miR-409-3p通过抑制PIK3CA介导的自噬增强耐药卵巢癌细胞对顺铂的敏感性

miR-409-3p sensitizes drug-resistant ovarian cancer cells to cisplatin by inhibiting PIK3CA-mediated autophagy

目的:探究微小RNA-409-3p(miR-409-3p)对耐药卵巢癌细胞顺铂敏感性的影响及其机制。方法:用RT-qPCR检测化疗敏感和耐药卵巢癌组织、耐顺铂卵巢癌细胞及其亲本细胞中miR-409-3p的表达情况。对耐顺铂卵巢癌细胞转染miR-409-3p mimics或miR-NC后,再给予顺铂处理,用WST-1法检测细胞活力,流式细胞术检测细胞凋亡比例,Western blot检测磷脂酰肌醇3-激酶催化亚基α(PIK3CA)、beclin-1、LC3和SQSTM1表达情况。生物信息学和双萤光素酶报告基因实验检测miR-409-3p的靶基因,并通过功能修复实验验证。构建荷瘤裸鼠模型,分析miR-409-3p及其靶基因在体内对耐顺铂卵巢癌细胞顺铂敏感性的影响。结果:耐药卵巢癌组织中miR-409-3p的表达水平显著低于敏感组织(P<0.05),耐药卵巢癌细胞中miR-409-3p的表达水平显著低于亲本细胞(P<0.05)。转染miR-409-3p mimics后,耐药卵巢癌细胞对顺铂的敏感性显著增强(P<0.05)。与miR-NC组比较,miR-409-3p mimic组PIK3CA和beclin-1表达水平及LC3-Ⅱ/LC3-Ⅰ比值显著降低(P<0.05),SQSTM1表达水平显著升高(P<0.05)。PIK3CA是miR-409-3p的靶基因。miR-409-3p能在体内增强耐药卵巢癌细胞对顺铂的敏感性(P<0.05)。结论:miR-409-3p通过抑制靶基因PIK3CA来抑制促生存自噬,进而恢复耐药卵巢癌细胞的顺铂敏感性。

AIM:To explore the effect of microRNA-409-3p(miR-409-3p)on cisplatin sensitivity of drug-resistant ovarian cancer cells and its mechanism.METHODS:The expression of miR-409-3p in chemosensitive and chemoresistant ovarian cancer tissues,and cisplatin-resistant ovarian cancer cells and their parents was detected by RT-qPCR.Cisplatin-resistant ovarian cancer cells were transfected with miR-409-3p mimics or miR-NC,and then treated with cisplatin.Cell viability was detected by WST-1 assay,apoptosis was detected by flow cytometry,and the expression levels of phosphatidylinositol 3-kinase catalytic subunitα(PIK3CA),beclin-1,LC3 and SQSTM1 were detected by Western blot.The target gene of miR-409-3p was detected by bioinformatics and luciferase reporter assay,and verified by functional repair experiments.A tumor-bearing nude mouse model was constructed to analyze the effect of miR-409-3p and its target gene on cisplatin sensitivity of cisplatin resistant ovarian cancer cells in vivo.RESULTS:The expression level of miR-409-3p in chemoresistant ovarian cancer tissues was lower than that in sensitive tissues(P<0.05),and that in drug-resistant ovarian cancer cells was lower than that in parental cells(P<0.05).The ovarian cancer cells were more sensitive to cisplatin after transfection with miR-409-3p mimics(P<0.05).Compared with miR-NC group,the expression of PIK3CA and beclin-1,and the ratio of LC3-Ⅱ/LC3-Ⅰin miR-409-3p mimic group was decreased(P<0.05),while the expression of SQSTM1 was increased(P<0.05).PIK3CA was the target gene of miR-409-3p.miR-409-3p could enhance the sensitivity of drug-resistant ovarian cancer cells to cisplatin in vivo(P<0.05).CONCLUSION:miR-409-3p inhibits pro-survival autophagy by inhibiting the target gene PIK3CA,and restores cisplatin sensitivity of drug-resistant ovarian cancer cells.