lncRNA XIST通过miR-32-5p/EZH2分子轴调控结直肠癌HCT-8细胞的恶性生物学行为

lncRNA XIST regulates the malignant biological behaviors of colorectal cancer HCT-8 cells through miR-32-5p/EZH2 molecular axis

目的:探讨lncRNA XIST通过miR-32-5p/果蝇Zeste基因增强子同源物2(enhancer of zeste homolog2,EZH2)分子轴调控结直肠癌HCT-8细胞的恶性生物学行为。方法:收集2014年7月至2018年8月中南大学湘雅医院直肠肛门外科资料完整的结直肠癌患者28例癌组织和配对的癌旁组织标本,采用qPCR检测结直肠癌组织及细胞系中lncRNA XIST和miR-32-5p的表达水平,双荧光素酶报告基因验证1ncRNA XIST、miR-32-5p和EZH2的靶向关系,并进一步通过WB检测EZH2的表达水平。CCK-8、Transwell及Annexin V-FITC/PI染色流式细胞术检测HCT-8细胞增殖、迁移及凋亡情况。结果:lncRNA XIST在结直肠癌组织及细胞系中高表达,且在HCT-8细胞中表达最高(P<0.05或P<0.01)。双荧光素酶报告基因证实,lncRNA XIST靶向负调控miR-32-5p(P<0.05),且EZH2是miR-32-5p的靶基因。敲降1ncRNA XIST抑制HCT-8细胞增殖和迁移并诱导其凋亡(P<0.05或P<0.01);进一步实验证明,敲降lncRNA XIST上调miR-32-5p的表达水平,从而下调EZH2表达水平,进而抑制HCT-8细胞增殖和迁移并诱导其凋亡。结论:lncRNA XIST通过miR-32-5p/EZH2分子轴促进HCT-8细胞增殖、迁移并抑制细胞凋亡。

Objective:To explore the mechanism of IncRNA XIST(XIST) regulating the biological behaviors of colorectal cancer HCT-8 cells via miR-32-5 p/EZH2(enhancer of Zeste homolog 2) axis.Methods:A total of 28 pairs of cancer tissues and corresponding para-cancerous tissues form colorectal cancer patients with complete clinical data were collected from the Colorectal and Anal Surgery,Xiangya Hospital of Central South University during July 2014 and August 2018.The expression levels of IncRNA XIST and miR-32-5 p in colorectal cancer tissues and cell lines were detected by qPCR.The targeted relationship between IncRNA XIST,miR-32-5 p and EZH2 was verified by dual luciferase reporter gene,and the expression level of EZH2 was further detected by WB.The proliferation,migration and apoptosis of HCT-8 cells were detected by CCK-8,Transwell and flow cytometry with Annexin V-FITC/PI staining,respectively.Results:lncRNA XIST was highly expressed in colorectal cancer tissues and cell lines with the highest expression in HCT-8 cells(P<0.05 or P<0.01).Dual luciferase reporter gene assay validated that IncRNA XIST negatively regulated miR-32-5 p(P<0.05),and EZH2 was a target gene of miR-32-5 p.Knockdown of IncRNA XIST inhibited proliferation and migration and induced apoptosis of HCT-8 cells(P<0.05 or P<0.01).Further experiments demonstrated that knockdown of IncRNA XIST up-regulated the expression of miR-32-5 p and further down-regulated the expression level of EZH2,thereby inhibiting the proliferation and migration of HCT-8 cells and inducing apoptosis.Conclusion:lncRNA XIST promotes proliferation,migration and inhibits apoptosis of HCT-8 cells via miR-32-5 p/EZH2 axis.