Department of Neurology and Clinical Research Center of Neurological Disease, The Second Affiliated Hospital of Soochow University, Suzhou, 215004, Jiangsu, China.
Jiangsu Key Laboratory of Neuropsychiatric Diseases and Institute of Neuroscience, Soochow University, Suzhou, Jiangsu, 215123, China.
J Neuroinflammation. 2024 Oct 26;21(1):275. doi: 10.1186/s12974-024-03268-4.
Autophagy dysfunction in glial cells is implicated in the pathogenesis of Parkinson's disease (PD). The previous study reported that α-synuclein (α-Syn) disrupted autophagy in cultured microglia. However, the mechanism of microglial autophagy dysregulation is poorly understood.
Two α-Syn-based PD models were generated via AAV-mediated α-Syn delivery into the mouse substantia nigra and striatal α-Syn preformed fibril (PFF) injection. The levels of microglial UNC-51-like kinase 1 (Ulk1) and other autophagy-related genes in vitro and in PD mice, as well as in the peripheral blood mononuclear cells of PD patients and healthy controls, were determined via quantitative PCR, western blotting and immunostaining. The regulatory effect of signal transducer and activator of transcription 1 (STAT1) on Ulk1 transcription was determined via a luciferase reporter assay and other biochemical studies and was verified through Stat1 knockdown or overexpression. The effect of α-Syn on glial STAT1 activation was assessed by immunohistochemistry and western blotting. Changes in microglial status, proinflammatory molecule expression and dopaminergic neuron loss in the nigrostriatum of PD and control mice following microglial Stat1 conditional knockout (cKO) or treatment with the ULK1 activator BL-918 were evaluated by immunostaining and western blotting. Motor behaviors were determined via open field tests, rotarod tests and balance beam crossing.
The transcription of microglial ULK1, a kinase that controls autophagy initiation, decreased in both in vitro and in vivo PD mouse models. STAT1 plays a critical role in suppressing Ulk1 transcription. Specifically, Stat1 overexpression downregulated Ulk1 transcription, while Stat1 knockdown increased ULK1 expression, along with an increase in LC3II and a decrease in the SQSTM1/p62 protein. α-Syn PFF caused toll-like receptor 4-dependent activation of STAT1 in microglia. Ablation of Stat1 alleviated the decrease in microglial ULK1 expression and disruption of autophagy caused by α-Syn PFF. Importantly, the ULK1 activator BL-918 and microglial Stat1 cKO attenuated neuroinflammation, dopaminergic neuronal damage and motor defects in PD models.
These findings reveal a novel mechanism by which α-Syn impairs microglial autophagy and indicate that targeting STAT1 or ULK1 may be a therapeutic strategy for PD.
神经胶质细胞中的自噬功能障碍与帕金森病（PD）的发病机制有关。先前的研究报道，α-突触核蛋白（α-Syn）破坏了培养的小胶质细胞中的自噬。然而，小胶质细胞自噬失调的机制尚不清楚。
通过 AAV 介导的α-Syn 递送至小鼠黑质和纹状体α-Syn 预形成纤维（PFF）注射，生成了两种基于α-Syn 的 PD 模型。通过定量 PCR、Western blot 和免疫染色法测定体外和 PD 小鼠以及 PD 患者和健康对照者外周血单个核细胞中微胶质细胞 UNC-51 样激酶 1（Ulk1）和其他自噬相关基因的水平。通过荧光素酶报告基因测定和其他生化研究确定信号转导和转录激活因子 1（STAT1）对 Ulk1 转录的调节作用，并通过 Stat1 敲低或过表达进行验证。通过免疫组化和 Western blot 评估α-Syn 对神经胶质细胞 STAT1 激活的影响。通过免疫染色和 Western blot 评估微胶质细胞 Stat1 条件性敲除（cKO）或使用 ULK1 激活剂 BL-918 治疗后 PD 和对照小鼠黑质纹状体中微胶质细胞状态、促炎分子表达和多巴胺能神经元丢失的变化。通过旷场试验、旋转棒试验和平衡木穿越试验测定运动行为。
体外和体内 PD 小鼠模型中，控制自噬起始的微胶质细胞 ULK1 激酶的转录均降低。STAT1 在抑制 Ulk1 转录中起关键作用。具体来说，Stat1 过表达下调了 Ulk1 转录，而 Stat1 敲低增加了 ULK1 的表达，同时 LC3II 增加，SQSTM1/p62 蛋白减少。α-Syn PFF 引起小胶质细胞 TLR4 依赖性 STAT1 激活。Stat1 缺失减轻了α-Syn PFF 引起的小胶质细胞 ULK1 表达降低和自噬破坏。重要的是，ULK1 激活剂 BL-918 和微胶质细胞 Stat1 cKO 减轻了 PD 模型中的神经炎症、多巴胺能神经元损伤和运动缺陷。
这些发现揭示了α-Syn 损害小胶质细胞自噬的新机制，并表明靶向 STAT1 或 ULK1 可能是 PD 的一种治疗策略。
