Inhibition of Vascular Smooth Muscle Cell Migration by Cytochrome P450 Epoxygenase-Derived Eicosanoids

Abstract

Abstract-Vascular smooth muscle cell (SMC) migration and proliferation contribute to neointimal hyperplasia and restenosis after vascular injury. The epoxyeicosatrienoic acids (EETs), which are products of cytochrome P450 (CYP) epoxygenases, possess vasodilatory, antiinflammatory, and fibrinolytic properties. To determine whether these compounds also possess antimigratory and antiproliferative properties, we stimulated rat aortic SMCs with either 20% serum or platelet-derived growth factor (PDGF-BB, 20 ng/mL). In a concentration-dependent manner, treatment with EETs, particularly 11,12-EET, inhibited SMC migration through a modified transwell filter by 53% to 60%. EETs, however, have no inhibitory effects on PDGF-stimulated SMC proliferation. Adenoviral-mediated overexpression of the CYP isoform, CYP2J2, in SMCs also inhibited serum-and PDGF-induced SMC migration by 32% and 26%, respectively; both effects of which were reversed by the CYP inhibitors SKF525A or clotrimazole, but not by the K Ca channel blocker, charybdotoxin, or the cyclooxygenase inhibitor, diclofenac. The effect of EETs correlated with increases in intracellular cAMP levels. Indeed, forskolin and 8-bromo-cAMP exert similar inhibitory effects on SMC migration as 11,12-EET and the effects of 11,12-EET were blocked by cAMP and protein kinase A (PKA) inhibitors. [1][2][3] The EETs have properties similar to those of endothelium-derived hyperpolarizing factor (EDHF) because they hyperpolarize and relax vascular smooth muscle cells (SMCs) by activating calcium-sensitive potassium (K Ca ) channels. 4,5 Recently, several CYP epoxygenases, including members of the CYP2B, CYP2C, and CYP2J subfamilies, have been identified in vascular endothelial cells. However, their relative importance in endothelial EET biosynthesis has not been determined. 1,6 -8 Treatment of porcine coronary artery endothelial cells in vitro with either ␤-naphthoflavone or nifedipine induces CYP2C expression, increases 11,12-EET biosynthesis, and enhances bradykinin-induced coronary artery relaxation via SMC membrane hyperpolarization. 1,9 Moreover, transfection of endothelial cells with an antisense oligonucleotide to CYP2C8/9 or treatment with the selective CYP2C9 inhibitor, sulfaphenazole, attenuates EDHFmediated vascular responses, thus providing supporting evidence that the EDHF synthase in the porcine vascular bed may be a CYP2C isoform. 1,9 In contrast to their vasodilatory effects, EETs have also been shown to possess important nonvasodilatory actions within the vascular system. For example, EETs which are produced by a member of the CYP2J family, CYP2J2, inhibit cytokine-induced endothelial cell adhesion molecule expression by inhibiting the proinflammatory transcriptional factor, NF-B. 6 Furthermore, in vascular endothelial cells, addition of physiologically relevant concentrations of EETs or overexpression of CYP2J2 increases tissue plasminogen activator (tPA) expression and fibrinolytic activity via a G␣ sdependent, cAMP-mediated mechanism. 10 However, neither the antiin...

Arachidonic Acid Epoxygenase Metabolites Stimulate Endothelial Cell Growth and Angiogenesis via Mitogen-Activated Protein Kinase and Phosphatidylinositol 3-Kinase/Akt Signaling Pathways

“…Together, these data indicate that the EET-mediated effects on BAEC proliferation, migration, and angiogenesis likely occur via NO-dependent mechanisms as well as via activation of PI3-kinase/Akt, MAPK, and possibly EGFR pathways as well. Interestingly, a recent study indicated that EETs inhibit vascular smooth muscle cell migration but not proliferation and that this effect is mediated by an EETinduced increase in intracellular cAMP and activation of the PKA signaling pathway (Sun et al, 2002). The differential response to EETs in vascular smooth muscle cells and endothelial cells may be due to tissue specificity of EET activity.…”

“…Indeed, 11,12-EET has been proposed to be identical to endothelium-derived hyperpolarizing factor (Bauersachs et al, 1997;Bolz et al, 2000;Matoba et al, 2000Matoba et al, , 2002Hamilton et al, 2001;Lacza et al, 2002;Miura et al, 2003;Morikawa et al, 2003;Tanaka et al, 2003;Yada et al, 2003). Exogenous application of EETs inhibits vascular smooth muscle cell migration, platelet aggregation, nuclear factor-B activation, and vascular cell adhesion molecule-1 expression (Node et al, 1999;Fleming et al, 2001;Sun et al, 2002;Krotz et al, 2004), suggesting an overall beneficial role for EETs within the vasculature and a protective role in the development of atherosclerosis.…”

Apolipoprotein E Binding to Low Density Lipoprotein Receptor-related Protein-1 Inhibits Cell Migration via Activation of cAMP-dependent Protein Kinase A

“…For example, the cAMP activator forskolin as well as stable cAMP analogues such as 8-bromo-cAMP are effective inducers of PKA activity and inhibitors of smooth muscle cell migration (17,19). In addition, adrenomedullin has also been shown to inhibit the migration of rat aortic smooth muscle cells by inducing intracellular cAMP accumulation (17).…”

“…These results indicated that apoE inhibition of fibroblast migration is also mediated via induction of cAMP accumulation and the consequential activation of PKA. Finally, to verify that the lack of apoE-induced cAMP accumulation in LRP-deficient cells is not a generalized phenomenon of abnormal cyclic nucleotide metabolism due to LRP-1 deficiency, we tested the effect of forskolin, a direct activator of adenylyl cyclase and inhibitor of smooth muscle cell migration (17,19), on the migration of MEF-1, PEA-10, and PEA-13 cells. Results, as shown in Fig.…”

Emerging role of epoxyeicosatrienoic acids in coronary vascular function

“…Also, EDHF is particularly important in modulating arteriolar tone in cardiovascular disease states [6,11,12] when endothelial NO synthase is impaired by excess asymmetric dimethylarginine (ADMA) [13] and NO is directly quenched by excess superoxide [14]. In addition, EDHF also possesses antiatherosclerotic properties similar to NO [15][16][17]. Hence, EDHF may represent an endogenous vascular protective mechanism, and has therefore received considerable attention in recent years as a potential therapeutic target for treatment of cardiovascular disease.…”