CHARACTERIZATION OF PSEUDOMONAS PHOSPHOLIPASES INVOLVED IN VIRULENCE

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 囊性纤维化(CF)患者最常见的致病和死亡原因是铜绿假单胞菌感染引起的肺系统衰竭。铜绿假单胞菌对肺损伤的一个潜在机制是破坏肺表面活性物质。肺表面活性物质功能障碍可导致小气道关闭，严重降低肺功能。虽然肺表面活性物质功能障碍在慢性阻塞性肺疾病中的临床意义尚不完全清楚，但肺表面活性物质功能障碍可能会在病情加重时导致肺功能下降。我们认为，铜绿假单胞菌分泌的几种毒力因子在感染过程中降解表面活性物质，从而导致肺功能下降。 来自小鼠肺部感染模型的初步证据表明，感染铜绿假单胞菌的小鼠表现出广泛的呼吸道闭合。部分闭合是由于分泌的溶血性磷脂酶C，plcH，但在plcH缺失株中仍存在显著的表面活性物质功能障碍。我们推测，PA4921编码的分泌型脂肪酶是铜绿假单胞菌肺部感染过程中表面活性物质降解的附加介质之一。在这项研究中，我们将验证预测的PA4921的酶活性，并确定其在肺表面活性物质中诱导的转录调控机制。在初步鉴定之后，我们将研究PA4921在野生型和囊性纤维化跨膜调节基因敲除(CFTRKO)小鼠肺力学改变中的作用。在铜绿假单胞菌感染期间，将测量肺力学、表面活性物质组成和表面活性物质功能。肺内滴注纯化的PA4921酶将被测试是否足以改变肺力学和催化表面活性物质的降解。这些实验将使我们能够确定PA4921是否是适用于CF恶化的潜在治疗靶点

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The most common cause of morbidity and mortality in patients with Cystic Fibrosis (CF) is pulmonary system failure caused by Pseudomonas aeruginosa infection. One potential mechanism of P. aeruginosa damage to the lung is destruction of lung surfactant. Surfactant dysfunction can lead to small airway closure and severely decreased lung function. While the clinical significance of surfactant dysfunction in the CF lung is not fully understood, surfactant dysfunction may result in reduced lung function during exacerbations. We propose that several virulence factors secreted by P. aeruginosa function to degrade surfactant during infection resulting in decreased lung function. Preliminary evidence from a mouse lung infection model shows that P. aeruginosa infected mice show extensive airway closure. Some of this closure is due to the secreted hemolytic phospholipase C, PlcH, but in a plcH deletion strain there remains significant surfactant dysfunction. It is our hypothesis that the secreted lipase encoded by PA4921 is one of the additional mediators of surfactant degradation during P. aeruginosa lung infection. In this study we will verify the predicted enzymatic activity of PA4921 and determine the transcriptional regulatory mechanism governing its induction in pulmonary surfactant. Following the initial characterization, we will examine the role of PA4921 in alteration of lung mechanics in both wild type and Cystic Fibrosis Transmembrane Regulator knock-out (CFTR KO) mice. Lung mechanics, surfactant composition, and surfactant function will be measured during P. aeruginosa infection. Pulmonary instillation of purified PA4921 enzyme will be tested for sufficiency to alter lung mechanics and catalyze surfactant degradation. These experiments will allow us to determine if PA4921 is a potential therapeutic target applicable to CF exacerbations