MITOCHONDRIAL FUNCTION IN OXIDATIVE INJURY

氧化损伤与肝脏的病理生理学有关。 异种代谢中的损伤、缺血/再灌注损伤和 中性粒细胞激活。氧化损伤通过多种机制发生。 已经用几个不同的模型进行了研究，这些模型涉及不同的 制剂和生物制剂。我之前的研究表明，果糖， 肝脏中一种有效的糖酵解底物，可防止多种 氧化剂化学品。这表明线粒体损伤参与了 氧化应激。酸化的胞外pH也提供了大量的 保护身体免受氧化化学物质的伤害。这些发现构成了以下结论的基础 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。具体来说， 我将评估线粒体通透性转变在 叔丁基氢过氧化氢(t-BuOOH)、甲萘二酮和丙二醛的毒性损伤 胞外三磷酸腺苷。在培养的大鼠肝细胞中，我将评估 假设如下：1)这些毒物导致线粒体 通过促进渗透率转变的开放来实现渗透率转变 线粒体膜内的小孔或巨细胞。不同 代理商可以通过不同的机制来促进渠道开放，例如 线粒体内钙离子或pH升高，线粒体氧化 谷胱甘肽(GSH)或吡啶核苷酸，或产生反应性 氧气物种。2)Megachannel开口导致线粒体 氧化磷酸化的去极化和解偶联。由此产生的 生物能量缺乏可能是通向细胞的共同的最终途径 因不同的氧化剂化学物质而死亡。糖酵解底物，如 果糖通过提供一种替代物使肝细胞免于致命性损伤 细胞三磷酸腺苷的来源。4)酸性酸碱度，可防止 由多种氧化剂化学物质引起的致命细胞损伤，抑制 孔洞的打开和防止线粒体通透性的开始 过渡。据此，L将以培养的细胞损伤为特征 T-BuOOH、甲萘二酮和细胞外ATP对大鼠肝细胞的影响 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将测量胞浆和 线粒体离子(Ca~(2+)、H~+、Na~+、Mg~(2+))、胞浆和线粒体GSH 和NAD(P)H，胞质和线粒体活性氧，以及 渗透率转变和胞体开始时的增量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.