Tuberculosis (TB) is a chronic infectious disease caused by the intracellular bacterial pathogen Mycobacterium tuberculosis. Hence, cell- mediated immunity rather than antibodies is the critical protective immune response against M. tb. Studies with animal models of TB suggest that the major histocompatibility complex (MHC) class I-restricted CD8+ T-cell response complements host defenses mediated by activated macrophages and CD4+ T cells, and has an important role in successful resolution of mycobacterial infections. However, in humans the role of CD8+ T cells has not been fully addressed. We have developed an in vitro assay that is based on a novel computer-driven algorithm predicting MHC class I ligands (peptides) with a proven high accuracy. Our assay can measure the presentation of predicted peptides by proper MHC class I alleles, and thus allows for identification of the epitopes recognized by CD8+ T cells. Thus, the overall objective of these studies is the analysis of the role of CD8+ T-cell response in resistance to TB. First, secreted mycobacterial proteins with known sequences will be analyzed by the computer program for the presence of potential CD8+ T-cell epitopes. Presentation of these predicted M. tb. -derived peptides by MHC class I will be determined in our peptide-binding assay with an antigen transporter-deficient cell line expressing HLA alleles of interest. This analysis will be extended to HLA alleles predominant in populations with high incidences of TB and HIV infection. Second, this methodology will be used in quantitative and qualitative analysis of the MHC class I-restricted CD8+ T-cell immune responses to M. tb. -derived peptides in PPD+ healthy donors and TB patients. The frequencies of memory CD8+ T cells in the peripheral blood and activated cytotoxic T lymphocytes in the bronchio-alveolar lavage will be measured. Comparison of CD8+ T-cell effector functions in TB patients vs. PPD+ healthy donors, and also in PPD+/HIV-infected and active TB/HIV- infected patients will determine the correlation between the CD8+ T-cell responses and resistance to M. tb. infection in these patients. Finally, M. tb. -specific CD8+ T cell clones will be isolated and functionally characterized. These T cell clones will be used to study the requirements for MHC class I-restricted presentation of M. tb. antigens by macrophages infected in vitro with viable bacilli.
| Dong, Yuzhi; Demaria, Sandra; Sun, Xuming et al. (2004) HLA-A2-restricted CD8+-cytotoxic-T-cell responses to novel epitopes in Mycobacterium tuberculosis superoxide dismutase, alanine dehydrogenase, and glutamine synthetase. Infect Immun 72:2412-5 |
| Dong, Yuzhi; Lieskovska, Jaroslava; Kedrin, Dmitriy et al. (2003) Soluble nonclassical HLA generated by the metalloproteinase pathway. Hum Immunol 64:802-10 |
| Demaria, S; Bushkin, Y (2000) Soluble HLA proteins with bound peptides are released from the cell surface by the membrane metalloproteinase. Hum Immunol 61:1332-8 |
| Demaria, S; DeVito-Haynes, L D; Salter, R D et al. (1999) Peptide-conformed beta2m-free class I heavy chains are intermediates in generation of soluble HLA by the membrane-bound metalloproteinase. Hum Immunol 60:1216-26 |
