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目的:研究miR-135a/b对肺癌耐顺铂细胞株A549/CDDP顺铂耐药的影响。方法:运用实时荧光定量PCR检测miR-135a/b在A549和A549/CDDP细胞株中的差异表达;MTT法检测转染后A549及A549/CDDP细胞对CDDP的敏感性;构建MCL1-3′-UTR荧光素酶报告质粒验证miR-135a/b的靶基因;Western blot检测转染前后细胞MCL1蛋白的表达差异;流式细胞术检测转染后耐药细胞对顺铂诱导凋亡的影响。结果:miR-135a/b在A549/CDDP细胞中表达量降低;在耐药株中上调miR-135a/b后显著增加细胞对顺铂的敏感性;荧光素酶实验证实MCL1是miR-135a/b的靶基因;抗凋亡蛋白MCL1在A549/CDDP细胞中呈高表达,上调miR-135a/b明显抑制耐药细胞中MCL1蛋白的表达;miR-135a/b显著增加A549/CDDP细胞对顺铂诱导的凋亡。结论:miR-135a/b通过靶向调控MCL1蛋白表达增加NSCLC细胞对顺铂的敏感性和凋亡。
Objective: To investigate the effect of miR-135a / b on cisplatin-resistant lung cancer A549 / CDDP cells. Methods: The differential expression of miR-135a / b in A549 and A549 / CDDP cell lines was detected by real-time fluorescence quantitative PCR. The sensitivity of A549 and A549 / CDDP cells to CDDP was detected by MTT assay. UTR luciferase reporter plasmid was used to verify the target gene of miR-135a / b. Western blot was used to detect the expression of MCL1 protein before and after transfection. Flow cytometry was used to detect the effect of drug-resistant cells on cisplatin-induced apoptosis. Results: The expression of miR-135a / b decreased in A549 / CDDP cells; the up-regulation of miR-135a / b in drug-resistant strains significantly increased the sensitivity of cells to cisplatin; luciferase assay confirmed that MCL1 is a miR- b target gene; anti-apoptotic protein MCL1 was highly expressed in A549 / CDDP cells, upregulation of miR-135a / b significantly inhibited MCL1 protein expression in drug-resistant cells; miR-135a / b significantly increased A549 / CDDP cells Shun Platinum-induced apoptosis. Conclusion: miR-135a / b can increase the sensitivity and apoptosis of NSCLC cells to cisplatin by regulating the expression of MCL1 protein.