Isolation of drugs which rescue insulin resistance by activating GLUT4 transport activity.

我们已经报道，慢性GH预处理抑制胰岛素诱导的葡萄糖摄取，而不影响胰岛素诱导的GLUT 4易位。本研究旨在确定调节GLUT 4转运活性的Akt底物和GLUT 4翻译后修饰。此外，为了挽救胰岛素抵抗，我们寻找调节GLUT 4转运活性的小分子化合物和抗体，首先，我们成功地分离了Akt底物AS 47，其胰岛素诱导的磷酸化被GH预处理抑制。敲低AS 47可抑制胰岛素诱导的葡萄糖摄取，但不影响GLUT 4的转运，表明AS 47在GLUT 4的转运活性中起重要作用。对GLUT 4的部分修饰位点进行定点突变，并将突变体转染HEK 293细胞。然后测量葡萄糖摄取。GLUT 4磷酸化位点突变（GLUT 4-P）显著增强葡萄糖转运活性，表明磷酸化损害GLUT 4转运活性。此外，我们已经成功地分离出了识别GLUT 4胞外区的抗体，本研究成功地鉴定了在GLUT 4转运活性中起重要作用的Akt底物和GLUT 4翻译后修饰。与Akt底物和GLUT 4相互作用的化学物质或抗体可能是治疗胰岛素抵抗的候选药物。

We have reported that chronic GH pretreatment inhibited insulin-induced glucose uptake without affecting insulin-induced GLUT4 translocation. This study was undertaken to identify Akt substrates and GLUT4 postranslational modification that regulate GLUT4 transport activity. In addition, in order to rescue insulin resistance we searched for the small molecular chemicals and antibodies that modulate GLUT4 transport activity.At first, we succeeded to isolate the Akt substrates, AS47, whose insulin-inducedphosphorylation was suppressed by GH pretreatment. Knockdown of AS47 inhibited insulin-induced glucose uptake without affecting GLUT4 translocation, suggesting that AS47 plays important roles in GLUT4 transport activity.Next, we searched for posttranslational modification motif in a GLUT4 molecule. GLUT4 was site-directed mutagenized in some modification sites and these mutants were transfected into HEK293 cell. And then glucose uptake was measured. Mutation at the phosphorylation site of GLUT4 (GLUT4-P) significantly enhanced glucose transport activity, suggesting that phosphorylation impaired GLUT4 transport activity. Moreover, we have already succeeded to isolate antibodies, which recognize GLUT4 extracellular domain.In this study we succeeded to identify the Akt substrate and GLUT4 posttranslational modification, which play important roles in GLUT4 transport activity. Chemicals or antibodies, which interact with the Akt substrate and GLUT4 could be candidates tocure insulin resistance.