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1、siRNA干扰骨形态发生蛋白2对肝癌SMMC7721细胞迁移和侵袭能力的抑制作用吴建兵,傅华群,黄龙璋,刘安文,张吉翔(南昌大学第二附属医院肿瘤科,江西省重点医学分子实验室,南昌330006)摘要:目的设计并化学合成针对骨形态发生蛋白2(bonemorphogeneticproteins,BMP-2)的siRNA分子片段,转染人肝癌SMMC7721细胞,观察其对SMMC7721细胞迁移和侵袭能力的影响。方法设计并化学合成针对BMP-2的3个siRNA分子序列,阳离子脂质体法瞬间转染SMMC7721细胞,运用半定量逆转录聚合酶链反应(
2、RT-PCR)和Western印迹法检测细胞中BMP-2在mRNA水平和蛋白水平的变化,体外侵袭实验检测转染后细胞侵袭能力的变化。结果RT-PCR和Westernblot显示3对特异性BMP-2-siRNA转染肝癌细胞SMMC7721之后,其BMP-2的基因和蛋白表达均受到抑制,以siRNA-B抑制效果最明显[(0.32±0.07)vs(0.92±0.14),(0.37±0.10)vs(0.89±0.17),P<0.01]。体外侵袭实验显示转染后肝癌细胞数明显低于未转染组和阴性对照组[分别为(6.48±3.62)、(23.72±3.
3、24)、(22.38±3.84),P<0.05]。结论BMP-2与肝癌的侵袭转移潜能相关,siRNA-BMP-2能明显抑制肝癌细胞SMMC7721的BMP-2表达,从而抑制肝癌细胞的迁移和侵袭能力。关键词:肝肿瘤;骨形态发生蛋白;RNA干扰;迁移;侵袭中图法分类号:文献标识码:AInhibitoryeffectofsiRNA-BMP-2onmigrationandinvasionofhumanhepatomacelllineSMMC7721WUJian-bing,FUHua-qun,HUANGLong-zhang,LIUAn-wen
4、,ZHANGJi-xiang(DepartmentofOncology,SecondAffiliatedHospitalofNanchangUniversity,Nanchang330006,China)Abstract:ObjectiveTodesignandpreparesiRNAstargetingbonemorphogeneticprotein2(BMP-2)geneandtoobserveitsinfluenceonmigrationandinvasionofSMMC7721cells.MethodsThreespecif
5、icsiRNAsofBMP-2weredesignedandsynthesized,andweretransientlytransfectedintoSMMC7721viacathodolyteliposometransfectionmethod.ReversetranscriptasePCRandWesternblottingwereusedtomeasureBMP-2expressionatmRNAandproteinlevels.Themigrationandinvasionabilityofcancercellwereass
6、essedbyinvasionassayinvitro.ResultsAfterAllthe3specificBMP-2-siRNA(A,B,C)transfectingintohumanhepatomacelllineSMMC7721,theexpressionofBMP-2mRNAandproteinweresignificantlyinhibitedbyRT-PCRandWesternblottingmethods,withBMP-2-siRNA-Bhavingthehighestinhibitionrate[(0.32±0.
7、07)vs(0.92±0.14),(0.37±0.10)vs(0.89±0.17),P<0.01].Theinvasionassaydemonstratedthatthenumberofhepatomacancercellintransfectiongroupwassignificantlylowerthanthatinuntransducedgroup,blankcontrolgroupandnegativecontrolgroup[(6.48±3.62)vs(23.72±3.24),(22.38±3.84),P<0.05],wh
8、ichsuggestedthattheinhibitioneffectofBMP-2geneexpressioncanobviouslydecreasethemigrationandinvasionabilityofhepatomac