miR-584下调CCN2对结直肠癌SW1116细胞增殖、侵袭的影响及机制研究

Effect of miR-584 down regulating CCN2 on proliferation and invasion of colorectal cancer SW1116 cells and its mechanism

目的探讨miR-584下调CCN2对结直肠癌SW1116细胞增殖、侵袭的影响及机制。方法2020年1—4月于辽宁省肿瘤医院中心实验室进行实验,设立SW1116直肠癌细胞组、miR-NC组(空白载体)、miR-584 mimics组(过表达)、miR-584抑制剂组(低表达);各组细胞每孔设6个平行样,培养72 h。采用MTT法、Transwell法和流式细胞术分别测定细胞增殖、侵袭、凋亡水平,RT-PCR及蛋白印迹法测定miR-584、CCN2 mRNA与蛋白表达水平。结果与SW1116直肠癌细胞组比较,miR-584 mimics组OD值、存活率、单克隆形成数目、穿膜数降低,凋亡率升高(P<0.05),miR-584抑制剂组OD值、存活率、单克隆形成数目、穿膜数升高,凋亡率降低(P<0.05);与miR-584 mimics组比较,miR-584抑制剂组OD值、存活率、单克隆形成数目、穿膜数升高,凋亡率降低(t=16.854、19.634、18.532、24.547、35.951,P均=0.000)。与SW1116直肠癌细胞组比较,miR-584 mimics组细胞miR-584 mRNA表达升高,CCN2 mRNA、蛋白表达降低(P<0.05),miR-584抑制剂组细胞miR-584 mRNA表达降低,CCN2 mRNA、蛋白表达升高(P<0.05);与miR-584 mimics组比较,miR-584抑制剂组细胞miR-584 mRNA表达降低,CCN2 mRNA和蛋白表达升高(t=28.654、26.254、19.965,P均=0.000)。SW1116直肠癌细胞组与miR-NC组上述指标水平比较,差异无统计学意义(P>0.05)。结论miR-584能抑制结直肠癌SW1116细胞增殖、侵袭,促进其凋亡,其机制可能与miR-584下调CCN2的表达有关。

Objective To explore the effect and mechanism of miR-584 down-regulation of CCN2 on the proliferation and invasion of colorectal cancer SW1116 cells.Methods From January to April 2020,experiments were carried out in the Central Laboratory of Liaoning Cancer Hospital,setting up SW1116 rectal cancer cell group,miR NC group(blank vector),miR-584 mimics group(overexpression),and miR-584 inhibitor group(low expression).Each group of cells set up 6 parallel samples per well and cultured for 72 h.MTT method,Transwell method and flow cytometry were used to determine the levels of cell proliferation,invasion,and apoptosis.RT PCR and Western blotting were used to determine the mRNA and protein expression levels of miR-584 and CCN2.Results Compared with the SW1116 rectal cancer cell group,the miR-584 mimics group OD value,survival rate,number of single clones,number of transmembrane formation decreased,and apoptosis rate increased(P<0.05),miR-584 inhibitor group OD value,survival rate,single clone formation The number and the number of penetrating membrane increased,and the apoptosis rate decreased(P<0.05).Compared with the miR-584 mimics group,the miR-584 inhibitor group increased the OD value,survival rate,number of colonies formed,and number of membrane penetration,and decreased apoptosis rate(t=16.854,19.634,18.532,24.547,35.951,P=0.000).Compared with the SW1116 rectal cancer cell group,the miR-584 mRNA expression in the miR-584 mimics group was increased,and the CCN2 mRNA and protein expression was decreased(P<0.05).The miR-584 mRNA expression in the miR-584 inhibitor group was decreased,and the CCN2 mRNA and protein expression was increased(P<0.05).Compared with the miR-584 mimics group,the miR-584 mRNA expression in the miR-584 inhibitor group decreased,and the CCN2 mRNA and protein expression increased(t=28.654,26.254,19.965,all P=0.000).There was no statistically significant difference in the levels of the above indicators between the SW1116 rectal cancer cell group and the miR NC group(P>0.05).Conclusion miR-584 can inhibit the proliferation and invasion of colorectal cancer SW1116 cells,and promote their apoptosis.The mechanism may be related to the down-regulation of CCN2 expression by miR-584.