CYTOKINE-INDEPENDENT DEFENSES AGAINST MYCOBACTERIUM TUBERCULOSIS

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The overall goal of this protocol is to determine the cytokine-independent mechanisms by which various lymphocyte populations mediate killing of intracellular Mycobacterium tuberculosis (M. tb) within human monocytes and alveolar macrophages. The specific aims are 1) to determine the contact-dependent mechanisms by which lymphocytes activate bactericidal functions of M. tuberculosis-infected mononuclear phagocytes; 2) to determine whether the maximal lymphocyte-mediated activation of M. tuberculosis-infected mononuclear phagocytes requires sequential or interactive effects of cell contact and cytokine production; and 3) to determine whether other lymphocyte subsets (including CD8+ T cells, gamma-delta T cells and natural killer (NK) cells) mediate contact-dependent activation of M. tuberculosis-infected mononuclear phagocytes, to compare the mechanisms used by these effector populations to those of CD4+ T cells, and to determine the role of fas/fasL-mediated cytotoxic effector mechanisms on killing of M. tuberculosis within human mononuclear phagocytes. Our studies are primarily based on an in vitro assay for assessment of intracellular growth of M. tb. We have been able to utilize this assay to demonstrate the ability of lymphocyte populations to mediate killing of intracellular M. tb. Various assays of cytotoxicity and apoptosis have been used to determine the correlation between these lymphocyte functions and killing of M. tb. The participation of human subjects in the study involves both blood drawing and bronchoscopy with bronchoalveolar lavage. Blood monocytes and various lymphocyte populations are isolated from peripheral blood. As M. tb is spread via the respiratory route, the most relevant mononuclear phagocytes for study are alveolar macrophages. Bronchoalveolar lavage is utilized to obtain these cells from healthy non-smokers.