摘要
目的观察微小RNA-590-3p(miR-590-3p)对结直肠癌(CRC)细胞顺铂(DDP)耐药的影响,并探讨其可能的机制。方法在CRC细胞系SW480、SW620、HCT116中选择miR-590-3p表达最高的SW620细胞构建DDP耐药株(DDP/SW620)。取SW620、DDP/SW620细胞,采用CCK-8法检测并计算DDP作用后的半数抑制浓度(IC50),采用实时荧光定量PCR法检测miR-590-3p表达。将DDP/SW620细胞分为miR-590-3p inhibitor组和NC组,分别转染miR-590-3p inhibitor和NC质粒,计算DDP作用后的IC50。采用TargetScan7.1软件和双荧光素酶报告实验预测miR-590-3p的靶基因为富含亮氨酸的重复序列和免疫球蛋白样结构域蛋白1(LRIG1)。取miR-590-3p inhibitor组和NC组细胞,采用流式细胞术检测细胞凋亡率,采用Western blotting法检测LRIG1、耐药相关基因MDR1和凋亡相关蛋白Bax、Bcl-2蛋白表达。结果DDP/SW620细胞DDP的IC50、miR-590-3p相对表达量均高于SW620细胞,二者比较P均<0.05。miR-590-3p inhibitor组DDP的IC50、miR-590-3p相对表达量均低于NC组,两组比较P均<0.05。miR-590-3p inhibitor组细胞凋亡率及LRIG1、Bax蛋白相对表达量均高于NC组,MDR1和Bcl-2蛋白相对表达量均低于NC组(P均<0.05)。结论miR-590-3p可促进CRC细胞发生DDP耐药,其机制可能与负性调控LRIG1进而促进MDR1表达及减少细胞凋亡有关。
Objective To observe the effect of microRNA-590-3p(miR-590-3p)on cisplatin(DDP)resistance in colorectal cancer(CRC)cells and to explore its possible mechanism.Methods SW620 cells with the highest miR-590-3p expression were selected from CRC cell lines SW480,SW620,and HCT116,to construct DDP-resistant strains(DDP/SW620).CCK-8 was used to detect the half inhibitory concentration(IC50)of SW620 and DDP/SW620 cells after DDP treatment,and miR-590-3p expression was detected by real-time fluorescent quantitative PCR.DDP/SW620 cells were divided into the miR-590-3p inhibitor group and NC group,which were transfected with miR-590-3p inhibitor and NC plasmids,respectively.IC50 of miR-590-3p inhibitor group and NC group was calculated after DDP treatment.TargetScan7.1 software and dual luciferase reporter assay predicted that the target genes of miR-590-3p were leucine-rich repeats and immunoglobulin-like domain protein 1(LRIG1).The cells were collected in the miR-590-3p inhibitor group and NC group,and the apoptosis rate was detected by flow cytometry.The expression levels of LRIG1,resistance-related gene MDR1,and apoptosis-related proteins Bax and Bcl-2 were detected by Western blotting.Results The IC50 of DDP and miR-590-3p expression in DDP/SW620 cells were higher than those in SW620 cells,with statistically significant difference(P<0.05).The IC50 and miR-590-3p relative expression of DDP in the miR-590-3p inhibitor group were lower than those in the NC group,with statistically significant difference between the two groups(P<0.05).The apoptosis rate and relative expression of LRIG1 and Bax proteins in the miR-590-3p inhibitor group were higher than those in the NC group,and the relative expression of MDR1 and Bcl-2 proteins were lower than those in the NC group(all P<0.05).Conclusion MiR-590-3p can promote DDP resistance in CRC cells by negatively regulating LRIG1,thereby promoting MDR1 expression and reducing apoptosis.
作者
徐明
唐澍
刘琦
黄姝娟
XU Ming;TANG Shu;LIU Qi;HUANG Shujuan(Chenzhou First People's Hospital,Chenzhou 423000,China)
出处
《山东医药》
CAS
2020年第11期6-10,共5页
Shandong Medical Journal
基金
湖南省自然科学基金资助项目(2017J0550)。
