以斑馬魚為動物模式探討過氧化酶體活化受體γ因硫乙胺誘發脂肪肝所扮演的角色

Abstract

中文摘要: 過氧化酶體增生活化受體γ(PPARγ)被認為是脂肪細胞的一個主要分化調節因子。 在最近的報告中發現，在瘦體素缺乏所導致脂肪肝老鼠模式中（Matsusue et al.,2003），肝臟專一性剔除過氧化酶體增生活化因子γ將改善老鼠脂肪肝的病灶。PPARγ2大量表現在成熟的脂肪細胞並且在脂肪肝動物模式中也有升高的現象。PPARγ調節脂肪生成和堆積在肝細胞 。本篇研究的目的在於肝臟專一性表現PPARγ是否有能力使肝臟脂肪化，以及在脂肪肝形成中扮演什麼樣的角色。為了了解PPARγ在正常及脂肪肝所扮演的角色，首先，選殖出斑馬魚的兩種長度PPARγcDNA分別為1533及1581鹼基對，接下來利用顯微注射的方式，使用肝臟專一性啟動子表現PPARγ在斑馬魚卵。反轉錄聚合鏈反應結果顯示轉殖PPARγ大量表現在肝臟，提高脂肪細胞相關基因，例如:脂肪細胞脂肪酸結合蛋白（aP2）、脂締素（adiponectin）以及脂肪生成相關基因，例如:固醇調控序列結合蛋白一型（SREBP-1c）。第二，利用硫乙胺誘發顯微注射PPARγ5天後之魚苗脂肪肝化。由我們實驗室發表的報告中，指出硫乙胺處理斑馬魚稚魚七天，將造成脂肪肝炎。反轉錄聚合鏈反應結果顯示轉殖PPARγ經TAA處理十天後，在第三有提高的現象，但是從硫乙胺處理控制組的結果顯示第9天時，脂肪生成相關基因才會上升，故PPARγ的表現加強了硫乙胺誘導脂肪肝的形成過程。

Abstract： Peroxisome proliferator-activated receptor gamma (PPARγ) is considered to be one of the master regulators of adipocyte differentiation. In recent report, liver-specific disruption of PPARgamma improves fatty liver in leptin-dificient mice（Matsusue et al.,2003）. PPARγ2 is abundantly expressed in mature adipocytes and is elevated in the liver of animals that develop fatty liver. PPARγ regulates lipogenesis and lipid accumulation in steatotic hepatocyte. The aim of this study was to study the role of PPARγin pathogenesis of fatty liver. And to examine over-expression of PPARγ in liver by liver-specific L-FABP promoter and ubiquitous expression by EF-1αpromoter could induce lipogenesis and result in steatosis ? To understand roles of PPARγ in normal and fatty liver in zebrafish, I first cloned two cDNAs with length of 1533 bp and 1581 bp encoding PPARγ1 and PPARγ2, respectively. Liver-specific over-expression of PPARγ in zebrafish larva resulted in up-regulation of adipogenetic genes such as, adipocyte Fatty Acid Binding Protein (AFABP/AP2), adiponectin, and lipogenetic gene such as, SREBP-1. Secondly, I tried to promote hepatosteatosis in PPARγ-injected 5 day post fertilization (dpf) zebrafish larva by hepatotoxin thioacetamid (TAA) treatment. We have previously showed that TAA treatment for 7 days leads to liver steatohepatitis of zebrafish larva at 10 dpf. After injected with pLF2.8-zfPPARγ, zebrafish larva at 5 dpf were treated with TAA for 10 days. The results of RT-PCR showed that zebrafish treated with TAA had adipogenetic and lipogenetic genes up-regulated on the third and 5th day, After injected pLF2.8-zfPPARγ 5-day postfertilization, treaded TAA 10 days. The results of RT-PCR show that zebrafish treaded TAA 10th day made adipogenetic and lipogenetic up-regulated on the 3th and 5th day, because on TAA treated control Data show that lipogenetic gene would up-regulate at 9th day, so overexpression of PPARγ enhance TAA induced fatty liver.