Lung-derived complement in pneumonia

点击翻译按钮获取中文摘要

PROJECT ABSTRACT The objective of this R01 proposal is to investigate the function of lung-derived complement proteins and harness their activities to mitigate the severity of pneumonia. Complement proteins C3 and Factor B (FB) comprise an early arm of the host immune response. They are primarily derived from the liver and function in the circulation by killing pathogens such as bacteria. However, our recent work has demonstrated the importance of local C3 expression in lung epithelial cell survival during stress. This proposal focuses on an emerging role for lung- derived FB, a ligand for C3, in reducing excessive tissue damage in the setting of an acute bacterial pneumonia. Our goal is to determine how FB promotes pulmonary host defense. We will investigate sources of FB in the lung and establish its putative protective effect relative to C3. A major hurdle for investigating tissue-specific roles of complement has been the limited availability of models and assays. We have developed novel transgenic mouse models and functional assays that distinguish the roles of liver- and lung-derived complement proteins, and specifically identify the role of lung-derived FB in pneumonia. Our mouse models are supplemented with data from in vitro human models that demonstrate the role of FB to protect against stress-induced epithelial cell death. Additionally, CRISPR-induced deletion of cell-derived complement proteins suggests active internalization of exogenous complement proteins. These combined results support our central hypothesis that lung-derived FB promotes host defense by mitigating epithelial cell death. This proposal will test our hypothesis by achieving two Specific Aims. Aim 1 compares global FB-deficient, targeted liver FB-deficient, and lung epithelial cell-derived FB-deficient mice to assess how lung-derived FB mitigates acute bronchopneumonia severity and cell death. We also will assess if augmenting FB in the lung using pharmacological and gene delivery approaches protects against pneumonia. Aim 2 analyzes whether lung-derived, intracellular FB activity mitigates cell death in vitro by leveraging a combination of human primary lung epithelial cells and FB-deficient cells to dissect the molecular and biochemical mechanisms responsible for complement function in the lung. The proposal integrates knowledge of pulmonary complement activation, intracellular complement protein trafficking, and structure-function relationships with gene therapy, cell imaging, proteomics, and CRISPR screens to determine how lung-derived FB promotes epithelial cytoprotection during stress. These approaches are independent but complementary for investigating the immunobiological role of lung-derived FB in mucosal barrier protection of the lung. The proposed work is important because understanding how the early host immune response modulates tissue damage is essential for designing and implementing urgently needed, new therapies for pneumonia. Thus, we will assess how lung-derived FB facilitates host defense at the site of infection and promotes tissue resilience. These efforts will help our long-term goal of developing a novel host-focused therapy for pneumonia, thus aligning with a priority area of the NHLBI Working Group Report on pneumonia research.