Sabato 27 ottobre 2012 - Pacini Editore

patHOlOGiCa 2012;104:267-358 COMUnICAzIOnI OrALI
BIOLOGIA MOLECOLArE
The alteration of lipid metabolism in Burkitt
lymphoma identifies a novel diagnostic marker:
the adipophilin
M.R. Ambrosio1 , P.P. Piccaluga2 , M. Ponzoni3 , B.J. Rocca1 ,
V. Malagnino1 , M. Onorati1 , G. De Falco1 ,V. Calbi4 , M. Ogwang4 ,
K.N. Naresh5 , S.A. Pileri2 , C. Doglioni3 , L. Leoncini1 , S. Lazzi1 1 Department of Human Pathology and Oncology, Anatomic Pathology
Section University of Siena, Italy; 2 Molecular Pathology Laboratory, Haematopathology
Unit, Department of Haematology and Oncology “L. and
A. Seràgnoli”, S. Orsola-Malpighi Hospital, University of Bologna, Italy;
3 Pathology Unit, Department of Oncology, University Scientific Institute
San Raffaele, Milan, Italy; 4 Saint Mary Hospital, Lacor Gulu, Uguanda;
5 Department of Histopathology, Hammersmith Hospital Campus, Imperial
College, London UK
Background. Burkitt lymphoma (BL) is listed in the WHO classification
of lymphoid tumours as an ‘aggressive B-cell non-Hodgkin
lymphoma’. None of the histological, immunohistochemical
or molecular parameters can be singly used for the diagnosis of
BL and the WHO classification suggests that a combination of
several techniques is necessary. BL has very characteristic cytologic
features on fine needle aspiration cytology (FNAC) where the
identification of the typical cytoplasmic vacuoles in the lymphoid
cells represents a diagnostic hallmark. These vacuoles containing
lipids cannot be seen on histological preparations. The formation,
maintenance, modification and involution of lipid droplets is crucially
regulated by a family of lipid droplet- associated proteins
(PAT-proteins), to which belong five members, usually referred to
as PLIN proteins. The presence of lipid vacuoles in the cytoplasm
may suggest that biosynthesis of lipids and other macromolecules
is altered in BL as there is increasing evidence of lipid metabolism
reprogramming in tumour cells. This study was designed to analyse
the lipid metabolism in BL.
Materials and methods. A total of sixty formalin-fixed and
paraffin-embedded specimens were investigated at the Department
of Human Pathology and Oncology, Anatomic Pathologic
Section, University of Siena, Italy. The initial diagnosis of these
cases was: BL in 43, diffuse large B-cell lymphoma (DLBCL)
in 30 and B-cell lymphoma unclassifiable with features intermediate
between DLBCL and BL (DLBCL/BL) in 7. The slides
were reviewed by two expert haematopathologists and classified
according to the scoring system recently designed by Naresh
et al. for aggressive B-cell lymphomas which distinguishes
BL and not BL. According to the algorithm, we identified 47
BL and 33 not BL. Histological sections were stained with
adipophilin antibody and the pattern of immunostaining as well
as the labelling intensity was evaluated. The statistical association
between the distribution of expression of adipophilin and
the diagnostic category (BL and not BL) was evaluated, with
P < 0.05 considered as being statistically significant. Moreover,
we analyzed Gene expression profiling (GEP) data of 13 BL
and 20 DLBCL, focusing on the expression of ADFP (PLIN2),
and other genes related to lipid metabolism (SCD, SCD5, FASN,
USF1, PPARA).
Results. We found adipophilin as the only member of the PATprotein
family expressed in BL. Moreover, supervised analysis
revealed that 5 other genes involved in lipid metabolism were
differentially expressed (fold change > 2; p < 0.05) in BL and
DLBCL. While SCD and PPARA, were up-regulated in DL-
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BCLs, SCD5, FASN and USF1 were up-regulated in BL. To
confirm GEP results, adipophilin expression was investigated
by immunohistochemistry in BL and not-BL cases. A strong immunoreactivity,
characterized by single or multiple droplets in
the cytoplasm and clustering of these to the outer nuclear membrane,
was observed in 45 out of 47 BL cases. Weak positivity
characterized by dispersed fine lipid droplets in the cytoplasm
of a small minority of cells was detected in 3 out of 33 cases
of not-BL. 30 of 33 not-BL cases did not show any expression
of adipophilin. Macrophages showed a granular staining within
the cytoplasm, and this served as an internal positive control.
The proportion of cases positive for adipophilin expression
was significantly higher (p < 0.05) in BL cases than the not-
BL cases. Interestingly, the three cases of not-BL category that
showed weak, fine positivity were characterized by a diffuse
proliferation of medium- to large-sized cells with irregular
nuclear contours and relatively large nucleoli. Starry-sky macrophages,
mitoses and apoptosis were prominent, and reactive
small lymphocytes were scanty. This morphological features
corresponded to score 1 (range: 0-3) on the algorithm for aggressive
B-cell lymphomas proposed by Naresh et al. and may
well represent B cell lymphoma, unclassifiable with features
intermediate between DLBCL and BL.
Conclusions. Our results suggest that accumulation of lipid
vacuoles is due to an altered lipid metabolism in BL and may
reflect a metabolic phenotype promoting tumourigenesis. Rapidly
dividing tumour cells require rapid generation and release
of adipophilin droplets. The increased number of adipophilin
lipid droplets may thus be related to the high proliferation rate
of this tumour, the fastest growing tumour in humans. Lipid
vacuoles may be specifically recognized by a monoclonal
antibody against adipophilin, which may therefore be a useful
marker for the diagnosis of BL because of its peculiar
expression pattern. This peptide might represent an interesting
candidate for interventional strategies (e.g. target therapy or
vaccine therapy), in fact, a recent preliminary study has investigated
the possible use of adipophilin as a T-cell epitope to
induce antigen-specific cytotoxic T-lymphocytes and mediate
tumour cell lysis.
A better knowledge of lipid metabolism alteration in BL can
potentially provide new markers to improve diagnosis and prognosis
as well as novel therapeutic approaches for BL treatment.
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