Hepatitis C virus infection down-regulates the expression of peroxisome proliferator-activated receptor α and carnitine palmitoyl acyl-CoA transferase 1A
Hepatitis C virus infection down-regulates the expression of peroxisome proliferator-activated receptor α and carnitine palmitoyl acyl-CoA transferase 1A
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DOI：
10.3748/wjg.v11.i48.7591
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发表时间：
2005-12-28
影响因子：
4.3
通讯作者：
Desreumaux, Pierre
中科院分区：
文献类型：
--
作者：
Cheng, Yang;Dharancy, Sebastien;Desreumaux, Pierre
目的：阐明过氧化物酶体增殖物激活受体α（PPARα）及其靶基因肉毒碱棕榈酰辅酶A转移酶1A（CPT1A）在丙型肝炎病毒（HCV）感染发病机制中的作用。方法：采集慢性HCV感染者和对照者的肝脏样本。用携带pEF352neo的载体转染HepG2细胞。分析了稳定表达HCV核心蛋白的两个独立克隆（克隆N3和N4）。从细胞和肝组织中提取总RNA。使用 SYBR Green Master 通过实时聚合酶链反应 (PCR) 对 PPAR α 和 CPT1A mRNA 进行定量。通过聚丙烯酰胺凝胶电泳分离总提取蛋白质并进行电印迹。将膜与抗 PPAR α 抗体一起孵育，然后与与 PPAR α 辣根过氧化物酶缀合的猪抗兔 IgG 一起孵育。通过 PPAR α 的增强化学发光反应显示蛋白质条带。对于 PPAR α 的免疫组织化学染色，切片与针对 PPAR α 的山羊一抗多克隆抗体在室温下孵育。 结果：实时 PCR 表明，丙型肝炎患者的 PPAR α 水平和 CPT1A 基因表达水平较对照组显着降低（1.8 +/- 2.8 vs 13 +/- 3.4，P = 0.0002；1.1 +/- 1.5 vs 对照组）。 7.4 +/- 1，P = 0.004）。 Western blot结果显示，丙型肝炎患者肝脏中PPARα蛋白水平低于对照组（2.3±0.3 vs 3.6±0.2，P=0.009）。慢性丙型肝炎患者的免疫组织化学染色结果表明，与对照肝脏相比，肝细胞中 PPAR α 染色减少。体外研究发现，在稳定表达HCV核心蛋白的N3和N4结肠中，PPARαmRNA水平显着低于对照。结论：肝内PPARα表达受损与慢性HCV感染时肝损伤的致病机制有关。 HCV感染不仅在mRNA水平而且在蛋白质水平上降低PPARα和CPT1A的表达。 PPARα在慢性HCV感染的发病机制中发挥着重要作用，但该核受体在HCV感染中的功能受损尚需进一步研究。 (C) 2005 The WJG Press 和 Elsevier Inc. 保留所有权利。
AIM: To elucidate the role of the peroxisome proliferator-activated receptor alpha (PPAR alpha) and its target gene carnitine palmitoyl acyl-CoA transferase 1A (CPT1A) in the pathogenesis of hepatitis C virus (HCV) infection.METHODS: Liver samples were collected from the patients with chronic HCV infection and controls. HepG2 cells were transfected with vector pEF352neo carrying. Two independent clones (clone N3 and N4) stably expressing HCV core protein were analyzed. Total RNA was extracted from cells and liver tissues. PPAR alpha and CPT1A mRNAs were quantified by real-time polymerase chain reaction (PCR) using SYBR Green Master. Total extracted proteins were separated by polyacrylamide gel electrophoresis, and electroblotted. Membranes were incubated with the anti-PPAR alpha antibody, then with a swine anti-rabbit IgG conjugated to horseradish peroxidase for PPAR alpha. Protein bands were revealed by an enhanced chemiluminescence reaction for PPAR alpha. For immunohistochemical staining of PPAR alpha, sections were incubated with the primary goat polyclonal antibody directed against PPAR alpha at room temperature.RESULTS: Real-time PCR indicated that the PPAR alpha level and expression level of CPT1A gene in hepatitis C patients lowered significantly as compared with the controls (1.8 +/- 2.8 vs 13 +/- 3.4, P = 0.0002; 1.1 +/- 1.5 vs 7.4 +/- 1, P = 0.004). Western blot results showed that the level of PPAR alpha protein in the livers of hepatitis C patients was lower than that in controls (2.3 +/- 0.3 vs 3.6 +/- 0.2, P = 0.009). The immunohistochemical staining results in chronic hepatitis C patients indicated a decrease in PPAR alpha staining in hepatocytes compared with those in the control livers. The in vitro studies found that in the N3 and N4 colon stably expressing HCV core protein, the PPAR alpha mRNA levels were significantly lower than that in the controls.CONCLUSION: The impaired intrahepatic PPAR alpha expression is associated with the pathogenic mechanism in hepatic injury during chronic HCV infection. HCV infection reduced the expression of PPAR alpha and CPT1A at the level of not only mRNAs but also proteins. PPAR alpha plays an important role in the pathogenesis of chronic HCV infection, but the impaired function of this nuclear receptor in HCV infection needs further studies. (C) 2005 The WJG Press and Elsevier Inc. All rights reserved.