【摘要】 背景:血管平滑肌细胞由收缩型向合成型的转变过程中线粒体可能参与了这个复杂的过程。目的:拟观察线粒体功能相关的PGC-1/NRF-1/mtTFA通路在人血管平滑肌细胞增殖中的作用。设计、时间及地点:分组设计,对比观察,于2007-01在中国医科大学附属一院中心实验室完成。材料:氧化型低密度脂蛋白由中科院协和研究所提供;叠氮钠由国药集团化学试剂有限公司沈阳分公司提供;人脐动脉平滑肌细胞由中美科技有限公司提供。方法:实验分为4组,对照组予以正常培养基;氧化型低密度脂蛋白组常规培养基中加入氧化型低密度脂蛋白使其终浓度达到10mg/L作用24h;PGC-1反义组在氧化型低密度脂蛋白组干预措施的基础上,将细胞以1×108L-1的密度接种于6孔板上,并将配置好的反义PGC-1寡核苷酸脂质体复合物70μL加入各孔,6h后补充小牛血清和培养基;叠氮钠组在氧化型低密度脂蛋白组干预措施的基础上,培养基中加入叠氮钠使其终浓度达到32mmol/L作用24h。主要观察指标:蛋白免疫印迹检测各组干预后培养细胞中PGC-1、NRF-1和mtTFA蛋白表达。观察各组干预完成后线粒体功能改变。四甲基偶氮唑盐比色法检测各组细胞活性。结果:与对照组相比,氧化型低密度脂蛋白组PGC-1、NRF、mtTFA蛋白表达、线粒体细胞色素C氧化酶酶活性及细胞增殖活性均增加(P<0.05)。与氧化型低密度脂蛋白组相比,PGC-1反义组PGC-1、NRF-1和mtTFA的蛋白表达减少、线粒体细胞色素C氧化酶酶活性及细胞增殖活性均下降(P<0.05);叠氮钠组PGC-1、NRF、mtTFA蛋白表达及细胞色素C氧化酶酶活性降低(P<0.05);细胞增殖受到抑制(P<0.05)。结论:PGC-1/NRF/mtTFA信号传导途径在人血管平滑肌细胞增生过程中通过改变线粒体功能影响细胞的生长状态。更多还原

【Abstract】 BACKGROUND:Mitochondria may participate in the complex process that vascular smooth muscle cells transform from contractile state to synthetic state. OBJECTIVE:To observe the effect of PGC-1/NRF-1/mtTFA pathway associated with mitochondrial function on the proliferation of vascular smooth muscle cells. DESIGN,TIME AND SETTING:Contrast observation was conducted in January 2007 at Central Laboratory,the First Affiliated Hospital of China Medical University. MATERIALS:Oxidized low-density lipoprotein(Xiehe Institute of Chinese Academy of Sciences,China) ;sodium azide(Shenyang Branch of Sinopharm Chemcial Reagent Co.,Ltd.,China) ;smooth muscle cells of human umbiblical artery(Wuhan Uscn Sciences Co.,Ltd.,China) . METHODS:There were 4 groups in the experiment,the control group was given normal medium;oxidized low-density lipoprotein group was treated by routine medium supplemented with 10 mg/L oxidized low-density lipoprotein for 24 hours;in antisense PGC-1 group,based on the oxidized low-density lipoprotein intervention,cells were seeded on a 6-well plate at the density of 1×108 L-1,and then added prepared 70 μL antisense PGC-1 oligonucleotide liposome complex,and added fetal bovine serum and medium 6 hours later;in sodium azide group,cells were treated by medium supplemented with 10 mg/L oxidized low-density lipoprotein and 32 mmol/L sodium azide for 24 hours. MAIN OUTCOME MEASURES:The expression of PGC-1,NRF-1 and mtTFA protein of cultured cells after intervention in each group were detected by western blotting. Changes of mitochondrial function were observed after intervention in each group. Cell viability was detected by methyl thiazolyl tetrazolium colorimetric assay. RESULTS:Compared with control group,the expression of PGC-1,NRF-1 and mtTFA protein,enzyme activity of mitochondrial cytochrome c oxidase and cell proliferation activity increased in the oxidized low-density lipoprotein group(P < 0.05) . Compared with oxidized low-density lipoprotein group,the expression of PGC-1,NRF-1 and mtTFA protein,enzyme activity of mitochondrial cytochrome c oxidase and cell proliferation activity decreased in the antisense PGC-1 group(P < 0.05) . In the sodium azide group,the expression of PGC-1,NRF-1 and mtTFA protein,enzyme activity of mitochondrial cytochrome c oxidase decreased(P < 0.05) ,cell proliferation was inhibited(P < 0.05) . CONCLUSION:PGC-1/NRF/mtTFA signal transduction pathway influences the growth status of cells through changing the mitochondrial function in the proliferation of human vascular smooth muscle cells.更多还原