Mitochondria and Pulmonary Endothelial Cell Death

线粒体和肺内皮细胞死亡

• 批准号: 6776070
• 负责人: JAMES PETERSON
• 金额: $ 28.02万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-04-01 至 2008-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6776070
• 关键词: NAD(P)H dehydrogenase; NADPH cytochrome c2 reductase; SDS polyacrylamide gel electrophoresis; animal tissue; cell death; cytochrome oxidase; enzyme activity; enzyme inhibitors; enzyme mechanism; enzyme substrate; free radical oxygen; free radicals; hypoxia; in situ hybridization; liquid chromatography mass spectrometry; mitochondria; molecular site; nitric oxide; nitrogen compounds; oxidative phosphorylation; oxidative stress; protein purification; pulmonary circulation; respiratory epithelium; tissue /cell culture; vascular endothelium

DESCRIPTION (provided by applicant): The pulmonary vasculature, particularly the endothelium, is known to be a key site responsive to both physiological and pathological changes in 02 delivery and exposure to endogenous NO. Mitochondria are critical loci of cellular respiration, biosynthesis, and metabolism of reactive oxygen species (ROS) and reactive nitrogen species (RNS). We have established that mitochondrial respiratory complexes I (NADH dehydrogenase) and Ill (cytochrome c reductase) are primary locations of irreversible inhibition by RNS and that complex IV (cytochrome c oxidase), in addition to its known cytochrome c:O2 oxidoreductase activity, may also function as an NO oxidase. Accordingly, we hypothesize that under normal physiological circumstances, the NO oxidase activity of complex IV limits formation of RNS by rapidly converting NO to the relatively innocuous nitrite ion. The efficiency of this catabolic reaction, which also consumes 02, depends upon the prevailing NO/O2 ratio. Under hyperoxic conditions, or in pathophysiological circumstances where the electron-transport chain has been compromised, RNS (and ROS) formation is not effectively suppressed. This results in irreversible inhibition of complexes I and III, which in turn exacerbates the situation by increasing the production of the damaging reactive species and ultimately, leads to cell death. In testing this broad hypothesis, we propose to determine: 1) that the NO oxidase activity of complex IV limits nitrosative stress in pulmonary endothelial cells; 2) the molecular sites at which RNS irreversibly inhibit complex I and complex III; 3) the extent to which nitrosative stress may affect the interaction of cytochrome c with complex Ill and/or complex IV; 4) the functional role of RNS-dependent modification of the interaction between cytochrome c and complexes III and/or IV in promoting the loss of cytochrome c from the intermembrane space during proapoptotic stimulation of cultured pulmonary endothelial cells.

描述（由申请人提供）：已知肺脉管系统，尤其是内皮，是对02递送和暴露于内源性NO的生理和病理变化的关键部位。线粒体是细胞呼吸，生物合成以及活性氧（ROS）和反应氮种（RNS）的定型的关键基因座。我们已经确定，线粒体呼吸复合物I（NADH脱氢酶）和ILL（细胞色素C还原酶）是RNS不可逆性抑制的主要位置以及该复合物IV（细胞色素C氧化酶），除了其已知的细胞染料C：O2氧化氧化氧化物酶活性外，也可能是氧化氧化酶的活性。因此，我们假设在正常的生理环境下，复杂IV的无氧化酶活性通过将NO迅速转化为相对无害的亚硝酸盐离子，从而形成了RN的RN。这种分解代谢反应的效率也消耗了02，取决于普遍的NO/O2比。在高氧化条件下，或者在电子传输链被损害的病理生理状况下，RN（和ROS）形成没有被有效抑制。这导致对复合物I和III的不可逆转抑制，这反过来又通过增加破坏性反应性物种的产生并最终导致细胞死亡加剧了这种情况。在检验这一广泛的假设时，我们建议确定： 1）复合静脉内的无氧化酶活性限制了肺内皮细胞中的硝化应激； 2）RN不可逆地抑制复合物I和复合物III的分子位点； 3）亚硝化应激可能影响细胞色素c与复杂疾病和/或复合物IV的相互作用； 4）在培养的肺内皮细胞的促凋亡刺激过程中，细胞色素C和复合物III和/或IV之间相互作用的变化在促进膜间空间中的细胞色素C丧失方面的相互作用的功能作用。