Regulation of 6-pyruvoyltetrahydropterin synthase activity and messenger RNA abundance in human vascular endothelial cells.

Abstract

The nitric oxide synthase cofactor tetrahydrobiopterin (BH4) is involved in the regulation of endothelium-dependent vascular functions mediated by nitric oxide. Vascular endothelial cells synthesize and secrete large amounts of BH4 on cytokine activation. There is scant knowledge about molecular mechanisms of cytokine-triggered BH4 production in endothelial cells. Pteridine production, mRNA expression of GTP cyclohydrolase (GTPCH) and 6-pyruvoyltetrahydropterin synthase (PTPS) (both key enzymes of BH4 biosynthesis), and PTPS activity were studied in human umbilical vein endothelial cells (HUVECs) exposed to inflammatory cytokines. BH4 levels were </=140-fold enhanced on treatment of HUVECs with combined interferon-gamma/tumor necrosis factor-alpha/interleukin-1 (IFN/TNF/IL-1). Specific PTPS activity was approximately 3-fold higher in cytokine-treated HUVECs than in untreated cells. Reverse-transcription/limiting-dilution polymerase chain reaction analysis showed that in response to IFN/TNF/IL-1, mRNA abundance of GTPCH and PTPS was increased approximately 64-fold and 10-fold, respectively. The present study demonstrates for the first time the cytokine-dependent regulation of PTPS, the second enzyme in BH4 synthesis. Although GTPCH is believed to be the rate-limiting step, control of endothelial PTPS expression by cytokines may play an important role in regulating BH4-dependent nitric oxide production in the vascular system.