Apoptotic Signaling Induces Hyperpermeability Following Hemorrhagic Shock

描述(由申请人提供)： 这份申请描述了一个全面的血管研究计划，旨在将蔡尔兹博士培养成一名独立的研究员。通过分子生物学、研究设计和统计学方面的课程工作，他将获得实现这一目标所需的分析技能。他还将被要求参加几次年度会议和一个月刊俱乐部。成立了一个由血管和细胞生物学方面经验丰富的研究人员组成的咨询委员会，让查尔兹博士获得实验室技术方面的实践经验。在K01奖项的整个颁奖期间，我们都强调资助人精神的发展，从申请当地基金开始，毕业到美国心脏协会的赠款，最后在该奖项的第三年向美国国立卫生研究院提交R01。该研究计划的重点是调节内皮细胞-细胞黏附连接，以防止失血性休克后微血管的高通透性。在创伤、烧伤、败血症和失血性休克后，循环中的液体丢失是影响心血管内环境平衡的主要因素。防止微血管屏障开放或重建屏障功能对于限制对心血管、肺和神经系统的有害影响至关重要。失血性休克引起的高通透性导致内皮细胞-细胞黏附连接复合体的改变。3-连环蛋白结构的特异性改变被认为是这些连接的一个重要的调节器。申请人假设失血性休克诱导的caspase激活导致内皮细胞黏附连接的(3-连环蛋白依赖的修饰)。为了解决这一假设，申请人提出了两个相互关联的特定目标：(1)确定细胞凋亡级联和失血性休克诱导的高通透性之间的关系；(2)确定caspase-3与黏附连接复合体的相互作用(S)。活体显微镜和一种新的体内蛋白转染法与细胞单层并行，将被用来确定微血管高通透性的变化。原子力显微镜将被用来评估内皮细胞的粘附力。(摘要结束)

DESCRIPTION (provided by applicant): This application describes a comprehensive vascular research program designed to develop Dr. Childs into an independent investigator. Through course work in molecular biology, research design, and statistics, he will acquire the necessary analytical skills to achieve this goal. He will also be required to attend several annual conferences and a monthly journal club. An Advisory Board of experienced investigators in vascular and cellular biology has been established for Dr. Childs to obtain hands on experience in laboratory techniques. Development of grantsmanship is emphasized throughout the duration of the K01 award, beginning with applications for local funds, graduating to an American Heart Association grant, and culminating in a R01 submission to the NIH in the third year of this award. The research plan focuses on the regulation of endothelial cell-cell adherens junctions to prevent microvascular hyperpermeability following hemorrhagic shock. Fluid loss from the circulation is a major factor in compromising cardiovascular homeostasis following trauma, burns, sepsis and hemorrhagic shock. Preventing microvascular barrier opening or reestablishing barrier function is crucial in limiting the deleterious effects on the cardiovascular, pulmonary and nervous systems. Hemorrhagic shock-induced hyperpermeability results in alteration of the endothelial cell-cell adherens junction complex. Site-specific alterations in the structure of (3-catenin have been implicated as an important modulator of these junctions. The applicant hypothesizes that hemorrhagic shock-induced activation of caspases results in a (3- catenin-dependent modification of endothelial adherens junction. To address this hypothesis, the applicant proposes two interlocking specific aims: (1) define the relationship between the apoptotic cascade and hemorrhagic shock-induced hyperpermeability and (2) identify the interaction(s) of caspase-3 with the adherens junction complex. Intravital microscopy with a novel in vivo protein transfection technique in parallel with cell monolayers will be utilized to determine changes in microvascular hyperpermeability. Atomic force microscopy will be used to assess endothelial cell adhesive forces. (End of Abstract)