Cellular and Molecular Dynamics of Retinal Microglial in the Context of Photoreceptor Degeneration

感光器变性背景下视网膜小胶质细胞的细胞和分子动力学

基本信息

批准号：
10223314
负责人：
Daniel Raphael Saban
金额：
$ 44.31万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-08-01 至 2024-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10223314
关键词：
Address Age Anaphylatoxins Blindness Bone Marrow Bone Marrow Purging C3AR1 gene Cell Death Cells Chemotaxis Chimera organism Clinical Complex Cytoprotection Data Data Set Defect Development Disease Environment Etiology Excision Explosion Eye Galectin 3 Genetic Genetic Transcription Immune Immune response In Situ Interruption Knock-in Mouse Knock-out Knockout Mice Knowledge Lead Left Letters Life Ligands Light Mediating Methods Microglia Modeling Molecular Molecular Profiling Mus Myeloid Cells Neuraxis Neurons Outcome Pathology Patients Phagocytes Phagocytosis Pharmacology Phenocopy Photoreceptors Physiological Physiology Play Population Process Property Protein Tyrosine Kinase Reagent Reporter Retina Retinal Degeneration Retinal Diseases Rhodopsin Role Series Signal Transduction Stargardt&apos s disease Structure of retinal pigment epithelium TREM2 gene Techniques Testing Time Tissues Transgenic Organisms Validation Vision experimental study gain of function imaging modality in vivo loss of function macrophage migration molecular dynamics monocyte new therapeutic target novel novel therapeutic intervention novel therapeutics patient population photoreceptor degeneration prenatal preservation programs receptor recruit response single-cell RNA sequencing small molecule tool

项目摘要

SUMMARY Many, if not most, forms of retinal degeneration involve ectopic accumulation of subretinal macrophages. However, the contribution of bona fide microglia to this immune response and their independent role in disease is poorly understood. It is now widely appreciated that macrophages in degenerating neuronal tissues are not only comprised of microglia but may also include monocyte-derived macrophages. The former are endogenous prenatal-derived cells maintained locally throughout life, whereas the latter represent transiently recruited passengers in disease states. Hence, these two distinct lineages have nonredundant activities in the disease process and should thereby be studied as distinct entities. However, distinguishing microglia is technically challenging since standard techniques such as immunolabeling, conventional reporter mice, or myeloablation bone marrow chimeras are insufficient to study these two populations. In fact, the only method to achieve such separation is through recently established Cx3cr1-CreER microglia lineage tracing mice. Yet, few studies have employed this approach, which has resulted in a knowledge gap in the field. As it is also now known that microglia are essential in establishing and preserving neuronal activity in physiological conditions, determining microglia-specific activities in retinal disease is now imperative. Using the lineage tracing approach and single- cell RNA sequencing (scRNA-seq), our lab recently identified a novel population of cytoprotective retinal microglia in photoreceptor degeneration models. We now wish to build upon these findings in our current proposal by applying novel tools to unravel these cells mechanistically, as well as to determine the significance of their cytoprotective program across etiologically distinct retinal degenerative diseases. We begin in Aim 1 by leveraging our scRNA-seq dataset to inhibit microglial chemotaxis that will allow us to determine whether cytoprotection is a subretinal-specific response. Aim 2 takes advantage of our scRNA-seq dataset as well for targeted conditional and global knockouts to establish the molecular underpinnings of microglia-mediated protection. Lastly, in Aim 3 we will apply loss- and gain-of-function studies to examine whether this cytoprotective microglial program is operative in both primary photoreceptor degeneration and retinal pigment epithelial pathology-related degeneration. In summary, our proposal is not only timely, but is also poised to unravel the innerworkings of this novel microglial population across etiologically distinct retinal degeneration models and perhaps help uncover novel therapeutic targets that can bolster their activities for vision preservation in photoreceptor degeneration.

概括许多（即使不是大多数）视网膜变性的形式涉及视网膜下巨噬细胞的异位积累。但是，真正的小胶质细胞对这种免疫反应的贡献及其在疾病中的独立作用理解很差。现在广泛认为，退化神经元组织中的巨噬细胞不是仅由小胶质细胞组成，但也可能包括单核细胞来源的巨噬细胞。前者是内源性的产前衍生的细胞在整个生命中保持局部维持，而后者表示暂时招募疾病状态的乘客。因此，这两个不同的谱系在疾病中具有非冗余活性过程，应该作为不同的实体进行研究。但是，在技术上区分小胶质细胞是具有挑战性，因为标准技术，例如免疫标记，常规记者小鼠或骨髓化骨髓嵌合体不足以研究这两个人群。实际上，实现此类的唯一方法分离是通过最近建立的CX3CR1-CREER小胶质细胞谱系追踪小鼠的。但是，很少有研究采用了这种方法，这导致了该领域的知识差距。现在也知道小胶质细胞对于在生理条件下建立和保留神经元活性至关重要，确定现在必须在视网膜疾病中进行小胶质细胞特异性活性。使用谱系跟踪方法和单一细胞RNA测序（SCRNA-SEQ），我们的实验室最近确定了新的细胞保护视网膜种群光感受器变性模型中的小胶质细胞。我们现在希望以当前的发现为基础提案通过应用新工具来机械地阐明这些细胞，并确定显着性其跨病原体视网膜退化性疾病的细胞保护程序。我们从AIM 1开始利用我们的SCRNA-Seq数据集抑制小胶质细胞趋化性，这将使我们能够确定是否是否细胞保护是视网膜下特异性反应。 AIM 2还利用我们的SCRNA-SEQ数据集有针对性的条件和全局敲除，以建立小胶质细胞介导的分子基础保护。最后，在AIM 3中，我们将应用损失和功能收益研究来检查是否细胞保护小胶质细胞程序在一级光感受器变性和视网膜色素中都是可操作的上皮病理相关变性。总而言之，我们的建议不仅是及时的，而且还准备揭示了这个新型小胶质细胞在病因上不同的视网膜变性的内部工作模型，也许有助于发现可以加强其活动活动的新型治疗靶标在光感受器变性中保存。