Limited recognition ofMycobacterium tuberculosis-infected macrophages by polyclonal CD4 and CD8 T cells from the lungs of infected mice

Abstract

20Immune responses following Mycobacterium tuberculosis (Mtb) infection or vaccination 21 are frequently assessed by measuring T cell recognition of crude Mtb antigens, 22 recombinant proteins, or peptide epitopes. We previously showed that not all Mtb-23 specific T cells recognize Mtb-infected macrophages. Thus, an important question is 24 what proportion of T cells elicited by Mtb infection recognize Mtb-infected macrophages. 25We answer this question by developing a modified elispot assay using viable Mtb-26 infected macrophages, a low multiplicity of infection and purified T cells. In C57BL/6 27 mice, CD4 and CD8 T cells were classically MHC restricted. Comparable frequencies of 28 T cells that recognize Mtb-infected macrophages were determined using interferon-γ 29 elispot and intracellular cytokine staining, and lung CD4 T cells more sensitively 30 recognized Mtb-infected macrophages than lung CD8 T cells. Compared to the numbers 31 of Mtb antigen-specific T cells for antigens such as ESAT-6 and TB10.4, low frequencies 32 of pulmonary CD4 and CD8 T cells elicited by aerosolized Mtb infection recognize Mtb-33 infected macrophages. Finally, we demonstrate that BCG vaccination elicits T cells that 34 recognize Mtb-infected macrophages. We propose that the frequency of T cells that 35 recognize infected macrophages could correlate with protective immunity and may be an 36 alternative approach to measuring T cell responses to Mtb antigens. 37with the failure of T cells specific for those antigens to recognize Mtb-infected 64 macrophages 4 . Importantly, following aerosol infection, Mtb disseminates to the 65 mediastinal lymph node, where T cells are first primed by dendritic cells, which then 66 expand and traffic to the lung 9, 10 . We speculate that there may be a mismatch in the 67 antigens presented (or cross-presented) by uninfected DC in the lymph nodes and 68 antigens presented by infected macrophages in the lung. Thus, T cells primed in the lymph 69 nodes during natural infection may not necessarily recognize antigens presented by Mtb-70 infected macrophages in the lung 11 . Regardless of the mechanism, we wondered whether 71 the inability of some T cells to recognize Mtb-infected macrophages might explain why the 72 number of antigen-specific T cells may not necessarily correlate with vaccine-induced 73 protection. 74To assess T cell recognition of Mtb-infected macrophages we developed a 75 modified elispot assay based on interferon (IFN)-g spot forming cells (SFC). Using a low 76 multiplicity of infection (MOI), we quantify the frequency of T cells that recognize Mtb-77 infected macrophages during primary infection in mice. We find that an unexpectedly low 78 frequency of ex vivo CD8 and CD4 T cells recognizes Mtb-infected macrophages. We 79 demonstrate that majority of the T cells from C57BL/6 mice that recognize Mtb-infected 80 macrophages are conventionally MHC-restricted T cells. Our data shows that CD4 T 81 cells efficiently detect Mtb-infected macrophages at a lower MOI, whereas CD8 T cells 82 on...

Spatial Organization and Recruitment of Non-Specific T Cells May Limit T Cell-Macrophage Interactions Within Mycobacterium tuberculosis Granulomas

“…We use our multi-scale model to determine the proportion of Mtb-specific T cells within granulomas over the course of infection as compared to non-specific T cells present by comparing which frequencies match the dataset derived from NHP. To do this, we varied the frequency of Mtb-specific T cell classes from 1%-25% to capture a potential larger range occurring within primates ( 70 ). For each day of virtual infection, we used GranSim to calculate median, numbers, and proportions of Mtb-specific T cells versus non-specific T cells found within simulated granulomas ( Figure 7 ).…”

“…Previous studies have shown that T cells localize to and are rapidly recruited into mycobacterial granulomas in the absence of antigen recognition ( 36 – 38 ). One study found around 5-20% of CD4+ T cells recognize Mtb-infected macrophages by 19- to 22-weeks post infection ( 70 ). Comparatively in GranSim , 10%–20% T cells are Mtb-specific T cells 7- to 28-weeks post infection, with only Mtb-specific T cells given activation capabilities.…”

“…Patankar et al. showed in mice granulomas that 5%–20% of T cells are Mtb-specific ( 70 ). Here, we varied the frequency of each Mtb-specific T cell class from 1%-25% to capture a potential larger range occurring within primates.…”

In silico analysis of epitope-based vaccine candidate against tuberculosis using reverse vaccinology

“…Therefore, the selection of epitopes fixes with MHC is an essential aspect in predicting potent T-cell epitopes 75 . Also, CD4 + and CD8 + T-cells' recognition is critical while developing multi-epitope-based vaccines 76 , 77 . We predicted B and T-cell epitopes from the nominated antigens and joined them using AAY and GPGPG linkers in order to construct epitope-based vaccine 78 .…”

Classical CD4 T cells as the cornerstone of antimycobacterial immunity

“…36 This original pool, named "MTB300", and versions thereof have been used by a number of studies to measure and phenotype Mtb-specific responses. 28,[37][38][39][40][41][42][43][44][45][46][47][48][49][50] Due to the overlap of epitopes recognized by different species MHC this pool has also been shown to capture T cell reactivity in mice and nonhuman primates. 38,41,44,49,50 These studies have contributed to our understanding of immunity against Mtb and other mycobacteria.…”