中华消化杂志
中華消化雜誌
중화소화잡지
Chinese Journal of Digestion
2013年
6期
399-402
,共4页
冯慧%姚玮艳%陈熹%张辰宇%王亚雷
馮慧%姚瑋豔%陳熹%張辰宇%王亞雷
풍혜%요위염%진희%장신우%왕아뢰
胰腺肿瘤%微RNAs%癌基因蛋白质v-erbB%聚合酶链反应%免疫印迹法
胰腺腫瘤%微RNAs%癌基因蛋白質v-erbB%聚閤酶鏈反應%免疫印跡法
이선종류%미RNAs%암기인단백질v-erbB%취합매련반응%면역인적법
Pancreatic neoplasms%microRNAs%Oncogene proteins v-erbB%Polymerase chain reaction%Immunoblotting
目的 在胰腺癌细胞中寻找并验证微小核糖核酸(miRNA)-148a的下游调控基因.方法 运用生物信息学方法预测miRNA-148a调控的目的基因,构建含目的基因的3’端非翻译区(3'-UTR)的荧光素酶报告质粒,用脂质体转染miRNA-148a类似物和抑制剂进入胰腺癌细胞BXPC-3,检测荧光素酶活性,确定miRNA-148a是否直接与靶基因3’-UTR结合.同时改变胰腺癌细胞BXPC-3中miRNA-148a表达水平,用Western免疫印迹法在蛋白水平检测靶基因ErbB3表达的变化.统计学处理采用单因素方差分析.结果 生物信息学发现ErbB3和miRNA-148a上有一个保守的结合靶位点.miRNA-148a直接与ErbB3的3’-UTR结合,使荧光素酶活性下降到阴性对照的[-(25.00±4.70)％,F=4.66,P＜O.01]；过表达miRNA-148a后,BXPC-3细胞中ErbB3蛋白水平灰度值下降到阴性对照的[(26.16±4.69)％,F=6.563,P＜O.05].结论 miRNA-148a在胰腺癌细胞株BXPC-3中直接靶向并调控ErbB3的表达.
目的 在胰腺癌細胞中尋找併驗證微小覈糖覈痠(miRNA)-148a的下遊調控基因.方法 運用生物信息學方法預測miRNA-148a調控的目的基因,構建含目的基因的3’耑非翻譯區(3'-UTR)的熒光素酶報告質粒,用脂質體轉染miRNA-148a類似物和抑製劑進入胰腺癌細胞BXPC-3,檢測熒光素酶活性,確定miRNA-148a是否直接與靶基因3’-UTR結閤.同時改變胰腺癌細胞BXPC-3中miRNA-148a錶達水平,用Western免疫印跡法在蛋白水平檢測靶基因ErbB3錶達的變化.統計學處理採用單因素方差分析.結果 生物信息學髮現ErbB3和miRNA-148a上有一箇保守的結閤靶位點.miRNA-148a直接與ErbB3的3’-UTR結閤,使熒光素酶活性下降到陰性對照的[-(25.00±4.70)％,F=4.66,P＜O.01]；過錶達miRNA-148a後,BXPC-3細胞中ErbB3蛋白水平灰度值下降到陰性對照的[(26.16±4.69)％,F=6.563,P＜O.05].結論 miRNA-148a在胰腺癌細胞株BXPC-3中直接靶嚮併調控ErbB3的錶達.
목적 재이선암세포중심조병험증미소핵당핵산(miRNA)-148a적하유조공기인.방법 운용생물신식학방법예측miRNA-148a조공적목적기인,구건함목적기인적3’단비번역구(3'-UTR)적형광소매보고질립,용지질체전염miRNA-148a유사물화억제제진입이선암세포BXPC-3,검측형광소매활성,학정miRNA-148a시부직접여파기인3’-UTR결합.동시개변이선암세포BXPC-3중miRNA-148a표체수평,용Western면역인적법재단백수평검측파기인ErbB3표체적변화.통계학처리채용단인소방차분석.결과 생물신식학발현ErbB3화miRNA-148a상유일개보수적결합파위점.miRNA-148a직접여ErbB3적3’-UTR결합,사형광소매활성하강도음성대조적[-(25.00±4.70)％,F=4.66,P＜O.01]；과표체miRNA-148a후,BXPC-3세포중ErbB3단백수평회도치하강도음성대조적[(26.16±4.69)％,F=6.563,P＜O.05].결론 miRNA-148a재이선암세포주BXPC-3중직접파향병조공ErbB3적표체.
Objective To look for and confirm the downstream regulated gene of microRNA (miRNA)-148a in pancreatic cancer cell line.Methods The target gene regulated by miRNA-148a was predicted through bio-informatics analysis.The plasmid containing desired gene and with luciferase 3'-untranslated region (3'-UTR) reporter was constructed.Pancreatic cancer cells BXPC-3 were transfected with analogs and inhibitors of miRNA-148a by liposomes.The activity of luciferase was measured to determine whether miRNA-148a directly connected with desired gene.The expression level of miRNA-148a was changed in BXPC-3 cells,and the changes of target gene v-verb-b2 erythroblastic leukemia viral oncogene homolog 3 (awian) (ErbB3) expression were detected by Western blot at protein level.The data were analyzed by one way ANOVA.Results There was a conservative binding site of ErbB3 with miRNA-148a detected by bio-informatics analysis,miRNA-148a directly combined with ErbB3 and the activity of luciferase decreased to (25.00+47.00) ％ of the negative control (F=4.66,P＜ 0.01).After miRNA-148a overexpression,the gray value of ErbB3 expression in BXPC-3 cells decreased to (26.16±4.69)％ of control group (F=6.563,P＜0.05).Conclusion miRNA-148a directly targeted and regulated the expression of ErbB3 in pancreatic cancer cell line BXPC-3.
