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Synergistic Interaction of FTD Genes in Neuroinflammation and Neurodegeneration

FTD 基因在神经炎症和神经变性中的协同相互作用

基本信息

批准号：
10393790
负责人：
Naznin Jahan
金额：
$ 4.37万
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：
2022
资助国家：
美国
起止时间：
2022-01-12 至 2023-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10393790
关键词：
3-Dimensional Affect Aging Autophagocytosis Biochemistry Biological Models Brain Brain Pathology Brain region C9ORF72 CRISPR/Cas technology Cell Nucleus Cells Cellular biology Clinical Research Coculture Techniques Cytoplasm Data Data Set Dementia Disease Disease Progression Exhibits Fellowship Foundations Frontotemporal Dementia Future GRN gene Gene Abnormality Genes Genetic Gliosis Goals Homeostasis Human Immune Immunohistochemistry In Situ Hybridization In Vitro Knockout Mice Lead Limbic System Longevity Lysosomes Mediating Microglia Modeling Mus Mutation Nerve Degeneration Neuraxis Neurodegenerative Disorders Neuroglia Neurons Organoids PGRN gene Pathogenesis Pathway interactions Patients Phase Phenotype Proteins RNA-Binding Proteins Reporting Research Scientist Signal Pathway TDP-43 aggregation Testing Thalamic structure Therapeutic Toxic effect Training Ubiquitin Western Blotting Wild Type Mouse Work age related aging brain astrogliosis base career cerebral atrophy cohort early onset immune activation induced pluripotent stem cell insight loss of function mouse model neuroinflammation neuron loss neurotoxicity null mutation protein TDP-43 protein aggregation therapeutic target transcriptome sequencing transcriptomics

项目摘要

Project Summary Frontotemporal dementia (FTD) is an early onset neurodegenerative disease, and the second most common cause of dementia in patients 60 years or younger. The majority of familial FTD are caused by intronic hexanucleotide (CCCCGG) repeat expansion in chromosome 9 open reading frame 72 (C9orf72) gene and by dominant mutations in the Progranulin (GRN) gene, causing haploinsufficiency in both genes, abnormal protein aggregate formation in neurons. Several functional and transcriptomic studies have shown that mice with null mutation in C9orf72 or Grn show abnormal microglia (resident CNS immune cells) activation mediated pathogenesis of neurodegeneration in FTD. While the exact functions for C9orf72 and Progranulin (PGRN [protein]) are still unclear, several studies have implicated both in autophagy and endolysosomal pathways in neurons and microglia, and concurrent mutations resulting in increased brain atrophy in patients. These results suggest a possible interaction between C9orf72 and PGRN in neurodegeneration during brain aging The goal of my project is to investigate the synergistic interaction of C9orf72 and Grn genes in glial homeostasis and neuronal degeneration using mouse models. In support of this, my preliminary data showed that C9orf72-/- ;Grn-/- DKO mice have significantly shortened lifespan, much shorter than C9orf72-/- mice, Grn-/- mice and control mice. Brain pathology examination in C9orf72-/-;Grn-/- DKO mice showed age-dependent gliosis as well as neuronal TDP-43 aggregates that are more pronounced and wide-spread than those in C9orf72-/- or Grn-/- mice. These results support my hypothesis and further indicate that loss of C9orf72 and Grn synergistically disrupt glia-neuron homeostasis and lead to more pronounced neurodegeneration phenotype. For the F00 phase, I propose to uncover the mechanism of C9 and PGRN in neurodegeneration in the aging brain via single-cell and bulk RNA-sequencing in 7 and 12 months old control, C9orf72-/-, Grn-/- and C9orf72-/-;Grn-/- DKO brain to determine how loss of these two FTD genes disrupts the homeostasis in glia-neuron interaction (Aim 2a). These transcriptomic data will be validated using in situ hybridization, immunohistochemistry and western blots (Aim 2b). Finally, I propose to develop in vitro cultures, including neuron-only cultures and glia-neuron co-cultures, which will provide more insights into the synergistic interaction between C9orf72 and PGRN in the autophagy- lysosome pathways and in glia-mediated toxicity to neurons (Aim 2c). For the K00 phase of this fellowship, I plan to develop induced pluripotent stem cells (IPSC)-derived 3D brain organoids as model systems to investigate disease mechanism and identify therapeutics for neurodegeneration. To identify signaling pathways that could be affected by diseases, I plan on using single-cell transcriptomics on patient IPSC derived glia-neuron organoids, followed by CRISPR/Cas9-based manipulation strategies on the signaling pathways dysregulated in these brain organoids to elucidate the diseases progression mechanisms.

项目摘要额颞叶痴呆（FTD）是一种早发性神经退行性疾病，老年痴呆症的病因在60岁或更年轻的患者。大多数家族性FTD是由内含子引起的，在9号染色体开放阅读框72（C9 orf 72）基因中六核苷酸（CCCCGG）重复扩增，颗粒蛋白前体（GRN）基因的显性突变，导致两个基因的单倍不足，异常蛋白在神经元中聚集形成。几项功能和转录组学研究表明， C9 orf 72或Grn中的突变显示异常的小胶质细胞（驻留CNS免疫细胞）活化介导的 FTD中神经变性的发病机制。虽然C9 orf 72和颗粒蛋白前体（PGRN）的确切功能 [蛋白质]）仍不清楚，一些研究表明自噬和内溶酶体途径都参与了神经元和小胶质细胞，以及导致患者脑萎缩增加的并发突变。这些结果提示C9 orf 72和PGRN在脑老化过程中神经变性中可能相互作用本课题的目的是研究C9 orf 72和Grn基因在胶质细胞内稳态中的协同作用和使用小鼠模型的神经元变性。为了支持这一点，我的初步数据显示，C9 orf 72-/- Grn-/- DKO小鼠的寿命显著缩短，比C9 orf 72-/-小鼠、Grn-/-小鼠和对照组短得多小鼠在C9 orf 72-/-;Grn-/- DKO小鼠中的脑病理学检查显示年龄依赖性胶质细胞增生以及神经元TDP-43聚集体比C9 orf 72-/-或Grn-/-小鼠中的那些更明显和更广泛分布。这些结果支持了我的假设，并进一步表明C9 orf 72和Grn的缺失协同破坏了神经胶质-神经元稳态并导致更明显的神经变性表型。对于F00阶段，本研究拟通过单细胞免疫组化技术，探讨C9和PGRN在老年脑神经退行性变中的作用机制。以及7和12月龄对照、C9 orf 72-/-、Grn-/-和C9 orf 72-/-;Grn-/- DKO脑中的大量RNA测序，确定这两个FTD基因的缺失如何破坏神经胶质-神经元相互作用的稳态（目的2a）。这些转录组学数据将使用原位杂交、免疫组织化学和蛋白质印迹进行验证（Aim 2b）。最后，我建议开发体外培养，包括仅神经元培养和神经胶质-神经元共培养，这将为C9 orf 72和PGRN在自噬中的协同相互作用提供更多的见解- 溶酶体途径和神经胶质介导的神经元毒性（目的2c）。在K 00阶段，我计划开发诱导多能干细胞（IPSC）衍生的3D大脑类器官作为模型系统来研究疾病机制和识别神经变性的治疗方法。为了确定可能受疾病影响的信号通路，我计划使用单细胞转录组学，患者IPSC衍生的胶质神经元类器官，然后在神经元上进行基于CRISPR/Cas9的操作策略。这些脑类器官中的信号通路失调，以阐明疾病进展机制。