Abstract TP273: Effect of Sex on Brain Microvascular Endothelial Cell Death Pathways: Impact of Hypoxia and Diabetes

Abstract

Diabetes exacerbates hemorrhagic transformation and worsens survival/recovery after ischemic stroke, especially in female patients. Brain microvascular endothelial cells (BMECs) are early targets in diabetes and ischemic injury. Differences in survival and reparative properties of BMECs may contribute to sex differences seen in stroke recovery. Recent evidence suggests that ferroptotic and necroptotic regulated cell death (RCD) mechanisms are activated in neurons after ischemic injury. Hypothesis: Diabetic conditions amplifies ferroptotic cell death and RCD pathways are differentially activated in female and male cells. Methods: Human male and female BMECs were cultured under normal and diabetic and hypoxic conditions. Cell viability, migration, barrier function and markers of apoptosis (Caspase-3), ferroptosis (IREB2), and necroptosis (RIPK3) were measured. Downstream signaling was assessed using ferroptosis and necroptosis inducers (erastin & TNFa) and inhibitors (Ferrostatin-1 & Necrostatin-1). Results: (Table): Hypoxia caused a greater decrease in cell viability and migration under diabetic conditions especially in male cells. Male cells had greater migratory properties. Diabetic conditions increased apoptotic, necroptotic, and ferroptotic gene expression in female BMECs, while only ferroptotic gene expression was increased in male BMECs. Ferroptosis was differentially regulated in male and females after erastin-1 challenge. Ferrostatin-1 inhibited erastin-induced death in male BMECs but it was not associated with expression of ferroptosis-related genes. Conclusion: Male BMECs are more susceptible to ferroptotic cell death than female BMECs. Diabetes and hypoxic conditions activate different RCD pathways and downstream signaling in male and female BMECs. Identification of sex and disease effects of RCD mechanisms in BMECS has the potential to develop vascular protection/restoration strategies for stroke recovery.