LDL Receptor Related Protein-1 in Metabolic and Cardiovascular Disease

描述（由申请人提供）：几项大规模人群研究已经确定了LDL受体相关蛋白-1 （LRP1）基因多态性与广泛的代谢性疾病之间的关联，包括动脉粥样硬化、肥胖/糖尿病、非酒精性脂肪性肝病（NAFLD）和肝细胞癌（HCC）预后不良。后一项观察结果表明，肝脏LRP1功能障碍可能加速NAFLD向脂肪性肝炎和肝硬化的进展，并增加HCC的风险。该项目过去资助期的研究表明，肝细胞特异性Lrp1基因失活或突变会改变小鼠Lrp1内吞作用和细胞表面再循环，从而增强脂肪诱导的脂肪变性和细胞死亡。由于LRP1基因缺失和影响LRP1细胞表面循环的突变在人类中尚未报道，我们开始研究肝细胞表面LRP1表达是否以及如何受到生理调节。我们的初步结果表明，脂肪酸和高脂肪饮食抑制了LRP1向肝细胞表面的易位，而胰岛素具有相反的作用，刺激了LRP1细胞表面的易位。这项更新应用的目的是破译LRP1循环和转运到细胞表面的调控机制。我们将验证肝细胞中LRP1细胞表面表达的减少直接导致脂肪诱导的肝脏炎症和肝硬化，并直接导致与高脂肪/胆固醇饮食和脂肪肝疾病相关的HCC加速进展的假设。此外，我们提出增加LRP1在细胞表面的再循环是缓解这些代谢异常的可行策略。特异性目的1将确定脂质抑制LRP1转运至质膜的机制，验证脂质积累改变LRP1胞质域磷酸化的假设，从而抑制其与转运至细胞表面所需的衔接蛋白的相互作用。我们还将评估两种增加LRP1循环的策略，并测试它们在改善肝细胞功能和预防脂质性肝功能障碍和肝硬化方面的有效性。特异性目的2将描述胰岛素激活LRP1易位到细胞表面的机制，验证胰岛素诱导脂激酶活性产生磷脂酰肌醇（聚磷酸）以增加LRP1接子复合物的募集到循环内体以易位到细胞表面的假设。后续实验将探索渗透性肌肽多磷酸盐作为克服胰岛素抵抗以改善肝脏代谢健康的策略的潜力。特异性Aim 3将验证肝细胞LRP1功能障碍促进脂质性肝硬化的假设，从而为加速肝肿瘤进展提供理想的局部环境。这些研究的完成将确定调节LRP1易位到细胞表面的机制，并提供抑制NAFLD进展为NASH、肝硬化和HCC的新策略。

DESCRIPTION (provided by applicant): Several large-scale population studies have identified the association between polymorphisms of the LDL receptor-related protein-1 (LRP1) gene with a wide spectrum of metabolic diseases, including atherosclerosis, obesity/diabetes, nonalcoholic fatty liver diseases (NAFLD), and poor prognosis of hepatocellular carcinoma (HCC). The latter observations imply that LRP1 dysfunction in the liver may accelerate the progression of NAFLD to steatohepatitis and cirrhosis with increased HCC risk. Studies from the past funding period of this project have shown that hepatocyte-specific Lrp1 gene inactivation or mutations that alter LRP1 endocytosis and cell surface recycling in mice potentiates fat-induced steatosis and cell death. Since total LRP1 gene deletion and mutations that affect LRP1 cell surface recycling have not been reported previously in humans, we initiated studies to examine if and how LRP1 expression at the cell surface of hepatocytes may be regulated physiologically. Our preliminary results showed that fatty acid and high fat diet suppress LRP1 translocation to the cell surface of hepatocytes, whereas insulin has the opposite effect and stimulates LRP1 cell surface translocation. The objective of this renewal application is to decipher the mechanisms underlying the regulation of LRP1 recycling and translocation to the cell surface. We will test the hypothesis that reduction of cell surface expression of LRP1 in hepatocytes directly contributes to fat-induced liver inflammation and hepatic cirrhosis, and is directly responsible for the accelerated HCC progression associated with high fat/cholesterol diet and fatty liver disease. Moreover, we propose that increasing LRP1 recycling to the cell surface is a viable strategy to alleviate these metabolic abnormalities. Specific Aim 1 will identify the mechanism underlying lipid suppression of LRP1 translocation to plasma membrane, testing the hypothesis that lipid accumulation alters phosphorylation of the cytoplasmic domain of LRP1, thereby suppressing its interaction with adaptor proteins required for its translocation to the cel surface. We will also evaluate two strategies to increase LRP1 recycling and test their effectiveness to improve hepatocyte functions and prevent lipid-induced liver dysfunction and cirrhosis. Specific Aim 2 will delineate the mechanism underlying insulin activation of LRP1 translocation to the cell surface, testing the hypothesis that insulin induces lipid kinase activit to generate phosphatidylinositol (poly)phosphates to increase the recruitment of LRP1-adaptor complex to the recycling endosomes for translocation to the cell surface. Follow up experiments will explore the potential of permeable inositide polyphosphates as a strategy to overcome insulin resistance to improve metabolic health of the liver. Specific Aim 3 will test the hypothesi that LRP1 dysfunction in hepatocytes promotes lipid-induced liver cirrhosis, thereby providing an ideal local environment to accelerate liver tumor progression. The completion of these studies will identify mechanisms regulating LRP1 translocation to the cell surface, and offers novel strategies to suppress NAFLD progression to NASH, liver cirrhosis, and HCC.