geminin基因rna干扰对大鼠血管平滑肌细胞增殖的影响

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1、Geminin基因RNA干扰对大鼠血管平滑肌细胞增殖的影响张园园,李林峪,周叶萍，舒茂琴（400038重庆，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）[摘要]目的以RNA干扰抑制血管平滑肌细胞(vascularsmoothmusclecells，VSMCs)Geminin基因的表达观测VSMCs增殖情况及表型标志物。方法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。分别用RT-PCR，Westernblot检测VSMCs中Geminin基因、蛋白表达的变化及相关标志性基因的表达；应用EdU掺入试验、[3H]-TdR掺入

2、试验检测VSMCs增殖的情况。结果①siRNA转染后，3个阳性siRNA组以第1亚组干扰效果最为显著，而另外2个阳性亚组较对照组无显著差异。②siRNA转染后,阳性干扰组的[3H]-TdR、EdU掺入量均较两对照组显著升高。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（smoothmuscleactin，α-Actin）表达水平明显降低，对照组之间无显著差异。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

3、有助于VSMCs的表型由收缩型向合成型转化。[关键词]Geminin；RNA干扰；血管平滑肌细胞；表型；细胞增殖[中图法分类号][文献标志码]ATheeffectsofRNAinterferencetargetinggeminingeneontheproliferationofvascularsmoothmusclecellsinratsZhangYuanyuan,LiLinyu,ShuMaoqin(DepartmentofCardiology,SouthwestHospital,ThirdMilitaryMedicalUniversity,Chon

4、gqing,400038,China)[Abstract]ObjectiveToanalyzetheeffectsofgeminingeneontheproliferationandphenotypeofratvascularsmoothmusclecells(VSMCs)byusingsiRNAmethod．MethodsVSMCsweretransfectedwithsiRNAtoknockdownofGeminingene．ThechangeofGemininproteinwasdetectedbyWesternblottingandRT-PC

5、R．ThebestsiRNA-Gemininofthethreepositiveinterferencegroupswasusedtoconstructthemodelofslowviruspackage.Thechangesofphenotype—dependentmarkerswasanalyzedbyWesternblot．TheproliferationofVSMCswasobservedby[3H]-TdRandEdUincorporation.ResultsAftertransfectedofthethreepairsofsiRNAtar

6、getingGeminingenesrespectively,asignificantdecreaseofGemininexpressionwasobservedinoneofthreepairs，comparedwiththatoftwocontrols(non-transfectionandnegativesiRNA).[3H]-TdRandEdUincorporationhadincreasedwasobservedinpositivegroup，andahigherproliferationindexwasalsofoundinpositiv

7、egroup．Theexpressionofsyntheticmarker一OsteopontinincreasedandThelevelofVSMCscontractilemarkers一α-actindecreased.ConclusionGeminingenesilencedmayraisetheAbilityofVSMCsproliferationandmediatethetransformationofphenotypefromcontractilephenotypetosyntheticphenotype.[Keywords]Gemini

8、n；siRNA；Vascularsmoothmusclecells(VSMCs)；Phenotypes；Pr