Objective: To investigate the effect CD36 deficiency on muscle insulin signaling in mice fed a normal-fat diet and explore the possible mechanism.
Methods: Wild-type (WT) mice and systemic CD36 knockout (CD36-/-) mice with normal feeding for 14 weeks (n=12) were subjected to insulin tolerance test (ITT) after intraperitoneal injection with insulin (1 U/kg). Real-time PCR was used to detect the mRNA expressions of insulin receptor (IR), insulin receptor substrate 1/2 (IRS1/2) and protein tyrosine phosphatase 1B (PTP1B), and Western blotting was performed to detect the protein expressions of AKT, IR, IRS1/2 and PTP1B in the muscle tissues of the mice. Tyrosine phosphorylation of IR and IRS1 and histone acetylation of PTP1B promoter in muscle tissues were detected using co-immunoprecipitation (Co-IP) and chromatin immunoprecipitation (ChIP), respectively.
Results: CD36-/- mice showed significantly lowered insulin sensitivity with obviously decreased area under the insulin tolerance curve in comparison with the WT mice (P < 0.05). CD36-/- mice also had significantly higher serum insulin concentration and HOMA-IR than WT mice (P < 0.05). Western blotting showed that the p-AKT/AKT ratio in the muscle tissues was significantly decreased in CD36-/- mice as compared with the WT mice (P < 0.01). No significant differences were found in mRNA and protein levels of IR, IRS1 and IRS2 in the muscle tissues between WT and CD36-/- mice (P>0.05). In the muscle tissue of CD36-/- mice, tyrosine phosphorylation levels of IR and IRS1 were significantly decreased (P < 0.05), and the mRNA and protein levels of PTP1B (P < 0.05) and histone acetylation level of PTP1B promoters (P < 0.01) were significantly increased as compared with those in the WT mice. Intraperitoneal injection of claramine, a PTP1B inhibitor, effectively improved the impairment of insulin sensitivity in CD36-/- mice.
Conclusion: CD36 is essential for maintaining muscle insulin sensitivity under physiological conditions, and CD36 gene deletion in mice causes impaired insulin sensitivity by up-regulating muscle PTP1B expression, which results in detyrosine phosphorylation of IR and IRS1.
目的: 探讨在正常饮食状态下，CD36基因缺失对小鼠肌肉胰岛素信号通路的影响及作用机制。
方法: 野生型小鼠（WT）和CD36基因敲除小鼠（CD36-/-）给予正常饮食喂养14周（n=12）。小鼠禁食4 h，腹腔注射胰岛素（1 U/kg）进行胰岛素耐量实验。Real-time PCR检测小鼠肌肉胰岛素受体(IR)、胰岛素受体底物1/2（IRS1/2）、蛋白酪氨酸磷酸酶1B（PTP1B）基因表达。Western blot检测小鼠肌肉蛋白激酶B(AKT)、IR、IRS1/2和PTP1B的蛋白表达。免疫共沉淀（Co-IP）检测肌肉IR和IRS1的酪氨酸磷酸化程度。染色质免疫共沉淀（ChIP）技术检测肌肉PTP1B启动子组蛋白乙酰化水平。
结果: 在正常饮食状态下，与WT小鼠相比，CD36-/-小鼠的胰岛素耐量显著增强（P < 0.05），血清胰岛素浓度升高（P < 0.01），胰岛素抵抗指数(HOMA-IR)升高（P < 0.05）。在肌肉组织中，CD36-/-小鼠与WT小鼠相比，p-AKT/AKT蛋白表达比值显著降低（P < 0.01）。Real-time PCR和Western blot结果表明，肌肉组织IR，IRS1，IRS2的mRNA和蛋白水平在WT和CD36-/-小鼠间无显著差异（P>0.05）。Co-IP实验显示IR和IRS1的酪氨酸磷酸化水平在CD36-/-小鼠中显著降低（P < 0.05）。CD36-/-小鼠肌肉中PTP1B mRNA和蛋白表达均高于WT小鼠（P < 0.05），ChIP实验显示PTP1B基因启动子的组蛋白乙酰化水平显著升高（P < 0.01）。腹腔注射PTP1B的抑制剂可改善CD36-/-小鼠的胰岛素敏感性。
结论: 在正常饮食条件下，CD36基因对于维持生理肌肉胰岛素敏感性十分重要，小鼠CD36基因缺失通过上调PTP1B基因表达，诱导IR、IRS1去酪氨酸磷酸化而使肌肉胰岛素信号传导受损。
Keywords: CD36; PTP1B; insulin sensitivity; muscle.