【摘要】 目的探索缺氧对B-淋巴细胞瘤蛋白9(BCL9)的调控关系及BCL9对结肠癌细胞的生物学作用。方法收取20例结肠癌标本及同一患者的正常结肠标本,利用免疫组化技术、实时荧光定量PCR技术检测BCL9的表达;利用常氧或缺氧培养HCT116细胞,实时荧光定量PCR及蛋白印迹法(western-blot)检测BCL9的表达;构建包含BCL9启动子序列的荧光素酶报告载体,通过缺氧处理或过表达HIF1α,检测荧光素酶活性变化;通过过表达质粒或siRNA干扰BCL9的表达,随后检测HCT116细胞的增殖活性及迁移能力。结果免疫组化及实时荧光定量PCR检测显示肿瘤组织中BCL9表达明显高于正常组织[(3.25±0.53)vs.(1.03±0.12),P <0.05];缺氧培养HCT116细胞后BCL9的表达显著提高,[(6.71±0.83)vs.(1.54±0.21),P <0.05];同时,缺氧培养或过表达HIF1α能显著提高荧光素酶的活性[(3.53±0.75)vs.(0.96±0.15),(4.83±0.62)vs.(1.02±0.14);P <0.05];干扰BCL9的表达抑制HCT116细胞的增殖及迁移能力[(1.23±0.12)vs.(1.87±0.15),P <0.05];提高BCL9表达能促进HCT116细胞的增殖及迁移能力[(2.43±0.16)vs.(1.81±0.14),P <0.05]。结论缺氧可诱导BCL9在结肠癌中高表达,并通过BCL9促进结肠癌细胞增殖、迁移。更多还原

【Abstract】 Objective The purpose of the paper was to explore the regulation relationship between hypoxia and BCL9 in colorectal cancer cells and whether the BCL9 can in?uence the function of colorectal cancer cells. Methods Detected BCL9 in 20 cases of colorectal cancer and the normal colorectal tissue by immunohistochemistry and RT-qPCR; Assay the BCL9 expression in colorectal cancer cells under hypoxia condition or normoxia condition by RT-PCR and western-blot; Constructed luciferase plasmid which contain of the promoter of BCL9, by given the hypoxia condition to induce endogenous HIF-1α or use over-expression plasmid induced exogenous HIF-1α in HCT116 cells to explored the regulate relationship between HIF-1α and LDHA; Middle the expression of BCL9 by over-expression plasmid and siRNA, and detected the proliferation and migration by CCK8 assay and transwell assay. Results BCL9 is always overexpression in colorectal cancer [(3.25±0.53) vs.(1.03±0.12), P < 0.05]; By given oxygen condition, we found that hypoxia can induce BCL9 over-expression in colorectal cancer cells [(6.71±0.83) vs.(1.54±0.21),P < 0.05]; Both hypoxia and over-expression exogenous HIF-1α leaded luciferase activity enhanced;Using RNA interfere technique to knockdown the expression of HIF-1α will suppression LDHA expression[(3.53±0.75)vs.(0.96±0.15),(4.83±0.62) vs.(1.02±0.14); P < 0.05]; Interfere the expression of BCL9 suppresion HCT116 cells proliferation and migration [(1.23±0.12) vs.(1.87±0.15), P < 0.05]; On the other hand, over-expression BCL9 leaded to the opposite result [(2.43±0.16) vs.(1.81±0.14), P < 0.05];Conclusion Hypoxia induce BCL9 over-expression and promote HCT116 cells proliferation and migration.更多还原