EphA2 in Non-Alcoholic Fatty Liver Disease

项目摘要 非酒精性脂肪性肝病（NAFLD）的特征在于由增强的游离脂肪肝引起的肝脂肪变性。 酸（FFA）摄取和脂质合成，通过线粒体脂肪酸氧化减少脂质清除， 脂蛋白输出。过量的肝脏脂质积聚可引起脂毒性，其促进局部 炎症、肝细胞气球样变和纤维化。这一过程导致了非酒精性的发展 脂肪性肝炎（NASH），如果留下，可能进一步进展为肝硬化、肝衰竭或肝细胞癌 未经检查。目前NAFLD/NASH的治疗寻求减少肝脂肪变性或减少纤维化 推动NASH发展的反应。虽然这些治疗在临床前试验中显示出很大的希望， 它们还没有提供明确的治疗益处。 受体酪氨酸激酶Eph家族是哺乳动物基因组中最大的家族， 受体相互作用（肝配蛋白）配体在相邻细胞介导细胞粘附/排斥信号在轴突 引导、组织图案化、白细胞归巢和转移。不同的配体依赖性和配体- 独立的磷酸化事件似乎介导EphA 2信号传导的这些通常相反的功能。EphA2 与邻近细胞上ephrinA 1的相互作用促进酪氨酸自磷酸化（Y 588，Y 772）和内源性 激酶活性，导致增殖和迁移的抑制。在没有配体的情况下，EphA 2可以 在S897上被磷酸化，典型地被Akt磷酸化，导致增殖和迁移增强， 大部分未知的机制。我们实验室的工作证明了EphA 2在多种细胞类型中表达， EphA 2缺失减少动脉粥样硬化斑块形成。我们的初步数据显示EphA 2基因敲除小鼠 显示在高脂肪饮食喂养后肝脏重量/体重比显著降低， 减少脂肪肝和减少炎症。此外，用高脂肪饮食处理的C57 B1/6 J小鼠 显示EphA 2配体ephrinA 1的表达增强，沿着EphA 2标记物的增强 配体依赖性信号传导。相比之下，NASH患者和NASH的小鼠模型均显示出降低的免疫应答。 ephrinA 1表达和增强的EphA 2配体非依赖性信号传导的标志物。所以我们 假设配体依赖性EphA 2信号传导作为早期EphA 2表达的初始保护机制， 随着疾病进展而减弱的NAFLD。使用细胞培养和小鼠模型的组合， 我们将确定EphA 2/EphrinA 1表达和信号传导在脂质稳态和代谢中的作用， (Aim 1），并确定EphA 2/EphrinA 1表达和信号传导在NAFLD/NASH进展中的作用（Aim 2）。如果成功，这些研究将阐明EphA 2信号传导在NAFLD/NASH发病机制中的新作用， 可能提供一种新的治疗靶点，以减少NAFLD/NASH的肝损伤。

PROJECT SUMMARY Non-alcoholic fatty liver disease (NAFLD) is characterized by hepatic steatosis driven by enhanced free fatty acid (FFA) uptake and lipid synthesis with reduced lipid clearance through mitochondrial fatty acid oxidation and lipoprotein export. Excessive hepatic lipid accumulation can cause lipotoxicity, which promotes local inflammation, hepatocyte ballooning, and fibrosis. This process leads to the development of nonalcoholic steatohepatitis (NASH), that can further progress to cirrhosis, liver failure, or hepatocellular carcinoma if left unchecked. Current treatments for NAFLD/NASH seek to either reduce hepatic steatosis or diminish the fibrosis response that drives NASH development. While these treatments have shown great promise in pre-clinical trials, they have yet to provide a clear therapeutic benefit. The Eph family of receptor tyrosine kinases, the largest family in the mammalian genome, interact with Eph receptor-interacting (ephrin) ligands on adjacent cells to mediate cell adhesion/repulsion signaling during axonal guidance, tissue patterning, leukocyte homing, and metastasis. Differential ligand-dependent and ligand- independent phosphorylation events appear to mediate these often-inverse functions of EphA2 signaling. EphA2 interactions with ephrinA1 on an adjacent cell promote tyrosine autophosphorylation (Y588, Y772) and intrinsic kinase activity, resulting in the inhibition of proliferation and migration. In the absence of ligand, EphA2 can become phosphorylated on S897, classically by Akt, resulting in enhanced proliferation and migration through largely unknown mechanisms. Work from our lab demonstrated EphA2 expression in multiple cell types and EphA2 deletion reduces atherosclerotic plaque formation. Our preliminary data show that EphA2 knockout mice show significantly reduced liver weight/body weight ratios following high fat diet feeding associated with diminished hepatosteatosis and reduced inflammation. Furthermore, C57Bl/6J mice treated with a high fat diet for 8 weeks show enhanced expression of the EphA2 ligand ephrinA1, along with enhanced markers of EphA2 ligand-dependent signaling. In contrast, both NASH patients and mouse models of NASH show decreased ephrinA1 expression and enhanced markers of EphA2 ligand-independent signaling. Therefore, we hypothesize that ligand-dependent EphA2 signaling serves as an initial protective mechanism in early NAFLD that weakens as the disease progresses. Using a combination of cell culture and mouse models, we will determine the role of EphA2/EphrinA1 expression and signaling in lipid homeostasis and metabolism (Aim 1), and determine the role of EphA2/EphrinA1 expression and signaling in NAFLD/NASH progression (Aim 2). If successful, these studies will elucidate a novel role for EphA2 signaling in NAFLD/NASH pathogenesis and may provide a novel therapeutic target to reduce liver damage in NAFLD/NASH.