Single Cell Characterization of FTLD-GRN

阿尔茨海默病(AD)和与之相关的痴呆症，额颞叶变性(FTLD)，有几个共同的 主要的临床和神经病理特征，一直被认为是疾病谱的两端。在……里面 支持这一点的遗传证据表明，具有前颗粒蛋白(GRN)基因显性突变的患者 无一例外地发展为伴有TDP-43蛋白病变的FTLD，其特征是RNA结合蛋白积聚 TDP-43在额叶皮质的第2-3层。有趣的是，3‘非编码区的单核苷酸多态(SNPs) GRN基因降低了原颗粒蛋白(PGRN)水平，并与TDP风险增加有关。 43.30%~40%的AD患者边缘有蛋白病变。尽管有这些耐人寻味的基因-表型 对于PGRN缺乏如何促进神经胶质和神经元病理的相关性，人们仍然知之甚少。至 为了探讨PGRN缺乏症的神经退变机制，我们进行了单核RNA- GRN-/-小鼠在衰老过程中丘脑的序列分析(SnRNA-seq)表明，在PGRN 缺乏，小胶质细胞是第一种表现出自我平衡基因进行性丧失并获得 促进神经细胞死亡的促炎状态的转录和组织病理学特征 TDP-43蛋白病。为了将这些结果与人类疾病联系起来，我们进行了一项初步研究，通过 用FTLD额叶皮质和丘脑的死后组织进行SnRNA-seq的结果比较- GRN病例与19个月大的Grn-/-小鼠来自相似脑区的病例。这种人-鼠SNRNA-seq 比较发现FTLD-GRN病例和GRN-/-小鼠共有的转录变化，包括细胞 小胶质细胞(胞吐、免疫激活和趋化)和星形胶质细胞(星形胶质细胞-血管)的反应 耦合、细胞黏附和突触组织)。此外，我们的结果揭示了 FTLD-GRN大鼠额叶皮质和丘脑兴奋性和抑制性神经元转录水平的变化 病例，表明人类特有的神经元脆弱性。总而言之，这些结果提出了这样的假设 PGRN缺乏扰乱了小胶质细胞和星形胶质细胞的基因调控网络，改变了复杂的神经胶质细胞 促进FLTD-GRN神经退变的相互作用。为了验证这一点，我们建议1)绘制转录组 以及定义额叶皮质神经胶质病理和神经元脆弱性的基因调控网络 丘脑FTLD-GRN；2)视觉神经元和胶质细胞的细胞弹性机制 绘制了FTLD-GRN的转录本和表观基因组图；3)描述了FTLD-GRN 使用基于IPSC的模型研究FTLD-GRN中转录和表观遗传修饰的后果。这 该项目将提供关键数据，填补有关神经胶质细胞和神经元轨迹的知识空白 病理学，以及FTLD-GRN中特定脑区的脆弱性和复原性。这个项目的结果将是 进一步提供重要的见解，并使预测疾病进展的生物标记物的发现成为可能。

Alzheimer’s disease (AD) and the related dementia, frontotemporal lobar degeneration (FTLD), share several key clinical and neuropathological features and have been considered as two ends of a disease spectrum. In support of this, genetic evidence shows that patients with dominant mutations in the Progranulin (GRN) gene invariably develop FTLD with TDP-43 proteinopathy, characterized by the accumulation of RNA binding protein TDP-43 in layers 2-3 of the frontal cortex. Interestingly, single nucleotide polymorphisms (SNPs) in the 3’UTR of the GRN gene reduce Progranulin (PGRN) protein levels and have been associated with increased risk of TDP- 43 proteinopathy in the limbic regions in 30-40% of AD patients. Despite these intriguing genotype-phenotype correlations, how PGRN deficiency promotes glial and neuronal pathology remains poorly understood. To investigate the mechanism of neurodegeneration in PGRN deficiency, we performed single-nuclei RNA- sequencing (snRNA-seq) in the thalamus of Grn-/- mice during the aging process and showed that, in PGRN deficiency, microglia are the first cell type to show progressive loss of homeostatic genes and acquire transcriptomic and histopathological features of a pro-inflammatory state that promotes neuronal cell death and TDP-43 proteinopathy. To connect these results with human disease, we’ve conducted a pilot study by comparing the results from snRNA-seq using postmortem tissues from the frontal cortex and thalamus of FTLD- GRN cases with those from similar brain regions in 19-month-old Grn-/- mice. This human-mouse snRNA-seq comparison revealed shared transcriptomic changes in FTLD-GRN cases and Grn-/- mice, including cellular responses in microglia (exocytosis, immune activation, and chemotaxis) and astrocytes (astrocyte-vascular coupling, cell adhesion, and synaptic organization) in both brain regions. Furthermore, our results uncovered transcriptomic changes in excitatory and inhibitory neurons in the frontal cortex and thalamus of FTLD-GRN cases, suggesting human-specific neuronal vulnerability. Together, these results broach the hypothesis that PGRN deficiency disrupts the gene regulatory network in microglia and astrocytes and alters intricate glia-neuron interactions to promote neurodegeneration in FLTD-GRN. To test this, we propose to 1) Map the transcriptome and gene regulatory network that define glial pathology and neuronal vulnerability in the frontal cortex and thalamus of FTLD-GRN; 2) characterize the mechanism of cellular resilience in neurons and glia in the visual cortex of FTLD-GRN by mapping their transcriptomes and epigenomes; and 3) delineate the functional consequences of transcriptomic and epigenetic modifications in FTLD-GRN using IPSC-based models. This project will provide critical data that fill the knowledge gaps regarding the trajectories of glial and neuronal pathology, and brain region-specific vulnerability and resilience in FTLD-GRN. Results from this project will further provide important insights and enable the discovery of biomarkers that predict disease progression.