Hepatosplenic γδ T-cell lymphoma associated with hepatitis B virus infection

Hepatosplenic γδ T-cell lymphoma associated with hepatitis B virus infection

Shuji Ozaki^a, Koji Ogasahara^a, Masaaki Kosaka^a, Toshi Inoshita^a, Shingo Wakatsuki^b,
Hisanori Uehara^c and Toshio Matsumoto^a

^aFirst Department of Internal Medicine,^bFirst Department of Pathology, and ^cSecond Department of Pathology, The University of Tokushima School of Medicine, Tokushima, Japan.

Abstract:Hepatitis B virus (HBV) infection has been implicated in the development of hepatocellular and hematopoietic malignancies. We describe a patient with chronic hepatitis B who developed hepatosplenic γδ T-cell lymphoma. A45-year-old woman presented with marked hepatosplenomegaly and hepatic failure during the course of chronic hepatitis B. Peripheral blood examination revealed57%abnormal lymphoid cells which expressed the γδ T-cell receptor. The cytogenetic analysis of tumor cells showed an abnormal karyotype;47, XX, -13, +2mar in all 20metaphases examined. A clonal rearrangement of the T-cell receptor genes was demonstrated by Southern blot analysis, showing monoclonal expansion of tumor cells. A liver biopsy specimen showed fibrosis of the portal areas and sinusoidal infiltration of tumor cells. HBV infection was documented by the presence of IgG anti-HBc and anti-HBs antibodies in serum. Although HBV-DNA was not detected in tumor cells by polymerase chain reaction analysis, there is a possibility that proliferation of γδ T cells in response to HBV infection played a role in the pathogenesis of hepatosplenic γδ T-cell lymphoma. J. Med. Invest. 44:215-217, 1998

Keywords:hepatosplenic lymphoma, γδ T-cell, hepatitis B virus, chronic hepatitis, hepatosplenomegaly

INTRODUCTION
Hepatosplenic γδ T-cell lymphoma is a rare and aggressive T-cell malignancy characterized by the neoplastic proliferation of γδ T cells in the hepatic sinusoids and splenic red pulp (1, 2). An increased number of γδ T cells has also been reported in the hepatic sinusoids of patients with chronic viral hepatitis (3). Human γδ T cells exhibit preferential homing to the sinusoidal areas of lymphoid tissues and can mediate cellular immune functions in response to antigenic stimulation (4). However, the exact mechanism of the proliferation of these cells remains unknown.
Hepatitis B virus (HBV) infection has been implicated in the development of hepatocellular carcinoma (5). Moreover, HBV has been reported to play a pathogenic role in some cases of hematopoietic and lymphoid malignancies (6, 7). Although the role of HBV in the development of malignancies has not been clarified, integration of HBV-DNA into the cellular DNA of neoplasm is frequently found and this has been postulated as essential to HBV-related oncogenesis because of transcriptional activation function (8).
We report a case of hepatosplenic γδ T-cell lymphoma associated with chronic B hepatitis. Although HBV-DNA was absent in lymphoma cells of this patient, proliferation of γδ T cells by an immune response to HBV infection might have played a role in the development of hepatosplenic γδ T-cell lymphoma.

CASE REPORT
A 45-year-old woman had suffered from liver dys-function since 1990. Although the opportunity of HBV infection was not known, a diagnosis of chronic hepatitis B was made on the basis of the clinical course and antibody responses against HBV. She was subsequently lost to follow-up for almost 6 years and there was no data on serological examination. In 1996 she presented with jaundice and abdominal fullness. Physical examination revealed ascites and marked hepatosplenomegaly associated with no lymphadenopathy. The hemoglobin level was10.8g/dl, platelet count39x109/l and white blood cell count 9.3x109/l with 57% abnormal lymphoid cells showing irregular nuclei with condensed chromatin. The lymphoid cells from the peripheral blood expressed CD3,CD7 and the γδ T-cell receptor detected by TCRδ1 (T Cell Sciences, Cambridge, MA), but not the αβ T-cell receptor detected by TCR1 (Becton Dickinson, San Jose, CA). Other T-cell antigens (CD1, CD2, CD4, CD5, CD8, CD25), NK-cell antigens (CD16, CD‚T6, CD57), B-cell antigens (CD19, CD20) or HLA-DR were negative. The cytogenetic analysis of the tumor cells showed an abnormal karyotype;47, XX, -13, +2mar in all 20 metaphases examined. A clonal rearrangement of the T-cell receptor (TCR)-β, -γ, -δ chain genes was demonstrated but the immunoglobulin heavy chain (IgH) gene was in a germ line state by Southern blot analysis (Fig1). Serum aspartate aminotransferase was 856IU/l, alanine aminotransferase1056IU/l, alkaline phosphatase 447IU/l, lactate dehydrogenase 1254IU/l, total bilirubin 10.1x10-2g/l, albumin 32g/l, ammonia1.35x10-3g/l and soluble interleukin (IL)-2receptor 4060U/ml (normal range187.6-637.7). Alpha-fetoprotein and carcino-embryonic antigen were not elevated. Serological tests for human T-cell lymphotropic virus type I and hepatitis C virus (HCV) were negative. Hepatitis B surface antigen was negative. HBV infection was documented by the presence of IgG anti-HBc and anti-HBs antibodies in serum. To explore the possibility that lymphoma cells may become infected with HBV, integration of HBV-DNA was examined by polymerase chain reaction analysis as described previously (9). However, HBV-DNA was not detected in tumor cells. Computed tomography showed an enlarged liver and spleen and peritoneal fluid. No malignant cells were found in ascites. Histological features of the bone marrow showed an infiltration of lymphoma cells. A liver biopsy specimen showed fibrosis of the portal areas and sinusoidal infiltration of tumor cells (Fig2). These cells were medium-sized lymphocytes with a regular or slightly indented nucleus, and positive for CD3and CD45RO (UCHL-1) while negative for CD20 (L-26) by immunohistochemistry (data not shown). These findings were compatible with those of hepatosplenic γδ T-cell lymphoma and chronic hepatitis. Chemotherapy was started, but shortly after she developed fungal meningitis and died. Examination at autopsy revealed lymphoid infiltration in the liver, spleen, kidney, lung, bone marrow, anterior mediastinum and esophagus. The spleen weighed 500g and the red pulp was massively infiltrated by medium-sized neoplastic cells.

DISCUSSION
We report a case of hepatosplenic γδ T-cell lymphoma in leukemic phase associated with HBV infection. Aggravation of liver functions in this case was considered as a rapid progression of chronic hepatitis B until the proliferation of γδ T cells was found in the peripheral blood and liver. Clonal expansion of neoplastic cells was also demonstrated by cytogenetic and genotypic studies. However, the pathological significance of this chromo-some abnormality is unknown and its possible role in the pathogenesis of γδ T-cell lymphoma remains to be elucidated. To our best knowledge, there was no case report of hepatosplenic γδ T-cell lymphoma associated with chronic hepatitis or HBV infection.
Because HBV is both a hepatotropic and lymphotropic virus, the possible pathogenic relationship between hepatitis B and malignant lymphoma has been suggested. Talamo et al. have reported an autopsy case of primary hepatic lymphoma in a patient with hepatocellular carcinoma associated with HBV infection and speculated that HBV could have been the common cause of both neoplasms in the liver (10). In addition, Pontisso et al have reported the detection of HBV-DNA in bone marrow cells of leukemia patients (6). In contrast, HBV gene products have been identified in endothelial cells of the tumor tissues of leukemia and lymphoma patients, suggesting that HBV-infected endothelial cells support and stimulate the tumor growth by producing cytokines (7). However, we were unable to demonstrate HBV-DNA in tumor cells by polymerase chain reaction analysis and there is no evidence to suggest a direct role of HBV in the development of hepatosplenic γδ T-cell lymphoma in the present patient.
Nevertheless, histological findings showed fibrosis of the portal areas, indicating a chronic inflammatory change of the liver in this case. Primary lymphomas may arise as a monoclonal outgrowth from a pool of proliferating polyclonal lymphocytes involved in a chronic inflammatory process. An increased number of γδ T cells together with CD3+ lymphocyte infiltration has been reported in the hepatic sinusoids of chronic viral hepatitis(3). Thus, HBV infection in the liver may provide a chronic stimulation inciting a γδ T-cell growth. Alternatively, Bukowski et al have reported the expansion of γδ T cells with the cytotoxic activity in response to virus-infected cells (11). This finding suggests that γδ T-cell malignancy may result from monoclonal expansion of γδ T cells with defined antigen specificity directed against virus-infected targets. Taken together, despite the absence of HBV-DNA in neoplastic γδ T cells, there is a possibility that the γδ T-cell expansion in response to HBV caused an evolution of neoplastic γδ T cells. Further investigations are required to elucidate the relationship between HBV infection and γδ T-cell malignancy.

ACKNOWLEDGMENT
The authors thank Otsuka Assay Laboratories, Otsuka Pharmaceutical Co. Ltd (Tokushima, Japan) for the help in analyzing HBV-DNA.

REFERENCES
1. Farcet JP, Gaulard P, Marolleau JP, Le Couedic JP, Henni T, Gourdin MF, Divine M, Haioun C, Zafrani S, Goossens M, Hercend T, Reyes F:Hepatosplenic T-cell lymphoma:sinusal/sinusoidal localization of malignant cells expressing the T-cell receptor γδ. Blood75:2213-2219, 1990
2. Cooke CB, Krenacs L, Stetler-Stevenson M, Greiner TC, Raffeld M, Kingma DW, Abruzzo L, Frantz C, Kaviani M, Jaffe ES:Hepatosplenic T-cell lymphoma: a distinct clinicopathologic entity of cytotoxic γδ T-cell origin. Blood88:4265-4274, 1996
3. Kanayama K, Morise K, Nagura H : Immunohistochemical study of T cell receptor γδ cells in chronic liver disease. Am J Gastroenterol87:1018-1022, 1992
4. Chien Y-H, Jores R, Crowley MP:Recognition by γ/δ T cells. Annu Rev Immunol14:511-532, 1996
5. Tiollais P, Pourcel C, Dejean A : The hepatitis B virus. Nature317:489-495, 1985
6. Pontisso P, Locasciulli A, Schiavon E, Cattoretti G, Schiro R, Stenico D, Alberti A:Detection of hepatitis B virus DNA sequences in bone marrow of children with leukemia. Cancer59:292-296, 1987
7. Galun E, Ilan Y, Livni N, Ketzinel M, Nahor O, Pizov G, Nagler A, Eid A, Rivkind A, Laster M, Ron N, Blum HE, Shouval D:Hepatitis B virus infection associated with hematopoietic tumors. Am J Pathol145:1001-1007, 1994
8. Spandau DF, Lee CH : Trans-activation of viral enhancers by the hepatitis B virus X protein. J Virol62:427-434, 1988
9. Matsubara K, Tokino T:Integration of hepatitis B virus DNA and its implications for hepatocarcinogenesis. Mol Biol Med7:243-260, 1990
10. Talamo TS, Dekker A, Gurecki J, Singh G:Primary hepatic malignant lymphoma:its occurrence in a patient with chronic active hepatitis, cirrhosis, and hepatocellular carcinoma associated with hepatitis B viral infection. Cancer46:336-339, 1980
11. Bukowski JF, Morita CT, Brenner MB:Recognition and destruction of virus-infected cells by human γδ CTL. J Immunol153:5133-5140, 1994

Received for publication December 2, 1997 ; accepted January 5, 1998.

1 Address correspondence and reprint requests to Shuji Ozaki, M.D., Ph.D., First Department of Internal Medicine, The University of Tokushima School of Medicine, Kuramoto-cho, Tokushima770-8503, Japan and Fax:+81-886-33-7121.