Molecular control of neurogenesis in the adult subventricular zone

成人室下区神经发生的分子控制

• 批准号: 8240502
• 负责人: KENNETH J CAMPBELL
• 金额: $ 45.3万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2015-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8240502
• 关键词: Address; Adult; Alleles; Animals; Ara-C; Astrocytes; Biological Assay; Biology; Brain; Brain Injuries; Brain Pathology; Cell Culture Techniques; Cell Differentiation process; Cell division; Cells; Corpus striatum structure; Data; Deoxyuridine; Development; Embryo; Gene Expression Profile; Generations; Goals; Growth Factor; Health; Heterogeneity; Human; In Vitro; Injury; Interneurons; Knock-out; Label; Life; Light; Location; Maintenance; Mediating; Mitotic; Molecular; Molecular Genetics; Mus; Natural regeneration; Neurobiology; Neuroglia; Neurology; Neuronal Differentiation; Neurons; Oligodendroglia; Outcome; Outcome Study; Physiological; Physiology; Play; Production; Property; Proteins; Quinolinic Acid; Research; Role; Social Welfare; Source; Specificity; Stream; Testing; Tetanus Helper Peptide; Transgenes; Transgenic Organisms; Travel; Virus; adult neurogenesis; base; chromatin immunoprecipitation; gain of function; genetic analysis; genome-wide; gliogenesis; homeodomain; in vivo; innovation; insight; lateral ventricle; loss of function; molecular marker; nerve stem cell; neuroblast; neurogenesis; neuronal replacement; novel; olfactory bulb; overexpression; progenitor; promoter; public health relevance; repaired; research study; subventricular zone; transcription factor

DESCRIPTION (provided by applicant): Active production of new neurons and glia continues in restricted regions of the adult mammalian brain, including the subventricular zone (SVZ) lining the lateral ventricle. In these so-called neurogenic niches, neural stem cells (NSCs) and other progenitors (collectively called NPCs) persist throughout life and serve as the source of new neurons and glia. Recent studies have revealed that this continuous neurogenesis in adults contribute to physiology and pathology of the brain. Moreover, regenerating specific types of neurons after damage by manipulating endogenous NPCs is one of the important goals in restorative neurology. Yet, our understanding of the mechanisms underlying this important phenomenon is still very limited, and further research is imperative. In this proposal, we will address this issue by studying the roles for the homeodomain transcription factors Gsx1 and Gsx2 in adult neurogenesis. Gsx1 and Gsx2 control many aspects of telencephalic development, including regional specification and proliferation of NPCs, and neuronal differentiation. Their function in adults, however, has not yet been investigated. Our preliminary data show that Gsx1 and Gsx2 are expressed in specific subpopulations of NPCs that reside in anatomically discrete subdomains of the adult SVZ. We hypothesize that Gsx1 and Gsx2 play crucial roles in controlling the maintenance and/or differentiation of discrete subpopulations of NPCs in the adult SVZ, thereby contributing to the generation of specific OB interneuron subtypes. We also hypothesize that Gsx1+ and Gsx2+ progenitors contribute to injury-induced neurogenesis in the adult brain. We will test these hypotheses by experiments outlined in the following four specific aims: Aim 1: To reveal the identity of Gsx1+ and Gsx2+ progenitors in the adult SVZ. Our preliminary data show that Gsx1 and Gsx2 are expressed in NPCs that reside in discrete subdomains of the adult SVZ. We will reveal the properties of these cells by 1) detailed molecular marker analysis using Gsx2flox/+ and Gsx1::GFP mice, 2) short-term and long-term labeling with the mitotic marker 5-bromo-2'deoxyuridine (BrdU) combined with treatment with the anti-mitotic agent b-D-arabinofuranoside (Ara-C) and various growth factors, and 3) clonal cell culture assays. Aim 2: To reveal the roles for Gsx1 and Gsx2 in neurogenesis and gliogenesis in the adult SVZ. To reveal the adult-specific function of Gsx1 and Gsx2, we will perform conditional gain-of-function (GOF) and loss-of-function (LOF) analyses in vivo. Mice carrying floxed alleles of Gsx1 and Gsx2 will be used for progenitor subtype-specific knockout in the adult brain. We will also perform conditional GOF experiments using mice carrying tet-O promoter-driven Gsx1 and Gsx2 transgenes. Virus-mediated overexpression and knockdown of Gsx1 and Gsx2 will be performed in parallel. Aim 3: To reveal the function of Gsx1 and Gsx2 using in vitro culture. To better understand the mechanisms of action of Gsx1 and Gsx2, we will perform GOF and LOF analyses using clonal culture of NPCs. We will also identify the transcriptional targets for Gsx1 and Gsx2 by genome-wide chromatin-immunoprecipitation (ChIP) and transcriptome assays. Aim 4: To reveal the roles for Gsx1 and Gsx2 in injury-induced neurogenesis in the adult brain The mechanisms underlying injury-induced neurogenesis in the adult brain remain largely unknown. We will examine if Gsx1+ and Gsx2+ progenitors participate in ectopic neurogenesis following quinolinic acid (QA)-induced excitotoxic injury in the striatum. We will also determine if Gsx1 and Gsx2 play any role in injury-dependent control of SVZ progenitors and neurogenesis through GOF and LOF analyses using the aforementioned transgenic and virus-mediated approaches. Revealing the roles of Gsx1/2 in the adult brain will provide novel insights into the mechanisms for adult neurogenesis, in particular, the molecular basis for the specificity of adult NPCs. Thus, this study will significantly advance our understanding of adult neurogenesis and the biology of NPCs. Given the importance of adult neurogenesis in physiology and pathology of the brain, the outcomes of this study will have significant contribution to the promotion of human health and welfare. PUBLIC HEALTH RELEVANCE: Continuous production of new neurons in the adult brain (adult neurogenesis) has been shown to play an important role in higher brain functions as well as repair of damaged brains. Yet, our understanding of the mechanisms underlying this important phenomenon is still limited. This study will address this issue by studying the roles of the transcription factors Gsx1 and Gsx2 in adult neurogenesis. Thus, the outcomes of the proposed research will have significant contribution to the promotion of human health and welfare.

描述（由申请人提供）：新神经元和神经胶质细胞的活跃产生在成年哺乳动物大脑的限制区域中持续进行，包括侧脑室内衬的室下区（SVZ）。在这些所谓的神经源性生态位中，神经干细胞 (NSC) 和其他祖细胞（统称为 NPC）终生持续存在，并作为新神经元和神经胶质细胞的来源。最近的研究表明，成人的这种持续的神经发生有助于大脑的生理和病理。此外，通过操纵内源性NPC来再生损伤后特定类型的神经元是恢复神经病学的重要目标之一。然而，我们对这一重要现象背后机制的理解仍然非常有限，进一步的研究势在必行。在本提案中，我们将通过研究同源域转录因子 Gsx1 和 Gsx2 在成人神经发生中的作用来解决这个问题。 Gsx1 和 Gsx2 控制端脑发育的许多方面，包括 NPC 的区域规范和增殖以及神经元分化。然而，它们在成人中的功能尚未得到研究。我们的初步数据表明，Gsx1 和 Gsx2 在 NPC 的特定亚群中表达，这些亚群位于成人 SVZ 的解剖学离散子域中。我们假设 Gsx1 和 Gsx2 在控制成人 SVZ 中 NPC 离散亚群的维持和/或分化中发挥关键作用，从而有助于特定 OB 中间神经元亚型的产生。我们还假设 Gsx1+ 和 Gsx2+ 祖细胞有助于成人大脑中损伤诱导的神经发生。 我们将通过以下四个具体目标概述的实验来检验这些假设： 目标 1：揭示成人 SVZ 中 Gsx1+ 和 Gsx2+ 祖细胞的身份。我们的初步数据表明，Gsx1 和 Gsx2 在位于成人 SVZ 离散子域的 NPC 中表达。我们将通过以下方式揭示这些细胞的特性：1) 使用 Gsx2flox/+ 和 Gsx1::GFP 小鼠进行详细的分子标记分析，2) 使用有丝分裂标记 5-bromo-2'deoxyuridine (BrdU) 进行短期和长期标记结合抗有丝分裂剂 b-D-阿拉伯呋喃糖苷 (Ara-C) 和各种生长因子的治疗，以及 3) 克隆细胞培养测定。目标 2：揭示 Gsx1 和 Gsx2 在成人 SVZ 神经发生和胶质细胞发生中的作用。为了揭示 Gsx1 和 Gsx2 的成人特异性功能，我们将在体内进行条件功能获得（GOF）和功能丧失（LOF）分析。携带 Gsx1 和 Gsx2 等位基因的小鼠将用于成人大脑中祖细胞亚型特异性敲除。我们还将使用携带 tet-O 启动子驱动的 Gsx1 和 Gsx2 转基因的小鼠进行条件 GOF 实验。病毒介导的 Gsx1 和 Gsx2 过表达和敲低将同时进行。目标 3：通过体外培养揭示 Gsx1 和 Gsx2 的功能。为了更好地了解 Gsx1 和 Gsx2 的作用机制，我们将使用 NPC 的克隆培养物进行 GOF 和 LOF 分析。我们还将通过全基因组染色质免疫沉淀 (ChIP) 和转录组测定来确定 Gsx1 和 Gsx2 的转录靶标。目标 4：揭示 Gsx1 和 Gsx2 在成人大脑损伤诱导的神经发生中的作用成人大脑损伤诱导的神经发生的机制仍然很大程度上未知。我们将检查 Gsx1+ 和 Gsx2+ 祖细胞是否参与喹啉酸 (QA) 诱导纹状体兴奋性毒性损伤后的异位神经发生。我们还将使用上述转基因和病毒介导的方法，通过 GOF 和 LOF 分析来确定 Gsx1 和 Gsx2 是否在 SVZ 祖细胞和神经发生的损伤依赖性控制中发挥任何作用。 揭示 Gsx1/2 在成人大脑中的作用将为成人神经发生机制提供新的见解，特别是成人 NPC 特异性的分子基础。因此，这项研究将显着增进我们对成人神经发生和 NPC 生物学的理解。鉴于成人神经发生在大脑生理学和病理学中的重要性，这项研究的结果将对促进人类健康和福祉做出重大贡献。 公共健康相关性：成人大脑中新神经元的持续产生（成人神经发生）已被证明在高级大脑功能以及受损大脑的修复中发挥着重要作用。然而，我们对这一重要现象背后机制的理解仍然有限。本研究将通过研究转录因子 Gsx1 和 Gsx2 在成人神经发生中的作用来解决这个问题。因此，拟议研究的成果将对促进人类健康和福祉做出重大贡献。