Several experimental and epidemiological studies have demonstrated cerebrovascular protective effects of estrogen that is associated with reduced risk, development and severity of cerebrovascular disease and events such as ischemic stroke. A central mechanism underlying this estrogen-related neuroprotection is believed to be enhanced endothelial-derived nitric oxide (NO) bioavailability. NO is critical for maintaining the vasodilatory capacity of the cerebral vasculature. In addition to its vasodilatory properties NO maintins blood brain barrier integrity and mitigates various atherogenic processes. The direct and indirect effects of estrogen on human brain endothelial cell NO production is not fully understood as most studies on the effects of estrogen on cerebral vascular function involve animal models and in vitro studies have used a non-cerebral endothelial cell line. The experimental aim of this study was to determine, in vitro, the effect of estrogen on cerebral endothelial cell NO synthase (NO) activation and NO production. Human cerebral microvascular endothelial cells (hCMECs) were cultured (3rd passage) and incubated with culture media in the absence and presence of 17β-estradiol (100 nM) for 24 hours (n=5 experimental units). hCMECs are a well validated, and clinically relevant, cell model for studying brain microvascular endothelial cell function. Cells were harvested and intracellular expression of eNOS and phosphorylated (p)-eNOS at Ser1177 and Thr495 sites were determined by capillary electrophoresis immunoassay. Phosphorylation is the primary posttranslational modification regulating eNOS enzyme activity and, in turn, NO production. Phosphorylation of Ser1177 confers the greatest activation of eNOS; whereas, phosphorylation at Thr495 inhibits eNOS activity. Although neither total eNOS (461.1+28.0 vs 461.6+34.0 AU; P=0.99) nor p-eNOS (Thr495) (13.3+1.2 vs 12.3+1.8 AU; P=0.65) expression was significantly altered; however, expression of p-eNOS (Ser1177) was markedly higher (31.4+2.0 vs 21.0+1.9 AU; P=0.006) in hCMECs treated with 17β-estradiol. Consequently, NO production was ~20% higher in 17β-estradiol-treated cells (6.8±0.7 vs 5.6±0.4 μmol/L; P=0.01). In summary, 17β-estradiol favorably affects eNOS activity and, in turn, NO production in brain endothelial cells in vitro. Increased NO bioavailability is central to cerebral blood flow regulation, maintaining vascular patency and reducing the risk and severity of cerebrovascular disease and ischemic stroke. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.
Valenti et al. (Fri,) studied this question.