Inhibition of endothelial nitric oxide synthase induces and enhances myocardial fibrosis

The endothelial nitric oxide synthase (eNOS) contributes to cardiac remodelling. We studied the role of eNOS in the development of myocardial fibrosis during cardiac overload. Ten-week-old male C57/Bl6 wildtype (WT) and eNOS mice (eNOS(/)) were subjected to transverse aortic constriction (TAC, 360 m) and WT were treated with l-N-G-nitroarginine methyl ester (l-NAME, 100 mg/kg/day) for 35 days. Inhibition of eNOS by l-NAME induced interstitial fibrosis, augmented replacement fibrosis, and induced apoptosis of cardiac fibroblasts and cardiomyocytes. l-NAME and eNOS(/) markedly increased the fibrosis induced by TAC and enhanced the myocardial prevalence of CXCR4(pos) fibroblasts. Myocardial stromal-derived factor-1 (SDF-1) expression was up-regulated by l-NAME and down-regulated after TAC. Blood pressure lowering by co-treatment with hydralazine (250 mg/L/day) did not reverse the l-NAME effects. In mice transplanted with green fluorescent protein (GFP)(pos) bone marrow, l-NAME increased the percentage of GFP(pos) fibroblasts in the myocardium to 4570. Strain-mismatched BMT of eNOS(/)-BM increased and of WT-BM decreased the percentage of CXCR4(pos) fibroblasts in all groups. The number of fibrocytes (CD45(pos) collagen I-pos cells) in the peripheral blood and in the bone marrow was increased both by TAC and l-NAME. l-NAME but not the inhibitor of inducible NOS 1400 W and of neuronal NOS 7-nitroindazole increased hydroxyproline and collagen I1. l-NAME up-regulated SDF-1 mRNA in cultured neonatal rat cardiac fibroblasts as well as their migratory capacity. eNOS inhibition induces and enhances cardiac fibrosis independently of blood pressure by activating SDF-1/CXCR4, extracellular matrix production in cardiac fibroblasts and by increasing recruitment of fibrocytes from the bone marrow.