Analysis of Mycobacterium tuberculosis-Specific CD8 T-Cells in Patients with Active Tuberculosis and in Individuals with Latent Infection

Caccamo, Nadia; Guggino, Giuliana; Meraviglia, Serena; Gelsomino, Giuseppe; Carlo, Paola Di; Titone, Lucina; Bocchino, Marialuisa; Galati, Domenico; Matarese, Alessandro; Nouta, Jan; Klein, Michél R.; Salerno, Alfredo; Sanduzzi, Alessandro; Dieli, Francesco; Ottenhoff, Tom H. M.

doi:10.1371/journal.pone.0005528

Cited by 87 publications

(89 citation statements)

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“…Primary data from seven TB patients (with PBMCs) stimulated with peptide 68 are shown in Supporting Information Figure 4. Thus, and differently from CD8 T cells recognizing M. tuberculosis peptides in the context of HLA-class Ia molecules [31,[34][35][36][37] In contrast, coculture of B cells with 9 of the 15 HLA-E-restricted CD8 T-cell lines resulted in a significant increase in the production of IgG and IgM. As a control, coculture of tonsillar B cells with sorted CD8 + T cells isolated from the PBMCs of healthy donors did not increase significantly Ab production.…”

Section: Functional Properties Of Hla-e-restricted and M Tuberculosimentioning

confidence: 90%

“…Tetramer staining was carried out as described in detail previously [31,37,54]. PBMCs (10 6 /mL) were incubated in U-bottom 96-well plates, washed twice in PBS containing 1% FCS (Sigma), and stained for 30 min at 4°C with PE-labeled tetramers (5 μL each) prepared as previously described [31,37,54]; they were washed and subsequently stained with FITC-labeled anti-CD8 mAb (RPA-TB, BD Biosciences) and analyzed by flow cytometry on a FACSCanto.…”

Section: Tetramer Stainingmentioning

confidence: 99%

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Human CD8 T lymphocytes recognize Mycobacterium tuberculosis antigens presented by HLA‐E during active tuberculosis and express type 2 cytokines

et al. 2015

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CD8 T cells contribute to protective immunity against Mycobacterium tuberculosis. In humans, M. tuberculosis reactive CD8 T cells typically recognize peptides associated to classical MHC class Ia molecules, but little information is available on CD8 T cells recognizing M. tuberculosis Ags presented by nonclassical MHC class Ib molecules.We show here that CD8 T cells from tuberculosis (TB) patients recognize HLA-E-binding M. tuberculosis peptides in a CD3/TCR αβ mediated and CD8-dependent manner, and represent an additional type of effector cells playing a role in immune response to M. tuberculosis during active infection. HLA-E-restricted recognition of M. tuberculosis peptides is detectable by a significant enhanced ex vivo frequency of tetramer-specific circulating CD8 T cells during active TB. These CD8 T cells produce type 2 cytokines upon antigenic in vitro stimulation, help B cells for Ab production, and mediate limited TRAIL-dependent cytolytic and microbicidal activity toward M. tuberculosis infected target cells. Our results, together with the finding that HLA-E/M. tuberculosis peptide specific CD8 T cells are detected in TB patients with or without HIV coinfection, suggest that this is a new human T-cell population that participates in immune response in TB.Keywords: CD8 T lymphocytes r HLA-E r Mycobacterium tuberculosis r TB r Tetramers r Type 2 cytokines Additional supporting information may be found in the online version of this article at the publisher's web-site Correspondence: Prof. Francesco Dieli e-mail: francesco.dieli@unipa.it * These authors share first authorship for this work.* * These authors share last authorship for this work. [2]. In humans, M. tuberculosis reactive CD8 T cells recognize peptides associated to classical HLA-A, HLA-B, and HLA-C class I (class Ia) molecules, glycolipids associated to group 1 CD1 molecules [5,6], and mycobacterial Ag associated to MHC class I related molecule (MR1) [7]. However, there is little information on the role that these cells play during infection. In mice, the MHC class Ib molecule H2-M3 binds formylated peptides derived from M. tuberculosis and induces H2-M3-restricted CD8 T cells [8,9] that are protective against M. tuberculosis infection [10]. In humans, CD8 T cells restricted by class Ib molecules comprise the very large majority of the overall M. tuberculosis specific CD8 T-cell response [11] and CD8 T cells recognizing M. tuberculosis Ags in the context of the class Ib molecule HLA-E have been isolated from subjects with latent M. tuberculosis infection [12,13]. However, the functions of this HLA-E-restricted population, as well as its contribution to the host response to M. tuberculosis during infection and disease, remain unknown.HLA-E is the least polymorphic of all the HLA molecules [14] with only two alleles in the Caucasian population, which differ at one aa position located outside the peptidebinding groove [15]. Physiologically, HLA-E binds nonamer peptides derived from the signal sequence of other HLA class I molecules [16,17], a...

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Section: Functional Properties Of Hla-e-restricted and M Tuberculosimentioning

confidence: 90%

Section: Tetramer Stainingmentioning

confidence: 99%

Human CD8 T lymphocytes recognize Mycobacterium tuberculosis antigens presented by HLA‐E during active tuberculosis and express type 2 cytokines

et al. 2015

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“…+ T cells contribute to M. tuberculosis control through: 1) IFN-g and TNF-a production (9,(15)(16)(17); 2) lysis of infected host cells (16)(17)(18); and 3) direct killing of mycobacteria (19)(20)(21). One study demonstrated clonal CD4 + and CD8 + T cell expansion in granulomas from subjects with latent TB infection (22), and similar changes in the TCR repertoire were reported in peripheral blood versus pleural fluid in TB patients (23).…”

Section: Following Recognition Of Mycobacterial Ags On Infected Cellsmentioning

confidence: 87%

“…In a later study, we reported similar findings for CD8 + T cells directed against six M. tuberculosis epitopes (two of which were newly identified). In that study, it was also found that M. tuberculosis peptide-specific IL-2 + /IFN-g + CD8 + T cell responses were associated with natural protection against developing TB disease (15). In parallel studies, Kaufmann and colleagues (25) found clonal expansion of effector memory CD8 + T cells in older children with TB, with potential impact on the course and severity of disease.…”

Section: Following Recognition Of Mycobacterial Ags On Infected Cellsmentioning

confidence: 88%

Genome-Based In Silico Identification of New Mycobacterium tuberculosis Antigens Activating Polyfunctional CD8+ T Cells in Human Tuberculosis

Tang

Meijgaarden

Caccamo

et al. 2011

The Journal of Immunology

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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 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-γ, IL-2, and TNF-α 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.

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“…Nonspecific background was extremely low when more than one cytokine was examined. A cutoff of 0.01% was used as described previously [48]; values below this were set to zero.…”

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confidence: 99%

Multifunctional CD4⁺ T cells correlate with active Mycobacterium tuberculosis infection

et al. 2010

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Th1 CD41 T cells and their derived cytokines are crucial for protection against Mycobacterium tuberculosis. Using multiparametric flow cytometry, we have evaluated the distribution of seven distinct functional states (IFN-c/IL-2/TNF-a triple expressors, IFN-c/IL-2, IFN-c/TNF-a or TNF-a/IL-2 double expressors or IFN-c, IL-2 or TNF-a single expressors) of CD4 1 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-c double and IFN-c single expressors as compared with the other CD4 1 T-cell subsets. Proportions of the other double or single CD4 1 T-cell expressors did not differ between TB and LTBI subjects. These distinct IFN-c, IL-2 and TNF-a profiles of M. tuberculosis-specific CD4 1 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 1 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. IntroductionInfections with Mycobacterium tuberculosis (M. tuberculosis) cause a global epidemic with almost 9 million new cases and over 1.6 million deaths per year [1,2]. Outcome of M. tuberculosis infection depends on early identification and proper treatment of individuals with active tuberculosis (TB), but the lack of accurate diagnostic techniques has contributed to the re-emergence of TB as a global health threat. More than 2 billion individuals are estimated to be latently infected with M. tuberculosis (LTBI). To date, however, Ã These authors have contributed equally to this work. There were a number of differences between TB patients and subjects with LTBI following stimulation with ESAT-6, Ag85B and the 16-kDa antigen (Fig. 2). Most notably, and in contrast with the previously reported results in chronic viral infections, we found a significantly higher proportion of 31 CD4 1 T cells simultaneously secreting IFN-g, IL-2 and TNF-a in patients with TB, as compared with LTBI subjects, upon stimulation with any of the three tested M. tuberculosis antigens (Fig. 2). Using a threshold of 0.01% to avoid systematic biases incurred by zeroing negative values (frequency values o0.01% were set to zero), we found that 31 CD4 1 T cells were detectable in very few LTBI subjects (3/18, 3/18 and 2/18 in response to Ag85B, ESAT-6 and 16 kDa, respectively), but were frequently detected in most TB patients (17/20, 18/20 and 17/20, in Eur. J. Immunol. 2010. 40: 2211-2220 Nadia Caccamo et al. 2212& 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.eji-journal.eu response to Ag85B, ESAT-6 and 16 kDa, respectively; see also Table 1 for comparison).In contrast, ...

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Analysis of Mycobacterium tuberculosis-Specific CD8 T-Cells in Patients with Active Tuberculosis and in Individuals with Latent Infection

Cited by 87 publications

References 35 publications

Human CD8 T lymphocytes recognize Mycobacterium tuberculosis antigens presented by HLA‐E during active tuberculosis and express type 2 cytokines

Human CD8 T lymphocytes recognize Mycobacterium tuberculosis antigens presented by HLA‐E during active tuberculosis and express type 2 cytokines

Genome-Based In Silico Identification of New Mycobacterium tuberculosis Antigens Activating Polyfunctional CD8+ T Cells in Human Tuberculosis

Multifunctional CD4⁺ T cells correlate with active Mycobacterium tuberculosis infection

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Analysis of Mycobacterium tuberculosis-Specific CD8 T-Cells in Patients with Active Tuberculosis and in Individuals with Latent Infection

Cited by 87 publications

References 35 publications

Human CD8 T lymphocytes recognize Mycobacterium tuberculosis antigens presented by HLA‐E during active tuberculosis and express type 2 cytokines

Human CD8 T lymphocytes recognize Mycobacterium tuberculosis antigens presented by HLA‐E during active tuberculosis and express type 2 cytokines

Genome-Based In Silico Identification of New Mycobacterium tuberculosis Antigens Activating Polyfunctional CD8+ T Cells in Human Tuberculosis

Multifunctional CD4+ T cells correlate with active Mycobacterium tuberculosis infection

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Multifunctional CD4⁺ T cells correlate with active Mycobacterium tuberculosis infection