Fisslthaler Beate, Benzing Thomas, Busse Rudi, Fleming Ingrid
Institut für Kardiovaskuläre Physiologie, Klinikum der J. W. Goethe-Universität, Theodor-Stern-Kai 7, Frankfurt am Main D-60590, Germany.
Nitric Oxide. 2003 Jun;8(4):253-61. doi: 10.1016/s1089-8603(03)00042-9.
Insulin-induced vasodilatation in vivo has been attributed to the activation of the endothelial nitric oxide (NO) synthase (eNOS). The present study addressed the effects of insulin on the activity and expression of eNOS in native and cultured endothelial cells. Insulin applied to native porcine aortic endothelial cells elicited the tyrosine phosphorylation of the insulin receptor and receptor substrate, the subsequent activation of phosphatidylinositol 3-kinase (PI 3-K), Akt (protein kinase B), and ERK1/2. Insulin did not activate eNOS in cultured endothelial cells nor relax endothelium-intact arterial segments. However, 4h after application of insulin to native endothelial cells eNOS mRNA was increased 2-fold. A comparable increase in eNOS protein was detected after 18-24h and associated with an increase in intracellular cyclic GMP. In native endothelial cells, insulin enhanced the DNA-binding activity of Sp1 and AP-1, but not that of NF-kappaB. The insulin-induced increase in eNOS expression was prevented by wortmannin as well as by AP-1 decoy oligonucleotides. The MEK1 inhibitor, PD 98059, also enhanced eNOS expression in native and cultured endothelial cells, an effect which was independent of ERK1/2 and associated with an increase in the DNA-binding activity of AP-1 and Sp1. These results demonstrate that insulin activates multiple signalling pathways in endothelial cells but does not acutely activate eNOS. Insulin however enhances eNOS mRNA and protein by a mechanism involving the combined activation of a PI 3-K- and AP-1-dependent pathway.
胰岛素在体内诱导的血管舒张被认为是由于内皮型一氧化氮（NO）合酶（eNOS）的激活。本研究探讨了胰岛素对天然和培养的内皮细胞中eNOS活性及表达的影响。将胰岛素应用于天然猪主动脉内皮细胞可引发胰岛素受体和受体底物的酪氨酸磷酸化，随后激活磷脂酰肌醇3激酶（PI 3-K）、Akt（蛋白激酶B）和ERK1/2。胰岛素在培养的内皮细胞中未激活eNOS，也未使内皮完整的动脉段舒张。然而，在将胰岛素应用于天然内皮细胞4小时后，eNOS mRNA增加了2倍。在18 - 24小时后检测到eNOS蛋白有类似的增加，并伴有细胞内环磷酸鸟苷的增加。在天然内皮细胞中，胰岛素增强了Sp1和AP-1的DNA结合活性，但未增强NF-κB的活性。渥曼青霉素以及AP-1诱饵寡核苷酸可阻止胰岛素诱导的eNOS表达增加。MEK1抑制剂PD 98059也增强了天然和培养的内皮细胞中eNOS的表达，这一效应独立于ERK1/2，且与AP-1和Sp1的DNA结合活性增加有关。这些结果表明，胰岛素激活内皮细胞中的多种信号通路，但不会急性激活eNOS。然而，胰岛素通过一种涉及PI 3-K和AP-1依赖性途径联合激活的机制增强eNOS mRNA和蛋白的表达。
