Characterizing the role of peroxiredoxin 6 in Alcoholic Liver Disease.

这项建议的长期目标是研究过氧化还蛋白6(PRX6)在心脏疾病中的抗氧化作用。 以及慢性酒精引起的氧化应激如何影响这种蛋白质的活性 消费。这个项目有三个拟议的具体目标。第一个具体目标是确定 并表征了4-羟基壬烯醛和4-羟基酮烯醛修饰的位置和生物学效应。 重组PRX6。加合物将在体外用液相色谱和 串联质谱仪。分子模拟将被用来研究任何构象变化 由于乙醛的修饰和生化分析，将评估其生物学意义 修改后的版本。第二个具体目的是评估PRX6在酒精中毒进展中的作用 利用各种啮齿动物模型制作肝病模型。野生型小鼠将被长期喂以含乙醇的饮食 醛-蛋白质加合物的数量将通过二维在不同的时间点进行评估 电泳法和蛋白质印迹法。此外，PrX6/-基因敲除小鼠将被长期喂食乙醇 为了研究PRX6在肝脏中是否是一种重要的抗氧化剂，我们进行了一项研究。最后， 转基因PRX6过表达小鼠将用于评估其对酒精性肝脏的保护作用 由于这种过度表达。最后一个具体目标是评估任何可能的抗氧化剂。 野生型和PrX6基因敲除小鼠肝脏慢性酒精消耗的代偿作用。 利用分离的肝细胞和肝组织匀浆，通过定量RT-PCR，蛋白质 通过Western blotting表达和酶活性将评估主要的细胞抗氧化剂 蛋白质，即过氧化氢酶、谷胱甘肽过氧化物酶和超氧化物歧化酶。长期大量饮酒是 在美国，肝病是导致疾病和死亡的主要原因。因此，酒精性肝病 (ALD)是一个主要的公共卫生问题。ALD是一种多因素疾病，其中氧化应激是 这是一个已知的促成因素。因此，通过深入研究PRX6在骨肉瘤发病机制中的作用。 ALD进展科学界可以向更好地了解这种疾病更近一步 更接近于设计预防ALD进展的治疗策略。

The long term objective of this proposal is to investigate the antioxidant role of peroxiredoxin 6 (PRX6) in the liver and how the activity of this protein might be affected by oxidative stress due to chronic ethanol consumption. There are three proposed specific aims for this project. The first specific aim is to identify and characterize the location and biological effect of 4-hydroxynonenal and 4-oxononenal modification of recombinant PRX6. Adducts will be identified and characterized in vitro using liquid chromatography and tandem mass spectrometry. Molecular modeling will be used to investigate any conformational changes due to aldehyde modification and biochemical assays will be performed to assess biological significance of the modification. The second specific aim will evaluate the role of PRX6 in the progression of alcoholic liver disease using various rodent models. Wild type mice will be chronically fed an ethanol containing diet and the number of aldehyde-protein adducts will be assessed at various time points via two dimensional electrophoresis and Western blots. Also, PRX6 -/- knockout mice will be chronically fed an ethanol containing diet in order to investigate whether or not PRX6 is an important antioxidant in the liver. Lastly, transgenic, PRX6 over-expressing mice will be used to evaluate the protective effects in the alcoholic liver due to this over-expression. The last specific aim is designed to evaluate any possible antioxidant compensation due to chronic ethanol consumption in the liver of wild type and PRX6 -/- knockout mice. Using isolated hepatocytes and liver homogenates, mRNA expression via quantitative RT-PCR, protein expression via Western blotting and enzymatic activity will be assessed for the major cellular antioxidant proteins, i.e. catalase, glutathione peroxidase, and superoxide dismutase. Long-term, heavy alcohol use is a leading cause of illness and death from liver diease in the United States. As such, alcoholic liver disease (ALD) represents a major public health concern. ALD is a multifactorial disease in which oxidative stress is a known contributing factor. Therefore, by thoroughly investigating the role of PRX6 in the mechanism of ALD progression the scientific community can move a step closer to a better understanding of the disease and closer to designing treatment strategies for preventing the advancement of ALD.