Insulin-stimulated cyclic guanosine monophosphate inhibits vascular smooth muscle cell migration by inhibiting Ca/calmodulin-dependent protein kinase II.
BACKGROUND
Insulin resistance is associated with vascular disease. Physiological concentrations of insulin inhibit cultured vascular smooth muscle cell (VSMC) migration in the presence of nitric oxide, and the failure to do so in insulin-resistant states may aggravate vascular disease. We sought to determine the molecular mechanisms by which insulin inhibits VSMC migration.


RESULTS
Insulin at 1 nmol/L stimulated cGMP production in cultured rat VSMCs that were induced to express inducible nitric oxide synthase (iNOS). VSMC migration was measured in a wound-closure assay, and the platelet-derived growth factor-AB (PDGF-AB)-stimulated component of VSMC migration after wounding was inhibited by insulin, 8-Br-cGMP, and 1-[N-0-bis(5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenylpiperazine (KN-62), a selective inhibitor of calcium/calmodulin-dependent protein kinase II (CaM kinase II). Wounding alone or incubating cells with only PDGF-AB stimulated CaM kinase II activity in an insulin- and 8-Br-cGMP-inhibitable manner. Transfecting VSMCs with a constitutively active CaM kinase II mutant blocked the inhibition by insulin of both wound-induced and wound plus PDGF-AB-induced VSMC migration. High intracellular Ca2+ ([Ca]i)-stimulated CaM kinase II activity was inhibited by 8-Br-cGMP by an okadaic acid-sensitive mechanism.


CONCLUSIONS
We conclude that in cultured rat VSMCs expressing iNOS, insulin, via stimulation of cGMP production, inhibits both wound alone-induced and the PDGF-AB-stimulated component of VSMC migration by inhibiting CaM kinase II activity. cGMP inhibits CaM kinase II at a post-[Ca]i step by a protein phosphatase-dependent mechanism.
Zhang
Sui
S
Division of Renal Diseases and Hypertension, Department of Medicine, the University of Texas Health Science Center, Houston, Tex, USA.
Yang
Yu
Y
Kone
Bruce C
BC
Allen
Julius C
JC
Kahn
Andrew M
AM
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics; D002465:Cell Movement / Q000187:drug effects; D002478:Cells, Cultured; D006152:Cyclic GMP / Q000031:analogs & derivatives* / Q000378:metabolism* / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007328:Insulin / Q000494:pharmacology*; D015759:Ionomycin / Q000494:pharmacology; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000201:enzymology / Q000502:physiology*; D009154:Mutation; D010766:Phosphorylation; D051381:Rats; D017207:Rats, Sprague-Dawley; D013912:Threonine / Q000378:metabolism
D004791:Enzyme Inhibitors; D007328:Insulin; D013912:Threonine; C016276:8-bromocyclic GMP; D015759:Ionomycin; C063302:KN 62; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D006152:Cyclic GMP