New Tools to Understand Microglial Function

了解小胶质细胞功能的新工具

基本信息

批准号：
8539639
负责人：
BEN A BARRES
金额：
$ 18.98万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-09-05 至 2014-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8539639
关键词：
Adult Alzheimer&apos s Disease Antibodies Antibody Specificity Blood - brain barrier anatomy Bone Marrow Brain C-terminal Candidate Disease Gene Cell Lineage Cell Surface Proteins Cells Central Nervous System Agents Chimera organism Cytoplasmic Tail Data Data Set Development Diphtheria Toxin Disease Drug Targeting Engineering Event Extracellular Domain Functional disorder Gene Expression Gene Expression Profile Gene Expression Profiling Genes Genetic Genetic Transcription Health Hematopoiesis Human Immune Immune response In Situ Hybridization Inflammation Injection of therapeutic agent Integral Membrane Protein Knock-in Mouse Knockout Mice Label Light Lipopolysaccharides Maps Microglia Molecular Profiling Monoclonal Antibodies Multiple Sclerosis Mus Myelogenous Myeloid Cells N-terminal Neuraxis PTPRC gene Pathologic Peripheral Physiology Population Publications RNA Relative (related person)Reporter Reporting Research Personnel Reverse Transcriptase Polymerase Chain Reaction Role Saline Sorting - Cell Movement Synapses Tamoxifen Testing Therapeutic Time Tissues Transgenic Mice Transplantation Work Yolk Sac aged base brain tissue cell type fractalkine receptor genetic manipulation injured interest macrophage molecular marker monocyte mouse model nervous system disorder novel promoter protein expression recombinase tool

项目摘要

DESCRIPTION (provided by applicant): Microglia is myeloid-derived resident cells within the brain but their exact roles in health and disease are still poorly understood. The inability to reliably distinguish microglia from closely related myeloid cells called macrophages confounds many studies of microglial function. Therefore we screened for a new microglial specific gene that would allow the reliable identification, targeting, and characterization of microglia. In our preliminary studies, we identify TM119, a highly expressed transmembrane protein, as a highly expressed microglia-specific marker that is not expressed by macrophages or other peripheral immune cells. In this application, we will develop several new tools based on TM119 expression to selectively identify, isolate, and manipulate microglia. In the first aim, we will develop two antibodies against TM119 for the identification of microglia by immunostaining and isolation of pure microglia from whole brain tissue by immunopanning. In the second aim, we will develop two mouse lines, a TM119/Cre recombinase knock-in mouse and a TM119/CreERT2 BAC transgenic mouse. These mice will drive constitutive (knock-in) or inducible (BAC) Cre expression selectively within microglia. Crossing these mice with other mouse lines that express Cre- dependent genes will allow selective and specific manipulation of microglial genes. In the final aim, we will use the antibody and genetic tools developed in Aims 1 & 2 to acutely purify microglia from adult, developing, or inflamed mouse tissues to generate gene profiles of pure microglia by gene array and RNAseq. These profiles and all tools developed will be made immediately available upon publication to interested researchers. The tools developed in the proposed studies will enable investigators to better understand the roles of microglia, which may prove critical for a better understanding of the pathophysiology and treatment of human neurological diseases.

描述（由申请人提供）：小胶质细胞是大脑中髓样衍生的居民细胞，但它们在健康和疾病中的确切作用仍然很少了解。无法可靠地将小胶质细胞与称为巨噬细胞的密切相关的髓样细胞区分开来，这混淆了许多小胶质功能的研究。因此，我们筛选了一个新的小胶质细胞基因，该基因将允许可靠的鉴定，靶向和表征小胶质细胞。在我们的初步研究中，我们将TM119（一种高度表达的跨膜蛋白）鉴定为一种高度表达的小胶质细胞特异性标记物，巨噬细胞或其他周围免疫细胞未表达。在此应用程序中，我们将基于TM119表达式开发几种新工具，以选择性地识别，隔离和操纵小胶质细胞。在第一个目标中，我们将通过免疫染色和通过免疫剂从全脑组织中进行免疫染色和分离，开发两种针对TM119的抗体，以鉴定小胶质细胞。在第二个目标中，我们将开发两种小鼠系，TM119/CRE重组酶敲入小鼠和TM119/Creert2 BAC转基因小鼠。这些小鼠将在小胶质细胞中选择性地驱动本构（敲入）或可诱导（BAC）CRE表达。将这些小鼠与表达依赖性基因的其他小鼠系跨越，将允许对小胶质细胞的选择性操纵。在最终目的中，我们将使用目标1和2中开发的抗体和遗传工具从成人，发育或发炎的小鼠组织中急性纯化小胶质细胞来通过基因阵列和RNASEQ产生纯小胶质细胞的基因谱。这些配置文件和所有开发的工具将立即向有兴趣的研究人员出版。拟议的研究中开发的工具将使研究人员能够更好地了解小胶质细胞的作用，这对于更好地理解人类神经疾病的病理生理学和治疗可能至关重要。