miRNA-192对非小细胞肺癌A549／DDP细胞顺铂耐药性的影响

目的 探讨抑制microRNA-192（miR-192）在非小细胞肺癌A549／DDP细胞对顺铂（DDP）耐药性方面的影响及可能机制。方法 通过实时荧光定量PCR（qRT-PCR）法检测人肺腺癌耐DDP细胞株A549／DDP及其亲本细胞株A549细胞中miR-192的表达水平，A549／DDP细胞转染miR-192抑制剂（inhibitor）和miR-阴性对照（NC）48h后采用qRT-PCR检测转染效率，分别采用MTT法、克隆形成实验及流式细胞术检测转染48h后A549／DDP细胞对DDP的药物敏感性、细胞增殖能力及细胞凋亡变化，Western blotting检测转染48h后细胞中Bax和Bcl-2的表达变化。结果 miR-192在A549／DDP细胞中的表达水平高于A549细胞（P＜0.05）；转染miR-192 inhibitor 48h后的A549／DDP细胞miR-192水平低于转染miR-NC者（P＜0.05）；转染miR-192 inhibitor后，A549／DDP细胞的增殖能力减弱、凋亡细胞增多、DDP对其半数抑制浓度降低、Bax蛋白水平升高和Bcl-2蛋白水平下降，与转染miR-NC者比较，差异均有统计学意义（P＜0.05）。结论 抑制miR-192能够降低A549／DDP细胞对DDP的耐药性，其作用机制可能是通过增加细胞凋亡以及下调Bcl-2蛋白和上调Bax蛋白表达来实现的。

Impact of miRNA-192 for cisplatin resistance of non-small cell lung cancer A549/DDP cells

Objective To explore the effect of microRNA-192（ miR-192） in enhancing cisplatin（ DDP） resistance of nonsmall cell lung（ NSCLC） A549 /DDP cell line and elucidate its related mechanism. Methods The real-time quantitative PCR（ qRTPCR） was used to detect miR-192 expression in A549 cells and A549 /DDP cells. The A549 /DDP cells were transfected with miR-192inhibitor or miR-negative control（ NC） and qRT-PCR was used to test the transfection efficiency. Drug sensitivity to DDP 48h after transfection was tested by MTT assay. Colony formation assay was used to detect the proliferation of A549 /DDP cells 48h after transfection. Flow cytometric analysis was used to observe apoptosis of A549 /DDP cells 48h after transfection on treatment with DDP. The protein expressions of Bax and Bcl-2 in A549 /DDP cells after transfection were detected by Western blotting method. Results The miR-192 expression in A549 /DDP cell line was significantly higher than that in A549 cell line（ P〈0. 05）. The miR-192 expression was significantly decreased at 48h after transfection of miR-192 inhibitor than that of miR-NC（ P〈0. 05）. The half inhibition concentration of DDP was significantly lower in A549 /DDP cells transfection of miR-192 inhibitor than miR-NC（ P〈0. 05）. Colony formation assay showed that the proliferation of miR-192 inhibitor transfected A549 /DDP cells was significantly inhibited（ P〈0. 05）. Compared with miR-NC A549 /DDP cells,the apoptosis of cells transfected with miR-192 inhibitor increased after treatment with DDP（ P〈0. 05）.Furthermore,the down-regulated Bcl-2 and up-regulated Bax were observed in A549 /DDP cells transfection of miR-192 inhibitor versus miR-NC（ P〈0. 05）. Conclusion Inhibiting expression of miR-192 could reduce the resistance of A549 /DDP cells to DDP with the possible mechanism of inducing apoptosis,down-regulating the expressions of Bcl-2 protein and up-regulating the expression of Bax protein.