CYTOKINE-INDEPENDENT DEFENSES AGAINST MYCOBACTERIUM TUBERCULOSIS

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。该方案的总体目标是确定各种淋巴细胞群介导人单核细胞和肺泡巨噬细胞内细胞内结核分枝杆菌（M. tb）杀伤的细胞因子独立机制。具体目的是：1)确定淋巴细胞激活结核分枝杆菌感染的单核吞噬细胞的杀菌功能的接触依赖机制；2)确定结核分枝杆菌感染单核吞噬细胞的最大淋巴细胞介导激活是否需要细胞接触和细胞因子产生的顺序或相互作用；3)确定其他淋巴细胞亚群（包括CD8+ T细胞、γ - δ T细胞和自然杀伤（NK）细胞）是否介导结核分枝杆菌感染的单核吞噬细胞的接触依赖性激活，比较这些效应群与CD4+ T细胞使用的机制，并确定fas/ fasl介导的细胞毒性效应机制在人单核吞噬细胞内杀死结核分枝杆菌中的作用。我们的研究主要是基于体外试验来评估结核分枝杆菌的细胞内生长。我们已经能够利用这个试验来证明淋巴细胞群介导细胞内结核分枝杆菌杀伤的能力。各种细胞毒性和细胞凋亡试验已被用来确定这些淋巴细胞功能与结核分枝杆菌杀伤之间的关系。参与研究的人类受试者包括抽血和支气管镜检查及支气管肺泡灌洗。血液单核细胞和各种淋巴细胞群从外周血中分离出来。由于结核分枝杆菌通过呼吸道传播，因此与研究最相关的单核吞噬细胞是肺泡巨噬细胞。利用支气管肺泡灌洗从健康的非吸烟者身上获得这些细胞。

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.