The functional preservation of the central nervous system (CNS) is based on the neuronal plasticity and survival. In this context, the neuroinflammatory state plays a key role and involves the microglial cells, the CNS-resident macrophages. In order to better understand the microglial contribution to the neuroprotection, microglia-derived extracellular vesicles (EVs) were isolated and molecularly characterized to be then studied in neurite outgrowth assays. The EVs, mainly composed of exosomes and microparticles, are an important cell-to-cell communication process as they exhibit different types of mediators (proteins, lipids, nucleic acids) to recipient cells. The medicinal leech CNS was initially used as an interesting model of microglia/neuron crosstalk due to their easy collection for primary cultures. After the microglia-derived EV isolation following successive methods, we developed their large-scale and non-targeted proteomic analysis to (i) detect as many EV protein markers as possible, (ii) better understand the biologically active proteins in EVs and (iii) evaluate the resulting protein signatures in EV-activated neurons. The EV functional properties were also evaluated in neurite outgrowth assays on rat primary neurons and the RNAseq analysis of the microglia-derived EVs was performed to propose the most representative miRNAs in microglia-derived EVs. This strategy allowed validating the EV isolation, identify major biological pathways in EVs and corroborate the regenerative process in EV-activated neurons. In parallel, six different miRNAs were originally identified in microglia-derived EVs including 3 which were only known in plants until now. The analysis of the neuronal proteins under the microglial EV activation suggested possible miRNA-dependent regulation mechanisms. Taken together, this combination of methodologies showed the leech microglial EVs as neuroprotective cargos across species and contributed to propose original EV-associated miRNAs whose functions will have to be evaluated in the EV-dependent dialog between microglia and neurons.

中文翻译：

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小胶质细胞外小泡的分离：迈向miRNA签名和神经保护。

中枢神经系统（CNS）的功能保存基于神经元的可塑性和生存能力。在这种情况下，神经炎症状态起关键作用，并涉及小胶质细胞，即中枢神经系统驻留巨噬细胞。为了更好地了解小胶质细胞对神经保护的作用，分离了小胶质细胞外囊泡（EVs）并进行了分子表征，然后在神经突生长试验中进行研究。电动汽车主要由外泌体和微粒组成，是重要的细胞间通讯过程，因为它们对受体细胞表现出不同类型的介体（蛋白质，脂质，核酸）。药用水primary中枢神经系统最初被用作小胶质细胞/神经元串扰的有趣模型，因为它们易于收集用于原代培养。在采用连续方法分离出小胶质细胞的EV后，我们开发了它们的大规模且无目标的蛋白质组学分析方法，以（i）尽可能多地检测EV蛋白标记，（ii）更好地了解EV中的生物活性蛋白，以及（iii ）评估EV激活神经元中产生的蛋白质标记。还在大鼠原代神经元的神经突生长试验中评估了EV的功能特性，并对小胶质细胞来源的电动汽车进行了RNAseq分析，以提出小胶质细胞来源的电动汽车中最具代表性的miRNA。这种策略可以验证电动汽车的隔离性，确定电动汽车的主要生物学途径，并证实电动汽车激活的神经元的再生过程。在平行下，最初在小胶质细胞来源的电动汽车中鉴定出6种不同的miRNA，其中3种直到现在才在植物中才知道。在小胶质细胞EV激活下对神经元蛋白的分析表明可能存在miRNA依赖性调节机制。综上所述，这种方法的组合显示了水ech小胶质细胞电动汽车可作为跨物种的神经保护货物，并有助于提出与电动汽车相关的原始miRNA，其功能必须在小胶质细胞和神经元之间的电动汽车依赖性对话中进行评估。