Expression of Epstein-Barr virus-encoded small RNA (by the EBER-1 gene) in liver specimens from transplant recipients with post-transplantation lymphoproliferative disease

Abstract

Background: Epstein-Barr virus (EBV)-associated post-transplantation lymphoproliferative disease (PTLD) develops in 1 to 10 percent of transplant recipients, in whom it can be treated by a reduction in the level of immunosuppression. We postulated that the tissue expression of the small RNA transcribed by the EBER-1 gene during latent EBV infection would identify patients at risk for PTLD.

Methods: We studied EBER-1 gene expression in liver specimens obtained from 24 patients 2 days to 22 months before the development of PTLD, using in situ hybridization with an oligonucleotide probe. Control specimens were obtained from 20 recipients of allografts with signs of injury due to organ retrieval, acute graft rejection, or viral hepatitis in whom PTLD had not developed 9 to 71 months after the biopsy.

Results: Of the 24 patients with PTLD, 17 (71 percent) had specimens in which 1 to 40 percent of mononuclear cells were positive for the EBER-1 gene. In addition, 10 of these 17 patients (59 percent) had specimens with histopathological changes suggestive of EBV hepatitis. In every case, EBER-1-positive cells were found within the lymphoproliferative lesions identified at autopsy. Only 2 of the 20 controls (10 percent) had specimens with EBER-1-positive cells (P < 0.001), and such cells were rare.

Conclusions: EBER-1 gene expression in liver tissue precedes the occurrence of clinical and histologic PTLD. The possibility of identifying patients at risk by the method we describe here and preventing the occurrence of PTLD by a timely reduction of immunosuppression needs to be addressed by future prospective studies.

Figure 1

Lymphocytic Infiltration of the Hepatic Portal Tracts by Mononuclear cells Positive for EBER-1 mRNA in a Patient with EBV-Associated Hepatitis. Panel A (×210) shows scattered EBER-1–positive nuclei in approximately 20 percent of the lymphocytes infiltrating the portal tract. Panel B (hematoxylin and eosin, ×210) shows an infiltrate of activated lymphocytes involving the portal vein and simulating graft rejection.

Figure 2

Lymphocytic Infiltration of the Hepatic Sinusoids by EBER-1–Positive Cells in a Patient with EBV-Associated Hepatitis. In Panel A (×210), the sinusoids contain aggregates of lymphocytes, and the hepatocytes show focal hydropic swelling, steatosis, and nuclear pyknosis. In Panel B (×210), the sinusoids also contain EBER-1–positive lymphocytes.

Figure 3

PTLD Involving a Portal Tract (× 420). Approximately 70 percent of the cells are EBER-1–positive, with both the small round lymphocytes and the large immature lymphocytes affected. The bile-duct epithelium in the center of the field is stained with anti–Cam 5.2 antibody and does not show evidence of EBV infection.

Figure 4

Fatal PTLD Involving Hepatocytes (×630). Rare EBER-1–positive nuclei were found In cells conclusively identified as hepatocytes, by simultaneous immunohistochemical evaluation with the anti-keratin antibody Cam 5. 2.

Figure 5

EBV-Infected Lymphocytes in the Lesions of PTLD (×420). Immunohistochemical evaluation with anti-CD20 antibody, combined with in situ hybridization, showed that in PTLD the majority of EBV-infected lymphocytes are B cells. Cells positive for CD20 are stained brown, and those positive for EBER-1 mRNA are stained blue-black.