临床误诊误治
臨床誤診誤治
림상오진오치
CLINICAL MISDIAGNOSIS & MISTHERAPY
2015年
1期
108-112
,共5页
孔祥玲%吴维光%佘野%葛红雨
孔祥玲%吳維光%佘野%葛紅雨
공상령%오유광%사야%갈홍우
宫颈肿瘤%组蛋白去乙酰化酶1%RNA干扰%化疗敏感性
宮頸腫瘤%組蛋白去乙酰化酶1%RNA榦擾%化療敏感性
궁경종류%조단백거을선화매1%RNA간우%화료민감성
Cancer of cervix%Histone deacetylase 1%RNA interference%Chemosensitivity
目的：探讨体外应用RNA干扰技术抑制组蛋白去乙酰化酶1(HDAC1)表达,提高人宫颈癌HeLa细胞对顺铂敏感性的作用及机制。方法体外合成针对HDAC1的小干扰RNA(siRNA),脂质体法转染HeLa细胞,将其分为正常对照组(无干预措施)、阴性对照组(转染阴性对照siRNA)及siRNA干扰组(转染HDAC1基因siRNA)。采用Western blot法检测各组细胞HDAC1蛋白和乙酰化组蛋白H4(Ac-H4)的表达,采用聚合酶链反应(PCR)法检测HDAC1基因和NF-κB基因mRNA表达。 HeLa细胞经不同浓度顺铂作用后,采用四甲基偶氮唑蓝比色法检测对顺铂的敏感性,以及采用流式细胞仪检测细胞凋亡率。结果与正常对照组及阴性对照组比较,siRNA干扰组HeLa细胞HDAC1基因mRNA和蛋白表达均降低( P<0．05或P<0．01),细胞内Ac-H4表达增加( P<0．05),同时NF-κB基因mRNA表达降低(P<0．05)。经不同浓度顺铂作用后,与正常对照组及阴性对照组比较,siRNA干扰组HeLa细胞对顺铂的50%抑制浓度降低(P<0．05或P=0．01),细胞凋亡率增加(P<0．01)。结论 HDAC1基因siRNA能够提高宫颈癌HeLa细胞对顺铂的敏感性,增加组蛋白乙酰化水平、抑制HDAC1基因及NF-κB基因表达可能是其作用机制。
目的：探討體外應用RNA榦擾技術抑製組蛋白去乙酰化酶1(HDAC1)錶達,提高人宮頸癌HeLa細胞對順鉑敏感性的作用及機製。方法體外閤成針對HDAC1的小榦擾RNA(siRNA),脂質體法轉染HeLa細胞,將其分為正常對照組(無榦預措施)、陰性對照組(轉染陰性對照siRNA)及siRNA榦擾組(轉染HDAC1基因siRNA)。採用Western blot法檢測各組細胞HDAC1蛋白和乙酰化組蛋白H4(Ac-H4)的錶達,採用聚閤酶鏈反應(PCR)法檢測HDAC1基因和NF-κB基因mRNA錶達。 HeLa細胞經不同濃度順鉑作用後,採用四甲基偶氮唑藍比色法檢測對順鉑的敏感性,以及採用流式細胞儀檢測細胞凋亡率。結果與正常對照組及陰性對照組比較,siRNA榦擾組HeLa細胞HDAC1基因mRNA和蛋白錶達均降低( P<0．05或P<0．01),細胞內Ac-H4錶達增加( P<0．05),同時NF-κB基因mRNA錶達降低(P<0．05)。經不同濃度順鉑作用後,與正常對照組及陰性對照組比較,siRNA榦擾組HeLa細胞對順鉑的50%抑製濃度降低(P<0．05或P=0．01),細胞凋亡率增加(P<0．01)。結論 HDAC1基因siRNA能夠提高宮頸癌HeLa細胞對順鉑的敏感性,增加組蛋白乙酰化水平、抑製HDAC1基因及NF-κB基因錶達可能是其作用機製。
목적：탐토체외응용RNA간우기술억제조단백거을선화매1(HDAC1)표체,제고인궁경암HeLa세포대순박민감성적작용급궤제。방법체외합성침대HDAC1적소간우RNA(siRNA),지질체법전염HeLa세포,장기분위정상대조조(무간예조시)、음성대조조(전염음성대조siRNA)급siRNA간우조(전염HDAC1기인siRNA)。채용Western blot법검측각조세포HDAC1단백화을선화조단백H4(Ac-H4)적표체,채용취합매련반응(PCR)법검측HDAC1기인화NF-κB기인mRNA표체。 HeLa세포경불동농도순박작용후,채용사갑기우담서람비색법검측대순박적민감성,이급채용류식세포의검측세포조망솔。결과여정상대조조급음성대조조비교,siRNA간우조HeLa세포HDAC1기인mRNA화단백표체균강저( P<0．05혹P<0．01),세포내Ac-H4표체증가( P<0．05),동시NF-κB기인mRNA표체강저(P<0．05)。경불동농도순박작용후,여정상대조조급음성대조조비교,siRNA간우조HeLa세포대순박적50%억제농도강저(P<0．05혹P=0．01),세포조망솔증가(P<0．01)。결론 HDAC1기인siRNA능구제고궁경암HeLa세포대순박적민감성,증가조단백을선화수평、억제HDAC1기인급NF-κB기인표체가능시기작용궤제。
Objective To observe histone deacetylase 1 (HDAC1) expression inhibited by siRNA in vitro on chemo-sensitivity to cisplatin in human cervical carcinoma cell line HeLa, and to investigate the possible mechanisms. Methods The siRNA targeting HDAC1 was generated in vitro, and transfected into human cervical carcinoma HeLa cell by lipo-fectamine. The HeLa cells were divided into normal control group ( no intervention) , negative control group ( transfected with negative control siRNA) , and siRNA group ( transfected with HDAC1 siRNA) . Western blot was used to detect the HDAC1 protein and acetyl level of histone H4 (Ac-H4), and real time PCR was used to detect mRNA of HDAC1 and NF-κB. The cells were treated under different density DDP. The cell viability was detected by methyl thiazolyl tetrazolium ( MTT) assay and 50% inhibitive concentration ( IC50 ) and sensitivity of DDP were examined, and the apoptosis rate was measured by flow cytometry. Results Compared with those in control groups and negative control group, the expressions of HDAC1 gene pro-tein and mRNA and NF-κB mRNA decreased (P<0. 05 or P<0. 01), and the Ac-H4 increased in siRNA group (P <0. 05). After being treated with different concentrations of DDP, the IC50 to DDP and apoptotic rates in siRNA group were higher than those in other groups (P<0. 05 or P≤0. 01). Conclusion The siRNA targeting HDAC1 can enhance chemo-sensitivity to DDP in cervical cancer HeLa cells. Suppressing the expression of HDAC1 and increasing acetyl level of histone to down-regulate NF-κB gene expression may be its mechanisms.
