Mechanisms of regulatory T-cell mediated endothelial repair following viral pneumonia in aged hosts

项目摘要/摘要 这项K08提案的广泛目标有两个：1)促进发展必要的技能， 将使候选人实现她的长期目标，即成为一名成功的内科医生-科学家，专注于 老年宿主病毒性肺炎康复的决定因素，以及2)调查指导 Tregs在整个生命周期内协调解决严重的肺损伤。候选人和她的导师 根据应聘者的具体需求和目标设计了详细的培训计划。该计划包括一项 严谨的研究部分，将为PI提供新的知识和研究技能，以更好地审查 在病毒性肺炎康复期间，关键免疫细胞群与内皮之间的联系。这个 建议涉及病毒性肺炎，特别是流感引起的肺损伤，其临床对应的急性 呼吸窘迫综合征(ARDS)以及它们如何对老年人口造成不成比例的影响。尽管 经过几十年的专注研究，只有少数几种抗病毒疗法的疗效有限，可以应对严重的 病毒性肺炎。Tregs已被证明在不同的小鼠中可以减少炎症和促进组织修复 肺损伤模型。ARDS患者肺中的Treg也增加，这表明它们可能起到了 在人类对肺损伤的适应性免疫反应中的作用。然而，导致Tregs的具体机制 在肺损伤后，如何执行他们的修复计划仍不清楚。我们的初步数据显示 流感诱导的肺损伤后年轻的修复性Treg细胞程序受生物过程控制 与毛细血管的发展和修复有关。这种修复性程序在细胞中的老化宿主中丢失- 自主的方式。因此，我们假设衰老导致Treg细胞特异性下调重要的 促血管生成因子表达，如VEGFA，导致受损的肺泡内皮细胞修复和恢复 死于病毒性肺炎。该提案的长期目标是确定新的基于小分子和细胞的 在我们日益老龄化的人口中控制炎症和促进组织修复的治疗方法。为了测试这一点 假设，我们提出了以下具体目标：1)确定与年龄相关的改变是否有利于 Tregs的内皮修复功能源于细胞自主或微环境驱动的变化，2) 确定Treg细胞产生的VEFGA对于恢复促内皮细胞修复是否必要和充分 青年中随年龄增长而丧失的功能，以及3)决定年龄相关VEGFA在 重症病毒性肺炎患者支气管肺泡灌洗液中的肺泡刺激物与30天死亡率相关 肺炎。我们将使用标准技术评估肺损伤的严重程度、内皮修复、异时性 (年龄不匹配)过继Treg细胞转移，多个转基因小鼠品系功能丧失和获得- 功能实验、流式细胞术、荧光激活细胞分类和转录图谱 RNA测序作为支持这一建议的实验设计的主要方法。

PROJECT SUMMARY/ABSTRACT The broad objectives of this K08 proposal are two-fold: 1) to facilitate development of the necessary skills that will allow the candidate to achieve her long-term goal of becoming a successful physician-scientist focusing on determinants of recovery from viral pneumonia in aged hosts, and 2) to investigate the mechanisms that direct the Tregs to orchestrate resolution of severe lung injury throughout the lifespan. The candidate and her mentors have designed a detailed training plan tailored to the candidate's specific needs and goals. The plan includes a rigorous research component that will afford the PI new knowledge and research skills to better examine the links between a critical immune cell population and the endothelium during recovery from viral pneumonia. The proposal concerns viral pneumonia, specifically influenza-induced lung injury, its clinical counterpart the acute respiratory distress syndrome (ARDS) and how they both disproportionately affect the elderly population. Despite decades of dedicated research, there are only a few anti-viral therapies with limited efficacy to manage severe viral pneumonia. Tregs have been shown to decrease inflammation and promote tissue repair in diverse murine models of lung injury. Tregs also increase in the lungs of patients with ARDS, suggesting that they may play a role in the human adaptive immune response to lung injury. However, the specific mechanisms that cause Tregs to execute their pro-repair program following lung injury remain unknown. Our preliminary data shows that the youthful reparative Treg cell program following influenza-induced lung injury is dominated by biologic processes linked to the development and repair of blood capillaries. This reparative program is lost in aged hosts in a cell- autonomous manner. Thus, we hypothesize that aging results in Treg cell-specific downregulation of important pro-angiogenic factors expression such as VEGFA, leading to impaired alveolar endothelial repair and recovery from viral pneumonia. The long-term goal of the proposal is to identify novel small molecule- and cell-based therapeutics to control inflammation and promote tissue repair in our increasingly older population. To test this hypothesis, we propose the following Specific Aims: 1) determine whether age-related alterations in the pro- endothelial repair function of Tregs results from cell-autonomous or microenvironmentally-driven changes, 2) determine whether Treg cell-generated VEFGA is necessary and sufficient to restore the pro-endothelial repair function present in youth that is lost with aging, and 3) determine whether age-related VEGFA expression in alveolar Tregs in bronchoalveolar lavage fluid is associated with 30-day mortality in patients with severe viral pneumonia. We will use standard techniques to assess severity of lung injury, endothelial repair, heterochronic (age mis-matched) adoptive Treg cell transfer, multiple transgenic murine strains for loss-of-function and gain- of-function experiments, flow cytometry, fluorescence-activated cell sorting and transcriptional profiling with RNA-sequencing as the primary methods to support the experimental design of this proposal.