miRNA-451逆转非小细胞肺癌A549／DDP细胞对顺铂的耐药性及其机制研究’

目的：探讨microRNA-451（miR-451）逆转非小细胞肺癌A549／DDP细胞对顺铂的耐药性及其可能的作用机制。方法：应用实时荧光定量PCR法（real～timefluorescentquantitativePCR，qRT—PCR）检测耐DDP细胞株A549／DDP及其亲本细胞株A549中miR-451的表达差异，同时检测A549／DDP细胞转染miR-451mimic后miR-451表达的变化；分别应用MTT法、克隆形成实验、流式细胞术检测转染后A549／DDP细胞对DDP药物敏感度，细胞增殖能力，联合DDP处理后细胞凋亡变化；Western印迹法检测转染后A549／DDP细胞Akt、P—Akt（Ser473）、bcl-2和bax的表达变化。结果：miR-451在耐DDP细胞株A549／DDP中的表达量显著低于敏感细胞株A549（P〈0．05），A549／DDP细胞转染miR-451mimic24h后，细胞中miR-451的表达水平较对照组明显提高（P〈0．05）。在A549／DDP细胞中过表达miR-451可产生以下效应：相较于对照组，DDP对A549／DDP／miR-451细胞的半数抑制浓度（half inhibitioncon centration，IC50）减低（P〈0．05），A549／DDP／miR-451细胞的增殖能力减弱，A549／DDP／miR-451经过DDP处理后细胞凋亡增多（P〈0．05）。Western印迹法结果显示，与对照组比较，转染miR-451mimic的A549／DDP细胞P—Akt（Ser473）、bcl-2表达水平降低；bax表达水平升高。结论：miR-451可能通过诱导细胞凋亡，抑制细胞增殖，上调bax蛋白及下调P—Akt（Ser473）、bcl-2蛋白表达而逆转A549／DDP细胞对DDP的耐药性。

miRNA -451 resensitizes non - small cell lung cancer A549/DDP cells to cisplatin resist- ance and the mechanisms involved

Objective：To explore the effect of microRNA -451（miR-451） in reversing eisplatin （DDP） resist-ance of non - small cell lung cancer （NSCLC） A549/DDP cell line and elucidate its related mechanisms. Methods ： The real-time fluorescent quantitative PCR （qRT-PCR） was used to detect miR-451 expression in A549 cells, DDP - resistant A549/DDP ceils, and A549/DDP cells transfeeted with miR-451 mimic or miR - NC. The difference in miR -451 expression was compared between the groups. MTT assay, colony formation assay and flow-cytometric analysis were sequentially used to detect transfection of A549/DDP cells to DDP drug sensitivity, cell proliferation ability, apoptosis on treatment with DDP. The protein expression of Akt, p -Akt （Ser473） , bcl-2 and bax in transfec-tion A549/DDP cells were detected by western blotting. Results：The miR -451 expression level in A549/DDP cell line was significantly lower than that in A549 cell line （ P 〈 0.05 ）. The miR - 451 expression was significantly in-creased at 24h after transfection than that of the control group （P 〈0.05）. Ectopic miR-451 expression in A549/ DDP ceils could produce the following effects ： compared to the control group, the half inhibition concentration （ IC50 ） of DDP in A549/DDP/miR - 451 cells lower （ P 〈 0.05 ）, proliferation of A549/DDP/miR - 451 cells diminished （ P 〈 0.05 ） , apoptosis on treatment with DDP increased （ P 〈 0.05 ）. Furthermore, the protein expressions of p - Akt （Ser473） , bcl -2 were downregnlated and expression of bax was upregnlated in miR -451 mimic transfected group than those in miR-NC group. Condusion：miR -451 reverses the resistance of A549/DDP cells to DDP through in-duction of apoptosis, diminished cell proliferation, down - regulating the expressions of p - Akt（ Ser473 ） , bcl - 2 pro-teins and up - regulating the expression of bax protein.