GSH MEDIATED DETOXIFICATION OF HNE IN MITOCHONDIRA

我们实验室和其他实验室的一致发现是， 对乙醇消耗的反应包括对线粒体（M）的损伤， 呼吸和氧化应激的诱导。我们最近的研究表明， 证明了脂质过氧化的有毒产物4-羟基壬烯醛 (HNE)抑制细胞色素c氧化酶（CO），一个关键组成部分， 呼吸链 这可能是乙醇抑制 能源生产。初步研究表明， 谷胱甘肽S转移酶（GST）介导的HNE与GSH的结合是一种免疫抑制剂。 抗HNE抑制CO的重要防线。乙醇 消耗GSH的M，它可以改变GST活性，并且乙醇相关 氧化应激在M内产生HNE。 因此，我们建议确定 乙醇消耗对GST介导的HNE解毒作用的影响 并确定GSH补充方案，这可能会防止这种情况。 假说. 我们假设一种重要的细胞防御机制 乙醇相关氧化应激对CO的抑制作用是 线粒体GST介导HNE和乙醇的缀合 消费会损害这种解毒系统。 具体目标。 1. 定量线粒体GST在细胞凋亡中的作用， 对M. 2. 确定影响 线粒体GST/HNE结合系统上的乙醇消耗， 确定在何种条件下可以作出这种防御机制 在乙醇暴露期间发挥最佳功能。 实验这些研究将利用来自大鼠肝脏的M来建立 急性和慢性E摄入对GST-HNE结合系统的影响， 终点是GSH-HNE结合物的产生，CO的抑制， 和形成HNE-CO加合物（HNE抑制CO的一种方式）。 此外，在乙醇处理过程中， 将确定这些参数的消耗量。 总之，乙醇相关的氧化应激在M内产生HNE， 或靠近CO的位置，并且该HNE抑制该酶。 此外，乙醇可以消耗细胞器中的谷胱甘肽所需的 GST介导的HNE结合可以防止CO的这种抑制。 我们认为，这是一个新的机制，有待记录， 可以保护关键的M功能免受乙醇诱导的氧化应激。 这是这一探索性建议的重点。

Consistent findings from our laboratory and others are that hepatoxic responses to ethanol consumption include damage to mitochondrial (M) respiration and induction of oxidative stress. Recent studies by us have documented that a toxic product of lipid peroxidation, 4-hydroxynonenal (HNE) inhibits cytochrome c oxidase (CO), a key component of the respiratory chain. This may be one mechanism by which ethanol inhibits M energy production. Preliminary studies, have illustrated that glutathione S transferase (GST)-mediated HNE conjugation with GSH is an important line of defense against the inhibition of CO by HNE. Ethanol depletes M of GSH, it can alter GST activity, and ethanol-related oxidative stress produces HNE within M. Thus, we propose to determine the effects of ethanol consumption on GST-mediated detoxification of HNE and define regimens of GSH repletion which may prevent this. Hypothesis. We hypothesize that an important cellular defense against the inhibition of CO by ethanol-related oxidative stress is mitochondrial GST-mediated conjugation of HNE and that ethanol consumption can impair this detoxification system. Specific Aims. 1. Quantitate the role of mitochondrial GST in the detoxification of HNE generated within M. 2. Determine the effects of ethanol consumption on the mitochondrial GST/HNE conjugation system and ascertain the conditions under which this defense mechanism can be made to function optimally during ethanol exposure. Experiments. The studies will utilize M from rat livers to establish effects of acute and chronic E intake on the GST-HNE conjugating system, with endpoints being production of GSH-HNE conjugates, inhibition of CO, and formation of HNE-CO adducts (a means by which HNE inhibits CO). Additionally, the effects of normalizing M pools of GSH during ethanol consumption on these parameters will be determined. In Summary, ethanol-related oxidative stress produces HNE within M at or near the location of CO and this HNE inhibits the enzyme. Additionally, ethanol can deplete the organelle of GSH needed for the GST-mediated HNE conjugation that can prevent this inhibition of CO. We submit that this is a new and yet to be documented mechanism by which a key M function can be protected from ethanol-induced oxidative stress. It is the focus of this exploratory proposal.