Role of GDNF in the regulation of hepatic steatosis

描述(由申请人提供)： 非酒精性脂肪性肝病(NAFLD)是退伍军人慢性肝病的常见病因，随着肥胖症的流行，其患病率持续上升。肝脏脂肪变性是NAFLD的一个组成部分，其中甘油三酯在肝脏中积聚。最近我们发现：(I)GDNF转基因小鼠在喂食高脂肪食物时，尽管食物摄入量和体力活动相似，但仍能抵抗体重增加和肝脏脂肪变性的发生。(Ii)GDNF-TG(在胶质细胞中表达)及其受体GFR-1a在人和小鼠肝细胞中表达(Iii)GDNF-TG小鼠表现出增强的能量利用能力，(Iv)与饲喂高脂饮食的WT小鼠相比，GDNF-TG小鼠具有较低的血糖水平和改善的糖耐量试验。GDNF的这些潜在有益作用的机制还有待探索。推测GDNF预防高脂饮食所致肝脏脂肪变性的机制可能是通过改善肝脏能量消耗和抑制肝脏脂肪生成来实现的。在这项提案中，我们将确立 GDNF在调节肝脏脂肪变性中的作用充分。目前，FDA批准的治疗肝脏脂肪变性的药物很少。我们已经确定了神经营养因子GDNF的一个新的作用，它可以改善脂肪酸b的氧化，同时减少脂肪生成；这是以前没有达到的结果。我们将利用体外和体内方法研究这些过程背后的机制，涉及其受体GFRa1的过度表达或下调，并剖析涉及的信号通路。在特定的目标1中，我们将确定GDNF在促进肝细胞中的脂肪酸β氧化中的作用。初步研究表明，GDNF可以激活肝细胞中的PGC-1a，导致与脂肪酸β氧化和线粒体生物发生相关的酶的上调。我们将利用体内和体外高表达GDNF的小鼠(GDNF-TG小鼠)来确定GDNF在调节肝脏脂肪酸氧化中的必要性和充分性。在具体目标2中，我们将阐述GDNF抑制肝脏脂肪生成的分子基础，重点是GDNF下调PPAR-g及其下游成脂基因FASN和CD36的作用。我们的初步结果表明，GDNF导致PPAR-g的表达减少。在具体目标3中，我们将把肝细胞特异性GDNF纳米粒输送到肝脏，用于靶向治疗HF饮食喂养的小鼠的肝脏脂肪变性，并评估GDNF预防或治疗HF饮食诱导的小鼠脂肪变性的疗效。对退伍军人医疗保健的影响：目前，超过25%的退伍军人患有肥胖症，更多的退伍军人超重。2008年，肥胖及其相关代谢综合征引起的并发症花费了美国医疗保健系统1470亿美元(CDC MMWR报告，2010年8月)。预防和治疗肥胖的策略可以减少与肥胖相关的并发症(如NAFLD)和医疗成本。确定治疗肝脏脂肪变性的新疗法将直接使退伍军人受益。综上所述，我们的数据可能会为治疗或预防肝脏脂肪变性提供新的靶点。

DESCRIPTION (provided by applicant): Non-alcoholic fatty liver disease (NAFLD) is a common cause of chronic liver disease in Veterans and its prevalence continues to increase, with the growing obesity epidemic. Hepatic steatosis is a component of NAFLD, wherein there is accumulation of triglycerides in the liver. Recently we discovered that (i) GDNF transgenic mice when fed a high fat diet resist weight gain and development of hepatic steatosis, despite similar food intake and physical activity. GDNF transgenic mice over express GDNF in glia under the GFAP promoter (GDNF-tg) (ii) GDNF (expressed in glia) and its receptor GFR-1a are expressed in human and murine hepatocytes (iii) GDNF-tg mice demonstrate enhanced energy utilization assessed by indirect calorimetry, (iv) compared to WT mice fed a high fat diet the GDNF-tg mice have lower serum glucose levels and improved glucose tolerance tests. The mechanisms for these potentially beneficial effects of GDNF have yet to be explored. We hypothesize that the mechanism of GDNF prevention of high fat diet induced hepatic steatosis is through improved hepatic energy expenditure and inhibition of hepatic lipogenesis. In this proposal we will establish the necessity and sufficiency of GDNF in regulating hepatic steatosis. Currently there are few FDA approved drugs for the treatment of hepatic steatosis. We have identified a novel role for the neurotrophic factor, GDNF, that improves fatty acid b-oxidation with a combined reduction in lipogenesis; an outcome that has not been previously achieved. We will examine the mechanism underlying these processes utilizing in vitro and in vivo approaches involving over expression or knock down of its receptor GFRa1 and dissect the signaling pathways involved. In Specific Aim 1 we will determine the role of GDNF in enhancing fatty acid beta oxidation in hepatocytes. Preliminary data suggests that GDNF can activate Peroxisome proliferator- activated receptor coactivator -1a (PGC-1a) in hepatocytes, resulting in up regulation of enzymes related to fatty acids beta-oxidation and mitochondrial biogenesis. We will establish the necessity and sufficiency of GDNF in regulating hepatic fatty acid oxidation using mice over expressing GDNF (GDNF-tg mice) in vivo and in hepatocytes in vitro. In Specific Aim 2 we will demonstrate the molecular basis of GDNF inhibition of hepatic lipogenesis, focusing on GDNF downregulation of PPAR-g and its downstream lipogenic genes, FASN and CD36. Our preliminary results indicate GDNF results in a reduced expression of PPAR-g. In Specific Aim 3 we will deliver hepatocytes-specific GDNF-encapsulated nanoparticles to the liver for targeted therapy of hepatic steatosis to HF-diet fed mice and assess the therapeutic efficacy of GDNF to prevent or treat HF-diet induced steatosis in mice. Impact on Veterans Health Care: Currently more than 25% of our Veterans are obese and a larger number are overweight. In 2008, complications arising from obesity and its related metabolic syndrome cost the US health care system $147 billion (CDC MMWR report, August 2010). Strategies to prevent and treat obesity can reduce obesity related complications (like NAFLD) and health care costs. Identification of new therapies for hepatic steatosis will directly benefit the veteran population. Taken together our data from this proposal may provide novel targets for the treatment or prevention of hepatic steatosis.