Understanding the role of astrocyte-derived extracellular vesicles in health and disease

我们的目标是构建一个全面的、多组学的星形胶质细胞分泌组(包括RNA/蛋白质/代谢物)，重点是细胞外小泡(EVS)的特征。星形胶质细胞EVS的含量将与非EV分泌部分(即细胞裂解物中的细胞内分析物)进行比较。然后，我们将确定星形胶质细胞衍生的EV在健康和疾病中调节星形胶质细胞反应性和神经元-星形胶质细胞通讯的机制。与神经退行性变广泛相关的是，我们将重点研究与肌萎缩侧索硬化症(ALS)相关的蛋白质(例如RNA结合蛋白、泛素结合蛋白)，并确定ALS突变在多大程度上会导致EV的加载和分泌功能障碍。为此，ALS的影响将通过自噬或氧化应激的遗传/药理学操作来模拟。将对神经元和星形胶质细胞进行分子和细胞评估，以进一步仔细检查功能障碍的EVS对反应性星形胶质细胞相关通路的影响(如转录调节、代谢应激等)。以及神经元-星形胶质细胞的通讯(如神经元死亡、轴突长度和星形胶质细胞摄取谷氨酸)。最后，我们将调查EVS是否代表具有独立于供体细胞发挥功能的‘代谢功能单位’，作为表型变化和治疗靶点的潜在生物标记物。

We aim to construct a comprehensive, multi-omics astrocyte secretome (including RNA/proteins/metabolites) with the focus on the characterisation of extracellular vesicles (EVs). The content of astrocyte EVs will be compared to that of non-EV secreted fraction (i.e. intracellular analytes in cell lysates). We will then determine the mechanism by which astrocyte-derived EVs regulate astrocyte reactivity and neuron-astrocyte communication in health and disease. Broadly relevant to neurodegeneration, we will focus on proteins associated to amyotrophic lateral sclerosis (ALS) (e.g. RNA binding, ubiquitin-binding proteins) and determine to what extend ALS mutations can result in dysfunction of the loading and secretion of EVs. To this end, the effect of ALS will be mimicked using genetic/pharmacological manipulation of autophagy or oxidative stress. Molecular and cellular assessments of neuron and astrocytes will be performed to further scrutinise the effect of dysfunction EVs on relevant pathways in reactive astrocytes (e.g. transcriptional regulation, metabolic stress, etc...) and neuron-astrocyte communication (e.g. neuronal death, axon length and glutamate uptake in astrocytes). Finally, we will investigate whether EVs represent 'metabolically functional units' with the ability to function independently of donor cells, as potential for biomarkers of phenotypic changes and therapeutic target.