Geminin基因RNA干扰对大鼠血管平滑肌细胞增殖的影响  

Effect of RNA interference targeting geminin on proliferation of vascular smooth muscle cells in rats

作　　者：张园园[1] 李林峪[1] 周叶萍[1] 舒茂琴[1] 

机构地区：[1]第三军医大学西南医院心血管内科重庆市介入心脏病学研究所,重庆400038

出　　处：《第三军医大学学报》2012年第3期210-213,共4页Journal of Third Military Medical University

基　　金：国家自然科学基金(30971228)~~

摘　　要：目的以RNA干扰抑制血管平滑肌细胞(vascular smooth muscle cells,VSMCs)Geminin基因的表达观测VSMCs增殖情况及表型标志物。方法分别利用RT-PCR,Western blot检测VSMCs中Geminin基因、蛋白表达的变化,筛选出干扰效果最为显著的序列用于构建慢病毒包裹的稳定干扰模型。应用[3H]-TdR、EdU掺入试验检测VSMCs增殖的情况;同时使用Western blot分别检测干扰前后相关标志性基因的表达。结果①siRNA转染后,以第1亚组干扰效果最为显著,与另外2个阳性亚组及对照组比较差异具有统计学意义(P<0.05)。②[3H]-TdR、EdU掺入实验显示:阳性干扰组的掺入量较两对照组明显升高(P<0.05)。③siRNA转染使Geminin基因表达减弱后,阳性组VSMCs合成型标志物骨桥蛋白(osteopontin,OPN)表达水平显著升高,收缩型标志物平滑肌肌动蛋白(smooth muscle actin,α-Actin)表达水平明显降低,与对照组间比较,差异均有统计学意义(P<0.05)。结论 RNA干扰介导的Geminin基因沉寂可显著增强VSMCs的增殖能力,并有助于VSMCs的表型由收缩型向合成型转化。Objective To investigate the proliferation and phenotypic muscle cells （VSMCs） by using RNA interference targeting geminin. Methods markers of rat vascular smooth VSMCs were transfected with siRNA for knockdown of geminin. Changes of geminin at protein and mRNA levels were detected by Western blotting and RT-PCR, and siRNA-geminin that exhibited the most significant inhibitory effect was used to con- struct lentivirus-packaged stable interference model. The proliferation of VSMCs was observed by [ 3H ]-TdR and EdU incorporation. The changes of phenotype-dependent markers were analyzed by Western blotting. Results After VSMCs transfected with three pairs of siRNA targeting geminin, a significant decrease of geminin expression was observed in one pair, as compared with that of the other two pairs and the controls （P 〈 0.05 ）. [ 3 H ] -TdR and EdU incorporation showed a higher proliferation index in the positive interference group than that in the control groups （ P 〈 0.05 ）. In the positive interference group, level of synthetic marker osteopontin （OPN） increased and level of contractile marker smooth muscle actin （cL-Actin） decreased, and the differences were significant as compared with those of the control groups （P 〈 0.05）. Conclusion Geminin silencing by siRNA can promote VSMCs proliferation and phenotype transformation from contractile phenotype to synthetic phenotvpe.

关 键 词：GEMININ RNA干扰 血管平滑肌细胞 表型 细胞增殖 

分 类 号：R322.12[医药卫生—人体解剖和组织胚胎学] R329.28[医药卫生—基础医学]