Tuberculosis disease (TB) may progress at different rates and have different outcomes. Neutrophils have been implicated in TB progression; however, data on their role during TB are controversial. Here we show that in mice, TB progression is associated with the accumulation of cells that express neutrophilic markers Gr-1 and Ly-6G, but do not belong to conventional neutrophils. The cells exhibit unsegmented nuclei, have Gr-1dimLy-6GdimCD11b+ phenotype and express F4/80, CD49d, Ly-6C, CD117, CD135 markers characteristic not of neutrophils, but of immature myeloid cells. The cells accumulate in the lungs, bone marrow, spleen and blood at the advanced (pre-lethal) stage of M. tuberculosis infection and represent a heterogeneous population of myeloid cells at different stages of their differentiation. The accumulation of Gr-1dimCD11b+ cells is accompanied by the disappearance of conventional neutrophils (Gr-1hiLy-6Ghi-expressing cells). The Gr-1dimCD11b+ cells suppress T cell proliferation and IFN-γ production in vitro via NO-dependent mechanisms, i.e. they exhibit characteristics of myeloid-derived suppressor cells (MDSCs). These results document the generation of MDSCs during TB, suggesting their role in TB pathogenesis, and arguing that neutrophils do not contribute to TB pathology at the advanced disease stage.

BackgroundInfection with Mycobacterium tuberculosis (Mtb) results in different clinical outcomes ranging from asymptomatic containment to rapidly progressing tuberculosis (TB). The mechanisms controlling TB progression in immunologically-competent hosts remain unclear.Methodology/Principal FindingsTo address these mechanisms, we analyzed TB progression in a panel of genetically heterogeneous (A/SnxI/St) F2 mice, originating from TB-highly-susceptible I/St and more resistant A/Sn mice. In F2 mice the rates of TB progression differed. In mice that did not reach terminal stage of infection, TB progression did not correlate with lung Mtb loads. Nor was TB progression correlated with lung expression of factors involved in antibacterial immunity, such as iNOS, IFN-γ, or IL-12p40. The major characteristics of progressing TB was high lung expression of the inflammation-related factors IL-1β, IL-6, IL-11 (p<0.0003); CCL3, CCL4, CXCL2 (p<0.002); MMP-8 (p<0.0001). The major predictors of TB progression were high expressions of IL-1β and IL-11. TNF-α had both protective and harmful effects. Factors associated with TB progression were expressed mainly by macrophages (F4-80+ cells) and granulocytes (Gr-1hi/Ly-6Ghi cells). Macrophages and granulocytes from I/St and A/Sn parental strains exhibited intrinsic differences in the expression of inflammatory factors, suggesting that genetically determined peculiarities of phagocytes transcriptional response could account for the peculiarities of gene expression in the infected lungs. Another characteristic feature of progressing TB was the accumulation in the infected lungs of Gr-1dim cells that could contribute to TB progression.Conclusions/SignificanceIn a population of immunocompetent hosts, the outcome of TB depends on quantitatively- and genetically-controlled differences in the intensity of inflammatory responses, rather than being a direct consequence of mycobacterial colonization. Local accumulation of Gr-1dim cells is a newly identified feature of progressing TB. High expression of IL-1β and IL-11 are potential risk factors for TB progression and possible targets for TB immunomodulation.

Following challenge with virulent Mycobacterium tuberculosis, mice of the I/St inbred strain exhibit shorter survival time, more rapid body weight loss, higher mycobacterial loads in organs, and more severe lung histopathology than mice of the A/Sn strain. We previously performed a genome-wide scan for quantitative trait loci (QTLs) that control the severity of M. tuberculosis-triggered disease in [(A/Sn ؋ I/St) F1 ؋ I/St] backcross-1 (BC1) mice and described several QTLs that are significantly or suggestively linked to body weight loss. In the present study we expanded our analysis by including the survival time phenotype and by genotyping 406 (A/Sn ؋ I/St) F2 mice for the previously identified chromosomal regions of interest. The previously identified 12-cM-wide QTL on distal mouse chromosome 3 was designated tbs1 (tuberculosis severity 1); the location of the QTL on proximal chromosome 9 was narrowed to a 9-cM interval, and this QTL was designated tbs2. Allelic variants of the tbs2 locus appeared to be involved in control of both body weight loss and survival time. Also, the data strongly suggested that a QTL located in the vicinity of the H-2 complex on chromosome 17 is involved in control of tuberculosis in mice of both genders, whereas the tbs1 locus seemed to have an effect on postinfection body weight loss in female mice. Interestingly, these loci appeared to interact with each other, which suggests that there might be a basic genetic network for the control of intracellular parasites. Overall, linkage data reported here for F2 mice are in agreement with, and add to, our previous findings concerning the control of M. tuberculosis-triggered disease in the BC1 segregation.Identification of genes and their alleles that confer resistance and/or susceptibility to tuberculosis (TB) provides deep insight into basic mechanisms of immunity and pathology. Variations in NRAMP1 (5) and/or NRAMP1-linked loci on human chromosome 2q35 (8), VDR (3, 27), and class II HLA genes (6, 7) were shown to be linked to or associated with susceptibility to TB in humans. However, linkage and association results vary between studies, which may be due to the genetic control being polygenic and ethnicity dependent and to the absence of clearly delineated phenotypes (4, 24).Mouse models of TB have proved to be valuable for studies of antimycobacterial immunity and of the genetic control of susceptibility and resistance (16). For example, the Nramp1 gene, which has provided great insight in our understanding of macrophage-mycobacterium relationships, was first discovered in a mouse model of susceptibility to Mycobacterium bovis BCG (9, 28). Numerous inbred mouse strains have been tested to determine their survival times after challenge with virulent Mycobacterium tuberculosis (1,17,20). Among these strains, I/StSnEgYCit (I/St) mice display the shortest survival time, while A/SnYCit (A/Sn) mice survive significantly longer. In addition, I/St mice display more severe and rapid disease progression than A/Sn mice in terms of body weight los...

Th1 lymphocytes are considered the main mediators of protection against tuberculosis (TB); however, their phenotypic characteristics and relationship with Th17 and Th1Th17 populations during TB are poorly understood. We have analyzed Th1, Th17, and Th1Th17 lymphocytes in the blood and pulmonary lesions of TB patients. The populations were identified based on the production of IFN-γ and/or IL-17 and the coexpression of CXCR3 (X3) and CCR6 (R6). In the blood, IL-17 and IFN-γIL-17 lymphocytes were barely detectable (median, <0.01% of CD4 lymphocytes), whereas IFN-γ lymphocytes predominated (median, 0.45%). Most IFN-γ lymphocytes (52%) were X3R6, suggesting their "nonclassical" (ex-Th17) nature. In the lungs, IL-17 and IFN-γIL-17 lymphocytes were more frequent (0.3%, < 0.005), yet IFN-γ cells predominated (11%). Phenotypically, lung CD4 cells were X3R6 The degree of differentiation of blood effector CD4 lymphocytes (evaluated based on CD62L/CD27/CD28 coexpression) increased as follows: X3R6 < X3R6 < X3R6, with X3R6 cells being largely terminally differentiated CD62LCD27CD28 cells. Lung CD4 lymphocytes were highly differentiated, recalling blood X3R6 populations. Following in vitro stimulation with anti-CD3/anti-CD28 Abs, X3R6CD4 lymphocytes converted into X3R6 and X3R6 cells. The results demonstrate that, during active TB, Th1 lymphocytes predominate in blood and lungs, document differences in X3/R6 expression by blood and lung CD4 cells, and link the pattern of X3/R6 expression with the degree of cell differentiation. These findings add to the understanding of immune mechanisms operating during TB and are relevant for the development of better strategies to control it.

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