eNOS gene transfer inhibits smooth muscle cell migration and MMP-2 and MMP-9 activity.
Vascular smooth muscle cell (SMC) migration is a critical step in the development of neointima after angioplasty. Matrix metalloproteinases (MMPs) degrade the basement membrane and the extracellular matrix, facilitating SMC migration. Transfer of the endothelial nitric oxide synthase (eNOS) gene to the injury site inhibits neointima formation. Neither the signaling pathways leading to NO-mediated inhibition of SMC migration and proliferation nor the alterations in these pathways have been characterized. We hypothesize that NO inhibits SMC migration in part by regulating MMP activity. To test this hypothesis, we transfected cultured rat aortic SMCs with replication-deficient adenovirus containing bovine eNOS gene and analyzed the conditioned medium for MMP activity. We observed that eNOS gene transfer significantly (P<0.05) inhibited SMC migration and significantly (P<0.05) decreased MMP-2 and MMP-9 activities in the conditioned medium. Similarly, addition of the NO donor DETA NONOate and 8-bromo-cGMP to the culture medium significantly decreased MMP-2 and MMP-9 activities in the conditioned medium collected 24 hours after treatment. Furthermore, Western blot analysis of the conditioned medium collected from eNOS gene-transfected SMCs showed a significant increase in tissue inhibitor of metalloproteinases-2 (TIMP-2) levels. Our data suggest that NO decreases MMP-2 and MMP-9 activities and increases TIMP-2 secretion, and this shifts the balance of MMP activity, which may favor the inhibition of cell migration because of inhibition of extracellular matrix degradation.
Gurjar
M V
MV
Department of Anatomy and Cell Biology, The University of Iowa College of Medicine, Iowa City 52242, USA.
Sharma
R V
RV
Bhalla
R C
RC
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001011:Aorta / Q000166:cytology; D015153:Blotting, Western; D002465:Cell Movement / Q000502:physiology*; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000166:cytology; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic; D018014:Gene Transfer Techniques; D006168:Guinea Pigs; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism*; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism*; D061965:Matrix Metalloproteinase Inhibitors; D009131:Muscle, Smooth, Vascular / Q000166:cytology*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000235:genetics* / Q000378:metabolism*; D052250:Nitric Oxide Synthase Type III; D009603:Nitroso Compounds / Q000494:pharmacology; D051381:Rats; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000032:analysis / Q000378:metabolism
D061965:Matrix Metalloproteinase Inhibitors; D020030:Nitric Oxide Donors; D009603:Nitroso Compounds; D019716:Tissue Inhibitor of Metalloproteinase-2; C094210:2,2'-(hydroxynitrosohydrazono)bis-ethanamine; C016276:8-bromocyclic GMP; D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III; C496316:Nos3 protein, rat; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9; D006152:Cyclic GMP