Estrogen increases glutathione level in brain endothelial cells exposed to ischemic injury by repression of ABCG2 activity

Abstract

The adenosine triphosphate (ATP)-binding cassette (ABC) efflux transporter ABCG2, which is located in the blood-brain barrier (BBB), limits the entry of endogenous compounds and xenobiotics into the brain. ABCG2 expression and activity are regulated by estrogen, a potent neuroprotectant, and are also affected by ischemic insult. However, the role of ABCG2 in estrogen-mediated protection in the ischemic brain is not clear. To define the action of ABCG2 in the brain after ischemic stroke, changes in ABCG2 protein levels in the brain of female mice and the effect of ABCG2 suppression in the brain endothelial cell line bEnd.3 were examined under ischemic injury. In the brain of estrogen-deficient ovariectomized mice, ABCG2 protein levels before and after the ischemic stroke were increased compared with ovary intact mice; this effect was reversed by estrogen replacement. Moreover, in bEnd.3 cells, estrogen reduced the basal ABCG2 protein level and efflux activity, and protected cells from ischemic injury without inducing ABCG2 expression. When bEnd.3 cells were transfected with ABCG2 small interfering RNA (siRNA), cell death from the ischemic insult was reduced and this protection was further enhanced by estrogen treatment. The intracellular concentration of glutathione, an antioxidant that is transported by ABCG2, was increased in cells transfected with ABCG2 siRNA after ischemic injury. These data indicated that the suppression of ABCG2 activity is associated with estrogen-mediated neuroprotective effects via the increase of intracellular glutathione. The findings further suggested that the modulation of ABCG2 activity offers a therapeutic target for brain diseases in estrogen-deficient aged women.