Abstract
Endothelial nitric oxide synthase (eNOS) is the enzyme that endogenously biosynthesize nitric oxide (NO) from L-arginine. Because NO serves important functions to maintain vascular homeostasis, it must be controlled tightly. If not, endothelial dysfunction occurs. The activity of eNOS is regulated by multiple interdependent control mechanisms and signaling pathways, and one of that is phosphorylation at Ser1177 residue of the enzyme. In this study, we figured out that orotic acid, an intermediate in pyrimidine synthesis, inhibits activation of Akt/eNOS signaling pathway by insulin. In ECV304 human endothelial cells, orotic acid inhibits insulin activation of eNOS. In ECV304 cells incubated with orotic acid, phosphorylation at Ser1177 of eNOS by insulin is decreased in dose-dependent manner. Orotic acid also reduces NO production by insulin. However, orotic acid didn’t affect eNOS expression or eNOS phosphorylation without insulin signaling. To investigate the effect of orotic acid on upstream signaling pathway, we also checked Akt, a well-known kinase which phosphorylates eNOS. Insulin activates Akt through PI3K/Akt signaling pathway, and Akt phosphorylates eNOS directly. Orotic acid also inhibits Akt phosphorylation by insulin. In conclusion, orotic acid inhibits insulin activation of Akt/eNOS signaling pathway and decreases NO production. Therefore, orotic acid breaks vascular homeostasis and induces endothelial dysfunction.
