12th Congress of the European Hematology ... - Haematologica

0969
COMPARISON OF CD25 IMMUNOHISTOCHEMISTRY AND CD25 FLOW CYTOMETRY
IN SYSTEMIC MASTOCYTOSIS
Ch. Baumgartner, M.-T. Krauth, K. Sonneck, S. Florian, M. Födinger,
A.W. Hauswirth, L. Müllauer, P. Valent
Medical University of Vienna, VIENNA, Austria
Background. Systemic mastocytosis (SM) is a clonal myeloid disorder
characterized by an abnormal accumulation and growth of mast cells
(MC) and their progenitors in one or more extracutaneous organs. In
most cases, the bone marrow is involved and thus is examined in suspected
disease. Expression of CD25 in bone marrow MC, with or without
co-expression of CD2, is an important minor criterion of systemic
mastocytosis. So far, most studies have examined CD25 expression on
MC by fow cytometry. Methods. We examined the expression of CD25
by bone marrow MC in patients with SM (n=30) by immunohistochemistry
(IHC) and compared these results with those obtained from flow
cytometric assessement of CD25 expression. Results. In the majority of
all patients (27/30; 90%), CD25 was detectable in MC by both staining
techniques. In one patient, CD25 was only detectable by IHC, but not
by flow cytometry, whereas in 2 patients, in whom no compact MC
infiltrates were detectable in the bone marrow, only the flow cytometric
evaluation revealed aberrant expression of CD25 in MC. Conclusions.
CD25 IHC is equally diagnostic and sensitive in SM compared to flow
cytometry and thus can be recommended as diagnostic test to document
the phenotype-related minor SM criterion. Our data also suggest,
however, that optimal assessment of CD25 expression in neoplastic MC
in all patients requires the application of both techniques, i.e. IHC and
flow cytometry.
0970
SELF-RENEWAL CAPACITY OF STROMAL PRECURSOR CELL IN MESENCHYMAL STEM
CELLS HIERARCHY IN MICE
N. Drize, I. Nifontova, L. Chertkov
National Reseach Center for Hematology, MOSCOW, San Marino
Background. Adult mesenchymal stem cell (MSC) as any other stem cell
is capable for self-renewal and differentiation. In the mice bone marrow
(BM) there are cells able to create hematopoietic microenvironment de
novo after transplantation of the BM plug under the renal capsule of syngeneic
animal. In hematopoietic ectopic foci formed 6 weeks after the
transplantation the stromal cells belong to a donor of BM and
hematopoietic cells are of recipient origin. The capability of these cells
to differentiate within all stromal cell types forming functional
hematopoietic microenvironment completely fits stem cells differentiation
criteria. MSC could be retransplanted 9 times without changes of
foci size formed de novo. It is possible to calculate the approximate number
of MSC per femur which comes to 5-10 per 106 BM cells. In irradiated
recipients the size of the foci enlarged significantly, but this phenomenon
is not due to MSC number increase as this effect doesn’t apply
to the next retransplantation to the non-irradiated recipients. So the category
of precursor cells, capable to differentiate to the cells of functional
stromal microenvironment, but not capable of self-renewal exists.
Aims. The position of CFU-F in hierarchy of MSC is still obscure. The aim
of the study was to investigate the capability of CFU-F to transfer
hematopoietic microenvironment. Methods. Çone marrow plugs were
transplanted under the renal capsule of syngeneic mice. In 6 weeks the
foci formed were either retransplanted under the renal capsule of secondary
recipients or nucleated cells in the foci were resuspended and counted.
CFU-F frequency was measured by standard protocol. Cloning was
performed using 3 concentrations of BM cells (30000, 40000 and 50000
cells per well of 96-well plate). Clones of fibroblasts were passaged to
24-wells plate, than to 6-well and finally to 25 cm2 flask in media containing
20% FCS and 5 ng/mL bFGF. The cells from the flask were transplanted
under the renal capsule both of non-irradiated and irradiated
mice. Results. CFU-F frequency in murine BM is 47,6±1,2 per 106 cells if
fider of irradiated guinea pig BM cells was added to system and 36,2±9,5
per 106 when the media with 5 ng/mL bFGF was used. Limiting dilution
analysis showed that the frequency of CFU-F equals 1 per 30000 cells,
in flasks it was estimated as 1 per 21000 cells on fider cells and 1 per
27000 cells in bFGF presence. CFU-F possesses limited proliferative
potential. Hundred percent of clones had grown up in 24-well plate (i.e.
5 mitoses were performed), 60% filled the well of 6-well plate (two
more mitoses) and only 45% reached the square of the flask (i.e. could
undergo 9 mitoses). After transplantation of these layers under the renal
capsule of both non-irradiated and irradiated syngeneic mice no ectopic
12 th Congress of the European Hematology Association
foci developed. So progeny of CFU-F are not able to transfer hematopoietic
microenvironment. Conclusions. The data suggest that CFU-F has
limited proliferative capacity and do not contain the MSC. CFU-F position
in MSC hierarchy seems to be lower than cells capable to develop
large foci in irradiated recipients.
0971
SPONTANEOUS FACTOR V AUTOANTIBODY-A CASE REPORT AND SYSTEMIC REVIEW
OF ITS NATURAL HISTORY AND MANAGEMENT
A.L. Ang, H.J. Ng
Singapore General Hospital, SINGAPORE, Singapore
Background. Factor V(FV) inhibitors are infrequently encountered. They
often arise as alloantibodies after topical bovine thrombin exposure, or
may develop in congenital FV deficiency patients after fresh frozen plasma(FFP)
transfusion. Less commonly, they are autoantibodies that develop
spontaneously in non-hemophiliac patients. FV autoantibody tends
to be associated with more serious haemorrhagic complications compared
to alloantibody associated with bovine thrombin exposure.
Aims.We report the spontaneous development of FV autoantibody in an
elderly lady who presented with melaena. We also performed a systematic
review of published cases of spontaneous FV autoantibody and our
patient, to attain a better understanding of the patient characteristics, natural
history and management of this condition. Methods. Published cases
of spontaneous FV autoantibody from 1950 till December 2006 were
searched with no language restriction on Ovid MEDLINE. Additional
articles were identified by reviewing reference lists. Cases associated
with topical thrombin exposure and congenital FV deficiencies were
excluded. Analysis was done on 71 cases(including our patient) with
available information. Results. A 92-year old female with diabetes mellitus
and dementia presented with haemetemesis and melaena. She was
usually on tolbutamide, chlorpromazine, carbamazepine and haloperidol.
Her prothrombin(PT) and activated partial thromboplastin
time(aPTT) were >100secs, and were not correctable on 1:1 plasma mixing
studies. There was no demonstrable lupus anticoagulant(LA) activity.
Her FV level was 900) days in all treated
and untreated patients. 48 patients(67.6%) had bleeding complications
haematologica/the hematology journal | 2007; 92(s1) | 361