Pyrrolidine dithiocarbamate inhibits immunostimulant-induced tetrahydrobiopterin synthesis in rat vascular smooth muscle.

GTP cyclohydrolase I is the first and rate-limiting enzyme in the synthesis of tetrahydrobiopterin, a cofactor of nitric oxide (NO) synthase. Immunostimulants increase NO and tetrahydrobiopterin synthesis in vascular smooth muscle cells by coinducing NO synthase and GTP cyclohydrolase I gene expression. Given that nuclear factor kappa(B) mediates the induction of NO synthase gene expression by lipopolysaccharide (LPS), the role of nuclear factor kappa(B) in the induction of GTP cyclohydrolase I in LPS-stimulated rat vascular smooth muscle cells was assessed by examining the effects of pyrrolidine dithiocarbamate, an inhibitor of the activation of nuclear factor kappa(B), on the abundance of GTP cyclohydrolase I mRNA and biopterin synthesis. Pyrrolidine dithiocarbamate inhibited both NO and biopterin synthesis induced by LPS in a dose-dependent manner with similar half-maximal inhibitory concentrations, 12 mu M for NO and 17 mu M for biopterin, respectively. At a concentration of 25 mu M, which inhibited NO and biopterin synthesis but caused no cytotoxicity, pyrrolidine dithiocarbamate substantially reduced the LPS-induced increase in the abundance of NO synthase and GTP cyclohydrolase I mRNAs. These results suggest that pyrrolidine dithiocarbamate inhibits LPS-induced NO and biopterin synthesis by inhibiting the expression of NO synthase and GTP cyclohydrolase I genes. Thus, the induction of both genes necessary for cellular NO synthesis in vascular smooth muscle appears to be regulated, at least in part, by a common mechanism: nuclear factor kappa(B) activation.