Proteomic and Functional Analyses of Astrocyte Exosomes

 描述（由申请人提供）：星形胶质细胞外泌体的蛋白质组学和功能分析星形胶质细胞是中枢神经系统（CNS）中最丰富的细胞类型。在过去的十年中，越来越多的注意力已经支付给关键的稳态和不同的作用，星形胶质细胞在中枢神经系统的健康和疾病。星形胶质细胞在CNS生理学和病理学中的新兴影响已经通过星形胶质细胞分泌的因子如何影响CNS发育、突触传递、神经变性或感染而突出。因此，了解星形胶质细胞如何响应细胞外信号并受细胞外信号调节，有望确定支配CNS如何适应和响应损伤或炎症的基本机制。外泌体是一类非常小的细胞外囊泡，已知其由许多细胞类型分泌。最近的研究已经确定了外泌体在介导细胞信号传导和炎症反应中的关键功能。然而，人们对星形胶质细胞释放的外泌体在炎症反应中的功能知之甚少，而且以前的研究也没有详细说明星形胶质细胞外泌体中含有哪些蛋白质。我们知识中的这一差距将在本申请中解决如下：具体目标1将使用尖端蛋白质组学分级分离平台，在基础和炎症条件下生成从星形胶质细胞分离的外泌体内所含蛋白质的蛋白质组学图谱。具体目标2将确定星形胶质细胞外泌体作为体外和体内星形胶质细胞对炎症刺激反应的推定介体的功能。我们提供的初步数据表明，培养的星形胶质细胞在白细胞介素-1 β治疗后，外泌体的释放在定性和定量上都有增加。重要的是，我们提供的数据表明，星形胶质细胞衍生的外泌体存在于体循环中，并且可以从血清中收集和定量。我们假设外泌体代表了星形胶质细胞对炎症和神经病理条件反应的基本组成部分。这些研究的结果将为理解外泌体如何定义星形胶质细胞功能提供新的功能基础。这项工作将为科学界提供两个共同的成果：（1）我们将在基础和炎症条件下产生星形细胞外泌体的蛋白质组学图谱，这可能导致高保真的生物标志物鉴定，以及（2）我们将确定星形细胞外泌体在体外和体内模型中的功能作用。因此，这些研究可能会激发广泛的兴趣，星形胶质细胞衍生的外泌体作为介质的CNS中的细胞间通讯的健康和疾病。

 DESCRIPTION (provided by applicant): Proteomic and Functional Analyses of Astrocyte Exosomes Astrocytes are the most abundant cell type in the central nervous system (CNS). Over the past decade increasing attention has been paid to the critical homeostatic and diverse roles astrocytes play in the CNS health and disease. The emerging influence of astrocytes in CNS physiology and pathology has been highlighted by how factors secreted by astrocytes impact CNS development, synaptic transmission, neurodegeneration, or infection. Hence, understanding how astrocytes respond to and are regulated by extracellular cues is expected to identify fundamental mechanisms that govern how the CNS adapts and responds to injury or inflammation. Exosomes are a class of very small extracellular vesicles that are known to be secreted by many cell types. Recent studies have now identified critical functions for exosomes in mediating cell signaling and inflammatory responses. However, little is known about the function of exosomes released from astrocytes in response to inflammation and no previous study has detailed what proteins are contained in astrocytic exosomes. This gap in our knowledge will be addressed in this application as follows: Specific Aim 1 will generate a proteomic map of the proteins contained within exosomes isolated from astrocytes, under basal and inflammatory conditions, using a cutting edge proteomic fractionation platform. Specific Aim 2 will determine the function of astrocyte exosomes as putative mediators of astrocytic responses to inflammatory stimuli both in vitro and in vivo. We provide preliminary data that demonstrate a qualitative and quantitative increase in the release of exosomes from cultured astrocytes in response to treatment with interleukin-1beta. Importantly, we present data to indicate that astrocyte-derived exosomes are present within systemic circulation and can be collected and quantified from blood serum. We hypothesize that exosomes represent a fundamental component of the astrocytic response to inflammatory and neuropathological conditions. Results from these studies will provide a new functional basis to understand how exosomes define astrocyte function. This work will provide the scientific community with two shared outcomes: (1) we will produce a proteomic map of astrocytic exosomes under basal and inflammatory conditions that might lead to high fidelity biomarker identification, and (2) we will determine the functional role of astrocytic exosomes in in vitro and in vivo models. Thus, these studies may stimulate widespread interest in astrocyte-derived exosomes as mediators of intercellular communication in the CNS in health and disease.