Development of Microglia Knockout Mouse

小胶质细胞敲除小鼠的研制

• 批准号: 10040196
• 负责人: Bogoljub Ciric
• 金额: $ 15.6万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-15 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10040196
• 关键词: Acquired Immunodeficiency Syndrome; Address; Adrenoleukodystrophy; Adult; Age; Alzheimer' s Disease; Amyotrophic Lateral Sclerosis; Animal Model; Biology; Blood - brain barrier anatomy; Bone Marrow; Brain Diseases; Cause of Death; Cells; Central Nervous System Infections; Communicable Diseases; Data Analyses; Dependence; Development; Discipline; Disease; Disease model; Encephalitis; Ensure; Enterobacteria phage P1 Cre recombinase; Experimental Autoimmune Encephalomyelitis; Experimental Models; Exposure to; Funding; Generations; Genes; Genetic Transcription; Health; Homeostasis; Huntington Disease; Immune; Inflammation; Intervention; Knock-out; Knockout Mice; Longevity; LoxP-flanked allele; Macrophage Colony-Stimulating Factor Receptor; Maintenance; Malignant Neoplasms; Malignant neoplasm of brain; Microglia; Modeling; Mononuclear; Multiple Sclerosis; Mus; Myeloid Cells; Neuraxis; Outcome; Parkinson Disease; Partner in relationship; Phagocytes; Phenotype; Physiological Processes; Play; Receptor Gene; Receptor Signaling; Research; Research Personnel; Rett Syndrome; Role; Speed; Tamoxifen; The Jackson Laboratory; Time; TimeLine; Transgenic Mice; Transgenic Organisms; astrogliosis; base; bone cell; experimental study; macrophage; mouse model; nervous system development; novel; painful neuropathy; progenitor; progenitor system; targeted treatment; tool; transcription factor; treatment duration

SUMMARY Microglia are central nervous system (CNS) resident macrophages involved in maintaining CNS homeostasis. Importantly, they are also involved in a number of diseases, including Alzheimer’s disease, Huntington’s disease, amyotrophic lateral sclerosis, Parkinson’s disease, multiple sclerosis, infectious diseases and cancer. Despite their importance in health and disease, researchers still lack an experimental model that would enable simple and permanent depletion of microglia to study their functions and their potential as targets for therapy. The principal difficulty in studying microglia is their extensive similarity to other macrophages, which precludes depletion of microglia without concomitant depletion of macrophages. Making use of recent advances in microglia biology, we propose to characterize a transgenic mouse model, termed here Mg-ipKO that enables inducible, permanent, and highly selective depletion of microglia only. The depletion is based on expression of Sall1, a transcription factor expressed in microglia but not in other macrophages/myeloid cells. Sall1-driven expression of Cre recombinase will lead to knockout of CSF-1 receptor (CSF-1R) in microglia. Given that microglia survival is absolutely dependent on CSF-1R, its knockout will lead to depletion of microglia. The depletion will be tamoxifen-inducible and permanent. A similar concept for short- term microglia depletion by Sall1-driven expression of Cre to knockout CSF-1R has recently been proven successful, supporting the feasibility of the approach that we propose. We have recently generated novel Sall1-FRT-Cre transgenic mouse line, and we are currently crossing these mice with two other transgenic mouse lines (obtained from The Jackson Laboratory) to generate final triple transgenic Mg-ipKO mice with the capacity for inducible microglia depletion. We anticipate that we will have generated Mg-ipKO mice by the time funding for this project may start, allowing for an immediate start of experiments with these mice. We propose the specific aim: To characterize microglia depletion and its effects in Mg-ipKO mice. To determine the usefulness of the model, we will induce microglia depletion in adult Mg-ipKO mice by tamoxifen treatment. We will then determine the extent of the depletion and optimize tamoxifen treatment if necessary. Furthermore, we will characterize potential consequences of the depletion on CNS homeostasis (blood-brain barrier integrity, CNS inflammation, astrogliosis) and determine if, over time, the CNS repopulates with microglia that originate from either CNS progenitors, or with macrophages of bone marrow origin. We expect that by the completion of this proposed research we will have developed and characterized an advanced transgenic mouse model for the depletion of microglia that overcomes the limitations of existing approaches and provides a highly useful research tool for multiple biomedical disciplines.

概括 小胶质细胞是中枢神经系统 (CNS) 常驻巨噬细胞，参与维持 CNS 稳态。 重要的是，它们还与许多疾病有关，包括阿尔茨海默病、亨廷顿舞蹈病、 肌萎缩侧索硬化症、帕金森病、多发性硬化症、传染病和癌症。 它们对健康和疾病的重要性，研究人员仍然缺乏一个实验模型来实现简单的 以及小胶质细胞的永久耗竭，以研究它们的功能及其作为治疗靶标的潜力。 研究小胶质细胞的主要困难是它们与其他巨噬细胞的广泛相似性，这排除了 消耗小胶质细胞而不同时消耗巨噬细胞。 利用小胶质细胞生物学的最新进展，我们建议表征转基因小鼠 模型，此处称为 Mg-ipKO，仅能够诱导、永久和高度选择性地消耗小胶质细胞。 这种耗竭是基于 Sall1 的表达，Sall1 是一种转录因子，在小胶质细胞中表达，但在其他细胞中不表达。 巨噬细胞/骨髓细胞中 Sall1 驱动的 Cre 重组酶表达将导致 CSF-1 受体的敲除。 鉴于小胶质细胞的存活绝对依赖于 CSF-1R，其敲除将导致小胶质细胞中的 (CSF-1R)。 小胶质细胞的消耗是他莫昔芬诱导的并且是永久性的。 最近已证实，通过 Sall1 驱动的 Cre 表达来敲除 CSF-1R 可以消除足月小胶质细胞 成功，支持我们提出的方法的可行性。 我们最近生成了新型 Sall1-FRT-Cre 转基因小鼠品系，目前正在杂交 这些小鼠与另外两个转基因小鼠系（从杰克逊实验室获得）产生最终的三重 我们预计我们将拥有具有诱导型小胶质细胞耗竭能力的转基因 Mg-ipKO 小鼠。 在该项目的资助开始时，我们已经生成了 Mg-ipKO 小鼠，从而可以立即开始 用这些老鼠做实验。 我们提出的具体目标是：表征 Mg-ipKO 小鼠中的小胶质细胞耗竭及其影响。 为了确定该模型的实用性，我们将通过他莫昔芬诱导成年 Mg-ipKO 小鼠小胶质细胞耗竭 然后，我们将确定消耗的程度，并在必要时优化他莫昔芬治疗。 此外，我们将描述中枢神经系统稳态（血脑 屏障完整性、中枢神经系统炎症、星形胶质细胞增生）并确定随着时间的推移，中枢神经系统是否会重新填充小胶质细胞 起源于中枢神经系统祖细胞或骨髓来源的巨噬细胞。 我们预计，通过完成这项拟议的研究，我们将开发并表征 一种先进的小胶质细胞耗竭转基因小鼠模型，克服了现有模型的局限性 方法并为多个生物医学学科提供了非常有用的研究工具。