Cytoprotective cytokine signaling and reperfusion injury

项目4是HL 42550的一个新项目，其长期目标是描述最近观察到的通过肿瘤坏死因子（TNF）受体或gp 130受体家族发出信号的促炎细胞因子在心肌细胞中赋予细胞保护/抗凋亡作用的机制。为此，项目4的直接具体目标将是描述这些细胞因子在实验性心肌缺血/再灌注（I/R）损伤中的细胞保护作用的机制。具体目标1将检验以下假设：小鼠I/R损伤期间诱导的活性氧中间产物负责通过线粒体功能障碍、细胞色素c释放、半胱天冬酶9和3活性增加以及DNA降解引起心肌细胞凋亡。具体目标2将检验TNF通过减弱和/或预防线粒体功能障碍来防止I/R诱导的心肌细胞凋亡的假设，方法是检查具有低水平TNF心脏限制性过表达的转基因小鼠和两种TNF受体敲除小鼠中I/R损伤的影响。 具体目标3将通过检查TRAF 2显性阴性构建体条件性过表达的转基因小鼠中I/R损伤的影响，检验TNF的细胞保护作用由TNF受体相关因子2（TAF 2）介导的假设。具体目标4将检验以下假设：gp 130通路的激活通过减弱半胱天冬酶9和3的活性来保护成年心肌细胞免受I/R诱导的细胞凋亡，以及gp 130损伤在具有白血病抑制因子心脏限制性表达的转基因小鼠和具有gp 130心室限制性敲除的小鼠中的细胞保护作用。上述研究将在全身I/R损伤的Langerdorff缓冲液灌注模型、I/R损伤的闭胸模型和缺氧/复氧的分离小鼠心肌细胞模型中进行。总之，这些研究将提供明确的新的信息有关的基本机制负责的促炎细胞因子的细胞保护作用后I/R损伤。

Project 4 is a new project of HL 42550, whose long-term objective is to delineate the mechanisms that are responsible for the recent observation that pro-inflammatory cytokines that signal through the tumor necrosis factor (TNF) receptors or the gp130 family of receptors confer cytoprotective /anti-apoptotic effects in cardiac myocytes. To this end the immediate specific objectives of Project 4 will be to delineate the mechanisms for the cytoprotective effects of these cytokines in the setting of experimental myocardial ischemia/reperfusion (I/R) injury. Specific Aim 1 will test the hypothesis that reactive oxygen intermediates induced during I/R injury in mice are responsible for provoking cardiac myocyte apoptosis through mitochondrial dysfunction, release of cytochrome c, increased activity of caspases 9 and 3, and DNA degradation. Specific Aim 2 will test the hypothesis that TNF protects against I/R induced myocyte apoptosis by attenuating and/or preventing mitochondrial dysfunction, by examining the effects of I/R injury in transgenic mice with low levels of cardiac restricted over-expression of TNF and knockout mice both TNF receptors. Specific Aim 3 will test the hypothesis that the cytoprotective effects of TNF are mediated by TNF receptor associated factor 2 (TAF2), by examining the effects of I/R injury in transgenic mice with conditional over-expression of a TRAF2 dominant negative construct. Specific Aim 4 will test the hypothesis that activation of the gp130 pathway protects the adult cardiac myocyte against I/R induced apoptosis by attenuating the activity of caspases 9 and 3, and that the cytoprotective effects of the gp130 injury in transgenic mice with cardiac restricted expression of leukemia inhibitory factor and mice with a ventricular restricted knockout of gp130. The above studies will be conducted in a Langerdorff buffer perfusion model of global I/R injury, a closed-chest model of I/R injury, and in isolated murine cardiac myocyte model subjected to hypoxia/reoxygenation. Taken together, these studies will provide definitive new information with regard to the basic mechanisms responsible for the cytoprotective effects of pro- inflammatory cytokines following I/R injury.