Single Cell Transcriptomic Profiling of Multiple System Atrophy Brain

多系统萎缩脑的单细胞转录组分析

• 批准号: 10799995
• 负责人: Un Jung Kang
• 金额: $ 59.33万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-04 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10799995
• 关键词: Affect; Age; Astrocytes; Atlases; Autopsy; Brain; Brain region; Cell Nucleus; Cell physiology; Cells; Cerebellar Diseases; Cerebellum; Classification; Clinical; Complex; Corpus striatum structure; Cytoplasmic Inclusion; Data; Data Set; Dysautonomias; Event; Exhibits; Experimental Models; Fluorescent in Situ Hybridization; Functional disorder; Gene Expression; Gene Expression Profile; Genes; Genetic Transcription; Human; Immunohistochemistry; In Situ Hybridization; Individual; Informatics; Mediating; Messenger RNA; Microglia; Modeling; Molecular; Molecular Profiling; Morphology; Multiple System Atrophy; Myelin; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Oligodendroglia; Olivopontocerebellar Atrophies; Parkinsonian Disorders; Pathologic; Pathologic Processes; Pathology; Pathway interactions; Patients; Phenotype; Physiology; Property; Sampling; Secondary to; Site; Stress; Striatonigral Degeneration; System; Techniques; Testing; Tissues; Up-Regulation; alpha synuclein; brain tissue; cell type; cerebral atrophy; differential expression; human tissue; insight; neuron loss; nonhuman primate; overexpression; proteostasis; putamen; response; single nucleus RNA-sequencing; single-cell RNA sequencing; synuclein; therapeutic target; transcriptome; transcriptomics

Project Summary/Abstract Multiple system atrophy (MSA) is a rare progressive neurodegenerative disease characterized by selective accumulation of α-synuclein in glial cytoplasmic inclusions (GCIs) within oligodendrocytes. Clinically, MSA patients present with various combinations of parkinsonism, cerebellar dysfunction, and dysautonomia. MSA is subclassified based on predominance of symptomology, which is associated with the primary site of neurodegeneration: MSA-P for parkinsonism and striatonigral degeneration or MSA-C for cerebellar features and olivopontocerebellar atrophy, though most cases involve both systems. The causes of α-synuclein accumulation within oligodendrocytes and the consequences for oligodendrocyte physiology in MSA are largely unknown. Likewise, how oligodendrocyte dysfunction causes neuronal death remains obscure. In this proposal, we will use single nucleus RNA sequencing (snRNA-seq) to generate transcriptomes of single cells from postmortem brain tissue from MSA patients to delineate the cell type specific transcriptional changes associated with MSA. We will probe striatal, cerebellar, and cortical tissue sets from the same patients for all patients of our sample set which contains both MSA-C and MSA-P cases. This allows us to capture the changes that occur in the primary site of pathology for each MSA subtype, striatum for MSA-P and cerebellum for MSA-C, along with the secondary sites, and a minimally affected brain region (cortex). In Aim 1, we will collect additional MSA cases and generate snRNA-seq profiles from all tissue sets. These data will be integrated and clustered to identify major cell types and subtypes from which informatic analysis of differentially-expressed genes will be used to identify regulatory networks and infer change in function. In Aim 2, we will validate the changes identified by snRNA-seq in tissue with immunohistochemistry and multiplexed fluorescence in situ hybridization with RNAscope, allowing the assessment of whether cells bearing dysregulated gene expression patterns have evidence of dysfunction. Initially focusing on oligodendrocytes, we will determine whether cells bearing GCI exhibit dysregulated transcriptomes, whether α-synuclein mRNA is overexpressed in these cells, and whether this affects myelin integrity. In Aim 3, we will test whether forced overexpression of α-synuclein in oligodendrocytes is sufficient to recapitulate the snRNA-seq profiles obtained from MSA tissues using a nonhuman primate MSA model. Upon completion, this proposal will generate an atlas of the MSA-dependent transcriptional changes of nearly all cell types in the striatum, cerebellum, and cortex, identify key alterations in gene expression and the pathways affected, determine whether α-synuclein expression is increased in cells with GCI, and whether de novo expression of α-synuclein mRNA fully recapitulates the cell states associated with MSA.

项目摘要/摘要 多系统萎缩（MSA）是一种罕见的进行性神经退行性疾病，其特征是选择性 α-突触核蛋白在神经胶质细胞包含（GCI）中的积累。临床上，MSA 患者出现了帕金森氏症，小脑功能障碍和功能障碍的各种组合。 MSA是 基于症状的占主导地位的子分类，这与 神经变性：用于帕金森氏症和纹状体变性的MSA-P或小脑特征的MSA-C 和橄榄球脑萎缩，尽管大多数情况都涉及这两个系统。 α-突触核蛋白的原因 在少突胶质细胞内积累，对MSA中的少突胶质细胞生理的后果是 在很大程度上未知。同样，少突胶质细胞功能障碍导致神经元死亡仍然晦涩难懂。在这个 提案，我们将使用单核RNA测序（SNRNA-Seq）生成单细胞的转录组 从MSA患者的死后脑组织到描绘细胞类型的特定转录变化 与MSA相关。我们将探测所有患者的纹状体，小脑和皮质组织。 我们的样本组患者均包含MSA-C和MSA-P病例。这使我们能够捕获 每种MSA亚型的病理学主要位点发生的变化，MSA-P和小脑的纹状体 对于MSA-C，以及次要部位以及受影响最小的大脑区域（皮层）。在AIM 1中，我们将 收集其他MSA病例，并从所有组织集中产生SnRNA-Seq曲线。这些数据将是 集成和聚集以识别主要的细胞类型和亚型 差异表达的基因将用于识别调节网络并推断功能变化。目标 2，我们将验证具有免疫组织化学和多重的组织中SnRNA-Seq鉴定的变化 与rnascope的荧光原位杂交，从而评估是否包含细胞 基因表达模式失调具有功能障碍的证据。最初专注于少突胶质细胞，我们 将确定带有GCI暴露失调转录组的细胞，α-突触核蛋白mRNA是否为 在这些细胞中过表达，这是否影响髓磷脂完整性。在AIM 3中，我们将测试是否强制 少突胶质细胞中α-核蛋白的过表达足以概括获得的SnRNA-Seq曲线 使用非人类灵长类动物MSA模型从MSA组织。完成后，该建议将产生 纹状体，小脑和 皮质，确定基因表达的键改变和受影响的途径，确定α-突触核蛋白是否是否 GCI细胞中的表达增加，以及α-突触核蛋白mRNA的从头表达是否完全 概括与MSA相关的细胞态。