MITOCHONDRIAL FUNCTION IN OXIDATIVE INJURY

氧化损伤与肝脏的病理生理有关 异生物质代谢过程中的损伤，缺血/再灌注损伤，以及 中性粒细胞活化氧化损伤的发生机制多种多样 并且已经用几种不同的模型进行了研究， 制剂和生物制剂。我之前的研究表明果糖， 肝脏中有效的糖酵解底物，可防止各种 氧化剂化学品。这表明线粒体损伤参与了 氧化应激 酸性细胞外pH值也提供了大量 对氧化剂化学品的保护。 这些发现构成了 这个应用程序。 我的总体目标是了解细胞 氧化应激过程中毒性损伤的潜在机制。 具体地说， 我将评估线粒体通透性转换在 毒性损伤诱导的叔丁基过氧化氢（t-BuOOH），甲萘醌和 细胞外ATP在培养的大鼠肝细胞中，我将评估 以下假设：1）这些毒物引起线粒体 通过促进渗透性转变的打开来实现渗透性转变 线粒体内膜中的孔或巨通道。不同 试剂可以通过不同的机制促进通道开放，例如 线粒体内Ca 2+或pH值升高，线粒体氧化 谷胱甘肽（GSH）或吡啶核苷酸，或产生反应性 氧物种。 2)巨通道开放导致线粒体 去极化和氧化磷酸化解偶联。 所得 生物能量缺乏可能作为一个共同的最终途径，导致细胞 不同的氧化剂化学物质的死亡。 糖酵解底物，如 果糖通过提供一种替代的 细胞ATP的来源。 4)酸性pH值，可防止 由多种氧化化学物质引起的致命细胞损伤，抑制 孔开放，并防止线粒体渗透性的发生 过渡因此，我将描述细胞损伤培养 大鼠肝细胞从t-BuOOH，甲萘醌和细胞外ATP方面 剂量反应、营养状况和pH依赖性。利用激光 扫描共聚焦显微镜，l将测量细胞质和 线粒体离子（Ca 2+、H+、Na+、Mg 2+）、胞质和线粒体GSH 和NAD（P）H，细胞溶质和线粒体活性氧，以及 响应于渗透性转变和细胞的起始的Δ-psi 死亡 该项目将提供有关以下方面的基本新信息： 氧化损伤导致细胞死亡的潜在机制。

Oxidative injury has been implicated in the pathophysiology of liver injury during xenobiotic metabolism, ischemia/reperfusion injury, and neutrophil activation. Oxidative injury occurs by a variety of mechanisms and has been studied with several diverse models involving different agents and biological preparations. My previous work shows that fructose, an effective glycolytic substrate in liver, protects against variety of oxidant chemicals. This suggests that mitochondrial injury is involved in oxidative stress. Acidotic extracellular pH also provides substantial protection against oxidant chemicals. These findings form the basis for this application. My overall goal is to understand the cellular mechanisms underlying toxic injury during oxidative stress. Specifically, I will evaluate the role of the mitochondrial permeability transition in toxic injury induced tert-butylhydroperoxide (t-BuOOH), menadione and extracellular ATP. In cultured rat hepatocytes, I will evaluate the following hypotheses: 1) These toxicants cause a mitochondrial permeability transition by promoting opening of permeability transition pores or megachannels in the inner mitochondrial membrane. Different agents may promote channel opening by different mechanisms, such as increases of intramitochondrial Ca2+ or pH, oxidation of mitochondrial glutathione (GSH) or pyridine nucleotides, or production of reactive oxygen species. 2) Megachannel opening leads to mitochondrial depolarization and uncoupling of oxidative phosphorylation. The resulting bioenergetic deficit may serve as a common final pathway leading to cell death for different oxidant chemicals. Glycolytic substrates like fructose rescue hepatocytes from lethal injury by providing an alternate source of cellular ATP. 4) Acidotic pH, which is protective against lethal cell injury caused by a wide variety of oxidant chemicals, inhibits the pore opening and prevents onset of the mitochondrial permeability transition. Accordingly, l will characterize cellular injury to cultured rat hepatocytes from t-BuOOH, menadione and extracellular ATP in terms of dose-response, nutritional status and pH-dependence. Utilizing laser scanning confocal microscopy, l will measure changes of cytosolic and mitochondrial ions (Ca2+, H+, Na+, Mg2+), cytosolic and mitochondrial GSH and NAD(P)H, cytosolic and mitochondrial reactive oxygen species, and delta-psi in response to the onsets of permeability transition and cell death. The project will provide fundamental new information concerning mechanisms underlying cell death resulting from oxidative injury.