Diversity and function of mononuclear phagocytes in the lung driven by mycobacterium tuberculosis infection

项目总结/摘要 结核分枝杆菌（Mtb）在单核吞噬细胞（MP）中存活和复制的能力是 这是其致病性和感染世界四分之一人口的能力的核心。乔尔博士的实验室 Ernst（共同指导者）和其他人已经证明了结核分枝杆菌群体之间的差异生长和复制。 宪兵。然而，MP子集的复杂性和多样性只是最近才在转录分辨率上被定义 在结核分枝杆菌感染期间没有很好地描述。因此，目前尚不清楚哪些亚群对结核分枝杆菌最容易感染 生长以及Mtb是否指导单核细胞分化。使用流式细胞术和单细胞RNA测序 从感染的小鼠的scRNA-seq，初步数据表明不同的感染率和对Mtb的反应 在两个募集的巨噬细胞亚群中。这就指导了结核分枝杆菌毒力的核心假设， 在感染期间，因子驱动肺中招募的MP的分化状态，为Mtb提供了肥沃的生态位 增长和复制。这项工作的长期目标是划定结核分枝杆菌和国会议员之间的关系 在最近理解肺免疫细胞异质性的革命背景下， 宿主导向疗法。利用荧光表达Mtb、流式细胞术和单细胞RNA测序 在感染实验室Mtb菌株（H37 Rv）和缺乏功能性ESX-1分泌系统的菌株的小鼠中， 特异性目的1将检验Mtb毒力系统驱动单核细胞分化为特异性 巨噬细胞亚群通过使用流式细胞术、免疫组织化学和离体感染模型， 目的2将检验募集的巨噬细胞亚群具有不同的限制结核分枝杆菌的能力的假设 复制的 培训计划的重点是发展对分枝杆菌、单核细胞 吞噬细胞和肺环境生物学，分枝杆菌的遗传操作，scRNA-seq和生物信息学， 领导力和负责任的研究实践。培训将包括教学课程、讲习班、研讨会， 地方和国际会议。主要导师和共同导师是博士奥伦罗森伯格和博士乔尔 恩斯特，这两个成功的物理学家，科学家与免疫学，结核病生物学和主机相结合的专业知识， 病原体相互作用一个总体导师团队还包括一名生物信息学科学顾问， 他是下一代测序的合作者，也是有成就的医生-科学家职业导师。查医生 实验室和办公空间在加州大学旧金山分校使命湾，与共享空间在BSL 2和BSL 3实验室，与 获得这项研究所需的设备和小鼠模型。职业发展奖 申请有一个创新的研究建议，严格的培训计划，指导领导人在分枝杆菌 免疫学和生物学，医生科学家顾问团队，有将科学家转变为 独立性，和一个特殊的候选人朝着一个独立的职业生涯在分枝杆菌宿主 相互作用和慢性肺病。

Project Summary/Abstract Mycobacterium tuberculosis’s (Mtb) ability to survive and replicate in mononuclear phagocytes (MPs) is central to its pathogenicity and capability to infect a quarter of the world’s population. The laboratory of Dr. Joel Ernst (co-mentor) and others have demonstrated differential Mtb growth and replication between populations of MPs. Yet, the complexity and diversity of MP subsets is only recently being defined at transcriptional resolution and not well described during Mtb infection. It is therefore unclear which subsets are most permissive to Mtb growth and if Mtb directs monocyte differentiation. Using flow cytometry and single cell RNA-sequencing (scRNA-seq) from infected mice, preliminary data demonstrates differential infection rates and responses to Mtb within two recruited macrophage subpopulations. This has directed the central hypothesis that Mtb virulence factors drive differentiation states of recruited MPs in the lung during infection, providing a fertile niche for Mtb growth and replication. The long-term goal of this work is to delineate the relationship between Mtb and MPs in the context of a recent revolution in understanding lung immune cell heterogeneity, enabling more effective host-directed therapies. Utilizing fluorescently expressing Mtb, flow cytometry, and single cell-RNA sequencing in mice infected with a laboratory strain of Mtb (H37Rv), and a strain lacking a functional ESX-1 secretion system, specific Aim 1 will test the hypothesis that an Mtb virulence system drives differentiation of monocytes to specific macrophage subsets. With use of flow cytometry, immunohistochemistry, and ex vivo infection models, specific Aim 2 will test the hypothesis that recruited macrophage subsets have differential ability to restrict Mtb replication. The training plan is focused to develop expertise in immunologic responses to mycobacterium, mononuclear phagocyte and lung milieu biology, genetic manipulation of mycobacterium, scRNA-seq and bioinformatics, leadership, and responsible research practices. Training will include didactic courses, workshops, seminars, local and international conferences. The primary mentor and co-mentors are Dr. Oren Rosenberg and Dr. Joel Ernst, both successful physician-scientists with combined expertise in immunology, Mtb biology, and host- pathogen interactions. An overarching team of mentors also include a scientific advisor in bioinformatics, collaborator in next generation sequencing, and accomplished physician-scientist career mentors. Dr. Zha has laboratory and office space at UCSF Mission Bay, with shared space in a BSL2 and BSL3 laboratories, with access to equipment and mouse models necessary for this research. This Career Development Award application has an innovative research proposal, rigorous training plan, mentorship of leaders in mycobacterial immunology and biology, team of physician-scientist advisors with a track record of transitioning scientists to independence, and an exceptional candidate driven toward an independent career in mycobacterial-host interaction and chronic lung disease.