BACTERIAL SURFACE PROTEINS: POTENTIAL TARGETS FOR SEPSIS

释放到血流中的细菌细胞壁成分被认为在引发导致革兰氏阴性脓毒症的炎症反应中起着重要作用。细菌外膜含有脂多糖(LPS)、外膜蛋白(OMP)和磷脂。脂多糖作为脓毒症的重要介质已被广泛研究，但OMPS在脓毒症中的重要性尚未得到系统评价。我们的初步数据表明，含有三个保守的OMPS和内毒素的复合体从细菌释放到人血清中，也释放到脓毒症大鼠的循环中。我们已经鉴定了这三种蛋白质，并发现至少有一种蛋白质具有生物活性。本项目的主要目标是进一步确定脓毒症时三种OMP的释放特征，评估它们在脓毒症发病机制中的作用，并评价抗OMP免疫作为一种治疗脓毒症的手段。第一个具体目标是评估人类革兰氏阴性脓毒症和两种实验性革兰氏阴性脓毒症大鼠模型中OMPS和OMP/LPS复合体的释放。第二个特异性目的是通过研究对内毒素抵抗和内毒素应答的小鼠的致死率、血液和器官中细胞因子的诱导以及肺中性粒细胞的隔离，来评价单个OMP、OMP/LPS复合体和完整细菌表面的OMP在体外对分离的免疫细胞和体内的生物活性。第三个具体目的是评价主动免疫对OMP制剂在小鼠和大鼠脓毒症感染模型中毒性的保护作用。申请者的长期职业目标是成为一名研究革兰氏阴性感染和败血症基本机制的独立调查员。职业发展计划旨在通过实践研究和教育活动扩大应聘者的研究知识库。这些将包括课程工作(实验室和教学)，出席和参加实验室会议和研究研讨会，以及与咨询委员会成员的正式和非正式互动。候选人的最终目标是应用败血症相关机制的基本知识来开发针对细菌毒素的抗败血症策略。

Bacterial cell wall components released into the bloodstream are believed to be important in eliciting the inflammatory response that causes Gram-negative Sepsis. The bacterial outer membrane contains lipopolysaccharide (LPS), outer membrane proteins (OMPs), and phospholipids. LPS has been studied extensively as an important mediator in sepsis; the importance of OMPs in sepsis has not been systematically evaluated. Our preliminary data indicate that complexes containing three conserved OMPs and LPS are released from bacteria into human serum and are also released into the circulation of septic rats. We have identified the three proteins and have found that at least one of the proteins has biological activity. The major goals of this project are to further characterize release of the three OMPs in sepsis, to assess their roles in the pathogenesis of sepsis, and to evaluate anti-OMP immunity as a means of treating sepsis. The first specific aim is to assess release of OMPs and OMP/LPS complexes in humans with Gram- negative sepsis and in two models of experimental Gram-negative sepsis in rats. The second specific aim is to evaluate individual OMPs, OMP/LPS complexes, and OMPs on the surface of intact bacteria for biological activity in vitro on isolated immune cells, an in vivo by studying lethality, induction of cytokines in blood and organs, and pulmonary neutrophil sequestration in LPS resistant and LPS responsive mice. The third specific aim is to evaluate protective effects of active immunity on toxicity of OMP containing preparations in mice and in the rat infection models of sepsis. The applicant's long-term career objective is to become an independent investigator studying basic mechanisms of Gram- negative infection and sepsis. The career development plan is designed to expand the candidate's research knowledge base through hands-on research and educational activities. These will include course work (laboratory and didactic), attendance at and participation in laboratory meetings and research seminars, and formal and informal interactions with the Advisory Committee members. The ultimate goal of the candidate is to apply the fundamental knowledge of mechanisms involved in sepsis to develop anti-sepsis strategies that target bacterial toxins.