具核梭杆菌通过调控miR-181b在结肠癌细胞中形成炎性微环境的机制  

Mechanism of Fusobacterium nucleatum via Regulating miR-181b for Forming An Inflammatory Microenvironment in Colon Cancer Cells

作　　者：林欣[1] 何杰[1] 魏芳[1] 赵冲[1] 曾利红[1] 吴琼[1] 黄惠康[1] 杜艳蕾[1] 王红[1] 

机构地区：[1]广州医科大学附属广州市第一人民医院消化内科广州消化疾病中心,510180

出　　处：《胃肠病学》2017年第10期592-598,共7页Chinese Journal of Gastroenterology

摘　　要：背景:具核梭杆菌(Fn)是口腔常见的致病菌,研究发现Fn与结直肠癌的发生、发展密切相关,尤其是炎症相关性结直肠癌。目的:探讨Fn感染在结肠癌细胞中形成炎性微环境的机制。方法:构建Fn感染Caco-2细胞的炎症模型,行miRNA测序。将miR-181b mimics或inhibitor转染Fn感染的Caco-2细胞。以qRT-PCR和蛋白质印迹法分别检测TNF-αmRNA和蛋白表达,ELISA法检测上清液中TNF-α含量,Transwell小室法检测穿膜淋巴细胞数。结果:Fn组TNF-αmRNA和蛋白表达均显著高于对照组(P<0.05),上清液中TNF-α含量显著升高(P<0.05),穿膜淋巴细胞数量明显增多(P<0.05)。miRNA测序和qRT-PCR结果均显示,Fn组miR-181b表达较对照组显著降低(P<0.05)。与对照组相比,miR-181b mimics+Fn组TNF-αmRNA和蛋白表达显著降低(P<0.05);而miR-181b inhibitor组TNF-αmRNA和蛋白表达均显著升高(P<0.05)。生物信息学工具和双荧光素酶检测证实TNF-α可能为Caco-2细胞中miR-181b的靶基因。结论:Fn通过抑制Caco-2细胞中miR-181b表达而上调TNF-α表达,募集淋巴细胞形成炎性微环境。Background： Fusobacterium nucleatum（ Fn） is a common oral pathogen. Studies have shown that Fn is closely related to the occurrence and development of colorectal cancer,especially the inflammation-related colorectal cancer.Aims： To investigate the mechanism of Fn in forming an inflammatory microenvironment in colon cancer cells. Methods：An inflammation model of Caco-2 cells infected by Fn was constructed,and miRNA sequencing was performed. miR-181 b mimics or inhibitor was transfected into Fn infected Caco-2 cells. mRNA and protein expressions of TNF-α were determined by qRT-PCR and Western blotting,respectively,and concentration of TNF-α in supernatant was measured by ELISA,number of lymphocyte penetrating the membrane was measured by Transwell chamber. Results： Compared with control group,mRNA and protein expressions of TNF-α were significantly increased（ P〈0. 05）,concentration of TNF-α in supernatant was significantly increased（ P〈0. 05）,and number of lymphocyte penetrating the membrane was significantly increased in Fn group（ P〈0. 05）. miRNA sequencing and qRT-PCR results showed that expression of miR-181 b was significantly decreased in Fn group than in control group（ P〈0. 05）. Compared with control group,mRNA and protein expressions of TNF-α were significantly decreased in miR-181 b mimics ＋ Fn group（ P〈0. 05）,however,mRNA and protein expressions of TNF-α were significantly increased in miR-181 b inhibitor group（ P〈0. 05）. Bioinformatics tools and Luciferase assay confirmed that TNF-α might be the target gene of miR-181 b in Caco-2 cells. Conclusions： Fn can upregulate the expression of TNF-α by inhibiting miR-181 b in Caco-2 cells and recruiting lymphocytes to form an inflammatory microenvironment.

关 键 词：具核梭杆菌 miR-181b 肿瘤坏死因子Α 结直肠肿瘤 

分 类 号：R735.3[医药卫生—肿瘤]