Estrogen Decreases Mitochondrial ROS Production in Human Brain Endothelial Cells

Mitochondria are major producers of reactive oxygen species (ROS) as a by‐product of oxidative phosphorylation. We have recently shown that estrogen (E) has profound effects on mitochondrial function in cerebral arteries, increasing efficiency of energy production and decreasing mitochondrial ROS. To further explore effects of E, we used human brain microvascular endothelial cells (HBMEC) treated with 10 nM E for 24 hours. Direct measurement of mitochondrial superoxide in live cells, as measured using the MitoSOX dye, showed that E significantly decreased mitochondrial superoxide production. We also measured the ratio of activities of aconitase to fumarase (A/F) as another indicator of mitochondrial ROS production in HBMEC. Aconitase, with an iron‐sulfur center, is inactivated by mitochondrial superoxide, while fumarase is unaffected. E increased the A/F ratio, indicating less mitochondrial ROS production. Subsequent enzyme reactivation with reducing agents showed that E had not affected total levels of aconitase activity, ruling out an effect of E on enzyme expression levels. A change in function of the mitochondrial antioxidant enzyme, manganese superoxide dismutase (MnSOD) could account for the ability of E to suppress superoxide production. However, there were no significant differences in MnSOD mRNA, protein levels, or enzyme activity after 24 hr E treatment. Future studies will explore mechanisms by which E may decrease mitochondrial ROS and protect against mitochondrial DNA damage. Nevertheless, the neuroprotective ability of E to decrease cerebral endothelial mitochondrial ROS may account for differences in stroke morbidity and mortality between men and women. Supported by NIH HL‐50775.