Assessment of clonality of rosetting T lymphocytes in Hodgkin's disease bysingle-cell polymerase chain reaction: detection of clonality in a polyclonal background in a case of lymphocyte predominance Hodgkin's disease

Rosetting of CD4+ T cells around the neoplastic Hodgkin and Reed-Sternberg (H&RS) cells is a characteristic feature of Hodgkin's disease (HD). To answ er the question whether this phenomenon is solely due to chemokine-mediated attraction of T cells or whether the rosetting T cells in addition recogni ze antigens presented by the H&RS cells, we examined the T cells adherent t o H&RS cells. Cells from five cases of HD [four classic HD and one lymphocy te-predominant (LP) HD] were examined by single-cell analysis for the T-cel l receptor (TCR) gamma gene. Between 5 and 17 rosettes containing one to te n rosetting lymphocytes and the corresponding H&RS cells were amplified in separate plastic tubes. Of the resulting 119 TCR gamma polymerase chain rea ction (PCR) products, 87 were sequenced. While no evidence of a clonal expa nsion was obtained in the lymph nodes from four of five patients with class ic HID, clonal TCR gamma sequences were found in the lymph node from the pa tient within LPHD in two independent experiments analyzing seven and ten di fferent rosetting complexes, respectively. Of 13 products, 11 showed identi cal V gamma9 sequences. Unrelated products were found in all other TCR gamm a family subgroups in this case. Single H&RS cells picked as controls were negative for TCR gamma rearrangements. Our results demonstrate that clonal proliferations on a polyclonal background can occur among the T cells formi ng rosettes with Hodgkin cells and lend support to the view that Hodgkin ce lls may also function as cells presenting antigens to the adhering T cells.