SOX21在胃癌中的表达和甲基化状态及对体外胃癌细胞的作用

The expression and methylation status of SOX21 in gastric cancer and its effect on gastric cancer cells in vitro

目的探讨性别决定区Y-BOX21(SOX21)在胃癌(GC)中的表达和甲基化状态及对体外胃癌细胞的作用。方法收集2016年4月~2017年5月于东莞市人民医院行胃癌根治术患者的癌组织及对应的癌旁组织各44例,以胃癌细胞株AGS和人胃粘膜上皮细胞株GES-1为进一步研究对象,将培养后的细胞分为空白对照组、NC组和SOX21组,甲基化特异性PCR检测SOX21甲基化状态;qRT-PCR、免疫组化(IHC)和Western blot检测SOX21和Wnt/β-catenin通路相关基因表达;CCK-8试剂盒和克隆形成实验检测细胞增殖能力;流式细胞术检测细胞凋亡;Transwell检测细胞迁移和侵袭能力。结果与癌旁组织相比,GC组织SOX21甲基化率明显升高(P<0.001),SOX21 IHC评分、mRNA表达和蛋白表达均明显降低(P<0.05)。SOX21在GES-1细胞中呈去甲基化状态,而在AGS中呈完全甲基化状态。与GES-1细胞相比,AGS细胞中SOX21 mRNA和蛋白表达均显著降低(P<0.05)。与NC组相比,SOX21组中SOX21 mRNA表达、蛋白表达和细胞凋亡率明显升高(P<0.05),细胞活力、克隆形成数、迁移和侵袭细胞数明显降低(P<0.05),β-catenin、c-myc、cyclinD1和MMP7蛋白表达明显降低(P<0.05)。结论SOX21在胃癌组织中呈高甲基化状态且表达降低,过表达SOX21可能通过抑制Wnt/β-catenin通路抑制胃癌细胞增殖、迁移和侵袭,促进细胞凋亡。

Objective To explore the effect of SOX21 on gastric cancer(GC)and its possible mechanism.Methods A total of 44 cases of GC tissue and corresponding adjacent tissues were collected.Gastric cancer cell line AGS and human gastric mucosal epithelial cell line GES-1 were used as further research objects.Methylation-specific PCR was used to detect SOX21 methylation status.qRT-PCR,immunohistochemistry(IHC)and Western blot were used to detect the expression of SOX21 and Wnt/β-catenin pathway related gene.CCK-8 kit and clone formation experiments were used to detect cell proliferation ability.The flow cytometry was used to detect cell apoptosis.Transwell was used to detect cell migration and invasion ability.Results Compared with the adjacent tissues,the methylation rate of SOX21 in GC tissue was significantly increased(P<0.001),and the SOX21 IHC score,mRNA expression and protein expression were significantly reduced(P<0.05).SOX21 was demethylated in GES-1 cells,but fully methylated in AGS.Compared with the GES-1 group,the SOX21 mRNA and protein expressions in the AGS group were significantly reduced(P<0.05).Compared with the NC group,SOX21 mRNA expression,protein expression and apoptosis rate were significantly increased in SOX21 group(P<0.05),cell viability,number of clone formation,number of migration and invasion cells were significantly reduced(P<0.05),β-catenin,c-myc,cyclinD1 and MMP7 protein expression was significantly reduced(P<0.05).Conclusion SOX21 is hypermethylated and its expression is reduced in gastric cancer tissues.Overexpression of SOX21 may inhibit the proliferation,migration and invasion of gastric cancer cells and promote cell apoptosis by inhibiting the Wnt/β-catenin pathway.