JAZF1在脂肪肝形成与发展中的作用及相关机制研究

Nonalcoholic fatty liver disease (NAFLD) covers a spectrum from simple fatty liver (hepatic steatosis), with benign prognosis, to a potentially progressive form, nonalcoholic steatohepatitis (NASH), which may lead to liver fibrosis , cirrhosis and even liver cancer. With the gradually increasing prevalence,NAFLD is considered to be the most common liver disorder in the most developed and developing countries. However, the potential pathogenesis of the formation and progress of NAFLD remains unclear. Given the pathogenesis of NAFLD, which is generally accepted (excess triglyceride accumulation in the liver, liver inflammation, endoplasmic reticulum stress and fibrosis), eight weeks old mice including transgenic mice of JAZF1 overexpressing and C57BL/6 WT mice were fed on a high-fat diet and normal diet for 3 periods (3 months, 6 months, 12 months); The direct interactin between JAZF1 and TGIF protein was verified by Co Immunoprecipitation and double yeast hybrid experimental.The role of JAZF1 overexpression on the accumulation of triglycerides (involved in the fatty acid uptake, fatty acid synthesis, fatty acid oxidation, and very low density lipoprotein (VLDL) synthesis and secretion)was explored in mouse liver and primary cultured hepatocytes; The role of JAZF1 overexpression on the fatty acids synthesis whether through AMPK / SREBP-1 or AMPK / ACC signal and on the oxidation of fatty acids whether through PPARα signal to reduce the accumulation of triglycerides in the liver were focused on,and explored whether TGIF is necessary ;The changes of endoplasmic reticulum stress signaling (IRE1α/XBP-1 或PERK/eIF2α/ATF4/CHOP signal )were determined with the aging and prolonged high-fat diet in the liver of JAZF1 overexpression mice and primary cultured hepatocytes,and also explored whether TGIF is necessary; The role and related mechanisms of JAZF1 overexpression in the progression of NAFLD to NASH were discussed.

非酒精性脂肪性肝脏疾病是一种可以由简单的预后良好的脂肪肝发展为非酒精性脂肪性肝炎(NASH)，进一步演化为肝纤维化，肝硬化，甚至最终发展为肝癌的慢性疾病。本课题组拟对8周左右小鼠高脂和普食喂养3个时间段，利用JAZF1过表达转基因鼠，研究JAZF1过表达在小鼠肝脏及其原代培养肝细胞中涉及甘油三酯累积的脂肪酸摄取、脂肪酸合成、脂肪酸氧化以及极低密度脂蛋白（VLDL）合成和分泌过程中的作用；利用免疫共沉淀（Co-IP）及双酵母杂交试验验证JAZF1与TGIF蛋白是否有直接相互作用；重点探讨了JAZF1过表达是否通过与TGIF相互作用后，调控AMPK/SREBP-1或者AMPK/ACC信号减少脂肪酸的合成，调控 PPARα信号促进脂肪酸的氧化从而减少甘油三酯在肝脏的累积,调控内质网应激IRE1α/XBP-1 或PERK/eIF2α/ATF4/CHOP信号。

JAZF1是一个与2型糖尿病、子宫内膜间质肉瘤和前列腺癌相关联的新基因，但是关于这个基因的功能研究不多。作为TR4的辅助抑制因子，本课题组一直着力于JAZF1在糖脂代谢中的作用。而JAZF1在NAFLD的形成和发展中的作用及作用机制是未知的。本研究将WT和JAZF1-Tg鼠普食和高脂喂养了3个时间段，并利用Ad-JAZF1或shJAZF1处理由小鼠原代肝细胞和人HepG2肝细胞诱导的肝脂肪变性细胞模型，结果表明：1在NAFLD的患者及动物模型中肝脏JAZF1基因/蛋白表达下降。2、揭示衰老过程中基因的表达变化——随着年龄的增长，JAZF1基因表达会逐渐下调。JAZF1-Tg鼠肝脏中甘油三酯累积较同年龄WT鼠少， 肝细胞内涉及脂肪酸合成相关基因的mRNA和蛋白表达减少，揭示了JAZF1-Tg鼠可抵抗老龄化诱导的NAFLD的形成。3、高脂喂养3个时间段，通过对脂肪酸合成相关基因以及脂肪酸氧化相关基因的研究较好的解释了JAZF1基因在从最初的肝脂肪变性到NASH的发展过程中JAZF1改善高脂喂养诱导的NAFLD的形成与发展的机制。4、小鼠原代肝细胞和人HepG2肝细胞成功诱导肝脂肪变性细胞模型，利用Ad-JAZF1或shJAZF1处理验证细胞内脂滴的累积，脂肪酸合成基因的mRNA和相关蛋白表达、AMPK及其下游靶蛋白ACC的磷酸化。结合体内以及体外研究说明JAZF1基因过表达可激活AMPK信号，通过AMPK/SREBP-1c和AMPK/ACC途径减少肝细胞内脂质的生成，抵抗脂肪酸诱导的肝细胞内脂质的累积。5、双荧光素酶报告基因分析在胰岛素刺激下，JAZF1基因对SREBP-1c基因启动子活性的调控作用。6.阐明JAZF1抵抗高脂喂养诱导的NAFLD形成与发展的分子机制——JAZF1通过激活AMPK，抑制SREBP-1c启动子内的LXREs转录活动抑制SREBP-1c的表达，这可抑制肝脂质合成相关基因的转录。本研究第一次证明了在NAFLD的患者及动物模型肝脏JAZF1基因/蛋白表达下降。并揭示了JAZF1减少年龄和高脂诱导的肥胖及肝脂肪变性可能机制。因此,JAZF1可能是一种新的治疗衰老和饮食相关的非酒精性脂肪肝的靶点。