Role of nuclear factor kappa B in cytokine-induced nitric oxide and tetrahydrobiopterin synthesis in rat neonatal cardiac myocytes.

The nitric oxide (NO) signalling pathway is thought to play a direct role in regulating the contractile properties of cardiac muscle both in vitro and in vivo. The inducible isoform of NO synthase (iNOS) mediates a sustained increase in NO production in response to cytokines in the cardiac myocytes; however, the regulation of NO synthesis in these cells remains poorly understood. Tetrahydrobiopterin (BH4) is an essential cofactor for NO formation. Cytokines induce the de novo synthesis of BH4 in cardiac myocytes, an event that is essential for the induction of NO synthesis. Activation of NO formation by cytokines in cardiac myocytes requires transcriptional induction of the genes that encode iNOS and guanosine triphosphate cyclohydrolase I (GTPCH), the first and rate-limiting enzyme in de novo BH4 synthesis. Given that nuclear factor kappa B (NF-kappa B) mediates the induction of iNOS gene expression in various cell types, the role of NF-kappa B in the induction of iNOS in cytokine-stimulated rat neonatal cardiac myocytes was assessed by examining the effects of pyrrolidine dithiocarbamate (PDTC), an inhibitor of NF-kappa B activation, on the abundance of iNOS mRNA and NO synthesis. The effects of PDTC on GTPCH mRNA abundance and biopterin synthesis were also investigated. PDTC inhibited in a dose-dependent manner both NO and BH4 synthesis induced by a combination of interleukin-1 alpha (IL-1 alpha) and interferon-gamma (IFN gamma), with a half-maximal inhibitory concentration of 22 muM. PDTC also prevented the accumulation of iNOS and GTPCH mRNAs induced by IL-1 alpha and IFN gamma. Cytokine-induced NO and BH4 synthesis was also inhibited by tosyl-lysine-chloromethyl ketone. another inhibitor of NF-kappa B activation. Results suggest that PDTC inhibits cytokine-induced NO and BH4 synthesis by inhibiting the expression of iNOS and GTPCH genes. Thus, the induction of both genes necessary for NO synthesis in cardiac myocytes appears to be regulated, at least in part, by a common mechanism: NF-kappa B activation.