Aim To explore the effect of sitagliptin on the expression of endothelin-1(ET-1) and endothelial nitric oxide synthase(eNOS) in human aortic endothelial cells(HAEC) and its underlying mechanism in high glucose environment. Methods HAEC were cultured in high glucose environment(25 mmol/L), and treated with different concentrations of sitagliptin(0, 5, 10 and 20 μmol/L, respectively). The mRNA and protein expressions of eNOS, ET-1, iNOS and phosphate nuclear factor-kappa B p65(p-NF-κB p65) were measured. The measurements for eNOS, ET-1, iNOS, NF-κB p65 on mRNA and protein levels in HAEC were evaluated after incubation with tumor necrosis factor-α(TNF-α) and sitagliptin. Results Compared with normal medium(glucose concentrations for 7 mmol/L), both the mRNA and the protein expression of eNOS in HAEC significantly decreased in high glucose medium, while those of ET-1, iNOS and p-NF-κB p65 protein significantly increased(P<0.05). Compared with 0 μmol/L sitagliptin, 20 μmol/L sitagliptin significantly increased mRNA and protein expressions of eNOS, while decreased those of ET-1, iNOS and p-NF-κB p65 protein(P<0.05). Compared with sitagliptin alone treated HAEC, both the mRNA and the protein expressions of eNOS significantly decreased in HAEC treated with TNF-α and sitagliptin, while those of ET-1, iNOS and p-NF-κB p65 protein expressions significantly increased(P<0.05). Conclusions Sitagliptin enhances eNOS, represses ET-1, iNOS expressions at the level of transcription and translation through inhibiting NF-κB p65 phosphorylation in HAEC in high glucose environment. This may contribute to the improvement of endothelial function and prevention of subsequent atherogenesis.