Interactive effects of insulin-like growth factor-1 and β-estradiol on endothelial nitric oxide synthase activity in rat aortic endothelial cells

Insulin-like growth factor-1 (IGF-1) and beta-estradiol (E2) have vasodilatory effects, in part, through stimulation of vascular nitric oxide (NO) production. However, their interactive effects on endothelial nitric oxide synthase (eNOS) and NO production have not been previously studied in endothelial cells (EC). Employing rat aortic EC (RAEC), the effects of acute (20 and 30 minutes) and prolonged (4 hours) stimulation with 100 nmol/L IGF-1 and 1 nmol/L E2 (alone or in combination) were assessed with respect to protein levels and enzymatic activities for phosphatidyl inositol 3-kinase (PI3K) and serine/threonine kinase Akt (Akt), enzymes involved in eNOS activation. Exposure to IGF-1 for 30 minutes or E2 for 20 minutes increased insulin receptor substrate-1 (IRS-1) association with the regulatory (p85) subunit of PI3K, enhanced tyrosine phosphorylation of p85, and increased PI3K activity. Combined treatment had a greater effect on p85 phosphorylation and PI3K activity then either agonist alone. Moreover, IGF-1 and E2 enhanced Akt Ser(473) phosphorylation, with the effect of IGF-1 being much greater. Acute expose to both E2 (20 minutes) and IGF-1 (30 minutes) were associated with an increase in eNOS activity. Prolonged exposure (4 hours) to either IGF-1 or E2 increased expression of the p85 subunit as well as eNOS activity. Pretreatment with PI3K antagonist wortmannin (WT) prevented this increase in eNOS activity. The results suggest that IGF-1 and E2 may interact through PI3K/Akt-related pathways to increase eNOS activity. Copyright 2003 Elsevier, Inc. All rights reserved.