中国肺癌杂志
中國肺癌雜誌
중국폐암잡지
CHINESE JOURNAL OF LUNG CANCER
2014年
5期
384-390
,共7页
张芳%李洋%吴蘅%齐康%尤嘉琮%李雪冰%祖玲玲%潘振华%王玉丽%李永文%李颖%王珉%沈旺%周清华
張芳%李洋%吳蘅%齊康%尤嘉琮%李雪冰%祖玲玲%潘振華%王玉麗%李永文%李穎%王珉%瀋旺%週清華
장방%리양%오형%제강%우가종%리설빙%조령령%반진화%왕옥려%리영문%리영%왕민%침왕%주청화
肺肿瘤%miRNA芯片%RT-PCR%miR-192%Bim
肺腫瘤%miRNA芯片%RT-PCR%miR-192%Bim
폐종류%miRNA심편%RT-PCR%miR-192%Bim
Lung neoplasms%miRNA microarray%RT-PCR%miR-192%Bim
背景与目的顺铂是非小细胞肺癌化疗的一线药物，但获得性耐药限制了其疗效的发挥。本研究的目的是筛选鉴定与肺癌顺铂耐药相关的microRNAs，探讨其与肺癌顺铂耐药的影响及分子机制。方法应用miRNA芯片及RT-PCR筛选鉴定A549与A549/DDP肺癌细胞间的差异表达miRNAs，将差异表达miR-192转染A549和A549/DDP细胞株，CCK-8检测miR-192对半数抑制浓度（half inhibition concentration, IC50）的影响，流式细胞术检测细胞凋亡，生物软件预测及双荧光素酶报告基因法寻找miR-192的靶基因，RT-PCR及Western blot检测靶基因表达水平在转染前后的变化。结果 MiR-192在A549/DDP中显著高表达，表达量是A549细胞表达量的37.59±0.35倍。在低表达miR-192的A549细胞中过表达miR-192，细胞对顺铂的IC50显著增高，顺铂引起的细胞凋亡率显著降低；反之，在高表达miR-192的A549/DDP中抑制miR-192的表达，顺铂的IC50显著降低，顺铂引起的细胞凋亡率显著增加。miR-192可靶向作用促凋亡基因Bim的3’-UTR，并在转录后水平负向调控Bim的表达。结论 MiR-192通过靶向负调控促凋亡基因Bim表达，诱导肺腺癌细胞株A549产生顺铂耐药，并减少顺铂引起的细胞凋亡。
揹景與目的順鉑是非小細胞肺癌化療的一線藥物，但穫得性耐藥限製瞭其療效的髮揮。本研究的目的是篩選鑒定與肺癌順鉑耐藥相關的microRNAs，探討其與肺癌順鉑耐藥的影響及分子機製。方法應用miRNA芯片及RT-PCR篩選鑒定A549與A549/DDP肺癌細胞間的差異錶達miRNAs，將差異錶達miR-192轉染A549和A549/DDP細胞株，CCK-8檢測miR-192對半數抑製濃度（half inhibition concentration, IC50）的影響，流式細胞術檢測細胞凋亡，生物軟件預測及雙熒光素酶報告基因法尋找miR-192的靶基因，RT-PCR及Western blot檢測靶基因錶達水平在轉染前後的變化。結果 MiR-192在A549/DDP中顯著高錶達，錶達量是A549細胞錶達量的37.59±0.35倍。在低錶達miR-192的A549細胞中過錶達miR-192，細胞對順鉑的IC50顯著增高，順鉑引起的細胞凋亡率顯著降低；反之，在高錶達miR-192的A549/DDP中抑製miR-192的錶達，順鉑的IC50顯著降低，順鉑引起的細胞凋亡率顯著增加。miR-192可靶嚮作用促凋亡基因Bim的3’-UTR，併在轉錄後水平負嚮調控Bim的錶達。結論 MiR-192通過靶嚮負調控促凋亡基因Bim錶達，誘導肺腺癌細胞株A549產生順鉑耐藥，併減少順鉑引起的細胞凋亡。
배경여목적순박시비소세포폐암화료적일선약물，단획득성내약한제료기료효적발휘。본연구적목적시사선감정여폐암순박내약상관적microRNAs，탐토기여폐암순박내약적영향급분자궤제。방법응용miRNA심편급RT-PCR사선감정A549여A549/DDP폐암세포간적차이표체miRNAs，장차이표체miR-192전염A549화A549/DDP세포주，CCK-8검측miR-192대반수억제농도（half inhibition concentration, IC50）적영향，류식세포술검측세포조망，생물연건예측급쌍형광소매보고기인법심조miR-192적파기인，RT-PCR급Western blot검측파기인표체수평재전염전후적변화。결과 MiR-192재A549/DDP중현저고표체，표체량시A549세포표체량적37.59±0.35배。재저표체miR-192적A549세포중과표체miR-192，세포대순박적IC50현저증고，순박인기적세포조망솔현저강저；반지，재고표체miR-192적A549/DDP중억제miR-192적표체，순박적IC50현저강저，순박인기적세포조망솔현저증가。miR-192가파향작용촉조망기인Bim적3’-UTR，병재전록후수평부향조공Bim적표체。결론 MiR-192통과파향부조공촉조망기인Bim표체，유도폐선암세포주A549산생순박내약，병감소순박인기적세포조망。
Background and objective Cisplatin is the ifrst-line drug for the chemotherapy of non-small cell lung cancer (NSCLC), but the acquired chemoresistance restricted the effect of its treatment. hTe aim of this study is to validate the miRNAs related to the Cisplatin resistance in lung cancer and elucidate the molecular mechanisms. Methods We performed miRNA microarray and RT-PCR to obtain the aberrant differential expressed miRNAs between A549 and its paired Cisplatin-resistant cell line A549/DDP cells, and then we investigated the biological functions of miR-192, which is the aberrant differen-tial expressed miRNA. Atfer transfection of the miR-192 into A549 cells, we measured the half inhibition concentration (IC50), cell apoptosis of the trasfectant cells, and then we used biological sotfwares and dual-luciferase report assay to explore the target gene of the miR-192, which was further validated by RT-PCR and Western blot. Result MiR-192 was highly over-expressed in A549/DDP cells , whose quantity was 37.59±0.35 fold higher than that in A549 cells. Overexpression of miR-192 in A549 cells signiifcantly conferred resistance to Cisplatin and inhibited apoptosis. By contrast, down-expression of miR-192 in A549/DDP cells remarkably restrained the Cisplatin resistance and induced apoptosis. MiR-192 binded to Bim 3’-UTR and negatively regulated Bim expression at the post-transcriptional level in lung adenocarcinoma cells. Conclusion Our data suggested that miR-192 induced Cisplatin-resistance and inhibited cell apoptosis in lung cancer via negative targeting Bim expression.
