TNFα-NF-κB activation through pathological α-Synuclein disrupts the BBB and exacerbates axonopathy

Dysfunction of the blood brain barrier (BBB) is recognized as a key factor in the progression of neurodegenerative diseases, 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 TNFα-NF-κB pathway. Further in vivo studies with PFF induced α-Synucleinopathy and 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 TNFα inhibitor, Etanercept. These findings suggest that targeting brain endothelial TNFα signaling could be a potential therapeutic approach for Synucleinopathy-related NDs. Primary human brain microvascular endothelial cells (HBMVECs) were cultured and treated with control, monomeric α-synuclein, or pre-formed fibrils (PFF) of α-synuclein for 24 or 72 hours. After treatment, RNA was extracted for transcriptomic analysis to investigate gene expression changes induced by different forms of α-synuclein.

血脑屏障（blood brain barrier, BBB）功能障碍被认定为神经退行性疾病进展的关键危险因素，但其发病的具体机制以及对神经退行性变的影响仍未阐明。本研究旨在明确突触核蛋白病（Synucleinopathy）中的聚集蛋白α-突触核蛋白（α-Synuclein, α-Syn）对BBB完整性与功能的病理作用，并筛选α-Syn相关血管病变的治疗靶点。
我们采用脑内皮细胞模型，结合生成式对抗网络（generative adversarial network, GAN）深度学习分析病理变化，探究预制纤维状α-突触核蛋白（preformed fibril α-Syn, PFF）对BBB完整性的病理影响。研究发现，PFF可激活免疫应答，通过肿瘤坏死因子α（TNFα）-核因子κB（NF-κB）通路增加内皮单层通透性。
进一步通过PFF诱导的突触核蛋白病模型与转基因动物模型（G2-3）开展的体内实验显示，α-Syn聚集会破坏BBB结构与功能，进而引发轴突变性；使用非血脑屏障穿透性TNFα抑制剂依那西普（Etanercept）进行干预，可缓解该病理进程。上述研究结果提示，靶向脑内皮细胞的TNFα信号通路，或可成为治疗突触核蛋白病相关神经退行性疾病的潜在策略。
我们培养了原发性人脑微血管内皮细胞（Primary human brain microvascular endothelial cells, HBMVECs），分别以对照组、单体α-突触核蛋白或预制α-突触核蛋白纤维（PFF）处理细胞24小时或72小时；处理结束后提取RNA，开展转录组学分析，以探究不同形式α-突触核蛋白诱导的基因表达变化。