Remodeling of the M. tuberculosis cell wall by the host microenvironment

描述（由申请人提供）：当结核分枝杆菌（M.tb）通过空气传播感染时，杆菌沉积在肺的肺泡腔中。在肺泡腔内及其近端存在许多先天免疫机制，这些机制在维持肺内稳态中至关重要。结核分枝杆菌是一种高度适应宿主的巨噬细胞内病原体，在感染过程中可能利用这些机制发挥其优势。Torrelles博士目前在Schlesinger博士实验室的研究重点是了解结核分枝杆菌如何在宿主识别和适应中使用其甘露糖基化表面。他正在研究表面甘露糖基化脂聚糖在人类巨噬细胞上的C型凝集素识别中的作用，以及结核分枝杆菌甘露糖帽脂阿拉伯甘露聚糖在这些细胞内的代谢。关于结核分枝杆菌如何受到巨噬细胞外肺泡微环境中免疫压力的影响，人们知之甚少。除了肺泡巨噬细胞之外，肺泡空间中肺防御的主要成分是I型和II型上皮细胞、单核细胞和嗜中性粒细胞，以及它们分泌到肺泡腔的产物（即，表面活性剂）。这些肺泡隔室细胞中的每一个都含有其自身独特的水解酶阵列，这些水解酶被释放到肺泡环境中并被隔离在表面活性剂中。当结核分枝杆菌最初沉积在终末细支气管和肺泡中时，以及从裂解的巨噬细胞释放后，杆菌与这些水解酶密切接触。在K99/R 00 NIH独立之路奖期间，Torrelles博士将研究人类肺泡环境对结核分枝杆菌细胞包膜的影响，以及这些影响如何决定结核分枝杆菌在宿主体内的命运。利用放射性标记的结核分枝杆菌H37 Rv强毒株，结合生物化学、分子生物学和细胞生物学的方法，我们提出：1）研究影响结核分枝杆菌H37 Rv细胞外膜的肺泡隔室细胞特异性水解酶和肺表面活性物质。为了确保研究保持集中，我们将优先考虑候选水解酶，并将我们的实验限制为总共3-5种水解酶的研究; 2）表征我们选择的人肺水解酶对毒性结核分枝杆菌H37 Rv细胞包膜完整性的影响;以及3）确定水解酶衍生的对结核分枝杆菌H37 Rv细胞包膜的修饰如何影响肺泡隔室细胞内的芽孢杆菌存活。鉴定影响结核分枝杆菌在宿主体内命运的肺泡水解酶将能够开发更具预测性的体外模型，并鉴定新的药物靶点。

DESCRIPTION (provided by applicant): When Mycobacterium tuberculosis (M.tb) infection occurs by airborne transmission, bacilli are deposited in the alveolar spaces of the lungs. Within the alveolar space and proximal to it exist a number of innate immune mechanisms that are critical in maintaining pulmonary homeostasis. M.tb, a highly host-adapted intracellular pathogen of macrophages, may use these mechanisms to its advantage during infection. Dr. Torrelles' current research in Dr. Schlesinger's laboratory is focused on understanding how M.tb uses its mannosylated surface in host recognition and adaptation. He is studying the role of surface mannosylated lipoglycans in recognition by C-type lectins on human macrophages and the metabolism of M.tb mannose-capped lipoarabinomannan within these cells. Little is known about how M.tb is affected by the immune pressure that it encounters in the alveolar microenvironment outside of the macrophage. In addition to alveolar macrophages, major constituents of lung defense in the alveolar space are type I and II epithelial cells, monocytes, and neutrophils, and their secreted products to the alveolar lumen (i.e., surfactant). Each of these alveolar compartment cells contains its own unique array of hydrolases that are released to the alveolar environment and sequestered in surfactant. When M.tb is initially deposited in the terminal bronchioles and alveoli, as well as following release from lysed macrophages, the bacilli are in close contact with these hydrolases. During the K99/R00 NIH Pathway to Independence Award, Dr. Torrelles will examine the effects of the human alveolar environment on the cell envelope of M.tb and how these effects dictate the fate of M.tb within the host. Using radiolabeled virulent M.tb H37Rv, and biochemical, molecular and cell biology approaches, we propose: 1) To characterize specific hydrolases derived from alveolar compartment cells and pulmonary surfactant that affect the cell envelope of M.tb H37Rv. To ensure that the studies remain focused, we will prioritize candidate hydrolases and restrict our experiments to the study of the 3-5 hydrolases in total; 2) To characterize the effects of our selected human lung hydrolases on the integrity of the virulent M.tb H37Rv cell envelope; and 3) To determine how hydrolase-derived modifications on the cell envelope of M.tb H37Rv affect the bacillus survival within alveolar compartment cells. The identification of alveolar hydrolases that affect M.tb fate within the host will enable more predictive in vitro models to be developed and novel drug targets to be identified.