CysLT受体在小胶质细胞介导的帕金森病神经炎症及神经元损伤中的作用

Neuroinflammation induced by microglial activation plays a critical role in the pathological processes of Parkinson’s disease (PD). Autophagy is involved in microglial activation and inflammation. 5-lipoxygenase (5-LOX) regulates PD microglial inflammation and autophagy. 5-LOX products cysteinyl leukotrienes (CysLTs) are potent proinflammatory mediators; and their actions are mediated by activating CysLT receptors. Modulation of microglial activation by CysLT receptors has been demonstrated in brain injury after cerebral ischemia. Nevertheless, whether CysLT receptors regulate microglial inflammation and neuronal injury associated with PD remains poorly unknown. Herein, we plan to study following problems：1) To explore whether CysLT receptors regulate microglia-mediated PD-like neuroinflammation and neuronal injury using in vitro cultured primary microglial cells and mouse microglial cell line BV2 cells as well as dopamine (DA) neurons; whether CysLT receptor activation promote microglial inflammation through activating autophagy, and what subtype(s) of CysLT receptors is involved in microglial modulation and what signal transduction occurs after CysLT receptor activation. 2) To reveal whether the activation of CysLT receptor is related to microglial autophagy, inflammation and DA neuronal damage in the mouse model of PD. 3) To observe the potential effects of interfering with microglial CysLT receptor-autophagy-inflammation pathways on neuroinflammation and DA neuronal injury in PD. Our goal is to study the possibility that CysLT receptors represent a potential therapeutic target for PD.

小胶质细胞激活诱导的神经炎症在帕金森病（PD）病理过程中起关键作用。自噬参与小胶质细胞炎症。5-脂氧酶（5-LOX）调节PD小胶质细胞炎症和自噬。5-LOX产物半胱氨酰白三烯（CysLT）是重要炎症介质，通过激动CysLT受体发挥生物效应。CysLT受体调节脑缺血后小胶质细胞激活已得到证明，但其与PD的关系尚未阐明。本项目拟研究：1）在原代小胶质细胞、BV2小胶质细胞和多巴胺（DA）神经元，阐明CysLT受体在小胶质细胞介导的PD神经炎症及神经元损伤中的调控作用，CysLT受体是否调节自噬进而调节小胶质细胞炎症，参与调节的受体亚型及信号特点；2）在PD动物模型，验证CysLT受体与小胶质细胞自噬、炎症反应及DA神经元损伤的关系；3）评价基因或药物手段干预CysLT受体-自噬-炎症通路对PD神经炎症和DA神经元损伤的作用及机制。本项目旨在阐明CysLT受体作为PD神经保护新靶点的意义。

小胶质细胞激活诱导的神经炎症在帕金森病（Parkinson’s disease, PD）病理过程中起关键作用。自噬机能障碍参与小胶质细胞炎症。CysLTs是重要炎症介质，通过激动CysLT受体发挥生物效应。CysLT受体调节脑缺血后小胶质细胞激活和神经炎症已得到证明，但其与PD的关系尚未阐明。本研究以体外培养的BV2小胶质细胞，原代小胶质细胞，神经元细胞以及PD动物为研究对象，建立了PD样小胶质细胞激活及神经元损伤模型，研究了CysLT受体在介导小胶质细胞激活、神经炎症以及调控神经元损伤中的关键作用。研究结果显示，在小胶质细胞条件培养液转移共培养或小胶质-神经元Transwell共培养模型，LPS和MPP+诱导体外培养的小胶质细胞激活、炎症细胞因子和氧化应激产物增加，并进一步导致神经元变性死亡。CysLT1受体和CysLT2受体拮抗剂（montelukast和HAMI3379）抑制LPS和MPP+诱导的小胶质细胞炎症及神经毒性。CysLT1受体和CysLT2受体基因敲低（CysLT1 siRNA，CysLT2 shRNA）产生相似的作用。在PD动物模型，PD神经毒素MPTP诱导小鼠黑质小胶质细胞激活，多巴胺（DA）神经元死亡，纹状体DA含量下降和小鼠行为学障碍。应用CysLT受体拮抗剂抑制MPTP诱导的小胶质细胞激活和DA神经元死亡，并一定程度改善小鼠行为学能力。在细胞和动物实验，CysLT1受体拮抗剂montelukast显示比CysLT2受体拮抗剂HAMI3379更强的抗炎和神经保护作用。此外，CysLT受体拮抗剂也抑制毒素诱导的小胶质细胞自噬底物SQSTM1/p62表达，改善自噬流，提示自噬调节参与CysLT受体拮抗剂的神经保护机制。本项目从新的角度阐明了CysLT受体介导PD发病中小胶质细胞激活和DA神经元死亡的功能作用，提示阻断CysLT受体可能作为一个有希望的PD治疗策略。