β3肾上腺素受体调节肝细胞核因子活性促进肝细胞载脂蛋白A-Ⅰ基因的转录

β3-AR promotes apoA-Ⅰgene transcription by regulating nuclear factor activity in hepatocytes

目的探讨β3肾上腺素受体(β3-AR)对肝细胞载脂蛋白A-Ⅰ(apoA-Ⅰ)基因转录的调控作用。方法不同浓度(10^-5~10^-10 mol/L)β3-AR激动剂(BRL37344)和拮抗剂(SR59230A)分别孵育HepG2细胞,CCK8检测细胞活性。将HepG2细胞分为对照组、激动剂组(10^-6 mol/L的BRL37344)和拮抗剂组(10^-7 mol/L的SR59230A),孵育6 h;RT-qPCR检测apoA-ⅠmRNA表达;ELISA检测细胞外基质apoA-Ⅰ分泌水平;染色质免疫共沉淀联合RT-PCR检测肝细胞核因子(HNF)-4和HNF-3及早期生长反应蛋白1(EGR-1)。结果HepG2细胞在10^-6~10^-8 mol/L的BRL37344和SR59230A区间内生长增殖良好,孵育6 h和24 h时细胞呈现较高活性。与对照组比较,激动剂组肝细胞apoA-ⅠmRNA和细胞外基质apoA-Ⅰ分泌水平上调[2.9±0.5 vs 1.0±0.2,(590.1±54.6)μg/ml vs(395.5±23.4)μg/ml,P<0.01],且HNF-4和HNF-3结合活性显著升高(90.4±13.4 vs 17.2±1.2,78.3±20.6 vs 19.7±2.5,P<0.01)。3组肝细胞EGR-1结合活性比较,无统计学差异(P>0.05)。结论激活β3-AR使肝细胞HNF-4和HNF-3活性增高,可能是β3-AR上调肝脏apoA-Ⅰ转录水平的分子机制之一。

Objective To study the role ofβ3-AR in regulating apoA-Ⅰgene transcription in hepatocytes.Methods HepG2 cells were incubated withβ3-AR agonist(BRL37344)and antagonist(SR59230A)at the concentrations of 10^-5 mol/L and 10^-10 mol/L.The HepG2 cells were divided into control group(cell culture medium),agonist group(10^-6 mol/L BRL37344),antagonist group(10^-7 mol/L SR59230A).After the HepG2 cells were incubated for 6 h,the apoA-ⅠmRNA expression was detected by RT-qPCR,the apoA-Ⅰsecretion volume was measured by ELISA,the expressions of HNF-4,HNF-3 and EGR-1 were detected by RT-PCR.Results The growth and proliferation of HepG2 cells were quite good in 10^-6 mol/L of BRL37344 and in 10^-8 mol/L of SR59230A.The activity of HepG2 cells was quite high after the HepG2 cells were incubated for 6 h and 24 h.The apoA-ⅠmRNA expression level was significantly higher(2.9±0.5 vs 1.3±0.4,P<0.01),the apoA-Ⅰsecretion volume was significantly larger in agonist group than in antagonist group(P<0.01).The binding activity of HNF-4 and HNF-3 was significantly higher in agonist group than in antagonist group(P<0.01).No significant difference was detected in binding activity of EGR-1 among the 3 groups(P>0.05).Conclusion Activation ofβ3-AR can increase the activity of HNF-3 and HNF-4 in hepatocytes,which is one of the molecular mechanisms ofβ3-AR in upregulating the apoA-Ⅰgene transcription.