Polymorphic MHC molecules dictate cytokine profile and clinical disease in mycobacterial diseases

Abstract

We have demonstrated an HLA linked control of susceptibility to leprosy and pulmonary tuberculosis (PTB), the associated allele being HLA-DR2. The present study has been extended to include lepromatous leprosy (LL)and multiple drug resistant tuberculosis (MDR-TB). PCRSSOP hybridization as well as sequence analysis of DRB1, DQA1 and DQB1 genes was carried out in Indian patients with leprosy, tuberculosis and healthy controls. Further, CD4+ T cell proliferation assay system was used to dissect the TH1FFH2 status of the immune response following M.leprae infection and the data was correlated with the MHC phenotype of the host. M.leprae heat shock proteins, HSP65, HSP18 and their trypsin digested fragments were utilized as the antigen source. Further, the profile of cytokine release and MHC restriction have been studied in M.leprae HSPs-primed CD4+ T cell cultures from polar leprosy patients to identify the relevant peptide responsible for the diversity in immune reaction elicited against the mycobacteria. The study revealed that HLA-DR2 was more strongly associated with LL leprosy and drug resistant PTB, both of which are clinically severe diseases with a number of immunological similarities including production of parasitespecific antibody, depressed CMI, chronic presence of bacilli and ultimately chronic illness. Molecular subtyping revealed that more than 97% of DR2+ves among patients and controls were either DRB1*1501 or 1502, suggesting that the presence of relevant epitope(s) on the DRB1*15 sequence may be involved in preferential binding of pathogenic mycobacterial peptides leading to the stimulation of pathogenic T cell clones that result in a detrimental immune response. Sequence analysis revealed a positive association of tuberculoid leprosy (TL) with HLA-DRB1 alleles that contain Arg13 or ARG70-Arg71 in the DRB1 first domain. Culture experiments showed that while the undigested HSPs (HSP65, HSP18) and their tryptic fragments of optimal digestion could stimulate CD4+T cell from TL patients and healthy contacts, only two fragments, TDB65-2 (18kd) and TDB18-3 (3kd) triggered CD4+T cells of anergic LL patients. These proteins were restricted by multiple HLA class II determinants with HLA-DR15 providing the strongest restriction. HSP65 and HSP18 induced TH2 like activity when presented in the context of HLA-DR1 and DR-7 respectively. However, when restricted by HLA-DR15 and other alleles like DR-5 and DR8, TH1 like cytokine profile was obtained as determined by the release of IL-2 and IFN-gamma. Further, TDB65-2 and TDB18-3 induced only TH1 activity both in TL as well as LL patients in the context of many DR alleles, DR15 being the main restriction element. The study provides direct evidence in the human system of the involvement of MHC class II phenotypes in governing the functional outcome of an immune response and ultimate disease phenotype in mycobactedal infectious diseases. The induction of TH1 like activity in LL patients is indeed very important for planning immune intervention therapy in leprosy.