Molecular mechanisms of insulin signal transduction and diabetes mellitus

已发现几种细胞因子和生长因子的受体的胞外域在血浆中循环。人血清中可溶性IR α的存在受到怀疑，而分泌可溶性IRα的转基因小鼠表现出慢性高血糖。我们建立了一种新的酶联免疫吸附试验（ELISA）系统，用于检测IR的胞外域（α-亚基和部分β-亚基）和全长IR。利用这些ELISA系统，我们发现人血浆中存在可溶性hIR α和hIRβ的部分胞外区，而不是完整的IR或完整的hIR β，并且在血糖升高的患者中其血浆水平升高。IR的胞外结构域可能被高血糖状态相关的机制切割，至少部分是这样。我们和其他人先前报道了Gaq蛋白偶联受体（GaqPCR）的激活也刺激了几种细胞系中GLUT 4的易位和葡萄糖摄取。在此，我们报道了GaqPCR的激活也促进了5 '-AMP激活的蛋白激酶（AMPK）对160 KDa的Akt底物（Rab GT3激活蛋白（GAP））AS 160的磷酸化。通过使用siRNA敲低AMPKα1亚基抑制AS 160磷酸化促进GLUT 4囊泡保留在细胞内区室，但不影响Gaq刺激的GLUT 4易位的倍数增加。这些结果表明，AS 160是胰岛素和GaqPCR激活刺激的GLUT 4囊泡运输的共同介质。AS 160调节GLUT 4囊泡的细胞内分布及其在细胞内的滞留。

The ectodomains of receptors for several cytokines and growth factors have been found to circulate in plasma. Thus, the existence of soluble IR in human serum has been suspected.Furthermore, transgenic mice secreting soluble IRα into the plasma showed chronic hyperglycemia. Here, we established novel enzyme-linked immunosorbent assay (ELISA) systems to measure both the ectodomain (α-subunit and a part of β-subunit) of IR and full length of IR.With these ELISA systems, we report that soluble hlRα with parts of extracellular region of hIRβ, but not as a whole IR or with intact hlRβ, is present in human plasma and that its plasma level is elevated in patients with elevated blood glucose. The ectodomain of IR may be cleaved, at least in part, by hyperglycemic state-associated mechanisms.We and others previously reported that the activation of Gaq protein- coupled receptors (GaqPCRs) also stimulates GLUT4 translocation and glucose uptake in several cell lines. Here, we report that the activation of GaqPCRs also promoted the phosphorylation of AS160, Akt substrate of 160 KDa which is Rab GTPase activating protein (GAP), by 5'-AMP activated protein kinase (AMPK). The suppression of AS160 phosphorylation by AMPKα1 subunit knockdown using siRNA promoted GLUT4 vesicle retention at intracellular compartments, but did not affect a fold increase of the Gaq- stimulated GLUT4 translocation. These results suggest that AS160 is a common mediator of the GLUT4 vesicle trafficking stimulated by insulin and by the activation of GaqPCR. The AS160 regulates the intracellular distribution of the GLUT4 vesicles and its retention inside the cells.