Mechanism of GDNF Regulation of Hepatic Steatosis

GDNF调控肝脏脂肪变性的机制

• 批准号: 10513309
• 负责人: Shanthi K Srinivasan
• 金额: --
• 依托单位: VETERANS HEALTH ADMINISTRATION
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-10-01 至 2025-09-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10513309
• 关键词: Apoptosis; Autophagocytosis; Biogenesis; CREB1 gene; Cell Death; Data; Development; Eating; Fatty Liver; Fatty acid glycerol esters; Fibrosis; Fructose; Genes; Genetic; Ginger; Glial Fibrillary Acidic Protein; Glucose; Health; Healthcare; Healthcare Systems; Hepatic; Hepatocyte; High Fat Diet; Human; In Vitro; Incidence; Knockout Mice; Lipids; Liver; Liver Fibrosis; Mediating; Metabolic syndrome; Mitochondria; Mus; Neuroglia; Obesity; Obesity Epidemic; Obesity associated liver disease; Overweight; PINK1 gene; Palmitates; Pathway interactions; Patients; Physical activity; Play; Prevalence; Prevention; Publishing; Regulation; Reporting; Role; Signal Pathway; Signal Transduction; Testing; Tissues; Transgenic Mice; Veterans; Wild Type Mouse; cell injury; chronic liver disease; cost; drinking water; experimental study; glial cell-line derived neurotrophic factor; improved; in vivo; knock-down; lipid nanoparticle; liver injury; military veteran; nanoparticle; neurotrophic factor; new therapeutic target; non-alcoholic fatty liver disease; novel; obese patients; overexpression; pharmacologic; pleiotropism; prevent; promoter; protein expression; receptor; stellate cell; western diet

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. Currently more than 25% of our veterans are obese and a larger number are overweight. In this proposal, we will examine the pleiotropic effects of Glial Derived Neurotrophic Factor (GDNF) on hepatocytes. We have demonstrated that GDNF transgenic (GDNF- Tg) mice fed a high fat diet are protected from obesity and the development of hepatic steatosis despite similar food intake and physical activity. GDNF-Tg mice overexpress GDNF under the control of the GFAP promoter expressed in glia and stellate cells. GDNF and its receptor GFRα−1 are expressed in human and murine hepatocytes. Thus, GDNF present in the liver can act locally on the hepatocytes. We recently demonstrated that GDNF can enhance autophagy and prevent liver injury. Our preliminary data demonstrate novel effects of GDNF on the liver that include: (i) Human liver tissue from patients with steatosis and fibrosis have reduced GDNF expression (ii) Western diet (WD)-fed mice have reduced level of Sirt3 and this is ameliorated in GDNF- Tg mice fed a WD; (iii) Human hepatocytes treated with GDNF have increased mitophagy. (iv) GDNF prevents apoptosis in primary human hepatocytes. The signaling pathway for these potentially beneficial effects of GDNF have yet to be explored. We hypothesize that the mechanism of GDNF prevention of hepatic injury is through promoting hepatocyte Sirt3 signaling, subsequent improved mitochondrial function leading to increased hepatocyte survival. Using both genetic and pharmacological approaches we will define the role of GDNF in inducing mitophagy to lead to improved mitochondrial function, fat reduction and reduced hepatic injury. To test the hypothesis and further investigate the underlying mechanism(s) of GDNF regulation of hepatic steatosis, we propose the following interrelated, but independently achievable aims: Specific Aim 1: To determine the mechanism of GDNF regulation of Sirt3 Preliminary data indicate that GDNF is a potent inducer of Sirt3 in vivo and in vitro. We have demonstrated that GDNF can activate the ERK signaling pathway in hepatocytes through its receptor GFRα1. We will establish if the mechanism of GDNF regulation of Sirt3 is through the GFRα1-ERK-CREB-PGC-1α pathway. WT and GDNF-Tg mice will be fed a Western diet together with fructose and glucose added in drinking water (WD/FG) for 16 weeks and ERK-CREB-PGC-1α-Sirt3 pathway assessed in hepatocytes. In vitro the necessity and sufficiency of ERK-CREB-PGC-1α in the GDNF regulation of Sirt3 expression will be determined using gene knock down and overexpression strategies. Specific Aim 2: To examine the role of mitochondrial function and mitophagy in GDNF-mediated hepatocyte survival. Our preliminary data demonstrate that GDNF increases mitochondrial function and mitophagy in hepatocytes. We will establish the role of GDNF in regulating mitochondrial function and mitophagy using the GDNF-Tg mice and in vitro experiments involving treatment of hepatocytes with GDNF. We will examine the mechanism of GDNF regulation of mitochondrial health using gene knock down and over- expression strategies. We will examine the role of GDNF-induced mitophagy in regulating hepatocyte survival. Specific Aim 3: To establish that GDNF-loaded ginger lipid nanoparticles can prevent hepatic lipid accumulation and promote hepatocyte survival. Our preliminary data demonstrate our ability to generate liver-specific natural ginger nanoparticles. We will examine the effect of GDNF-loaded ginger nanoparticles on hepatic steatosis and hepatocyte cell death and injury. GDNF-loaded ginger nanoparticles will be injected once a week into WT, Sirt3 KO mice and PINK1 KO to further establish the mechanism of action of GDNF involving Sirt3 and PINK1. Taken together our data from this proposal may provide novel targets for the treatment or prevention of hepatic steatosis and injury.

非酒精性脂肪性肝病(NAFLD)是退伍军人慢性肝病的常见原因，其 随着肥胖症的流行，患病率继续上升。目前我们25%以上的退伍军人 肥胖的人和更多的人超重。在这项提案中，我们将研究神经胶质细胞的多效性效应。 衍生神经营养因子对肝细胞的影响。我们已经证明了GDNF转基因(GDNF- 高脂肪饲料喂养的小鼠可以预防肥胖和肝脏脂肪变性的发生，尽管 食物摄入量和体力活动。GFAP启动子调控GDNF-TG小鼠过表达GDNF 表达于神经胶质细胞和星状细胞。胶质细胞源性神经营养因子及其受体α−-1在人和小鼠体内的表达 肝细胞。因此，肝脏中存在的GDNF可以局部作用于肝细胞。我们最近展示了 GDNF具有促进自噬、预防肝损伤的作用。我们的初步数据显示了新的效果 GDNF对肝脏的影响包括：(一)人肝组织脂肪变性和纤维化患者的肝组织有所减少 西方饮食(WD)喂养的小鼠降低了SIRT3水平，这在GDNF-1中得到了改善。 喂食WD的TG小鼠；(Iii)GDNF处理的人肝细胞增加了有丝分裂吞噬能力。(四)GDNF预防 原代人肝细胞的凋亡。这些潜在有益影响的信号通路 GDNF尚待探索。我们推测GDNF预防肝损伤的机制可能是 通过促进肝细胞SIRT3信号转导，随后线粒体功能的改善导致 提高肝细胞存活率。使用遗传学和药理学方法，我们将定义 GDNF诱导有丝分裂，改善线粒体功能，降脂降肝 受伤。检验假说并进一步研究神经节细胞源性神经营养因子调控的潜在机制(S) 对于肝脏脂肪变性，我们提出以下相互关联但可独立实现的目标：具体目标1： 为确定GDNF对SIRT3的调节机制，初步数据表明GDNF是一种有效的 体内外SIRT3的诱导剂。我们已经证明GDNF可以激活ERK信号通路 通过其受体GFRSIRT1在肝细胞中的表达。我们将确定α的调控机制是否 GfRCREBERKCREBPGC-1α-1α通路。WT和GDNF-TG小鼠将一起喂食西餐 在饮用水中添加果糖和葡萄糖(WD/FG)16周，ERK-CREB-PGC-1α-SIRT3 在肝细胞中评估的途径。体外研究ERK-CREB-PGC-1α在胶质细胞源性神经营养不良中的必要性和充分性 SIRT3表达的调节将通过基因敲除和过表达策略来确定。 特定目的2：研究线粒体功能和有丝分裂吞噬在GDNF介导中的作用 肝细胞存活。我们的初步数据显示，GDNF增加了线粒体功能和 肝细胞中的有丝分裂。我们将确定GDNF在调节线粒体功能和 使用GDNF-TG小鼠进行有丝分裂吞噬，以及涉及GDNF处理肝细胞的体外实验。 我们将利用基因敲除和过表达来研究GDNF对线粒体健康的调节机制。 表达策略。我们将研究GDNF诱导的有丝分裂吞噬在调节肝细胞存活中的作用。 特异性目标3：建立GDNF姜脂纳米粒预防肝脂变的实验研究 蓄积，促进肝细胞存活。我们的初步数据证明了我们有能力产生 肝脏特有的天然生姜纳米颗粒。我们将研究负载GDNF的生姜纳米颗粒对 肝脏脂肪变性和肝细胞死亡和损伤。加载GDNF的生姜纳米颗粒将注射一次 WT、SIRT3 KO小鼠和PINK1 KO小鼠一周后进一步建立GDNF参与的作用机制 SIRT3和PINK1。综上所述，我们来自这项提案的数据可能会为治疗或 预防肝脏脂肪变性和损伤。