Abstract WP015: Hypoxia and Ferroptosis: A Double Hit Driving Pericyte Dysfunction and Pathological Transition After Stroke.

Pericyte regulates cerebral blood flow, promotes blood–brain barrier (BBB) integrity, and orchestrates reparative angiogenesis after stroke. Ferroptosis, an iron-dependent regulated cell death pathway, has been implicated in driving pericyte dysfunction and pathological transition of healthy type-1 pericytes (PC1) into type-2 pericytes (PC2). PC2 are characterized by upregulated expression of inflammatory, BBB-disruptive, and phagocytic genes, as well as enhanced Alpha-smooth muscle actin (α-SMA) expression, all of which worsen stroke outcomes. We proposed that a combined insult of hypoxia and ferroptosis accelerates the PC1-to-PC2 transition and aggravates stroke injury. To test our hypothesis, Cre-lox P mice expressing rhodamine red in pericytes underwent a transient ferric chloride/middle cerebral artery occlusion (MCAO) stroke model. Ferroptosis was inhibited in select groups with the iron chelator deferoxamine (I.P 100 mg/kg body weight) daily for 5 days after stroke. Infarct size, edema, and motor function (grip strength test) were assessed post-stroke. In vitro, human brain pericytes were exposed to hypoxia with or without ferric chloride to induce ferroptosis, confirmed by lipid peroxidation assays. Immunohistochemical studies were used to assess pericyte transition (α-SMA expression), and pericyte dysfunction (BBB integrity, tight junction proteins, and pericyte capillary coverage). Results: Hypoxia increased ferroptosis-induced PC1-to-PC2 transition, evidenced by elevated α-SMA expression in pericytes (P