2,4-Diamino-6-hydroxypyrimidine (DAHP) suppresses cytokine-induced VCAM-1 expression on the cell surface of human umbilical vein endothelial cells in a BH 4-independent manner

Abstract

2,4-Diamino-6-hydroxypyrimidine (DAHP) is considered a specific inhibitor of BH 4 biosynthesis and is widely used in order to elucidate the possible biological function of BH 4 in various cells. In the present study, we found that both the synthesis of tetrahydrobiopterin (BH 4) and expression of vascular cell adhesion molecule 1 (VCAM-1) were increased in human umbilical vein endothelial cells (HUVEC) treated with proinflammatory cytokines. Thus we examined the effects of DAHP to clarify whether BH 4 might be involved in the expression of VCAM-1 in HUVEC. DAHP reduced the levels of both BH 4 and VCAM-1 induced by TNF-α and IFN-γ. However, the dose–response curves of DAHP for the suppression of the VCAM-1 level and that of BH 4 level were markedly different. Supplementation with sepiapterin failed to restore the depressed VCAM-1 level, although it completely restored the BH 4 level. Furthermore, DAHP significantly reduced the VCAM-1 level under the experimental conditions using TNF-α alone, which failed to induce BH 4 production. Taken together, these results indicate that DAHP inhibited the expression of VCAM-1 in a BH 4-independent manner in HUVEC. In the present study, we also found that DAHP significantly suppressed the accumulation of cytokine-induced NF-κB (p65) in the nucleus as well as the mRNA levels of VCAM-1 and GTP cyclohydrolase I (GTPCH), the rate-limiting enzyme of BH 4 synthesis. The data obtained in this study suggest that DAHP reduced VCAM-1 and GTPCH protein synthesis at least partially via suppressing the NF-κB level in the nucleus of HUVEC.