Highly Focused T Cell Responses in Latent Human Pulmonary Mycobacterium tuberculosis Infection

Abstract

The elucidation of the molecular and immunological mechanisms mediating maintenance of latency in human tuberculosis aids to develop more effective vaccines and to define biologically meaningful markers for immune protection. We analyzed granuloma-associated lymphocytes (GALs) from human lung biopsies of five patients with latent Mycobacterium tuberculosis (MTB) infection. MTB CD4+ and CD8+ T cell response was highly focused in the lung, distinct from PBL, as assessed by TCR-CDR3 spectratyping coupled with a quantitative analysis of TCR VB frequencies. GALs produced IFN-γ in response to autologous macrophages infected with MTB and to defined MTB-derived HLA-A2-presented peptides Ag85a242–250, Ag85b199–207, early secreted antigenic target 6 (ESAT-6)28–36, 19-kDa Ag88–97, or the HLA-DR-presented ESAT-61–20 epitope. Immune recognition of naturally processed and presented MTB epitopes or the peptide ESAT-61–20 could be linked to specific TCR VB families, and in two patients to unique T cell clones that constituted 19 and 27%, respectively, of the CD4+ and 17% of the CD8+ GAL population. In situ examination of MTB-reactive GALs by tetramer in situ staining and confocal laser-scanning microscopy consolidates the presence of MHC class I-restricted CD8+ T cells in MTB granuloma lesions and supports the notion that clonally expanded T cells are crucial in immune surveillance against MTB.

MHC class II Tetramer Guided Detection of Mycobacterium tuberculosis‐specific CD4+ T Cells in Peripheral Blood from Patients with Pulmonary Tuberculosis

“…7) showed a strong CD8 + Mycobacterium-reactive Ag85B-specific T-cell response, despite low numbers of CD4 + T cells reactive to the same antigen. Recent reports suggested that functional were tested against the corresponding epitope from ESAT-6 which has been described to be recognized in freshly isolated CD4 + GAL (3,14) and against APC pulsed with the ESAT-6 protein (APC+Ag+ T cells). T-cell reactivity determined by IFN production could be blocked using HLA-DR, but not with an anti-MHC class I-specific MoAb.…”

“…Data are either presented as per cent tetramer-positive staining T cells in the CD4 + T-cell population or as absolute CD4 + Tetramer+ events/50,000 CD4 + T cells. Granuloma-associated lymphocytes (GAL) from granuloma tissue obtained from patients who effectively contained the infection were prepared as described recently [14]. Patients underwent thoracotomy to rule out lung cancer.…”

“…Tissue sections were incubated on a shaker at 4°C overnight with DR4 tetramer complexes loaded with the appropriate peptides (final concentration 1 lg/ml) and a goat anti-CD4 antibody (DAKO, Hamburg, Germany). Staining was performed as described in detail recently [14]. Incubation and processing of the samples with all the reagents, except the class II tetramer complex served as the negative control.…”

Mycobacterium tuberculosis-Specific CD8+ T Cells and Their Role in Immunity

“…Interestingly, IL-4 production by CD8 + T cells seems to be associated with active disease in individuals and may be a surrogate marker for the infectious stage (194). M tuberculosis-specific CD8 + T cells in the lungs of latently infected people produce GM-CSF and IFNγ (178). Although GM-CSF is not thought to function as an effector molecule, it may have a role in immunoregulation.…”

Diagnosis of Mycobacterium tuberculosis

“…Based on these findings, different diagnostic assays have been developed to measure INF-γ released by peripheralblood mononuclear cells cells in response to Mtb-specific antigens. Two such antigens are the early secreted antigenic target 6-kDa (ESAT-6) protein and culture filtrate protein 10-kDa (CFP-10) (Andersen et al, 2000;Tully et al, 2005). Both antigens have been shown to be important for the growth, survival, and pathogenesis of Mtb (Brodin et al, 2005;Munk et al, 2001).…”