Cell-Produced α-Synuclein Is Secreted in a Calcium-Dependent Manner by Exosomes and Impacts Neuronal Survival
Cell-Produced α-Synuclein Is Secreted in a Calcium-Dependent Manner by Exosomes and Impacts Neuronal Survival
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DOI：
10.1523/jneurosci.5699-09.2010
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发表时间：
2010-05-19
影响因子：
5.3
通讯作者：
Vekrellis, Kostas
中科院分区：
文献类型：
--
作者：
Emmanouilidou, Evangelia;Melachroinou, Katerina;Vekrellis, Kostas
α-突触核蛋白是帕金森病发病机制的核心。虽然最初α-突触核蛋白被认为是一种纯粹的细胞内蛋白，但最近的数据表明，它可以在人的血浆和CSF中以及神经元细胞的培养基中检测到。为了解决分泌的α-突触核蛋白在神经元稳态中的作用，我们已经产生了野生型α-突触核蛋白和β-半乳糖苷酶诱导的SH-SY 5 Y细胞。在这些细胞的条件培养基（CM）中容易检测到α-突触核蛋白的可溶性寡聚体和单体物质，其浓度与在人CSF中观察到的浓度相似。我们已经发现，在这个模型中，α-突触核蛋白是由外化囊泡以钙依赖的方式分泌的。电子显微镜和液相色谱-质谱蛋白质组学分析表明，这些囊泡具有外泌体的特征性标志，即多泡体的分泌型腔内囊泡。应用含有分泌的α-突触核蛋白的CM导致受体神经元细胞的细胞死亡，这可以在α-突触核蛋白从CM免疫耗竭后逆转。从该CM分离的高分子量和低分子量α-突触核蛋白种类显著降低细胞活力。重要的是，在应用前用寡聚体干扰化合物处理CM可使受体神经元细胞免于观察到的毒性。我们的研究结果首次表明，细胞产生的α-突触核蛋白是通过外泌体，钙依赖性机制分泌的，并表明α-突触核蛋白分泌有助于放大和传播帕金森病相关的病理。
alpha-Synuclein is central in Parkinson's disease pathogenesis. Although initially alpha-synuclein was considered a purely intracellular protein, recent data suggest that it can be detected in the plasma and CSF of humans and in the culture media of neuronal cells. To address a role of secreted alpha-synuclein in neuronal homeostasis, we have generated wild-type alpha-synuclein and beta-galactosidase inducible SH-SY5Y cells. Soluble oligomeric and monomeric species of alpha-synuclein are readily detected in the conditioned media (CM) of these cells at concentrations similar to those observed in human CSF. We have found that, in this model, alpha-synuclein is secreted by externalized vesicles in a calcium-dependent manner. Electron microscopy and liquid chromatography-mass spectrometry proteomic analysis demonstrate that these vesicles have the characteristic hallmarks of exosomes, secreted intraluminar vesicles of multivesicular bodies. Application of CM containing secreted alpha-synuclein causes cell death of recipient neuronal cells, which can be reversed after alpha-synuclein immunodepletion from the CM. High- and low-molecular-weight alpha-synuclein species, isolated from this CM, significantly decrease cell viability. Importantly, treatment of the CM with oligomer-interfering compounds before application rescues the recipient neuronal cells from the observed toxicity. Our results show for the first time that cell-produced alpha-synuclein is secreted via an exosomal, calcium-dependent mechanism and suggest that alpha-synuclein secretion serves to amplify and propagate Parkinson's disease-related pathology.