Human vascular smooth muscle cells express a constitutive nitric oxide synthase that insulin rapidly activates, thus increasing guanosine 3' :5'-cyclic monophosphate and adenosine 3' :5'-cyclic monophosphate concentrations

• Published in: Diabetologia 1999-07, Vol.42 (7), p.831-839
• Main Authors: TROVATI, M, MASSUCCO, P, MATTIELLO, L, COSTAMAGNA, C, ALDIERI, E, CAVALOT, F, ANFOSSI, G, BOSIA, A, GHIGO, D
• Format: Article
• Language: English
• Subjects: Adenosine; Biological and medical sciences; Blood vessels and receptors; Calcium - pharmacology; Cells, Cultured - drug effects; Citrulline - metabolism; Cyclic AMP - analysis; Cyclic GMP - analysis; Diabetes; Endothelium; Fundamental and applied biological sciences. Psychology; Gene Expression; Humans; Hypotheses; Insulin; Insulin - pharmacology; Ionomycin - pharmacology; Muscle, Smooth, Vascular - cytology; Muscle, Smooth, Vascular - enzymology; Nitric oxide; Nitric Oxide - biosynthesis; Nitric Oxide - physiology; Nitric Oxide Synthase - chemistry; Nitric Oxide Synthase - drug effects; Nitric Oxide Synthase - genetics; Nitric Oxide Synthase - metabolism; Nitric Oxide Synthase Type III; Nitroprusside - pharmacology; RNA, Messenger - analysis; Smooth muscle; Transcription, Genetic; Vertebrates: cardiovascular system
• ISSN: 0012-186X; 1432-0428
• DOI: 10.1007/s001250051234

Insulin incubation of human vascular smooth muscle cells (hVSMC) for 120 min increases both guanosine 3':5'-cyclic monophosphate (cGMP) and adenosine 3':5'-cyclic monophosphate (cAMP) and these effects are blocked by inhibiting nitric oxide synthase (NOS). These data suggest that insulin activates a constitutive Ca2+-dependent NOS (cNOS), not described at yet in hVSMC. To test this hypothesis, we evaluated in hVSMC: i) the kinetics of the insulin-induced enhancement of the two cyclic nucleotides; ii) the ability of nitric oxide (NO) to increase both cyclic nucleotides; iii) NO involvement in the short-term influence of insulin on both cyclic nucleotides; iv) the ability of insulin to increase NO production in a few minutes; v) the presence of a cNOS activity; vi) the expression of mRNA for cNOS. In hVSMC incubated with insulin, NO donors and the Ca2+ ionophore ionomycin, we measured cAMP and cGMP (RIA); in hVSMC incubated with insulin and ionomycin we measured NO, evaluated as L-(3H)-citrulline production from L-(3H)-arginine; by northern blot hybridization, we measured the expression of cNOS mRNA. i) By incubating hVSMC with 2 nmol/l insulin for 0-240 min, we observed an increase of both cGMP and cAMP (ANOVA: p = 0.0001). Cyclic GMP rose from 0.74 +/- 0.01 to 2.62 +/- 0.10 pmol/10(6) cells at 30 min (p = 0.0001); cAMP rose from 0.9 +/- 0.09 to 11.65 +/- 0.74 pmol/10(6) cells at 15 min (p=0.0001). ii) Sodium nitroprusside (100 mol/l) and glyceryltrinitrate (100 micromol/l) increased both cGMP and cAMP (p = 0.0001). iii) The effects of insulin on cyclic nucleotides were blocked by NOS inhibition. iv) An increase of NO was observed by incubating hVSMC for 5 min with 2 nmol/l insulin (p = 0.0001). v) Ionomycin (1 micromol/l) enhanced NO production (p = 0.0001) and increased both cyclic nucleotides (p = 0.0001). vi) hVSMC expressed mRNA of cNOS. Human VSMC express cNOS, which is rapidly activated by insulin with a consequent increase of both cGMP and cAMP, suggesting that insulin-induced vasodilation in vivo is not entirely endothelium-mediated.