T-CELL RECEPTOR DIVERSITY EXPRESSED BY CD4(-CELLS ACTIVATED BY PRIMARY ALLOGENEIC HLA-DR STIMULATION - ESTIMATION OF THE DEGREE OF CDR3 DIVERSITY() T)

In order to analyse the diversity of T-cell receptors (TCRs) expressed by the T-cell population activated by allogeneic HLA-DR stimulation, TCR beta cDNA was synthesized from mRNA of human CD4(+) T cells that h ad been stimulated in a primary mixed lymphocyte reaction (MLR). The T CR beta cDNA was amplified by the polymerase chain reaction (PCR), sub jected to bacterial cloning, and sequenced from V beta through J beta. Twenty-six different V beta genes and 10 different J beta segments we re detected among 56 randomly selected cDNA clones. Occurrences of V b eta 17.1 and J beta 1.5 were higher than those found in the CD4(+) T-c ell population activated with a CD3-specific antibody. A total of 53 d ifferent CDR3 sequences, two of them occurring more than once, were de tected among the 56 cDNA clones. In order to estimate the degree of CD R3 diversity, amino acid similarity in the CDR3 region of the cDNA was calculated and compared with those of the anti-CD3-activated T-cell s equences as well as those of various published T-cell clone sequences, each directed to either alloantigens or single antigenic peptides. It was found that the similarity score among CDR3 sequences obtained fro m the MLR (56.4 +/- 10.3) was comparable to those of anti-CD3-activate d T cells (55.7 +/- 10.7) and those of T-cell clones directed toward a lloantigens (range, 48.4 + 12.4-59.4 +/- 13.1), but significantly smal ler than those of T-cell clones directed toward single antigenic pepti des such as those derived from myelin basic protein (75.6 +/- 17.9) an d cytochrome c (76.9 +/- 20.5). These results provide quantitative pro of that TCRs of T cells activated by primary allogeneic HLA-DR stimula tion have a larger diversity than those recognizing single antigenic p eptides.