Aims: Estrogen induces nitric oxide (NO) in the endothelium and appears to protect against inflammation and atherosclerosis. NO can induce post-translational protein modifications such as cysteine S-nitrosylation in the cellular proteins which may exert anti-inflammatory effects. However, whether estrogen can induce protein S-nitrosylation in the endothelium is not known. Given this background, we investigated the role of 17beta-estradiol (E2beta), the major form of estrogen in the body, on endothelial protein S-nitrosylation.
Methods and results: Experiments were performed in human umbilical vein endothelial cells (HUVECs). S-nitrosylation was detected by immunostaining for nitrosocysteine and further confirmed by biotin switch method. Ovariectomized 12-month-old Sprague-Dawley rats with/without estradiol supplementation were used for in vivo validation of findings. We found that physiologically relevant doses of E2beta increased protein S-nitrosylation in HUVECs through estrogen receptor-alpha (ERalpha) and endothelial nitric oxide synthase (eNOS). Interestingly, specific agonists for both ERalpha and ERbeta increased eNOS protein expression, while only the former could activate eNOS through phosphorylation. S-nitrosylation by E2beta prevented angiotensin II-induced upregulation of intercellular cell adhesion molecule-1, suggesting a potential anti-inflammatory mechanism. Finally, we showed that exogenous E2beta could increase endothelial S-nitrosylation in vivo in a rat model.
Conclusion: Our results demonstrate for the first time that E2beta increases protein S-nitrosylation in the vascular endothelium, which might be a novel pathway to mediate the protective effects on the vasculature.