Cystic fibrosis (CF) is a genetic disorder associated with a wide range of systemic complications, including vascular endothelial dysfunction. Nitric oxide (NO) plays a major role in maintaining vascular function and tetrahydrobiopterin (BH4) is a critical determinant of NO bioavailability. Thus, the purpose of this study was to investigate the effects of oral administration of BH4 on endothelial function in patients with CF. Methods: 29 patients with CF (18 ± 8 yrs old) and 29 healthy matched controls were recruited. Patients with CF participated in a randomized trial where they received either a 5 mg/kg dose of oral BH4 (BH4-5; n=17) or a 20 mg/kg dose of oral BH4 (BH4-20; n=12). On a separate visit, a subset of patients from each group were retested following a placebo (PLC, n=9). Brachial artery flow-mediated dilation (FMD) was used to evaluate vascular endothelial function before and 3 h after treatment. Results: Baseline FMD was lower in patients compared to controls. No change in FMD was observed following either PLC or BH4-5 (∆FMD: -0.8 ± 0.0% and -0.5 ± 2.5%; p=0.273 and 0.132, respectively). Treatment with BH4-20, however, resulted in a significant improvement in FMD (∆FMD: 1.1 ± 1.4%) when compared to BH4-5 (p=0.023) and PLC (p=0.017). Conclusion: These data suggest that a single oral dose of BH4 at 20 mg/kg can improve vascular endothelial function in patients with CF. These findings support the hypothesis that BH4 bioactivity contributes, in part, to endothelial dysfunction in patients with CF.