circUBE2D2靶向miR-376a-3p调控结直肠癌SW620细胞的增殖、迁移及侵袭

circUBE2D2 Regulates Proliferation,Migration and Invasion of SW620 Colorectal Cancer Cells via miR-376a-3p

目的 探讨circUBE2D2对结直肠癌SW620细胞增殖、迁移及侵袭的影响及其可能作用机制.方法 收集2020年1月至2020年5月大连大学附属新华医院肛肠外科收治的45例结直肠癌患者的癌组织及其相应癌旁组织标本,采用qRT-PCR法检测circUBE2D2、miR-376a-3p的表达量;以人结直肠癌细胞SW620为研究对象,随机分为si-NC组、si-circUBE2D2组、miR-NC组、miR-376a-3p组、si-circUBE2D2+anti-miR-NC组和si-circUBE2D2+anti-miR-376a-3p组;CCK-8法、平板克隆形成实验、划痕实验与Tran-swell实验分别检测细胞增殖、克隆形成、迁移及侵袭;双荧光素酶报告实验检测miR-376a-3p过表达对野生型载体WT-circUBE2D2的荧光素酶活性;Western印迹法检测上皮型钙黏蛋白(E-cadherin)、神经型钙黏蛋白(N-cadherin)蛋白表达量.结果 与癌旁组织比较,结直肠癌组织中circUBE2D2的表达量升高(P<0.05),miR-376a-3p的表达量降低(P<0.05);与si-NC组比较,si-circUBE2D2组细胞活力、划痕愈合率和N-cadherin蛋白水平降低(P<0.05),细胞克隆形成数和侵袭细胞数减少(P<0.05),E-cadherin蛋白水平升高(P<0.05);与miR-NC组比较,miR-376a-3p组细胞活力、划痕愈合率和N-cadherin蛋白水平降低(P<0.05),细胞克隆形成数和侵袭细胞数减少(P<0.05),E-cadherin蛋白水平升高(P<0.05);miR-376a-3p过表达可抑制野生型载体WT-circUBE2D2的荧光素酶活性(P<0.05);与si-circUBE2D2+an-ti-miR-NC组比较,si-circUBE2D2+anti-miR-376a-3p组细胞活力、划痕愈合率和N-cadherin蛋白水平升高(P<0.05),细胞克隆形成数和侵袭细胞数增多(P<0.05),E-cadherin蛋白水平降低(P<0.05).结论 干扰circUBE2D2表达可通过靶向调控miR-376a-3p表达而降低结直肠癌细胞增殖、克隆形成、迁移及侵袭能力.

Objective To explore the effect of circUBE2D2 on the proliferation,migration and invasion of SW620 colorectal cancer cells and its underlying mechanism.Methods The cancer tissues and the corresponding para-cancerous tissues from 45 patients with colorectal cancer admitted to our hospital from January 2020 to May 2020 were collected.The expression levels of circUBE2D2 and miR-376a-3p were detected by qRT-PCR.Human colorectal cancer cell line SW620 was used in this research and were divided into groups as follows:si-NC group,si-circUBE2D2 group,miR-NC group,miR-376a-3p group,si-circUBE2D2+anti-miR-NC group,and si-circliBE2D2+anti-miR-376a-3p group.The CCK-8 and plate colony formation assay were used to measure the proliferation and colony formation of cells.The migration and invasion of cells were determined by the wound healing assay and the transwell assay.The dual luciferase reporter experiment was used to assay the effect of miR-376a-3p overexpression on the luciferase activity of the wild-tvpe vector WT-cir-cUBE2D2.The expression levels of E-cadherin and N-cadherin proteins were detected by Western blotting.Results The expression level of circUBE2D2 was increased in colorectal cancer tissues,while that of miR-376a-3p was decreased when compared with adjacent tissues(P<0.05).The cell viability and scratch healing rate were reduced,the number of colonies formed,the number of invasive cells and the expression level of N-cadherin were decreased in the si-circUBE2D2 group and the miR-376a-3p group when compared with the si-NC group and the miR-NC group,while the expression level of E-cadherin was increased respectively(P<0.05).Overexpression of miR-376a-3p could inhibit the luciferase activity of wild-type vector WT-circUBE2D2(P<0.05).The cell viability,the scratch healing rate,the number of cell colony formed,the number of invasive cells,and the expression level of N-cadherin were increased in the si-circUBE2D2+anti-miR-376a-3p group,while the expression level of E-cadherin was decreased when compared with the si-circUBE2D2+anti-miR-NC group(P<0.05).Conclusion Interference of circUBE2D2 expression could reduce the proliferation,colony formation,migration and invasion of colorectal cancer cells by targeting miR-376a-3 p.