Summary: Dysfunction of the blood-brain barrier (BBB) is recognized as a key factor in the progression of neurodegenerative diseases (NDs), but the detailed mechanisms behind its pathogenesis and impact on neurodegeneration remain elusive. This study aimed to reveal the pathological effects of α-Synuclein (α-Syn), an aggregation protein in synucleinopathy, on BBB integrity and function and identify therapeutic targets for α-Syn-related vasculopathy. Using a brain endothelial cell model, we investigated the pathological effect of preformed fibril α-Syn (PFF) on BBB integrity, employing generative adversarial network (GAN) deep learning to analyze pathological changes. We found that PFF activates immune responses, increasing endothelial monolayer permeability via the TNF-α-NF-κB pathway. Further in vivo studies with PFF induced α-synucleinopathy, and a transgenic animal model (G2-3) revealed that α-Syn aggregation disrupts the BBB, leading to axonal degeneration that was mitigated by treatment with a non-BBB-penetrating TNF-α inhibitor, etanercept. These findings suggest that targeting brain endothelial TNF-α signaling could be a potential therapeutic approach for synucleinopathy-related NDs.
