Abstract P802: Potential Sex Differences in Endothelial Cell Death Pathways: Relevance to Stroke Recovery

Abstract

Introduction: We have shown that iron chelation with deferoxamine (DFX) prevents stroke-induced cerebral vasoregression and improves sensorimotor and cognitive deficits after ischemic stroke in diabetic male rats. Diabetic female rats, on the other hand, develop greater hemorrhagic transformation (HT) but no vasoregression and DFX improves outcomes. HT breakdown products like hemin contribute significantly in secondary inflammation and neuronal death. Given the importance of brain microvascular endothelial cells (BMVECs) on neurovascular restoration in stroke recovery and our previous findings of increased ferroptotic cell death in endothelial cells from male animals, the objectives of this study were to determine 1) the impact of hemin on ferroptotic death in BMVECs of female and male origin, and 2) effect of DFX on ferroptosis. Methods: Male (HBEC5i) and female (BMECd3) human endothelial cell lines were grown in normal conditions. In some experiments cells were treated with hemin (50μM) for 6 hours with and without ferroptosis inhibitor ferrostatin-1 (20μM) or DFX (100μM). Cell viability (MTT and CytoSelect assays), ferroptosis marker proteins and lipid peroxidation (LPO) were assessed. by immunoblotting and, respectively. Results: GPX4 and NRF2 content was decreased with hemin in both sexes while 4HNE immunofluroscence for LPO, a key feature of ferroptosis, was increased. DFX did not prevent hemin-induced changes in GPX4 and ferritin heavy chain in female cells while it was effective in male cells. DFX and ferrostatin-1 improved cell viability in both male and female BMVECs. There were differences in ferrostatin 1 and DFX effects (Table). Conclusions: These results suggest there may be sex differences in how ferroptosis occurs in response to hemin. Better understanding of endothelial ferroptotic cell death regulation and the effect of iron chelation on cell survival can provide novel insights with respect to sex differences in stroke recovery.