鸢尾素通过磷酸腺苷酸活化蛋白激酶/哺乳动物雷帕霉素靶蛋白信号通路调控自噬抑制血管平滑肌细胞炎性反应和增殖的研究  

Inhibitory effect of irisin on vasular smooth muscle cell inflammation and proliferation by regulating autophagy through adenosine monophosphate-actived protein kinase/mammalian target of rapamycin pathway

作　　者：牟仁奎 向水 黄进启 郑勇 蔡彦力 李旭 姚元波 Mou Renkui;Xiang Shui;Huang Jinqi;Zheng Yong;Cai Yanli;Li Xu;Yao Yuanbo(Medical Department of Hubei Minzu University,Enshi 445000,China;Deparment of Cardiothoracic Surgery,Central Hospital of Enshi Tujia and Miao Prefecture,Enshi 445000,China)

机构地区：[1]湖北民族大学医学部,湖北恩施445000 [2]恩施土家族苗族自治州中心医院心胸外科,445000

出　　处：《中华实验外科杂志》2019年第6期1020-1023,共4页Chinese Journal of Experimental Surgery

基　　金：湖北省自然科学基金(2017CFB736).

摘　　要：目的探讨鸢尾素抑制血管平滑肌细胞(VSMC)增殖和炎性反应的机制。方法培养原代大鼠胸主动脉VSMC,加入鸢尾素预孵育后,再经血小板衍生生长因子BB(PDGF-BB)刺激,酶联免疫吸附试验(ELISA)检测细胞上清液炎性细胞因子白细胞介素(IL)-1β、IL-6、单核细胞趋化因子-1(MCP-1)、细胞间黏附分子-1(ICAM-1)、肿瘤坏死因子-α(TNF-α)表达水平,噻唑蓝(MTT)检测VSMC增殖情况。蛋白质印迹法(Western blot)检测磷酸腺苷酸活化蛋白激酶(AMPK)/哺乳动物雷帕霉素靶蛋白(mTOR)通路相关分子AMPK/p-AMPK(Thr172)、乙酰辅酶A羧化酶(ACC)/磷酸化(p)-ACC(Thr308)、mTOR/p-mTOR(Ser2448)、核糖体蛋白S6激酶(p70SK6)/p-70SK6(Thr389)蛋白表达及其磷酸化水平,细胞自噬相关分子微管相关蛋白1轻链3(LC3)Ⅰ、LC3Ⅱ、核孔糖蛋白P62(P62)蛋白表达水平。组间比较采用t检验。结果与对照组比较,经PFGF-BB刺激后,VSMC中AMPK(0.39±0.11比0.83±0.18,t=14.250,P<0.01)、p-AMPK(0.25±0.03比0.49±0.12,t=12.370,P<0.01)、ACC(0.54±0.16比1.07±0.24,t=10.670,P<0.01)、p-ACC(0.38±0.07比0.97±0.30,t=18.130,P<0.01)表达下调,mTOR(2.57±0.53比1.23±0.25,t=21.350,P<0.01)、p-mTOR(1.35±0.0.37比0.57±0.12,t=8.870,P<0.01)、p70SK6(1.85±0.39比0.81±0.19,t=7.170,P<0.01)、p-70SK6(0.69±0.22比0.38±0.11,t=9.260,P<0.01)表达上调,LC3Ⅰ(0.67±0.21比0.67±0.21,t=5.780,P<0.01)、LC3Ⅱ(0.42±0.13比0.93±0.38,t=6.820,P<0.01)表达下调,P62(1.24±0.38比0.58±0.14,t=5.270,P<0.01)表达上调,VSMC增殖明显增加[(146.62±11.96)%比100.00%,t=13.250,P<0.01],炎性细胞因子IL-1β(37.9±8.5比10.8±2.9,t=18.620,P<0.01)、IL-6(57.4±14.1比32.4±8.7,t=6.250,P<0.01)、MCP-1(158.3±39.1比57.2±12.5,t=14.280,P<0.01)、ICAM-1(371.8±77.3比160.4±39.8,t=10.850,P<0.01)、TNF-α(16.3±3.8比7.8±2.4,t=7.390,P<0.01)水平升高。与PDGF-BB刺激组比较,经鸢尾素预孵育后,再经PDGF-BB刺激,VSMC中AMPK(0.72±0.26,t=2.670,P<0.05)、p-AMPK(0.46±0.13,t=3.280,P<0.05)、ACC(0.95±0.28Objective To explore the underlining mechanisms of irisin inhibiting inflammation and proliferation of vascular smooth muscle cells (VSMCs). Methods The cultured primary rat aortic VSMCs were pre-incubated with irisin and then treated with platelet derived growth factor-BB (PDGF-BB). Enzyme linked immunosorbent assay (ELISA) and methyl thiazol tetrazolium (MTT) assay were used to measure the VSMCs inflammation and proliferation, respectively. The protein expression and phosphorylation of adenosine monophosphate-actived protein kinase (AMPK)/mammalian target of rapamycin (mTOR) signaling pathway related molecule AMPK/p-AMPK (Thr172), acetyl-CoA carboxylase (ACC)/phosphorylated (p)-ACC (Thr308), mTOR/p-mTOR (Ser2448), ribosomal protein S6 kinase (p70SK6)/p-70SK6 (Thr389), and autophay related molecule microtubule-associated protein 1 light chain 3 (LC3)Ⅰ, LC3Ⅱ and nucleoporin p62 (P62) were detected by Western blotting. Results After treatment with PDGF-BB, VSMCs proliferation dramatically increased, and the inflammatory cytokines IL-1β(37.9±8.5 vs. 10.8±2.9, t=18.620, P<0.01), IL-6 (57.4±14.1 vs. 32.4±8.7, t=6.250, P<0.01), MCP-1 (158.3±39.1 vs. 57.2±12.5, t=14.280, P<0.01), ICAM-1 (371.8±77.3 vs. 160.4±39.8, t=10.850, P<0.01), tumor necrosis factor-α(TNF-α)(16.3±3.8 vs. 7.8±2.4, t=7.390, P<0.01) in the cell supernatant were significantly up-regulated. The protein expression levels of AMPK (0.39±0.11 vs. 0.83±0.18, t=14.250, P<0.01), p-AMPK (0.25±0.03 vs. 0.49±0.12, t=12.370, P<0.01), ACC (0.54±0.16 vs. 1.07±0.24, t=10.670, P<0.01), p-ACC (0.38±0.07 vs. 0.97±0.30, t=18.130, P<0.01), LC3Ⅰ(0.67±0.21 vs. 0.67±0.21, t=5.780, P<0.01) and LC3Ⅱ(0.42±0.13 vs. 0.93±0.38, t=6.820, P<0.01) were down-regulated, mTOR (2.57±0.53 vs. 1.23±0.25, t=21.350, P<0.01), and p-mTOR (1.35±0.0.37 vs. 0.57±0.12, t=8.870, P<0.01), p70SK6 (1.85±0.39 vs. 0.81±0.19, t=7.170, P<0.01), p-70SK6 (0.69±0.22 vs. 0.38±0.11, t=9.260, P<0.01) and P62 (1.24±0.38 vs. 0.58±0.14, t=5.270, P<0.01) were up-regulated

关 键 词：鸢尾素 腺苷酸活化蛋白激酶/哺乳动物雷帕霉素粑蛋白信号通路 自噬 血管平滑肌细胞增殖 静脉移植物再狭窄 

分 类 号：R54[医药卫生—心血管疾病]