CAJ | 학술논문

目的探讨肿瘤细胞微RNA(miRNA)对化疗药物敏感性的作用.方法通过miRNA芯片技术检测顺铂(DDP)耐药细胞株A549/DDP与非耐药细胞株A549的miRNA表达的差异,利用荧光定量聚合酶链反应(PCR)技术验证相应miRNA的表达情况,通过在细胞株中抑制或过表达目标miRNA,研究其对细胞化疗药物敏感性的影响.结果 A549/DDP细胞对DDP的耐药为A549细胞的18倍.A549/DDP细胞与A549细胞存在51个表达水平差异在4倍以上的miRNA,其中24个表达上调,27个表达下调.PCR进一步证实miR-376c、miR-31、miR-29a、miR-221在A549/DDP细胞中显著上调,miR-196a、miR-20a、miR-20b、miR-17、miR-451在A549/DDP细胞中显著下调.在提高A549/DDP细胞中miR-17的表达后,细胞对DDP的敏感度增加了11.7％,提高miR-451的表达或者抑制miR-29a的表达后,对DDP的敏感度分别下降了15.5％、12.9％,抑制miR-376c、miR-31、miR-221或过表达miR-196a、miR-20a、miR-20b均不影响A549/DDP细胞对DDP的敏感度.结论非小细胞肺癌DDP耐药细胞与非耐药细胞的miRNA表达谱有差异,miRNA参与肺癌化疗耐药,miR-17具有逆转非小细胞肺癌DDP耐药的潜力.
목적 탐토종류세포미RNA(miRNA)대화료약물민감성적작용.방법 통과miRNA심편기술검측순박(DDP)내약세포주A549/DDP여비내약세포주A549적miRNA표체적차이,이용형광정량취합매련반응(PCR)기술험증상응miRNA적표체정황,통과재세포주중억제혹과표체목표miRNA,연구기대세포화료약물민감성적영향.결과 A549/DDP세포대DDP적내약위A549세포적18배.A549/DDP세포여A549세포존재51개표체수평차이재4배이상적miRNA,기중24개표체상조,27개표체하조.PCR진일보증실miR-376c、miR-31、miR-29a、miR-221재A549/DDP세포중현저상조,miR-196a、miR-20a、miR-20b、miR-17、miR-451재A549/DDP세포중현저하조.재제고A549/DDP세포중miR-17적표체후,세포대DDP적민감도증가료11.7％,제고miR-451적표체혹자억제miR-29a적표체후,대DDP적민감도분별하강료15.5％、12.9％,억제miR-376c、miR-31、miR-221혹과표체miR-196a、miR-20a、miR-20b균불영향A549/DDP세포대DDP적민감도.결론 비소세포폐암DDP내약세포여비내약세포적miRNA표체보유차이,miRNA삼여폐암화료내약,miR-17구유역전비소세포폐암DDP내약적잠력.
Objective To analyze the differences in microRNA (miRNA) expression between A549 and A549/DDP cells and explore the association between miRNA expression and drug resistance in non-small cell lung cancer (NSCLC).Methods The drug resistance of A549/DDP cells was evaluated using CCK-8 assay and flow cytometry.Microarray technique and RT-PCR were used to analyze the differential expression of the miRNA between A549 and A549/DDP cells.Enforced or inhibited target miRNA expression in cisplatin resistant cell was used to investigate whether miRNA involve in modulating the sensitivity of NSCLC cells to chemotherapeutic agent,exploiting the emerging knowledge of miRNA for the development of new human therapeutic applications for overcoming anticancer drug resistance and trying to discover biomarkers that were better able to predict the cancer chemotherapy sensitivity.Results The drug resistance index of A549/DDP cells relative to the parental A549 cells was 18.Microarray analysis of A549 and A549/DDP cells identified 51 differentially expressed genes (≥4-fold),including 24 up-regulated and 27 down-regulated genes in A549/DDP cells.RT-PCR identified 9 miRNA that were differentially expressed between A549 and A549/DDP cells.Of these differentially expressed miRNA,miR-376c,miR-31,miR-29a,miR-221 showed significantly increased expression,and miR-196a,miR-20a,miR-20b,miR-17,miR-451 showed significantlylowered expression in A549/DDP cells as indicated by the results of microarray analysis and RT-PCR.DDP sensitivity was increased 11.7 ％ in A549/DDP cells transfected with miR-17,but the chemosensitivity was decreased when miR-451 was over-expressed or miR-29a was inhibited by selective inhibitor,the reduction was 15.5 ％,12.9 ％,respectively,whereas chemosensitivity did not change when miR-376c,miR-31,miR-221 were inhibited or miR-196a,miR-20b,miR-20a were over-expressed.Conclusion A549/DDP cells show a different miRNA expression profile from its parental A549 cells,suggesting the involvement of miRNA in tumor cell drug resistance.miR-17 has the potential to be an efficient agent for preventing and reversing DDP-resistance in NSCLC.These results provide a strong rationale for the development of miRNA-based therapeutic strategies aiming to overcome cancer cell resistance.