Differential Inhibition of Constitutive and Inducible Nitric Oxide Synthase in Vascular Endothelial Cells by Analogues of Tetrahydrobiopterin

Abstract In the vasculature, a physiologic production of nitric oxide (NO) is maintained by endothelial nitric oxide synthase (eNOS). Induction of inducible nitric oxide synthase (ÍNOS) under inflammatory conditions (e.g. septic shock) resulting in high levels of nitric oxide (NO) is believed to be partly responsible for the pathophysiologic changes in the vascular system that occur under inflammatory conditions (e.g. septic shock). Both NOS isoforms are dependent on the obligatory cofactor tetrahydrobiopterin (BH 4 ). We investigated the selectivity and potency of the BH 4 analogues 4-amino-BH 4 and 5-methyl-BH 4 in inhibiting eNOS and iNOS in a murine vascular endothelial cell (MVEC) model expressing either eNOS or iNOS under physiologic and inflammatory conditions, respectively. Exogenous BH 4 and its precursor sepiapterin both enhanced physiologic eNOS activity in resting MVEC, while 4-amino-BH 4 slightly inhibited eNOS. 5-methyl-BH 4 did not have any effect on eNOS. BH 4 , sepi - apterin, and 5-methyi-BH 4 had no effect on iNOS in inflammatory activated MVEC. In contrast, 4-amino-BH4 selectively inhibited iNOS with a potency comparable to the unselective NOS inhibitor Νω-monomethyl-L-argimne (L-NMMA). The present study demonstrates that 4-amino-BH 4 selectively and potently inhibits iNOS in vascular endothelial cells, while its effect on eNOS is minimal. The selective inhibition of iNOS is a promising strategy for the treatment of inflammatory conditions with high output of NO. Further in vivo studies are required to determine whether inhibition of NO production by analogues of BH 4 offers any advantage compared to inhibition by L-arginine analogues.