Effect of HIV protease inhibitor on alcohol induced ER stress and liver injury.

描述(申请人提供)：HIV蛋白酶抑制剂(HIV PI)用于高效抗逆转录病毒疗法(HAART)。然而，HIV PI通常与肝脏损害的发生有关。HIV蛋白水解酶抑制剂引起肝损伤的机制尚不清楚。新的证据表明，HIV PIs诱导内质网(ER)应激反应，已被证明在包括坏死性炎症、肝细胞死亡和脂肪肝在内的肝功能障碍中起重要作用。我们以前观察到，饮酒是公认的感染艾滋病毒和进展的易感性的辅助因素，它会诱导肝脏中的高同型半胱氨酸血症和内质网应激反应。相当大比例的艾滋病毒感染者酗酒，认为酒精和艾滋病毒PIs之间的相互作用会导致严重的肝脏脂肪变性和损伤，这是合乎逻辑的。我们假设HIV、PI和酒精对内质网应激和脂代谢有增强作用，从而加重肝损伤。我们打算在原代培养的小鼠和人肝细胞中探索HIV PI诱导的内质网应激反应，并在体内研究其与酒精对喂饲酒精的动物的致病作用的相互作用。我们的具体目标是：(1)比较单独或联合使用HIV PI在慢性配对和酒精喂养的小鼠体内引起的ER应激和肝损伤，并通过检测HIV PI在体外诱导的原代小鼠和人肝细胞的ER应激反应，以及通过监测HIVPI在血浆和肝细胞培养上清液中的生物利用度来评估相加或协同效应；(2)证实在存在或不存在模仿人类抗逆转录病毒治疗的单一或联合HIVPI的情况下，口服高蛋氨酸低叶酸饮食的小鼠的ER应激反应；(3)通过监测HIVPI诱导的原代小鼠和人肝细胞胞浆内钙释放，以及检测HIVPI诱导的肝细胞蛋白酶体活性、葡萄糖转运体的表达和活性，研究HIVPI和酒精诱导的ER应激反应的协同机制；(4)分离非实质细胞(肝星状细胞、Kupffer细胞和内皮细胞)，并在这些细胞类型中区分HIVPI或酒精诱导的ER应激反应；(5)确定保护性分子伴侣(如4-苯基丁酸酯)在HIVPI诱导的ER应激和肝损伤中的作用。该项目将更好地了解艾滋病毒蛋白水解酶抑制剂的肝脏毒性，并提供更好的战略，以改善对艾滋病毒感染患者的护理。

DESCRIPTION (provided by applicant): HIV protease inhibitors (HIV PIs) are used in the highly ctive antiretroviral therapy (HAART). However, HIV PIs are often associated with development of liver damages. The mechanisms of the HIV protease inhibitor- induced liver injury are poorly defined. Emerging evidence indicates that the HIV PIs induce endoplasmic reticulum (ER) stress response which has been shown to play an essential role in liver dysfunction including necroinflammation, hepatic cell death and fatty liver. We previously observed that alcohol consumption, which is a well-recognized co-factor in susceptibility to the infection and progression of HIV, induced hyperhomocysteinemia and ER stress response in the liver. A significant proportion of HIV infected patients abuse alcohol and it is logical to consider that an interplay between the effects of alcohol and HIV PIs contributes to severe hepatic steatosis and injury. We hypothesize that HIV PI and alcohol exert a potentiated effect on ER stress and lipid metabolism which worsens liver injury. We propose to explore the HIV PI-induced ER stress response in both primary mouse and human hepatocytes and to investigate in vivo its interactions with pathogenic effects of alcohol in animals fed alcohol. Our specific aims are: (1) to compare ER stress and hepatic injury induced by single or combined HIV PI treatments in vivo in chronic pair- and alcohol-fed mice, and to assess additive or synergistic effects by examining HIV PI-induced ER stress response in vitro in both primary mouse and human hepatocytes and by monitoring bioavailability of HIV PIs in plasma and culture medium of hepatocytes; (2) to confirm ER stress response in mice fed orally a high methionine low folate diet in the presence or absence of single or combined HIV PIs that mimic the antiretroviral therapy in human; (3) to study synergistic mechanisms of ER stress response by HIV PIs and by alcohol through monitoring calcium releasing into the cytosol in primary mouse and human hepatocytes and through examining proteasome activities and expression and activities of glucose transporters in hepatocytes administered HIV PIs; (4) to isolate non-parenchymal cells (hepatic stellate cells, Kupffer cells, and endothelial cells) and differentiate HIV PI- or alcohol-induced ER stress response in these cell types; (5) to determine in vitro and in vivo the effectiveness of protective molecular chaperones (e.g. 4-phenylbutyrate ) in HIV PI- induced ER stress and liver injury. This project will provide a better understanding of the hepatotoxicity of HIV protease inhibitors and better strategies to improve care for HIV-infected patients.