miRNA-21-5p通过pdcd4靶向调控胃癌细胞对顺铂敏感性的机制研究

目的:探究微小RNA-21-5p（miRNA-21-5p）通过下调pdcd4基因表达调控胃癌耐药细胞系SGC-7901/DDP对顺铂敏感性机制的研究。方法:采用四甲基偶氮唑蓝（MTT）法检测不同浓度的顺铂（0、1、2、4、8、16、32 μmol/L）对胃癌细胞系SGC7901及胃癌耐药细胞系SGC7901/DDP增殖抑制的影响。通过实时荧光定量PCR检测SGC7901/DDP细胞中瞬时转染miRNA-21-5p inhibitors后miRNA-21-5p相对表达量的变化。采用Western blot及实时荧光定量PCR检测转染组、无关序列转染组（NC组）及空白转染组（CON组）分别转染miRNA-21-5p inhibitors及无关序列后对靶基因pdcd4蛋白及mRNA表达量的影响。结果:MTT实验结果显示耐药细胞SGC7901/DDP相对于亲本细胞SGC7901对顺铂不敏感（P<0.05）。实时荧光定量PCR结果表明，转染组miRNA-21-5p表达量均明显低于NC组和CON组（P<0.01），转染组pdcd4 mRNA表达量均明显高于NC组和CON组（P<0.01）。Western blot检测结果示，转染组pdcd4蛋白表达量均明显高于NC组和CON组（P<0.01）。结论:miRNA-21-5p表达水平的升高在一定程度上有利于胃癌细胞增殖，并通过下调pdcd4的表达，降低胃癌细胞对顺铂的敏感性。

The mechanism research of miRNA-21-5p in the targeted regulation of the sensitivity of gastric cancer cells to cisplatin by pdcd4

Objective:To investigate the mechanism of microRNA-21-5p(miRNA-21-5p) in the targeted regulation of sensitivity of gastric cancer resistant cell line SGC7901/DDP to cisplatin by down-regulating the expression of pdcd4 gene.Methods:The inhibition effects of different concentrations of cisplatin(0,1,2,4,8,16,32 μmol/L) on the proliferation of gastric cancer cell line SGC7901 and gastric cancer drug-resistant cell line SGC7901/DDP were detected using the MTT.The relative expression of miRNA-21-5 in SGC7901/DDP cells after transient transfecting of miRNA-21-5p inhibitors was detected using real-time fluorescence quantitative polymerase chain reaction(qRT-PCR).The expressions of pdcd4 protein and mRNA in transfection group,irrelevant sequence transfection group(NC group) and blank transfection group(CON group) were detected after transfecting with miRNA-21-5p inhibitors and irrelevant sequence,respectively.Results:The results of MTT assay showed that the drug-resistant cells SGC7901/DDP were insensitive to cisplatin compared with the parent cells SGC7901(P<0.05).The results of qRT-PCR showed that the expression level of miRNA-21-5p in transfection group was significantly lower than that in NC group and CON group(P<0.01),the expression level of pdcd4 mRNA in transfection group was significantly higher than that in NC group and CON group(P<0.01).The results of Western blot showed that the expression level of pdcd4 protein in transfection group was significantly higher than that in NC group and CON group(P<0.01).Conclusions:The increasing of miRNA-21-5p expression level is beneficial to the proliferation of gastric cancer cells,and can reduce the sensitivity of gastric cancer cells to cisplatin by down-regulating the expression of pdcd4.