Netrin-5 Preserves Blood-Brain Barrier Integrity via Wnt3a/β-Catenin Pathway Activation in Murine Cerebral Ischemia

Blood-brain barrier compromise represents a pivotal pathological mechanism in ischemic stroke, driving neurological deterioration. Netrin-5, an axon guidance protein family member, demonstrates regulatory potential for BBB integrity. Employing middle cerebral artery occlusion (MCAO) mice and oxygen-glucose deprivation/reperfusion (OGD/R) in human brain microvascular endothelial cells (HBMVECs), we found that Netrin-5 was significantly downregulated in the murine cortex post-MCAO and was also downregulated in HBMVECs upon OGD/R exposure. Adenoviral Netrin-5 delivery in MCAO mice attenuated cerebral infarction, improved functional outcomes, reduced edema, and preserved BBB integrity, evidenced by diminished Evans blue extravasation and albumin leakage. Furthermore, Netrin-5 restored tight junction protein ZO-1 expression and activated Wnt3a/β-catenin signaling. In HBMVECs, Netrin-5 overexpression counteracted OGD/R-induced endothelial permeability, elevated transepithelial electrical resistance (TEER), and increased ZO-1, Wnt3a, and β-catenin levels. Critically, Wnt3a knockdown abrogated these protective effects, establishing Wnt3a/β-catenin signaling as indispensable for Netrin-5-mediated BBB preservation. In contrast, knockdown of Netrin-5 exacerbated BBB disruption in MCAO mice and increased endothelial permeability in HBMVECs. These results position Netrin-5 as a potential therapeutic intervention for ischemic stroke.