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Cellular and Genetic Origins of Astrocytes

星形胶质细胞的细胞和遗传起源

基本信息

批准号：
8214621
负责人：
DAVID H ROWITCH
金额：
$ 72.57万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-02-01 至 2014-01-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): The human brain is made up of neurons and glia. Glia comprise about 90% of brain cells and are involved in devastating disorders such as multiple sclerosis, seizures and Alzheimer's disease. We know little about development of the most prevalent glial cells called astrocytes. This proposal will identify fundamental mechanisms by which neural stem cells give rise to astrocytes. It is also intended to generate useful markers to investigate roles for astrocytes during development and in human disease. The goal of the proposed research is to identify a set of genetic markers for development of astrocytes and to map their cellular origins in the central nervous system. The underlying hypothesis to be tested is that astrocytes develop from heterogeneous locations in the developing CMS. We have three specific aims: Specific Aim 1 is to identify and characterize activity of transcription factors that uniquely mark and may regulate astrocyte development. In preliminary work, we have interrogated the mammalian transcriptome to identify transcription factors that co-localize with astrocytes in the developing spinal cord. Expression analysis and gain- and Ioss-of-function screens will be conducted to prioritize generation of antibodies to transcription factors that mark and may regulate astrocyte lineage heterogeneity in the CNS. Specific Aim 2 is to develop cell type- and stage-specific markers for fibrous and protoplasmic astrocytes. We have developed fluorescent activated cell sorting (FACS) protocols for acute harvest of gray matter astrocytes and analysis by expression profiling. By sorting cells from Gensat transgenic mice that express green fluorescent protein (GFP) in (1) pan-astroglial and (2) white matter astrocyte compartments, we will identify markers specific for fibrous astrocytes, protoplasmic astrocytes and astrocyte precursors in various CNS regions. Specific Aim 3 is to determine whether astrocyte diversity correlates with developmental site of origin in the embryonic spinal cord and brain. We will use a cohort of cre/lox transgenic mice to fate map heterogeneous progenitor domains for astrocytes and their ultimate cellular origins in the embryonic CNS. Based on information obtained in Aims1/2, we will generate a floxed conditional reporter transgenic mouse Iine that will express the FACS-selectable marker GFP exclusively in astroglia. This will enable acute purification of astrocyte subsets for further discrimination of astrocyte heterogeneity. PUBLIC HEALTH RELEVANCE These studies will elucidate the mechanisms by which neural stem cells give rise to astrocytes and provide useful genetic tools for understanding the diverse roles of astrocytes in human development, with practical implications for diagnosis and treatment of human disorders such as epilepsy, brain cancer and neurodegeneration.

描述（由申请人提供）：人脑由神经元和神经胶质组成。神经胶质细胞占脑细胞的90%，与多发性硬化症、癫痫和阿尔茨海默病等破坏性疾病有关。我们对最常见的神经胶质细胞星形胶质细胞的发育知之甚少。这项提案将确定神经干细胞产生星形胶质细胞的基本机制。它还旨在产生有用的标记物，以研究星形胶质细胞在发育和人类疾病中的作用。这项研究的目的是确定一组星形胶质细胞发育的遗传标记，并绘制它们在中枢神经系统中的细胞起源。待检验的基本假设是星形胶质细胞从发育中的CMS中的异质位置发育。我们有三个具体目标：具体目标1是识别和表征独特标记和调节星形胶质细胞发育的转录因子的活性。在前期工作中，我们已经询问了哺乳动物的转录组，以确定与发育中的脊髓星形胶质细胞共定位的转录因子。将进行表达分析和功能获得和丧失筛选，以优先生成标记并可能调节CNS中星形胶质细胞谱系异质性的转录因子的抗体。具体目标2是开发纤维和原生质星形胶质细胞的细胞类型和阶段特异性标志物。我们已经开发了荧光激活细胞分选（FACS）方案急性收获的灰质星形胶质细胞和分析的表达谱。通过分选Gensat转基因小鼠中在（1）泛星形胶质细胞和（2）白色星形胶质细胞区室中表达绿色荧光蛋白（GFP）的细胞，我们将鉴定不同CNS区域中纤维星形胶质细胞、原生质星形胶质细胞和星形胶质细胞前体的特异性标志物。具体目标3是确定星形胶质细胞的多样性是否与胚胎脊髓和大脑的发育部位有关。我们将使用一组cre/lox转基因小鼠的命运映射星形胶质细胞及其最终的细胞起源在胚胎中枢神经系统的异质祖域。基于在目的1/2中获得的信息，我们将产生一个floxed条件性报告基因转基因小鼠品系，其将仅在星形胶质细胞中表达FACS选择性标记GFP。这将使急性纯化的星形胶质细胞亚群进一步辨别星形胶质细胞的异质性。这些研究将阐明神经干细胞产生星形胶质细胞的机制，并为理解星形胶质细胞在人类发育中的不同作用提供有用的遗传工具，对癫痫、脑癌和神经变性等人类疾病的诊断和治疗具有实际意义。