Gelsolin modulation of airway hyperresponsiveness and inflammation

项目摘要/摘要 文献中有新的证据表明，不同类别的治疗哮喘的新型配体可以显著 松弛气道平滑肌(ASM)，尽管细胞内钙浓度短暂增加，这是 传统上一直与收缩联系在一起，而不是放松。明胶是一种钙激活的肌动蛋白 使肌动蛋白细胞骨架解聚，导致肌浆网松弛的蛋白质。因此，明胶蛋白的激活通过 一过性细胞内钙升高可能是ASM松弛的统一机制 通过这些不同的配体。在这项提案中，我们提供了令人兴奋的初步数据，证明了老鼠 缺乏遗传的明胶在体外、体外和体内都表现出ASM松弛受损。除了切断 肌动蛋白，细胞内的明胶蛋白具有与磷脂酰肌醇4，5二磷酸(PIP2)的结合域，这是 肌醇三磷酸的磷脂酶Cβ合成底物 钙的释放和收缩。我们将进一步证明，控制细胞内明胶蛋白的表达和 激活是慢病毒介导的全长过表达ASM细胞松弛的一种治疗策略 通过引入含有PIP2的结合域的短的明胶蛋白多肽，这将 选择性地抑制GQ介导的细胞内钙的增加。这项提议也证明了明胶的作用 在肺部炎症的发展过程中，这是哮喘的另一个关键病理特征。我们建议 为了证明大量表达明胶蛋白的肺巨噬细胞可以成为 过敏性肺部炎症中的免疫调节。我们将演示局部递送多肽片段 明胶能减少呼吸道平滑肌收缩和过敏性肺部炎症。 一个由成功的科学家和内科科学家组成的指导委员会将提供科学 研究计划各个方面的专业知识，包括离子通道生理学、钙信号、Th2肺 炎症、肺免疫学和巨噬细胞生物学。校内校外的综合规划 课程和培训辅以更多具有精确切割肺切片和 磁扭转细胞细胞术将拓展研究、培训和职业发展。研究课题 是在呼吸道平滑肌病理生理学和肺免疫领域进行独立研究的理想途径 过敏性肺部炎症的细胞生物学。作为一个整体，这份K08提案勾勒出了一条稳健的道路 科学独立和作为一名内科科学家成功和持续职业生涯的基础。

Project Summary/Abstract There is emerging evidence in the literature that diverse classes of novel ligands for asthma can acutely relax airway smooth muscle (ASM) despite a transient increase in intracellular calcium concentrations, which has classically been associated with contraction, not relaxation. Gelsolin is a calcium-activated actin severing protein that depolymerizes the actin cytoskeleton leading to ASM relaxation. Thus, activation of gelsolin by transient intracellular calcium increases may be the unifying mechanism that would account for ASM relaxation by these diverse ligands. In this proposal, we present exciting preliminary data demonstrating that mice genetically lacking gelsolin exhibit impaired ASM relaxation in vitro, ex vivo and in vivo. In addition to severing actin, intracellular gelsolin has a binding domain for phosphatidylinositol 4,5 bisphosphate (PIP2), which is the substrate for phospholipase Cβ synthesis of inositol triphosphate (IP3), the critical regulator of smooth muscle calcium release and contraction. We will further demonstrate that controlling intracellular gelsolin expression and activation is a therapeutic strategy for ASM cell relaxation using lentiviral-mediated overexpression of full length gelsolin and by the introduction of a short gelsolin peptide containing the binding domain for PIP2, which would selectively impair Gq-mediated increases in intracellular calcium. This proposal also demonstrate gelsolin’s role in the development of lung inflammation, which is another key pathological characteristic in asthma. We propose to demonstrate that pulmonary macrophages, in which gelsolin is abundantly expressed, can be a target for immune modulation in allergic lung inflammation. We will demonstrate that local delivery of a peptide fragment of gelsolin decreases airway smooth muscle contraction and allergic lung inflammation. A mentoring committee composed of successful scientists and physician-scientists will provide scientific expertise in each aspect of the research plan including ion channel physiology, calcium signaling, Th2 lung inflammation, lung immunology and macrophage biology. A comprehensive plan of intramural and extramural coursework and training complemented by additional collaborators with expertise in precision cut lung slices and magnetic twisting cell cytometry will expand the research training and career development. The research topic is ideal for a path to independent research in areas of airway smooth muscle pathophysiology and lung immune cell biology in allergic lung inflammation. Taken as a whole, this K08 proposal outlines a robust pathway to scientific independence and the foundation of a successful and sustained career as a physician-scientist.