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Identification of Mycobacterium tuberculosis CTL epitopes restricted by HLA-A*0201 in HHD mice
Alireza Rafiei1, Yiukito Kiutade2
1Dept. of Immunology and Microbiology, Sari Medical School, Mazandaran University of Medical Science, Mazandaran, Iran.
Background:
CD8 T cells are thought to play an important role in protective immunity to tuberculosis. The major histocompatibility complex class I subtype HLA-A*0201 is one of the most prevalent class I alleles, with a frequency of over 30% in most populations. HLA-A*0201 transgenic, H-2D(b)/mouse beta2-microglobulin double-knockout mice (HHD) which express human HLA-A*0201 but no mouse class 1, was shown to provide a powerful model for studying induction of HLA-A*0201-restricted immune responses in vivo.
Methods:
HHD mice were immunized with plasmid DNA encoding MPB51 by using a gene gun, and IFN-gamma production from the immune spleen cells was analyzed in response to a synthetic overlapping peptide library covering the mature MPB51 sequence. catatonic T lymphocytes (CTL) activity was measured using cytotoxicity assay and the three-color flowcytometry was used to reveal IFN-gamma-producing immune spleen cells.
Results:
Our findings were shown that only one peptide, p51-70, appeared to stimulate the immune splenocytes to produce IFN-gamma. Flow cytometric analysis with intracellular IFN-gamma and the T-cell phenotype revealed that the p51-70 peptide contains an immunodominant CD8+ T-cell epitope. Further analysis with computer-assisted algorithms permitted identification of a T-cell nona mer epitope, p54-62. Finally, we proved that the p54-62/HLA-A*0201 complex is strongly recognized by HLA class I-restricted CD8+ MPB5 1-specific CTL cells.
Conclusion:
These results suggest that vaccination with MPB51 gene elicited MPB51-specific CTL. In addition, the P54-62 epitope thus represent potential subunit component for the design of vaccines against tuberculosis.
