Dipartimento di Biologia e Patologia Cellulare e Molecolare, Università di Napoli Federico II, Via Sergio Pansini, 5, Naples 80131, Italy.
Diabetologia. 2010 May;53(5):955-65. doi: 10.1007/s00125-010-1676-1. Epub 2010 Feb 18.
AIMS/HYPOTHESIS: Glucosamine, generated during hyperglycaemia, causes insulin resistance in different cells. Here we sought to evaluate the possible role of endoplasmic reticulum (ER) stress in the induction of insulin resistance by glucosamine in skeletal muscle cells.
Real-time RT-PCR analysis, 2-deoxy-D: -glucose (2-DG) uptake and western blot analysis were carried out in rat and human muscle cell lines.
In both rat and human myotubes, glucosamine treatment caused a significant increase in the expression of the ER stress markers immunoglobulin heavy chain-binding protein/glucose-regulated protein 78 kDa (BIP/GRP78 [also known as HSPA5]), X-box binding protein-1 (XBP1) and activating transcription factor 6 (ATF6). In addition, glucosamine impaired insulin-stimulated 2-DG uptake in both rat and human myotubes. Interestingly, pretreatment of both rat and human myotubes with the chemical chaperones 4-phenylbutyric acid (PBA) or tauroursodeoxycholic acid (TUDCA), completely prevented the effect of glucosamine on both ER stress induction and insulin-induced glucose uptake. In both rat and human myotubes, glucosamine treatment reduced mRNA and protein levels of the gene encoding GLUT4 and mRNA levels of the main regulators of the gene encoding GLUT4 (myocyte enhancer factor 2 a [MEF2A] and peroxisome proliferator-activated receptor-gamma coactivator 1alpha [PGC1alpha]). Again, PBA or TUDCA pretreatment prevented glucosamine-induced inhibition of GLUT4 (also known as SLC2A4), MEF2A and PGC1alpha (also known as PPARGC1A). Finally, we showed that overproduction of ATF6 is sufficient to inhibit the expression of genes GLUT4, MEF2A and PGC1alpha and that ATF6 silencing with a specific small interfering RNA is sufficient to completely prevent glucosamine-induced inhibition of GLUT4, MEF2A and PGC1alpha in skeletal muscle cells.
CONCLUSIONS/INTERPRETATION: In this work we show that glucosamine-induced ER stress causes insulin resistance in both human and rat myotubes and impairs GLUT4 production and insulin-induced glucose uptake via an ATF6-dependent decrease of the GLUT4 regulators MEF2A and PGC1alpha.
目的/假设：在高血糖期间产生的葡萄糖胺会导致不同细胞的胰岛素抵抗。在这里，我们试图评估内质网（ER）应激在葡萄糖胺诱导骨骼肌细胞胰岛素抵抗中的可能作用。
在大鼠和人肌肉细胞系中进行实时 RT-PCR 分析、2-脱氧-D：-葡萄糖（2-DG）摄取和 Western blot 分析。
在大鼠和人肌管中，葡萄糖胺处理导致内质网应激标志物免疫球蛋白重链结合蛋白/葡萄糖调节蛋白 78kDa（BIP/GRP78[也称为 HSPA5]）、X 框结合蛋白-1（XBP1）和激活转录因子 6（ATF6）的表达显著增加。此外，葡萄糖胺损害了大鼠和人肌管中胰岛素刺激的 2-DG 摄取。有趣的是，用化学伴侣 4-苯丁酸（PBA）或牛磺熊脱氧胆酸（TUDCA）预处理大鼠和人肌管，完全阻止了葡萄糖胺对 ER 应激诱导和胰岛素诱导的葡萄糖摄取的影响。在大鼠和人肌管中，葡萄糖胺处理降低了编码 GLUT4 的基因的 mRNA 和蛋白水平，以及编码 GLUT4 的主要调节因子的基因（肌细胞增强因子 2a[MEF2A]和过氧化物酶体增殖物激活受体-γ共激活因子 1α[PGC1α]）的 mRNA 水平。同样，PBA 或 TUDCA 预处理可防止葡萄糖胺诱导的 GLUT4（也称为 SLC2A4）、MEF2A 和 PGC1α（也称为 PPARGC1A）抑制。最后，我们表明 ATF6 的过度产生足以抑制 GLUT4、MEF2A 和 PGC1α 的基因表达，并且用特异性小干扰 RNA 沉默 ATF6 足以完全防止葡萄糖胺诱导的骨骼肌细胞中 GLUT4、MEF2A 和 PGC1α 的抑制。
结论/解释：在这项工作中，我们表明葡萄糖胺诱导的 ER 应激导致人和大鼠肌管中的胰岛素抵抗，并通过 ATF6 依赖性降低 GLUT4 调节因子 MEF2A 和 PGC1α 来损害 GLUT4 的产生和胰岛素诱导的葡萄糖摄取。
