CD4-CELL RESPONSES TO RECOMBINANT HSP65 AND HSP18 OF MYCOBACTERIUM-LEPRAE AND THEIR TRYPSIN-DIGESTED FRAGMENTS IN LEPROSY - DIVERSITY IN HLA-DR RESTRICTION( T)

Mycobacterium leprae heat-shock proteins hsp65 and hsp18 have received immense attention as major T-cell target antigens in leprosy. Both of these hsps and their tryptic fragments were characterized for their a bility to stimulate CD4+ T cells derived from polar leprosy cases and healthy contacts. The optimal digestion of hsps with trypsin yielded f our fragments of hsp65 - TDB65-1 (24 kDa), TDB65-2 (18 kDa), TDB65-3 ( 17 kDa), TDB65-4 (14 kDa) - and three of hsp18 - TDB18-1 (10 kDa), TDB 18-2 (5 kDa), TDB18-3 (3 kDa). While all of these tryptic fragments an d undigested hsps triggered CD4+ T cells from tuberculoid (TT) leprosy patients and healthy contacts (SI > 2), only two fragments - TDB65-2 and TDB18-3 - were found to be stimulatory in anergic lepromatous (LL) leprosy patients (SI = 5.27 and 3.0, respectively). Blocking studies using allele-specific anti-DR monoclonal antibodies revealed multiple HLA-Dr restriction, with DR2 providing the strongest restriction in bo th TT as well as LL leprosy. These findings indicate that M. leprae hs ps and their trypsin-digested fragments are promiscuous and recognizab le in the context of diverse HLA alleles, of which DR2 is the most eff icient restriction element. The 18-kDa fragment of hsp65 and the 3-kDa fragment of hsp18 are the most versatile fragments that could elicit in vitro proliferation in both polar forms of leprosy.