免疫受体NOD1在人脂肪细胞胰岛素抵抗形成中的作用

摘要

目的 研究免疫受体核苷酸结合寡聚化结构域蛋白1(NOD1)在人脂肪细胞胰岛素抵抗形成中的作用,并探讨其可能的作用机制.方法 将人前脂肪细胞诱导分化为成熟的脂肪细胞,加入NOD1受体激动剂iE-DAP,利用双萤光素酶报告系统检测NF-κB转录活性,酶联免疫吸附法检测炎症细胞因子水平,2-脱氧-[3H]-D-葡萄糖摄入法观察脂肪细胞的葡萄糖摄取率,蛋白免疫印迹检测胰岛素受体底物1(IRS-1)、蛋白激酶B(Akt)磷酸化及磷酸肌醇3激酶(PI-3K)p85的蛋白表达水平.结果 iE-DAP促进人脂肪细胞NF-κB转录活性(P＜0.05),刺激炎性细胞因子白细胞介素(IL)-6、IL-8及单核细胞趋化因子1的分泌(均P＜0.01).iE-DAP显著抑制胰岛素刺激状态下人脂肪细胞的葡萄糖转运(P＜0.05),呈时间剂量效应.iE-DAP处理脂肪细胞,IRS-1 307位丝氨酸磷酸化水平明显升高,PI-3K p85蛋白表达显著降低,并抑制胰岛素诱导的Akt 473位丝氨酸和308位苏氨酸磷酸化水平(均P＜0.05).结论 NOD1介导的炎症反应参与人脂肪细胞胰岛素抵抗的发生,其机制可能与其抑制IRS-1、PI3-K、Akt信号通路有关.%Objective To investigate the role of nucleotide-binding oligomerization domain-containing protein 1 (NOD1) in inducing insulin resistance in differentiated human adipocytes.Methods Human preadipocytes obtained via liposuction were induced to differentiate into mature adipocytes.iE-DAP,a specific ligand for NOD1,was administered to human adipocytes in culture.NF-κB transcriptional activity and proinflammatory cytokines production were determined by luciferase assay and enzyme-linked immunosorbent assay.Glucose uptake in adipocytes was measured by 2-deoxy-D-[3 H] glucose uptake (P＜0.05).The expression of phosphatidylinositol-3-kinase p85 (PI-3K p85),insulin receptor substrate-1 (IRS-1) and Akt phosphorylations were detected by Western blotting.Results NF-κB transcriptional activity and cytokines such as interleukin (IL)-6,IL-8,and monocyte chemotactic protein 1 secretion were markedly increased after stimulation with iE-DAP (P＜0.05 or P＜0.01).Insulin-induced glucose uptake was decreased with the activation of NOD1 in a dose-and time-dependent fashion.NOD1 activation weakened insulin signal transduction as being revealed by increasing IRS-1 Ser307phosphorylation,reducing protein expression of PI-3K p85,and attenuating insulin-induced phosphorylation of Akt on Ser473 and Thr308in human adipocytes.Conclusion These results indicate that NOD1 activation induces inflammatory response and insulin resistance via IRS-1/PI3-K/Akt signaling pathway in human adipocytes.