外周突触核蛋白通过血脑屏障激活小胶质细胞的分子机制研究

It is quite clear thatα-synuclein is essential in neurodegeneration of synucleinopathies including Parkinson disease (PD). To date, most of studies have been focused on the neurotoxicity of α-synuclein aggregates derived from neurons, whereas few attentions have been paid to the α-synuclein present in the extracelluar space or even peripherally derived α-synuclein. In this regard, our recent studies showed that peripheral synuclein, of which the concentrations are hundreds of times higher than those in the CNS, could enter the CNS, regional specifically, especially when the blood brain barrier (BBB) was damaged. Additionally,exogenous α-synuclein has been reported to activate microglia and stimulate their directional migration toward the source of α-synuclein. Therefore, we hypothesize that peripheral α-synuclein may get into the CNS via the damaged BBB, followed by stimulating microglial activation and initiating neuroinflammation that is highly related to PD development and progression. To test our hypotheses, we design a variety of in vitro and in vivo experiments, aiming at: 1) verifying the ability of peripheral α-synuclein to enter the CNS crossing the BBB, and ascertaining the distribution pattern of α-synuclein that moves into different regions in the brain; 2) determining the predominant form(s) (e.g. monomers, oligomers or polymers) of α-synuclein that gets access to the CNS; 3) confirming whether peripheral α-synuclein is capable of recruiting activated microglia; 4) elucidating the mechanisms by which peripheral α-synuclein activates microglia once it gets into the CNS. This proposal will potentially uncover the cellular and molecular basis that peripheral α-synuclein stimulates microglia to cause neuroinflammation, likely establishing a novel theoretical foundation for the early diagnosis and therapy for synucleinopathies like PD.

α型突触核蛋白在帕金森病(PD)中的作用已比较明确，但是以往研究多关注神经元中的α型突触核蛋白产生错义折叠及沉积所造成的神经毒性作用。近期实验表明，细胞外α型突触核蛋白可激活小胶质细胞并使之趋化迁移，引起神经炎症反应，促进PD发生和发展。我们最近研究发现，外周α型突触核蛋白不但可进入神经中枢，而且在血脑屏障受损时、PD敏感脑区选择性明显增加。因此我们推测: 外周α型突触核蛋白可透过开放的血脑屏障进入中枢, 进而激活小胶质细胞并引发PD相关的炎症反应。为此，我们将开展系列体内体外研究: 进一步明确不同形式的外周α型突触核蛋白穿过血脑屏障进入中枢及其在不同脑区的分布情况；阐明外周α型突触核蛋白进入中枢的优先形式；确定外周α型突触核蛋白能否吸引并激活小胶质细胞；分析外周α型突触核蛋白活化小胶质细胞的分子机制。本研究将揭示外周源性α型突触核蛋白进入中枢激活小胶质细胞的机制, 为诊断和治疗提供依据。

背景：小胶质细胞功能对于维持大脑健康至关重要，其激活是神经退行性变的重要组成部分。在伤害或疾病条件下引起“反应性”或“炎症性”表型的因素已有大量研究。暴露于聚集的或低聚形式的α-突触核蛋白（一种丰富的脑蛋白）就是其中一个因素，在驱动小胶质细胞活化中起着至关重要的作用。包括在路易氏体（LB）疾病（如帕金森氏病）中趋化性迁移和炎性介质的产生。另一方面，人们越来越认识到小胶质细胞也会发生变化，取决于细胞环境，其主要促进重建和抗炎功能，即在生理状态下小胶质细胞的大多数所需功能。维持小胶质细胞处于这种生理状态的本质是未知的。.方法：在这项研究中，我们使用体外和体内模型，用LPS +IFN-γ，IL-4 + IL-13，α-突触核蛋白单体和α-突触核蛋白寡聚体对原发性小胶质细胞或BV2小胶质细胞进行了刺激，并通过RT-PCR，Western印迹，ELISA，IF，IHC，Co-IP分析了小胶质细胞表型和潜在机制。.结果：我们描述了α-突触核蛋白的新生理功能，其中它通过与细胞外信号调节激酶（ERK）相互作用和募集ERK，核因子κB（NF-κB）来调节小胶质细胞向抗炎表型的方向以及过氧化物酶体增殖物激活的受体γ（PPARγ）途径。.结论：这些发现表明，鉴于脑组织中α-突触核蛋白的丰富性和多功能性，以前未知的单体α-突触核蛋白功能，可能为路易氏体相关疾病的发病机理和潜在疗法及其他方面提供了新的见解。