Table_4_Single-Cell Transcriptomics and In Situ Morphological Analyses Reveal Microglia Heterogeneity Across the Nigrostriatal Pathway.docx

Microglia are the resident immune effector cells of the central nervous system (CNS) rapidly reacting to various pathological stimuli to maintain CNS homeostasis. However, microglial reactions in the CNS may also worsen neurological disorders. Hence, the phenotypic analysis of microglia in healthy tissue may identify specific poised subsets ultimately supporting or harming the neuronal network. This is all the more important for the understanding of CNS disorders exhibiting regional-specific and cellular pathological hallmarks, such as many neurodegenerative disorders, including Parkinson’s disease (PD). In this context, we aimed to address the heterogeneity of microglial cells in susceptible brain regions for PD, such as the nigrostriatal pathway. Here, we combined single-cell RNA-sequencing with immunofluorescence analyses of the murine nigrostriatal pathway, the most affected brain region in PD. We uncovered a microglia subset, mainly present in the midbrain, displaying an intrinsic transcriptional immune alerted signature sharing features of inflammation-induced microglia. Further, an in situ morphological screening of inferred cellular diversity showed a decreased microglia complexity in the midbrain when compared to striatum. Our study provides a resource for the identification of specific microglia phenotypes within the nigrostriatal pathway, which may be relevant in PD.

小胶质细胞 (Microglia) 是中枢神经系统 (central nervous system, CNS) 的固有免疫效应细胞，可快速响应各类病理刺激以维持中枢神经系统稳态。然而，中枢神经系统内的小胶质细胞活化反应也可能加重神经功能障碍。因此，对健康组织中小胶质细胞的表型分析，或可识别出特定的预就绪亚群 (poised subsets)，这些亚群最终可对神经元网络起到支持或损伤作用。对于表现出区域特异性与细胞病理特征的中枢神经系统疾病（如众多神经退行性疾病，包括帕金森病 (PD)）的研究而言，这一点尤为关键。在此背景下，本研究旨在探究帕金森病易感脑区（如黑质纹状体通路 (nigrostriatal pathway)）内的小胶质细胞异质性。本研究针对帕金森病中受累最严重的脑区——小鼠黑质纹状体通路，整合了单细胞RNA测序 (single-cell RNA-sequencing) 与免疫荧光 (immunofluorescence) 分析。我们发现了一类主要存在于中脑的小胶质细胞亚群：该亚群具有内在的转录水平免疫警觉特征，且兼具炎症诱导型小胶质细胞的相关特性。此外，对推断的细胞多样性进行原位形态学筛查后发现，与纹状体 (striatum) 相比，中脑内的小胶质细胞复杂度有所降低。本研究为黑质纹状体通路内特定小胶质细胞表型的识别提供了研究资源，该表型或与帕金森病的发生发展相关。