Microglia contribution to disease pathogenesis in C9orf72 ALS/FTD

项目摘要 小胶质细胞在C9orf72(C9)肌萎缩侧索硬化症/额颞叶痴呆中的作用 疾病谱仍然知之甚少。早期的研究发现，小胶质细胞的激活 在患有痴呆和执行功能受损的ALS患者中显著升高，这表明小胶质细胞激活 与肌萎缩侧索硬化症的FTD样症状相关。FTLD中小胶质细胞的最新神经病理学检查 原颗粒蛋白与C9orf72基因突变的患者尸检得出结论，观察到的小胶质细胞 两个基因不同的患者亚组之间的功能障碍不同，表明 小胶质细胞功能障碍取决于患者群体的病因。小胶质细胞有一个有趣的方面-- 神经元通讯是小胶质细胞通过以下途径在维持和完善突触网络中的作用 突触的选择性修剪，主要发生在发育过程中，但已被证明也 在阿尔茨海默病(AD)和包括FTD在内的相关痴呆症中被触发。突触丢失的程度 AD与认知衰退密切相关，甚至比斑块、缠结或神经元丢失的量更大。 最近对肌萎缩侧索硬化症死后组织的研究证实，阿尔茨海默病患者前额叶皮质突触丢失增加 报告有认知障碍的患者。我们实验室有初步数据支持这一假设 C9orf72患者大脑皮质前脑区存在神经免疫相互作用改变 证实了FTD，在FTD中，小胶质细胞和神经元相互调节功能。使用患者派生的HiPSC 小胶质细胞和皮质神经元，我们能够证明C9患者来源的HiPSC小胶质细胞单一培养具有 具有固有的表型，包括改变的基因图谱、吞噬活性和溶酶体功能。多数 有趣的是，初步数据表明，C9小胶质细胞确实调节神经元的兴奋性和C9的存活率。 IPSC神经元。为了进一步研究小胶质细胞在C9皮质变性中的作用和贡献，我们 建议深入研究C9 HiPSC-小胶质细胞的内在特性(来自所有患者亚组：FTD， FTD/ALS，ALS；Aim1)。然后，我们将第一次将这些小胶质细胞与C9和健康对照组共同培养 HIPSC皮质神经元，以更好地了解这两种细胞类型之间的共同调节(目标2)。最后，在 第三个目的，我们将研究C9患者尸检组织中小胶质细胞的激活和病理。这将是 包括来自现有SnRNA序列数据集特定细胞类型的遗传简档、免疫组织化学 小胶质细胞增生症和小胶质细胞特异性候选基因/蛋白的多标记免疫染色 C9神经元病病理标记物(TDP-43和C9 dprs)以获得关于小胶质细胞是否 在神经细胞病变附近优先改变。

Project Abstract The role of microglia in the C9orf72 (C9) amyotrophic lateral sclerosis (ALS)/ frontotemporal dementia (FTD) disease spectrum remains poorly understood. Early investigations found that microglia activation was significantly higher in ALS with dementia and impaired executive function, suggesting that microglia activation correlates with FTD-like symptoms in ALS. More recent neuropathologic examinations of microglia in FTLD patient autopsy brains with mutations in progranulin versus C9orf72 concluded that the observed microglia dysfunction was different between the two genetically different patient subgroups suggesting specificity of microglia dysfunction depending on the etiology of the patient population. One interesting aspect of microglia- neuron communication is the role of microglia in the maintenance and refinement of synaptic networks through the selective pruning of synapses, which occurs predominantly during development but has been shown to also be triggered in Alzheimer's disease (AD) and related dementias, including FTD. The degree of synapse loss in AD strongly correlates with cognitive decline, even more than the amount of plaque, tangles or neuronal loss, and a recent study of ALS postmortem tissue confirmed increased synapse loss in the prefrontal cortex of patients with reported cognitive impairments. Our laboratory has preliminary data supporting the hypothesis that there is an altered neural-immune interaction in the cortical forebrain regions of C9orf72 patients with confirmed FTD in which microglia and neurons modify each other's function. Using patient-derived hiPSC microglia and cortical neurons, we are able to show that C9 patient-derived hiPSC microglia mono-cultures do have intrinsic phenotypes, including altered gene profiles, phagocytic activities and lysosomal function. Most interestingly, preliminary data suggests that C9 microglia do regulate neuronal excitability and survival of C9 iPSC neurons. To further investigate the role and contribution of microglia in C9 cortical degeneration, we propose to thoroughly investigate the intrinsic properties of C9 hiPSC-microglia (from all patient subgroups: FTD, FTD/ALS, ALS; Aim1). For the first time, we will then co-culture these microglia with C9 and healthy control hiPSC cortical neurons to better understand the co-regulation between these two cell types (Aim 2). Finally, in the third aim, we will study microglia activation and pathology in C9 patient postmortem autopsy tissue. This will include cell-type specific genetic profiling from existing snRNA seq data sets, immunohistochemistry of microgliosis and multi-label immunostaining for microglial-specific candidate genes/proteins in conjunction with C9 neuronal disease pathology markers (TDP-43 and C9 DPRs) to gain novel knowledge on whether microglia are preferentially altered in close vicinity to neuronal pathologies.