Presence of Epstein‐Barr virus harbouring small and intermediate‐sized cells in Hodgkin'S disease. Is there a relationship with Reed‐Sternberg cells?

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Abstract
Forty‐four cases of Hodgkin's disease (HD), mostly of the nodular sclerosing type, were investigated for the presence of Epstein‐Barr virus (EBV) by polymerase chain reaction (PCR) and DNA and RNA in situ hybridization (DISH, RISH), as well as by immunohistochemistry for the detection of latent membrane protein‐1 (LMP‐1) of EBV. in situ hybridization (ISH) was combined with immunohistochemistry to correlate the presence and activity of the virus at the cellular level. In 18/34 (53 per cent) cases, EBV‐DNA sequences could be detected with the PCR method. In 12/18 positive cases, DISH and RISH were also positive. In the remaining six EBV‐PCR positive cases, two were also positive with RISH and LMP‐1, whereas no positive signal with DISH could be obtained. All DISH and/or RISH positive cases were also positive for LMP‐1. With RISH, not only the Reed‐Sternberg cells and their mononuclear variants (RS cells) stained positive, but also small and intermediate cells frequently reacted with the EBV‐specific probes (EBER‐1 and ‐2). Double staining with cellular markers (CD3, CD20, CD45, CD45RO, CD68, and the lectin PNA) revealed that most of the smaller EBER‐positive cells frequently did not express T, B, or histiocytic markers, but that they, as well as the RS cells, showed cytoplasmic and membranous staining with PNA. These smaller EBER‐positive cells were not found in EBV‐PCR negative HD. EBER‐positive RS cells were almost always LMP‐1 positive, as well as a substantial proportion of the intermediate‐sized cells, whereas the majority of the small EBER‐positive cells remained LMP‐1 negative. In EBV‐PCR positive non‐malignant lymph nodes, only a few EBER‐1 and ‐2 positive cells could be observed. As in infectious mononucleosis, these cells frequently expressed the B‐cell marker CD20. Although we cannot exclude the fact that the majority of the smaller EBV‐positive cells in HD belong to reactive EBV‐infected lymphocytes, our data favour the hypothesis that at least some of these smaller cells may belong to the reservoir of neoplastic cells in HD.

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