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目的:通过建立人胃癌细胞SGC-7901的顺铂耐药细胞株SGC-7901/DDP,探讨miR-497对SGC-7901/DDP耐药性的影响及其机制。方法:体外研究采用顺铂体外逐步加量诱导法建立人胃癌细胞SGC-7901耐药株,并通过检测药物半抑制浓度和耐药基因MDR1、BCRP、MRP1的表达以鉴定耐药细胞株;检测在亲本及耐药细胞中miR-497、MDR1、MRP1、BCRP和凋亡相关基因Bax、Bcl-2的表达水平;miR-497模拟物分别转染SGC-7901/DDP细胞株,利用SRB法和流式细胞术检测转染miR-497模拟物后细胞对顺铂醇的敏感程度和细胞的周期、凋亡的变化,并检测耐药基因和凋亡相关基因的表达。结果:成功建立人胃癌SGC-7901/DDP耐药细胞株,耐药细胞株中耐药基因MDR1、BCRP、MRP1表达均升高,抗凋亡基因Bcl-2升高,凋亡基因Bax下降,miR-497表达下降(P<0.05);miR-497模拟物提高耐药细胞株对顺铂的药物敏感性,凋亡水平增加,细胞周期G0/G1期细胞增多(P<0.05),并可抑制耐药细胞中耐药基因的表达,降低Bcl-2/Bax比值(P<0.05)。结论:人胃癌耐药细胞株SGC-7901/DDP的miR-497表达下调;上调miR-497的表达可逆转人胃癌SGC-7901/DDP细胞株对化疗药物顺铂的耐药性。
OBJECTIVE: To investigate the effect of miR-497 on the drug resistance of SGC-7901 / DDP and its mechanism by establishing a cisplatin-resistant cell line SGC-7901 / DDP of human gastric cancer cell line SGC-7901. Methods: SGC-7901 resistant human gastric cancer cell lines were induced by cisplatin in vitro and gradually. The drug-resistant cell lines were identified by detecting the concentrations of half-inhibitory drugs (MDR1, BCRP and MRP1) The expression levels of miR-497, MDR1, MRP1, BCRP and apoptosis-related genes Bax and Bcl-2 in parental and drug resistant cell lines were determined by Western blotting. The miR-497 mimics were transfected into SGC-7901 / Flow cytometry was used to detect the sensitivity of cells to cisplatin and the changes of cell cycle and apoptosis after transfected with miR-497 mimics. The expression of multidrug resistant genes and apoptosis related genes were detected. Results: The resistant cell lines SGC-7901 / DDP were established successfully. The expression of multidrug resistance gene MDR1, BCRP and MRP1 were all increased, the anti-apoptotic gene Bcl-2 was increased and the apoptosis gene Bax was decreased. miR-497 decreased (P <0.05); miR-497 mimics increased the drug sensitivity of cisplatin to drug-resistant cell lines, increased apoptosis and increased cell number in G0 / G1 phase (P <0.05) Inhibition of drug-resistant cell resistance gene expression, reduce Bcl-2 / Bax ratio (P <0.05). Conclusion: miR-497 expression is down-regulated in human gastric cancer cell line SGC-7901 / DDP. Upregulation of miR-497 expression can reverse the drug resistance of human gastric cancer cell line SGC-7901 / DDP to cisplatin.