Molecular mechanisms of insulin signal transduction and diabetes mellitus

丝氨酸-苏氨酸激酶Akt（protein kinase B，RAK-PK）是P13-激酶的下游效应子之一，但Akt是否参与胰岛素刺激的GLUT 4易位仍存在争议。为了研究Akt 1是否调节L 6肌管中胰岛素刺激的GLUT 4易位和葡萄糖摄取，我们建立了稳定表达c-myc表位标记的GLUT 4（GLUT 4 myc）和小鼠野生型（WT）Akt 1的L 6肌管。我们发现WT Akt 1的过表达促进了胰岛素刺激的p70 S6激酶（p70 S6 K）活性，并增加了GSK 3 β的基础活性，但不促进胰岛素刺激的GLUT 4易位或葡萄糖摄取。这些数据支持Akt不是胰岛素刺激的GLUT 4转位或胰岛素激活的P13-激酶摄取葡萄糖信号的主要信号分子的结果。为了明确血小板衍生生长因子（PDGF）对3 T3-L1脂肪细胞葡萄糖转运的影响，我们研究了PDGF和胰岛素诱导的葡萄糖摄取 关于我们 3 T3-L1、3 T3-L1 GLUT 4 myc和3 T3-L1 GLUT 1 myc脂肪细胞中葡萄糖转运蛋白转位和PI 3-激酶活性。在3 T3-L1和3 T3-L1 GLUT 4 myc脂肪细胞中，胰岛素和PDGF分别刺激葡萄糖摄取9-10倍和5.5-6.5倍。外源性GLUT 4 myc表达增强PDGF诱导的葡萄糖转运。在3 T3-L1 GLUT 4 myc脂肪细胞中，胰岛素和PDGF分别诱导GLUT 4 myc易位增加8倍和5倍，在细胞表面抗c-myc抗体结合试验中测定。用荧光检测进一步证实了PDGF诱导的GLUT 4 myc易位。相反，PDGF刺激3 T3-L1 GLUT 1 myc脂肪细胞中GLUT 1 myc易位增加2倍，葡萄糖摄取增加2.5倍。PDGF诱导的GLUT 4 myc易位、葡萄糖摄取和PI 3-激酶活性在5-10 mm时达到最大（100%），此后迅速下降，在60 mm内分别为40%和12%，此时胰岛素的作用最大。Wortmannin（0.1 μM）可阻断PDGF诱导的3 T3-L1 GLUT 4 myc脂肪细胞中GLUT 4 myc易位和葡萄糖摄取。这些结果表明，PDGF可以瞬时触发GLUT 4的易位，并通过在3 T3-L1脂肪细胞中的磷脂酰肌醇3-激酶依赖性信号通路中的GLUT 4和GLUT 1两者的易位来刺激葡萄糖摄取。少

One of the down stream effectors of P13-kinase is serine-threonine kinase Akt (protein kinase B, RAK-PK), but the involvement of Akt in insulin-stimulated GLUT4 translocation is controversial. To investigate whether Akt 1 regulates insulin-stimulated GLUT4 translocation and glucose uptake in L6 myotubes, we established L6 myotubes stably expressing c-myc epitope-tagged GLUT4 (GLUT4myc) and mouse wild type (WT) Akt 1 . We found that overexpression of WT Akt 1 promoted insulin-stimulated p70S6 kinase (p70S6K) activity and increased the basal activity of GSK3β, but did not promote insulin-stimulated GLUT4 translocation or glucose uptake. These data supported the result that Akt is not a main signaling molecule to transmit the signal of insulin-stimulated GLUT4 translocation or glucose uptake from insulin-activated P13-kinase. To define the effect of platelet-derived growth factor (PDGF) on glucose transport in 3T3-L1 adipocytes, we investigated the PDGF- and insulin-induced glucose uptake … More , translocation of glucose transporters and PI 3-kinase activity in 3T3-L1, 3T3-L1GLUT4myc and 3T3-L1GLUT1myc adipocytes. Insulin and PDGF stimulated glucose uptake by 9-10 and 5.5-6.5-fold, respectively, in both 3T3-L1 and 3T3-L1GLUT4myc adipocytes. Exogenous GLUT4myc expression led to enhanced PDGF-induced glucose transport. In 3T3-L1GLUT4myc adipocytes, insulin and PDGF induced an 8-fold and 5-fold increase in GLUT4myc translocation, respectively, determined in a cell surface anti-c-myc antibody binding assay. This PDGF-induced GLUT4myc translocation was further demonstrated with fluorescent detection. In contrast, PDGF stimulated a 2-fold increase of GLUT1myc translocation and 2.5-fold increase of glucose uptake in 3T3-L1GLUT1myc adipocytes. The PDGF-induced GLUT4myc translocation, glucose uptake and PI 3-kinase activity were maximal(100%) at 5-10 mm, and thereafter rapidly declined to 40, , and 12%, respectively, within 60 mm, a time when effects of insulin were maximal. Wortmannin (0.1 μM) abolished PDGF-induced GLUT4myc translocation and glucose uptake in 3T3-L1GLUT4myc adipocytes. These results suggest that PDGF can transiently trigger the translocation of GLUT4 and stimulates glucose uptake by translocation of both GLUT4 and GLUT 1 in a phosphatidylinositol 3-kinase-dependent signaling pathway in 3T3-L1 adipocytes. Less