Innate immune defense against community associated methicillin-resistant S.aureus

描述（由申请人提供）：社区相关性耐甲氧西林金黄色葡萄球菌（CA-MRSA）经常引起坏死性感染，比其他SA更严重。它们的流行蔓延和在健康个体中的高发病率和死亡率引起了生物医学界的警惕。由于CA-MRSA感染的临床特征反映了其与宿主相互作用的独特特征，我们的注意力将集中在CA-MRSA（对照SA）与防御SA的先天免疫元件之间的动态相互作用上。关键的决定因素可能在最初的相互作用中表现出来，后来在感染期间幸存的细菌逃离吞噬细胞，或两者兼而有之。我们最近发现了依赖于o2的吞噬细胞NADPH氧化酶和细胞外不依赖于o2的IIA组磷脂酶A2 （GpIIA-PLA2）在杀死和降解SA中的一种新的协同作用。我们的数据表明CA-MRSA不仅对多形核中性粒细胞（PMN）具有相对抗性，而且对GpIIA-PLA2也具有相对抗性。此外，我们发现CA-MRSA更容易从PMN中逃逸，并且逃逸的CA-MRSA对GpIIA-PLA2的抗性比初始CA-MRSA更强。被吞噬的SA对PMN的摄入表现出强大的代谢反应，包括编码细胞壁修饰的基因的表达，这可能对细胞内存活、PMN的溶解和持续炎症很重要。我们的数据表明，CA- MRSA对GpIIA-PLA2的抗性增加需要dlt操纵子的表达，dlt操纵子是细胞壁合成的关键决定因素。在此背景下，我们提出了以下新的假设：(1)PMN在解决SA感染中的作用取决于PMN内在抗菌作用与动员的细胞外抗SA因子活性的综合作用，包括GpIIA-PLA2和更长寿的吞噬细胞如巨噬细胞；(2)初始SA-PMN相互作用的差异改变了CA-MRSA在PMN内的命运以及感染的迅速解决，表现为存活的CA-MRSA对GpIIA-PLA2或巨噬细胞作用的易感性改变；(3) CA-MRSA感染的坏死特征反映了PMN、GpIIA-PLA2和巨噬细胞的集体作用不太有效。为了验证这些假设，我们将采用新颖的实验设计来追求以下目标：确定CA-MRSA对吞噬细胞（PMN）和细胞外（IIA PLA2组）细胞毒性抗性增强的机制。2. 确定PMN-CA-MRSA相互作用对每个细胞命运的影响3。评估CA-MRSA对PMN- GpIIA-PLA2的特异性反应对其与PMN相互作用的结果或随后的命运的贡献。最近发现的一种葡萄球菌（社区相关性耐甲氧西林金黄色葡萄球菌，CA-MRSA）已成为社区（即非住院个体）感染的重要和常见原因。受影响的个体通常是年轻人，以前健康，并遭受严重感染，通常是坏死性肺炎和软组织疾病。我们的研究将进一步了解CA-MRSA如何抵抗人类宿主的防御，从而产生如此严重和毁灭性的疾病。

DESCRIPTION (provided by applicant): Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) frequently cause necrotizing infections that are more severe than those seen with other SA. Their epidemic spread and high morbidity and mortality in otherwise healthy individuals have raised alarm in the biomedical community. Reasoning that the clinical characteristics of CA-MRSA infections reflect unique features of its interactions with the host, our attention will be focused on the dynamic interplay between CA-MRSA (vs control SA) and elements of innate immunity that defend against SA. Critical determinants may be manifested during the initial interaction, later during infection as surviving bacteria escape the phagocyte, or both. We have recently identified a novel synergy between the O2-dependent phagocyte NADPH oxidase and the extracellular O2-independent Group IIA phospholipase A2 (GpIIA-PLA2) in the killing and degradation of SA. Our data suggest that CA-MRSA are relatively resistant not only to polymorphonuclear neutrophil (PMN) but also to GpIIA-PLA2. Furthermore, we found that CA-MRSA more readily escape from PMN and that escaped CA-MRSA are more resistant to GpIIA-PLA2 than are naive CA-MRSA. Phagocytosed SA exhibit robust metabolic responses to ingestion by PMN, including expression of genes encoding cell wall modifications that may be important for intracellular survival, lysis of PMN, and persistent inflammation. Our data demonstrate that the increased resistance of CA- MRSA to GpIIA-PLA2 requires expression of the dlt operon, a critical determinant of cell wall synthesis. Within this context, we propose the following novel hypotheses: (1) the role of PMN in resolution of SA infection depends on the integrated effects of intrinsic PMN antibacterial action with the activity of mobilized extracellular anti-SA factors, including GpIIA-PLA2 and longer lived phagocytes such as macrophages; (2) differences in initial SA-PMN interactions alter the fate of CA-MRSA within PMN as well as the prompt resolution of infection, manifested by altered susceptibility of surviving CA-MRSA to the actions of GpIIA-PLA2 or macrophages; and (3) the necrosis characteristic of CA-MRSA infection reflects the less effective collective actions of PMN, GpIIA-PLA2 and macrophages. To test these hypotheses, we will employ novel experimental design to pursue the following aims: 1. To determine the mechanism(s) underlying the increased resistance of CA-MRSA to phagocyte (PMN) and extracellular (Group IIA PLA2) cytotoxicity. 2. To determine the effects of PMN-CA-MRSA interaction on the fate of each cell 3. To assess the contribution of specific CA-MRSA responses to PMN- GpIIA-PLA2 to the outcome of their interaction with PMN or subsequent fate. Project Narrative A recently identified type of staphylococcus (community-associated methicillin-resistant Staphylococcus aureus, CA-MRSA) has emerged as a significant and common cause of infection in the community (i.e. in non hospitalized individuals). Affected individuals are generally young and previously healthy, and suffer severe infection, often necrotizing pneumonia and soft-tissue disease. Our studies will advance understanding of how CA-MRSA resist human host defenses and thereby produce such severe and devastating disease.