Although CD8(+) T cells help control Mycobacterium tuberculosis infection, their M. tuberculosis Ag repertoire, in vivo frequency, and functionality in human tuberculosis (TB) remains largely... Show moreAlthough CD8(+) T cells help control Mycobacterium tuberculosis infection, their M. tuberculosis Ag repertoire, in vivo frequency, and functionality in human tuberculosis (TB) remains largely undefined. We have performed genome-based bioinformatics searches to identify new M. tuberculosis epitopes presented by major HLA class I supertypes A2, A3, and B7 (covering 80% of the human population). A total of 432 M. tuberculosis peptides predicted to bind to HLA-A*0201, HLA-A*0301, and HLA-B*0702 (representing the above supertypes) were synthesized and HLA-binding affinities determined. Peptide-specific CD8(+) T cell proliferation assays (CFSE dilution) in 41 M. tuberculosis-responsive donors identified 70 new M. tuberculosis epitopes. Using HLA/peptide tetramers for the 18 most prominently recognized HLA-A*0201-binding M. tuberculosis peptides, recognition by cured TB patients' CD8(+) T cells was validated for all 18 epitopes. Intracellular cytokine staining for IFN-gamma, IL-2, and TNF-alpha revealed mono-, dual-, as well as triple-positive CD8(+) T cells, indicating these M. tuberculosis peptide-specific CD8(+) T cells were (poly) functional. Moreover, these T cells were primed during natural infection, because they were absent from M. tuberculosis-noninfected individuals. Control CMV peptide/HLA-A*0201 tetramers stained CD8(+) T cells in M. tuberculosis-infected and noninfected individuals equally, whereas Ebola peptide/HLA-A*0201 tetramers were negative. In conclusion, the M. tuberculosis-epitope/Ag repertoire for human CD8(+) T cells is much broader than hitherto suspected, and the newly identified M. tuberculosis Ags are recognized by (poly) functional CD8(+) T cells during control of infection. These results impact on TB-vaccine design and biomarker identification. The Journal of Immunology, 2011, 186: 1068-1080. Show less
Caccamo, N.; Guggino, G.; Joosten, S.A.; Gelsomino, G.; Carlo, P. di; Titone, L.; ... ; Dieli, F. 2010
Th1 CD4(+) T cells and their derived cytokines are crucial for protection against Mycobacterium tuberculosis. Using multiparametic flow cytometry, we have evaluated the distribution of seven... Show moreTh1 CD4(+) T cells and their derived cytokines are crucial for protection against Mycobacterium tuberculosis. Using multiparametic flow cytometry, we have evaluated the distribution of seven distinct functional states (IFN-gamma/IL-2/TNF-alpha triple expressors, IFN-gamma/IL-2, IFN-gamma/TNF-alpha or TNF-alpha/IL-2 double expressors or IFN-gamma, IL-2 or TNF-alpha single expressors) of CD4(+) T cells in individuals with latent M. tuberculosis infection (LTBI) and active tuberculosis (TB). We found that triple expressors, while detectable in 85-90%TB patients, were only present in 10-15% of LTBI subjects. On the contrary, LTBI subjects had significantly higher (12- to 15-fold) proportions of IL-2/IFN-gamma double and IFN-gamma single expressors as compared with the other CD4(+) T-cell subsets. Proportions of the other double or single CD4(+) T-cell expressors did not differ between TB and LTBI subjects. These distinct IFN-gamma, IL-2 and TNF-alpha profiles of M. tuberculosis-specific CD4(+) T cells seem to be associated with live bacterial loads, as indicated by the decrease in frequency of multifunctional T cells in TB-infected patients after completion of anti-mycobacterial therapy. Our results suggest that phenotypic and functional signatures of CD4(+) T cells may serve as immunological correlates of protection and curative host responses, and be a useful tool to monitor the efficacy of anti-mycobacterial therapy. Show less
