Role of mitochondrial calcium in the pathogenesis of alcoholic liver disease

描述（由申请人提供）：酒精滥用对身体几乎每一个组织都有有害影响，是世界范围内发病率和死亡率的主要原因。酒精对细胞功能的急性作用是完全可逆的，而长期酒精中毒可导致不可逆的组织损伤。导致不可逆细胞损伤发生和进展的分子机制仍然知之甚少。在这项应用中，我们提出研究磷酸肌苷依赖信号通路的适应性变化在酒精诱导的组织损伤发病机制中起关键作用的假设。我们的初步数据表明，给大鼠喂食含乙醇的液体饲料（即DeCarli-Lieber） 60天，可以增强肝脏对磷脂酶C-¿偶联激素的敏感性。与对照组相比，添加低浓度的激素可引起从酒精喂养动物分离的肝细胞中更持久的胞质钙增加。此外，酒精动物的细胞在低激素刺激下，肌醇-1,4,5-三磷酸（InsP3）的形成有较大的增加，这表明长期饮酒改变了磷脂酶C-¿的活性。正常情况下，胞质钙的增加激活线粒体生理，使ATP的形成与利用相匹配；然而，长期或不适当的钙增加也会导致基质钙超载和线粒体功能障碍。线粒体损伤是慢性酒精患者和酒精滥用动物模型中观察到的共同特征。线粒体功能障碍可能增加组织对其他类型损伤或凋亡刺激的易感性。这可能对慢性酒精中毒患者尤其重要，慢性酒精中毒患者的促炎细胞因子TNFa水平升高，可通过线粒体依赖途径引起肝细胞凋亡。在这一提议中，我们将(1)表征酒精诱导的磷酸肌醇依赖信号通路的改变，(2)确定对照和长期乙醇喂养大鼠中insp3依赖性钙增加对线粒体钙水平和能量代谢的影响，(3)确定Ca2+动员激素对酒精喂养大鼠及其对偶喂养对照组中tnfa诱导的肝细胞凋亡的影响。拟议的研究将为持续存在的乙醇引起的适应性反应及其对肝脏的相关损伤后果提供新的见解。

DESCRIPTION (provided by applicant): Alcohol abuse has deleterious affects on almost every tissue in the body and is a major cause of morbidity and mortality worldwide. The acute actions of alcohol on cellular function are fully reversible whereas long-term alcohol intoxication can lead to irreversible tissue damage. The molecular mechanisms contributing to the onset and progression of irreversible cell injury are still poorly understood. In this application, we propose to investigate the hypothesis that adaptive changes in the phosphoinositide-dependent signaling pathway play a key role in the pathogenesis of alcohol-induced tissue injury. Our preliminary data indicate that feeding rats an ethanol-containing liquid diet (i.e., DeCarli-Lieber) for 60 days enhances the liver' sensitivity to hormones coupled to phospholipase C-¿. The addition of low concentrations of hormones evoked more sustained cytosolic calcium increases in hepatocytes isolated from alcohol-fed animals compared to their pairfed controls. Moreover, cells from alcoholic animals had a larger increase in inositol-1,4,5-trisphosphate (InsP3) formation following low hormone stimulation suggesting that chronic alcohol consumption altered phospholipase C-¿ activity. Normally, cytosolic calcium increases activate mitochondrial physiology to match ATP formation with utilization; however prolonged or inappropriate calcium increases can also lead to matrix calcium overload and mitochondrial dysfunction. Mitochondrial damage is a common feature observed in chronic alcoholic patients and animal models of alcohol abuse. Mitochondrial dysfunction may increase the tissues' susceptibility to other types of injury or apoptotic stimuli. This may be particularly important in chronic alcoholics that have elevated levels of TNFa, a proinflammatory cytokine, which evokes apoptotic cell death in hepatocytes through the mitochondrial-dependent pathway. In this proposal, we will (1) characterize the alcohol-induced alterations in phosphoinositide-dependent signaling pathway, (2) determine the effects of InsP3-dependent calcium increases on mitochondrial calcium levels and energy metabolism in control and chronically ethanol-fed rats and (3) determine the effects Ca2+-moblizing hormones on TNFa-induced apoptosis in hepatocytes from alcohol-fed rats and their pair-fed controls. The proposed studies will provide new insights into the adaptive responses evoked by the sustained presence of ethanol and the associated injurious consequences for the liver.