COUP-TFⅡ在胃癌组织中表达及对胃癌SGC7901细胞中VEGFR3-NRP2轴转录活性的调控  

Expression of COUP-TF Ⅱin gastric cancer tissues and its regulation of transcriptional activity of VEGFR3-NRP2 axis in gastric cancer SGC7901 cells

作　　者：李琼羽 王凯[1] 孙华波 王彩艳 许瑞雪 丛培佳 连海峰[1] LI Qiongyu;WANG Kai;SUN Huabo;WANG Caiyan;XU Ruixue;CONG Peijia;LIAN Haifeng(Department of Gastroenterology,Affiliated Hospital of Binzhou Medcal College,Binzhou 256603,Shandong,China;Department of Gastroenterology,Yantai Affiliated Hospital of Binzhou Medical College,Yantai 264100,Shandong,China)

机构地区：[1]滨州医学院附属医院消化内科,山东滨州256603 [2]滨州医学院烟台附属医院消化内科,山东烟台264100

出　　处：《中国肿瘤生物治疗杂志》2023年第3期230-236,共7页Chinese Journal of Cancer Biotherapy

基　　金：山东省自然科学基金项目(No.ZR2020MH060);2021年大学生创新创业训练计划立项项目(No.202110440120)。

摘　　要：目的:探讨COUP-TFⅡ在胃癌中对淋巴转移相关因子VEGFR3-NRP2轴的调控分子机制。方法:收集2015年3月至2015年8月在滨州医学院附属医院手术切除的60例胃癌组织和相应的癌旁组织及正常胃黏膜活检组织,用免疫组化和qPCR法检测COUP-TFⅡ、VEGFR3和NRP2的表达;培养胃癌细胞SGC7901和BGC823,构建过表达和siRNA-COUP-TFⅡ质粒后转染SGC7901细胞,用WB和qPCR检测转染后SGC7901细胞中COUP-TFⅡ、VEGFR3、NRP2的表达,用免疫共沉淀(CHIP)和双荧光素酶报告基因实验验证COUP-TFⅡ与VEGFR3-NRP2轴的靶向关系。结果:免疫组化检测显示,VEGFR3、NRP2、COUP-TFⅡ在胃癌组织中呈高表达(P<0.01);q PCR结果显示,与癌旁组织和正常组织相比,胃癌组织VEGFR3、NRP2和COUPTFⅡ的mRNA呈高表达(P<0.05或P<0.01);WB和qPCR法结果显示,与对照组相比,过表达COUP-TFⅡ组SGC7901细胞中COUP-TFⅡmRNA和蛋白水平表达均显著升高(均P<0.01);敲减组SGC7901细胞中COUP-TFⅡmRNA和蛋白水平均显著下降(P<0.05或P<0.01),且VEGFR3和NRP2 mRNA水平也均显著下降(均P<0.01);CHIP结果显示,SGC7901和BGC823细胞中COUP-TFⅡ抗体的免疫共沉淀物中含有VEGFR3和NRP2启动子DNA序列;双荧光素酶报告基因实验结果显示,COUP TFⅡ表达水平与VEGFR3和NRP2表达水平呈正相关。结论:COUP-TFⅡ在胃癌组织中高表达且对VEGFR3-NRP2轴存在正性调控作用,且在胃癌中高表达,COUP-TFⅡ可能为胃癌治疗的新靶点。Objective:To explore the molecular mechanism of COUP-TFⅡregulating lymphatic metastasis-related factor VEGFR3-NRP2 axis in gastric cancer.Methods:Sixty cases of gastric cancer tissues and paracancerous tissues surgically resected at the Binzhou Medical University Hospital between March 2015 and August 2015 were collected,and the expression of COUP-TFⅡ,VEGFR3 and NRP2 in the tissues was detected by immunohistochemistry and qPCR.Gastric cancer SGC7901 and BGC823 cells were cultured,and COUP-TFⅡoverexpression and siRNA-COUP-TFⅡplasmids were constructed and further transfected into SGC7901 cells.The expression of COUP-TFⅡ,VEGFR3 and NRP2 in SGC7901 cells after transfection was detected by WB and qPCR,and the targeting relationship between COUP-TFⅡand VEGFR3-NRP2 axis was verified by immunoprecipitation(CHIP)and dual-luciferase reporter assay.Results:Immunohistochemistry showed that VEGFR3,NRP2,and COUP-TFⅡwere highly expressed in gastric cancer tissues(all P<0.01).qPCR results showed that the mRNA levels of VEGFR3,NRP2 and COUP-TFⅡwere also highly expressed in gastric cancer tissues compared with paraneoplastic and normal tissues(P<0.05 or P<0.001).WB and qPCR methods showed that compared with the control group,both the mRNA and protein levels of COUP-TFⅡwere significantly higher in the COUP-TFⅡoverexpression group(both P<0.01);while the mRNA and protein levels of COUP-TFⅡwere significantly decreased in the SGC7901 cells of COUP-TFⅡknockdown group(P<0.05 or P<0.01),and the VEGFR3 and NRP2 mRNA levels were also significantly decreased(both P<0.01).CHIP results showed that the immunoprecipitates of COUP-TFⅡantibodies in SGC7901 and BGC823 cell lines contained VEGFR3 and NRP2 promoter DNA sequences.The results of dual-luciferase reporter assay showed that COUP-TFⅡexpression level was positively correlated with VEGFR3 and NRP2 levels.Conclusion:COUP-TFⅡhas a direct positive regulatory effect on the VEGFR3-NRP2 axis and is highly expressed in gastric cancer.COUP-TFⅡmay serve as a new target

关 键 词：胃癌 SCG7901细胞 BGC823细胞 鸡卵清蛋白上游启动子转录因子2 VEGFR3 神经纤毛蛋白2 淋巴转移 

分 类 号：R735.2[医药卫生—肿瘤] R730.2[医药卫生—临床医学]