NOS gene transfer inhibits expression of cell cycle regulatory molecules in vascular smooth muscle cells.
The mechanisms of nitric oxide (NO)-mediated inhibition of vascular smooth muscle (VSM) cell proliferation are still obscure. Cyclins A and E in association with cyclin-dependent kinase 2 (cdk2) serve as positive regulators for mammalian cell cycle progression through the G1/S checkpoint of the cell cycle and subsequent cell proliferation. Therefore, we have tested the effect of adenovirus-mediated transfection of the endothelial nitric oxide synthase (eNOS) gene into guinea pig coronary VSM cells on platelet-derived growth factor (BB homodimer) (PDGF-BB)-stimulated cell proliferation and the expression of cell cycle regulatory molecules. Transfection of the eNOS gene (eNOS) into VSM cells significantly inhibited (P < 0.05) [3H]thymidine incorporation into the DNA in response to PDGF-BB stimulation compared with lacZ-transfected control cells. The eNOS transfer significantly inhibited (P < 0.05) PDGF-BB-induced proliferating cell nuclear antigen (PCNA) and cyclin A expression in VSM cells compared with cells transfected with the control vector. The time course of cyclin E expression in response to PDGF-BB stimulation was delayed in eNOS-transfected cells. Levels of cyclin-dependent kinase inhibitors p21 and p27 were not significantly affected by eNOS transfer. eNOS transfer did not decrease PDGF-beta receptor number, affinity, and autophosphorylation measured by radioreceptor assay and Western analysis. These results suggest that inhibition of PDGF-stimulated expression of cyclin A, cyclin E, and PCNA is the target of NO action. These findings could explain, at least in part, NO-mediated inhibition of VSM cell proliferation.
D000256:Adenoviridae; D000818:Animals; D000077214:Becaplermin; D002455:Cell Division / Q000235:genetics; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000166:cytology / Q000201:enzymology; D019925:Cyclin A / Q000378:metabolism; D019927:Cyclin E / Q000378:metabolism; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D015967:Gene Expression Regulation, Viral / Q000187:drug effects; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D006168:Guinea Pigs; D007763:Lac Operon; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000201:enzymology*; D019001:Nitric Oxide Synthase / Q000235:genetics*; D052250:Nitric Oxide Synthase Type III; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000378:metabolism; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D020574:Proto-Oncogene Proteins c-sis; D013936:Thymidine / Q000378:metabolism / Q000494:pharmacology; D014316:Tritium