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. 2011 Mar;17(3):372-6.
doi: 10.1038/nm.2299. Epub 2011 Feb 20.

Dominant TNF-α+ Mycobacterium tuberculosis-specific CD4+ T cell responses discriminate between latent infection and active disease

Alexandre Harari  1 , Virginie Rozot, Felicitas Bellutti Enders, Matthieu Perreau, Jesica Mazza Stalder, Laurent P Nicod, Matthias Cavassini, Thierry Calandra, Catherine Lazor Blanchet, Katia Jaton, Mohamed Faouzi, Cheryl L Day, Willem A Hanekom, Pierre-Alexandre Bart, Giuseppe Pantaleo
Affiliations

Dominant TNF-α+ Mycobacterium tuberculosis-specific CD4+ T cell responses discriminate between latent infection and active disease

Alexandre Harari et al. Nat Med. 2011 Mar.

Abstract

Rapid diagnosis of active Mycobacterium tuberculosis (Mtb) infection remains a clinical and laboratory challenge. We have analyzed the cytokine profile (interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α) and interleukin-2 (IL-2)) of Mtb-specific T cells by polychromatic flow cytometry. We studied Mtb-specific CD4+ T cell responses in subjects with latent Mtb infection and active tuberculosis disease. The results showed substantial increase in the proportion of single-positive TNF-α Mtb-specific CD4+ T cells in subjects with active disease, and this parameter was the strongest predictor of diagnosis of active disease versus latent infection. We validated the use of this parameter in a cohort of 101 subjects with tuberculosis diagnosis unknown to the investigator. The sensitivity and specificity of the flow cytometry-based assay were 67% and 92%, respectively, the positive predictive value was 80% and the negative predictive value was 92.4%. Therefore, the proportion of single-positive TNF-α Mtb-specific CD4+ T cells is a new tool for the rapid diagnosis of active tuberculosis disease.

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Conflict of interest statement

COMPETING FINANCIAL INTERESTS

The authors declare no competing financial interests.

Figures

Figure 1
Figure 1
Quantitative and qualitative analysis of Mtb-specific T cell responses in the test cohort. (a) IFN-γ ELISPOT responses after stimulation with ESAT-6 or CFP-10 peptide pools in a cohort of 283 participants with latent Mtb infection (n = 272) or active tuberculosis disease (n = 11, Supplementary Fig. 1). Shown are the numbers of spot-forming units (SFU) per 106 mononuclear cells. Statistical significance (P values) of the results was calculated by unpaired two-tailed Student’s t test using GraphPad Prism 5. Bonferroni’s correction for multiples analyses was applied. (b) Qualitative analysis of Mtb-specific CD4+ T cell responses by polychromatic flow cytometry. Shown are representative flow cytometry analyses of the functional profile of Mtb-specific CD4+ T cell responses in participants with either latent Mtb infection (Subject L5, left) or active tuberculosis disease (Subject A2, right). Profiles are gated on live CD3+CD4+ T cells, and the various combinations of IFN-γ, IL-2 and TNF-α are shown following stimulation with ESAT-6 and CFP-10 peptide pools or PPD. NS, not significant; Neg, negative control (unstimulated). (c) Simultaneous analysis of the functional profile of Mtb-specific CD4+ T cells on the basis of IFN-γ, IL-2 or TNF-α production. ESAT-6–, CFP-10– and PPD-specific CD4+ T cell responses are shown (as indicated by the six colored boxes to the right of the panel) from 48 participants with latent Mtb infection and eight participants with active tuberculosis (TB) disease. Representative examples from subject L5 and subject A2 are also identified. All the possible combinations of the various functions are shown on the x axis, whereas the percentages of the distinct cytokine-producing cell subsets within Mtb-specific CD4+ T cells are shown on the y axis. The pie charts summarize the data, and each slice corresponds to the proportion of Mtb-specific CD4+ T cells positive for a certain combination of functions. Colors in the pie charts are indicated by the seven colored boxes at the bottom of the panel. (d) Distribution of CFP-10– and/or ESAT-6–specific CD4+ T cell responses among subjects with latent Mtb infection or active tuberculosis disease.
Figure 2
Figure 2
Analysis of Mtb-specific T cell responses in the validation cohort after unblinding of the clinical status. (a) IFN-γ ELISPOT responses after stimulation with ESAT-6 or CFP-10 peptide pools. Shown are the numbers of SFU per 106 mononuclear cells. Statistical significance (P values) of the results was calculated by unpaired two-tailed Student’s t test in GraphPad Prism 5. Bonferroni’s correction for multiples analyses was applied. (b) Analysis of Mtb-specific IFN-γ ELISPOT T cell responses in subjects enrolled in Switzerland (CH) and SA. (c) Distribution of CFP-10– and/or ESAT-6–specific CD4+ T cell responses among subjects from the validation cohort with positive and concordant Mtb-specific CD4+ T cell responses (Supplementary Fig. 5).
Figure 3
Figure 3
Percentages of CFP-10– or ESAT-6–specific single-positive TNF-α–producing CD4+ T cells of the 94 subjects from the validation cohort with concordant responses against CFP-10 and ESAT-6. Dashed line represents the cutoff of 37.4% of single-positive TNF-α. (a) Subjects with active disease or latent infection are identified with blue and red dots, respectively. (b) Subjects from South Africa (SA) or Switzerland (CH) are identified with orange and green dots, respectively.
Figure 4
Figure 4
Longitudinal analysis of the functional profile of Mtb-specific CD4+ T cells from five subjects analyzed during untreated active tuberculosis disease and then after tuberculosis treatment. Shown is the full functional profile on the basis of IFN-γ, IL-2 and TNF-α production of a total of 7 Mtb-specific CD4+ T cell responses. All the possible combinations of the different functions are shown on the x axis, whereas the percentages of the distinct cytokine-producing cell subsets within Mtb-specific CD4+ T cells are shown on the y axis. The pie charts summarize the data, and each slice corresponds to the proportion of Mtb-specific CD4+ T cells positive for a certain combination of functions.
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References

    1. Kaufmann SH How can immunology contribute to the control of tuberculosis? Nat. Rev. Immunol 1, 20–30 (2001). - PubMed
    1. Flynn JL & Chan J Immunology of tuberculosis. Annu. Rev. Immunol 19, 93–129 (2001). - PubMed
    1. Lalvani A et al. Enhanced contact tracing and spatial tracking of Mycobacterium tuberculosis infection by enumeration of antigen-specific T cells. Lancet 357, 2017–2021 (2001). - PubMed
    1. Ewer K et al. Comparison of T cell-based assay with tuberculin skin test for diagnosis of Mycobacterium tuberculosis infection in a school tuberculosis outbreak. Lancet 361, 1168–1173 (2003). - PubMed
    1. Meier T, Eulenbruch HP, Wrighton-Smith P, Enders G & Regnath T Sensitivity of a new commercial enzyme-linked immunospot assay (T SPOT-TB) for diagnosis of tuberculosis in clinical practice. Eur. J. Clin. Microbiol. Infect. Dis 24, 529–536 (2005). - PubMed

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