Introduction/Aim: Circadian rhythm disruption is emerging as a risk factor for metabolic disorders and particularly, alterations in clock genes circadian expression have been shown to influence insulin sensitivity. Recently, the reciprocal interplay between the circadian clock machinery and HPA axis has been largely demonstrated: the circadian clock may control the physiological circadian endogenous glucocorticoids secretion and action; glucocorticoids, in turn, are potent regulator of the circadian clock and their inappropriate replacement has been associated with metabolic impairment. The aim of the current study was to investigate in vitro the interaction between the timing-of-the-day exposure to different hydrocortisone (HC) concentrations on muscle insulin sensitivity. Methods: Serum-shock synchronized mouse skeletal muscle C2C12 cells were exposed to different HC concentrations recapitulating the circulating daily physiological cortisol profile (standard cortisol profile), the circulating daily cortisol profile that reached in adrenal insufficient (AI) patients treated with once-daily MR-HC (flat cortisol profile) and treated with thrice-daily of conventional IR-HC (steep cortisol profile). The 24 hrs spontaneous oscillation of the clock genes in synchronized C2C12 cells was used to align the timing for in vitro HC exposure (Bmal1 acrophase, midphase and bathyphase) with the reference times of cortisol peaks in AI treated with IR-HC (8 am, 1 pm, 6 pm). A panel of 84 insulin sensitivity related genes and intracellular insulin signaling proteins were analyzed by RT-qPCR and western blot, respectively. Results: Only the steep profile, characterized by a higher HC exposure during Bmal1 bathyphase, produced significant downregulation in 21 insulin sensitivity-related genes. Among these, Insr, Irs1, Irs2, Pi3kca and Adipor2 were downregulated when compared the flat to the standard or steep profile. Reduced intracellular IRS1 Tyr608, AKT Ser473, AMPK Thr172 and ACC Ser79 phosphorylations were also observed. Conclusions: The current study demonstrated that is late-in-the-day cortisol exposure that modulates insulin sensitivity-related genes expression and intracellular insulin signaling in skeletal muscle cells.


中文翻译：

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肌肉中的皮质醇昼夜节律和胰岛素抵抗：氢化可的松暴露的剂量和时间对同步肌肉细胞中胰岛素敏感性的影响。

简介/目标：昼夜节律紊乱正在成为代谢紊乱的一个危险因素，特别是，时钟基因昼夜节律表达的改变已被证明会影响胰岛素敏感性。最近，生物钟机制和 HPA 轴之间的相互作用已得到广泛证实：生物钟可能控制生理性昼夜节律内源性糖皮质激素的分泌和作用；反过来，糖皮质激素是生物钟的有效调节剂，它们的不当替代与代谢障碍有关。本研究的目的是在体外研究不同时间的氢化可的松 (HC) 浓度对肌肉胰岛素敏感性的影响。方法：血清休克同步小鼠骨骼肌 C2C12 细胞暴露于不同的 HC 浓度，概括了每日循环生理皮质醇曲线（标准皮质醇曲线），在接受每日一次 MR-HC 治疗的肾上腺功能不全 (AI) 患者中达到的每日循环皮质醇曲线（平坦的皮质醇曲线）并用每天三次的常规 IR-HC（陡峭的皮质醇曲线）治疗。同步 C2C12 细胞中时钟基因的 24 小时自发振荡用于将体外 HC 暴露的时间（Bmal1 顶相、中期和深海相）与用 IR-HC 处理的 AI 中皮质醇峰值的参考时间（上午 8 点，下午 1 点、下午 6 点）。分别通过 RT-qPCR 和蛋白质印迹分析了一组 84 个胰岛素敏感性相关基因和细胞内胰岛素信号蛋白。结果：只有在 Bmal1 深水期以较高的 HC 暴露为特征的陡峭曲线在 21 个胰岛素敏感性相关基因中产生了显着下调。其中，Insr、Irs1、Irs2、Pi3kca 和 Adipor2 在与标准或陡峭剖面相比时被下调。还观察到细胞内 IRS1 Tyr608、AKT Ser473、AMPK Thr172 和 ACC Ser79 磷酸化减少。结论：目前的研究表明，当天晚些时候的皮质醇暴露可调节骨骼肌细胞中胰岛素敏感性相关基因的表达和细胞内胰岛素信号传导。Pi3kca 和 Adipor2 在与标准或陡峭轮廓相比时被下调。还观察到细胞内 IRS1 Tyr608、AKT Ser473、AMPK Thr172 和 ACC Ser79 磷酸化减少。结论：目前的研究表明，当天晚些时候的皮质醇暴露可调节骨骼肌细胞中胰岛素敏感性相关基因的表达和细胞内胰岛素信号传导。Pi3kca 和 Adipor2 在与标准或陡峭轮廓相比时被下调。还观察到细胞内 IRS1 Tyr608、AKT Ser473、AMPK Thr172 和 ACC Ser79 磷酸化减少。结论：目前的研究表明，当天晚些时候的皮质醇暴露可调节骨骼肌细胞中胰岛素敏感性相关基因的表达和细胞内胰岛素信号传导。