lncRNA CASC9通过调控miR-424-5p/SOX9分子轴对食管鳞状细胞癌细胞恶性生物学行为的影响

Effect of lncRNA CASC9 on Malignant Biological Behavior of Esophageal Squamous Cell Carcinoma Cells by Regulating miR-424-5p/SOX9 Axis

目的探讨长链非编码RNA(lncRNA)癌症易感性候选基因9(CASC9)通过调控miR-424-5p/SOX9分子轴对食管鳞状细胞癌(ESCC)细胞恶性生物学行为的影响。方法采用RT-qPCR检测CASC9在食管鳞状细胞癌组织、配对癌旁组织、食管鳞状细胞癌细胞系(KYSE450、KYSE150、EC190、EC9706)及正常人食管鳞状上皮细胞株(Het-1A)中的表达;敲降CASC9在KYSE450细胞中的表达水平,检测下调CASC9对细胞增殖、侵袭及其细胞周期的影响;采用Western blotting法检测SOX9蛋白及周期蛋白2(CCND1、CCNA、CDK4、CDK6)的表达;通过CCK-8和Transwell方法检测KYSE450细胞的增殖和侵袭能力,通过双荧光素酶报告基因法验证miR-424-5p与CASC9或SOX9的靶向关系。结果食管鳞状细胞癌组织与细胞系高表达CASC9(P<0.01),敲降CASC9可抑制KYSE450细胞增殖和侵袭能力、并阻滞细胞周期G1/S期(P<0.01)。双荧光素酶报告基因法证实,CASC9靶向miR-424-5p、并下调其表达,SOX9是miR-424-5p的靶基因;敲降CASC9可上调miR-424-5p、并抑制SOX9的表达(均P<0.01)。结论lncRNA CASC9可通过下调miR-424-5p,并促进SOX9表达而刺激KYSE450细胞的增殖、侵袭。

Objective To investigate the effect of lncRNA cancer susceptibility candidate 9(CASC9)on malignant biological behavior of esophageal squamous cell carcinoma cells by regulating miR-424-5p/SOX9 axis.Methods The expression of CASC9 was detected by RT-qPCR in esophageal squamous cell carcinoma,paracancerous tissues,esophageal squamous cell carcinoma cell lines(KYSE450,KYSE150,EC190 and EC9706),and normal esophageal squamous epithelial cell line Het-1A.The effects of CASC9 down-regulation on cell proliferation,invasion and cell cycle were measured in KYSE450 cells.The expression of SOX9 protein and cyclins(CCND1,CCNA,CDK4 and CDK6)was determined by Western blotting.The proliferation and invasion of KYSE450 cells were detected by CCK-8 and Transwell assay.The targeted relationship between miR-424-5p and CASC9 or SOX9 was verified by dual luciferase reporter gene assay.ResultsCASC9 was highly expressed in esophageal squamous cell carcinoma tissue and cell lines(P<0.01).Knockdown of CASC9 inhibited the proliferation and invasion of KYSE450 cells,and blocked the G1/S phase(P<0.01).Dual-luciferase reporter gene confirmed that CASC9 targeted miR-424-5p and down-regulated its expression,and that SOX9 was the target gene of miR-424-5p.Knockdown of CASC9 up-regulated the expression of miR-424-5p and inhibited the expression of SOX9(P<0.01).Conclusion LncRNA CASC9 can stimulate the proliferation and invasion of KYSE450 cells by down-regulating miR-424-5p expression and promoting SOX9 expression.