WIF-1, DKK-3, that are involved directly and indirectly in WNT 

pathway, in order to demonstrate WNT-pathway activation in 

the absence of ß-catenin and/or APC/Axin mutations during oral 

carcinogenesis. 

Methods. Methylation-specific PCR (MSP) was performed to 

study inactivation of SFRP-1, SFRP-2, SFRP-4, SFRP-5, WIF-1, 

DKK-3 genes in 37 cases of paraffin embedded oral cancer. 

Results. This study showed that the methylation is an important 

epigenetic alteration in oral cancer. In particular, SFRP-2, SFRP- 

4, SFRP-5, WIF-1, DKK-3 revealed methylation status of their 

promoter in OSCC, whereas SFRP-1 showed demethylation in 

cancer. Fisher’s exact test revealed statistically significant results 

(p < 0.05) for all genes. The Wald test confirmed the statistically 

significant association between SFRP2-4-5 gene methylation 

and OSCC (p < 0.05). SFRP-1 was also characterized by a different 

statistically significant epigenetic behaviour, because of it 

was demethylated in cancer (p < 0.05). Statistical regression test 

showed high levels of sensitivity, specificity and accuracy for 

SFRP genes, while WIF-1 and DKK-3 have reportedly high specificity, 

moderate accuracy but low sensitivity. This study suggests 

that a cause of catenin delocalization in oral cancer could be due 

to WNT pathway activation, by epigenetic alterations of SFRP, 

WIF-1 and DKK-3 genes.


epigenetic profile in endometrial carcinogenesis 

1)Bufo P. 2)Pannone G. 3)Santoro A. 4)Sanguedolce F. 5)Losito 

S. 6)Pasquali D. 7)Guida M. 

1)1. department of surgical sciences - section of an, Riuniti, Foggia, Italy 




5)2. istituto nazionale per lo studio e la cura dei, Fondazione “G. Pascale”, 

Napoli, Italy 6)3. department of clinical and experimental medicin, 

Sun, Napoli, Italy 7)4. department of gynaecology and obstetrics, University 

of naples “Federico II”, Napoli, Italy 

Background. Transcriptional silencing by CpG island hypermethylation 

plays a critical role in endometrial carcinogenesis. In 

a collection of benign, premalignant and malignant endometrial 

lesions, a methylation profile of a complete gene panel, such 

steroid receptors (ER_, PR), DNA mismatch repair (hMLH1), 

tumour-suppressor genes (CDKN2A/P16 and CDH1/E-CAD- 

HERIN) and WNT pathway inhibitors (SFRP1, SFRP2, SFRP4, 

SFRP5) was investigated in order to demonstrate their pathogenetic 

role in endometrial lesions. 

Methods. Methylation-specific PCR (MSP) was performed to assess 

gene inactivation. P53 and steroid receptors expression were 

evaluated by LSAB/HRP immunohistochemistry. 

Results. Our results indicate that gene hypermethylation may be 

an early event in endometrial endometrioid tumorigenesis. Particularly, 

ER_, PR, hMLH1, CDKN2A/P16, SFRP1, SFRP2 and 

SFRP5 revealed a promoter methylation status in endometrioid 

carcinoma, whereas SFRP4 showed demethylation in cancer. P53 

immunostaining showed weak-focal protein expression level both 

in hyperplasic lesions and in endometrioid cancer. Non endometrioid 

cancers showed very low levels of epigenetic methylations, 

but strong P53 protein positivity. Fisher exact test revealed a statistically 

significant association between hMLH1, CDKN2A/P16 

and SFRP1 genes methylation and endometrioid carcinomas and 

between hMLH1 gene methylation and peritumoral endometrium 

(p < 0.05). Our data confirm that the methylation profile of the 

peritumoral endometrium is different from the altered molecular 

background of benign endometrial polyps and hyperplasias. 

Therefore, our findings suggest that the methylation of hMLH1, 

CDKN2A/P16 and SFRP1 may clearly distinguish between benign 

and malignant lesions. Finally, this study assessed that the 

employment of an epigenetic fingerprint may improve the current 

diagnostic tools for a better clinical management of endometrial 

lesions. 

Vulvar angiomyofibroblastoma: a clinicopathological 

study of nine cases, including the lipomatous 

variant 

1)G. Vecchio, 1)R. Caltabiano, 1)A. Gurrera, 2)D. Kacerovska, 

3)M. Bisceglia, 2)M. Michal, 1)G. Magro 


Azienda Ospedaliero-Universitaria Policlinico Vittorio Emanuele, 

Catania, Italia; 2)Sikl’s Department of Pathology, Medical Faculty Hospital, 

Pilsen, Czech Republic 3)Dipartimento di Anatomia Patologica, 

IRCCS Casa Sollievo della Sofferenza, San Giovanni Rotondo, Italia 

Background. Angiomyofibrobastoma (AMF) is an uncommon, 

benign stromal tumour usually arising in the vulva. Other sites, 

including vagina, urethra, perineum, fallopian tube, inguino-scrotal 

and para-rectal region in male, may be involved. Clinically, 

most of the tumours present as a slowly-growing painless mass, 

often misdiagnosed as a Bartholin’s gland cyst, hydrocele, or aggressive 

angiomyxoma. Clinical behaviour is benign with a low 

tendency for local recurrence. 

Methods. The clinicopathological features of 9 cases of AMFs of 

the vulva are presented with emphasis on unusual morphological 

features. 

267 

Results. The lesions usually presented as painless masses located 

in the superficial vulvar region of women ranging in age from 46 

to 60 years. They were well circumscribed and ranged in size from 

2 to 3.5 cm in greatest diameter. Histologically, they were composed 

predominantly of medium-sized spindle to epithelioid cells 

variably arranged in cords or nests, and embedded in a fibrous 

to only focally myxoid stroma. In most cases neoplastic cells 

exhibited a perivascular arrangement around small to mediumsized 

hyalinized blood vessels. Mitotic activity ranged from 0 to 

2 mitoses per 50 HPF. Atypical mitoses, nuclear atypia and necrosis 

were not observed. Interestingly 3 cases, labelled “AMFs, 

lipomatous variant”, contained an abundant intratumoral fatty 

component, ranging from 20% to 70% of the entire tumour. In 

one case, adipocytes focally exhibited a lipoblast-like appearance. 

Additional unusual findings were the presence of neoplastic cells 

with vescicular nuclei and CD68+ giant multinucleated osteoclast-like 

cells. Immunohistochemically the cells were positive to 

vimentin, and variably to α-smooth muscle actin, desmin, CD34, 

and estrogen/progesterone receptors. No local recurrence was observed 

after a follow-up period ranging from 3 to 20 years. 

Myofibroblastoma of the lower female genital 

tract: expanding the morphologic spectrum, 

including the mammary-type variant 

1)P. Amico, 1)R. Caltabiano, 2)D. Kazakov, 2)D. Kacerovskà, 

2)M. Michal, 1)G. Magro 


Azienda Ospedaliero-Universitaria-Policlinico Vittorio Emanuele, 

Catania, Italia; 2)Sikl’s Department of Pathology, Charles University Medical 

Faculty Hospital, Pilsen, Czech Republic 

Background. Over the last decade, a benign myofibroblastic 

tumour with distinctive clinicopathological features has emerged 

from the category of the stromal tumours of the lower female 

genital tract, including aggressive angiomyxoma, angiomyofibroblastoma 

and cellular angiofibroma. The terms “superficial 

cervicovaginal myofibroblastoma (MFB)” or “MFB of the lower 

female genital tract” have been used interchangeably for this entity 

which characteristically arises from the sub-epithelial stroma 

of the vagina, and less frequently, of the vulva or cervix. 

Materials. We herein report the clinicopathological features of 

10 cases of MFB of the lower female genital tract to expand the 

morphologic spectrum. 

Results. Tumours clinically presented as polypoid or nodular 

masses of variable size (1-3 cm) located in vagina (7 cases) and 

vulva (3 cases). Age at diagnosis ranged from 19 to 69 years (). 

Histologically, all tumours were well circumscribed and unencapsulated, 

with the typical localization in the sub-epithelial connective 

tissue. Unlike previously reported, a band of native connective 

tissue, separating tumours from the overlying squamous 

epithelium, was missing in 5 cases, with tumour cells extending 

up to the epithelium. Neoplastic cells, from stellate to ovoid to 

spindle in shape, were embedded in a finely fibrous to focally 

myxoid stroma. Five tumours, being predominantly composed of 

spindle-shaped cells arranged in short fascicles with intervening 

thick collagen bands, were closely reminiscent of mammary MFB. 

Mitoses were rare. Only focally mild nuclear pleomorphism was 

seen. Interestingly, 6 cases showed hyalinized thick-walled blood 

vessels. Immunohistochemically, tumours were variably positive 

for desmin, α-smooth muscle actin, CD34, CD10, ER and PR. 

The present study first identifies the mammary-type variant of 

MFB of the lower female genital tract. Based on morphological 

and immunohistochemical findings, we postulate that MFB of 

the breast and MFB of the lower female genital tract arise from 

a common precursor stem cell which typically resides in the hormonally 

active stroma of women.

268 

The mosaic pattern of INI1/SMArCB1 protein 

expression is a reliable marker of sporadic 

schwannomatosis: an immunohistochemical study 

in a series of 10 cases 

1)A. Torrisi, 2)R. Caltabiano, 3)M. Ruggieri, 4)P. Nozza, 5)A. 

Ortensi, 5)V. D’Orazi, 2)S. Lanzafame, 2)G. Magro 

1)Dipartimento G.F. Ingrassia, Registro Tumori Integrato-Messina-Catania-Siracusa, 

Catania, Italia; 2)Dipartimento G.F. Ingrassia, Anatomia 

Patologica, Università di Catania, Azienda Ospedaliero-Universitaria 

Policlinico-Vittorio Emanuele, Catania, Italia; 3)Istituto Scienze Neurologiche 

CNR, Azienda Ospedaliero-Universitaria Policlinico-Vittorio Emanuele, 

Catania, Italia; 4)Anatomia Patologica, Ospedale Gaslini,Genova, 

Italia; 5)Microchirurgia e Chirurgia della Mano, Università La Sapienza, 

Fabia Mater, Roma, Italia 

Background. Schwannomatosis is characterized by the development 

of multiple spinal, peripheral, and cranial nerve schwannomas 

in the absence of diagnostic bilateral vestibular schwannomas 

of NF2. The majority of cases of schwannomatosis are 

sporadic, but familial cases do exist with an autosomal dominant 

pattern of inheritance. The INI1/SMARCB1 is a tumour suppressor 

gene that maps to chromosome band 22q11.2. It affects 

the expression of genes that regulate cell cycle, growth, and differentiation. 

It is also involved in the development of malignant 

rhabdoid tumours. 

Methods. The immunohistochemical expression of INI1/ 

SMARCB1 was assessed in a series of 10 cases of sporadic 

schwannomatosis and compared with the immunohistochemical 

profile of 5 cases of solitary sporadic schwannomas. 

Results. As expected, all sporadic schwannomas showed diffuse 

nuclear positivity ranging from 97% to 100% of neoplastic cells. 

On the contrary schwannomas from sporadic schwannomatosis 

showed a mosaic pattern, namely alternating positive and negative 

nuclei, consistent with the loss of INI1/SMARCB1 expression 

in a subset of tumour cells, ranging from 10% to 70% in the 

different cases. The little data available in the literature showed 

that only 55% of schwannomas from sporadic schwannomatosis 

exhibit the mosaic pattern of INI1/SMARCB1 expression, as 

commonly observed in familial schwannomatosis. 

We first report that 100% of schwannomas from sporadic 

schwannomatosis have a mosaic pattern of INI1/SMARCB1 

expression. Accordingly, apart from familial schwannomatosis, 

loss of immunohistochemical INI1/SMARCB1 expression, albeit 

with an interlesional variability, is a reliable marker of sporadic 

schwannomatosis. 

Analysis protocol of the retroareolar margin in the 

nipple sparing mastectomy: our experience 

1)Costarelli L. 1)Campagna D. 2)Amini M. 3)Fortunato L. 4)Poccia 

I. 

1)Anatomia ed istologia patologica, Az. osp S. Giovanni-Addolorata, 

Roma, Italia 2)Anatomia ed istologia patologica, Az. osp S. Giovanni- 

Addolorata, Roma, Italia 3)I chirurgia, Az. osp S. Giovanni-Addolorata, 

Roma, Italia 4)I chirurgia, Az. osp S. Giovanni-Addolorata, Roma, Italia 

Introduction. The nipple-sparing mastectomy (NSM) has become 

an accepted treatment for appropriately selected breast 

cancers to improve the aesthetic results and the patient’s satisfaction. 

The cosmetic results of the mastectomy followed by immediate 

reconstruction or by prosthetic implant have been judged to be 

good to excellent in 82% of the cases. 

The principal postoperative complication requiring a second surgical 

treatment is the necrosis of the nipple and the retro-areolar 

margin positive. To avoid doing a second surgery may be performed 

the intraoperative frozen sections and HE histopathologic 

examination of the retro-areolar tissue. 


Methods. The authors performed 43 nipple-sparing mastectomies 

on 37 patients (6 bilateral) during 2009/2010. The retro-areolar 

tissue obtained was serially sectioned throw 5-6 parallel slices at 

the time of the intraoperative frozen section, after painting surgical 

margin with India ink. 

The ablation of nipple-areola complex (NAC) was performed in 

the same time of the subcutaneous mastectomy if the neoplasia 

was close to margin (< mm 1). 

Results. The average age of the patients was 46 years (46 ± 9; 

range 32-67). The indications into account to decide to performed 

the nipple-sparing mastectomy were multifocality in 

36 cases (6 bilateral), locally advanced stage in 5 cases and 

familiarity in 2 cases. The rate of positive retroareolar margin 

was 11.6 percent (5 cases) at the frozen sections following the 

excision of the NAC in the same time. The rate of ablation of 

NAC in a second time was 11.6 percent (5 cases) for post-operative 

necrosis and 2.4 percent (1 case) for false negative at 

the intraoperative examination. Finally, the intraoperative protocol 

to examinate the retroareolar tissue reduce the percentage 

of reintervention improving cosmetic results with a high level 

of surgeons’ and patients’ satisfaction. 

Nut midline carcinoma: report of a case with 

unusual immunoprofile 

1)A. Canesso, 2)R. Alaggio, 3)E.G.S. D’Amore, 4)E. Gaio, 4)R. 

Artico, 1)S. Agabiti, 1)F. Sonego, 1)M. Guido 

1)Anatomia Patologica, A.o.ulss15 Alta Padovana, Cittadella, Italia, 

2)Anatomia Patologica, Università degli Studi di Padova, Padova, Italia; 

3)Anatomia Patologica, Ospedale San Bortolo, Vicenza, Italia; 4)ORL, 

A.o.ulss15 Alta Padovana, Cittadella, Italia 

Background. NUT midline carcinoma is a rare, highly aggressive 

neoplasia associated with rearrangement of the NUT gene on 

chromosome 15q14 most commonly in a balanced translocation 

with the BRD4 gene on chromosome 19p13, originally reported 

in head and neck and mediastinum in young females. Subsequently 

NUT-carcinoma has been identified in all age groups 

(from 3 to 78 years). We report a case of NUT midline carcinoma 

arising in a 52 year-old woman presenting with a left neck mass 

and displaying a challenging immunophenotype. 

Methods. A 52 year-old woman with no remarkable medical 

history and completely asymptomatic presented with a left neck 

mass of three months duration. A TC scan showed enlarged, 

centrally necrotic lymph nodes that were surgically removed and 

submitted for pathology evaluation. The specimen was formalinfixed, 

paraffin embedded and routinely stained (e.e), then a wide 

panel of immunostains was performed. 

Results. The neoplasia was composed of undifferentiated cells of 

medium size with scant eosinophilic cytoplasm, irregular nuclei 

and prominent nucleoli. Mitoses and areas of coagulative necrosis 

were common. Immunohistochemistry showed reactivity for 

CK7, MNF116, 34βE12, CD34 and p63. An unusual dot-like 

reactivity for WT1 and Vimentin was noted, as well as cytoplasmic 

positivity for κ and γ light chain. All the other immunostains 

were negative, thus excluding lymphoma, sarcomas, melanoma 

or metastatic carcinoma. NUT immunostaining showed a strong 

and diffuse nuclear staining. FISH analysis was positive for NUT 

rearrangement, but not for BRD4 rearrangement, consistent with 

a NUT-variant carcinoma. The patient died two months later for 

disseminated disease. NUT-variant carcinoma is an under-recognized 

entity and should be considered in the spectrum of differential 

diagnoses in metastatic carcinomas with unknown primary 

tumor. Moreover aberrant positive immunostains, like κ and γ in 

the present case may represent a potential diagnostic pitfall.


Diagnostic value of automated Her2 evaluation in 

breast cancer. A study on 272 equivocal (score 2+) 

Her2 immunoreactive cases using an fda approved 

system 

1)Cantaloni C. 2)Eccher C. 3)Morelli L. 4)Leonardi E. 5)Bragantini 

E. 6)Aldovini A. 7)Fasanella S. 8)Ferro A. 9)Dalla palma P. 

10)Barbareschi M. 

1)Anatomia patologica, S Chiara, Trento, Italia 2)Statistica, Fondazione 

Bruno Kessler, Trento, Italia 3)Anatomia Patologica, S Chiara, Trento, 

Italia 4)Anatomia Patologica, S Chiara, Trento, Italia 5)Anatomia Patologica, 

S Chiara, Trento, Italia 6)Anatomia Patologica, S Chiara, Trento, 

Italia 7)Anatomia Patologica, S Chiara, Trento, Italia 8)Oncologia, 

S Chiara, Trento, Italia 9)Anatomia Patologica, S Chiara, Trento, Italia 

10)Anatomia Patologica, S Chiara, Trento, Italia 

Backgroung. Accurate immunohistochemical Her2 evaluation is 

fundamental for treatment of breast cancer (BC). The U.S. Food 

and Drug Administration (FDA) approved the Aperio IHC Her2 

Breast Tissue Image Analysis application for the detection and 

semi-quantitative measurement of Her2. 

Methods. To validate computer assisted analysis (CAA) in 

clinical practice we analyzed 292 equivocally (score2+) Her2 immunoreactive 

BC; all cases were stained with Dako Herceptest, 

evaluated by an experienced pathologist and analyzed with FISH. 

The automatic Aperio categorization and the percentage of immunoreactive 

cells as evaluated by the computer (CPV) and by the 

pathologist (PPV) were recorded. CAA classified 7 (2.4%) cases 

as negative (0), 136 (46.6%) as score 1+, 134 (40.5%) as score 

2+ and 15 (5.1%) as score 3+. CCA classification is associated 

with Her2 amplification (p < 0.0001). The mean CPV is 18.44% 

sd ± 19.00 (range 0.01-76.10). 

Results. CPV and PPV are significantly associated and correlated 

(p < 0.001), have similar sensitivity and specificity in 

identifying Her2 FISH amplified cases. The difference in CPV in 

amplified and non amplified subgroups is statistically significant 

(p < 0.001). ROC analysis indicates that CPV is good at separating 

FISH not-amplified from amplified cases (p < 0.001). The 

optimal cut-off value maximizing both sensitivity and specificity 

is 17.6% (sensitivity = 73.3%, specificity = 71.6%). Reducing 

the cut-off value to 0.67% it is possible to reach the sensitivity 

of 100% with 16.2% specificity. CCA Her2 IHC evaluation is 

feasible and reliable: automated classification is not satisfactory 

as some amplified cases might be erroneously clustered as score 

1+. Lower CPV cut-off values should be used. CAA can reduce 

the number of cases unnecessarily submitted to FISH. 

The role of geminin, a DNA replication factor, 

in oral squamous cell carcinoma. Preliminary 

report of a tissue micro array based 

immunohistochemical study 

1)Cantile M. 2)Franco R. 3)Aquino G. 4)Losito S. 5)Botti G. 

6)Santoro A. 7)Mattoni M. 8)Bufo P. 9)Pannone G. 

1)Istituto Nazionale Per Lo Studio E La Cura Dei Tum, Fondazione ‘G. 

Pascale’, Napoli, Italy 2)Istituto Nazionale Per Lo Studio E La Cura Dei 

Tum, Fondazione ‘G. Pascale’, Napoli, Italy 3)Istituto Nazionale Per Lo 

Studio E La Cura Dei Tum, Fondazione ‘G. Pascale’, Napoli, Italy 4)Istituto 

Nazionale Per Lo Studio E La Cura Dei Tum, Fondazione ‘G. Pascale’, 

Napoli, Italy 5)Istituto Nazionale Per Lo Studio E La Cura Dei 

Tum, Fondazione ‘G. Pascale’, Napoli, Italy 6)Department Of Surgical 

Sciences, Institute Of Path, Riuniti, Foggia, Italy 7)Department Of Surgical 

Sciences, Institute Of Path, Riuniti, Foggia, Italy 8)Department Of 

Surgical Sciences, Institute Of Path, Riuniti, Foggia, Italy 9)Department 

Of Surgical Sciences, Institute Of Path, Riuniti, Foggia, Italy 

Background. The DNA replication licensing machinery is integral 

to the control of proliferation differentiation, and maintenance of 

genomic stability in human cells. Geminin is a licensing repressor 

and prevents re-initiation of cell replication by blocking re-loading 

of MCM proteins at replication origins. The recent literature has 

269 

proposed that Geminin could be used as sensitive proliferative and 

prognostic marker. The aim of this study is the evaluation of Geminin 

expression in oral squamous cell carcinomas (OSCCs) by Tissue 

Micro Array based immunohistochemistry (TMA based IHC). 

Methods. We performed TMA based IHC on 10 specimens of 

normal oral squamous epithelia and 150 OSCCs. IHC was performed 

by standard streptavidinin-biotin immunoperoxidase method 

(LSAB-HRP) using specific monoclonal Ab against Geminin. 

Results. Geminin is a 25kDa nuclear protein involved in regulation 

of the initiation of DNA replication. DNA replication requires the 

association of Cdc-6 and minicromosome maintenance (MCM) 

protein with chromatin. Geminin blocks this assembly of the 

MCM into the pre-replication complex. Expression of Geminin is 

regulated throughout the cell cycle with Geminin levels lowest at 

G1. Throughout S, G2 and M phases, Geminin levels are elevated 

followed by a decrease during mitosis as the protein is targeted for 

degradation by the anaphase-promoting complex (APC). Our study 

showed Geminin over-expression in the most of OSCCs as compared 

to normal oral epithelia. In details, this proteins was strongly 

up-regulated in OSCCs characterized by high mitotic index. Our 

preliminary results indicate that assessment of Geminin may be 

useful as prognostic factor in patients with OSCCs. 

Adenoid-cystic carcinoma of the breast with 

sebaceous and adenosquamous differentiation. 

A clinicopathologic study of an aggressive case 

1)M. Carlucci, 1)M. Iacobellis, 1)F. Colonna, 2)M. Marseglia, 

3)M. Gambarotti, 4)M. Bisceglia, 5)C. Giardina 

1)Anatomia Patologica, Ospedale Umberto I, Altamura, Italia; 2)Chirurgia, 

Ospedale Umberto I, Altamura, Italia; 3)Anatomia Patologica, Istituto 

Ortopedico Rizzoli, Bologna, Italia; 4)Anatomia Patologica, IRCCS 

Ospedale Casa Sollievo della Sofferenza, San Giovanni Rotondo, Italia; 

5) Anatomia Patologica, Università degli Studi di Bari, Bari, Italia 

Background. Adenoid cystic carcinoma (ACC) of the breast represents 

about 0.1% of breast carcinomas and, in contrast to the aggressive 

nature of the homonymous tumor arising in the head and 

neck region, usually has a favorable prognosis. This tumour has 

well-demarcated margins, and can be over 10 cm diameter size and 

multifocal. ACC is “a morphologically heterogeneous neoplasm” 

with trabecular-tubular, cribriform and solid pattern, occasionally 

associated with sebaceous and adenosquamous differentiaton. 

Case report. An 84-year old woman was admitted with an ulcerated 

12 cm tumor mass in her left breast, without palpable axillary 

lymph nodes. A simple mastectomy was performed. On histological 

examination a mixed type of adenoid-cystic carcinoma 

of the breast with sebaceous and adenosquamous differentiation 

was apparent, with trabecular, cribriform and solid patterns, and 

in places sharing features of ordinary invasive ductal carcinoma. 

Immunohistochemically the cribriform areas were focally positive 

for actin and for CK34beta12. The pseudocystic spaces were 

partly positive for type IV collagen. EMA strongly decorated 

areas of sebaceous differentiation and c-kit immunoreactivity was 

also focally documented. Estrogens and progesterone receptors 

were totally negative. 

After mastectomy a total body CT scan showed pulmonary and 

osseous metastases that partially responded to chemotherapy. 

About 7 months later an intramuscular mass rapidly growing up 

to > 10 cm was also noticed in her right thigh. Following a needle 

biopsy-based diagnosis of malignancy, the tumor mass was excised, 

and the histological diagnosis established was “metastasis 

of ductal carcinoma G3 with sebaceous differentiation”. 

Conclusion. Adenoid-cystic carcinoma has the potential to 

differentiate toward skin adnexal structures giving rise to both 

sebaceous and adenosquamous cells. However similar tumors 

may also be interpreted as “mixed invasive carcinoma” with both 

usual features and features commonly seen in salivary gland type 

and adnexal skin type carcinomas.

270 

Incidence of O6-methylguanine DNA 

methyltransferase expression in pituitary 

adenomas: our experience at the “regina elena” 

National Cancer Institute 

Carosi M., 1Baldelli R., Panichi D., 1Barnabei A., 2Telera S., 1Ap petecchia M., 2Pompili A., Pescarmona E. 

Pathology, 1Endocrinology and 2 Neurosurgery, “Regina Elena” National 

Cancer Institute 

Clinically significant pituitary tumours occur in approximately 

in every 1000 individuals. The majority of pituitary tumours are 

benign adenomas; however, between 35% and 55% of adenomas 

demonstrate invasion into bone, dura or adjacent structures such 

as the cavernous or sphenoid sinuses or brain. Although it is a 

rare phenomenon, a subset of invasive adenomas display aggressive 

behaviour and become resistant to medical therapy, causing 

substantial morbidity; these tumours require multiple operations 

and radiotherapy in an attempt to control tumour growth. Various 

chemotherapeutic regimes have been tried in the management of 

pituitary carcinoma. Although occasional temporary responses 

are reported, the results are usually disappointing. Recent case 

studies have successfully used temozolomide, an alkylating chemotherapeutic 

drug, in the management of pituitary carcinoma 

and aggressive pituitary tumours. Temozolomide is widely used 

in the management of glioblastoma multiforme and is effective in 

other neuro-oncological tumours as well as other neuroendocrine 

tumours. Temozolomide is administered orally, readily crosses 

the blood–brain barrier and is not cell-cycle specific, advantageous 

when treating relatively slow-growing pituitary tumours. 

O-methylguanine-DNA methyltransferase (MGMT) is a DNA repair 

protein that reverses alkylation at the O position of guanine. 

As such, MGMT counteracts the effect of temozolomide, which 

alkylates DNA at this position. Low tumour MGMT expression 

has been shown in some studies to correlate with temozolomide 

response and increased survival in patients with brain tumours. A 

commonly proposed mechanism of reduced MGMT expression is 

methylation of its promoter, although different tumour types vary 

widely in the frequency of methylation. 

The aim of this study was to evaluate the possible relationship 

between MGMT hypermethylation and clinical response to chemotherapy 

in pituitary adenomas; the method to determine the 

hypermethylation status of MGMT, namely methylation-specific 

PCR, allowing the selection of patients most likely to benefit 

from temozolomide treatment 

fine needle aspiration cytology of thyroid lesions: 

cytohistologic correlation and accuracy. Overwiew 

of 15 years experience 

1)Casadei G.P. 2)Crucitti P. 3)Lega S. 4)Bondi A. 

Anatomia Patologica, Dipartimento di Oncologia, Ospedale Maggiore, 

Bologna 

Objective: The aim of this study was to evaluate the accuracy of 

the fine needle aspiration cytology (FNAC) and its contribution 

to tumor diagnosis. 

Methods. In the period from 1995 to 2009, a total of 12.989 

thyroid FNAC were performed at Maggiore hospital and in 

two referring smaller hospitals, and examined in a pathology 

laboratory at one site. Cytologic diagnoses were re-classified 

according to the Italian Society of Pathology (SIAPEC) 5-tiered 

category system, as THY 1 unsatisfactory, THY 2 benign, THY 3 

indeterminate, THY 4 suspicious, THY 5 malignant. Patients who 

underwent surgical treatment were 3.944 (30%). Sample with histologic 

discrepancy were rewieved, and clinically re-evaluated. 

Results. The distribution of cytologic samples by the 5 diagnostic 

categories was 3.088 (24%) inadequate (THY 1), 68% THY 2, 5% 

THY 3, 1,2% THY 4, and 1,4% THY 5. 


Histological examination was performed in 2% of inadequate 

samples, 4,7% of benign lesions, 22% in THY 3, and in 33% and 

42% of THY 4 and THY 5 category respectively. 

Malignancy was histologically observed in 22% of inadequate 

FNAC, 16% of benign lesions, 38% of undeterminate lesions, 

82% of suspicious lesions and 93% of malignant ones. 

Diagnostic discrepancy rate between cytologic and histologic 

diagnosis was about 15%. The overall sensitivity and specificity 

of thyroid FNAC was 77% and 95% respectively. Wrong 

diagnostic results arise from errors of sampling, as for smaller 

than 1 cm nodules, inadequate smearing, and cytological 

equivocal features. 

Conclusions. Although FNAC of thyroid nodules can be performed 

with high sensitivity and specificity, it needs of application 

of firm rules and guidelines in performing the procedure in 

such a way to reduce the rate of false-negative and false-positive 

diagnoses. The study of clinical correlation in single case and the 

direct involvement of the pathologist with the clinicians on taking 

the sample is advisable. 

The human claustrum: a microanatomical and 

immunohistochemical study 

1)Castagna M. 2)Fattori S. 3)Castelluccio E. 4)Quilici F. 5)Perrini 

P. 6)Pirone A. 

1)Dipartimento di Chirurgia, Ospedale S. Chiara, Pisa, Italia 2)Dipartimento 

di Chirurgia, Ospedale S. Chiara, Pisa, Italia 3)Dipartimento di 

Chirurgia, Ospedale S. Chiara, Pisa, Italia 4)Dipartimento di Chirurgia, 

Ospedale S. Chiara, Pisa, Italia 5)Neurochirurgia, Ospedale S. Chiara, 

Pisa, Italia 6) Dipartimento di Produzione animale, Università di Pisa, 

Pisa, Italia 

Background. The claustrum is a thin collection of gray matter located 

deep with respect to the insula. While numerous investigations 

focused on the chemo- and cytoarchitecture of the claustrum 

specific to the localization of calcium-binding proteins (CBPs) 

and neuropeptides (Nps) in a variety of mammalian species, few 

studies examined the microanatomy and the immunohistochemistry 

of human claustrum. 

Methods. Two normal human cerebral hemispheres were fixed 

in a 10% formalin solution for two months and then frozen at -10 

to -15C for two to four weeks. The lateral surface of the brain 

was dissected by applying Klinger’s fiber dissection technique 

under microspcope. One additional brain of a 65-year old male 

who died of a myocardial infarction provided the claustrum for 

immunohistochemical characterization. Individual sections were 

processed for Nissl, parvalbumin (PV) or neuropeptide-Y (NPY) 

staining. Tissue was processed with either monoclonal anti-PV 

mouse ascites fluid clone PA-235 (P-317; 1:1000: Sigma) or 

rabbit anti-NPY porcine serum (IHC 7172, 1:800, Peninsula). 

PV-positive neurons were viewed with a confocal microscope using 

indirect immunofluorescence (Leica TCS-NT, krypton-argon 

laser), and NPY-positive neurons were viewed using standard 

light microscopy (Leitz Diaplan). 

Results. The claustrum presents a ventral (fragmented) and a 

dorsal (compact) part which are, respectively, anteroinferior and 

posterosuperior. The ventral part of the external capsule forms 

the uncinate and occipito-frontal fascicles. The dorsal part of the 

external capsule forms the claustrocortical fibers. 

The immunoistochemical investigation disclosed evidence of PV- 

and PNY-immunoreactivity in the dorsal claustrum. PV-positive 

neurons were generally round, fusiform or pyramidal in shape, often 

multipolar, with well-filled axonal arborizations. They ranged 

in diameter from 10 to 20 µm. NPY-positive neurons were generally 

round or fusiform in shape, ranging in diameter from 15 to 

30 µm. Like previous studies in other mammals, we characterized 

claustral PV- and NPY-positive neurons at the light-microscopic 

level. In addition, we provided the anatomical bases for a topographical 

organization of the human claustrum. Further studies


are necessary to investigate the immunospecific and topographic 

subdivision within the claustrum, as well as colocalization and 

coexpression patterns with various other CBPs and NPs. 

Multifaceted Her-2 and topoisomerase-IIa status in 

gastric carcinoma with potential clinical impact 

1) Cataldo I. 2) Brunelli M. 3) Barbi S. 4) Pecori S. 5) Beghelli 

S. 6) Tomezzoli A. 7) Bersani S. 8) Brunello E. 9) Martignoni G. 

10) Scarpa A. 

1) Department of pathology and diagnostic, Policlinico G.B. Rossi, Verona, 

Italy 2) Department of pathology and diagnostic, Policlinico G.B. Rossi, 

Verona, Italy 3) Department of pathology and diagnostic, Policlinico G.B. 

Rossi, Verona, Italy 4) Department of pathology and diagnostic, Policlinico 

G.B. Rossi, Verona, Italy 5) Department of pathology and diagnostic, 

Policlinico G.B. Rossi, Verona, Italy 6) Anatomic pathology, Ospedale civile 

maggiore, Verona, Italy 7) Department of pathology and diagnostic, 

Policlinico G.B. Rossi, Verona, Italy 8) Department of pathology and diagnostic, 

Policlinico G.B. Rossi, Verona, Italy 9) Department of pathology 

and diagnostic, Policlinico G.B. Rossi, Verona, Italy 10) Department of 

pathology and diagnostic, Policlinico G.B. Rossi, Verona, Italy 

Background. In gastric carcinoma, Her-2/neu gene amplification 

is predictive of responsiveness to Trastuzumab whereas Topoisomerase-IIa 

(Topo-IIa) to anthracycline. A subset of patients 

does not respond to these drugs. The heterogeneity may in part 

justify the lack of clinical efficacy. 

Methods. 172 gastric carcinomas (60 diffuse-type, 98 intestinaltype 

and 14 mixed) were recruited and 7 tissue micro-array were 

built by punching three neoplastic cores per case. Hercept Test 

(HER-2) was performed and cases scored by using the TOGA 

system (3+, 2+, 1+, 0). Her-2/neu and Topo-IIa gene amplification 

was assessed by FISH analysis. In all cases, HER-2 heterogeneity 

was evaluated among each cores per single case; a cases 

was registered as heterogeneous when at least one core did differ 

from the others. We also evaluated the discrepancy between the 

overall score (summing up the values from the three cores) vs the 

single observed on whole tissue sections. 

Results. Amplification of Her-2/neu and Topo-IIa was respectively 

observed in 10% and 21% of the cases. Strong 3+ Her-2 

immunoexpression was found in 8% of cases and both Her-2/neu 

and Topo-IIa resulted amplified in 43% into this group. Among 

the 2+, 1+ and 0 groups, Her-2/neu was amplified in respectively 

12%, 14% and 1% and Topo-IIa in 25%, 30% and 17%. Polysomy 

of chromosome 17 (with no amplification) was observed in 

7% (Her-2) and 11% (Topo-IIa) of cases. 

We found a prevalence of HER-2 immunoexpression and Topo- 

IIa amplification among the intestinal subtype. Heterogeneity 

among cores was found in 40% of cases for Her-2; in 46% of this 

subset we observed discrepancy among the values in between 

sum of the cores vs whole sections. 

In conclusion, heterogeneity of HER-2 gene exists in a subset of 

gastric carcinomas with potential clinical relevance; the status 

of Topo-IIa does not strictly match with that of Her-2/neu. The 

impact of these patterns on treatment outcome in gastric cancer 

need further investigation. 

Cutaneous polypoid atypical leiomyoma of the 

scrotum: a case report and a review of literature 

1) Cataldo I. 2) Brunelli M. 3) Grosso G. 4) Pedica F. 5) Magro 

G. 6) Menestrina F. 7) Martignoni G. 

1) Department of Pathology and Diagnostic, Policlinico G.B. Rossi, Verona, 

Italia 2) Department of Pathology and Diagnostic, Policlinico G.B. 

Rossi, Verona, Italia 3) Urologia, clinica pederzoli, peschiera del garda, 

italia 4) Department of Pathology and Diagnostic, Policlinico G.B. Rossi, 

Verona, Italia 5) Anatomic pathology, G.F. Ingrassia, Policlinico-Vittorio 

Emanuele, Catania, Italia 6) Department of Pathology and Diagnostic, 

Policlinico G.B. Rossi, Verona, Italia 7)Department of Pathology and Diagnostic, 

Policlinico G.B. Rossi, Verona, Italia 

271 

Atypical leiomyoma of the scrotum is a rare benign entity arising 

from the muscular dartos tunica. To date fourteen cases of 

atypical leiomyoma of the scrotum have been reported in literature. 

They have been named as atypical, bizarre or symplastic 

leiomyoma alternatively, remarking the atypical leiomyomatous 

characteristic of the lesion with scanty or no mitosis nor necrosis. 

We describe a new case of atypical leiomyoma of the scrotum 

with polypoid appearance in a 52 years old man. At microscopic 

examination the lesion was constituted by fascicles of leiomyomatous 

spindle cells, with cellular atypia, without mitosis nor 

necrosis. Immunohistochemically, the spindle cells were positive 

for desmin and α-smooth muscle actin and negative for CK8-18, 

CD34, S100, androgen, progesteron and estrogens receptors. The 

immunohistochemical assays confirmed the leiomuscular differentiation 

of the lesion and rule out any suspicion of malignancy. 

Clear cell adenocarcinoma of the colon: 

report of a case with 2 years follow-up 

1)Cusatelli P. 2)Fiscon V. 3)Pizzi S. 4)Becherini F. 5)Canova E. 

1)Anatomia Patologica, ULSS 15 Alta Padovana, Camposampiero (PD), 

Italia 2)Chirurgia, ULSS 15 Alta Padovana, Cittadella (PD), Italia 3)Anatomia 

Patologica, ULSS 15 Alta Padovana, Camposampiero (PD), Italia 

4)Anatomia Patologica, ULSS 15 Alta Padovana, Campoasampiero (PD), 

Italia 5)Anatomia Patologica, ULSS 15 Alta Padovana, Camposampiero 

(PD), Italia 

Bakground. Clear cell adenocarcinoma is a very rare entity in 

the colon and its prognosis is not clear, since follow-up data are 

not available. Clear cell adenocarcinoma usually occurs in the 

left colon as a part of a large conventional adenoma. So far, only 

one case occurring in the right colon and not associated with 

adenoma has been reported. We describe a case of pure clear cell 

adenocarcinoma occurring in the left colon without any evidence 

of associated adenoma and followed-up for more than 2 years. 

Methods. A 63 years old man presented in September 2007 with 

melena. Endoscopy showed 3 polyps and an ulcerated, 3 cm diameter, 

lesion. Polyps were removed endoscopically and all were 

conventional adenomas. A biopsy obtained from the ulcerated 

lesion showed a small fragment of adenocarcinoma. Radiological 

examination did not show evidence of tumour elsewhere in the 

body and a left colon resection was performed. 

Results. At histology, the entire lesion was composed of clear 

cell adenocarcinoma infiltrating the muscularis propria (pT2). 

Lymph nodes were not metastatic (15 lymph nodes assessed). 

Immunohistochemistry showed the following profile: CK 7-, 

CK20+, CDX2+, CD10-, p53+ (strong and diffuse positivity), 

hMLH1+/hMSH2+ (consistent with microsatellite stability). 

Ki67 was positive in nearly all neoplastic cells. 

Despite the high proliferative activity of the tumour, the patient 

did not show recurrence or distant metastasis at the last control 

in March 2010. 

Conclusion. This case confirm that clear cell is a variant of colic 

adenocarcinoma and does not necessarily occur in association 

with conventional adenoma. Its course does not seem particularly 

aggressive. 

Intraparenchymal leiomyoma of the breast: 

report of a case with emphasis on needle core 

biopsy-based diagnosis 

G.M. Vecchio, 1)A. Cavaliere, 1)F. Cartaginese, 1)A. Lucaccioni, 

2)T. Lombardi, 3)A. Bosco, 3)A. Sabino, 3)G. Magro 



Catania, Italia; 2)Istituto di Anatomia Patologica, Azienda Ospedaliera di 

Perugia, Perugia, Italia; 3)U.O. di Senologia, Ospedale Città di Castello, 

Città di Castello, Italia 

Background. Leiomyomas are benign smooth muscle tumours 

that can potentially occur anywhere, including breast. In this site

272 

leiomyomas are usually found both in the skin and periareolar 

region, whereas only rarely they involve breast parenchyma. Only 

23 cases of intraparenchymal leiomyomas have been reported, 

exclusively in women, to date. Histogenesis of these tumours 

is still controversial and an origin from vascular smooth muscle 

cells or stromal stem cells, or from embryologically displaced 

smooth muscle cells, has been postulated. 

Materials. We herein report the first case of an intraparenchymal 

leiomyoma of the breast diagnosed by needle core biopsy. 

Tumour was incidentally discovered, on routine ultrasonography, 

in the right breast of a 36 year-old woman. Sonographically, 

tumour presented as a solid, hypoechoic, 2cm-mass with well 

circumscribed margins. 

Results. Needle core biopsy showed interlacing bundles of blandlooking 

eosinophilic spindle cells, closely reminiscent of leiomyoma. 

A lumpectomy was performed. Cut section showed a firm 

and white nodule with smooth external surface. Histologically, 

an unencapsulated tumour with the typical features of a classic 

leiomyoma was observed. Mitoses, nuclear pleomorphism or 

necrosis were absent. Immunohistochemical analyses, revealing a 

diffuse staining for desmin, α-smooth muscle actin, h-caldesmon 

and ER/PR, confirmed the diagnosis. 

The present case emphasizes that the diagnosis of intraparenchymal 

leiomyoma may be confidentially rendered on needle core 

biopsy. In this regard, it should be stressed that making a correct 

diagnosis is primarily dependent on awareness that this tumour 

may occur in the breast parenchyma. 

Diaphragmatic myositis causing unexplained 

neonatal sudden death 

1)Cesari S. 2)Dal bello B. 3)Perotti G. 4)Silini EM. 

1)Anatomia Patologica, Fondazione IRCCS San Matteo, Pavia, Italia 

2)Anatomia Patologica, Fondazione IRCCS San Matteo, Pavia, Italia 3)Patologia 

neonatale, Fondazione IRCCS San Matteo, Pavia, Italia 4)Anatomia 

patologica, Azienda Ospedaliero-Universitaria, Parma, Italia 

Background. The definition of sudden infant death syndrome 

(SIDS) requires a full post-mortem investigation to exclude 

identifiable causes of death according to detailed protocols. We 

describe the pathologic findings of a clinically unexplained sudden 

death in the perinatal/neonatal period. 

Methods. We performed autopsy on a 2 months old female newborn 

who suddenly died in her cot by an unexplained breathing 

arrest. She was born at 34 weeks of pregnancy by vaginal delivery 

and was apparently healthy until the acute event. Heart rhythm 

was restored after 30’ of cardiopulmonary resuscitation, but brain 

death by anoxia occurred after 6 days of mechanical ventilatory 

support. All tissues were sampled for histology, including the 

diaphragm, and samples from heart and spleen were frozen for 

long QT syndrome genetic analysis. 

Results. The newborn showed growth retardation (weight 3200 

g, crown-rump length 35 cm; total length cm 46). Macroscopic 

examination was negative except from pleural and peritoneal 

effusions. Histology showed: 1) brain ischemic necrosis and 

focal inflammatory meningeal infiltrates; 2) bilateral diffuse 

bronchopneumonia with acute alveolar damage, abscesses and 

focal organizing areas; 3) lympho-histiocytic myositis with 

extensive necrosis of fibres and dystrophic calcification of skeletal 

muscles, in particular the diaphragm. No bacterial or viral 

infections were identified. Long QT syndrome was excluded by 

genetic analysis. 

We concluded that the main cause of death was necrotizing 

myositis specifically involving diaphragm; bronchopneumonia 

was likely caused by abnormal respiratory movements and brain 

necrosis was due to anoxia during prolonged cardiac arrest. 

In conclusion, sudden neonatal death can be caused by diaphragmatic 

inflammatory pathology, as previously described in 5 newborns 

(Sundararajan, Med Sci Law 2005, 45 110-114). Sampling 


of diaphragm should be included in the post-mortem evaluation 

of these events. 

recurrent giant keloid of the sacral region treated 

with post-excisional radiotherapy 

1)Chiaramonte A. 2) Scaramuzzi G. 3)Tancredi A. 4)Troiano A. 

5)Bisceglia M 

1)Unit of Surgery, IRCCS Casa Sollievo della Sofferenza Hospital, San 

Giovanni Rotondo, Italy 2)Unit of Surgery, IRCCS Casa Sollievo della 

Sofferenza Hospital, San Giovanni Rotondo, Italy 3) Unit of Surgery, IRC- 

CS Casa Sollievo della Sofferenza Hospital, San Giovanni Rotondo, Italy 

4)Unit of Radiotherapy, IRCCS Casa Sollievo della Sofferenza Hospital, 

San Giovanni Rotondo, Italy 5)Unit of Anatomic Pathology, IRCCS Casa 

Sollievo della Sofferenza Hospital, San Giovanni Rotondo, Italy 

Background. Keloid is an abnormal pattern of dermal reaction to 

injury, resulting in excess collagen deposition. Various types of 

injuries are on record, such as surgery, trauma, burns, inflammatory 

skin diseases (folliculits, acne), viral dermatological diseases 

(chicken pox), vaccinations (Calmette-Guerin/BCG, small pox, 

and hepatitis B), fish stings (catfish), foreign bodies. Occasionally 

the injury may be clinically inapparent. The pathogenesis is unknown, 

but genetic, hormonal, or local factors may be involved. 

Black people are more frequently affected. It is usually sporadic, 

but familial occurrences have also been described 1 . There’s no 

sex prevalence among affected individuals. They can be either 

solitary or numerous, and may vary in size from small papules to 

large masses. Symptoms may vary from mild local distress (pain, 

pruritus) to cosmetic discomfort or anatomic disabilities, even to 

disfiguring deformities, associated with dramatic psychological 

and social side-effects. Keloids may occur at any age, but they are 

more common in the young. No universally accepted treatment 

protocol has been standardized, but several choices are available 

according to several factors (site, size, clinical history, prior treatments). 

Keloids are often resistant to treatment and have a high 

rate of recurrence 2 . 

Objectives. To report on a case of a recurrent giant (monstrous) 

keloid affecting a young patient, which eventually was treated 

with post-surgical radiotherapy. 

Case Report. A 22-year old Caucasian short man, 155 cm high 

(weight 65 kilos) was hospitalized, complaining of medical, 

anatomical, and psychosocial problems relating to a history of 

10 year duration of a recurrent keloid. The patient’s standing 

and walking were impaired and he needed assistance in coping 

with stairs. At physical examination a huge, bulging, oval mass 

with a knobby surface 45 cm in length (20 cm wide; 10 cm 

thick) was apparent on his lower back. The mass was firm in 

consistency and covered by skin which was focally eroded or 

moist with evil-smelling secretions, and occluding the anus. 

Physical examination also revealed a nodular exophytic mass 

5 cm in size, protruding from the umbilical scar, which appeared 

two years earlier. No other physical deformities were seen. His 

hands and fingers as well as his feet and toes were normal. Past 

medical history revealed excision of an intergluteal and perianal, 

subcutaneous fibrolipoma of 2 cm in size at the age of 11, which 

was histologically examined. At the age of 12 and at the age of 

14, he was hospitalized in specialized centers for surgical plastic 

reconstruction and underwent second and third surgical excisions 

due to keloid formations of 4 cm and 10 cm in size, respectively. 

A new keloid became evident shortly afterwards, which was at 

times treated with steroid injections without success. The tumor 

mass progressively enlarged reaching the above dimensions. The 

preoperative clinical suspicion was that of a sarcomatous growth, 

which was confirmed by means of PET-CT investigation, but 

needle biopsy did show a reactive proliferation of fibroblasts and 

myofibroblasts alternating with abundant acidophilic bands of 

collagen, typical of keloid. Simultaneoulsy the patient received 

genetic counseling and endocrinological evaluation, which


showed a normal male karyotype, and excluded keloid-associated 

genetic syndromes, familial history of keloidal formation, 

and diabetes mellitus. The patient also underwent neurological 

examination since he had been diagnosed with primary epilepsy, 

at the age of 5, which was confirmed, and was taking oral anticonvulsivant 

drugs (Depakine, Gardenal) for prophylaxis and 

maintenance since. The tumor mass was surgically extirpated 

en bloc (weight 3.400 gr) and sent for pathological examination. 

The surgical wound was repaired with a plastic reconstruction 

operation. Histological examination confirmed the diagnosis of 

keloid, which was only focally present at the lateral excision 

margins. 60 days after surgery radiotherapy was undertaken 

using photon 8MW (total dose delivered 22Gy in 11 days, with 

a daily fraction of 2 Gy). The umbilical keloid was not treated 

since surgery was not necessary and due to the patient’s predisposition 

to keloid formation. Follow-up: no recurrence has been 

noticed so far 20 months after this combined treatment (surgery 

plus post-surgical radiotherapy). 

Discussion. Hypertrophic scar and keloids are common occurrences, 

estimated to affect 5 to 15% of general population. Giant 

keloids are extremely rare with only 6 cases recorded in the 

literature, the largest reaching the size of 20 cm in its greatest 

diameter, all of which with a known history of injury, including 

unusual etiologies (chicken pox 3 , cat-fish sting, vaccination 

with BCG 1 case each), except 1 case with no attributed inciting 

cause 4 : the latter case was also the only arising in a familial context. 

Three cases showed multiple lesions of various dimensions. 

Although unique as to the size and deformity caused, our case 

can be categorized as one of the common sporadic cases: non-endocrine, 

since no endocrinological abnormalities was recognized, 

non-familial, since no keloidal inheritance pattern in his pedigree 

was ascertained, non-syndromic, since no stigma of keloid-associated 

syndromes (e.g., Rubinstein-Taybi syndrome) or of the 

disfiguring (infantile) hyaline fibromatosis were seen, and nondruggable, 

since the absence of any plausible role in keloidal proliferations. 

Instead the inciting factor was well identified as the 

first surgical trauma which triggered a likely individual genetic 

predisposition to keloid formation. The histological differential 

diagnosis include keloidal dermatofibroma, desmoplastic fibroblastoma 

(collagenous fibroma), hyaline fibromatosis. The case 

presented herein is the largest one ever observed, and one with 

most dramatic psychological impact (the patient lived almost 

in isolation due to shame of the disease) 5 . Keloids have been 

shown to respond to radiotherapy, pressure therapy, cryotherapy, 

intralesional and topical injections of corticosteroids, interferon 

and bleomicin or fluorouracil, topical silicone or other dressings, 

and laser treatment used alone or in various combinations, with 

variable but largely transient success 6 7 . Surgery has been used in 

case of necessity. Primary radiation therapy has been used for unresectable 

keloids 8 . The combination of surgery and postsurgical 

radiotherapy has already been proposed for cases where surgery 

is required, and has already been effectively used (follow-up > 2 

years) in another case of giant keloid 4 . 

Conclusion. Giant keloids are extremely rare, and they may 

respond to a combined approach (surgery plus postsurgical radiotherapy). 

references 

1 Marneros AG, Norris JE, Olsen BR, et al. Clinical genetics of familial 

keloids. Arch Dermatol. 2001;137:1429-34. 

2 Alster TS, Tanzi EL. Hypertrophic scars and keloids: etiology and 

management. Am J Clin Dermatol 2003;4:235-43. 

3 Gathse A, Ibara JR, Obengui Moyen G. Gigantic keloïds after chickenpox. 

A case report. Bull Soc Pathol Exot 2003;96:401-2. 

4 Jones K, Fuller CD, Luh JY, et al. Case report and summary of literature: 

giant perineal keloids treated with post-excisional radiotherapy. 

BMC Dermatol 2006;6:7. 

5 Furtado F, Hochman B, Ferrara SF, et al. What factors affect 

the quality of life of patients with keloids? Rev Assoc Med Bras 

2009;55:700-4. 

273 

6 Juckett G, Hartman-Adams H. Management of keloids and hypertrophic 

scars. Am Fam Physician 2009;80:253-60. 

7 Mutalik S. Treatment of keloids and hypertrophic scars. Indian J Dermatol 

Venereol Leprol 2005;71:3-8. 

8 Malaker K, Vijayraghavan K, Hodson I, et al. Retrospective analysis 

of treatment of nresectable keloids with primary radiation over 25 

years. Clinical Oncology 2004;16:290-8. 

fibro-myofibroblastic proliferation in the 

gallbladder wall. report of two cases 

1)S. Russo, 1)A. Cimmino, 1)A. Napoli, 1)M. Palumbo, 1)M. 

Colagrande, 1)M. Silecchia, 1)M. Stolfa, 1)R. Ricco, 2)V. Ninfo 

1)Dipartimento di Anatomia Patologica, Azienda Policlinico-Università 

di Bari, Bari, Italia; 2)Dipartimento di Scienze Oncologiche e Chirurgiche 

Azienda Ospedaliera-Università di Padova, Italia 

Background. Fibro-myofibroblastic proliferation is a lesion 

described in various organs over the last two decades. It is also 

called inflammatory pseudosarcomatous fibro-myxoid tumor and 

it can mimic sarcoma. Only three cases have been previously 

described in gallbladder. 

Methods. We report two cases: a 76-year-old female and a 

32-year-old male. Both presented with symptoms of chronic 

gallstone cholecystitis with recurrent episodes of fever, vomiting, 

nausea and epigastric pain. 

Ultrasound examination of the abdomen showed diffuse wall 

thickening, some calculi, without expansion of intra and extrahepatic 

bile ducts. 

Then both have undergone cholecystectomy. 

Results. Macroscopic examination of the gallbladder showed 

increased size, and the presence of many calculi.; the gallbladder 

wall was thickened. 

Histological examination showed in muscle layer and in perimuscular 

connective tissue fibroblastic and myofibroblastic proliferation 

with very mild nuclear atypia, associated with diffuse 

chronic inflammatory process composed of lymphocytes, plasma 

cells, macrophages, and neutrophil granulocytes. 

The distinction from solitary fibrous tumor, malignant fibrous 

hystiocitoma, fibrosarcoma and other similar entities was based 

upon the presence of mild nuclear atypia and the immunohistochemistry. 

In both cases the spindle cells stain positive for HHF35 and 

vimentin, rarely presented nuclear staining for Ki67 and were 

negative for S100, desmin and CD68. 

Our final diagnosis in both cases was fibro-myofibroblastic proliferation. 

Conclusions. The heterogeneicity in clinical behavior of the 

fibro-myofibroblastic proliferations previously described in various 

sites is probably related to different etiological factors: surgical 

trauma, infections and autoimmune disorders. 

The fibro-myofibroblastic proliferation of gallbladder is a very 

rare entity, always related with cholecystitis. Its uncertain malignant 

potential requires careful follow-up. 

A rare case of histiocytoid cardiomiopathy 

1)Cocca MP. 2)Nozza P. 3)Marzullo A. 4)Caruso G. 

1)Anatomia patologica, Università di bari, Bari, Italia 2)Anatomia patologica, 

Istituto giannina gaslini, Genova, Italia 3)Anatomia patologica, 

Policlinico, Bari, Italia 4)Anatomia patologica, Policlinico, Bari, Italia 

Background. Histiocytoid cardiomiopathy (HC) is a rare (about 

100 cases reported in literature), genetic cardiac disorder of 

infancy or childhood, predominantly affecting girls (M:F = 3:1) 

below the age of 2 years, which manifests clinically as severe 

cardiac arrhythmias or dilated cardiomiopathy and edema with 

heart failure and sudden death. Autosomal recessive, X–linked, 

and maternal inheritance has been described. Meanwhile, several 

reports indicate that HC is a cardiac manifestation of a mitochon-

274 

drial disorder which involves the Purkinje cells of the conduction 

tissue. 

Methods. We have seen in consultation a right atrial subendocardial 

mass of a child aged nine, Arabic and presenting Blackfan–Diamond 

anemia, which underwent allogeneic bone marrow 

transplantation. He then showed an acute GVHD, which became 

chronic and was complicated by repeated infections. 

Results. We observed nests of enlarged, polygonal, histiocyte– 

like cells with foamy granular or vacuolar, weakly eosinophilic 

cytoplasm, separated by fibrous branches, with calcium and hemosiderin 

deposition, but not mitoses. Immunohistochemistry 

showed expression of desmin and mithocondrial protein; S100 

protein and CD68–PGM1 resulted negative. 

Differential diagnosis must be effected between HC and cardiac 

rhabdomyoma (CR), the most common pediatric heart tumor. In 

CR clinical features includes arrhytmias, outflow tract obstruction, 

heart failure and hydrops fetalis and is often associated with 

tuberous sclerosis complex. It appears as a well demarcated mass, 

usually in the ventricles that consists in nodules of enlarged cardiomyocites 

with cleared cytoplasm PAS + for glycogen content, 

with vacuolization and myofilaments. CR has a natural history of 

spontaneous regression, without surgery. 

Our case deserves to be reported since our patient was male, 

older than typical cases for this condition and, finally, because 

the diagnosis was made on biopsy material rather than autopsy, 

as is usual. 

Jugular paraganglioma: report of a case 

1)Cocca MP. 2)Palumbo M. 3)Resta L. 4)Cimmino A. 5)Ninfo 

V. 

1)Anatomia Patologica, Policlinico Di Bari, Bari, Italia 2)Dipartimento 

Di Anatomia Patologica, Policlinico Di Bari, Bari, Italia 3)Dipartimento 



Di Anatomia Patologica, Policlinico Di Padova, Padova, Italia 

Background. Paraganglioma of the head and neck (HNP) represent 

rare tumors that arise from extraadrenal chromaffin cells 

of neural crest origin. They represent 10-18% of all chromaffin 

tissue-related tumors which are reported at a rate of 2-8 cases/ 

million·yr They are highly vascular neoplasms that are benign 

in the majority of the case. Common sites of origin are carotid 

bifurcation, jugular bulb, timpanic plexus and vagal nerve. It is a 

common cancer in women between 50 and 70 years. 10% of cases 

are familial paraganglioma (autosomal dominant with variable 

penetrance) plurifocal, secreting. 

Methods. We describe the case of a big jugular paraganglioma in 

a 13 year old boy who presented about a month hearing loss, ear 

pain and a swollen left temporal mass that at RMI was occupying 

the middle cranial fossa, extending to epicranic soft tissues 

without infiltrate the brain parenchyma. 

Results. We received some fragments of firm consistency, graybrownish 

in colour. Histologically we observed a mesenchymal 

neoplasm with organoids growth pattern, with nests or cords of 

large monomorphic cells, round or polygonal, without atypia 

nor mitosis; vesicular nuclei and prominent nucleoli, intensely 

eosinophilic cytoplasm and finely granular or clear appearance, 

bordered by connective tissue containing a rich network of 

capillaries. The tumor invaded the bone. Morphological appearance 

seemed compatible with an alveolar sarcoma. At immunohistochemistry, 

we found expression for desmin, muscle actin 

- smooth, CD10, NSE, synaptophysin and focally for chromogranin; 

negative for actin HHF - 35, EMA and cytokeratin. We 

performed antibody for S-100 protein that revealed a cell population 

not made immediately obvious in hematoxylin-eosin: a 

network of cells sustentacular support. The approach to the study 

of this type of tumor was made difficult by the size and extent 

unusual to epicranic soft tissue. 


Secretory meningioma of the orbit: 

report of a rare case 

1)Cocca MP. 2)Piscitelli D. 3)Palumbo M. 4)Fiore MG. 5)Rossi 

R. 6)Resta L. 

1)Dipartimento Di Anatomia Patologica, Policlinico Di Bari, Bari, Italia 






Background. Meningiomas account for approssimately 15- 

25% of all primary intracranial tumors. They are derived from 

arachnoid cap cells and are most often seen in middle-aged and 

elderly patients. The vast majority of these tumors are benign; 

however, complete removal can be difficult and recurrence 

is an issue. Orbital meningiomas represent between 0,4 and 

2% of all intracranial meningiomas and account for 3 to 9% 

of all orbital tumors. Primary orbital meningiomas arise from 

the optic nerve sheath and may infiltrate the sphenoid wing 

producing hyperostosis. The secretory meningioma is a welldifferentiated 

variant, relatively rare. Clear cell meningioma 

is an uncommon, aggressive variant of meningioma usually 

affecting younger females. 

Methods. We describe the case of a woman aged 75 who presented 

exophtalmia and a severely impaired vision. Computed 

tomographic scan of the upper wall of the orbit showed a wellcircumscribed 

mass measuring 35 × 30 × 20 mm. The resected 

specimen consisted of a white-grey fragment, firm in consistency. 

For histological analysis, the specimen was fixed in 10% formaldehyde 

and embedded in paraffin blocks. Four – micrometer 

sections of the paraffin blocks were stained with Hematoxylin 

– eosin and periodic acid – Schiff (PAS). 

Results. We observed lobules of cells, surrounded by thin collagenous 

septae. Tumor cells were largely, uniform, with oval 

nuclei, with focal epithelial differentiation, in the form of intracellular 

lumina containing PAS-positive, eosinophilic material. 

In some areas, tumor cells were polygonal, with clear, glycogenrich 

cytoplasm. These structures stain immunoistochemically for 

CEA. The surrounding tumor cells stain for cytokeratin, EMA 

and Progesterone Receptor. 

Conclusions. This case illustrates unusual features of an admixture 

of two rare variant of meningioma in an adult woman with 

an unusual supratentorial location. 

Adventitial cystic disease of the popliteal artery. 

Case report and review of the literature 

1)Colacchio G. 2)Ferrante A. 3)Palena G. 4)Coggia M. 5)Bisceglia 

M. 

1)Unit of Vascular Surgery M.A.S., IRCCS “Casa Sollievo della Sofferenza” 

Hospital , S. Giovanni Rotondo, Italy 2) Unit of Vascular Surgery 

M.A.S., IRCCS “Casa Sollievo della Sofferenza” Hospital, S. Giovanni 

Rotondo, Italy 3)Unit of Vascular Surgery M.A.S., IRCCS “Casa Sollievo 

della Sofferenza” Hospital, S. Giovanni Rotondo, Italy 4)Department of 

Vascular Surgery, Ambroise Paré University Hospital, Boulogne-Billancourt, 

France 5)Unit of Anatomic Pathology, IRCCS “Casa Sollievo della 

Sofferenza” Hospital, S. Giovanni Rotondo, Italy 

Background. Adventitial cystic disease (ACD) is a rare condition 

involving arterial segments. It represents an uncommon 

cause of intermittent claudication, secondary to external arterial 

lumen compression. ACD is estimated to account for less 

than 0.1% of all patients evaluated for intermittent claudication. 

Around 400 cases have been described to date 1 2 . ACD typically 

afflicts young to middle-aged men without a history of trauma, 

systemic arterial disease, and risk factors for atherosclerosis. 

The microscopical appearance of the involved arterial tract is 

that of mucinous degeneration, analogous to that of soft tissue


ganglion. The most commonly affected artery is the popliteal 

artery, although other arteries, such as the external iliac artery, 

femoral artery, radial artery, and even veins (e.g. saphenous vein) 

may be involved. 

Objectives. To report a case of severe ACD, involving the 

popliteal artery which was surgically treated and histologically 

examined. 

Case Report. The patient was a 44-year old healthy, male, moderate 

smoker, who complained of intermittent claudication in the 

left calf for six months. At physical examination no distal pulses 

were found in the left leg. MRI scan demonstrated reduction of 

the arterial lumen due to compression by a fluid-filled cystic 

lesion developing in the vascular adventitia, in the absence of 

luminal thrombosis. The surgical treatment consisted of resection 

of the affected tract of popliteal artery with direct arterial reconstruction 

obtained using an inverted external saphenous vein. 

Results. The resection specimen was 5 cm long and had a fusiform, 

sausage-shaped aspect. The artery was encased by a 

circumferential cystic lesion 2.5 cm in diameter, filled by jellylike 

material and bulging into the arterial lumen which at places 

was occluded. No communication was demonstrated between 

the cystic lesion and the arterial lumen. Histological examination 

showed myxoid degeneration of the connective tissue of 

the arterial adventitial layer, analogous to the changes seen in a 

ganglion of soft tissue, and the diagnosis of ACD of the popliteal 

artery was made. Distal pulse restoration was almost immediate, 

the postoperative course was uneventful, and the patient was discharged 

on the eighth day following surgery, on oral anticoagulant 

therapy. At 1-year follow-up the patient reported complete 

remission of his claudication. 

Discussion. ACD is a disease of males (male to female sex ratio 

15:1), affecting middle-aged individuals (age range 30-50). The 

fluid-filled cystic lesion develops in the adventitial layer of the 

artery and is responsible for the narrowing of the vascular lumen 

and limitation of arterial flow. The distal arterial pulses are 

usually normal at rest, although they typically disappear with 

exercise (Ishikawa’s sign). The disease is progressive and in 

severely advanced cases the compression may lead to total occlusion 

of the arterial lumen. In any case the impact on arterial 

flow is intermittent, explaining the discontinuous nature of the 

claudication. Some patients can experience a variable symptomfree 

interval, plausibly due to redistribution of pressure between 

the various compartments in cases with multilocular cysts and 

in cases with cysts communicating with adjacent tendon cysts; 

however, even spontaneous lymphatic drainage of the cyst or 

leakage by rupture into the surrounding connective tissues can 

also relieve obstruction, if only temporarily. In other cases, the 

progressive compression on the arterial lumen produces progressive 

worsening of the claudication, leading up to leg pain at 

rest. The pathogenesis of ACD is similar to that of the soft tissue 

ganglion. The clinical differential diagnosis includes popliteal 

artery entrapment syndrome, popliteal aneurysm, Buerger’s 

disease and embolism. Resection of the affected tract of the 

artery and in situ graft (mostly using external saphenous vein) 

is the most widely accepted modality of therapy. Alternative 

but questionable or ineffective proposed therapeutic procedures 

are aspiration of the cyst contents, incision and cyst evacuation, 

and endovascular stent placement. The pathological diagnosis 

is easy. Analogies can be made also to ganglion cysts developing 

in the sheath of a peripheral nerve, which parenthetically 

is also seen in the popliteal fossa usually involving herein the 

peroneal nerve 3 . 

references 

1 Mino MJ, Garrigues DG, Pierce DS, et al. Cystic adventitial disease of 

the popliteal artery. J Vasc Surg 2009;49:1324. 

2 Meka M, Hamrick MC, Wixon CL. Cystic Adventitial Disease of the 

Popliteal Artery: A Potential Cause of Lower Extremity Claudication. 

Am Surg 2009;75:86-9. 

275 

3 Rawal A, Ratnam KR, Yin Q, et al. Compression neuropathy of common 

peroneal nerve caused by an extraneural ganglion: a report of 

two cases. Microsurgery 2004;24:63-6. 

follicular dendritic cell tumor 

1)Colagrande A. 2)Scivetti A. 3)Scamarcio R. 4)Angelotti U.F. 

5)Traversi C. 6)Rossi R. 7)Cimmino A. 8)Resta L. 

1)Anatomia patologica, Policlinico, Bari, Italia 2)Anatomia patologica, 





Policlinico, Bari, Italia 

Background. The antigen-presenting accessory cells, collectively 

know as reticulum cells, comprising the mononuclear phagocyte 

and immunoregulatory effector system of the lymphoid tissue 

include three different types of cells: 1) the follicular dendritic 

cell (FDC) confined exclusively to the B-cell follicular compartment, 

which plays a major role in the induction and maintenance 

of humoral immune responses; 2) the interdigitating dendritic 

cell (IDC) located in the T-cell zones of lymph nodes, tonsil, and 

spleen, which is a potent antigen-presenting cell responsible for 

stimulating resting T cells in the primary immune response; 3) the 

fibroblastic reticular cell (FBRC), which is a stromal support cell 

located in the parafollicular areas and deep cortex usually dislaying 

myofibroblastic features. 

All three cell types can undergo neoplastic transformation. 

Our report concerns a splenic tumor with morfhological characteristics 

of FDC. 

Methods. Case report: a 55 year-old woman with enlarged spleen 

and lymphoadenopathy. 

Diagnosis: tumor composed by oval to spindled cells with eosinophilic 

cytoplasm arranged in clusters or fascicles in a storiform 

growth pattern. 

The tumor cells are characteristically admixed with small lymphocytes. 

Their nuclei are elongated witht small eosinophilic nucleoli, and 

finely dispersed chromatin. 

Results. Immunoistochemically, the neoplastic cells express 

CD21, CD68 PGM1, fascin and vimentin; CD34, CD35, CD23, 

S-100 and EMA are negative. 

Ultrastructurally, the tumor show complex intergitating cytoplasm 

processes joined by desmosome like structures. 

FDC neoplasms are rare low grade malignant tumors. The behaviour 

is more akin to that of soft tissue sarcomas than lymphomas. 

The majority of cases involve lumph nodes of the neck, axilla, 

and mediastinum, although approximately 30% tumors involve 

extranodal sites such as liver, oral cavity, tonsils and intraabdominal 

soft tissues. 

The splenic site is very infrequent. 

Can claudin-5 protein be a good marker in the 

assessment of papillary thyroid carcinoma? 

1)Colato C. 2)Dardano A. 3)Brazzarola P. 4)Monzani F. 5)Menestrina 

F. 6)Chilosi M. 7)Ferdeghini M. 

1)Patologia & Diagnostica, Università di Verona, Verona, Italia 2)Medicina 

Interna, Università di Pisa, Pisa, Italia 3)Scienze Chirurgiche, Università 

di Verona, Verona, Italia 4)Medicina Interna, Università di Pisa, Pisa, 

Italia 5)Patologia & Diagnostica, Università di Verona, Verona, Italia 

6)Patologia & Diagnostica, Università di Verona, Verona, Italia 7)Patologia 

& Diagnostica, Università di Verona, Verona, Italia 

Background. Claudins (CLDNs), the main components of tight 

junction proteins, play a role in cell proliferation, adhesion and 

tumorigenesis. Their expression profile is complex and appears 

to be organ dependent.

276 

CLDN5 is claimed to be specific for endothelial cells but its 

expression has been found in several cancers and may also be 

associated with an aggressive behaviour. 

In thyroid tissue, CLDN5 immunostaining is not well characterized. 

Aim. To test CLDN5 protein expression in different papillary 

thyroid carcinoma (PTC) samples with an aggressive course 

as well as in corresponding regional lymph node metastases 

(LNM), compared with that of normal adult and fetal thyroid 

tissue. 

Methods. 83 PTC samples [48 classic, 22 follicular, 9 tall cell 

and 4 solid variants] and 27 LNM were selected for immunohistochemical 

analysis. 

Results. CLDN5 was negative in 45,7% of primary tumors and in 

59,3% of LNM. In the positive cases, a discontinuous pericellular 

staining was found in only a few samples while weak and focalized 

reactivity was observed in most of the cases. No correlation 

with histological subtypes, the nodal status, or tumor size was 

found. In fetal thyroid glands CLDN5 showed a discontinuous 

linear or dot staining on the lateral membrane similar to normal 

adult thyroid tissue. The vascular endothelium displayed strong 

positivity. 

Conclusions. CLDN5 was similarly expressed both in fetal and 

adult thyroid tissue. Its exact physiological role is still not clear. 

In a previous study, CLDN5 expression has been described in 

all thyroid tumors evaluated except one, suggesting a potential 

involvement of this molecule in carcinogenesis and metastasis. 

Conversely, we found only a faint or moderate CLDN5 immunostaining 

in 54,3% of PTC and in 40,7% of LNM; therefore, our 

data does not support a significant role of CLDN5 in thyroid cell 

tumorigenesis or metastasis. 

Claudin-1 expression in solid cell nests of the 

thyroid: further evidence for a histogenetic link 

with papillary carcinoma 

1)Colato C. 2)Fierabracci A. 3)Gobbato M. 4)Martignoni G. 

5)Dardano A. 6)Monzani F. 7)Chilosi M. 8)Menestrina F. 9)Ferdeghini 

M. 

1)Patologia & Diagnostica, Università di Verona, Verona, Italia 2) Laboratorio 

di autoimmunità e di cellule staminali d’organo, Ospedale Pediatrico 

Bambino Gesù;, IRCCS, Roma, Italia 3)Patologia & Diagnostica, 

Università di Verona, Verona, Italia 4)Patologia & Diagnostica, Università 

di Verona, Verona, Italia 5)Medicina Interna, Università di Pisa, Pisa, 

Italia 6)Medicina Interna, Università di Pisa, Pisa, Italia 7)Patologia & 

Diagnostica, Università di Verona, Verona, Italia 8)Patologia & Diagnostica, 

Università di Verona, Verona, Italia 9)Patologia & Diagnostica, 

Università di Verona, Verona, Italia 

Background. Solid cell nests (SCNs), a normal component of the 

human thyroid gland, are considered remnants of the ultimobranchial 

body. The immunohistochemical profile of SCNs is well 

characterized and, as they apparently harbor the minimal properties 

of a stem cell phenotype, can be regarded as a pool of stem 

cells of the adult thyroid. Moreover, the immunohistochemical 

detection of p63 in SCNs and BRAF mutation in solid cell nest 

hyperplasia suggest a link between these embryonic remnants and 

papillary thyroid carcinoma (PTC). 

However the biological significance of SCNs remains disputable. 

Claudins (CLDNs), a family of tight junction proteins, play a 

role in adhesion, cell proliferation, and tumorigenesis. Recently, 

CLDN1 was found to be up-regulated in PTC both at the gene 

and protein level. 

Aim. To assess the immunohistochemical expression of CLDN1 

in a collection of SCNs. 

Methods. 15 cases of SCNs with solid or cystic features, found 

incidentally in specimens resected for PTC, follicular adenomas 

and multinodular goiter, were selected. 


Results. In all cases, SCNs displayed a strong, diffuse and linear 

membrane staining, forming a honeycomb-like pattern. In contrast, 

the C cells were constantly negative and in normal thyroid 

tissue only scattered positive cells were observed. 

Conclusions. We report for the first time CLDN1 expression in 

SCNs. This marker appears a highly sensitive tool in detecting 

SCNs, as described for p63 and Galectin-3. We confirm that 

CLDN1 is frequently up-regulated in PTC and may represent a 

novel marker for this tumor as well as Galectin-3. This finding 

supports the hypothesis of a histogenetic link between SCNs and 

PTC. 

Finally, CLDN1 immunoreactivity may also be useful in separating 

SCNs from C cells, as normal C cells do not express this 

membrane protein. 

Analysis of tyrosine-kinase receptors expression in 

endometrial stromal sarcoma 

1)Cossu Rocca P. 2)De Miglio M.R. 3)Mura A. 4)Uras M.G. 

5)Contini M. 6)Maricosu E. 7)Manca A. 8)Carru C. 9)Bosincu 

L. 10)Massarelli G. 

1)Anatomia patologica, Università di Sassari, Sassari, Italia 2)Scienze 

biomediche, Università di Sassari, Sassari, Italia 3)Anatomia patologica, 

Università di Sassari, Sassari, Italia 4)Anatomia patologica, Università di 

Sassari, Sassari, Italia 5)Anatomia patologica, Università di Sassari, Sassari, 

Italia 6)Anatomia patologica, Università di Sassari, Sassari, Italia 

7) Anatomia patologica, Università di Sassari, Sassari, Italia 8)Scienze 

biomediche, Università di Sassari, Sassari, Italia 9)Anatomia patologica, 

Università di Sassari, Sassari, Italia 10)Anatomia patologica, Università 

di Sassari, Sassari, Italia 

Background. Endometrial stromal sarcomas (ESS) are rare neoplasms, 

which are currently classified in low grade (LG) ESS, 

with indolent growth, tendency to local recurrences and, rarely, 

to metastasize, and undifferentiated endometrial sarcomas (UES), 

with a very aggressive behavior. Surgery remains the treatment of 

choice for ESS. Whilst hormonal therapy has been claimed as a 

successful therapy to decrease recurrences in LG-ESS, nonetheless, 

effective adjuvant therapy to prolong survival has not yet 

been established. Thus, alternative approaches such as molecularly 

targeted therapies need to be investigated. 

Aim of our study was to analyze immunohistochemical expression 

of tyrosine kinase receptors in a series of ESS, to evaluate 

their potential role as molecular targets. 

Methods. Immunohistochemistry was performed in 10 ESS, 

including 7 LG-ESS and 3 UES. Specific antibodies against 

ABL, CD117, EGFR, PDGFR-α have been utilized. Staining 

intensity and percentage of positive cells were scored for each 

case. 

Results. ABL expression was detected in 70% of cases, i.e. 5 

out of 7 LG-ESS and 2 out of 3 UES, with % of positive cells 

ranging from 10 to 50%, and staining intensity ranging from 1+ 

to 2+. EGFR expression was observed in 90% of cases, i.e. 6 out 

of 7 LG-ESS and all the 3 UES, with % of positive cells ranging 

from 50 to 80%, and staining intensity ranging from 1+ to 

3+. PDGFR-α expression was appreciable in 90% of the cases, 

i.e. 6 out of 7 LG-ESS and all the 3 UES, with % of positive 

cells ranging from 30 to 70%, and staining intensity ranging 

from 1+ to 2+. CD117 expression was consistently negative in 

all the cases. 

Conclusion. Our study confirms that tyrosine kinase receptors 

expression is frequently observed in ESS. Further studies are 

needed to identify specific genetic abnormalities, potentially 

useful to select patients who might benefit from current targeted 

therapeutic options.


Metastasis of colon cancer to the thyroid gland: 

a case diagnosed on fine-needle aspirate by 

a combined cytological, immunocytochemical 

and molecular approach 

1)Cozzolino I. 2)Malapelle U. 3)Carlomagno C. 4)Varone V. 

5)Palombini L. 6)Troncone G. 


II”, Napoli, Italia 2)Scienze biomorfologiche e funzionali, Università 

di Napoli “Federico II”, Napoli, Italia 3)Oncologia ed endocrinologia 

molecolare e clinica, Università di Napoli “Federico II”, Napoli, Italia 




Università di Napoli “Federico II”, Napoli, Italia 

Background. Fine-needle aspiration (FNA) with cytological 

evaluation reliably diagnoses primary and secondary thyroid neoplasms. 

However, identifying the primary origin of a metastatic 

process involving the thyroid gland is challenging. In particular, 

metastasis of colon cancer to the thyroid gland is very rare. 

Comparison between the SIAPeC-IAP and 

the Bethesda systems for reporting thyroid 

cytopathology: experience in two hospitals 

1)Crippa S. 2)Bongiovanni M. 

1)Institute of Pathology, Locarno, Switzerland 2)Pathology, Geneva University 

Hospitals, Geneva, Switzerland 

Backgroung. The 5-tiered SIAPEC-IAP thyroid FNA System 

and the new-6-tiered Bethesda thyroid FNA System offer two approaches 

to the problem of reporting thyroid FNA resultas. In this 

study, we present the combined experience from our instituions 

for reporting thyroid FNAs using these two different systems and 

evaluate their efficacy based upon surgical follow-up. 

Design. Data on thyroid FNAs and their corresonding surgical 

specimens were collected at our two instituions over a two-year 

period. We compared the sensitivity and specificity for each of 

the component group within the 2 systems where a diagnosis lead 

to surgery (CAT 3, 4, 5, 6 and TIR 3, 4, 5 versus benign). 

Results 

SIAPEC-IAP 

system 

Bethesda 

system 

277 

Methods. In this case report, a right lobe solid thyroid nodule 

in a 66-year-old male was aspirated. FNA cytology showed necrosis 

and atypical tall columnar cells; since, the patient at age 

60 had undergone surgery for a sigmoid-rectal cancer metastasizing 

to the liver and subsequently to the lung, a suspicion of 

metastasis from colon cancer was raised. This was corroborated 

by cell-block immunocytochemistry showing a cytokeratin (CK) 

7 negative/CK20-positive staining pattern; thyreoglobulin and 

TTF-1 were both negative. Since KRAS codon 12/13 mutations 

frequently occur in colon cancer, whereas they are extremely 

uncommon in primary thyroid tumors, DNA was extracted from 

the aspirated cells, and KRAS mutational analysis was carried 

out. The codon 12 G12D mutation was found; the same mutation 

was evident in the primary cancer of the colon and in its liver and 

lung metastasis. 

Conclusion. Thus, a combined cytological, immunocytochemical 

and molecular approach unquestionably correlated metastatic 

adenocarcinoma cells aspirated from the thyroid to a colo-rectal 

origin. 

category fna % malignant Benign category fna % malignant Benign 

tir1 34 8.8 2 4 cat1 31 10.5 0 4 

tir2 306 79.3 2 24 cat2 156 53 2 15 

cat3 4 1.4 / / 

The sensitivity for TIR 3, 4, 5 and for CAT 3, 4,5, 6 were almost 

equal in the two reporting systems, SIAPEC-IAP and Bethesda 

(91,6% vs 90%), while specificity was higher in the British (60% 

vs 31,9%). Interestingly, the number of cases in the TIR 3 and 

CAT 4 categories (indeterminate/suspicious) differed significantly 

between the two reporting systems (6,2 and 27%). For these two 

categories, the rate of malignancy on surgical excision is similar 

(21% and 18%), with a PPV of 17% and 15% respectively. 

Conclusions. The two reporting systems, the SIAPEC and the 

Bethesda, show similar sensitivities but the SIAPEC-IAP system 

exhibited better specificity, possibly related to differences in the 

use of the so called grey zone (TIR3 and CAT 4) categories. 

Larger studies will be useful to further confirm these data. 

tir3 24 6.2 3 14 cat4 79 27 5 28 

tir4 6 1.6 5 1 cat5 13 4.4 4 4 

tir5 16 4.1 14 1 cat6 11 3.7 9 0 

total 386 26 (37%) 44 (63%) totaltal 294 20 (28%) 51 (72%) 

effect of multidisciplinary work and training in 

fine-needle aspiration of the thyroid 

1)S. Crippa, 2)S. Suriano, 1)L. Mazzucchelli, 2)L. Giovanella 

1)Institute of Pathology, Locarno, Switzerland; 2)Nuclear Medicine and 

PET/CT Centre, Oncology Institute of Southern Switzerland, Bellinzona, 

Switzerland 

Background. Fine needle aspiration cytology (FNAC) is an effective 

diagnostic tool for patient with thyroid nodules. Specifity 

and sensitivity of this technique depend on operator’s experience, 

and lead to better results when performed in a multidisciplinary 

setting. Clinical significance of FNAC can be further enhanced 

by adopting standardized reporting, as proposed by the British 

Thyroid Association – Royal College of Physicians in 2002 

(Thy1 to Thy5) and SIAPEC-IAP in 2007 (TIR1 to TIR 5). 

Methods. We evaluated the activity over the past two years of our 

multidisciplinary Thyroid Unit ambulatory for thyroid FNAC. 

When available, definitive histological diagnosis were compared 

to the respective cytological diagnostic categories. Data concerning 

3 rd year of activity are in progress.

278 

Results 

1 st year 

(179 nodules) 

Multicentricity in lobular carcinoma of the breast 

is related to the histological grade 

1)Cucchi M.C. 2)Foschini M.P. 

1)Department of oncology, unit of surgical pathology, Bellaria hospital, 

Bologna, Italy 2)Department of pathology, Bellaria hospital, Bologna, 

Italy 

Background. Incidence of multicentricity in cases of Invasive 

Lobular Carcinoma (ILC) of the breast is a matter of debate. The 

overall rate of multiple foci of ILC is considered about 10% of 

the cases. 

ILC is classically considered a grade II lesion. Nevertheless several 

histological variants of ILC have been described, some of 

which are histologically grade III lesions. 

The purpose of the present study is to evaluate the relation between 

histological grade and presence of multicentricity in ILC. 

Methods. All cases of ILC diagnosed in the period January 1997- 

December 2007 were retrieved from the files of the Department 

of Anatomic Pathology of the University of Bologna at Bellaria 

Hospital. Slides were reviewed and ILC classified and graded according 

to currently accepted criteria 1 2 . Multicentricity was considered 

when foci of ILC were detected in different quadrants. 

Results. 164 cases of ILC constituted the basis of the present 

study. Cases were subdivided as follows: ILC classic variant 

grade II(ILC-cl): 102 cases; ILC grade III (ILC-grIII): 22 cases; 

ILC pleomorphic variant, grade III, 40 cases (ILC-PgrIII). 

Multicentricity was detected in 13\102 ILC-cl (12,7%), in 8\22 

ILC-grIII (36.3%) and in 11\40 ILC-PgrIII (27.5%). 29/40 cases 

(72,5%) of ILC-PgIII were treated with mastectomy, while the 

same procedure was applied in 55/102 cases of ILC-cl (53,9%). 

Multicentricity was about three times more frequent in cases of 

ILC grade III (including the pleomorphic variant). Consequently 

mastectomy was more frequently carried out in of ILC-PgIII, 

while breast conserving treatment (quadrantectomy) was applied 

most frequently in ILC-cl. 

references 

1 Elston CW, Ellis IO. Pathological prognostics factors in breast 

cancer. The value of histological grade in breast cancer: experience 

from a large study with a long-term follow-up. Histopathology 

1991;19:403-10. 

2 Tavassoli F, Eusebi V. Tumors of the Mammary Gland, AFIP series 

2009. 

2 nd year 

(207 nodules) 


category cases % benign malignant category cases % benign malignant 

tir1 18 10.0 2 1 tir1 16 7.7 2 1 

tir2 142 79.3 14 2 tir2 164 79.2 10 0 

tir3 10 5.6 4 1 tir3 14 6.8 10 2 

tir4 3 1.7 1 2 tir4 3 1.5 0 3 

tir5 6 3.4 0 5 tir5 10 4.8 1 9 

Over the period examined, we detected 2/386 false negative and 

1 /386 false positive nodules. The positive predictive value for 

TIR5 was 92.8%, and for TIR4 83.3%. The negative predictive 

value for TIR3 was 82.3%. Sensitivity and specificity for TIR4 

and TIR5 categories were 90.5% and 92.3%, respectively. The 

percentage of TIR1 categories diminished from the first to the 

second year. Preliminary results of the 3 rd year of activity confirm 

this trend. 

Conclusions. 1. Standardized diagnostic categories for FNAC 

enhance diagnostic reproducibility and facilitate communication 

with clinicians. 2. A multidisciplinary integrative diagnostic approach 

for thyroid nodules allows the formulation of clear recommendations 

for subsequent surgical procedures. 3. Preliminary 

results of the 3 rd year of activity confirm this trend. 

High prevalence of B-rAf mutation in papillary carcinoma 

of the thyroid in north east italy 

1)Cuorvo L.V. 2)Girlando S. 3)Bonzanini M. 4)Morelli L. 

5)Amadori P. 6)Dalla palma P. 7)Barbareschi M. 

1)Anatomia Patologica, S.Chiara, Trento, Italia 2)Anatomia Patologica, 

S.Chiara, Trento, Italia 3)Anatomia Patologica, S.Chiara, Trento, Italia 

4)Anatomia Patologica, S.Chiara, Trento, Italia 5)Ambulatorio Di Endocrinologia, 

Apss, Trento, Italia 6)Anatomia Patologica, S.Chiara, Trento, 

Italia 7)Anatomia Patologica, S.Chiara, Trento, Italia 

Background. B-RAF is one of the three isoforms of the serine-threonine 

kinase RAF (A-RAF, B-RAF and C-RAF) that is 

regulated by RAS and activates downstream effectors molecules. 

B-RAF V600E mutation is frequently observed in several tumors, 

including papillary thyroid carcinomas (PTC), where it is considered 

of potential diagnostic and prognostic value. The reported 

prevalence of B-RAF mutation in PTC in different Italian populations 

varies from 14% to 69%. 

Methods. We investigated the prevalence and utility of the B-RAF 

V600E mutation in a series of 91 fine needle aspiration biopsies of 

the thyroid and in 60 histologically proven PTC in a well defined 

North Italian population. Every sample was selected and processed 

for DNA extraction. Successively, Exon 15 B-RAF mutations were 

identified by direct DNA sequencing analysis using the AB PRISM 

310 (Applied Biosystems, Foster City, CA). 

Results. In our series B-RAF mutation has been detected in 43 

(72%) PTC, and was more frequent in classic (34 out of 44, 77%) 

vs follicular PTC (9 out of 16, 56%). Forty one (46%) FNAB 

showed B-RAF mutation and corresponded to histologically 

proven PTC (33 classic type and 8 PTCVF), which had been citologically 

classified as: malignant (28 cases), atypical/suspicious 

(10), inadequate (1) and benign (2). B-RAF mutations were never 

seen in non PTC/PTCVF FNAB cases, implying a 100% positive 

predictive value. Our data demonstrate a high prevalence of B- 

RAF mutation in our study population, underscoring the possibility 

of strong regional differences in B-RAF mutation prevalence 

in PTC, and further confirm its high diagnostic on FNAB. 

In search for correlation among markers for 

limbal stem cells niche 

1)Curcio (C). 2)Calienno (R). 3)Lanzini (M). 4)Iezzi (M). 5)Mariotti 

(M). 6)Colesanti (M). 7)Musiani (P). 8)Nubile (M). 

1)Oncologia E Neuroscienze/Oftalmologia, SS. Annunziata/CESI, Chieti, 

Italia 2)Oftalmologia Clinica, SS. Annunziata, Chieti, Italia 3)Onco-


logia E Neuroscienze/Oftalmologia, Ss. Annunziata/CESI, Chieti, Italia 

4)Anatomia Patologica/ Oncologia E Neuroscienze, SS. Annunziata/CESI, 

Chieti, Italia 5)Oncologia E Neuroscienze/Oftalmologia, SS. Annunziata/ 

CESI, Chieti, Italia 6)Oftalmologia Clinica, SS. Annunziata, Chieti, Italia 

7)Anatomia Patologica/ Oncologia E Neuroscienze, Ss. Annunziata/CESI, 

Chieti, Italia 8)Oftalmologia Clinica, SS. Annunziata, Chieti, Italia 

Background. The corneoscleral limbus is known to be the site of 

corneal epithelial stem cells (SC). Several molecules have been 

proposed as SC markers but none of them is able to univocally 

identify them. The aim of this study was to evaluate co-expression 

of different SC markers in human limbus. 

Methods. In this work five corneoscleral specimens from normal 

human donor eye-bank eyes (age 52-73 years) were fixed in 

formalin, divided in 8 segments, embedded in paraffin and examined 

by immunohistochemistry and immunofluorescence for 

p63, vimentin (vim), laminin 5, integrin (Int) α6, int β1, int β4, 

connexin 43, ki67 and N-cadherin positivity. We firstly analyzed 

the distribution and the anatomical structure of limbal crypts in 

each of the segments. Then we evaluated the percentage of positive 

areas in the niches. Finally we looked for colocalizations and 

possible correlations among markers. 

Results. We confirmed a different number of niches among the 

segments of the same corneoscleral rim. Moreover we observed 

high variability of niches number among patients which interestingly 

correlates with the percentage of p63 positivity of niche 

cells. Confocal microscopy double staining for p63 and vim did 

not show evident colocalization and vim + cells were seen in the 

superficial layers rather than in the deep layer of crypts. Int β1 

staining directly correlated with p63 positivity while the remaining 

proteins appeared variably and widely distributed. 

Colocalization was evident at least for two SC markers (Int _1/ 

p63) within the basal layers, while vim, expressed mainly in the 

superficial layers could act as late progenitor cell marker 

Microsatellite analysis of rare serous tumors 

of the fallopian tubes. Comparison with serous 

ovarian neoplasms 

D'Adda T., Manni S., Giordano G. 

Dipartimento di Patologia e Medicina di Laboratorio, Sezione di Anatomia 

Patologica, Università degli Studi di Parma, Parma, Italia 

Background. Primary tubaric serous carcinomas are rare neoplasms 

morphologically similar to high-grade (HG) ovarian carcinomas. 

Lately, several evidences suggested that Fallopian tubes 

may be the site of origin of most HG ovarian serous carcinomas. 

Also benign neoplasms develop in the Fallopian tubes, among 

them metaplastic papillary tumor (MPT), an extremely rare lesion 

with indolent behavior showing morphologic similarities 

with borderline ovarian tumors (BOT). Aim of this study was to 

evaluate molecular similarities between rare tubaric neoplasms 

and ovarian tumors of similar degree of malignancy. 

Methods. A microsatellite analysis was performed in parallel 

on the following serous neoplasms: one tubaric vs 3 ovarian HG 

carcinomas; one MPT vs 2 low-grade ovarian carcinomas and 4 

BOT. Sixteen microsatellites located at 8 chromosomal regions 

frequently involved in ovarian carcinogenesis were PCR-amplified 

from DNA extracted from formalin-fixed, paraffin-embedded 

tumoral and normal tissues; amplimers were analyzed in an 

automated DNA sequencer to reveal allelic losses. 

Results. Tubaric carcinoma showed alterations in 88% of informative 

regions, in particular large deletions at 8p, Xp and 17p/q 

and sporadic losses at 1p, 9p and 10q. Ovarian HG carcinomas 

showed an heterogeneous behavior, with a median of 63% altered 

regions (range 50-71%). The regions deleted in the tubaric carcinoma 

were similarly altered in one or more ovarian carcinomas. 

MPT did not reveal alterations in the investigated regions, as 

did 50% of BOT, while the remaining 50% showed only limited 

losses at X chromosome. 

279 

Conclusions. The frequently shared alterations are in favor of the 

common origin of tubaric and ovarian HG carcinomas, though 

more studies will be needed to further support this hypothesis. 

The molecular similarities shared by MPT and BOT suggest the 

need to correctly recognize and monitor such a rare lesion with 

nuclear atypia, at potential risk of development of local recurrences. 

Oral microhistology: an opportunity for dentists to 

take advantage of an innovative technique that 

provides a first level diagnosis in oral oncology 

1)Navone R. 1)D’Angelo G. 2)Marsico A. 3)Rostan I. 4)Pentenero 

M. 5)Gandolfo S. 

1)Scienze biomediche e oncologia umana, Sez. anatomia patologica, Torino, 

Italia 2), Ospedale koelliker, Torino, Italia 3)Scienze biomediche e oncologia 

umana, Sez. anatomia patologica, Torino, Italia 4)Scienze cliniche 

e biologiche, Sez. medicina e oncologia orale osp. S. Luigi, Orbassano 

(TO), Italia 5)Scienze cliniche e biologiche, Sez. medicina e oncologia 

orale osp. S. Luigi, Orbassano (TO), Italia 

Background. Due to late diagnosis and lack of tests able to identify 

early stage precancerous lesions, squamous carcinoma of the 

oral cavity (OSCC) still has a low survival rate. Potentially malignant 

lesions (PMLs) of the oral cavity are divided into classes 

I and II: the former are those with a manifest clinical suspicion 

and the latter those with an apparently innocent clinical appearance. 

To date, oral cavity OSCC and PMLs have been assessed 

by scalpel biopsy, an invasive method generally indicated only 

for class I lesions. 

Methods. An original less invasive sampling method, using a 

dermatological curette, has given results comparable to the scalpel 

biopsy (Navone R et al.: Oral Potentially Malignant Lesions: 

First Level Microhistological Diagnosis from Tissue Fragments 

Sampled in Liquid-Based Diagnostic Cytology. J Oral Pathol 

Med 2008, 37: 358-363). However, in our research only dental 

experts in specialised centres sampled with this method. As the 

territorial (private practise) dentist is the first to observe an apparently 

innocent lesion, as in class II PMLs, a clinical trial was set 

up with them after a brief training period. Samples were obtained 

according to our instructions with the curette technique by 50 

dentists and histologically treated as normal microbiopsies. 

Results. There were 119 samples, 7 were inadequate (5.8%), 103 

negative (hyperkeratosis, parakeratosis or simple hyperplasia), 7 

low/medium grade dysplasia (OIN 1-2), 1 high grade dysplasia 

(OIN 3) and 1 OSCC. 

In our previous study in specialised centres, there was a 3.6% rate 

of inadequate samples (6/164). Although the results of this field 

trial are slightly higher (5.8%, 7/119), they are still very good. 

as this is a 1 st level test, meaning that it is feasible to entrust the 

sampling to private sectorial dentists on the field. This may well 

be an effective method to assess non-tumoral lesions that require 

only follow-up, whilst the positive lesions are to be sent to the 

specialised centres. Therefore, adopting this methodology, even 

apparently innocent lesions (class II) that, to date, have not been 

considered for biopsy, will be managed adequately by the dentist. 

Are rhabdoid tumor and mucinous carcinoma 

of thyroid gland variants of anaplastic carcinoma? 

A clinicopathological study and molecular analysis 

of two cases 

1)D’Antonio A. 2)Russo R. 3)Caleo A. 4)Orabona P. 5)Addesso 

M. 6)Angrisani B. 7)Liguori G. 8)Angrisani P. 

1)Anatomia Patologica Ed Oncologia, A.U.O. San Giovanni Di Dio E 

Ruggi D’aragona, Salerno, Italia 2)Anatomia Patologica Ed Oncologia, 

A.U.O. San Giovanni Di Dio E Ruggi D’aragona, Salerno, Italia 3)Anatomia 

Patologica Ed Oncologia, A.U.O. San Giovanni Di Dio E Ruggi 

D’aragona, Salerno, Italia 4)Anatomia Patologica, Ospedale San Sebastiano, 

Caserta, Italia 5)Anatomia Patologica, Asl Sa1, Scafati, Italia

280 

6)Anatomia Patologica Ed Oncologia, Università Di Medicina, Roma, 

Italia 7)Anatomia Patologica, Int Pascale, Napoli, Italia 8)Anatomia Patologica 

Ed Oncologia, A.U.O. San Giovanni Di Dio E Ruggi D’aragona, 

Salerno, Italia 

Anaplastic carcinomas of thyroid gland (AC) are histologically 

an heterogeneous group of malignant epithelial neoplasms 

with poor outcome. Some cases of AC are characterized by the 

presence of focal area of differentiated carcinoma (papillary or 

follicular) suggesting a possible origin from “dedifferentiation” 

of a previous carcinoma. We describe two rare type of thyroid 

carcinoma characterized by prevalent mucinous and rabdoid differentiation. 

The first case was composed of nests and sheets of 

malignant epithelial cells associated with extensive extracellular 

mucin that substituted and entrapped the follicular parenchyma 

of the thyroid. Thyroglobulin and focally thyroid transcription 

factor (TTF) 1 were positive. From these findings, we classified 

this tumour as primary mucinous thyroid carcinoma. The second 

case was composed mainly of “rhabdoid” cells with abundant 

cytoplasm and eosinophilic globular inclusions that displaces 

vesicular nuclei with central prominent nucleoli resulting in a 

plasmacytoid appearance. Rhabdoid cells co-expressed vimentin 

and cytokeratins with overexpression of p53 protein but. were 

thyroglobulin and TTF1 negative. No areas of differentiated 

thyroid carcinoma were detected in both cases. We have also 

performed a molecular studies for RET/PTC1-RET/PTC3 fusion 

genes and BRAF mutation. No RET/PTC gene rearrangement and 

BRAF point mutation were identified in the rhabdoid and mucinous 

carcinoma. Despite radiotherapy and chemotherapy, these 

neoplasm rapidly progressed and patients died few months after 

presentation. MC and MRT should be considered as a variant of 

anaplastic carcinoma predominantly or exclusively composed of 

epithelial mucin-producing and rhabdoid cells. The absence of 

RET/PTC gene rearrangement and residual differentiated thyroid 

carcinoma al histological level, may be due to de novo origin of 

tumor and rather of dedifferentiation of a previous carcinoma 

Hepatoid differentiation in ovarian neoplasms: 

a diagnostic dilemma 

1)D’Antonio A. 2)Sparano L. 3)Addesso M. 4)Russo R. 5)Angrisani 

B. 6)Angrisani P. 

1)Anatomia Patologica Ed Oncologia, A.U.O. San Giovanni Di Dio E 

Ruggi D’Aragona, Salerno, Italia 2)Anatomia Patologica, Asl Sa1 Ospedale 

Scarlato, Scafati, Italia 3)Anatomia Patologica, Asl Sa1, Ospedale 

Scarlato,Scafati, Italia 4)Anatomia Patologica Ed Oncologia, A.U.O. San 

Giovanni Di Dio E Ruggi D’Aragona, Salerno, Italia 5)Anatomia Patologica, 

Università Di Medicina, Roma, Italia 6)Anatomia Patologica Ed 

Oncologia, A.U.O. San Giovanni Di Dio E Ruggi D’Aragona, Salerno, 

Italia 

Hepatoid differentiation, characterized by the presence of polygonal 

cells with abundant eosinophilic cytoplasm and prominent nucleoli 

resembling hepatocarcinoma, is a rare and unusual features 

of ovarian neoplasms. Focal hepatoid differentiation occurring in 

22% of cases yolk sac tumors in one series. Rarely, hepatoid cells 

represents the predominant pattern of YST (Hepatoid YST) or 

an exclusive component of epithelial tumor as primary hepatoid 

carcinoma (HC). 

We report two cases of adnexal neoplasms composed prevalently 

of hepatoid cells. 

In one case the patient, a 42-year-old woman with ovarian mass 

(FIGO Stage I) that histologically was a primary hepatoid carcinoma 

concurrent with Sertoli-type. The second case was a YST 

tumor arising in fallopian tube of an elderly woman composed 

mainly of hepatoid cells associated with an endometrioid pattern. 

Both cases were characterized by elevated serum levels of AFP. 

The occurrence of the epithelial neoplasms with hepatoid differentiation 

may represent a problem for of differential diagnosis 

and therapeutic management of these patients. Although no dif- 


ferences in survival have been noted among the different histological 

pattern no sufficient data is currently available regarding 

hepatoid YST and pure EC. Surgery combined with a cisplatinbased 

therapy might be able to improve the survival of patients 

in the majority of cases of YST. Measurement of serum AFP are 

useful to monitor this respond to treatment. 

A diagnosis of ovarian metastasis from hepatocellular carcinoma 

is easy in patients with known primary tumor of liver and should 

be always excluded in case of HC as an hepatoid variant of yolk 

sac tumor. Immunohistochemistry is not useful in these cases. 

However, a combination of clinical and pathological features is 

necessary for a correct diagnosis. 

Major problems in the diagnosis and treatment of 

small (≤ 2 cm) cirrhotic liver nodules 

1)D’Errico AD. 2) Banchelli IB 

1)Unita di Anatomia Patologica Istituto “F. Addarii”, Policlinico S. Orsola 

-Malpighi, Bologna, Italia 

Background. Small nodules (≤ 2 cm) in cirrhotic liver are classified 

as MRN (macro-regenerative-nodule), LGDN and HGDN 

(low and high grade dysplastic nodules) and early HCC (EHCC, 

vaguely nodular and distinctly nodular type). 

The most challenging differential diagnosis is between HGDN 

and EHCC, particularly in biopsy samples. The diagnosis of 

small hepatic nodules is based firstly on the radiological findings: 

The main international clinical guidelines currently state that a 

small cirrhotic nodule showing intense arterial uptake followed 

by washout in the venous phase is diagnosed as HCC. The diagnosis 

of HCC ≤ 2 can be therefore established without a positive 

biopsy if at least two imaging techniques are conclusive. If just 

one imaging technique is conclusive of EHCC the biopsy has to 

be performed. 

Methods. We currently perform a combined morphological and 

immunohistochemical analysis involving immunostaining for 

CD34, Ki67 and Glypican 3 on every biopsy of cirrhotic nodules 

suspicious but not conclusively diagnostic of HCC at imaging. 

Conclusions. Distinction between preneoplastic nodules and 

early HCC has critical implications according to the current 

guidelines regarding HCC in Europe, United States and Japan. 

Dysplastic lesions should be followed by regular imaging studies 

since one third of them will develop a malignant phenotype. Early 

tumors must be treated by curative procedures such as resection, 

transplantation and percutaneous ablation. 

Pathological characterization of wnt-activated 

hepatocellular carcinomas (HCC) as assessed by 

glutamine synthetase (gs)immunostaining 

1)Dal Bello B. 2)Campanini N. 3)Tinelli C. 4)Froio E. 

5)D’ambrosio G. 6)Soliani P. 7)Maestri M. 8)Silini EM. 

1)Anatomia Patologica, Fondazione Irccs Policlinico San Matteo, Pavia, 

Italia 2)Anatomia Patologica, Azienda Ospedaliero-Universitaria Di Parma, 

Parma, Italia 3)Biostatistica, Fondazione Irccs Policlinico San Matteo, 

Pavia, Italia 4)Anatomia Patologica, Arcispedale Santa Maria Nuova, 

Reggio Emilia, Italia 5)Anatomia Patologica, Ircss Multimedica, Milano, 

Italia 6)Chirurgia, Azienda Ospedaliero-Universitaria Di Parma, Parma, 

Italia 7)Chirurgia Epato-Pancreatica, Ircss Fondazione Policlinico San 

Matteo, Pavia, Italia 8)Anatomia Patologica, Azienda Ospedaliero-Universitaria 

Di Parma, Parma, Italia 

Background. HCCs showing wnt pathway activation have been 

identified as a specific subtype by gene expression profiling. 

Wnt-activated HCCs are characterized by beta-catenin (CTNN1) 

mutation, and nuclear CTNN1 or GS immunostaining. Distinct 

morphology and improved survival have been reported, but a 

detailed pathological description is lacking. We systematically 

analysed a large series of resected HCCs stratified by CTNN1/GS 

immunostaining.


Methods. We collected a retrospective series of 161 BCLC early 

stage HCCs resected from 146 patient. Main clinical features 

were: male sex 74%, mean age 66 yrs, anti-HCV 85%, multiple 

nodules 20%, mean diameter 3.0 cm. Specimens were examined 

by standard histology according to a predefined set of variables 

and by CTNN1/GS immunostaining. CTNN1 mutations were investigated 

in 76 nodules by sequencing. 

Results. Sixty-six HCCs (41%) were GS+, 26 of which (16%) 

were also CTNN1+. 

GS+ was significantly associated with: early component 

(p = 0.008); capsulated and mononodular (p = 0.001) lesion; expansile 

growth pattern (p = 0.014); microtrabecular architecture 

(p = 0.007); pseudoacinar structure (p < 0.001); dilated sinusoidal 

pattern (p < 0.001); lack of fibrosis (p = 0.002), steatosis 

(p = 0.004) and inflammation (p = 0.003); small cell (p = 0.003); 

bile staining (p = 0.000); low nuclear grade (p = 0.024); absence 

of ballooning, ialine and cytoplasmic inclusions (p = 0.014). 

The pathology of non-neoplastic liver were similar between 

groups (cirrhosis 72%, chronic hepatitis 87%). There were no 

differences according to age, sex, etiology, number of nodules, 

tumor diameter, presence of satellites and vascular invasion. 

CTNN1 mutations were identified in 13 (17%) HCCs, all were 

GS+ (25%, p = 0.006). Among GS- HCCs, fibrosis and steatosis 

identified two further subgroups with distinct morphology. 

In conclusion, Wnt-activated HCCs show specific pathological 

features that can help to explain differences in biologic behaviour. 

A survey of human papillomavirus (HPV) type 

distribution and multiple infections entering into 

the vaccine era 

1)Dal Bello B. 2)Cesari S. 3)Alberizzi P. 4)Gardella B. 5)Iacobone 

D. 6)Spinillo A. 7)Silini EM. 


Italia 2)Anatomia Patologica, Fondazione Irccs Policlinico San Matteo, 

Pavia, Italia 3)Anatomia Patologica, Fondazione Irccs Policlinico San 

Matteo, Pavia, Italia 4)Ostetricia E Ginecologia, Fondazione Irccspoliclinico 

San Matteo, Pavia, Italia 5)Ostetricia E Ginecologia, Fondazione 

Irccs Policlinico San Matteo, Pavia, Italia 6)Ostetricia E Ginecologia, 

Fondazione Irccs Policlinico San Matteo, Pavia, Italia 7)Anatomia Patologica, 

Azienda Ospedaliero-Universitaria Di Parma, Parma, Italia 

Background. A large proportion of HPV infections is sustained 

by genotypes that are not targeted by currently available multivalent 

vaccines and/or by multiple viral types. The impact of HPV 

type distribution and multiple infections on the potential efficacy 

of current vaccines is controversial. We investigated the type and 

number of HPVs present in women referred to colposcopy in a 

single tertiary institution and we evaluated their clinicopathologic 

correlations. 

Methods. Viral typing was performed by SFP 10 -LIPA on a consecutive 

series of cervical scrapings from 3166 women (mean age 

37yrs, 4.4% HIV+) undergoing colposcopy for abnormal cytology 

over a four years period (2005-2009). 62% of the women had 

targeted and/or cone cervical biopsy. CIN severity was correlated 

with the type and number of HPVs. 

Results. Overall prevalence of HPV-DNA was 70%, 98% in 

CIN1 and 98,6% in CIN≥2; specific HPV types were identified 

in 89% of cases. Twenty-eight different types were detected, 

HPV16 (34%), 31 (20%), 52 (23%) and 51 (15%) being the most 

frequent. Frequencies of HPV-6, 11 and 18 were 17%, 9% and 

12% respectively. Other types were HPV-53 (9%), 33 (6%), 39 

(7%) and 56 (5%). Multiple types were detected in 57.2% of 

women, of whom 41.6% had CIN1 and 48% CIN ≥ 2. Overall, 

multiple infections were diagnosed in 54.1% of CIN1, 78.4% of 

CIN≥2 and 44.5% of negative biopsies (p < 0.001). Infections 

by HPV-16 or 18 occurred in 43.3% of CIN, including 33.9% 

of CIN1 and 56% of CIN ≥ 2. Infections by HPV-6, 11, 16 or 18 

281 

occurred in 55.1% of CIN, including 48.4% of CIN1 and 64.2% 

of CIN ≥ 2. Finally, 44.9% of CIN and 35.8% of CIN ≥ 2 were 

entirely sustained by HPV types that are not targeted by currently 

available multivalent vaccines 

In conclusion, the distribution of HPV types and the risk of CIN 

correlated with multiple viral infections highlight the importance 

of genotyping in the clinical management of women with abnormal 

cytology and point to potential limitations in current vaccine 

strategies. 

Polyoma virus dna integration in extracutaneous 

merkel cell –like carcinoma 

1)De Biase D. 2)Ragazzi M. 3)Asioli S. 4)Eusebi V. 

1)Dipartimento di Ematologia e Scienze Oncologiche “L & A Seragnoli”, 

Sezione di Anatomia Patologica, Ospedale Bellaria, Università di Bologna, 

Bologna, Italy 2)Dipartimento di Ematologia e Scienze Oncologiche 

“L & A Seragnoli”, Sezione di Anatomia Patologica, Ospedale Bellaria, 

Università di Bologna, Bologna, Italy 3) Dipartimento di “Scienze 

biomediche e oncologia umana”, Università di Torino, Torino, Italy 4) 

Dipartimento di Ematologia e Scienze Oncologiche “L & A Seragnoli”, 

Sezione di Anatomia Patologica, Ospedale Bellaria, Università di Bologna, 

Bologna, Italy 

Background. Merkel cell carcinoma (MCC) is a neuroendocrine 

tumour, with typical morphological features. Originally reported 

as primary carcinoma of skin, it has been described in numerous 

other sites such as lymph-nodes, breast and salivary glands. 

Cytogenetic studies have shown trisomy of chromosome 6 in 

about 50% of MCC of both skin and lymph nodes indicating a 

strict similarity of these two forms. Recent molecular studies revealed 

in up to 80% of cases a clonally integrated polyomavirus, 

named Merkel cell polyomavirus (MCPV). It seems that MCPV 

is restricted to MCC as no positivity was found in 74 cases of 

visceral neuroendocrine carcinomas [Am J Surg Pathol. 2009 

Dec;33(12):1771-7]. Aim of the present study was to verify the 

presence of MCPV in MCC of lymph nodes and parotid to further 

investigate similarities and differences between the two groups. 

Methods. Cases of primary MCC studied were: 7 of lymph 

nodes, 2 of parotid, 13 of skin. 13 cases of small cell carcinoma 

(SCC) of lung (11 primaries and 2 brain metastases) were also 

analyzed. Immunohistochemistry for keratin 20, chromogranin, 

synaptophysin and TTF1 was obtained in all cases. Tumour cells 

were microdissected and DNA extracted. Viral DNA was studied 

with PCR assay using primers previously described by Duncavage 

et al. [Mod Pathol. 2009 Apr;22(4):516-21]. The PCR products 

were evaluated in a 3% agarose gel and sequenced. 

Results and conclusions. MCPV was detected in 4 cases of 

MCC primary of lymph node (in 3 cases DNA was not evaluable) 

and in all cases of parotid and cutaneous MCC. Keratin 

20 was positive in all cases of MCC. On the contrary, all cases 

of pulmonary SCC were negative for both MCPV and CK20. It 

appears that cutaneous and extracutaneous MCC share similar 

histological, immunohistochemical and molecular features. This 

is a further evidence that Merkell cell origin is no longer tenable 

as Merkel cells have not been described in lymph nodes and 

parotid glands. 

Kras and braf genotyping in colorectal cancer: 

pyrosequencing approach 

1)De Maglio G. 2)Aprile G. 3)Guarrera G.M. 4)Cernic S. 

5)Mazzer M. 6)Foltran L. 7)Falconieri G. 8)Fasola G. 9)Pizzolitto 

S. 

1)Department of Pathology, University Hospital S.M. della Misericordia, 

Udine, Italy 2)Department of Oncology, University Hospital S.M. 

della Misericordia, Udine, Italy 3)Health Technology Assessment Unit, 

University Hospital S.M. della Misericordia, Udine, Italy 4)Department 

of Pathology, University Hospital S.M. della Misericordia, Udine, Italy 

5)Department of Oncology, University Hospital S.M. della Misericordia,

282 

Udine, Italy 6)Department of Oncology, University Hospital S.M. della 

Misericordia, Udine, Italy 7)Department of Pathology, University Hospital 

S.M. della Misericordia, Udine, Italy 8)Department of Oncology, 

University Hospital S.M. della Misericordia, Udine, Italy 9)Department of 

Pathology, University Hospital S.M. della Misericordia, Udine, Italy 

Background. Oncogenic mutations in KRAS and BRAF genes 

have been reported to be predictive of resistance to anti-EGFR 

monoclonal antibodies. To detect mutations, RealTime-PCR and 

direct sequencing are the most widely used systems. An alternative 

promising method is pyrosequencing, a highly specific and 

sensitive technology. 

Methods. Pyrosequencing with CE-IVD marked kits for KRAS 

and BRAF genotyping was applied on 346 tissues obtained 

from 210 cases of advanced colorectal carcinoma. Tissues were 

routinely processed and macrodissected to obtain samples with 

> 70% of tumor cells. DNA extraction was thereafter accomplished. 

One third of samples consisted in colon, hepatic biopsies 

or tissues with tumoral infiltrates < 6 mm 2 . Anti-EGFR MoAb 

response® (KRAS status) and Anti-EGFR MoAb response® 

(BRAF status) (Diatech, Italy) were used accordingly to manufacturer’s 

instructions. PCR reactions and pyrosequencing assays 

were performed on Rotor-Gene TM 6000 (Corbett Research, Australia) 

and PyroMark TM Q96 ID instrument (Biotage, Sweden), 

respectively. 

Results. KRAS/BRAF mutations were found in 121 (57,6%) 

patients. Mutations were distributed for KRAS within codon 12 

(76, 36,2%), G13D in codon 13 (20, 9,6%), Q61H in codon 61 (5, 

2,4%) and A146T in codon 146 (9, 4,3%). Mutations in codon 12 

were: G12D (26, 34,2%), G12V (24, 31,6%), G12A (13, 17,6%), 

G12S (6, 8,1%), G12C (5, 6,8%), G12F (1 case), G12R (1 case). 

BRAF has mutations in exon 11 (1 case) and V600E in exon 15 

(11, 5,3%). 

Our data confirm the pattern of mutations reported in the literature. 

Pyrosequencing proves to be an appealing technique 

of feasible and effective implementation in routine practice. In 

particular, accurate and fast (2,5 days-turnaround time) KRAS/ 

BRAF status detection even in small endoscopic biopsies or 

limited tumoral infiltrates appears valuable in the perspective of 

patient management. 

Complex rare mutation G12f in KRAS gene. Two 

complementary methods: pyrosequencing and 

allele specific quantitative PCr 

1)De Maglio G. 2)Morandi L. 3)Aprile G. 4)Falconieri G. 

5)De Biase D. 6)Visani M. 7)Guarrera G.M. 8)Fasola G. 

9)Pizzolitto S. 

1)SOC Anatomia Patologica, Az. Ospedaliero-Univ. S.M. della Misericordia, 

Udine, Italia 2)Dipartimento di Ematologia e Scienze Oncologiche, 

Università-ASL Ospedale Bellaria, Bologna, Italia 3)Dipartimento di 

Oncologia, Az. Ospedaliero-Univ. S.M. della Misericordia, Udine, Italia 


Udine, Italia 5)Dipartimento di Ematologia e Sienze Oncologiche, 

Università-ASL Ospedale Bellaria, Bologna, Italia 6)Dipartimento di 

Ematologia e Scienze Oncologiche, Università-ASL Ospedale Bellaria, 

Bologna, Italia 7)Unità per la Valutazione delle Tecnologie Sanitarie, Az. 

Ospedaliero-Univ. S.M. della Misericordia, Udine, Italia 8)Dipartimento 

di Oncologia, Az. Ospedaliero-Univ. S.M. della Misericordia, Udine, Italia 


Udine, Italia 

Background. Among mutations affecting KRAS gene, G12D and 

G12V occur more frequently. G12V was found to have a significant 

effect on failure-free and overall survival. Only few sporadic 

studies report more than one mutation in the same sample. Rare 

complex missense mutation G12F was only reported in 4 cases 

of colorectal cancer, 16 lung cancers, 2 pancreatic cancers and 1 


soft tissue tumor by the Catalog of Somatic Mutations in Cancer 

(http://www.sanger.ac.uk/genetics/CGP/cosmic). 

Methods. An endoscopic colon biopsy from a patient with 

advanced colon cancer was analysed for KRAS mutations by 

pyrosequencing with Anti-EGFR MoAb response® kit (Diatech, 

Italy) and confirmed by conventional direct sequencing. 

Moreover, the sample was tested by locked nucleic acid (LNA)modified 

Allele Specific primers and internal molecular beacon 

probes (ASLNAqPCR), a technique that detects the most frequent 

mutations of KRAS (G12A, G12C, G12D, G12R, G12S, 

G12V, G13D). 

Results. Pyrosequencing revealed a pattern consistent with a 

c.34-35GG > TT mutation. However, this method alone could not 

demonstrate whether the sample has a G12F mutation (resulting 

from a two nucleotides substitution GG > TT) or it was a mix of 

two tumoral cell clones that harbored c.34G > T or c.35G > T 

reflecting two independent G12C and G12V mutations. Direct 

sequencing confirmed pyrosequencing results showing two atypical 

T peaks over G peaks. ASLNAqPCR detected only G12C 

genotype with a balanced ratio between mutant and wild type 

allele. Due to allele specific primer sequence it was possible to 

demonstrate that c.34G > T variant was in cis with c.35G > T, 

giving heterozygote mutation for G12F. 

In conclusion, pyrosequencing could detect with a high sensitivity 

a wide range of KRAS mutations. However it should be coupled 

with other methods like ASLNAqPCR to certainly identify 

complex genotypes such as G12F. 

Helicobacter pylori (HP) Infection rate in Molise 

Carbone A., De Ninno M., Guerriero M. 

Anatomia Patologica, Centro Università Cattolica “Giovanni Paolo II”, 

Campobasso, Italia 

Background. HP infection is related to gastric diseases such as 

erosive gastritis, ulcer, cancer and lymphoma. To date, in clinical 

practice, histological examination of gastric tissue samples 

remains the gold standard test for evaluation of HP presence. This 

test, indeed, presents a low cost-benefit ratio, good specificity and 

sensibility, and allows, at same time, an histological diagnosis. 

We studied the frequency of gastric HP infection in the Molise 

population. 

Methods. A population sample of 3.271 sequentially observed, 

unselected subjects, entered in this study in a 56 months observation 

period. All were at their first endoscopy at the “John Paul II” 

Center in Campobasso. This population sample represents about 

1% of total population resident in Molise in the observation period 

(320.907 subjects). All biopsies were studied at microscope 

after routine H&E staining for diagnosis, and after Giemsa staining 

for HP evaluation. 

Results. Patients showed a median age of 51 years with ages 

normally distributed among decades (range: 14-90). Females 

were more represented than males with 1.955 vs 1.316 subjects 

(60% and 40%, respectively). Interestingly, females resulted 

also significantly younger than males (median values: 50 and 

53, respectively; p = 0.00004). An overall HP+ proportion of 

38% was registered in this population sample with no significant 

difference between females and males (Pearson chi-square test 

for overall frequencies distribution: p = 0.74; ns). Patients with 

positive results were slightly but significantly older than those 

with no evidence of HP at histology (mean age 51.8±14.0 and 

50.4±16.4, respectively; p = 0.016). In the HP+ group, females 

and males presented no significant difference in age (51.4±14.0 

and 52.3±14.1, respectively; p = 0.27, ns). An estimation of the 

incidence of H. pylori in Molise returns an overall incidence of 

2.5l.


lymphangiogenesis in cutaneous melanoma: 

prognostic significance and correlation with 

sentinel lymph node status 

1)R. Del Sordo, 1)G. Bellezza, 1)R. Colella, 1)M.G. Mameli, 

1)M. Giansanti, 2) A. Sidoni, 1)A. Cavaliere 

1)Istituto di Anatomia Patologica, Università di Perugia, Italia; 2)Istituto 

di Anatomia Patologica, Terni, Università di Perugia, Italia 

Background. Tumour-induced lymphangiogenesis is a predictive 

indicator of metastasis to lymph node in cutaneous 

melanoma 1 but the prognostic significance of lymphangiogenic 

factor as vascular endothelial growth factors (VEGF)-C and its 

receptor VEFGR3 is still controversial. The aim of our study is 

to determine the intra and peritumoral lymphatic vessel density 

(LVD), the expression of VEGF-C and VEGFR-3 and correlate 

the findings with sentinel lymph node (SLN) status and clinicopathological 

data. 

Materials. 22 cases of cutaneous melanoma with metastasis to 

SLN (SLN+) were enrolled and matched with a group of 22 

cases without SLN metastasis (SLN-). Lymphatic vessels and 

lymphangiogenic factor were detected by immuhistochemistry 

using D2-40 (clone D2-40), VEGF-C (clone C-1) and VEGFR3 

(clone KLT9) antibodies. LVD was defined as the number of 

vessels/mm 2 . The staining was analyzed semiquantitatively using 

a scoring system which considered the intensity (grades 0-3) 

and extent (0: negative; 1:≤33%; 2: 34-66%;; 3: 67-100%). The 

results obtained were multiplying together: points 0, 1, 2-low 

staining, 3, 4, 6, 9-high staining. 

Results. Intratumoral LVD was higher in melanomas with SLN+ 

(p = 0.038) and was significantly associated with nodular melanoma 

(p = 0.008), Clark’s level IV-V (p = 0.003), lymphocytic 

infiltrate (non brisk vs brisk; p = 0.03), type of SLN metastasis 

(macrometastasis vs isolated cells; p = 0.052) and location of metastasis 

(central vs subcapsular; p = 0.013). VEGF-C and VEGR3 

were equally detected in SLN- and SLN+ cases. No correlations 

was found between peritumoral LVD, VEGF-C, VEGFR3 and 

clinico-pathological data. 

In conclusion our finding suggest that high intratumoral LVD is 

associated with SLN metastasis but overexpression of VEGF-C 

and VEGFR-3 seem to be not predict SLN status. 

references 

1 Massi D et al. J Clin Pathol 2006;59:166-73. 

Mir-205 expression levels in non-small 

cell lung cancer do not always distinguish 

adenocarcinomas from squamous cell carcinomas 

1)Del Vescovo DR. 2)Cantaloni DR. 3)Bragantini DR. 4)Morelli 

DR. 5)Silvestri DR. 6)Fasanella DR. 7)Cuorvo DR. 8)Dalla 

palma PROF. 9)Barbareschi DR. 

1)Cibio, Università Trento, Trento, Italia 2)Anatomia Patologica, Ospedale 

Di Trento, Trento, Italia 3)Anatomia Patologica, Ospedale Di Trento, 

Trento, Italia 4)Anatomia Patologica, Ospedale S.Chiara Di Trento, 

Trento, Italia 5)Chirurgia B, Ospedale S.Chiara Di Trento, Trento, Italia 

6)Anatomia Patologica, Ospedale S.Chiara Di Trento, Trento, Italia 




Background. Classification and therapy of lung tumours is 

based on precise histological diagnosis. Recent data suggest that 

quantification of micro RNA expression may distinguish adenocarcinoma 

(ADC) from squamous carcinoma (SQC). Aim of the 

present study is to analyse a series of well characterized lung 

tumours using real time PCR to quantify the expression of mir 

205 in relation to histotype. 

Materials and methods. 40 formalin-fixed and paraffin-embedded 

consecutive lung carcinomas (20 ADC and 20 SQC) 

283 

were analyzed. Quantification of microRNA expression was 

carried out using TaqMan MicroRNA Assay kits according to 

Applied Biosystems protocol. Hsa-miR-205, hsa-miR-21, and 

U6snRNA were measured by qRT-PCR in triplicate. Normalized 

threshold cycle data (Ct) of hsa-miR-205 and hsa-miR-21 were 

calculated by subtracting AvgCt U6 from AvgCt mir205 or AvgCt mir21, 

respectively, Ct 205 = AvgCt mir205 - AvgCt U6 and Ct 21 = AvgCt mir21 

– AvgCt U6. Sample score was then obtained using the formula 

Score ≡ AvgCt mir205 - [(AvgCt mir21 + AvgCt U6)/ 2] = Ct 205 - (Ct 21/2), 

according to Lebanony et 2009. 

Results. The relative level of miR-205 was generally lower in AC 

as compared with SQC. Most SQC showed high levels of miR- 

205 and their sample score was below the proposed cut-off value 

of 1.5 to classify tumors in AC and SQC. Interestingly 4 out of 

20 ADC would have been classified as SQC and 3 out of 20 SQC 

would have been misclassified as ADC. 

Discussion. Our present results, although confirming that miR205 

expression levels are generally different in SQC as compared 

with ADC, clearly show that this approach may still misclassify a 

non negligible (7 out of 40, 17,5%) number of cases. The discrepancies 

between our study and the one of Lebanony et al. 2009 and 

Bishop et al. 2010 may be related to technical differences, tumor 

heterogeneity and tissue preservation, and underscore the need 

for an integrated diagnostic approach to lung tumors. 

Managment of histopathology laboratory in 

Africa: the St. raphael St francis & Nsambya 

Hospital experience 

1)Dell’Antonio G. 2)Colantoni A. 3)Bonanno E. 4)Othieno E. 

5)Aloi F.S. 

1)Anatomia Patologica, San Raffaele, Milano, Italia 2)Anatomia Patologica, 

Policlinico Universitario Tor Vergata, Roma, Italia 3)Anatomia Patologica, 

Policlinico Universitario Tor Vergata, Roma, Italia 4)Anatomic 

Pathology, Nsambya Hospital, Kampala, Uganda 5)Aispo, Nsambya Hospital, 

Kampala, Uganda 

Background. Nsambya Hospital in Kampala (Uganda), accredited 

by the Uganda Catholic Medical, is a tertiary child-maternal 

referral hospital with a capacity of 361 beds which has played a 

pioneering role in HIV/AIDS activities. It is involved in patient 

care, research and teaching for graduates of any of Uganda’s four 

medical schools. Here they spend a year of internship in Surgery, 

Internal Medicine, Pediatrics and Obstetrics-Gynecology under 

the supervision of local specialists and consultants. 

A modern histopathology laboratory (HL) has special challenges 

because prevention, diagnosis and clinical practice relies on morphological 

and qualitative (biomarkers) characteristic of pathological 

tissues and more and more therapeutics decisions are based on 

specific immunostainings (IHC) (i.e.hormonal receptors). 

Methods. In Nsambya HL are mainly processed cytologic samples 

(PAP test and fine needle aspirates), biopsies and surgical 

specimen from gynecologic pathologies. The human resources 

are a pathologist and two technologists with expertise in cyto/ 

histopathology. Existing procedures have been reviewed and formalized 

as guide lines, some new procedures, such as “thin layer 

cytology”, histochemistry and IHC have been introduced. 

Results. In the first five months of 2010 in the HL were performed 

about 500 PAP test, 30 fine needle aspirations, 680 histological 

specimens some of which combined with IHC improving both the 

number and the quality of specimens examined. In march 2010 

AISPO, with APOF and Tor Vergata University counselling, 

has opened a new HL with western standards and machineries. 

The most critical issues are technicians’ training made inside the 

lab, the written protocols approved by the pathologist to ensure 

the continuity and a quality control in sample preparation and 

diagnosis. We think that telepathology with internet broad band, 

as demonstrated by other experiences done by APOF, will be the 

best solution to these problems.

284 

Aberrant S-100 protein expression in metanephric 

stromal tumour: report of a case 

1)A, Gurrera, 2)A. Di Cataldo, 1)G. Leone, 3)V. Di Benedetto, 

1)E. Vasquez, 1)G. Magro 



Catania, Italia; 2)Dipartimento Ematologia e Oncologia Pediatrica, 

Azienda Ospedaliero Universitaria Policlinico-Vittorio Emanuele, Catania, 

Italia; 3)Dipartimento Chirurgia Pediatrica, Azienda Ospedaliero- 

Universitaria Policlinico-Vittorio Emanuele, Catania, Italia 

Abstract. Metanephric stromal tumour (MST) is a rare pediatric 

renal-specific neoplasm preferentially occurring in children. It is 

currently included in the spectrum of metanephric tumour along 

with metanephric adenoma and metanephric adenofibroma of 

which it is regarded as the pure stromal variant. 

Materials. We herein report a case of MST occurring in a 9-yearold 

boy who complained recurrent pain in his right flank. TC 

showed a mass in the lower pole of kidney 

Results. Grossly, tumour presented as a 5.5.cm well-demarcated, 

fibrous nodule, with a solid-microcystic appearance. Histological 

examination revealed an unencapsulated tumour composed of 

bland-looking ovoid- to spindle- to stellate-shaped cells, embedded 

in an abundant fibrous, frequently hyalinized, stroma. Alternating 

hypercellular and hypocellular areas imparted tumour a 

vague nodular low-power appearance. Notably, entrapped native 

kidney ducts, focally surrounded by onion-skin collarettes and 

angiodysplasia of renal arterioles were scattered throughout the 

tumour. Heterologous glial or cartilage components were lacking. 

Immunohistochemically, neoplastic cells showed diffuse immunoreactivity 

for vimentin, CD34 and, surprisingly, for S-100 protein. 

A focal staining was also obtained with CD99 and α-smooth 

muscle actin. Pancytokeratins, WT1, bcl-2, EMA, GFAP and 

CD117 were all negative. Morphological and immunohistochemical 

features were consistent with the diagnosis of “MST, fibrous 

variant, with aberrant S-100 protein expression”. Although S-100 

protein stains the glial and/or cartilaginous components that may 

be occasionally encountered in MST, this marker has not been 

previously reported in the fibroblastic component. Pathologist 

should be aware of this possibility to avoid potential confusion 

with other benign or malignant S-100 protein tumours. 

Difficulty diagnosis of a glucagonomas on biopsy 

1)Di Clemente D. 2)Castorani L. 3)Sabbà C. 4)Gentile A. 


2)Dip. Med. Interna. E Med. Pubblica, Policlinico Di Bari, Bari, Italia 

3)Dip. Med. Interna. E Med. Pubblica, Policlinico Di Bari, Bari, Italia 


Background. The glucagonoma is a rare well-differentiated pancreatic 

endocrine neoplasm (PEN), associated with clinical manifestations 

of inappropriate secretion of glucagon, as necrolytic 

migratory erythema, stomatitis, diabetes, weight loss and anemia. 

Methods. 47 years old woman was hospitalized for cachectic, 

with a history of diabetes mellitus and recurrent infections for 

desquamative lesions in the skin of the face and lower limbs and 

oral and anal mucosa. Acknowledgement biochemical investigations 

of normochromic normocytic anemia, hypoproteinemia, 

hypocalcemia, insulin, hyperglycemia, increase in HbA1c, c-peptide, 

indices of inflammation. CT abdomen: increased volume of 

the pancreas. Biopsied pancreatic of two sclerohyalinosis cores 

and minute neoplastic aggregate consists of epithelial cells with 

large cytoplasm and small nuclei. The morphology and immunohistochemistry 

(synaptophysin, CD56, CAM 5.2 and glucagon: 

positive, chromogranin: low positive, Ki67: 10%, TTF1: 

negative) supported the diagnosis of neuroendocrine tumor of the 

pancreas, consistent with the clinical diagnosis of glucagonoma, 

subsequently upheld by the dose of glucagon (49 600 ng/l). The 


patient died after 4 months by debulking surgery to metastases in 

vital organs. 

Conclusions. The glucagonoma diagnosed in time may be susceptible 

to treatment with good prognosis, but the diagnostic 

delay represents a risk to the patient. Furthermore, glucagon may 

be sought with the IIC, but is less intensely expressed than other 

hormones expressed by the PENS and although were detected reactive 

peptides derived from proglucagone (glicentin and peptide 

1 and glucagon-like 2) antibodies against these proteins are not 

used and available usually. 

Acinar cell carcinoma of the pancreas in children 

1)Di Clemente D. 2)Palumbo M. 3)Alaggio R. 4)Santoro N. 

5)Gentile A. 


2)Dipartimento Di Anatomia Patologica, Policlinico Di Bari, Bari, 

Italia 3)Ass. Italiana Di Ematologia E Oncologia Pediatrica, Comitato 

Anatomo-Patologico, Padova, Italia 4)Pediatria Generale E Specialistica, 

Policlinico Di Bari, Bari, Italia 5) Dipartimento Di Anatomia Patologica, 

Policlinico Di Bari, Bari, Italia 

Background. The pancreatic acinar cell carcinoma is a malignant 

epithelial neoplasm with exocrine enzyme production by neoplastic 

cells. Represents 1-2% of pancreatic tumors of the adult and 

15% of pancreatic tumors in childhood and is associated with 

increased of α-fetal protein and hypersecretion of trypsin and 

lipase, responsible for the necrosis in the subcutaneous fat and 

polyarthralgias and polyarthritis 

Methods. Child of 6 years was admitted for swelling of the face and 

hyperchromasia skin at the joints. Investigations biochemical detection 

of α-fetal protein high (6644 ng/ml). CT abdomen: wide ring 

neoformation around the duodenum of pancreatic origin. Biopsied, 

the four fragments were part of a carcinoma histologically composed 

of solid nests epiteliomorfi elements with abundant eosinophilic 

cytoplasm and presence of PAS positive granules, the polar 

nucleus with prominent nucleolus and dispersed the chromatin, cell 

aggregates and large foamy histiocytic,neoplastic permeation endolymphatic. 

The histology and investigations IIC (α-1-antitrypsin, 

C8, CK 18, CKAE1-AE3: positive; vimentin, CD10, CD56, chromogranin, 

synaptophysin, NSE, PgR, CK19: negative; Ki67: 30% 

α-fetus-protein positive in rare and isolated cells) did give evidence 

for the diagnosis of pancreatic exocrine acinar carcinoma. The histological 

slides were reviewed (AIEOP) and the positive exclusive 

feedback of the membrane of β-catenin confirmed our diagnosis. 

Child was subsequently subjected to surgical resection of the mass, 

did not have cancer treatments and currently enjoys good health. 

Conclusions. We report this case for its rarity and the importance 

of accurate differential diagnosis with well differentiated PEN not 

working (which is absent intracytoplasmic PAS positivity), the 

pancreatoblastoma, the solid tumor pseudopappilare (expressing 

positive nuclear β-catenin). 

Metaplastic chondrosarcomatoid breast cancer 

1)Di Clemente D. 2)Palumbo M. 3)Fiore G. 4)Ingravallo G. 

5)D’Eredità G. 6)Giardina C. 





5)Dipartimento Di Chirurgia Generale E Speciale, Policlinico Di Bari, 

Bari, Italia 6)Dipartimento Di Anatomia Patologica, Policlinico Di Bari, 

Bari, Italia 

Introduction. Metaplastic breast carcinoma is a rare and heterogeneous 

neoplasm often showing areas of mesenchymal differentiation 

type spindle cells, myxoid, chondroid or osseous. 

Methods. 62 year old woman presenting a lump in the SE quadrant 

of her left breast, with a prewious positive FNAC (C5), was


hospitalized and underwent mastectomy and axillary dissection. 

The tumour had a diameter of up to cm. 3.3, browning colour and 

regular contours. Histologically the tumour showed lobulated 

contours and nodular growing pattern with poorly differentiated 

cancerous cell proliferation peripherally and chondrosarcomatous 

proliferation centrally with a smooth transition between the two 

parts. Myxoid areas were also present. 

An high number of mitosis was evident. Peripherally there were 

also areas of ductal carcinoma in situ of solid type. No vascular 

invasion was detected around the tumour.The axillary lymph 

nodes resulted metastases free. Investigations IIC: Estrogen and 

progesterone receptors were totally: negative, proliferative activity 

tested by Ki67 antigen was very high, (80%) and HER2/neu 

resulted “1 +”. 

CK CAM5.2, CK34 β E12 and CK pool: resulted strongly expressed 

in the epithelial component, while P63 positive only in 

rare myoepithelial cells. 

Conclusions. The diffuse positivity for CK and especially for 

CK34 E12 β argue in favor of tumor histogenesis from basal 

cells with broad differentiation spectrum. Negativity for axillary 

lymph node metastases along with the high proliferative activity 

suggests a biological behaviour closer to a sarcomatous than a 

carcinomatous tumour. 

The vascular microdensity in prostate cancer 

1)Di Cristofano C. 2)Biolcati M. 3)Leopizzi M. 4)Miraglia 

A. 5)Sardella B. 6)Marangi G. 7)Chiappetta C. 8)Petrozza V. 

9)Laghi A. 10)Della rocca C. 

1)Department of Experimental Medicine, Sapienza University of Rome, 

Polo Pontino, I.C.O.T, Rome, Italy 2)Department of Experimental Medicine, 

Sapienza University of Rome, Polo Pontino, I.C.O.T, Rome, Italy 3)Department 

of Experimental Medicine, Sapienza University of Rome, Polo 

Pontino, I.C.O.T, Rome, Italy 4)Department of Experimental Medicine, 








of Radiological Sciences, Sapienza University of Rome, Polo 


Sapienza University of Rome 

Introduction. Neoangiogenesis involving the growth of new 

vessels in the microcirculation and in this environment the tumor 

cells can proliferate. Morphological aspects of neoagiogenesis is 

linked to raising of microvascular density (MVD). The increased 

expression of markers for blood vessels and the presence of proangiogenic 

factors were observed in many tumors and they are 

often correlated with a worse prognosis, and these factors are 

considered the target of anti-angiogenic therapies. In prostate 

cancer (PC), some studies have shown that MVD is increased in 

PC areas and it’s correlated with disease progression. 

Computed tomography (CT) perfusion is a noninvasive tool for 

the analysis of some quantitative indices of tissue perfusion such 

as tissue blood volume (BV), tissue blood flow (BF), mean transit 

time (MTT) and permeable surface tissue (PS) which could be 

associated to MDV in PC. 

The aim of this study was to evaluate MDV in PC using immunohistochemical 

analysis (IHC) and compare it with the flow 

parameters of CT perfusion. 

Materials and Methods. We were selected 10 patients with PC 

who underwent a CT with perfusion for preoperative clinical 

staging of the tumor and then a radical prostatectomy. We prepared 

serial macrosection of the entire prostate that correspond 

to the levels made during the TC perfusion. MDV was evaluated 

by IHC using Ab anti-CD34 (clone QBEnd/10, Leica) and it was 

quantified by an automatic image analyzer (D-Sight, Menarini). 

285 

The IHC results were compared with parameters identified by CT 

perfusion analyzing 333 total areas. 

Results. In our study emerged a statistical correlation between 

MVD and perfusion parameters CT (PS;BV;BF) (

Moreover, we found an increase of both MVD and parameters of 

perfusion CT in adenocarcinoma compared to prostate tissue and 

the MTT is resulted to be statistically associated to adenocarcinoma 

(p = 0,002). 

Immunohistochemistry expression of TNf-alpha 

and TNf-r2 before and after treatment with 

biological drugs in psoriatic lesions 

1)Di Cristofano C. 2)Cacciotti J. 3)Leopizzi M. 4)Chiapetta C. 

5)Miraglia A. 6)Sardella B. 7)Marangi G. 8)Petrozza V. 9)Potenza 

C. 10)Della rocca C. 

1)Department of Experimental Medicine, Sapienza University of Rome, 

Polo Pontino, I.C.O.T, Rome, Italy 2)Department of Experimental Medicine, 











of Dermatology, Sapienza University of Rome, Polo Pontino, 

Rome, Italy 10)Department of Experimental Medicine, Sapienza University 

of Rome, Polo Pontino, I.C.O.T, 

Introduction. Psoriasis is a chronic skin disorder with multifactorial 

etiology, affecting 3% of the general population and it is 

highly disabling in its severe form. 

Histopatologically plaque psoriasis can be classified into three 

phases that probably reflect the stages of disease pathogenesis: 

initial stage with unclear and fleeting histopathological features, 

steady stage with early and late forms, and regressive stage. It’s 

been shown that high levels of TNF-alpha are involving in the 

development, proliferation and maintenance of psoriasis plaques 

and for this reason modern biological therapy considered TNFalpha 

a molecular target for treatment of psoriasis using TNF-alpha 

antagonists (Etanercept) or by the use of monoclonal antibody 

(Efalizumab). 

The aim of this study was to observe the clinic and histology of 

plaque psoriasis in patients before and after treatment with Etanercept 

evaluating the immunohistochemistry (IHC) expression 

and localization of TNF alpha and of its receptor TNF-R2(p75). 

Materials and Methods. We were selected 16 patients with 

histopathologic diagnosis of plaque psoriasis treated with Etanercept. 

We were performed biopsies of psoriatic plaques pre- and 

post-treatment and of healthy skin. For ICH analysis we were 

used antibodies anti-TNFalpha (clone 52B83, Hycult biotech.) 

and anti-TNFR2(p75) (Santa Cuz biotech.). 

Results. TNF-alpha and TNF-R2 receptor (p75) expression 

change during the development of plaque psoriasis, however this 

expression does not seem associated with plaque progression, in 

the transition between early and late phase. Etanercept modulates 

the expression of two biological markers and particularly the 

expression of TNF-R2 receptor (p75). 

endosonography guided fine needle and lesion 

intramural very difficultie: how adequacy increase 

M. Di Maso, V. Nirchio*, N. Muscatiello, C. Panella, E. Ierardi, 

Gastroenterology Unity Univ., Ospedali Riuniti, Foggia, Italy; *U.O.S 

Cytopathology, departments of Pathology, Ospedali Riuniti, Foggia, Italy 

Background. Endoscopic ultrasound (EUS)-guided fine needle 

(FN) has increease the diagnosis capability of endosonogra-

286 

phy white respect to lesion intamural to gastroentestinal tract. 

Limitation of this technique are that it is often difficult to obtain 

adequate tissu for diagnosis. We have examined the use of a 22 

Gauge needle and use 19 Q.C.needle Wilson Cook. 

Aim. To evalutate adequate tissue obtained the use needle in patients 

whit intramural masses in the gastrointestinal tract. 

Methods. 96 patients underwent we have examined fine needle 

the use of a 22 G and 19 Q.C. which obtained both cytology ande 

core biopsy specimen. Lesion biopsied include intramural lesion 

from esophagus (31), stomach (46), rectal (19). Number of FN 

passes / patients range from 3 to 9 the use of a 22 G. Number of 

F.N. passes/patients it is 1 the use of a 19 Q.C. 

Results. Overall adequate tissue the use of a 21 needle it is 66 

patient and 19 needle Q.C. 81 patients. 

NEEDLE Adequacy 

22 g eus-n 3 68,75% 

19 Q.c. 84,37% 

Complication were minor and included 2 bleeding and 1 addominal 

pain. 

Conclusions. Needle 19 G Q.C a good yield of obtaining core 

biopsies as well as cytology specimens 

eBV-positive mucocutaneous ulcer of the large 

intestine: report of a case associated with 

diverticulosis 

A. Di Napoli, M. Giubettini, S. Scarpino, A. Ferrari*, S. Uccini, 

and L. Ruco 

Dipartimento di Medicina Clinica e Molecolare, UOC di Anatomia Patologica 

e UOC di Ematologia*, II Facoltà di Medicina, Ospedale Sant’Andrea, 

Università “La Sapienza” Roma 

Background. In a recent publication Dojcinov et al. (1) described 

a previously unrecognized lymphoproliferative disorder 

associated with EBV infection. They reported that 26 patients, 

who underwent iatrogenic immunosuppression (9 cases) or immunosenescence 

(17 cases, age > 65), developed isolated, sharply 

circumscribed ulcers involving oropharyngeal mucosa (16 cases), 

skin (6 cases) and gastrointestinal tract (4 cases). Lesions were 

histologically characterized by a polymorphous infiltrate and 

atypical large B cell blasts with Hodgkin/Reed-Sternberg-like 

morphology. The atypical B cells showed strong staining for 

EBER, CD30 and CD15, reduced expression of CD20, and were 

sited in a background of CD3+ reactive T cells. Polymerase chain 

reaction revealed 39% clonal Ig rearrangements and 31% clonal 

and restricted T-cell patterns; a monoclonal Ig rearrangement 

was accompanied by a restricted T cell response in 3 cases. The 

disease showed a self-limited, indolent clinical course in most of 

the patients and 45% of the cases regressed spontaneously with 

no treatment. 

In the present report we describe a case of EBV+ mucocutaneous 

ulcer occurring in the large intestine of a 81-years-old female. 

The clinicopathologic and molecular features observed in our 

case are slightly different from those previously reported, thus 

contributing to a further characterization of this newly recognized 

entity. 

Methods. Clinical history. A 81-years-old female patient was 

admitted at the emergency room of the Sant’Andrea Hospital of 

Rome because of abdominal pain due to intestinal perforation, 

probably related to diverticulosis. In the clinical history the patient 

had diabetes, vascular hypertension, ischemic cardiopathy, 

a previous cerebral infarction, and an autoimmune thrombocytopenia 

since 2006; this latter was treated with steroids and azathioprine 

for one year with no response; more recently the patient 

received Romiplostin obtaining a complete response. The patient 

underwent an intervention of left colectomy and the surgical 

specimen was sent to the Pathology Lab. The patient did not re- 


cover completely after the surgical intervention, and in the postoperative 

period suffered of progressive liver dysfunction. She 

died one month after the intervention; an autopsy was performed. 

Immunohistochemistry. Immunostaining on paraffin sections was 

done for CD3, CD4, CD8, CD20, CD79a, CD30, CD15, CD45, 

CD10, BCL-2, BCL-6, MUM-1, CD34, CD117 and LMP-1 

with antigen retrieval and antibody dilutions per manufacturer’s 

recommendation. The immune reaction product was devoleped 

using LSAB-2 plus, Dako, Denmark. In situ hybridization. In situ 

hybridization for EBV-encoded RNA (EBER) was conducted on 

formalin-fixed paraffin-embedded sections using Epstein-Barr 

Virus (EBER) PNA Probe/Fluorescein,and FITC/HRP (Dako, 

Denmark). Polymerase Chain Reaction (PCR) for immunoglobulin 

(IG) and T-cell receptor (TCR) gene rearrangements. 

Total DNA was extracted from paraffin tissue sections with a 

lysis buffer containing proteinase K. Molecular evaluation of 

T cell receptor gamma (TCR gamma) gene rearrangement was 

performed by PCR amplification (Polimerase Chain Reaction) 

of DNA fragments corresponding to the VJ segment of hypervariable 

region of the TCR range. For the amplification reaction 

was used “kit for Molecular Diagnosis Analysis of clonal 

rearrangements of TCR-gamma (Master Diagnostica, Spain). 

Molecular evaluation of immunoglobulin heavy chain (IgH) gene 

rearrangements was performed by PCR amplification of DNA 

fragment corresponding to the VD-JH segment of hyper-variable 

region of the IgH. Results of TCR-gamma and IgH gene rearrangements 

were analysed according to the BIOMED-2 report 

(Leukemia, 2003, 17:2257-2317) with 3100 Genetic Analyzer 

(Applied Biosystem). 

Results. Gross pathology. The sigmoid colon (length 9.3 cm) 

was involved by two distinct types of lesions: multiple diverticula 

and a 6 cm wide ulceration of the mucosa surface, with 

transmural penetration of the lesion and perforation of the visceral 

serosa. Seven mesenteric lymph nodes (diameter of the 

largest 0.5 cm) were detected on gross inspection. Histology. The 

ulceration corresponded to a wide shallow area of mucosal dysepithelisation 

associated with multiple fissures deeply penetrating 

through the intestinal wall. A transmural inflammatory-like 

reaction was associated with intestinal fissures and was present 

in the perivisceral adipose tissue; acute serositis was observed in 

correspondence of the serosal perforation. The inflammatory-like 

infiltrate was polymorphous, and was composed mainly of mature 

CD3+ T lymphocytes with CD4+ cells largely prevailing on 

CD8+ cells; some CD20+/CD79a+ B cell nodules were sparsely 

present; numerous polyclonal CD138+ plasma cells and CD68+ 

histiocytes were admixed with lymphocytes. The inflammatory 

background was associated with scattered large pleomorphic 

cells, reminiscent of Hodgkin and Reed-Sternberg cells (H/RS); 

these latter were CD30+/CD15+/MUM-1+, were negative for 

CD3, CD4, CD8, CD20, CD79a, CD10, bcl-6, CD45, CD34, 

CD68 and CD117, and were infected by EBV as suggested by 

positive staining for EBER and LMP-1. The seven regional 

lymph nodes were completely effaced and showed a similar 

histological picture. Some of the intestinal diverticula were associated 

with the lymphoproliferative disease. PCR performed 

on DNA extracted from paraffin blocks of the intestine revealed 

the concomitant presence of a monoclonal IG rearrangement and 

of a monoclonal TCRg rearrangement. Autopsy findings. The 

patient died one month after the surgical intervention and an 

autopsy was performed. The ultimate cause of death was liver 

failure due to diffuse involvement of the portal spaces by B cell 

lymphoma; moreover, multiple nodular areas of coagulative 

necrosis were found; some lymphomatous angiocentric lesions, 

could be demonstrated. Spleen, and aortic lymph nodes were also 

involved by the disease; no evidence of bone marrow involvement 

was found. 

Discussion. In the present report we describe a case of EBV+ 

mucocutaneous ulcer of the large intestine occurring in a 81


years-old female patient. It seems likely that the advanced age of 

the patient, and perhaps the pharmacological treatment that she 

received for the autoimmune thrombocytopenia may have contributed 

to weaken her immunocompetence status thus favouring 

EBV lymphomagenesis. 

The histological, immunohistochemical and molecular features 

of our case are closely similar to those described by Dojcinov 

et al. (1). In fact, the presence of CD15+/CD30+ H/RS-like cells 

associated with an inflammatory-like background was highly 

suggestive for a diagnosis of Hodgkin’s lymphoma; however, 

the observation of a clonal IgH rearrangement and the recent description 

of EBV+ mucocutaneous ulcer allowed us to establish 

the correct diagnosis. A number of untreated cases reported by 

Dojcinov et al. 1 exhibited either spontaneous regression, or had 

persistent but limited disease confined to the skin or mucosa; 

moreover, the histological picture was often characterized by lesions 

well-demarcated from the surrounding structures. All these 

evidence contributed to suggest that EBV+ mucocutaneous ulcer 

is a disease characterized by an indolent behaviour and by a self 

limited clinical course. The behaviour of our case was much more 

aggressive, ultimately resulting in the death of the patient due to 

hepatic failure caused by extensive liver infiltration. In fact, at autopsy 

evidence of the disease was found in the regional and aortic 

lymph nodes, in the spleen, and diffusely in the liver; moreover, 

the lesion were not histologically well-demarcated, but showed 

features of an infiltrative malignant disease. 

To provide an explanation for the localized mucocutaneous manifestation 

of the disease it was speculated that a minor mucosal 

irritation or a mucocutaneous injury might be responsible for a 

lowered local resistance favouring proliferation of EBV-infected 

cells. In the series of Dojcinov et al. 1 three cases developed in 

the intestine; a colonic mass in a 69/F with rheumatoid arthritis; 

an anorectal ulcer in a 78/M with a long history of ulcerative 

colitis; a small ulcer at the rectosigmoid junction in 64/F with a 

history of essential thrombocytemia. In our case we have found 

a close spatial association between mucocutaneous ulcer and diverticula 

in the sigma. Thus, it cannot be ruled out that the local 

irritation due to the diverticular disease represented the trigger 

that favoured development of EBV+ mucocutaneous ulcer at 

that site. 

Finally, concomitant TCR and IG gene rearrangements were 

found by Dojcinov et al. in 3 of the 18 investigated cases; IG 

clonal rearrangement in 7 cases, and TCR gene rearrangement 

in 6 cases. It was speculated that the monoclonal IG rearrangements 

were indicative of the clonality of the EBV-driven B cell 

proliferation, and that the TCR gene rearrangements were indicative 

of a “restricted” T cell response; this latter might reflect the 

prevalence of oligoclonal T cell responses in elderly subjects 

that are markedly restricted and deficient in their epitope specific 

repertoire, rendering the host at increased risk of infection. In the 

DNA extracted from our case clonal IG and TCR-gamma rearrangements 

could both be demonstrated; the finding might be 

consistent with a clonal EBV+ B cell disease associated with a 

“restricted” T cell response, as previously proposed. In alternative, 

the possibility that concomitant IG and TCR-gamma gene 

rearrangements occurred in the neoplastic cell clone should also 

be considered. As a matter of fact this would not be unique to 

EBV+ mucocutaneous ulcer since concomitant IG and TCR rearrangements 

were already demonstrated in a consistent number of 

cases of precursor-B-ALL 2 . 

references 

1 Dojcinov, et al. EBV positive mucocutaneous ulcer – A study of 26 

cases associated with various sources of immunosuppression. Am J 

Surg Pathol 2010;34:405-17. 

2 van der Velden VH, et al. TCRB gene rearrangements in childhood 

and adult precursor-B-ALL: frequency, applicability as MRD- 

PCR target, and stability between diagnosis and relapse. Leukemia 

2004;18:1971-80. 

287 

fast fISH protocol for Her-2/neu analysis in breast 

cancer 

1)Di Oto E. 2)Pession A. 3)Tallini G. 

1)Sez. di anatomia, istologia e citologia patologica, Bellaria, Bologna, 

Italia 2)Sez. di anatomia, istologia e citologia patologica, Bellaria, Bologna, 

Italia 3)Sez. di anatomia, istologia e citologia patologica, Bellaria, 

Bologna, Italia 

Background. Techniques to evaluate HER-2/neu status in breast 

cancer include IHC and FISH. FISH is an established, reproducible, 

and extremely reliable method for detecting HER-2/neu 

gene status on archival paraffin blocks. It has long been known 

that HER-2/neu gene amplification determined by FISH is an 

independent predictor of outcome in breast cancer patients and it 

is the prerequisite to treat patients with Herceptin. We routinely 

perform FISH in cases that are undetermined after IHC evaluation 

(2+ score, 2007 ASCO/CAP HER-2/neu guidelines) with an 

average turnaround time of 7 days. To decrease this time we have 

validated a faster hybridization protocol. 

Methods. We evaluated different pre-treatment procedures and 

hybridization times in a series of 20 breast cancers previously 

analyzed with the standard method that includes overnight hybridization. 

Of the 20 cases, 5 were scored as HER-2/neu amplified, 

7 as having chromosome 17 polysomy without HER-2/neu 

amplification, and 8 as being non-amplified and diploid for 

chromosome 17. Short hybridization between 1 h and 4 h and different 

heat pre-treatment times were assessed. Signal counts and 

intensity with the modified procedures were compared for each 

case with the standard method. The Chi Square test was used for 

analysis of signal counts ratio. 

Results. Decreasing hybridisation time while slightly increasing 

heat pre-treatment time gave satisfactory results in terms of 

intensity, counts and overall signal quality, with no change in 

HER-2/neu reporting. The shortest hybridization time that did 

not compromise FISH results was 2 h and 30’. With 2 h and 30’ of 

hybridization, the Chi square test for comparison of ratios showed 

the best concordance between the fast and the standard protocol 

with 14’ of heat pre-treatment (χ 2 = 0.08; 8 LD). 

Conclusion. We have developed and statistically validated a 

FISH protocol with a short hybridization time (2 h and 30’) to 

reduce turnaround for HER-2/neu reporting. 

role of intratumoral KrAS heterogeneity in the 

response to eGfr-targeted therapy of metastatic 

colorectal cancer (mCrC) patients 

1)Dono M. 2)Massucco C. 3)Cerruti G. 4)Truini M. 5)Chiara 

S. 6)Sonaglio C. 7)Canobbio L. 8)Anselmi L. 9)Margallo E. 

10)Zupo S. 

1)Tecnologie diagnostiche avanzate, Istituto nazionale ricerca cancro, 

Genova, Italia 2)Tecnologie diagnostiche avanzate, Istituto nazionale ricerca 

cancro, Genova, Italia 3)Tecnologie diagnostiche avanzate, Istituto 

nazionale ricerca cancro, Genova, Italia 4)Tecnologie diagnostiche avanzate, 

Istituto nazionale ricerca cancro, Genova, Italia 5)Oncologia medica 

integrata, Istituto nazionale ricerca cancro, Genova, Italia 6)Oncologia 

medica integrata, Istituto nazionale ricerca cancro, Genova, Italia 

7)Uo oncologia medica, Asl3, Genova, Italia 8)Sc anatomia aptologica, 

Asl3, Genova, Italia 9)Tecnologie diagnostiche avanzate, Istituto nazionale 

ricerca cancro, Genova, Italia 10)Tecnologie diagnostiche avanzate, 

Istituto nazionale ricerca cancro, Genova, Italia 

Background. EGFR targeted therapy has proven efficacious in 

the treatment of mCRC but clinical benefit is partially achieved 

only in patients with wild type KRAS gene (wt). Accordingly, 

EMEA made mandatory KRAS analysis before drug administration. 

Although several methods with different detection limits are 

available for KRAS mutation testing, validation and standardization 

of these procedures in a clinical setting are still lacking. 

Moreover, it is unknown if minimal presence of mutated clones in

288 

a bulk of wt tumor could impact on the outcome of EGFR treated 

patients: which is the % of mutated DNA/wt DNA able to identify 

the non responder patients? 

Methods. 75 patients considered KRAS wt by sequencing were 

treated by EGFR therapy. These patients were subsequently analyzed 

for KRAS mutations by a more sensitive real time method 

(LNA-PCR) in order to achieve amplification of rare mutated 

sequences present in the tumor DNA (sensitivity of sequencing 

and of LNA-PCR > 10% and £0.5% mutant DNA/wt DNA, respectively). 

Results. In a cohort of 75 patients, 60% (45/75) have wt KRAS 

with both techniques (wt seq/wt LNA-PCR), whereas 40% 

(30/75) have discordant results (wt seq/mutated LNA-PCR) demonstrating 

that LNA-PCR detected very rare mutated subclones. 

Presently, clinical data of efficacy of EGFR targeted agent are 

available for 27 patients out of the 75 studied for KRAS status. 

We did not find difference in clinical outcome among wt seq/wt 

LNA-PCR patients compared to wt/mut LNA-PCR ones (30% 

PR vs 23.5% PR, respectively), demonstrating that the capacity to 

detect less than 10% of mutated cells in a wt tumor do not seem 

to impact the efficacy of therapy. Although the n° of patients 

studied is small, this observation suggests an important clinical 

information: a very sensitive method not only is not necessary 

but it might exclude from therapy patients who could still benefit 

of it. We are extending clinical analysis on the total number of 

mCRC cases of this study. 

Correlations between intralobular interstitial 

morphological changes and epithelial changes in 

ageing testis 

1)I.E. Plesea, 2)S.D. Enache, 2)F.C. Popescu, 3)O.T. Pop, 1)C.G. 

Cotoi, 1)M.A. Enache, 1)R.M. Plesea 

1)Pathology, University of Medicine and Pharmacy, Craiova, Romania; 

2)Pathology, Emergency County Hospital, Craiova, Romania; 3)Histology, 

University of Medicine and Pharmacy, Craiova, Romania 

Background. Authors studied possible influences of intra (i) 

lobullar (l) stromal compounds [intertubullar spaces and seminiferous 

(s) tubule (t) wall (w)] morphologic changes on s epithelium 

(e) during ageing process. 

Methods. The material consisted of surgical samples of testicular 

tissue from 192 pacients with orchidectomy for prostate carcinoma. 

7 age groups were designed, from 50 to 80 years. Tissue 

samples were fixed in neutral buffered formalin, embedded in 

paraffin stained with HE, Goldner and Gömöri and immunomarked 

(in a subgroup of 28 cases) for smooth muscle actin, 

collagen IV, and CD34. 

Results. SE had an uneven involution, both individually and 

interindividually, but with normal spermtogenesis in manny of 

ST. E degenerative changes were seen mainly in L periphery. 

Different stages of maturation arresting were more frequent in 

older patients. 

IL septae had changes with extremely variable intensity, dispersed 

mainly in L periphery, without significant spread and 

without extensive trend with ageing. Leydig cells showed focal 

hyperplasia without extensive trend related with ageing. 

STW presented strictly in the internal layer of lamina propria 

(aposed to basement membrane of ES) a focal sclerosis, with 

variable extension concerning its presence, thickness and T circumference 

(T without sclerosis, with focal sclerosis and with 

fibro-hyaline “collar” - FHyC) but not related with ageing. 

IL arteriolae showed focal areas of degeneration with a wide individual 

and interindividual range of intensity and extention, but 

not related with age. 

Capillary network (CN), with both its peri-T and intramural segments, 

was present in all age groups, with no quantitative endothelial 

changes and decreasing only in very old cases. 

FHyC was often associated with E atrophy. 


Conclusions. STW focal sclerosis could explain focal degeneration 

of SE in senescence, although CN undergoes no significant 

changes. 

Acknowledgement: Work supported from Research Project 41- 

015/2007 financed by CNMP. 

fine-needle cytology in nodular thyroid 

pathology. Cytopathologic and clinical correlations 

using SIAPeC recommended diagnostic categories. 

An italian peripheral hospital experience 

1)Erra S. 2)Colombo M. 3)Modena S. 4)Bocchio C. 5)Butera M. 

6)Pastormerlo M. 7)Pavesi M. 

1)Soc anatomia pat, Santo spirito, Casale monferrato, Italia 2)Soc anatomia 

patologica, Santo spirito, Casale monferrato, Italia 3)Soc anatomia 

patologica, Santo spirito, Casale monferrato, Italia 4)Soc anatomia pat., 

Santo spirito, Casale monferrato, Italia 5)Soc anatomia patologica, Santo 

spirito, Casale monferrato, Italia 6)Soc anatomia pat., Santo spirito, Casale 

monferrato, Italia 7)Soc anatomia patologica, Santo spirito, Casale 

monferrato, Italia 

Introduction. Fine-Needle Aspiration Cytology is a consolidated 

procedure in the diagnosis and clinical management of the 

thyroid palpable or US detected nodule.In past years cytological 

diagnosis obtained from FNC of thyroid nodules were descriptive 

and in the most of cases they lack objectivity,having only some 

few grade in reproducibility. In November 2007 Italian Society 

of Pathologists (SIAPEC) decides to adopt diagnostic categories 

for the diagnosis in thyroid cytology, recommending its use in 

routine diagnostic activity. SIAPEC categories are similar to 

British Thyroid Association ones, whereas Tir 1 indicates inadequate 

sample, Tir 2 negative for malignancy, Tir 3 presence of 

cells with indeterminate significance(microfollicular or oncocitic 

proliferation), Tir 4 suspected for malignancy, Tir 5 positive for 

malignant cells. In the present report, we evaluate our experience 

with Fine-needle Cytology for the diagnosis of the thyroid nodules 

adopting SIAPEC recommended diagnostic categories. 

Methods. A retrospective search using database from our Pathology 

Department was performed to identify patients who underwent 

FNC for palpable or non-palpable thyroid nodules from 

January 2008 to April 2010 at Casale Monferrato Santo Spirito 

Hospital. In our structure, thyroid pathology is studied and managed 

in an equipment contest, with the presence of a specialist 

surgeon, an endocrinologist, a pathologist and an US-experienced 

radiologist. 382 US-guided FNC were performed in the reported 

period; the aspirate samples were treated in double way: a part 

material was shuffled and stained with Papanicolau and May- 

Grunwald Giemsa stain, while a part was fixed in an alcoholic 

fixative to prepare monolayer slide with Cytic Thin Prep 2000 

Processor and coloured with PAP stain. FNC specimens were 

examined and every diagnosis was expressed using SIAPEC 

recommended diagnostic categories. The obtained results were 

correlated to the final diagnosis on surgical specimen in operated 

patients, while in the most of negative or inadequate results (Tir 

2 or Tir 1 on cytological sample) a clinical correlation was made, 

with follow-up of the patient or the repetition of FNC exam. 

Results. 382 FNC was performed, with the prevalence of female 

patients (329 cases in front of 53 males). These was the results: 

133Tir 1 (34%), 178 Tir 2 (46%),57 Tir 3 (15%),11 Tir 4(2.8%),3 

Tir 5 (0.8%). Histological evaluation was possible for 54 operated 

patients, with a good value of specificity and sensibility obtained 

from cyto-histological correlation.Only 3/9 Tir1 cases were neoplastic 

lesions and 2 Tir2 cases were histological malignant on 8 cytological 

negative samples with surgical removal; the most of Tir3 

cases (23/27 operated cases) corresponded to follicular iperplasia 

or adenoma, while the other 4 cases were follicular carcinoma (2 

cases) or follicular variant of papillary carcinoma (2 ones). All Tir4 

and Tir5 samples were malignant with surgical removal in our or 

in other hospitals, except to one Tir4 case corresponding to nodular


iperplasia. Tir2 not operated patients are in follow-up without actual 

significant pathological reports; most of Tir1 cases correspond 

to cystic nodules on US exam and not operated Tir3 patients are in 

follow-up or will go to surgical removal in few time. The use of 

diagnostic categories in cytopathological reports of nodular thyroid 

pathology gives objectivity to FNC exam and improves the equipment 

collaboration among different medical specialists. 

GIST: differential diagnosis using 

immunohistochemical panel of antibodies in 

selected cases 

1)Erra S. 2)Modena S. 3)Colombo M. 4)Mazzoni E. 5)Costamagna 

D. 6)Pavesi M. 

1)Soc Anatomia Patologica, Santo Spirito, Casale Monferrato, Italia 2)Soc 

Anatomia Patologica, Santo Spirito, Casale Monferrato, Italia 3)Soc Anatomia 

Patologica, Santo Spirito, Casale Monferrato, Italia 4)Soc Anatomia 

Patologica, Santo Spirito, Casale Monferrato, Italia 5)Soc Chirurgia 

2, Azienda Ospedaliera Ospedale Maggiore Della Carità, Novara, Italia 

6)Soc Anatomia Patologica, Santo Spirito, Casale Monferrato, Italia 

Background. GIST (Gastrointestinal Stromal Tumor) is the 

most common mesenchymal tumor in gastrointestinal tract. It 

originates from the neoplastic transformation of the interstitial 

cell of Cajal, a neuron-derived cell migrating from the neural 

crest to the intestine during fetal life. Mechanisms involved in 

cancerogenesis of GIST have been identified in oncogenic point 

mutation of cKit, immunohistochemical detectable trough CD117 

monoclonal antibody. Recent studies report cKit-negative GIST. 

DOG-1 monoclonal antibody is used in differential diagnosis of 

these cases, in alternative to genetic analysis of cKit point mutation. 

Combination of CD117 and DOG-1 mAbs is useful in correct 

differential diagnosis of GIST and in determining prognostic 

factors and response to target therapy. 

Methods. 18 surgical specimens of mesenchymal tumors of 

gastroenteric district have been studied. Male:female ratio of 

these cases is 6:12. 13 cases are intraparietal gastric tumors, 3 

ileal mesenchymal masses, 1 digiunal tumor and only one case 

is localized in peritoneal serosa, without localizations in digestive 

tract. Immunohistochemical profile of each case has been 

determined using a panel of mAbs, including CD34, CD117 and 

DOG-1 in addition to Actins and S100 protein. 

Results. 14/18 (83%) cases are resulted positive for DOG-1 mAb. 

10 of these DOG-1 positive GISTs overexpress cKit; in 3 stromal 

tumors cKit overexpression has been detected, with immunohistochemical 

negativity for DOG-1. Only one case has resulted 

negative for each of the monoclonal antibodies investigated, with 

a scanty and focal expression of DOG-1. We attribute this result 

to problems in fixation of the surgical specimen. 89% of the selected 

cases (15/18) are resulted uncommetted GISTs (CD34+), 

one case has corresponded to GIST with neural differantiation 

and in two cases the only immunohistochemical positivity has 

been respectively for DOG-1 in a case and for CD117 in the other 

one. In conclusion, we remarke the utility of a complete immunohistochemical 

panel of monoclonal antibodies in differential 

diagnosis of mesenchymal gatrointestinal tumors, including the 

combined use of CD117 and DOG-1, in alternative to genetical 

investigation of cKit point mutation. 

risk management: correct patient and specimen 

identification in a surgical pathology laboratory. 

The experience of Infermi Hospital, rimini, Italy 

Fabbretti G., Sampaoli I, Fiumana M, *Busatto F, *Santucci L. 

Surgical Pathology Unit, Department of Clinical Pathology and Radiology, 

*Information Thecnology Departement, Infermi Hospital Rimini 

Because of its complex nature, surgical pathology practice is 

error prone. In this article we describe our methods for reducing 

error as much as possible in the pre-analytic and analytic phases. 

289 

This was achieved by revising procedures, using computer technology 

and automation. 

Most mistakes are the result of human error in the identification 

and matching of patient and sample. To avoid faulty data 

interpretation, we employed a new comprehensive computer system 

that acquires all patient ID information directly from the 

hospital’s database with a remote order entry; it also provides 

label and request forms via-Web. Patient and sample are identified 

directly and immediately and at the site where the surgical 

procedures are performed. Barcode technology is used to input 

information at every step and automation is used for cassettes and 

slides to avoid errors that occur when information is recorded or 

transferred by hand. Quality control checks occur at every step of 

the process so as to ensure that nothing is left to chance and that 

no phase is dependent on a single person, no matter how able. 

The system also provides statistical analysis of errors so that new 

strategies can be implemented in order to avoid their repetition. 

In addition, staff receive frequent training on error avoidance and 

new developments. 

The results have been good, with a very low error rate recurrence 

(0,27%). None of these compromised patient health and all errors 

were detected before the release of the diagnosis report. 

Cancerization of cutaneous flap reconstruction 

for oral squamous cell carcinoma: three cases 

studied with the MTDNA D-loop sequence analysis 

1)Farnedi A. 2)Marchetti C. 3)Morandi L. 4)Cocchi R. 5)Badiali 

G. 6)Montebugnoli L. 7)Pennesi M.G. 8)Eusebi L.H. 9)Foschini 

M.P. 

1)Section of Anatomic Pathology, Department of Haematology and Oncology 

“L. and A. Seràgnoli”, University of Bologna, Bellaria hospital, 

Bologna, Italy 2)Department of Oral and Maxillofacial surgery, S. Orsola-Malpighi 

hospital, University of Bologna, Bologna, Italy 3)Section of 

Anatomic Pathology, Department of Haematology and Oncology “L. and 

A. Seràgnoli”, University of Bologna, Bellaria hospital, Bologna, Italy 

4)Department of Maxillo-facial surgery, Bellaria hospital, Bologna, Italy 

5)Department of Oral and Maxillofacial surgery, S. Orsola-Malpighi hospital, 

University of Bologna, Bologna, Italy 6)Department of Oral sciences, 

University of Bologna, Bologna, Italy 7)Department of Maxillo-facial 

surgery, Bellaria hospital, Bologna, Italy 8)Section of Anatomic Pathology, 

Department of Haematology and Oncology “L. and A. Seràgnoli”, 

University of Bologna, Bellaria hospital, Bologna, Italy 9)Section of Anatomic 

Pathology, Department of Haematology and Oncology “L. and A. 

Seràgnoli”, University of Bologna, Bellaria hospital, Bologna, Italy 

Background. Tissue defects, resulting from surgical resection 

of Oral squamous cell carcinoma (OSCC), are routinely reconstructed 

with skin grafts. OSCCs arising from the grafted skin 

have been described, however, it is still unclear whether they are 

recurrences or second primary tumours. 

The clonal relationship, based on the high frequency rate of Dloop 

region mtDNA mutations in tumors, can be a reliable marker 

for clonality assays from microdissected paraffin-embedded tissue 

samples. 

Methods. By screening mitochondrial DNA D-loop region, we 

evaluated the clonal relationship between the primary OSCCs and 

the tumor appearing in the skin graft in three patients. 

In all the three cases, primary tumors and the second tumors appearing 

on the skin grafts had been formalin fixed and paraffin 

embedded. All the tumors were classified according to grading 

and TNM staging. Pertinent lesions were microdissected using 

the laser assisted SL µcut microtest GmbH distributed by Nikon 

and DNA was extracted. The mtDNA D-loop sequence analysis 

was performed by amplifying four overlapping segments. PCR 

products were directly sequenced using CEQ2000 XL instrument. 

Results. Tumors arising in the skin graft showed a clonal relationship 

with the previous OSCC and, on the basis of the results 

obtained with the mtDNA analysis, could be considered related

290 

to the primary OSCC and developed from a common genetically 

altered field. 

Data here obtained suggest the possibility of a spreading of the 

clonal neoplastic cell population of the primary OSCC to the 

cutaneous flap that might be stimulated by cytokines produced 

by the grafted skin. 

More studies are needed to evaluate the molecular relationship 

between primary and second OSCC in order to identify patients at 

higher risk of developing a second tumor of the skin graft. 

Proliferative activity in human breast cancer: 

assessment of KI67 Automated evaluation And The 

Influence Of Different KI67 equivalent Antibodies 

1)Fasanella S. 2)Cantaloni C. 3)Eccher C. 4)Cuorvo LV. 5)Leonardi 

E. 6)Bragantini E. 7)Morelli L. 8)Aldovini D. 9)Dalla palma 

P. 10)Barbareschi M. 

1)Anatomia Patologica, S.Chiara, Trento, Italia 2)Anatomia Patologica, 

S.Chiara, Trento, Italia 3)Statistica, Fondazione Bruno Kessler, Trento, 

Italia 4)Anatomia Patologica, S.Chiara, Trento, Italia 5)Anatomia Patologica, 

S.Chiara, Trento, Italia 6)Anatomia Patologica, S.Chiara, Trento, 

Italia 7)Anatomia Patologica, S.Chiara, Trento, Italia 8)Anatomia Patologica, 

S.Chiara, Trento, Italia 9)Anatomia Patologica, S.Chiara, Trento, 

Italia 10)Anatomia Patologica, S.Chiara, Trento, Italia 

Background. Ki-67 labeling index (Ki67LI) is a measure of 

tumor proliferation, with important clinical relevance in breast 

cancer, and it is extremely important to standardize its evaluation. 

To test the efficacy of computer assisted microscopy (CAM) applied 

to completely digitized slides and to assess its feasibility in 

routine practice. 

Methods. 315 consecutive breast cancer routinely immunostained 

for Ki-67 (223 with SP6 (Labvision) and 92 with MM1 

(Novocastra) antibodies) previously examined by an experienced 

pathologist, have been re-evaluated using Aperio Scanscope Xs. 

Results. Mean human Ki67LI values were 35.91% ± 14.35% 

and 28.37% ± 18.29% respectively for SP6 and MM1 antibodies; 

mean CAM Ki-67LI values were 30.97% ± 19.19% and 

22.12% ± 18.36% respectively for SP6 and MM1. Human and 

CAM evaluation are statistically highly correlated (Pearson: 

0.859, p < 0.0001), although human LI are systematically higher. 

Cases have been subdivided in three groups based on tertile 

distribution; cut-offs varied depending on antibody used and on 

evaluation methods: for human evaluation using SP6 and MM1 

they were £ 30, 31-42, ≥ 43, and £ 20, 21-40, ≥41 respectively; 

for CAM evaluation they were £ 19, 20-37, ≥ 38 and £ 12, 13-24, 

≥ 25. Our study shows that a) CAM can be easily adopted in routine 

practice, b) human and CAM Ki67LI are highly correlated, 

although human LI are systematically higher, c) Ki67LI grouping 

on the basis of tertiles obtained using different evaluation 

methods and different antibodies shows important differences in 

cut-off values. These cut-off may differ from the ones suggested 

in literature (e.g.: 2009 Sant Gallen Consensus), underscoring 

that it is not correct to adopt general cut-off values to subgroup 

cases without taking in consideration the evaluation methods and 

antibody used. 

Concurrent pheochromocytoma and cortical 

carcinoma of the adrenal gland 

1)Fassina A. 2)Cappellesso R. 3)Tricarico P. 4)Bombonato F. 

5)Schiavi F. 

1)Scienze medico diagnostiche e terapie speciali, Università di padova, 

Padova, Italia 2)Scienze medico diagnostiche e terapie speciali, Università 

di padova, Padova, Italia 3)Scienze medico diagnostiche e terapie speciali, 

Università di padova, Padova, Italia 4)Scienze medico diagnostiche 

e terapie speciali, Università di padova, Padova, Italia 5)Istituto veneto di 

oncologia, Università di padova, Padova, Italia 


Background. The concurrent occurrence of pheochromocytoma 

and adrenal cortical carcinoma (ACC) has never been reported so 

far in western literature, except for the association of pheochromocytoma 

with benign cortical conditions such as hyperplasia or 

adenoma. 

Methods. A lesion in the left adrenal gland was excised in a 46year-old 

woman, and immunohistochemistry for MIB-1, inhibin 

α, CD56, chromogranin A, synaptophysin, S100, serotonin, somatostatin, 

CK7, and CK20 was performed. 

Exons and intronic flanking regions of SDHB, SDHC, SDHD, 

VHL, RET (exons 8, 10, 11, 13, 14, 15 and 16) and TMEM127 

genes were screened by direct sequencing in DNA extracted 

from peripheral blood leukocytes. Gross deletion analysis was 

performed using multiple ligation-dependent probe amplification 

method (MLPA) for SDHB, SDHC, SDHD and VHL genes, 

and using a quantitative multiplex PCR for TMEM127 gene. 

Results. The lesion was consistent with a diagnosis of pheochromocytoma, 

with scant nuclear atypia, few spindle cells, low Ki67 

index, and a PASS score < 4. A concurrent second cell population 

was found adjacent to the pheochromocytoma, consisting of small 

cells with oval nuclei, occasional nucleoli, dispersed chromatin, 

and eosinophilic cytoplasm reminiscent of zona reticularis cells. 

There was neither necrosis nor broad collagen bands. Immunostaining 

of this second cell population was positive for inhibin α 

and CD56, and negative for chromogranin A, synaptophysin, 

S100, serotonin, somatostatin, CK7, and CK20. The Ki67 index 

was high ( > 4%). Capsular and vascular invasion could not be 

assessed due to sample fragmentation. The final diagnosis was 

concurrent ACC and pheochromocytoma. Genetic testing was 

negative for all familial mutations confirming the sporadic nature 

of these tumors. 

Conclusion. Paracrine stimulation of adrenal cortex proliferation 

by factors secreted by the pheochromocytoma has been so far the 

only explanation for the coexistence of these tumors. 

mirNA signature and atherosclerotic plaque 

instability 

1)Felicioni L. 2)Malatesta S. 3)Mammarella C. 4)Ucchino S. 

5)Spigonardo F. 6)Del grammastro M. 7)Cipollone F. 8)Mezzetti 

A. 9)Marchetti A. 10)Buttitta F. 

1)Dipartimento Di Oncologia E Medicina Sperimentale, Università “G. 

D’Annunzio”, Chieti, Italia 2)Dipartimento Di Oncologia E Medicina 

Sperimentale, Università “G. D’Annunzio”, Chieti, Italia 3)Centro Di Ricerca 

Clinica, Ce.S.I., Fondazione Università “G. D’Annunzio”, Chieti, 

Italia 4)Dipartimento Di Chirurgia Vascolare, Università “G. D’Annunzio”, 

Chieti, Italia 5)Dipartimento Di Chirurgia Vascolare, Università 

“G. D’Annunzio”, Chieti, Italia 6)Dipartimento Di Oncologia E Medicina 

Sperimentale, Università “G. D’Annunzio”, Chieti, Italia 7)Centro Di 

Ricerca Clinica, Ce.S.I., Fondazione Università “G. D’Annunzio”, Chieti, 

Italia 8)Centro Di Ricerca Clinica, Ce.S.I., Fondazione Università “G. 

D’Annunzio”, Chieti, Italia 9)Dipartimento Di Oncologia E Medicina 

Sperimentale, Università “G. D’Annunzio”, Chieti, Italia 10)Dipartimento 

Di Oncologia E Medicina Sperimentale, Università “G. D’Annunzio”, 

Chieti, Italia 

Background. The natural history of atherosclerotic plaque, in 

particular its evolution toward rupture and consequent thrombosis, 

is still unclear and to date remains impossible to identify 

the single patient in whom the disruption of a vulnerable plaque 

leading to myocardial infarction or stroke is likely to occur. 

The identification of new predictive markers of future atherothrombotic 

events (i.e. myocardial infarction and stroke), is still 

a main challenge for scientific research. MicroRNAs (miRNAs) 

are small non-coding endogenous RNAs and represent a new 

important class of gene regulators. They have been shown implicated 

in the pathogenesis of various neoplastic diseases and 

more recently in some cardiovascular disease. The aim of this


study was to investigate the expression level of miRNAs in human 

plaques and to correlate it with clinical features of plaque 

destabilization. 

Methods and Results. Two separate groups of plaques were 

collected from patients who underwent carotid endarterectomy 

in the Chieti (n = 15) and Ancona (n = 38) hospitals. All the 

plaques were subdivided in symptomatic (n = 22) and asymptomatic 

(n = 31) according to the presence/absence of stroke. Firstly, 

on the plaques collected at the Chieti hospital, we performed 

large-scale analysis of miRNA expression by Real Time-PCR. 

Between the miRNAs examined, we discovered profound differences 

in the expression of 5 miRNAs (miRNA-100, miRNA-127, 

miRNA-145, miRNA-133a and miRNA-133b) in symptomatic 

vs asymptomatic plaques. Remarkably, when we repeated the 

analysis on the 41 cases of the Ancona plaque sub-set, all these 

5 miRNAs confirmed to be significantly more expressed in the 

symptomatic plaques. 

These results are the first to report alterations in the expression of 

specific miRNAs in human atherosclerotic plaques and suggest 

that miRNAs may have an important role in regulating the evolution 

of atherosclerotic plaque toward instability and rupture. 

Suitability of clear cell renal cell carcinoma to heat 

shock proteins-inhibitors 

1) Ferrero S. 2) Gazzano G. 3)Brunelli M. 4) Bisaro C. 5)Martignoni 

G. 6) Bosari S. 

1) Anatomia Patologica, Università Di Milano, S. Paolo, Milano, Italia 2) 

Anatomia Patologica, Università Di Milano, S. Paolo, Milano, Italia 3) 

Anatomia Patologica, Università Di Verona, Policlinico Gb Rossi, Verona, 

Italia 4) Anatomia Patologica, Università Di Milano, S. Paolo, Milano, 

Italia 5) Anatomia Patologica, Università Di Verona, Policlinico Gb Rossi, 

Verona, Italia 6) Anatomia Patologica, Università Di Milano, S. Paolo, 

Milano, Italia 

Background. Heat shock proteins (HSP) HSP27 and HSP90 are 

expressed in response to a wide variety of physiological and environmental 

insults thus allowing the cell to survive to lethal conditions. 

In cancer cells, both HSP27 and HSP70 may participate in 

oncogenesis thus the inhibition of HSP90 and HSP27 has become 

a novel strategy of cancer therapy. Few cases have been studied 

among renal cell carcinoma (RCC) at a tissue level. 

Methods. 388 RCCs with clear cell histology available on 10 tissue 

microarrays were recruited. Each case was represented by 3 

neoplastic cores. All these cases were stratified according to the 

Mayo Clinic SSIGN (Size, Staging, Grading, Necrosis) score into 

five prognostic groups with increasing worse prognosis (1 to 5). 

Immunostainings for HSP90 and 27 were performed. Scoring was 

performed by multiplying % of positivity (0 = 0, 1 ≤ 30%; 2 = 31- 

60%; 3 = 61-100%) x intensity (0 = 0, 1 = fant, 2 = moderate, 

3 = strong) of neoplastic cells. A mean immunoscore number 

from the three cores was set per case. Results were expressed as 

immunoscore per number of scorable cases per SSIGN category. 

Results. 117 and 122 RCCs were respectively available for 

HSP90 and HSP27 scoring. The remaining were lost due to loss 

of tissue sections or few neoplastic cells scorable. HSP90 scored 

4,9 in 32 with SSIGN1, 3,5 in 41 SSIGN2, 4,8 in 11 SSIGN3, 

4,2 in 22 SSIGN4 and 5 in 3 SSIGN5 patients. HSP27 scored 4,6 

in 33 with SSIGN1, 3,1 in 43 SSIGN2, 2,6 in 11 SSIGN3, 3,6 in 

24 SSIGN4 and 2,7 in 3 SSIGN5 patients. A significant trend of 

increasing value for HSP90 has been observed when comparing 

cases tabled SSIGN1-2 vs SSIGN3-5 (4,2 to 4,6 mean values) 

(p = 0.05); a minor stratification has been observed for HSP27 

(3,8 to 3,9) (p = 0.08). 

In conclusion, HSP90 and HSP27 are variable immunoexpressed 

in a subset of clear RCCs, with a trend to higher prognostic 

SSIGN score. At light of new emerging tailored therapies in kidney 

cancer, we report an increasing suitability of these patients to 

inhibitor of heat shock protein molecules. 

evaluation of Her2 overexpression, gene 

amplification and chromosome 17 centromere 

copy number in primary pancreatic 

adenocarcinoma, metastatic lymph node and 

metastasis 

291 

1)S. Salvi, 2,3)P. Ferro, 1)S. Boccardo, 3)N. Gorji, 3)P. Dessanti, 

4)D. Gianquinto, 5)A. Vigani, 3,6)M. Franceschini, 1)M. Truini, 

7)M.P. Pistillo, 3)F. Fedeli, 3)S. Roncella 

1)SC Anatomia Patologica e Citoistologia, IST, Genova; 2)AIL “F. Lanzone”, 

La Spezia, Italia; 3)SC Anatomia ed Istologia Patologica e Citodiagnostica, 

ASL5, La Spezia, Italia; 4)SC Chirurgia, ASL5, La Spezia, 

Italia; 5)SC Oncologia, ASL5, La Spezia, Italia; 6)Comitato Assistenza 

Malati e Lotta contro i Tumori, Sarzana, Italia; 7)SC Genetica dei Tumori, 

Laboratorio Tumori Mammari, IST, Genova, Italia 

Background. Pancreatic ductal adenocarcinoma (PDA) is one of 

the most lethal cancer with increasing incidence in the Western 

world. Thus, the development of novel therapeutic strategies 

is a main focus to improve PDA patient survival. Trastuzumab 

targeted therapy for p185 HER2 has been proposed although the 

percentage of cases with HER2 overexpression remains unclear, 

as well as the biological role of the gene status. The aim of this 

study was to evaluate, at protein and genomic expression level, 

the status of HER2 in PDA. 

Methods. We analyzed 60 PDA including 22 tumours at initial 

diagnosis, 15 matched nodal metastasis and 23 tru-cut needle or 

tumours from recurrent distant metastasis. On paraffin-embedded 

tissues, we performed IHC staining of p185 HER2 with 4B5 mAb, 

by the Benchmark XT system (Ventana). FISH to evaluate HER2 

amplification and copy number of chromosome 17 centromere 

(CEP17) was performed by Pathvision kit (Abbott) and/or Zyto- 

Light kit (ZytoVision). 

Results. We found p185 HER2 overexpression in 8/60 (13%) of 

tissues, gene amplification in 3/60 (5%) and increased CEP17 in 

6/60 (10%) of tissues. In particular, p185 HER2 overexpression was 

found in 1/22 (5%) of primary PDA, 7/23 (29%) of metastasis 

but in none of metastatic lymnh nodes (0/15). HER2 amplification 

was restricted to the recurrent distant metastasis 3/23 (12%), 

whereas increased CEP17 was found in 4/23 (17%) of distant 

metastasis and in 2/15 (13%) of metastatic lymph nodes. All 3 

cases of HER2 amplified tumours showed IHC score 2+ whereas 

only one case of those with increased CEP17 was associated to 

overexpression of p185 HER2 (score 2+). 

Conclusions. In PDA, HER2 amplification and increased CEP17 

copies was mainly related to recurrent metastasis or lymph node 

infiltration. All HER2 amplified cases showed p185 HER2 overexpression, 

but at lower extent than that observed in breast cancer 

(score 2+ vs score 3+), while the increased CEP17 was not related 

to overexpression of p185 HER2 . 

Globet cell carcinoid of the ovary: 

primary or secondary? A case report 

1)Fiore MG. 2)Rossi R. 3)Palumbo M. 4)Clemente D. 5)Piscitelli 

D. 6)Resta L. 


Popoliliclinico, Bari, Italia 3)Anatomia patologica, Policlinico, Bari, Italia 



Background. Globet cell carcinoid of the ovary is a very rare 

neoplasm with uncertain biological behavior. It is, usually, a metastatic 

tumor from the gastrointestinal tract, especially from the 

appendix. Exceptionally it is primary of the ovary. The criteria in 

favour of the primitive origin are the unilaterality, the absence of 

multiple ovarian nodules, the presence of teratomatous elements. 

Materials and methods. 18-year-old woman with unilateral 

tumor of the left ovary wich was sligtly enlarged measuring cm 

6 × 4,5 × 3. The ovarian mass was oval, firm, smooth and ap-

292 

peared to be encapsulated. On cut section, it was gray-yellow, entirely 

solid and lobulated. Histologically, ovarian neoplasm was 

composed of numerous small cords and nests of epithelial cells 

some of which with finely granular eosinophilic cytoplasm, whilst 

others with clear microvacuolated cytoplasm or with globet cell 

appearance. The nuclear pleomorphism and mitotic activity were 

minimal. The tumor cells were present within lymphatic spaces 

of the parenchyma as well as of the mesoovarian soft tissues. The 

tumor was in pure form and no associated with teratomatous elements. 

The neoplastic cells appeared to be strongly positive for 

periodic acid-Schiff, Alcian blue, cytokeratin; some individual 

neoplastic cells were positive for synaptophysin and chromogranin. 

Electron microscopy demonstrated the presence of two 

types of cells: first type with mucinous vacuoles of varying sizes 

and the second type with few membrane-bound neuroendocrine 

granules. A bioptic specimen of the controlateral ovary was free 

of tumor as well as the appendix on microscopic examination. 

Results. Further careful radiologic investigation were negative 

and the patient, after 1 year of follow-up, was alive and free of 

tumor. These findings much more supporte the hypothesis of primary 

globet cell carcinoid tumor originating in the ovary. 

Intraoral “animal-type melanoma”: 

description of a case 

Fiore MG. Rossi R. Palumbo M. Colagrande A. Piscitelli D. 

Resta L. 


Background. Animal type melanoma is a rare histological variant 

of melanoma of uncertain biological behavior, so termed 

because its similarity to a variant of melanoma seen in grey 

horses. It frequently shows an indolent course without mortality, 

but sometimes is highly aggressive, with a rapidly fatal outcome, 

similarly to common melanoma. Recently it has been included in 

a new entity denominated “Pigmented epithelioid melanocytoma” 

(PEM), a spectrum of melanocytic tumors, such as the epithelioid 

blue nevus, malignant blue nevus and other type, characterized by 

very similar features, sometimes associated and overlapped one 

to the other, but still with a very different biological behaviour, 

difficult to be predicted on morphological grounds. 

Materials and methods. 68-year-old male with a large, pigmented, 

warty plaque-like lesion of the palatal mucosa. Microscopically, 

the neoplastic cells were heavily pigmented with an epithelioid 

morphology, vertical-type growth pattern with infiltration 

of the dermis and of the skeletal muscular plan. The nuclear 

pleomorphism was minimal and occasional mitotic figures were 

identified. The cellular density, very strong in the central zones 

of the proliferation, typically is reduced at the periphery of the 

lesion where the cells show dendritic appearance, morphologically 

reminiscent of a blue nevus. The immunohistochemical and 

ultrastructural studies confirmed the nature of neoplasia formed 

by melanosoma-producing cells. 

Results. The morphology and highly pigmented nature of the 

tumor was suggestive of animal-type melanoma, also referred 

as pigmented epithelioid melanocytoma, where the vertical-type 

growth pattern, with precocious involvement of the depth structures, 

rather than the histological features has been the determinant 

factor of a poor prognosis, evidenced by the presence of 

pulmonary metastases. 

Inflammatory and atrophic lesions of the prostate: 

a study of concordange among histopathologists 

1)Fiorentino M. 2)Giunchi F. 3)Bollito E. 4)Stark J. 

1)Anatomia Patologica, Policlinico S. Orsola-Malpighi, Bologna, Italia 

2)Anatomia Patologica, Policlinico S. Orsola-Malpighi, Bologna, Italia 

3)Anatomia Patologica, Orbassano, Torino, Italia 4) Department Of Epidemiology, 

Harvard School Of Public Health, Boston, U.S.A. 


Background. Inflammatory and atrophic lesions of the prostate 

are widely underestimated. According to the current European 

guidelines the mention of atrophic lesions in the pathology reports 

is not mandatory. A classification of prostate atrophic 

lesions in simple atrophy (SA), cystic atrophy (SACF), postatrophic 

hyperplasia (PAH) and partial atrophy (PA) has been 

recently proposed, but its application in the histopathological 

practice is still debated, particularly in the European countries. 

Recent epidemiological and biological evidences pointed out a 

causal relationship between the presence of proliferative atrophic 

lesions (particularly PAH) and the development of PIN and 

prostate cancer. Due to their variable histological appearance the 

diagnosis of prostate atrophic lesions is extremely subjective and 

might be skewed by inter-observer variability. 

Methods and Results. In order to optimize the rate of agreement 

among pathologists on the diagnosis of atrophic lesions we 

planned a concordance study. The feasibility of the study was 

analysed calculating the sample size required for the inter-rater 

reliability assessment considering the following variables: the 

number of categories; the number of raters; the number of subjects 

being rated and the possible distribution of the categories. 

To maximize the power of the analysis we decided to enroll 15 

dedicated uro-pathologists and 15 general practicing pathologist 

using 5 sets of 120 slides. Each set included atrophic lesions classified 

according to the criteria proposed by Demarzo et al. and 

stratified according to the known distribution of each lesion in 

popolation. The kappa concordance rate among pathologists with 

different background on the diagnosis of prostate atrophic lesions 

will definitely shed light on the introduction of the atrophic nomenclature 

in the routine patology reporting. 

The prognostic value of resection margins in oral 

squamous cell carcinoma 

1)Flamminio F. 2)Cocchi R. 3)Pennesi M. G. 4)Eusebi L.H. 

5)Foschini M.P. 

1)Dip. di Ematologia e Scienze Oncologiche “L. e A. Seràgnoli”, Sezione 

di Anatomia, Istologia e Citologia Patologica “Marcello Malpighi”, 

Università-ASL Ospedale Bellaria, Bologna, Italia 2)U.O.C. Chirurgia 

Maxillo Facciale, Dip. Neuroscienze, Ospedale Bellaria, Bologna, Italia 

3) U.O.C. Chirurgia Maxillo Facciale, Dip. Neuroscienze, Ospedale Bellaria, 

Bologna, Italia 4)Dip. di Ematologia e Scienze Oncologiche “L. 

e A. Seràgnoli”, Sezione di Anatomia, Istologia e Citologia Patologica 

“Marcello Malpighi”, Università-ASL Ospedale Bellaria, Bologna, Italia 

5)Dip. di Ematologia e Scienze Oncologiche “L. e A. Seràgnoli”, Sezione 

di Anatomia, Istologia e Citologia Patologica “Marcello Malpighi”, Università-ASL 

Ospedale Bellaria, Bologna, Italia 

Background. Surgical resection of oral squamous cell carcinoma 

(OSCC) with appropriate margins of uninvolved tissue is critical 

to prevent local recurrences and for making correct therapeutical 

decisions. 

Aim of the present study was to evaluate several histologic features 

useful for prognostic purposes. 

Materials and methods. 141 surgically resected OSCCs, with 

a follow-up from 6 to 120 months (media 40 months), were histologically 

reviewed in order to assess: depth of invasion, nerve 

invasion, pattern of tumour invasion, minimum distance from 

margins and presence of dysplasia on resection margins. 

Results. Lymph-node metastasis were strictly related to depth of 

invasion and were present in 46.5% of cases which showed depth 

of invasion > 5mm and in 10% of cases with depth of invasion 

< 5mm. Furthermore, lymph-node metastasis were strictly related 

to nerve invasion, and were present in 60.6% of cases with nerve 

invasion and in 27.7% of cases without nerve invasion. 

Recurrences were strongly related to the tumour invasion pattern. 

They developed in 13.9% of cases, which showed infiltrative 

pattern of invasion and did not develop in cases with a compact 

pattern of invasion. In T1/T2 cases, recurrences were strongly 

related to the distance of OSCC from surgical margins. They


developed in 23.1% of cases with minimum distance < 1mm, in 

11.1% of cases with minimum distance of 1-5mm and in 3.3% of 

cases with minimum distance > 5mm. Finally, second OSCC was 

strictly related to the presence of high grade dysplasia on surgical 

margins. It developed in 30% of cases with high grade dysplasia 

on surgical margins and in 14% of cases with low grade dysplasia 

or without dysplasia on surgical margins. 

Conclusions. The present data confirm that an accurate evaluation 

of the histological features of resected primary OSCCs and 

of the surgical margins has a strong prognostic impact on lymphnode 

metastases, recurrences and appearance of second OSCC. 

Gastrointestinal autonomic nerve tumors: a case 

report and short review of literature 

S.A. Senatore, F. Floccari, P. Rizzo, A. D’Amuri 

U.O.C. Anatomia Patologica, “Ospedale Sacro Cuore di Gesù”, P.O. Gallipoli 

ASL Lecce, Gallipoli, Italia 

Background. A 78 year-old-woman underwent gastric biopsy 

which discovered a gastrointestinal autonomic nerve tumor 

(GANT) with high malignancy potential. One month later an 

explorative laparotomy for a peritoneal carcinosis was performed. 

The omentum was removed and a diagnosis of GANT was done. 

Methods. Immunohistochemical stains (IHC) were performed for 

CD117, CD45 (LYMPH T), CD45 LCA, CK AE1/AE3, Desmin, 

EMA, NSE, Ki-67, S-100 protein, Sinaptofisin and Vimentin. 

Results. GANTs are uncommon stromal tumours accounting 

for 0,1% of benign tumors of the gastrointestinal tract. It is a 

subgroup of GISTs with specific ultrastructural differences suggesting 

its origin from the myenteric plexus. Common sites for 

GANTs include the stomach, duodenum, jejunum and ileum and 

to date literature search has revealed only twenty cases of colonic 

schwannomas and four cases of rectal shwannomas. The exact 

biological behaviour of GANTs is not yet fully elucidated due to 

the limited number of reported cases and as a result. Mitotic activity 

( > 5 mitoses per high power fields) and tumor size ( > 5cm) 

are associated with high risk of metastases or recurrence. Treatment 

of CD117 positive GANTs with tyrosin kinase inhibitors 

have been shown to beneficial and could in the future, represent 

an appropriate form of palliative therapy in those patients with 

unresectable as well as metastatic tumors. Radical surgical resection 

of GANT seems to be the only available curative approach 

and long term survival is possible even in large metastasized 

tumors. Our histologic examination showed an encapsulated 

tumor made of spindle cells with several nuclear pleomorphism 

and high mitotic activity. Immunohistochemically it was strongly 

positive for CD117, NSE, Sinaptofisin, S-100 but negative for 

desmin. Ki-67 proliferative index was high. In our case report the 

high malignant potential evaluated in the gastric biopsy caused 

peritoneal metastases and the patient died later for neoplasm 

complications. 

Primary multiple extragastrointestinal stromal 

tumors of the greater omentum: a case report 

A. D’Amuri, F. Floccari, P. Rizzo, L. Peschiulli, S.A. Senatore 

U.O.C. Anatomia Patologica, “Ospedale Sacro Cuore di Gesù”, P.O. Gallipoli 

ASL Lecce, Gallipoli, Italia 

Background. A 74 year-old-man underwent surgery for the 

removal of a voluminous multinodular mass inglobating ileum, 

spleen, pancreas tail, omentum, round ligament of liver, gastric 

fundus and greater curvature. Four years before he was cystectomyzed 

because of an infiltrating bladder carcinoma. A diagnosis 

of extragastrointestinal stromal tumor (EGIST) with a probable 

great omentum origin was performed. 

Methods. Histochemistry and Immunohistochemical stains (IHC) 

were performed for α smooth muscle actin, CD117, CD34, CK 

293 

AE1/AE3, Desmin, GFAP, Ki-67, Neurofilament, NSE, PAS, S- 

100 protein, Sinaptofisin, Somatostatin and Vimentin. 

Results. Gastrointestinal stromal tumors (GISTs) are the most 

common mesenchymal tumors of the gastrointestinal tract. 

Recent studies revealed that GISTs are associated with gain-offunction 

mutations of c-kit gene and platelet derived growth factor 

receptor α gene (PDGRFA). The kit protein can be detected 

by immunohistochemical assays for CD117 antigen. GISTs are 

believed to be derived from interstitial cells of Cajal which are 

present in muscolar layer of gastrointestinal wall. Mesenchymal 

tumors of the omentum, mesentery and retroperitoneum with 

immunohistochemical features of the GISTs are classified as 

extragastrointestinal stromal tumors (EGISTs). The incidence of 

EGISTs is less than 10% of the GISTs group. EGISTs are histologically 

and immunohistochemically similar to their gastrointestinal 

counterpart. The size, cellularity, mitotic activity, age and 

location were reported as the most accurate predictors of adverse 

outcome. Our histologic examination showed spindle cells irregularly 

disposed with a mesenchimal origin. Immunohistochemically 

it was strongly positive for CD117, CD34, neurofilament 

and vimentin, but negative for desmin, S-100 protein, smooth 

muscle actin and p53. Although we found a low Ki-67 labelling 

index, the large size of the tumor and its multiple locations may 

predict an aggressive course. 

low expression of heparin-binding eGf-lIKe 

growth factor and epidermal growth factor 

receptor/erBB1 in human advanced melanoma 

1)Fondi C. 2)Benemei S. 3)Baroni G. 4)Innocenti S. 5)Apicella 

P. 6)Cecchi R. 7)Biancalani M. 8)Santucci M. 9)Massi D. 

1)Area Critica Med.Chir. Sez. Anatomia Patologica, A.O.U. Careggi, Firenze, 

Italia 2)Dip. Farmacologia Clinica E Preclinica, A.O.U. Cereggi, 

Firenze, Italia 3)Area Critica Med.Chir. Sez. Anatomia Patologica, A.O.U. 

Careggi, Firenze, Italia 4)Unità Di Patologia, Usl 3, Pistoia, Italia 5)Unità 

Di Patologia, Usl 3, Pistoia, Italia 6)Unità Di Dermatologia, Usl 3, 

Pistoia, Italia 7)Unità Di Patologia, Empoli, Firenze, Italia 8)Area Critica 

Med.Chir. Sez. Anatomia Patologica, A.O.U. Careggi, Firenze, Italia 

9)Area Critica Med.Chir. Sez. Anatomia Patologica, A.O.U. Careggi, Firenze, 

Italia 

Background. The identification of molecules involved in the 

process of melanoma progression is critical for the development 

of novel therapies and give the therapeutic breakthrough that has 

been long overdue. Heparin-binding epidermal growth factor-like 

growth factor (HB-EGF) is a member of the EGF growth factor 

family and is a potent stimulator of proliferation and migration 

in a variety of cells. Among membrane-anchored growth factors, 

HB-EGF is of special interest because it exhibits dual specificity 

for EGFR and ErbB4functions as a receptor for Diphtheria Toxin 

(DT). The potential contribution of HB-EGF/EGFR in melanoma 

immunobiology is presently unclear. 

Methods. Paraffin sections from 52 melanoma samples were 

immunohistochemically analyzed for HB-EGF and EGFR expression 

using a standard avidin-biotin technique. The series 

included: in situ melanomas (n = 6), primary invasive melanomas 

(n = 32), and melanoma metastases (n = 14). Cytoplasmic 

and nuclear HB-EGF staining as well as EGFR cytoplasmic and 

membranous staining were separately assessed. 

Results. Mean age of patients with invasive melanomas was 63.3 

years; 19 patients were females and 27 males. Tumors occurred 

on the head and neck (n = 1), extremities (n = 13), trunk (n = 17), 

acral region (n = 1). Overall, melanomas samples did not show 

evidence of nuclear HB-EGF staining. The proportion of melanomas 

showing cytoplasmic HB-EGF positivity in > 50% of cells 

significantly decreased from in situ melanoma, invasive melanomas 

and metastases (83.3% vs. 43.7% vs. 21.4%, p = 0.007 

Fisher’s exact test). Membranous EGFR staining was observed 

only in 4/52 invasive melanoma samples, with weak or moder-

294 

ate intensity. Lack of EGFR and nuclear HB-EGF expression in 

advanced melanomas do not support increased HB-EGF/EGFRmediated 

signaling in melanoma tumor progression. Our findings 

limit the potential application of DT treatment as melanoma immunotherapy. 

Diagnostic accuracy of endoscopic-ultrasound 

guided fine needle aspiration (euS-fNA) cytology 

of solid and cystic lesions of the pancreas 

1)A. Fornelli, 2)P. Baccarini, 1)S. Lega, 3)C. Fabbri, 3)A.M. 

Polifemo, 4)E. Jovine, 4)M. Masetti, 4) F. Martuzzi, 4)N. Zanini, 

1)A. Bondi 

1)Anatomia Patologica, Dipartimento Oncologico, Ospedale Maggiore, 

Bologna, Italia; 2)Anatomia Patologica, Dipartimento Oncologico, Ospedale 

Bellaria, Bologna, Italia; 3)Chirurgia, Ospedale Bellaria, Bologna, 

Italia; 4)Chirurgia, Ospedale Maggiore, Bologna, Italia 

Background. Endoscopic ultrasound guided fine needle aspiration 

(EUS-FNA) is increasingly utilized in the work-up of 

pancreatic neoplasms. EUS represents a cost-effective tool in 

the management of all patients with pancreatic cancer through 

its ability to pre-operatively stage pancreatic cancers by imaging 

the neurovascular axis and to sample even small lesions with fine 

needle aspiration. 

According to the literature the diagnostic sensitivity varies from 

65% to 94% and the specificity is around 100%. False-negative 

and inadequate diagnoses are relatively common, thus the role of 

cytology for a patient who is supposed to have a radiologically 

resectable cancer is controversial. 

Methods. Pancreatic resection specimens examined between 

January 2008 and April 2010 were retrieved from the files of the 

Institute of Anatomic Pathology of Maggiore Hospital in Bologna 

and matched with EUS-FNA pre-operative cytology examined at 

the Institute of Anatomic Pathology of Bellaria Hospital in Bologna, 

where EUS examinations are currently carried out, often 

with the assistance of a cytopathologist for on-site interpretation. 

Results. A total of 112 pancreatic resection specimens were 

retrieved. Pre-operative cytology was taken in 51 out of 112 patients. 

The overall sensisitivity and specificity were respectively 

78% and 100%. The reported sensitivity was based on a calculation 

that includes suspicious and atypical cases as true positives; 

these were respectively 20% and 3%. Atypical cases were proved 

to be IPMN at histology; the positive predictive value of the suspicious 

category was 100%. 

No increased morbidity, such as acute pancreatitis or tumour 

seeding, was documented. 

Our data confirm the diagnostic efficacy of the EUS-FNA technique, 

which seems to facilitate timely diagnosis and management 

of the patients. 

Small cell neuroendocrine carcinoma of the right 

vocal cord 

1)Fornelli A. 2)Dall’Olio D. 3)Bondi A. 

1) Anatomia Patologica, Dipartimento Oncologico, Ospedale Maggiore, 

Bologna, Italia 2) Dipartimento di Neuroscienze, Ospedale Maggiore, 

Bologna, Italia 3)Oncologico, Ospedale Maggiore, Bologna, 

Italia 

Background. Small cell neuroendocrine carcinoma is a very 

rare laryngeal tumour occurring in less than 0,5% of cases. It 

usually presents in the 6th and 7th decades in heavy smokers, 

the supraglottis being the most common location. At least 

50% of the patients harbours cervical metastases at the time of 

presentation and the reported 5-year survival rate is 5%. It is 

morphologically identical to its pulmonary counterpart; neuroendocrine 

activity is easily demonstrated with immunohistochemical 

stains. As in other sites, it may be pure or associated 

with other patterns. 


Case report. The patient is a 64 year-old man, ex-smoker, with 

a history of cough and dysphonia of few months duration. At 

laryngoscopy the right vocal cord was mobile and showed an 

ulcerated lesion measuring 1 cm. Following the pre-operative 

diagnosis of squamous cell carcinoma the patient underwent subtotal 

laryngectomy with righ lateral neck dissection. At histology 

the tumour showed a superficial component of squamous cell 

carcinoma, representing less than 5% of the whole lesion, merging 

with moderate dysplasia of the superficial epithelium. It was 

composed of sheets of small pleomorphic cells with interspersed 

foci of necrosis. Mitoses were numerous, up to 50/10 high 

power fields. Neither infraglottic, nor controlateral involvment 

was present. The lateral neck dissection contained nine reactive 

lymph-nodes. Synaptophysin and CD56 were positive while 

chromogranin, CK20 and TTF-1 were negative. 

Conclusions. The presence of a minimal component of superficial 

squamous cell carcinoma, along with TTF-1 negativity, 

allowed us to rule out a metastatic disease from the lung. The 

limited stage of disease at the time of diagnosis (pT2 N0) represents 

a further peculiarity. Post-operative total body CT and PET 

scans were negative for metastatic deposits, thus no radiotherapy 

was administrated. The patient is alive with no evidence of recurrence 

6 months after surgery, however a longer follow-up is 

mandatory. 

Comparative study of different antibody clones to 

assess prognostic and predictive factors of breast 

cancer 

1)Fortunato C. 2)Colantoni A. 3)Chiesa F. 4)Spagnoli LG. 

5)Bonanno E. 

1)Anatomia patologica, Policlinico universitario tor vergata, Roma, Italia 

2)Anatomia patologica, Policlinico universitario tor vergata, Roma, Italia 

3), Roche diagnostics, Monza, Italia 4)Anatomia patologica, Policlinico 

universitario tor vergata, Roma, Italia 5)Anatomia patologica, Policlinico 

universitario tor vergata, Roma, Italia 

Background. Pathologists today are discriminating hormoneresponsive 

from non hormone-responsive breast cancers almost 

exclusively by immunohistochemical means. This has led to a 

search for novel antibodies whose application may facilitate a 

more tailored therapy, thereby avoiding unnecessary toxicity. 

Tissue microarrays (TMAs) have emerged as a high-throughput 

technology for protein evaluation in large cohorts of paraffin 

embedded tissues. The purpose of this study was to characterize 

the sensitivity and specificity of different antibodies for 

detection of prognostic and predictive markers by immunohistochemistry 

(IHC) of formalin-fixed paraffin breast carcinoma 

sections. 

Methods. We set up a TMA with 6 different tissue cores for each 

of the following phenotype of human breast cancer: ER + PR + 

HER2-, ER + PR-HER2-, ER-PR-HER2 +, ER-PR-HER2. Fortyseven 

TMA slides were stained using both automated Ventana 

Roche Bench Mark XT apparatus or performing manually reactions. 

The antibody clones tested were: estrogen SP1 (Ventana, 

Rabbit Monoclonal), 6F11 (Novocastra, Mouse Monoclonal), 

1D5 (Dako, Mouse Monoclonal); progesteron 1E2 (Ventana, 

Rabbit Monoclonal), 1A6 (Novocastra, Mouse Monoclonal), 

16 (Novocastra, Mouse Monoclonal), PgR636 (Dako, Mouse 

Monoclonal); Ki67: K2 (Ventana, Mouse Monoclonal), Mib1 

(Dako, Mouse Monoclonal) e 30-9 (Ventana, Rabbit Monoclonal); 

HER2: A4085 (Dako, Rabbit Policlonal), Pathway Her2 

(Ventana, Rabbit Monoclonal), Hercep Test (Dako, Rabbit 

Policlonal). The slides were digitalized and analyzed with iScan 

Bioimagene. 

Results. Anti estrogen SP1, anti progesteron 1E2, anti Ki67 

clone 30-9 and Pathway Her2 showed the highest scores by visual 

analysis. Likewise, these antibodies showed higher staining 

intensity by the automated method of analysis. This new antibod-


ies provided stronger and sharper nuclear immunohistochemical 

signals as compared with most commonly used antibodies, with 

no non-specific cytoplasm staining. 

Chromosomal abnormalities of the fetus and 

placenta: fish diagnostic usefulness 

1)Franceschetti I. 2)Piazzola E. 3)Brunelli M. 4)Martignoni G. 

1)Anatomia patologica, Ospedale San Bortolo, Vicenza, Italia 2)Anatomia 

patologica, Ospedale civile maggiore, Verona, Italia 3)Anatomia 

patologica, Policlinico G.B. Rossi, Verona, Italia 4)Anatomia patologica, 

G.B. Rossi, Verona, Italia 

Background. All clinical signs and features of the different 

syndromes correlated to chromosomal alteration are well described 

in literature, but the same elements are hardly assessable 

when performing a macroscopic examination of the fetus dead 

in uterus or of the abortive product. Therefore the presence of a 

fetal karyotype is extremely useful. In all sine causa fetal deaths, 

Fish technique could be of some help for a complete genetic 

counselling. Our scope is to evaluate the possible overlapping 

of FISH investigation on material in paraffin over the classical 

cytogenetics, the expression of the signals in the different organs, 

to find out the most suitable organs and tissues to this kind of 

investigation and to assess the possibility of introducing it in the 

diagnostics of the MEF, of the repeated spontaneous abortions 

and in the IUGR sine causa. 

Methods. We selected all abortion with genetic data present 

in our archive. We collected 46 cases, reviewed the slides and 

evaluated the degree of tissue maceration and autolysis, the 

entity of the sampling and the presence of both fetal and placental 

material. We chose 8 cases characterized by complete 

sampling and different types of chromosomal abnormality and 

we performed FISH using probes for chromosomes 18, X, Y, 

13 and 21. 

Results. About the signal reading, the best preparations are: 

lungs, kidneys, adipose tissue, muscular tissue, cartilage and 

placenta. In the case of probes X, Y, 18 the signal is complete for 

the major part of the nuclei (from 70 to 97% of the nuclei). In the 

case of probes 13,21the signal is weaker (from 55 to 100% of the 

nuclei). FISH has proved to be overlappable and more precise in 

the case of mosaicism. Several questions can be solved by FISH: 

most frequent chromosomopathies in abortive material; in MEF; 

presence of material from Y chromosome in Turner’s syndrome; 

placental mosaicism in IUGR cases sine causa. 

role of fine needle cytology in the diagnosis 

of lymphnode metastases in a cancer center. 

experience with 661 cases in three years (2007- 

2009) 

1)Fulciniti F. 2)Butera D. 3)Staiano M. 4)La Vecchia F. 5)Curcio 

M.P. 

1)S.S.D. Di Citopatologia, A.F. Di Anat. Patol., Istituto Nazionale Tumori 

“Fondazione G. Pascale”, Napoli, Italia 2)S.S.D. Di Citopatologia, A.F. 

Di Anat. Patol., Istituto Nazionale Tumori “Fondazione G. Pascale”, Napoli, 

Italia 3)S.S.D. Di Citopatologia, A.F. Di Anat. Patol., Istituto Nazionale 

Tumori “Fondazione G. Pascale”, Napoli, Italia 4)S.S.D. Di C, Istituto 

Nazionale Tumori “Fondazione G. Pascale”, Napoli, Italia 5)S.S.D. Di 

C, Istituto Nazionale Tumori “Fondazione G. Pascale”, Napoli, Italia 

The cytologic reports in which a lymphnode metastasis had been 

diagnosed by Fine Needle Cytology (FNC) in the last 3 years 

were collected and analyzed in order to verify the diagnostic accuracy 

of the method and its informative value with respect to 

the clinician and patients. The series consisted of 661 cases (338 

Males, 323 Females, median age 63, range 14-90). A previous diagnosis 

of malignancy was known in 491/661 cases (74%), while 

295 

the site of the primary tumor was correctly predicted by FNC in 

the remaining 177(24%) cases with cyto-histologic correlation. 

15 lymphnode metastases (0.02%) were confirmed histologically 

but a primary site was never found during the diagnostic work-up. 

A cytological diagnosis preceeded and correctly oriented the diagnosis 

of the primary tumor in 26/72 (36%) cases of thyroid carcinoma, 

11/63 (17%) cases of oro/rhynopharyngeal carcinoma, 

14/122 breast carcinomas (11%), 9/87 (10%) lung carcinomas, 

26/175 (14%) melanomas, 13/48 (27%) of the gastro-intestinal 

tract carcinomas, 4/33 (12%) of endometrial carcinomas, 7/17 

(41%) ovarian carcinomas, 7/17 (41%) genito-urinary tract carcinomas. 

A timely and correct cytological report had an important 

role in the correct diagnostic work-up and therapeutic management 

of out- and inpatients with known and previously unknown 

history of malignant neoplasms. 

Analysis of immunoglobulin gene rearrangements 

with BIOMeD-2 multiplex PCr protocol in lymphoid 

proliferations 

1)Gafà R. 2)Maestri I. 3)Ulazzi L. 4)Cappella ED. 5)Cavazzini 

L. 6)Nenci I. 

1)Unità Operativa di Anatomia Patologica, Azienda Ospedaliero-Universitaria 

di Ferrara, Ferrara, Italia 2)Unità Operativa di Anatomia 

Patologica, Azienda Ospedaliero-Universitaria di Ferrara, Ferrara, 

Italia 3)Unità Operativa di Anatomia Patologica, Azienda Ospedaliero- 

Universitaria di Ferrara, Ferrara, Italia 4)Unità Operativa di Anatomia 


Italia 5)Unità Operativa di Anatomia Patologica, Azienda Ospedaliero- 

Universitaria di Ferrara, Ferrara, Italia 6)Unità Operativa di Anatomia 


Italia 

Background. The differential diagnosis between malignant B 

cell lymphoma and reactive proliferations can be challenging. In 

such cases analysis of immunoglobulin (Ig) gene rearrangements 

is a useful additional diagnostic tool. 

Methods. Analysis of IgH and IgK gene rearrangements was performed 

on 60 formalin-fixed paraffin-embedded tissue samples 

from 48 patients with various lymphoid disorders using commercially 

available kits according to the BIOMED-2 multiplex 

PCR approach. The PCR products were analyzed using the Gene 

Mapper software after capillary electrophoresis. 8 samples were 

excluded because of poor quality DNA. The remaining 52 cases 

included: 23 nodal and extranodal B-cell lymphomas (defined 

according to the WHO 2008 classification), 3 suspected B-cell 

lymphomas, 3 atypical B-cell proliferations, 5 reactive lymphadenopaties, 

10 gastric samples with suspicious lymphoid infiltrate 

and 8 lymphoid proliferations including Kikuchi lymphadenitis, 

progressively transformed germinal centres, Castleman disease, 

skin B-cell pseudolymphoma. 

Results. In all the B-cell lymphomas, the suspected B-cell 

lymphomas and the atypical B-cell proliferations presence of 

unequivocal and reproducible clonal products in multiple loci was 

demonstrated. Reactive lymphadenopaties resulted polyclonal 

in every locus and tube. Four samples with suspicious gastric 

lymphoid infiltrates resulted monoclonal and all 4 patients subsequently 

developed a MALT lymphoma clonally related to the 

previous lymphoid proliferation. Only 1 of the three skin pseudolymphomas 

turned out to be monoclonal and the remaining 

lymphoid proliferations resulted clearly polyclonal with all the 

tubes examined. 

Conclusion. In our experience analysis of immunoglobulin gene 

rearrangements with the BIOMED-2 protocol is reliable and 

reproducible with high sensitivity and specificity. Furthermore it 

can detect clonal relationship between lymphomatous processes 

that are separated anatomically and/or chronologically.

296 

Analysis of microsatellite instability and dna 

mismatch repair protein expression in endometrial 

carcinoma 

1)Gafà R. 2)Maestri I. 3)Ulazzi L. 4)Lanza G. 

1)Sezione di Anatomia Patologica, Università di Ferrara, Ferrara, Italia 




Background. Microsatellite instability (MSI) occurs in sporadic 

and hereditary endometrial carcinoma (EC). In both cases a functional 

inactivation of the DNA mismatch repair (MMR) genes 

with lack of protein expression is seen. Aim of this study was 

to evaluate MSI and MMR protein expression in a consecutive 

series of ECs. 

Methods. The study includes 123 patients with EC (age, 39-83 

years). Tumors were predominantly of endometrioid type (87.8%) 

and FIGO stage 1 (69.1%). In all tumors MMR protein expression 

(MLH1, MSH2, MSH6) was evaluated by immunohistochemistry 

using commercially available monoclonal antibodies. MSI was 

analyzed on DNA obtained from formalin-fixed paraffin-embedded 

tumor tissue samples by a fluorescence-based PCR method 

using the five Bethesda panel markers and BAT40. Tumors were 

classified as MSI-H, MSI-L and MSS according to the Bethesda 

guidelines. 

Results. MSI-H was observed in 37 tumors (30.1%), whereas 4 

tumors (3.2%) were classified as MSI-L and 82 (66.7%) as MSS. 

Within the 114 samples with reliable immunohistochemistry, 76 

(66.7%) showed normal protein expression (MMRP+) and 38 

(33.3%) demonstrated loss of at least one protein (MMRP-). In 

detail, 29 tumors showed complete loss of MLH1 expression, 

4 tumors displayed loss of MSH2 and MSH6 expression and 5 

tumors showed selective loss of MSH6 protein. There was a statistical 

correlation between the two methods (p < 0.001). In fact 

of the 76 MMRP+ tumors 74 were MSS, 1 was MSI-L and 1 was 

MSI-H. Conversely, 33 of the 38 MMRP- tumors were MSI-H, 

3 were MSI-L and 2 were MSS. No significant association was 

found between MMR and MSI status with patient’s age, histologic 

type, tumor grade and stage. 

Conclusion. Our results indicate that MMR deficit occurs frequently 

in EC and that a significative number of ECs develops 

on hereditary basis due to alterations of MSH2 and MSH6 genes. 

Unlike colorectal cancer, MSI-L ECs frequently show loss of 

MMR protein expression. 

embryonal tumors with abundant neuropil and 

true rosettes (eTANTr): unusual histopathological 

findings 

1)Gardiman Mp. 2)Fassan M. 3)Danieli D. 4)D’Amore Egs. 

5)Opocher E. 6)Faggin R. 7)Perilongo G. 8)Rugge M. 

1)Medical Diagnostic Sciences & Special Therapies, University Of Padova, 

Padova, Italia 2)Medical Diagnostic Sciences & Special Therapies, 

University Of Padova, Padova, Italia 3)Pathology Unit, S.Bortolo Hospital 

Vicenza, Vicenza, Italia 4)Pathology Unit, S.Bortolo Hospital Vicenza, 

Vicenza, Italia 5)Department Of Pediatrics, University Of Padova, Padova, 

Italia 6)Department Of Neurosurgery, University Of Padova, Padova, 

Italia 7)Department Of Pediatrics, University Of Padova, Padova, Italia 


Padova, Italia 

Background. Embryonal Tumor with Abundant Neuropil and 

True Rosettes (ETANTR) constitute a distinctive entity inside 

the group of Central Nervous System (CNS) Primitive Neuroectodermal 

Tumors (PNET). Most of ETANTR occur in children 

younger than 3 years of age, arising in the posterior fossa region 

or in the frontal lobes. Prognosis of ETANTR is very poor with 

an overall survival of less than 2 years. 


Cases presentation. We present 4 cases of ETANTR, 2 of them 

with unusual histologic characteristics. Usually these neoplasms 

are characterized by the presence of undifferentiated neuro-epithelial 

cells with broad zones of well-differentiated neuropil and 

ependymoblastic rosettes arising abruptly from paucicellular 

regions of neoplastic neuropil. Immunohistochemical reactions 

show a negative stain for GFAP, neurofilament and synaptophysin 

in the undifferentiated blue cells while the neuropil is positive 

for synaptophysin. In one of our cases, a sarcomatous, reticulinrich 

component of elongated spindle cells showing diffuse immunoreactivity 

for smooth muscle actin and focal desmin staining 

was present, a combination of large “anaplastic” elements 

with round to oval highly pleomorphic nuclei, as it is seen in 

anaplastic medulloblastoma, and pseudo-stratified glandular-like 

and epythelial-like cells aggregate in cord or tubular strucutures, 

reminding a medulloepythelioma, was found in another case. 

An abundant neuropil background with true ependymoblastic 

rosettes confirmed the ETANTR diagnosis among both of these 

two unusual cases. 

Conclusions. These peculiar hystological features of ETANTR 

suggest these neoplasms are undifferentiated neuroepithelial tumours 

which may display divergent differentiation towards other 

type of embryonal tumours or mesenchimal component with 

acquisition of sarcomatous phenotype. 

Congenital brain tumours: a report of four cases 

1)Gardiman Mp. 2)Fassan M. 3)Mannicci D. 4)D’Avella D. 

5)Calderoni M. 6)Perilongo G. 7)Rugge M. 


Padova, Italy 2)Medical Diagnostic Sciences & Special Therapies, 

University Of Padova, Padova, Italy 3)Gynecological Sciences And Human 

Reproduction, University Of Padova, Padova, Italy 4)Neurosurgery, 

University Of Padova, Padova, Italy 5)Neuroradiological Service, University 

Of Padova, Padova, Italy 6)Pediatrics, University Of Padova, Padova, 

Italy 7)Medical Diagnostic Sciences & Special Therapies, University Of 

Padova, Padova, Italy 

Background. Congenital Central Nervous System Tumors (SNC) 

(i.e. tumours relate to diagnosis during the fetal life or in the first 

28 days of life) are now more often diagnosed through the more 

widespread and accessible neuroimaging. Malignant histology 

is common with persistently poor outcomes despite evolving 

adjuvant therapy. 

Methods. A retrospective review was performed of paediatrics tumours 

surgically treated at Paediatric Oncology Department of the 

University Hospital of Padova over a 10-year period (1999-2009). 

Results. Four cases of congenital SNC tumors were identified; 

all of them were supratentorial. There was hydrocephalus in 2 

cases and all cases presented with an increasing head circumference. 

Histology revealed one case of ETANTR (Embrional 

Tumor with Abundant Neuropil and True Rosettes), two cases 

of glioblastoma multiforme (GBM), and one case of teratoma. 

After the birth, the patient with ETANTR and the two with GBM 

underwent surgical resection. The infant with ETANTR and one 

of those with GBM died within few days after the diagnosis. The 

other patient with GBM received chemotherapy and he is tumor 

free 30 months after the end of therapy with good outcome. The 

child with teratoma died within the first post-natal week and an 

autopsy was performed after death. Histological examination revealed 

a huge teratoma of the suprasellar region with mature and 

immature component. 

Conclusions. Central Nervous System tumors in infants are 

suggestive of some oncogenic prenatal factors. Scant literature 

remains about management and ethics of the treatment of babies 

diagnosed at this very fragile stage of development.


Pleomorphic xanthoastrocytoma with multiple 

localizations and subdural dissemination: case 

report and review of the literature 

1)Gardiman MP. 2)Iaria L. 3)Calderoni M. 4)Faggin R. 5)Perilongo 

G. 6)Rugge M. 


Padova, Italy 2)Medical Diagnostic Sciences & Special Therapies, 

University Of Padova, Padova, Italy 3)Neurosurgery, University Of Padova, 

Padova, Italy 4)Neurosurgery, University Of Padova, Padova, Italy 

5)Pediatrics, University Of Padova, Padova, Italy 6)Medical Diagnostic 

Sciences & Special Therapies, University Of Padova, Padova, Italy 

Background. Pleomorphic xanthoastrocytoma (PXA) is a rare 

astrocytic tumor that usually occurs in the superficial cerebral 

hemispheres of children and young adults and has usually a favorable 

prognosis. We report on an unusual case of multicentric 

PXA with subdural dissemination. 

Case presentation. A 14-year-old girl was admitted to Paediatric 

Neurosurgery Department of the Hospital University of Padua 

with a recent history of sciatic pain syndrome and atactic gait. 

MRI revealed multiple lesions: two large cystic lesions in the 

cerebellar vermis and in the left cerebellar hemisphere, a solid 

intramedullary enhancing nodule at the dorsal level with diffuse 

enlargement of the spinal cord with extensive edema and 

multiple sub-meningeal nodules in both acoustic inner meati and 

in the sellar region. There was also a diffuse enhancement along 

the cerebellar and brainstem surface. After surgery, histological 

examination showed spindle cells intermingled with pleomorphic 

mono- or multinucleated giant cells. We observed abundant eosinophilic 

granular bodies, Rosenthal fibers and perivascular lymphoid 

infiltration, and absence of necrosis. The tumor cells were 

surrounded by basement membranes that stained positively for 

reticulin. Glial fibrillary acid protein (GFAP) confirmed the astrocytic 

origin, whereas synaptophysin and neurofilament stains 

were negative. MIB1 showed a very low proliferation index 

(< 3%). The pathologic diagnosis was multicentric pleomorphic 

xanthoastrocytoma with subdural dissemination. 

Conclusions. To our knowledge, this is the third case of multicentric 

PXA with disseminated disease reported in the literature. 

Hence, it was considered important to describe this case in order 

to add further information regarding the spectrum of the presenting 

clinical features of this rare neoplasm. 

The “masters” project (massa carrara advanced 

screening technologies evaluation in routine 

setting): a preliminary report 

1)Gatteschi S. 2)Pieraccini L. 3)Nicolai C. 4)Pietrini F. 5)Tozzini 

S. 6)Lombardi S. 7)Bertacca G. 8)Casalini S. 9)Migliorini P. 

10)Cavazzana A. 

1)Oncologia, Asl1 Massa E Carrara, Carrara, Italia 2)Oncologia, Massa 

E Carrara, Carrara, Italia 3)Oncologia, Massa E Carrara, Carrara, Italia 

4)Oncologia, Massa E Carrara, Carrara, Italia 5)Oncologia, Massa E 

Carrara, Carrara, Italia 6)Diagnostica, Massa E Carrara, Massa, Italia 

7)Diagnostica, Massa E Carrara, Massa, Italia 8)Oncologia, Massa E 

Carrara, Carrara, Italia 9)Materno-Infantile, Massa E Carrara, Massa, 

Italia 10)Oncologia, Massa E Carrara, Carrara, Italia 

Background. Cervical Cancer Screening programs based on Paptest 

cytology are largely operator dependant. HPV genotyping 

and p16 immunocytochemical detection may help to increase the 

level of accuracy of cytologic diagnoses. 

Methods. Women in screening age interval living in Massa-Carrara 

province are enrolled in this project. All cervical specimens 

are processed with the LBC Thinprep method through a TP 3000 

Processor. All positive samples (Asc-us to Carcinoma) are tested 

for HPV infection, through PCR technique and pyro-sequencing, 

and for p16 over-expression (CINtec cytology). 5% of negative 

re-screened Pap-test samples are also tested. Biopsy are routinely 

stained with the p16 CINtec histology test. 

297 

Results. At present, 191/3420 screened LBC samples (136 positive 

and 55 negative cases) were enrolled in the study. HPV and 

p16 results are available for 118 positive and 14 negative cases 

respectively; confirmatory biopsies are available in 29 cases. 91% 

(107/118) of positive pap-tests showed an HPV infection; 65,4% 

(70/107) of HPV+ cases with positive cytology were High risk 

genotypes while 30% of cases were LR and undetermined risk 

(UR). The p16 nuclear over-expression was detected in 19,5% of 

cases (36/184) with a higher rate in the HR HPV than in LR and 

UR HPV groups [30% vs 13%]. Interestingly, 92% (12/13) of cytological 

p16 + cases showed high grade histological confirmed 

lesions (CIN II-III) whereas cytology (High-SIL) alone identified 

50% of these lesions (8/16). P16 PPV (predictive positive value) 

to identify high grade lesions was 92%, whereas NPV (negative 

predictive value) was 71.4%. 

Preliminary conclusions. HPV genotyping and p16 immunocytochemistry 

are easily accessible routine tests in a LBC setting. 

P16 testing seems to improve the cytology diagnostic accuracy in 

detecting squamous high grade lesions. 

CDKN1B/P27 expression in peripheral T-cell 

lymphoma not otherwise specified 

Anna Gazzola 1 , Maria Teresa Sista 1 , Claudio Agostinelli 1 , Simona 

Righi 1 , Maria Rosaria Sapienza 1 , Claudia Mannu 1 , Maura 

Rossi 1 , Francesco Bacci 1 , Elena Sabattini 1 , Philip Went 2 , Pier 

Luigi Zinzani 1 , Stefano A. Pileri 1 *, Pier Paolo Piccaluga 1 * 

1 Department of Haematology and Oncology “L. and A. Seràgnoli”, Haematopathology 

and Haematology Units, S. Orsola-Malpighi Hospital, 

University of Bologna, Bologna, Italy; 2 Institute of Pathology, 8063 Zürich, 

Switzerland; *SAP and PPP equally contributed to this work 

Background. Peripheral T-cell lymphoma not otherwise specified 

(PTCL/NOS) is the commonest T-cell lymphoma. Recently, gene 

expression profiling (GEP) allowed the identification of PTCL/ 

NOS-associated molecular signatures, leading to better understanding 

of their histogenesis, pathogenesis and prognostication. In particular, 

proliferation control turned out to be strongly affected, being 

a high proliferation index associated to unfavourable prognosis. 

In this study we aimed to evaluate the possible occurrence of 

CDKN1B/p27 aberrations in PTCL/NOS, as CDKN1B/p27 is 

a main regulator of proliferation and is often involved in lymphomagenesis. 

Methods. We studied CDKN1B/p27 expression at RNA and protein 

level, by DNA- and tissue-microarrays, in 28 and 98 cases, 

respectively. Additionally, we performed direct sequencing of 

CDKN1B in 81 PTCLs/NOS. 

Results. We found that CDKN1B mRNA is similarly expressed 

in PTCL/NOS and normal T-lymphocytes. In addition, we did not 

found structural abnormalities, including mutations, deletions or 

possible SNPs, in any exons, exon-intron junction or regulatory 

region. Similarly, the CDKN1B locus was found to be intact in 

the course of a high density karyotyping study carried out by the 

Affymetrix 250k SNPs arrays in a series of PTCLs/NOS. Furthermore, 

we demonstrated by immunohistochemistry physiological 

expression of p27 in neoplastic cells, which was mutually exclusive 

with Ki-67, as expected when the system is intact. Interestingly, 

on the other hand, 6 out of 98 PTCLs/NOS presented with 

p27 positivity and Ki67 ≥ 80%; however, a double immunofluorescence 

staining showed that the two molecules were never really 

co-expressed revealing p27 physiological activity. In addition, 

we studied the expression of other molecules which are functionally 

related to CDKN1B/p27 in controlling cell cycle (including 

CCNE1); notably, they did not appear to be affected at either 

mRNA or protein level. Finally, we found that p27 expression did 

not appear to be significantly related with overall survival, though 

a possible favorable trend was recorded in p27 positive cases. 

Conclusion. In conclusion, CDKN1B/p27 aberrations seem to be 

uncommon in PTCL/NOS pathogenesis.

298 

IDH-1/IDH-2, TP53, and C-MYC/N-MYC status in 10 

cases of supratentorial PNeT (S-PNeT) in adult 

patients 

1)M. Gessi, 2)M. Bisceglia, 3)L. Lauriola, 1)A. Waha, 4)F. 

Giangaspero, 1)T. Pietsch 

1)Institute of Neuropathology, University of Bonn Medical Center, Bonn, 

Germany; 2)Dipartimento di Patologia, IRCCS Casa Sollievo della Sofferenza, 

S. Giovanni Rotondo, Italia; 3)Istituto di Anatomia Patologica, 

Università Cattolica, Roma, Italia; 4)Dipartimento di Medicina Sperimentale 

e Anatomia Patologica, Università di Roma “La Sapienza”, 

Roma, Italia 

Background. Advances in understanding the molecular basis of 

CNS-PNET biology are critical to improve patient’s outcome. 

Recently, new data on their molecular features have been reported, 

suggesting that s-PNET in adult patients may represent 

a specific tumor entity among the CNS-PNET. In this view, we 

evaluated the mutational status of IDH-1/IDH-2 and TP53 as 

well as the c-myc and N-myc amplification status in 10 cases of 

adult s-PNET. 

Methods. After DNA extraction from paraffin embedded tissue, 

10 cases of adults s-PNET (median age 54.6 years), were 

screened for mutation of TP53 (exon 4 > exon 9) as well as 

IDH-1 (codon 132) and IDH-2 using SSCP-based and sequencing 

assays. The cases presenting shifts anomalies in the SSCP-assay 

were cloned and sequenced. To quantify c-myc and N-myc gene 

copy numbers, we utilized a quantitative PCR-based assay using 

exogenous DNA standard competitors. 

Results. The SSCP-based mutational screening and sequencing 

of the TP53 gene revealed five different point mutations affecting 

exons 4, 5, 7, 8. In two cases, a mutation at codon 132 of the 

IDH-1 gene has been also found. No mutations of IDH-2 gene as 

well as c-myc or N-Myc amplifications have been found. 

Conclusion. Our data, showing a high incidence of TP53 and 

IDH-1 mutations and the absence of amplification of the c-myc 

and N-myc genes, strengthen the hypothesis that adult s-PNETs 

may represent a specific subset of tumor among the CNS–PNET, 

differentiating also them from their paediatric counterpart. 

Serum levels of CA 15-3, CA 125 and CA 19.9 in 

triple negative breast cancer at time of diagnosis: 

preliminary report 

1)Giacometti C. 2)Marchesini C. 3)Callea M.R. 4)Zanin E. 5)Zuliani 

M. 6)Baseggio C. 7)Busolin R. 8)Pasini L. 

1)Patologia Clinica, Casa Di Cura Giovanni Xxiii, Monastier Di Treviso, 

Treviso, Italy 2)Patologia Clinica, Casa Di Cura Giovanni XXIII, Monastier 

Di Treviso, Treviso, Italy 3)Patologia Clinica, Casa Di Cura Giovanni 

XXIII, Monastier Di Treviso, Treviso, Italy 4)Patologia Clinica, Casa 

Di Cura Giovanni XXIII, Monastier Di Treviso, Treviso, Italy 5)Patologia 

Clinica, Casa Di Cura Giovanni XXIII, Monastier Di Treviso, Treviso, 

Italy 6)Patologia Clinica, Casa Di Cura Giovanni XXIII, Monastier Di 

Treviso, Treviso, Italy 7)Chirurgia Generale, Casa Di Cura Giovanni 

Xxiii, Monastier Di Treviso, Treviso, Italy 8)Patologia Clinica, Casa Di 

Cura Giovanni XXIII, Monastier Di Treviso, Treviso, Italy 

Introduction. Triple-negative breast cancers (TNBC) are negative 

for estrogen receptor, progesterone receptor, and the human 

epidermal growth factor receptor 2 (HER2neu) and have worse 

prognosis. Cancer antigen 15-3 (CA 15-3) is often used in follow-up 

care of breast cancer and provide important clues to the 

clinicians for disease progression in metastatic and recurrent 

breast cancer. 

Methods. We retrospectively analyzed serum CA 15-3, CA 

125 and CA 19.9 levels (DxI 800, Beckman Coulter) at time of 

primary surgery in 51 consecutive non-metastatic breast cancer 

patients presenting at Casa di Cura Giovanni XXIII (Monastier 

di Treviso, Treviso, Italy) between November 2008 and October 

2009. The 13.7% of the patients (7/51) were TNBC. The 

negative control group consisted of 51 women with histologi- 


cally proven benign breast disease (27 fibroadenomas – FAs; 

8 papilloma/papillomatosis and 16 miscellaneous (fibrocystic 

disease – FCD, ductal hyperplasia/atypical ductal hyperplasia 

– DH/ADH). 

Results. TNBC group have higher histological grade (mean ± SD 

3 ± 0 vs 2.36 ± 0.61, p = 0.01) and higher serum levels of CA 

15-3 and CA125 compared to non-triple negative group. Patients 

with breast cancer were statistically significantly older 

than patients with benign disease (mean age yrs ± SD, range: 

56.5 ± 11.7, 32-88 vs 46.9 ± 12.7, range 21-77, p = 0.0001). No 

statistically significant difference were noted between breast cancer 

group and benign breast disease group in the serum levels of 

the CA 15-3 and CA 19.9, while CA 125 was significantly higher 

in benign disease group. 

Conclusions. The difference in serum levels of TNBC may be 

partly explained by highest histological grade of TNBC and serum 

levels of CA 15-3 and CA 125 may differentiate this cancer 

subgroups. Our findings need to be confirmed in further studies, 

in order to define the possible role of combined use of multiple 

serum markers in the individuation of particular breast cancer 

subgroups. 

Her2 in gastric cancer: a comparative study of 

protein expression and gene amplification in 

gastric biopsy and corresponding surgical samples 

1)Giannatiempo R. 2)Baron L. 3)Franco R. 4)Postiglione M. 

1)U.O.S. Anatomia Patologica, Ospedale Evangelico Fondazione Betania, 

Napoli, Italia 2)S.O.C. Anatomia Ed Isrtologia Patologica, P.O. San 

Leonardo, Castellammare Di Stabia, Italia 3)A.F. Anatomia Patologica, 

I.N.T. Fondazione G. Pascale, Napoli, Italia 4)U.O.S. Anatomia Patologica, 

Ospedale Evangelico Fondazione Betania, Napoli, Italia 

Background. No targeted modality of treatment has so far been 

incorporated to gastric cancer (GC) strategy of therapy. Her2 

overexpression is increasing recognized as a frequent molecular 

abnormality, driven as in breast cancer by gene amplification. 

Aim. Evaluating the frequency of Her overexpression by IHC and 

its concordance with gene amplification by FISH in surgical and 

endoscopic biopsy GC specimens using two commercial kits. 

Materials and methods. Sections of routine endoscopic biopsies 

and the subsequently surgically resected whole tumors of 84 GC 

with underwent surgery. 

IHC analysis was performed using Pathway Her2,Ventana and 

the expression grading were extimated according to FDA approved 

scoring system for breast cancer;Her2 gene amplification 

was evaluated using PathVysion Her2 DNA Probe kit (Vysis, 

dual color). 

Results. Among these 84 surgical specimens, 20 GC(23.8%) 

were evaluated as Her2 overexpression. The following IHC 

scores were obtained:0,52(62%); 1+,12(14%); 2+,6(7%) and 

3+,14(17%). 

Gene amplification was observed in 18 GC(21.4%). The concordance 

rate IHC/FISH in our surgical serie GC was 79% (50% for 

2+ and 86% for 3+). 

For 84 endoscopic biopsy specimens, Her2 protein was demonstrated 

in 22.6%(19 GC). The IHC score for biopsy specimens 

was 0 in 50 GC(59.5%), 1+ in 15 GC(18%), 2+ in 8 GC(9.5%), 

3+ in 11 GC(13%). 

Gene amplification was observed in 17 GC(20.2%).The concordance 

rate IHC/FISH in our biopsy serie GC was 78% (50% for 

2+ and 91% for 3+). 

Both Her2 overexprssion and Her2 gene amplification rates were 

similar in specimens obtained by surgery and biopsy (20/84;23.8 

vs.19/84;22.6) and (18/84; 21.4 vs.17/84;20.2). 

Her2 positivity differed significantly by tumor location(16% 

stomach vs.30% gastro-oesophageal junction) and histological 

type according to Lauren’s classification (intestinal 28%, diffuse 

6%, mixed type 15.5%).


Conclusions. IHC/FISH differences were largely due to FISH 

positive GC that were IHC 0/1+. Furthermore IHC/FISH discrepancies 

were attributed to basolateral membrane staining of 

glandular cells(resulting in incompletely stained membranes) as 

well as a higher % of heterogeneous tumors in GC in comparison 

with breast cancer. 

epidermal growth factor receptor gene mutations 

in thyroid carcinomas 

Capodanno A., Giannini R., Torregrossa L., Proietti A., Basolo F. 

Department of Surgery, Santa Chiara Hospital, Pisa, Italy 

Background. The epidermal growth factor receptor (EGFR) is 

involved in cancer development and progression and is frequently 

overexpressed in a variety of human cancers representing an 

attractive target for selective anticancer therapy. The response 

rate to EGFR inhibitors in non small-cell lung cancer is strongly 

associated with somatic activating mutations in the EGFR tyrosine 

kinase (TK) domain. EGFR and EGF have been detected in 

thyroid carcinomas (TCs) and they have been proposed to play a 

key role in TC proliferation and progression. Moreover, a high 

frequency (30.4%) of EGFR mutations has been recently found 

in papillary TC (PTC) of Japanese patients. 

Methods. In the present study, we investigated the EGFR mutational 

status (exons 18-21) by direct gene sequencing analysis in 

98 TCs, including 40 well differentiated (30 PTC and 10 follicular 

TCs, FTC), 40 poorly differentiated (PDC) and 18 anaplastic 

(ATC) TCs. 

Results. The frequency of EGFR gene mutation was 18/98 

(18.4%) with 14 mutations in the exon 19 and 4 in the exon 

21. In detail, mutations were found in 4/30 (13.3%) of PTC, 

5/10 (50%) of FTC, 6/40 (15.0%) of PDC and 3/18 (16.7%) of 

ATC. All the mutations found in PTC were the in-frame deletion 

E746_E750delELREA in the exon 19 that is the most common 

drug-sensitizing mutation. The E746_E750delELREAinsVP in 

the exon 19 was the only mutation found in FTC. While in well 

differentiated TCs all the mutations involved exon 19 of the EG- 

FR gene, in PDC and ATC we observed both exon 19 deletions 

(4 E746_E750delELREA and 1 E746_E750delELREAinsP) and 

exon 21 missense mutations (T847I, L858R, P848S, E829K/ 

V834A/G857E). Our results confirm the high frequency of the 

EGFR-activating mutations in thyroid cancer suggesting an important 

role of EGFR in thyroid tumorigenesis. Moreover, these 

findings might offer a new strategy for the treatment of the more 

aggressive thyroid cancer histotypes or of the tumors that are 

resistant to the conventional therapies. 

ultrasound technology in rapid tissue processing 

D.Gusolfino, R. Clarini, R. Giardini 

U.O di Anatomia e Istologia Patologica e Citodiagnostica, Istituti Ospitalieri 

di Cremona, Italia 

Background. An acceptable compromise between high level 

quality of histological slides and short answer time is always a 

challenge for Pathologists. Rapid tissue processing is required in 

the evaluation of organs transplantation and surgery specimens 

and to reduce answer time in routine evaluations. High technology 

tools are now available to short the answer time without 

compromise the quality of the specimens. Aim of this study is 

to evaluate the ability of rapid tissue processor HistraQS, using 

ultrasound, to achieve these goals. 

Methods. In the Pathology Unit of Cremona Hospital 20 non 

fixed specimens from breast cancer were processed with HistraQS. 

Immunoistochemical (IIC) was performed in tissue biopsies 

fragments 3 to 5 mm thick and the results compared with 

parallel specimens conventionally evaluated to check the reliability 

of the new technique. 

299 

Results. Only 58 minutes were required to process specimens and 

obtain a diagnosis on HE slides. IIC analyses of prognostic and 

predictive factors (ER, PgR, and Her2) were better evaluated in 

the thinner specimens (3 mm). Between the 20 thinner biopsies, 

18/20 showed a strong nuclear reaction to ER and PgR receptors 

comparable to the conventional procedure. The remaining two 

biopsies showed some artefact reactions probably due to incomplete 

fixation and deidratation. The evaluation of the thicker 

specimens (> 5 mm) showed a lower percentage of reliable 

results, suggesting a better performance of thinner specimens. 

The Her 2 membrane receptor, more sensible to fixation defects 

compared to nuclear receptors, showed positive results in about 

half of the thinner slides. 

Conclusions. The HistraQS is a reliable and reproducible system 

to reduce specimens processing time, useful to allow rapid diagnosis 

in thin biopsies. An “intra-day” organization of diagnostic 

processes can be achieved optimizing the processing technique. 

The fields of interest could include not only breast biopsies, but 

also other very important chapters of oncological pathology such 

as prostate or bronchial biopsies. 

Clinico-pathological implications of the epidermal 

growth factor receptor, COX-2 expression and 

HPV status in the squamous cell carcinoma of the 

uterine cervix in elderly 

1)Giordano G. 2)D’Adda T. 3)Dal bello B. 4)Bersiga A. 5)Brigati 

F. 6)Campanini N. 7)Berretta R. 8)Rocco A. 9)Merisio C. 

1)Department Of Pathology And Medicine Of Laborator, Parma, Parma, 

Italy 2)Department Of Pathology And Medicine Of Laboratory, Parma, 

Parma, Italy 3)Department Of Pathology, Pavia University, Pavia, Pavia, 

Italy 4)Pathology Section Of Cremona Hospital, Cremona, Cremona, Italy 

5)Department Of Pathology And Medicine Of Laboratory, Parma, Parma, 

Italy 6)Department Of Pathology And Medicine Of Laboratory, Parma, 

Parma, Italy 7)Department Of Obstetric And Gynecologic Sciences A, 

Parma, Parma, Italy 8)Department Of Clinical And Experimental Medicine,, 

Napoli, Napoli, Italy 9)Department Of Obstetric And Gynecologic 

Sciences A, Parma, Parma, Italy 

Background. To find information for invasive squamous cervical 

carcinoma in the elderly, 110 tissue specimens collected at three 

Institutions and obtained from two groups of patients (< 60 years 

of age and > 60 years of age), were analyzed for human papilloma 

virus (HPV) status, for the Cylooxygenase-2 (Cox 2), the 

epidermal growth factor receptor (EGFR) expression and clinicopathological 

features 

Material and Methods. The surgical pathology files of the Pathology 

Department of Parma University, of Pavia University and 

Cremona hospital were searched for cases of invasive squamous 

cervical carcinomas, from the years 1993-2009. 

The presence of HPV DNA was evaluated in neoplastic tissue 

using polymerase chain reaction (PCR). 

Immunohistochemical staining was performed using antibodies 

COX-2, and EGFR. 

All these parameters in the younger and older women were compared, 

using appropriate statistical tests. Overall survival curves 

were drawn using Kaplan-Meir estimates 

p < 0.05 was taken as level of significance. 

Results and Conclusions. Our study demonstrated that the invasive 

squamous cervical carcinoma in older women is characterized 

by following features: 1) More advanced stage of development 

than in the young group (p = 0.04). 

2) Higher mortality (p = 0.006). 3) Presence of HPV DNA in the 

65% of cases, which, in the absence of sexual activity, could be 

due to reactivation of latent HPV infection and to an impairment 

of host immunologic response. 4) Over-expression of Cox-2 in a 

number of cases significantly higher than younger group, but this 

immunoreactivity does not relate to EGFR expression neither to 

the presence of HPV.

300 

Thus, it is possible that the higher positivity to Cox- 2 in older 

women could be related to additional factors associated with aging, 

such as a high incidence of chronic inflammation and the 

decline in ovarian function which can increase pro-inflammatory 

cytochine. 5) Over-expression of EGFR higher than younger patients 

although this is not significant (p = 0,073). 6) Simultaneous 

immunoreactivity to Cox-2 and EGFR in a number cases significantly 

higher than younger group (p = 0.01) and this finding 

seems to have prognostic significance showing lower of survival 

than cases without this immunoreactivity (p = 0.002). 

Objectives. The number of neoplasms with more advanced stage 

of development was significantly higher than the number of cases 

in the younger group (p = 0.04). 

Moreover, we observed that in the older group the mortality was 

higher than younger patients (p = 0.006). 

The presence of HPV DNA in invasive squamous cervical carcinoma 

was significantly higher in younger group of women 

(p = 0.0095). Cox-2 staining was significantly higher in older 

patients than younger patients (p value: 0.032). The Cox-2 

immunoreactivity was significantly correlated with presence 

of lymph nodal metastases (p value: 0.05). Instead, in both 

groups of women, Cox-2 expression did not correlate with the 

presence of HPV DNA (p = 0.8158), stage of development of 

neoplasm at diagnosis (p = 0.5824) and and survival (p = no 

significant). 

Although no differences in expression of EGFR was observed between 

the younger and older groups (p value: 0,073), in the older 

women this marker showed higher expression. The expression of 

this marker was significantly related to the stage of development 

of the neoplasm (p = 0.0033), but did not relate to the presence of 

HPV DNA (p = 0.6405). 

The EGFR positivity had prognostic significance. In fact, cases 

with extensively staining (+++) (positivity present in > 50% 

of neoplastic elements) and cases with moderately positivity 

(++) (immunostaining in 10-50% of neoplastic cells) revealed a 

significant correlation with poor prognosis than negative cases 

(no reactivity in any neoplastic cells) and cases focally positive 

(+) (immunoreactivity < 10% of neoplastic elements) (p:0.03). 

In the older women the co-expression of Cox-2 and EGFR was 

significantly higher in the older group than the younger group 

(p = 0.01, Fisher exact test). Moreover, the simultaneous expression 

of these markers had poor prognostic significance, showing 

lower of survival than cases without this immunoreactivity 

(p = 0.002). 

Cribriform-morular variant of papillary thyroid 

carcinoma in a man with Gardner’s syndrome: 

pathological features and clinical implications 

1)S. Crippa, 2)L. Giovanella, 2)S. Suriano, 3)P. Saletti, 1)M. 

Frattini, 4)V. Nosé, 1)L. Mazzucchelli 

1)Institute of Pathology, Locarno, Switzerland; 2)Nuclear Medicine and 

PET/CT Centre, Oncology Institute of Southern Switzerland, Bellinzona, 

Switzerland; 3)Medical Oncology, Oncology Institute of Southern Switzerland, 

Bellinzona, Switzerland; 4) Department of Pathology, Brigham 

and Women’s Hospital, Boston MA, USA 

Background. Papillary thyroid carcinoma (PTC) is an extraintestinal 

manifestation of patients with familial adenomatous 

polyposis (FAP), mostly occurring young women. Recent publications 

highlight that FAP-associated PTC represents a distinct 

type of follicular cell neoplasm histologically characterized by 

cribriform-morular aspects, of which the incidence probably has 

been underestimated so far. We report a case history of PTC occurring 

in 55-year-old man with Gardner’s syndrome. 

Clinical history. Screening thyroid ultrasonography revealed a 

solid and well-circumscribed nodule in the left lobe of 3,5 cm, 


hard in clinical examination. Fine needle aspiration (3 passes) 

was not representative (Tir1 according to “Consensus citologico” 

SIAPEC-IAP 2007). The patient underwent left hemithyroidectomy. 

The thyroid lesion consisted of neoplastic cells in several 

small nests dispersed in an encapsulated nodule characterized 

by highly hyalinized fibrous stroma with calcifications and bone 

metaplasia. Immunohistochemical staining revealed focal but 

strong nuclear beta-catenin expression restricted to nests with 

morular aspects but not in adjacent normal appearing follicular 

structures. 

Conclusions. PTC is a rare extraintestinal manifestation of 

Gardner’syndrome that should not be disregarded. Thyroid 

screening ultrasound examination is appropriate in women as 

well as in men. We suggest that any thyroid nodule in FAP 

individuals should be considered as suspicious until proved otherwise. 

A “negative” fine needle aspiration should be interpreted 

with caution, due to the extensive fibrosis and lack of characteristic 

nuclear changes of the cribriform-morular variant of PTC. 

Finally, we suggest that morular nests with focal beta-catenin 

expression within a thyroid nodule represents a putative precursor 

lesion of PTC. 

lactoferrin expression in colorectal cancer: 

relationships with proliferation indices and 

survival 

1)Giuffrè G. 2)Ieni A. 3)Simone A. 4)Scarfì R. 5)Caristi N. 

6)Tuccari G. 

1)Patologia Umana, A.O.U. Policlinico G. Martino, Messina, Italia 2)Patologia 

Umana, A.O.U. Policlinico G. Martino, Messina, Italia 3)Patologia 

Umana, A.O.U. Policlinico G. Martino, Messina, Italia 4)Patologia 

Umana, A.O.U. Policlinico G. Martino, Messina, Italia 5)Patologia Umana, 

A.O.U. Policlinico G. Martino, Messina, Italia 6)Patologia Umana, 

A.O.U. Policlinico G. Martino, Messina, Italia 

Background. Lactoferrin (LF), an iron binding glycoprotein, 

is found in exocrine secretions and secondary granules of polymorphonuclear 

neutrophils. Oral administration of bovine LF 

suppresses experimental carcinogenesis in the colon; recently 

it has been shown that LF inhibits the growth of adenomatous 

polyps also in humans. This anti-tumoral activity of LF seems 

to be due to a wide range of functions such as inhibition of cell 

proliferation, induction of apoptosis, modulation of immune system, 

decrease of angiogenesis. Taking into consideration these 

findings as well as reports about LF immunolocalization in human 

tumours, we have investigated the immunoexpression LF in 

a series of surgically-resected colorectal cancers (CRC) to verify 

its possible correlations with proliferation indices (Ki-67 and 

AgNOR), p53 status and survival time. 

Methods. From 77 untreated patients formalin-fixed paraffinembedded 

CRC samples were selected and serial sections were 

cut. The immunoreactions were performed utilising monoclonal 

antibodies against LF (1A1; Meridian Life Science), Ki-67 

(MIB-1; Dako) and p53 (DO-7; Dako). To determinate the proliferation 

rate, the AgNOR technique was performed according to 

guidelines of the Committee on AgNOR. Statistical analysis was 

made by non-parametric methods; the Kaplan-Meier method was 

utilized for survival analysis. 

Results. A variable degree of cytoplasmatic immunoreactivity for 

LF was encountered in 71 cases (92%) of CRC; 28 cases (36%) 

showed a high expression of LF and this finding was significantly 

correlated with low expressions of Ki-67 as well as p53 and with 

a low AgNOR quantity. No relationships were demonstrated 

between LF and clinico-pathological parameters such as sex, age, 

site, histotype, grade, stage, and clinical course. These findings 

suggest that high levels of LF are associated with a low proliferative 

activity in CRC.


Her-2 overexpression in advanced node-positive 

gastric carcinomas: is there any relationship 

with other clinico-pathological histoprognostic 

parameters? 

Giuffrè G., Ieni A., Colonese F., Barresi V., Scarfì R., 

Simone A., Branca G., Adamo V., Tuccari G. 

Dipartimento Di Patologia Umana, AOU Policlinico G.Martino, Messina, 

Italia 

Background. Survival rate of patients with advanced/metastatic 

resectable gastric and gastroesophageal carcinomas remains poor 

despite chemotherapy or chemoradiation; consequently new targeted 

therapies are needed. Recent studies indicate HER-2 overexpression/amplification 

in many human cancers; in particular 

Trastuzumab, a monoclonal antibody targeting HER-2, enhances 

survival rate in both primary and metastatic HER-2 positive breast 

cancer patients. Therefore, we have immunohistochemically analyzed 

the HER-2 status in advanced gastric adenocarcinomas in 

order to identify patients eligible to trastuzumab therapy as well 

as to reveal possible relationships with other clinico-pathological 

parameters and prognosis. 

Methods. Fifty-eight formalin-fixed paraffin embedded gastric 

surgical specimens were obtained from an equal number 

of patients (M 35 - F 23); mean age 66.7 (range 37-95 yrs); 

mean follow-up value 47.4 months. Clinico-pathological histoprognostic 

data concerning site and dimensions of carcinomas, 

histotype, lymphatic invasion, grading, nodal status, staging, 

growth fraction (Ki-67) and proliferation rate (AgNOR) were 

also available. HER-2 status (AO485 DAKO, Denmark) has 

been evaluated by a score: 0 (no staining), 1+ (faint and discontinous 

staining in ≤ 10% of neoplastic elements), 2+ (light to 

moderate lateral, basolateral or complete staining in ≥ 10% of 

neoplastic elements), 3+ (strong, intense lateral, basolateral or 

complete staining in ≥ 10% of neoplastic elements). All cases 

considered equivocal (2+) have been furtherly assessed by FISH 

test (pharmDx DAKO). Statistical analysis was performed by 

Chi-square test. 

Results. 31 cases showed intestinal histotype, 19 were diffuse 

and 8 gastric carcinomas were mixed-type; moreover, 18 were 

stage II, 27 stage III, 13 stage IV. HER-2 overexpression was 

found in 10/58 cases (17.24%) and it was significantly related 

only to intestinal histotype, high Ki-67 and AgNOR values, presence 

of distant metastases. 

High frequency of codon 61 and 146 mutations at 

first diagnosis in a series of colorectal cancer: a 

mono-institutional experience 

1)Giusti A. 2)Bertacca G. 3)Lombardi S. 4)Orlandi M. 5)Del freo 

A. 6)Culli M. 7)Bigini D. 8)Tornaboni D. 9)Tozzini S. 10)Cavazzana 

A. 

1)Oncologia, Carrara, Carrara, Italia 2)Diagnostica, Massa, Massa, Italia 

3)Diagnostica, Massa, Massa, Italia 4)Oncologia, Carrara, Carrara, 

Italia 5)Oncologia, Carrara, Carrara, Italia 6)Oncologia, Carrara, Carrara, 

Italia 7)Oncologia, Carrara, Carrara, Italia 8)Oncologia, Carrara, 

Carrara, Italia 9)Oncologia, Carrara, Carrara, Italia 10)Oncologia, Carrara, 

Carrara, Italia 

Background. Patients with metastatic colorectal cancer may benefit 

from therapy with the monoclonal antibodies (MoAb) direct 

against epidermal growth factor receptor (EGFR) cetuximab and 

panitumumab. Colorectal cancer (CRC) with KRAS and BRAF 

mutations are resistant to antibodies anti-EGFR; their mutations 

occur in approximately 40% and 10% respectively of CRCs. 

However up to 50% of wild type KRAS CRC do not benefit from 

such therapies. According to ASCO 2009 guidelines candidate 

patients should be tested for KRAS mutations in codons 12 and 

301 

13 to predict response to anti-EGFR MoAb. Little is known about 

the frequency of mutation in codon 61 and 146. The aim of this 

study is to characterize the mutational status of both KRAS and 

BRAF in a series of 69 consecutive and unselected CRC and to 

correlate the results with clinicopathologic parameters. 

Methods. 69 consecutive CRC cases were enrolled in the study. 

DNA was extracted from representative, formalin-fixed, paraffin-embedded 

tissue blocks containing at least 20% of tumor 

cells. KRAS (codons 12, 13, 61, 146) and BRAF (exons 11 and 

15) mutational analysis was performed by Pyrosequencing assay. 

Complete clinical information were available in 68 patients. 

Results. Codon 12 and 13 were mutated in 23 out of 69 (33%) 

cases, while the mutational rate for codon 61 and 146 was 6% 

(4/69) and 9% (6/69) respectively, raising the frequency of 

KRAS mutation in our series to 48%. RAF was mutated in 6/69 

cases (9%). The overall frequency of KRAS/BRAF mutation was 

56.5% (39/69). No difference was found as far as KRAS mutation 

was concerned regarding site of origin of the tumor, stage, 

sex, and age of patients. Viceversa, BRAF mutated tumors were 

all located in the right colon and trasversum, no mutation was 

found on left sided tumors (p = 0.002). From the analysis of these 

preliminary results it seems mandatory to include in the analysis 

codon 61 and 146 since they account up to 30% of all KRAS 

mutation in our series. 

Deletion of chromosome 3p in clear cell renal 

cell carcinoma, a comparison between classical 

cytogenetic and fluorescence in situ hybridization 

1)Gobbo S. 2)Brunelli M. 3)Segala D. 4)Bersani S. 5)Menestrina 

F. 6)Martignoni G. 

1)Anatomia Patologica, Universitaà Di Verona, Verona, Italia 2)Anatomia 

Patologica, Università Di Verona, Verona, Italia 3)Anatomia Patologica, 

Università Di Verona, Verona, Italia 4)Anatomia Patologica, Università Di 

Verona, Verona, Italia 5)Anatomia Patologica, Università Di Verona, Verona, 

Italia 6)Anatomia Patologica, Università Di Verona, Verona, Italia 

Background. Genetic aberrations of gene VHL are considered 

an early pathogenetic event in clear cell renal cell carcinoma 

development. In most of cases one allele of VHL is mutated and 

the other is lacking due to deletion of its genomic locus in the 

short arm of chromosome 3. This has been confirmed by several 

cytogenetic studies based on metaphase karyotyping. Nowadays, 

the most diffuse cytogenetic analysis on diagnostic practice is 

fluorescence in situ hybridization and few data are available on 

formalin-fixed and paraffin embedded tissues. Increasing experience 

in its interpretation must be achieved. For this reason we 

tested the detection efficacy of 3p deletion on clear cell renal cell 

carcinoma comparing the standard classical metaphase to interphase 

fluorescence in situ hybridization analysis. 

Methods. We selected 9 cases of clear cell renal cell carcinoma 

showing a standard karyotype characterized by deletion of chromosome 

3p. We performed on these cases fluorescence in situ 

hybridization analysis using centromeric probe for chromosome 

3 and a locus specific probe for 3p (Olympus). We considered 

deleted the cases where the ratio between the centromeric signals 

and the locus specific signals (3/3p) was higher than the mean 

ratio plus 3 standard deviations observed in the adjacent renal 

parenchyma. 

Results. Five out of 9 cases of clear cell renal cell carcinoma 

(55%) showed 3p deletion with fluorescence in situ hybridization. 

Normal renal parenchyma showed a mean ratio 3/3p of 1,23 

with a standard deviation of 0,05. Differences in between the two 

cytogenetic analyses could be explained by divergent clonal proliferation 

occurring when metaphase cell cultures are prepared. 

Overall, the concordance between the two techniques must be 

kept in mind when FISH analysis is requested as diagnostic tool.

302 

feasibility of fluorescence in situ hybridization 

analysis in “finefixed” human tissue 

1)Gobbo S. 2)Brunelli M. 3)Segala D. 4)Eccher A. 5)Ghimenton 

C. 6)Scarpa A. 7)Martignoni G. 8)Menestrina F. 

1)Anatomia patologica, Università di verona, Verona, Italia 2)Anatomia 

patologica, Università di verona, Verona, Italia 3)Anatomia patologica, 

Università di verona, Verona, Italia 4)Anatomia patologica, Ospedale 

civile maggiore, Verona, Italia 5)Anatomia patologica, Ospedale civile 

maggiore, Verona, Italia 6)Anatomia patologica, Università di verona, 

Verona, Italia 7)Anatomia patologica, Università di verona, Verona, Italia 

8)Anatomia patologica, Università di verona, Verona, Italia 

Background. Because formalin fixation results in poor preservation 

of DNA and RNA, there are efforts to introduce new molecular 

friendly fixatives. Recently a new ethanol based fixative 

named FineFIX (Milestone, Sorisole, Italy) has been introduced 

and several articles exist about its advantages in improving molecular 

analysis based on tissue extraction. Nowadays Fluorescence 

in situ hybridization is the most diffuse cytogenetic method 

to discern diagnostic, prognostic and predictive factors in human 

neoplasms. Its popularity is mostly due to the feasibility on formalin 

fixed tissue whereas there is no report about its efficiency 

on FineFIXed tissues. 

Methods. We collected fresh samples (4ml in volume) from 

different human tissues (spleen, liver and kidney), divided each 

sample in two specular parts and put one in 50 ml of formalin and 

the other in 50 ml of FineFIX for 24 hours. Standard processing 

and paraffin embedding was performed. Fluorescence in situ 

hybridization was applied on 4 µm slides according to the homemade 

protocol used in our cytogenetic laboratory. In brief, the 

slides were deparaffinised and treated with 0.1mM citric acid at 

95ºC for 10 minutes. Digestion of the tissue was performed with 

0.4 ml of pepsin at 37ºC for 40 minutes. FISH was performed 

with 5ml of diluted probe (1:100). Denaturation was achieved 

incubating 83ºC for 12 minutes and hybridization at 37ºC overnight. 

We tested centromeric probes for chromosome 7 and 17 

on each sample. 

Results. Fluorescence signals were bright in formalin fixed tissue 

as in the samples fixed in FineFIX. The number of signals as 

expected was double for the great majority of the cells and the 

rate of false monosomic cells was the same. As single difference 

the tissue fixed in FineFIX seamed to be more digested by the 

10 minutes of pepsin used in our protocol suggesting the need of 

decreasing the digestion time for sample fixed with FineFIX. 

Mammary myofibroblastoma: report of a new 

series of seven cases with emphasis on unusual 

features and needle core biopsy-based diagnosis 

1)Amico P. 1)Greco P. 2)Kazakov D. 3)Kacerovskà D. 4)Michal 

M. 5)Puzzo L. 6)Magro G. 

1)Dipartimento G.F. Ingrassia Anatomia Patologica-Università Di Catania, 

Azienda Osp. Univ. Policlinico-Vittorio-Emanuele, Catania, Italia 

2)Sikl’s Department Of Pathology, Charles University, Medical Faculty 

Hospital, Pilsen, Czech Republic 3)Sikl’s Department Of Pathology, 

Charles University, Medical Faculty Hospital, Pilsen, Czech Republic 

4)Sikl’s Department Of Pathology, Charles University, Medical Faculty 

Hospital, Pilse, Czech Republic 5)Dipartimento G.F. Ingrassia Anatomia 

Patologica-Università Di Catania, Azienda Osp. Univ. Policlinico-Vittorio-Emanuele, 

Catania, Italia 6)Dipartimento G.F. Ingrassia Anatomia 

Patologica-Università Di Catania, Azienda Osp. Univ. Policlinico-Vittorio-Emanuele, 

Catania, Italia 

Background. Myofibroblastoma (MFB) of the breast is an uncommon 

benign stromal tumour, which exhibits a wide variety 

of cytomorphologic features and architectural patterns. This is 

mainly due to the fact that neoplastic cells, showing a variable 

fibro-myofibroblastic differentiation, may adopt marked intralesional 

and interlesional variability in morphology. In this regard, 

several morphological variants, including cellular, infiltrative, 


epithelioid, deciduoid-like, collagenized/fibrous, lipomatous, and 

myxoid variants, have been recognized over time. 

Materials. We report the clinico-pathological features of a new 

series of 7 cases of mammary MFBs, with emphasis on unusual 

morphological features and core biopsy-based diagnosis 

Results. There were 6 female and 1 male patients. Age at diagnosis 

ranged from 59 to 93 years. All tumours were well-circumscribed 

and ranged in size from 2.5 to 5 cm in greatest diameter. 

Histologically they were classified into the following variants: i) 

classic (2 cases); ii) collagenized/fibrous (2 cases); iii) epithelioid 

(2 cases); iv) lipomatous (1 case). Two cases (collagenized/fibrous 

and epithelioid cell variant) were preoperatively diagnosed 

by core biopsy. Immunohistochemically all tumors, albeit with a 

variable extension, were positive to desmin, α-smooth muscle actin, 

CD34, bcl-2, CD99, CD10, ER/PR/AR. The most intriguing 

morphological features were observed in one case of epithelioid 

cell variant and in the single case of lipomatous variant. The former 

tumour was composed of medium- to large-sized epithelioid 

cells exhibiting a mild to moderate degree of nuclear pleomorphism 

and a multinodular arrangement. This latter finding has 

not been previously reported in mammary MFB. The lipomatous 

variant was composed of a predominant fatty component (80%) 

with a minority of spindle cells intermingling with mature adipocytes, 

frequently embedded in a myxoid stroma containing thin 

and thick collagen bands. The overall picture of this tumour was 

closely reminiscent of a spindle cell lipoma. 

Although recognition of morphological variants and unusual features 

in mammary MFB could seem to be of academic interest, 

we think they may represent a diagnostic challenge, especially 

when evaluating needle core biopsies. 

Prevalence of intratubular germ cell neoplasia 

in testis cancer: implications in the testis sparing 

surgery 

1)F. Pierconti, 2)G. Gulino 

1)Istopatologia, Ucsc, Roma, Italia; 2)Urologia, Ucsc, Roma, Italia 

Background. Despite ITGCN (Intra Tubular Germ Cell Neoplasia) 

has been described in testicular tissue adiacent to invasive 

cancer. As far both multifocality and ITGCN are the major limitations 

to simple enucleation of germ cell testis tumours. Aim of 

this study was to assess the prevalence and topographic localization 

of ITGCN in patients with testis germ cell tumours in order 

to detect any safety margin from primary cancer. 

Methods. 41 orchiectomy specimens have been analyzed. Multiple 

5 mm sections have been obtained including the primary 

tumour and the “healthy” tissue of the whole testis. The mean 

diameter of the mass, the ratio between tumour and testis volume, 

the presence of foci of ITGCN and the distance of ITGCN from 

the primary mass have been evaluated. Multifocality such as synchronous 

foci of cancer, vascular invasion or involvement of the 

rete testis were analyzed. 

Results. Classic seminoma was found in 26 patients (19/26 stage 

I,7/26 stage II or more), non seminomatous or mixed tumours 

in 15 (11/15 stage I,4/15 stage II or more). The average diameter 

of the tumor masses was 23 mm, mean testicular diameter 

45 mm and mean ratio between cancer mass and testis 0.51. In 

18 specimens out of 41(43%) was documented ITGCN synchronous, 

with single focus in 8 out of 41(19.5%), with multiple foci 

in 10 out of 41(24.3%). The average distance between ITGCN 

and the primary mass was equal to 18 mm. In 3 specimens out 

of 41(7.3%) was documented evidence of multifocality with foci 

of synchronous invasive germ cell tumor size up to 8 mm. Our 

preliminary data indicate that about half of patients with germ cell 

tumors present histologically as well as the mass also documented 

ITGCN located up to 2 cm from the primary mass. Has not been 

identified, therefore a safety margin that allows to perform such 

an organ-sparing radical surgery.


role of a DNA vaccine targeting erBB2 in 

a hamster model of chemical lnduced oral 

carcinogenesis 

1)Hysi (A). 2)Berta (GN). 3)Stramucci (L). 4)Ascione (P). 5)Curcio 

(C). 6)Musiani (P). 7)Iezzi (M). 

1)Anatomia Patologica/Oncologia E Neuroscienze, SS. Annunziata/CESI, 

Chieti, Italia 2)Scienze Biologiche E Cliniche, Unità Farmacologia, Università 

Di Torino, Torino, Italia 3)Anatomia Patologica/Oncologia E 

Neuroscienze, SS. Annunziata/CESI, Chieti, Italia 4)Anatomia Patologica/Oncologia 

E Neuroscienze, SS. Annunziata/CESI, Chieti, Italia 5)Oncologia 

E Neuroscienze/Oftalmologia, SS. Annunziata/CESI, Chieti, Italia 


Chieti, Italia 7)Anatomia Patologica/Oncologia E Neuroscienze, SS. Annunziata/CESI, 


Background. Vaccines against pathogens prevent cancer onset 

in several cases. For instance, the vaccination against hepatitis B 

virus reduces the incidence of hepatocellular carcinoma, whereas 

vaccines against human papilloma viruses are expected to greatly 

reduce the incidence of cervical carcinoma. Immunoprevention 

of viral tumors is now becoming implemented at the population 

level, thanks to the development of effective vaccines that are 

expected to decrease human tumor burden in the near future. 

Vaccines addressing tumor associated molecules with a causal 

role in the promotion of carcinogenesis unrelated to infections 

(oncoantigens) halt the progression of early stages of neoplastic 

lesions in several cancer prone genetically engineered mice. The 

effectiveness of these anti-oncoantigen vaccines lies in their ability 

to target molecules delivering signals that play an essential 

role in driving the progression of neoplastic lesions. 

Methods. Here, we evaluated whether a DNA vaccine targeting 

an archetypal oncoantigen (ERBB2) also protect against 7,12-di 

methylbenz[α]anthracene (DMBA)-induced oral carcinogenesis 

in random bred hamsters. 

Results. The number, size and severity of oral lesions were significantly 

reduced in ERBB2 immunized hamsters. The intensity 

of inhibition directly correlated with the titer of vaccine-elicited 

anti-ERBB2 antibodies. These findings suggesting that vaccination 

against selected oncoantigens can interfere with the progression 

of chemical carcinogenesis opens a new scenario in the use 

of vaccine in hampering the progression of chemically-induced 

cancer. 

KrAS mutation analysis on cytological specimens 

of non small cell lung cancer 

Iaccarino A., Malapelle U., Desiderio D., Cembrola S., Palombini 

L., Troncone G. 

Scienze biomorfologiche e funzionali, Università di Napoli “Federico II”, 

Napoli, Italia 

Background. Recent evidences showed that advanced non 

small cell lung cancer (NSCLC) patients with tumors harboring 

a KRAS gene mutation do not derive benefit from the administration 

of epidermal growth factor receptor-antagonists. To 

date, the specimens tested for KRAS mutational analysis are 

formalin-fixed paraffin-embedded (FFPE) primary tumor tissue 

blocks. However, in patients with NSCLC the source of FFPE 

material is limited. In this setting, NSCLC cytological samples 

may be exploited. However, these specimens are mostly routine 

archivial slides; thus, their suitability for the KRAS assay needs 

to be tested. 

Methods. K-ras analysis by direct gene sequencing, was carried 

out on DNA extracted from cytological tumor samples after microdissection. 

Results. Here, we show that 25/26 (96.1%) NSLC smears were 

perfectly adequate for codon 12 and 13 KRAS mutational analysis 

by direct gene sequencing. Only one case (3.9%) showing 

abundant necrotic debris and poor cellular preservation was not 

303 

informative for KRAS status. Codon 12 gene mutations were 

found in 2/25 (8%) of the adequate cases (c35G > T). 

Conclusions. Thus, whenever histological specimens of NCSLC 

are not available, KRAS testing may be reliably performed on 

cytological specimens. 

Morphological and biomolecular characteristics 

of subcentimetric invasive breast carcinomas: a 

Sicilian multicentric retrospective analysis 

1 Ieni A, 1 Giuffrè G, 2 Lanzafame S, 3 Nuciforo G, 3 Curduman M, 

4 Villari L, 5 Roz E, 6 Certo G, 7 Cabibi D, 4 Salomone E, 6 Labate 

A, 8 Messina D, 7 Franco V, 1 Adamo V and 1 Tuccari G on behalf 

of the Sicilian Sections SIAPEC-IAP-AIOM (Gruppo di lavoro 

sulla Terapia Molecolare dei Tumori) 

1 Dipartimento di Patologia Umana, Università di Messina, A.O.U. “Policlinico 

G. Martino”, Messina; 2 Dipartimento Anatomia, Patologia diagnostica, 

Medicina legale, Igiene e Sanità Pubblica, Università di Catania, 

“Policlinico G. Rodolico”, Catania; 3 Centro Catanese Oncologia 

Humanitas, Catania; 4 A.O.U. Vittorio Emanuele II, Catania; 5 Casa di 

Cura “La Maddalena”, Palermo; 6 Casa di Cura “Cappellani”, Messina; 

7 Dipartimento di Patologia Umana, Università di Palermo, A.O.U. “Policlinico 

Giaccone”, Palermo; 8 U.O.C. Anatomia Patologica A.O. Sant’Antonio 

Abate, Trapani. 

Background. Data concerning human epidermal growth factor 

receptor 2 gene (HER-2) in pT1a,bN0M0 breast cancers are conflicting 

and heterogeneous. In subcentimetric invasive breast carcinomas 

(SIBC), high tumor grade is the most consistent factor 

associated with poor prognosis together with younger age, estrogen/progesterone 

receptor–negative status, high Ki-67; however, 

cases HER-2-positive pT1a,bN0M0 carcinomas seem to have an 

higher risk of relapse and related death, although the decision to 

use trastuzumab in SIBC is still debatable. In order to analyze the 

morphological and biomolecular characteristics of SIBC, we have 

collected a cohort of 410 cases from Sicilian anatomopathological 

units, also in the attempt to verify the existence of some relationships 

among HER-2 status, hormone receptor status, Ki67 values, 

grade, histotype and node involvement. 

Methods. From four-hundred ten formalin-fixed paraffin-embedded 

tissue blocks of SIBC 4 µm thick parallel sections were 

cut, mounted on silane-coated glasses and, after routine retrieval 

procedure, immunostained for ER (ID5, DBA, 1:10), PR (PgR- 

ICA, Abbott, 1:10), Ki-67 antigen (MIB-1, DAKO Cytomation, 

1:200). HER-2 status was scored according to manufacturer’s 

recommendations (HercepTest AO485 DAKO); cases considered 

equivocal (2+) have been successively assessed by FISH test 

(pharmDx DAKO). Statistical analysis was performed by Chisquare 

test. 

Results. In SIBC, a significant correlation (p < 0.001) between 

high hormonal receptors expression and not-amplified HER-2 

status as well as between high Ki-67 values and amplified HER- 

2 status have been found. One-hundred thirteen cases were pT1a 

(N0 86%; N1 14%) and two-hundred ninety seven were pT1b 

(NO 82%, N1 18%): between these two groups no statistical differences 

were encountered for all examined parameters, except 

for the grade. Moreover, when all pT1a,bN0 were compared with 

corresponding N+ cases, only HER-2 status was significantly 

different (p = 0.014). 

An immunohistochemical investigation of premetastatic 

niche (PMN) in node-negative invasive 

breast carcinomas 

1)Ieni A. 2)Simone A. 3)Giuffrè G. 4)Grosso M. 5)Scarfì 

R. 6)Plutino FM. 7)Colonese F. 8)Adamo V. 9)Tuccari G. 

1)Dipartimento Di Patologia Umana, Azienda Ospedaliera Universitaria 

G.Martino, Messina, Italia 2)Dipartimento Di Patologia Umana, Azienda 

Ospedaliera Universitaria G.Martino, Messina, Italia 3)Dipartimento Di 

Patologia Umana, Azienda Ospedaliera Universitaria G.Martino, Mes-

304 

sina, Italia 4)Dipartimento Di Patologia Umana, Azienda Ospedaliera 

Universitaria G.Martino, Messina, Italia 5)Dipartimento Di Patologia 

Umana, Azienda Ospedaliera Universitaria G.Martino, Messina, Italia 

6)Dipartimento Di Patologia Umana, Azienda Ospedaliera Universitaria 

G.Martino, Messina, Italia 7)Dipartimento Di Patologia Umana, Azienda 

Ospedaliera Universitaria G.Martino, Messina, Italia 8)Dipartimento Di 

Patologia Umana, Azienda Ospedaliera Universitaria G.Martino, Messina, 

Italia 9)Dipartimento Di Patologia Umana, Azienda Ospedaliera 

Universitaria G.Martino, Messina, Italia 

Background. Invasive breast cancers (IBC) usually metastasize 

into regional lymph nodes, so their involvement has been considered 

one of the major morphological parameters in clinical 

management. 80% of women with lymph node-negative invasive 

breast cancers are expected to be alive and free from distant 

metastases at ten years but, unfortunately, 20% of women with 

node-negative breast cancer develop metastases. Moreover, the 

recruitment of the bone marrow-derived hematopoietic clusters 

with a maintained progenitor cell status (HPCs) represents an 

essential cellular event in the process of metastasis and these cellular 

clusters may identify sites of future metastases, representing 

a pre-metastatic niche. Herein we have immunohistochemically 

analyzed negative axillary lymph nodes from patients affected by 

IBC in order to verify if HPCs may represent the first step of the 

metastatic spread. 

Methods. 626 lymph nodes (603 pN0 and 23 pN1a), obtained 

from 51 patients (mean age 62.05; range 41-85 yrs), surgically 

treated in the period 1998-2007 for invasive breast carcinomas, 

the mean follow-up was 62.59 months (6-136 mo.). Endocrine 

receptor status (ER, PgR) as well as growth fraction (Ki67 LI) 

were also available. Formalin-fixed paraffin-embedded 4-µmthick 

serial sections were treated with the following polyclonal 

antisera: CD133 (Abgent, w.d. 1:80), CD117 (DAKO, w.d. 

1:500), VEGFR1 (Flt1) (Santa Cruz Biotechnology, w.d. 1:400) 

and with mouse monoclonal anti-human CD34 (DAKO, w.d. 

1:50). The possible correlation between immunohistochemical 

data and clinico-pathological characteristics of breast carcinomas 

was investigated using non-parametric methods. 

Results. CD34 and CD117 appeared the most useful HPCs 

markers, whereas Flt-1 and CD133 exhibited a variable rate of 

immunostained elements. A significant relationship (p < 0.001) 

was found between immunohistochemical detection of HPCs and 

development of distant metastases, high Ki67 values as well as 

exitus for IBC. 

Correlations between cytology and 

immunocytochemistry on cellular blocks in the 

assessment of mediastinal tumors 

1)F.C. Popescu, 2)A.M. Ioncica, 3)I.E. Plesea, 2)A. Saftoiu, 4)M. 

Comanescu 

1)Pathology, Emergency County Hospital, Craiova, Romania; 2)Research 

Center of Gastroenterology and Hepatology, University of Medicine and 

Pharmacy, Craiova, Romania; 3)Pathology, University of Medicine and 

Pharmacy, Craiova, Romania 4)Pathology, National Institute “Victor Babes”, 

Bucharest, Romania 

Background. This preliminary study is assessing the combined 

citology and immunocitochemistry on cell blocks examinations 

of the samples obtained by endoscopic ultrasound transesophageal 

fine needle aspiration (EUS-FNA) and/or endoscopic ultrasound 

transbronchial needle aspiration (EBUS-TBNA) in order 

to confirm the diagnosis of malignancy and stage the pulmonary 

tumors. EBUS-TBNA allowed the tissue sampling in real time in 

levels 2, 4 and 7 of mediastinal lymph nodes and EUS – FNA at 

levels 7 and 5. 

Methods. The study included 40 cases suspected of pulmonary 

cancer. Smears were stained with Papanicolaou and May 

Grunwald Giemsa. The citology exam assessed the presence of 

malignant cells and their disposal. Cell blocks were obtained by 


wax embedding of sediments. First section from cell blocks was 

stained with Hematoxilin-Eosine and the next ones were immunomarked 

with specific antibodies for identification of tumoral 

fenotype. Tumor biopsy was possible only in few cases for correlating 

the cytology with the histopathological results. 

Results. In 27 cases, FNA was performed on lymph nodes, in 4 

cases on tumors in 9 cases on both lymph nodes and tumor. Metastasis 

were diagnosed in 12 patients. Cell blocks were obtained 

in 16 cases and tumoral cells were present in 5 of them. The tumor 

fenotype was of scuamos carcinoma in 3 cases (ie AE1/AE3, 

CK5/6 and p63 positive) and of adenocarcinoma in 2 cases (ie 

CK7 and TTF-1 positive) all these 5 cases being confirmed by 

biopsy. 

Conclusions. Citology on FNA smears is very important for confirming 

tumoral malignancy and for staging pulmonary tumors. 

The tumor fenotype can be established by immunocitochemistry 

on cell blocks. However, immunocitochemistry is limited because 

of the small quantity of FNA cell samples and subjective 

because of the tumor heterogenity. 

Acknowledgement: Work supported from Research Project 41- 

079/2007 financed by CNMP. 

Triple negative breast cancer and histological 

subtypes, immunohistochemical and 

clinicopathological analysis. 

1)Ishikawa Y. 2)Nakajima H. 3)Katano M. 4)Koibuchi Y. 

5)Horiguchi J. 6)Hayashi M. 7)Oyama T. 

1)Department of diagnostic patholgy, Gunma university graduate 

school of medicine, Maebashi, Japan 2)Department of diagnostic patholgy, 

Gunma university graduate school of medicine, Maebashi, Japan 

3)Department of diagnostic patholgy, Gunma university graduate 

school of medicine, Maebashi, Japan 4)Thoracic and visceral organ 

surgery, Gunma university graduate school of medicine, Maebashi, 

Japan 5)Thoracic and visceral organ surgery, Gunma university graduate 

school of medicine, Maebashi, Japan 6)Breast oncology, Tokyo 

medical university hachioji medical center, Tokyo, Japan 7)Department 

of diagnostic patholgy, Gunma university graduate school of medicine, 

Maebashi, Japan 

Introduction. Estrogen receptor and Progestrone receptor negative 

and HER2 negative (triple negative: TN) tumors have 

focused as a prognostic group with aggressive behavior that currently 

lack benefit of available systemic therapy. In this study, 

immunohistochemical and clinicopathological examination was 

carried out to TN breast cancer from the histopathological stand 

point of view. We also examine TN ductal carcinoma in situ (TN 

DCIS), compared with both ductal (dcIC) and invasive (iIC) components 

of TN invasive breast cancer. 

Materials and methods. Tumor tissues were obtained from 98 

patients with TN invasive carcinoma and 10 patients with TN 

DCIS at Gunma and Dokkyo University Hospital from 1995 to 

2007. Two characteristic histological subclassification was added 

to invasive carcinoma, one of them was so called “atypical” medullary 

carcinoma (type A) and the other one was TNIDC with 

central acellular zone (type B). Conventional carcinoma were 

classified as type C (conventional IDC) and other special types 

were classified as type D (special types). 

Results and Comments. We compared the correlation of each 

immunohistochemical result in DCIS, dcIC and iIC. The positive 

rates of EGFR expression were higher than that in dcIC. The 

positive rates of CK 14 expression in DCIS was lower than that 

in dcIC and iIC. There was a tendency that MIB-1 positivity increases 

from DCIS, dcIC to iIC. In cases of invasive carcinoma, 

EGFR expression was significantly correlated with CK5/6 expression, 

and CK5/6 with CK14. 

Type A and B have a few ductal components and they have also 

high MIB-1 index. The relapse rate in Type B is highest (36.4%). 

Overall survival (OS) in type B is shortest than in other groups


(p = 0.019). TN breast cancer is a heterogeneous group and in this 

study, it was suspected based on the tissue subtype classification 

that prognosis of type B is significantly worse compared with that 

of other types. 

Post-traumatic inflammatory pseudotumour of 

the breast with atypical/bizarre cells: a potential 

diagnostic pitfall 

1)G.M. Vecchio, 2)G. La Greca, 3)G. Grasso, 1)E. Vasquez, 1)E. 

Giurato, 1)A. Torrisi, 1)G. Magro 



Catania, Italia; 2)Dipartimento Scienze Chirurgiche-Università di Catania, 

Ospedale Cannizzaro, Catania, Italia; 3)Servizio Anatomia Patologica, 

Ospedale Cannizzaro, Catania, Italia 

Background. The terms “inflammatory pseudotumour” (IPT) or 

“inflammatory myofibroblastic tumour” (IMT) are usually used 

interchangeably in the literature to describe controversial fibroinflammatory 

entities, ranging from reactive lesions to potentially 

metastasizing tumours. However, despite its morphological and 

immunophenotypical overlapping with IPT, over the last years 

IMT has emerged as a distinct tumour entity that characteristically 

occurs in the lung, abdomen, pelvis and retroperitoneum of 

children and adolescents (). Although 19 cases of IMT/IPT have 

been reported in the breast, no history of previous trauma/infection 

has been documented. Accordingly most authors refer to 

these lesions as IMTs. 

Materials. We herein report the first case of an IPT of the 

breast parenchyma with atypical/bizzare cells, developing after 

a recent mechanical trauma in a 22year-old male patient. Ultrasonography 

revealed an ill-defined 7-cm mass. Lumpectomy 


Results. Grossly, the cut surface showed a solid-cystic mass, 

whitish in colour. Histologically the lesion consisted of a 

proliferation of spindle-shaped cells with palely eosinophilic 

cytoplasm and vesicular nuclei, predominantly arranged in a 

fascicular growth pattern. An inflammatory infiltrate of lymphocytes, 

plasma cells and eosinophils was variable associated. 

Focally spindle- to polygonal-shaped mono- or multi-nucleated 

cells exhibited marked cytological atypia, namely large vesicular 

or hyperchromatic nuclei with prominent nucleoli. Mitotic 

activity was low (up 3 mitoses × 10 HPF) and neither atypical 

mitoses nor necrosis was seen. The overall picture of the lesion 

was indistinguishable from that of an “IMT with atypical 

features”. Immunohistochemically the cells were positive only 

to vimentin, α-smooth muscle actin and focally to desmin. No 

immunoreactivity was obtained with ALK protein. Our case emphasizes 

that when dealing with an ALK-negative spindle cell 

tumour associated with inflammation, occurring in an unusual 

site for IMT, including breast, and/or in middle-aged or older 

adult, alternative diagnoses, especially IPT should be seriously 

considered. 

Immunocolorazione per Her-2 (HerCeP-test) e 

fISH per Her-2. uno studio di validazione su 100 

campioni paralleli di citologia per ago sottile di 

carcinoma mammario 

1)La Vecchia F. 2)Curcio M.P. 3)Staiano M. 4)Gioioso A. 

5)Butera D. 6)Demuru A. 7)Fulciniti F. 8)Botti G. 

1)Anatomia Patologica, Ist. Fon. Sen. Pascale, Napoli, Italy 2)Anatomia 

Patologica, Ist. Fon. Sen. Pascale, Napoli, Italy 3)Anatomia Patologica, 

Ist. Fon. Sen. Pascale, Napoli, Italy 4)Anatomia Patologica, Ist. Fon. Sen. 

Pascale, Napoli, Italy 5)Anatomia Patologica, Ist. Fon. Sen. Pascale, Napoli, 

Italy 6)Anatomia Patologica, Ist. Fon. Sen. Pascale, Napoli, Italy 

7)Anatomia Patologica, Ist. Fon. Sen. Pascale, Napoli, Italy 8)Anatomia 

Patologica, Ist. Fon. Sen. Pascale, Napoli, Italy 

305 

Introduzione. Uno dei problemi connessi all’immunocolorazione 

per Her-2 dei campioni citologici con metodica Hercep-test o 

similare è la possibile interferenza della fissazione alcolica con 

i risultati della immunoreazione. Diverse riserve si registrano in 

letteratura sull’uso di fissativi differenti dalla formalina acquosa 

nella processazione immunocitochimica di campioni citologici 

di biopsia per ago sottile da neoplasie mammarie. Ne consegue 

che pazienti vengono spesso sottoposte a biopsie al solo scopo di 

valutare lo score di Her-2, esponendole così a stress aggiuntivo e 

a possibili complicanze. 

Materiali e Metodi. Si sono sottoposti 100 strisci paralleli, da 

campioni di biopsia per ago sottile da carcinomi mammari, a 

colorazione immunocitochimica per Her-2 con metodica Herceptest 

e a FISH per Her-2. I vetrini sono stati prima colorati sec 

Papanicolaou per verificarne cellularità e fissazione e poi sottoposti 

ad immunocolorazione per Hercep-test previo smascheramento 

in tampone a pH basico a 99°C. Vetrini citologici paralleli 

sono stati fissati all’aria e congelati a -20°C per essere sottoposti 

a FISH. Tutti i casi sono stati validati tramite confronto tra i risultati 

dell’Hercep-test e FISH con quelli dei corrispondenti 

campioni istologici. 

Risultati. Su un totale di 44 casi con score citologico 0/1+, 43 

(97.7%) hanno presentato lo stesso score con metodica Herceptest 

sul campione istopatologico; 1 (2.3%) score 3+ sul campione 

istopatologico, con FISH non amplificata; 2 (4.5%) score 

istologico 1+ con FISH amplificata. Tutti i 25 casi con score 

citologico 2+ hanno presentato lo stesso score anche in istologia; 

17 (68%) hanno mostrato una FISH non amplificata sul campione 

citologico, mentre 8 (32%) sono risultati polisomici per il cromosoma 

17. Dei 31 casi con score citologico 3+, 28 (90.3%) hanno 

mostrato score 3+ anche sul campione istopatologico, con FISH 

amplificata; 1 (3.3%) score istologico 1+ con FISH su campione 

istopatologico indicante polisomia per cromosoma 17; 2 (6.6%) 

score istologico tra 0 e 1 con FISH su campione citologico non 

amplificata. 

Conclusioni. Si è registrato un elevato grado di correlazione 

tra lo score di Hercep-test effettuato su vetrini citologici fissati 

in alcool al 95% ed i corrispondenti campioni istologici. I 

risultati non giustificano l’ostracismo di numerosi Autori all’uso 

di preparati citologici fissati in alcool al 95% per la valutazione 

dell’Hercep-test score. 

Appendicular mucocele: a case report 

1)Labate A. 2)Mazzon E. 3)Certo G. 4)Mazzitelli R. 

1)Anatomia Patologica, Clinica Cappellani Giomi Spa, Messina, Italia 

2)Medicina e Farmacologia, Università Di Messina, Messina, Italia 

3)Anatomia Patologica, Clinica Cappelani Giomi Spa, Messina, Italia 

4)Chirurgia Generale, Casa Di Cura Caminiti, Villa S. Giovanni (Rc), 

Italia 

Introduction. Appendiceal mucocele (AM) is a rare entity that 

can present with a variety of clinical symptoms or occur as an 

incidental surgical finding. The incidence is 0.2%-0.4% of patients 

undergoing appendectomy. AM is a progressive dilatation 

of the appendix from the intraluminal accumulation of the mucoid 

substance. 

We present the case of a patient who developed subacute intestinal 

obstruction secondary to appendiceal mucocele. 

Case report. a 73 years old male patient was admitted to our 

hospital because of pain, nausea, vomiting and palpable right 

lower quadrant mass. 

Analysis of intestinal segments (ileum-colon) revealed the presence 

of diverticula are and expansion of appendix. Also we 

observate stenotic areas cm 3.5 × 2. Were sampled fragments of 

the peritoneum and omentum. The samples were processed for 

histological evaluation. 

Results. The morphological features shows intraluminal mucina 

who compress the lining epithelium with epithelial displacement.

306 

The histological analysis highlights the almost complete absence 

of the mucosal layer and a residual mucosal strate exibite a low 

grade dysplasia. 

Histopathological examination of the lesion was consistent with 

gigant appendicular cistoadenoma. 

Discussion. Appendiceal mucocele is a rare entity characterized 

by a gross enlargement of the appendix from accumulation of 

mucoid substance within the lumen.AM is often incidentally discovered 

either during surgery or on radiologic examination. Frequently, 

AM is associated with colorectal cancer and appendiceal 

cystadenocarcinoma. Because of its anatomic position, it should 

be considered in the differential diagnosis of adnexal masses. The 

authors emphasize that anatomo patologic and clinic diagnosis of 

an underlying malignancy in a mucocele is important for patient 

management. 

references 

Gastroenterol 2008;14(14):2280-3. 

Rev Col Bras Cir. 2009;36(2):180-2. 

G Chir 2007;28:274-6. 

Gastrointestinal stromal tumor of the caecum and 

omentum: report of a case 

1)Labate A. 2)Mazzon E. 3)Lo verde P. 4)Certo G. 5)Straci S. 

1)Anatomia Patlogica, Clinica Cappellani Giomi Spa, Messina, Italia 

2)Clinico Sperimentale Di Medicina E Farmacologia, Policlinico 

G.Martino, Messina, Italia 3)Anatomia Patologica, Clinica Cappellani 

Giomi Spa, Messina, Italia 4)Anatomia Patologica, Clinica Cappellani 

Giomi Spa, Messina, Italia 5)U.F. Di Chirurgia Generale, Iomi Giomi, 

Messina, Italia 

Introduzione. GISTs are mesenchymal tumors, that occur most 

frequently in the stomach (60-70%), jejunum and ileum (20-25%), 

and less frequently in the duodenum, colorectal ((5%), esophagus 

and appendix (1%). Most GIST are positive for CD117. 

We present the case of a patient who developed acute intestinal 

obstruction. 

Case report. A 61 years old male patient was admitted to our 

hospital with an intra-abdominal mass and developed acute intestinal 

obstruction. 

We evaluated samples of a small intestine, caecum and omentum 

biopsy. The specimens were fixed in 10% buffed formalin, and 

paraffin embedded. Parraffin sections were stained with hematoxilyn 

and eosin for histological evaluation. For immunohystochemical 

studies, section were incubated with anti Ki67 and 

c-Kit. 

Results. Gross features: the lesion shows a mass of 8 × 6 cm 

surrouds the bowel histologically, the tumor is composed of 

spindle-shaped cells with an interlacing bundle pattern. the nuclei 

are larger and particulary in omental node, show numerous mitoses.(> 

15-20HPF). Immunohistochemical examination showed 

that it was positive for c-kit and hight expressin for Ki67 The 

lesion was consistent with caecum GIST. 

Discussion. Gastrointestinal stromal tumors (GIST) are the most 

common mesenchymal tumors of the gastrointestinal tract. Surgical 

resection is the primary treatment of GISTs. Radiotherapy 

and chemotherapy are generally ineffective. The classic GIST 

is a small lesion (< 5 cm) and usually were dividet in to six 

categories. Significants prognostic fetures included tumor size 

(< 5cm), mitosis (< 5/10 HPF), presence of necrosis and epithelioid 

aspects. We describe very unusual case of a gastrointestinal 

stromal tumor (GIST) of caecum and omentum who shoved hight 

maglignat features, 

references 

Surg Today 2001;31(8):715-8. 

World J Surg Oncol 2007;5:66. 


MlH1 promoter methylation and BrAf mutation 

in colorectal carcinomas with defective DNA 

Mismatch repair 

1)Lanza G. 2)Ulazzi L. 3)Maestri I. 4)Gafà R. 





Background. Recent studies indicate that analysis of MLH1 promoter 

methylation and BRAF gene mutation can be employed to 

differentiate hereditary (Lynch syndrome) from sporadic MSI-H 

MLH1-negative colorectal carcinomas. 

Methods. Mismatch repair (MMR) status has been prospectively 

evaluated by immunohistochemical analysis of MMR 

protein expression (MLH1, MSH2, MSH6 and PMS2) and 

microsatellite instability (MSI) investigation in 1978 colorectal 

adenocarcinomas surgically resected from January 2004 to December 

2009. MSI status was determined by a fluorescent PCR 

method using the Bethesda panel markers (BAT25, BAT26, 

D2S123, D5S346, D17S250) plus BAT40. In MMR-deficient 

(MMR-D) tumors, MLH1 promoter methylation was assessed 

by methylation specific PCR and V600E BRAF mutation by 

direct sequencing. 

Results. 290 (17.2%) carcinomas were classified as MMR-D 

(loss of MMR protein expression and/or MSI-H). Most MMR-D 

tumors showed loss of MLH1 expression (237, 81.7%). MLH1 

methylation was detected in 191/214 (89.3%) MLH1-negative 

carcinomas and in 2/46 (4.3%) MMR-D MLH1-positive carcinomas. 

V600E BRAF mutations were observed in 102/152 (67.1%) 

MLH1-negative and in only 1 of 36 (2.8%) MLH1-positive 

MMR-D cancers. BRAF mutations were identified only in tumors 

showing MLH1 promoter methylation. 

Conclusions. Our data confirm that assessment of MLH1 promoter 

methylation and of BRAF mutation status might be used 

in the selection of colorectal cancer patients with presumptive 

Lynch syndrome. 

Oncocytic lipoadenomatous tumours of salivary 

glands 

1)Lega S. 2)Casadei GP. 3)Collina G. 4)Salerno A. 5)Berni canani 

A. 6)Dall’Olio D. 

1) Anatomia Patologica, Dipartimento Oncologico, Maggiore, Bologna, 

Italia 2) Anatomia Patologica, Dipartimento Oncologico, Maggiore, Bologna, 

Italia 3) Anatomia Patologica, Dipartimento Oncologico, Maggiore, 

Bologna, Italia 4) Anatomia Patologica, Dipartimento Oncologico, Maggiore, 

Bologna, Italia 5) Dipartimento di Neuroscienze, Maggiore, Bologna, 

Italia 6) Dipartimento di Neuroscienze, Maggiore, Bologna, Italia 

Context. Oncocytic change in salivary gland is a frequent event 

in neoplastic and non-neoplastic conditions. A recently described 

peculiar rare type of salivary gland tumor is the oncocytic lipoadenoma. 

Histologically it shows an adenomatous epithelial component 

of oncocytic cells intermingled with abundant adipous 

tissue, delimited by a delicate fibrous capsule. Its first description 

was reported in 1998, in submandibular gland. 

Case report.We report three cases of oncocytic lipoadenoma observed 

in our institution. The median age of the patients was 66,6 

and the male/female ratio was ½. Two patients had the tumor in 

the left parotid gland and a female patient had the tumor in right 

submandibular gland. All the tumor were soft, echographically 

well defined with the exception of a submandibular tumor that 

showed unomogeneous mass. All tumors had a fine, delicate 

fibrous capsule and showed a population of epithelial cells with 

oncocytic features consisting in large, brightly eosinophilic granular 

cytoplasm and ovoid nuclei with an evident nucleolus, intermingled 

with mature adipous tissue, in clusters or sheets separated 

from the oncocytic cells, or intimately intermingled with them.


No necrosis, atipia, pleomorphism and mitosis were evident. The 

oncocytic cells were positive for CKpan, CK7, CK5/6 and for 

mitochondrial antigens. In one case focal peripheral cells were 

p63 positive. CK 20 was negative too. Immunostains for SMA, 

S-100 protein were negative: scattered adipocytes were positive 

for S-100 protein. PTAH was performed in case one g 

Conclusion. Our cases recapitulate the histologic findings described 

in the literature. 

Preoperative studies were not specific and fine needle aspiration 

cytology was useful in two cases when oncocytic cells were evident 

suggesting an oncocytic tumor. Surgical resection was not 

complicated and the follow-up was uneventful after three years in 

oldest case, confirming the benign nature of the tumor. 

Trabecular papillary carcinoma of thyroid with 

metaplastic squamous change. A case report 

1)Lega S. 2)Collina G. 3)Cremonini N. 4)Casadei G.P. 

5)Bondi A. 

1) Anatomia Patologica, Dipartimento Oncologico, Maggiore, Bologna, 


Italia 3) Dipartimento Medico, Maggiore, Bologna, Italia 4)Anatomia 

Patologica Dipartimento Oncologico, Ospedale Maggiore, Bologna, 


Italia 

Context. Papillary carcinoma of thyroid (PTC) is a well-differentiated 

malignant tumor of follicular cells that shows a set of 

characteristic nuclear features. In two thirds of tumors, papillary 

growth predominates, whereas another one third of tumors have a 

predominantly follicular architecture. In addition to classic PTC, 

more than 10 different histologic variants has been described. 

Case report. We report an unusual case of thyroid carcinoma 

which occurred in a 43 year-old woman. She had a cystic lesion 

measuring 2 cm in diameter with a mural nodule of 0,8 cm. A 

FNAC was performed and a follicular lesion was suggested (TYR 

4). The mural nodule had unusual trabecular pattern but nuclear 

features of PTC. At perifery of the tumor there were spindled 

simil-sarcomatous stroma and squamous areas. 

On immunoistochemistry the PTC was positive with TTF1 which 

stained also the squamous areas, whereas the simil-sarcomatous 

stroma was negative for this antibody, while it stained for smooth 

muscle actin. The squamous areas also were positive for CK17 

and CK19. 

Conclusions. We report an unusual case of thyroid carcinoma 

with trabecular pattern associated with squamous metaplasia of 

thyroid cells immersed in a simil-sarcomatous stroma. The possibility 

of follicular carcinoma was ruled out for the presence of 

nuclear features consistent with PTC. The eventuality of a true 

squamous cell carcinoma, which is a rare thyroid tumor composed 

entirely of cells with squamous differentiation, was taken 

in to account, but the squamous cells present in this case were laking 

of nuclear atipia and/or necrosis and therefore a metaplastic 

process is favoured. Nevertheless cases of spindle cell squamous 

carcinoma of the thyroid have been reported associated to tall cell 

variant of PTC and it has been suggested that they may have a 

worse prognosis. Therefore is important to assess the metaplastic 

nature of squamous cells in a thyroid carcinoma to better define 

the therapeutic strategies. 

The virtual slide in a digital atlas: regional PACS for 

pathologists 

Lega S., Pierotti P., Crucitti P., Bondi A. 

Anatomia Patologica, Dipartimento Oncologico, Maggiore, Bologna, Italia 

Background. A project on Virtual Microscopy and Digital Pathology 

has been conducted in Emilia-Romagna and it has been 

planned for the promotion and the quality assessment in screening 

cytology and histology for the prevention of the tumors of 

307 

uterine cervix, breast and colon-rectum cancers. Object of this 

study is to realize a software to manage cytological and histological 

whole-slides digital images and related clinical data and 

to build a picture archive and communication system (PACS) 

among pathologists of the Region. The cases collected and 

cataloged in an online, systematic digital archive of slides, can 

be used as diagnostic reference tool in order to create a casistic 

Atlas online. 

Methods. Rare specimens and slides are recorded in digital Spectrum 

Aperio, a program that provides three levels of hierarchical 

organization, Project or Case⇒Specimens⇒Digital Slides. A 

project can contain many specimens connected to several slides. 

File attachments is allowed at any level: images, documents, 

tracks, broadcasts specimens and slides, according to logical diagnostic 

hierarchies. Cases are cataloged and indexed with NAP 

codes, a Nomenclature derived from SNOMED. 

Results. More than 500 cases, covering a wide range of histopathology 

(especially cancer) are already saved into our archive 

with the Spectrum Aperio interface and then viewed and coded 

according to NAP system. Retrievals to find slides and cases 

associated with a coded diagnosis are very simple and can be 

made either in Italian or English language. Once selected a case, 

the related digital slides can be viewed online and discussed 

simultaneously by different distant users, and complemented 

with additional clinical and pathological data (as gross and radiological 

images), added to perform various types of further 

processing. Atlas is a versatile instrument, indeed it can be used 

for teleconsultation, education, research and quality control, and 

it can be continuously improved and enriched with new cases or 

new attachements. 

The pathologist and colorectal cancer: 

an integrated multidisciplinary prospective analysis 

1)S.A. Senatore, 2)G. Leo, 2)M. Pisanò, 2)S. Malerba, 3)E. Molina, 

1)F. Floccari, 1)E. Villani, 1)A. Caldarazzo, 1)A. D’Amuri 

1)U.O.C. Anatomia Patologica, “Ospedale Sacro Cuore di Gesù”, P.O. 

Gallipoli ASL Lecce, Gallipoli, Italia; 2)Laboratorio di Biologia Molecolare 

e Oncologia Sperimentale, “Oncologico Giovanni Paolo II”, P.O. 

Vito Fazzi ASL Lecce, Lecce, Italia; 3) Dipartimento di Anatomia Umana, 

Farmacologia e Scienze Medico-Forensi, “Università degli Studi di Parma”, 

Parma, Italia 

Background. Colorectal carcinoma (CRC) is a malignant neoplasia 

which frequently develops a fatal course caused by 

metastases. EGFR, frequently overexpressed in CRC with an 

important role in tumor aetiology and progression, lead to apply 

anti-EGFR targeted therapies as a potent strategy in the treatment 

of metastatic CRC (mCRC). These therapies are effective only in 

a subset of patients. KRAS gene mutations that activate the RAS/ 

RAF/MEK/ERK pathway are associated with a poor response 

to these treatments. KRAS status can predict wich patient may 

or may not benefit of anti-EGFR therapy. The aim of this study 

was the relationship between morphological and functional markers 

in mCRCs, mainly the oncogenic activation of the signaling 

pathway that impairs the tumor response to anti-EGFR antibody 

therapies, in order to clarify Pathologist’s role. 

Methods. In surgical tissue specimens from patients with histologically 

confirmed CRCs metastasis before anti-EGFR targeted 

therapies application, Tumor grading, staging, clinical and 

pathological characteristics identified by the EGFR and ERK ½ 

detections by immunohistochemical (IHC) methods, KRAS and 

BRAF mutational status using Pyrosequencing technology were 

assessed. 260 and 89 mCRCs samples were analized respectively 

for KRAS and BRAF. 

Results. ERK ½ IHC reactions revealed positive staining in 

28,2% of cases. KRAS mutations occurred in 78/260 cases (30%), 

particularly in codon 12 (77%) and 13 (21,8%). BRAF mutations 

occurred in 6/89 samples at codon 600 (V600E) (6,7%). Our data

308 

demonstrated no significative relations between morphologic and 

EGFR IHC staining and molecular obtained results. IHC EGFR 

staining data obtained, suggest to consider it as a step in multiparametric 

evaluation, not predictive of treatment efficacy. We 

observed KRAS and unrelated BRAF mutations with IHC EGFR 

status in each patient. IHC ERK ½ staining appear as a functional 

prognostic and predictive parameter strictly related with KRAS 

mutated forms. 

The biological bank of malignant mesothelioma 

1)Libener R. 2)Orecchia S. 3)Bensi T. 4)Trincheri N. 5)Arnolfo 

E. 6)Salvio M. 7)Ugo F. 8) Mariani N. 9) Betta PG. 

1)Pathology, A.O., Alessandria, Italy 2)Pathology, A.O., Alessandria, Italy 




9) Pathology, A.O., Alessandria, Italy 

Background. Due to its relatively low incidence rate in the past 

when compared with other major cancers, MM has been neglected 

by the medical community for nearly 40 years. However, it is 

now necessary, especially in the light of the recent steep rise in 

the incidence of MM, that a databank be established in order 1. to 

understand the natural history of the disease and the outcome both 

of treatments currently in use and of those still to be developed, 

2. to better evaluate the benefits of the various current treatment 

approaches, and 3. to better test and evaluate the effectiveness 

of novel therapies in the future. In addition, a parallel biobank 

would be invaluable in the analysis of tumours for more rational 

treatment planning in the future. 

Methods. The MM biobank established at the Alessandria City 

Hospital with the contribution of S. Spirito Hospital of Casale 

Monferrato aims at providing MM pleural, pericardial and peritoneal 

tissue samples along with blood and DNA samples and 

with associated clinical, pathology, recurrence, follow-up and 

treatment data to meet the growing need of cancer research community 

especially in the setting of translational studies. 

Results. At the moment, the bank holds 288 consecutive paraffin-fixed 

tissue specimens of “definite” MMs (epithelioid: 211, 

mixed: 47, sarcomatoid: 30), 195 blood specimens stored at 

-80°C and 65 MM cell lines (established from serous effusions) 

in liquid nitrogen.The main use of this material is to investigate 

biological mechanisms of MM and to develop tools and strategies 

for both early diagnosis and targeted therapy of this neoplasm. 

In particular the following molecules are currently investigated 

at molecular levels using cell lines and tissue specimens: 1. 

AZD 6474 - Vandetanib (Zactima TM ), a tyrosine kinase inhibitor 

targeting VEGFR and EGFR; 2. AZD1152, a selective inhibitor 

of Aurora B Kinase; 3. RAD001 (Everolimus), an inhibitor of 

the mammalian Target of Rapamycin (mTOR), a key regulatory 

kinase. 

Perivascular epithelioid cell tumor (PeComa) 

of the gallbladder. first case report 

1)Liberati F. 2)Ventura L. 

1)Anatomia patologica, San camillo de lellis, Rieti, Italia 2)Anatomia patologica, 

San salvatore, L’Aquila, Italia 

Background. PEComas have been described in many different 

locations, including kidney, bladder, prostate, uterus, lung, pancreas 

and liver. We describe the presence of a PEComa arising in 

the gallbladder wall. 

Methods. The patient was a 58-year-old woman, with abdominal 

pain, nausea and vomiting. An abdominal ultrasound scan 

showed a distended gallbladder with a suspect stone embedded 

in the neck region and a laparoscopic cholecystectomy was then 

performed. 


The gallbladder measured 6.5 cm in length and presented a 

2 × 1.5 × 1.2 cm white-grayish nodule in the wall of the neck 

region. No stones were found. Routine processing and embedding 

was performed to obtain sections stained with hematoxylin-eosin, 

α-smooth muscle actin, desmin, S100 protein, HMB45, CD117, 

CD34 and Ki67. 

Results. A well-circumscribed but unencapsulated tumor was 

located outside the muscularis propria, composed by sheets of 

spindle and plump epithelioid cells with abundant pale eosinophilic 

cytoplasm, with a rich vascular network. Tumor cells were 

positive for HMB-45, actin and desmin; negative for CD117 

and CD34. S100 was positive in rare mature fat cells within 

the neoplasm. Ki67 proliferation index was 1%. These features 

were consistent with those of an extrarenal angiomyolipoma 

(PEComa). Chronic cholecystitis and pseudopyloric metaplasia 

were also present. No additional lesion was identified after a 

subsequent abdominal CT scan. 

Conclusions. We reported the first case of a PEComa originating 

in the gallbladder, suggesting that mesenchymal tumors of 

this organ may show PEC differentiation. The identification of 

a PEComa in the gallbladder is not surprising because similar 

lesions are well known in liver and pancreas locations and occasionally 

described in the common bile duct. The presence of a stenosing 

tumor in the neck region may have induced the symptoms 

in the absence of obstructing stones. One year after the diagnosis 

the patient remains free of disease. 

Human papillomavirus (HPV) infection in head and 

neck squamous cell carcinomas (HNSCC) 

1)Lombardi M. 2)Campanini N. 3) D’Adda T. 4) Pizzi S. 5) 

Corcione L. 6) Dal bello B. 7) Sesenna E. 8) Tinelli C. 9)Lanfranco 

D. 10)Poli T. 11)Silini EM. 

1)Patologia e medicina di laboratorio sez anat patol, Azienda ospedaliero-universitaria, 

Parma, Italia 2)Patologia e medicina di laboratorio sez 

anat patol, Azienda ospedaliero-universitaria, Parma, Italia 3)Patologia 

e medicina di laboratorio sez anat patol, Azienda ospedaliero-universitaria, 

Parma, Italia 4)Patologia e medicina di laboratorio sez anat 

patol, Azienda ospedaliero-universitaria, Parma, Italia 5)Patologia e 

medicina di laboratorio sez anat patol, Azienda ospedaliero-universitaria, 

Parma, Italia 6)Unità operativa anatomia patologica, Fondazione 

irrcs policlinico san matteo, Pavia, Italia 7)Scienze ot-od-oft e cerv-facc 

uo maxillo, Azienda ospedalieo-universitaria, Parma, Italia 8)Unità 

operativa Biostatistica, Fondazione irrcs policlinico san matteo, Pavia, 

Italia 9)Scienze ot-od-oft e cerv-facc uo maxillo, Azienda ospedalieouniversitaria, 

Parma, Italia 10)Scienze ot-od-oft e cerv-facc uo maxillo, 

Azienda ospedalieo-universitaria, Parma, Italia 11)Patologia e medicina 

di laboratorio sez anat patol, Azienda ospedaliero-universitaria, 

Parma, Italia 

Background. Variable prevalences of HPV DNA have been 

reported in HNSCCs. Available data for oral SCCs are limited 

in particular for SCCs of the mobile tongue whose incidence is 

increasing among young subjects. 

Methods. We analyzed a retrospective series of 203 SCCs (133 

males, mean age 66 yrs, stage 3-4 54%) of the oropharynx (36) 

and oral cavity (167). HPV typing was performed by the INNO- 

LiPA HPV assay on DNA extracted from archival tissue. Expression 

of p16, p53 and EGFR was detected by immunostaining. 

p16 promoter methylation was evaluated by methylation-specific 

PCR; loss of heterozygosity (LOH) was assessed with microsatellites 

D9S157 and D9S171. Outcome measures were analyzed by 

conditional regression analysis. 

Results. The use of a highly sensitive HPV typing assay evidenced 

a higher prevalence (69,4%, 59,7% HR-HPV) and a wider spectrum 

of types than previously reported. The most common genotypes 

were HPV31, 33, 52, 35, 6, 16 and 39; 38% of infections were 

multiple. Reactivity was observed in 23% of cases for p16, 70% 

p53 and 57% EGFR; p16 was methylated in 56% of cases. p16 im-


munostaining was not a reliable surrogate for HR-HPV infections. 

HPV-DNA status did not correlate with demographical variables, 

grade or stage, site, p16 staining, p16 methylation, LOH or survival. 

HR-HPVs were more frequent among men in the oropharynx 

(p < 0.001) whereas LR-HPVs were more frequent among 

women (< 0.005), > 40 years (p < 0.05) and in oral sites other 

than the tongue (p < 0.01). p16 staining correlated with distinct 

morphology including basaloid features. p16 methylation was 

more frequent in the tongue (p < 0.005) and correlated with HPV 

status (p < 0.005). The overall 10 year mortality was 39% and 

correlated with established pathological variables and lack of p16 

methylation in both univariate (p 0.017) and multivariate analysis 

(HR 1.8, p 0.026). 

HPV types and modes of p16 inactivation vary according to 

sex and site suggesting difference in pathogenetic mechanisms 

among HNSCCs. 

Solitary fibrous tumour of the breast parenchyma: 

report of a case with emphasis on needle core 

biopsy-based diagnosis 

1)G.M. Vecchio, 1)L. Salvatorelli, 1)F. Longo, 2)B. Cavanaugh, 

3)J. Palazzo, 1)G. Magro 



Catania, Italia; 2)Department Radiology, Division of Breast Imaging, 

Thomas Jefferson University Hospital, Philadelphia, USA; 3)Department 

of Pathology, Division of Pathology Thomas Jefferson University Hospital, 

Philadelphia, USA 

Background. Solitary fibrous tumour (SFT) is a relatively uncommon 

spindle cell neoplasm that typically occurs in the pleura. 

Currently SFT is believed to occur anywhere, including soft tissues 

and viscera. The diagnosis of SFT at extra-pleural sites may 

be challenging, especially when evaluating small biopsies and 

thus surgical excision is usually required for a correct interpretation. 

Only a few cases of SFT have been reported in the breast 

parenchyma, but in none of these the diagnosis was rendered by 

needle core biopsy. 

Materials. We report the first case of SFT of the breast diagnosed 

by needle core biopsy, discussing differential diagnosis with 

other spindle cell tumour- and tumour-like lesions. 

Results. A 62-year-old woman presented, within the upper 

outer left breast, a 1 cm mass with obscured margins on spot 

compression views. Ultrasound demonstrated hypoechoic mass 

with circumscribed margins and internal vascularity. Ultrasound 

guided percutaneous vacuum assisted biopsy was performed 

and demonstrated a proliferation of bland-looking, CD34 + 

spindle cells with intervening with thick collagen bands and 

medium-sized blood vessels with hyalinization of their walls. 

Some of these vessels had a hemangioperictyoma-like pattern. 

Based on morphological and immunohistochemical profile, a 

diagnosis of SFT was proposed. A lumpectomy was performed 

and the diagnosis of SFT was confirmed. Histologically, an 

unencapsulated, well circumscribed spindle cell tumour was 

seen. Neoplastic cells had pale cytoplasm and oval- to spindleshaped 

nuclei with one small nucleolus. Notably, thick collagen 

bands were scattered among cells which were closely packed 

and haphazardly arranged (patternless). Apart diffuse CD34 

immunoreactivity, neoplastic cells were variably stained with 

CD99, bcl-2 and ER. Differential diagnosis included a wide 

list of benign and malignant spindle cell lesions that can occur 

in the breast, especially including nodular fasciitis, spindle 

cell lipoma, myofibroblastoma, inflammatory myofibroblastic 

tumour, leiomyosarcoma, fibrosarcoma/malignant fibrous histiocytoma 

and fibromatosis/nodular fasciitis-like low-grade 

sarcomatoid/metaplastic carcinoma. 

309 

The diagnosis of most SFT is straightforward, but some lesions 

are not easy to diagnose if seen out of the usual anatomic context 

in which SFT is not expected to occur. Although radiologic 

images of SFT of the breast are non-specific, the present case 

emphasizes the diagnostic role of needle core biopsy. 

Vascular changes in 14 autopsies of children 

with HIV infection 

1)V.G.S. Lopes, 2)G.V.H. Herdy, 1)E.P. Dias, 1)R. Granato, 3)R. 

S. Gomes, 3)G.H. Nascimento, 3)A.C.S. Lopes, 3)R.F.N. Abreu, 

3)P. Casquilho, 1)A.C.D. Silva 

1)Departamento de Patologia, Hospital Universitário Antônio Pedro, Niterói, 

Brasil; 2)Departamento Materno Infantil, Hospital Universitário 

Antônio Pedro, Niterói, Brasil; 3)Departamento de Clínica Médica, Hospital 

Universitário Antônio Pedro, Niterói, Brasil 

Background. The manifestations of human immunodeficiency 

virus (HIV) infection vasculitides are one of the less common 

but nonetheless important consequences. The vasculopathy in 

patients with acquired immunodeficiency syndrome (AIDS) have 

a multifactorial etiology, however research have shown that infection 

results in marked functional and morphologic changes to 

the vascular endothelium. 

Methods. We evaluated 14 children autopsies performed in the 

decade of 1990 with clinical and laboratotial of HIV infection. 

We analyzed the macro and microscopy with emphasis on vascular 

changes. 

Results. There were no vascular changes in the macroscopic 

point of view. Light microscopy revealed vascular lesions in 

10 cases. Of these, six had lesions consistent with HIV vasculopathy. 

In one, was described pulmonary involvement and, in 

others, changes in various organs like heart, pancreas, kidney and 

thyroid. Vascular changes consisted of lymphocytic infiltrate, 

vascular calcification of the middle layer, fragmentation of the 

elastic layer and sub-intimal fibrosis. 

Conclusions. Vasculitis in an HIV positive patient is an uncommon 

but important disease that might manifest as an organ based 

disease process. In the cases observed, vascular changes reflect 

evolutionary stages of the same process. 

Pulmonary pathologic findings in autopsy 

of newborn with influenza A/H1N1 viral infection: 

case report 

1)V.G.S. Lopes, 2)S.H. Villela, 2)J.A.M. Junior, 2)S. Sias, 1)E.P. 

Dias, 3)R.S. Gomes, 3)G.H. Nascimento, 2)A.C.S. Lopes, 1)C.L. 

Lopéz, 1)R.A. Granato 

1)Patologia, Hospital Universitário Antônio Pedro, Niterói, Brasil; 2)Pediatria, 

Hospital Universitário Antônio Pedro, Niterói, Brasil; 3)Clínica 

Médica, Hospital Universitário Antônio Pedro, Niterói, Brasil 

Background. In March 2009, a novel swine-origin influenza A/ 

H1N1 virus was identified and the World Health Organization declared, 

in June, the first influenza pandemic in 41 years. There are 

few reports of the pathologic findings, although the H1N1 virus 

infection has assumed pandemic proportions. We described the 

clinicopathological characteristics with emphasis on pulmonary 

pathological findings of one premature newborn autopsy who 

had died by respiratory failure due to H1N1 infection confirmed 

by real-time reverse-transcription polymerase chain reaction on 

nasopharyngeal swab sample. 

Methods. We analyzed the clinical record, laboratory tests and 

the autopsy study of one premature newborn who had died for 

respiratory failure for H1N1 virus infection. 

Results. Were noted bronchitis, bronchiolitis, pneumonitis, diffuse 

alveolar damage proliferative phase, emphysema and secondary 

pulmonary hypertension. Multiple infarctions myocardial, 

fibrosis and hypoplastic thymus, spleen congestion and hemor-

310 

rhage, acute tubular necrosis, metabolic disease of the liver, 

kidney and heart, cerebral edema and hypoxic neuronal changes 

also were observed in autopsy. 

Conclusions. The pathologic findings of pulmonary disease 

caused by H1N1 infection are similar to findings of injuries 

caused by the influenza virus group, thus, corroborating with 

what was identified in past pandemics. 

Nodular lymphocyte predominant Hodgkin 

lymphoma in two siblings affected by Hermansky 

Pudlak type 2 syndrome 

1)Lorenzi L. 2)Vermi W. 3)Badolato R. 4)Lonardi S. 5) Rossini 

C. 6)Medicina D. 7)Fappani L. 8)Bossini P. 9)Fisogni S. 10)Parolini 

S. 11)Facchetti F. 

1)I Servizio Anatomia Patologica, Spedali Civili Di Brescia, Università 

degli Studi di Brescia, Italia 2)I Servizio Anatomia Patologica, Spedali Civili 

Di Brescia, Università degli Studi di Brescia, Italia 3)Clinica Pediatrica, 

Spedali Civili Di Brescia, Università degli Studi di Brescia, Italia 4)I 

Servizio Anatomia Patologica, Spedali Civili Di Brescia, Università degli 

Studi di Brescia, Italia 5)I Servizio Anatomia Patologica, Spedali Civili 

Di Brescia, Università degli Studi di Brescia, Italia 6)I Servizio Anatomia 

Patologica, Spedali Civili Di Brescia, Università degli Studi di Brescia, 

Italia 7)I Servizio Anatomia Patologica, Spedali Civili Di Brescia, Università 

degli Studi di Brescia, Italia 8)I Servizio Anatomia Patologica, 

Spedali Civili Di Brescia, Università degli Studi di Brescia, Italia 9) I 


Studi di Brescia, Italia 10)Dipartimento Scienze Biomediche E Biotecnologie, 

Università Di Brescia, Università degli Studi di Brescia, Italia 11)I 


Studi di Brescia, Italia 

Background. Primary immunodeficiencies (PID) are associated 

with higher risk to develop lymphoproliferative diseases, predominantly 

high grade non-Hodgkin B-cell lymphomas. Nodular 

Lymphocyte Predominant Hodgkin Lymphoma (NLPHL) has 

been reported in patients with Autoimmune Lymphoproliferative 

Syndrome (ALPS), a PID resulting from anomalies of the apoptotic 

cascade, particularly in cases with Fas gene exon 9 mutations. 

Here we report the first two cases of NLPHL occurring in 

Hermansky Pudlak type 2 syndrome (HPS2), a rare PID due to 

mutations on the β3A gene, causing impairment of the adaptor 

protein 3 complex and protein sorting to lysosomes. Since defects 

of cytotoxicity mediated by NK cells [Fontana S, Blood 2006] 

and CTL [Clark RH, Nat Immunol 2003] have been reported in 

HPS2, we analyzed the expression of cytotoxic proteins in these 

patients. 

Patients and Methods. Two HPS2 siblings, female and male, 

with proven defective NK cytotoxicity [Fontana et al., Blood 

2006], developed at the age of 10 and 8 years cervical and retroperitoneal 

lymphadenopathy with typical features of NLPHL. 

Germline mutations of exon 9 of the Fas gene were investigated 

on peripheral lymphocytes, while FAS expression, as well as 

Perforin (Prf) and Granzyme B (GrB) content in CD56+ NK 

cells and CD8+ CTL were evaluated with single or double immunostains; 

5 NLPHL cases from immune competent subjects 

were used as controls. 

Results. HPS2-NLPHL lacked germline mutations of the Fas 

gene and showed expression of FAS on LP cells similar to controls. 

In contrast, Prf and GrB content in CD56+ NK cells and 

CD8+ CTLs was significantly higher in HPS2-NLPHL compared 

to controls (p < 0.01). The abnormal accumulation of cytotoxic 

proteins in NK/CTLs confirms, in vivo, their defective release in 

HPS2; this observation discloses novel immunological mechanisms 

of cancer immune surveillance in NLPHL. 


A novel G48r/D92N H-rAS mutation identified in a 

child with an atypical spitzoid tumor 

1)Lorenzoni A. 2)Simi L. 3)Pinzani P. 4)Tomasini C. 5)Orlando 

C. 6)Santucci M. 7)Massi D. 

1)Dip Area Critica Med Chir Sez Anat Patologica, Univ Di Firenze Aouc 

Careggi, Firenze, Italia 2)Clinical Physiopathology,Clin Biochemistry 

Unit, Univ Di FirenzeAouc Careggi, Firenze, Italia 3) Physiopathology,Clin 

Biochemistry Unit, Univ Di Firenze Aouc Careggi, Firenze, Italia 4)Depart 

Of Med Sciences And Human Oncol,Dermat Sect, Università Di Torino, 

Torino, Italia 5)Clinical Physiopathology Clin Biochemistry Unit, 

Univ Di Firenze Aouc Careggi, Firenze, Italia 6)Dip Area Critica Med 

Chir Sez Anat Patologica, Univ Di Firenze Aouc Careggi, Firenze, Italia 

7)Dip Area Critica Med Chir Sez Anat Patologica, Univ Di Firenze Aouc 

Careggi, Firenze, Italia 

Background. Atypical spitzoid tumors, also reported as “spitzoid 

tumors of uncertain malignant potential” exhibit some of 

the morphological features of typical Spitz nevus and some of 

spitzoid melanoma, and are often difficult to classify accurately. 

In light of the diagnostic difficulties, ancillary molecular techniques 

are particularly needed. About 10% of typical Spitz nevi 

contain a copy number gain of 11p, and two-thirds of such cases 

show H-RAS mutations. There have been no reports of H-RAS 

mutations in spitzoid melanomas and, recently, H-RAS mutations 

were found in 14% of atypical Spitz nevi and in 20% of spitzoid 

tumors of uncertain malignant potential associated with a benign 

clinical behaviour. 

Methods. A 6-year-old female child presented with a slightly 

pigmented cutaneous papule 5 mm in diameter on her right leg. 

The lesion was surgically excised. The histopathological features 

were consistent with an atypical Spitz tumor, with free margins. 

No further treatment was performed. The patient is alive with no 

further evidence of disease 3 yrs and 6 months after diagnosis. 

Formalin-fixed, paraffin-embedded tissues underwent proteinase 

K overnight digestion at 56°C. DNA was extracted using the 

DNA FFPE Tissue kit (QIAGEN, Milan, Italy). BRAF V600E mutation 

was detected by real time PCR allele-specific method. DNA 

sequencing for H-RAS analysis was performed, as previously 

described. 

Results. The tissue sample carried a new pathogenic H-RAS 

double mutation on the exon 3, specifically G48R/D92N, while 

no H-RAS mutations were identified in the exon 2. The tumor was 

found B-RAF V600E mutated. Our findings support the diagnostic 

importance of H-RAS mutational status evaluation in spitzoid 

melanocytic proliferations in pediatric age, where the presence 

of a H-RAS mutation can be regarded as an additional decisive 

criterion that labels an atypical spitzoid tumor as a lesion that will 

pursue a benign clinical behaviour. 

Cribriform-morular variant of thyroid carcinoma: 

distinct immunohistochemical profile from other 

papillary thyroid carcinoma variants 

1)Lovitch S.B., 2)Faquin W., 3)Crippa S, 3)Mazzucchelli S., 

1)Nosé V. 

1)Department of Pathology, Brigham and Women’s Hospital, Boston, MA, 

USA; 2)Department of Pathology, Massachusetts General Hospital, Boston, 

MA, USA; 3)Institute of Pathology, Locarno, Switzerland 

Background. The cribriform-morular variant of thyroid carcinoma 

(CMv-TC) typically occurs as an extraintestinal manifestation 

of familial adenomatous polyposis (FAP), although rare sporadic 

cases have been reported. It occurs almost exclusively in young 

females, is well-differentiated, often multifocal, is characterized 

by cribriform, solid, and morular areas lacking typical nuclear 

features of papillary thyroid carcinoma, and is associated with 

germline and somatic mutations in the APC and beta catenin 

genes. In contrast to conventional PTC, CMv TC rarely metastasizes 

and carries a good prognosis.


Methods. We reviewed nine cases of CMv-TC seen at our two 

institutions, and performed immunohistochemical analysis on a 

total of fourteen lesions from six cases. Clinical history and genetic 

test results were obtained from the medical record. 

Results. All patients with CMv-TC were female and ranged in 

age from 18 to 51. All CMv-TC tumors showed strong aberrant 

nuclear and cytoplasmic accumulation of beta-catenin, were positive 

for CK19, p53 and Bcl-2, and showed very weak or absent 

immunoreactivity for HBME-1 and galectin-3. 

Conclusions. Immunohistochemical features of CMv TC- are 

distinct from PTC, including classical, tall cell, and diffuse 

sclerosing variants. Expression of Bcl-2 and p53 in a well-differentiated 

tumor is particularly surprising, as these markers in 

thyroid tumors indicate dedifferentiation in PTC. Our findings 

suggest that CMv-TC should be classified as a distinct category 

of thyroid carcinoma arising in a familial setting, rather than as 

a variant of PTC. 

HCV-related subcutaneous “lipoma-like” B-cell 

lymphoma: a new presentation of primary 

extranodal marginal zone B-cell lymphoma 

1)Lucioni M. 2)Boveri E. 3)Arcaini L. 4)Capello D. 5)Riboni R. 

6)Fiandrino G. 7)Berti E. 8)Gaidano G. 9)Lazzarino M. 10)Paulli 

M. 


Italia 2)Anatomia Patologica, Fondazione Irccs Policlinico San Matteo, 

Pavia, Italia 3)Ematologia, Università Di Pavia/Irccs Policlinico San 

Matteo, Pavia, Italia 4)Ematologia, Università Del Piemonte Orientale, 

Novara, Italia 5)Anatomia Patologica, Fondazione Irccs Policlinico San 

Matteo, Pavia, Italia 6)Anatomia Patologica, Università Di Pavia/Irccs 

Policlinico San Matteo, Pavia, Italia 7)Dermatologia, Università Milano 

Bicocca, Milano, Italia 8)Ematologia, Università Del Piemonte Orientale, 

Novara, Italia 9)Ematologia, Università Di Pavia/Irccs Policlinico San 

Matteo, Pavia, Italia 10)Anatomia Patologica, Università Di Pavia/Irccs 

Policlinico San Matteo, Pavia, Italia 

Background. Chronic antigenic stimulation from infectious 

agents is pathogenetically related to various lymphoproliferative 

disorders which often primarily arise at extranodal sites. Hepatitis 

C virus (HCV) infection has been linked to increased incidence 

of lymphoid neoplasms. Among these, marginal zone B-cell 

lymphoma (MZL) represents one of the most frequent entities, 

particularly in its splenic variant. 

Methods. We describe a series of 12 HCV+ patients (median 

age 69,5 years) with extranodal MZL presenting in the form of 

single or multiple subcutaneous nodules, most of which mimicked 

subcutaneous lipomas clinically. Diagnosis was established 

according to the criteria for extranodal MZL established 

by the 2008 WHO lymphoma classification. PCR analysis of 

IGHV genes status was performed with subsequent automated 

sequencing. 

Results. In all cases histological examination documented MZL 

infiltration that was strictly confined to subcutaneous tissue, both 

with diffuse or nodular and diffuse pattern. Epidermis, dermis 

and skin adnexa were entirely spared. Molecular analysis on 17 

biopsy samples (obtained from 9 patients) showed functional 

IgH gene rearrangements in all cases, with detection of somatic 

mutations in 82%, thus suggesting that this subset of subcutaneous 

lymphomas are histogenetically related to B-cells that have 

experienced germinal center reaction. Staging procedures at 

diagnosis did not show any other MALT-site or nodal localization. 

In 2 patients response was achieved with antiviral treatment. 

Subsequent extra-cutaneous spread to MALT sites occurred in a 

single case and the patient actually died of disease. Our observations 

expand the spectrum of HCV-associated lymphomas, to 

include a subset of extranodal MZL characterized by a novel 

primary “lipoma-like” subcutaneous presentation and indolent 

clinical course. 

Detection of eGfr mutation in histological and 

cytological samples of non small cell lung cancer 

311 

Lupi C., Sensi E., Capodanno A., Alì G,, Giordano M., Boldrini 

L., Fontanini G. 

Dept. of surgery, S. chiara hospital, Pisa, Italy 

Background. Lung cancer is the leading cause of cancer deaths 

worldwide for both men and women. Epidermal growth factor 

receptor (EGFR) is a tyrosine kinase transmembrane receptor 

that plays a role in survival and cell proliferation. Somatic mutations 

in EGFR are present in 10% of nonsmall cell lung cancers 

(NSCLC), are preferentially found in never-smokers, women, 

east Asians and adenocarcinomas, and predict response to the 

EGFR-specific tyrosine kinase inhibitors (EGFR-TKI) in patients 

with NSCLC. Lung cancer diagnosis is often based on cytology 

alone. However, almost all published data on EGFR gene mutation 

analyses were obtained from surgical samples and biopsies. 

This study tested the feasibility of EGFR gene mutation analyses 

on cytological specimens. 

Methods. EGFR mutation analyses were performed by direct 

gene sequencing on 40 histological specimens, including 25 

surgical samples and 15 biopsies, and 21 cytological samples, 

including 15 fine needle aspirations (FNA), 2 brushing, 1 bronchoalveolar 

lavage, 2 pleural fluid and 1 cell block sample. 

Results. EGFR mutation was detected in 10 of 61 samples 

(16.4%); particularly 5 (5/40, 12.5%) histological samples and 

5 (5/21, 23.8%) cytological specimens were mutated. These 

mutations included: the well-described deletion E746_A750 in 

exon 19 (3 cases, 2 surgical and 1 FNA specimens); the point 

mutation L858R in exon 21 (1 cell block sample) and the previously 

described E746_E749delinsY (2 cases, 1 surgical and 1 

FNA samples) and L747_E749delA750P (2 surgical samples) in 

exon 19. Finally, 2 samples (1 FNA, 1 pleural fluid) showed two 

different known insertions in exon 20: P772_H773dupinsA and 

H773_V774insNPH. 

This study demonstrated that sequence-based testing of cytological 

material can show equivalent, if not higher, sensitivity for mutation 

detection when compared with histological specimen. 

Villous adenoma of the urinary bladder: 

morphological features predicting of recurrence 

1) Maccio L. 2)De Gaetani C. 3)Reggiani Bonetti L. 4)Schirosi L. 

5)Sartori G. 6)Bagni I. 7)Sighinolfi M.C. 8)Bianchi G.P. 

1)Dipartimento ad Attività Integrata di Laboratori, Anatomia Patologica 

e Medicina Legale; Policlinico, Modena, Italia 2)Dipartimento ad Attività 

Integrata di Laboratori, Anatomia Patologica e Medicina Legale; Policlinico, 

Modena, Italia 3)Dipartimento ad Attività Integrata di Laboratori, 

Anatomia Patologica e Medicina Legale; Policlinico, Modena, Italia 

4)Dipartimento ad Attività Integrata di Laboratori, Anatomia Patologica 

e Medicina Legale; Policlinico, Modena, Italia 5)Dipartimento ad Attività 

Integrata di Laboratori, Anatomia Patologica e Medicina Legale; Policlinico, 

Modena, Italia 6)Dipartimento ad Attività Integrata di Laboratori, 

Anatomia Patologica e Medicina Legale; Policlinico, Modena, Italia 7) 

Dipartimento di Urologia, Az Universitaria Policlinico, Modena, Italia 8) 

Dipartimento di Urologia, Az Universitaria Policlinico, Modena, Italia 

Background. The occurrence of villous adenomas in urinary 

tract is uncommon. Prognosis is excellent, being transurethral 

resection curative. However, local recurrences such as malignant 

transformation have been described. 

Methods. We collected all cases of villous adenoma of the 

urinary bladder, diagnosed during the period 1991-2009 in the 

Institute of Pathologic Anatomy of Modena. We evaluated morphology 

(growth pattern, grade of dysplasia and mitoses) and the 

presence of concomitant foci of carcinoma in specimens obtained 

from transurethral resection. Non-neoplastic changes such as 

glandular or squamous metaplasia and inflammatory process are 

noted. Immunostaining for CDX2, CK7, CK2, CEA, PSA were

312 

performed in order to exclude tumor from other sites. MIB-1 

index was evaluated in all cases. 

Results. There were 4 male and 3 female (middle age, 77 years). 

Villous adenoma were sessile in 5 cases and semi-peduncolated 

in 2. Dimensions varied between 0,7cm to 3cm. Lesions were 

isolated in 4 cases, dislocated at the lateral wall of the bladder in 

3 case and at the trigone in 1. Multiple adenomas (≥2) occurred 

in 3 cases, simultaneously localized at the lateral-posterior walls 

and at the trigone. Histologically, all tumors showed papillary 

architecture with low grade of dysplasia (1 case), moderate 

(in 2) and severe (in 4). Closely-packed branched anatomized 

papillae, with multilayer stratificated columnar cells (> 3 cells) 

were observed in 4 cases (3 had high grade dysplasia, and 1 

moderate dysplasia). These 4 lesions were the grater in dimension 

and showed elevated MIB1 index (> 15%). Their follow-up 

revealed recurrences in 3 cases. All recurrences occurred during 

the 2 years after resection. One patient, affected by villous 

adenoma with moderate grade dysplasia developed adenocarcinoma 

within 1 year. 

Menopausal status and risk of thick melanoma 

in overweight women:correlation with estrogen 

receptor-beta expression 

1)Maio V. 2)Lorenzoni A. 3)Gori A. 4)Simoni A. 5)Crocetti E. 

6)De giorgi V. 7)Santucci M. 8)Massi D. 

1)Area Critica Med: Chirurgica Sez Anatomia Patologi, Careggi, Firenze, 

Italia 2)Area Critica Med Chirurgica Sez Anatomia Patologic, Careggi, 

Firenze, Italia 3)Divisionr Di Dermatologia Universita Di Firenze, Santa 

Maria Nuova, Firenze, Italia 4)Dipartimento Area Critia Med.Chirurgica 

Sez Anatom, Careggi, Firenze, Italia 5)Clinical And Descriptive Epidemiology 

Unit, Ispo, Firenze, Italia 6)Divisione Di Dermatologia Università 

Di Firenze, Santa Maria Nuova, Firenze, Italia 7)Dipartimento Di 

Area Critica Med Chirurgica Sez An, Careggi, Firenze, Italia 8)Dip Area 

Critica Med Chirurgica Sez Anatomia Patol, Careggi, Firenze, Italia 

Background. Increasing epidemiological evidence underlines 

the role of estrogens in melanoma aetiology. Previous studies 

suggested that estrogens may have a direct inhibitory effect 

on melanoma tumour growth, through the binding of estrogen 

receptor-beta (ERβ). In this study we investigated the relationship 

between body weight (body mass index, BMI), and Breslow 

thickness in patients affected by primary cutaneous melanoma. 

Methods. Patients with primary melanomas observed between 

1998-2009 (n = 605) were included in the analysis (332 females 

and 273 males). The effect of body mass index (BMI), 

≥ 25 vs < 25 kg/m 2 , on the risk of having a thick melanoma diagnosis 

was estimated in terms of Odds Ratio (OR) by means of a 

logistic regression analysis stratified for sex and age-classes with 

adjustment for age within each age-decade. ERβ protein expression 

was evaluated by immunohistochemistry on the melanoma 

tissues from a representative series of 66 patients (21 prenopausal 

women, 23 postmenopausal women and 22 males). 

Results. The incidence of thick melanomas was greater in 

overweight women than in non-overweight ones (OR = 1.64). 

When considering only postmenopausal women, the odds ratio 

increased further (OR = 2.50). Given that thickness is the main 

prognostic factor in melanoma, excess bodyweight may be considered 

a negative modifiable determinant of melanoma prognosis 

in postmenopausal women. Mean ERβ expression in melanoma 

cells was 65% ± 27; 50% ± 32 and 35% ± 17 in the categories 

of prenopausal women, postmenopausal women and males, respectively 

(p < 0.01). ERβ expression was inversely correlated 

with melanoma thickness (p < 0.05), supporting the hypothesis 

that ERβ expression may play a role in melanoma progression. 

Further research will elucidate whether ERβ expression plays an 

independent role in the prognosis and therapy of melanoma. 


KrAS testing on colo-rectal carcinoma cytological 

imprints 

Malapelle U., Bellevicine C., Russo A., Salatiello M., Palombini 


Scienze biomorfologiche e funzionali, Università di Napoli “Federico II”, 


Background. Anti-EGFR monoclonal antibodies, cetuximab, 

and panitumumab, are administrated under the condition that advanced 

colorectal cancer (CRC) carries a wild-type KRAS gene. 

Thus, clinicians request pathologists to genotype KRAS before 

treatment. In the near future routine mutation testing at the same 

time of the surgery may be implemented. The reliability of a rapid 

KRAS testingon ex vivo cytological samples obtained by direct 

scraping of the colon tumour tissue is here evaluated. 

Methods. A consecutive series of 20 surgically resected, primary 

CRC specimens was analysed. 

Fresh tissue from CRC was scraped with a scalpel blade, smeared 

on uncoated glass slides, air-dried and Diff-Quik stained to 

ensure malignant cell presence. The same tissue area was also 

histologically processed. Exon 2 KRAS gene mutations were 

evaluated on both cytological and histological specimens by 

dideoxy sequencing and by the Thera-Screen KRAS Kit (DxS, 

Manchester, England). Data obtained on imprint cytology and 

matched histological 

samples showed full concordance; however, the mutation frequency 

was slightly higher (35%) by the Thera-Screen KRAS Kit 

than by the dideoxy sequencing (30%). 

Conclusion. Thus, colon cancer imprint cytology sample is a biospecimen 

reliable for both dideoxy-sequencing and Thera-Screen 

KRAS analysis and it may be useful to abbreviate the KRAS 

assay turnaround time. 

G13V: a novel K-rAS mutation in colorectal cancer 

1)Malapelle U. 2)Cozzolino I. 3)Iaccarino A. 4)Maria R. 5)Di 

grazia M. 6)Troncone G. 




Università di Napoli “Federico II”, Napoli, Italia 4)Unità operativa 

di oncologia, Civile di caserta, Caserta, Italia 5)Scienze biomorfologiche 


Background. Activating mutations in the K-ras oncogene mainly 

occurr in codons 12 and 13 and may be predictive of response to 

drugs directly linked to the K-ras signaling pathway, such as panitumumab 

and cetuximab. Tumors harboring a KRAS gene mutation 

do not derive benefit from this therapy. Most of these mutations 

are observed in codon 12 (GGT) or 13 (GGC) of exon 2. 

Methods. K-ras analysis was carried out on DNA extracted from 

paraffin-embedded tumor samples after microdissection of a 

transverse colon tumour and its liver metastasis occurring in a 

59 years old femal. Exons 1 and 2 were amplified by PCR and 

then sequenced. Results. A never-reported K-ras mutation with a 

novel tandem double GC ‡ TT mutation, in the first and second 

positions of codon 13 of Kras exon 2, substituting valine (TTC) 

for normal glycine (GGC), occurred. BRAF and p53 were not 

found to be modified and microsatellite instability was not present. 

To confirm this mutation, the PCR product of Kras exon 2 

was subcloned, and 6 subclones were sequenced. Four out of six 

subclones confirmed the occurrence of a double base-pair change 

from Guanine-Cytosine to Thymine-Thymine, whereas the other 

2 were wilde-type for the codons 12 and 13. 

Conclusion. This study is the first report of a novel G13V K-ras 

mutation in a patient with metastatic colorectal cancer.


Application of the KrAS StripAssay TM to the 

analysis of KrAS gene mutations in colorectal 

cancer: assessment of the sensitivity and 

specificity of the method 

Malatesta S., Felicioni L., Viola P., Del grammastro M., Sciarrotta 

M., Pullara C., Buttitta F., Marchetti A. 

Dipartimento Di Oncologia E Medicina Sperimentale, Università “G. 

D’Annunzio”, Chieti, Italia 

Background. Therapeutic agents targeting the epidermal growth 

factor receptor (EGFR) have improved outcome for patients with 

metastatic colorectal carcinoma (mCRC). However, only a subset 

of patients benefits from these treatments. Somatic mutations at 

codons 12 and 13 of the KRAS gene are frequent events in CRC 

and are associated with poor response to anti-EGFR therapy. Direct 

sequencing (DS), the conventional method adopted for mutational 

analysis, is not able to detect low prevalence mutations due to its 

low sensitivity. Therefore, more sensitive methods are needed to 

correctly identify patients resistant to anti-EGFR therapy. 

Methods. We applied the KRAS StripAssay TM technique (codons 

12-13) (Vienna Lab Diagnostics GmbH) to detect KRAS mutations 

on a series of 76 consecutive CRCs. The same series was 

previously investigated by DS and mutant-enriched sequencing 

(ME-sequencing), a very sensitive method that increases the detection 

sensitivity of mutations at codon 12 and 13 of the KRAS gene. 

We also evaluated the sensitivity and the specificity of the KRAS 

StripAssay TM using the ME-sequencing as reference method. 

Results. The KRAS StripAssay TM was able to identify KRAS 

mutations in 37 (49%) of 76 tumor samples. DS and ME-sequencing 

detected KRAS mutations in 36 (47%) and 38 (50%) 

cases, respectively. When compared with ME-sequencing, the 

KRAS StripAssay TM showed a specificity of 100% and a sensitivity 

of 97%, by only missing an uncommon case of double 

mutation at codon 12 on the same allele. In comparison with DS, 

the KRAS StripAssay TM demonstrated higher sensitivity (97% vs 

95%). Our results indicate that the KRAS StripAssay TM is a reliable 

and sensitive method for the detection of KRAS mutations 

in colorectal cancer patients. 

Her2 assessment in gastric cancer: proposal 

of a work-flow for practical routine use 

1)Asioli S. 2)Maletta F. 3)Verdun di Cantogno L. 4)Satolli MA. 

5)Schena M. 6)Pecchioni C. 7)Botta C. 8)D’Angelo G. 9)Recupero 

D. 10)Sapino A. 

1)Scienze biomediche e oncologia umana, Molinette, Torino, Italia, 

2)Scienze biomediche e oncologia umana, Molinette, Torino, Italia 

3)Scienze biomediche e oncologia umana, Molinette, Torino, Italia 

4)Scienze biomediche e oncologia umana, Molinette, Torino, Italia 5)Oncologia 

medica, Molinette, Torino, Italia 6)Scienze biomediche e oncologia 

umana, Molinette, Torino, Italia 7)Scienze biomediche e oncologia 

umana, Molinette, Torino, Italia 8)Scienze biomediche e oncologia umana, 

Molinette, Torino, Italia 9)Scienze biomediche e oncologia umana, 

Molinette, Torino, Italia 10)Scienze biomediche e oncologia umana, Molinette, 

Torino, Italia 

Background. In gastric cancer (GC) the expression of HER2 is 

known as a marker of prognosis and recently it has been confirmed 

as a predictive marker of response to trastuzumab. 

Methods. GC specimens of 42 patients were collected. Representative 

samples from both primary tumors (42 samples) and lymph 

node metastases (23 samples), were selected. In each case, 4B5 

(Ventana), CB11 (kit Oracle Menarini), HercepTest (Dako) antibodies 

were tested in immunohistochemistry (IHC) and scored 

as proposed for GC. HER2 gene status was studied by double 

probe fluorescence in situ hybridization (FISH) in all cases. 

Concordance among IHC scoring results of the 3 antibodies and 

313 

between FISH results and IHC (0/1+ and 2+/3+), independently 

from the percentage of positive cells, were evaluated using the 

Cohen-Fleiss’ kappa statistic (K). Then, the number of specimens 

needed to be tested in cases with < 10% of HER2 overexpression 

was assessed. Finally, influence of gain of CEP17 (copies number 

> 3) on the results of FISH ratio was considered. 

Results. The 3 antibodies showed a K of 0,76 (p < 0,05). In the 

primary tumor, the overall concordance of FISH/IHC, taking 

into account the results of at least one antibody, was of 0,95 

(p < 0,05). FISH/IHC concordance decreased to 0,82 (p < 0,05) 

when correlated with lymph node metastases. Five cases showed 

2+/3+ score values in < 10% of cells. In 4 of these cases the percentage 

increased to > 10% adding 2 more sections from different 

tissue blocks of the primary tumor. In our results, the gain of 

CEP17 did not influence the final score ratio of FISH analysis. In 

conclusions, the HER2 analysis in GC needs a specific protocol 

avoiding working over-load and to solve equivocal cases. 

Identification of genetic determinants underlying 

coronary microvascular remodelling 

1)Mancini M. 2)Varela-carver A. 3)Petretto E. 4)Leopizzi 

M. 5)Parker H T. 6)Kleinert C. 7)Rimoldi O. 8)D’Amati G. 

9)Camici PG. 

1)Medicina Sperimentale, Policlinico Umberto I, Roma, Italia 2)Clinical 

sciences, Hammersmith hospital, Londra, Regno unito 3)Clinical sciences, 

Hammersmith hospital, Londra, Regno unito 4)Medicina Sperimentale, 

Policlinico Umberto I, Roma, Italia 5)Clinical sciences, Hammersmith 

hospital, Londra, Regno unito 6)Clinical sciences, Hammersmith hospital, 

Londra, Regno unito 7)Clinical sciences, Hammersmith hospital, Londra, 

Regno unito 8)Medicina Sperimentale, Policlinico Umberto I, Roma, Italia 

9)Università Vita-Salute, Ospedale San Raffaele, Milano, Italia 

Background. There is current evidence that abnormalities in the 

function and structure of the coronary microcirculation occur in 

many clinical conditions, this has led to the concept of “coronary 

microvascular dysfunction” (CMD). We studied the histologic 

and histomorphometric phenotypes of coronary arterioles in 30 

Recombinant Inbred (RI) strains derived from spontaneously 

hypertensive (SHR) and the normotensive Brown Norway (BN) 

and mapped them to the genome. 

Methods. Histological and histomorphometric studies were carried 

out on HE and Picrosirius red sections of 3-6 hearts from 

each RI strain using Metamorph 6.2 software. Vessel and lumen 

diameters and the medial area of arterioles were measured. Genome-wide 

correlation analysis was carried out on expression 

QTLs (eQTLs) identified in the heart with medial area and other 

phenotypes. Transcripts that were both under cis-acting genetic 

regulation and significantly correlated with medial area, but not 

with blood pressure indices, were prioritized. 

Results. We identified a set of 60 transcripts under genetic control 

and significantly associated with medial area. Among these 

we prioritized 2 genes because of their biological function: Rcn2 

(Reticulocalbin 2) that encodes for a calcium-binding protein and 

Hsp40 (Heat shock protein 40). Both proteins expression (4.9 fold 

decrease for Rcn2 and 4.3 fold decrease for Hsp40, p = 0.0001, 

n = 4-6) and mRNA level (0.80 fold decrease for Rcn2 and 0.74 

fold decrease for Hsp40, p = 0.0001, n = 4-6) were reduced in 

SHR compared to BN rats. Finally, further experiments in congenic 

animals demonstrated that both Rcn2 and Hsp40 mRNA 

levels returned to BN expression levels as compared to SHR 

(p = 0.001 and p = 0.05 respectively, n = 4). 

The identification of these genetic determinants suggests that 

coronary microvascular remodelling is not exclusively determined 

by environmental factors such as blood pressure, and 

advocates more complex genetic regulation and mechanisms for 

these traits.

314 

use of IGK gene rearrangement analysis for 

clonality assessment of lymphoid malignancies. 

A single center experience 

Mannu C., Sagramoso C., Gazzola A., Rossi M., Sapienza M.R., 

Laginestra A., Bacci F., Sabattini E., Agostinelli C., Artioli P., 

Chilli L., Da Pozzo G., Piccioli M., Righi S., Sandri F., Pileri 

S.A. * , Piccaluga P.P. * 

Department of Hematology and Oncology “L. and A. Seràgnoli”, Hematopathology 

Unit, S. Orsola-Malpighi Hospital, University of Bologna, 

Italy; * SAP and PPP equally contributed to this work 

Background. The diagnosis of B-non Hodgkin lymphomas 

(NHLs) is based on clinical, morphological and immunohistochemical 

features. However, in up to 10-15% of cases, analysis 

of immunoglobulin heavy (IGH@) or light (IGH@/IGL@) chains 

genes rearrangements is required to discriminate between malignant 

and reactive lymphoproliferations. Clonality assessments is 

basically performed by IGH@ analysis; however, IGK@ study is 

sometime required. 

In this study, we evaluated the role of IGK@ analysis in the routine 

diagnostic of lymphoproliferative processes. 

Methods. Clonality patterns were studied in 63 B-cell lymphoproliferative 

disorders by using the BIOMED-2 protocols for 

IGH@ and IGK@ assays. PCR products was evaluated by both 

hetroduplex and GeneScan analysis. 

Results. IGK@ analysis was technically successful in all 63 cases. 

Overall, it supported the histopathological suspicion in 48/63 

cases, while in 15/63 it guided a different diagnosis (N = 7), suggested 

a specific clinical monitoring (N = 2), or was considered 

as false negative (N = 6). Specifically, in 2/11 suspected RCs, 

IGK@ study converted the final diagnosis highlighting the existence 

of a lymphomatous clone. In one case, conversely, though 

the IGK@ pattern was definitely clonal, the final diagnosis was 

RC, a close follow-up being suggested. IGK@ genes were clonally 

rearranged in 41/52 cases of sLy. In 11/52, in contrast, molecular 

analyses documented a polyclonal pattern, guiding a final 

diagnosis of RC in 5/11 cases. In the other 6, morphological and 

immunophenotypical evidences definitely supported the initial 

suspect and results were interpreted as false negative. Interestingly, 

MZL and FL appeared to be the entities most frequently 

requiring IGK@ analysis. 

Conclusions. IGK@ study appeared to be extremely useful in 

supporting challenging diagnosis and even discriminating a proportion 

of unsolved cases (~15%). Thus, basing on our series, 

when NHL is suspected, negative results at IGH analysis should 

not be considered as conclusive and further investigation of IGK 

is appropriate. 

Association between single nucleotide 

polymorphisms in the COX-2, TNf-α 

and VeGf-A genes and susceptibility to 

hepatocellular carcinoma(hcc) and pancreatic 

adenocarcinoma(PA) 

1)Marasà L. 2)Montalto G. 3)Marasà S. 4)Balasus D. 5)Giacalone 

A. 

1)Pathology department, Arnas civico hospital, Palermo, Italy 2)University 

of palermo, Chair of internal medicine, Palermo, Italy 3)Department of 

pathology, Arnas civico hospital, Palermo, Italy 4)Department of pathology, 

Arnas civico hospital, Palermo, Italy 5)University of palermo, Chair of 

internal medicine, Palermo, Italy 

Background. Several studies have demonstrated the association 

between single nucleotide polymorphisms(SNPs) and 

susceptibility to the development of various diseases including 

cancer. These allelic variations fall in the promoters or coding 

regions of genes influencing their transcriptional activity 

and post-transcriptional.The HCC and the PA share a stage of 

chronic inflammatory disease as a risk factor, in which are in- 


volved both environmental and genetic factors. TNF-α, COX-2 

and VEGF are mediators of inflammation and angiogenesis. 

High levels of these mediators were found in serum and tissues 

of patients with HCC and PA.Evaluate SNPs in these genes 

could be a potential biomarker of susceptibility to cancer development.These 

molecules are potential therapeutic targets for 

HCC and PA. 

Methods. DNA extracted from whole blood belonging to controls, 

HCC and PA subjects all of Sicilian origin, was used for the 

evaluation of SNPs with Restriction Fragment Length Polymorphisms 

method in -1195 G/A position of COX2 gene promoter 

(A allele is associated with higher levels of the cytokine); -308 

G/A position of TNF-alpha gene promoter (G allele is associated 

with lower levels of the cytokine); 936 G/T position of the coding 

region of VEGF-A gene(T allele is associated with high levels 

of VEGF-A).The expression levels of COX2 and VEGF were 

verified by IHC. 

Results. IHC analysis showed an overexpression of COX2 and 

VEGF both on HCC and PA. Analysis of SNPs reported that the 

936 T is more frequent in patients with HCC than in controls 

(p = 0.0215),confirming the data in IHC,whereas for other SNPs 

a statistical difference hasn’t yet been found.However, in our 

previous work we have tested throught Laser capture microdissection 

and Methylation Specific PCR in the PA that the methylation 

status of CpG islands that were hypomethylation, which 

confirms the data in IHC.This suggests the study of genetic and 

epigenetic mutations to identify new biomarkers of susceptibility 

to cancer. 

Triple metachronous multiple tumours: 

a case report 

1)Margiotta G. 2)Calvisi G. 3)Ciuffetelli V. 4)Damiani D. 5)Vitale 

AR. 

1)Pathology Unit, “San Salvatore” Hospital, L’Aquila, Italy 2)Pathology 

Unit, “San Salvatore” Hospital, L’Aquila, Italy 3)Pathology Unit, “San 

Salvatore” Hospital, L’Aquila, Italy 4)Pathology Unit, “San Salvatore” 

Hospital, L’Aquila, Italy 5)Pathology Unit, “San Salvatore” Hospital, 

L’Aquila, Italy 

We report a case of triple metachronous multiple tumour in a 66 

years old woman admitted to the hospital of L’Aquila, Italy. The 

first tumour was a transitional menigioma that she developed 

at the age of 58 years. After two years, the patient developed a 

gastric carcinoma and after six years she developed a follicular 

adenoma of the thyroid. The treatment of each tumour was chirurgical. 

To date, the patient is in good conditions of health and 

without recurrence. Also if this entity is not very rare, and also if 

two lesions are benign tumour (and for this reason we report the 

case as “metachronous multiple tumour” instead of “metachronous 

multiple malignancies”) we report this case for underlye that 

the possibility that metachronous multiple tumours exist must 

always be considered during evaluation of a patient. Patient must 

be informed that there is the risk of developing secondary tumour 

after the first treatment and that is imperative to reporting any 

new symptom which might occur. 

references 

Irimie A, Achimas-Cadariu P, Burz C, et al. Multiple primary malignancies 

- epidemiological analysis at a single tertiary institution. J Gastrointestin 

Liver Dis 2010;19(1):69-73. 

Macrì A, Saladino E, Basile A, et al. Quintuple primitive malignant neoplasms. 

A case report. Acta Chir Belg 2010;110(1):95-7.


uterine tumor resembling ovarian sex cord tumor. 

A case report 

1)Margiotta G. 2)Crisman G. 3)Coletti G. 4)Carta G. 5)Leocata 

P. 

1)Dept of experimental medicine, University of L’Aquila, L’Aquila, Italy 

2)Dept of experimental medicine, University of L’Aquila, L’Aquila, Italy 

3)Pathology unit, “san salvatore” hospital, L’Aquila, Italy 4)Gynecological 

unit, University of L’Aquila, L’Aquila, Italy 5)Dept of health’s sciences, 

University of L’Aquila, L’Aquila, Italy 

Background. Uterine tumors resembling ovarian sex cord tumors 

(UTROSCT) are rare neoplasms (only 77 cases of UTROSCT 

have been reported in literature to date), described for the first time 

in 1976 by Clement and Scully 1 . Morphologic and immunohistochemical 

findings indicate that UTROSCT arise from pluripotential 

uterine mesenchymal cells, which mainly differentiate into sex 

cord cells. We report a case of a 74 years old woman, who was 

admitted to our hospital for vaginal bleeding and uterine enlargement. 

According to all clinical data a standard total abdominal hysterectomy 

and bilateral salpingo-oophorectomy was performed. 

Methods. On histologic examination the tumor was composed of 

sweeping fascicles of smooth muscle cells surrounding a diffuse 

proliferation of tubular and gland-like structures lined by plump 

cells with indistinct cytoplasm. Immunohistochemistry was 

imperative for a correct diagnosis. Sections previously formalinfixed 

and paraffin-embedded were stained with calretinin, CD10, 

CD99, inhibin, actin, CAM 5.2, estrogen receptors. The tumor 

cells were strongly positive for CD99, calretinin, CAM 5.2, inhibin, 

estrogen receptors. A focal positive reaction with actin was 

observed. Stain for CD10 was negative. Immunohistochemical 

stains of the sex cord elements may show positivity for vimentin, 

cytokeratin, actin and desmin in variable proportions. Inhibin is a 

more specific marker for these cells 2 . 

Results. Finally, a diagnosis of uterine tumor resembling ovarian 

sex cord tumors (UTROSCT) was posed. After 8 months from 

the hysterectomy, the patient is in good condition of health and 

without recurrence. 

references 

1 Clement PB, Scully RE. Uterine tumors resembling ovarian sex-cord 

tumors. A clinicopathologic analysis of fourteen cases. Am J Clin 

Pathol 1976;66(3):512-25. 

2 Czernobilsky B. Uterine tumours resembling ovarian sex tumours: an 

update. Int J Gynecol Pathol 2008;27(2):229-35. 

Malignant fibrous histiocytoma of the corpora 

cavernosa: a case report 

1)G.Crisman 1)Margiotta G. 2)Discepoli S. 3)Leocata P. 

1)Dept of Experimental Medicine, “San Salvatore” Hospital, University 

of L’Aquila, L’Aquila, Italy 2)Pathology Unit, P.O. “Ss. Filippo e Nicola”, 

Avezzano (L’Aquila), Italy 3)Dept of Health’s Sciences, University of 

L’Aquila, L’Aquila, Italy 

Background. Malignant fibrous histiocytoma (MFH) has been 

regarded as the most common soft tissue sarcoma of the adulthood, 

with a male sex predilection. MFH could arise everywhere 

throughout the body, but it is rarely observed in the genitourinary 

tract, usually involving the bladder or the kidney. Penis represents 

an extremely rare site of involvement: up to now, only six cases 

have been reported so far. 

Methods. The authors report on a case of a 61-year-old Caucasian 

man, presented with a slowly enlarging, painless mass sited 

on the upper part of corpora cavernosa. Clinical examination 

of penis skin, scrotum, testicles, spermatic cord and inguinal 

lymph nodes was unremarkable. An biopsy of about 1,5 cm in 

diameter was performed, and a gray-white cut surface mass with 

focal hemorrhage was observed. All specimens were routinely 

processed, and histopathological features revealed a spindle cells 

lesion, mostly organized in a storiform pattern, within abundant 

315 

myxoid areas. Several atypical mitoses and focal areas of necrosis 

were observed as well. In addition, Vimentin, CD68, S100, 

Desmin, Actin, Ki67, α-1 Antitrypsin stains were performed, 

showing a strong positivity for Vimentin and α-1 Antitrypsin, 

with scattered CD68 positive cells. 

Finally, a diagnosis of malignant fibrous histiocytoma, myxoid 

type, arisen from the fibrous septa of the corpora cavernosa, was 

made, thus leading the patient to a penectomy. 

Results. Because of the rarity of this particular site of involvement, 

a diagnosis of MFH of penis represents a great challenge 

for both clinicians and pathologists. The authors briefly discuss 

and deepen differential diagnoses of this kind of lesions. 

loss of heterozigosity (lOH) as molecular marker 

of progression in oral squamous carcinogenesis 

A. Marsico * , M. Micheletti ** , I. Rostan ** , M. Pentenero *** , 

S. Gandolfo *** , R. Navone ** 

* ** UO di Anatomia Patologica, Ospedale Koelliker, Torino; Dipartimento 

di Scienze Biomediche e Oncologia Umana dell’Università di Torino (Sez. 

di Anatomia Patologica), *** Dipartimento di Scienze Cliniche e Biologiche 

dell’Università di Torino (Sez. di Medicina e Oncologia Orale) 

Background. Oral squamous carcinoma (OSCC) is characterised 

by genetic alterations of the epithelial cells. The loss of 

heterozygosity (LOH) is an event considered fundamental for 

carcinogenesis i.e. the disappearance of a more or less large 

area of DNA in one or two members of a couple of homologue 

chromosomes. This is the mechanism whereby the genetic loci 

containing oncosuppressor genes involved in tumoral progression, 

are lost. The LOH has evidenced the involvement of oncosuppressor 

genes situated in determined chromosomal regions 

e.g. 3p14.2, 3p24, 3p21.3, 9p21, 17p13, 4q, 8p, 11q and 13q. The 

loss of specific chromosomal regions that contain oncosuppressor 

genes represents an early predictor of potentially malignant oral 

lesion (PML) progression (as high as 36% also in the absence 

of dysplasia). The presence of LOH has been observed at 3p 

and/or 9p in 50% of oral leukoplakias, with a 3.8 fold increase in 

the risk of malignant transformation. Further LOH (4q, 8p, 11q, 

13q and/or 17p) lead to a 33-fold increase in the risk of tumoral 

progression. 

Methods. We selected polymorphic microsatellite markers, situated 

in chromosomal loci with a higher evidence of LOH and a 

significant heterozygosity in oral PMLs and OSCC. We focused 

our attention on chromosome 3 (D3S1234 e D3S1300, locus 

3p14.2, gene FHIT i.e. the fragile histidine triad gene, D3S1317, 

locus 3p26, gene VHL i.e. von-Hippel Lindau) and chromosome 

9 (IFNA, locus 9p22, gene IFNA, D9S171 and D9S1751, locus 

9p21), that had the highest number of LOH and on cases involving 

progression. 

We describe a case of a 62-year-old female, diagnosed with verrucous 

carcinoma on the border of the tongue. 

Results. The analysis of the 6 markers was carried out on DNA 

samples extracted from the neoplasia using a DNA sample of 

normal mucosa of the same subject as control. There was a loss 

of heterozygosity on the short arm of chromosome 3, evidenced 

by the analysis of the D3S1300 and D3S1234 markers. Whilst 

there was a normal allelic profile in the 3 heterozygote points 

of the markers IFNA, D9S171, D9S1751 on the short arm of 

chromosome 9. 

The incorporation of the molecular data as to the loss of heterozygosity 

at histopathologic diagnosis of PMLs or OSCC may 

predict the evolution of these lesions, thus distinguishing cases 

with a high probability of progression or worsening from those 

with a lower risk.

316 

evaluation of DNA ploidy in carcinoma and 

potentially malignant lesions of the oral cavity 

on samples obtained with a dermatolgical curette 

1)Marsico A. 2)Rostan I. 3)Pentenero M. 4)Gandolfo S. 

5)Navone R. 

1)Anatomia Patologica, Koelliker, Torino, Italia 2)Scienze Biomediche E 

Oncologia Umana Uni. To, Molinette, Torino, Italia 3)Scienze Cliniche E 

Biologiche Uni To, S. Luigi Gonzaga, Orbassano, Italia 4)Scienze Cliniche 

E Biologiche Uni To, S. Luigi Gonzaga, Orbassano, Italia 5)Scienz 

Biomediche E Oncologia Umana Uni To, Molinette, Torino, Italia 

Introduction. The late stage diagnosis of oral squamous carcinoma 

leads to a low survival rate. This may be due to the difficulty 

in diagnosis and to the fact that there may be lesion development 

without morphological evidence of dysplasia. As it is known, 

also on the basis of our group research 1 that the DNA content 

may be a good marker of neoplastic and preneoplastic lesions, we 

used two different techniques to compare the DNA ploidy with 

microhistology 2 . 

Methods. 211 samples were obtained by a dermatological curette 

(AcuDispo Curette, Acuderm inc) for lesions suspicious 

for carcinoma or potential malignant lesions (PMLs) of the oral 

cavity. The samples were suspended in saline solution for flow 

cytometry (FCM), using a cytofluorimeter FACSCalibur (Becton 

Dickinson) and a Cycletest kit, Plus DNA Reagent. The second 

sample of the same site was fixed in ThinPrep® for 40 of these 

cases. Static cytometry was then performed (ICM), by Perceptronix, 

Vancouver BC, Canada (Dr. A. Doudkine). 

Results. Aneuploidy was observed in 54.8% of the carcinoma 

with FCM and in 71.4% of those with ICM. 50% of the PMLs 

with dysplasia and FCM and in 72.7% of those with ICM; in 

15.1% of the PMLs without dysplasia with FCM and in 12.5% 

with ICM. 

.Compared to FCM, ICM had a higher percentage of aneuploidy 

in invasive carcinoma and dysplasia. Whilst, the ploidy results 

for the PMLs without dysplasia were superimposable on both 

techniques. 

In conclusion, the investigation of DNA with FCM or ICM on 

cells sampled from oral mucosa may offer useful information as 

to the preneoplastic or neoplastic processes. This is particularly 

true in the presence of aneuploidy in lesions without histo-cytological 

evidence of dysplasia, where it may have an important 

prognostic role. The sampling technique set up by our group is efficacious 

in providing cytological (for ctyo-diagnosis and ploidy) 

and histological material (for microhistology). 

references 

1 Donadini et al. Oral cancer genesis and progression: DNA near-diploid 

aneuplodization and endoreduplication by high resolution flow 

cytometry. Cell Oncol 2010 (in press). 

2 Navone R, et al. Oral Potentially Malignant Lesions: First Level Microhistological 

Diagnosis from Tissue Fragments Sampled in Liquid- 

Based Diagnostic Cytology. J Oral Pathol Med 2008;37:358-63. 

epigenetic silencing of SOCS3 identifies a subset of 

prostate cancer with an aggressive behavior 

1)F. Pierconti 1)Martini M. 2)Larocca LM. 3)Pinto F. 

1)Istopatologia, Ucsc, Roma, Italia 2)Istopatologia, Ucsc, Roma, Italia 

3)Urologia, Ucsc, Roma, Italia 

Background. Chronic inflammation and subsequent tissutal 

alterations may play a key role in prostate carcinogenesis. In this 

way, molecular alterations of the suppressor of cytokine signaling 

(SOCS)3, one of the most important inhibitory molecule of 

inflammatory signal transduction circuitries, could contribute to 

explain the pleiotropic role of interleukin (IL)-6 in this type of 

cancer. 

Methods. We analyzed the methylation status and mRNA expression 

of SOCS3 in 20 benign prostate hyperplasias (BPH) 


and in 51 prostate cancer specimens. We analyzed the SOCS3 

methylation status using Methylation-Specific PCR. Hypermethylation 

was confirmed by sequencing after subcloning. 

Epigenetic silencing of this gene was also demonstrated by 

real-time PCR. Results and correlation with clinical data were 

statistically analyzed. 

Results. We found that the promoter of SOCS3 was methylated 

in 39.2% of prostate cancer. On the contrary, all BPH and 

normal controls had an unmethylated pattern. Real-time analysis 

showed that in methylated cases SOCS3 mRNA expression was 

reduced by 3 and 4 folds as compared to BPH and unmethylated 

cases, respectively. Interestingly, SOCS3 mRNA level was 

higher in unmethylated prostate cancer than in BPH. Moreover, 

methylation of SOCS3 promoter significantly associated with 

intermediate-high grade Gleason score (p = 0.0007) and with an 

unfavorable clinical outcome (p = 0.0019). Our data suggest that 

SOCS3 hypermethylation may be involved in the pathogenesis of 

prostate cancer and could identify a tumor subset with an aggressive 

behavior. 

Nogo-A: a useful marker in diagnosis of 

oligodendroglioma, with a possible complementary 

role in identifying 1p19q codeletion 

G. Marucci, R. Panzacchi, E. Di Oto, A. Farnedi, C. Ligorio, 

M.P. Foschini 

Sezione di Anatomia Patologia “M. Malpighi”, Dipartimento di ematologia 

e scienze oncologiche “L .e A. Seragnoli”, Ospedale Bellaria, Università 

di Bologna, Italia 

Background. Differential diagnosis between oligodendrogliomas 

and other gliomas remains a critical issue. Aim of the study 

is to verify the diagnostic value of Olig-2 and Nogo-A and their 

relation with 1p19q codeletion detected by FISH analysis. 

Materials and methods. 158 cases of Central Nervous System 

(CNS) tumors were immunostained with Olig-2 and Nogo-A 

(24 oligodendrogliomas (O), 23 anaplastic oligodendrogliomas 

(AO), 2 oligoastrocytomas (OA), 2 anaplastic oligoastrocytomas 

(AOA), 30 glioblastoma multiforme (GBM), 2 diffuse astrocytomas 

(A), 4 anaplastic astrocytomas (AA), 10 pilocytic astrocytomas 

(PA), 9 ependymomas, 12 anaplastic ependymomas (AE), 

10 neurocytomas, 10 meningiomas, 10 choroid plexus papillomas 

and 10 metastases). Thus FISH analysis was performed in areas 

showing Nogo-A immunopositivity. 

Results. Olig-2 strong positivity: O 91,6%, AO 86,9%, OA 

100%, AOA 100%, GBM 83,3%, AA 50%, A 50%, PA 80% 

and AE 8,3%. Nogo-A strong positivity: O 75%, AO 78,2%, OA 

100%, AOA 50%, GBM 20%, AA 25%, AE 8,3% and10% of 

neurocytomas. Nogo-A driven FISH analysis evidenced 1p19q 

codeletion in 4 further cases of O, in 2 further cases of AO, in 1 

case of OA and in 1 further case of GBM. 

Conclusion. Nogo-A is more useful and specific than Olig-2. 

Furthermore, using a Nogo-A driven FISH analysis, it is possible 

to identify a larger number of 1p19q codeletion in O, mixed gliomas 

and in GBMs with oligo component. 

Significato dei disordini del giro dentato 

nella sclerosi temporale mesiale 

1)Marucci (G). 2)Rubboli (G). 3)Giulioni (M). 

1)Sezione di Anatomia Patologica “M. Malpighi”, Dipartimento di ematologia 

e scienze oncologiche “L .e A. Seragnoli”, Ospedale Bellaria, 

Bologna, Italia 2)Divisione di neurologia, Ospedale Bellaria, Bologna, 

Italia 3)Divisione di neurochirurgia, Ospedale Bellaria, Bologna, Italia 

Razionale. L’epilessia temporale cronica farmaco-resistente è 

la forma più comune di epilessia sottoposta a terapia chirurgica. 

Quadri istopatologici frequentemente osservati in questi pazienti 

sono rappresentati dalla displasia corticale e dalla sclerosi temporale 

mesiale (STM). La STM è caratterizzata dalla perdita di


neuroni nei vari settori del Corno di Ammone. L’obiettivo del 

presente lavoro è quello di verificare se anche le alterazioni del 

giro dentato giocano un ruolo nel quadro della STM. 

Metodi. Sono stati studiati 14 pazienti con diagnosi preoperatoria 

di epilessia temporale cronica farmaco-resistente, sottoposti a terapia 

chirurgica presso l’ospedale Bellaria di Bologna. Sono stati 

arruolati esclusivamente pazienti con STM, senza altra patologia 

concomitante. E’ stata valutata la presenza di dispersione delle 

cellule granulari (GCD) secondo la classificazione proposta da 

Blümcke et al. nel 2009. Il follow-up andava da 12 a 84 mesi (48 

mesi in media) e la prognosi epilettologica è stata valutata adottando 

la classificazione di Engel. 

Risultati. GCD era presente in 7 casi (50%). E’ stata osservata 

una correlazione statisticamente significativa tra GCD e numero 

medio di crisi epilettiche per mese. La percentuale di pazienti 

con follow-up epilettologico post-chirurgico non ottimale (ovvero 

in classe di Engel diversa dalla 1A) è stata il 57,14% nei 

pazienti senza GCD, mentre scendeva al 14,29% nei pazienti 

con GCD. I due pazienti con follow-up peggiore non mostravano 

GCD. 

Conclusioni. I risultati del presente studio suggeriscono che le 

alterazioni del giro dentato giocano un ruolo nell’epilessia temporale 

farmaco-resistente. In particolare nei pazienti con MTS 

si è evidenziata una associazione tra la presenza di disordini del 

giro dentato e una prognosi epilettologica post-chirurgica più 

favorevole. 

Necrotizing peripartum myocarditis 

1)Marzullo A. 2)Solarino B. 3)Maselli E. 4)Serio G. 

1)Anatomia patologica, Università di bari, Bari, Italia 2)Medicina legale, 

Università di bari, Bari, Italia 3)Medicina legale, Università di bari, Bari, 

Italia 4)Anatomia patologica, Università di bari, Bari, Italia 

Background. Peripartum heart disease is a group of conditions 

occurring during the last trimester of pregnancy through the first 

6 months post partum with unknown etiology and pathogenesis. 

The myocardial involvement includes myocarditis, coronary artery 

dissection and peripartum cardiomyopathy. 

Methods. We report the case of a pregnant, aged 27, that after 

the delivery at the 37 th week of gestational age, complained weakness 

and abdominal pain. Laboratory data revealed hypocromic 

anaemia, elevated VES, non specific repolarization anomalies 

on ECG and normal cardiac profile at echocardiography. For the 

worseness of general conditions the patient died 24 days later and 

the last echocardiography showed a severe hypocontractility of 

the left ventricle. 

Results. At autopsy, the heart had normal shape and volume, 

lowered consistency; on cut surface yellowish areas were intermingled 

with pale red ones. The histology showed a diffuse 

inflammatory infiltration made predominantly by eosinophils and 

severe myocite necrosis; on the base of the histological findings 

a diagnosis of peripartum eosinophilic myocarditis was made. 

Systemic and pulmonary diseases were excluded. In this report 

are discussed the possible etiopathogenesis and the differential 

diagnosis. 

BAG3 protein delocalization in prostate carcinoma 

1)Mascolo M. 2)Vecchione ML. 3)Ilardi G. 4)Siano M. 5)Nugnes 

L. 6)De rosa G. 7)Staibano S. 

1)Department of Biomorphological and Functional Sciences, Pathology 

Section, University “Federico II”, Naples, Italy 2)Department of 

Biomorphological and Functional Sciences, Pathology Section, University 

“Federico II”, Naples, Italy 3)Department of Biomorphological 

and Functional Sciences, Pathology Section, University “Federico 

II”, Naples, Italy 4)Department of Biomorphological and Functional 

Sciences, Pathology Section, University “Federico II”, Naples, Italy 

5)Department of Biomorphological and Functional Sciences, Pathology 

Section, University “Federico II”, Naples, Italy 6)Department of Bio- 

317 

morphological and Functional Sciences, Pathology Section, University 

“Federico II”, Naples, Italy; Oncology Research Center of Basilicata 

(C.R.O.B.), Rionero in Vulture, Potenza, Italy 7)Department of Biomorphological 

and Functional Sciences, Pathology Section, University “Federico 

II”, Naples, Italy 

Background. Prostatic cancer (PC) currently ranks as the second 

cause of death for malignancy among men of Western countries. 

Despite the progressive increase of early diagnosis, mainly due to 

the widespread diffusion of the prostate-specific-antigen (PSA) 

screening and to the improvement of ultrasound diagnostic techniques, 

a subset of PC shows a metastasizing and lethal course, 

not predictable upon the traditional prognostic parameters. The 

data of the recent literature indicate that, as it has been found for 

the large majority of solid human malignancies, BAG3 protein, 

a member of the BAG family of heat shock protein (HSP) 70 

cochaperones,is involved in the regulation of proliferation and 

apoptosis in normal as in neoplastic cells. 

Methods. Formalin-fixed, paraffin-embedded surgical specimens 

of 55 cases of PC and 15 cases of benign prostatic hyperplasia 

(BPH) and normal prostate tissue obtained from areas surrounding 

BPH were retrieved from the files of the Department of 

Biomorphological and Functional Sciences, Section of Pathology, 

University Federico II of Naples. BAG3 expression was 

evaluated by immunohistochemistry. Results were compared 

with clinicopathological features of tumours and the outcome of 

patients. 

Results. We found BAG3 expressed in all the prostatic tissues 

of the study: non-neoplastic prostate tissue showed a cytoplasmatic 

staining with apical reinforcement, a finding which appears 

consistent with the reported connection of the protein with 

the membrane focal cell-adhesion complexes. In PC, BAG3 was 

generally overexpressed, but showed a linear decrease from 

low-grade (Gleason score < 7) to high grade tumors (Gleason 

score > 7), coupled with the loss of polarization of the signal 

in metastasizing cases. These results indicate that BAG3 intracytoplasmic 

delocalization is a specific feature of cancer vs 

non-neoplastic prostate; moreover, the protein looks as promising 

new marker for prediction of prostate cancer invasiveness 

and behavior. 

Possible implication of local immune response 

in Darier’s disease. An immunohistochemical 

characterization of lesional inflammatory infiltrate 

1)Mastrogiulio M.G. 2)Onorati M. 3)Ambrosio M.R. 4)Mourmouras 

V. 5)Rocca B.J. 6)Pieronudo F. 7)Miracco C. 8)Sacchini 

A. 9)Bartolomei S. 10)Luzi P. 

1)Deprtment of Human Pathology and Oncology-Section of Anatomic 

Pathology, University of Siena, Italy2)Department of Human Pathology 

and Oncology-Section of Anatomic Pathology, University of Siena, 

Italy3)Department of Human Pathology and Oncology-Section of Anatomic 

Pathology, University of Siena, Italy 4)Department of Human 

Pathology and Oncology-Section of Anatomic Pathology, University of 

Siena, Italy5)Department of Human Pathology and Oncology-Section 

of Anatomic Pathology, University of Siena, Italy 6)Department of Human 

Pathology and Oncology-Section of Anatomic Pathology, University 

of Siena, Italy 7)Department of Human Pathology and Oncology- 

Section of Anatomic Pathology, University of Siena, Italy 8)Department 

of Human Pathology and Oncology-Section of Anatomic Pathology, 

University of Siena, Italy 9) Department of Human Pathology and Oncology-Section 

of Anatomic Pathology, University of Siena, Italy 10) 

Department of Human Pathology and Oncology-Section of Anatomic 

Pathology 

Background. Darier’s disease (DD) is an infrequent autosomal 

dominantly inherited skin disorder caused by mutations in the 

ATP2A2 gene, which encodes a calcium pump highly expressed 

in epidermal keratinocytes. This defect leads to acantholysis. Skin 

infections that complicate the disease, are thought to depend on 

the compromised skin integrity, whereas cell-mediated immunity

318 

is considered to be normal. To date, there are no investigations 

on the local inflammatory infiltrate in DD skin lesions. In this 

immunohistochemical study we characterized and quantified it, 

making comparisons with two other inflammatory skin disorders, 

i.e. pemphigus vulgaris (PV), and lichen ruber planus (LRP), and 

with the normal skin (NSk). 

Methods. We analyzed 16 skin biopsies of patients with DD, and 

for comparison, 14 patients with PV, and 10 with LRP. As negative 

control, we examined NSk excised from the abdomen of 12 

healthy subjects who underwent aesthetic surgery. In all cases, 

leukocyte subsets were characterized using a panel of antibodies 

for CD3, CD4, CD8, CD20, granzyme B, CD 25, FOXP3, CD1a, 

CD 68 and CD123. 

Results. We found a significant (p < 0.05) decrease of CD1a+ 

Langerhans cells (LCs) in DD, compared to PV, LRP, and NSk, 

and of CD123+ plasmacytoid dendritic cells (pDCs), compared 

to PV and LRP. 

Conclusions. We hypothesize that the genetic damage of keratinocytes 

typical of DD, might result in a loss of some subsets of 

dendritic cells, and, consequently, in an impaired local immune 

response, which might worsen the infections that inevitably occur 

in this disease. 

low foxp3 expression in negative sentinel lymph 

nodes is associated with node metastases in 

colorectal cancer 

1)Cassenti A. 2)Matera L. 3)Sandrucci S. 4)Castellano I. 5)Cassoni 

P. 

1)Scienze biomediche e oncologia umana, Molinette, Torino, Italia 2)Medicina 

interna, Molinette, Torino, Italia 3)Chirurgia oncologica, Molinette, 

Torino, Italia 4)Scienze biomediche e oncologia umana, Molinette, Torino, 

Italia 5)Scienze biomediche e oncologia umana, Molinette, Torino, 

Italia 

Background. Tregs (Foxp3+) play a pivotal role in maintaining 

immune system homeostasis through their ability to suppress immunological 

responses, including tumor immunity against tumorassociated 

antigens. In vivo immunosuppressive effect of these 

cells in colorectal cancer still remains controversial; actually, 

we hypothesized that Tregs (Foxp3+) do not contribute to CRC 

escape from host immunity. In fact, a strong association between 

Foxp3 expression and tumor regression has been reported in 

CRC recently, suggesting that Tregs are not playing an immunosuppressive 

role but may contribute to homeostatic control of a 

robust immune response. 

Methods. We here investigated if Foxp3 expression in sentinel 

lymphnode (SLNs) in CRC may correlate with node metastases 

and patient outcome. The expression of Foxp3 was determined by 

immunohistochemistry in histology negative SNLs of 30 patients 

with CRC (18 pT2 and 12 pT3). The expression was then correlated 

with nodal status and patient outcome. 

Results. All pT2, and 3/12 pT3 patients, had > 10% Foxp3+ 

cells in SLNs. 6/12 pT3 patients had lymphnode metastases and 

all showed < 10% Foxp3+ lymphocytes in their negative SLNs. 

Thus, Foxp3 expression in > 10% of the microenvironment cells 

of SNs seems to correlate with lack of migration of tumor cells 

to the downstream lymphnodes (non-SLNs). Positive correlation 

between Foxp3 staining and tumor protection was further 

confirmed by the significantly longer survival of patients with 

high vs low Foxp3 expression in SLNs. In fact, 20/21 (95%) 

patients with high Foxp3, but 1/9 (11%) with low Foxp3 were 

still alive after 7 years. SLN Foxp3 positivity may represent both 

an immunological marker associated to pN0 status and a positive 

prognostic factor for CRC; its use here confirms the emerging 

view that Treg expression is correlated with increased tumor 

protection and survival and is indicative of a successful immune 

response taking place. 


A mirNA expression profiling identifies mir-145 as 

a key player in malignant pleural mesothelioma 

Eliseo Mattioli 1 , Domenico Di Marzo 2 , Iris Forte 1 2 , Baharak 

Khadang 1 , Donatella Spina 1 , Marisa De Feo 3 , Paola Indovina 1 , 

Antonio Giordano 1 2 4 2 4 

and Francesca Pentimalli 

1 Dipartimento di Patologia Umana ed Oncologia, Università di Siena, 

Siena, Italy; 2 Centro Ricerche Oncologiche di Mercogliano “Fiorentino 

Lo Vuolo” (C.R.O.M.), Mercogliano (AV), Italy; 3 Department of Cardiothoracic 

and Respiratory Sciences, Second University of Naples and 

Department of Cardiovascular Surgery, V. Monaldi Hospital, Naples, 

Italy; 4 Sbarro Institute for Cancer Research and Molecular Medicine, 

Temple University, Philadelphia, PA (USA) 

Background. Aim of this research is to analyze the miRNA 

expression profile of pleural malignant mesothelioma (MM) in 

tissues and cell lines. 

Methods. We collected 14 MM cases from the archives of the 

Department of Human Pathology of the University of Siena and 

6 normal pleural samples from patients undergoing coronaric 

surgery (screening set). Paraffin sections were dissected to enrich 

the tissue components of interest in the samples; total RNA 

was extracted and hybridized on an Exiqon miRCURY LNA TM 

platform. 

In order to validate the results from the microarray screening, 22 

additional MM cases and 22 normal controls were subsequently 

added to the study (validation set) and subjected to dissection, 

total RNA extraction and Real Time RT-PCR. Additionally, 

expression of a subset of miRNAs was assessed also in 10 mesothelial 

cell lines. 

Results. 48 miRNAs showed differential expression: 16 were 

upregulated whereas 32 were downregulated in mesotheliomas 

compared to normal pleuras. Principal Component Analysis 

showed clearcut separation between tumors and controls. Real 

Time PCR has so far confirmed the differential expression of 6 

out of 48 miRNAs; of these 6, miR-145 has proved to be highly 

sensitive and specific tumor biomarker through ROC curve 

analysis. Preliminary results in the MSTO211H cell line suggest 

that this miRNA is controlled by p53 in MM, consistently with 

previously published data in breast and colon cancer cell lines. 

miR-145 is reported to target genes involved in invasiveness and 

metastasis; among these, MUC1 has been found upregulated in 

MM and indicated as a potential therapeutic target. 

We are now aiming at identifying other biological targets of 

miR-145 to detail its functions in mesothelial cell line models, 

in order to contribute to an improved clinical management of 

mesothelioma. 

ThinPrep© cytological specimens in detection of 

eGfr mutations: a comparison with histological 

specimens in NSClC 

1)Mattioli E. 2)Daprile R. 3)Rubini V. 4)Petriella D. 5)Pinto R. 

6)Galetta D. 7)Simone S. 8)Tommasi S. 

1)Pathological anatomy unit, Nci “Giovanni Paolo II”, Bari, Italy 2)Pathological 

anatomy unit, Nci “Giovanni Paolo II”, Bari, Italy 3)Pathological 

anatomy unit, Nci “Giovanni Paolo II”, Bari, Italy 4)Clinical 

experimental oncology laboratory, Nci “Giovanni Paolo II”, Bari, Italy 

5)Clinical experimental oncology laboratory, Nci “Giovanni Paolo II”, 

Bari, Italy 6)Medical and experimental oncology department, Nci “Giovanni 

Paolo II”, Bari, Italy 7)Pathological anatomy unit, Nci “Giovanni 

Paolo II”, Bari, Italy 8)Clinical experimental oncology laboratory, Nci 

“Giovanni Paolo II”, Bari, Italy 

Background EGFR mutations (exons 19-21) are mandatory to 

set target therapy in NSCLC. Surgical specimens are suitable for 

diagnosis and biological characterization of NSCLC in less than 

30% of cases. Therefore, cytological samples or small biopsies 

should be used.


Methods. 18 Liquid Based Cytology (LBCs): 13 percutaneous 

US guided FNC of neoplastic lung nodules, 3 Pleural liquid, 1 

bronchial washing and 1 sputum and 20 histological samples 

entered the study. After cytological diagnosis, LBC samples with 

more that 50% of neoplastic cells have been routinely stored at 

-20°C for molecular analyses. Immunophenotyping has been 

performed on cell block of the same material. Histology has been 

performed on paraffin embedded specimens after lung surgery. 

DNA has been extracted by commercial kit (Qiagen DNAmicro 

kit) and analyzed for K-RAS (exon 2) and EGFR mutations (exons 

19-21) by direct sequencing. 

Results. FNC material was suitable to analyze 15/18 LBC 

(83.3%) and 17/20 (85%) histological specimens. Lung cancer 

diagnoses were classified (WHO, 2004) as follows: A) LBC: 10 

Adenocarcinomas (ADC), 1 SCLC, 3 Squamous Cell Carcinomas 

(SCC), 1 metastatic (colon) ADC and 3 NSCLC/NOS; B) Histology: 

12 ADC (2 mucinous), 5 SCC, 2 ADC/NOS, 1 BAC. 

2/15 LBCs (13.3%) and 2/17 (11.8%) histological evaluable 

samples showed EGFR gene mutation: all cases were ADCs; 1 

deletion in exon 19 and 1 missense mutation in exon 21 were 

detected in LBC samples, 1 deletion in exon 19 and 1 insertion 

in exon 20 were present in histological specimens. Only 1/12 

analyzed patients presented K-RAS mutation in lung ADC/ 

NOS. It is to be outlined that when tissue DNA resulted unamplifiable, 

it was possible to determine mutations on cytological 

specimen. 

In conclusion, LBC samples stored at -20°C are useful for molecular 

detection directed to therapy setting. This algorithm appears 

as a feasible strategy for storing FNC material cell banks. 

references 

Savic S, et al. Comprehensive epidermal growth factor receptor gene 

analysis from cytological specimens of non-small-cell lung cancers. 

Br J Cancer 2008;98(1):154-60. 

Paradiso A, et al. Exhaled breath condensate is not suitable to detect 

EGFR somatic mutations. Eur Respir J 2008;32(4):1126-7. 

Her2-mediated epigenetic control in human 

breast cancer: CPT1A as a novel biomarker and 

target for therapy 

1)Mazzarelli P. 2)Pucci S. 3)Zonetti M.J. 4)Spagnoli L.G. 

1)Biopatologia, Policlinico di Tor Vergata, Roma, Italia 2)Biopatologia, 

Policlinico di Tor Vergata, Roma, Italia 3)Biopatologia, Policlinico di Tor 

Vergata, Roma, Italia 4)Biopatologia, Policlinico di Tor Vergata, Roma, 

Italia 

Background. The altered metabolism of tumor cells may be a 

potential means by which these cells evade programmed cell 

death, favoring survival and tumoral growth. In particular, lipid 

metabolism is markedly altered in the tumoral context. Fatty acids 

synthase (FASN), the major enzyme required for the synthesis 

of fatty acids, is up-regulated in a wide array of solid tumors and 

ErbB2 (HER2) receptor, amplified in 25% of breast cancers, has 

been recognized as activator of FASN promoter. On the other 

hand, fatty-acids β-oxidation is inhibited in the tumoral context. 

We previously showed that the carnitine palmitoyl transferase 

I (CPT I), rate-limiting enzyme in the transport of long-chain 

fatty acids for β-oxidation, was significantly decreased in the 

mitochondria and it strikingly localized in the nuclei of tumor 

samples, where it could be implicated in the epigenetic regulation 

of transcription by its link to HDAC1. 

Methods. Here we analyze human breast carcinomas and breast 

cancer cell lines (SK-BR3, BT474, MCF7) correlating the HER2 

status with FASN protein expression. Moreover, we transfected 

MCF7 cells with small interfering RNAs (siRNAs) to silence 

CPT1A nuclear expression and analyzed the histone and non 

histone acetylation and the gene expression downstream effects, 

by microarray analysis. 

319 

Results. We confirmed that FASN was over-expressed in a high 

percent of breast cancer samples and it could be indicator of 

HER2 transduction activity. Then we displayed that the silencing 

of nCPT1A was a sufficient condition to induce apoptosis 

in MCF7 cells. The cell death triggered by RNA interference 

correlated with decreased HDAC activity and hyperacetylation 

of histone- and non histone-proteins involved in cancer-relevant 

death pathways. Gene array studies showed that pro-apoptotic 

genes such as BAD and CASP9 were up-regulated, whereas 

invasion and metastasis-related genes were down-modulated at 

transcriptional level. In breast cancer, the activation of Her2/Neu 

signaling and the altered metabolism indirectly induce nCPT1A 

that regulates pro-survival genes at epigenetic level, representing 

a potential target for anti-cancer therapy. 

Valutazione dello stato Her 2 nel carcinoma 

della mammella con anticorpo CB11: metodiche 

immunoistochimiche a confronto 

G. Mazzoleni, M. Herz, M. Lüthy, E. Hanspeter, A. Kasal 

Anatomia Patologica Ospedale Regionale di Bolzano 

Introduzione. La determinazione dello stato HER 2 è essenziale 

per una corretta strategia terapeutica nel carcinoma della mammella. 

Sono approvati e comunemente utilizzati sia metodi immunoistochimici 

con evidenziazione del recettore sulla membrana, 

che metodi FISH con valutazione dell’amplificazione del gene. 

L’anticorpo dal Clone CB11 per il dominio interno del recettore 

è comunemente utilizzato nei laboratori di anatomia patologica, 

solitamente come screening per selezionare i casi ++ (equivoci) 

da verificare con FISH, considerata dai più il gold standard 1 2 . 

Oracle è un kit in fase di approvazione FDA con anticorpo CB11, 

utilizzando il quale abbiamo avuto risultati controversi con i controlli 

di qualità NordiQc e Ringversuch Quip. Abbiamo pertanto 

voluto confrontare diverse metodiche utilizzando l’anticorpo 

CB11 (Novocastra) e il kit Oracle con diversi protocolli di 

smascheramento antigenico paragonandoli al risultato ottenuto 

con la FISH. 

Materiali e metodi. È stato valutato lo stato HER 2 su 101 casi 

consecutivi di carcinoma mammario sia in situ che infiltrante, 

utilizzando le seguenti metodiche su apparecchiatura completamente 

automatica Leica Bond Max. 

cB11 (novocastra) 1:400 senza antigen retrival 

Kit oracle leica prediluito er1(ph6) 25’ 



L’amplificazione del gene è stata valutata in tutti i casi mediante 

FISH utilizzando le sonde HER 2 Vysis. 

Risultati. Per lo score delle reazioni immunoistochimiche si sono 

seguite le indicazioni ASCO/CAP. Per la FISH negativo con rapporto 

< 1.8 e positivo > 2.2 (equivoco fra i due valori) 

I risultati sono rappresentati nella seguente tabella: 

nr. casi % CB11 ER1 ER2 10 ER2 30 FISH 

63 62,4 - / + - / + - / + - / + - 

16 15,8 - / + - / + - / + ++ - 

4 3,9 - / + - / + ++ ++ - 

4 3,9 - / + + ++ +++ - 

5 4,9 +++ +++ +++ +++ + 

5 4,9 - / + +++ +++ +++ + 

1 0,9 - / + ++ ++ +++ - 

1 0,9 ++ ++ ++ +++ + 

1 0,9 ++ +++ +++ +++ + 

1 0,9 ++ + + + - 

101

320 

Conclusioni. Dall’analisi delle reazioni risultava chiaramente 

un trend di progressiva intensità della reazione dal CB11 verso 

ER2 30’. Anche nei casi concordanti negativi appariva evidente 

una maggiore intensità di colorazione con pH9 sia nelle strutture 

normali che negli elementi tumorali. Dallo studio emerge che 

nessun caso amplificato alla FISH sfuggiva alla determinazione 

con Oracle ER1 (un solo caso ++) e ER2 (un solo caso ++ con 

10’) contrariamente a quanto avvenuto nei controlli di qualità 

esterni (NodiQc e Quip 2009). Tuttavia con ER2 30’ risultavano 

5 casi iperespressi +++ negativi alla FISH. Per contro con CB11 

in 5 casi le pazienti sarebbero state etichettate negative (-/+) e 

routinariamente non inviate alla FISH. Si dimostra quindi una 

maggiore affidabilità del kit Oracle rispetto all’uso di CB11 

standardizzato in casa. Il pH9 tende ad aumentare l’intensità di 

reazione aumentando i casi da controllare con FISH già con ER2 

10’. Con ER2 30’ si ha un numero inaccettabile di falsi positivi 

(metodica approvata nel run B8 Nordiqc2009). 

Bibliografia 

1 Purdie CA, et al. Histopathology 2010;56:702-7. 

2 Sauter GJ. Clin Oncol 2009;27:1323-33. 

K-ras mutational testing before and after 

neoadjuvant chemo-radiation (NCrT) in patients 

(PTS) with locally advanced rectal cancer (lArC) 

1)Molinari F. 2)Zanellato E. 3)Nucifora M. 4)Riva A. 5)Saletti 

P. 6)Franzetti-pellanda A. 7)Crippa S. 8)Mazzuchelli L. 9)Frattini 

M. 

1)Laboratorio diagnostica molecolare, Istituto cantonale di patologia, 

Locarno, Svizzera 2)Laboratorio diagnostica molecolare, Istituto cantonale 

di patologia, Locarno, Svizzera 3)Laboratorio diagnostica molecolare, 

Istituto cantonale di patologia, Locarno, Svizzera 4)Laboratorio 

diagnostica molecolare, Istituto cantonale di patologia, Locarno, Svizzera 

5)Istituto oncologico della svizzera italiana, Ente ospedaliero cantonale, 

Bellinzona, Svizzera 6)Istituto oncologico della svizzera italiana, Ente 

ospedaliero cantonale, Lugano, Svizzera 7)Patologia clinica, Istituto cantonale 

di patologia, Locarno, Svizzera 8)Patologia clinica, Istituto cantonale 


Istituto cantonale di patologia, Locarno, Svizzera 

Background. K-Ras testing represents the prerequisite before 

the administration of EGFR-targeted therapies to metastatic 

colorectal cancer (mCRC) pts. The correct choice of tumor 

blocks on which K-Ras sequencing must be performed is 

fundamental. For patients with LARC, an adequate amount of 

primary tumor tissue either before or after NCRT is not always 

available. In addition information on K-Ras testing after NCRT 

are not available 

Methods. Tumor biopsies, obtained before and after NCRT, of 

61 pts with LARC were examined for K-Ras status. A careful 

microdissection was performed at microscope in all cases. K-Ras 

was evaluated both by direct sequencing (DS, with a sensitivity 

of 10-20%) and mutant-enriched PCR (ME-PCR, with a sensitivity 

of 0.01%) 

Results. DNA was amplifiable in all cases. In pre-treatment biopsies, 

DS revealed K-Ras mutations in 22 cases (36%), ME-PCR 

in 26 (43%). In post-treatment biopsies, DS found K-Ras mutations 

in 13 cases (21%), ME-PCR in 24 (39%). Six cases showed 

a discordant pattern between pre and post-therapy biopsies: 4 pts 

showed the K-Ras mutation limited to the pre-treated biopsy (in 2 

cases detected by DS and ME-PCR, in 2 cases only by ME-PCR), 

2 pts showed the mutation in post-treatment biopsy (detected only 

by ME-PCR). 

Conclusions. The use of ME-PCR enhances the detection of 

K-Ras mutations in pts with LARC treated with NCRT. Discrepancies 

between pre- and post-treatment biopsies may occur. 

If pre-treatment biopsy is available, DS can be routinely used. 

Post-treatement biopsies should be preferentially investigated by 

means of high sensitive methodologies, such as ME-PCR. 


Simultaneous analysis of KrAS and BrAf mutations 

by allele specific quantitative PCr using locked 

nucleic acid modified primers and beacon probes 

1)Morandi L. 2)De biase D. 3)Visani M. 4)Demaglio G. 5)Pizzolitto 

S. 6)Pession A. 7)Tallini G. 

1)Ematologia e scienze oncologiche, Bellaria, Bologna, Italy 2)Patologia 

sperimentale, Bellaria, Bologna, Italy 3)Patologia sperimentale, Bellaria, 

Bologna, Italy 4)SOC Anatomia patologica, Azienda ospedaliero-universitaria, 

Santa maria della misericordia, Udine, Italy 5)SOC Anatomia 

patologica, Azienda ospedaliero-universitaria, Santa maria della misericordia, 

Udine, Italy 6)Patologia sperimentale, Bellaria, Bologna, Italy 

7)Ematologia e scienze oncologiche, Bellaria, Bologna, Italy 

Background. KRAS exon 2 mutations occur in 35-45% of 

metastatic colorectal cancers (mCRC) and preclude responsiveness 

to EGFR-targeted therapy with cetuximab or panitumumab. 

BRAF mutations at V600E are present in 5-10% of mCRC and 

have been inversely correlated with progression free and overall 

survival in KRAS wild type mCRC patients, underscoring the 

importance of testing mCRC for both KRAS and BRAFV600E. 

We report a novel allele specific qPCR assay to simultaneously 

detect KRAS exon2 and BRAFV600E mutations using locked 

nucleic acid (LNA)-modified Allele Specific primers and internal 

molecular beacon probes (ASLNAqPCR). 

Materials. LNA-modified allele specific primers were designed 

to identify the vast majority of KRAS mutations (G12A, G12C, 

G12D, G12R, G12S, G12V, G13D) in CRC. To define the 

dynamic range we diluted mutated DNA from the SW620 and 

CAL62 (KRAS+), and the ARO and OCUT (BRAFV600E+) cell 

lines. We analyzed 127 consecutive tumor samples from patients 

with mCRC. 

Results. Our ASLNAqPCR assay has a sensitivity of 0.1% for 

KRAS exon 2 mutations and of 0.01% for BRAFV600E. There 

were no false positive results in non-neoplastic controls. A KRAS 

mutation was detected in 52 (40.9%) and a BRAF mutations in 

8 (6.3%) tumors. Direct sequencing and the ASLNAqPCR assay 

generated discordant result in 12 samples (9.4%). These were due 

to higher sensitivity of the ASLNAqPCR assay in samples with 

low tumor cell percentage (10 cases) and to the occurrence of 

mutations not covered by the ASqPCR assay (2 cases with KRAS 

Q61H and G12F, respectively). 

High sensitivity, specificity and quantification of the mutant/ 

wild type allele ratio give to this assay very good performances. 

The ability of ASLNA-qPCR to identify KRAS or BRAF 

mutations with a complete assay time of 2 hours since DNA 

purification, compares extremely favorably with standard direct 

sequencing. 

Clonality analysis by aCGH and d-loop mtDNA 

direct sequencing of multicentric ductal 

carcinomas in situ of the breast 

Luca Morandi # , Federica Flamminio # , Dario De Biase # , Michela 

Visani # , R. Masetti § , Maria Pia Foschini # , Vincenzo Eusebi # 

# Dipartimento di Ematologia e Scienze Oncologiche, Sezione di Anatomia 

Istologia e Citologia Patologica “M. Malpighi” Università -ASL Ospedale 

Bellaria, Bologna; § Centro Interdipartimentale di Senologia, Università 

Cattolica Policlinico “A, Gemelli”, Roma 

Background. It has been suggested that ductal carcinoma in 

situ (DCIS) develops within a single lobe. This view has been 

challenged by a study of DCIS using large sections. It was 

found that well differentiated G1 DCIS/DIN1 is a multicentric 

condition in 76.9% of the cases, while poorly differentiated 

duct carcinoma in situ (G3 DCIS) is often unifocal. Aim of 

the study was to find out common or different DNA mutations 

among different DIN which might reflect clonal (unicentric) or 

polyclonal (multicentric) origin of multiple foci when present 

in the same case.


Materials and methods. 15 randomly selected patients (5 

DCIS/DIN1, 5 G2 DCIS/DIN2, 5 G3 DCIS/DIN3) with multiple 

DCIS foci, were studied with large macro-sections. Two to 4 

foci of DIN from each case were laser-microdissected. DNA was 

purified and sequenced for mtDNA D-loop region and evaluated 

by array comparative genomic hybridization (aCGH). Genetic 

distance among different foci was visualized by phylogenetic 

analyses using the neighbor-joining (NJ) method. 

Results. Patients were all females ranging in age from 36 to 87 

years (mean 65.06). mtDNA and aCGH data indicated that all 

five cases of multiple DIN1, located at a distance of 12 mm or 

superior, showed a remarkable genetic distance among them. 

Multiple foci microdissected from 3 out of 5 G2 and 4 out of 5 

G3 cases, showed a closed phylogenetic relationship. 

Conclusions. 100% of DIN1 cases were polyclonal that has suggested 

a multicentric origin (12-55 mm). On the contrary most 

of the G2 and G3 DCIS/DIN cases revealed a possible common 

ancestor cell (clonality) among different lesions. These data are 

consonant with previous findings that demonstrated that poorly 

differentiated DCIS spreads along the affected duct in a continuous 

fashion, whereas well-differentiated DCIS shows a multifocal 

type of growth with gaps of intervening noninvolved gland in 

most of the cases. 

mtDNA d-loop sequence analysis as a tool to 

distinguish between recurrences and second 

primary tumours in oral squamous cell carcinoma 

1)Leonardi E. 1)Morandi L. 2)Marchetti C. 3)Badiali G. 4)Montebugnoli 

L. 5)Foschini MP. 

1)Haematology and Oncology “L. and A. Seragnoli”, Bellaria-University 

of Bologna, Bologna, Italy 2)Oral and Maxillofacial Surgery, S.Orsola- 

Malpighi-University of Bologna, Bologna, Italy 3)Oral and Maxillofacial 

Surgery, S.Orsola-Malpighi-University of Bologna, Bologna, Italy 4)Oral 

Sciences, University of Bologna, Bologna, Italy 5)Haematology and Oncology 

“L. and A. Seragnoli”, Bellaria-University of Bologna, Bologna, 

Italy 

Background. One main issue in the prognosis and management 

of oral squamous cell carcinoma (OSCC) concern the development 

of secondary tumours and the possibility to distinguish 

recurrences from true second primaries. Molecular analysis of genetic 

alterations can be performed for a better characterization of 

neoplastic features and to investigate clonal origin of the tumours 

and their developing pathway. 

Methods. Nine patients presenting at least two locoregional neoplastic 

lesions were selected for the study. Mitochondrial DNA 

was extracted from formalin-fixed, paraffin-embedded tissue 

sections and D-loop region was analyzed to evaluate the genetic 

relationship between subsequent tumours. 

Results. A clear clonal relationships between two subsequent 

neoplastic presentations was observed in 4 of 9 cases. On the 

other hand, remaining patients showed more complex phylogenetic 

trees, where the second tumour cell population could carry 

mutations evident also in the next healthy tissue or, more rarely, 

gain completely different genetic alterations. 

Discussion. Our data suggested that recurrent tumours could 

develop from clonal cell populations or, when multiple genetic 

patterns are observed, from further altered cells of the oral mucosa, 

consistently with a cancerization field hypothesis. More 

studies are needed to evaluate the genetic relationship between 

primary and second OSCC to develop an effective molecular 

tool to identify patients at higher risk of a recurrence or a second 

primary tumour. 

321 

Is tumour budding reproducible as prognostic 

factor? 

1)Morichetti D. 2)Pusiol T. 3)Piscioli F. 

1)Institute of Anatomic Pathology, S.Maria Del Carmine Hospital Rovereto, 

Rovereto, Italy 2)Institute of Anatomic Pathology, S.Maria Del Carmine 

Hospital Rovereto, Rovereto, Italy 3)Institute of Anatomic Pathology, 

S.Maria Del Carmine Hospital Rovereto, Rovereto, Italy 

Background. Tumour Budding (TB) has been shown to be a 

negative prognostic factor in patients with stage II colorectal 

carcinoma (CRC) 1 and may be useful for identifying the 

subset of T3N0M0 patients at high risk of recurrence who 

may benefit from adjuvant therapy 2 . TB has been reported 

as a pathological marker suggesting high malignant potential 

and decreased postoperative survival in patients with well- or 

moderately- differentiated pT3 CRC 3 . Shinto et al. 4 suggest 

that TB involves two independent processes: the loss of cellular 

cohesion and the cellular activation leading to increased 

invasiveness. Regarding the prognosis of CRC, the impact of 

TB may be accepted, when the definition is appropriate, reproducible 

and precise. 

Method. We have examined the definitions of TB reported in the 

published literature relating to TB as prognostic factor. 

Result. The term TB itself did not appear in the literature until 

Hase et al. 5 used it and it was not reported in other organs. These 

authors defined TB as “microscopic clusters of undifferentiated 

cancer cells just ahead of the invasive front of the lesion”. The 

number of malignant cells was not specified. This definition has 

been accepted in numerous studies regarding the value of TB 

as prognostic factor. Other authors reported TB as “single or a 

group of < 5 detached tumour cells, usually but not always at the 

invasive front” 2 6 7 . 

Discussion and Conclusions. TB may be accepted as prognostic 

factor only when the number of neoplastic cells is specified. 

Conversely, the reproducible prognostic budding scoring system 

is problematic. Consequently the prognostic conclusions of the 

reported studies may be accepted only when the number of tumour 

cells is specified. Moreover, TB may be found adjacent to 

glandular malignant structures. In these cases it may be misinterpreted 

and over diagnosed because it may represent a feature of 

the infiltrative front itself. 

references 

1 Nakamura T, Mitomi H, Kanazawa H, et al. Tumor budding as an index 

to identify high-risk patients with stage II colon cancer. Dis Colon 

Rectum 2008;51:568-72. 

2 Wang LM, Kevans D, Mulcahy H, et al. Tumor budding is a strong 

and reproducible prognostic marker in T3N0 colorectal cancer. Am J 

Surg Pathol 2009;33:134-41. 

3 Okuyama T, Oya M, Ishikawa H. Budding as a useful prognostic 

marker in pT3 well- or moderately-differentiated rectal adenocarcinoma. 

J Surg Oncol 2003;83:42-7. 

4 Shinto E, Mochizuki H, Ueno H, et al. A novel classification of tumour 

budding in colorectal cancer based on the presence of cytoplasmic 

pseudo-fragments around budding foci. Histopathology 2005;47:25- 

31. 

5 Hase K, Shatney C, Johnson D, et al. Prognostic value of tumor 

“budding” in patients with colorectal cancer. Dis Colon Rectum 

1993;36:627-35. 

6 Prall F, Nizze H, Barten M. Tumour budding as prognostic factor in 

stage I/II colorectal carcinoma. Histopathology 2005;47:17-24. 

7 Ueno H, Murphy J, Jass JR, et al. Tumour ‘budding’ as an index to 

estimate the potential of aggressiveness in rectal cancer. Histopathology 

2002;40:127-32.

322 

Pelvic epidermal cyst of the round ligament 

with seborrheic keratosis – like changes in its wall 

1)Morichetti D. 2)Pusiol T. 3)Piscioli F. 

1) Institute of Anatomic Pathology, S.Maria del Carmine Hospital Rovereto, 

Rovereto, Italy 2)Institute of Anatomic Pathology, S.Maria del Carmine 

Hospital Rovereto, Rovereto, Italy 3)Institute of Anatomic Pathology, 

S.Maria del Carmine Hospital Rovereto, Rovereto, Italy 

Background. We report herein the first case of Epidermal Cist 

(EC) of Round Ligament (RL), characterized by seborrheic keratosis-like 

change in the wall. 

Methods. A 30-years-old female was referred to our institution 

with abdominal pain. 

Results. Ultrasonography showed an hypoechoic heterogeneous, 

round mass adjacent to the lower extremity of the left ovary, measuring 

4.5 cm in maximum diameter. Contrast-enhanced Computed 

Tomography of the pelvis in the venous phase revealed a 

round cystic lesion with inhomogeneous fluid content (4.7-cm in 

diameter) in the side of the left large ligament and anteriorly to 

the omolateral adnexa. A laparoscopic resection of the mass was 

performed. Macroscopically the mass measured 6 cm. × 6 cm. × 

3.5 cm. On cut section the mass was an unilocular cyst filled with 

soft, yellow, amorphous material. Histologically the cystic wall 

was lined by a stratified squamous epithelium with a granular 

cell layer. The cavity contained keratin material. The cystic wall 

showed numerous areas with close-set basaloid cells and pseudohorn 

cysts. The latter aspect consisted of cystic invaginations of 

the epithelium filled with surface keratin, which in a given microscopic 

section may be cut in cross-section, thereby appearing 

as “cysts” within the involved epithelium. 

Discussion. Tumors of the RL may present intra-abdominally, 

in the inguinal canal or in the labium. Common RL lesions are 

leiomyoma 1 2 , endometriosis and mesothelial cyst 3 . Only one 

case of EC of RL has been reported in the literature to day. In 

1968 Lecca and Belvederi described a mass arising from the left 

RL in a 23 years old-woman with histological features of EC 4 . 

Few cutaneous ECs exhibit seborreheic keratosis-like changes in 

their wall 5 . In all extracutaneous reported ECs this feature is not 

found. The present case is unique in that it is an extracutaneous 

EC with seborreheic keratosis-like changes in cystic wall arising 

from a very unusual site as RC. 

references 

1 Bakotic BW, Cabello-Inchausti B, Willis IH, Suster S. Clear-cell epithelioid 

leiomyoma of the round ligament. Mod Pathol 1999;12:912- 

8. 

2 Lösch A, Haider-Angeler MG, Kainz C, et al. Leiomyoma of the round 

ligament in a postmenopausal woman. Maturitas 1999;31:133-5. 

3 Breen JL, Neubecker RD. Tumors of the round ligament. A review of 

the literature and a report of 25 cases. Obstet Gynecol 1962;19:771- 

80. 

4 Lecca U, Belvederi GD. Considerations on a case of epidermoid cyst 

of the round ligament. Minerva Ginecol 1968;30:1782-3 

5 Rahbari H. Epidermoid cysts with seborrheic verruca-like cyst walls. 

Arch Dermatol 1982;118:326-8. 

Molecular analysis of extra-neuraxial 

hemangioblastoma. A study of 6 cases 

L.A. Muscarella * , J. Lamovec † , P. Parrella * , N. Zidar ± , L. 

D’Agruma § , M. Michal ‡ , V. Guarnieri § , M. Coco * , J.C. Fanburg- 

Smith , L. Zelante § , M. Bisceglia^ . 

* § ^ Laboratory of Oncology, Medical Genetics Service, and Unit of Anatomic 

Pathology, IRCCS “Casa Sollievo della Sofferenza” Hospital, San 

Giovanni Rotondo, Foggia, Italy; † Department of Pathology, Institute of 

Oncology, Ljubljana, Slovenia; ± Institute of Pathology, Medical Faculty, 

University of Ljubliana, Slovenia; ‡ Department of Pathology, Charles University 

Medical Faculty Hospital, Pilsen, Czech Republic; Department of 

Soft Tissue Pathology, Armed Forces Institute of Pathology, Washington, 

D.C., USA. 


Background. Hemangioblastoma (HGB) is a discrete, solid or 

cystic, potentially curable tumor made up of variable combination 

of interstitial stromal cells of uncertain histogenesis and a 

rich network of thin-walled vessels (HGB). While capillary HGB 

is the most frequent manifestation of von Hippel-Lindau (VHL) 

disease, an autosomal dominant condition, the majority of cases 

(70%) are of the nonfamilial, sporadic type. VHL-associated 

HGB may occur in any part of the central nervous system (neuraxial 

HGB), including optic nerve and retina. Rarely, HGB may 

also affect spinal nerve roots 1 , filum terminale, and cauda equina 

2 (perineuraxial HGB), either in the context of VHL syndrome 

or sporadically. Even more rare are capillary HGB which have 

been observed either in internal organs, such as liver, pancreas, 

and kidney, almost always in a clinical setting of VHL, or in soft 

tissue 3-5 , or even in bone 6 (peripheral HGB). Patients with VHL, 

usually exhibit mutations of a tumor suppressor gene mapped to 

chromosome 3p25-26 (VHL gene). According to the “two hit” 

theory of Knudson, HGB in VHL syndrome is due to a second 

“hit” occurring in the tissue, where the tumor arises (somatic 

mutation as a second hit). To the best of our knowledge, only 2 

cases of extra-axial HGB have been analyzed at the molecular 

level, one was on multifocal, recurrent lesions, arising from different 

spinal nerve roots of midcervical medullary segments in 

a 57 year-old man 1 , and the other involved the soft tissue of the 

ankle in a 74 year-old woman 4 . 

Objectives. To report our molecular investigation on 6 cases of 

extra-neuraxial HGB. 

Materials and Methods. Patients and tissue specimens. 6 cases 

of extraneuraxial HGB were found in the joint files of the institutions 

involved. 5 patients were females and 1 was a male. All 

tumors were extraneuraxial, 4 in intraspinal/paraspinal locations, 

and 2 in peripheral soft tissue. Case 1. A 40-year old female with 

VHL disease, affected by 2 extradural intraspinal HGB, arising 

from L1-L2 and L5-S2 nerve roots, respectively, the latter 

extending into the retroperitoneal space. The patient was also 

operated on and diagnosed with adrenal pheochromocytoma and 

renal cell carcinoma. Case 2. A 58 year-old female diagnosed 

with an intraspinal and extraspinal, dumbbell HGB, arising from 

T9-T10 nerve roots, of 4.5 cm in size, largely protruding in the 

posterior mediastinum. Case 3. A 65-year-old female, affected 

by a T6-T9, intraspinal, extradural, tumor of 4 cm in size. Case 

4. A 74 year-old female, with a 2.5 cm tumor nodule located in 

the soft tissue on her right ankle. Case 5. A 74 year-old female, 

diagnosed with a C2-C3 intraspinal extradural tumor. Case 6. A 

47-year old male with a 7.0 cm retroperitoneal tumor. All tumors 

were surgically excised and histologically examined. Normal 

and tumoral tissue samples were available for this study from all 

cases. For all samples an extensive molecular characterization of 

the VHL gene was performed by Mutation, Fluorescent Loss of 

Heterozygosity (LOH) and Methylation Analysis. Mutation analysis 

was performed by searching for point mutations in the entire 

coding sequence and promoter region of the gene, including 

the exon-intron boundaries. LOH analysis was performed using 

three microsatellite markers flanking the VHL gene: D3S1335, 

D3S1038, D3S1317, and LOH or allelic imbalance (AI) value 

was quantified using the formula (peak 1 height/peak2 height 

in tumour DNA)/(peak 1 height/peak 2 height in normal DNA), 

and the cut-off for defining an AI was lowered to 0.5 whereas an 

AI > 0.5 and < 0.65 defined a borderline case. The methylation 

status in the promoter region of the VHL gene was determined by 

Methylation Specific PCR (MSP), and PCR products were analyzed 

on 3% agarose gels with ethidium bromide and visualized 

under UV illumination. 

Results. Patients. Case 1, case, 2, case 3, and case 5 are new 

cases. Case 4 and case 6 had already been the subject of previous 

reports 4 5 . Case 1 was clinically affected by VHL disease. Mutation 

analysis documented in the HGB tumor tissue of case 1 two 

nucleotide substitutions, which had previously been reported by


others in syndromic HGB, precisely, the germline c.452T > A 

substitution in exon 2, creating the aminoacid change I151N, 

and the somatic c.450G > A substitution in exon 3, resulting in 

the aminoacid change R167Q. Furthermore, most notably, by 

the same technique a new somatic nucleotide deletion in exon 

3 (c.598delA) was found in case 5, causing a frameshift with a 

premature predicting stop codon at position 201. Microsatellite 

analysis showed LOH for at least one informative marker in 

the following 3/6 tumours: in case 5, herein to be intended as a 

second hit, and therefore consistent with inactivation of both alleles; 

and in case 4 and case 6 as single hit. Methylation analysis 

did not disclose promoter methylation in any of the six tumour 

samples analyzed. No VHL genetic alteration was demonstrated 

in both case 2 and case 3 by means of any of the techniques 

employed. 

Discussion. VHL syndrome is due to germline inactivating mutations 

of the VHL tumor suppressor. The VHL gene is presumed 

to be involved also in the development of sporadic HGB. In this 

study we investigated the spectrum of VHL gene alterations in 6 

cases of extra-axial HGB. Concerning case 1 our genetic findings, 

including one germline (I151N) and one somatic (R167Q) 

VHL gene mutations, fully reflected the clinical manifestations 

of a clinically evident syndromic condition. As far as the 5 sporadic 

cases are concerned: in case 5 a somatic point mutation 

(c.598delA) and an LOH was identified, this being consistent 

with somatic inactivation of both alleles in a “two-hit” manner; 

in both case 4 and case 6 LOH only in the VHL gene was documented, 

and this is again in support of VHL gene involvement 

in the development of sporadic extra-axial HGB. The absence 

of any documented genetic alteration in the remaining sporadic 

tumors (case 2 and case 3) might be explained according to one 

of these reasons: 1. the genetic change is localized in the intronic 

or regulatory regions of the VHL gene, which were not examined; 

2. the genetic anomaly involves other genes, which interplay 

with the VHL expression and that were not investigated in these 

context. Previous molecular analyses performed on HGB tumor 

tissue did not document genetic alterations in VHL gene: in one 

case both complete sequence analysis and LOH analysis had been 

performed, and the failing to document any genetic alterations led 

to the conclusion that a molecular event directly involving the 

VHL gene may not be the causative factor in the tumorigenesis of 

extra-axial HGB 1 ; in another study, employing Mutation analysis 

only, no change in the coding sequence of VHL gene was found 4 , 

and this is one of the 2 cases in which this study disclosed LOH. 

Conclusion. This is the first report of VHL gene alterations 

identified in extra-axial HGB. Particularly, the genetic findings 

demonstrated in case 5 (double somatic hit) strongly confirm the 

hypothesis that the VHL gene is involved in the development of 

extra-axial HGB. 

references 

1 Raghavan R, Krumerman J, Rushing EJ, et al. Recurrent (nonfamilial) 

hemangioblastomas involving spinal nerve roots: case report. Neurosurgery 

2000;47:1443-8. 

2 da Costa LB Jr, de Andrade A, Braga BP, et al. Cauda equina hemangioblastoma: 

case report. Arq Neuropsiquiatr 2003;61:456-8. 

3 Fanburg-Smith JC, Gyure KA, Michal M, et al. Retroperitoneal peripheral 

hemangioblastoma: a case report and review of the literature. 

Ann Diagn Pathol 2000;4:81-7. 

4 Michal M, Vanecek T, Sima R, et al. Primary capillary hemangioblastoma 

of peripheral soft tissues. Am J Surg Pathol 2004;28:962-6. 

5 Nonaka D, Rodriguez J, Rosai J. Extraneural hemangioblastoma: a 

report of 5 cases. Am J Surg Pathol. 2007;31:1545-1. 

6 Panelos J, Beltrami G, Capanna R, et al. Primary Capillary Hemangioblastoma 

of Bone: Report of a Case Arising in the Sacrum. Int J Surg 

Pathol 2008 Jul 8. doi:1177/1066896908320549. 

Therapeutic impact of endoscopic ultrasound 

guided fine needle aspiration/biopsy 

of mediastinal lesions 

323 

N. Muscatiello, M. Di Maso, V. Nirchio * , C. Panella, E. Ierardi 

Gastroenterology Unity Univ., Ospedali Riuniti, Foggia, Italy; * U.O.S 

Cytopathology, departments of Pathology, Ospedali Riuniti, Foggia, Italy 

Background. Transoesophageal endoscopic ultrasuond whit fine 

needle is a minimally invasive procedure to demostrate unresecactability 

in lung cancer patients with our without enlarge mediastinal 

lesions whit our without lymphonodes invasions. 

Methods. The study was a prospective controlled study. EUS-FN 

was performed in 53 patients, 18 females and 35 males whit a 

curve array endosonography (Hitachi/Pentax Fg 36 UA) and 21 

G needle and 19 QC (W.C.), developed at Foggia University. 22 

Patients with lung cancer were refered to EUS-FN either due to 

suspected mediastinal tumor invasion or enlarged lymph nodes 

suspect by CT. 31 patients were referred with either a solid lesion 

or enlarged lymph nodes of unknown origin located adjacent to 

the esophagus demostrated by CT. 

Results. All diagnoses were confirmed either by toracotomy 

or clinical follow-up. 49 patients had lung cancer (6 patients in 

stadium IB, 5 patients in stadium IIB, 21 patients in IIIA and 16 

patients in staium IIIB after final classification), one patient had 

an abscess of the lung, one patient had Hodgkin lymphoma, one 

patient had non Hodgkin lymphoma one patient had leiomyosarcoma 

of the esophagus, one patients had sarcoidosis as final 

diagnosis. The cytological diagnosis obtained by EUS-FN was 

conclusive for cancer in 36 lesion and consistent whit a benign 

lesion in 17 patients. There were 4 false negative diagnoses and 

no false positive diagnoses. The clinical impact of EUS-FN was 

as follow: in 4 patients with a negative mediastinoscopy EUS- 

Fn demostrated cancer in the mediastinum. AT operation one 

of the patients had a positive lymph nodes by the esophagus 

and 3 patients had ingrowt of the aortic arc. The sensitivity of 

cytology was 68%, within EUS-FN was 90% and the specificity 

was 100%. The positive predictive value was 100% and the 

negative predictive value was 76%- The where one minimally 

complication. 

Conclusion. EUS-FN is a safe and reliable method in the evaluation 

of patients whith a solid lesion of the mediastinum- Has a 

huge therapeutic impact and should be considered for diagnosis 

of cancer spread to the mediastinum as well as for primary 

diagnosis of solid lesion located in the mediastinum. Citology 

diagnosis would are associated with istology. 

Hemorragic endovasculitis of chorionic villi in 

association with haemophilia a: a case report 

Musizzano Y., Pacella E., Malachina S., Traverso V., Fulcheri 

E. 

“U.O. Anatomia Patologica Universitaria, Azienda Ospedaliero Universitaria 

“San Martino”, Genova, Italia” 

Background. Hemorrhagic endovasculitis (HEV) is a disruptive 

lesion of placental vessels of any order, showing endothelial injury, 

hemorrhage, and extravasation of fragmented and deformed 

red blood cells (RBC). It is sometimes considered as a distinct 

idiopathic lesion, while according to others it overlaps with fetal 

thrombotic vasculopathy. HEV is seen in less than 5-6% of all 

pregnancies, but its prevalence is much higher in stillbirth, IUGR, 

fetal neurologic injuries and nonimmune hydrops. 

Case report. A 35 years old pregnant woman at 10 gestational 

weeks, affected by Haemophilia A, presented with sudden and 

unexpected uterine bleeding. Gynecological examination, including 

hysteroscopy, confirmed miscarriage; hence, curetting 

of the uterine cavity was performed and the specimen was sent 

to the pathology lab. On histopathological examination, mas-

324 

sive recent hemorrhage was seen in decidual and retroplacental 

site and in the chorionic plate. The villi, in particular, showed 

unusual diffusely hemorrhagic stroma, with fragmented and 

often immature RBC. Anti-CD34 revealed discontinuous endothelia 

and fragmented endothelial cells around vascular lumens, 

while Perls was focally positive in villous stroma, sometimes 

reminding of dystrophic mineralization of the basement membrane. 

Conclusions. HEV can be very harmful in cases with possible hereditary 

transmission of a hypocoagulative state, as can be seen in 

cases with known haemophilic parents and conceptus of unknown 

sex. Unfortunately, fetal karyotype is usually not assessed following 

the first miscarriage, nor accurate histopathological examination, 

if any, is performed. Conversely, the latter should include 

the evaluation of both maternal and fetal structures; moreover, 

the use of two easy-to-do stainings, can disclose the presence of 

underlying HEV, hence contributing to identify high-risk women 

and to correct manage their further pregnancies, in the setting of 

reproductive pathology. 

Chronic hystiocytic intervillositis: report of two 

cases 

Musizzano Y., Pacella E., Traverso V., Malachina S., Fulcheri 

E. 

U.O. Anatomia Patologica Universitaria, Azienda Ospedaliero Universitaria 

“San Martino”, Genova, Italia 

Background. Chronic hystiocytic intervillositis (CHI) is an idiopathic 

placental lesion showing uniform intervillous hystiocytic 

infiltrate and lacking chronic villitis or polymorphous inflammation; 

it is associated with often recurrent and kariotypically 

normal early spontaneous abortions (ESA), fetal growth retardation 

(FGR), intrauterine fetal death, and maternal autoimmunity 

(AI). It is much more rare in the 2 nd and 3 rd trimester; anyway, we 

present here two cases of CHI occurring at 38 and 17 gestational 

weeks (GW). 

Case #1. A 42 years old woman underwent operative delivery at 

38 GW, giving birth to a male of 2100 g with single umbilical 

artery and cerebral blood flow modifications. Histological examination 

of the placenta revealed massive uniform intervillous 

infiltrate, strongly and diffusely CD68+, and perivillous fibrinoid 

deposition. Both the child and his mother are alive and well at 

13-month follow-up. 

Case #2. A 31 years old recurrent aborter had her third pregnancy 

loss at 17 GW; ultrasound performed 24 h before miscarriage 

discovered FGR. Placental histopathology showed perivillous 

fibrinoid, dense CD68+ monocyte-macrophagic infiltrate; moreover, 

thrombosed vessels, stromal edema and thickened vascular 

walls were seen in the decidua. Fetal autopsy confirmed FGR 

consistent with 14 GW and isolated paraombelical gastroschisis. 

One year later, the patient underwent ESA at 9 GW; the histopathological 

examination of this fourth miscarriage disclosed 

decidual sclerosing vasculopathy associated to endometritis, villitis 

and intervillitis. 

Conclusions. Although more common in the 1 st trimester 

and in recurrent aborters, CHI can be seen at variable gestational 

age, and in women experimenting previous successful 

pregnancies. Anyway, in both cases considered here, some 

accompanying histopathological features suggested maternal 

AI; In our opinion, following a diagnosis of CHI, even subclinical 

AI should be investigated, in order to prevent further 

miscarriages. 


Mineralization of trophoblastic basement 

membrane as a marker of chronic villitis in early 

spontaneous abortion 

Musizzano Y., Traverso V., Malachina S., Fulcheri E. 

“U.O. Anatomia Patologica Universitaria, Azienda Ospedaliero Universitaria 

“San Martino”, Genova, Italia” 

Background. Mineralized trophoblastic basement membrane 

(MBM) can be seen in a number of conditions, including stillbirth, 

hypertension, congenital and chromosomal abnormalities, prematurity, 

fetal thrombotic vasculopathy, and infections. In the latter, 

MBM can represent a marker of chronic villitis (CV), particularly 

when torpid and associated with longer fetal survival. Anyway, 

the term MBM alone does not specificate if iron, calcium or both 

accumulated, since calcification can either follow iron deposition, 

or be primary. Our aim was to investigate the incidence and nature 

of MBM in early spontaneous abortions (ESA) with CV and its 

association with various histopathological pictures. 

Methods. We reviewed all ESAs diagnosed with villitis from 

2004 to 2009; ESAs featuring CV devoid of any inflammatory 

activity were retrieved and subclassified based on clinics and 

morphology and on the finding of MBM on routine histological 

examination. Consecutive sections from each specimen were 

stained with Perls and von Kossa in order to point out MBM, and 

the results were evaluated and compared. 

Results. Of 1095 ESAs, 359 (32.8%) were diagnosed with villitis, 

among which 28 cases (7.8%) of CV; in this group, 18 cases showed 

CV as the only cause of abortion (64.3%), while the remaining 10 

were associated to other significant morphological findings (e.g. 

karyotype abnormalities, autoimmune diseases). Perls confirmed 

MBM in 71.4% of cases and von Kossa in 39.3%, at least one stain 

being positive in 75% and both methods in 35.7%. 

Conclusions. Together with stromal fibrosis, perivillous fibrinoid 

and inflammation, MBM is a reliable marker of CV, seen in 75% 

of cases; anyway, it is very rarely seen on routine histology, 

and this basic histochemistry is mandatory. In MBM, iron is 

twice as frequent as calcium, and the latter is almost never seen 

alone. Hence, MBM could be a marker of previous hemorrhage, 

eventually undergone dystrophic calcification, in cases showing 

long-standing CV. 

ebstein’s anomaly 

1)A. Scivetti, 1)A. Napoli, 1)A. Marzullo, 1)A. Colagrande, 1)R. 

Scamarcio, 1)U. Angelotti, 2)M. Marinaccio, 2)R. Catacchio, 

3)A. Tito, 1)G. Caruso 

1)DAP, Policlinico Bari, Italia; 2)DOG, Policlinico, Bari, Italia; 3)Università 

degli Studi di Bari, Italia 

Background.Complex congenital heart and great vessels diseases 

represent an important field of cardiovascular patology, 

especially in childhood, in which they play an important role 

as the most recurrent and significative malformations. Ebstein’s 

anomaly in particular is a rare malformation (0,5-1%) of the 

tricuspid valve with multifactorial etiology (genetics, reproductive 

or environmental factors). It represents an extreme form of 

dysplastic valve cusps with incomplete differentiation from the 

endocardial wall, whose anatomic spectrum is sufficiently variable 

to cause a different morpholgy of the right ventricle, with 

different hemodynamic consequences. 

Methods. Case 1: fetus in a 33 years old woman with miscarriage 

at 24th weeks of pregnancy, whose only information we received 

relates to clinical coagulopathy. Case 2: fetus in a 39 years old 

woman who performed an abortion ex art. 6 law 194/78 at 22th weeks of pregnancy, because of Ebstein’s anomaly associated 

with pulmonary artery atresia, confirmed by ultrasonography. 

Results. Case 1 macroscopic: the septal and posterior leaflets 

were dysplastic, displaced towards the right ventricle and


stuck to its wall, they were partially welded at the commissure 

both among themselves and with the anterior one (Ebstein’s 

anomaly), there was also a huge dilatation and abnormal wall 

thinning of right heart cavities. The right lung appeared diffusely 

brownish because of marked edema and congestive 

phenomena histologically identified. Case 2 macroscopic: it 

showed Ebstein’s anomaly with lower insertion of posterior 

and septal leaflets of the tricuspid valve, right ventricular 

hypoplasia, marked dilatation of the right atrium with patent 

foramen ovale and hypoplasia of the pulmonary artery. 

Normal the left ventricle and aorta in its emergency, intact 

interventricular septum. 

Complex congenital heart disease 

1)A. Scivetti, 1)A. Napoli, 2)G. Napoli, 1)A. Marzullo, 3)N. 

Blasi, 3)R. Catacchio, 1)A. Colagrande, 1)R. Scamarcio, 1)U. 

Angelotti, 4)A. Tito, 1)G. Caruso 

1)DAP, Policlinico, Bari, Italia; 2)Anatomia patologica Ospedale San 

Paolo, Bari, Italia; 3)DOG, Policlinico, Bari, Italia; 4)Università degli 

Studi di Bari 

Background. Complex Congenital Heart Disease show structural 

abnormalities of the heart and great vessels present since the birth 

with obvious or potential functional significance. They have an 

incidence among children of 8 0 / 00. Isolated or associated with 

other defects, it’s really difficult to establish their aetiology and 

pathogenesis. 

Methods. We observed three fetuses in 26, 36 and 36 years old 

women, who underwent to abortion ex art. 6 law 194/78 at 21st, 

23rd,18th weeks of pregnancy, because of the ultrasound reports. 

Case 1: single atrioventricular canal with a large ventricular 

septal defect (VSD), tricuspid atresia, pulmonary artery does 

not appear. Case 2: transposition of great arteries with VSD and 

pulmonary atresia. Case 3: hypertrophic aortic valve, large VSD, 

hypoplastic mitral valve, prevalence of right heart wall, reported 

trisomia 18. 

Results. Case 1 macroscopic: dextrocardia, atrial situs solitus, interatrial 

septal defect, atrioventricular (AV) and ventriculoarterial 

(VA) discordance (corrected transposition), atresic mitral valve 

placed on the right, double access of the tricuspid valve placed 

on the left in both ventricles corresponding to a riding a large 

ventricular septal defect, aorta lies ahead and to the right pulmonary 

artery, pulmonary atresia, mirror aortic arch, left descending 

aorta, persistent left superior vena cava draining into left atrium. 

Case 2 macroscopic: dysmorphic and quadrangular heart, with 

tip curved to the left, large VSD, right ventricular hypertrophy, 

anterior aorta, pulmonary atresia, patent ductus arteriosus. Case 3 

macroscopic: large ventricular septal defect, atresic mitral valve 

and patent ductus arteriosus; cystic adenomatoid malformation of 

the left lung; small intestine in the right upper quadrant and colon 

in the left iliac fossa. 

Primary non-Hodgkin lymphoma of ureter, 

large b-cell type 

1) D. Di Clemente, 1) A. Napoli, 2) G. Salerno, 1) A. Cimmino, 

1) M.G. Fiore, 1) M. Palumbo, 1) G. Fiore, 1) G. Arborea, 1) R. 

Ricco 

1) DAP, Policlinico, Bari, Italia; 2) Divisione di Urologia, Policlinico, 

Bari, Italia 

Background. Non-Hodgkin Lymphoma (NHL) is typical of 50- 

60 year-old people and prevail among men more than women 

(ratio 2:1). Primary NHL of ureter is rare and only 15 cases has 

been reported in literature. Infact there isn’t lymphoid tissue in 

urinary tract. The most frequent subtype of NHL of ureter is large 

B-cell type(45%), followed by Burkitt’s, follicular, marginal zone 

lymphoma (15% each one), T-cell lymphoma (10%). 

325 

Methods. We report the case of a 72 year-old man who had his 

gallbladder surgically removed (62 year-old), TIA (64 year-old) 

and LUTS about 4-5 years ago. He was admitted to our hospital 

with macroscopic hematuria as the main symptom. Abdominopelvic 

CT showed right-side dilatation of renal pelvis, proximal 

hydroureter and thickening of its wall, with omolateral lymph 

node enlargement. A lumbar lymphadenectomy and partial right 

ureterectomy was performed. The intraoperative examination of 

a lymph node showed white areas at the macroscopic examination 

and a poorly differentiated cell malignant lymphoma, largetype, 

at the microscopic examination. Histology of others lymph 

nodes and ureter’s tract showed “poorly differentiated malignant 

lymphoma of ureter localization, infiltrating the wall”. The immunoistochemical 

analysis showed spread CD20 positivity in 

large cells, CD30 positivity, CD3 and CD45Ro positivity in 

reactive T cells, CD68 (PG-M1) positivity in histiocytes, CD15, 

CD21, CK-pool negativity, Ki67 positive (95%) in large cells. 

According to this immunoistochemical analysis we made diagnosis 

of “NHL, large B-cell type, with a T-cell and histiocytes 

population. 

Results. NHL of ureter is a rare disease and it has to be included 

in the differential diagnosis of all cases of uncertain origin ureter’s 

stenosis. The final diagnosis requires immunoistochemical 

analysis in order to identify the correct subtypes of lymphoma 

and to plan the appropriate treatment. 

Bladder endometriosis (case report) 

1)G. Arborea 1)Napoli A. 2)Salerno G. 3)D’Eredità G. 4)Giardina 

C. 5)Di Clemente D. 6)Palumbo M. 7)Fiore G. 8)Maiorano E. 

1)Dap, Policlinico, Bari, Italy 2)Urologia, Policlinico, Bari, Italy 3)Chirugia 

G. Marinaccio, Policlinico, Bari, Italy 4)Dap, Policlinico, Bari, 

Italy 5)Dap, Policlinico, Bari, Italy 6)Dap, Policlinico, Bari, Italy 7)Dap, 

Policlinico, Bari, Italy 8)Dap, Policlinico, Bari, Italy 

Background. Endometriosis is a common disorder which affects 

up to 10-20% women of reproductive age, involving bladder and 

urinary tract only in 0.1-0.4% cases. The most frequent clinical 

presentations of bladder endometriosis include urinary urgency 

and frequent micturition, pelvic pain, microscopic and macroscopic 

haematuria. Diagnosis of bladder endometriosis is based 

on cystoscopic suspicion and requires biopsy with histological 

confirmation. 

Methods. We report the case of a 33-year-old woman who 

showed menstrual mictalgia for the last four years. Pain persisted 

for all menstrual period and the ten days after. The patient 

underwent to gynecological exam, cistoscopy, ultrasonography 

and CT which showed bladder-uterus expansive lesion. Hence 

in September 2009 she was admitted to hospital with suspicion 

of bladder-uterus endometriosis. The hystero-cistoscopy showed 

oedematous mucosa and a 2 cm nodule on the bladder trigone. A 

biopsy was performed. Histology showed Von-Brunn nests and 

glandular metaplasia. This diagnosis was confirmed by immunohistochemical 

analysis: CD10 was negative. Because of a persisting 

pain the patient got to a new hospitalization in December 

2009. A new cystoscopy showed presence of “chocolate cysts”. 

The cysts was removed by TURB and a biopsy was performed. 

Histology showed endometrial mucosa and histiocytes containing 

haemosiderin pigment in the bladder wall. Therefore endometriosis 

was diagnosed. The symptoms got better but didn’t disappear 

and this required a new hospitalization in April 2010. The last 

cistoscopy the patient performed showed trigone’s scarring areas 

caused by previous TURB, lifted mucosa and some 2-4 mm 

nodules containing gelatinous material similar to chocolate. 

Histological examination confirmed again diagnosis of bladder 

endometriosis. 

Results. Bladder endometriosis is a pelvic pain syndrome that 

causes urinary problems which may get better but not completely 

regress with endoscopic surgery.

326 

expression of HMlH1 and HMSH2 in the prognosis 

of papillary urothelial carcinoma 

1)D. Di Clemente, 1)A. Napoli, 2)G. Salerno, 3)A. Stella, 

1)M. Palumbo, 1)G. Fiore, 1)G. Arborea, 1)C. Caporusso, 4)G. 

D’Eredità1)E. Maiorano 

1)DAP, Policlinico, Bari, Italia; 2)Divisione di Urologia, Policlinico, 

Bari, Italia; 3)Genetica, Policlinico, Bari, Italia; 4)Chirurgia “ G. Marinaccio”, 

Policlinico, Bari, Italia 

Background. Transitional cell carcinoma is a papillary tumor 

spread, which favors the male, aged between 30 and 60 years. 

Predisposing factors for the onset of this cancer are: cigarette 

smoking, alcohol, exposure to arylamine and genetic predisposition. 

This seems to be correlated with the loss of heterozygosity 

on chromosome 9, or association of nonsense mutations in the 

gene for the protein hMSH6 and p53 gene, or the failure to protein 

expression, such as hMLH1, hMSH2, hMSH6 or instability 

of micro-satellites MSI EMAST. 

Methods. Two cases of papillary urothelial carcinoma of the 

bladder, respectively belonging to father and son, aged 67 and 

37 years, came to our attention. The son, in 2008, is hospitalized 

for renal colic. Cystoscopy showed minute papillary lesion 

in the trigone, removed during trans-urethral endoscopic intervention. 

Histologic diagnosis of this lesion was: “high-grade 

urothelial carcinoma of bladder malignancy, focally infiltrating 

the tunica propria (T1-G3) and minute fragments of muscularis 

free of tumor”. The father, in 2009, is admitted to the same 

hospital for haematuria, underwent cystoscopy, which revealed 

the presence of neoformation vegetans, apparently not over 

the trigone removed with invasive endoscopic transurethral 

resection. We examinated minutes and multiple irregular fragments, 

brownish-gray and histologic diagnosis was “papillary 

urothelial bladder carcinoma with spindle cell areas, low-grade 

malignancy with focal areas of senior issues and ulceration (T1 

G2-3)”. 

Results. We wanted to study the two cases with MLH1 and 

MSH2 proteins and genetic investigation of chromosome 9. The 

results of our study suggest that failure to protein expression of 

hMLH1 and hMSH 2 can be used as markers for urothelial malignancy 

and loss of heterozigosyty explains the importance of the 

genetic etiology to this disease. 

Malacoplachia uterine 

1)R. Scamarcio, 1)A. Napoli, 1)A. Scivetti, 1)A. Colagrande, 

1)U. Angelotti, 2) G.Napoli, 1)L. Resta 

1)DAP, Policlinico, Bari, Italia; 2)Anatomia patologica Ospedale San 

Paolo, Bari, Italia 

Background. The malakoplakia is a xanthogranulomatous chronic 

inflammatory that most commonly affects the urinary and 

gastrointestinal tract of middle-aged women. 

Rarely affects the female genital tract. It begins with brown 

plaques, sometimes ulcerated, consisting of infiltrates of lymphocytes 

and macrophages with inclusions rich in calcio (Michaelis- 

Gutmann). 

Methods. Case report: 76 years old woman with repente occurrences 

of metrorragia. 

Endometrial cytology washings: presence of numerous 

histiocytic cells with large granular cytoplasm. At hysteroscopy: 

atrophic endometrium on the anterior wall. 

Macroscopic findings: very minute fragment. 

Diagnosis: small fragments of endometrial mucosa with glandular 

atrophy and massive infiltration of epiteliomorfi histiocytes 

with PAS-positive broad cytoplasm. 

Himmunohistochemically: positive reaction for CD68. 

IIC:positive reaction for CD68.The morphological aspects suggest 

uterine malakoplakia. 


Results. The malakoplakia should be considered a disease characterized 

by impaired patient’s response to infection, for inability 

to complete elimination of swallowed material. 

Patients present usually irritative symptoms with a history of 

recurrent infections, treated with various antibiotics, without 

significative results. 

It ‘was noted an association between impaired immune defense 

of the patient caused by systemic diseases and the development 

of malakoplakia. 

Intestinal endometriosis case report 

1)Colagrande A. 1)Napoli A. 2)Scivetti A. 3)Scamarcio R. 4)Angelotti 

U. 5)Napoli G. 6)Russo S. 7)Resta L. 8)Ricco R. 

1)Dap, Policlinico, Bari, Italy 2)Dap, Policlinico, Bari, Italy 3)Dap, Policlinico, 

Bari, Italy 4)Dap, Policlino, Bari, Italy 5)Ap, Ospedalòe 

san paolo, Bari, Italy 6)Dap, Policlinico, Bari, Italy 7)Dap, Policlinico, 

Bari, Italy 8)Dap, Policlinico, Bari, Italy 

Background. Endometriosis is defined as the presence of endometrial 

mucosa outside the uterine corpus. It is a common benign 

disease among women of reproductive age.Various structures 

may be involved by this process, as ovaries, tubes, uterine ligaments, 

portions of the intestine such as the small bowel, appendix, 

cecum, sigmoid, and rectum, the peritoneum, the rectovaginal 

septum, vagina, vulva, cervix, inguinal canals, umbilicus and 

abdominal wall. Each month, under the effects of hormones of 

the menstrual cycle, the implanted in the endometrium undergoes 

abnormal bleeding, with irritation of surrounding tissue,and subsequent 

scar tissue and adhesions. 

Methods. 32 years old woman after repeated occurrences of 

proctorragia performes colonoscopy which shows in the rectum 

a large polypoid lesion. A histological diagnosis of tubular adenoma 

was made. Following surgery with resection of segment 

of large intestine (rectum) of the lenght of 10 cm and ileocecal 

segment lenght 16 cm. Macroscopically clusters of multiple 

erythematous polyps are observed in the two segments. Microscopic 

findings: endometriosis of the rectum and small intestine, 

extending from sierosa to the mucosa with polypoid aspects and 

muscular hyperplasia; foci of endometriosis are also present in 5 

perivisceral lymphnodes. 

Results. Endometriosis affects the intestinal tract in 15% to 37% of 

patients with pelvic endometriosis. However, the correct diagnosis 

is often delayed because the lesion may masquerade clinically as 

regional enteritis, appendicitis, ischemic enteritis or colitis, diverticulitis, 

acute self-limited colitis or a neoplasm. The desease may 

also cause a diagnostic confusion for the surgical pathologist when 

the endoscopically obtained sampling is superficial while endometriosis 

usually involves the deeper layers of the bowel wall. 

How to make a diagnosis of Burkitt’s lymphoma 

with a limited panel of antibodies? 

results from a review of 252 cases 

1)Naresh K.N. 2)Lazzi S. 3)Bellan C. 4)Onorati M. 5)Ambrosio 

M.R. 6)Rocca B.J. 7)Malagnino V. 8)Ginanneschi C. 9)Raphael 

M. 10)Leoncini L. 

1)Histopathology, Imperial College, London, United Kingdom 2)Human 

Pathology And Oncology-Ant. Pathol.Section, Santa Maria Delle Scotte, 

Siena, Italy 3)Human Pathology And Oncology-Anat. Pathol. Section, 

Santa Maria Delle Scotte, Siena, Italy 4)Human Pathology And Oncology-Anat. 

Pathol. Section, Santa Maria Delle Scotte, Siena, Italy 5)Human 

Pathology And Oncology-Anat. Pathol. Section, Santa Maria Delle Scotte, 

Siena, Italy 6)Human Pathology And Oncology-Anat. Pathol. Section, 

Santa Maria Delle Scotte, Siena, Italy 7)Human Pathology And Oncology-Anat. 

Pathol. Section, Santa Maria Delle Scotte, Siena, Italy 8)Human 

Pathology And Oncology-Anat. Pathol. Section, Santa Maria Delle Scotte, 

Siena, Italy 9)Service D’Hematologie, University Paris South, Paris, 

France 10)Human Pathology And Oncology-Anat. Pathol. Section, Santa 

Maria Delle Scotte, Siena, Italy


Background. Burkitt lymphoma (BL) is one of the most studied 

human malignancies. Improvements in therapeutic options for 

adult aggressive B-cell lymphomas make distinction of BL from 

diffuse large B-cell lymphoma (DLBCL) and other lymphomas 

extremely critical. Distinguishing BL from B cell lymphoma, 

unclassifiable with features intermediate between DLBCL and 

BL (DLBCL/BL), and DLBCL poses diagnostic problems. To 

address this issue, we propose an immunohistochemistry and 

FISH based scoring system. 

Methods. We evaluated 251 aggressive B-cell lymphomas that 

included 121 and 103 DLBCL samples from Europe and from 

sub-Saharan Africa. The score system was employed in three 

phases. Phase 1 evaluated morphology (typical -3, consistent but 

not diagnostic -2, overlapping features between BL/ DLBCL -1, 

DLBCL -0), CD10 (positive -1, negative -0) and BCL2 (absent 

-2, weak -1, moderate to strong -0). Phase 2 considered Ki-67 

(> 95% -2, 90-95% -1, others -0), CD38 (positive -1, negative -0) 

and CD44 (negative -1, positive -0). Phase 3 used FISH (positive 

for MYC-IGH translocation and negative for BCL2 & BCL6 

translocations -2, others -0). Cumulative score at phase 3 ≥ 8 is 

consistent with diagnosis of BL whereas score 6-7 suggests that 

BL may be not excluded; in the latter case, the diagnostic impact 

of each of the different parameters need to be assessed and complete 

karyotype and array CGH would be useful. 

Results. Using the scoring based algorithm, we were able to arrive 

at a specific diagnosis of BL in 82%, 93% and 97% cases at 

phases 1, 2 and 3 respectively. This approach also led to specific 

diagnosis of DLBCL or DLBCL/BL in 84% of cases that were 

not diagnosed as BL. 

Conclusions. Our diagnostic algorithm/scoring would immensely 

improve the diagnostic ability of pathologists and would also 

improve the usage of the WHO classification of lymphomas 

across the world. 

Papillary serous tumor of low malignant potential 

(P.S.T.l.M.P.) Paratesticular: a case report 

1)Nenna r. MD. 2)Inchingolo c. d. MD. 3)Albino g. MD. 4)Cirillo 

m. e. MD. 

1)Anatomia Patologica, L. Bonomo, Andria, Italy 2)Anatomia Patologica, 

L. Bonomo, Andria, Italy 3)Urologia, L. Bonomo, Andria, Italy 4)Urologia, 

L. Bonomo, Andria, Italy 

Background. Papillary Serous Tumour of Low Malignant Potential 

(P.S.T.L.M.P.) of the paratesticular structures is morfologically 

and immunophenotypically identical to ovarian serous 

borderline tumor. Its histogenesis is under discussion. 

Since this tumour is similar to that seen in the female genital 

tract and especially in the ovary, this tumour belongs to the group 

of Mullerian Epithelial Tumour or Ovarian-Type Epithelial Tumours 

(O.T.E.T.). 

To date, no serous borderline tumour of paratestis reported in 

literature has recurred or metastasized after complete resection or 

a radical orchiectomy. 

Methods. A 64-year-old man whit a thirty-year history of cronic 

left hydrocele, progressively increasing, presented himself to the 

urologist complaining of discomfort for the inguinal zone. 

Physical examination revealed left hydrocele transilluminabile 

without tension. 

Scrotal ultrasound showed “hydrocele corpusculated with hypertrophy 

of the vaginal tunic in the head of epididymis of perhaps 

inflammatory reactive nature, worthy of surgical exploration during 

surgery for hydrocele”. 

Preoperative blood levels of tumor markers (CA-125; BetaHCG; 

Alfa-fetoprotein) were not available. 

After five months, the patient was operated for resection and 

eversion of the left vaginal tunic. After the aspiration of a citrine 

yellow liquid, a papillary mass whit a size of 4 × 2 cm was found 

in the vaginal opening at the paratesticular site, between the up- 

327 

per pole of the testis and the head of epididymis. During intraoperative 

examination this tumour was described as “Papillary 

Serous Epitelial Tumour Borderline”, while in the final histologic 

report as “Papillary Serous Tumor of Low Malignant Potential 

(PSTLMP) Mullerian-type”. 

After obtaining the informed consent of patient, radical orchiectomy 

was done. 

Results. Grossly, the tumor appears as a greyish papillary solid 

mass,whit the size of 4 × 2 cm and no necrotic areas. 

Microscopic sections revealed well-formed papillae with a fibrovascular 

core lined by serous cuboidal or columnar epithelial 

cells, often in many layers including apical cilia. The epithelium 

was bland, mitotic figures were present, but rare, no microinvasion 

and no frank nuclear anaplasia were identified. Psammoma 

bodies were not observed. The tumour wasn’ t associated with 

ITGCN or with teratomatous elements of testis. 

Epithelial cells displayed immunoreactivity identical to borderline 

papillary serous tumors of the ovary: strong and diffuse 

positive staining with a broad-spectrum Cytokeratins 

AE1/AE3, Cytokeratin 7, EMA, CA125, WT1, Estrogen 

Receptor/1D5, Progesteron Receptor/PgR636 and Vimentin; 

negative staining with CEA, Cytocheratin 20, Cytokeratins 5/6, 

Calretinin, CD15, and PLAP. Proliferative activity by MIB1 

staining was 5%. 

Conclusions. Papillary serous tumours of low malignant potential 

(P.S.T.L.M.P.) may occur in the tunica vaginalis, testis, 

spermatic cord and epididymis, and grossly, microscopically and 

immunohistochemically identical to its ovarian counterpart. It is 

usually unilateral. Patients range in age from 6 to 77 years (mean, 

56 years). Proliferative activity by MIB-1 staining ranges from 

1% to 10% (mean, 5,5%). 

The differential diagnosis includes papillary serous carcinoma 

(typically consisting of invasive papillae), papillary cystoadenoma 

of the epididymis (benign neoplasm that arises from the 

efferent duct epithelium; often bilateral and associated with von 

Hippel-Lindau syndrome) and benign and malignant papillary 

mesothelioma (asbestos exposure correlated neoplasm with a 

biphasic histologic pattern and positive staininig for Calretinin 

and CK 5/6). 

A radical orchietomy is the treatment of choice. To date, no serous 

borderline tumour of the paratestis reported in literature has 

recurred or metastasized after complete resection. 

Its histogenesis is under discussion. Early in development, tissues 

have the potential to develop into either male or female structure. 

Bilateral urogenital ridges grow from coelomic epithelium 

around week 5 of development. If no signals occur to trasform the 

structure into testis, the organ develops into an ovary. The same 

coelomic epithelium is responsible for both male and female 

structure; therefore, a tumor affecting this tissue could affect 

either sex. 

Epithelial tumours of testis that resemble ovarian tumours may 

be seen in either testis and paratestis. Testicular disease is less 

common than paratesticular disease, and the etiology is unknow. 

It has been hypothesized that these lesions might development 

from the remnants of the mullerian duct (for example from appendix 

testis, a vestigial remnant of the male mullerian duct) 

or from mesothelial inclusions of the tunica vaginalis by the 

process of mullerian neometaplasia. Intratesticular tumours are 

hypothesized to result from areas of coelomic epithelium thah 

became trapped within the testicular tissue. An additional theory, 

although less popular, is that the tumor develops in the ovarian 

component of a hermaphrodite. 

references 

Carano KS, Soslow RA. Immunophenotypic analisis of ovarian and 

testicular mullerian papillary serous tumours. Modern Pathology 

1997;10(5):414-20. 

Jones M, Young RH, Srigley JR, et al. Paratesticular serous papillary 

carcinoma. A report of six cases. Am J Surg Pathol 1995;19:1359-66.

328 

Kaushik D, Gulamjat SM, Thangjam DS. Papillary cystadenoma of the 

epididymis. Kuwait Medical Journal 2005;37(2):122-4. 

McClure R, Keeney G, Sebo T, et al. Serous borderline tumour 

of the paratestis: a reprt of seven cases. Am. J. Surg. Pathol 

2001;25(3):373-8. 

Sanchez BC, Baez PJM, Beltran AV, et al. Mullerian-type papillary serous 

tumor: exceptional pathology of the testis. Report of a case and 

discussion. Actas Urol. Esp 2000;24(3):256-9. 

Sumrall A, Puneky L, Brown A, et al. Ovarian cancer in a man? Clinical 

Ovarian Cancer 2009;2;57-9. 

van der Putte SCJ, Toonstra J, Sie-Goi D. Mullerian Serous Cystadenoma 

of the strotum following orchiopexy. ADv Urol 2009. 

The cytologic evaluation of pleural fluid 

in the diagnosis of malignant mesothelioma 

Nirchio V., Clemente C.**, Ardò NP°, Castriota M^, Bufo P*., 

Sollitto F.° 

Struttura Semplice Dipartimentale di Citologia Diagnostica, Dipart. di 

Patologia Clinica OO.RR; °Struttura Complessa di Chirurgia Toracica 

Universitaria, OO.RR-Foggia; *Struttura Complessa di Anatomia Patologica 

Universitaria, OO.RR; ^Struttura Complessa di Anatomia Patologica 

Ospedaliera, OO.RR; **Struttura Complessa di Anatomia Patologica, 

IRCCS Casa Sollievo della Sofferenza, S.G.R 

Introduction. Mesothelioma represents 1% of all tumors and 

pleura is the most frequent localization. 

In 90% of cases work or environment expositive risk factors related 

to asbestos are found. 

In this study we report the cases of mesothelioma occured in the 

district of Foggia in the last 10 years and compared cytologic and 

hystopatologic diagnosis. 

Materials and methods. This review is based on the archives of 

the Hystopathology Institute of Foggia (for the period 2000-2009) 

and the Hystopathology Institute of S. Giovanni Rotondo (from 

1992 to 2009). 

From 2000 to 2009 we observed at Servizio di Citologia Diagnostica, 

Azienda Ospedaliero-Universitaria OO.RR of Foggia, 13 (7 

male and 6 female) patients, with clinical history suggestive for 

mesothelioma. 

We performed cytologic examination for all patients, 9 on pleural 

fluid, 3 on ascitis, 1 on needle aspiration specimens. 

Results. The mean age was 64.5 (range 49-80), with median age 

68,9. 

We observed two positive cytologic findings, one on pleural fluid, 

one on the needle aspiration specimen. The other cases were 

negative or showed metastasis from ephitelial tumor. 

During the same period, at our Institution, 21 (15 male, 6 female) 

patients had hystologic finding of mesothelioma, with mean age 

of 62,5 and median age of 68,8. 

In the last 10 years, in the district of Foggia, cytologic diagnosis 

was reached in 27 cases. 

Conclusions. Data analysis suggests that hystopathology is the 

“gold standard” for diagnosis of mesothelioma, however cytology, 

together with clinical remarks, give the first diagnostic guidance, 

necessary to further advanced assessments for the diagnosis 

of this life-threatening disease. 

references 

Renshaw AA, et al. The role of cytologic evaluation of pleural fluid in the 

diagnosis of malignant mesothelioma. Chest 1997;111(1):106-9/ 

Moore AJ, et al. Malignant Mesothelioma. Orphanet J rare Dis 2008; 

Dec 19:3-34. 

Rakha EA, et al. The sensitivity of cytologic evaluation of pleural fluid in 

the diagnosis of malignant mesothelioma. Diagn Cytopathol 2010. 

Pericardial effusion neoplastic metastasis 

to lung adenocarcinoma. Description of a case 

1)Nocita A. 2)Pizzi G. 3)Filardo A. 4)Squillaci S. 5)Tallarigo F. 

1) Anatomia Patologica, Ospedale S. Giovanni di Dio, Crotone, Italia; 2) 


Anatomia Patologica, Ospedale Pugliese-Ciaccio, Catanzaro, Italia; 3) 

Anatomia Patologica, Ospedale Vallecamonica, Esine, Italia 

Introduction. Payments neoplastic of serous cavities are usually 

bleeding and reform over a short time since drainage. They are 

characterized by the presence of many tumor cells that have either 

individually or in three-dimensional aggregates. Payments can be 

divided into primary (mesothelioma) and secondary (metastatic). 

Tumors that most commonly cause payments metastatic cancer 

are lung cancers (35%) than breast (25%), primitive neoplasms 

not known (12%), and finally lymphomas and leukemias (10%). 

From the standpoint of incidence, the pleura is the most affected, 

followed by the peritoneum, while payments are extremely rare 

cancer of the pericardium. 

Case report. A man of 65 years, admitted to emergency at 

the O.U Internal Medicine Hospital of Crotone diagnosed with 

massive effusion and subsequent cardiac tamponade. Subsequently, 

the patient is transferred to the Hospital Pugliese-Ciaccio 

Catanzaro where you ultrasound guided pericardiocentesis, 

whereby some are drained (800 cc) of fluid and blood 

serum, and that is made cytology test that the microbial test. 

Material and Methods. For cytology test were used 40 ml of 

payment. This was evenly distributed in conical tubes and prepared 

after centrifugation and smears of the sediment, other cytological 

preparations stained, then, hematoxylin-eosin. From the 

morphological point of view, on a background blood, it highlights 

many aspects of tubular aggregates sometimes branched neoplastic 

elements consisting of medium and large size with rounded 

nuclei, chromatin fienemente granular cytoplasm and more or 

less abundant. Immunocytochemical investigation is performed 

by testing the following antibody panel: (Cytokeratin AE1/AE3, 

CK 7, TTF-1, CD 56, Chromogranin, P63). 

Results. The neoplastic cells showed positive for TTF1, Cytokeratin 

AE1/AE3 and CK7, but were negative for CD 56, P63 

and Chromogranin. Thus was diagnosed localization, metastatic 

pericardial of non-small cell lung cancer whose morphologic and 

immunophenotypic profile is consistent with adenocarcinoma. 

Discussion and conclusions. The neoplastic involvement of the 

pericardium is observed in 3-4% of autopsies in general and in 2- 

31% in autopsy series of patients cancer. Carcinoma of the lung, 

breast, lymphomas and leukemias are the most common causes 

of malignant pericardial effusion. The diagnosis of malignant 

pericardial effusion can be extremely difficult because the clinical 

manifestations are insidious and may mimic more common 

diseases. When establishing a pericardial effusion onset may be 

gradual or rapid and the symptoms are related to the rate of accumulation 

of liquid. Regarding diagnostic methods, the survey 

represents the fundamental pericardiocentesis with the resulting 

fluid cytology. 

Intraoperative frozen section technique for breast 

cancer: end of an era 

1)Nottegar A. 2)Manfrin E. 3)Remo A. 4)Pollini GP. 5)Falsirollo 

F. 6)Parisi A. 7)Molino A. 8)Dalfior D. 9)Reghellin D. 

10)Bonetti F. 

1)Patologia e Diagnostica, sez. Anatomia patologica, Università di Verona-Policlinico 

GB Rossi, Verona, Italia 2)Patologia e Diagnostica, sez. 

Anatomia patologica, Università di Verona-Policlinico GB Rossi, Verona, 

Italia 3)Istituto di Anatomia patologica, Ospedale Mater salutis, Legnago 

(Verona), Italia 4)Chirurgia, U.O. Chirurgia generale A, Università 

di Verona-Policlinico GB Rossi, Verona, Italia 5)Centro di prevenzione 

senologica, Ospedale di Marzana, Verona, Italia 6)Patologia e Diagnostica, 

sez. Anatomia patologica, Università di Verona-Policlinico GB Rossi, 

Verona, Italia 7)Oncologia medica, Università di Verona-Ospedale civile 

maggiore, Verona, Italia 8)Istituto di Anatomia patologica, Ospedale G. 

Fracastoro, S. Bonifacio (Verona), Italia 9)Istituto di Anatomia patologica, 

Ospedale Cazzavillan, Arzignano (Vicenza), Italia 10)Patologia e 

Diagnostica, sez. Anatomia patologica, Università di Verona-Policlinico 

GB Rossi, Verona, Italia


Background. Intraoperative frozen section (FS) technique has 

played an important role in the diagnosis of breast cancer aiding 

the surgeon to choose the best therapeutic approach on discrete 

palpable lesions, but its use has been discouraged in lesions 

inferior to 1 cm. Over the years, the flow-chart referred to the 

assessment of breast lesions has been progressively shifted from 

intraoperative procedures to pre-surgical diagnostic techniques, 

as fine needle aspiration cytology (FNAC) and automated or 

vacuum-assisted gun needle biopsy. The diffusion of mammography 

has increased the detection of small sized cancers 

(< 1 cm) as well as proliferative low grade atypical lesions for 

which intraoperative FS technique has to be considered not 

mandatory. 

Methods. Data on 2436 breast carcinoma diagnosed between 

1992 and 2006 were collected. The rate of intraoperative procedures 

was calculated in each year and results correlated with 

tumor size and pre-operative diagnostic procedures. 

Results. Over the years, there was a decreasing use of FS. The 

rate of cancers diagnosed with FS was 51.2% (1992), 48.2% 

(1993), 48.5% (1994), 44.0% (1995), 39.1% (1996), 29.0% 

(1997), 25.6% (1998), 22.8% (1999), 12.0% (2000), 1.0% (2001), 

0% (2002), 6.2% (2003), 3.7% (2004), 0% (2005-2006). The 

decreasing use of FS was indistinctly extended to all pT cancer 

categories, whatever the cancer size. In the same period, the 

adoption of cytology and core biopsy increased as pre-surgical 

diagnostic accuracy was. FNAC positive predictive value for a 

malignant diagnosis was 99.3%, the inadequate rate from cancer 

was 2.4% and the false-positive rate was 0.5%. 

After one hundred years from its first adoption, FS technique 

is no more considered on primary breast lesions. In an audited 

diagnostic activity on breast pathology, FS on primary lesion is 

generally inappropriate, particularly in the assessment of clinically 

impalpable lesions. 

An unusual female breast metastasis from urinary 

bladder sarcomatoid carcinoma 

1)Nottegar A. 2)Manfrin E. 3)Remo A. 4)Vasori S. 5)Pollini GP. 

6)Pellini F. 7)Martignoni G. 8)Bonetti F. 

1)Patologia e Diagnostica, sez. Anatomia patologica, Università di Verona-Policlinico 

GB Rossi, Verona, Italia 2)Patologia e Diagnostica, sez. 

Anatomia patologica, Università di Verona-Policlinico GB Rossi, Verona, 

Italia 3)Istituto di Anatomia patologica, Ospedale Mater salutis, Legnago 

(Verona), Italia 4)U.O. di Radiologia, Università di Verona-Policlinico 

GB Rossi, Verona, Italia 5)Chirurgia, Unità Operativa Chirurgia generale 

A, Università di Verona-Policlinico GB Rossi, Verona, Italia 6)Chirurgia, 

Unità Operativa Chirurgia generale A, Università di Verona-Policlinico 

GB Rossi, Verona, Italia 7)Patologia e Diagnostica, sez. Anatomia patologica, 

Università di Verona-Policlinico GB Rossi, Verona, Italia 8)Patologia 

e Diagnostica, sez. Anatomia patologica, Università di Verona-Policlinico 

GB Rossi, Verona, Italia 

Background. Breast metastases are uncommon and account for 

about 2% of female breast malignancies. Most frequently, the primary 

tumor is a haematological disease or a controlateral breast 

carcinoma. Solid metastatic cancers most frequently originate 

from lung, kidney, stomach, intestinal tract, ovary, uterine cervix 

and thyroid gland. Breast metastases due to the systemic diffusion 

of urinary bladder neoplasm have been less frequently reported 

and the described histotype has been transitional cell carcinoma. 

A female breast metastasis from urinary bladder sarcomatoid carcinoma 

(SC), diagnosed with FNAC and CNB, is described. 

Material and methods. A 66-years-old female presented with 

macroscopic haematuria. Abdominal ultrasound (US) showed a 

thickened area of the bladder wall suspicious for bladder neoplasm. 

Cystoscopic examination revealed a broad-based tumour 

of 5.5 cm in diameter located in the anterior and right bladder 

wall. The histological examination of lesion biopsies showed a 

mixed epithelioid and spindle cells neoplasia (CK7+; CK5/6 +; 

CK8/18/19+; Vimentin-/+; Desmin-;CD34-) with invasion of the 

329 

muscle wall associated with in situ transitional cell carcinoma. 

The diagnosis of urinary bladder SC was confirmed by cystectomy. 

Six months later, an asymptomatic 15mm sized focal lesion 

was detected in the left breast on mammograms. The lesion 

was assessed with fine needle aspiration cytology and a highly 

atypical spindle cells population was on smears. The immunohistochemical 

typing of the lesion on core needle biopsy favored the 

diagnosis of SC metastasis. 

Results. SC of the urinary bladder is a rare neoplasm as breast 

metastases are. The clinical history and immunohistochemistry 

are useful to pose a correct differential diagnosis with primitive 

breast spindle cells neoplasm (pure spindle cell carcinoma, fibromatosis, 

phylloides tumour, melanoma and primitive sarcomas). 

Gastrointestinal metastasis from breast lobular 

cancer: a hurdle run 

1)C. Mignogna, 2)L. Nugnes, 1)A. Giambalvo, 1)D. Ientile 

1)Anatomia Patologica, Ospedale Buccheri la Ferla “Fatebenefratelli”, 

Palermo, Italia; 2)Dipartimento di Scienze biomorfologiche e funzionali, 

Università degli studi di Napoli “Federico II”, Napoli, Italia 

Extrahepatic gastrointestinal localization from breast cancer is an 

uncommon event, often occurring after a prolonged disease-free 

interval and long time after the primitive tumor. Therefore, the 

remote neoplastic history is often underestimated. Symptoms are 

frequently non specific, so endoscopic biopsies and /or frozen 

sections with a misleading suspect of inflammatory disease are 

usually the first specimens received by the pathologist. 

Given the poor prognosis of this particular localization, most of 

the literature reports autoptical series. Infiltrating lobular carcinoma 

is the most common histotype observed; gastric lesions 

seems to be more frequent than colo-rectal ones. 

We herein describe two cases of lobular beast cancer metastasing 

gastrointestinal tract: a colonic one presenting in emergency with 

abdominal pain 24 years after the first diagnosis and a gastric one 

thee years later. 

A rare association: gastrointestinal stromal tumor 

of the stomach and malignant mixed Müllerian 

tumor of the uterus 

1) Onorati M. 2) Ambrosio M.R. 3)Rocca B.J. 4) Mourmouras 

V. 5) Mastrogiulio M.G. 6) Vindigni C. 7)Carducci A. 8) Santopietro 

R. 

1)Department of Human Pathology and Oncology-Section of Anatomic 

Pathology, University of Siena, Italy 2)Department of Human Pathology 

and Oncology-Section of Anatomic Pathology, University of Siena, Italy 

3)Department of Human Pathology and Oncology-Section of Anatomic 

Pathology, University of Siena, Italy 4) Department of Human Pathology 

and Oncology-Section of Anatomic Pathology, University of Siena, 

Italy 5) Department of Human Pathology and Oncology-Section of 

Anatomic Pathology, University of Siena, Italy 6)Department of Human 

Pathology and Oncology-Section of Anatomic Pathology, University of 

Siena, Italy 7) Department of Human Pathology and Oncology-Section 

of Anatomic Pathology, University of Siena, Italy 8) Department of Human 

Pathology and Oncology-Section of Anatomic Pathology, University 

of Siena, Italy 

Background. Malignant gastrointestinal stromal tumors (GIST) 

are rare mesenchymal tumors which originate from the wall of the 

gastrointestinal tract. Most of them are sporadic in nature, affecting 

individuals in their 5 th or 6 th decade of life. The coexistence 

with other tumors is uncommon. We observed an association 

between gastric GIST and malignant mixed müllerian tumor 

(MMMT) of the uterus. 

Methods. A 68-year old female who presented with an epigastric 

mass, associated with uterine bleeding, underwent gastrectomy 

and hysterectomy. Morphological and immunohistochemical 

studies, as well as mutation analysis for c-kit gene on gastric lesion 

were carried out.

330 

Results. The gastric lesion (7 cm in greater dimension) was located 

in the antrum and infiltrated the serosa; it appeared whitish, 

elastic and containing areas of hemorrhage. Within the uterine 

cavity a large polypoid mass with fleshy cut surface was observed. 

Microscopically, the gastric lesion consisted prevalently of atypical 

epithelioid cells, intermingled with a spindle cell component, 

mitoses were numerous (13/50 HPF) and Mib-1 was high (40%); 

the tumor cells showed strong positivity for CD117 and CD34. The 

uterine lesion showed two components, sharply demarcated and 

histological malignant. The epithelial one was represented by an 

high grade endometrioid adenocarcinoma (CK AE1/AE3 positive), 

the stromal component was heterologous and vimentine positive. 

The diagnosis was high risk gastric GIST (according to Fletcher) 

co-existing with a MMMT of the uterus (pTIcN0Mx, Ic FIGO). 

Mutation analysis for c-kit gene showed a mutation in exon 11. 

Conclusion. GISTs and MMMTs are two very rare tumours 

and their simultaneity in patients with no cancer predisposition 

syndrome, is uncommon. To the best of our knowledge this is the 

first case of a gastric GIST associated to an uterine MMMT. 

role of plakoglobin immunohistochemistry 

in diagnostic evaluation of juvenile sudden 

cardiac death 

1)Orlandi M. 2)Pisano A. 3)Zachara E. 4)Di gioia CRT. 5)Gallo 

P. 6)D’Amati G. 

1)Medicina Sperimentale, Policlinico Umberto I, Roma, Italia 2)Medicina 

Sperimentale, Nico Umberto I, Roma, Italia 3)Medicina Speri, Policlinico 

Umberto I, Roma, Italia 4)Medicina Sperimentale, Policlinico Um, Roma, 

Italia 5)Medicina Sperimentale, Policlinico Um, Roma, Italia 6)Medicina 

Sperimentale, Policlinico Um, Roma, Italia 

Background. Juvenile sudden cardiac death (SCD) can be due 

to a variety of acquired and inherited conditions, and is often the 

first manifestation of a hidden genetic disease. Arrhythmogenic 

right ventricular cardiomyopathy (ARVC) due to mutations in 

desmosomal proteins is one of the most frequent causes of SCD. 

Autopsy diagnosis of ARVC is based on the findings of myocardial 

atrophy and fatty/fibro-fatty replacement. However, cases 

with mild and segmental fibro-fatty replacement still represent 

a diagnostic grey zone between desmosomal-related ARVC and 

non-specific myocardial changes. Immunohistochemical (IH) 

detection of plakoglobin (PKG), a protein of intercalated disks, 

has been recently proposed as a diagnostic tool for histologic 

diagnosis of ARVC. We studied the usefulness of this method 

to rule out ARVC in cases of juvenile SCD with morphologic 

features suggestive but not conclusive for the disease. 

Methods. We selected 4 cases with autopsy features suggestive 

of ARVC in which a clinical family screening, along with 

a molecular autopsy of the proband had allowed a post-mortem 

diagnosis of channelopathy. 

As positive controls, we used 3 explanted hearts with clinically 

and genetically proven ARVC (Table 1). IH was performed on 

paraffin slides from both ventricles, with antibodies to PKG and 

N-Cadherin as internal control, using immunoperoxidase with 

conventional labeled polymer technology, with a 1:50.000 antibody 

dilution. 

Results. Plakoglobin was intensely expressed at myocyte intercalated 

disks, both in the right and left ventricles, in all cases of 

channelopathies with morphologic changes suggestive of ARVC. 

In contrast, it was markedly reduced or absent in explanted ARVC 

hearts, confirming the clinical and morphologic findings. According 

to our preliminary results, in cases of SCD with ambiguous 

morphologic features, diffuse positive stain of intercalated disks 

is useful to rule out the diagnosis of desmosomal-related ARVC. 

PKG immunohistochemistry can be an additional tool for autopsy 

diagnosis, that is crucial to guide the genetic screening of SCD, 

expecially in absence of a significant clinical history or previous 

instrumental findings. 


Quality assurance in Veneto breast cancer 

screening program: histopathology on virtual slides 

1)E. Orvieto, 2)E. Bianchini, 2)G. Briani, 2)A. Caneva, 2)G. 

Capitanio, 2)D. Della Libera, 2)S. Dante, 2)R. Di Pietro, 2)P. 

Gasparini, 2)I. Pavon, 2)L. Laurino, 2)G. Leo, 2)M. Lo Mele, 

2)S. Paolino, 2)Q. Piubello, 2)F. Sonego, 3)S. Guzzinati, 4)A. 

Rizzo 

1)U.O. di Anatomia Patologica, Azienda Ospedaliera di Padova, Padova, 

Italia; 2) Gruppo Regionale Veneto dei Patologi dello Screening Mammografico; 

3)Registro tumori del Veneto, Istituto Oncologico Veneto, Padova, 

Italia; 4)U.O. Anatomia e Istologia Patologica, Ulss 8 - Ospedale S. Giacomo, 

Castelfranco Veneto, Italia 

Background. The workgroup, purposed itself to test the reproducibility 

regarding diagnosis of invasive breast cancer by virtual 

slides. 

Methods. The original slides were sent from 15 Institutes which 

are part of the workgroup of the screening of breast and were 

scanned using Aperio Scan Scope. In the first round, it was tested 

the reproducibility regarding Histotype & Grading (using E&E 

criteria) in 32 cases. In the second round it was tested the reproducibility 

about Grading, evaluating the single scores (tubules 

formation, nuclear atypia, mitoses). 

Results. In the first round 399 diagnoses were made. Analysis 

of the result evidenced a reproducibility regarding histotype of 

more than 90% of the participants in 13 cases, of 75-90% in 10 

cases, of < 75% in 9 cases. The overall k of concordance was 

0.46. Certain lack of concordance was seen in diagnoses of ductal 

vs tubular, and ductal vs lobular vs mixed histotypes. K of concordance 

of ductal, tubular and lobular were respectively 0.43; 

0.77; 0.44. In regard to grading, the overall k of concordance 

was 0.51, with better reproducibility in valuating G3 (0.62) and 

G1 (0.57) and worse in G2. In the second round it was asked to 

the participants to diagnose, separately, the single scores (tubules 

formation, nuclear atypia, mitoses). To count mitoses, areas corresponding 

to 10 HPF were identified on the virtual slide. The k 

of concordance showed lack of reproducibility for mitoses (0.27) 

rather than tubules formation (0.46) and nuclear atypia (0.35); the 

overall k regarding grading was 0.35, while coefficient of Kendall 

was 0.71. These results encourage us to pursue this workgroup, 

able to involve the participants in screening, with a very good 

cost/benefit ratio. 

Veneto Breast Cancer Screening Group: E Bianchini, G Briani, 

A Caneva, G Capitanio, D Della Libera, S Dante, R Di Pietro, P 

Gasparini, I. Pavon, L Laurino, G Leo, M Lo Mele, S Paolino, Q 

Piubello, F Sonego. 

Mutant-enriched PCr and reverse hybriditation 

assay in KrAS testing of colorectal cancer 

1)Pagano L. 2)Ammirabile M. 3)Malapelle U. 4)Della Ragione 

C. 5)Mitilini N. 6)Troncone G. 7)Nappi O. 

1)Dipartimento medicina di laboratorio e anatomia patologica, Aorn 

Cardarelli, Napoli, Italia 2)Dipartimento medicina di laboratorio e 

anatomia patologica, Aorn Cardarelli, Napoli, Italia 3)Dipartmento di 

Scienze Biomorfologiche e funzionali, Università Federico II, Napoli, 

Italia 4)Dipartimento medicina di laboratorio e anatomia patologica, 

Aorn Cardarelli, Napoli, Italia 5)Dipartimento medicina di laboratorio 

e anatomia patologica, Aorn Cardarelli, Napoli, Italia 6)Dipartmento di 

Scienze Biomorfologiche e Funzionali, Università Federico II, Napoli, Italia 

7)Dipartimento medicina di laboratorio e anatomia patologica, Aorn 

Cardarelli, Napoli, Italia 

Background. Epidermal growth factor receptor (EGFR) is currently 

a major target in cancer therapy. In the last few years it has 

become apparent that it very hard to predict response to a given 

EGFR drug basing only on the receptor IHC expression. Thus, 

currently, there is a focus on gene mutation assays to predict 

response to EGFR antagonists. In particular, recent evidences


showed that metastatic colorectal cancer (CRC) patients with 

tumors harboring a KRAS gene mutation do not derive benefit 

from the administration of EGFR-directed monoclonal antibodies. 

Recently, a sensitive nonquantitative novel assay for the 

detection of KRAS mutations in FFPE tissue combining mutantenriched 

PCR and reverse hybriditation (KRAS-strip assay) has 

been proposed 1 . 

Methods. Here the clinical performance of this novel test is 

evaluated on 48 CRC paraffin embedded and compared to the 

direct sequencing one. 

Results. The frequency of mutations was 40% (n = 19) for KRAS: 

G12D 37% (7), G13D 25%(5), G12V 18%(4), G12C 12%(2), 

G12A 6% (1). Results were confirmed by direct sequencing. 

Morever, two cases showing mutations by the KRAS-strip were 

confirmed by direct sequencing only when this latter method was 

performed by microdissecting a wider area of neoplastic tissue. 

In conclusion, the mutant-enriched PCR and reverse hybriditation 

has highly clinical accuracy. 

references 

1 Ausch et al. Journal of Molecular Diagnostics 2009. 

Bilatera ureteral chronic schistosomiasis with 

direct renal infection: case report of rare direct 

renal infestation 

Palumbieri G. 

U.O.C. Istologia patologica e Citodiagnostica, Ospedale “Mons. R. Dimiccoli”, 

Barletta, Italia 

Background. Schistosomiasis, also know as bilharzias, comprises 

a group of parasitic infections caused by waterborne trematode 

worms. It is one of the most prevalent parasitic infections in the 

world. Schistosoma haematobium, endemic in sub-Saharan Africa 

and in the Middle East (in Italy it is rather unusual), mainly 

affects the urinary system, where it leads to hematuria, chronic 

cystitis and chronic obstructive uropathy with bacterial pyelonephritis 

or immune-complex-mediated glomerulonephritis; on 

the contrary the direct renal infestation by S. haematoobium is 

very rare. 

Clinico-pathologic case. A 25-year-old African (Nigeria) woman 

with hematuria, renal colic and with a history of exposure to the 

infections; a chronic obstructive uropathy is demonstrated on 

urogram, showing constrictions with dilatation of ureters, renal 

pelvices and caliceal system. 

Methods. The specimens (left kidney and bilateral ureteral 

stump) were fixed in 10% neutral buffered formalin and embedded 

in paraffin. Sections were processed for routine staining, 

including hematoxylin and eosin, Pas, von Kossa. For immunoistochemistry, 

the avidin-biotin peroxidase complex method 

was used; antibodies employed are CD68 (cl. KP1), CD45 and 

pankeratin. 

Results. Hydroureter and hydronephrosis of the chronic obstructive 

uropathy of schistosomiasis with aspecific chronic pyelonephritis 

is demonstrated, with site of ureteric stricture and with the 

dilatation of the ureters and of the pelvi-calyceal system, with 

compression of the papillae and parenchymal thinning (only a 

thin rim of parenchyma is present). Multiple calcified eggs of 

S. haematobium with their characteristic terminal spine are found 

in all layers of the ureter and in the periureteral soft tissues, and 

cause schistosomal granuloma and mural fibrosis with focal 

stricture. 

The pelvi-calyceal system and focally parenchimal tissue of the 

kidney showed numerous deposits of calcified schistosomal eggs 

within inflammatory infiltration with granulomas and fibrosis. 

Conclusion. Direct schistosomal infection of kidney is very rare 

(only a few cases have been published). 

Clear cell sarcoma of soft tissue. A case report 

331 

1)Palumbieri G. 



Background. This rare neoplasm is a clear cell sarcoma tipically 

involving tendons, fascia or aponeuroses of the distal lower extremities 

of young adults (the median age is 29 years old); it can 

occur in patients of any age, but it is extremely rare in children 

and in the old. It behaves like a high-grade soft tissue sarcoma 

with poor overall survival. It derived from neural crest cells with 

melanocitic differentiation: neoplastic immunophenotype is positive 

with HMB-45, S-100 protein and other melanoma antigens; 

furthermore melanosomes in varyng stages of development are 

detected in the cytoplasm by electron microscopy. However in 

contrast to malignant melanoma and other malignancies, clear 

cell sarcoma of soft tissue show a reciprocal chromosomal 

translocation t(12;22)(q13;q12), that leads to the fusion of the 

EWS gene (22q12) to ATF1 gene (12q13) in up to 90% of cases 

(EWSR1-ATF1 fusion gene), demonstrated by RT-PCR and 

FISH; in addition, the clear cell sarcoma of the gastrointestinal 

tract may have a variant fusion gene EWSR1-CREB1. Metastasis 

occurs mainly to regional lymph node, lungs, bone, liver, skin, 

heart and brain. Poor prognosis is associated with tumour size 

more than 5 cm, necrosis, local recurrence and metastasis. Clinicopathologic 

case: a 73 year-old female presents a soft tissue 

tumour of the ankle, near the tendon, for over a year. At gross 

examination, the tumour is a circumscribed, lobulated, firm, 

white-yellow mass measuring 8 × 6.7 × 4 cm, with omogeneous 

cut surface, distorted by focal necrosis, hemorrhage or pseudocystic 

change. 

Methods. The surgical specimen was fixed in 10% neutral buffered 

formalin and embedded in paraffin; sections were stained 

with haematoxylin-eosin, pas/ pas-diastasi stain, Gomori stain 

and Van Gieson stain; for immunohistochemistry, the avidin-biotin 

peroxidase complex method was used: antibodies employed 

are Ki67-MIB1, S-100 protein, HMB-45, melan-A, vimentin, 

desmin, actin-HHF35, CD34, CD117, CD10, EMA, CEA, CK- 

MNF116, CK7, CK20, 34betaE12, p63. 

Results. Histopathological and immunohistochemical features: 

a poorly circumscribed uniform neoplasm positive for HMB45 

and S-100 protein, with low pleomorphism and mitotic activity, 

composed of polygonal cells or fusiform cells with clear or eosinophilic 

cytoplasm, vesicular nuclei and prominent nucleolus; 

they were arranged in nests or short fascicles, separated by thin or 

dense fibrocollagenous septa, sometimes with focal hemorrhage, 

necrosis and microcystic aspect. 

The mainstay of treatment is wide excision of tumour. The use 

of sentinel lymph node biopsy is important in detecting regional 

early micrometastasis and guiding the extent of surgery. 

Hyperfunctioning lipoadenoma of thyroid gland: 

first case report 

1)Palumbieri G. 

1)U. O. C. Istologia patologica e Citodiagnostica, Ospedale “Mons. R. 

Dimiccoli”, Barletta, Italia 

Background. Lipoadenoma of thyroid gland is an extremely rare 

thyroid follicular adenoma with mature adipose cells interspersed 

throughout the tumour; to date, nearly 13 cases have been published 

in the literature worldwide, but I describe the first case of 

the hyperfunctioning thyroid lipoadenoma, to my knowledge. 

Clinico-pathologic case. A 64-year-old woman with a history of 

hyperfunctioning thyroid nodule of the left lobe and, on scintigraphic 

radionuclide scan, a “hot” nodule of the left thyroid lobe 

with an inhibition of activity of the right thyroid lobe. A left 

thyroid lobectomy was performed.

332 

Methods. Macroscopically, left lobectomy specimen of thyroid 

measured 33 × 25 × 24 mm; the esternal surface was smooth 

and encapsulated, while the cut surface shown a solitary oval 

tan-brownish nodule of 27 × 16 mm. The surgical specimen was 

fixed in 10% neutral buffered formalin and embedded in paraffin. 

Sections were processed for routine staining, including hematoxylin-eosin 

and Pas; for immunohistochemistry, the avidin-biotin 

peroxidase complex method was used: antibodies employed are 

pankeratin, thyroglobulin, TTF1 and S100 protein. 

Results. Microscopically, the nodule consisting of both thyroid 

follicular cells and mature adipose tissue and it was enclosed 

in a thin fibrous capsule; the tumour predominantly was found 

to be composed of thyroid follicular cells arranged in a normomacrofollicular 

pattern (90% of the tumour) intermixed with 

mature adipose tissue (10% of the tumour); the fatty component 

predominantly was found adjacent to the capsule of the tumour; 

degenerative haemorrhagic changes were seen within thyroid 

follicular component and focally within the fatty component. No 

cellular atypia and no signs of capsular or vascular invasion were 

seen in the tumour. 

Results. In the literature all patients with thyroid lipoadenoma 

were euthyroid, and all those who had a scintigraphic scan had an 

absence of activity at the site of the nodule. 

The present paper reports the first case of the hyperfunctioning 

thyroid lipoadenoma, to my knowledge. 

Multinodular paraganglioma of thyroid gland. 

A case report 

Palumbieri G. 



Background. Thyroid paragangliomas are very rare neuroendocrine 

tumors of paraganglionic origin, with 26 cases reported in 

the literature, and multinodular macroscopic pattern is exceptionally 

rare. 

A 63-year-old female had presented with euthyroid multinodular 

thyroid gland. The patient was treated with total thyroidectomy: 

at gross examination, the gland has distorted shape with multiple 

brownish nodules of variable size (range, 0.2-2.5). 

Methods. Tissue were fixed in buffered formalin and routinely 

processed for inclusion in paraffin; sections were stained with 

haematoxylin-eosin, with PAS stain and with Gomori stain; for 

immunohistochemistry, the avidin-biotin peroxidase complex 

method was used. 

Results. Histopathological features: a poorly circumscribed 

multiple macro-micronodular neoplasm with an organoid nesting 

pattern (“zellballen”), sometimes with trabecular or confluent 

sheet pattern, composed of slightly atypical monomorphic polygonal 

chief cells with elongated sustentacular cells at periphery 

of the nests or intermingled with the chief cells. Unlike malignant 

neoplasms elsewhere, local infiltration is not indicative of malignancy 

in thyroid paraganglioma. 

Immunohistochemical features: the chief cells are positive for 

chromogranin and NSE (cell proliferation marker Ki67-MIB1 

was low: less than 3-4% of chief cells showed nuclear staining), 

whereas no immunoreactivity was detected for calcitonin, CEA, 

TTF-1, cytokeratins, calcitonin, thyroglobulin, galectin-3 e TPO; 

sustentacular cells are positive for S-100 protein. 

Differential diagnosis includes hyalinizing trabecular tumour of 

the thyroid gland, medullary thyroid carcinoma and metastatic 

carcinoid tumour. 


Neurothekeoma: a case report 

1)Palumbo M. 2)Cocca MP. 3)Fiore G. 4)Di Clemente D. 

5)Arborea G. 6)Montrone T. 7)Cimmino A. 

1)Dipartimento di anatomia patologica, Policlinico di Bari, Bari, Italia 


3)Dipartiemnto di anatomia patologica, Policlinico di Bari, Bari, Italia 

4)Anatomia patologica, Policlinico di Bari, Bari, Italia 5)Anatomia patologica, 

Policlinico di Bari, Bari, Italia 6)Anatomia patologica, Policlinico di 

Bari, Bari, Italia 7)Anatomia patologica, Policlinico di Bari, Bari, Italia 

Background. Neurothekeomas (nerve sheath myxomas) are uncommon 

benign tumors of nerve sheath origin, most commonly 

located on the upper extremities and the head and neck. They 

also occurred on the trunk, the lower extremities, and mucosa. 

Histologic variants of neurothekeomas include classical, cellular, 

and mixed tumors. 

Methods. We describe the case of a 11 year old boy who presents 

a nodular formation in fifth finger of his left hand, looking cartilage, 

not adherent to the flexor tendons. 

Results. We observed a multinodular tumor with a variably myxoid 

stroma; these nodules are round or ovoid, of varying size and 

demarcated by usually thin collagen bands containing delicate 

blood vessels. The cells, often spindle, were aggregated and in 

some cases form whorls; stromal mucin, on Alcian blue stain, 

was focal. At immunohistochemistry, this tumor was reactive for 

Vimentin, negative for S-100 protein and EMA. 

Several processes enter into the differential diagnosis: myxoid 

schwannoma, neurofibroma, low-grade myxofibrosarcoma. 

Neurothekeomas should be included in the differential diagnosis 

of dermal nodules in infants and children. 

Duodenal somatostatinoma: case report 

1)Palumbo M. 2)Cocca MP. 3)Piscitelli D. 4)Fiore MG. 5)Rossi 

R. 6)Resta L. 







Background. Neuroendocrine tumors (NETs) of the stomach, intestine, 

and pancreas are heterogeneous, as far as their morphology, 

function, and biology are concerned. NETs producing mainly 

somatostatin have been observed in the duodenum, pancreas, bile 

ducts, and ovaries. In the duodenum, SOM-NETs have been reported 

in the setting of both the multiple endocrine neoplasia type 

1 (MEN1), the neurofibromatosis type 1 (NF1) and von Hippel 

–Lindau(VHL) syndromes. According to the WHO criteria (site, 

size, angioinvasion, infiltration level, proliferation index, immunohistochemical 

phenotype, and evidence of metastatic spread), 

NETs were classified as well-differentiated, of uncertain biological 

behavior,well-differentiated neuroendocrine carcinomas or 

poorly differentiated neuroendocrine carcinomas. 

Methods. We describe the case of a woman aged 70 who presented 

a duodenal mass, firm in consistency and grey in colour, 

measuring 2,2 cm. 

The specimen was fixed in 4% formaldehyde; from paraffin-embedded 

tissue blocks, 3-4 µm thin sections were cut and stained 

with hematoxylin and eosin and periodic acid-Schiff. Preparation 

of tissues and immunohistochemical expression analysis were 

performed as described previously in detail. 

Results. We observed a neuroendocrine tumor of the duodenum 

arranged in nests with dysmetric, sometimes atypical, nuclei, 

infiltrating the duodenal wall up to the visceral serosa, with 

ulceration of the overlying mucosa. We have not observed psammomatosi 

bodies and the tumor proliferation index (Ki67/MIB-1) 

was < 1/10 HPF.


This tumor was immunostained for chromogranin A, synaptophysin, 

somatostatin, NSE and CD56: all these investigations 

were positive. We report the ultrastructural findings of granules 

and vescicles appearing in the tumor cells, membrane-bounded, 

rounded with clear peripheral halo and dense core. 

A novel case of rhabdoid colorectal carcinoma 

associated with a CIMP+ phenotype and BRAF 

mutation 

1)Pancione M. 2)Di blasi A. 3)Sabatino L. 4)Fucci A. 5)Dalena 

AM. 6)Palombi N. 7)Carotenuto P. 8)Daniele B. 9)Normanno N. 

10)Colantuoni V. 

1)Department of Biological and Environmental Science, University of 

Sannio, Benevento, Italy 2)Departments of Oncology and Pathology, 

Azienda Ospedaliera “G. Rummo”, Benevento, Italy 3)Department of 

Biological and Environmental Science, University of Sannio, Benevento, 

Italy 4)Department of Biological and Environmental Science, University 

of Sannio, Benevento, Italy 5)Departments of Oncology and Pathology, 

Azienda Ospedaliera “G. Rummo”, Benevento, Italy 6)Departments of 

Oncology and Pathology, Azienda Ospedaliera “G. Rummo”, Benevento, 

Italy 7)Pharmacogenomic Laboratory, Center For Oncology Research, 

Mercogliano, Avellino, Italy 8)Departments of Oncology and Pathology, 

Azienda Ospedaliera “G. Rummo”, Benevento, Italy 9)Pharmacogenomic 

Laboratory, Center For Oncology Research, Mercogliano, Avellino, Italy 

10)Department of Biological and Environmental Science, University of 

Sannio, Benevento, Italy 

Background. Colorectal carcinoma with rhabdoid features is a 

rare tumor only five cases of which have been described so far. 

The molecular alterations underlying this rare phenotype have 

not been clarified. 

Methods. Immunohistochemical staining for a panel of twenty 

different markers was performed on paraffin embedded tissues. 

BRAF and KRAS mutations at codon 600 in exon 15 and codons 

12/13 in exon 2, respectively, were evaluated by PCR/sequencing 

and Real-Time PCR. CpG island methylator phenotype (CIMP) 

genes and additional loci were assessed by methylation specific 

PCR after DNA bisulphite modification. 

Results. The tumor was extremely aggressive displaying rapid 

growth and metastasis spreading to the liver and other distant 

organs. The patient, a 71-year-old woman, died within only eight 

months from surgery despite target chemotherapy. Histologically, 

the tumor was enriched in cells with a typical rhabdoid-type morphology 

showing intense and diffuse vimentin staining. Tumor 

cells were variably positive for EGFR, p53, Ki67 and β-catenin 

and negative for CK20/CK7, E-cadherin and CDX2. In addition, 

a marked reduction or loss of MSH2/MLH1 expression was detected, 

suggesting a high microsatellite instability. Genetic analysis 

revealed the presence of the V600E BRAF mutation and absence of 

KRAS mutations. Remarkably, DNA methylation was observed at 

4 out of 5 (80%) CIMP specific markers such as: MLH1, CDKN2A, 

IGF2, NEUROG1, RUNX3, whereas, no methylation at four additional 

tumor suppressor gene promoters (CDH1, CDKN1B, CD- 

KN1C and MGMT). This is the first case of a colorectal carcinoma 

with rhabdoid features associated with microsatellite instability, 

CIMP+ phenotype and BRAF mutation. The present findings indicate 

that genetic and epigenetic events contribute to the occurrence 

of this rare phenotype, suggesting potential implications for the 

clinical management of this highly aggressive malignancy. 

HCG hastens both the spontaneous development 

of mammary carcinom and the metastatization 

of erBB-2+cells in mice 

1)Pannellini (T). 2)Mariotti (M). 3)Hysi (A). 4)Toto (V). 

5)Stramucci (L). 6)Musiani (P). 7)Iezzi (M). 


Chieti, Italia 2)Oncologia E Neuroscienze/Oftalmologia, SS. Annunziata/CESI, 

Chieti, Italia 3)Anatomia Patologica/Oncologia E Neuroscienze, 

CESI, Chieti, Italia 4)Anatomia Patologica/Oncologia E Neuroscienze, 

333 

SS. Annunziata/CESI, Chieti, Italia 5)Anatomia Patologica/Oncologia E 

Neuroscienze, CESI, Chieti, Italia 6)Anatomia Patologica/Oncologia E 

Neuroscienze, SS. Annunziata/CESI, Chieti, Italia 7)Anatomia Patologica/Oncologia 

E Neuroscienze, SS. Annunziata/CESI, Chieti, Italia 

Background. Breast cancer is more frequent in human nulliparae, 

whereas its incidence is reduced by early full-term pregnancy. 

Rodent studies have suggested that normal hCG secretion during 

pregnancy may afford protection by inducing breast structure differentiation. 

The opposite effect, however, has been observed in 

transgenic mice that overexpress the hCG-β subunit or LH (hCG 

and LH interact with the same hCG/LH receptor) and develop 

breast cancers. 

Methods. We have assessed the effect of administration of hCG 

for 21 days (corresponding to the duration of pregnancy) in young 

virgin transgenic mice carrying the activated rat HER-2/neu oncogene 

(BALB-neuT). These mice develop atypical mammary 

duct hyperplasia at four weeks of age and then multiple mammary 

tumors. 

Results. We found that hCG accelerates this development. In addition, 

examination of a tumor cell line from BALB-neuT mice 

indicated that hCG acts both indirectly through the production 

of ovarian hormones, and directly by enhancing the proliferation 

and metastasization of cells expressing the hCG/LH receptor as 

well as the HER-2/neu protein product (r-p185 neu ). These findings 

suggest that hCG favours the growth and progression of p185 neu 

and hCG/LH receptor-positive breast tumors. 

WIP null mice display a progressive immunological 

disorder that resembles Wiskott-Aldrich syndrome 

1)Pannellini (T). 2)Toto (V). 3)Liberatore (M). 4)Curcio (C). 

5)Anton (IM). 6)Musiani (P). 




SS. Annunziata/CESI, Chieti, Italia 4)Oncologia E Neuroscienze/ 

Oftalmologia, SS. Annunziata/CESI, Chieti, Italia 5)Centro de Biología 

Molecular Severo Ochoa, Csic-uam, Universidad Autónoma de Madrid, 

Madrid, Spagna 6)Anatomia Patologica/Oncologia E Neuroscienze, SS. 

Annunziata/CESI, Chieti, Italia 

Background. The Wiskott-Aldrich syndrome (WAS) is an Xlinked 

immunodeficiency syndrome caused by mutations in the 

WAS protein (WASP). This participates in signalling and cytoskeletal 

homoeostasis, and some of its activities are regulated 

by its binding to the WASP interacting protein (WIP). WIP deficiency, 

however, has not yet been shown to be of pathological 

significance in humans. 

Methods. Here we show that, in WIP null (WIP(-/-)) mice, it produces 

haematological alterations and anatomical abnormalities in 

several organs, most probably as a consequence of autoimmune 

attacks. 

Results. Granulocytosis and severe lymphopenia are associated 

with a proportional increase in segmented cells and fewer bone 

marrow erythrocytes and lymphocytes. Splenomegaly is accompanied 

by an increase of haematopoietic tissue and red pulp, 

reduction of the white pulp, and fewer B (B220(+)) lymphocytes 

(also apparent in the lymph nodes and Peyer’s patches). Ulcerative 

colitis, interstitial pneumonitis, glomerular nephropathy 

with IgA deposits, autoantibodies, and joint inflammation are 

also evident. These progressive immunological disorders closely 

mimic those seen in WAS. WIP deficiency may thus be implicated 

in some cases in which mutations in the gene encoding 

WASP are not detected.

334 

Spontaneous cutaneous cholesterol crystal 

embolism with focal clinical symptomatology. 

report of a case in unusual location with secondary 

histological changes reminiscent of atypical 

decubital fibroplasia 

1)Panniello G. 2)Fenizi G. 3)Amicarelli V. 4)Sanguedolce F. 

5)Grasso M.A. 6)Bisceglia M.. 

1)Unit of Clinical Dermatology, Ospedali Riuniti, Foggia, Italy 2)Unit of 

Clinical Dermatology, Ospedali Riuniti, Foggia, Italy 3)Unit of Clinical 

Dermatology, Ospedali Riuniti, Foggia, Italy 4)Unit of Anatomic Pathology, 

Ospedali Riuniti, Foggia, Italy 5)Hospital Pharmacy Program, School 

of Pharmacy, “La Sapienza” University, Rome, Italy 6)Unit of Anatomic 

Pathology, IRCCS Casa Sollievo della Sofferenza, San Giovanni Rotondo, 

Italy 

Background. Cholesterol crystal embolization (CCE), also 

called arterial embolism of atheromatous origin, is one of the 

many complications of atherosclerosis. The clinical history of 

these patients usually includes either hypertension, ischemic 

heart disease, renal failure, aortic aneurysm, cerebrovascular 

disease, congestive heart failure, and/or diabetes mellitus. CCE is 

usually a iatrogenic event occurring either after vascular (mostly 

aortic) surgery (e.g. grafting of an aneurysm) or invasive angiographic 

investigations (mainly coronarography) or in the course 

of anticoagulant or thrombolytic therapy. More rarely it occurs 

after trauma or even in absence of any inciting cause. CCE may 

manifest as a systemic disease, characterized by constitutional 

symptoms (fever, myalgia, anaemia) and clinical signs due to 

single or multiorgan involvement (renal failure, gastrointestinal 

ischemia, central nervous system infarcts or retinal disturbances). 

CCE may also be subclinical, especially in pelvic organs, where 

collateral circulation is more extensive. The skin is one of the 

most commonly involved organs 1 . In decreasing order of frequency 

the cutaneous manifestations described include livedo 

reticularis, gangrene, cyanosis (“blue toe syndrome”), ulcers, 

nodules, or purpura 2-4 . Skin manifestations are usually seen in the 

context of a systemic disease and are mostly confined to the lower 

extremities and lower trunk, with the upper extremities involved 

in only 8% of cases 2 3 . 

Objectives. To report a rare case of spontaneous focal cutaneous 

involvement of the upper limb, with peculiar reactive histological 

changes. 

Case Report. A 65-year old man with a clinical history of ischemic 

heart disease, nephroangiosclerosis, and peripheral lower 

limbs arterial atherosclerosis was admitted with a complaint of a 

non-healing, painful skin ulceration on his left elbow of 5 months 

duration. At physical examination this chronic lesion, which had 

already been treated with topical conventional medications in 

outside hospitals, appeared as an elevated, poorly circumscribed, 

reddish and infiltrated plaque of 6 cm in its main diameter with a 

central ulceration of 3 cm. Chemical laboratory tests documented 

an elevated erythrocyte sedimentation rate, mildly elevated levels 

of blood urea nitrogen and creatinine, and an elevated serum 

level of uric acid. The clinical differential diagnosis included 

gout inflammatory arthritis, for which colchicine treatment 

was started, local atheroembolic disease confined to the upper 

left arm, and a soft tissue tumor or pseudotumor. Radiologic 

examination of the elbow excluded joint and bone destruction if 

any due to (gout) arthritis. Funduscopic examination of the eyes 

did not reveal retinal emboli. A skin biopsy, including subcutis, 

taken from the border of the ulcer, was sent for histological 

examination. Microscopically liquefactive and coagulative, ulcerative, 

necrosis of the upper dermis with underlying reactive 

fibrosis extending to the subcutaneous fat were the main findings. 

Capillary and fibroblast proliferation, small vessels wall 

pseudovasculitic changes with fibrin thrombi, and secondary 

acute and chronic inflammatory cells were clearly evident. Urate 


crystal deposits were not seen. A peculiar finding noted was the 

presence of large and even bizarre fibroblasts with abundant 

cytoplasm and large hyperchromatic nuclei. At the deep border 

of the biopsy an arteriole was seen with occlusion of the central 

lumen by a needle-shaped cholesterol crystal cleft. Taking all the 

evidence into account, the diagnosis of “cholesterol embolism 

with fibroblastic pseudotumoral reaction” was established. Then 

the patient underwent wedge resection of the ulceration and the 

wound was sutured and normally repaired. At 1-year follow-up 

the patient has no evidence of local disease. 

Discussion. The incidence of CCE is difficult to estimate and 

not firmly established due to the polymorphism of the clinical 

presentation and the subclinical and asymptomatic presentation 

in some instances 3 . An autopsy study on unselected individuals 

discovered a frequency of 4% for spontaneous (asymptomatic) 

CCE 3 . The disease is due to the detachment of cholesterol crystals 

from atheromatous plaques which disseminate to and occlude 

small arteries or arterioles. Cutaneous manifestations mostly occur 

on the lower limbs. It is extremely difficult to suspect spontaneous 

CCE involving the skin in isolation. Individuals who may 

experience such an event are elderly patients with atherosclerotic 

problems. The main clinical skin findings are cyanosis, gangrene, 

ulcerations and pseudovasculitic nodules 4 5 . The diagnosis is histologically 

established by documenting atheromatous debris with 

cholesterol crystals occluding arteriole(s). To our best knowledge, 

focal cutaneous involvement of the upper limb by spontaneous 

CCE has not been reported so far. Skin biopsy is a sensitive 

tool for diagnosis revealing the diagnostic finding of biconvex 

cholesterol crystal clefts occluding emboli in up to 90% of cases 5 . 

The rest of the histological changes in cases of long standing 

skin ulcer are mainly represented by necrosis of the upper dermis 

with underlying deep dermal and subcutaneous fibrotic changes, 

pseudovasculitic vessel changes and nonspecific inflammatory 

infiltrates. The atypical fibroblastic reaction described above was 

the subject of speculation and analogies were made with the 

atypical decubital fibroplasia seen in debilitated patients 6 , which 

in our case should be attributed to prolonged impaired circulation 

in proximity of a bony protuberance. 

references 

1 Scolari F, Tardanico R, Zani R, et al. Cholesterol crystal embolism: A 

recognizable cause of renal disease. Am J Kidney Dis 2000;36:1089- 

109. 

2 Falanga V, Fine MJ, Kapoor WN. The cutaneous manifestations of 

cholesterol crystal embolization. Arch Dermatol 1986;122:1194-8. 

3 Donohue KG, Saap L, Falanga V. Cholesterol crystal embolization: an 

atherosclerotic disease with frequent and varied cutaneous manifestations. 

J Eur Acad Dermatol Venereol 2003;17:504-11. 

4 Hirschmann JV, Raugi GJ. Blue (or purple) toe syndrome. J Am Acad 

Dermatol 2009;60:1-20. 

5 Carlson JA, Chen KR. Cutaneous Pseudovasculitis. Am J Dermatopathol 

2007;29:44-55. 

6 Montgomery EA, Meis JM, Mitchell MS, et al. Atypical decubital 

fibroplasia. A distinctive fibroblastic pseudotumor occurring in debilitated 

patients. Am J Surg Pathol 1992;16:708-15. 

Novel virus other than HPV 16 in laryngeal cancer: 

a case report with an innovative molecular 

virology profile 

1)Pannone G. 2)Sanguedolce F. 3)Santoro A. 4)Mattoni M. 5)De 

maria S. 6)Lo muzio L. 7)Bufo P. 

1)Department of Surgical Sciences, Institute of Path, Riuniti, Foggia, Italy 




5)Cnr, Institute of Genetics and Biophysics “Adriano Buz, Napoli, Italy 

6)Department of Surgical Sciences, Irccs Crob - Centro Di Riferimento 

Oncologico Di B, Rionero In Vulture, Italy 7)Department of Surgical 

Sciences, Institute of Path, Riuniti, Foggia, Italy


Background. Human Papilloma Virus infection is thought to 

play a role in laryngeal carcinogenesis; the variable association 

reported in literature may be due to wide range of HPV genotypes. 

We report the case of a 51-year-old man affected by laryngeal 

squamous cell carcinoma. 

Methods. Analysis of DNA extracted by cancer cells was performed 

by an innovative molecular virology assay (INNO-LiPA 

HPV Genotyping Extra). Immunohistochemical staining with 

anti-HPV-capsid antigen monoclonal antibody (clone K1H8; 

DAKO), which allows detection of a main capsid-epitope of HPV 

6, 11, 16, 18, 31, 33, 42, 51, 52, 56, and 58, and with anti-E7 viral 

antigen (polyclonal; Zymed) by a standard LSAB-HRP technique 

was also valued. 

Results. Our study showed the presence of two high-risk HPV 

genotypes, HPV-73 and -82. Immunohistochemical examination 

confirmed positivity for both capsid protein and viral oncogenic 

protein E7. Such association has never been reported in literature 

so far, and a brief discussion on the importance of assessing HPV 

status in laryngeal cancer is provided. 

evaluation of the risk of neoplastic transformation 

in cases of premalignant oral lesions 

1)Panzacchi R. 2)Cocchi R. 3)Eusebi LH. 4)Pennesi MG. 5)Foschini 

MP. 

1)Dipartimento di ematologia e scienze oncologiche “L.e A. 

Seragnoli”,Sezione di Anatomia Patologica, Ospedale Bellaria, Università 

di Bologna, Italia 2)Unità operativa di Chirurgia Maxillo-Facciale, 

Ospedale Bellaria, Bologna, Italia 3)Dipartimento di ematologia e scienze 

oncologiche “L.e A. Seragnoli”,Sezione di Anatomia Patologica, Ospedale 

Bellaria, Università di Bologna, Italia 4)Unità operativa di Chirurgia 

Maxillo-Facciale, Ospedale Bellaria, Bologna, Italia 5)Dipartimento di 

ematologia e scienze oncologiche “L.e A. Seragnoli”,Sezione di Anatomia 

Patologica, Ospedale Bellaria, Università di Bologna, Italia 

Background. Oral squamous cell carcinoma (OSCC) is the most 

common malignancy of the oral cavity. Usually it is preceded by 

precancerous lesions. Clinically, the most common premalignant 

lesion is leucoplakia corresponding to a wide spectrum of histological 

features. The aim of the present study is to assess the risk 

to develop OSCC according to the various grades of histological 

premalignant lesions. 

Materials and methods. All oral biopsies diagnosed as hyperkeratosis 

or dysplasia, in the period 1992-2003 were retrieved 

from the files of the Section of Anatomic Pathology of the University 

of Bologna at Bellaria Hospital. Results were compared 

with those presented in 20 published papers. 

Results. 112 cases were selected. In 53 cases the histological 

diagnosis was “hyperkeratosis without dysplasia”, in 21 cases 

“mild dysplasia”, in 19 cases “moderate dysplasia” and in 19 

cases “severe dysplasia or in situ carcinoma”. An OSCC arose in 

2/53 cases of hyperkeratosis (3.7%), in 2/21 cases of mild dysplasia 

(9.5%), in 2/19 cases of moderate dysplasia (10.5%) and in 

9/19 cases of severe dysplasia (48%). 

The scientific articles selected reported a total of 4114 cases 

with a clinical diagnosis of leucoplakia. In 1176 cases (28.6%) 

dysplasia was present at pathological examination. A follow-up 

period of 2.6-20 years revealed a carcinoma in 90/1176 cases of 

dysplasia (7.6%) and in 112/2938 cases with no dysplasia (3.8%). 

Furthermore 1039 cases with a histological diagnosis of dysplasia 

were reported: in 112/1039 cases a follow-up period of 0.5-20 

years revealed a carcinoma (10.7%). 

Discussion and conclusion. Data here presented demonstrate 

that hyperkeratosis without dysplasia has a low risk (< 4%) of 

neoplastic transformation. The risk increases when dysplasia is 

present and appears related to the grade of dysplasia. 

335 

Molecular markers of human prostate cancer in 

different stage of progression: an in situ study 

1)Paoloni S. 2)Colantoni A. 3)Perfetti A. 4)Ciafrè SA. 5)Spagnoli 

LG. 6)Bonanno E. 

1)Anatomia Patologica, Policlinico Universitario Tor Vergata, Roma, Italia 

2)Anatomia Patologica, Policlinico Universitario Tor Vergata, Roma, 

Italia 3)Anatomia Patologica, Policlinico Universitario Tor Vergata, 

Roma, Italia 4)Medicina Sperimentale E Scienze Biochimiche, Università 

di Roma Tor Vergata, Roma, Italia 5)Anatomia Patologica, Policlinico 

Universitario Tor Vergata, Roma, Italia 6)Anatomia Patologica, Policlinico 

Universitario Tor Vergata, Roma, Italia 

Background. Prostate cancer (PCa) is the most commonly diagnosed 

non-cutaneous cancer in men and is the second leading 

cause of cancer death. PCa is a highly heterogeneous disease, 

both in terms of pathology and clinical presentation. To better 

stratify prostate cancer patients we propose an in situ study to 

verify in patients’ neoplastic tissue the expression of molecules 

that commonly affected tumor-suppressive and tumorigenic pathways 

in prostate. Formalin fixed and paraffin embedded tissue 

(FFPE) collections will allow us to stratify CaP according to its 

different stages. 

Tissue microarrays (TMAs) offer the potential to rapidly translate 

basic science research findings to practical clinical application. 

Methods. In this study we compared the following experimental 

groups: Lethal phenotype (high grade, metastatic); Dormant phenotype 

(latent prostatic carcinoma); Pre-cancerous lesion (PIN); 

Benign prostatic tissue. TMA were built up using the semiautomatic 

apparatus Galileo CK3500 BioRep. The expression of 

p27, p63, CD44 and racemase has been studied by immunohistochemistry. 

Slides wer digitalised with I-Scan BioImagene digital 

microscope and analyzed with the “Tissue Mine” software for the 

evaluation of immunohistochemical reactions. 

Results. CD44, p63, Ki67 and racemase expression did not show 

any significant difference in among the subgroups of carcinomas, 

while discriminating between benign hyperplasia and cancer 

tissues. The p27 protein was expressed in the nucleus of benign 

prostatic cells whereas it was almost absent in the nucleus of 

“Lethal” phenotype tumours. In this group p27 expression was 

highly expressed in the cytoplasm. It is worth to note that the p27 

expression in “Dormant phenotype” PCa was similar to those 

observed in benign prostate tissue. The translocation of p27 in 

the cytoplasm would be correlated with PCa progression and 

its quantity could differentiate between an aggressive or latent 

carcinoma phenotype. 

Temporal lobe epilepsy model, neural stem cells 

and gene therapy 

1-2-3) B. Paradiso, 2-3) S. Zucchini, 4-3) P. Marconi, 4-3) E. 

Berto, 2-3) A. Binaschi, 1) E. Magri, 5) G. Navarro Mora, 5) P. 

Fabene, 1) A. Marzola, 2-3) M. Simonato. 

1) Department of Experimental and Diagnostic Medicine, Section of Anatomic 

Pathology, University of Ferrara, Ferrara, Italy. 2) Department of 

Clinical and Experimental Medicine, Section of Pharmacology, and Neuroscience 

Center, University of Ferrara. 3) National Institute of Neuroscience, 

Italy. 4)Department of Experimental and Diagnostic Medicine, 

Section of Microbiology, University of Ferrara. 5) Department of Morphological 

and Biomedical Sciences, Section of Anatomy, University of 

Verona, Verona, Italy 

Background. The effects of neurotophins on seizures are quite 

debated, and most of the data actually support the notion that they 

favour epileptogenesis 1 2 . At the same time it may be said that 

neurotrophins (especially in combination with other neurotrophic 

factors) can actually prevent seizure-induced damage 2 for their 

well-known neuroprotective effects. Therefore, the neurotrophic 

factors may be both “angels” and “devils” as regards epilepsy. An 

open question then becomes if neuroprotection may prevent epi-

336 

leptogenesis 3 but this possibility remains at the level of academic 

discussion, because it is obviously impossible to administer any 

treatment before occurrence of spontaneous seizures caused by 

epileptogenic lesion. 

Methods. In this work, therefore, we decided to administer the 

neurotrophic factor-producing vector when seizure-induced damage 

was already in place. 

Results. We provide evidence that the anti-epileptogenic effect 

is mediated by an increase in neuronogenesis rather then by 

prevention of ongoing damage. Very little positive results can be 

found in the literature with reference to successful prevention of 

epileptogenesis 4 and this is the first evidence of a disease-modifying 

effect based on a “treatment” of endogenous neurogenesis 

by neurotrophic factors. 

references 

1 Binder DK, Croll SD, Gall CM, et al. TiNS 2001;l24(1):47-53. 

2 Simonato M, Tongiorgi E, Kokaia M. TiPS 2006;27(12):631-8. 

3 Pitkanen A, Sutula TP. Lancet Neurology 2002;1(3):173-81. 

4 Pitkanen A, Kubova H. Expert Opin Pharmacother 2004;5(4):777-98. 

Primary breast amyloid tumor in a screening 

program 

1)Parisi A. 2)Manfrin E. 3)Brunelli S. 4)Brunelli M. 5)Nottegar 

A. 6)Bonetti F. 

1)Anatomia patologica, Policlinico G.B.Rossi, Verona, Italia 2)Anatomia 

patologica, Policlinico G.B.Rossi, Verona, Italia 3)Radiologia, Centro screening 

Marzana, Verona, Italia 4)Anatomia patologica, Policlinico G.B.Rossi, 

Verona, Italia 5)Anatomia patologica, Policlinico G.B.Rossi, Verona, Italia 

6)Anatomia patologica, Policlinico G.B.Rossi, Verona, Italia 

Backgrounds. Breast involvement by amyloid is rare and it is 

usually a late presentation of a previously diagnosed underlying 

disease in patients in whom diffuse visceral amyloid depositions 

are known. Localized amyloidosis of the breast constitute an exceptional 

phenomenon, with very few cases having been reported 

in the literature, sometimes associated with cancer too. 

We present a case in a screening program context. 

Methods and Results. We report the features of an amyloid 

tumor of the breast presenting as bilateral breast masses in a 52 

year old woman in 2002. Clinically and mammographically, the 

masses simulated metastatic or multifocal carcinoma. Fine-needle 

aspiration cytology (FNAC) revealed irregular globules of acellular 

amorphous material and numerous multinucleated giant cells. 

A core biopsy (CNB) showed deposition of amorphous material 

with calcifications and osseus metaplasia. The patient was lost to 

follow-up and presented eight years later with a slight increase of 

both breast lesions. 

The woman turned out to be affected by sclerodermia. First of all 

a FNA on one side and a CNB on the other were performed, with 

findings suspicious for amyloid deposit. Than a huge vacuum 

assisted biopsy with mammotome was done and the histology 

confirmed the presence of extensive vascular, interstitial, and 

periductal deposits of acellular amorphous material which occasionally 

formed large confluent patches in both breasts, with diffuse 

calcifications associated with no evidence of cancer. Congo 

red staining in polarized light showed the characteristic green 

birefringence of amyloid. 

Conclusions. The correct diagnosis and the properly management 

of patients with a primary diagnosis of breast amyloidosis 

are major problems. The comprehensive knowledge of all clinical 

data help to achieve the right diagnosis in a screening context. 

A multidisciplinary approach and a triage to evaluate the risk 

of cancer associated are determinant in the management of the 

patient. 


Hepatoid carcinoma in pancreas: report of a pure 

case 

1)Parisi A. 2)Nottegar A. 3)Pedica F. 4)Salvia R. 5)Morelli L. 

6)Bonzanini M. 7)Menestrina F. 8)Capelli P. 


patologica, Policlinico G.B.Rossi, Verona, Italia 3)Anatomia patologica, 

Policlinico G.B.Rossi, Verona, Italia 4)Chirurgia generale B, Policlinico 

G.B.rossi, Verona, Italia 5)Anatomia patologica, Ospedale S.Chiara, 

Trento, Italia 6)Anatomia patologica, Ospedale S.Chiara, Trento, Italia 


patologica, Policlinico G.B.Rossi, Verona, Italia 

Backgrounds. Hepatoid carcinoma have been described in many 

organs but very rarely in pancreas and more often in association 

with other neoplasia (endocrine tumor, ductal adenocarcinoma) 

than in pure form. 

We describe the features of a case of a pure hepatoid carcinoma. 

Methods and results. A 52 year old man presented in June 2008 

with a mass in the head of the pancreas. 

Endoscopic ultra sound fine needle aspiration showed single 

or acinar-like clusters of atypical epithelial cells, with low cromoghranine 

A expression and strong CD56 positivity at immunohistochemistry. 

The final report was suspicious for endocrine 

tumor. 

The patient underwent selective embolization followed by 4 

cycles of radiomethabolic therapy. 

The TAC at July 2009 showed a reduction of the mass, with no 

evidence of secondary lesions. The patient underwent pancreatoduodenectomy. 

Sectioning the head of the pancreas a 5 cm solid 

tumor was identified, well circumscribed with psuedolobulated 

apparence, composed predominantly of sheets of large amphophilic 

to eosinophilic polygonal tumor cells separated by bands 

of fibrous tissue or with thin fibrovascular cores. Extremely focal 

identification within the cells of a yellowish-brown pigment consistent 

with bile clearly suggested hepatocellular differentiation, 

confirmed by immunohistochemistry with strong and diffuse 

positivity for anti-Hepatocyte monoclonal antibody. Tumor cells 

stained also for cytokeratin and CD56, while were completely 

negative for the other common endocrine markers. 

After surgery the patient is fine and free of disease at the first 6 

months control. 

Conclusions. Even rare hepatoid carcinoma is an interesting entity 

to keep in mind, as we can find it in many different organs and 

in pancreas too. We don’t believe it belong to some heterotopic 

liver tissue in pancreas, as we’ve never seen normal liver tissue 

in pancreas. Recognise these tumor is important trying to better 

understand their origin and their prognostic significance. 

Hepatocellular carcinoma with intrabile duct 

growth: report of 3 cases 

1)Pedica F. 2)Pecori S. 3)Cataldo I. 4)Pachera S. 5)Ruzzenente 

A. 6)Campagnaro T. 7)Guglielmi A. 8)Daniele I. 9)Piccoli P. 

10)Capelli P. 

1)Patologia E Diagnostica-Sez. Anatomia Patologica, Policlinico G.B. 

Rossi-Università di Verona, Verona, Italia 2)Patologia E Diagnostica-Sez. 

Anatomia Patologica, Policlinico G.B. Rossi-Università di Verona, Verona, 

Italia 3)Patologia E Diagnostica-Sez. Anatomia Patologica, Policlinico 

G.B. Rossi-Università di Verona, Verona, Italia 4)Dipartimento di Scienze 

Chirurgiche, Policlinico G.B. Rossi-Università di Verona, Verona, Italia 

5)Dipartimento di Scienze Chirurgiche, Policlinico G.B. Rossi-Università 

di Verona, Verona, Italia 6)Dipartimento di Scienze Chirurgiche, Policlinico 

G.B. Rossi-Università di Verona, Verona, Italia 7)Dipartimento di 

Scienze Chirurgiche, Policlinico G.B. Rossi-Università di Verona, Verona, 

Italia 8)Patologia E Diagnostica-Sez. Anatomia Patologica, Policlinico 

G.B. Rossi-Università di Verona, Verona, Italia 9)Patologia E Diagnostica-Sez. 

Anatomia Patologica, Policlinico G.B. Rossi-Università di Verona 

Introduction. Hepatocellular carcinoma (HCC) is the fifth most 

common cancer in the world and rarely jaundice is caused by


tumor invasion into the bile duct. HCC with intrabile duct growth 

(IG) was called “icteric type” hepatoma and “cholestatic type 

HCC”. 

We describe 3 cases of HCC with IG and studied the immunofenotype 

because there are no data in literature, to our knowledge. 

Methods. We observed 3 cases of HCC with IG in our institution 

in these last 2 years, which correspond to 3 male patients with age 

variable between 61-68 years old. 

The first was a Chinese affected by neonatal HBV, while the second 

was positive for HbcAb and the third last one was negative 

for viral hepatitis markers. Liver parenchyma was not cirrhotic. 

We applied a panel of 8 immunohistochemical markers including 

CK8-18, Hep par1 (OCH1E5), CK7, CK19, alfa-fetoprotein 

(AFP), CD133 (prominin-1), CD56 (NCAM) and OV-6. 

Results. All 3 cases were positive for CK8-18 and Hep par1, but 

variabily expressing the other markers. 

HCC of the first patient was strongly AFP and focally OV-6 positive 

in some neoplastic intraductal cells. 

The third case had almost 50% CK7 and 30% CK19 positive 

cells; some of them were OV-6 strongly positive. Moreover a few 

tumoral cells were also positive for CD56. 

In all 3 cases perilesional ductal proliferations were CD133, OV- 

6, CK7 and CK19 positive. 

In conclusion, the immunophenotype was slighly different in 

these 3 cases. 

The first case was AFP positive, differently from other 2 cases 

and this marker is well known to be a poor prognostic marker and 

to be also progenitor cell marker. 

The last case espressed CK7, CK19 and OV-6. 

We can suggest that also this rare kind of HCCs has a heterogenous 

phenotype and is composed by different cell populations. 

Smooth muscle cell layer in inflammatory bowel 

diseases 

1)Pedica F. 2)Pecori S. 3)Pedron S. 4)Montagna L. 5)Capelli P. 

6)Meneestrina F. 7)Chilosi M. 

1)Patologia e Diagnostica-Sez. Anatomia Patologica, Policlinico G.B. 

Rossi-Università di Verona, Verona, Italia 2)Patologia e Diagnostica-Sez. 

Anatomia Patologica, Policlinico G.B. Rossi-Università di Verona, Verona, 

Italia 3)Patologia e Diagnostica-Sez. Anatomia Patologica, Policlinico 

G.B. Rossi-Università di Verona, Verona, Italia 4)Patologia e Diagnostica-Sez. 

Anatomia Patologica, Policlinico G.B. Rossi-Università di Verona, 

Verona, Italia 5)Patologia e Diagnostica-Sez. Anatomia Patologica, 

Policlinico G.B. Rossi-Università di Verona, Verona, Italia 6)Patologia e 

Diagnostica-Sez. Anatomia Patologica, Policlinico G.B. Rossi-Università 

di Verona, Verona, Italia 7)Patologia e Diagnostica-Sez. Anatomia Patologica, 

Policlinico G.B. Rossi-Università di Verona, Verona, Italia 

Background. The lymphatic system consists of a network of 

thin-walled vessels that drain fluid and particles from the interstitial 

spaces. Nowadays, our understanding of the lymphatic system 

is still scanty, especially in humans. Lymphatic capillaries consist 

of a single layer of non-fenestrated endothelial cells resting on 

incomplete basal lamina, while collectors posses a smooth muscle 

cell layer. 

Their role in inflammatory bowel diseases (IBDs) is under investigation. 

Especially in Crohn’s disease (CD), lymphatics have 

been hypothized to be fundamental in its etiopathogenesis. 

We hypothized that in CD lymph vessels could express SMA 

because of high pressure which they are forced to. Our aim was 

to investigate lymph vessels in IBDs and normal colon applying 

immunohistochemical analysis for smooth muscle actin (SMA) 

and podoplanin. 

Methods. We collected 17 surgical samples including 11 cases of 

CD, 2 cases of ulcerative colitis (UC) and 4 resection margins for 

colic adenocarcinoma as controls. 

We applied a double staining with SMA (in brown) and podoplanin 

(in red) to analyze their possible concomitant expression in 

337 

some lymphatics. We count their mean number expressing SMA 

in 3 “hot spots” (characterized by major density of lymph vessels) 

and the ratio SMA positive/total number of lymphatics. The 

“hot spots” were chosen between subserosa, muscolaris propria 

e submucosa. 

Results. The mean number of SMA positive lymph vessels was 

7,32 in CD (ratio = 0,86), 2 in UC (ratio = 0,26) and 0,075 (ratio 

= 0,017) in normal controls. 

Conclusions. These preliminary datas demonstrate that lymph 

vessels are different in number and in structure in CD, UC and 

normal controls and suggest that lymph vessels in CD may 

develop smooth muscle cell layer when they are forced to high 

pressure. 

These data needs to be verified on a larger number of cases but 

suggest that CD may be biologically and structurally different 

from UC. 

Compound BRCA1 and HMlH1 mutation in a young 

woman 

1)Pedroni M. 2)Di Gregorio C. 3)Cortesi L. 4)Botticelli L. 

5)Priore Oliva C. 6)Simone ML. 7)Medici V. 8)Federico M. 

9)Viale G. 10)Ponz de Leon M. 

1)Dipartimento di Medicina Interna, Università di Modena e Reggio Emilia, 

Modena 2)Dipartimento integrato di Laboratori, Anatomia Patologica 

e Medicina legale, Az. ospedaliero universitaria policlinico, Modena 

3)Dipartimento di Oncologia ed Ematologia, Az. ospedaliero universitaria 

policlinico, Modena, 4)Dipartimento integrato di Laboratori, Anatomia 

Patologica e Medicina legale, Az. ospedaliero universitaria policlinico, 

Modena 5)Dipartimento di Scienze Biomediche, Università di Modena e 

Reggio Emilia, Modena, 6)Dipartimento di Scienze Biomediche, Università 

di Modena e Reggio Emilia, Modena, Italia 7)Dipartimento di Scienze 

Biomediche, Università di Modena e Reggio Emilia, Modena 8)Dipartimento 

di Oncologia ed Ematologia, Az. ospedaliero universitaria policlinico, 

Modena 9)Anatomia Patologica e MDL, Istituto Europeo Oncologia, 

Milano, 10)Dipartimento di Medicina Interna, Università di Modena e 

Reggio emilia, Modena 

Background. Germline mutations in BRCA1 and BRCA2 genes 

are responsible for a large proportion of hereditary breast and 

ovarian cancers. Carriers of mutations in Mismatch Repair genes 

predispose to Lynch Syndrome, characterized by early malignancies 

of the large bowel and others epithelial tumours, including 

endometrial and urological cancers. 

Methods. We describe a case showing early onset unilateral 

breast cancer at 35 years, who subsequently developed endometrial, 

ovarian cancer and renal clear carcinoma at age 39. Moreover, 

the patient was affected by an infiltrating carcinoma of the 

controlateral breast at age 46. 

Results. We found in this woman a heterozygous state for a 

BRCA1 mutation (c.300T > G). The extended genealogic tree 

showed a suspect history of breast cancer in the paternal branch 

and a strong positive history of colorectal cancer in the maternal 

branch. Consequently, endometrial cancer was investigated for 

Microsatellite Instability (MSI) and expression of Mismatch Repairs 

proteins. An elevated MSI and altered expression of MLH1 

protein were detected. The further sequencing of hMLH1gene 

revealed the mutation c.1489dupC, ex13. The same mutation 

was found in the mother of the patient. To investigate whether 

the others tumours were associated with Lynch Syndrome, we 

performed immunohistochemistry and MSI. All tumours, including 

breast cancer showed no expression of MLH1 protein and 

positivity for Microsatellite Instability. Loss of the wild type 

hMLH1 allele was detected in the breast cancer, suggesting that 

Mismatch repair defect contributed to the development of breast 

cancer phenotype. In conclusion, we describe a proband –from 

two families, one with BRCA1 and the other with hMLH1 mutations- 

with multiple tumors of various organs. Molecular data 

suggest that inactivation of both genes contribute to the development 

of breast cancer.

338 

Complete cytoreduction and hyperthermic 

intraperitoneal chemotherapy in patients 

with peritoneal carcinomatosis: 

our experience focusing on ovarian cancer 

1)E. Penitente, 1)P. Viola, 1)R. Claudi, 1)S. Malatesta, 2)R. Massari, 

3)M. De Tursi, 1)D. Angelucci, 1)A. Colasante 

1)UOC Anatomia patologica, Ospedale Clinicizzato SS Annunziata, Chieti, 

Italia; 2)UOC Chirurgia ,Ospedale Clinicizzato SS Annunziata, Chieti, 

Italia; 3)UOC Oncologia, Ospedale Clinicizzato SS Annunziata, Chieti, 

Italia 

Background. Peritoneal cacinomatosis is considered the ending 

stage for many epithelial abdominal tumors, with a poor 

prognosis. In this stage encouraging results have been reported 

with complete cytoreduction (CC) and hyperthermic intraperitoneal 

chemotherapy (HIPEC). Many studies have been reported 

encouraging results on overall survival (OS) and in disease free 

survival (DFS). We report our experience from September 2006 

to December 2009, focusing on ovarian cancer and correlating 

nuclear tumor grading with patients outcome. 

Methods. 39 patients were enrolled: 2 stomach carcinomas, 3 

sarcomas, 9 colon carcinoma and 23 ovarian carcinomas. For the 

ovarian serous carcinomas we used a 2-tier grading system based 

on nuclear grade: low (nuclear grade 1) and high grade (nuclear 

grade 3). 

Results. We studied 23 pts, median age of 63 years (range 42-76 

yrs): 3 endometrioid and 20 serous histotype; according to the 

reported grading system we have 9 G1 carcinomas (6 serous G1) 

and 16 G3 serous carcinomas. Survival curves were calculated 

with the Kaplan-Meier method and compared with the long-rank 

test. In our experience, OS was estimated to be about 70% at 2 

years and there wasn’t significant correlation with nuclear grading. 

Significant correlation was reached correlating CC with OS. 

Concerning DFS, we observed only two months difference in 

DFS G1 vs G3 patients, but it wasn’t significant. 

Conclusion. Although our data correlating nuclear grading and 

clinical evolution are not significant, several physio-pathological 

issues remain to clarify (primary cancer cell heterogeneity 

evaluation, effects of previous therapies, improvement of targeted 

therapies.); but, our data further enforce that CC and HIPEC is a 

feasible option in these ending stage patients, young adults, even 

if a randomized trials are required. 

fNA’s diagnostic role after the beginning of breast 

screening program: experience of the breast unit 

(B.u.) in Trieste 

1)Petris M. 2)Martellani F. 3)Ober E. 4)Giudici F. 5)Zacchi A. 

6)Torelli L. 7)Bonifacio D. 8)Romano A. 9)Tonutti M. 10)Di 

bonito L. 

1)A.C.A.D.E.M., AOU Ospedali Riuniti di Trieste, Trieste, Italia 2)A. 

C.A.D.E.M., AOU Ospedali Riuniti di Trieste, Trieste, Italia 3)A. 

C.A.D.E.M., AOU Ospedali Riuniti di Trieste, Trieste, Italia 4)Matematica 

E Informatica, Università, Trieste, Italia 5)A.C.A.D.E.M., AOU Ospedali 

Riuniti di Trieste, Trieste, Italia 6)Matematica E Informatica, Università, 

Trieste, Italia 7)A.C.A.D.E.M., AOU Ospedali Riuniti di Trieste, Trieste, 

Italia 8)A.C.A.D.E.M., AOU Ospedali Riuniti di Trieste, Trieste, Italia 

9)A.C.A.D.E.M., AOU Ospedali Riuniti di Trieste, Trieste, Italia 10)A. 

C.A.D.E.M., AOU Ospedali Riuniti di Trieste, Trieste, Italia 

Background. Many authors argue that FNA looses the role of 

first level morphological investigation with screening,being 

replaced by microhistology (tru-cut, VAB). We evaluated the 

role of FNA in the definition of breast lesions in Trieste after the 

beginning of screening. 

Methods. Comparison of two-year periods: 2004-2005 (before 

screening activation), 2008-2009 (coinciding with the screening 

second round). The B.U. widely respected the quality standards 

required by European guidelines for diagnostic cytology in 


both two-year periods (Cs: 96.4% -98.5%; Spe: 76,6%, PPV 

C5: 100% -99, 8%, PPV C4: 92.7% -79.1%, PPV C3: 12.3% 

-8.5%; Ina: 7.9% -4.5%). Results. The FNA was used as the first 

morphological investigation in 1384 cases (91.1%) of 1521 lesions 

studied in the first two years and in 1835 cases (88.6%) of 

2091 in the second period. Advanced investigation with tru-cut 

was necessary in 84 cases, respectively (6.1%) and in 154 cases 

(8.4%) cytologically approached, the use of VAB was necessary 

respectively in 28 cases (2%) and in only 20 cases (1.1%). The 

lesions studied exclusively with tru-cut were 10 (0.7%) in the 

first period and 28 (1.3%) in the second, those studied with VAB 

were 46 (3%) in the first and 119 (5.7%) in the second period. 

81 lesions (5.3%) were referred directly for surgery without a 

diagnostic support in the first two years and only 61 (2.9%) in 

the second period. Thanks to FNA respectively 701 (50.7%) and 

953 (51.9%)were resolved as benign lesions. The increase of all 

diagnostic methods (FNA: +32.5%; tru-cut: +93.6% and VAB: 

+87.8%) resulted in an increase in surgical activity (+19.8%) allowing 

to diagnose and treat more than 156 cancers (28.9%). In 

our Unit, due to the high standards, FNA still mantains the role 

of first choice investigation. 

Chromosome (Ch17) disorder and Ki67 expression: 

negative prognostic factors in invasive breast 

cancer 

1)Petroni S. 2)Addati T. 3)Giotta F. 4)Quero C. 5)Caponio M.A. 

6)Rubini V. 7)Palma F. 8)Mossa G. 9)Simone G. 

1)Pathological Anatomy Unit, NCI “Giovanni Paolo II”, Bari, Italy 2)Pathological 

Anatomy Unit, NCI “Giovanni Paolo II”, Bari, Italy 3)Clinical 

Experimental Oncology Unit, NCI “Giovanni Paolo II”, Bari, Italy 4)Pathological 

Anatomy Unit, NCI “Giovanni Paolo II”, Bari, Italy 5)Pathological 

Anatomy Unit, NCI “Giovanni Paolo II”, Bari, Italy 6)Pathological 

Anatomy Unit, NCI “Giovanni Paolo II”, Bari, Italy 7)Pathological Anatomy 

Unit, NCI “Giovanni Paolo II”, Bari, Italy 8)Pathological Anatomy 

Unit, NCI “Giovanni Paolo II”, Bari, Italy 9)Pathological Anatomy Unit, 

NCI “Giovanni Paolo II”, Bari, Italy 

Background. Her2 protein status and aneusomy effects of Ch17 

still remain controversial. Ch17 polysomy has been identified in 

20-40% of the invasive breast cancer, probably related to Her2 

protein expression 1 . Other studies 2 failed to confirm this relation, 

showing that it’s a rare event and that coincident CEP17 amplification 

may be due to overestimation of the Chromosome in FISH. 

The aim of this study was to evaluate polysomy/amplification 

of Cep17,in invasive breast cancer, related to Ki67, hormonal 

receptor(ER and PgR) and Her2 expression. 

Methods. 661 cases of invasive breast cancer were collected. Immunohystochemical 

staining for detection of Ki67 and hormonal 

receptors was performed on 646; HER2 protein was valued on 621 

samples.FISH (Vysis) analysis was performed on all histological 

cases. The HER2 gene was considered amplified in tumours with 

ratio≥2.2, Ch17 was considered polysomic/amplified(aneusomy) 

with CEP17≥3 copy/nucleus. 

Results. Ch17 polysomy was observed in 179/661 cases (27.1%),in 

176 protein overexpression (HercepTest-HT) was evaluated: 

39(22.2%) were scored 3+, 106 (60.3%) 2+, 31(17.6%) as 0/1+. 

HER2 gene amplification was detected in 23/179 cases(12.9%), 

enclosing 17 overexpressed 3+ and 5 scored as 2+; in 1 case HT 

was not valuable. 

Four aneusomic cases were low grade(G1) and 80 (44.7%) out 

of 179 were N+. Moreover, 120/174 evaluable tumours (70%) 

showed high Ki67 and 114(65.5%) resulted ER+/PgR+. 

Ch17 aneusomy showed a significantly higher incidence in HER2 

not amplified cases and confirmed that HER2 gene amplification 

is not associated to Ch17 disorder (aneusomy vs amplification 

p = 0.174).In this study, the high average of the patients who presented 

Ch17 aneusomy with an histological high grade(2-3),high 

proliferation index and metastatic activity, suggests a relation 

between Ch17 and negative prognostic factors leading to the


hypothesis that these patients could be a clinical high risk group, 

not responsive to the conventional therapy. 

references 

1 Shah SS, et al. Diagn Mol Pathol. March 2009;8:1. 

2 Yen I-T, et al. Modern Pathology 2009;22:1169-75. 

Biological characterization of primary breast 

cancer and corresponding abdominal and pelvic 

metastasis: report of 21 cases 

1)Petroni S. 2)Falco G. 3)Caponio M.A. 4)Altieri R. 5)Centrone 

M. 6)Simone G. 7)Giotta F. 

1)Pathological Anatomy Unit, NCI “Giovanni Paolo II”, Bari, Italy 2)Gynecology 

Unit, NCI “Giovanni Paolo II”, Bari, Italy 3)Pathological Anatomy 

Unit, NCI “Giovanni Paolo II”, Bari, Italy 4)Senology Unit, NCI 

“Giovanni Paolo II”, Bari, Italy 5)Pathological Anatomy Unit, NCI “Giovanni 

Paolo II”, Bari, Italy 6)Pathological Anatomy Unit, NCI “Giovanni 

Paolo II”, Bari, Italy 7)Medical and Experimental Oncology Department, 

NCI “Giovanni Paolo II”, Bari, Italy 

Background. Metastatic breast cancer is a heterogeneous disease 

with distinctive histological and biological features, clinical behaviours 

and therapy response. 

Abdominal and pelvic metastasis of breast origin are extremely 

rare and have a very poor prognosis because of their resistance 

to the therapy. 

The objective of this retrospective study was to analyzed biological 

characteristics in primary breast cancer and in corresponding 

metacronous metastatic tumour. 

Methods. 21 primary invasive breast cancer and corresponding 

abdominal or pelvic recurrences were collected (1988-2009). Metastasis 

were localized: 16 in ovary, 1 in cervix, 1 in endometrium 

and 3 in omentum. 

Detection of hormonal receptors was performed on 18/21 primary 

breast cancer and on 20/21 metastatic samples. Ki67 immunoreactivity 

was valuable on 13/21 primary breast cancer and on 

19/21 metastasis. HER2 (Hercep Test) was performed on 18/21 

metastatic samples. 

Results. Twelve out of 18 (66.6.1%) primary valuable cases were 

ER+/PgR+ and 6 (33.4%) ER-/PgR-; whereas 3/20 of metastatic 

sites resulted ER+/PgR+ (15%), 5 (25%) ER-/PgR+, 3 (10%) 

ER+/PgR- and 9 (45%) ER-/PgR-. 4/13 (30.7%) primary breast 

cancer and 7/19 (36.8%) metastatic cases had an high Ki67; 

moreover, 14 metastases were HER2 negative whereas in 4 cases 

HER2 was overexpressed. 

Six patients (mean FU: 64 months; range 12-120 months) had 

follow-up data: after the first event, 5 were treated with Chemotherapy 

(CT) and Tamoxifene (TAM), whereas 1 was treated 

with Radiotherapy and TAM. Receptor expression was higher in 

primary than in secondary lesions and receptor-negative primary 

tumours showed receptor-negative recurrences. 

Our data suggest that loss of ER and PgR expression in abdomen 

and pelvic recurrent breast cancer have high incidence. Moreover, 

30% of metastatic sites evidenced a triple negative (ER, 

PgR, HER2) status suggesting that this group of patients do not 

respond to hormonal and immunologic therapy. 

references 

Ellis MJ, et al. JNCI 2008;100:1380-88. 

Musgrove EA, Sutherland RL. Nature 2009;9:631-43. 

Histopathological features and cytopathological 

correlation of thyroid nodules after percutaneous 

laser ablation 

1)Piana S. 2)Gardini G. 3)Froio E. 4)Riganti F. 5)Valcavi R. 

1)Anatomia Patologica, Arcispedale Santa Maria Nuova, Reggio Emilia, 

Italia 2)Anatomia Patologica, Arcispedale Santa Maria Nuova, Reggio 

Emilia, Italia 3)Anatomia Patologica, Arcispedale Santa Maria Nuova, 

Reggio Emilia, Italia 4)Endocrinologia, Arcispedale Santa Maria Nuova, 

339 

Reggio Emilia, Italia 5)Endocrinologia, Arcispedale Santa Maria Nuova, 

Reggio Emilia, Italia 

Background. Ultrasound-guided percutaneous laser ablation 

(PLA) is a new therapeutic approach aimed to reduce the volume 

of benign thyroid nodules causing local discomfort or cosmetic 

complaints. It is a minimally invasive and a well-tolerated procedure 

and the effects induced by the thermic energy can be 

controlled with no or minimal damage on the surrounding tissue. 

The aim of our study is to describe the histopathological features 

of the treated nodules and to correlate them with the cytological 

results before and after the PLA. 

Methods. Among 302 patients who are clinically followed after 

one or more applications of PLA, thirteeen patients (2 men and 11 

women) underwent partial or total thyroidectomy either because 

the procedure was ineffective, or because the cytological evaluation 

cannot rule out a follicular neoplasm. 

Results. On histology, the basic pattern of laser-induced lesions 

was a quite well-defined area with central cavity, filled with 

dark amorphous material and necrotic debris, surrounded by a 

capsule of fibrous connective tissue and scattered inflammatory 

cells. These histological features were in accordance with the 

cytological findings after PLA. The morphological differences of 

the ablation zones from the remaining thyroid tissue depends on 

the variable effects that PLA can induce on blood vessels, on the 

presence of different amounts of colloid areas or of connective 

tissue which my influence the energy transmission to the tissue. 

PLA can induce a significant volume reduction and an improvement 

of local symptoms without significant collateral effects; in 

the future this mini-invasive technique could be considered alternative 

to surgery in symptomatic patients with non functioning 

and functioning thyroid nodules, in patients with high surgical 

risk and, evenly, for palliative treatment in thyroid carcinoma 

or in recurrent thyroid carcinoma untreatable with surgery or 

radioiodine therapy. 

Variable activation of canonical and alternative 

Nf-Kappa B pathway in peripheral T-cell lymphoma 

not otherwise specified 

Pier Paolo Piccaluga1 , Claudio Agostinelli1 , Anna Gazzola1 , 

Maria Teresa Sista1 , Simona Righi1 , Francesco Bacci1 , Elena 

Sabattini1 , Maura Rossi, Claudia Mannu, Philip Went2 , Stefano 

A. Pileri1 1Department of Hematology and Oncology “L. and A. Seràgnoli”, Hematopathology 

and Hematology Units, S. Orsola-Malpighi Hospital, University 

of Bologna, 40138 Bologna, Italy; 2Institute of Pathology, 8063 

Zürich, Switzerland 

Background. Peripheral T-cell lymphomas not otherwise specified 

(PTCL/NOS) is the commonest subtype of PTCL. Recently, 

NFKB pathway was suggested as possibly involved in PTCL/ 

NOS molecular pathogenesis. Purpose of the study. We studied 

gene and protein expression of NFKB related molecules in PTCL 

not otherwise specified (PTCL/NOS) in order to assess 1) the 

expression pattern of NFKB among PTCLs/NOS, 2) the NFKB 

cellular localization, and 3) the prognostic relevance of NFKB 

expression. 

Methods. In order to address the above mentioned issues, we 

first studied by immunohistochemistry the expression and cellular 

localization of the NFKB effectors molecules of p50, p52, 

RELA/p65, RELB/p50, and c-REL on tissue-microarrays containing 

98 PTCL/NOS cases. In addition, we performed gene 

expression analysis of 88 PTCL cases and 20 samples of normal 

T-cells, by focusing on the expression of NFKB molecules and 

transcriptional targets. 

Results. First, we found that the vast majority of PTCL/NOS 

did express RELA, RELB, and c-REL at protein level. However, 

the 71% of cases showed only cytoplasmic expression of such 

molecules, suggesting a general shut off of both canonical and

340 

alternative NFKB pathways. On the other hand, 25% of cases 

showed an activation of the canonical pathway, while, in few instances 

(3%) the alternative pathway was activated. Interestingly, 

one case presented with both canonical and alternative pathways 

apparently activated. Double immunofluorescent staining was 

then used to confirmed the nuclear/cytoplasmic NFKB molecules 

expression in neoplastic T-cells. Noteworthy, gene expression 

profile analysis carried on a large panel of cases including most of 

those studied by IHC, confirmed such patterns. Finally, we found 

that NFκB molecules expression (either at RNA and protein 

level) did not significantly correlate with the clinical outcome. 

Conclusions. Basing on comprehensive gene expression data 

and cellular localization detected by double immunostains, we 

found that PTCLs/NOS are characterized by variable expression 

of NFΚB molecules, the activation of the pathway being apparently 

restricted to a limited fraction of cases (28%). In particular, 

the canonical, the alternative or both pathways were activated in 

25%, 3% and 1% of cases, respectively. 

Promoting quality in cytology and histology 

for oncological screening programs (uterine 

cervix, colon-rectum, breast). regional PACS 

for pathologists 

Pierotti P., Lega S., Crucitti P., Bondi A. 

Anatomia Patologica, Dipartimento Oncologico, Maggiore, Bologna, 

Italia 

Background. A quality assessment method employed in cytology 

and histology, is based on the circulation of set standard of 

slide among laboratories. As a consequence of technological evolution 

in information systems and in particular with the introduction 

of Picture Archiving and Communication System (PACS), 

we propose a complementary model based on this technology. 

Methods. The entire sample is transferred into a file, diagnosis 

is performed on a screen and a digital file replaces the slide. A 

digital slide can be considered equal to a conventional slide on a 

quality level, with the benefit that it can be reproduced in many 

copies, easily and rapidly shared on CD, DVD or on the Web. In 

order to do that, 2 Aperio® SCANSCOPE CS scanner is used, 

together with 2 servers for data storage, ImageScope TM a software 

for visualization and editing of images and SPECTRUM TM 

a software which, installed on every server, permits to share 

slides. Cytological slides have been scanned at 40X objective 

magnification, while histological slides have been scanned at 

20X objective magnification. This must be backed by software 

for the realization of network slide seminar to perform periodic 

test of diagnostic reproducibility and proficiency test. We started 

a quality assessment program for the uterine cancer screening, involving 

10 Regional Centers which provided to send 5 cytologic 

cases and corresponding histological slides or confirmation at the 

follow-up. Every case sent had to be representative of specific 

morphological cases and fit into medical report protocols. 

Results. The digital slide is especially avaiabile at a distance and 

from multiple locations simultaneously with drastic reduction of 

time needed to archieve proficienty test reproducibility.We aim 

to test individual reproducibility and evaluating the diagnostic interobserver 

concordance in the oncologic screening and building 

an online Atlas, which can be used for didactic reasons, as well 

as to perform comparisons. 

early onset and late onset carcinoma of left colon 

show different clinical, pathological and molecular 

features 

1)Pilozzi E. 2)Ciolfi A. 3)Duranti E. 4)Allevato F. 5)Giustiniani 

MC. 6)Ferri M. 7)Zardo G. 8)Ruco L. 

1)Clinical and Molecular Medicine, Sant’andrea, Roma, Italy 2)Cellular 

Biotechnologies and Haematology, Sant’andrea, Roma, Italy 3)Clinical 


and Molecular Medicine, Sant’andrea, Roma, Italy 4)Clinical and Molecular 

Medicine, Sant’andrea, Roma, Italy 5)Pathological Anatomy, San 

Giovanni Calabite Fatebenefratelli, Roma, Italy 6)Surgery, Sant’andrea, 

Roma, Italy 7)Cellular Biotechnologies and Haematology II Facult, Sant’andrea, 

Roma, Italy 8)Clinical and Molecular Medicine, Sant’andrea, 

Roma, Italy 

Background. Most sporadic colorectal carcinoma develop in an 

advanced age (late onset). However is reported that about 17% 

develop in subject ≤ 50 years. Early onset colorectal carcinomas 

are suggestive of hereditary predisposition, most occur in the 

context of Lynch syndrome and show specific features: right 

colon localization, mucinous histology, microsatellite instability 

(MSI+). However studies on early onset colorectal carcinoma 

MSI-, mainly localized on the left colon, have been reported. 

The aim of our study was to compare clinical, pathological 

and molecular feature of early onset MSI- left colorectal carcinoma 

(LCC < 50) and late onset MSI- left colorectal carcinoma 

(LCC > 70). 

Methods. 22 MSI- LCC < 50 and 21 MSI- LCC > 70 were selected. 

In all cases were evaluated: pathological (growth pattern, 

grading, stage, vascular invasion, tumor-infiltrating lymphocytes) 

and molecular features (MSI, K-RAS mutation, BRAF mutation). 

Genome methylation status of CpG islands was evaluated using 

Restriction Landmark Genome Scanning (RLGS). 

Results. LCC < 50 showed advanced stage (III or IV) and infiltrative 

pattern of growth in about 60% of cases compared to 

28% of LCC > 70. No significant differences were found in mucinous 

differentiation, grading or lymphocytic infiltrate. KRAS 

was mutated in 38% of LCC < 50 and in 19% of LCC > 70. All 

mutations but one affected codon 12. No mutation of BRAF 

were identified in either groups. RLGS showed that even if the 

number of methylation events did not differ between the 2 groups 

however the sequences of DNA methylated in each group were 

different. 

Conclusions. Early onset left colorectal carcinomas show a more 

aggressive pathological and clinical features. The higher percentage 

of KRAS mutation supports this hypothesis. Moreover the 

different pattern in DNA methylation between the two groups 

suggest that they may differ in their molecular pathogenesis. 

Thyroid fine-needle aspiration classification 

system: our experience using terminology and 

morphologic criteria similar to that proposed in 

bethesda system 

1)Pinarello A. 2)Gasparoni P. 3)Giordano G. 4)Lo giudice C. 

5)Rizzo A. 

1)Anatomia ed Istologia Patologica, S. Giacomo, Castelfranco Veneto, 

Italia 2)SSD Endocrinologia, S. Giacomo, Castelfranco Veneto, Italia 

3)SSD Endocrinologia, S. Giacomo, Castelfranco Veneto, Italia 4)SSD 

Endocrinologia, S. Giacom, Castelfranco Veneto, Italia 5)Anatomia ed 

Istologia Patologica, S. Giacomo, Castelfranco Veneto, Italia 

Background. The objective of this study was to report our experience 

in using this reporting system to review the distribution of 

diagnosis categories and to evaluate the specificity of the system 

based on the cytologic-histologic correlation. 

Methods. A total of 3134 thyroid nodules underwent FNA, that 

is, 3134 FNAs from 2864 patients were examined at our institution 

between july 1, 2004 and february 28, 2009. All FNAs were 

classified prospectively into unsatisfactory, benign, indeterminate 

(cells of undetermined significance), follicular neoplasm (FN), 

suspicious for malignancy, and positive for malignancy. The 

IND category was used for 2 subsets of cases: (a) those that morphologically 

fall into the gray zone between adenomatoid nodule 

and FN, for Hurthle cell nodule (hyperplasia vs neoplasm), and 

chronic lymphocytic thyroiditis with concern for neoplasia; and 

(b) for suboptimal specimens due to low epithelial cellularity or 

collection artifacts.


Results. The distribution of these categories from 3134 evaluated 

nodules was as follows: 627 (21.9%) unsatisfactory, 1697 

(59.4%) benign, 248 (8.6%) indeterminate, 142 (4.9%) FN, 51 

(1.8%) suspicious, and 99 (3.5%) malignant. Patients who underwent 

surgery with suspicious and malignant diagnosis were 

as follows: 47 (91%) suspicious, and 87 (88%) malignant: the 

positive predictive value for suspicious diagnosis was 90%. False 

positive rate for malignant diagnosis was 0%. Cases with malignant 

diagnosis were representative of: 67 papillary carcinoma, 

5 follicular carcinomas, 5 medullary carcinomas, 5 anaplastic 

carcinomas, 3 non-Hodgkin lymphomas and one metastatic 

tumor. At this time, no malignancy are found in cases with indeterminate 

category. Our results shown an excellent association 

between the categories and in predicting benign vs malignant 

thyroid nodules. 

Ileal adenocarcinoma associated with jejunal 

gastrointestinal stromal tumor in a patient 

with neurofibromatosis 1 

Pireddu A., Bellezza G., Guerriero A., Cavaliere A. 

Institute of pathological anatomy, Santa maria della misericordia, Perugia, 

Italy 

Background. Patients with neurofibromatosis 1 (NF-1) are at 

increased risk of developing various tumours, particularly gastrointestinal 

stromal tumor (GIST), instead the coexistence of neurofibromatosis 

with small-bowel adenocarcinoma is exceedingly 

rare. We present a case of ileal adenocarcinoma and jejunum 

GIST in a patient with NF-1. 

Methods. A 75 years old woman with medical history of NF-1 

was admitted to the Surgical Department with acute abdomen. 

Abdominal computed tomography demonstrated a solid round 

mass of 3 cm. in diameter in the ileum. Exploratory laparotomy 

revealed an area of stenosis in association with a jejunum small 

serosal nodule. Eight centimetres of ileum were resected and 

submitted to our department. On opening the bowel, an ulcerating 

stenosing lesion was observed. The histologic examination 

showed it to be a moderately differentiated adenocarcinoma infiltrating 

the intestinal wall up to the serosa. The jejunum serosal 

nodule was shown to consist of relatively uniform spindle cells, 

with moderate atypia and with mitotic rate 1/50 HPF. The cells 

were immunoreactive for CD117, CD34 and vimentin, while they 

were negative for smooth muscle actin. The histologic and immunohistochemical 

features were consistent with GIST. 

Results. Involvement of gastrointestinal tract in NF-1 varies 

from 10% to 25% with majority of the neoplasms located in the 

small intestine (72%). The frequency of the GIST in NF1 is about 

6,5% and common genetic points mutations between the two 

entities have been found. On the other hand the adenocarcinoma 

of small-bowel in patients with NF-1 is very rare with only eight 

cases described 1 and until now an oncogenic relationship, such as 

described for GIST, has not been demonstrated. The pathogenic 

mechanism proposed for intestinal carcinogenesis is the longer 

transit time of the intestinal flow due to NF-1. 

references 

1 Stratopoulos C, et al. Eur J Canc Care 2009;18:466-9. 

Her2 overexpression in gastric cancer: 

a comparison between surgical and bioptic 

specimens 

1)Pirrelli M. 2)Valentini AM. 3)Di maggio M. 4)Armentano R. 

5)Caruso ML. 

1)Anatomia Patologica, IRCCS De Bellis, Castellana Grotte, Italia 2)Anatomia 

Patologica, IRCCS De Bellis, Castellana Grotte, Italia 3)Anatomia 

Patologica, IRCCS De Bellis, Castellana Grootte, Italia 4)Anatomia Patologica, 

IRCCS De Bellis, Castellana Grotte, Italia 5)Anatomia Patologica, 

IRCCS De Bellis, Castellana Grotte, Italia 

341 

Background. The ToGA trial, showed that Herceptin, a monoclonal 

antibody against HER2-protein, has been proven to be 

efficient in patients with metastatic gastric cancer with HER2 

overexpression (IHC 3+ or IHC 2+/FISH+). The overall survival 

raised from 11.1 to 13.8 months. A HER2-scoring system modified 

from the protocol in breast cancer was used. 

Methods. Seventy surgical specimens and 13 endoscopic biopsies 

from 72 patients with cancer of the stomach or gastroesophageal 

junction (GJ) were tested for HER2 immunohistochemical 

overexpression by using the polyclonal antibody DAKO. In 11 

patients either surgical or bioptic specimen were tested. 

The 70 surgical samples were: 40 intestinal, 22 diffuse and 8 

mixed and special type; 3 cases were from GJ. The 13 bioptic 

specimens were intestinal type only; 2 were from GJ. 

The HER2 expression was scored negative (0/+) or positive 

(++/+++) according with Hofmann criteria and some modifications 

recently added in a consensus meeting held in Catania. 

Results. The overall prevalence of HER2 positive cases was 25/83 

(30.1%); the 3+ samples were 10 (12%). There were 17/70 (24,3%) 

positive surgical and 8/13 (61,5%) positive bioptic specimens: the 

3+ samples were 7/70 (10%) and 3/13 (23%), respectively. 

Histological type: 22/53 (41,5%) intestinal type cancers were 

positive. Only 1/22 (4,5%) diffuse and 2/8 (25%) mixed and 

special type were positive. All the 3+ cases, either surgical or 

bioptic, were intestinal type. All the five GJ cancer were positive, 

two of them were 3+. 

Finally, the comparison between the eleven surgical and bioptic 

cases showed this finding: 7 negative surgical cases when evaluated 

on biopsies were negative in 5 cases, positive in 2. All 4 

positive surgical cases were positive also in bioptic specimens. 

Considering the results on the surgical specimens as the gold 

standard the sensitivity was 100%, the specificity 71,4%, the 

positive predictive value 66,7%. 

A rare case of primitive leiomyosarcoma 

of the conjunctiva 

Piscitelli D., Rossi R., Palumbo M., Fiore MG., Resta L. 


Background. Leiomyosarcomas are malignant tumors of smooth 

muscle origin and can occur in any anatomic site. Primary ocular 

leiomyosarcomas are extremely rare tumors. Although a few 

primary ocular leiomyosarcomas have been described in the 

literature, there is only one case previously reported of primary 

leiomyosarcoma of the conjunctiva. We present an additional 

case in a 57-year-old woman who developed an enlarging conjunctival 

lesion at the lower fornice of the left eye. 

Materials and methods. Gross appearance was of a large polipoid 

mushroom-like friable mass measuring 6,5 × 6x4 cm adherent to 

the fornical and bulbar conjunctiva. Histologic examination revealed 

a malignant tumor composed of cells predominantly epithelioid 

shaped, admixed with a smaller number of spindle cells. The 

neoplastic cells had an abundant lightly eosinophilic cytoplasm, 

or clear-cell appearance, were moderately pleomorphic and had 

a high mitotic index (> 10 × 10 HPF), including some atypical 

forms. A few multinucleated giant tumor cells were observed. The 

predominant pattern of growth was in the form of nodules and 

broad ill-defined intersecting fascicles. There were small necrotic 

foci. The neoplastic cells stained positively with vimentin, calponina, 

actina m.l and reacted negatively with CKpool, S100, HMB45, 

CD99, CD20, CD79, CD3,CD138. Electron microscopy disclosed 

abundant thin cytoplasmic filaments with dense bodies. 

Results. We have reported an unusual tumor of the conjunctiva 

in which the combined morphological, immunohistochemical and 

ultrastructural studies have contributed in clarifying the diagnosis 

of the leiomyosarcoma. Tipically, this tumors occurs in older 

female patients and are a high incidence of local recurrence and 

distant metastasis.

342 

Primary synovial sarcoma of kidney: a case report 

1)A. Pitino, 1)D. Ricci, 1)E. Feyles, 2)M. Graziano, 2)F. Bardari, 

3)F. Cesarani, 4)C. Spairani, 5)K. Bei, 5)K.M. Murphy, 6)S. 

Squillaci 

1)Divisione di Anatomia Patologica, Cardinal Massaia, Asti, Italia; 2)Divisione 

di Urologia, Cardinal Massaia, Asti, Italia; 3)Divisione di Radiologia, 

Cardinal Massaia, Asti, Italia; 4)Divisione di Anatomia Patologica, 

Novi Ligure, Italia; 5)Dipartimento di Patologia, John Hopkins Medical 

Institutions, Baltimore, Usa; 6)Divisione di Anatomia Patologica, Ospedale 

di Vallecamonica, Esine, Italia 

Introduction. Synovial sarcoma (SS) is a rare high-grade soft 

tissue tumor that most commonly occurs in para-articular regions 

of the extremities. These tumors have been described in other 

unusual locations, including the pleura, lung and mediastinum. 

SS is usually divided into three different subgroups depending on 

histologic appearance: biphasic SS, composed of epithelial-like 

and spindle cells, monophasic SS, with only spindle cell component 

and poorly differentiated SS, with variable histomorphological 

appearance. 

We describe the clinicopathologic, immunohistochemical and 

molecular features of a case of SS, located in the kidney. 

Methods. A 67 year old man underwent preoperative clinical 

assessment in the Surgical Division for pulmonary adenocarcinoma. 

Abdominal computed tomographic scans (CT) disclosed 

a 6,7 cm partly cystic asymptomatic mass in the left kidney with 

polylobular solid areas. One year later the patient presented with 

episodes of gross hematuria. Subsequent ultrasound study and 

CT showed remarkable enlargement of the left kidney mass. 

The patient subsequently underwent a left nephrectomy. Seven 

months later CT detected intra-abdominal tumor recurrence with 

iliopsoas nodules. 

Results. The kidney weighed 327 g. and measured 11 × 6x4 cm. 

There was a single, firm, yellow-gray to brown-colored, variegated 

partly cystic mass in the lower pole measuring 4 cm. in 

its maximum diameter. Histologically, the tumor was highly 

cellular with irregularly fasciculated, monotonous spindled 

cells with sparse mitoses and moderate to severe nuclear atypia. 

Immunohistochemical studies demonstrated the tumor cells to 

be reactive for vimentin, CD99 and Bcl-2 and focally faintly 

for EMA and cytokeratins (AE1/AE3, Cam 5.2). There was no 

reactivity for other antibodies. Ki 67 index was high (60%). 

RT-PCR was carried out and SYT/SSX2 fusion transcript was 

detected. 

Conclusions. Primary SS of kidney is a rare entity firstly described 

in 1999. Diagnosis is difficult due to the rarity of this 

lesion and its similar presentations as compared to other renal 

tumors. The differential diagnosis includes adult Wilms tumor, 

sarcomatoid renal cell carcinoma, hemangiopericytoma, congenital 

mesoblastic nephroma and primitive neuroectodermal tumor. 

Surgical treatment is considered the procedure of choice and the 

value of chemotherapy is to be proven but SS may be sensitive 

to high doses of ifosamide and doxorubicin based regimens. An 

accurate diagnosis including cytogenetic and/or molecular studies 

is mandatory. 

Continuous, high-throughput and rapid tissue 

processing: reality in udine 

1)Pizzolitto S. 2)Guarrera G.M. 3)Angione V. 4)Arpinelli S. 

5)D’Alessandro E. 6)De Maglio G. 7)Falconieri G. 8)Rocco M. 

9)Zagami M. 10)Morales A.R. 

1)SOC Anatomia Patologica, Az. Ospedaliero-Univ. S.M.della Misericordia, 

Udine, Italia 2)Unità per la valutazione delle tecnologie sanitarie, 

Az. Ospedaliero-Univ. S.M.della Misericordia, Udine, Italia 3)SOC Anatomia 

Patologica, Az. Ospedaliero-Univ. S.M.della Misericordia, Udine, 

Italia 4)SOC Anatomia Patologica, Az. Ospedaliero-Univ. S.M.della Misericordia, 

Udine, Italia 5)SOC Anatomia Patologica, Az. Ospedaliero- 

Univ. S.M.della Misericordia, Udine, Italia 6)SOC Anatomia Patologica, 


Az.Ospedaliero-Univ. S.M.della Misericordia, Udine, Italia 7)SOC Anatomia 

Patologica, Az. Ospedaliero-Univ. S.M.della Misericordia, Udine, 

Italia 8)SOC Anatomia Patologica, Az. Ospedaliero-Univ. S.M.della Misericordia, 

Udine, Italia 9)SOC Anatomia Patologica, Az. Ospedaliero- 

Univ. S.M.della Misericordia, Udine, Italia 10)University of Miami Miller 

School of Medicine, Miami, Usa 

Background. In Anatomic Pathology, changes toward automatization 

have reluctantly been accepted in the last few years. 

Although Continuous-Throughput Processing (CTP) is now 

available, Conventional Processing (CP) and laboratory organization 

tend to remain unchanged. Since 2004, on the base of Miami 

experience, our group appraised the opportunity of introducing 

an innovative CTP system with important clinical advantage for 

patient management. 

Methods. A Health Technology Assessment study was performed 

to analyze the Sakura technology and new possible workflows for 

quality improvement, turnaround-time (TAT) reduction, procedure 

standardization added to potential replacing of formalin by 

molecular fixatives. On February 2010 Sakura LEAN equipment 

(Tissue-TEC ® Xpress ® , AutoTEC ® and Prisma ® ) was introduced 

in our routine practice. All specimens were formalin-fixed. We 

analyzed CTP impact evaluating laboratory TAT in a time of 2 

months (March-April) compared with the same period of 2009. 

Technicians team was equally represented and case mix was 

analogous in number and type; 8100 cassettes were processed 

with an average of 180/day. 

Results. CTP reduced processing time from approximately 

10 hours of CP to 2 hours. Embedding time decreased from 

an average of 6 to 2 hours/day. With a continuous workflow, 

slides were available for microscopic examination in about 4 

hours. Quality of histologic slides, histochemical and immunohistochemical 

stains, FISH testing and DNA preservation 

were comparable with that of CP. Laboratory TAT decreased 

from 4.5 to 2.3 days for large specimens and from 2.1 to 1.4 for 

small biopsies. 

Our experience represents an interesting preliminary step to a 

successful implementation of express technologies. Team participation, 

motivation and a substantial change in organization habits 

are essential for the expected optimal outcome along to the LEAN 

principle in the near future. 

Multidrug resistance related proteins (MrPS) 

expression in primary breast tumor and 

circulating tumor cells (CTCs) 

1)Porta N. 2)Gradilone A. 3)Leopizzi M. 4)Cacciotti J. 5)Di Cristofano 

C. 6)Gazzaniga P. 7)Cortesi E. 8)Della Rocca C. 9)Naso 

G. 10)D’Amati G. 

1)Department of experimental medicine, Sapienza university of rome, 

Rome, Italy 2)Department of experimental medicine, Sapienza university 

of rome, Rome, Italy 3)Department of experimental medicine, Sapienza 

university of rome, Rome, Italy 4)Department of experimental medicine, 

Sapienza university of rome, Rome, Italy 5)Department of experimental 

medicine, Sapienza university of rome, Rome, Italy 6)Department of experimental 

medicine, Sapienza university of rome, Rome, Italy 7)Department 

of experimental medicine, Sapienza university of rome, Rome, Italy 

8)Department of experimental medicine, Sapienza university of rome, 

Rome, Italy 9)Department of experimental medicine, Sapienza university 

of rome, Rome, Italy 10)Department of experimental medicine, Sapienza 

university of rome, Rome, Italy 

Background. Metastatic breast cancer still represents an incurable 

disease. The cancer cells in peripheral blood, circulating 

tumor cells (CTCs), are probably an early sign of metastatic 

tumor. The expression of ATP-binding cassette (ABC) transporters 

on CTCs is predictive of response to chemotherapy in cancer 

patients. We tested the hypothesis that ABC transporters (MRP-1 

and MRP-7) expression in primary tumors and in CTCs might 

have predictive value in breast cancer (BC). 

Methods. We evaluated the multidrug resistance related proteins


(MRP-1 and MRP-7) RNA expression in primary tumors and 

CTCs in BC patients. We investigated the correlation of MRPs 

expression between primary tumors and CTCs. Furthermore we 

studied the prognostic value of MRPs expression. 

Results. The CTCs were found in 77% of BC patients. The MRP- 

1 expression was found in 44% of primary tumors and 11% of 

CTCs. The MRP-7 expression in primary tumors and CTCs was 

88% and 66%, respectively. We found a correlation between the 

MRPs expression in primary tumors and CTCs. The presence of 

CTCs and the expression of MRPs is predictive of response to 

chemotherapy. 

Availability of a new formalin-free fixative green 

fix for morpho-molecular purposes 

Postiglione M., Maglione A., Russo A., Nicastro A., Oppressore 

D., Maione M.P., Giannatiempo R. 

U.O.S. Anatomia Patologica, Osepdale Evangelico Fondazione Betania, 


Background. Despite excellent morphology for routine diagnostics, 

NBF is a poor preserver for nucleic acids and facilities the 

formation of protein-protein crosslinnks rendering tissue refractory 

to many protein studies. 

Aim. We evaluated an alternative and novel no toxic fixative, 

Greenfix regarding its effects on histomorphology, as well as on 

preservation of protein immunoreactivity and quality of genomic 

DNA. 

Materials and methods. Parallel tissue blocks from 60 breast 

cancers were fixed in 10%NBF and in Greenfix for 24h at RT and 

4-µm-thick sections were prepared for routine HE. To evaluate 

histomorphology, special attention was paid to the overall pattern 

of tissue preservation, cellular and extracellular structures as well 

as to cell and nuclear morphology and tinctorial of cell component. 

Immunostainig was performed automatically with Benchmark 

Ventana for ER, PR, HER2, KI67,E-Caderin.The intensity 

pattern and specifity of the immunohistochemical reactions were 

assessed and compared on all slides. HER2 gene amplification 

was evaluated by FISH dual color (PathVysion,Vysis). 

Results. HE stained slides of Greenfix and NBF fixed tissue 

showed no significant differences in tissue architecture, cellular 

and nuclear morphology or tinctorial reaction.Greenfix preserved 

tissue integrity and showed a better detail than chromatin.Although 

Greenfix required some optimization of immunostaining 

procedures including antigen retrieval, the % of stained cells was 

similar for ER, PR and HER2 as compared to NBF fixative. Succefful 

immunostainig was also obtained for KI67 and E-Caderin. 

Furthermore, HER2 gene amplification could be performed by 

FISH using Greenfix suggesting DNA integrity and preservation. 

Conclusions. Greenfix showed great potential for performing 

both morphological and molecular analysis on the same fixed 

tissue sample and so could represent an easy to use alternative 

to NBF compatible with both current diagnostic pathology and 

tissue ancillary investigations. 

fOXP3 expression in aggressive thyroid carcinom 

1)Ugolini C., 2)Proietti A., 1)Pelliccioni S., 2)Basolo F. 

1) Anatomia patologica sperimentale, Medicina di laboratorio e diagnostica 

molecolare, Ospedale S. Chiara, Pisa, Italia; 2) Anatomia patologica 

sperimentale, Dipartimento di Chirurgia, Università di Pisa, Pisa, Italia 

Background. Thyroid carcinoma (TC) is increasing in prevalence 

in the last years. Anaplastic thyroid carcinoma (ATC) are 

estimated to comprise 1-2% of thyroid malignancies. Unfortunately, 

their rapid onset and awful course have not altered their 

detection or outcomes. 

The immune system plays an important role in the tumor progression 

of different type of cancer. Thyroid cancer shows an inflammatory 

cell infiltrate whose role is still not completely understood. 

343 

An important immunitary factor, observed in several kind of cancers 

is the forkhead transcription factor Foxp3, that plays an important 

role in regulatory T cell (Treg) function and it is the only 

marker of CD4+CD25+ Treg. It has been reported that Foxp3 is 

highly expressed in differentiated thyroid tumors and it has been 

correlated with more aggressive disease. 

Methods. 40 cases of aggressive TC (8 papillary (PTC), 11 

poorly differentiated (PDC), 21 ATC), comparable for gender, 

age and tumor size, were considered. All PTC and PDC cases had 

developed lymph-node and/or distant metastasis. All TC were 

analyzed with immunohistochemistry for Foxp3. The Foxp3 expression 

were evaluated in epithelial tumor cells. For all patients 

clinico-pathological features were considered and the results 

analyzed by statistical tests. 

Results. TC were categorized, on the basis of the presence of 

areas of differentiation, in two groups: differentiated cancer 

(DC) and not-differentiated cancer (NOT-DC). Comparing the 

Foxp3 expression in the 2 groups we found a significative presence 

in DC (p = 0.004). Moreover we found a high correlation 

between the presence of Foxp3 in tumor tissue and its extrathyroid 

infiltration (p = 0.003). The Foxp3 expression in tumor 

tissue was not correlated with other clinico-pathological features 

including distant metastasis. In conclusion, Foxp3 acts at local 

level in differentiated cancer but it does not play a role in tumor 

progression. 

sClu, VeGf-A165 And Il-6 in human colon 

carcinogenesis: a molecular network leading 

to cell death escape through micrornas 

dysregulation 

1)Pucci S. 2)Mazzarelli P. 3)Zonetti M.J. 4)Spagnoli L.G. 

1)Biopatologia, Università di Roma Tor Vergata, Roma, Italy 2)Biopatologia, 

Università di Roma Policlinico di Tor Vergata, Roma, Italy 3)Biopatologia, 

Università di Roma Policlinico di Tor Vergata, Roma, Italy 4)Biopatologia, 

Università di Roma Policlinico di Tor Vergata, Roma, Italy 

Background. Cooperation through the sharing of diffusible factors 

of tumor microenvinoment and the redirection of some specific 

guardian pathways raises new questions about tumorigenesis 

and has implication on designing new therapeutic approaches. 

Recent studies suggest a potential role of IL6-sIL6R in the 

pathogenesis of colon cancer, although data are still conflicting. 

Increased formation of IL6-sIL6R complexes that interact with 

gp130 on the cell membrane leads to increased expression and 

nuclear translocation of STAT3, which induces VEGFexpression 

and the activation of anti-apoptotic genes, such BclxL. Methods. In human colorectal carcinomas (n = 50) at different 

stages of disease we observed, by IHC, an increase of IL-6 and 

VEGF165A production correlated to the aggressiveness of the tumor. 

The IL6 and VEGFA165 in vitro treatment of colon cancer 

cell lines, modulated the expression of genes involved in tumor 

invasion and apoptosis, as observed by microarrays. 

Results. In particular, IL-6 downmodulated Bax expression at 

mRNA level. Concomitantly, IL-6 exposure influenced Bax also 

at protein level acting on the Bax-Ku70-sCLU physical interactions 

in the cytoplasm, by affecting the Ku70 acetylation and 

phosphorylation state. Moreover, we demonstrate that IL6 together 

with VEGF are able to inhibit Bax-dependent cell death also by 

increasing the production of the pro-survival form of Clusterin, 

shifting death into survival. Strikingly we observed that the cooperation 

between IL-6 and VEGFA influenced the expression 

of tumor suppressing miRNAs affecting the epigenetic HDAC1 

activity and the epithelial to mesenchymal transition, turning 

the neoplastic cell from epithelial to mesenchimal, strongly correlated 

to the malignization of many types of cancers.These still 

obscure molecular interactions underlie the relevant role of these 

microenvironmental factors, in the complicated cross talk among 

molecules that could effectively turn the cell fate.

344 

Solitary extramedullary plasmacytoma of the 

thyroid gland associated with multinodular goiter: 

a case report and review of the literature 

1)Puliga G. 2)Olla L. 3)Bellisano G. 4)Di naro N. 5)Ganau M. 

6)Lai M.L. 7)Faa G. 8)Tolu G.A. 

1)U.O. Anatomia Patologica, San Martino, Oristano, Italia 2)U.O. Anatomia 

Patologica, San Martino, Oristano, Italia 3)U.O. Anatomia Patologica, 

San Martino, Oristano, Italia / Anatomia Patologica - Università di 

Cagliari, San Giovanni di Dio, Cagliari, Italia 4)U.O. Anatomia Patologica, 

San Martino, Oristano, Italia 5)S.C. Neurochirurgia, Cattinara, Trieste, 

Italia 6)Anatomia Patologica - Università di Cagliari, San Giovanni 

di Dio, Cagliari, Italia 7)Anatomia Patologica - Università di Cagliari, 

San giovanni di Dio, Cagliari, Italia 8)U.O. Anatomia Patologica, San 

Martino, Oristano, Italia 

Background. Solitary Extramedullary Plasmacytoma (SEP) is 

a rare malignant neoplasm arising from plasma cells, most commonly 

occurring in the nasal cavity, nasopharynx and larynx. 

Thyroid involvement is rare: less than 75 cases of SEP of the 

thyroid gland have been reported to date. 

Methods. A 74 year old woman, with an history of multinodular 

goiter, presented with dysphonia and a painful neck swelling, related 

to a rapidly growing nodule in the right thyroid lobe. Thyroid 

function tests showed a subclinical hypothyroidism; no evidence 

of Hashimoto’s disease was found. Ultrasound scan confirmed 

the presence of an isoechoic nodule, 35 mm in diameter, with 

CDIII vascular pattern. FNAC showed a monotonous population 

of atypical cells, interpreted as suspicious for malignant neoplasia 

(Thy4). The patient underwent total thyroidectomy. 

Results. Histopathological examination showed an unencapsulated 

neoplasm composed of atypical tumor cells characterized by abundant 

cytoplasm and eccentric nuclei. At immunohistochemistry, 

tumor cells revealed diffuse reactivity for CD138 and CD45RB and 

predominant staining for kappa chains. Pan-cytokeratins, TTF1, 

Thyreoglobulin, Calcitonin, CD20 and CD79a were negative. 

Clinically, a complete multiple myeloma workup was negative. 

On this basis, the conclusive diagnosis of SEP was made. At a 12 

months follow-up, the patient refers good clinical conditions without 

evidence of multiple myeloma. In conclusion, SEP should be 

considered in the differential diagnosis of a rapidly enlarging thyroid 

nodule. Clinical correlation and immunocytochemistry are crucial in 

avoiding pitfalls. Surgery indeed remains the best modality of treatment 

whenever the lesion is localised and easily removable. 

references 

Avila A, et al. Clinical features and differential diagnoses of solitary 

extramedullary plasmocytoma of the thyroid: a case report. 

Ann Diagn Pathol 2009;13(2):119-23. 

Kuo SF, et al. Extramedullary plasmocytoma of the thyroid. N Z 

Med J 2006;119(1235):U2005. 

Orbital cellular fibroblastic/myofibroblastic 

neoplasm 

1)Pusiol T. 2)Morichetti D. 3)Piscioli F. 

1)Institute of anatomic pathology, S.maria del carmine hospital, Rovereto, 

Italy 2)Institute of anatomic pathology, S.maria del carmine hospital, 

Rovereto, Italy 3)Institute of anatomic pathology, S.maria del carmine 

hospital, Rovereto, Italy 

Background. Fibroblastic/Myofibroblastic Tumours (FMTs) 

represent a large subset of mesenchymal tumours. Many lesions 

in this category contain cells with both fibroblastic and myofibroblastic 

features, which may represent functional variants of a 

single cell type. We report an unusual orbital FMT. 

Materials and methods. In January 2010 a 66-year-old man presented 

with a 3-months history of proptosis and decreased visual 

acuity. Computed Tomography (CT) and complete removal of the 

tumour were performed. After 5 months of follow-up the patient 

is free of disease. 


Result. The CT showed retrobulbar mass. Grossly the tumour 

was tan-ivory in color and measured 2.2 × 1.5 × 1 cm. Histologically 

the neoplasm was composed by spindle cells with tapering 

nuclei and indistinct palely eosinophilic cytoplasm. For the most 

part the lesions have a well-developed fascicular growth pattern. 

The patternless architecture and varying cellularity of solitary 

fibrous tumour were not found. There was no significant atypia 

nor pleomorphism and mitoses were not frequent. Immunostains 

show multifocal positivity for SMA and CD34, while S-100 

protein, GFAP and desmin are negative. No convincing features 

of malignancy were present. The diagnosis of “cellular FMT” 

was performed according to Prof. Christopher D.M. Fletcher’s 

consultation. 

Discussion. Mixofibrosarcoma 1 , solitary fibrous tumour 2 and 

inflammatory myofibroblastic tumour 3 are well-knowed orbital 

neoplasm with predominant myofibroblastic spindle cells component, 

accompagned by fibroblastic and other cellular types. 

The present case not showed typical proliferative pattern of typical 

orbital FMT. The label “cellular fibroblastic/myofibroblastic 

tumor” is a descriptive terminology because the growth pattern 

was not classified in the category of other well-known orbital 

neoplasm tumours with similar histogenesis. Further studies are 

necessary to establish if this neoplasm is a new entity of orbital 

FMTs. 

references 

1 Zhang Q, Wojno TH, Yaffe BM, et al. Myxofibrosarcoma of the 

orbit: a clinicopathologic case report. Ophthal Plast Reconstr Surg 

2010;26:129-31. 

2 Brunnemann RB, Ro JY, Ordonez NG, et al. Extrapleural solitary 

fibrous tumor: a clinicopathologic study of 24 cases. Mod Pathol 

1999;12:1034-42 

3 Ahmad SM, Tsirbas A, Kazim M. Inflammatory myofibroblastic tumour 

of the orbit in a 7-year-old child. Clin Experiment Ophthalmol 

2007;35:160-2. 

Peritoneal malignant psammomatous 

mesothelioma 

D.Morichetti, M.G. Zorzi, T. Pusiol, F. Pisciol 

Istituto di Anatomia Patologica, Ospedale S. Maria del Carmine, Rovereto, 

Italia 

Background. Psammoma Bodies (PBs) are concetrically laminated 

calcific spherules that occasionally appear cracked (psammos 

[“sand”] + oma [“tumor”]). PBs are observed in malignancies 

and in a number of benign non-neoplastic conditions. We 

report one case of peritoneal Malignant Mesothelioma (MM) with 

massive deposition of PBs with emphasis to diagnostic differentiation 

with similar neoplasms. 

Matherial and Methods. A 72 years-old man presented with 

abdominal swelling and marked weight loss. A cytologic analysis 

of ascites was performed. The explorative laparoscopy showed 

diffuse minute parietal peritoneal nodules. No history of exposure 

to asbestos was found. 

Results. The cytology of peritoneal cavity effusion showed 

malignant cells. The peritoneal biopsy revealed a superficial 

papillary growth of malignant epithelial-like cells with diffuse 

involvement of submesothelial tissues. Massive deposition 

of PBs was observed. Nuclear and cytoplasmic calretinin 

immunoreactivity was present in neoplastic cells, along with 

membranous D2-40 and membranous/cytoplasmic cytokeratin 

5 staining. 

Discussion. PBs may be seen in approximately 5% to 10% cases 

of peritoneal MM, especially in the well differentiated papillary 

MM 1 2 . To date massive deposition of PBs have not been reported 

in peritoneal MM. The pathogenesis of extensive presence of PBs 

is unknown. We believe that single necrotic cells constitute seed 

crystals that become incrusted with the mineral deposits and the 

progressive acquisition of outer layers may create its lamellated


configurations. The neoplasm may simulate serous psammocarcinoma 

of the peritoneum. The immunophenotipic profile was 

conclusive of MM. The behaviour of serous psammocarcinoma 

is more closely silimar to borderline serous tumor than of serous 

carcinoma. The presence of PBs should have favourable impact 

to prognosis. Further studies are necessary to estabilish if massive 

deposition of PBs may define a new variant of MM with better 

prognosis. 

references 

1 Daya D, McCaughey WT. Well-differentiated papillary mesothelioma 

of the peritoneum. A clinicopathologic study of 22 cases. Cancer 

1990.15;65:292-6. 

2 Attanoos RL, Gibbs AR. Pathology of malignant mesothelioma. Histopathology 

1997;30:403-18. 

A significative immunohistochemical application 

for therapeutic management about a case of skin 

dermatofibrosarcoma protuberans with lung 

metastasis 

1)A. D’Amuri, 2)G. Quarta, 2)L. Paolelli, 2)G. Scavelli, 1)F. 

Floccari, 1)L. Aiello, 1)M.R. Pede, 1)S.A. Senatore 

1)U.O.C. Anatomia Patologica, “Ospedale Sacro Cuore di Gesù”, P.O. 

Gallipoli ASL Lecce, Gallipoli, Italia; 2)U.O. Oncologia, “Ospedale Sacro 

Cuore di Gesù”, P.O. Gallipoli ASL Lecce, Gallipoli, Italia 

Background. A 59 year-old-man (Albany) underwent skin 

nodule biopsy in the deltoid area with an unknown histological 

diagnosis. 8 months later he came in Italy and was admitted in 

the Oncology Division for a mediastinic syndrome, mediastinal 

lymphoadenomegaly and lung multiple nodules. A mediastinal 

radiotherapy was needed. After a symptom-free interval a new 

lesion occured in the scar of the previous operation and the biopsy 

confirmed a Dermatofibrosarcoma Protuberans (DFSP). Bronchial 

biopsies revealed a lung DFSP metastasis. 7 months later 

two recurrencies, one in the same deltoid region and the other one 

in the scalp, were diagnosed. 

Methods. Immunohistochemical stains (IHC) were performed for 

alpha-1AT, BCL2, CD20, CD45, CK AE1/AE3, Desmin, Ki-67, 

Lisozime, S-100, Vimentin in skin and lung tissues samples. 

Results. DFSP is a rare nodular cutaneous tumor with tendency 

to recur locally after excision and low metastatic potential. DFSP 

is characterized by specific chromosomal abnormalities involving 

platelet derived growth factor B locus (PDGFB). In current 

literature there are only a few reports of surgical excision of 

pulmonary metastases and cure has rarely been achieved despite 

resection. In our study we compared IHC and morphological 

patterns in skin and bronchial biopsies. Histologic examination 

showed spindle cells in a storiform patterns, as well as multinucleated 

tumor giant cells and few mitotic figures. IHC revealed 

a strong vimentin and alpha-1AT staining with low proliferative 

index Ki-67 (10%). A chromosomal abnormality mutation 

of PDGFRB was found. On the basis of our observations we 

confirmed a common origin between skin and lung lesions. The 

patient was succesfully treated with Imatinib mesylate at 800mg/ 

day, a tyrosine kinase inhibitor with activity against activated 

PDGFR in DFSP patient, for 9 months. He received an Imatinib 

manteining dose of 400 mg/day. Until now a complete remission 

was confirmed by PET/TC. 

Oncocytic carcinoma of the breast: a histological, 

immunohistochemical, ultrastructural and 

molecular study 

1)M. Ragazzi, 1)D. De Biase, 1)A. Farnedi, 2)C.M. Betts, 3)F. 

C. Geyer, 3)M.B. Lambros, 1)G. Tallini, 3)J.S. Reis-Filho, 4)V. 

Eusebi 

1)Dipartimento di Ematologia e scienze oncologiche “L. e A. Seràgnoli”, 

Sezione di Anatomia Patologica, Ospedale Bellaria, Università di Bo- 

345 

logna, Bologna, Italia; 2)Dipartimento di Patologia Sperimentale, Università 

di Bologna, Bologna, Italia; 3)The Breakthrough Breast Cancer 

Research Centre, Institute of Cancer Research, London, UK; 4)Anatomia, 

istologia e citologia Patologica, Ospedale Bellaria – Università di Bologna 

- ASL, Bologna, Italia 

Background. Oncocytic breast carcinomas (OC) are tumours 

composed of ≥70% of oncocytic cells (WHO). Purpose of this 

study is to determine the frequency, morphological and clinical 

features of invasive OC in a large series together with a molecular 

study. 

Methods. 28 putative OC (selected cases) and 76 consecutive 

invasive breast carcinomas were studied. Immunohistochemistry 

for mitochondria, GCDFP-15, chromogranin, oestrogen (ER), 

progesterone and androgen receptors, c-erb-B2, CK7, CK14, 

EMA and CD68 was performed. A semi-quantitative assessment 

based on intensity and extent of the reactivity for mitochondria 

was employed and three classes were defined: OC ≥70% 3+; mitochondrion 

rich (mt-richC) 50-70% 3+ or > 50% 2+; all the other 

cases (IBC). Ultrastructure was available for six cases of OC. For 

overall survival analysis Kaplan-Maier curves were compared 

using Wilcoxon and logrank tests (p < 0.05). Morphological features 

of the three groups were compared using Fisher’s exact test 

(p < 0.05). Ten OC, 8 mt-richC and 36 IBC matched for grade 

and ER were microdissected and analyzed with aCGH. Results 

were subjected to unsupervised hierarchical clustering analysis 

(UHCA). The patterns of copy number gains, losses and amplifications 

between OC/mt-richC and IBC were compared using a 

multi-Fisher’s exact test corrected by the false discovery rate. 

Results and conclusions. A total of 32 cases of OC was identified: 

17 from the selected cases (60.7%) and 15 from the consecutive 

cases (19.7%). OC appears to be a morphological entity 

with features distinct from those of IBC. Clinical features are not 

distinctive. UHCA based on aCGH demonstrates that OC and 

mt-richC form a cluster separate from IBC. Moreover, multi- 

Fisher’s exact test reveals that OC and mt-richC are significantly 

different from IBC in their pattern of copy number aberrations. 

Taken together our results demonstrate that OC and mt-richC 

may constitute an entity distinct from IBC at both histological 

and genomic levels. 

Her2 in gastric carcinomas: a pathological 

and clinical overview 

1)Ramieri MT. 2)Pica E. 3)Mansueto G. 4)Gamucci T. 5)Murari 

R. 6)Alò PL. 

1)Anatomia Patologica, Umberto I, Frosinone, Italia 2)Anatomia Patologica, 

Umberto I, Frosinone, Italia 3)Oncologia, Umberto I, Frosinone, 

Italia 4)Oncologia, Umberto I, Frosinone, Italia 5)Anatomia Patologica, 

Umberto I, Frosinone, Italia 6)Anatomia Patologica, Umberto I, Frosinone, 

Italia 

Background. To evaluate HER2 expression in gastric cancer in 

order to assess amplification prevalence and associations between 

clinical and pathological features. 

Methods. 160 gastric carcinomas (96 intestinal-type, 64 diffusetype) 

were enrolled in the study. All samples were evaluated with 

IHC (4B5) and SISH (HER2/Ch17) to assess HER2 gene expression 

and/or amplification. The IHC score was in accordance with 

Hofmann guidelines (2). 

Results. HER2 amplification was detected in 20 of 96 (20,8%) 

intestinal-type and in 2 of 64 diffuse-type (3%) carcinomas 

(p < 0.01). About all samples have a 3+ IHC score. Only 7 

samples showed a 2+ IHC score, of which 3 showed amplified 

status. Data revealed a prevalence of proximal tract tumors in 

amplified vs non-amplified specimens (57% vs 41%). Histotype, 

site of primary (GEJ vs Gastric) and prevalence of amplification 

(20,8%) were in accordance with previous studies (3). 

About the 53 cases (31 intestinal-type, 22 diffuse-type) with 

clinical data we have only 6 amplified cases, all of intestinal

346 

type (p < 0.05), with a prevalence in the proximal region of 

the stomach (66%) and most with distant metastasis (66%). 

An association between amplified vs not-amplified intestinaltype 

carcinomas has shown a more advanced disease stage 

(66% vs 24% stage IV, p < 0.05) and a worse survival rate 

(33% vs 56%), with or without chemotherapy. An association 

between amplified intestinal-type and diffuse-type carcinomas 

has shown a more advanced disease stage (66% vs 50%) with 

similar survival rate (33% vs 27%). A comparison between 

old and new AJCC cancer staging system was performed. Our 

study revealed 32 cases where both staging system produced 

similar results and any difference between amplified state and 

survival rate. 

These data showed a similar behaviour of amplified intestinaltype 

and diffuse-type carcinomas and suggest to consider appropriate 

a molecular morphology test to study HER2 status in all 

intestinal-type carcinomas. 

lymph node micrometastasis and survival 

of patients with stage I colorectal carcinoma 

1)Reggiani Bonetti L. 2)Di Gregorio C. 3)Pedroni M. 4)Barresi 

G. 5)De Gaetani C. 6)Ponz de leon M. 

1)Dip. Attività Integrata di Laboratori, Anatomia Patologica e Medicina 

Legale; Az. Universitaria Policlinico di Modena, Modena, Italia 2) Dip. 

Attività Integrata di Laboratori, Anatomia Patologica e Medicina Legale; 

Az. Universitaria Policlinico di Modena, Modena, Italia 3)Dipartimento 

di Medicina Interna, Az. Universitaria Policlinico di Modena, Modena, 

Italia 4)Anatomia patologica, Az. Universitaria Policlinico G. Martino, 

Messina, Italia 5)Dip. Attività Integrata di Laboratori, Anatomia Patologica 

e Medicina Legale; Az. Universitaria Policlinico di Modena, Modena, 

Italia 6) Dipartimento di Medicina interna, Policlinico di Modena, 

Italia 

Background. Patients with Stage I colorectal cancer (DUKES 

A) usually show an excellent prognosis. However, few of them 

die of metastatic disease. Occult micrometastasis in lymph node 

could explain cancer progression. In a previous study, through the 

Colorectal Cancer Registry of Modena, we selected patients with 

Stage I disease who died of metastatic tumor (25 cases) and, using 

pan-cytokeratin antibody, we detected lymph node micrometastasis 

in 18 of theme (72%). Micrometastasis were significantly 

correlated with vascular invasion and tumor budding. In order to 

reinforce our findings, these cases were matched with 70 Stage 

I patients with favourable prognosis (controls) selected from the 

same Registry. 

Methods. Likewise the previous study, resected lymph nodes 

from controls were entirely cut at 200 µm intervals, yielding 4 

µm thick sections. Alternate sections were evaluated with HE and 

IHC staining with pan-cytokeratin antibodies. 

Results. Micrometastasis were almost absent among controls (1 

of 70, 1.4%), p < 0.001 by χ2 test, vs cases. Vascular invasion 

and tumor budding were more frequent among Stage I patients 

with an unfavourable prognosis than in controls. By regression 

analysis, occult malignant cells and vascular invasion maintained 

an independent association with prognosis (C.I. 4.75-32.04 and 

1.41-8.54). In conclusion, our data confirm the role of occult micrometastasis 

in affecting clinical outcome of patients with stage 

I colorectal cancer. 

Attenuated familial adenomatous polyposis 

in Northern Italy 

1)Reggiani Bonetti L. 2)Di Gregorio C. 3)Urso E. 4)Pucciarelli 

S. 5)Pedroni M. 6)Balsamo A. 7)Laudi C. 8)Viel A. 9)Venesio T. 

10)Ponz de Leon M. 

1)Dip. Attività Integrata di Laboratori, Anatomia patologica e Medicina 

Legale; Policlinico di Modena, Italia 2)Dip. Attività Integrata di Laboratori, 

Anatomia patologica e Medicina Legale; Policlinico di Modena, 


Italia 3)Scienze Oncologiche e Chirurgiche, Az. Ospedaliero-Universitaria, 

Padova, Italia 4)Scienze Oncologiche e Chirurgiche, Az. Ospedaliero-Universitaria, 

Padova, Italia 5)Dipartimento di Medicina Interna, Az. 

Universitaria Policlinico di Modena, Modena, Italia 6)Anatomia Patologica 

e Gastroenterologia, Istituto per la Ricerca e la Cura del Cancro, 

Candiolo - Torino, Italia 7)Anatomia Patologica e Gastroenterologia, 

Istituto per la Ricerca e la Cura del Cancro, Candiolo - Torino, Italia 

8)Oncologia Sperimentale 1, Centro di Riferimento Oncologico, Aviano 

(PN), Italia 9) Anatomia Patologica e Gastroenterologia, Istituto per la 

Ricerca e la Cura del Cancro, Candiolo - Torino, Italia 10)Dipertimento 

di Medicina Interna, Az. Universitaria Policlinico di Modena, Mo 

Background. Attenuated Familial Adenomatous Polyposis 

(AFAP) is featured by the presence of 10 to 99 colorectal 

adenomatous polyps. The disease may be associated with mutations 

in either APC or MutYH genes. We purposed to assess 

the contribution of APC and MutYH constitutional alterations 

to the AFAP phenotype, and to find out genotype/phenotype 

correlations. 

Methods. As part of counselling for Familial Adenomatous Polyposis 

(FAP). 91 probands were identified meeting the criteria 

of AFAP. Genetic testing was offered to all probands, and APC 

and MutYH constitutional mutations were searched by DNA 

sequencing. 

Results. 107 affected individuals were identified. MutYH mutations 

were detected in 21 families (30.4% of the 69 tested), 

and APC mutations in 7 (10.1%). Thus, constitutional alterations 

were found in 40.5% of the probands. Patients with APC 

mutations showed an earlier age of cancer onset and a higher 

mean number of polyps (48.5 ± 33.0 vs 35.7 ± 24.9 in MutYH + 

individuals, and 33.2 ± 18.4 in the “no mutation” group). Clinical 

features rendered the “no mutation” group closer to MutYH 

+ than to the APC + group. Colorectal cancer at diagnosis was 

detected in 40% of AFAP individuals. In conclusion, AFAP is 

a new clinical entity whose frequency in the general population 

remains undefined. The number of adenomas shows a wide range 

of values, with an average of 30 to 40 lesions. The molecular basis 

of AFAP can be established in approximately half of patients; 

both MutYH and APC genes are implicated in AFAP, though the 

role of MutYH seems of major relevance. 

Cytomegalovirus infection of the upper 

gastrointestinal tract: a clinical and pathological 

study of 30 cases 

1)Reggiani Bonetti L. 2)Merighi A. 3)Losi L. 4)Bertani A. 5)Bettelli 

S. 6)Maiorana A. 

1)Dipartimento ad attività integrata di laboratori, Anatomia patologica 

e medicina legale; policlinico, Modena, Italia 2)Medicine e specialità 

mediche gastroenterologia, Ed endoscopia digestiva;policlinico, Modena, 

Italia 3)Dipartimento ad attività integrata di laboratori, Anatomia patologica 

e medicina legale; policlinico, Modena, Italia 4)Gastroenterologia, 

Ed endoscopia digestiva;policlinico, Modena, Italia 5)Dipartimento ad 

attività integrata di laboratori, Anatomia patologica e medicina legale; 

policlinico, Modena, Italia 6)Dipartimento ad attività integrata di laboratori, 

Anatomia patologica e medicina legale; policlinico, Modena, Italia 

Background. Upper gastrointestinal (UGI) tract involvement 

in Human cytomegalovirus (HCMV) infection is largely documented 

in both healthy and immunocompromised patients. Endoscopic 

findings vary from erythema to hypertrophic mucosal 

changes to ulcers. 

Methods. We reviewed 30 UGI biopsies of HCMV-infected 

patients diagnosed in the Department of Pathologic Anatomy of 

Modena in a 10-year period. Clinical and endoscopic data were 

correlated to histological findings of HCMV infection, based on 

the identification of viral inclusion bodies in routine hematoxylin 

and eosin-stained sections and confirmed by immunohistochemical 

staining using primary anti-HCMV monoclonal antibodies 

(Clone CCH2-DAKO, Glostrup, Denmark).


Results. There were 20 male and 10 female patients, aged 27 to 

91 years; 10 were immunocompromised (6 HIV+, 3 with liver 

transplantation and 1 with renal transplantation); 4 had malignancies 

previously treated with surgery and chemotherapy (2 

gastric MALT lymphoma, 1 bile duct carcinoma, 1 hepatocellular 

carcinoma) and 1 was affected by common variable immunodeficiency. 

Mucosal alterations were endoscopically observed in 

the stomach (19 cases), esophagus (9), cardias region (6) and 

duodenum (1). We observed single ulcers in 10 cases (larger than 

2 cm in 4), multiple ulcers (3 or more) in 8, mucosal thickenings 

in 10 and polyps in 3. In non-immunocompromised patients, distal 

esophagus and antropyloric region were mostly involved. All 

HIV+ individuals showed synchronous involvement of multiple 

areas of the UGI tract. Histologically, hyperplastic changes were 

mostly associated with mucosal thickenings and polyps and/or 

with glandular epithelial localization of viral inclusions. Ulcerated 

lesion were characterized by endothelial and/or stromal cells 

inclusions. All cases of atypical inclusions were detected in HIV+ 

patients. Follow-up data revealed a persistent HCMV infection 

only in a 1 HIV+ patient. 

Silver In Situ Hybridization (SISH) in determination 

of Her-2 gene status: our experience 

1) Reghellin D. 2) Rucco V. 3) Schiavo N. 4) Paniccià Bonifazi 

A. 5)Lestani M. 

1) U.O.C. Anatomia Patologica, Ulss 5 “Ovest Vicentino”, Arzignano 

(VI), Italia 2) U.O.C. Anatomia Patologica, Ulss 5 “Ovest Vicentino”, 

Arzignano (VI), Italia 3) U.O.C. Anatomia Patologica, Ulss 5 “Ovest Vicentino”, 

Arzignano (VI), Italia 4) U.O.C. Anatomia Patologica, Ulss 5 

“Ovest Vicentino”, Arzignano (VI), Italia 5) U.O.C. Anatomia Patologica, 

Ulss 5 “Ovest Vicentino”, Arzignano (VI), Italia 

Background. Knowledge of HER2 status in breast cancer is a 

prerequisite for Trastuzumab therapy. Immunohistochemistry 

(ICH) and Fluorescence in situ hybridization (FISH) are the two 

most used methods for such purpose. Recently a new device, 

Silver in situ hybridization (SISH), has been proposed for such 

intent. 

Methods. We started performing SISH(INFORM®, Ventana) in 

breast cancer specimens in 2007. Each case had been previously 

studied by ICH. In 12 cases FISH was performed. ICH and FISH 

results were interpreted as suggested by ASCO guidelines. SISH 

samples were interpreted as indicated in Ventana interpretative 

guide (positive: HER2/C17 ratio> 2.2, negative: HER2/C17 ratio 

< 1.8, equivocal: HER2/C17 ratio comprised between 1.8 and 

2.2). Polysomy for C17 was assessed when HER2/C17 ratio was 

1 in tumors having 3 or more HER2 and C17 signals. 

Results. We performed SISH in 75 breast cancer cases; in 

2/75 cases (3%) polysomy for C17 was found; in 7/75 cases 

(9%) SISH results were not assessable. In all cases with positive 

ICH(3+) (n = 11) SISH was positive, except for 3/11 cases 

with not assessable SISH and 1/11 case with negative SISH. In 

all cases with negative ICH(0/1+) (n = 6) SISH was negative. 

Among cases with equivocal ICH(2+) (n = 55), SISH was negative 

in 40/55 cases (73%), positive in 8/55 cases (14%), equivocal 

in 1/55 case (2%) and not assessable in 4/55 cases (7%); in 2/55 

cases (4%) a polysomy for C17 was demonstrated. SISH and 

FISH results were identical, except for a case with negative SISH 

and positive FISH. 

Conclusions. In our experience, SISH is reliable in HER2 gene 

status assessment. Not assessable cases mostly dued to pre-staining 

problems (inadequate fixation). SISH is fast, automated, 

rather simply to interpret and archivable. In small to medium 

size Anatomical Pathology Units it could be the ideal method in 

HER2 gene status evaluation. When a polysomy for C17 is present 

and in equivocal or dubious cases, we advise a second opinion 

performing FISH. 

Comparison between gynecological pap smear 

and thin layer cytology with double decantation 

automatic technology: novaprep system 

347 

G. Rella, L. Mossuto, V. Soli * , L. Fabbiani * , F. Rivasi * 

U.O.C. Anatomia Patologica e Citodiagnostica, P.O.”Sarcone” Terlizzi. 

ASL, Bari; * Università di Modena e Reggio Emilia, Modena. Dipartimento 

ad Attività Integrata di Laboratori, Anatomia Patologica e Medicina 

Legale. Struttura Complessa di Anatomia Patologica 

500 gynecological samples have been analyzed from December 

2009 to May 2010 using double methods: traditional pap smear 

and thin layer cytology. 

The samples refer to women with different age and also to first 

and second level screenings, coming from different hospital sampling 

centers. 

Thin layer cytology noted: cellular /nuclear morphology in accordance 

with standard conventionals; inflammatory and haemorrhagic 

cellular/extracellular and microorganisms decreasing 

presence in comparison with traditional pap smear and that high 

and low grade lesions have accordance with traditional pap smear 

diagnosis. 

Samples thin layer collection allows to repeat the samples processing 

more times and to use the specimens in secondary analysis 

such as immunohistochemistry and molecolar biology. 

The brush with detachable head for the collection of the samples 

shall be used by trained staff to avoid hypocellular samples or 

vials completely lacking in material. 

There is accordance between the two methods and a good versatility 

of Novaprep technology. 

It is necessary to apply Novaprep system thin layer cytology 

study to further cases. 

P16 expression in prostate and its lesions: a 

potential useful diagnostic marker. a comparison 

with p504s (racemase) in 111 cases 

1)Remo A. 2)Zanella C. 3)Bortuzzo G. 4)Seghetto I. 5)Bellotti 

A. 6)Foresto A. 7)Pelegatti S. 8)Parise G. 9)Fasolin A. 10)Vendraminelli 

V. 

Partment of Pathology, “Mater Salutis” Hospital, Azienda Ulss21, Legnago 

(VR), Italy 2)Department of Pathology, “Mater Salutis” Hospital, 

Azienda Ulss21, Legnago (VR), Italy 3)Department of Pathology, “S. 

Giacomo Apostolo” Hospital, Azienda Ulss 8, Asolo (Tv), Italy 4)Department 

of Pathology, “Mater Salutis” Hospital, Azienda Ulss21, Legnago 

(VR), Italy 5)Department of Pathology, “Mater Salutis” Hospital, Azienda 

Ulss21, Legnago (VR), Italy 6)Department of Pathology, “Mater Salutis” 

Hospital, Azienda Ulss21, Legnago (VR), Italy 7)Department of Pathology, 

“Mater Salutis” Hospital, Azienda Ulss21, Legnago (VR), Italy 8)Department 

of Pathology, “Mater Salutis” Hospital, Azienda Ulss21, Legnago 


Azienda Ulss21, Legnago (VR), Italy 10)Department of Pathology, “Mater 

Salutis” Hospital, Azienda Ulss21, Legnago (VR), Italy 

Background. P16 is a inhibitor of the progression during the G1 

phase of the cell cycle. In prostatic tumors p16 is rarely mutated 

and loss of expression occurs frequently through hypermethylation 

or deletion. The aim of this study was to evaluate p16 

expression in prostatic carcinoma (PC) and in normal prostatic 

glands (BP). 

Methods. A total of 111 cases, including 52 cases of PC and 59 

cases of BP, were studied for p16 and p504S. 

Results. P16 showed strong cytoplasmatic expression in 85% 

(44/52) of PC, regardless Gleason Score, and weekly positivity 

in 4% (2/52). 81% of BP (91/111) were negative, including 

benign prostate cases and benign glands adjacent to cancers, and 

19% (20/111) weakly positive. 67% of PIN (14/21) were positive 

and 5% case (1/21) weakly. Atrophic glands were positive in 

12% (2/16) and weakly in 43% (7/16). Cystic dilatation (in IPB) 

were weakly positive in all 7 cases. Urothelial metaplasia were 

weakly positive in 8% case (1/12) and negative in 92% (11/12).

348 

Squamous metaplasia was weakly positive in 1 case and negative 

in 1. 

P504S showed strong cytoplasmatic granular staining in 92% 

(48/52) of PC and weakly positive in 4% (2/52). 71% of BP 

(79/111) were negative, 28% (31/111) weakly positive and 1% 

(1/111) positive. All cases of PIN (100%) were positive. Atrophic 

glands were weakly positive in 25% (4/16) and negative in 75% 

(12/16). Cystic dilatation (in IPB) were weakly positive in 10% 

(1/10) and negative in 90% (9/10). Urothelial metaplasia were 

negative in all 12 cases. Squamous metaplasia was weakly positive 

in 1 case and negative in 1 case. Seminal vesicles (n = 2) and 

verum montanum (n = 2) were negative for both markers. 

None carcinoma was negative simultaneously for p16 and 

p504s. 

Sensitivity were 85% (p16) and 92% (p504s), Specificity 100% 

(p16) and 92% (p504s), positive predictive value 100% (p16) 

and 98% (p504s), negative predictive value 88% (p16) and 93% 

(p504s). 

In conclusion, p16 is a potential useful diagnostic marker particularly 

in addition to p504s. 

Metastasis in intramammary node: occult breast 

carcinoma? A case report 

1)Remo A. 2)Zanella C. 3)Sandrini R. 4)Zaninelli M. 5)Bellini P. 

6)Fasolin A. 7)Bonetti A. 8)Campostrini F. 9)Lanza F. 10)Vendraminelli 

R. 

1)Department of Pathology, Mater Salutis” Hospital, Azienda ULSS21, 

Legnago (Vr), Italy 2)Department of Pathology, Mater Salutis” Hospital, 

Azienda ULSS21, Legnago (Vr), Italy 3)Department of Surgery, Mater Salutis” 

Hospital, Azienda ULSS21, Legnago (Vr), Italy 4)Department of 

Oncology, Mater Salutis” Hospital, Azienda ULSS21, Legnago (Vr), Italy 

5)Department of Pathology, Mater Salutis” Hospital, Azienda ULSS21, 


Azienda ULSS21, Legnago (Vr), Italy 7)Department of Oncology, Mater 

Salutis” Hospital, Azienda ULSS21, Legnago (Vr), Italy 8)Department of 

Radiotherapy, Mater Salutis” Hospital, Azienda ULSS21, Legnago (Vr), 

Italy 9)Department of Surgery, Mater Salutis” Hospital, Azienda ULSS21, 


Azienda ULSS21, Legnago (Vr), Italy 

Background. A 60-year-old woman postmenopausal patient was 

seen to “Mater Salutis” Hospital in Legnago (VR), when she 

presented with a history of a lump in the right breast. On clinical 

examination there was a small (about 10 mm in size) unfixed lesion 

in the lower sagittal area of the right breast. The ultrasound 

revealed a well-defined solid hypoechogenic nodule measuring 

12 mm with some vascular structures in the same area. The axilla 

was negative. The cytologic examination showed neoplastic epithelial 

cells mixed to lymphoid cells (C5). 

Methods. The patient underwent nodulectomy in addition to sentinel 

node biopsy. On histology, the nodule consisted of a ductal 

carcinoma well-differentiated (G1) associated to an organized 

lymphoid tissue comprehensive of germinal centres. On immunohistochemistry 

the tumour was estrogen receptor positive but progesterone 

receptor negative. Labelling index (ki67) was 10-20% 

and Her-2 score 0. Differential diagnosis was with a medullary or 

a carcinoma associated to lymphoid tissue. CD21, CD23 and S- 

100 confirmed the diagnosis of metastatic intrammamary node of 

occult carcinoma, showing follicular dendritic and interdigitating 

cells. Surgical margins were cut and sentinel node negative. The 

multidisciplinary group proposed an MRI of the breast to detect a 

primary carcinoma but this was negative. Staging investigations 

(CT and bone scan) were within normal limits. 

Results. In this case evaluate the patient’s risk was very difficult 

and the options available included: 1) Mastectomy and axillary 

dissection, followed by adjuvant treatment 2) Only radiotherapy 

and/or chemiotherapy. 

We proposed an adjuvant ormonotherpy associated to radiotherapy. 


In literature only two cases have been reported and in both the 

primary tumor, at time of diagnosis, was not detected. In one of 

these 5 years later did became detectable the primary tumor as a 

new lump. 

endocrine breast carcinoma in androgen insensivity 

syndrome (testicular feminization)(morris 

syndrome). A case report. 

1)Remo A. 2)Zanella C. 3)Zaninelli M. 4)Filippini M. 5)Raisa G. 

6)Pozzani S. 7)Bortuzzo G. 8)Fasolin A. 9)Bonetti A. 10)Vendraminelli 

R. 

1)Department of Pathology, “Mater Salutis” Hospital, Azienda ULSS21, 

Legnago (VR), Italy 2)Department of Pathology, “Mater Salutis” Hospital, 

Azienda ULSS21, Legnago (VR), Italy 3)Department of Oncology, 

“Mater Salutis” Hospital, Azienda ULSS21, Legnago (VR), Italy 4)Department 

of Pathology, “Mater Salutis” Hospital, Azienda ULSS21, Legnago 


Azienda ULSS21, Legnago (VR), Italy 6)Department of Pathology, “Mater 

Salutis” Hospital, Azienda ULSS21, Legnago (VR), Italy 7)Department 

of Pathology, “S. Giacomo Apostolo” Hospital, Azienda ULSS 8, Asolo 

(Tv), Italy 8)Department of Pathology, “Mater Salutis” Hospital, Azienda 

ULSS21, Legnago (VR), Italy 9)Department of Oncology, “Mater Salutis” 

Hospital, Azienda ULSS21, Legnago (VR), Italy 10)Department of Pathology, 

“Mater Salutis” Hospital, Azienda ULSS21, Legnago (VR), Italy 

Background. Androgen insensitivity syndrome or testicular 

feminization syndrome (Morris syndrome)(MS) is a type of male 

pseudohermaphroditism (46,XY karyotype) and is one of the 

several intersex conditions inherited according to an X-linked recessive 

(or incompletely recessive) modality of transmission. The 

testes in androgen insensitivity syndrome are at risk of tumors, 

which usually occur after puberty and therefore, bilateral orchiectomy 

is strongly recommended immediately before puberty. Testicular 

tumors seen in this syndrome are hamartomas, germ cell 

tumors, sex-cord tumors, and seminoma plus Sertoli cell tumor in 

the same gonad. Tumors in other organs has not been reported. 

We report the first case of endocrine breast carcinoma in MS 

Methods. A phenotipic woman (46,XY karyotype), at age of 5, 

has been diagnosed MS. She underwent prophylactic gonadectomy 

at age of 21 and subsequent continously ormonal substitutive 

therapy associated to bisphosphonates. At 42-year-old she was 

referred to the radiology for a lump in right breast. 

Results. On physical examination a palpable, painful and mobile 

nodule was present. The ultrasound revealed a well-defined solid 

hypoechogenic nodule measuring 22 mm. The cytologic examination 

showed atypical epithelial cells suggesting a carcinoma 

(C4).The patient underwent surgery (lumpectomy plus sentinel 

node biopsy). Histologically, the tumor was an invasive carcinoma, 

poor differentiated (G3), with a 2,2 cm in size. Estrogen 

and Progesteron receptors were negative Endocrine markers 

expression ranging from 20% (synaptophysin) to 60% (CD56) 

and proposed the tumor as an endocrine carcinoma. Proliferating 

index (ki67) was 90% and Her-2 score 0. Surgical margins were 

cut and sentinel node negative. Adjiuvant chemotherapy (FEC + 

taxotere) was proposed followed by radiotherapy. The patient is 

alive and free from recurrence at 17months from diagnosis. 

This case is the first of breast carcinoma in MS and suggests an 

increased risk of breast cancer due to use of estrogens in these 

individual. 

Solitary tracheal localization of Kaposi’s sarcoma 

in an italian endemic area 

1)Resta L. 2)Martella C. 3)Stomeo S. 4)Napoletano P. 5)Palumbo 

M. 6)Rossi R. 

1)Anatomia Patologica, Policlinico, Bari, Italia 2)Istopatologia, Lab. Pignatelli, 

Lecce, Italia 3)Pneumologia I, P.O. San Cesareo, Lecce, Italia 

4)Anatomia Patologica, Policlinico, Bari, Italia 5)Anatomia Patologica, 

Policlinico, Bari, Italia 6)Anatomia Patologica, Policlinico, Bari, Italia


Background. Salento, a country in the Southern of Apulia, is 

a region area with an high incidence of the endemic form of 

Kaposi’s sarcoma. The disease affects old patients, almost of 

male gender, is localized to the cutis of arms and legs, often 

symmetrically involved, with an indolent clinical course and rare 

visceral progression. The primary and unique localization in the 

trachea is exceptional. 

Methods. L.A., 80-year-old man, underwent to bronchoscopy 

and broncho-alveolar washing for a suspicious of lung neoplasm. 

The examination reveals a small (6 mm) polypoid lesion in the 

lower third of trachea. 

Results. Histologically, the polypoid lesion was covered by 

normal tracheal mucosa and constituted by a combined proliferation 

of small vessels and spindle cells. Small foci of diapedetic 

hemorrage, hemosiderin rich histiocytes, and rare inflammatory 

cells were present. Immunohistochemistry revealed the 

expression for factor VIII-related antigen in the proliferating 

endothelial cells, CD34 in endothelial and spindle cells, and cytokeratins 

only in superficial epithelium. A diagnosis of Kaposis’ 

sarcoma was performed. Cytology of the broncho-alveolar 

fluid was negative. 

The subsequent clinical and radiological examination of the patient 

did not revealed other localization of the disease. 

The tracheal localization of Kaposi’s sarcoma is referred in 34 

patients with immuno-deficiency (AIDS, organ transplantation), 

with multi-visceral involvement. Exclusive and/or primitive 

tracheal localization in an endemic area was not referred in literature. 

Wegener’s granulomatosis: a case report 

Arborea G., Resta L., Palumbo M., Fiore G., Diclemente D. 

1)Anatomia patologica, Policlinico, Bari, Italia 

Background. Wegener’s Granulomatosis (WG) is a necrotizing 

vasculitis associating with extravascular granulomatosis. The 

disease is not so rare and its incidence is 4-8 cases/1,000,000 inhabitans/year. 

The mean age of occurrence is 45 years but forms 

have been described in very elderly subjects. The most frequent 

localizations of WG are upper respiratory tract, lung and kidney. 

Methods. We report the case of a 70-year-old man who has been 

complaining of alveolar-maxillary pain for about two months. 

The maxillary dentalscan showed a “paramedian structural reshuffle 

with cortical vestibular lytic break”. This damage was 

then confirmed with CT and MR. Hence the patient underwent 

surgery to remove the damaged areas. Hyperplastic mucosa’s 

specimens was taken from left maxillary sinus and nasal cavity 

and sent to histological analysis. All histologic specimens showed 

necrosis, chronic inflammation, necrotizing vasculitis and granulomatosis. 

According to these histopathological elements we 

made diagnosis of WG. 

Results. WG is a cronic severe desease that is fatal if not treated. 

However, currently available immunosuppressant therapies can 

cure most cases of this disease with a five-year survival which 

exceed 80%. 

Primary intracranial Hodgkin’s lymphoma. 

A case report and review of the literature 

1)Riccioni (L). 2)Morigi (FP). 3)Gessaroli (M). 4)Giovannini 

(A). 5)Guiducci (G). 

1)U.O. Di Anatomia Patologica, Ospedale “M. Bufalini”, Cesena, Italia 

2)U.O. Di Anatomia Patologica, Ospedale “M. Bufalini”, Cesena, Italia 

3)U.O. Di Chirurgia Maxillo-Facciale, Ospedale “M. Bufalini”, Cesena, 

Italia 4)U.O. Neuroradiologia, Ospedale “M. Bufalini”, Cesena, Italia 

5)U.O. Di Neurochirurgia, Ospedale “M. Bufalini”, Cesena, Italia 

Background. Central nervous system (CNS) involvement occurs 

in 0.2-0.5% of patient with Hodgkin’s lymphoma (HL) and is 

349 

usually secondary to a disseminated disease outside the CNS or 

manifests at time of relapse, more frequently in human immunodeficiency 

virus (HIV)-positive patients. Primary intracranial HL 

at presentation is exceedingly rare, with only 10 case reported to 

date. Among these cases only 6 were an isolated localization of 

the disease. 

Case history. We report a rare case of isolated primary intracranial 

HL occurring in a 30-year-old immunocompetent male, 

who presented with a progressive left exophthalmos. He had 

a serious cranial-facial trauma six years before, that had been 

surgically treated. Magnetic resonance imaging showed a left 

frontal-ethmoidal-orbital mass, showing contrast enhancement on 

T1 weighted images. The patient underwent craniotomy and the 

lesion was successfully resected. 

Results. Histologically the tumor was a dense lymphoid tissue 

with a diffuse fibrillar background, infiltrating at the periphery 

the cerebral parenchyma. It was characterized by the presence 

of numerous large blastic cells, with a large, round, vesicular 

nucleus and prominent eosinophilic nucleoli, embedded within 

an inflammatory background composed of small lymphocytes, 

plasma cells and eosinophils. Scattered classical lacunar and 

binucleate Reed-Sternberg cells were present. On immunohistochemistry 

mononuclear and binucleate large cells were positive 

for CD30, CD15, HLA-DR, PAX-5 (faint) and EBV-LMP1. In 

situ hybridization for EBV encoded mRNA (EBER) resulted 

positive. The overall results were consistent with the diagnosis of 

a lymphocyte-depleted classical HL. 

Staging procedures gave a negative result. The patient was 

subsequently treated with four cycles of ABVD-chemotherapy 

followed by 30.6 Gy radiotherapy to the left orbital region. He 

is alive and disease free at 3-year follow-up, with no systemic 

manifestations of HL. 

expression of P63 in merkel cell carcinoma is 

related to prognosis: an immunohistochemical 

and molecular analysis 

1)S. Asioli,1)A. Righi, 2)D. De Biase, 2)L. Morandi, 3)V. Caliendo, 

2)M. Ragazzi, 1)C. Botta, 1)L. Verdun di Cantogno, 1)F. 

Maletta, 3)G. Macripò, 2)V. Eusebi, 1)G. Bussolati 

1)Scienze biomediche e oncologia umana, Ospedale Molinette, Torino, 

Italia; 2)Ematologia e scienze oncologiche, Ospedale Bellaria, Bologna, 

Italia; 3)Divisione di Dermatologia, San Giovanni Battista-San Lazzaro, 

Torino, Italia 

Background. p63 expression in Merkel cell carcinoma (MCC) 

indicates an aggressive behaviour of the tumour. At least three 

TA variants (TAp63α,β,γ) and three ∆N variants (∆Np63α,β,γ) 

by alternative splicing from p63 gene have been identified. 

Recently it has been suggested that presence of polyomavirus 

(MCPyV) in MCC tumour tissue is an indicator of adverse prognosis. 

To better define the role of p63 and its variants in MCC and 

the possible relation to MCPyV, we examined a series of MCC 

from 45 patients collected from different Institutions. 

Methods. 50 cases of MCC from 45 patients (6 cases showed 

nodal metastases and 1 case brain metastasis) were investigated 

for p63 expression by immunohistochemistry (IHC) and by reverse-transcription 

polymerase chain reaction (RT-PCR) using 

isoform-specific primers to evaluate the p63 mRNA expression 

patterns. Probes for p63 gene (3q28) were used for FISH analysis 

to value the p63 gene status. The presence of MCPyV in the MCC 

tumour genome was also investigated by PCR in all cases. 

Results. p63 expression was detected in 62% of cases by IHC and 

it was associated with decreasing overall survival (p = 0.003). All 

these cases but one presented at least one of the p63 isoforms by 

RT-PCR, both in the primary MCC (25 cases) and in metastases 

(5 cases), with a variable expression pattern of the isoforms 

(TAp63γ was present in 76.7% of cases, ∆Np63β in 16.7%, 

∆Np63α in 36.7%, TAp63β in 16.7%, TAp63γ in 6.7%, ∆Np63γ

350 

in 3.3%). P63 gene gain was found by FISH analysis in only one 

case. Clonal integration of MCV DNA sequences was observed 

in 86.6% of cases. The present IHC and molecular data confirm 

p63 expression in a group of MCC with aggressive clinical behaviour 

and suggest that a transcriptional dysregulation of p63 

gene is involved in the pathogenesis of MCC. IHC analysis is 

less specific than the molecular analysis to value p63 expression 

in MCC. Clonal integration of MCPyV DNA sequences does not 

seem related to prognosis. 

BuBr1 expression in oral squamous cell carcinoma 

and its relationship to tumor stage and survival 

1)Rizzardi C. 2)Torelli L. 3)Barresi E. 4)Schneider M. 5)Canzonieri 

V. 6)Melato M. 

1)Patologia e medicina legale, Università di trieste, Trieste, Italia 2)Matematica 

ed informatica, Università di trieste, Trieste, Italia 3)Patologia e 

medicina legale, Università di trieste, Trieste, Italia 4)Anatomia ed istologia 

patologica, Ass2 isontina, Gorizia, Italia 5)Anatomia patologica, Cro, Aviano, 

Italia 6)Patologia e medicina legale, Università di trieste, Trieste, Italia 

Background. Defects in the mitotic spindle checkpoint have been 

proposed to contribute to the chromosomal instability observed in 

human cancers, including oral squamous cell carcinoma. BUBR1 

is a key component of the spindle checkpoint, whose role in oral 

carcinogenesis still needs to be clarified. 

Methods. We have analyzed the expression of BUBR1 in 49 

cases of oral squamous cell carcinoma by immunohistochemistry, 

and compared the findings with clinicopathological parameters, 

proliferative activity, and DNA ploidy. 

Results. BUBR1 was overexpressed in 11 (22.4%) cases. BUBR1 

overexpression was significantly associated with a less advanced 

pathological tumor stage (p = 0.05), possibly as a consequence 

of a less tendency to metastasize and to relapse, although recurrences 

seemed to occur earlier than in the group without overexpression 

of the protein. Despite the relatively limited number of 

cases analyzed, our data imply the possibility that BUBR1 may 

be involved in the progression of squamous cell carcinoma of the 

oral cavity, although the mechanism of action and significance 

remain unknown, and are controversial. Furthermore, our data 

suggest that BUBR1 may be a promising prognostic marker in 

patients with oral squamous cell carcinoma. 

Histopathology quality control in breast cancer 

screening program 

1)A. Rizzo, 2)A. Farnedi, 3)C. Naldoni, 4)S. Guzzinati, 5)Emilia 

Romagna breast cancer screening group, 6)V. Eusebi 

1)Anatomia ed Istologia Patologica, Ospedale S. Giacomo, Ulss 8 Castelfranco 

Veneto, Italia; 2)Sezione di Anatomia Patologica, Ospedale Bellaria 

- Università di Bologna, Bologna, Italia; 3)Assessorato alla Salute, Regione 

Emilia Romagna, Bologna, Italia; 4)Registro Tumori del Veneto, Istituto 

Oncologico Veneto - IRCCS, Padova, Italia; 6)Sezione di Anatomia Patologica, 

Ospedale Bellaria – Università di Bologna, Bologna, Italia 

Background. The aim of this study was that of evaluating the 

diagnostic reproducibility on core biopsies (NCB) of complex 

breast lesions, among several pathologists from Emilia Romagna 

and other Italian regions. 

Methods. Fifty-four slides of NCB performed for breast lesions 

were selected from 24 slide seminars (2002-10) among 14 pathology 

units of Emilia Romagna and other regions involved in 

screening programmes, according to the B classification for NCB 

(Tumors breast Pathology, AFIP 2009). Immunohistochemical 

features were known after discussion. Each case had to be labelled 

along the major lesion present in the slide corresponding 

to the B category. The final diagnosis was accepted when the 

majority of presents agreed (MD). 

Results. According to MD, 7 cases were classified as B2, 35 B3, 

1 B4, 11 B5. 21 cases (48%) were atypical ductal hyperplasia 


vs DIN 1c; other cases were also representative of proliferative 

myositis, low grade angiosarcoma, low grade adenosquamous 

carcinoma, infiltrating epitheliosis. All pathologists reached 

the same diagnosis in six cases (11%); in five cases only one 

institution proposed a different diagnosis. Individual weighted 

kappa coefficients in comparison to MD are good-excellent (0.62 

- 0.78: p < 0.001) for 8 Institutes (57%), moderate (0.40-0.57) 

for 4, low (kappa 0.34 and 0.36) for 2 Institutes. Overall, individual 

weighted kappa is 0.577. Kendall’s coef. of concordance 

was 0.55. 

Cases were selected for the study when represented a diagnostic 

problem which was shown by the high prevalence of B3 diagnoses 

(60%) where there are the lowest levels of diagnostic agreement. 

It concluded that in complex lesions a second opinion is 

recommended. 

Myeloid sarcoma and synchronous 

adenocarcinoma of the colon 

1)Rocca B.J. 2)Ambrosio M.R. 3)Onorati M. 4)Mourmouras V. 

5)Di Mari N. 6)Bellan C. 7)Leoncini L. 8)Lazzi S. 

1)Department of Human Pathology and Oncology -Anatomic Pathology 

Section, Santa Maria delle Scotte, Siena, Italy 2)Department of Human 

Pathology and Oncology -Anatomic Pathology Section, Santa Maria delle 

Scotte, Siena, Italy 3)Department of Human Pathology and Oncology 

-Anatomic Pathology Section, Santa Maria delle Scotte, Siena, Italy 




Scotte, Siena, Italy 6)Department of Human Pathology and Oncology 

-Anatomic Pathology Section, Santa Maria delle Scotte, Siena, Italy 




Scotte, Siena, Italy 

Background. Myeloid sarcoma is defined as a tumor mass composed 

of myeloid blasts, arising in extramedullary sites. It may 

precede or coincide with an acute myeloid leukemia (AML), 

often being the first manifestation or represent acute blastic 

transformation of myelodysplastic syndromes (MDS), myeloproliferative 

neoplasms (MPN) or MDS/MPN. Bone, lymph nodes 

and skin are the most common localizations, involvement of the 

large bowel is rare. 

Methods. A 76-year old woman underwent surgery for acute 

abdomen. Routine haematological and biochemical investigation 

were normal. Sections from formalin fixed, paraffin embedded 

samples were stained with haematoxylin and eosin and a panel of 

antibodies were checked. 

Results. The surgical specimen consisted of 33 cm right colon 

showing a polypoid ulcerated lesion (5 cm in greater dimension) 

perforating the wall. Microscopically, a medium grade 

adenocarcinoma infiltranting the muscular layer was observed. A 

diffuse infiltration of cells with blastic appearance, round nuclei, 

finely dispersed chromatin and a single or multiple small central 

nucleolus, with scant eosinophilic or basophilic cytoplasm, was 

found. The cells were: CD10-, CD20-, CD3-, MPO+, CD68+, 

CD68PGM1+, CD56+, CD123-, BDCA-2/CD303-, TCL1-; proliferative 

rate (Mib-1) was of 50-60%. Scattered mitotic figures 

were also observed. A diagnosis of synchronous myeloid sarcoma 

and adenocarcinoma (G2) was made. Lymph nodes were infiltrated 

by both tumors. Two months later the patient developed an 

AML as confirmed by bone marrow biopsy. 

Conclusion. The peculiarity of this case is the clinical presentation 

as an acute abdomen due to the infiltration of serosa 

by myeloid sarcoma that lead to discover an adenocarcinoma. 

Hematological malignancies coexistence with solid tumor are 

uncommon and histopathological description of synchronous 

large bowel myeloid sarcoma and adenocarcinoma has never 

been previously made.


leiomyosarcoma of the uterus with focal 

rhabdomyosarcomatous differentiation and 

urinary symptoms 

1. Luca Roncati 2. Giuseppe Barbolini 3. Giuliana Sartori 4. 

Elena Siopis 5. Laura Marra 6. Francesco Rivasi 

1, 2, 3, 6 Department of Laboratory Services, Pathology and Forensic Medicine, 

Section of Pathology, University of Modena and Reggio Emilia, 

Modena, Italy; 4 Department of Diagnostic and Imaging Services, Section 

of Radiology, University of Modena and Reggio Emilia, Modena, Italy; 5 

Department of Oncology, Hematology and Respiratory Diseases, Section 

of Oncology, University of Modena and Reggio Emilia, Modena, Italy 

Background. Leiomyosarcomas are the most common histologic 

type of uterine sarcomas. Heterologus mesenchymal leiomyosarcoma 

is the rarest variant, in which the tumor may exhibit lipoleiomyosarcomatous, 

osteosarcomatous or rhabdomyosarcomatous 

differentiation. We report a case of uterine leiomyosarcoma 

with focal rhabdomyosarcomatous differentiation, the fourth 

proved case of the Literature. 

Methods. The patient was a nulligravid 51-years-old woman 

who noticed an increase of lower abdominal fullness with appearance 

of urinary symptoms. Radiology revealed an uterus 

of 12 × 10 × 8 cm with an intramural mass displacing the bladder 

and the right ureter. Total hysterectomy with bilateral salpingo-oophorectomy 

was performed; loco-regional hypertrophic 

lymph nodes were not found. On gross examination a 7,5 firm, 

centrally soft, tumor was found infiltrating the posterior-lateral 

myometrium up to perimetrium. The tumor was composed of a 

predominant leiomyosarcomatous component and a minor rhabdomyosarcomatous 

component without forms of transition between 

them. Besides phosphotungstic acid haematoxylin (PTAH) 

immunohistochemistry for desmin, α-smooth muscle actin, sarcomeric 

actin, myoglobin, CD10, CD56 and P16 was performed. 

Molecular biology techniques for the state of methylation in the 

promoter region of oncosuppressor CDKN2A gene, encoding for 

P16 protein, were also performed after microdissection of both 

neoplastic components. 

Results. Leiomyosarcomatous cells were immunoreactive for 

desmin and α-smooth muscle actin, only. Rhabdomyosarcomatous 

cells, provided with cytoplasmic cross striations, were immunoreactive 

for desmin, sarcomeric actin, myoglobin, CD10, 

CD56, P16. Moreover a loss of heterozygosity (LOH) was found 

only in the microdissected specimens of rhabdomyosarcomatous 

cells. The patient is alive and well two years after surgery. 

Vascular endothelial growth factor / receptor 

systems expressed by basophils in vaginal 

angiomyofibroblastoma 

1 Luca Roncati 2 Giuseppe Barbolini 3 Francesco Rivasi 

1 Department of Laboratory Services, Pathology and Forensic Medicine, 

Section of Pathology, University of Modena and Reggio Emilia, Modena, 

Italy 2 Vascular endothelial growth factor / receptor systems 3 expressed 

by basophils in vaginal angiomyofibroblastoma 

Background. Angiomyofibroblastoma (AMFB) is a rare well 

circumscribed benign tumor principally occurring in vulvovaginal 

soft tissue, mainly composed of blood vessels and alternating 

zone of cellularity. Its angiogenesis is not clearly understood 

since mesenchymal stem cells, tumor cells and mast cells have 

been considered to be involved in this role. 

Methods. We report four cases of angiomyofibroblastoma of the 

vaginal wall. Three patients were previously affected by breast 

cancer (treated with tamoxifen), while the fourth by lipoma of 

the right arm. Besides toluidine blue (pH 4,5) immunohistochemistry 

for vimentin, desmin, alpha isoform (smooth muscle) actin, 

CD10, estrogen and progesteron receptors, CD31, CD34, D2-40 

(podoplanin), CD117 (c-KIT), vascular endothelial growth factor 

(VEGF) and its receptor (Flt-4) was performed. 

351 

Results. The spindle / ovoid neoplastic cells were immunoreactive 

for vimentin, desmin, alpha isoform actin, CD10, estrogen 

and progesteron receptors. Besides small to medium-size blood 

vessels, occasionally ectatic and branching, immunoreactive for 

CD31 and CD34, thin walled lymphatics immunoreactive for D2- 

40 were also noticed. 

Moreover numerous perivascular basophils orthochromatic with 

toluidine blue and immunoreactive for CD117, vascular endothelial 

growth factor and its third receptor (Flt-4) were observed. Our 

findings imply that activated basophils may play a crucial role in 

angiogenesis of AMFB. 

Applicazione dell’immunoistochimica in citologia 

in fase liquida e su citoincluso nelle neoplasie 

tiroidee 

Rossi E.D., Fadda G., Visca E., Zannoni G.F., Vellone V.G., 

Rindi G. 

Istituto di anatomia e istologia patologica, Università cattolica s. cuore, 

Roma, Italia 

Introduzione. La citologia agoaspirativa rappresenta un fondamentale 

strumento nella diagnostica delle lesioni tiroidee. 

Essa presenta tuttavia dei limiti che non consentono la massima 

accuratezza diagnostica: a) imprevedibile quantità di materiale 

ottenibile dal prelievo; b) difficoltà di distinguere le cellule 

benigne dalle maligne. Alcune tecniche si sono affiancate alla 

morfologia convenzionale per fornire informazioni utili per la 

succitata diagnosi differenziale e per la selezione dei pazienti che 

necessitano di terapie più aggressive o di un follow-up più accurato: 

tra queste in particolare l’immunocitochimica ha mostrato 

interessanti sviluppi. 

Obiettivo. L’obiettivo del presente studio è quello di mostrare 

l’efficacia diagnostica dell’immunoistochimica per HBME-1 e 

galectina-3 applicata a casi di proliferazione follicolare tiroidea 

(PF - TIR 3 sec. la classificazione SIAPEC-IAP del 2008) con 

successivo controllo istologico. La metodica immunoistochimica 

è stata eseguita sia su preparati allestiti in fase liquida che su preparati 

ottenuti dall’inclusione del materiale residuo della citologia 

in fase liquida. 

Materiali e metodi. Nel periodo novembre 2009-aprile 2010 

sono stati esaminati 1493 agoaspirati tiroidei presso l’U.O.C. 

di Istopatologia e Citodiagnosi del Policlinico “A.Gemelli” 

di Roma, 134 dei quali (9%) classificati come PF. Su questi 

casi è stata eseguita l’indagine immunocitochimica per valutare 

l’espressione di HBME-1 e galectina-3 sui preparati allestiti in 

fase liquida secondo la metodica Thin Prep 2000 della Hologic 

Italia (Roma). Su 22 casi con successivo intervento chirurgico è 

stato anche allestito il citoincluso dal materiale residuo della fase 

liquida con il sistema Cytoblock (Shandon) e gli stessi anticorpi 

sono stati testati per valutare le eventuali differenze rispetto alla 

citologia. Di questi ultimi 15 (68,1%) hanno avuto una diagnosi 

di proliferazione follicolare (PF -TIR 3), 4 (18,2%) sono stati 

classificati come carcinomi papillari (PC - TIR 5), 1 (4,5%) come 

sospetto per carcinoma (SC- TIR 4) e 2 (9,1%) come strumi colloideo-cistici 

(SCC - TIR 2). 

Risultati. Nove casi (40,5%) non hanno evidenziato sufficiente 

cellularità (in 6 di essi era identificabile solo colloide). Dei 13 

casi con cellularità adeguata 7 (53,8%) sono stati perfettamente 

confermati (IIC su citologia e istologia concordante), 5 (38,5%) 

hanno avuto la discordanza su un solo anticorpo, 1 solo caso con 

discordanza su entrambi gli anticorpi. Le conferme sono state 

quasi esclusivamente su casi di proliferazione follicolare (TIR 

3) mentre nessuno dei PC ha avuto materiale sufficiente nel 

citoincluso. 

Conclusioni. Lo studio dimostra come sia possibile allestire il 

citoincluso anche dal materiale residuo dopo la citologia in fase 

liquida e che i risultati dell’immunoistochimica sui preparati citologici 

e microistologici siano sovrapponibili.

352 

BCl10 expression in peripheral T-cell lymphoma 

not otherwise specified 

Maura Rossi, Claudio Agostinelli, Anna Gazzola, Claudia Mannu, 

Maria Rosaria Sapienza, Maria Antonella Laginestra, Carlo 

Sagramoso, Elena Sabattini, Francesco Bacci, Patrizia Artioli, 

Luigi Chilli, Federica Sandri, Milena Piccioli, Gianpaolo Da 

Pozzo, Simona Righi, Stefano A Pileri, Pier Paolo Piccaluga 

Department of Hematology and Oncology “L. and A. Seràgnoli”, Hematopathology 

Unit, S. Orsola-Malpighi Hospital, University of Bologna, 

Italy; *SAP and PPP equally contributed 

Background. BCL10 encodes for a TCR-signaling downstream 

protein with apoptotic properties which was found to be expressed 

in several B-non Hodgkin lymphomas, while few data are 

available for peripheral T cell lymphomas not otherwise specified 

(PTCLs/NOS). 

We analyzed BCL10 expression in PTCL/NOS in order to establish 

the prevalence of BCL10 expression and its potential 

prognostic significance in this setting. 

Methods. Gene expression profile (GEP)analysis of 40 PTCLs 

[28 PTCLs/NOS, 6 angioimmunoblastic lymphomas (AITLs), 

and 6 anaplastic large cell lymphomas (ALCLs; 3 ALK+ and 3 

ALK-)], 4 non-neoplastic reactive lymph-nodes, and 20 samples 

of normal T-lymphocytes, was performed with the Affymetrix 

HG-U133 2.0 plus microarray; immunohistochemical expression 

of BCL10 was led in 52 PTCLs/NOS on tissue microarrays. 

Results. GEP showed significantly lower BCL10 expression 

in all PTCLs in comparison to normal samples. No significant 

differences emerged among PTCL types. The mean expression 

value was 418.92 in PTCLs/NOS; 402.1 in AITLs, 234.0 in AL- 

CLs and 742,1 in normal samples (p < 0.05). 

BCL10 expression was positive in 16/52 cases (31%), not showing 

any correlation with the expression of either Ki-67 (≥80% in 

12% of cases) or T-cell associated molecules (CD3, CD5, CD7, 

CD52). On the other hand, reactive lymph-nodes presented with 

consistent expression of BCL10 in paracortical T-lymphocytes in 

all instances. 

Finally, we investigated whether BCL10 expression was associated 

with PFS or OS. Indeed, we did not observe significant differences 

in BCL10 + vs. BCL10 - cases. However, a favorable trend 

in BCL10 + cases was recorded. 

Conclusion: BCL10 turned out to be frequently down-regulated 

in PTCLs, in comparison to normal T-lymphocytes, suggesting 

possible abnormalities in TCR signaling cascade. In addition, 

though in our series BCL10 expression did not significantly correlate 

with patients’ survival, a favorable trend in BCL10 + cases 

was observed, indicating the opportunity of testing this marker 

in larger series. 

Cervical lesions in young women living in reggio 

emilia 

T. Rubino, L. Bulgarelli, R. Bio, L. Campioli, C. Fodero, S. 

Prandi 

U. O. Centro di Citologia Cervicovaginale di screening- Dipartimento 

Oncologico Arcispedale S. Maria Nuova, Reggio Emilia, Italia 

Introduction. Cervical screening programmes targets a female 

population of between 25 and 64 years, however there is no 

unanimous agreement on the age to begin performing pap smears. 

While AA declare that cervical lesions discovered before 25 

years cause the use of unnecessary treatment to prevent invasive 

carcinoma (a rare event because most such lesions as CIN I and 

CIN II regress), on the contrary incidences of CIN III in the last 

two decades have increased for women younger than 35 years 

in comparison with the older one. If we assume a conservative 

progression rate for CIN III to invasion of 1% per year and there 


is some suggestion that progression may be higher in younger 

women, significant numerous of younger woman are at risk of 

developing invasive cervical cancer. For these reasons we wanted 

to rate the incidence of cervical lesions in a population of young 

women engaged in spontaneous screening for both pap smears 

and biopsies. 

Methods and results. From January 2008 to May 2010, 2337 

pap smears were made in Consultori Giovani of Azienda USL- 

RE. A study was conducted on women between 14 and 24 years 

and 196 (8,38%) pap smears were positive ≥ ASCUS, while 

2141 (91,61%) were negative/inflammatory. Positive pap smears 

were very divided: ASCUS/AGC N°52 (26,53%), LSIL N° 119 

(60,71%), ASCH N°5 (2,55%), HSIL N°20 (10,20%). This study 

revealed that 43 (22,93%) positive women did not perform any 

colposcopy, 53 (27,4%) performed colposcopy with negative/not 

performed biopsies, 67 (34,18%) CIN I, 12 (6,12%) CIN II, 21 

(10,71%) CIN III. 

Conclusions. This high number of refusal to perform colposcopy 

is worrying because behind low-grade lesions, may be high-grade 

CIN. We found 21 CIN III, the first one at 17 years, which were 

permanent lesions. The question is: are we going to find new 

prospects? Something is changing: the new American guidelines 

show us that Cervical screening programmes have to start at 

age 21 with pap smears, because the HPV test is not specific 

enough. 

Chromosomal abnormalities in paraffin-embedded 

first trimester spontaneous abortions detected by 

fISH 

1)Russo R. 2)Fumo R. 3)Gaeta S. 

1)Anatomia Patologica, S Giovanni Di Dio E Ruggi D’aragona, Salerno, 

Italia 2) Patologica, S Giovanni Di Dio E Ruggi D’aragona, Salerno, 

Italia 3)Anatomia Patologica, S Giovanni Di Dio E Ruggi D’aragona, 

Salerno, Italia 

Background. A chromosomal abnormality is present in the 

majority of early spontaneous abortions and most of them are 

due to numerical chromosome abnormalities. Cytogenetic study 

of spontaneous abortions is not always performed and a relatively 

high rate of culture failures is reported. The use of FISH 

to identify chromosomes in interphase cells has been successfully 

applied to uncultured fetal cells, chorionic villus tissue 

or dysmorphic fetus. The purpose of the present study was to 

evaluate the efficiency of FISH in understanding the etiology of 

spontaneous abortions. 

Method. FISH was performed on paraffin-embedded first trimester 

spontaneous abortions using 15, 16, 18, X, Y CEP probes and 

13, 21 and 22 LSI probes panel. One hundred interphasic nuclei 

were analyzed for each probe. Both the mesenchymal and trophoblastic 

cells from the placental villous samples were scored. The 

case was considered pathologic when at least 80% of the nuclei 

scored showing more (trisomic/tetrasomic) or less (monosomic) 

of two copies for only one probe. Reliability of the FISH method 

was demonstrated in control samples in which karyotype was 

available. Moreover, cells from decidual tissue were not counted, 

but served as an internal diploid control. 

Results. Four hundred fifty-five cases from first trimester spontaneous 

abortion were examined for both pathological and FISH 

analysis. 300/455 (66%) cases presented numerical chromosome 

abnormalities. Trisomies were detected in 78%, poliploidy (triploidy 

and tetraploidy) in 11.3%, X monosomy in 6%, mosaic in 

3,6% and sexual trisomy (XXX,XXY) in 1% of all pathological 

cases. Of 235 trisomies, 55.3% was represented by +16, 22.1% 

+15, 9.8% +21, 9% +22, 2.1% +18 and 1.7% +13. 

Conclusions. The lack of peculiar morphological criteria to recognize 

the chromosomal etiology of early spontaneous abortions 

makes FISH analysis with use of an appropriate probe panel, a 

surprising test to discover genetically caused abortions.


Sarcomatoid variant of anaplastic large cell 

lymphoma of the ovary: report of a unique case 

occurring as a stromal nodule in a mucinous 

cystoadenofibroma 

1)Russo S. 2)Baldassarre F. 3)Siciliano A. 4)Maiello F.M. 5)Facchetti 

F. 

1)Anatomia Patologica Ospedale Maresca, ASLNA3sud, Naples, Italy. 

2-3-4)Anatomia Patologica, Ospedale Pellegrini, ASLNA1, Napoli, 

Italy. 5)Director, Department of Pathology I Spedali Civili - University 

of Brescia. 

Background. Primary ovarian lymphoma are extremely rare and 

usually composed of B-cell non-Hodgkin lymphomas. Here we 

report a case of ALK1+ anaplastic large cell lymphoma (ALCL) 

occurring in the ovary and arising within a septum of a mucinous 

cystoadenofibroma. 

Methods. A 49 years old woman with a history of breast carcinoma 

was admitted because of a mass in the left ovary. The 

patient underwent a salpingooophorectomy; the gross specimen 

consisted of an ovoid mass measuring 10 × 6 × 5 cm with a cystic 

appearance, containing a 2 cm grey mural nodule; the fallopian 

tube was normal. Paraffin sections were stained with hematoxylin 

and eosin and immunohistochemistry was performed using an 

avidin-biotin-peroxidase technique. The patient died few days 

after surgery because of an increasing persistent hyperpyrexia. 

No autopsy was performed. 

Results. Histological examination revealed a cystoadenofibroma; 

the mural nodule contained a polymorphic cell population 

including large atypical cells with interweaving fascicles 

of plump spindle cells (reminiscent of storiform malignant 

fibrous histiocytoma), scattered neutrophils and histiocytes. 

The atypical cells were strongly positive for CD45RO, CD30 

(membranous and Golgi), ZAP70 and ALK1 protein (nuclear 

and cytoplasmic), while they were negative for CD3, CD20, 

CD21, clusterin, cytokeratins, and S100 protein. At the best of 

our knowledge this represents the first case of ALK1+ ALCL 

primarily occurring in the ovary. Immunophenotyping was fundamental 

not only to identify the nature of the atypical cells, but 

also to exclude sarcoma-like mural nodules of mucinous cystic 

ovarian tumors, as well as a myofibroblastic tumor, which may 

express ALK1. 

Solitary plasmacytoma of the thyroid: a case report 

1)Russo S. 2)De Gregorio A. 3)Annunziata S. 4)Baron L. 5)Fiore 

L. 6)La Provitera A. 7)Salvati A. 

1-2-3)Anatomia Patologica, P.O. Maresca ASLNA3sud, Torre del greco 

(Napoli), Italy. 4)Anatomia Patologica P.O.S.Leonardo di Castellammare 

di Stabia (Napoli), Italy. 5-6)Chirurgia generale e d’urgenza, P.O. di Boscotrecase 

ASLNA3sud, Boscotrecase (NA), Italy. 7)Anatomia patologica, 

P.O. Maresca ASLNA3sud, Torre del Greco(NA), Italy. 

Background. We report a case of solitary plasmacytoma of the 

thyroid arisen in a background of Hashimoto thyroiditis. Most 

patients with plasma cell neoplasia have generalized disease at 

diagnosis, i.e. multiple myeloma (MM). However, a minority of 

patients with plasma cell malignancies present with either a single 

bone lesion, or less commonly, a soft tissue mass, of monoclonal 

plasma cells: solitary bone plasmacytoma (SBP) or solitary 

extramedullary plasmacytoma (SEP). SBP has a high risk of progression 

to MM. In contrast, SEP is nearly always truly localized 

and has a high cure rate with local treatment. Although SEP can 

arise throughout the body almost 90% arise in the head and neck, 

especially in the upper respiratory tract including the nasal cavity, 

sinuses, oropharynx, salivary glands and larynx. The next most 

frequent site is the gastro-intestinal tract. Other sites can rarely 

be involved, including the thyroid. 

353 

Methods. A 77-year-old man presented to an outpatient clinic 

with a large painless neck mass without other symptoms. The entire 

thyroid was surgically removed, and his postsurgical course 

was uneventful. The specimen measured 14 X 6 X 5 cm. The cut 

surface was lobulated, soft and white. 

Results. Intraacinar clusters of macrophages, lymphocytes, and 

plasma cells were present. Some acini were compressed, and represented 

only by a cluster of closely grouped pale vesicular nuclei 

with little cytoplasm. The distorted acini were surrounded and 

separated by a diffuse, sheet-like infiltrate of large round-oval 

CD20+, CD138+, CD79a+, TTF1 negative cells with eccentric 

nuclei, often showing chromatin clumps radially arranged in ‘cart 

wheel’ fashion, involving the entire parenchyma. Haematological 

work up for MM remains negative at the time of writing. Immunohistochemical 

staining revealed evidence of monoclonalism of 

plasma cells with light chain restriction and predominant staining 

for kappa chains. In our case only surgical extirpation was done 

as the margins were clear. 

Biphasic large cell neuroendocrine carcinoma 

– pure mucinous carcinoma of the gallbladder: 

a unique combination. Case report 

1)Russo S. 2)De gregorio A. 3)Maiello F.M. 4)Paolini B. 5)Fiore 

L. 6)Laprovitera A. 7)Sepe J. 8)Carrabba A. 9)Salvati A. 

1)Pathology, Maresca Hospital, ASLNA3sud, Naples, Italy 2)Pathology, 

Maresca Hospital, ASLNA3sud, Naples, Italy 3)Pathology, National Cancer 

Institute, Milan, Italy 4)Boscotrecase Hospital, General and emergency 

surgery, ASLNA3 sud, Naples, Italy 5)Boscotrecase Hospital, General 

and emergency surgery, ASLNA3 sud, Naples, Italy 6)University of Maryland, 

University College, Naples, Italy 7)Biological Science, University 

of Naples, Naples, Italy 8)Pathology, Maresca Hospital, ASLNA3sud, 

Naples, Italy. 

Background. We report a case of primary combined large cell 

neuroendocrine carcinoma – pure mucinous carcinoma of the 

gallbladder, which represents the first description of this entity. 

Large Cell Neuroendocrine Carcinoma (LCNEC) shares some 

features of the well-differentiated neuroendocrine tumor, such as 

the “organoid” growth pattern and rosette formation, while also 

manifesting characteristics of the poorly differentiated small cell 

carcinomas, including necrosis, high mitotic rate, and salt-andpepper 

chromatin. Mucinous adenocarcinomas of the gallbladder 

are extremely rare: only 29 cases have been reported in the 

literature. They are morphologically similar to those that arise in 

other anatomic sites. By definition, more than 50% of the tumour 

contains extracellular mucin. 

Methods. The patient is a 59-year-old Italian man who underwent 

cholecystectomy under the preoperative diagnosis of cholecystitis 

with gallstones and gallbladder tumour. At laparotomy, 

cholecystectomy, liver wedge resection, and regional lymph node 

dissection were performed. The resected gallbladder showed 

thickened wall, gallstones and a 4 cm gelatinous, cauliflower-like 

soft tissue mass. Following surgery, the gallbladder tumor was 

diagnosed as a mixed endocrine–exocrine carcinoma. There was 

evidence of lymph node metastasis and direct liver invasion. The 

mucin-producing carcinoma was composed of poorly differentiated 

glandular cells with mucin lakes. The LCNEC was characterized 

by large cells with prominent nucleoli, coarse chromatin, 

and a high mitotic rate. The cells showed an “organoid” growth 

pattern with rosette formation and frequent areas of necrosis. 

Chromogranin A, synaptophysin and CD56 were diffusely and 

strongly expressed in the LCNEC component. 

Results. This case may provide helpful insights regarding the 

histogenesis of this unusual combination of tumors: the concept 

of a collision tumor between two neoplasms that have arisen in 

adjacent areas may be the best explanation for the pathogenesis.

354 

Breast lump in a male, first manifestation of 

occult pulmonary oat cell carcinoma: a rare case 

report. Cytological and immunocytochemical 

diagnosis in previously Hematoxylin and eosin 

stained cytologic material 

1)Russo S. 2)De sio A.L. 3)Artiola G. 4)Sepe J. 5)Paolini B. 

6)Maiello F.M. 

1)Pathology Dpt., Maresca Hospital, ASLNA3sud, Naples, Italy 2)Pathology 

Dpt., Pellegrini ASLNA1, Naples, Italy 3)Pathology Dpt., Pellegrini 

Hospital, ASLNA1, Naples, Italy 4)Biology, University of Maryland University 

College, Naples, Italy 5)Pathology Dpt., National Cancer Institute, 

Milan, Italy 6)Pathology Dpt., Pellegrini Hospital, ASLNA1, Naples, Italy. 

Background. Metastatic disease to the breast from extramammary 

sites is uncommon. It can be difficult to differentiate between 

primary breast cancer and a metastatic disease. It is important to 

make an accurate diagnosis as this has an impact on the therapeutic 

planning: an incorrect diagnosis can lead to unnecessary 

surgical interventions. This report describes a case in which a 

cytopathological diagnosis of blood-borne metastatic disease to 

the breast from primary lung oat cell carcinoma was carried out 

on Fine Needle Cytology (FNAC) samples. The patient was a 

male who presented initially with a palpable lump in the breast, 

without history of malignant disease. 

Methods. Primary site was identified on morphological and 

immunocytochemical (ICC) bases. ICC has a significant role in 

identifying the primary origin of tumor and has to be considered 

in the presence of unusual clinical and cytologic patterns. FNAC 

was carried out on the outpatient, using a 21-gauge needle, with 

suction: a 10 ml syringe and a Cameco (Cameco AB, Taby, Sweden) 

needle holder were used to perform the aspiration on the 

palpable lesion. The obtained material was smeared onto seven 

slides: smears were stained with Hematoxylin and eosin after wet 

fixation in 95% alcohol for immediate adequacy assessment onsite. 

Six of them were then processed for immunocytochemistry. 

Destaining was not required because the procedures for immunohistochemical 

staining remove the previous stain. 

Results. ICC stains demonstrated cytoplasmic positivity (even dotlike) 

for cytokeratins, chromogranin, synaptophisin and diffuse 

and strong nuclear staining for TTF1. Neoplastic cells showed no 

reactivity for p63 and CD45/LCA. A final diagnosis of metastatic, 

pulmonary oat cell carcinoma was reached that was confirmed 

radiologically. The distinction between metastatic small cell or 

poorly differentiated squamous pulmonary carcinoma, Merkel 

carcinoma and lymphoma can present a diagnostic problem. 

Preventive vaccination with telomerase controls 

tumor growth in genetically engineered and 

carcinoge-induced mouse models of cancer 

1)Sabatini (F). 2)Liberatore (M). 3)Pannellini (T). 4)Lazzaro 

(D). 5)Ciliberto (G). 6)Bronte (V). 7)Scarselli (E). 8)Iezzi (M). 

9)Musiani (P). 




SS. Annunziata/CESI, Chieti, Italia 4)Istituto Di Ricerca Di Biologia 

Molecolare, Istituto Di Ricerca Di Biologia Molecolare, Pomezia, 

Italia 5)Istituto Di Ricerca Di Biologia Molecolare, Istituto Di Ricerca Di 

Biologia Molecolare, Pomezia, Italia 6)Istituto Oncologico Veneto, Istituto 

Oncologico Veneto, Padova, Italia 7)Istituto Di Ricerca Di Biologia 

Molecolare, Istituto Di Ricerca Di Biologia Molecolare, Pomezia, Italia 




Background. The telomerase reverse transcriptase, TERT, is an 

attractive target for human cancer vaccination because its expression 

is reactivated in a conspicuous fraction of human tumors. 

Genetic vaccination with murine telomerase (mTERT) could 


break immune tolerance in different mouse strains and resulted 

in the induction of both CD4+ and CD8+ telomerase-specific 

T cells. The mTERT-derived immunodominant epitopes recognized 

by CD8+ T cells were further defined in these mouse strains 

and used to track immune responses. 

Methods. Antitumor efficacy of telomerase-based vaccination 

was investigated in two cancer models closely resembling human 

diseases: the TRAMP transgenic mice for prostate cancer and a 

carcinogen-induced model for colon cancer. TERT overexpression 

in tumor lesions was shown in both models by immunohistochemistry, 

thus reinforcing the similarity of these tumors to their 

human counterparts. 

Results. Repeated immunizations with mTERT-encoding DNA 

resulted in a significant delay of tumor formation and progression 

in both the prostate cancer and the colon cancer models. Moreover, 

evaluation of the intratumoral infiltrate revealed the presence 

of telomerase-specific T cells in vaccinated mice. The safety 

of vaccination was confirmed by the absence of histomorphologic 

changes on postnecropsy analysis of several organs and lack of 

adverse effects on blood cell counts. These results indicate that 

TERT vaccination can elicit antigen-specific immunosurveillance 

and imply this antigen as a potential candidate for preventive 

cancer vaccines. 

Avidinox for highly efficient tissue-pretargeted 

radionuclide therapy 

1)Sabatini (F). 2)Mariotti (M). 3)Ascione (P). 4)Leoni (B). 5)Chinol 

(M). 6)Carminati (P). 7)De Santis (R). 8)Musiani (P). 

1)Anatomia Patologica/Oncologia e Neuroscienze, SS. Annunziata/CESI, 

Chieti, Italia 2)Oncologia e Neuroscienze/Oftalmologia, SS. Annunziata/ 

CESI, Chieti, Italia 3)Anatomia Patologica/Oncologia e Neuroscienze, SS. 

Annunziata/CESI, Chieti, Italia 4)Department Of Immunology, Sigma-Tau 

Spa R&D, Roma, Italia 5)Istituto Europeo Tumori, Istituto Europeo Tumori, 

Milano, Italia 6)Department Of Immunology, Sigma-Tau Spa R&D, 

Roma, Italia 7)Department Of Immunology, Sigma-Tau Spa R&D, Roma, 

Italia 8)Anatomia Patologica/Oncologia e Neuroscienze, SS. Annunziata/ 

CESI, Chieti, Italia 

Background. Avidin is widely used in vitro for its capacity to 

bind biotin. However, avidin’s in vivo use is limited by its short 

residence in blood and tissues. 

Methods. An avidin variant, named AvidinOX, has been recently 

described. This product is obtained by 4-hydroxyazobenzene-20carboxylic 

acid-assisted sodium periodate oxidation of avidin. 

This method generates aldehyde groups from avidin carbohydrates, 

sparing biotin-binding sites from inactivation. AvidinOX 

binds cellular and interstitial protein amino groups through 

Schiff’s bases, resulting in a tissue halflife of 2 weeks, compared 

with 2 hours of native avidin. Binding of AvidinOX occurs in 

normal and neoplastic tissues. 

Results. Data show that AvidinOX, administered intranipple in 

the breast of transgenic BALB = neuT mice, is highly efficient 

for capturing 90Y-biotinDOTA, intravenously injected after 48 

hours, leading to eradication of multifocal cancer lesions. Efficacy 

data, together with good tolerability results, indicate that 

AvidinOX is a highly innovative reagent for tissue-pretargeted 

radionuclide therapy 

Pheno-genotypic identification of circulating 

tumor cells in uveal melanoma patients 

1)Salvianti F. 2)Pinzani P. 3)Pepi M. 4)Mazzini C. 5)Pazzagli M. 

6)Santucci M. 7)Massi D. 

1)Department of Clinical Physiopathology, AOUC, Florence, Italy 2)Department 

of Clinical Physiopathology, AOUC, Florence, Italy 3)Division 

of Pathological Anatomy, Department of Critical Care Medicine and 

Surgery, AOUC, Florence, Italy 4)Department of Oto-neuro-ophthalmology, 

AOUC, Florence, Italy 5)Department of Clinical Physiopathology, 

AOUC, Florence, Italy 6)Division of Pathological Anatomy, Department


of Critical Care Medicine and Surgery, AOUC, Florence, Italy 7)Division 

of Pathological Anatomy, Department of Critical Care Medicine and Surgery, 

AOUC, Florence, Italy 

Background. Analysis of circulating tumor cells (CTC) in the 

peripheral blood of uveal melanoma patients provides clinically 

useful information. The isolation by size of epithelial tumor 

cells (ISET/ScreenCell) is a direct method for CTC identification, 

in which tumor cells are collected by filtration, because 

of their large size. Upon isolation, the identity of tumor cells as 

melanoma cells can be supported by immunohistochemistry and 

their presence indirectly supported by mRNA tyrosinase levels. 

Recently a Four-color FISH probe targeting the loci 6p25, 6q23, 

11q13 and the centromeric region of chromosome 6 (CEP6) has 

been devised by Abbott Molecular Laboratories-USA to identify 

chromosomal abnormalities in melanocytic lesions. We herein 

investigated the presence and clinical significance of ISET/ScreenCell-isolated 

CTC in uveal melanoma patients. The identification 

of CTC was corroborated by suitable immunohistochemical 

and FISH analysis. 

Methods. Forty-one patients with uveal melanoma were longitudinally 

investigated over a period of 5 years. Blood tyrosinase 

mRNA levels were assessed by quantitative RT-PCR. Results 

were correlated with clinical data and, in a subgroup of patients, 

with the number of CTC assessed by ISET/ScreenCell. The identity 

of cells, trapped in filters, as CTC was supported by positivity 

for immunohistochemical markers (S-100 protein, HMB-45, 

MART-1/Melan A) and, in selected cases, by presence of chromosomal 

abnormalities by FISH analysis. 

Results. Increased tyrosinase mRNA levels were found in 20 of 

41 (49%) uveal melanoma patients and mRNA tyrosinase levels 

correlated with tumor dimension (p < 0.01), disease-free and 

overall survival (p < 0.05). CTC were isolated by ISET/Screen- 

Cell in 5/16 patients and a direct correlation was found between 

CTC values and tyrosinase levels. Our findings encourage further 

exploration of immunohistochemistry and FISH analysis for CTC 

identification and support the clinical significance of melanoma 

circulating cells for the work-up and choice of appropriate therapies 

in uveal melanoma patients. 

The diagnostic challenge of gastrointestinal 

melanomas: a retrospective analysis of 42 cases 

Raffaella Santi1 , Paola Apicella2 , Mauro Biancalani3 , Camilla Eva 

Comin1 , Morena Doria2 , Augusto Giannini4 , Vincenza Maio1 , Luca 

Messerini1 , Clelia Miracco7 , Luca Novelli1 , Milena Paglierani1 , 

Lavinia Pugliese4 , Armando Rossi5 , Marco Santucci1 , Carmelo 

Urso6 , Carla Vindigni7 , Federica Zolfanelli8 , Daniela Massi1 1Division of Pathological Anatomy, Department of Critical Care Medicine 

and Surgery, University of Florence; 2Pathology Unit, ASL 3 Pistoia; 3Pa thology Unit, ASL 11 Empoli; 4Pathology Unit, ASL 4 Prato; 5Pathology Unit, ASL 9 Grosseto; 6Dermatopathology Section, S.M. Annunziata Hospital, 

ASL 10, Florence; 7Department of Human Pathology and Oncology, 

University of Siena; 8Pathology Section, Ospedale Nuovo S. Giovanni 

di Dio, ASL 10, Florence 

Background. Malignant melanoma involving the gastrointestinal 

(GI) tract is mainly related to metastatic disease while primary mucosal 

melanomas are exceedingly rare. Despite multimodal therapeutic 

approach, primary GI melanomas are associated with a poor 

prognosis. Recently it has been emphasized that affected patients 

may benefit from imatinib mesylate (Gleevec) targeted therapy. 

Methods. Forty-two GI melanomas were retrospectively analyzed. 

Selected cases were submitted to appropriate immunohistochemistry 

(c-KIT, S100 protein, HMB-45 and/or MART-1) and 

7 cases with clear cell features were submitted to FISH analysis 

to exclude the presence of a t(12;22)(q13;q12) translocation (r/o 

clear cell sarcoma). 

Results. There were 25 females and 17 males, with a mean age 

of 70.8 years (range 45-89 years). Clinical presentation included 

355 

abdominal pain, palpable mass, diarrhea, GI bleeding, obstruction 

or weight loss. The most common anatomical location was 

the anorectal region (n = 26), followed by the large bowel (n = 6), 

small bowel (n = 6), stomach (n = 3) and oesophagus (n = 2). 

Mean tumour size was 4.4 cm (range 0.4-12.5 cm). Most cases 

presented as exophytic, diffusely ulcerated lesions. An adjacent in 

situ melanoma component was detected in 6/26 anorectal melanomas. 

Twenty cases were amelanotic; melanin pigment, at least 

focally, was present in 22 cases. In 34 patients submitted to surgical 

resection, 17 showed invasion of the perivisceral fat, whereas 

in the remaining cases melanomas were confined to the visceral 

wall. In the 19 patients with loco-regional nodal resection, 15/19 

patients (79%) showed metastatic disease at presentation. On the 

basis of the morphological appearance and available clinical history, 

the cases were tentatively classified as primitive (n = 6, 14%), 

metastatic or putative metastatic GI melanomas (n = 36, 86%). In 

conclusion, the identification of GI melanomas results in problems 

relating to their histogenesis, lack of conventional histopathological 

prognostic factors for appropriate staging and determination of 

their primary or secondary nature. A potential metastatic nature 

remains difficult to be formally excluded, because GI localization 

can precede the identification of a primary site or may results from 

an unknown or fully regressed primary cutaneous melanoma. 

evaluation of Notch receptors in a case of adult 

Wilms tumour 

1)Santi R. 2)Paglierani M. 3)Villari D. 4)Pepi M. 5)Nicita G. 

6)Massi D. 7)Nesi G. 

1)Divisione di Anatomia Patologica, Università di Firenze, Azienda Ospedaliero 

Universitaria Careggi, Firenze, Italia 2)Divisione di Anatomia 

Patologica, Università di Firenze, Azienda Ospedaliero Universitaria Careggi, 

Firenze, Italia 3)Clinica Urologica, Università di Firenze, Azienda 

Ospedaliero Universitaria Careggi, Firenze, Italia 4)Divisione di Anatomia 

Patologica, Università di Firenze, Azienda Ospedaliero Universitaria 

Careggi, Firenze, Italia 5)Clinica Urologica, Università di Firenze, 

Azienda Ospedaliero Universitaria Careggi, Firenze, Italia 6)Divisione di 

Anatomia Patologica, Università di Firenze, Azienda Ospedaliero Universitaria 

Careggi, Firenze, Italia 7)Divisione di Anatomia Patologica, Università 

di Firenze, Azienda Ospedaliero Universitaria Careggi, Firenze, 

Italia 

Background. Wilms tumour (nephroblastoma) is the most common 

renal neoplasm of childhood, rarely occurring in adults. 

With respect to histological and immunophenotypic features, 

no differences emerge between adults and children. Both the 

morphology and the genetic profile of Wilms tumour suggest 

an intimate relationship with kidney development. Since Notch 

signalling is known to be implicated in many developmental processes, 

including nephrogenesis, a role for Notch pathway components 

in the oncogenesis of this tumour may be hypothesised. 

We investigated Notch receptor expression in an adult case of 

nephroblastoma. 

Methods. A 34-year-old female patient developed lung metastasis 

48 months after radical nephrectomy for Wilms tumour. 

Haematoxylin-eosin and immunohistochemically stained sections 

of both primary and metastatic tumours were reviewed. Additionally, 

immunohistochemical analysis of Notch-1 and Notch-2 was 

performed. 

Results. Microscopically, the renal neoplasm consisted of a 

tubular and glandular proliferation, strongly immunoreactive for 

WT-1, confirming the previously established diagnosis of Wilms 

tumour with a predominant epithelial component. The tumour 

maintained the same morphology in the pulmonary metastasis. 

Immunohistochemical assay showed diffuse and intense Notch-1 

and Notch-2 expression both in the primary and in the metastatic 

lesions. 

Conclusions. Preliminary results of the ongoing pilot study we 

are carrying out on a series of paediatric nephroblastomas indicate 

that Notch-1 and Notch-2 expression is decreased in the

356 

blastema compared to the neoplastic epithelial component and the 

adjacent kidney parenchyma. These results are consistent with the 

protein expression findings in the adult case herein described, and 

suggest that Notch pathway may be implicated in the oncogenesis 

of Wilms tumour. 

A mediastinal time bomb in a young man, 

a rare case of coronary aneurysm 

1)Santise G. 2)Minervini MI. 3)Schicchi R. 4)D’Ancona G. 

5)Sciacca S. 6)Turrisi MA. 7)Pilato M. 

1)Chirurgia cardiotoracica, Ismett, Palermo, Italia 2)Anatomia patologica, 

Ismett, Palermo, Italia 3)Cardiologia, Buccheri la ferla “fatebenefratelli”, 

Palermo, Italia 4)Chirurgia cradiotoracica, Ismett, Palermo, Italia 

5)Chirurgia cradiotoracica, Ismett, Palermo, Italia 6)Chirurgia cradiotoracica, 

Ismett, Palermo, Italia 7)Chirurgia cradiotoracica, Ismett, Palermo, 

Italia 

Introduction. A 34 year old young man was referred to our 

institution with history of chest pain, cardiac enzymes raising, 

episodes of arrhythmias and a not better specified diagnosis of 

pericardial mass at the echocardiography. 

Method. The patient underwent a chest CT scan that confirmed 

a huge coronary aneurysm of the right coronary compressing the 

right ventricle. The patient was anyway referred for surgical resection 

of the coronary aneurysm and coronary artery bypass graft. 

Pre-operatively, the surgical strategy was carefully planned. 

Intra-operatively, an aneurysm 10 cm in greatest dimension was 

found, with a thin wall and intramural hematoma ready to tear. 

No important signs of atherosclerosis nor arteritis were seen. 

Specimens were submitted to pathology. 

Results. Macroscopic examination revealed laminar fragment 

measuring 12.5 × 8.5 × 0.2 cm in maximum diameter with a focal 

presence of laceration of the endothelium and a thrombotic lesion 

within the wall measuring 3.0 × 3.0 cm. 

Histological findings showed coronary aneurysm associated with 

mild atherosclerosis, mural necrosis and cystic medial degeneration. 

Pools of mucin material, highlighted by Mucicarmine positive 

stain, were found within the wall. Intramural hematoma and 

marked adventitial hemorrage were also seen. No arteritis was 

appreciated. 

Overall, the findings described appeared very similar to the ones 

commonly present in the aortic wall of patients with Marfan’s 

syndrome. 

The postoperative course was straight forward: the patient was 

discharged in postoperative day 6 and referred for genetic analysis 

to rule out Marfan’s disease. 

Conclusion. This case highlights the possibility of a very rare 

localization of an aneurysm most likely due to a connective disorder. 

A multidisciplinary approach and a strict follow-up is highly 

recommended to achieve the diagnosis of such rare occurrences. 

Cryptococcal infection presenting as cellulitis 

in a renal transplant recipient 

1)Santoro A. 2)Giallella M. 3)Punzi A. 4)Corsi F. 5)Pennella A. 

6)Serio G. 

1)Acienze chirurgiche, Università di foggia, Foggia, Italy 2)Scienze 

chirurgiche, Università di foggia, Foggia, Italy 3)Anatomia patologica, 

Università di Bari, Bari, Italy 4)Anatomia patologica, Università di 

Bari, Bari, Italy 5)Scienze chirurgiche, Università di foggia, Foggia, Italy 

6)Anatomia patologica, Università di Bari, Bari, Italy 

Background. Cryptococcus neoformans is an encapsulated, 

basidiomycetous yeast that is present in the environment worldwide. 

Cryptococcosis is a significant opportunistic infection in 

solid-organ transplant recipient, with a reported incidence of 

1-5% and mortality of 20-40%. Immunodepressed hosts with disseminated 

Cryptococcosis usually present with central nervous 

system or pulmonary involvement. Skin lesions occur in approxi- 


mately 10-15% of cases. Cellulitis is rare, having been reported 

in only 16 patients. 

Methods. We describe a case of Cryptococcal cellulitis in a 

renal transplant recipient. A 45.year-old man with chronic renal 

allograft underwent renal transplantation in 1999 for end-stage renal 

disease of unknown cause. After 11 years, in February 2010, 

patient was admitted to Foggia Hospital, in the Nephrology Unit, 

with a 10-day history of aspecific fever and with several erythematous, 

tender macules measuring 0,5 cm in diameter in right 

thigh. The first clinic suspicious was for a lympho-proliferative 

disease. A skin and a bone-marrow biopsy were performed. 

Results. Bone-marrow biopsy was negative. Histologycal examination 

of skin biopsy specimen disclosed a panniculitis with extension 

into the overlying dermis. Haematoxylin and eosin stains 

demonstrated a subacute-chronic granulomatous inflammation. 

The periodic acid Schiff (PAS) and Grocott Gomori’s methenamine 

silver showed round to oval 3-6µ budding yeast within the 

granulomatous inflammation. In addition, Mucicarmine, Alcian 

Blue, Methylene Blue and Fontana-Masson stains were positive 

showing encapsulated organisms most consistent with Cryptococcus 

neoformans. This report underscores that patients with 

cutaneous Cryptococcosis should be thoroughly evaluated, as it 

may be the first manifestation of a systemic disease. A prompt 

histological diagnosis may allow for an early therapy and improve 

patient survival. 

Aurora B expression as a prognostic marker 

and therapeutic target in oral cancer 

1)Santoro A. 2)Pannone G. 3)Hindi SAH. 4)Sanguedolce F. 

5)Rubini C. 6)Tortorella S. 7)Cagiano S. 8)Pedicillo C. 9)Lo 

muzio L. 10)Bufo P. 

1)Department of surgical sciences - section of anato, Riuniti, Foggia, Italy 


3)2 section of oral pathology, Babylon university, Babylon, Iraq 4)Department 

of surgical sciences - section of anato, Riuniti, Foggia, Italy 5)Section 

of anatomic pathology, Università politecnica delle marche, Ancona, 

Italy 6)Section of anatomic pathology, Irccs crob - centro di riferimento 

oncologico di b, Irccs crob - centro di riferimento oncologico di b, Italy 



9)Department of surgical sciences, Irccs crob - centro di riferimento oncologico 

di b, Irccs crob - centro di riferimento oncologico di b, Italy 10)Department 

of surgical sciences - section of anato, Riuniti, Foggia, Italy 

Background. Aurora B serine-threonine kinase is a member of 

the chromosomal passenger family of proteins, along with inner 

centromere protein (INCENP) and survivin. Several evidences 

have suggested that Aurora B overexpression is related to invasiveness 

and clinical outcome in many solid tumours. 

Methods. The aim of this study was to investigate the expression 

of the chromosomal passenger protein Aurora B and its phosphorylated 

form in a large series of human oral squamous cell cancers 

(OSCC) and to evaluate its clinical and prognostic significance. 

Western blotting analysis revealed overexpression of both Aurora 

B and Thr-232 Phopsho-Aurora B in OSCC lines as compared 

to normal keratinocytes and bladder cancer cells. Furthermore, 

protein expression was analysed by immunohistochemistry in 

101 OSCC of different site, stage and histological grade and in 

normal peritumoural areas. 

Results. The intracellular localization of Aurora B in tumour 

cells was mainly nuclear, especially in proliferative areas, and 

significant overexpression was found in tumours in comparison 

to normal peritumoural areas (p = 0,012). Staining results were 

correlated with clinicopathological parameters and long-term 

follow-up, and a significant association was found between 

protein expression and tumour stage (stage II, III and IV vs. 

stage I, p = 0,030) and size (< 2 cm vs > 2 cm, p = 0.010). Cox 

regression analysis confirmed a poorer disease-free survival in 

cases with high expression of Aurora B protein. Kaplan-Meier


curves showed shorter time to progression in patients with high 

levels of Aurora B expression (p < 0.05). Moreover, the tumoral 

group with nuclear Aurora B immunolocalization had the worst 

prognosis (p = 0.0364 in disease free survival). Our results suggest 

that assessing Aurora B expression might help in patients’ 

risk stratification and serve and as a novel therapeutic target in 

advanced OSCCs. 

Bone-marrow tubercular involvement after 

intravescical BCG instillation for bladder cancer 

1)Santoro A. 2)Punzi A. 3)Giallella M. 4)Corsi F. 5)Pennella A. 

6)Serio G. 

1)Scienze chirurgiche, Università di foggia, Foggia, Italy 2)Anatomia patologica, 

Università di bari, Bari, Italy 3)Scienze chirurgiche, Università 

di foggia, Foggia, Italy 4)Anatomia patologica, Università di bari, Bari, 

Italy 5)Scienze chirurgiche, Università di foggia, Foggia, Italy 6)Anatomia 

patologica, Università di bari, Bari, Italy 

Background. Bacillus Calmette-Guérin intravescical administration 

was introduced as prophylaxis and treatment in bladder high 

risk superficial cancer and carcinoma in situ by Morales et al. in 

1976. While it is generally well tolerated, BCG instillation is not 

without complications, the most frequent problems being local side 

effects such as granulomatous cystititis. Systemic adverse reactions, 

such as sepsis, hepatitis, pulmonary granulomatosis as well 

as bone marrow involvement have been reviewed by Lamm et al. 

Methods. We report the case of a tubercular granuloma in a 59 

year-old man with a history of renal chronic insufficiency, pulmonary 

squamous carcinoma (surgically removed and chemotherapeutically 

treated) and of bladder in situ carcinoma, treated with 

intravescical BCG. The first six BCG instillation were given on a 

weekly basis. Thereafter, the BCG was instilled every month. After 

instillation n.8, fever and severe weakness developed. On admission 

to Foggia Hospital, at the Nephrology Unit, laboratory exams 

revealed leucopoenia, eritropoenia, microhematuria and proteinuria. 

Koch bacillus was negative in the urine. At the immunofixation 

a seric monoclonal IgGγ component and a urinary monoclonal κ 

spike were observed, thus enforcing a clinic suspicious for a lympho-proliferative 

disorder. A bone-marrow biopsy was performed. 

Results. Pathologic evidence of granulomatous myelitis was confirmed 

on routinary Haematoxylin and Eosin sections. A small 

granuloma containing giant and epithelioid cells showed a typical 

central caseous necrosis. Ziehl-Neelsen stains confirmed the presence 

of acid fast organisms in the focal granuloma. Moreover, 

CD138 immunostaining revealed a faint increase of plasma cells. 

Finally, Gomori stain showed a mild fibrosis. The case report 

reminds that this rare but life threatening bone marrow involvement 

should be considered when systemic symptoms develop 

after BCG treatment. 

Identification of novel cryptic chromosomal 

abnormalities in primary myelofibrosis by 

single-nucleotide polymorphism oligonucleotide 

microarray 

Maria Rosaria Sapienza1 , Giuseppe Visani2* , Alessandro Isidori2 , 

Simona Righi1 , Antonella Laginestra1 , Claudio Agostinelli1 , Elena 

Sabattini1 , Michele De Nictolis3 , Massimo Valentini4 , Meris 

Donati4 , Roberto Emiliani4 , Anna Gazzola1 , Claudia Mannu1 , 

Maura Rossi1 , Carlo Finelli1 , Nicola Vianelli1 , Stefano A. Pileri1 , 

Pier Paolo Piccaluga1 1Department of Hematology and Oncology “L. e A. Seràgnoli”, Hematopathology 

and Hematology Sections, Molecular Pathology Laboratory, 

Sant’Orsola-Malpighi Hospital, University of Bologna, Bologna, Italy; 

2Hematology and Hematopoietic Stem Cell Transplant Center, San Salvatore 

Hospital, Pesaro, Italy; 3Department of Pathology, San Salvatore 

Hospital, Pesaro, Italy; 4Clinical Pathology Laboratory, San Salvatore 

Hospital, Pesaro, Italy. 

357 

Background. The molecular genetics of primary myelofibrosis 

(MF) is poorly known at present. 

In this study we performed high resolution karyotyping by SNP 

oligonucleotide microarray by using the most updated Affymetrix 

array (Genome-Wide Human SNP Array 6.0) in 20 cases of 

myelofibrosis (MF) in order to identify novel cryptic genomic 

aberrations. 

Methods. DNA was extracted from lymphocytes-depleted 

PBMNC of 14 primary and 6 secondary MF patients. DNA was 

then processed and hybridized to the Affymetrix SNP arrays 6.0 

as for manufacturer instruction. A whole-genome copy number 

variation (CNV), was performed using the Partek Suite 6.0. Ten 

lab-specific as well as 90 HapMap samples relative to Caucasian 

healthy donor were used as control reference. Genomic abnormalities 

were defined as recurrent when occurring in at least 25% 

of cases. JAK2 mutational status was assessed by alle-specific 

PCR. Clinical information and complete follow-up were retrieved 

for all cases. Direct sequencing, FISH, qPCR and immunohistochemistry 

(IHC) has been chosen for validation. 

Results. In all patients we could detect several CNV. The median 

number of CNV was 60 (range, 34-72), including 46 amplifications 

(A) and 14 deletions (D). All commonest previously described 

abnormalities were detected. In addition, several formerly 

uncovered recurrent lesions were identified, mainly involving 1p, 

1q, 2p, 4p, 4q, 5q, 6p, 6q, 7q, 8p, 9q 10q, 11p 11q, 12p, 14q, 15q, 

16p, 16q, 17q, 18q, 19q, 20p, 22q. Of note, numerous definite 

aberrations (A or D) distinguished JAK2 + vs. JAK2 - cases, specifically 

affecting 16q23.1, 1p36.13, 3q26, 14q13.2, 5q33.2, 6q14.1, 

7q33, 8p23.1, and 9p11.2.Grippingly, several genes of potential 

interest for PMF pathogenesis were identified within the involved 

loci, including RET, SCAPER, WWOX and SIRPB1. Among others, 

the product of such genes has been selected for validation 

by IHC. Similarly, many miRNA were recognized, which may 

deserve further investigation. 

Conclusions. By using a newly developed highly sensitive array 

we identified novel cryptic lesions in patients affected by MF. 

Future studies on larger series, as well as functional analyses will 

definitely assess their role in the pathogenesis of the disease. Of 

note, consistent differences were recorded in JAK2 + vs. JAK2 - , 

supporting the hypothesis of different genetic mechanisms occurring 

in the two sub-groups. 

Presacral myelolipoma 

Scamarcio R., Colagrande A., Angelotti UF., Scivetti A., Ingravallo 

G., Cimmino A. 


Background. Myelolipoma (ML) is a rare benign tumour that 

can usually originate in the surrenal glands, but also in extraadrenal 

sites like pelvis or thorax. It is hormonally inactive, and is 

symptomatic only in case of large neoplasm for extrinsic compression 

on close organs. 

Methods. A 71 years old female patient, during follow-up examination 

for untreated hepatocellular carcinoma of hepatic segment 

VIII, underwent to CT scans that detected large pelvic mass 

apparently clivable from rectum. Subsequently, a laparoscopy 

surgical resection of pelvic presacral tumor (8 cm in its largest 

diameter) was performed. 

Results. Overall, the tumor was well encapsulated and constituted 

by lobules of typical lipomatous cells intermingled with normal 

haematopoietic tissue (myeloid and eritroid cells and rare megacarioblasts 

and megacariocytes). The final diagnosis was ML. Presacral 

MLs require an accurate histopathological characterization 

because are radiologically confused with malignant retroperitoneal 

tumors, which are more common, and as well, they should be differentiated 

from mass-forming extramedullary hematopoiesis (i.e. 

extramedullary hematopoietic tumors), which are ill defined and

358 

lack fat and are associated with symptomatic status like myeloproliferative 

diseases and haemolytic anemia. At the moment, the 

pathogenesis of ML is unclear and it is supposed that continuous 

inflammatory conditions may stimulate mesenchymal stem cells 

to differentiate to adipocytes and hematopoietic cells. 

umbilical endometriosis 

Scamarcio R., Colagrande A., Angelotti U.F., Scivetti A., 

Traversi C., Cimmino A. 


Background. Endometriosis is a very common gynaecological 

pathology, even though the localization in the abdominal wall is a 

rare clinical problem (0,5-1%). Its rarity explains the fragmentariness 

of reports in literature. 

Methods. We report a clinical case of umbilical endometriosis in 

a patient who has never been treated with surgery or laparoscopy. 

V.I., 25 years old, presents an umbilical neoformation of brownish 

color since March 2009, increased in volume day by day. A 

sporadic siero hematic secretion is present in correspondence 

with the menstrual occurrence. In December 2009, because of 

the suspect of a neoplastic process she is subjected to incisional 

biopsy. The macroscopic report consists in an irregular brownish 

fragment of the diameter of 0,7 cm. The diagnosis is of umbilical 

polyp including embryonic residual in the form of a tubular structure 

delimited by columnar epithelium (referable to a residual of 

vitellin duct). She then performs an echographic inspection and 

RMN, through which a polypoid formation of the diameter of 

15 mm is observed, contracting relations of contiguity with the 

intestinal ansa. A radical removal of the umbilicus is performed. 

The macroscopic report consists in a polypoid neoformation of the 

diameter of 3 cm. The diagnosis: cutis and subcutis of polypoid 

appearance including a remarkable endometriosic focus with deposition 

of hemosiderinic pigment. This shows the importance of a 

complete exsection of the neoformation for a correct diagnosis. 

Results. Theory regarding the pathogenesis of primitive umbilical 

endometriosis: the umbilical cord removed at the moment of 

birthing process can be contaminated by other endometrial cells 

of the mother, which have been released during the phase of 

birthing in the same way reported in literature during caesarean 

sections, laparoscopies and amniocentesis. 

Comparison of different pathologic protocols 

for evaluation of sentinel lymph node in breast 

cancer 

1)Scarpellini F. 2)Vitali P. 3)Nuzzo F. 4)Nigrisoli E. 

1)Anatomia patologica, Bufalini, Cesena, Italia 2)U.o. epidemiologia e 

comunicazione, Ausl, Cesena, Italia 3)Anatomia patologica, Bufalini, Cesena, 

Italia 4)Anatomia patologica, Bufalini, Cesena, Italia 

Background. The sentinel lymph node (SLN) biopsy is actually 

considered the ideal procedure for breast cancer staging and represent 

a fundamental step in the management of this neoplasia. 

Different protocols in pathologic evaluation of SLN are reported 

in literature. The aim of our study is to compare the detection 

of sentinel lymph node metastases by using three different stepsectioning 

methods. 

Material and Methods. 248 SLN were examined in our Department 

in the period 2005-2008, according to FONCAM protocol. 

The FONCAM protocol used in our Department consists in completely 

sectioning the SLN at 50 micron intervals for 15 sections 

and successively at 100 micron intervals. 

All cases were present in our archive and were reviewed, selecting 

sections according to the two regional different protocols. 

In the “optimal” protocol proposed by the Regione Emilia Romagna 

(RER), the block is completely sampled by steps sectioning 

at 200 micron intervals. 


In the “essential” method proposed by the RER 4 levels at 200 

micron intervals are provided. 

Results. In our series of 248 SLN examined according FONCAM 

protocol 172 negative and 76 positive lymph nodes were obtained. 

Positive cases were 45 metastasis and 31 micrometastasis. 

Adopting the two alternative protocols (RER guidelines) for SLN 

examination (optimal and essential) all the original diagnosis 

were confirmed, excepting for 1 micrometastasis. 

The sensibility of the two new methods was 98.7%, the specificity 

was 100%. 

There were non statistic differences between FONCAM and regional 

protocols in Mc Nemar test (p = 1.0). 

Conclusions. The diagnostic accuracy of the thee compared 

methods for SLN pathologic processing is equivalent. 

Not so the time consuming and the work load for Pathologists 

and Technicians. 

Double p16 INK4A /KI67 staining and interobserver 

agreement in cin diagnosis 

Schiavo N., Barresi E., Paniccià bonifazi A., Reghellin D., Rucco 

V., Lestani M. 

U.O.C. Anatomia Patologica, ULSS 5 “Ovest Vicentino”, Arzignano (Vi), 

Italia 

Background. Reproducibility in diagnosis of CIN on cervical 

biopsies are linked to different factors (i.e. experience of pathologist 

and quality of sections). Previous studies have shown p16 

role in cervical lesions and utility of p16 INK4A in the assessment of 

cervical dysplasia, on citological smears and biopsies. Target of 

this study was the evaluation of inter-observer concordance, using 

H&E alone or a double p16 INK4A /Ki67 staining, grading CIN 

in cervical biopsies. 

Material and methods. 57 specimens, subdivided in negative 

(11 cases), HPV infection (19 cases), CIN-1 (16 cases), CIN-2 

(7 cases), CIN-3 (4 cases) were examined by 4 pathologists; CIN 

grade (sec. WHO 2003) and differentiation between CIN-1 and 

HPV infection were evaluated. Different cut-off for p16 INK4A / 

Ki67 positivity were previously estabilished in order to evaluate 

intraepithelial lesions. Agreement between all 4 pathologist was 

then verified for each case. 

Results. Significant improvement of complete agreement (4 on 

4 pathologists) in CIN lesions was obtained using immunohistochemestry 

(from 72% to 93%). 

Most of disagreements, using H&E sections, were in distinction 

between negative vs focal HPV infection and HPV vs CIN-1 

(72,5% of total disagreements). 

Using double p16 INK4A /Ki67 immunostaining distinction between 

HPV vs CIN-1 was clear (92% of agreement) but not between 

negative vs HPV cases. 

Conclusions. Double p16 INK4A /Ki67 staining is an useful marker 

for cervical neoplasia grading and it helps in the distinction between 

HPV infections from CIN-1. 

Poor agreement was observed in the distinction of negative cases 

from focal HPV infection, either morphologically either with immunohistochemestry. 

Sarcomatous malignant peritoneal mesothelioma: 

a case report 

Schiavo N., Paniccià Bonifazi A., Rucco V., Reghellin D., 

Barresi E., Lestani M. 

U.O.C. Anatomia Patologica, Ulss 5 “Ovest Vicentino”, Arzignano (Vi), 

Italia 

Background. Malignant peritoneal mesothelioma (MPM) is a 

rare neoplasm (20% of all mesotheliomas; incidence of 1 per 

1,000,000) with a rapid fatal course (median survival: 6-12 

months). The sarcomatous subtype is considered exceptional.


High level of asbestos exposure has been reported in only 30- 

50% of cases. 

Methods. A 45 years old woman presented with fever and generalized 

abdominal pain, abdominal swelling, anorexia, and asciteshttp://www.ncbi.nlm.nih.gov/pubmed/15849996. 

No history 

of asbestos exposure was known. A voluminous solid lesion at 

the right ovary with multiple abdominal-pelvic localizations was 

radiologically identified. Hysterectomy, bilateral annessiectomy, 

resection of rectum, omentectomy and multiple peritoneal biopsies 

were performed. 

Results. Macroscopically right ovary was completely replaced 

by a solid grey mass with large areas of necrosis. Uterine surface, 

parametrium, left ovary, omentum, and rectum were massively 

involved. 

Microscopically neoplasm showed a double pattern: a sarcomatous-undifferentiated 

component (more than 95% of the lesion) 

composed by epitelioid-spindled anaplastic cells with massive 

haematic embolization and large areas of necrosis; focal betterdifferentiated 

neoplastic foci were identified on peritoneum and 

omentum. They were composed by cubical cells with eosinofilic 

cytoplasm and evident nucleoli organized in a tubular, cord-like 

and “pseudo glandular” pattern. 

Immunohistochemestry revealed positivity of the common mesothelial 

markers in the better-differentiated epihelioid component 

(calretinin, WT1, EMA, CK7, vimentin) and negativity for 

BerEp4 and E-caderine. Sarcomatous component was strongly 

positive for vimentin, disomogenously for EMA, with weak-focal 

positivity for calretinin. 

Conclusions. We described an unusual form of peritoneal mesothelioma 

with a sarcomatous/indifferentiated hystology. Only 

accurate sampling and examination of multiple neoplastic foci 

allowed a correct diagnosis, by identification of more differenziated 

tubular/papillary patterns. 

erCC1 expression study in metastatic gastric cancer 

patients treated with fOlfOX6 

1)Schirosi L. 2)Sartori G. 3)Fontana A. 4)Losi L. 5)Luppi G. 

6)Reggiani bonetti L. 7)Del giovane C. 8)Bertolini F. 9)Conte 

PF. 10)Maiorana A. 

1)Integrated activity n 7-sec. pathologic anatomy, Policlinico of modena, 

Modena, Italy 2)Integrated activity n 7-sec. pathologic anatomy, Policlinico 

of modena, Modena, Italy 3)Integrated activity of oncology and haematology, 

Policlinico of modena, Modena, Italy 4)Integrated activity n 7-sec. 

pathologic anatomy, Policlinico of modena, Modena, Italy 5)Integrated 

activity of oncology and haematology, Policlinico of modena, Modena, 

Italy 6)Integrated activity n 7-sec. pathologic anatomy, Policlinico of modena, 

Modena, Italy 7)Integrated activity of oncology and haematology, 

Policlinico of modena, Modena, Italy 8)Integrated activity of oncology 

and haematology, Policlinico of modena, Modena, Italy 9)Integrated activity 

of oncology and haematology, Policlinico of modena, Modena, Italy 

10)Integrated activity n 7-sec. pathologic anatomy, Policlinico of modena, 

Modena, Italy 

Background. Gastric cancer is the second leading cause of cancer-related 

deaths. The newly developed cytotoxic drugs have 

partially improved response rates, often with adverse events. On 

this base, diagnostic techniques that can predict clinical outcomes 

are fundamental. Aim of this study was to determine the expression 

of Excision Repair Cross-Complementing gene 1 (ERCC1) 

and correlate it to clinical outcome. 

Methods. Between May 2005 and February 2009, 54 patients 

(pts), affected by metastatic gastric adenocarcinoma, were enrolled 

and treated with FOLFOX6 regimen. From 1 to 5 representative 

primary tumor sections from paraffin embedded formalin 

fixed samples were used to evaluate mRNA ERCC1 levels by 

quantitative Real Time PCR (qRT-PCR). Relative gene expression 

quantifications were calculated according to 2-∆∆Ct method 

using β-actin and β2-microglobulin gene as endogenous control 

and normal gastric mucosa as calibrator. Moreover ERCC1 and 

359 

Thymidylate Synthase (TS) expression was evaluated on representative 

tumor sections by immunohistochemistry (IHC). 

Results. Of 54 pts enrolled (M/F 40/14, median age at diagnosis 

66 years: range, 22-83), 53 underwent at least one cycle of chemotherapy 

and were evaluable for toxicity and response. ERCC1 

qRT-PCR was performed on 39 pts. The median value (0.78, 

range 0.17-2.67) was assigned as the cut-off value to divide pts 

into two groups (high or low ERCC1 mRNA values). ERCC1 and 

TS IHC were performed in 44 cases. In IHC, ERCC1 was positive 

in 32 pts (72.7%), negative in 12 (27.3%), while TS positive in 38 

pts (86.4%), negative in 6 (13.6%). We did not find any statistically 

significant correlation of ERCC1 (in IHC or qRT-PCR) and 

TS (in IHC) with response rate, disease control, time to progression 

and overall survival. 

Conclusion. No correlation with ERCC1 and TS expression was 

found in pts treated with FOLFOX6 as first line chemotherapy in 

metastatic gastric cancer. No correlation was also found between 

mRNA and protein levels of ERCC1. 

A malignant mesenteric extra-gastrointestinal 

stromal tumor (eGIST): a case report 

Scivetti A., Angelotti U.F., Colagrande A., Scamarcio R., 

Traversi C., Cimmino A. 


Background. GIST is the designation for a major subset of gastrointestinal 

mesenchymal tumors that histologically, immunohistochemically, 

and genetically differ from typical leiomyomas, 

leiomyosarcomas and schwannomas. They affect with higher 

frequency the stomach and small bowel. In fewer than 5% of 

cases, they originate primarily from the mesentery, omentum or 

peritoneum. 

Methods. A 73 years-old man was referred to our hospital for 

abdominal pain and dyspnea. On computed tomography (CT) 

we observed a huge abdominal mass, of irregular shape, with 

heterogeneous low-density, located between the bowel loops and 

occupying the entire abdomen. At laparotomy, a large solid/cystic 

mass measured 15 cm at its largest point, arising from the mesentery 

and parietal peritoneum, was present. A suspect subcutaneous 

nodule was removed in the same operative session. 

The abdominal mass consisted of intermingling solid and cystic 

component. Histopathologically, it was composed of atypical sort 

spindle-shaped epithelioid cells with an interlacing bundle pattern 

with the nuclei showing a focal palisading disposition. Immunohistochemically, 

the tumor was positive for c-KIT (CD117) and 

vimentin, weakly and slightly positive for alpha-smooth muscle 

actin (SMA), but negative for CD34, CK pool, S-100 protein, 

WT-1, EMA, GFAP, calretinin, synaptophysin, chromogranin. 

The Ki-67 labeling index was 15%, MI 8/50 HPF. The patient 

died for accidents related to chemotherapy. 

Results. In our case EGIST of the mesentery showed clinicopathological 

and immunohistochemical characteristics similar 

to those erosen from the digestive tract. It supports the hypothesis 

that these tumors originate from extragastrointestinal c-KIT 

positive cells. Mesenteric location appears to be associated with 

a poorer prognosis. 

Clinico-pathological and molecular findings in 

carcinoid tumors of the kidney 

Segala D., Gobbo S., Bersani S., Brunelli M., Martignoni G. 

Department of Pathology, University of Verona, Policlinico “G.B. Rossi”, 

Verona, Italy 

Background. Renal carcinoid tumors are rare neoplasms. Small 

series have been reported with emphasis on histopathology 

and clinical outcomes. Few studies regarded their cytogenetic 

features. Due to the high incidence of 11q13 riarrangements in

360 

neuroendocrine tumors arising in other human organs, we sought 

to assess 11q13 status in a serie of such a cases. 

Methods. Clinico-pathologic information on seven cases were 

recruited from in house and consultation cases at University of 

Verona. Follow-up was available on 5 patients ranging from 6 

months to 4 years. FISH analysis was performed with 11q13 

break-apart locus specific probes. 

Results. Patient age ranged from 35 to 74 years (average 56 y) 

(M:F; 5:2). All patients underwent radical nephrectomy. Lymph 

node dissection was performed in two cases. Tumors size ranged 

from 2.6 to 12 cm (average 5 cm). All cases presented a various 

pattern including tightly packed cords and trabeculae (80%) with 

minimal stroma, trabecular growth with prominent stroma, focal 

solid nests (15%), focal glandlike lumina (5%). Capsular invasion 

and infiltration of the renal sinus was seen in two cases, two 

with renal vein invasion and one with lymph-nodal metastases. 

Distant metastases was reported in two cases. One tumor showed 

an intraparenchymal metastasis. Calcifications were present in 3 

cases. Mitoses measured 0-12 (mean 4) per 10 HPF, necrosis was 

always absent. Immunostains were positive for synaptophysin 

(6/7), chromogranin (4/7), CK8-18 (7/7). CK7 was focally positive 

and CK20 stained in 1 case. TTF-1 and DOG-1 were negative 

in all cases. One patient died of disease at 8 months after surgery, 

two patients were alive and well and one died without disease. 

11q13 riarrangements was observed in 3/7 cases (one breaked 

signals, one loss of q arm, one gains of signals). 

We concluded that: 1) a subset of renal carcinoid tumours harbour 

11q13 riarrangements; 2) a subset of renal carcinoid tumors show 

an aggressive behavior. 

expression of aspartic protease NAPSIA-A among 

renal cell neoplasms 

Segala D., Gobbo S., Brunelli M., Pedron S., Chilosi M., Menestrina 

F., Martignoni G. 

Pathology, University Of Verona, Policlinico “G.B. Rossi”, Verona, Italy 

Background. Napsin-A is an aspartic protease reported to be 

expressed in the normal kidney and in type-II pneumocytes, 

recently recognized as a valid marker for the primitivity of 

pulmonary adenocarcinoma. Considering the importance of lysosomal 

aspartic proteases during renal cell carcinogenesis, we 

sought to evaluate the expression of napsin-A in a serie of renal 

cell neoplasms. 

Methods. We examined the immunoexpression of napsin-A 

in consecutive 133 renal cell epithelial neoplasms, including 

45 clear cell, 47 papillary, 20 chromophobe renal carcinomas 

(RCCs) and 23 renal oncocytomas. The immunohistochemical 

analysis was evaluated considering the intensity of the reaction 

scored in three grades and the percentage of immunolabeled 

neoplastic cells. Positive result was considered when more than 

10% of the neoplastic cells showed a cytoplasmic/membranous 

staining. We performed Western blot analysis to confirm the 

expression of this protein in the cases immunoreactive for 

napsin-A. Both immunohystochemical and western blot analyses 

were tested on samples of pulmonary adenocarcinoma as 

control. 

Results. Napsin-A immunoreactivity was detected in 6/ 45 (13%) 

clear cell RCCs, in 37/47 (79%) papillary RCCs, in 1/20 (5%) 

chromophobe RCCs and in 7/23 (34%) oncocytomas. The mean 

percentage of immunoreactive cells was respectively 27%, 47%, 

10% and 33%. The grade of intensity was overall higher in papillary 

RCCs than in the other histotypes. Western blot analysis 

confirmed the presence of napsin-A in clear cell and papillary 

RCCs, whereas the protein was not detected in oncocytoma. All 

the pulmonary adenocarcinomas tested as control showed both 

strong immunohystochemical reaction and positive western blot 

result. 


In conclusion, the aspartic protease napsin-A is variably immunoexpressed 

among renal cell neoplasms and, althought is not a 

specific marker, napsin-A is prevalently expressed in the papillary 

RCCs with higher intensity and percentage of reactive cells. 

Malignant lymphomas of the head and neck: 

review of a single-institute experience on 136 

cases 

1)G. Crisman, 2)G. Siniscalchi, 3)U. Falcone, 3)S. Guastafierro, 

2)G. Colella, 4)P. Leocata 

1)Dipartimento di Medicina Sperimentale, Ospedale Civile “San Salvatore”, 

Università degli Studi dell’Aquila, L’Aquila, Italia 2)Dipartimento 

di Patologia Testa e Collo, Seconda Università di Napoli, Napoli, Italia; 

3)Dipartimento di Ematologia, Medicina Trasfusionale e Immunologia, 

Seconda Università di Napoli, Napoli, Italy; 4)Dipartimento di Scienze 

della Salute, Università dell’Aquila, L’Aquila, Italia 

Background. Slowly enlarging masses of the head and neck area 

usually represent a great challenge for the clinicians, and several 

diagnostic tools could be used for diagnosis, from imaging to 

fine-needle aspiration biopsy. Malignant lymphomas represent 

the most common non-epithelial malignancy of the head and neck 

region, and they are classically divided by the World Health Organization 

(WHO 2008) into two subgroups, namely Hodgkin’s 

Lymphomas (HL) and non-Hodgkin’s Lymphomas. Usually, 

histological examination represents the main source for achieving 

the correct diagnosis. 

Methods. We retrospectively analyzed 136 cases from 136 patients 

with Hodgkin’s Lymphomas (HLs) and non-Hodgkin’s 

Lymphomas (NHLs) of the head and neck region and at least 5 

years of follow-up (or dead of disease). 

Results. Of 136 patients, 68 were males and 68 females, with an 

age range at the time of diagnosis of 5 to 79 years (mean, 44,2 

years). Cervical lymphadenopathy represents the most frequent 

clinical presentation (69 patients), followed by parotid masses 

(18 patients), and submandibular masses (16 patients). 98 NHL 

(72%) and 38 HL (28%) have been diagnosed. Among the NHL 

lymphomas, 89 ceses were from B-cell origin, whereas only 9 

T-cell lymphomas have been recognized. Follicular Lymphomas 

have been diagnosed in 31 cases, Large Cell B-Cell Lymphomas 

in 12 cases, mantle-cell lymohomas in 8 cases, CD30+ lymphomas 

in 4 cases, Burkitt lymphomas in 3 cases, and marginal-zone 

lymphoma only in one case. Among HL, the nodular-sclerosis 

variant is the most frequent, followed by the lymphocyte rich 

type. 

Because of their numerous histological variants, lymphoid neoplasms 

could represent a diagnostic challenge, and clinical and 

histopathological correlation and therapeutic approaches are 

presented and discussed. 

Positive urine cytology for renal collecting duct 

carcinoma: a case report 

Kapoula A., Skagias L., Politi E. 

Cytopathology Department, Aretaieio University Hospital, Athens, Greece 

Background. Collecting duct carcinoma (CDC), also known as 

Bellini duct carcinoma is a rare and aggressive renal neoplasm, 

arising from the epithelium of the distal segment of the collecting 

duct in the renal medulla pyramids. Since poor prognosis and 

early metastasis at the time of diagnosis are common, this tumor 

must be recognized and differentiated from conventional renal 

cell carcinoma and urothelial carcinoma. Case. Voided urine 

from a 76 year old male presented with gross hematuria, were 

examined. Computed tomography revealed a mass in the central 

part of the left kidney, along with a single nodule in the left lower 

pulmonary lobe. Cytological smears were very cellular and composed 

of numerous highly atypical, malignant cells, lying singly 

or in small, loosely cohesive groups. The tumor cells were round


to oval, spindle-shaped or tadpole-like and had large, irregular 

nuclei, single or multiple distinct nucleoli and cytoplasm that 

ranged from homogenous to vacuolated. Based on morphologic 

criteria a diagnosis of poorly differentiated carcinoma with sarcomatoid 

features was made. Subsequent immunocytochemical 

examination revealed positivity for high-molecular-weight cytokeratins 

(CK19, cytokeratin 34βE12), low-molecular-weight 

cytokeratin (Cam 5.2), Ulex europeaus agglutinin lectin (UEA- 

1) and vimentin. RCC antibody showed focal positive staining, 

whereas staining for CEA, cytokeratin 20, thrombomodulin, 

TTF-1 and CD10 was negative. The immunocytochemical findings, 

along with the morphology supported a diagnosis of renal 

collecting duct carcinoma, sarcomatoid variant. Conclusion. 

Because the few reports of CDCs in urine samples have been 

diagnosed either as renal cell carcinoma or as urothelial carcinomas, 

we conclude that it is mandatory to complement our morphologic 

study with immunocytochemical findings to achieve a 

more precise diagnosis. 

evaluation of accuracy of fine needle aspiration of 

the thyroid: a retrospective study on 245 cases 

1)Sollima L. 2)Crisman G. 3)Margiotta G. 4)Discepoli S. 5)Ciuffetelli 

V. 6)Saltarelli S. 

1)Dept of experimental medicine, University of l’aquila, L’Aquila, Italy 



4)Pathology unit, “ss. filippo e nicola” hospital, Avezzano, Italy 5)Pathology 

unit, “san salvatore” hospital, L’Aquila, Italy 6)Pathology unit, “san 

salvatore” hospital, L’Aquila, Italy 

Introduction. Due to its simplicity, low cost, and absence of 

major complications, FNA (fine needle aspiration) biopsy is the 

most accurate and sensitive diagnostic tool for the initial screening 

of patients with thyroid nodules, also if it has two major 

limitations: inadequate results and suspicious or indeterminate 

results. The aim of our study was to correlate FNAB with histological 

results to evaluate the sensitivity and specificity of this 

diagnostic procedure. 

Methods. a retrospective study on 245 patients affered to the 

Hospital of L’Aquila and Avezzano for FNAB from 2004 to 

2009 was perfomed. Subsequently, clinical and surgical data 

were correlated. The numbers of true-positive (TP), true-negative 

(TN), false-positive (FP) and false-negative (FN) results 

were calculated. These statistical values have been calculated: 

sensitivity in percentage: (TP/TP+FN) * 100; specificity in percentage: 

(1-(FP/FP+TN))(TN/TN+FP) * 100; positive predictive 

value (PPV) in percentage: (TP/TP+FP) * 100; negative predictive 

value (NPV) in percentage: (TN/TN+FN) * 100; diagnostic 

accuracy in percentage: (TP+TN/TP+TN+FP+FN) * 100. 

Results. According to the scientific literature, we obtained these 

data: sensitivity: 75%, specificity: 97,6%, PPV: 92,3%, NPV: 

91%, diagnostic accuracy: 91,3%. Most of the misdiagnosed 

cases were lesions diagnosed as THY3 for British Thyroid Association 

(BTA). 

Conclusions. FNAB of thyroid is highly accurate and has a low 

rate of false-negative and false-positive diagnoses. 

references 

Cáp J, Ryska A, Rehorková P, et al. Sensitivity and specificity of the fine 

needle aspiration biopsy of the thyroid: clinical point of view. Clin 

Endocrinol (Oxf) 1999;51(4):509-15. 

Amrikachi M, Ramzy I, Rubenfeld S, et al. Accuracy of Fine-Needle Aspiration 

of Thyroid. Arch Pathol Lab Med 2001;125:484-8. 

361 

expression of Il-32 in human lung cancer is related 

to the histotype and metastatic phenotype 

1)C. Sorrentino, 2)T. D’Antuono, 1)S. Di Meo, 1)P. Musiani, 1)E. 

Di Carlo 

1)Dipartimento di Oncologia e Medicina Sperimentale, Sezione di Anatomia 

Patologica, Università “G. d’Annunzio”, Via dei Vestini, 66100, 

Chieti, Italia; Centro Scienze dell’Invecchiamento (Ce.S.I.), Fondazione 

Università “G. d’Annunzio”, Via Colle dell’Ara, 66100, Chieti, Italia; 


Patologica, Università “G. d’Annunzio”, Via dei Vestini, 66100, 


Background. A strong link has been recently demonstrated 

between inflammation and lung cancer. Thus, we investigated 

whether the new proinflammatory cytokine IL-32 may be involved 

in the physiopathology of the main lung cancer histotypes 

and hence provide a novel therapeutic target. 

Methods. IL-32 expression, as visualized by immunohistochemistry 

on 23 premalignant and 148 malignant lesions, was 

correlated with clinico-pathological and survival data. Confocal 

microscopy, microdissection and real-time reverse transcription- 

PCR were used to identify cell sources and expression levels of 

IL-32. 

Results. IL-32 immunoreactivity was absent in normal lung tissue, 

while it could be found in lung cancers in two distinct sites: 

tumor cells (TC), and tumor infiltrating lymphocytes (TIL). 

IL-32 was strongly expressed by TC in the vast majority of 

Small Cell Lung Cancers (SCLC) (77%) and, among Non Small 

Cell Lung Cancers (NSCLC), by Large Cell Carcinomas (LCC) 

(64%), Adenocarcinomas (AC) (73%) and its precursor lesion. 

Conversely, IL-32 was absent from most (76%) Squamous Cell 

Carcinomas (SCC) and their precursors. 

Lymph node metastases frequently developed from IL-32-expressing 

lung cancers, and especially (82%) from those endowed 

with IL-32-expressing TIL mainly composed of CD68 + macrophages, 

CD4 + T lymphocytes, and DC-SIGN + dendritic cells. 

Expression levels of IL-32 by both TIL and TC in the primary 

tumor, particularly in AC and SCC, were paralleled by those of 

IL-6, IL-8 and VEGF, in the same cell population, and correlated 

with high intratumor microvessel density and poor clinical outcome 

as revealed by Kaplan-Meyer survival curves. 

Conclusions. Our findings suggest that both TC- and TIL-derived 

IL-32 are deeply involved in lung carcinogenesis by favoring invasion 

and metastasis. Indeed, TC expression of IL-32 could be 

a useful and easy detectable prognostic biomarker, especially for 

AC and SCC histotypes. 

Neo-adjuvant hormone therapy boosts intraprostatic 

infiltration of both cytotoxic-effector 

and regulatory T lymphocytes in prostate cancer 

patients 

1)C. Sorrentino, 1)S. Di Meo, 2)T. D’Antuono, 1)P. Musiani, 

1)E. Di Carlo 


Patologica, Università “G. d’Annunzio”, Chieti, Italia; Centro 

Scienze dell’Invecchiamento (Ce.S.I.), Fondazione Università “G. d’Annunzio”, 

Chieti, Italia; 2)Dipartimento di Oncologia e Medicina Sperimentale, 

Sezione di Anatomia Patologica, Università “G. d’Annunzio”, 


Background. The value of neoadjuvant hormone therapy (NHT) 

prior to radical prostatectomy (RP) as a means of restraining 

prostate cancer (PCa) and strengthening its immunotherapy is still 

uncertain. Here, we ask whether it subverts immunoregulatory 

pathways governing tumor microenvironment. 

Methods. We microdissected epithelium and stroma from cancerous 

and normal prostate specimens from 118 prostatectomized 

patients of whom 64 had received NHT, to detect immunoregu-

362 

latory cytokine/chemokine gene expression levels by real-time 

reverse transcription-PCR. Confocal microscopy was used to 

assess cytokine/chemokine cell sources, and double or triple immunostainings 

to characterize immune cell infiltrates. 

Results. NHT boosted the expression of IL-7 in stromal fibroblasts 

and smooth muscle cells and that of IFNγ-inducible 

protein (IP-10)/CXCL10 in the glandular epithelium of normal 

prostate tissue close to neoplastic foci, and massively increased 

the CD8 + and CD4 + T lymphocyte infiltrate. Lymphocytes mostly 

expressed T-cell intracellular antigen-1 (TIA-1), granzyme-B and 

perforin, which are typical of cytotoxic-effector T (Teff) cells. 

NHT also induced thymus and activation-regulated chemokine 

(TARC)/CCL17 production by monocytes/macrophages in the 

prostate and draining lymph nodes, and increased the number 

of their forkhead box P3 (foxp3) + CD25 + CD127 - T regulatory 

(Treg) cells. 

Kaplan-Meyer curves revealed a significant improvement in the 

biochemical disease-free survival of the NHT-treated patients 

with an high Teff / Treg cell ratio in their prostate cancers. 

Conclusions. Androgen withdrawal displays immunological 

effects by regulating different cytokine/chemokine gene expression 

in normal prostate and lymphoid tissues, and this probably 

favors intra-prostatic infiltration of both Treg- and especially 

Teff- lymphocytes. High levels of PCa infiltrating cytotoxic T 

lymphocytes predicts a better clinical outcome in NHT treated 

patients. 

Iatrogenic Kaposi’s sarcoma of the stomach 

in a HIV-seronegative pemphigus patient. 

Clinico-pathologic study of a case 

1) L. Sparano, 1)P. Riccio, 2)G. Magro, 3)M. Vairo, 3)M. Bisceglia 

1)Department of Pathology, “M. Scarlato” Hospital, Scafati, Italy; 2)Department 

of Pathology, University School of Medicine, Catania, Italy; 


Giovanni Rotondo, Italy 


histogenesis and viral etiology (gammaherpesvirus HHV- 

8-driven), occurring primarily in the skin 1 . A variety of clinical 

forms have been identified: sporadic, endemic, iatrogenic, and 

epidemic 1-3 . The iatrogenic form develops in immunocompromised 

hosts, i.e. transplant patients who are immunosuppressed, 

patients receiving immunosuppressive therapies for hematological 

malignancies (mainly chronic lymphocytic leukemia, and 

non-Hodgkin’s and Hodgkin’s lymphomas), solid tumors (mainly 

carcinomas of the breast, followed by lung, colon, larynx, liver, 

pancreas, and kidney, in decreasing order of frequency), or inflammatory 

and autoimmune diseases after long-term steroid 

treatment 1-3 . Non-neoplastic medical conditions treated with 

immunosuppressive drugs (mainly corticosteroids), which are on 

record as associated with KS (though rarely) are: inflammatory 

chronic intestinal diseases (ulcerative colitis, and Crohn’s disease), 

rheumatoid arthritis, giant cell arteritis (Horton arteritis), 

relapsing polychondritis, systemic lupus erythematosus, Behçet’s 

disease, and pemphigus. 

Objectives. 1. To report on a case of iatrogenic KS involving the 

stomach in an elderly female patient receiving immunosuppressive 

therapy for pemphigus. 2. To review the world literature with 

regards to iatrogenic KS complicating (autoimmune) pemphigus 

recorded between January 1960 and May 2010. 

Case report. A 78-year old lady underwent emergency gastrectomy 

for massive upper gastrointestinal hemorrhage. Histological 

examination of the surgical specimen revealed widespread 

involvement of the stomach by a diffuse intramural 

hemorrhagic tumor. Following the histological diagnosis of a 

vascular Kaposiform malignant neoplasm (immunopositive for 

CD31 and CD34, and negative for S100 protein, alpha-SMA 


and CD117), with “metastases” in 5 of 14 perigastric lymph 

nodes, the clinical history of medical immunosuppressive treatment 

with corticosteroids and azathioprine for 5 years due to 

autoimmune pemphigus came to light. Physical examination 

of the skin did not disclose cutaneous signs consistent with KS 

manifestations. Additional immunohistochemical testing with 

antibody to LNA-1 HHV-8 revealed strong and diffuse nuclear 

positivity in most of the “neoplastic” spindle cells. The final 

diagnosis was that of KS involving the stomach and regional 

lymph nodes. 

Discussion. Patients who receive immunosuppressive therapy 

are at increased risk for KS (immunosuppression-associated 

KS), the majority of whom are renal transplant patients. In 

addition to the role of immunologic impairment, other etiologic 

factors also play a role in this form of KS, and one of 

the recognized risk factors for this type of KS is ethnicity 

(Eastern European and Mediterranean people as well as Jews 

of European and North African origin are at higher risk). The 

immunosuppression-associated form of KS occurs between a 

few months and a few years after starting therapy. Clinical and 

experimental evidence have demonstrated the causative role 

of the human herpesvirus type 8 (HHV-8), a new member of 

the gamma-herpesvirus family, in the etiology of KS. HHV- 

8 has been detected in all epidemiological forms of KS, and 

has also been implicated in the pathogenesis of other diseases, 

mostly primary effusion lymphomas (“body cavity based-lymphomas”), 

multicentric Castleman’s disease, and lymph node 

based plasmablastic lymphoma. Other, but more controversial, 

conditions in which HHV-8 has a pathogenetic role are multiple 

myeloma, Waldenström’s macroglobulinemia, and pemphigus. 

About 10 cases of KS in the setting of autoimmune pemphigus 

(vulgaris, foliaceus) have been reported so far 4-8 . Pemphigus 

lesions have been found to contain HHV-8 DNA sequences, 

and this suggests that HHV-8 might have a role in their development. 

However, iatrogenic KS in the setting of pemphigus 

vulgaris has been infrequently observed, and it is more likely 

that KS associated with pemphigus is related (in certain predisposed 

individuals) to therapeutically induced immunosuppression, 

rather than primarily to pemphigus itself. All cases 

of KS associated with pemphigus which have been reported to 

date are cutaneous KS. The case presented herein, to the best 

of our knowledge and based on a computerized bibliography 

search, is the first case of gastric KS associated with pemphigus. 

The stomach may be frequently involved in all forms of KS 

(AIDS-associated, immunosuppressed transplant-associated, 

iatrogenically immunosuppressed non-transplant associated, 

and also in the classical form), and after the skin is the most 

frequent anatomic site of involvement by KS. Whatever the 

clinical and epidemiological setting, immunosuppression-associated 

KS involving the stomach may also affect the skin, 

but can be limited to the gastrointestinal tract. In our case KS 

was limited to the stomach. The differential diagnosis of KS of 

the stomach includes other types of spindle cell sarcomas such 

as angiosarcoma, GIST, leiomyosarcoma, and inflammatory 

myofibroblastic tumor: immunohistochemical testing with anti- 

HHV-8 antibody is critical, given its high sensitivity (almost 

100%) and specificity (100%) 9 . 

Conclusion. HHV-8 is found in the cutaneous lesions of pemphigus. 

Pemphigus is an autoimmune disease which is treated 

with immunosuppressive drugs. KS may arise in the setting of 

pemphigus, and this is more likely attributable to the impairment 

of cellular immunity rather than to the pemphigus itself. 

references 



2002;47:641-55. 


clinico-pathologic overview. Tumori 1991;77:291-310.


3 Ablashi DV, Chatlynne LG, Whitman JE Jr, et al. Spectrum of 

Kaposi’s sarcoma-associated herpesvirus, or human herpesvirus 8, 

diseases. Clin Microbiol Rev 2002;15:439-64. 

4 Amblard P, Reymond JL, Beani JC, et al. Pemphigus vulgaris and Kaposi 

sarcoma. Report of a new case (author’s transl). Dermatologica. 

1981;163:58-62. 

5 Kaplan RP, Israel SR, Ahmed AR. Pemphigus and Kaposi’s sarcoma. 

J Dermatol Surg Oncol 1989;15:1116-21. 

6 Avalos-Peralta P, Herrera A, Ríos-Martín JJ, et al. Localized Kaposi’s 

sarcoma in a patient with pemphigus vulgaris. J Eur Acad Dermatol 

Venereol 2006;20:79-83. 

7 Saggar S, Zeichner JA, Brown TT, et al. Kaposi’s sarcoma resolves 

after sirolimus therapy in a patient with pemphigus vulgaris. Arch 

Dermatol 2008;144:654-7. 

8 Serdaroglu S, Antonov M, Demirkesen C, et al. Iatrogenic Kaposi’s 

sarcoma in a patient with pemphigus vulgaris. Clin Exp Dermatol 

2009;34:839-40. 




Increasing trend of thyroid cancer: a populationbased 

study in the south of Switzerland 

1)Spitale A. 2)Crippa S. 3)Mazzola P. 4)Frattini M. 5)Mazzucchelli 

L. 6)Bordoni A. 

1)Registro Tumori Canton Ticino, Istituto Cantonale di Patologia, Locarno, 

Svizzera 2)Patologia Clinica, Istituto Cantonale di Patologia, Locarno, 

Svizzera 3)Registro Tumori Canton Ticino, Istituto Cantonale di 

Patologia, Locarno, Svizzera 4)Laboratorio di Diagnostica Molecolare, 

Istituto Cantonale di Patologia, Locarno, Svizzera 5)Patologia Clinica, 

Istituto Cantonale di Patologia, Locarno, Svizzera 6)Registro Tumori 

Canton Ticino, Istituto Cantonale di Patologia, Locarno, Svizzera 

Background. The incidence of thyroid cancer, particularly papillary 

carcinoma, has been increasing during the past decades. Recent 

studies in the US showed an increased trend, predominantly 

in women. The objective of the current study was to investigate 

incidence trends of thyroid cancer according to sex, age, histological 

type and tumour size. 

Methods. All thyroid cancers, occurred between 1996 and 2007, 

were selected from the files of Ticino Cancer Registry. World 

age-standardised incidence rates were calculated. Trends analysis 

was performed through the joinpoint regression, stratifying 

cases by sex, age, histological type and tumour size (< 10 mm, 

11-20 mm, 21-40 mm, > 40 mm). Annual Percentage Changes 

(APC) were reported. Cancer specific survival was performed by 

the Kaplan-Meier method and the Log-rank test was invoked to 

detect significant differences. 

Results. A total of 238 thyroid cancers were selected, 53 and 185 

in males and females respectively. Incidence rates increased in 

men (APC = 4.4; 95%CI: -22.2;40.1) and markedly in women 

(APC = 7.0; 95%CI: 1.4;13.0), particularly in the age group 

20-49 (APC = 13.1;95%CI: 5.1;21.6). Significant increase was 

observed in papillary carcinoma smaller than 10mm (APC = 8.9; 

95%CI: 0.3;18.3). Concerning cancer specific survival,a worst 

prognosis was observed in papillary tumours wider than 40mm 

(p = 0.002). 

Discussion. The widespread and aggressive use of ultrasound 

and image-guided fine needle aspiration, and changes in tumour 

classification seem to be recognised factors of the increase of 

thyroid cancer incidence. However, in our study we observed 

strong increase predominantly in women, and across all tumour 

size groups, suggesting that other factors (e.g. environmental 

agents and molecular pathways) need to be explored. Additional 

European population-based data are needed in order to confirm 

this results. 

Malignant carcinoid of the extrahepatic biliary 

tract: report of two cases 

363 

1)Squillaci S. 2)Marchione R. 3)Piccolomini M. 4)Colombo F. 

5)Bucci F. 6)Bruno M. 7)Bisceglia M. 

1)Division of Anatomic Pathology, Hospital of Vallecamonica, Esine, Italy 



4)Division of General Surgery, Hospital of Vallecamonica, Esine, Italy 

5)Division of General Surgery, Madonna Delle Grazie Hospital, Matera, 

Italy 6)Division of Anatomic Pathology, Madonna Delle Grazie Hospital, 

Matera, Italy 7)Department of Pathology, IRCCS “Casa Sollievo Della 

Sofferenza”, San Giovanni Rotondo, Italy 

Introduction. Extrahepatic bile duct (EHBD) carcinoid tumors 

are extremely uncommon. The latest classification edited by 

WHO in 2000 divides neuroendocrine neoplastic epithelial lesions 

of the EHBDs into four types: well-differentiated endocrine 

tumor (carcinoid tumor), well-differentiated endocrine carcinoma 

(malignant carcinoid), poorly differentiated endocrine carcinoma 

(small cell carcinoma) and mixed endocrine-exocrine carcinoma 

1 . To date only 70 cases of carcinoids have been reported and 

almost all publications contain only one such case with often 

insufficient follow-up data 2 . 

Objectives. To report the clinicopathologic study of two personal 

cases. 

Methods. Case 1. A 52-year-old man presented with clinical 

jaundice of 2 weeks duration. At imaging, there was an irregular 

bulging mass in the EHBD wall by MRI and CT. ERCP showed 

a stricture 2 cm in diameter in the distal third of the common 

hepatic duct. Cholecystectomy and extended transection of the 

involved tract of EHBD with portal lymphadenectomy was 

performed. The patient is alive with no evidence of disease 41 

months after surgery. Case 2. A 70-year-old man was investigated 

for acute recurrence of sharp abdominal pain, which was 

firstly ascribed to gallbladder lithiasis. No sign of jaundice was 

noted, but slightly dilated intrahepatic bile ducts were seen. The 

patient was submitted to cholecystectomy and during the procedure 

a mass was noted in the common hepatic duct. Resection of 

the involved tract of EHBD associated with left hepatectomy and 

extended portal lymphadenectomy was performed. The patient is 

alive with no evidence of disease 59 months after surgery. 

Results. The main tumor diameter was 2 and 4.5 cm in case 1 

and case 2, respectively. Histologically in both cases the tumor 

was comprised of medium-sized polygonal cells with nesting, 

palisades and adenoid-like trabecular growth patterns, with 

minimal pleomorphism and sparse mitoses. The tumor was 

infiltrative and extended into adjacent periductal soft tissue, 

showing vascular and perineural permeation. No metastases 

were found in the three excised portal lymph nodes in case 1. 

Metastases with the same cytological and architectural features 

were present in a hilar lymph node and in the subcapsular liver 

tissue of the quadrate lobe. In both cases the neoplastic cells 

were strongly immunoreactive for keratins and pan-endocrine 

markers such as chromogranin and synaptophysin, whilst negative 

for TTF-1, gastrin, insulin, glucagon, VIP, and prolactin. 

Somatostatin and pancreatic polypeptide were negative in case 

1, and focally positive in case 2. Proliferation index was low 

(< 2%) in both. 

Discussion. EHBD carcinoids are lesions with a well-recognized 

capacity for local recurrence and metastasis (malignant carcinoids), 

but, in contrast to other sites, tend to have indolent biological 

behavior, even when metastatic. The differential diagnosis must 

essentially be made vs other types of neuroendocrine neoplasms 

as poorly differentiated adenocarcinomas with numerous endocrine 

cells and small cell carcinomas, which are clinically more 

aggressive. Pathologists must be aware of the possible occurrence 

of EHBD carcinoids. Complete surgical resection is often curative 

depending on the location and the degree of tumor extension.

364 

references 

1 Capella C, Solcia E, Sobin LH, et al. Endocrine Tumors of the Gallbladder 

and extrahepatic bile ducts. In: Hamilton SR, Aaltonen LA 

(eds). Pathology and Genetics-Tumours of the Digestive System. Lyon: 

IARC Press 2000. 

2 Squillaci S., Marchione R., Piccolomini M., et al. Well differentiated 

neuroendocrine carcinoma (malignant carcinoid) of the extrahepatic 

biliary tract. Report of two cases and literature review. APMIS, 2010, 

in press. 

Cytologic diagnosis of primary effusion lymphoma 

in an elderly patient 

1)Squillaci S. 2)Marchione R. 3)Spairani C.* 4)Piccolomini M. 

5)Tondini M.° 6)Cirelli R. 7)Tallarigo F.** 

1)Division of anatomic pathology, Hospital of vallecamonica, Esine, Italy 

2)Division of anatomic pathology, Hospital of vallecamonica, Esine, Italy 

3)*Division of anatomic pathology, Hospital , Novi ligure, 

Italy 4)Division of anatomic pathology, Hospital of vallecamonica, 

Esine, Italy 5)°Division of pneumology, Hospital of vallecamonica, Esine, 

Italy 6)Division of anatomic pathology, Hospital of vallecamonica, Esine, 

Italy 7)**Division of anatomic pathology, S.giovanni di dio hospital, 

Crotone, Italy 

Background. Primary effusion lymphoma (PEL) is a subgroup 

of non-Hodgkin’s lymphoma predominantly described in HIVinfected 

individuals in the absence of a clinically identifiable 

contiguous tumor mass. It typically arises as pleural, peritoneal 

or pericardial effusion usually involving only one body site. PEL 

rarely develops in HIV-negative immunocompetent populations. 

Methods. A 89-year old man under treatment for severe aortic 

stenosis, hypertension, diabetes mellitus, dyslipidosis and paroxysmal 

atrial fibrillation was admitted to Hospital of Vallecamonica 

for dyspnea, nonproductive cough and thoracic compliant. Past 

medical history also included a hematologic deficiency of factor 

VIII. Clinical examination revealed left pleural effusion associated 

with diffuse interstitial pulmonary fibrosis. The patient did 

not have hepatitis B or C but HIV status was not available. 

Results. Examination of the pleural fluid specimen showed a 

hypercellular smear composed of large atypical cells. The nuclei 

with irregular outlines and one or more prominent nucleoli, were 

partly surrounded by abundant cytoplasm with peripheral digitations. 

These cells proved to be strongly immunoreactive to CD45 

and HHSV-8, whilst negative for calretinin and claudin-4. 

Conclusion. The morphological and immunophenotyping of the 

tumor cells are those of PEL with some of the appearances bridging 

immunoblastic and anaplastic large-cell lymphomas. The possibility 

of an epithelial metastatic effusion and a mesothelioma 

must always be excluded. This variant of lymphoma is extremely 

rare and our patient was referred to Haematology department 

for a more comprehensive investigation but died for disease 

two months later. Recently very few cases of HHV8-unrelated 

PEL-like lymphoma have been described. We believe the term 

PEL should be restricted to those HHV-8 positive cases only, as 

originally proposed in the 2001 WHO classification. 

references 

1) Carbone A, Gloghini A. PEL and HHV8-unrelated effusion lymphomas: 

Classification and diagnosis. Cancer. 2008;114:225-7. 

The proliferation marker Chromatin Assembly 

factor-1 is of clinical value in predicting the 

biological behaviour of cellular leiomyomas? 

1)Staibano S. 2)Nugnes L. 3)Mascolo M. 4)Vecchione ML. 

5)Ilardi G. 6)Siano M. 7)De rosa G. 

1)Dpt of Biomorphological and Functional Sciences-Pathology Section, 

University “Federico II”, Naples, Italy 2)Dpt of Biomorphological and 

Functional Sciences-Pathology Section, University “Federico II”, Naples, 

Italy 3)Dpt of Biomorphological and Functional Sciences-Pathology Section, 

University “Federico II”, Naples, Italy 4)Dpt of Biomorphological 


and Functional Sciences-Pathology Section, University “Federico II”, 

Naples, Italy 5)Dpt of Biomorphological and Functional Sciences-Pathology 

Section, University “Federico II”, Naples, Italy 6)Dpt of Biomorphological 

and Functional Sciences-Pathology Section, University 

“Federico II”, Naples, Italy 7)Dpt of Biomorphological and Functional 

Sciences-Pathology Section, University “Federico II”, Naples, Italy; 

Oncology Research Center of Basilicata (C.R.O.B.), Rionero in Vulture, 

Potenza, Italy 

Background. Leiomyomas (LM) are the most frequent benign 

tumour of the female genital tract. They represent the most common 

indication for hysterectomy or myomectomy during the 

reproductive age. The “cellular” cases are defined as LM showing 

an increased number of smooth muscle cells per unit area compared 

with the surrounding myometrium and/or with the ordinary 

histotype of LM and may constitute a challenging clinical and 

histological diagnostic problem. Recently, a new proliferation 

and prognostic marker, the Chromatin Assembly Factor–1 (CAF- 

1), a trimeric protein histone chaperone, has been found overexpressed 

in a series of human malignancies, in close association 

with their biological aggressiveness. This work focused on the 

role of CAF-1/p60 protein as a marker of clinical value for cellular 

leiomyomas (CLM). 

Methods. The expression of CAF-1/p60 was evaluated by immunohistochemistry 

on a selected series of CLM, with or without 

a history of treatment with GnRH. The resulting data were 

compared with traditional prognostic parameters, including the 

expression of the routine proliferation marker ki67/MIB1. 

Results. Comparing the tumours of the treated and the untreated 

group, we found a similar expression of CAF-1/p60, while the 

immunoexpression of ki67 resulted significantly higher in the 

untreated group. This allows us to speculate on the biological 

significance of these benign lesions. Our results suggest that these 

CLM show a high proliferation rate, but their low expression 

of CAF-1/p60 places them in the lower end of the spectrum of 

uterine lesions ranging from typical leiomyomas to leiomyosarcomas. 

These data support the idea that CAF-1/p60 may have a 

possible diagnostic and prognostic role for uterine smooth muscle 

tumors. 

relationship between histological features and 

epigenetic regulation of endothelial proliferation 

in a selected series of skin malignant melanoma 

1)Staibano S. 2)Siano M. 3)Mascolo M. 4)Ilardi G. 5)Nugnes L. 

6)Vecchione ML. 7)De rosa G. 

1)Dpt of Biomorphological and Functional Sciences-Pathology Section, 

University “Federico II”, Naples, Italy 2)Dpt of Biomorphological and 

Functional Sciences-Pathology Section, University “Federico II”, Naples, 

Italy 3)Dpt of Biomorphological and Functional Sciences-Pathology Section, 

University “Federico II”, Naples, Italy 4)Dpt of Biomorphological 

and Functional Sciences-Pathology Section, University “Federico II”, 

Naples, Italy 5)Dpt of Biomorphological and Functional Sciences-Pathology 

Section, University “Federico II”, Naples, Italy 6)Dpt of Biomorphological 

and Functional Sciences-Pathology Section, University “Federico 

II”, Naples, Italy 7)Dpt of Biomorphological and Functional Sciences- 

Pathology Section, University “Federico II”, Naples, Italy; Oncology Research 

Center of Basilicata (C.R.O.B.), Rionero in Vulture, Potenza, Italy 

Background Cutaneous melanoma (CM) is the most lethal form 

of skin malignancy, showing a progressive increase in incidence 

rate worldwide. 

Although recent improvements in diagnostic techniques allowed 

the diagnosis of most CM at an early stage, the number of patients 

diyng for CM remains still substantially unchanged. Neo-angiogenesis 

and tumor angioinvasivity are strictly related to the aggressiveness 

of malignant tumors, and a high microvessel density 

(MVD) has been correlated with an adverse clinical behavior of 

deeply invasive CM. We evaluated these parameters in CM, correlating 

MVD of each tumor with the proliferating status of microvessels, 

using the routine proliferation marker ki67/MIB1 and


the Chromatin Assembly Factor-1 (CAF-1)/p60 protein. CAF-1 is 

a trimeric protein complex fundamental for the epigenetic regulation 

of both cell proliferation and DNA repair. The p60 subunit 

of CAF-1 is frequently overexpressed in malignant tumors. Up to 

now, data concerning the relationship between CAF-1/p60 and 

ki67 expression, MVD and outcome of CM are not available. 

Methods. We evaluated by immunohistochemistry the micromacrovessels 

density, the ki67 index and the CAF-1/p60 expression 

in a selected series of CM The results were compared 

with the clinical and pathological features of each case and with 

patient’s outcome. 

Results. We found a variable extent of MVD and an heterogeneous 

expression of ki67 and CAF-1/p60 among our cases of 

invasive CM. The statistical analysis showed that biological aggressiveness 

of CM, besides Breslow thickness, was related to the 

deregulated proliferation rate of endothelial cells, and positively 

correlated with the overexpression of CAF-1/p60. On the contrary, 

we failed to show any significant correlation between MVD 

and clinical outcome of CM. Our results strongly suggest that a 

deregulated growth rate of endothelial cells, rather than MVD, 

characterized aggressive CM. 

role of MuTYH and MSH2 in the control of oxidative 

DNA damage, genetic instability, and tumorigenesis 

1)Stramucci (L). 2)Sabatini (F). 3)Hysi (A). 4)Liberatore (M). 

5)Molatore (S). 6)Barone (F). 7)Mazzei (F). 8)Bignami (M). 

9)Pannellini (T). 10)Musiani (P). 

1)Anatomia Patologica/Oncologia e Neuroscienze, Ss. Annunziata/CESI, 

Chieti, Italia 2)Anatomia Patologica/Oncologia e Neuroscienze, Ss. Annunziata/CESI, 

Chieti, Italia 3)Anatomia Patologica/Oncologia e Neuroscienze, 

Ss. Annunziata/CESI, Chieti, Italia 4)Anatomia Patologica/Oncologia 

e Neuroscienze, Ss. Annunziata/CESI, Chieti, Italia 5)Genetica 

e Microbiologia, Università di Pavia, Pavia, Italia 6)Environment and 

Primary Prevention, Iss, Roma, Italia 7)Environment and Primary Prevention, 

Iss, Roma, 8)Environment and Primary Prevention, Iss, Roma, 


Chieti, Italia 10)Anatomia Patologica/Oncologia e Neuroscienze, Ss. Annunziata/CESI, 


Background. Mismatch repair is the major pathway controlling 

genetic stability by removing mispairs caused by faulty replication 

and/or mismatches containing oxidized bases. Thus, inactivation 

of the Msh2 mismatch repair gene is associated with a 

mutator phenotype and increased cancer susceptibility. The base 

excision repair gene Mutyh is also involved in the maintenance 

of genomic integrity by repairing premutagenic lesions induced 

by oxidative DNA damage. 

Methods. Because evidence in bacteria suggested that Msh2 and 

Mutyh repair factors might have some overlapping functions, 

we investigated the biological consequences of their single and 

double inactivation in vitro and in vivo. 

Results. Msh2(-/-) mouse embryo fibroblasts (MEF) showed a 

strong mutator phenotype at the hprt gene, whereas Mutyh inactivation 

was associated with a milder phenotype (2.9 × 10(-6) and 

3.3 × 10(-7) mutation/cell/generation, respectively). The value 

of 2.7 × 10(-6) mutation/cell/generation in Msh2(-/-)Mutyh(-/-) 

MEFs did not differ significantly from Msh2(-/-) cells. When 

steady-state levels of DNA 8-oxo-7,8-dihydroguanine (8-oxoG) 

were measured in MEFs of different genotypes, single gene inactivation 

resulted in increases similar to those observed in doubly 

defective cells. In contrast, a synergistic accumulation of 8-oxoG 

was observed in several organs of Msh2(-/-)Mutyh(-/-) animals, 

suggesting that in vivo Msh2 and Mutyh provide separate repair 

functions and contribute independently to the control of oxidative 

DNA damage. Finally, a strong delay in lymphomagenesis was 

observed in Msh2(-/-)Mutyh(-/-) when compared with Msh2(-/- 

) animals. The immunophenotype of these tumors indicate that 

both genotypes develop B-cell lymphoblastic lymphomas displaying 

microsatellite instability. This suggests that a large frac- 

365 

tion of the cancer-prone phenotype of Msh2(-/-) mice depends on 

Mutyh activity. 

Jejunal T cell lymphoma associated with colonic 

intraepithelial lymphocytes with aberrant 

phenotype 

1)Tabanelli V. 2)Valli R. 3)Sabattini E. 4)Pileri SA. 

1)U.O. Emolinfopatologia, Dipartimento di Ematologia e Scienze Oncologiche 

“Lorenzo e Ariosto Seràgnoli”, Università di Bologna, Policlinico 

S.Orsola-Malpighi, Bologna, Italy 2)Dipartimento di Anatomia patologica, 

Arcispedale S. Maria nuova, Reggio Emilia, Italy 3)U.O. Emolinfopatologia, 

Dipartimento di Ematologia e Scienze Oncologiche “Lorenzo e 

Ariosto Seràgnoli”, Università di Bologna, Policlinico S.Orsola-Malpighi, 

Bologna, Italy 4) U.O. Emolinfopatologia, Dipartimento di Ematologia 

e Scienze Oncologiche “Lorenzo e Ariosto Seràgnoli”, Università di 

Bologna, Policlinico S.Orsola-Malpighi, Bologna, Italy 

Background. We present the case of a jejunal NK-like T-cell 

lymphoma associated with multifocal increase of intraepithelial 

lymphocytes (IELs) in the large bowel in a 48-year-old woman 

without evidence of coeliac disease (CD), as demonstrated by 

clinical history, endoscopy, serology and HLA genotypic analysis. 

Methods. The analysis were performed on formalin-fixed paraffin-embedded 

tissue; histological sections were stained for H&E 

and Giemsa; immunohistochemistry (IHC) was performed partly 

with a Lab Vision Autostainer 720 and partly with APAAP technique. 

In situ hybridization for EBV integration was performed 

(PNA detection kit, DAKO). TCR gene rearrangement was performed 

according to Biomed protocol. 

Results. Histologically, the jejunal wall was infiltrated by a 

proliferation of medium to large monomorphic lymphocytes with 

elevated mitotic ratio. The colonic wall reached after perforation 

was infiltrated ab extrinseco up to the muscolaris propria, while 

the spare colonic mucosa showed a mild bland lymphocytic infiltrate 

in the lamina propria and aggregates of atypical IELs. The 

uninvolved jejunal mucosa did not show either villous atrophy or 

increase of IELs, excluding a coexistent CD. 

At IHC, the neoplastic cells showed expression of CD3, CD7, 

CD56, CD8, TIA-1 and perforin, being negative for CD2, 

CD5, CD4, Granzyme B, CD20 and CD30. The Ki67 value 

was about 60%. Molecular analysis revealed monoclonal rearrangement 

of the TCR genes and negativity for EBER. 

The colonic IELs shared the same immunophenotype as the 

tumoural cells but the negativity for CD8 and a lower Ki67 

value, while the mild lymphocytosis in the colonic lamina 

propria showed regular expression of the T cell markers. PCR 

analysis performed in the dissected IELs presented the same 

monoclonal TCR rearrangement observed in the jejunal sample. 

Conclusions. To our knowledge, this is the first case of a small 

bowel NK like T-cell lymphoma associated with neoplastic colonic 

IELs in a patient without CD. 

Prostatic cancer metastatic to the stomach: a case 

report 

1)Tacchini D. 2)Vassallo L. 3) Peccetti A. 

1)Patologia Umana Ed Oncologia, S. Maria alle Scotte, Siena, Italia 

2)Patologia Umana Ed Oncologia, S.Maria alle Scotte, Siena, Italia 3) 

Sezione di Endoscopia Digestiva, Azienda U.S.L. 7, Ospedale di Nottola, 

Siena, Italia 

Background. Metastatic cancer of the prostate has a poor prognosis 

with survival rates ranging from 1 to 3 years. Common sites 

of metastases are bone, lung and lymphnodes; gastrointestinal 

tract is a very rare site whose occurrence can cause problems in 

diagnosis. 

Objective.We report a case of carcinoma of the prostate metastatic 

to the stomach in a 64 year old man with anemia, melena 

and bone pain.

366 

Methods. Gastric biopsy routinely processed and stained (HE). 

Morphological diagnosis confirmed by immunohistochemistry 

(PSA, proliferative activity evaluated with Ki67). 

Results. We report a case of 64 year old man with upper, nonspecific, 

gastrointestinal tract symptoms, accompained by abdominal 

and bone pain of moderate intensity. His medical record reports 

prostatic cancer diagnosed three years before, with appearance 

of lung and bone metastases one year later. At endoscopical 

examination, “volcano-like”, small ulcers, mostly in the lesser 

curvature and angular region of the stomach, were noted. Microscopical 

examination showed clusters of pleomorphic cells of 

medium- large size, with eosinophilic cytoplasm, large nuclei and 

prominent nucleoli arranged in solid “glandular” clusters, quite 

different from gastric adenocarcinoma. Immunohistochemistry 

for PSA confirmed prostatic origin of the lesion. 

Conclusion. Gastrointestinal metastases indicates advanced disease 

with poor prognosis. A correct diagnosis is important to 

direct therapy. 

A multicenter external quality control (eQC) study 

in the lazio region for the immunohistochemical 

determination of Her2 in breast cancer: the 

importance to define a strict protocol 

1)Terrenato I. 2)Perracchio L. 3)Costarelli L. 4)Bonanno E. 

5)Arena V. 6)Pizzamiglio S. 7)Muti P. 8)Paradiso A. 9)Verderio 

P. 10)Mottolese M. 

1)Epidemiologia, Istituto Nazionale Tumori Regina Elena, Roma, Italia 

2)Anatomia Patologica, Istituto Nazionale Tumori Regina Elena, Roma, 

Italia 3)Anatomia Patologica, Azienda Ospedaliera S.Giovanni Addolorata, 

Roma, Italia 4)Anatomia Patologica 1, Università Di Tor Vergata, 

Roma, Italia 5)Anatomia Patologica, Università’ Cattolica Del Sacro 

Cuore, Roma, Italia 6)Statistica Medica Biometria E Bioinformatica, 

Fondazione Irccs Istituto Nazionale Tumori, Milano, Italia 7)Statistica 

Medica Biometria E Bioinformatica, Fondazione Irccs Istituto Nazionale 

Tumori, Milano, Italia 8)Direzione Scientifica, Istituto Nazionale Tumori 

Regina Elena, Roma, Italia 9)Direzione Scientifica, Istituto Nazionale 

Tumori, Bari, Italia 10)Anatomia Patologica, Istituto Nazionale Tumori 

Regina Elena, Roma, Italia 

Background. The increasing evidence of Trastuzumab efficacy 

in breast cancer (BC) patients means that an accurate and reproducible 

evaluation of HER2 status is of paramount clinical 

importance. Within the framework of the Italian network for 

Quality Assessment of Tumor biomarkers (INQAT), an External 

Quality Assessment (EQA) program was developed to investigate 

the state of the art of immunohistochemical (IHC) HER2 determination 

in the Lazio region and to monitor the performance of 16 

laboratories (CP) carrying out this assay. 

Methods. To this end, each CP received and fulfilled a questionnaire 

about their routine HER2 assay. The analysis of the 

questionnaires evinced a variability in the time of tissue fixation, 

in the reagents used, in the immunostaining method and in 

the evaluation criteria which indicated a wide methodological 

heterogeneity among laboratories. Furthermore, the discrepancy 

observed in the number of samples assayed per year, showing a 

different experience among the CP (from 100 to 500 samples), 

might affect the accuracy of HER2 testing. Aims of this project 

is to evaluate both the staining and interpretative reproducibility 

of the HER2 IHC assay within the 16 CP in order to develop a 

shared operating procedure. The Regina Elena Cancer Institute 

(Rome) is the Coordinating Center (CC) and one of the four Reviser 

Centers (CR) that select the cases and define the reference 

distribution of HER2 IHC score. 

Results. This study will allow to explore the potential influence 

of pre-analytical and analytical factors on laboratory performance 

of HER2 determination in the Lazio region. 

On the Behalf of the Regione Lazio-L’Aquila Network for HER2 

immunohistochemical Quality Control. 

Supported by Roche. 


“Triple negative”, “basal-like” and “non triple 

negative” breast cancers: clinical, histopathological 

and immunophenotypic comparison 

1)Thai E. 2)Campanini N. 3)Camisa R. 4)Boggiani D. 5)Musolino 

A. 6)Silini E. 

1)Anatomia Patologica, Azienda ospedaliera-universitaria di Parma, 

Parma, Italia 2)Anatomia Patologica, Azienda ospedaliera-universitaria 

di Parma, Parma, Italia 3)Unità operativa di oncologia medica, Azienda 

ospedaliera-universitaria di Parma, Parma, Italia 4)Unità operativa di 

oncologia medica, Azienda ospedaliera-universitaria di Parma, Parma, 

Italia 5)Unità operativa di oncologia medica, Azienda ospedaliera-universitaria 

di Parma, Parma, Italia 6)Anatomia Patologica, Azienda ospedaliera-universitaria 

di Parma, Parma, Italia 

Background. “Triple negative” (TN) and “basal-like” breast 

carcinomas (BLBC) are a poorly defined group of lesions characterized 

by heterogeneous histological features, poor clinical 

outcome and lack of specific therapies. BLBC and TN tumors 

share several clinical and pathological features, although they 

cannot be considered synonyms. Few studies have analyzed 

systematically the pathological features of TN compared with 

grade-matched non-triple-negative (NTN) carcinomas. We aimed 

to provide a detailed clinico-pathological comparison between 

TN and NTN tumours and to examine the prognostic impact of 

the BLBC phenotype. 

Methods. Two retrospective, consecutive series of 194 TN and 

101 NTN breast cancers (mean age: 61; grade 3: 84%; ductal 

type, 92%; stage 2+, 60%, mean follow-up: 7,5 years) matched 

for age, grade, stage and year of diagnosis were compared by 

case-control analysis. Tumors were evaluated according to a 

predefined set of validated pathological variables (Fulford et al., 

Histopathology. 2006). Immunostains for CK5/14, SMA, EGFR, 

ER and PgR were performed. Outcome data (survival and timeto-relapse, 

TTR) were available for 230 patients and were analyzed 

according to Kaplan-Meier. 

Results. Pathological features associated with TN tumors were 

pushing borders (p < 0.0001), necrosis (p < 0.0001), central 

scar (p < 0.0001), lymphocyte infiltrate (p < 0.0001), vascular 

invasion (p = 0.0019) and prominent nucleoli (p = 0.03). Basal 

markers, CK5-14, SMA and EGFR, were more frequent in the 

TN group (69%, 20% and 45% respectively, vs. 2%, 1% and 2% 

in NTN)(p < 0.0001). TN patients had shorter survival compared 

to controls (survival 73% vs 86%, p = 0.04), only TNM stage 

(p < 0.001) was prognostically relevant. BLBC showed no differences 

in survival or TTR, neither on the whole series nor within 

the TN group. 

Basal-like features are frequent among TN carcinomas, but do not 

correlate with outcome when controlling for tumor grade, stage 

and receptor status. 

A unique case of ovarian psammocarcinoma 

with omolateral serous cystoadenofibroma and 

thecoma associated with brenner tumour and 

cystoadenofibroma of the controlateral ovary 

1)Thai E. 2)Lombardi M. 3)Brigati F. 4)Giordano G. 

1)Adepartment of pathology and medicine of laborator, Parma, Parma, 

Italy 2)Adepartment of pathology and medicine of laborator, Parma, Parma, 

Italy 3)Adepartment of pathology and medicine of laborator, Parma, 

Parma, Italy 4)Adepartment of pathology and medicine of laborator, Parma, 

Parma, Italy 

Background. A unique case of ovarian psammocarcinoma with 

omolateral serous cystoadenofibroma and thecoma associated with 

Brenner tumour and cystoadenofibroma of the controlateral ovary 

was reported with clinical, macroscopic and microscopic features 

Material and methods A 78-year-old nulliparous woman underwent 

bilateral salpingo-oophorectomy because of the presence a 

right mass measuring 3.5 × 3 cm with a cystic area, simulating


a malignant neoplasm. The left ovary, instead, presented small 

cystic lesions. 

The surgical specimens were fixed in 10% neutral-buffered formalin 

for a routine light microscopic examination. 

Results. On sectioning, the right ovary disclosed the presence 

of three lesions. Two of these were solid, while the third was 

cystic. One of the solid lesions was very small, heavily calcified, 

small grey sub-capsular nodule measuring 1.5 × 0.5 × 1.5 cm. 

Histologically, this lesion corresponded to a psammocarcinoma 

showing numerous psammoma bodies occasionally surrounded 

by papillary and tubular structures lined by cytological bland 

cuboidal or low columnar epithelium. 

The cystic neoplasm containing serous fluid and papillary projections, 

histologically, corresponded to a serous cystoadenofibroma. 

The other solid lesion was yellow and, microscopically, 

corresponded to a thecoma. 

Two lesions of left ovary, histologically, corresponded to cystoadenofibroma 

and benign Brenner tumour. 

Conclusions. To the best of the author’s knowledge, ours is a 

unique example of ovarian psammocarcinoma with omolateral simultaneous 

thecoma and serous cystoadenofibroma, which were 

associated with Brenner tumour and cystoadenofibroma of the 

controlateral ovary. 

Moreover, this example of malignant serous epithelial tumour is 

most peculiar, since it shows: 

– a coexistence with multiple benign epithelial neoplasms and a 

benign stromal tumour; 

– measured only 1.5 × 0.5 × 1.5 cm and, to the best of the author’s 

knowledge, represents the smallest case of psammocarcinoma 

described in the literature. 

Tissue markers of stemness in high grade breast 

carcinomas: prevalence and correlations with 

triple negative and basal-like status 

1)Thai E. 2)Musolino A. 3)Camisa R. 4)Grondelli C. 5)Campanini 

N. 6)Silini E. 

1)Anatomia patologica,Azienda ospedaliera-universitaria di Parma, 

Parma, Italy 2)Unità operativa di oncologia medica, Azienda ospedaliera-universitaria 

di Parma, Parma, Italy 3)Unità operativa di oncologia 

medica, Azienda ospedaliera-universitaria di Parma, Parma, Italy 4)Unità 

operativa di oncologia medica, Azienda ospedaliera-universitaria di 

Parma, Parma, Italy 5)Anatomia patologica, Azienda ospedaliera-universitaria 

di Parma, Parma, Italy 6)Anatomia patologica, Azienda ospedaliera-universitaria 

di Parma, Parma, Italy 

Background. CD44high/CD24low phenotype and positivity for 

CD133 and aldehyde dehydrogenase (ALDH) have been proposed 

as markers of “stemness” in breast carcinomas. How the 

expression of stem cells markers correlates with pathological 

features and outcome is poorly known. 

Methods. The base of the study was a retrospective series of 

295 high grade invasive breast carcinomas (mean age: 61; grade 

3: 84%; ductal type, 92%; stage 2+: 60%, mean follow-up: 7,5 

years). Tumors were evaluated according to a predefined set of 

validated pathological variables (Fulford et al., Histopathology. 

2006). Immunostains for CD44, CD24, CD133, ALDH, CK5/14, 

SMA, EGFR, ER and PgR were performed. Outcome data (survival 

and time-to-relapse, TTR) were available for 230 patients 

and were analyzed according to Kaplan-Meier. 

Results. CD44 and CD133 expression correlated with “triple-negative” 

status (p = 0.0015 and p < 0.0001, respectively). 

CD44high/CD24low phenotype correlated with expression of 

CD133 (p = 0.0019) but not ALDH. CD44high/CD24low tumors 

had pushing borders (p = 0.024) and were associated with “basallike” 

status (p = 0.0005), as assessed by CK5/14 and/or SMA 

positive staining. Neither stem cell-like nor basal-like features 

significantly correlated with clinical outcome in the whole series 

and in TN tumors. 

367 

Stem cell-like, basal-like, and “triple negative” features largely 

overlap. In this series of high grade, mostly ductal carcinomas, 

markers of stemness had no prognostic value. New emerging 

traits of breast cancer, such as stemness and basal-like features, 

are only a part of a wider spectrum of biological attributes yet to 

be explored in their clinical implications. 

Computational analysis of systemic sclerosis 

microarray data and chronic graft versus host 

disease gene expression 

1)Tinazzi I. 2)Colato C. 3)Biasi D. 4)Caramaschi P. 5)Emery P. 

6)Del Galdo F. 

1)Medicina Clinica & Sperimentale, Università di Verona, Verona, Italia 

2)Patologia & Diagnostica, Università di Verona, Verona, Italia 3)Medicina 

Clinica & Sperimentale, Università di Verona, Verona, Italia 4)Medicina 

Clinica & Sperimentale, Università di Verona, Verona, Italia 5)Molecular 

Medicine, Leeds University, Leeds, United kingdom 6)Molecular 

Medicine, Leeds University, Leeds, United kingdom 

Background. Systemic Sclerosis (SSc) is a chronic disease with 

autoimmune activation, fibroproliferative vasculopathy and tissue 

fibrosis. The mechanisms linking immune activation and 

fibrosis are still not well characterized. A widely accepted model 

of immune mediated skin fibrosis is chronic Sclerodermatous 

Graft vs Host Disease (SclGVHD). Recent histological studies of 

cGVHD skin biopsies confirmed the presence of both fibroproliferative 

vasculopathy and fibrosis whereas vessels rarefaction is 

peculiar of SSc. 

Aim. To identify which of the genes differentially expressed 

in SSc skin samples are shared in the specific transcriptome of 

cGVHD skin samlpes and therefore of potential relevance in 

bridging the immune activation and the skin fibrosis. 

Methods. Metanalysis of all microarray data published in the 

literature identified a set of 80 genes whose differential expression 

is highly reproduced in SSc skin samples. mRNA expression 

level of these genes was measured in 9 cGVHD skin samples by 

RT-qPCR, compared to normal and SSc skin mRNA. All the 

genes found to be significantly differentially expressed (p < 0.05) 

in univariate analysis were tested in multivariate analysis. 

Results. 9 cGVHD pts were studied. Of the 80 genes analyzed, 

46 were differentially expressed in cGVHD samples of wich 36 

had a similar down or up-regulation in SSc. Of these, 9 genes 

were similarly expressed in all cGVHD, whereas 6 were specific 

of SclGVHD and 21 of nonScl GVHD. 

Conclusions. Computational analysis of SSc gene signature 

throught the expression levels of the same genes in the cGVHD 

variants allowed to identify which subset of genes may be involved 

in immune activation and immune mediated fibrosis in 

SSc and which ones may be responsible of peculiar tissue reaction 

of the SSc because differentially expressed only in SSc. 

Trap syndrome 

1)A. Napoli, 2)A. Tito, 1)D. Di Clemente, 1)G. Arborea, 3)C. 

Jezzoni, 3)R. Catacchio, 4)G. Napoli, 1)G. Caruso, 1)R. Ricco 

1)DAP, Policlinico, Bari, Italia; 2)Università degli Studi di Bari, Italia; 

3)DOG, Policlinico, Bari, Italia; 4)Anatomia Patologica Ospedale San 

Paolo, Bari, Italia 

Background. Twin-reversed arterial perfusion syndrome (TRAP 

syndrome) is a rare condition that may complicate 1% of monochorionic 

and monoamniotic twin pregnancies. In one of the 

twins there are congenital malformations, the most severe and 

rare of which is the acardia (1 case per 35,000 births) associated 

with anencephaly and single umbilical artery. Acardic fetus receives 

oxygen poor blood from the healthy twin with an umbilical 

artery where there is a reverse flow. Reversed arterial perfusion 

is lethal for the acardic twin, but it can also cause death of the 

normal twin in 50% of cases. An early prenatal diagnosis is an es-

368 

sential prerequisite to make an appropriate management strategy 

of these pregnancies. 

Methods. A corpse comes to our observation with secondary 

sexual characteristics of female type, weighing 450 grams, clearly 

under anasarcatic state. Clinical News: Albanian mother, with a 

history of repeated miscarriages (six), hypertensive, with “rubeo”, 

“toxo” and “CMV” tests negative. The transvaginal ultrasound 

performed at 26 weeks of gestation shows: “twin monochorionic 

pregnancy complicated by “TRAP sequences”. Acardic fetus 

consists of soft and dropsy tissue mass (about 15cm) in which we 

recognize the spine and outline of the rib cage and limbs. The 

healthy fetus has no structural abnormalities detectable by ultrasound 

and presents a moderate cardiomegaly”. 

Results. Autopsy report. External examination: Head and neck: 

presence of soft-tissue formation, about 18 cm in diameter, 

we do not identify structures related to the skull, ear, eye and 

choanae structures. There are sketches of the upper limbs, with 

absence of phalanges bilaterally, and chest wall. The legs are 

recognizable by the presence of four fingers. Opening does not 

show the thoracic organs, stomach, liver, spleen, right kidney 

and internal genitalia. The macroscopic finding agrees with the 

literature. 

fine needle aspiration cytology of pilomatrixoma: 

a series of 25 cases with clinical correlations and 

differential diagnosis 

Todaro P, Bonanno AM, Speciale G, Ieni A, Branca G, *Catalano 

F, *Catalano A, Tuccari G. 

Department of Human Pathology (Section of Pathological Anatomy) and 

*Department of Surgical Specialties (Section of Plastic Surgery), A.O.U. 

Policlinic G.Martino, University of Messina, Italy 

Background. Pilomatrixoma (PMX) is uncommon benign skin 

tumour, generally slow-growing, with a predilection for the neck 

and face as well as extremities; it may occur at any age and may 

have an unusual clinical presentation, causing some problems in 

the differential diagnosis. Few reports regarding cytological features 

of PMX have been reported in the literature; in the present 

study, we report pre-operative diagnostic cytological characteristics 

based on a series of 25 PMX, underlining the differential 

diagnosis in relation to other cutaneous lesions. 

Methods. Twenty-five PMX were pre-operatively aspired by a 

23 G needle attached to a 10-ml disposable syringe. Cases of 

PMX (11 male, 14 female; mean age 32,72 years; ager range 

3-78 yrs) were successively surgically treated and subjected to 

routine histology in order to confirm the cytological diagnosis. 

The smears collected were air-dried and stained with May-Grunwald-Giemsa; 

additional samples were fixed in 95% ethanol and 

stained with Papanicolaou method. 

Results. All patients had a single tumour, which was localized on 

the pre-auricular (2), face (10), neck (1), scalp (4), chest (1), arm 

(3), leg (2), foot (2) Generally, all aspirates were cellular, consisting 

of clusters of small and medium-sized basaloid cells; their 

nuclei were round, regular, to ovoid, vesicular, with dispersed 

chromatin and occasional large nucleoli. Sheets or isolated ghost 

(shadow) cells were found in all smears, sometimes associated 

with multinucleated giant cells, keratin fragments, cellular debris 

and naked nuclei; in seven cases inflammatory cells were observed. 

Moreover, the cytological differential diagnosis for other 

small cell and keratinizing skin lesions was also analyzed, taken 

into consideration the clinical presentation and the final outcome. 

Finally, in all cases, the histopathology of resected tumours confirmed 

the cytological diagnosis of PMX. 


Surgery or follow-up in pt1 adenocarcinoma ex 

adenoma? An hypothesis of further markers of 

risk in a series from colorectal cancer screening 

files in fVG 

1) Avellini C. 2)Toffoli S. 3)Marzinotto S. 4)Orsaria M. 5)Zanier 

L. 6)Beltrami CA. 

1)Scienze Mediche e Morfologiche, Azienda S. Maria Della Misericordia, 

Udine, Italia 2)Scienze Mediche e Morfologiche, Azienda S. Maria Della 

Misericordia, Udine, Italia 3)Scienze Mediche e Morfologiche, Azienda S. 

Maria Della Misericordia, Udine, Italia 4)Scienze Mediche e Morfologiche, 

Azienda S. Maria Della Misericordia, Udine, Italia 5)Agenzia Regionale 

Sanità FVG, Agenzia Regionale Sanità FVG, Udine, Italia 6)Scienze 

Mediche e Morfologiche, Azienda S. Maria Della Misericordia, Udine, 

Italia 

Background. Colorectal cancer screening from 2008 in Friuli 

Venezia Giulia showed pT1 cases. Surgery vs follow-up choice 

relies on the distance between infiltration and resection margin, 

tumoral budding, microstaging, grading, vascular invasion. 

Microsatellite instability (MSI), Apaf-1, Clusterin, m-TOR and 

Pdcd4 are markers of early cancer with various expression; BRaf 

mutation has adverse effect in MSI tumors. 

Aim. of the study is to test a panel of risk markers in pT1 polyps 

to confirm morphologic criteria. 

Methods. First studied cases are from the Institute of Pathology 

of Udine: pT1 screening cases are 18, 8 without surgery. 22 cases 

from screening have polipectomy alone (9 cases) or surgical 

specimen (13 cases). G2, G3, vascular invasion, > than 50% of 

carcinoma in adenoma, budding, according to polypectomy margins 

status, define > risk lesions. pT1 polyps have been analyzed 

with above mentioned markers; BRaf study is ongoing, firstly in 

operated pT1 cases. 

Results. 7 polypectomy cases had + margins and low risk, while 

4 cases were at high risk with + margins. 8 cases with - margins 

showed low risk and 1 case high risk. 8 surgical specimen had 

residual neoplasia (pT1), 15 cases were – and 1 case had lymph 

node metastases. BRaf study showed mutation in 1 screening case 

(+ margins, high grade budding, G2). 

IHC panel: Apaf-1 + in high and low risk cases and + margins 

(60%); 25% + in high risk cases with - margins. Clusterin + in 

45% of high and low risk cases and + margins; 65% of high and 

low risk cases with + margins and 45% of high risk cases with 

- margins show mTOR reactivity in invasive component. + margins 

were associated with Pdcd-4 reactivity in 70% of high risk 

cases. 80% of low risk and - margin cases are MSI. 

Conclusion. Preliminary results show an association between 

IHC, residual submucosal invasion and shorter survival in non 

screening cases. pT1 cases on Regional ground will be collected 

and studied. This panel may be useful for patient management. 

epidermoid cyst of the thyroid: report of a case 

Toraldo M., Pietropaoli N., Lupi C., Cavaliere A. 

Institute Of Pathological Anatomy, Santa Maria Della Misericordia, 

Perugia, Italy 

Background. Thyroid epidermoid cyst is a rare epithelial lesion 

with only few cases reported in literature 1 . In this report we describe 

a new case in an older woman where an epidermoid cyst 

was an incidental finding in the thyroid gland. 

Case History. A 75 year old woman presented with a mass on 

the left side of the neck. An ultrasound examination demonstrated 

a solitary, hypoechoic nodule inside the left lobe of the thyroid 

gland. Fine needle aspiration of the nodule showed only rare follicular 

epithelial cells without atipia; total thyroidectomy was performed. 

The macroscopic exam evidenced a nodule of 4.5 cm in 

the left lobe; histopathological examination revealed a follicular 

adenoma and an intraparenchymal cyst lined by benign stratified 

squamous keratinized epithelium, resembling the epidermoid cyst


of the skin. Neither cutaneous adnexal structure nor lymphoid 

tissue was found. 

Discussion. Thyroid epidermoid cyst is an extremely rare lesion 

with only occasional cases reported. There are no differences in 

sex distribution; the age range from 4 to 60 years, with a mean of 

42 yr. The pathogenesis in not known and it is probably due to 

inclusions of epithelial tissue during embryological development 

(remnant of the ultimobranchial body, branchial pouch) 2 . Histologically, 

it can be easily differentiated from dermoid cyst by the 

absence of cutaneous adnexal structures and from branchial cleft 

cyst by the absence of lymphoid infiltrate. It is more difficult to 

differentiate from thyroglossal duct cyst which can be lined only 

by squamous epithelium without columnar epithelium 2 . The cyst 

may be asymptomatic and incidentally found only after thyroidectomy. 

For this reason it may be discovered in older patients, as 

in our case. 

references 

1 Chen KTK. Diagn Cytopathol 2007;35:123-4. 

2 Magro G, et al. Pathologica 2006;98:126-8. 

Immunohistochemical expression of interferonstimulated 

genes in hepatitis C: a possible 

predictive role? 

1)Tornillo L. 2)Brand R. 3)Sarasin-filippowicz M. 4)Dill M. 

5)Baumhoer D. 6)Heim M. 7)Terracciano LM. 


of pathology, University of basel, Basel, Switzerland 3)Department of 

biomedicine, University of basel, Basel, Switzerland 4)Department of biomedicine, 

University of basel, Basel, Switzerland 5)Institute of pathology, 

University of basel, Basel, Switzerland 6)Department of biomedicine, 


of basel, Basel, Switzerland 

Background. The standard therapy for chronic hepatitis C is a 

combination of interferon (IFN) and Ribavirin, but up to 40% 

of treated patients are nonresponders. IFN achieves its antiviral 

effects through the regulation of hundreds of IFN-stimulated 

genes (ISGs) and it has been shown that the dysregulation of the 

expression of 29 genes, mainly ISGs, may correctly predict the 

response to therapy. 

Aim. To correlate the immunohistochemical (IHC) expression of 

selected ISGs and the features of inflammatory infiltrate with the 

response to IFN-therapy. 

Methods. 116 liver biopsies of 80 HCV-patients were taken at 

different times after the beginning of IFN-therapy (4, 16, 48, 96, 

144 hours, 4, 8, 12 and 96 weeks) and stained immunohistochemically 

(IHC) for glypican 3 (GPC3), pSTAT1 and SOCS3 (ISGs), 

CD3, CD20, CD56 and CD68. The number of positive cells for 

each marker was counted and correlated with the response to IFN 

at different times. 

Results. There was a significant difference in the number of 

GPC3+ cells (p < 0.001) between responders and not-responders 

at 4, 12 and 96 weeks. No significant difference (p = 0.619) was 

found before and after the therapy. By a cut-off of 60 GPC3+ 

cells, the response to therapy could be predicted in 84.1% of the 

cases. An association between pSTAT1-positivity and activation 

of macrophages was seen (p < 0.001). 

Conclusions. We observed a „preactivation” of some ISGs 

(GPC3 and pSTAT1) in nonresponders. GPC3 IHC positivity was 

a strong predictor of the response to therapy. It is not clear how 

GPC3 may influence the response to IFN. It may be involved in 

the control of the receptor-ligand interactions or play a role in 

macrophage recruiting, as suggested by the dramatic increase of 

CD68+ cells in biopsies with high pSTAT1 count. It is tempting 

to speculate that the determination of ISGs may help to identify 

responders to IFN therapy. 

Incidence and histotypes of thyroid carcinoma 

in eastern Sicily 

369 

Torrisi A., Castaing M., Sciacchitano S., Fidelbo M., Benedetto 

G., Madeddu A., Vasquez E., Ieni A., Leone A., Sciacca S. 

Dipartimento “G.F Ingrassia”, Registro Tumori Integrato Catania-Siracusa-Messina, 

Azienda Ospedialiero-Universitaria Policlinico Universitario 

Vittorio Emanuele, Catania, Italia 

Background. The incidence of papillary thyroid carcinoma (PTC) 

has significantly increased over the past three decades especially 

in areas with active volcanoes including Hawaii, Philippines and 

Iceland. The aim of our study is to evaluate thyroid carcinomas 

(TCs) incidence and the distribution of different histotypes in 

three Sicilian provinces: Catania (CT), Siracusa (SR) and Messina 

(ME) and to assess the relationships between concentrations 

of elements such as boron (B), iron (Fe), manganese (Mn) and 

vanadium (V) in drinking water of CT province and increased 

risk for TC incidence as suggested by recent studies. 

Methods. Data was extracted from the Cancer Registry of CT- 

ME-SR-EN from the period 2003-2005. Incidence data was 

expressed in cases for 100.000 residents per year. There were 

calculated standardized rates (Italian census of 2001) and their 

relative confidence intervals at 95%. Resident population was 

taken from ISTAT for each year of registration and the mean 

was computed on the whole period. Distribution differences were 

assessed through Chi-square test. For water analysis, only the 25 

most important towns of the CT province were selected. Data on 

water concentrations of chemicals were furnished by ARPA. Correlation 

between incidence rates and concentrations was assessed 

through Spearman correlation test. 

Results. Standardized incidence rates in CT province were 9.2, 

95%CI = 7.7-10.7 in men and 36.7 in women, 95%CI = 33.8- 

39.6, per 100.000 per year. In ME there were respectively 4.8, 

95%CI = 3.4-6.2 and 25.5, 95%CI = 22.4-28.6 and in SR 2.8, 

95%CI = 2-3.6 and 8.8, 95%CI =7 .7-9.9. Ratio of papillary and 

follicular carcinomas were respectively 50.8, 6.9 and 14.4 in CT, 

ME and SR. Microcarcinomas distribution among provinces was 

statistically different being 32.3% in CT, 14.9% in ME and 12.7% 

in SR (p < 0.0001). Notably, no correlation was found between B 

concentrations (ρ = 0.29, p = 0.16), Fe (ρ = -0.18, p = 0.38), Mn 

(ρ = 0.09, p = 0.67) and V (ρ = -0.11, p = 0.61) and incidence of 

TCs in CT. An extended analysis on radon and iodio-131 is in 

progress. 

Our findings confirm higher incidence of TCs, especially PTC, 

in CT province. Additionally, there were statistically significant 

differences in the distribution of PTC ≤ 1 cm between CT vs 

the other two. Whether this finding could be explained by a 

more accurate gross and microscopy examination is to be established. 

What’s dominant nodule in Hashimoto’s thyroiditis: 

a clinico-pathologic study of 219 patients 

1)P. Amico, 2)A. Torrisi, 1)L. Salvatorelli, 2)M. Castaing, 1)G. 

M. Vecchio, 1)P. Gangemi, 3)M.G. Tranchima, 4)M. Cannizzaro, 

1)G. Magro 



Catania, Italia; 2)Dipartimento G.F. Ingrassia, Registro Tumori Integrato-Messina-Catania-Siracusa, 

Catania, Italia; 3)Servizio Anatomia Patologica, 

Azienda Ospedaliera Garibaldi, Catania, Italia; 4)Dipartimento 

Scienze Chirurgiche-Endocrino-Chirurgia, Azienda Ospedaliero-Universitaria 

Policlinico-Vittorio Emanuele, Catania, Italia 

Background. Although most thyroids affected by Hashimoto’s 

thyroiditis (HT) are diffusely enlarged, with a micronodular appearance, 

some patients may occasionally develop one palpable 

or non-palpable nodule of at least 1 cm in diameter, overgrowing 

from thyroid parenchyma. This nodule, labeled as “dominant

370 

nodule (DN)”, is commonly believed to represent a thyroid carcinoma 

arising in HT. Unfortunately the incidence, nature and 

clinical significance of DN are still controversial because the little 

data available in the literature are quite confusing. 

Materials. We selected retrospectively a series of 219 patients 

with histologically proven HT. The revision of gross pathology 

reports and original H&E stained slides led us to identify a subset 

of patients with “DN”, namely a nodule measuring at least 1.5 cm 

in diameter. The morphological features of “DN” were statistically 

correlated with clinical parameters, including preoperative 

FNAC diagnosis, whenever possible. 

Results. We found that 30% of patients with HT had “DN” (1.5 to 

4.5 cm). In most cases (88%) the nodule was single, while in 12% 

two nodules were identified. Histologically 89.7% of single DNs 

resulted to be “hyperplastic follicular lesions (HFL)”, whereas only 

10.3% were PTC. Among patients with two dominant nodules, 

62.5% had one HFL plus PTC, while in the remaining 37.5% both 

nodules were HFLs. With regard to HFL, 72% were composed of 

non-Hurthle cells, while 28% contained exclusively Hurthle cells. 

Interestingly 57% of HFLs, regardless of cytological composition, 

lacked an associated inflammatory component. 

Our findings show that most DNs in HT are HFLs and not PTCs 

or follicular neoplasms as commonly believed. Although diagnosis 

of HT is straightforward when both lymphoid and Hurthle cells are 

present, we found that most HFLs are composed of non-Hurthle 

cells and lack inflammation. Accordingly, pathologist should be 

aware of this possibility to avoid the misdiagnosis of follicular neoplasms 

in HT, either preoperatively (FNAC) or post-surgically. 

Microsatellite instability DNA testing in routinely 

processed colorectal carcinomas: correlation 

with clinicopathologic and survival data in 340 

consecutive cases 

1)Tosi AL. 2)Morandi L. 3)De Biase D. 4)Pession A. 5)Maestri 

A. 6)Turchetti D. 7)Baccarini P. 8)Brulatti M. 9)Tallini G. 

1)Dipartimento di Ematologia e Scienze Oncologiche “L&A Seragnoli”, 

sezione di Anatomia Patologica, Ospedale Bellaria, Università di Bologna, 

Bologna, Italia 2)Dipartimento di Ematologia e Scienze Oncologiche 

“L&A Seragnoli”, sezione di Anatomia Patologica, Ospedale Bellaria, 

Università di Bologna, Bologna, Italia 3)Dipartimento di Ematologia e 

Scienze Oncologiche “L&A Seragnoli”, sezione di Anatomia Patologica, 

Ospedale Bellaria, Università di Bologna, Bologna, Italia 4)Patologia 

sperimentale, Ospedale Bellaria, Bologna, Italia 5)Dipartimento di Oncologia, 

U.O.C. Oncologia Medica Ospedale Bellaria Azienda AUSL di 

Bologna, Italia 6)Cattedra e U.O. Genetica Medica Università di Bologna-Policlinico 

Sant’Orsola-Malpighi, Italia 7)Dipartimento di Scienze 

Oncologiche, U.O. di Anatomia Patologica, AUSL di Bologna, Ospedale 

Bellaria 8) U.O. Chirurgia generale ad indirizzo oncologico, Dipartimento 

di Scienze Oncologiche, AUSL di Bologna, Ospedale Bellaria, Bologna, 

Italia 9)Dipartimento di Ematologia e Scienze Oncologiche “L&A Seragnoli”, 

sezione di Anatomia Patologica, Ospedale Bellaria, Università di 

Bologna, Bologna, Italia 

Background. Colorectal cancer (CRC) is characterized by a 

multistep progression of genetic errors. Two different pathways 

are identified, that of chromosomal instability (CIN) and the microsatellite 

instability (MSI) pathway. Widespread microsatellite 

instability (MSI high, MSI-H phenotype) is present in 10-20% 

of sporadic colorectal cancers. MSI-H tumors have distinctive 

pathologic features and are believed to behave less aggressively 

when compared with tumors that lack microsatellite instability 

(MS stable, MSS) or that show instability at a few loci (MSI low, 

MSI-L phenotype). 

Methods. We studied 340 consecutive CRCs for MSI using 

multiplexed polymerase chain reactions (PCR) for 12 microsatellite 

markers. DNA was extracted from routinely processed 

formalin-fixed tissue. All surgical specimens underwent routine 

histopathological analysis for grading and staging. Tumors were 

considered MSI-H when 4 or more of the 12 loci were mutated, 


MSI-L when 1-3 loci were mutated and MSS when no mutations 

were identified. 

Results and Conclusions. 40 CRCs were MSI-H (12%), 45 

CRCs were MSI-L (13%) and 255 CRCs were MSS (75%). 

Correlation with clinicopathologic features confirms previous 

findings, MSI-H CRCs are more common in the right colon, 

display poorly differentiated histology and show prominent lymphocytic 

infiltration. Survival analysis after a median follow-up 

of 45 months shows that MSI-H tumors have a better prognosis in 

patients with stage 1 and 2 disease, but the prognosis is considerably 

worse for patients with stage 3 and 4 disease (p < 0.01). 

MSI-H CRCs show distinctive clinical and pathological features. 

In our series patients survival depended on tumor stage at presentation. 

The results of the study argue for MSI testing on routinely 

processed CRCs. 

A case of carcinosarcoma of the breast: 

hypothesis for its origin 

1)V. Toto, 2)G. Castrilli, 1)R. Claudi, 1)F. Sabatini, 1)D. Angelucci 


Chieti, Italia; 2)Anatomia Patologica, G. Bernabeo, Ortona, Italia 

Background. Carcinosarcoma is a rare, aggressive form of breast 

cancer. The diagnosis of carcinosarcoma is strictly defined by the 

presence of both epithelial and mesenchymal neoplastic cells, 

without a transition zone between the two lines. 

Methods. We reported here our experience with a case of carcinosarcoma 

of the breast in a 41-years old woman with a previous 

history of bilateral benign lesions, admitted to our hospital due to 

the rapid increasing of a right breast mass. 

Despite its round and regular form, needle biopsy was performed, 

showing the etheroplastic origin of the lesion. Bilateral quadrantectomy 

was done. 

Results. Microscopically the tumour consisted of neasts of invasive 

ductal carcinoma,surrounded by neoplastic stroma with 

sarcomatous features. These two components interlocked with 

each other without transition areas. Central necrosis and malignant 

calcification were seen; despite great vascular invasion, no 

metastasis to axilllary node was found. 

Immunostained sections reveal positive reaction in the epithelial 

component for CK5, AE1 and AE3 and both estrogen and progesterone 

receptors, while sarcomatous cells were positive for Cd10 

and vimentin, both markers of myoepithelial origins. Epithelial 

cells were Her-2 negative. 

Carcinosarcoma of the breast was then diagnosed. 

The positivity for Cd10 and vimentin, besides a previous history 

of fibroadenomas, support the hypothesis that this tumour could 

arise from pre-existing fibroadenoma or phylloid tumour. 

At the other side the absence of a global structure resembling 

phyllodes pattern, suggest instead that the two neoplastic population 

might have arised from two different kinds of cancer stem 

cells, epithelial and mesenchymal. 

Carcinosarcoma are rare, aggressive tumour, often null-subtype, 

with high recurrence rate. In our case, the node negative state and 

the hormonal receptors positivity might be considered a favorable 

prognostic element. 

essential role of the p110β subunit of 

phosphoinositide 3OH-kinase in male fertility 

1)Toto (V). 2)Liberatore (M). 3)Ascione (P). 4)Ciraolo (E). 

5)Morello (F). 6)Xiaoyun (L). 7)Pandolfi (PP). 8)Hirsch (E). 

9)Musiani (P). 10)Iezzi (M). 


Chieti, Italia 2)Anatomia Patologica/Oncologia e Neuroscienze, Ss. Annunziata/cesi, 

Chieti, Italia 3)Anatomia Patologica/Oncologia e Neuroscienze, 

Ss. Annunziata/CESI, Chieti, Italia 4)Mbc, Genetica, Biologia e


Biochimica, Torino, Italia 5)Mbc, Genetica, Biologia e Biochimica, Torino, 

Italia 6)Mbc, Genetica, Biologia e Biochimica, Torino, Italia 7)Beth 

Israel Decanes, Medical Center, Boston, Usa 8)Mbc, Genetica, Biologia 

e Biochimica, Torino, Italia 9)Anatomia Patologica/Oncologia e Neuroscienze, 

Ss. Annunziata/CESI, Chieti, Italia 10)Anatomia Patologica/Oncologia 

e Neuroscienze, Ss. Annunziata/CESI, Chieti, Italia 

Background. Phosphoinositide 3-kinases (PI3K) are key molecular 

players in male fertility. However, the specific roles of 

different p110 PI3K catalytic subunits within the spermatogenic 

lineage have not been characterized so far. 

Methods. Herein, we report that male mice expressing a catalytically 

inactive p110β develop testicular hypotrophy and impaired 

spermatogenesis, leading to a phenotype of oligo-azoospermia 

and defective fertility. The examination of testes from p110β-defective 

tubules demonstrates a widespread loss in spermatogenic 

cells, due to defective proliferation and survival of pre- and postmeiotic 

cells. In particular, p110β is crucially needed in c-Kitmediated 

spermatogonial expansion, as c-Kit-positive cells are 

lost in the adult testis and activation of Akt by SCF is blocked by 

a p110β inhibitor. 

Results. These data establish that activation of the p110β PI3K 

isoform by c-Kit is required during spermatogenesis, thus 

opening the way to new treatments for c-Kit positive testicular 

cancers. 

rare cancers in italy: the results of the 

“surveillance of rare cancers in italy” (rITA) project 

1)G. Gatta, 1)A.Trama, 2)S. Ferretti, 1)L. Licitra, 1)P. Casali, 

3)R. Capocaccia, 3)R. De Angelis, 3)S. Mallone, 3)M. Santaquilani, 

3)A. Tavilla, 4)P.A. Dei Tos and the RITA working group 

1)Fondazione IRCSS Istituto Nazionale dei Tumori, Milano, Italia; 2)Registro 

Tumori di Ferrara, Università di Ferrara, Italia; 3)Istituto Superiore 

di Sanità, Roma, Italia; 4)Ospedale di Treviso, ASL, Treviso, Italia 

Background. The “Surveillance of Rare Cancers in Italy” (RITA) 

project aimed at providing a definition of “rare cancer”, a list of 

cancers and rare cancer burden indicators, based on Italian population-based 

cancer registry data. RITA integrates the European 

Project “Surveillance of rare cancers in Europe” RARECARE 

(co-funded by the European Commission). 

Definition and list of rare cancers. An international consensus 

group developed a list of clinically relevant cancer entities. Accordingly, 

a tentative threshold for rarity (≤ 6/100.000/year) was 

chosen by clinical consensus leading to a list of “rare cancers”. 

The list of rare cancers was based on the International Classification 

of Diseases for Oncology (3 rd edition). The list was hierarchically 

structured in 2 layers: layer 1) families of tumours (relevant 

for the health care organisation) and layer 2) tumours clinically 

meaningful (relevant for clinical decision making and research). 

The list is available on the project website: www.rarecare.eu. 

The list includes 194 rare cancers. The Italian data came from 19 

population-based cancer registries. 

Results. According to our estimates 450,000 patients are living 

today with a diagnosis of rare cancers in Italy and every year 

there are 67,300 new diagnoses (22% of all new malignant cancers 

diagnosed). Rare cancers had, on average, worse relative 

survival than common cancers. Five years relative survival was 

51% for rare cancers and 69% for the common ones. 

Conclusions. Our results disclose the burden of rare cancers 

in Italy, represent an important baseline for future research and 

confirm that despite the rarity of each individual cancer type, 

rare tumours significantly contribute to the total cancer burden 

in Italy. Our data confirm that population-based CRs and databases 

are key instruments to increase knowledge on rare tumours 

and develop clinical research. In addition, our study proposed a 

definition and a list of rare cancers which provides a common 

language for rare cancers. 

371 

Pheochromocytomas and paragangliomas scoring 

systems: advantages and limits 

1)Tricarico P. 2)Cappellesso R. 3)Guzzardo V. 4)Schiavi F. 

5)Fassan M. 

1)Scienze Medico Diagnostiche e Terapie Speciali, Università di Padova, 

Padova, Italia 2)Scienze Medico Diagnostiche e Terapie Speciali, Università 

di Padova, Padova, Italia 3)Scienze Medico Diagnostiche e Terapie 

Speciali, Università di Padova, Padova, Italia 4)Dipartimento di Oncologia 

Clinica e Sperimentale, Istituto Oncologico Veneto, Padova, Italia 

5)Scienze Medico Diagnostiche e Terapie Speciali, Università di Padova, 

Padova, Italia 

Background. Pheochromocytoma and paraganglioma are rare 

tumors arising from chromaffin cells of the adrenal gland 

and paraganglia. Their malignancy is defined by direct local 

invasion of sites that do not typically have chromaffin tissue, 

occurring from 13 to 26% of all cases. Two scoring systems 

have been proposed: Pheochromocytoma of the Adrenal Gland 

Scaled Score (PASS, 2002) and Kimura (2005), aiming to separate 

benign from malignant neoplasms; PASS was conceived for 

adrenal tumors only and was based on 12 histological features, 

while Kimura was proposed for all chromaffin tissue-derived 

tumors and included histological, immunohistochemical and 

clinical features. 

Materials. We compared 217 cases of pheochromocytomas 

(n = 160) and paragangliomas (n = 57) between 1988 and 2009, 

including 11 malignant pheochromocytomas, retrieved from the 

archives of the Department of Pathology of Padova University. 

Each sample was evaluated using both PASS and Kimura scoring 

systems. 

Results. PASS scoring system detected 77 tumors with high 

risk of malignancy (score > 4) while Kimura detected 16 tumors 

(score 7-10, corresponding to poorly differentiated tumor). 

Conclusions. Both scoring systems correctly identified the 

11 malignant pheochromocytomas, but PASS scoring system 

clearly overestimated malignancy as it recognized 66 other 

cases as high risk tumors (specificity = 55%). Kimura scoring 

system demonstrated to be more precise as it detected only 5 

other tumors with high risk of malignancy (specificity = 97%). 

Nevertheless, none of the two systems seemed definitely reliable 

as a diagnostic tool for malignancy, and a more ample clinical, 

imaging, genetic, immunohistochemical, and molecular characterization 

is required. 

role of the stromal master regulator SPArC in 

myeloproliferation-induced bone marrow stroma 

disarrangement 

C. Tripodo1 , S. Sangaletti2 , C. Guarnotta1 , P.P. Piccaluga3 , 

A. Carè4 , M.P. Colombo2 , A.M. Florena1 1Dipartimento di Patologia Umana, Università di Palermo, Italia 

2Unità di Immunologia Molecolare, Istituto Nazionale Tumori, Milano, 

Italia 

3Dipartimento di Ematologia ed Oncologia, Università di Bologna, Italia 

4Dipartimento di Ematologia, Istituto Superiore di Sanità, Roma, Italia 

Background. In myeloproliferative neoplasms, the bone marrow 

stroma may be driven towards dynamic changes that might eventually 

cause the disarrangement of the functional hematopoietic 

niches thus emerging as a clinical and prognostic determinant. 

The secreted protein acidic and rich in cysteine (SPARC) has 

emerged as a master regulator of the stromal homeostasis in 

tissues undergoind remodelling, through its ability to modulate 

TGF-beta and other epithelial-to-mesenchymal transition signalling 

pathways and to influence the fate of mesenchymal cell differentiation 

programs (e.g. WNT/beta-catenin). 

Purpose of the study. In this study we investigated the contribution 

of the matricellular protein SPARC to the BM stroma remodelling 

associated with myeloproliferative neoplasms.

372 

Summary of results. By studying the expression of SPARC in 

58 cases of myeloid neoplasms (including Ph- MPN, MDS and 

AML) with or without overt stromal changes (i.e. marked increase 

in agiogenesis and fibrosis), and in 16 control BM samples, we 

found that SPARC protein expression showed a striking increase 

in samples undergoing stromal changes, as compared to those 

with preserved stromal architecture. Unlike in control samples 

and in samples without stromal changes, where SPARC expression 

was confined to megakaryocytes and endosteal fibroblasts, 

in cases with stromal modifications SPARC was expressed also 

by spindle-shaped and stellate stromal cells intermingling with 

hematopoietic cells, and surrounding vessels. 

Notably, by double immunofluorescence on confocal microscopy, 

and by the means of BM stromal cell culture analysis, we 

could demonstrate that SPARC expression also characterized the 

BM CD146+mesenchymal stem cell component (BMMSCs), 

which we recently found expanded in the advanced phases of 

myeloproliferative neoplasms with myelofibrosis. Moreover, we 

found that SPARC induction in BMMSCs preferentially oriented 

BMMSCs towards an osteblastic fate and hampered adipocytic 

differentiation. 

Conclusion. Altogether, our preliminary results suggest a role 

for SPARC in disregulating the balance between the normal BM 

hematopoietic niches by favouring the expansion of an altered BM 

stroma prone to ECM deposition, angiogenesis and new bone formation. 

They also candidate SPARC as a possible target of interference 

in the setting of patients with advanced stromal changes. 

Ameloblastic fibrosarcoma of the mandible. 

A case report 

1)Unti E. 2)Scibetta N. 3)Marasà L. 

1)Department of pathology, Arnas civico hospital, Palermo, Italy 2)Department 

of pathology, Arnas civico hospital, Palermo, Italy 3)Department 

of pathology, Arnas civico hospital, Palermo, Italy 

Background. Ameloblastic fibrosarcoma(AFS),is a rare mixed 

odontogenic tumour,with fewer than 50 cases reported in the 

world literature,consisting of a benign epithelial and malignant 

mesenchymal component.Two-thirds of AFSs seem to arise 

de novo,but the other has developed in recurrent ameloblastic 

fibromas or ameloblastic fibroodontomas.This tumor occurs 

chiefly in the mandible of the male young adults.Metastasis 

is not a feature of the lesion and fatal cases usually have been 

associated with uncontrollable local infiltration,following numerous 

recurrences.We present a case of AFS arising in ameloblastic 

fibroma. 

Methods. A 20-year-old man presented with painless mass in 

the left posterior region of the mandible.The panorex radiograph 

revealed a radiolucent lesion in the bone, measuring 20 mm in 

maximum diameter.Surgical removal and curettage of the lesion 


Results. The tumor was composed of anastomosing islands and 

strands of benign epithelial cells,reminiscent of the early stages of 

enamel organ development, including a central component with 

stellate reticulum type morphology,within a background of connettive 

immature and loosely cellular,reminiscent of the dental 

papilla,with foci of a more dense population of spindle cells,with 

nuclear pleomorphism and frequent mitotic figures.There was no 

evidence of cell pleomorphism in the epithelial component.The 

spindle cells were positive for vimentin,but negative for smooth 

muscle actin,S100 protein,MyoD1,CD68,c-kit,CD34.The benign 

looking ameloblastic epithelium showed immunoreactivity for 

pancytokeratin.This is the typical histological change occurring 

during malignant transformation from ameloblastic fibroma to 

AFS.The patient is now doing well without recurrence of disease 

24 months after surgery.While cases of AFS have been observed 

as arising de novo,our case appears to substantiate the transformation 

of ameloblastic fibroma to AFS. 


Desmoplastic small round cell tumor. A case report 





Background. Desmoplastic small round cell tumor(DSRCT) is 

a rare but highly aggressive neoplasm with poor outcome with 

predilection for adolescent male.It usually affects the abdominal 

cavity,with widespread peritoneal involvement at the time 

of diagnosis.It is composed of nests of small,undifferentiated 

round hyperchromatic cells,embedded in abundant desmoplastic 

stroma,with co-expression of epithelial,mesenchymal and neural 

antigens in the same cell.Cytogenetically it harbors a specific 

karyotypic abnormality,namely t(11;22)(p13;q12).We report a 

case of DSCT. 

Methods. A 12-year–old woman presented with abdominal 

pain lasting for 4 months.The abdominal CT scan revealed 

wellenhanced large retroperitoneal mass,8 cm in maximum 

diameter,with widespread peritonealand ovaric involvement.Ascites 

was also found.She was submitted to debulking surgery. 

Results. The gross appearance consists of multiple tumour 

nodules with cut surface firm,grey-white.The pathologic results 

included a poorly differentiated tumor composed of well-defined 

nests of small round blue cells,with scanty cytoplasm,separated by 

abundant desmoplastic stroma.The tumor cells were uniform and 

the chromatin was evenly dispersed with inconspicuous nucleoli. 

Apoptosis and nuclear molding,mitotic figures were identificable. 

The tumor cells were immunoreactive for keratin,EMA,vimentin 

,desmin,NSE,WT1(+),negative for CD99,chromogranin, synapt 

ophysin,HMB45,S100,CD3,CD20,CD117,Myf4.RT-PCR studies 

have demonstrated a characteristic reciprocal chromosomal 

translocation,t(11;22)(p13;q12)in the mass.The bone marrow 

biopsy was negative.Histologic diagnosis was DSRCT. DSRCT 

requires differential diagnosis for various neoplasms such as Ewing 

sarcoma/PNET,neuroendocrine tumor,rhabdomyosarcoma. 

Molecular genetic studies can clarify the diagnosis in seemingly 

straightforward as well as in overtly problematic cases.These 

diagnostic distinctions are now critical as disease-specific and 

risk-directed therapies have emerged. 

follicular carcinoma in struma ovarii.A case report 


1)Department of pathology, Arnas civico hospital, Palermo, Palermo 



Background. Struma ovarii(SO)is an uncommon monodermal 

form of ovarian teratoma in which > 50% of the tumor is composed 

by thyroid tissue.Its malignant transformation is even 

rarer and is seen in only 5% of those cases.The most common 

thyroid-type malignancies to arise in SO are papillary and follicular 

carcinomas.We report a case of FTC minimally invasive 

in SO in a 37-year-old woman,who presented with ovarian right 

mass. 

Methods. The thyroid hormonal profile as well serum levels of 

CA125,CEA were regular at moment of diagnosis.At laparotomy 

a Ø 7 cm mass was seen with variegated appearance and intact 

surface.There was no ascites and no visible peritoneal seedlings. 

Monolateral salpingo-oophorectomy was performed.The total 

body TAC didn’t shows metastasis.19 months after the diagnosis 

the patient was free from disease. 

Results. The neoplastic cut surface appeared solid,translucid. 

Histopathology revealed near-complete transformation of 

ovarian tissue into mature thyroid tissue.Other ectodermal teratomatous 

elements were seen.In the thyroid tissue were some 

areas of cells arranged in a follicular pattern,showing mild 

nuclear pleomorphism,high mitotic index(> 5 mitoses xHPF).


There was focal capsular invasion and no vascular invasion. 

The tumor was positive for CK7,CKAE1,Ecadherin,TTF1,thy 

roglobulin, vimentin,EMA,BCL2,negative for CK20,ER,PGR, 

calretinin,cromogranin,TP53.These findings were consistent 

with a diagnosis of follicular carcinoma minimally invasive in 

SO. In these cases metastases are usually blood borne rather 

than to regional nodes and occur in ~5% of the minimally 

invasive tumors with blood vessels invasion,and developed in 

~1% of the tumors diagnosed as carcinoma only on the basis 

of minimal capsular invasion.Consequently in regard to the 

occurrence of thyroid–type carcinoma on SO,precise terminology 

should be used,and the diagnostic term”malignant SO”was 

avoided. 

Malignant solitary fibrous tumor of the pleura. 

A case report 





Background. Solitary fibrous tumor(SFT)is a rare spindle cell 

mesenchymal neoplasm of probable submesothelial derivation. 

The majority of these tumors have a benign course,the malignant 

form still remains enigmatic.We report a case of malignant 

pleuric SFT. 

Methods. A 44-year-old woman presented with dispnea,during 

the past months.A CT scan of the chest,following a chest-× 

rays,revealed a marginated,elliptical mass,of 17 cm Ø in the lower 

lobe of the left lung.A transegmetary resection of the inferior 

lobe was performed.Seven months after the diagnosis the patient 

was free from disease. 

Results. The tumor presents as a pedunculated mass lying within 

the pleural cavity, circumscribed,with the free surface bosselated. 

It measured 17 × 11 × 6,5 cm.The cut surface is firm,gray,with 

whorled appearance,foci of hemorrhage necrosis and softening. 

The neoplasm is characterized by hypo and hypercellular areas 

separated by fibrous stroma haemangiopericytoma-like.The hypercellular 

areas are composed of closely packed spindled or oval 

cells with moderate cellular atypia and high mitotic activity(> 4 

mitoses x10HPF)and scant intervening stroma.In the hypocellular 

areas spindle or oval cells are scattered among strands of 

collagen.The surface appears denuded of its mesothelial layer. 

The cells were positive for vimentin,CD34,Bcl-2,focally for 

desmin,negative for cytokeratin,calretinin,EMA,S100,CD57,N 

SE,CD99,NF,c-KIT,CEA,SMA and showed lower expression 

of progesterone receptors,high expression of p53.The tumor 

was determined to be a malign pleural SFT. The behaviour of 

these tumors is often unpredictable.Recent evidence suggests 

that this tumor derives from long-lived”fibroblastic”stem cell 

and successive mutations may lead to the malignant form. 

The complete resection is accepted as the most important 

single prognostic factor,but some parameters(tumor size,mitotic 

index,necrosis,hypercellularity,p53 expression)are related to 

outcome. 

epiregulin (ereG), amphiregulin (AreG) expression 

and BrAf v600e mutation as predictive biomarkers 

in metastatic colorectal cancer (MCrC) patients 

treated with cetuximab 

1)Valentini AM. 2)Cavallini A. 3)Lippolis C. 4)Campanella D. 

5)Pirrelli M. 6)Armentano R. 7)Caruso ML. 8)Campanella G. 

9)Lolli I. 

1)Anatomia Patologica, IRCCS “S. De Bellis”, Castellana Grotte, Italia 

2)Laboratorio Di Biochimica, IRCCS “S. De Bellis”, Castellana Grotte, 

Italia 3)Laboratorio Di Biochimica, IRCCS “S. De Bellis”, Castellana 

Grotte, Italia 4)Laboratorio Di Biochimica, IRCCS “S. De Bellis”, 

373 

Castellana Grotte, Italia 5)Anatomia Patlogica, IRCCS “S. De Bellis”, 

Castellana Grotte, Italia 6)Anatomia Patlogica, IRCCS “S. De Bellis”, 

Castellana Grotte, Italia 7)Anatomia Patologica, IRCCS “S. De Bellis”, 

Castellana Grotte, Italia 8)Servizio Di Oncologia, IRCCS “S De Bellis”, 

Castellana Grotte, 9)Servizio Di Oncologia, IRCCS “S De Bellis”, Castellana 

Grotte 

Background. The wild type (wt) KRAS patients respond to treatment 

with cetuximab in 20% only. 

To overcome this problem, other predictive biomarkers are 

needed. We have considered: BRAF mutation, as second factor 

after KRAS mutation in MAPK activation pathway, and EREG 

and AREG gene expression that may play an important role, as 

EGFR ligands, in CRC growth by autocrine stimulation. 

Methods. Ten wt KRAS mCRC patients treated with cetuximab 

were enrolled in this retrospective study. A standard cetuximab 

dosing regimen was used. Tumor response was evaluated by 

CEA serum level and imaging. Seven of ten patients were responders: 

3 partial responses, 4 stable diseases. Nucleic acids 

were extracted from formalin-fixed paraffin-embedded (FFPE) 

tissue sections by FFPE Qiagen kits. BRAF mutation was 

detected by PCR-RFLP method (Ampli-set-BRAF kit; Bird, 

Italy). AREG and EREG mRNA expression was measured by 

RT-qPCR. The gene expression was expressed as number of 

molecules/1 µg RNA. 

Results. All patients had not mutated BRAF gene. The EREG 

and AREG mRNA expression increased from non-responder to 

responder patients: 18.0 ± 2.5 × 10 2 vs 27.5 ± 2.8 × 10 2 mol./µg 

RNA for AREG and 7.2 ± 1.5 × 10 2 vs 11.7±1.9 × 10 2 mol./µg 

RNA for EREG. 

In conclusion, patients with no mutations in both KRAS and BRAF 

genes but with higher AREG and EREG gene expression seem to 

better respond to cetuximab treatment. This finding suggests that 

AREG an EREG gene expression could be used as predictive 

biomarkers for cetuximab therapy in mCRC. However, further 

studies with a higher number of patients needs for this purpose. 

Work was supported by Fondazione Cassa di Risparmio di Puglia, 

Italy. 

Breast fna cytology: an approach to thinprep 

slides using standardized cytological criteria 

1)Van Eeckhout P. 2)Arisio R. 

1)Anatomic pathology laboratory, CHR Mons–Warquignies, Mons, Belgium 

2)Anatomic pathology laboratory, Sant’Anna hospital, Torino, Italy 

Objectives. To evaluate the learning process of experienced 

breast cytopathologists when they pass to ThinPrep slides. To 

investigate if the criteria used for conventional breast cytology 

need to be adapted for liquid based cytology (LBC). 

Study design. We reviewed 234 breast FNA ThinPrep slides with 

histological correlations. The samples were blindly evaluated 

by two cytopathologists, (FF and PVE), with experience only 

in conventional breast cytology (CBC). After their independent 

evaluations the discordant diagnoses were discussed at the multihead 

microscope. Six months later, one of the two pathologists, 

(FF), blindly reviewed the same cases, after he had used LBC in 

cervico-vaginal cytology. At each round the slides were evaluated 

according to the probabilistic approach, and to the four criteria 

proposed by Wang HH and Ducatman BS (1998). Each of the 

four diagnostic criteria was quantitatively evaluated in order to 

allow assessment of its strength in LBC. 

Preliminary Results. Forty-three cases were excluded: cysts, 

abscesses, fat necrosis, cases with biopsies obtained in other institutions 

and C1 split samples with a representative CBC. Even 

after reviewing together their discordant diagnoses, the pathologists 

with no experience in LBC did not meet al.l of the current 

accuracy standards. However training with LBC resulted in acceptable 

results: Complete Sensitivity: 95.2%, Specificity 63.3%, 

False Positives 0%, False Negatives 4.8%. Absolute Sensitivity

374 

(C5) was 60%, because C4 cases, with both benign and malignant 

cells, were frequent. 

Comments. Specific training may be useful in LBC. Many cases 

showed some benign groups together with clearly malignant 

cells: we wonder if C5 diagnoses can be established even in 

such cases. False negative interpretations were 1 case: tubular 

carcinoma, 1 case: G1 and 1 case: G2 IDC, 1 case: malignant and 

1 case: borderline Phyllodes tumors; 1 case: ILC, 1 case: tubulolobular 

carcinoma. Five out of 13 fibrodenomas received C4 or 

C3 diagnoses. Future work: our cases with discordant histological 

correlations will be mixed with difficult cases that received variable 

interpretations. They will be blindly validated by FF and RA 

at the multihead microscope in order to allow a statistical assessment 

of the Wang and Ducatman criteria in LBC. 

references 

Feoli F, Paesmans M, Van Eeckhout P. Fine needle aspiration cytology 

of the breast. Impact of experience on accuracy, using standardized 

cytologic criteria. Acta Cytol 2008;52:145-51. 

Gornstein B, Jacobs T, Bédard Y, et al. Interobserver agreement of a 

probabilistic approach to reporting breast fine-needle aspirations on 

ThinPrep. Diagn Cytopathol 2004;30:389-95. 

ubcH10 expression on thyroid fine-needle 

aspirates. 

Varone V., Iaccarino A., Cozzolino I., Bellevicine C., Palombini 


Anatomia patologica, Policlinico università federico ii, Napoli, Italia 

Background. Thyroid fine-needle aspiration (FNA) samples 

belonging to the follicular neoplasm/suspicious for malignancy 

classes are controversial. We identified UbcH10 as a marker useful 

in the diagnosis of several neoplasms, including thyroid cancer. 

Here, analysis of UbcH10 expression by quantitative RT-PCR 

and immunohistochemistry was applied to FNAs. Methods. A 

series of 84 follicular neoplasm/suspicious for malignancy FNAs 

with histological follow-up (30 malignant) was prospectively 

collected. UbcH10 imunostaining was carried out on cell blocks 

and compared to that of the proliferation marker Ki-67. At the 

mRNA level, UbcH10 was compared with CCND2 and PCSK2 

expression, these latter being the most performing components of 

the previously reported 3-gene assay; to determine the diagnostic 

accuracy the area under the curve (AUC) of the receiver operating 

characteristic (ROC) curve for each gene individually and in 

combination was evaluated. Results. UbcH10 and Ki-67 shared a 

similar pattern; although UbcH10 expression was higher in malignant 

than in benign lesions (p < 0,001), staining was sporadic 

and the cut-off value derived by the ROC analysis was too low 

(1,25%) for routine application. Conversely, UbcH10 expression 

assessment by qRT-PCR was effective. UbcH10 mRNA levels 

associated to malignant histology were significantly higher than 

those associated to benign histology (p = 0.02). The AUC was 

0.74 for UbcH10, 0.81 for CCDN2, 0.62 for PCSK2 and 0,84 for 

UbcH10 and CCND2 combination. Conclusions. UbcH10 qRT- 

PCR analysis, rather than immunohistochemistry, is useful to 

increase the suspicion of malignancy in thyroid FNAs. UbcH10 

may be added as a panel component in qRT-PCR based assays. 

TNM staging of GISTs 

Vassallo L., Mastrogiulio M.G., Tacchini D., Di Mari N. 

Patologia umana ed oncologia, Azienda ospedaliera universitaria senese, 

Siena, Italia 

Background. Gastrointestinal stromal tumours (GIST) are the 

most common (80%) mesenchymal neoplasia of the gastrointestinal 

tract that have been only recently included in the TNM 

classification of malignant tumours. Reported incidence is 10-20 

cases per million per year. Approximately 85% of GISTs harbor 


activating mutation in KIT and most of the cases respond to 

kinase hinibitors. Nonetheless, response to terapy is not homogeneous. 

Accurate staging and evaluation of prognostic features are 

crucial to address properly the treatment. 

Objective. Thirty-three cases of GIST diagnosed in our Departement 

between 2001-2010 have been staged according to the seventh 

edition of UICC-TNM 2009-2010 and have been evaluated 

for prognostic factors according to Fletcher et al. (2002) 

Methods. Surgical speciments routinely processed and stained 

(HE). Morphological diagnosis confirmed by immunohistochemistry 

(CD117 MoAb with no Ag-unmasking). 

Results. Males were 18. Median age was 68 (range 26-90). The 

anatomical sites were: stomach 18/33(54,5%), small intestine 

12/33(36,4%) and other sites 3/33(9,1%). Morphological pattern 

was spindle cells 70%, epithelioid 26% and mixed 4%. According 

to Fletcher none was very low risk, 9/33 (27,3%) was low 

risk, 8/33 (24,2%) was intermediate risk and 16/33 (48,5%) was 

high risk. TNM stage was: stage I 14/33 (42,4%), stage II 4/33 

(12,1%), stage III 13/33 (39,4%) and stage IV 2/33 (6,1%). 

Conclusions. TNM staging and prognostic groups according to 

Fletcher show significant differences. TNM takes account of 

several factors as nodes, distant metastases and anatomical site 

incorporating most of the proposals of Miettinen et al. (2006). 

Ileal metastases from lung carcinoma: a case report 

1)Vassallo L. 2)Tacchini D. 3)Spina D. 4)De martino A. 

5)Malatesti R. 

1)Patologia Umana ed Oncologia, Policlinico S.Maria Alle Scotte, Siena, 

Italia 2)Patologia Umana ed Oncologia, Policlinico S.Maria Alle Scotte, 

Siena, Italia 3)Patologia Umana ed Oncologia, Policlinico S.Maria Alle 

Scotte, Siena, Italia 4)U.O.C. Chirurgia Generale 3, Policlinico S.Maria 

Alle Scotte, Siena, Italia 5)U.O.C. Chirurgia Generale 3, Policlinico 

S.Maria Alle Scotte, Siena, Italia 

Background. Intestinal metastases from lung cancer have been 

rarely reported in literature and usually occurr in patients with 

terminal stage disease. They are usually asymptomatic but may 

present as perforation, obstruction and bleeding. 

Case report. We describe a case of 68 year old man who underwent 

right pneumonectomy 15 month prior to the diagnosis for a 

double lung carcinoma: one diagnosed as combined carcinoma, 

consisting of anaplastic large cell carcinoma (70%), tubulo-acinar 

adenocarcinoma (20%) and squamous carcinoma (10%), and the 

other as acinar adenocarcinoma. 

The abdominal CT, performed during the follow-up controls, 

revealed a mass in the terminal ileum. Resected specimen of 

the ileum, 27 cm in lenght, was recived. The segment showed a 

single transmural ulceronodular lesion of 9 × 7 cm with mesenteric 

extension. 

Microscopically, the mass showed an alveolar pattern and consisted 

of large poligonal cells with vescicular nuclei, prominent 

nucleoli and abundant cytoplasm. Immunohistochemical studies 

revealed positivity for cytocheratin AE1-AE3, negativity for 

CK7, CK 20, TTF1, p63, as well as negativity for neuroendocrine 

markers (cromogranin, synaptophisin, CD56). 

The neoplasia was morphologically and immunohistochemically 

similar to the anaplastic large cell component of the lung 

carcinoma. 

Post radio-chemotherapy residual of uterine 

cervical carcinoma on surgical specimens: 

differences in overall survival. A preliminary report 

VG Vellone, G. Amodio*, F. Morassi, B. Angrisani, G. Scambia*, 

G. Rindi, V. Masciullo*, GF Zannoni 

Istituto di Anatomia e Istologia Patologica. Pol. A Gemelli. Università 

Cattolica del Sacro Cuore. Roma *Istituto di Ginecologia ed Ostetricia. 

Pol. A Gemelli. Università Cattolica del Sacro Cuore. Roma


Background. Radio-chemotherapy followed by radical hysterectomy 

with lymphadenectomy is a well established therapeutic option 

for advanced cervical carcinoma. Pathological examination 

of the surgical specimens allows an accurate assessment of the 

actual response to radiochemotherapy. Impact of local residual 

(pR) and pathological restaging (ypTNM and FIGO) on overall 

survival is investigated 

Methods. Local response to therapy were classificated as follow: 

pR0: Pathological Complete Response; pR1: Pathological Partial 

Response; pR2: Pathological No Response. All patients were 

restaged according to ypTNM and FIGO staging; FIGO 0 were 

considered as No Residual Disease (NRD), FIGO I-II as Local 

Residual Disease (LRD), FIGO III-IV as Metastastic Disease 

(MD). 72 patients were enrolled in the present study. 

Results. 29 patients (40,3%) achieved pR0; 18 patients (25,0%) 

achieved pR1; 25 patients (34,7%) achieved pR2. Kaplan-Meyer 

survival curves showed a significant worse survival for pR2 

compared to both pR0 (p = 0,003) and pR1 (p = 0,045). No 

significant difference was observed between pR1 and pR0. 28 

patients (38,9%) resulted NRD; 30 patients (41,7%) resulted 

LRD; 14 patients (19,4%) resulted MD. Kaplan-Meyer survival 

curves showed a significant worse survival for MD compared to 

both NRD (p = 0,0003) and LRD (p = 0,0034). LRD showed a 

significant worse survival compared to NRD (p = 0,0349). 

These preliminary data suggest an important prognostic role 

of pathological evaluation of residual cancer both locally and 

systemic with patients with no residual cells having the best 

results. 

Metastases to uterine cervix. A rare desaese and 

potential diagnostic pitfalls 

V.G. Vellone, G. Petrone, L. Santoro, S. Moncelsi, E.D. Rossi, 

G Fadda, G.F. Zannoni 


Cattolica del Sacro Cuore. Roma 

Background. Metastasis to uterine cervix are uncommon. Literature 

contains references only to small numbers of cases, many 

of these are old, poorly documented and often without the aid of 

immunohistochemistry. This single–center study investigates the 

frequency of this diseaase and highlights the pitfalls in differential 

diagnosis with primary tumors. 

Methods. The computerized archive of the Department of Pathology 

of Policlinico Gemelli, Rome, was reviewed for the period 

1999-2009. A total of 1140 uterine cervical malignance was 

found. All the cases of uterine cervix cancer were critically revised. 

We selected 95 cases of metastatic involvement of uterine 

cervix. 

Results. Metastatases to cervix the represented the 8.3% of all 

cervical malignancies. Mean age was 60 years, older than patients 

with primary tumor (p < 0,0001). The primary site were: endometrium 

(67%), ovary (14%), myometrium (4%), large bowel (4%), 

breast (3%), stomach (2%) and cervical involvement of lymphoma 

(6%). The main diagnostic pitfall was between metastatic 

endometroid adenocarcinoma and intestinal adenocarcinoma with 

primary cervical adenocarcinoma, expecially in small biopsies. 

The morphological feature are similar but the recognition of the 

pre-cancerous lesions and a proper immunoistochemistry panel 

(Vimentin, CK7, CK20, CDX2, ER, PR and p16) are diagnostic. 

In a small number of case can be useful the research of HPV. 

Metastasis from mammary glands, in particular with lobular 

histotype, must be distinguished from lymphoma, clinical history 

and the immunochemistry can distinguish the two lesions. 

The extragenitalia tumors usually have multiple localization 

other than cervix. The genitalia tumors (endometrial carcinoma, 

ovarian carcinoma and uterine leyomiosarcoma and low grade 

stromal sarcoma) may show the cervix as the only localization of 

a metastatic disease. 

fluorescence microscopy of Pap-stained cervical 

cytology specimens 

Ventura L., Dal mas A. 

Anatomia patologica, San Salvatore, L’Aquila, Italia 

375 

Background. Fluorescence microscopy (FM) has been used to 

study routinely stained histologic and cytologic slides for many 

years. The method is based on autofluorescence and fluorescence 

induced by dyes (mainly eosin) and was successfully applied to 

demostrate various structures (elastic fibres, microorganisms, 

metaplastic cells) in different organs but, to the best of our knowledge, 

no one ever used it in cervical cytology. 

Methods. Selected cases of Papanicolaou-stained slides from 

cervical conventional smears (CS) or liquid-based preparations 

(LBP) were retrieved from the archive, including representative 

examples of non-neoplastic findings and cell abnormalities. The 

original slides were reviewed with FM to evaluate the fluorescence 

pattern of each single finding. 

Results. Fungal organisms appeared strongly fluorescent and 

readily visible in both pseudohyphae and yeast forms. Delicate, 

green fluorescence was observed in single filaments within 

clusters of Actinomyces, whereas weak/absent fluorescence was 

identified in lactobacilli and coccobacilli. Trichomonas vaginalis 

was recognized by fluorescence of cytoplasmic eosinophilic 

granules, while associated Leptothrix showed a pale signal. 

Superficial and keratotic cells displayed strong cytoplasmic 

fluorescence. Intermediate, parabasal and inflammatory cells 

showed low/absent cytoplasmic fluorescence. Nuclei were invariably 

invisible. No significant remark could be done about cell 

abnormalities, including ASC, HPV effect and SIL. Additional 

findings enclosed strong fluorescence of starch granules and hair 

fragments contaminating the smears. 

LBP allowed a better visualization of fluorescent elements. 

Conclusions. FM evaluation of Pap-stained cervical cytology 

specimens is very easy to perform, rapid and unexpensive. Such 

method can be useful in routine diagnosis of infectious conditions, 

expecially for fungal infections. Further studies may be 

of help in determining the real value of this technique in daily 

practice. 

The renal stone of Pandolfo III Malatesta (1370-1427), 

lord of fano (Marche, Central Italy) 

1)Ventura L. 2)Giuffra V. 3)Lunardini A. 4)Minozzi S. 5)Quaresima 

R. 6)Arrizza L. 7)Fornaciari G. 

1)Anatomia patologica, San salvatore, L’Aquila, Italia 2)Divisione di paleopatologia, 

Università, Pisa, Italia 3)Divisione di paleopatologia, Università, 

Pisa, Italia 4)Divisione di paleopatologia, Università, Pisa, Italia 

5)Chimica, ingegneria chimica e materiali, Università, L’Aquila, Italia 

6)Centro di microscopia elettronica, Università, L’Aquila, Italia 7)Divisione 

di paleopatologia, Università, Pisa, Italia 

Background. The natural mummy of Pandolfo III Malatesta 

(1370-1427), prince of Fano and leading figure of the Italian 

Renaissance, was exhumed from his monumental tomb in Fano 

in 1995. Previous studies revealed the typical ergonomic picture 

of a horseman and a soldier, as well as the presence of prostatic 

nodular hyperplasia. We present the extensive study of a large 

staghorn calculus of the left kidney. 

Methods. The specimen surface was examined by binocular stereoscopic 

microscopy (BSM) and scanning electron microscopy 

(SEM). Multiple fragments from the surface and inner portions of 

the calculus were submitted to X-Ray diffraction (XRD) analysis 

and scanning electron microscopy for elemental distibution 

analysis (SEM-EDAX). 

Results. The stone surface, mulberry similar, was honey brown 

in color and consisted of aggregated large crystals, evident at 

BSM and SEM level and suggesting a calcium oxalate dihydrate 

stone. XRD analysis demonstrated that the calculus was mainly

376 

composed of ammonium acid urate and calcium oxalate dihydrate 

(weddellite). Along with the organic constituents (C, O, N), 

SEM-EDAX detected the following chemical elements: K in the 

core; K, S, Si, Cl, Ca, P, Na and Ba in the surface. 

Conclusions. The chemical composition of the stone, as demonstrated 

by XRD analysis, supported the hypothesis of high animal 

protein and sugar intake by the subject. Moreover, the presence of 

ammonium acid urate, infrequently found in kidney stones, may 

indicate recurrent urinary tract infections. Most of the elements 

detected are usual constituents of renal stones. Among unusual 

elements found, the presence of silica and barium may be related 

to contamination as it is rarely found in urinary calculi. 

The value of biopsy laterality in association 

with PSA and gleason score for the identification 

of subjects at high risk of recurrence in prostate 

cancer 

1)Ventura L. 2)Gravina GL. 3)Festuccia C. 4)Marampon F. 

5)Fileni A. 6)Di clemente L. 7)Bonfili P. 8)Di staso M. 9)Fardella 

C. 10)Tombolini V. 

1)Anatomia patologica, San salvatore, L’Aquila, Italia 2)Radioterapia, 

San salvatore, L’Aquila, Italia 3)Radioterapia, San salvatore, L’Aquila, 

Italia 4)Radioterapia, San salvatore, L’Aquila, Italia 5)Urologia, San salvatore, 

L’Aquila, Italia 6)Urologia, San salvatore, L’Aquila, Italia 7)Radioterapia, 

San salvatore, L’Aquila, Italia 8)Radioterapia, San salvatore, 

L’Aquila, Italia 9)Radioterapia, San salvatore, L’Aquila, Italia 10)Radioterapia, 

San salvatore, L’Aquila, Italia 

Background. predicting patients with prostate cancer (Pca) at 

high risk of recurrence (HRR) is a major challenge for clinicians. 

Clinical T-stage poorly predicts the pathological stage and 

understaging occurs in up to 60% of cases. Here we determine if 

needle biopsy parameters improve the value of NCCN criteria for 

predicting men at HRR. 

Methods. A retrospective survey of 488 men who underwent 

RP was undertaken. Univariate and multivariate logistic regression 

with receiver operating characteristic (ROC) curves were 

generated to test which parameters were able to best individuate 

men at HRR when histopathologic findings were used as the 

reference standard. The parameters were: PSA, biopsy laterality, 

total number of positive biopsy cores, clinical T stage, and 

Gleason score. The combination of best predictors then was 

compared with the standard NCCN criteria in terms of ability to 

predict HRR. 

Results. At univariate analysis all clinical parameters [biopsy 

laterality (OR=2.389; 95%CI 1.49 to 3.82; p < 0.0001); Gleason 

score (OR = 1.678; 95%CI 1.37 to 2.046; p < 0.0001), total 

number of positive biopsy cores (OR = 1.488; 95%CI 1.27 to 

1.74; p < 0.0001) and PSA (OR = 1.329; 95%CI 1.26 to 1.53; 

p < 0.0001)] except the clinical T-stage (OR = 1.136; 95%CI 

0.86 to 1.49; p = 0.343) significantly predicted men at HRR. At 

multivariate analysis only biopsy laterality (OR = 2.453; 95%CI 

1.07 to 5.61; p = 0.033), Gleason score (OR = 1.847; 95%CI 

1.38 to 2.46; p < 0.0001) and PSA (OR = 1.490; 95%CI 1.29 to 

1.71; p < 0.0001) were predictors of HRR. The association of 

PSA, Gleason score and biopsy laterality achieved a significant 

larger AUC (AUC = 0.835; 95%CI 0.791 to 0.873; p < 0.0001) 

than the association of standard parameters used in the NCCN 

criteria (clinical T-stage, PSA and Gleason score) (AUC = 0.685; 

95%CI 0.630 to 0.736; p < 0.0001) in the prediction of HRR. So 

the biopsy laterality as replacement of clinical T stage contributes 

significantly to improve the value of NCCN criteria for predicting 

subjects at HRR. 


fluorescence microscopy of hematoxylin-eosin 

slides for the identification of gastric Helicobacter 

pylori infection 

1)Ventura L. 2)Rossi M. 

1)Anatomia patologica, San salvatore, L’Aquila, Italia 2)Chirurgia e diagnostica 

endoscopica, San salvatore, L’Aquila, Italia 

Background. Fluorescence microscopy (FM) has been used to 

study routine histological slides for many years. This technique 

is based on the autofluorescence of some structures and the fluorescence 

induced by eosin and other dyes. The method has been 

successfully applied to demostrate various structures (elastic 

fibres, microorganisms, metaplastic cells) in different organs 

but, to the best of our knowledge, no one ever used it to identify 

Helicobacter pylori (Hp) in gastric biopsies. 

Methods. Routine gastric endoscopic biopsies from 50 consecutive 

and unselected patients were observed in this preliminary, 

prospective study. Beside conventional microscopic examination, 

hematoxylin-eosin (H-E) stained slides were also observed with 

an epifluorescence microscope. Conventional microscopic examination 

of additional slides stained with Giemsa was employed 

as golden standard. 

Results. At FM observation Hp appeared fluorescent and distinctly 

visible at 400x magnification. Among the 50 cases tested, 

13 were positive for Hp in H-E slides, 16 resulted positive at 

FM observation of H-E slides and 17 were positive in Giemsa 

slides. When compared to Giemsa, the sensitivity and specificity 

of the FM method for the detection of Hp were 94% and 100%, 

respectively. FM observation of H-E sildes also improved the 

recognition of intestinal metaplasia, red blood cells and chief 

cells cytoplasmic granules. 

Conclusions. FM evaluation of H-E slides is very easy to perform, 

rapid and unexpensive. Our preliminary study reveals that 

it is a highly sensitive and specific method and therefore may represent 

a good alternative to histochemical stains for detecting Hp. 

Unfortunately, coccoid forms of Hp could not be distinguished 

from granules and other microorganisms that may be present 

on the luminal surface. Further advantages of this method are 

represented by an improved recognition of intestinal metaplasia 

and the possibility to perform retrospective studies on archival 

material. 

Diagnosis of systemic amyloidosis: 

role of minor salivary gland biopsy 

1)Verga L. 2)Foli A. 3)Morbini P. 4)Palladini G. 5)Caporali R. 

6)Obici L. 7)Russo P. 8)Lanzarini P. 9)Merlini G. 10)Paulli M. 

1)Anatomia Patologica E Centro Amiloidosi, Fondazione IRCCS Policlinico 

San Matteo, Pavia, Italia 2)Centro Amiloidosi, Fondazione IRCCS 

Policlinico San Matteo, Pavia, Italia 3)Anatomia Patologica, Fondazione 

IRCCS Policlinico San Matteo, Pavia, Italia 4)Centro Amiloidosi, Fondazione 

IRCCS Policlinico San Matteo, Pavia, Italia 5)U.O. Reumatologia, 

Fondazione IRCCS Policlinico San Matteo, Pavia, Italia 6)Centro Amiloidosi, 

Fondazione IRCCS Policlinico San Matteo, Pavia, Italia 7)Centro 

Amiloidosi, Fondazione IRCCS Policlinico San Matteo, Pavia, Italia 8)U. 

O. Malattie Infettive, Fondazione IRCCS Policlinico San Matteo, Pavia, 

Italia 9)Centro Amiloidosi, Fondazione IRCCS Policlinico San Matteo, 

Pavia, Italia 10)Anatomia Patologica, Fondazione IRCCS Policlinico San 

Matteo, Pavia, Italia 

Background. The diagnosis of amyloidosis requires histological 

demonstration of amyloid deposits and characterization of 

amyloid protein. The risk of bleeding risk and ready accessibility 

of alternative sites discourages organ biopsy in patients with suspected 

amyloidosis. Fine-needle aspiration of abdominal fat and 

salivary gland biopsy represent valid alternatives to organ biopsy. 

We reported that in AL amyloidosis the sensitivity of abdominal 

fat is 87%.


Methods. Here we report the results of a sequential diagnostic 

approach, with salivary gland biopsy as a second step in 62 consecutive 

patients referred to Policlinico San Matteo between 2002 

and 2007 for suspected systemic amyloidosis, in whom amyloid 

deposit were not detected in the abdominal fat aspirates. Seventy-four 

percent of patients had > 1 sign or symptom suggesting 

systemic amyloidosis. 

Results. Light microscopy examination of salivary gland biopsies 

detected amyloid deposits in 7 patients (11%). In all these samples 

immune-electron microscopy was performed and allowed to 

characterize the amyloid protein: AL λ deposits were detected in 

4 cases (57%), AL κ in 2 (29%) and AA in 1 (14%). In the remaining 

patients amyloidosis was eventually diagnosed in 5 additional 

subjects (8%) based on renal (2 AL λ) and cardiac (3 AL κ) 

biopsies. In all the patients hereditary amyloidosis was excluded 

by DNA analysis, in the AL patients a monoclonal component 

of the same type of that observed in the deposits was detected in 

serum and/or urine, the AA patient had persistently elevated SAA 

concentration, but the underlying cause of inflammation remained 

undetermined. At the end of two years follow-up, the diagnosis of 

amyloidosis was excluded in the remaining 50 patients. Overall, 

the diagnostic sensitivity of the salivary gland biopsy in patients 

with negative fat aspirate was 58%, the specificity was 100% and 

the negative predictive value was 91%. 

A sequential diagnostic approach based on second step salivary 

gland biopsy after negative abdominal fat aspirate can spare the 

biopsy of the organ involved to more than half the patients with 

systemic amyloidosis. 

Immuno-electron microscopy: 

diagnostic performance of analysis 

of abdominal fat in systemic amyloidoses 

1)Verga L. 2)Palladini G. 3)Morbini P. 4)Sarais G. 5)Foli A. 

6)Russo P. 7)Zenone bragotti L. 8)Lanzarini P. 9)Merlini G. 

10)Paulli M. 

1)Anatomia Patologica E Centro Amiloidosi, Fondazione IRCCS Policlinico 

San Matteo, Pavia, Italia 2)Centro Amiloidosi, Fondazione IRCCS 

Policlinico San Matteo, Pavia, Italia 3)Anatomia Patologica, Fondazione 

IRCCS Policlinico San Matteo, Pavia, Italia 4)Centro Amiloidosi, Fondazione 

IRCCS Policlinico San Matteo, Pavia, Italia 5)Centro Amiloidosi, 

Fondazione IRCCS Policlinico San Matteo, Pavia, Italia 6)Centro Amiloidosi, 

Fondazione IRCCS Policlinico San Matteo, Pavia, Italia 7)Centro 

Amiloidosi, Fondazione IRCCS Policlinico San Matteo, Pavia, Italia 8)U. 

O. Malattie Infettive, Fondazione IRCCS Policlinico San Matteo, Pavia, 

Italia 9)Centro Amiloidosi, Fondazione IRCCS Policlinico San Matteo, 

Pavia, Italia 10)Anatomia Patologica, Fondazione IRCCS Policlinico San 

Matteo, Pavia, Italia 

Background. The diagnosis of systemic amyloidosis rely on 

identification and characterization of amyloid deposits in tissues; 

abdominal fat aspirate (AFA) is a convenient alternative to organ 

biopsy. 

Methods. Here we report the diagnostic performance of IEM on 

AFA in 597 consecutive patients referred to Policlinico San Matteo 

between 2003 and 2008 for suspected systemic amyloidosis. 

All the patients were followed for at least 18 months and then the 

diagnosis of amyloidosis was definitely established or rejected. 

The AFA were stained with Congo red and IEM study was 

performed as previously described (Arbustini, Amyloid 2002). 

Mutations for hereditary amyloidosis were searched by DNA 

analysis. Organ involvement was defined according to the 2005 

International Society of Amyloidosis criteria. 

Results. A diagnosis of systemic amyloidosis was eventually 

established in 334 cases (56%). At light microscopy AFA was 

positive in 495 patients with 83% sensitivity (Se), 77% specificity 

(Sp), 81% positive predictive value (PPV) and 79% negative 

predictive value (NPV). At IEM amyloid fibrils were detected in 

459 patients, with 77% Se, 100% Sp, 100% PPV and 80% NPV. 

Characterization by IEM was possible in all the positive AFA 

377 

samples and was AL λ in 230 patients (50%), AL κ in 161 (27%), 

AA in 119 (20%), ATTR in 18 (3%). Characterization was confirmed 

by the clinical picture, follow-up and DNA analysis in 

all cases. In 90 patients with negative AFA the diagnosis was 

established on organ or minor salivary gland biopsies and characterized 

as AL λ in 39 patients (43%), AL κ in 17 (19%), AA 

in 17 (19%), ATTR in 15 (17%), AFib in 2 (2%), by IEM, light 

microscopy immunohistochemistry or DNA analysis. Immunoelectron 

microscopy increases the specificity of light microscopy 

examination of AFA and can correctly characterize the amyloid 

deposits in all cases. 

endoscopic ultrasound-guided fine needle 

aspiration (euS-fNA) in lymph nodal and 

mediastinal lesions: a multicenter experience 

1)Vetrani A. 2)Zeppa P. 3)Barra E. 4)Napolitano V. 5)Cozzolino 

I. 6)Malapelle U. 7)Troncone G. 8)Palombini L. 



Napoli “Federico II”, Napoli, Italia 3)Chirurgia generale e specialistica, 

Azienda ospedaliera monaldi, Napoli, Italia 4)Chirurgia generale e specialistica, 

Azienda ospedaliera monaldi, Napoli, Italia 5)Scienze biomorfologiche 





Università di Napoli “Federico II”, Napoli, Italia 

Background. Endoscopic ultrasound-guided fine needle aspiration 

(EUS-FNA) is an established procedure in lung cancer (LC) 

staging and in the diagnosis of mediastinal masses. Most of the 

experiences reported refer to single specialized centers where dedicated 

teams of endoscopists and pathologists perform the procedure. 

We report the EUS-FNA experience of a cooperation group 

involving clinicians and cytopathologists from three hospitals. 

Methods. Fifthy-seven consecutive EUS-FNA of mediastinal 

nodes in LC patients, 8 mediastinal and 2 sub-diaphragmatic masses 

were collected in three years. EUS-FNA was performed by two 

endoscopists and three experienced pathologists; on-site evaluation 

was performed in all cases by the three cytopathologists. Lymph 

node negative cases underwent surgery, which confirmed the cytological 

diagnoses but also detected two false negatives. Four of 

the 10 EUS diagnoses of mediastinal masses were histologically 

confirmed. All EUS diagnoses were blindly reviewed by three pathologists 

to assess intra and interpersonal reproducibility. 

Results. FNA-EUS diagnoses were: 10 inadequate (17%), 10 

negative (17%), 4 suspicious (7%) and 33 positive (59%). Diagnoses 

of mediastinal and sub-diaphragmatic masses were: relapse 

of LC (3), mesenchimal tumour NOS (3), gastrointestinal stromal 

tumor (GIST) (1), esophageal carcinoma (2) and paraganglioma 

(1). Attained sensitivity and specificity were 85% and 100% with 

an high interpersonal diagnostic reproducibility (p < 0.5). 

Conclusion. The sensitivity and specificity attained were similar 

to those reported in the literature suggesting that experienced 

cytopathologists and endoscopists from different Institutions can 

employ the same procedure reaching comparable results. 

Subcutaneous CD34+ spindle cell tumours: 

a continuous spectrum from spindle cell lipoma 

to myofibroblastoma 

1)A. Gurrera, 2)L. Villari, 3)G. Bruno, 2)G.M. Bruno, 1)P. 

Amico, 1)G.M. Vecchio, 1)G. Magro 



Catania, Italia; 2)Servizio Anatomia Patologica, Azienda Ospedaliero- 

Universitaria Policlinico-Vittorio Emanuele, Catania, Italia; 3)Divisione 

di Ortopedia, Azienda Ospedaliero-Universitaria Policlinico Vittorio 

Emanuele, Catania, Italia

378 

Background. Spindle cell lipoma (SCL) is a benign tumour 

composed of a variable admixture of CD34 + spindle cells and mature 

adipocytes, that frequently occurs in superficial soft tissues. 

Myofibroblastoma is a spindle cell tumour that characteristically 

expresses, in addition to CD34, myogenic markers, including desmin 

and α-smooth muscle actin. This tumour, morphologically 

reminiscent of SCL, is characteristically detected in the breast, 

even if a few cases have also been reported in soft tissues. Notably, 

some cases of SCL may be composed exclusively of spindle 

cells without an associated fatty component (fat-free SCL) and 

they are morphologically indistinguishable from MFB. Additionally, 

rare cases (< 2%) of SCL may express desmin. 

Materials. We discuss the morphological and immunophenotypical 

features of two cases of SCL with focal expression of desmin, 

one case of fat-free SCL and one case of soft tissue MFB to support 

the hypothesis that these tumors belong to the same category 

of neoplasms. 

Results. Among a large series of SCLs, we identified: i) two 

cases that, apart CD34, CD10, bcl-2 immunostaining, showed focal 

expression of desmin; ii) one case of SCL, completely devoid 

of fatty component (fat-free SCL), showing CD34, CD10 and bcl- 

2 immunoreactivity. In addition, a rare case of soft tissue MFB 

occurring in the wrist of a 36-year old woman was included. This 

tumour was positive for desmin, α-smooth muscle actin, CD34, 

CD10 and bcl-2. Interestingly, the fat-free SCL and MFB shared 

several morphological features, including CD34 + /CD10 +/ /bcl2 + 

bland-looking spindle cells, focally arranged in short bundles, and 

intervening thick collagen bands. They could be distinguished in 

that the latter expressed both desmin and α-smooth muscle actin. 

We propose a unifying histogenetic concept for SCL and MFB 

of soft tissues, suggesting that they be regarded as a continuous 

morphological and immunophenotypical spectrum, likely arising 

from a common CD34 + precursor mesenchymal cell with the capability 

of an exclusive or predominant fibroblastic (classic SCL 

or SCL with desmin expression, respectively) vs myofibroblastic 

(MFB) differentiation. The recent findings, showing that soft tissue 

MFB and SCL display the same chromosomal aberrations, 

namely the loss of RB/13q14 and FKHR/13q14 loci, seem to 

support our hypothesis. 

Her2 alterations in brenner tumors 

1)Viola P. 2)Felicioni L. 3)Malatesta S. 4)Del grammastro M. 

5)Sciarrotta M. 6)Pullara C. 7)Gadducci A. 8)Liberati M. 9)Marchetti 

A. 10)Buttitta F. 

1)Oncologia E Medicina Sperimentale, Università “G. D’Annunzio”, 

Chieti, Italia 2)Oncologia E Medicina Sperimentale, Università “G. D’Annunzio”, 

Chieti, Italia 3)Oncologia E Medicina Sperimentale, Università 

“G. D’Annunzio”, Chieti, Italia 4)Oncologia E Medicina Sperimentale, 

Università “G. D’Annunzio”, Chieti, Italia 5)Oncologia E Medicina 

Sperimentale, Università “G. D’Annunzio”, Chieti, Italia 6)Oncologia E 

Medicina Sperimentale, Università “G. D’Annunzio”, Chieti, Italia 7)Medicina 

Della Procreazione, Università Di Pisa, Pisa, Italia 8)Ostetricia 

E Ginecologia, Università “G. D’Annunzio”, Chieti, Italia 9)Oncologia 

E Medicina Sperimentale, Università “G. D’Annunzio”, Chieti, Italia 

10)Oncologia E Medicina Sperimentale, Università “G. D’Annunzio”, 


Background. Transitional cell tumors of the ovary represent 1-2% 

of all ovarian cancers and include two distinct clinicopathologic 

entities: Brenner tumors (BT) (benign, borderline and malignant) 

and transitional cell carcinomas. Brenner tumors are thought to 

derive from the surface epithelium and underlying stroma through 

transitional cell metaplasia. Recently an immunohistochemical 

study for epidermal growth factor receptor (EGFR), Ras, Cyclin 

D1, p16, Rb, and p53, as well as mutational analysis of K-Ras, 

B-Raf, CTNNB1, PIK3CA, and p53 genes have been conducted, 

but no data are available on HER2 mutations. 

Methods. We analyzed HER2 mutations in 110 ovarian tumors 

(70 serous, 19 endometrioid, 10 mucinous, 3 clear cell, 3 BT, 3 


undifferentiated and 2 mixed mullerian tumors) from as many 

patients surgically treated at the Department of Gynecology, 

University of Pisa. The analysis was performed by Single Strand 

Conformation Polymorphism (SSCP) analysis followed by direct 

sequencing of the PCR products. 

Results. One (0.9%) of the 110 cases examined showed a double 

mutation of HER2. The mutations were both located at exon 20: 

T2413C (Cys804Arg) and A2437G (Asn813Asp). From a pathological 

point of view, the mutated tumor was a BT monolateral, 

non metastatic, grade 1, stage IA. We therefore decided to investigate 

additional cases of ovarian BT for HER2 mutations. To this 

aim, we selected 8 patients affected by BT surgically treated at 

the Department of Gynecology, University of Chieti, from January 

2000 till January 2010. The mutational study of these cases, 

as well as additional analyses, including HER2 immunohistochemical 

staining and Fluorescent In Situ Hybridization (FISH) 

are in progress. 

Dedifferentiated endometrioid adenocarcinoma. 

Clinicopathologic study of a case 

1)Vita G. 2)Borgia L. 3)Di Giovannantonio L. 4)Bisceglia M. 

1)Department of Pathology, IRCCS Institute of Cancer, Rionero in Vulture, 

Italy 2)Department of Pathology, IRCCS Institute of Cancer, Rionero 

in Vulture, Italy 3)Department of Pathology, IRCCS Institute of Cancer, 

Rionero in Vulture, Italy 4)Department of Pathology, IRCCS Casa Sollievo 

della Sofferenza Hospital, San Giovanni Rotondo, Italy 

Background. Dedifferentiation, which is microscopically defined 

as the presence of high grade areas abruptly emerging 

from low-grade tumors, was firstly described in bone and soft 

tissue tumors, such as chondrosarcoma, giant cell tumor, parosteal 

osteosarcoma, central intraosseous well-differentiated 

osteosarcoma, well-differentiated liposarcoma, and chordoma. 

Subsequently, this phenomenon has also been described in epithelial 

malignancies, mainly of salivary glands (epimyoepithelial 

carcinoma, low grade polymorphous carcinoma, and acinic cell 

carcinoma), but also in the thyroid (follicular and papillary carcinoma), 

and kidney (clear cell renal cell carcinoma, chromophobe 

renal cell carcinoma). In 2006 a previously unrecognized association 

of undifferentiated carcinoma with low grade endometrioid 

adenocarcinoma of the uterus and ovary, was described for the 

first time 1 . Dedifferentiation confers more aggressive malignant 

behaviour than would be otherwise shown by the original tumor 

if present alone. Only 2 papers have appeared in the literature so 

far on the topic of dedifferentiated endometrioid carcinoma, both 

from the same institution 1 2 . 

Objectives. To report a cases of dedifferentiated endometrial 

endometrioid carcinoma. 

Case Report. A 45-year old lady was admitted for abdominal 

pain and signs of intestinal subocclusion. Physical examination 

and imaging studies demonstrated the presence in the right 

ovary of a solid mass measuring 10 cm in its greatest diameter. 

Another tumor mass was also noticed in the uterus. The patient 

underwent surgical intervention and the ovarian mass was sent 

for intraoperative consultation. On frozen section an undifferentiated 

ovarian malignant tumor, unspecified, was diagnosed. 

Total hysterectomy and bilateral adnexectomy was performed 

and the surgical specimen was sent for pathological examination. 

On sectioning the uterine corpus was involved by an 8 cm vegetating 

tumor, extending from the fundus to the cervix, deeply 

infiltrating the myometrium with extension into the parametria. 

On permanent sections, morphologically, the right ovarian tumor 

was confirmed as an undifferentiated malignant neoplasm with 

foci of necrosis and hemorrhages, also infiltrating the ipsilateral 

mesosalpynx. The tumor in the uterus showed both large areas 

of a low grade FIGO endometrioid carcinoma (60% of the entire 

tumor mass) and a solid tumor component (40%), comprised of 

sheets of malignant large round to polygonal cells, focally ex-


hibiting rhabdoid features, without any specific growth pattern. 

The left ovary was the site of a small microscopic focus not seen 

grossly (< 5 mm) of low grade FIGO endometrioid carcinoma 

– likely a second primary. Immunohistochemically the solid 

tumor component was diffusely positive for vimentin with a few 

scattered tumor cells (< 5%) found only in some of the blocks 

positive for cytokeratins (cocktail) and EMA, and negative for 

hematolymphoid markers, melanoma markers, smooth and skeletal 

muscle markers, neuroendocrine markers, and CD34. Taking 

all evidence into account the tumor was diagnosed as “endometrial 

dedifferentiated endometrioid carcinoma”, metastatic to 

the right ovary, likely coexisting with low grade endometrioid 

adenocarcinoma in the left ovary (FIGO stage IIIA). Follow-up. 

This is a recent case and the patient is currently being given 

courses of chemotherapy, including cisplatinum, anthracycline, 

and taxanes. 

Discussion. FIGO grade 1 and FIGO grade 2 endometrioid adenocarcinomas 

are low grade tumors with excellent prognosis, 

FIGO grade 3 endometrioid adenocarcinoma has intermediate 

prognosis, and undifferentiated endometrial carcinoma is high 

grade with poor prognosis. Undifferentiated carcinoma is characterized 

by a proliferation of monotonous, highly atypical, epithelial 

cells, at times with rhabdoid features, growing in solid sheets 

with no specific pattern 2 3 . FIGO grade 3 endometrioid carcinoma 

always exhibits a solid growth pattern with (focal) glandular 

differentiation. 3 Some tumors are of mixed type, endometrioid 

adenocarcinoma FIGO grade 1 or 2 associated with undifferentiated 

carcinoma: for these tumors the diagnosis of dedifferentiated 

(endometrial or ovarian) carcinoma is warranted 1 3 . The biologic 

behaviour in mixed tumors is determined by the undifferentiated 

component. However, the undifferentiated component of these 

tumors can be misdiagnosed as the solid component of FIGO 

grade 3 in a pure endometrioid carcinoma. Patients with undifferentiated 

carcinoma often (> 50%) present with advanced stage 

disease and > 60% die of disease within 5 years after diagnosis 2 . 

In comparison 30% of patients with high FIGO grade (grade 3) 

present with advanced stage disease and around 35% die within 

5 years after diagnosis 2 . In consequence of that the recognition 

of an undifferentiated carcinoma component related to a low 

FIGO grade is essential in posing the correct diagnosis. If systematically 

searched for, dedifferentiation is found in 6-7% of all 

endometrial carcinomas, however they likely are misinterpreted 

as FIGO grade 3 endometriod carcinoma 2 . Dedifferentiated 

carcinoma more often involves the uterus in isolation, but may 

also involve both the uterus and one or both ovaries or only the 

ovaries 1 . Dedifferentiation may occur either in the primary or in 

the recurrence. Immunohistochemically in the undifferentiated 

component: vimentin is consistently positive, and cytokeratin 

(cocktail) and EMA are positive in 5-10% and 25% of cells, 

respectively. Neuroendocrine markers can be expressed in 10% 

of cells in more than a third of cases. The undifferentiated 

component of dedifferentiated carcinoma may be confused with 

other tumors of both epithelial and mesenchymal lineage, and the 

differential diagnosis includes not only endometrioid adenocarcinoma 

of high FIGO grade, as previously discussed, but also large 

cell neuroendocrine carcinoma, unclassified sarcomas, malignant 

mixed mullerian tumors, and non-Hodgkin’s lymphomas. The 

recognition of an undifferentiated component in an otherwise low 

grade endometrioid carcinoma is very important. 

references 

1 Silva EG, Deavers MT, Bodurka DC, et al. Association of low-grade 

endometrioid carcinoma of the uterus and ovary with undifferentiated 

carcinoma: a new type of dedifferentiated carcinoma? Int J Gynecol 

Pathol 2006;25:52-8. 

2 Silva EG, Deavers MT, Malpica A. Undifferentiated carcinoma of the 

endometrium: a review. Pathology 2007;39:134-8. 

3 Altrabulsi B, Malpica A, Deavers MT, et al. Undifferentiated carcinoma 

of the endometrium. Am J Surg Pathol 2005;29:1316-21. 

379 

Immunohistochemical analysis for Actin, Vimentin 

and S100 protein in lymphomas: review of 37 cases 

1)Crisman G. 2)Vitale AR. 3)Lucioni M. 4)Leocata P. 5)Hansmann 

ML. 

1)Dept of Experimental medicine, “San Salvatore” Hospital, University 

of L’Aquila, L’Aquila, Italy 2)Dept of Health’s Sciences, University 

of L’Aquila, L`Aquila, Italy 3) Anatomic Pathology Section, Foundation 

IRCCS Policlinico San Matteo, Pavia, Italy 4)Dept of Health’s 

Sciences, University of L’Aquila, L`Aquila, Italy 5) Senckenbergisches 

Institute for Pathology, University of Frankfurt, Frankfurt am Main, 

Germany 

Background. Splenic Marginal Zone Lymphoma (SMZL) is 

defined by the World Health Organization (WHO 2008) as a 

B-cell neoplasm composed by small lymphocytes that surround 

and replace the white pulp follicle and merge with a peripheral 

zone of larger marginal zone-like cells. Less is known about the 

background and the relationship between tumor cells and normal 

spleen. The goal of this study was to evaluate the expression of 

Actin, Vimenti and S-100 protein in SMZL, compared with other 

variants of B-cell lymphomas and normal spleen. 

Methods. 37 cases have been selected as following: 14 cases of 

SMZL, 8 Follicular Lymphomas (FL), 6 Hodgkin Lymphomas 

(HL) and 3 Large Cell Lymphomas (LCL), and 6 normal spleen. 

All cases have been immonohistochemically stained by the labeled 

avidin-biotin-peroxidase complex technique using Actin, 

Vimentin and S100 and reviewed from at least two pathologists. 

Afterwards, clinical, histopathological and immunohistochemical 

findings were correlated. 

Results. The analysis of the expression of Actin, Vimentin and 

S-100 among lymphomas and normal spleen revealed a characteristic 

SMZL-specific pattern of expression of Vimentin and S-100. 

The authors herein discuss and deepen their results. 

A controversial case of hypertrophic gastropathy 

Zaccaria M., Ingravallo G., Marzullo A. 


Background. Hypertrophic hyperplastic gastropathies are clinico-pathologic 

entities which are difficult to be classified by an 

histological point of view. The histological diagnosis needs to be 

correlated with clinical, endoscopic and laboratory data. 

Four well-defined hypertrophic gastropathies exist: a) Classic 

Menetrier disease with protein loss and hypoclorhydria, b) hypertrophic-hypersecretory 

protein-losing gastropathy, c) hypertrophic 

hypersecretory gastropathy, and d) Zollinger-Ellison 

syndrome (Fenoglio-Preiser; Gastrointestinal pathology. Third 

edition, 2008). Hyperplastic polyps are the most common lesions 

confused with hypertrophic gastropathies. 

Methods. Herein, we describe a case of hyperplastic gastropathy 

in 72 years-old man with anemia, severe protein loss and 

hypoprotidemia. The endoscopy showed several and prominent 

giant mucosal polyps on the greater and small curvature ranging 

in size from few mm to several cm, intermingled with areas of 

apparently normal mucosa. The most striking histologic feature 

was the presence of typical hyperplastic polyps characterized by 

marked elongation and branching of the gastric pits leading to 

a serrated appearance, tall mucin-secreting foveolar cells line, 

exaggerated and distorted pits, mild atrophy of the glandular 

compartment and a modest inflammatory infiltrate of eosinophils 

and plasma cells. 

Results. The histological pattern do not allowed to include 

this form in any of the above cited types, due to the prominent 

polypoid shape that was not in agree with the giant rugal folds 

described in typical cases. Indeed, we may propose the diagnosis 

of hypertrophic-hyperplastic gastropathy with loss of protein of 

“polypoid-type”.

380 

TCl1A and MNDA expression in splenic marginal 

zone lymphoma 

Zamò; A., Munari E., Chilosi M., Menestrina F. 

Patologia Sezione di Anatomia Patologica, Azienda Ospedaliera Universitaria 

Integrata, Verona, Italia 

Background. Splenic marginal zone lymphoma (SMZL) is a 

low-grade lymphoma showing a rather non-specific immunophenotype. 

Gene expression profiling studies suggested that TCL1A 

could be a marker of SMZL, but data reported from a very small 

series of SMZL indicate the contrary. MNDA has been suggested 

as a possible marker for SMZL, but only one study has been 

published. 

Our aim was to evaluate TCL1A and MNDA expression on a 

series of spleens, and to correlate findings with other immunophenotypical 

and morphological data. 

Methods. We collected 25 cases of SMZL for which both spleen 

and bone marrow samples were available. Splenic involvement 

was evaluated morphologically and classified as nodular, nodular/diffuse 

and diffuse. The immunophenotyping included CD20, 

CD5, CD3, Ki67, TCL1, T-bet, CCND1, EBER, CD123, DBA44, 

BCL2, BCL6, CD23, CD27, IgM, IgD, ANXA1, MNDA and 

GCET1. 

Results. We found a nodular pattern of infiltration in 6/25 (24%) 

cases, a nodular/diffuse pattern in 13/25 (52%) cases and a purely 

diffuse pattern of infiltration in 6/25 (24%) cases. MNDA was 

positive in 96% of cases and T-bet in 16% of cases. ANXA1, 

CCND1, GCET1 and CD123 were always negative while TCL1A 

was negative in 20/25 cases (80%). We stained bone marrow biopsies 

with TCL1A and found that, interestingly, 4/5 TCL1-positive 

cases evaluated in the spleen were negative. We did not find 

any statistically significant correlation between TCL1 expression 

and any histological pattern. 

Conclusions. TCL1A is rarely expressed in SMZL cases, and its 

expression is nearly always lost in bone marrow involvement. 

We suggest that TCL1A might be useful in determining the phenotype 

of SMZL, which is predominantly negative, in contrast to 

others entities such MCL and B-CLL, which are nearly always 

positive. Although rarely negative, MNDA also proved useful 

for the diagnosis of SMZL and the two markers together may be 

helpful in the diagnosis. 

rhabdoid carcinoma of the colon in polyposis: 

a case report 

1)Zanella C. 2)Remo A. 3)Bortuzzo G. 4)Molinari E. 5)Tollini 

F. 6)Talamini A. 7)Corradini I. 8)Pirani P. 9)Bonetti A. 10)Vendraminelli 

R. 

1)Department of Pathology, “Mater Salutis” Hospital, Azienda ULSS21, 

Legnago (Vr), Italy 2)Department of Pathology, “Mater Salutis” Hospital, 

Azienda ULSS21, Legnago (Vr), Italy 3)Department of Pathology, “S. 

Giacomo Apostolo” Hospital, Azienda Ulss 8, Asolo (Tv), Italy 4)Department 

of Surgery, “Mater Salutis” Hospital, Azienda ULSS21, Legnago 

(Vr), Italy 5)Department of Surgery, “Mater Salutis” Hospital, Azienda 

ULSS21, Legnago (Vr), Italy 6)Department of Surgery, “Mater Salutis” 

Hospital, Azienda ULSS21, Legnago (Vr), Italy 7)Department of Pathology, 

“Mater Salutis” Hospital, Azienda ULSS21, Legnago (Vr), Italy 

8)Department of Surgery, “Mater Salutis” Hospital, Azienda ULSS21, 

Legnago (Vr), Italy 9)Department of Oncology, “Mater Salutis” Hospital, 

Azienda ULSS21, Legnago (Vr), Italy 10)Department of Pathology, “Mater 

Salutis” Hospital, Azienda ULSS21, Legnago (Vr), Italy 

Background. A 73-year-old woman was recovered to Legnago’s 

Hospital complaining of rectal haemorrhage and abdominal 

mass in the right lower quadrant. Laboratory data on admission 

showed elevated levels of S-CA125 (49,7 U/ml) and a decreased 

hemoglobin level (6,6 g/dl). Abdominal ultrasound revealed a 

voluminous mass, 10 × 4 cm in size, involving intestinal loop. 

Anamnestic history revealed meningioma at 31 years-old and 

essential hypertension. 


Results. The surgical presentation (big mass without nodes involvement) 

pushed at intraoperative pathologic consultation that 

showed a small blue cells neoplasia. 

Grossly, the tumor measuring 10 × 8 cm sited in right colon 

involving other intestinal loops (T4). It presented as a well-demarcated 

mass lesion with central deep ulceration. In remaining 

intestinal loops were present several tubular polyps with severe 

dysplasia. Histologically, the tumour was an heterogeneous 

neoplasia consisting of a poor differentiated (G3) glandular 

component associated to prominent rabdoid and undifferentiated 

blue cells infiltration (see frozen section). Tumour budding 

was present high level. Metastasis was observed in lymph nodes 

(2/23) (N1). 

The rhabdoid and neoplastic cells were negative for CK7; focally 

immunoreactive to CK20, and diffusely positive for vimentin;. 

The rhabdoid cells were not immunoreactive to desmin, S-100, 

HMB45, myogenin and CD45. Strong expression of MSH2 protein 

was noted but MLH1 was negative. 

Mutation was not found in exon 2 of the K-ras gene (“Wild-type”). 

The patient underwent to adjuvant chemotherapy (capecitabin 

and oxaliplatin). 5 months later the patient referred to surgery for 

recurrence and peritoneal metastasis. 

In literature has been reported 5 cases of rhabdoid colon carcinoma 

but this in the first associated to several polyps. All cases 

has a overall survival time maximum 12 months. 

For these cases the histology is the most important prognostic 

factor and could be hypothesized to used every therapy ab initio 

comprising biologica therapy when possible. 

extensive characterization of eGfr pathways 

may help in integrating the use of egfr-targeted 

therapies in patients with squamous cell anal 

cancer 

1)Zanellato E. 2)Martin V. 3)Molinari F. 4)Crippa S. 5)De dosso 

S. 6)Franzetti-pellanda A. 7)Movilia A. 8)Paganotti A. 9)Deantonio 

L. 10)Frattini M. 


Locarno, Svizzera 2)Laboratorio diagnostica molecolare, Istituto cantonale 


Istituto cantonale di patologia, Locarno, Svizzera 4)Patologia 

clinica, Istituto cantonale di patologia, Locarno, Svizzera 5)Oncology 

institute of southern switzerland, Ente ospedaliero cantonale, Bellinzona, 

Svizzera 6)Oncology institute of southern switzerland, Ente ospedaliero 

cantonale, Lugano, Svizzera 7)Pathology, Civil hospital, Legnano, 

Italia 8)Mediacal sciences, University school of medicine, Novara, 

Italia 9)Radiotherapy, University school of medicine, Novara, Italia 


Locarno, Svizzera 

Background. Locoregional squamous cell anal cancer (SCAC) 

is a rare disease, and definitive chemo-radiation represents the 

standard curative approach. Monoclonal antibodies (MoAbs) targeting 

EGFR are synergistic with radiotherapy in squamous cell 

carcinomas. In colorectal cancer, it has been demonstrated that 

the efficacy of anti-EGFR MoAbs is achieved both in the presence 

of copy number gain (CNG) of EGFR gene and absence of 

mutations in EGFR downstream members. We described alterations 

occurring in EGFR pathway, to evaluate whether MoAbs 

against EGFR can be integrated in the management of SCAC 

patients. 

Methods. Thirty-six SCAC biopsies were collected in the 

Departments of Pathology in Locarno, Legnano and Novara. 

EGFR gene status was assessed by fluorescent in-situ hybridization, 

whilst K-Ras, BRAF and PIK3CA mutations by direct 

sequencing. 

Results. Biopsies were evaluable in all but one cases. EGFR 

CNG was observed in 5 cases (20%). No BRAF mutations were 

detected. K-Ras was mutated in 1 case (3%, G12V change), 

PIK3CA in 8 cases (23%, 6 changes in exon 9 and 2 in exon


20). The K-Ras mutation occurred in a patient with EGFR CNG. 

One patient showed a concomitant PIK3CA exon 9 mutation and 

EGFR CNG. The 3 remaining cases with EGFR CNG were characterized 

by absence of any gene mutations in EGFR downstream 

members. 

Conclusions. In addition to EGFR gene deregulation, K-Ras and 

PIK3CA mutations are involved in SCAC carcinogenesis. It has 

been demonstrated in colorectal cancer that K-Ras wt/PIK3CA 

exon 9 mutations/EGFR CNG are associated with clinical benefit 

from EGFR-targeted therapies. Therefore 4 out of 36 SCAC 

patients (11%) might have a proficient pattern for anti-EGFR 

MoAbs treatment. Our results suggest a possible role of EGFRtargeted 

therapies in integrated treatment of locoregional SCAC, 

and emphasize the need of an early analysis to identify patients 

who might benefit from these drugs. 


high grade endometrioid carcinomas (G3 fIGO) 

in comparison with low grade endometrioid 

carcinoma (G1 and G2 fIGO) and non-endometrioid 

carcinoma 

GF Zannoni, VG Vellone, V. Arena, G. Chiarello, A. Carbone, 

MG Prisco*, G. Scambia*, D. Gallo* 



Pol. A Gemelli. Università Cattolica del Sacro Cuore 

Background. Endometrial cancers have long been classified into 

two major categories (Type I and II); Type I estrogen-dependent 

adenocarcinoma, with an endometrioid morphology, and Type 

II non-estrogen-dependent adenocarcinoma, mainly showing a 

serous papillary or clear cell morphology. However, not all tumors 

fit into this dualistic model. The current study was aimed at 

comparing clinical-pathological and molecular features of High 

Grade Endometrioid Carcinomas (G3 FIGO, HGECs) with those 

of Low Grade Endometrioid Carcinoma (G1-2 FIGO, LGECs) 

and of Non-Endometrioid Carcinoma (NECs). 

Methods. A total of 98 cases with hysterectomy, bilateral salpingo-oophorectomy 

and lymph nodal dissection, with more than 

10 lymph nodes, were included in this evaluation. To this end, a 

panel of clinical, morphological and molecular parameters, including 

histological type and grade, myometrial invasion, lymphvascular 

space invasion, lymphonodal and extra-lymphonodal 

metastases, staging and expression of steroid hormone receptors, 

p53, ki67, Bcl-2, and HER-2/neu, were evaluated in the three 

different populations: LGECs (n = 57), HGECs (n = 26), and 

NECs, (n = 15). 

Results. Data showed that PDECs exhibit mixed or overlapping 

morphological and biological features of both Type I and Type 

II endometrial carcinomas. HGECs appeared similar to LGECs 

in morphological appearance, and p53 expression level; findings 

strongly different from LGECs, included a higher local aggressiveness, 

and a higher invasion of lymph-vascular spaces, with 

a greater rate of lymph nodal metastases; HGECs had a lower 

expression of both ERα and PR, and a significantly higher proliferative 

index, compared to LGECs. HGECs were uniquely similar 

to NECs for invasion rate of lymph-vascular spaces, lymph 

nodal metastases incidence, ERα positivity, and proliferative 

index. HGECs, however, infiltrate more than NECs, retained a 

significant higher positivity for PR, and showed a lower expression 

of p53. 


poorly differentiated endometrioid carcinomas 

(G3 fIGO) in comparison with well differentiated 

endometrioid carcinoma (G1 and G2 fIGO) and 

non-endometrioid carcinoma 

381 

GF Zannoni, VG Vellone, V. Arena, G. Chiarello, A. Carbone, 

MG Prisco*, G. Scambia*, D. Gallo* 



Pol. A Gemelli. Università 

Background. Endometrial cancers have long been classified into 

two major categories (Type I and II); Type I estrogen-dependent 

adenocarcinoma, with an endometrioid morphology, and Type 

II non-estrogen-dependent adenocarcinoma, mainly showing a 

serous papillary or clear cell morphology. However, not all tumors 

fit into this dualistic model. The current study was aimed at 

comparing clinical-pathological and molecular features of High 

Grade Endometrioid Carcinomas (G3 FIGO, HGECs) with those 

of Low Grade Endometrioid Carcinoma (G1-2 FIGO, LGECs) 

and of Non-Endometrioid Carcinoma (NECs). 

Methods. A total of 98 cases with hysterectomy, bilateral salpingo-oophorectomy 

and lymph nodal dissection, with more than 

10 lymph nodes, were included in this evaluation. To this end, a 

panel of clinical, morphological and molecular parameters, including 

histological type and grade, myometrial invasion, lymphvascular 

space invasion, lymphonodal and extra-lymphonodal 

metastases, staging and expression of steroid hormone receptors, 

p53, ki67, Bcl-2, and HER-2/neu, were evaluated in the three 

different populations: LGECs (n = 57), HGECs (n = 26), and 

NECs, (n = 15). 

Results. Data showed that PDECs exhibit mixed or overlapping 

morphological and biological features of both Type I and Type 

II endometrial carcinomas. HGECs appeared similar to LGECs 

in morphological appearance, and p53 expression level; findings 

strongly different from LGECs, included a higher local aggressiveness, 

and a higher invasion of lymph-vascular spaces, with a greater 

rate of lymph nodal metastases; HGECs had a lower expression 

of both ERα and PR, and a significantly higher proliferative index, 

compared to LGECs. HGECs were uniquely similar to NECs for 

invasion rate of lymph-vascular spaces, lymph nodal metastases incidence, 

ERα positivity, and proliferative index. HGECs, however, 

infiltrate more than NECs, retained a significant higher positivity 

for PR, and showed a lower expression of p53. 

Cytoplasmic expression of estrogen receptor beta 

(erβ) as a prognostic factor in vulvar squamous 

cell carcinoma in elderly women 

Gian Franco Zannoni, Valerio Gaetano Vellone, Maria Grazia 

Prisco*, Ilaria De Stefano*, I. Pennacchia, Anna Fagotti*, 

Giovanni Scambia*, G. Rindi, Daniela Gallo* 



Pol. A Gemelli. Università Cattoli 

Background. Epidemiological data suggest that there are two 

types of vulvar squamous carcinoma (SCC), with two etiologic 

paths at work in carcinogenesis. The first type is often seen in 

women over the age of 50 and is associated with non-neoplastic 

disorders, such as hyperplasia or lichen sclerosus; the second type 

is seen in women under the age of 50 and is associated with HPV 

infection. We recently found that a shift from a mainly nuclear to 

a mainly cytoplasmic localization of Estrogen Receptor β (ERβ) 

tightly characterizes the transition from normal epithelium to 

invasive cancer in elderly patients with vulvar SCC non HPVrelated. 

In the present study we investigated the prognostic value 

of cytoplasmic ERβ in a series of thirty-three untreated vulvar 

cancer patients.

382 

Methods. Immunohistochemistry was carried out by using the 

polyclonal rabbit anti-human ERβ antibody (clone H-150). 

Nuclear and cytoplasmic staining was evaluated and correlated 

with histopathologic characteristics and molecular parameters 

(i.e. Ki67 and p21), overall survival (OS), and disease-free survival 

(DFS). 

Results. The expression of cytoplasmic ERβ was found to be significantly 

associated with FIGO grade (p = 0.006), while no association 

was found with any of the remaining variables examined. 

Cases with high cytoplasmic ERβ expression showed a lower 

disease-free survival (DFS) and a lower overall survival (OS) 

with respect to cases with low cytoplasmic ERβ (p = 0.007 and 

p = 0.01, respectively). There was also a progressive decline in 

both the DFS and OS with increasing tumor size (DFS, p = 0.05 

and OS, p = 0.07), and with increasing depth of infiltration (DFS, 

p = 0.14 and OS, p = 0.07). In multivariate analysis, only tumor 

size and cytoplasmic ERβ staining retained an independent negative 

prognostic role for DFS and OS. Our study suggests that the 

assessment of cytoplasmic ERβ expression could be helpful to 

identify poor prognosis in elderly patients with vulvar squamous 

cell carcinoma. 

role of IGH fISH DNA PrOBe, split signal, in pleural 

and peritoneal involvment of non-Hodgkin 

lymphoma on cytological effusion samples 

1)Zeppa P. 2)Genesio R. 3)Iaccarino A. 4)Cozzolino I. 5)Bianco 

A. 6)Plaiato F. 7)Fernandez L. 8)Vigliar E. 9)Nitsch L. 10)Palombini 

L. 

1)Scienze Biomorfologiche e Funzionali, Università di Napoli “Federico 

II”, Napoli, Italia 2)Chirurgia Generale e Specialistica, Azienda Ospedaliera 

V. Monaldi, Napoli, Italia 3)Scienze Biomorfologiche e Funzionali, 

Università di Napoli “Federico II”, Napoli, Italia 4)Scienze Biomorfologiche 

e Funzionali, Università di Napoli “Federico II”, Napoli, Italia 

5)Scienze Biomorfologiche e Funzionali, Università di Napoli “Federico 

II”, Napoli, Italia 6)Scienze Biomorfologiche e Funzionali, Università di 

Napoli “Federico II”, Napoli, Italia 7)Scienze Biomorfologiche e Funzionali, 

Università di Napoli “Federico II”, Napoli, Italia 8)Biologia e Patologia 

Cellulare e Molecolare, Università di Napoli “Federico II”, Napoli, 

Italia 9)Scienze Biomorfologiche e Funzionali, Università di Napoli “Federico 

II”, Napoli, Italia 10)Scienze Biomorfologiche e Funzionali, Un 

Background The human IGH (immunoglobulin heavy) locus at 

chromosome 14q32 is the most frequently involved in different 

translocations of non-Hodgkin lymphoma (NHL). The IGH FISH 

DNA probe, split signal, is a mixture of two fluorochrome-labeled 

DNA: the green fluorescein-labeled DNA probe (IGH-Flu) 

that binds to a 612 kb segment telomeric, and the red labelled 

(IGH-TR) that binds to a 460 kb segment centromeric, to the IGH 

breakpoint respectively. Therefore IGH FISH DNA probe, split 

signal should detect any translocation involving the IGH locus at 

chromosome 14q32. The cytological diagnosis of pleural or peritoneal 

involvement by NHL may be hampered by scanty cellularity, 

concomitant reactive infiltrate and good differentiation of the 

cells. The aim of this study was to evaluate if the IGH FISH DNA 

probe, split signal may be helpful in the cytological diagnosis of 

pleural and peritoneal involvement by NHL. 

Methods. We retrospectively tested the IGH FISH DNA Probe, 

split signal on 15 cytological samples of NHL pleural and peritoneal 

effusions and 5 negative samples to assess the sensitivity and 

specificity of the probe. 

Results. The IGH FISH DNA probe detected split signals in a 

significant number of cells in 12 out 15 positive cases and in none 

of the 5 negative controls. 

Conclusions. The IGH FISH DNA Probe, is highly sensitive 

for the detection of translocations involving the IGH locus on 

effusion’s cytological samples, whereas it is not specific for the 

single pathological sub-types. The procedure is highly effective 

in mixed polymorphous cell populations and in scantily cellulated 

samples in which other procedures may be hampered. 


Immunohistochemical study of global DNA 

methylation and histone H3 (lysine 9) acetylation 

in myelodysplastic syndrome on paraffinembedded 

bone marrow trephine biopsy 

specimens 

1)Zizzi A. 2)Poloni A. 3)Giantomassi F. 4)Stramazzotti D. 

5)Leoni P. 6)Goteri G. 

1)Neuroscienze, Sezione di Anatomia Patologica, Univ. Politecnica delle 

Marche-Ospedali Riuniti, Torrette - Ancona, Italia 2)Clinica Ematologica, 

Univ. Politecnica delle Marche-Ospedali Riuniti, Torrette - Ancona, 

Italia 3)Neuroscienze, Sezione di Anatomia Patologica, Univ. Politecnica 

delle Marche-Ospedali Riuniti, Torrette - Ancona, Italia 4)Neuroscienze, 

Sezione di Anatomia Patologica, Univ. Politecnica delle Marche-Ospedali 

Riuniti, Torrette - Ancona, Italia 5)Clinica Ematologica, Univ. Politecnica 

delle Marche-Ospedali Riuniti, Torrette - Ancona, Italia 6)Neuroscienze, 

Sezione di Anatomia Patologica, Univ. Politecnica delle Marche-Ospedali 

Riuniti, Torrette - Ancona, Italia 

Background. Myelodysplastic syndrome (MDS) are a heterogeneous 

group of clonal haematopoietic disorders with ineffective 

haematopoiesis leading to pancytopenia and an increased risk 

of transformation to acute myeloid leukaemia (AML). Clonal 

cytogenetic abnormalities have prognostic significance. More 

recently the importance of epigenetic events which regulate gene 

expression at post-translational level (alterations in DNA methylation 

status and modification of histone tails) has allowed the 

introduction of drugs inducing DNA hypomethylation or histone 

acetylation in MDS treatment. 

Methods. To study these biologic targets at histological level, we 

immunostained bone marrow sections from 55 MDS patients with 

two specific antibodies directed to epitopes related to the global 

DNA methylation and histone acetylation. Cases were stratified 

the International Prognostic Scoring System (IPSS), blast count 

and karyotype. Immunohistochemistry was performed on formalin-fixed, 

paraffin-embedded and EDTA decalcified bone marrow 

sections, using anti-5-methylcytosine/5mc and anti-Acetyl-Histone 

H3 (Lys9)/AcH3K9 antibodies. Immunoreactivity was evaluated 

by counting the percentage of positive cells and expressing 

the intensity of staining by a three point-scoring systems. For 

each case counts were repeated three times and the mean values 

were considered to calculate a final “H-score” by multiplying the 

percentage of positive cells and the intensity score. 

Results. The 5mc immunostaining score correlated significantly with 

the IPSS, the blast count and the karyotype, whereas the AcH3K9 

immunostaining did not, suggesting that global DNA hypermethylation 

correlate with MDS aggressiveness and providing a molecular 

explanation for the therapeutic success of hypomethylation-inducing 

agents in MDS and why patients with a poor karyotype respond best. 

Future studies have to analyse whether these parameters may serve 

as a new predictive marker for therapy response. 

foxP3: a novel prognostic marker of hepatocellular 

carcinoma 

Zonetti M.J., Mazzarelli P., Schiaroli S., Spagnoli L.G., Pucci S. 

Biopatologia, Tor Vergata, Roma, Italia 

Background. FOXP3 is a member of the forkhead family of 

nuclear transcription factors and is the main modulator of the 

immune response regulating the development and the function 

of regulatory T-cells (TregCD4 +CD25+Foxp3+), effectors of 

peripheral tolerance. Recently it was shown that Foxp3 is expressed 

in pancreatic and breast carcinomas and associates with 

worse overall survival probability. Three splice variant of FOXP3 

mRNA, have been recently identified. It has been demonstrated 

that TGF-β2, cytokine involved in the development of hepatitis 

and progression of hepatocellular carcinoma, can induce the 

nuclear expression of Foxp3 in pancreatic ductal adenocarcinoma 

cells. Therefore we analyze the correlation of Foxp3 and TGF-β2


expression in histological samples of hepatocellular carcinoma 

and in a hepatocarcinoma cell line, Hep-G2. 

Methods. FOXP3 and TGF-β2 expression was analyzed by immunohistochemistry 

on specimens of liver cell dysplasia (LCD) 

(n = 10), non-alcoholic steatosis (NASH) (n = 10) and hepatocellular 

carcinoma (HCC) HCV-related and not-related (n = 20). 

The modulation of Foxp3 splicing isoforms in liver tissue and in 

TGF-β2-treated Hep-G2 cell line, has been evaluated by western 

blot. FOXP3 mRNA isoforms were analyzed by RT-PCR. 

Results. We observed by IHC a positive correlation between 

nuclear Foxp3 expression in hepatocytes and the presence of 

TGF-β2 in tumoral microenvironment. In vitro experiments 

confirmed that TGF-β2 induces a strong selective modulation 

of Foxp3 isoforms in Hep-G2. Moreover in HCC was evident 

the up-regulation of the ∆2∆3 isoform, not expressed in normal 

liver tissues and in Treg lymphocytes, indicating the involvement 

of this factor in the neoplastic disease. The upregulation of 

this specific Foxp3 isoform only in HCC suggests the important 

involvement of this transcription factor in hepatocarcinogenesis 

and that it could be considered a new molecular marker in the 

natural development of the liver neoplastic disease. 

Pseudoepitheliomatous hyperplasia arising from 

hypertrophic lichen planus 

D. Morichetti, Zorzi M.G., Pusiol T., Piscioli F. 

Institute of Anatomic Pathology, S. Maria del Carmine Hospital, Rovereto, 

Italy 

Background. Hypertrophic lichen planus (HLP) shows prominent 

hyperplasia and overlying orthokeratosis of the epidermis. 

To day 50 cases of squamous cell carcinoma (SCC) have been 

reported as neoplastic transformation of HLP. We report pseudoepitheliomatous 

hyperplasia (PH) simultaneously found in two 

hypertrophic lichen planus (HLP) lesions simulating SCC with 

emphasis to diagnostic differentiation. 

Materials and Methods. A 73-years old woman presented numerous 

hyperkeratotic nodules of 14 months duration located 

on the middle surface of both lower legs end on the fingers. The 

biopsies of two lesions showed typical features of HPL in continuity 

with and adjacent to PH. The shave biopsy of the smallest 

383 

hyperkeratotic nodule was diagnosed as “well-differentiated 

SCC”. The histology of the latter nodule consisted of a benign 

irregular hyperplasia of the epidermis with gross acanthosis, 

downward proliferation with moderate dyskeratosis and horn 

cyst formation. These features are typical of PH. Review of the 

first biopsy, considering further clinical findings, suggested a 

diagnosis of PH rather than SCC. After two years the patient is 

free of recurrence. 

Discussion. PH is a histopathological reaction pattern rather than 

a disease sui generis. It is characterized by irregular hyperplasia 

of the epidermis which also involves follicular infundibula and 

acrosyringia. This proliferation occurs in response to a wide 

range of stimuli. Distinguishing PH and SCC may be challenging 

for pathologists. If the epidermal hyperplasia is severe it may 

mimic a SCC on a shave biopsy 1 . Numerous reported SCCs arising 

in HLP may not be accepted as such because the HLP-SCC 

sequence is not illustrated 2-7 . Since PH may simulate SCC, accurate 

criteria should be used in the differential diagnosis. In our 

case the infiltrative pattern is alarming and a precise diagnosis is 

problematic. We believe that the presence of multiple lesions, follow-up 

and proliferation from follicular infundibula are valuable 

criteria indicating HP rather than SCC. 

references 

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3 Singh SK, Saikia UN, Ajith C, et al. Squamous cell carcinoma arising 

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ruber planus of the leg complicated by a squamous cell carcinoma. Int 

J Dermatol 2003;42:415-6.

Pathologica 2010;102:385-390 AuTHOr INDeX 

Abreu R.F.N., 309 

Adamo V., 301, 303 

Addati T., 338 

Addati T., 237 

Addesso M., 279, 280 

Agabiti S., 268 

Agati P., 237 

Agati R., 237 

Aglietta M., 131 

Agostinelli C., 297, 314, 339, 

352, 357 

Agozzino A., 168 

Aiello L., 345 

Al Omoush T., 177 

Alaggio R., 263, 268, 284 

Albanese V., 244 

Alberghini M., 256, 259 

Alberizzi P., 281 

Albino G., 327 

Albrizio M., 259 

Aldovini A., 269 

Aldovini D., 290 

Alessandri G., 244, 247 

Alessandrini L., 237 

Alì G., 237 

Allevato F., 340 

Alò P.L., 345 

Aloi F.S., 283 

Altavilla A.M., 238 

Altieri R., 339 

Amadori P., 278 

Ambrosini Spaltro A., 238 

Ambrosio M.R., 187, 239, 317, 

326, 329, 350 

Ambu R., 247 

Amicarelli V., 334 

Amico P., 239, 255, 267, 302, 

369, 377 

Amini M., 268 

Ammirabile M., 330 

Amodio G., 374 

Anastasio A., 240 

Andorno A., 174 

Andreozzi M.C., 239, 240 

Angeli G., 193 

Angelini A., 167 

Angeloni C., 151 

Angelotti U., 324 




Angelotti U.F., 240, 275, 357, 358 

Angelucci D., 338, 370 

Angiero F., 247, 248 

Angione V., 220, 342 

Angrisani B., 279, 280, 374 

Angrisani P., 279, 280 

Annunziata S., 353 

Anselmi L., 287 

Antinori S., 172 

Anton I.M., 333 

Antoniazzi S., 247 

Antonini D., 240 

Apicella P., 293, 355 

Appetecchia M., 270 

Aprile G., 281, 282 

Aquino G., 269 

Arborea G., 325, 326, 332, 349, 

367 

Arbustini E., 168 

Arcaini L., 185, 311 

Ardò N.P., 328 

Arena V., 241, 366, 381 

Arisio R., 373 

Armentano R., 341, 373, 194, 195 

Armiraglio E., 256 

Arnolfo E., 308 

Arpinelli S., 342 

Arrizza L., 375 

Artico R., 268 

Artiola G., 354 

Artioli P., 314, 352 

Ascione P., 303, 354, 370 

Ascoli V., 241 

Asioli S., 242, 281, 313, 349 

Asselti M., 243, 261 

Avellini C., 368 

Azzoni C., 243 

Baccarini P., 294, 370 

Bacci F., 297, 314, 339, 352 

Badiali G., 289, 321 

Badolato R., 310 

Baggiani A., 135 

Bagni I., 311 

Balasus D., 314 

Baldassarre F., 353 

Baldelli R., 270 

Baldin P., 243 

Baldoni C., 244 

Balsamo A., 346 

Balzarini P., 244, 247, 248 

Banchelli I.B., 280 

Bandiera V., 177 

Barbagallo G.M., 244 

Barbagli L., 238 

Barbareschi M., 269, 278, 283, 

290 

Barbi S., 271 

Barbolini G., 351 

Bardari F., 342 

Barnabei A., 270 

Basolo F., 343 

Baron L., 245, 298, 353 

Barone F., 365 

Baroni G., 293 

Barra E., 377 

Barresi E., 245, 350, 358 

Barresi G., 245, 346 

Barresi V., 227, 245, 301, 317 

Baseggio C., 298 

Basolo F., 135, 298 

Baumhoer D., 369 

Bazzoli F., 164 

Beccati M.D., 246 

Becherini F., 271 

Beghelli S., 271 

Bei K., 342 

Bellan C., 187, 326, 350 

Bellei E., 248 

Bellevicine C., 246, 312, 374 

Bellezza G., 246, 283, 341 

Bellini P., 348 

Bellisano G., 247, 344 

Bellizzi A., 261, 347 

Beltrami C.A., 368 

Ben Dor D., 250, 254, 255, 258 

Benedetti F., 234 

Benedetto G., 369 

Benemei S., 293 

Benerini Gatta L., 244, 247, 248 

Benetti A., 244, 247, 248 

Bensi T., 308 

Berenzi A., 244, 247, 248 

Bergamini S., 248 

Berni Canani A., 306 

Berretta R., 299 

Bersani S., 264, 265, 271, 301, 

359 

Bersiga A., 299 

Berta G.N., 303 

Bertacca G., 297, 301 

Bertani A., 346 

Berti E., 311 

Berti P., 135 

Berto E., 335 

Bertolini F., 248, 358 

Betta P.G., 308 

Bettelli S., 248, 346 

Betts C.M., 345 

Bevilacqua G., 200 

Biancalani M., 293, 355 

Bianchi G., 248 

Bianchi G.P., 311 

Bianchini E., 330 

Bianco A., 382 

Bianco M., 251 

Biasi D., 367 

Bigini D., 301 

Bignami M., 365 

Bihl M.P., 239, 240 

Binaschi A., 335 

Bio R., 352 

Biolcati M., 285 

Bisaro C., 291 

Bisceglia M., 249, 250, 251, 253, 

254, 255, 256, 257, 258, 259, 

260, 265, 267, 269, 272, 274, 

298, 322, 334, 362, 363, 378 

Bisceglia M.L., 249, 258 

Bisceglia S., 249 

Bisceglie D., 261 

Blasi N., 325 

Bleiweiss I., 250 

Boccardo S., 291 

Bocchio C., 288 

Boggiani D., 366 

Boldrini L., 237, 261 

Bollito E., 292 

Bombonato F., 290 

Bonanno A., 262 

Bonanno A.M., 368 

Bonanno E., 283, 294, 335, 366 

Bonazza D., 177 

Bondi A., 244 

Bondi A., 340 

Bondi A., 153, 215, 244, 270, 

294, 307, 340 

Bondi F., 262 

Bonetti A., 348, 380 

Bonetti F., 265, 328, 329, 336 

Bonfadini M.G., 262, 263 

Bonfili P., 375 

Bongiovanni M., 277 

Bonifacio D., 177, 338 

Bonin S., 133 

Bonoldi E., 192, 194, 195 

Bonora E., 135 

Bonzanini M., 278, 336 

Bordi C., 243 

Bordoni A., 363 

Borgia L., 378 

Bortul M., 177 

Bortuzzo G., 347, 348, 380 

Bosari S., 291 

Bosco A., 271 

Bosco D., 241 

Bosincu L., 276 

Bosisio F.M., 263 

Bossini P., 310 

Botta C., 134, 313, 349 

Bottarelli L., 243 

Botti G., 266, 269, 305 

Botticelli L., 337 

Boveri E., 311 

Bragantini E., 264, 269, 283, 290 

Branca G., 301, 368 

Brancato F., 263 

Brand R., 369 

Brazzarola P., 275 

Briani G., 330 

Brigati F., 299, 366 

Brisigotti M., 238 

Bronte V., 354 

Brulatti M., 370 

Bruna R., 162 

Brunelli M., 143, 148, 264, 265, 

271, 291, 295, 301, 302, 336, 

359, 360 

Brunelli S., 336 

Brunello E., 264, 265, 271 

Bruno F., 241 

Bruno G., 377 

Bruno G.M., 377 

Bruno M., 265, 363 

Bruscaggin A., 162 

Bucci F., 363 

Bufo P., 266, 267, 269, 328, 334, 

356 

Buglioni S., 154. 

Bulgarelli L., 352 

Busatto F., 289 

Busolin R., 298 

Bussolati G., 242, 349 

Butera D., 295, 305 

Butera M., 288 

Buttitta F., 290, 313, 378 

Cabibi D., 303 

Cacciotti J., 285, 342 

Cadei M., 247 

Cagiano S., 356 

Caio G., 174 

Caldarazzo A., 307 

Calderoni M., 296, 297 

Caleo A., 279 

Caliandro D., 238 

Caliendo V., 349 

Calienno R., 278 

Callea M.R., 298 

Caltabiano R., 267, 268 

Calvisi G., 314 

Camici P.G., 313 

Camisa R., 366, 367 

Campagna D., 268 

Campagnaro T., 336 

Campanella D., 373 

Campanella G., 373 

Campanini N., 280, 299, 308, 

366, 367 

Campioli L., 352 

Campostrini F., 348 

Canavese G., 139 

Canesso A., 268 

Caneva A., 330 

Cannazza V., 254 

Cannizzaro M., 369 

Canobbio L., 287 

Canova E., 271 

Cantaloni C., 269, 283, 290 

Cantile M., 269 

Canzonieri V., 350 

Capelli A., 241 

Capelli P., 336, 337 

Capello D., 311 

Capitanio G., 330 

Capocaccia R., 371 

Capodanno A., 237, 261, 298 

Caponio M.A., 237, 338, 339 

Caporali R., 375

386 

Caporusso C., 326 

Cappella E.D., 295 

Cappellesso R., 237, 290, 371 

Caramaschi P., 367 

Carbone A., 224, 241, 282, 381 

Carcangiu M.L., 141 

Cardone P., 164 

Carducci A., 329 

Carè A., 371 

Caristi N., 300 

Carlomagno C., 277 

Carlucci M., 269 

Carluccio L., 238 

Carminati P., 354 

Carosi I., 250 

Carosi I., 250 

Carosi M., 270 

Carossa S., 242 

Carotenuto P., 333 

Carotenuto V., 251 

Carrabba A., 353 

Carru C., 276 

Carta G., 315 

Cartaginese F., 271 

Caruso G., 273, 324, 325, 367 

Caruso M.L., 341, 373 

Caruso R., 262 

Casadei G.P., 270, 306, 307 

Casali P., 371 

Casalini S., 297 

Casquilho P., 309 

Cassenti A., 164, 242, 318 

Cassoni P., 164, 242, 318 

Castagna M., 270 

Castaing M., 244, 369 

Castellano I., 139, 164, 318 

Castelli F., 247 

Castelluccio E., 270 

Castorani L., 284 

Castrilli G., 370 

Castriota M., 328 

Catacchio R., 324, 325, 367 

Catalano A., 368 

Catalano F., 368 

Cataldo I., 239, 271, 336 

Cavaliere A., 246, 271, 283, 341, 

368 

Cavallini A., 373 

Cavanaugh B., 309 

Cavazza A., 238 

Cavazzana A., 297, 301 

Cavazzini L., 295 

Cecchi R., 293 

Cembrola S., 303 

Cenacchi G., 229 

Centrone M., 339 

Cernic S., 281 

Cerruti G., 287 

Certo G., 303, 305, 306 

Cesarani F., 342 

Cesari S., 272, 281 

Cesinaro A.M., 182 

Cheng L., 188 

Chiapetta C., 285 

Chiappetta C., 285 

Chiara S., 287 

Chiaramonte A., 272 

Chiarello G., 381 

Chieco P., 153 

Chiesa F., 294 

Chilli L., 314, 352 

Chilosi M., 264, 275, 276, 337, 

360, 380 

Chinol M., 354 

Ciafrè S.A., 335 

Ciliberto G., 354 

Cimmino A., 240, 273, 274, 275, 

325, 332, 357, 358, 359 

Ciolfi A., 340 

Cipollone F., 290 

Ciraolo E., 370 

Cirelli R., 364 

Cirillo M.E., 327 

Ciuffetelli V., 314, 360 

Clarini R., 299 

Claudi R., 338, 370 

Clemente C., 328 

Clemente D., 291 

Cocca M.P., 273, 274, 332 

Cocchi R., 289, 292, 335 

Coco M., 322 

Coggi G., 192, 194 

Coggia M., 274 

Colacchio G., 274 

Colagrande A., 240, 275,, 292324, 

325, 326, 358 

Colagrande M., 273 

Colantoni A., 283, 294, 335 

Colantuoni V., 333 

Colasante A., 338 

Colato C., 275, 276, 367 

Colecchia M., 192 

Colella G., 360 

Colella R., 246, 283 

Colesanti M., 278 

Coletti G., 315 

Collina G., 181, 182, 306, 307 

Colombo F., 363 

Colombo M., 288, 289 

Colombo M.P., 371 

Colonese F., 301, 303 

Colonna F., 269 

Comanescu M., 304 

Comin C.E., 355 

Concardi M., 168 

Conte P.F., 248, 358 

Contini M., 276 

Corcione L., 308 

Corradini I., 380 

Corsi F., 356, 357 

Cortesi E., 342 

Cortesi L., 337 

Cossu Rocca P., 276 

Costamagna D., 289 

Costarelli L., 268, 366 

Cotoi C.G., 288 

Cozzolino I., 277, 312, 374, 377, 

382 

Cremonini N., 307 

Cricca M., 153 

Crippa S., 363, 380, 277, 300, 

310, 320 

Crisafulli C., 245 

Crisman G., 315, 360, 379 

Crocetti E., 312 

Crucitti P., 215, 270, 307, 340 

Cucchi M.C., 243, 278 

Culli M., 301 

Cuorvo L.V., 278, 283, 290 

Curcio C., 278, 303, 333 

Curcio M.P., 295, 305 

Curduman M., 303 

Cusatelli P., 271 

D’Amico C., 243 

D’Adda T., 243, 279, 299, 308 

D’Agruma L., 322 

D’Alessandro E., 342 

D’Amati G., 313, 330, 342 

D’Ambrosio G., 280 


D’Amore E.G.S., 268, 296 

D’Amuri A., 293, 307, 345 

D’Ancona G., 356 

D’Angelo G., 313, 279 

D’Angelo V., 251 

D’Antona G.I., 265 

D’Antonio A., 279 

D’Antonio A., 280 

D’Antuono T., 361 

D’Avella D., 296 

D’Eredità G., 284, 326 

D’Errico A.D., 280 

D’Errico M., 250 

D’Orazi V., 268 

Da Pozzo G., 314, 352 

Dal Bello B., 272, 280, 281, 299, 

308 

Dal Mas A., 375 

Dalena A.M., 333 

Dalfior D., 328 

Dall’Olio D., 294, 306 

Dalla Palma P., 269, 278, 283, 

290 

Damiani D., 314 

Damiani S., 238 

Daniele B., 333 

Daniele I., 336 

Daniele L., 176 

Danieli D., 296 

Dante S., 330 

Daprile R., 243, 261, 318 

Dardano A., 275, 276 

David E., 231 

De Angelis R., 371 

De Biase D., 135, 160, 281, 282, 

320, 345, 349, 370 

De Dosso S., 380 

De Falco G., 187 

De Feo M., 318 

De Gaetani C., 248, 311, 346 

De Giorgi V., 312 

De Giorgio F., 241 

De Gregorio A., 353 

De Luca C., 246 

De Luca F., 238 

De Maglio G., 281, 282, 342 

De Maria S., 334 

De Martino A., 374 

De Mattia D., 241 

De Miglio M.R., 276 

De Nictolis M., 357 

De Ninno M., 282 

De Pellegrin A., 177 

De Rosa G., 141, 317, 364, 364 

De Santis R., 354 

De Sio A,L 354 

De Stefani S., 248 

De Stefano I., 381 

De Tursi M., 338 

Deantonio L., 380 

Dei Tos P.A., 371 

Del Freo A., 301 

Del Galdo F., 367 

Del Giovane C., 358 

Del Grammastro M., 290, 313, 

378 

Del Sordo R., 246, 283 

Del Vescovo, 283 

Dell’Antonio G., 283 

Della Libera D., 330 

Della Mea V., 213 

Della Ragione C., 330 

Della Rocca C., 285, 342 

Demaglio G., 320 

Demuru A., 305 

Desiderio D., 246, 303 

Dessanti P., 291 

Dessy E., 244, 247, 248 

Destro A., 231 

Di Benedetto A., 154. 

Di Benedetto V., 284 

Di Bernardo A., 195 

Di Blasi A., 333 

Di Bonito L., 177, 338 

Di Carlo E., 361 

Di Cataldo A., 284 

Di Clemente D., 284, 325, 326, 

332, 367 

Di Clemente L., 375 

Di Cristofano C., 285, 342 

Di Gioia G.R.T., 330 

Di Giovannantonio L., 378 

Di Girolamo R., 237 

Di Grazia M., 312 

Di Gregorio C., 245, 337, 346 

Di Maggio M., 341 

Di Mari N., 350, 374 

Di Marzo D., 318 

Di Maso M., 285, 323 

Di Meo S., 361 

Di Napoli A., 286 

Di Napoli M., 177 

Di Naro N., 344 

Di Oto E., 287, 316 

Di Pietro R., 330 

Di Staso M., 375 

Di Tommaso L., 231 

Dias E.P., 309 

Dicandia L., 254, 265 

Diclemente D., 349 

Diegoli M., 168 

Diete M., 157 

Dill M., 369 

Dimitri L., 251 

Discepoli S., 315, 360 

Donati M., 357 

Dono M., 287 

Doria M., 355 

Dudine S., 177 

Duranti E., 340 

Eccher A., 264, 269, 290, 302 

Egarter-Vigl E., 247 

Elisei R., 135 

Elmberger G., 256 

Emery P., 367 

Emiliani R., 357 

Enache M.A., 288 

Enache S.D., 288 

Erra S., 288 

Erra S., 289 

Eusebi L.H., 164, 289, 292, 335 

Eusebi V., 130, 225, 238, 281, 

320, 345, 349, 350 

Faa G., 247, 344 

Fabbretti G., 289 

Fabbri C., 294 

Fabene P., 335 

Facchetti F., 310, 353 

Fadda G., 351 

Faggin R., 296, 297 

Fagotti A., 381 

Falco G., 339 

Falcone U., 360 

Falconieri G., 220, 222, 281, 282, 

342 

Falsirollo F., 328 

Fanburg-Smith J.C., 322 

Fappani L., 310

author indeX 

Faquin W., 310 

Faralli C., 199 

Fardella C., 375 

Farnedi A., 237, 289, 316, 345, 

350 

Fasanella 

283 

Fasanella S., 269, 290 

Fasola G., 281, 282 

Fasolin A., 347, 348 

Fassan M., 226, 296, 371 

Fassina A., 237, 290 

Fattori S., 270 

Faustini-Fustini M., 237 

Fedele F., 262 

Fedeli F., 291 

Federico M., 337 

Felicioni L., 290, 313, 378 

Fenizi G., 334 

Fenocchio D., 203 

Ferdeghini M., 275, 276 

Fernandez L., 382 

Ferrante A., 274 

Ferrara G., 180 

Ferrari A., 286 

Ferrero S., 291, 371 

Ferri I., 246 

Ferri M., 340 

Ferro A., 269 

Ferro P., 291 

Festuccia C., 375 

Feyles E., 342 

Fiandrino G., 185, 196, 311 

Fidelbo M., 369 

Fierabracci A., 276 

Filardo A., 328 

Fileni A., 375 

Filippini M., 348 

Finelli C., 357 

Fiordelisi F., 254 

Fiore G., 284, 325, 326, 332, 349 

Fiore L., 353 

Fiore M.G., 274, 291, 292, 325, 

332, 341 

Fiorentino M., 292 

Fiscon V., 271 

Fisogni S., 310 

Fiumana M., 289 

Flamminio F., 320 

Flamminio F., 292 

Floccari F., 293, 307, 345 

Florena A.M., 371 

Fociani L., 172 

Fodero C., 352 

Foesrster A., 239 

Foli A., 375 

Foli A., 377 

Foltran L., 281 

Fondi C., 293 

Fontana A., 248, 358 

Fontanini G., 237, 261 

Foresto A., 347 

Fornaciari G., 375 

Fornelli A., 294 

Fornelli A., 294 

Forte I., 318 

Fortunato C., 294 

Fortunato L., 268 

Foschini M.P., 237, 243, 278, 

289, 292, 316, 320, 321, 335 

Franceschetti I., 295 

Franceschetti S., 162 

Franceschini M., 291 

Franchi A., 139 

Franco R., 266, 269, 298 

Franco V., 303 

Frank G., 237 

Franzetti-Pellanda A., 320, 380 

Frattini M., 300, 320, 363, 380 

Froio E., 280 

Froio F., 339 

Fucci A., 333 

Fulcheri E., 211, 323, 324 

Fulciniti F., 295, 305 

Fumo R., 352 

Fusconi M., 256 

Fadda G., 375 

Gadducci A., 378 

Gaeta S., 252 

Gafà R., 233, 295, 296, 306 

Gaidano G., 162, 311 

Gaio E., 268 

Galetta D., 318 

Galliani C., 253 

Gallo D., 381 

Gallo P., 330 

Gambarotti M., 269 

Gamucci T., 345 

Ganau M., 344 

Gandolfo S., 263, 279, 315, 316 

Gangemi P., 369 

Garagnani P., 153 

Gardella B., 281 

Gardiman M., 264 

Gardiman M.P., 226, 296, 297 

Gardini G., 137, 339 

Gasparini P., 330 

Gasparoni P., 340 

Gasparre G., 135 

Gatta G., 371 

Gatteschi S., 297 

Gazzaniga P., 342 

Gazzano G., 291 

Gazzola A., 297, 314, 339, 352, 

357 

Genesio R., 382 

Gentile A., 284 

Gessaroli M., 349 

Gessi M., 228, 298 

Geuna E., 131 

Geyer F.C., 345 

Ghimenton C., 302 

Giacalone A., 314 

Giacometti C., 298 

Giallella M., 356, 357 

Giambalvo A., 329 

Gianelli U., 195 

Giangaspero F., 298 

Giannatempo G., 251 

Giannatiempo R., 298, 343 

Giannini A., 355 

Giannini R., 135, 298 

Giannone G., 237 

Gianquinto D., 291 

Giansanti M., 283 

Giantomassi F., 382 

Giardina C., 269, 284 

Giardini R., 195, 299 

Ginanneschi C., 326 

Gioioso A., 305 

Giordano A., 318 

Giordano G., 279, 299, 340, 366 

Giotta F., 243, 338, 339 

Giovanella L., 277, 300 

Giovannini A., 349 

Giovenali P., 203 

Girlando S., 278 

Giubettini M., 286 

Giudici F., 177, 338 

Giuffra V., 375 

Giuffrè G., 300, 301, 303 

Giulini S.M., 244 

Giulioni M., 316 

Giunchi F., 292 

Giurato E., 305 

Giusti A., 301 

Giustiniani M.C., 340 

Gobbato M., 276 

Gobbo S., 359 

Gobbo S., 264, 301, 302, 359, 360 

Gomes R.S., 309 

Gori A., 312 

Gorji N., 291 

Goteri G., 382 

Gradilone A., 342 

Granato F., 239 

Granato R., 309 

Granato R.A., 309 

Grasso G., 305 

Grasso M., 168 

Grasso M.A., 250, 334 

Gravina G.L., 375 

Graziano M., 342 

Greco P., 302 

Grigolato P., 244, 248 

Grillo L., 241 

Grondelli C., 367 

Grosso G., 271 

Grosso M., 303 

Guarnieri V., 322 

Guarnotta C., 371 

Guarrera G.M., 281, 282, 342 

Guastafierro S., 360 

Guerriero A., 341 

Guerriero M., 282 

Guglielmi A., 336 

Guglielmi G., 256 

Gugliotta P., 134 

Guida M., 267, 268 

Guiducci G., 349 

Gulino G., 302 

Gurrera A., 255, 263, 267, 284, 

377 

Gusolfino D., 299 

Guzzardo V., 371 

Guzzetti S., 202, 203 

Guzzinati S., 330, 350 

Hansmann M.L., 379 

Hanspeter E., 319 

Havlat M.F., 260 

Haxhijmeri O., 177 

Hayashi M., 304 

Heim M., 369 

Herdy G.V.H., 309 

Herz M., 319 

Hindi S.A.H., 356 

Hirsch E., 370 

Horiguchi J., 304 

Hysi A., 303, 333, 365 

Iaccarino A., 303, 312, 374, 382 

Iacobellis M., 269 

Iacobone D., 281 

Iannitto E., 197 

Iaria L., 297 

Ieni A., 245, 300, 301, 303, 368, 

369 

Ientile D., 195 

Ientile D., 329 

Ierardi E., 285, 323, 333 

Iezzi M., 278. 354, 370, 303 

Ilardi G., 317, 364 

Inchingolo C.D., 327 

Indovina P., 318 

Ingravallo G., 284, 357, 379 

Innocenti S., 293 

Ioncica A.M., 304 

Ishikawa Y., 304 

Isidori A., 357 

Isidoro E., 177 

Jezzoni C., 367 

Jovine E., 294 

Junior J.A.M., 309 

Kacerovskà D., 255, 267, 302 

Kapoula A. 360 

Kardashi A., 237 

Kasal A., 247, 319 

Katano M., 304 

Kazakov D., 255, 267, 302 

Khadang B., 318 

Kindl S., 196 

Kleinert C., 313 

Koibuchi Y., 304 

387 

La Greca G., 305 

La Provitera A., 353 

La Vecchia F., 295 

La Vecchia F., 305 

Labate A., 303, 305, 306 

Laghi A., 285 

Laginestra A., 314, 357 

Laginestra M.A., 352 

Lai M.L., 344 

Lambros M.B., 345 

Lamovec J., 322 

Lanfranco D., 308 

Lanza F., 348 

Lanza G., 233, 296, 306 

Lanzafame S., 244, 268, 303 

Lanzarini P., 375, 377 

Lanzini M., 278 

Laprovitera A., 353 

Larocca L.M., 316 

Lastilla G., 253 

Laudi C., 346 

Laurino L., 330 

Lauriola L., 298 

Lazzarino M., 311 

Lazzaro D., 354 

Lazzi S., 187, 239, 326, 350 

Lega S., 270, 294, 306, 307, 340 

Leo G., 307, 330 

Leocata P., 315, 360, 379 

Leonardi E., 177, 269, 290, 321 

Leoncini L., 187, 239, 326, 350 

Leone A., 369 

Leone B.E., 263 

Leone G., 284 

Leone O., 165 

Leoni B., 354 

Leoni P., 382 

Leopizzi M., 285, 313, 342 

Lestani M., 245, 347, 358 

Libener R., 308 

Liberati F., 308 

Liberati M., 378 

Liberatore M., 333, 354, 365, 

370 

Licitra L., 371 

Ligorio C., 316 

Liguori G., 279 

Lippolis C., 373 

Lo Giudice C., 340 

Lo Mele M., 330 

Lo Muzio L., 266, 334, 356 

Lo Verde P., 306

388 

Lolli I., 373 

Lombardi M., 308, 366 

Lombardi S., 297, 301 

Lombardi T., 271 

Lonardi S., 310 

Longo F., 255, 309 

Lopes A.C.S., 309 

Lopes V.G.S., 309 

Lopéz C.L., 309 

Lopez-Beltran A., 188, 189 

Lorenzi L., 310 

Lorenzoni A., 310, 312 

Loreti E., 246 

Losi L., 248, 346, 358 

Losito S., 266, 267, 269 

Lovitch S.B., 310 

Lucaccioni A., 271 

Lucianò R., 245 

Lucioni M., 379 

Lucioni M., 185, 196, 311 

Lugli A., 240 

Lunardini A., 375 

Lupi C., 135, 368 

Lupo R., 242 

Luppi G., 248, 358 

Lüthy M., 319 

Luzi P., 317 

Maccio L., 311 

Macrì L., 139 

Macripò G., 349 

Madeddu A., 369 

Maestri A., 370 

Maestri I., 233, 295, 296, 306 

Maestri M., 280 

Maglione A., 343 

Magnani C., 262, 263 

Magri E., 335 

Magro G., 239, 250, 255, 263, 26, 

267, 268, 271, 284, 302, 305, 

309, 362, 369, 377 

Maiello F.M., 245, 353, 354 

Maio V., 312, 355 

Maione M.P., 343 

Maiorana A., 217, 218, 248, 346, 

358 

Maiorano E., 326 

Malachina S., 323, 324 

Malagnino V., 326 

Malapelle U., 246, 277, 303, 312, 

330, 377 

Malatesta S., 290, 313, 338, 378 

Malatesti R., 374 

Malerba S., 307 

Maletta F., 242, 313, 349 

Malfettone A., 261 

Mallone S., 371 

Mameli M.G., 283 

Mammarella C., 290 

Manca A., 276 

Mancini M., 313 

Manfrin E., 265, 328, 329, 336 

Mangia A., 261 

Manni S., 279 

Mannicci D., 296 

Mannu C., 297, 314, 339, 352, 

357 

Mansueto G., 345 

Mantovani V., 153 

Marampon F., 375 

Marangi G., 285 

Marasà L., 314, 372, 373 

Marasà S., 314 

Marasco E., 153 

Marchesini C., 298 

Marchetti A., 290, 313, 321, 378 

Marchetti C., 289 

Marchiò C., 134, 146 

Marchione R., 363, 364 

Marconi P., 335 

Margallo E., 287 

Margiotta G., 314, 315, 360 

Maria R., 312 

Mariani N., 308 

Maricosu E., 276 

Marinaccio M., 324 

Marinelli L., 241 

Marini M., 170 

Mario M., 238 

Mariotti M., 278, 333, 354 

Marra L., 351 

Marseglia M., 269 

Marsico A., 240, 262, 279, 315, 

316 

Martella C., 348 

Martellani F., 177, 338 

Martignoni G., 143, 148, 264, 

265, 271, 276, 291, 295, 301, 

302, 329, 359, 360 

Martin V., 380 

Martini M., 316 

Martuzzi F., 294 

Marucci G., 160, 229, 237, 316 

Marzano A.L., 243 

Marzinotto S., 368 

Marzola A., 335 

Marzullo A., 273, 317, 324, 325, 

379 

Masciullo V., 374 

Mascolo M., 317, 364 

Maselli E., 317 

Masetti M., 294 

Masetti R., 320 

Massarelli G., 276 

Massari R., 338 

Massi D., 293, 310, 312, 354, 355 

Massi., D., 355 

Massucco C., 287 

Mastrogiulio M.G., 317, 329, 374 

Matera L., 318 

Mattioli E., 318 

Mattoni M., 269, 334 

Mazzarelli P., 319, 343, 382 

Mazzarello P., 204 

Mazzatenta D., 237 

Mazzei F., 365 

Mazzer M., 281 

Mazzini C., 354 

Mazzitelli R., 305 

Mazzola P., 363 

Mazzoleni G., 319 

Mazzon E., 306, 305 

Mazzoni E., 289 

Mazzucchelli L., 277, 300, 363 

Mazzucchelli S., 310 

Mazzucco G., 158 

Mazzuchelli L., 320 

McCallum D., 260 

Medici V., 337 

Medicina D., 310 

Melato M., 350 

Melotti F., 219, 224 

Melucci E., 154. 

Meneestrina F., 337 

Menestrina F., 234, 264, 265, 

271, 275, 276, 301, 302, 336, 

360, 380 

Menetti F., 237 

Merighi A., 346 

Merisio C., 299 


Merlini G., 375, 377 

Messerini L., 355 

Messina D., 303 

Mezzetti A., 290 

Micali S., 248 

Miccoli M., 135 

Miccoli P., 135 

Michal M., 255, 267, 302, 322 

Micheletti M., 315 

Miettinen M., 250 

Migliorini P., 297 

Mignogna C., 329 

Milani A., 131 

Milano L., 238 

Milione M., 219, 224 

Minervini M.I., 356 

Minozzi S., 375 

Miracco C., 183, 317, 355 

Miraglia A., 285 

Mitilini N., 330 

Modena S., 288 

Molatore S., 365 

Molina E., 307 

Molinari E., 380 

Molinari F., 320, 380 

Molino A., 328 

Monari E., 248 

Moncelsi S., 375 

Monga G., 158 

Montagna L., 337 

Montalto G., 314 

Monte V., 251 

Montebugnoli L., 289, 321 

Montemurro F., 131 

Montironi R., 188 

Montrone T., 332 

Monzani F., 275, 276 

Morales A.R., 342 

Morandi L., 135, 160, 238, 282, 

289, 320, 321,349, 370 

Morassi F., 374 

Morbini P., 375, 377 

Morelli L., 269, 278, 283, 290, 

336 

Morello F., 370 

Morichetti D., 321, 322, 344, 383 

Morigi F.P., 349 

Moro A., 195 

Moss G., 338 

Mossetti G., 256 

Mottolese M., 154, 366 

Mourmouras V., 239, 317, 329, 

350 

Movilia A., 380 

Munari E., 380 

Mura A., 276 

Murari R., 345 

Muraro L., 244 

Murphy K.M., 342 

Muscarella L.A., 322 

Muscatiello N., 323 

Muscatiello N., 285 

Musiani P., 278, 303, 333, 354, 

361, 365, 370 

Musizzano Y., 211, 323, 324 

Musolino A., 366, 367 

Muti P., 366 

Nakajima H., 304 

Naldoni C., 350 

Napoletano P., 348 

Napoli A., 273, 324, 325, 326, 

367 

Napoli G., 325, 326, 367 

Napoli P., 262 

Napolitano V., 377 

Nappi O., 202, 330 

Nardi F., F., 242 

Naresh K.N., 326 

Nascimento G.H., 309 

Naso G., 342 

Navarro Mora G., 335 

Navone R., 316 

Navone R., 240, 242., 262, 263, 

279, 315 

Negri G.G., 247 

Negrini G., 214 

Nenci I.J., 246 

Nenci I., 295 

Nenna R., 327 

Nesi G., 355 

Nicastro A., 343 

Nicita G., 355 

Nicola M., 185 

Nicolai C., 297 

Nigrisoli E., 358 

Ninfo V., 273, 274 

Nirchio V., 285, 323, 328 

Nitsch L., 382 

Nocita A., 328 

Normanno N., 333 

Nosé V., 300 

Nosé V., 310 

Nottegar A., 328, 329, 336, 265 

Novara F., 196 

Novelli L., 355 

Nozza P., 268, 273 

Nubile M., 278 

Nucifora M., 320 

Nuciforo G., 303 

Nugnes L., 317, 329, 364 

Nuzzo F., 358 

Ober E., 177, 338 

Obici L., 375 

Olla L., 344 

Onorati M., 187, 239, 317, 326, 

329, 350 

Opocher E., 296 

Oppressore D., 343 

Orabona P., 279 

Orecchia S., 308 

Orlandi M., 301, 330 

Orlando C., 310 

Orsaria M., 368 

Ortensi A., 268 

Orvieto E., 330 

Othieno E., 283 

Oyama T., 304 

Pacchioni D., 242 

Pacella E., 323, 324 

Pachera S., 336 

Pagano L., 330 

Paganotti A., 380 

Paglierani M., 355 

Paglierani M., 355 

Palazzo J., 309 

Palena G., 274 

Palladini G., 375, 377 

Palma F., 243, 338 

Palmiotti G., 259 

Palombi N., 333 

Palombini L., 277, 303, 312, 374, 

377, 382 

Palumbieri G., 332 

Palumbo M., 273, 274, 284, 291, 

292, 325, 326, 332, 341, 348, 

349 

Pancione M., 333

author indeX 

Pandolfi P.P., 370 

Panella C., 323 

Panella C., 285 

Paniccià Bonifazi A., 245, 347, 

358 

Panichi D., 270 

Pannellini T., 333, 354, 365 

Panniello G., 255, 258, 334 

Pannone G., 266, 267, 269, 334, 

356 

Panzacchi R., 316, 335 

Paolelli L., 345 

Paolini B., 192, 353, 354 

Paolino S., 330, 335 

Papotti M., 136, 138, 156, 176 

Paradiso A., 261, 366 

Paradiso B., 335 

Parafioriti A., 192, 194, 195, 256 

Parenti R., 239 

Parise G., 347 

Parisi A., 328, 336 

Parisi A., 336 

Parker H.T., 313 

Parolini C., 264 

Parolini S., 310 

Parravicini C., 172 

Parrella P., 322 

Pasini L., 298 

Pasquali D., 267 

Pasquinelli G., 250, 256, 257, 259 

Pastormerlo M., 288 

Paulli M., 185, 196, 311, 376, 377 

Pavesi M., 288, 289 

Pavon I., 330 

Pazzagli M., 354 

Peccetti A., 365 

Pecchioni C., 313 

Pecori S., 336, 337 

Pecori S., 271 

Pede M.R., 345 

Pedica F., 271, 336, 337 

Pedicillo C., 356 

Pedron S., 264, 337, 360 

Pedroni M., 337, 346 

Peer I., 247 

Pelegatti S., 347 

Pelliccioni S. 261, 343 

Pellini F., 329 

Pelosi G., 219, 223, 224 

Penitente E., 338 

Pennacchia I., 241, 381 

Pennella A., 356, 357 

Pennesi M., 292 

Pennesi M.G., 289, 335 

Pentenero M., 263, 279, 315, 316 

Pentimalli F., 318 

Pepi M., 354, 355 

Perfetti A., 335 

Perilongo G., 296, 297 

Perotti G., 272 

Perracchio L., 154, 366 

Perri F., 257 

Perrini P., 270 

Perris R., 239 

Perrone E., 256 

Perrone F., 219 

Perrone G., 254 

Pescarmona E., 270 

Peschiulli L., 293 

Pession A., 160, 287, 320, 370 

Petretto E., 313 

Petriella D., 318 

Petris M., 177, 338 

Petrone G., 375 

Petroni S., 237, 243, 338, 339 

Petrozza V., 285 

Piana S., 137, 339 

Piazzola E., 295 

Pica E., 345 

Piccaluga P.P., 187, 297, 314, 

339, 352, 357, 371 

Piccioli M., 314, 352 

Piccoli P., 336 

Piccolomini M., 363, 364 

Pieraccini L 297 

Pierconti F., 302, 316 

Pieronudo F., 317 

Pierotti P., 340 

Pierotti P., 307 

Pietrini F., 297 

Pietropaoli N., 368 

Pietsch T., 298 

Pilato M., 356 

Pileri P., 187 

Pileri S., 185 

Pileri S.A., 314, 339, 352, 357, 

365 

Pileri S.A., 297 

Pilotti S., 219 

Pilozzi E., 340 

Pinarello A., 340 

Pinto F., 316 

Pinto R., 318 

Pinzani P., 310, 354 

Pirani P., 380 

Pireddu A., 341 

Pirone A., 270 

Pirrelli M., 341, 373 

Pisano A., 330 

Pisanò M., 307 

Piscioli F., 321, 322, 344, 383 

Piscitelli D., 258, 274, 291, 292, 

332, 341 

Pistillo M.P., 291 

Pitino A., 342 

Piubello Q., 330 

Pizzamiglio S., 366 

Pizzi G., 328 

Pizzi S., 243, 271, 308 

Pizzolitto S., 320 

Pizzolitto S., 342 

Pizzolitto S., 220, 222, 281, 282 

Plaiato F., 382 

Platten M.A., 260 

Plesea I.E., 288, 304 

Plesea R.M., 288 

Plutino F.M., 303 

Poccia I., 268 

Podo F., 127 

Poli T., 308 

Polifemo A.M., 294 

Politi E., 360 

Politano M., 238 

Pollini G.P., 328, 329 

Poloni A., 382 

Pompili A., 270 

Ponz de Leon M., 245, 337, 346 

Pop O.T., 288 

Popescu F.C., 288, 304 

Popescu O., 237 

Porta N., 342 

Portolani N., 244 

Postiglione M., 245, 298, 343 

Potenza C., 285 

Pozzani S., 348 

Prandi S., 352 

Priore Oliva C., 337 

Prisco M.G., 381 

Proietti A., 298, 343 

Pucci S., 319, 343, 382 

Pucciarelli S., 346 

Pugliese L., 355 

Puliga G., 344 

Pullara C., 313, 378 

Punzi A., 356, 357 

Pusiol T., 321, 322, 344, 383 

Puzzo L., 302 

Quaresima R., 375 

Quarta G., 345 

Quarto F., 245 

Quattrocch E., 262 

Quero C., 243, 338 

Quilici F., 270 

Ragazzi M., 281, 345, 349 

Raisa G., 348 

Ramieri M.T., 345 

Rapa I., 136 

Rapezzi R., 153 

Raphael M., 326 

Rasi S., 162 

Recupero D., 313 

Reggiani Bonetti L., 245, 248, 

311, 346, 358 

Reghellin D., 245, 328, 347, 347, 

358 

Reis-Filho J.S., 345 

Remo A., 328, 329, 347, 348, 380 

Rendina D., 256 

Resta L., 240, 274, 275, 291, 292, 

326, 332, 341, 348, 349 

Riboni R., 196 

Riboni R., 311 

Riccardo G., 237 

Ricci D., 342 

Riccio P., 362 

Riccioni L., 349 

Ricco R., 273, 325, 326, 367 

Ridolfo A.L., 172 

Riganti F., 339 

Righi A., 349 

Righi A., 237, 242, 349 

Righi L., 156 

Righi S., 297, 314, 339, 352, 357 

Riminucci M., 216 

Rimoldi O., 313 

Rindi G., 243, 351, 374, 381 

Risio M., 164 

Riva A., 320 

Riva C., 142 

Rivasi F., 351 

Rizzardi C., 350 

Rizzo A., 330, 340, 350 

Rizzo P., 293 

Robbins P., 256 

Rocca B.J., 187, 239, 317, 326, 

329, 350 

Rocco A., 299 

Rocco M., 222, 342 

Rogena E., 187 

Romano A., 177, 338 

Romeo G., 135 

Roncalli M., 231 

Roncati L., 217, 218, 351 

Roncella S., 291 

Rosai J., 253 

Rossi A., 355 

Rossi D., 162 

Rossi E.D., 351, 375 

Rossi M., 297, 314, 339, 352, 

357, 375 

Rossi R., 274, 275, 291, 292, 332, 

341, 348 

Rossi S., 246 

389 

Rossini C., 310 

Rostan I., 262, 263, 279, 315, 316 

Rostan M.I., 240 

Rotondo M.I., 237, 261 

Roz E., 303 

Rubbol G., 316 

Rubini C., 356 

Rubini V., 237, 318, 338 

Rubino T., 352 

Rucco V., 245, 347, 358 

Ruco L., 286, 340 

Rufle A., 239 

Rugge M., 296, 297 

Ruggieri M., 268 

Russo A., 312, 343 

Russo P., 375, 377 

Russo R., 279, 280, 352 

Russo S., 273, 326, 353, 354 

Ruzzenente A., 336 

Sabatini F., 354, 365, 370 

Sabatino L., 333 

Sabattini E., 314 

Sabattini E., 297, 339, 352, 357, 

365 

Sabbà C., 284 

Sabino A., 271 

Sacchini A., 317 

Saftoiu A., 304 

Sagramoso C., 314, 352 

Salatiello M., 312 

Salerno A., 210, 306 

Salerno G., 325, 326 

Salerno V., 262 

Saletti P., 300, 320 

Salmaso R., 237 

Salomone E., 303 

Saltarelli S., 361 

Salvati A., 353 

Salvatorelli L., 309, 369 

Salvi S., 291 

Salvia R., 336 

Salvianti F., 354 

Salvio M., 308 

Sampaoli I., 289 

Sandri F., 314, 352 

Sandrini R., 348 

Sandrucci S., 318, 371 

Sanguedolce F., 255, 258, 266, 

267, 334, 356 

Santaquilani M., 371 

Santi R., 355 

Santise G., 356 

Santopietro R., 329 

Santoro A., 266, 267, 269, 334, 

356, 357 

Santoro F., 127 

Santoro L., 375 

Santoro N., 284 

Santucci L., 289 

Santucci M., 293, 310, 312, 354, 

355 

Sapienza M.R., 297, 314, 352, 

357 

Sapino A., 134, 139, 146, 313 

Sarais G., 377 

Sarasin-Filippowicz M., 369 

Sardanelli F., 127, 128 

Sardella B., 285 

Sartori G., 311, 351, 358 

Satolli M.A., 313 

Scamarcio R., 240, 275, 324, 325, 

326, 357, 358 

Scambia G., 374, 381 

Scarabelli L., 248

390 

Scaramuzzi G., 272 

Scarf&igrave R., 301, 303 

Scarfì R., 300 

Scarpa A., 232, 271, 302 

Scarpellini F., 358 

Scarpino S., 286 

Scarselli E., 354 

Scavelli G., 345 

Schena M., 313 

Schiaroli S., 382 

Schiavi F., 290, 371 

Schiavo N., 245, 347, 358 

Schicchi R., 356 

Schirosi L., 311, 358 

Schneider M., 350 

Schneider S., 240 

Sciacca S., 356, 369 

Sciacchitano S., 369 

Sciarrotta M., 313, 378 

Sciarrotta M., 313 

Scibetta N., 371 

Scillitani A., 256 

Scivetti A., 240, 275, 324, 325, 

326, 357, 358 

Seckl M.J., 207 

Segala D., 359 

Segala D., 360 

Segala D., 143, 148, 264, 301, 

302 

Seghetto I., 347 

Senatore S.A., 293, 307, 345 

Senatore 

S.A., 293 

Sensi E., 135 

Sepe J., 353, 354 

Serio G., 259, 317, 356, 357 

Servadio A., 237, 261 

Serviddio G., 258 

Sesenna E., 308 

Shriam R., 203 

Siano M., 317, 364 

Sias S., 309 

Siciliano A., 353 

Sickel J., 250 

Sidoni A., 246, 283 

Sighinolfi M.C., 248, 311 

Silecchia M., 273 

Silini E.M., 243, 272, 280, 281, 

308, 366, 367 

Silva A.C.D., 309 

Silvestri, 283 

Simi L., 310 

Simonato M., 335 

Simone A., 237, 243, 261, 300, 

301, 303, 338, 339 

Simone M.L., 337 

Simone S., 312, 318 

Siniscalchi G., 360 

Siopis E., 351 

Sista M.T., 297, 339 

Skagias L. 360 

Solarino B., 317 

Soliani P., 280 

Sollima L., 360 

Sollitto F., 328 

Sonaglio C., 287 

Sonego F., 268, 330 

Sorrentino C., 361 

Spagnoli L.G., 294, 319, 335, 

343, 382 

Spagnolo D.V., 238, 260 

Spagnolo D., 256 

Spairani C., 342, 364 

Sparano L., 362, 280 

Speciale G., 368 

Spigonardo F., 290 

Spina D., 239, 318, 374 

Spinillo A., 281 

Spitale A., 363 

Squillaci S., 328, 363, 364 

Squillaci S., 342 

Staiano M., 295,, 305 

Staibano S., 317, 364 

Stanta G., 133, 199 

Stark J., 292 

Stella A., 326 

Stigliano E., 241 

Stolfa M., 273 

Stomeo S., 348 

Straci S., 306 

Stramazzotti D., 382 

Stramucci L., 303, 333, 365 

Suriano S., 277, 300 

Tabanelli V., 365 

Tacchini D., 365, 374 

Taffon C., 241 

Talamini A., 380 

Tallarigo F., 328, 364 

Tallini G., 135, 160, 287, 320, 

345, 370 

Tamburini E., 243 

Tancredi A., 272 

Tardanico R., 264 

Tardio M., 257 

Tavani E., 174, 195 

Tavilla A., 371 

Telera S., 270 

Terracciano L., 165, 230, 231 

Terracciano L.M., 239, 240, 369 

Terrenato I., 366 

Testi A., 219 

Thai E., 366, 367 

Tibiletti M.G., 143 

Tinazzi I., 367 

Tinelli C., 280, 308 

Tironi A., 248 

Tito A., 324, 325, 367 

Todaro P., 368 

Todeschini G., 234 

Toffoli G., 159 

Toffoli S., 368 

Tollini F., 380 

Tolu G.A., 247, 344 

Tomasi A., 248 

Tomasini C., 310 

Tombolini V., 375 

Tomezzoli A., 271 

Tommasi S., 318 

Tondini M., 364 

Tonutti M., 177, 338 

Toraldo M., 368 

Torelli L., 177, 338, 350 

Tornaboni D., 301 


Tornillo L., 239, 240, 369 

Torregrossa L., 135, 298 

Torrisi A., 239, 268, 305, 369 

Tortorella S., 356 

Tosi A.L., 370 

Toto V., 333, 370 

Tozzini S., 297, 301 

Trabucco S., 259 

Trama A., 371 

Tranchima M.G., 369 

Traversi C., 240, 275, 358 

Traverso V., 323, 324 

Tricarico P., 290, 371 

Trincheri N., 308 

Trinchero E., 178 

Tripodo C., 197, 371 

Troiano A., 272 

Trombetta C., 245 

Troncone G., 147, 277, 303, 312, 

246, 330, 374, 377 

Truini M., 287, 291 

Tuccari G., 300, 301, 303, 368 

Tugnoli Pàttaro S., 206 

Turchetti D., 370 

Turrisi M.A., 356 

Ucchino S., 290 

Uccini S., 286 

Ugo F., 308 

Ugolini C., 343 

Ulazzi L., 233, 295, 296, 306 

Ungari M., 198 

Unti E., 371 

Uras M.G., 276 

Urso C., 355 

Urso E., 346 

Vago G.L., 172 

Vairo M., 257, 362 

Valcavi R., 339 

Valentini A.M., 341, 373 

Valentini M., 357 

Valerio L., 241 

Valli R., 365 

Van Eeckhout P., 373 

Varela-Carver A., 313 

Varenna M., 256 

Varone V., 246, 277, 374 

Vasori., S., 329 

Vasquez E., 263, 284, 305, 369 

Vassallo L., 365, 374 

Vecchio F.M., 241 

Vecchio G., 267 

Vecchio G.M., 239, 271, 305, 

309, 369, 377 

Vecchio F.M., 241 

Vecchione A., 133 

Vecchione M.L., 317, 364 

Vellone V.G., 351, 374, 375, 381 

Vendraminelli R., 348, 380 

Vendraminelli V., 347 

Venesio T., 346 

Ventura L., 308, 375 

Verderio P., 366 

Verdun di Cantogno L., 134, 313, 

349 

Verga L., 375, 377 

Vergine M., 265 

Vermi W., 310 

Vetrani A., 377 

Viale G., 337 

Vianelli N., 357 

Viel A., 346 

Vigani A., 291 

Vigliar E., 307, 382 

Villari D., 355 

Villari L., 303, 377 

Villela S.H., 309 

Vincenzo A., 241 

Vindigni C., 172, 238, 329, 355 

Viola P., 313, 338, 378 

Visani G., 357 

Visani M., 320, 282 

Visca E., 351 

Vita G., 265, 378 

Vitale A.R., 314, 379 

Vitali P., 358 

Vitarelli E., 245 

Viti R., 256 

Vittadello F., 247 

Vitti P., 135 

Vlajnic T., 240 

Volante M., 136, 156, 176 

Volante., M., 138 

Volta U., 174 

Waha A., 298 

Went P., 297, 339 

Wong D.D., 260 

Xiaoyun L., 370 

Zaccaria M., 379 

Zacchi A., 177, 338 

Zachara E., 330 

Zagami M., 342 

Zamò A., 234, 380 

Zanconati F., 177 

Zandonà L., 177 

Zanella C., 347, 348, 380 

Zanellato E., 320, 380 

Zanier L., 368 

Zanin E., 298 

Zaninelli M., 348 

Zanini N., 294 

Zannoni G.F., 351, 374, 375, 

381 

Zanotti R., 234 

Zardo G., 340 

Zelante L., 322 

Zenone Bragotti L., 377 

Zeppa P., 377, 382 

Zerbini M., 153 

Zidar N., 322 

Zinzani P.L., 297 

Zironi S., 248 

Zizzi A., 382 

Zlobec I., 240 

Zolfanelli F., 355 

Zonetti M.J., 319, 343, 382 

Zorzi M.G., 383 

Zucchini S., 335 

Zuffardi O., 196 

Zuliani M., 298 

Zupo S., 287

Issue 4 - August 2010 - Pacini Editore

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