自噬相关蛋白ATG16L1在肠上皮细胞炎症因子表达中的作用

Effect of Autophagy-related Protein ATG16L1 on Expression of Inflammatory Cytokines in Intestinal Epithelial Cells

背景:自噬相关基因ATG16L1单核苷酸多态性(SNP)与克罗恩病(CD)发病风险密切相关。目的:探讨ATG16L1在肠上皮细胞炎症因子表达中的作用。方法:利用人结肠腺癌细胞株Caco-2构建体外肠上皮细胞模型,予ATG16L1 siRNA干预,并以白细胞介素-1β(IL-1β)诱导炎症因子表达。采用real-time PCR和蛋白质印迹法验证ATG16L1 siRNA转染的有效性,real-time PCR和ELISA法检测Caco-2细胞IL-6、IL-8和单核细胞趋化蛋白-1(MCP-1)表达。结果:研究使用的siRNA能有效抑制ATG16L1表达。IL-1β可诱导Caco-2细胞IL-6、IL-8、MCP-1 mRNA和蛋白表达上调(P<0.05),沉默ATG16L1可使IL-1β诱导的上述炎症因子表达进一步增高(P<0.05)。结论:在肠上皮细胞中,ATG16L1可负性调控IL-1β诱导的炎症因子表达,在维持肠黏膜免疫系统稳态中发挥重要作用。

Background： Single nucleotide polymorphism（ SNP） in autophagy-related 16-like 1 gene（ ATG16 L1） has been shown to be related to the increased risk of Crohn＇s disease（ CD）. Aims： To investigate the effect of ATG16 L1 on expression of inflammatory cytokines in intestinal epithelial cells（ IECs）. Methods： In vitro intestinal epithelial model was established by using human colon adenocarcinoma cell line Caco-2. ATG16 L1 in Caco-2 cells was inhibited by targeted siRNA transfection,and expression of inflammatory cytokines was induced by interleukin-1β（ IL-1β）. The efficiency of transfection was confirmed by real-time PCR and Western blotting,while real-time PCR and ELISA were used to detect the expressions of IL-6,IL-8,and monocyte chemoattractant protein-1（ MCP-1） in Caco-2 cells. Results： ATG16 L1 could be effectively down-regulated by RNA interference. Expressions of IL-6,IL-8 and MCP-1 were significantly increased at the mRNA and protein level in Caco-2 cells treated with IL-1β（ P〈0. 05）. Knockdown of ATG16 L1 further increased the expressions of these inflammatory cytokines induced by IL-1β（ P〈0. 05）. Conclusions： ATG16 L1 negatively regulates the expression of inflammatory cytokines induced by IL-1β in IECs, which plays an important role in maintaining homeostasis of the gut mucosal immune system.