干扰肝细胞核因子-1α对HepG2细胞载脂蛋白M及胆固醇相关代谢酶表达的影响  

Effect of interfering hepatocyte nuclear factor-1 alfa in HepG2 on the expressions of apoM, apoA-I and the correlative key enzyme of cholesterol metabolism

作　　者：章尧[1] 陈昌杰[2] 杨清玲[2] 程龙强[3] 王惠[3] 黄丽珠[3] 

机构地区：[1]皖南医学院生物化学与分子生物学教研室,安徽省芜湖市241000 [2]蚌埠医学院生物化学与分子生物学教研室 [3]蚌埠医学院临床检验诊断实验中心

出　　处：《中华肝脏病杂志》2011年第2期121-126,共6页Chinese Journal of Hepatology

基　　金：基金项目：安徽省自然科学基金（090413111）

摘　　要：目的检测肝细胞癌和癌旁组织中肝细胞核因子（HNF）1α、肝受体同系物（LRH）-1与载脂蛋白（apo）M表达的相关陛，探讨HNF-1α对apoM、apoAI和胆固醇合成与转化关键酶表达的影响。方法RT—PCR检测人肝细胞癌组织和癌旁组织中HNF-1α、LRH-1apoM的表达水平。采用小干扰RNA（siRNA）在人肝癌细胞株HepG2细胞中干扰HNF1d表达后，RTPCR检测apoM、apoA—I及羟甲基戊二酰辅酶A还原酶（HMGCR）、胆固醇7α-羟化酶1（CYP7A1）、法尼醇X受体（FXR）、小异源二聚体伴侣（SHP）-1的变化情况；Westernblot检测apoM在蛋白水平的变化情况。对不同数据进行单因素方差分析、配对t检验或直线相关分析。结果HNF-1α、LRH1、apoMmRNA在人肝细胞癌组织中明显高于癌旁组织（t值分别为-7．075，-8．803，-7．167，P值均〈0．01）；HNF-1α和LRH-1与apoM的mRNA表达水平呈正相关（r=0．353，P〈0．01；r=0．523，尸〈0．01）。干扰HNF-1α后，与阴性对照相比，apoM、FXR、SHP1mRNA水平分别下降47．4％（F=77．627，P〈0．01）、47．9％（F=76．363，P〈0．01）、65．2％（F=365．772，P〈0．01），apoM蛋白水平下调54．3％（F=43．482，P〈0．01）；HMGCR、CYP7A1mRNA水平分别升高101．1％（F=218．666，P〈0．01）、138．5％（F=171．926，P〈0．01）；而对apoAI无明显影响（，==0．170，P〉0．05）。结论HNF-1α可提高apoM的表达水平，并通过促进胆固醇代谢关键酶的表达和抑制胆固醇代谢相关抑制因子的生成而促进胆固醇的转化，对心血管疾病具有保护作用。Objective To determine wether there were connections among hepatocyte nuclear factor- I alfa （HNF-1α ）, liver receptor homolog-1 （LRH-1）, apolipoprotein M （apoM） and to investigate the effects of HNF-1α in HepG2 on the expressions of apoM, apolipoprotein A-I （apoA-I） and the key enzymes in cholesterol metabolism and biotransformation. Methods The mRNA expressions of apoM, LRH-1 and HNF-1α were detected by RT-PCR. HNF-1α was interfered and RT-PCR was used to detect the changes of apo M, apo A-I, Cyp7A1, farnesoid X receptor （FXR） and small heterodimer partner-I （SHP-1）. Western blot was used to detect the change of apo M protein. Results The expressions of apoM, LRH-1 and HNF-1α mRNA were obviously higher in HCC tissue than that in para-cancer tissue （the vaule of t is -7.167, -7.075, -8.803, P 〈 0.01 respectively）. HNF-1αand LRH-1 positively correlated with the expression of apoM （r = 0.353, P 〈 0.01; r = 0.523, P 〈 0.01 respectively）; RT-PCR and western blot results showed that the expressions of apoM, FXR and SHP-1 mRNA, could be obviously suppressed by HNF-1 ct interfering as compared to the negative controls by 47.4%, 47.9%and 65.2% （P 〈 0.01） respectively, and the expression of apoM protein also decreased by 54.3% （F = 43.482, P 〈 0.01）. The expressions of HMGCR and CYP7A mRNA increased by 101.1% and 138.5% （P 〈 0.01） respectively as compared to the negative control. But there is no effect on expression of apoA-I mRNA （F = 0.170, P 〉 0.05）. Conclusions HNF-1 0t could promote cholesterol biotransformation by increasing the expression of apoM and the key enzymes in cholesterol metabolism and decreasing inhibiting factor. So HNF-1α provided protection against cardiovascular disease.

关 键 词：载脂蛋白类 肝细胞核因子1α RNA干扰 

分 类 号：R735.7[医药卫生—肿瘤]