Mechanism of microglia proliferation

小胶质细胞增殖机制

基本信息

批准号：
10185588
负责人：
Zhonghui Guan
金额：
$ 42.28万
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：
2021
资助国家：
美国
起止时间：
2021-06-01 至 2026-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10185588
关键词：
Adult Afferent Neurons Alzheimer&apos s Disease Brain CSF1R gene Cell Differentiation process Development Down-Regulation Embryo Fluorescence Gene Expression Profile Genes Immunohistochemistry Infection Infiltration Macrophage Colony-Stimulating Factor Maintenance Membrane Proteins Mental disorders Microglia Modeling Motor Neurons Multiple Sclerosis Nerve Degeneration Nervous System Physiology Neurodegenerative Disorders Neurologic Neurological Models Newborn Infant Optic Nerve Optic Nerve Injuries Pathway interactions Pattern Peripheral Nerves Phase Phosphorylation Play Population Proliferating Proliferation Marker Protein Tyrosine Kinase Proto-Oncogene Proteins c-myc Psyche structure Quantitative Reverse Transcriptase PCR Regulation RiboTag Role Seizures Signal Pathway Spinal Spinal Cord Stroke System TREM2 gene TYROBP gene Testing Time Transcriptional Regulation Up-Regulation cellular imaging experimental study injured irradiation monocyte mouse model neonate nerve injury nervous system disorder overexpression prevent sciatic nerve injury self-renewal single-cell RNA sequencing spinal cord and brain injury transcription factor

项目摘要

ABSTRACT Microglia proliferation occurs in numerous neurological disorders, including peripheral and optic nerve injuries, neurodegenerative diseases, stroke, traumatic brain and spinal cord injuries, multiple sclerosis, seizure, infection, irradiation, and mental disorders. Parabiotic studies, along with studies with single-cell imaging and multicolor fluorescence fate mapping system, have demonstrated that the self-renewal of resident microglia is responsible for the expansion of microglia population in mouse models of neurological diseases. In fact, the newborn microglia generated from microglial self-renewal have significant neurological function with unique transcriptional profile. Moreover, dysregulation of microglia proliferation significantly impacts the development of many neurological diseases, indicating that microglia proliferation plays critical roles in the development of neurological diseases. However, the mechanism of microglia proliferation is largely unknown. We did a group of preliminary experiments to explore the mechanisms of microglia proliferation, and identified a group of embryonic genes that have similar expression pattern as proliferation marker Mki67 during microglia development. With single-cell RNA sequencing (scRNA-Seq), we found that these embryonic microglial proliferation-associated genes were upregulated in lumbar cord microglia around the time they proliferate. Our qRT-PCR from the sorted microglia and microglial RiboTag confirmed the transient upregulation of transcription factor Myc, one of the embryonic microglial proliferation-associated genes, in lumbar cord microglia shortly after sciatic nerve injury. We also found that the deletion of Myc from adult microglia significantly prevented the early phase of nerve injury-induced microglia proliferation, with the late phase of microglia proliferation intact, suggesting that adult microglia proliferation has a Myc-dependent initiation phase and a Myc-independent maintenance phase. To explore the signaling pathway upstream of microglial Myc upregulation, we analyzed our scRNA-Seq and found that the genes encoding DAP12 and its associated membrane protein TREM2, which is required for microglia proliferation in Alzheimer's disease, are upregulated in lumbar cord microglia after sciatic nerve injury. In addition, we found that Syk, a protein tyrosine kinase downstream of TREM2 and DAP12, and upstream of Myc, is phosphorylated in lumbar cord microglia shortly after sciatic nerve injury. In searching for Myc independent mechanism for the maintenance phase of microglia proliferation, we also found that Mafb, a transcription factor expressed at much higher level in adult than in embryonic microglia, inhibits microglia proliferation in adult but not in neonate, and is downregulated in lumbar cord microglia after nerve injury-induced Myc upregulation. We now propose a set of experiments to further delineate the mechanism of microglia proliferation. Aim 1: To test the hypothesis that Myc is required for microglia proliferation in general. Aim 2: To test the hypothesis that Mafb contributes to microglia proliferation. Aim 3: To test the hypothesis that CSF1R-TREM2-DAP12-Syk pathway contributes to the expression regulation of microglial Myc and/or Mafb and to microglia proliferation.

抽象的小胶质细胞增殖发生在许多神经系统疾病中，包括周围神经和视神经损伤，神经退行性疾病、中风、脑部和脊髓损伤、多发性硬化症、癫痫发作、感染、辐射和精神障碍。联体生物研究，以及单细胞成像和多色研究荧光命运图谱系统，已证明常驻小胶质细胞的自我更新是负责的用于扩大神经系统疾病小鼠模型中的小胶质细胞数量。事实上，新生儿小胶质细胞自我更新产生的小胶质细胞具有重要的神经功能，具有独特的转录功能轮廓。此外，小胶质细胞增殖的失调会显着影响许多细胞的发育。神经系统疾病，表明小胶质细胞增殖在神经系统疾病的发展中起着至关重要的作用疾病。然而，小胶质细胞增殖的机制在很大程度上尚不清楚。我们做了一组初步的实验探索小胶质细胞增殖的机制，并鉴定了一组胚胎基因在小胶质细胞发育过程中与增殖标志物 Mki67 具有相似的表达模式。与单细胞 RNA测序（scRNA-Seq），我们发现这些胚胎小胶质细胞增殖相关基因腰脊髓小胶质细胞在增殖时上调。我们对分选的小胶质细胞进行 qRT-PCR 小胶质细胞 RiboTag 证实了转录因子 Myc 的短暂上调，Myc 是胚胎细胞中的一种坐骨神经损伤后不久腰脊髓小胶质细胞中的小胶质细胞增殖相关基因。我们还发现成年小胶质细胞中 Myc 的缺失显着阻止了神经损伤引起的早期阶段小胶质细胞增殖，晚期小胶质细胞增殖完整，表明成年小胶质细胞增殖具有 Myc 依赖性起始阶段和 Myc 独立维持阶段。探索小胶质细胞 Myc 上调的信号通路上游，我们分析了我们的 scRNA-Seq 并发现编码 DAP12 及其相关膜蛋白 TREM2 的基因，这是小胶质细胞所需的阿尔茨海默氏病中的增殖，在坐骨神经损伤后腰脊髓小胶质细胞中的表达上调。此外，我们发现 Syk 是 TREM2 和 DAP12 下游以及 Myc 上游的一种蛋白酪氨酸激酶，坐骨神经损伤后不久腰脊髓小胶质细胞中磷酸化。在寻找Myc独立对于小胶质细胞增殖维持阶段的机制，我们还发现转录因子Mafb 在成人中的表达水平比在胚胎小胶质细胞中高得多，抑制成人中小胶质细胞的增殖，但在新生儿中不存在，并且在神经损伤诱导的 Myc 上调后在腰脊髓小胶质细胞中下调。我们现在提出一组实验来进一步描述小胶质细胞增殖的机制。目标 1：测试 Myc 是小胶质细胞增殖所必需的假设。目标 2：检验 Mafb 的假设有助于小胶质细胞增殖。目标 3：检验 CSF1R-TREM2-DAP12-Syk 通路的假设有助于小胶质细胞 Myc 和/或 Mafb 的表达调节以及小胶质细胞增殖。