Single-nucleus RNA Sequencing Reveals GABAergic Vulnerability and Reactive Gliosis Driven by Loss of TDP-43

TDP-43 is an RNA-binding protein important for RNA processing, whose loss-of-function is involved in multiple neurodegenerative disorders, including frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), and Alzheimer's disease (AD). We performed single-nucleus RNA-sequencing to study the convergent and divergent molecular signatures of short-term and long-term TDP-43 depletion in the medial prefrontal cortex (mPFC) using Tdp-43F/F mice, compared with that of 5xFAD mice, a well-established AD mouse model with ß-amyloid plaque pathology. Our results demonstrated a significant loss of GABAergic neurons in the mPFC after short-term TDP-43 depletion. This was accompanied by a remarkable reactive gliosis in the mPFC. Our results revealed a strong GABAergic and glial involvement during early stages of TDP-43 loss-of-function, suggesting that the GABAergic system is vulnerable to TDP-43 pathology and could be considered a potential target for developing therapeutic strategies and biomarkers for early detection in TDP-43 linked AD-related dementia. Overall design: TDP-43 loss-of-function in the medial prefrontal cortex (mPFC) was investigeted using a conditional knockout mouse model Tdp-43 F/F with focally injected AAV viruses expressing either Cre recombinase or GCaMP6f control, alongside 5xFAD mice as AD controls. Short-term (2 months) and long-term (6 months) knockout cohorts, with corresponding age-matched controls, were analyzed. Freshly dissected mPFC tissues were pooled and subjected to single-nucleus RNA sequencing (10x Genomics 3' platform).

TDP-43（TDP-43）是一种参与RNA加工过程的重要RNA结合蛋白，其功能丧失与多种神经退行性疾病相关，包括额颞叶痴呆（FTD）、肌萎缩侧索硬化（ALS）以及阿尔茨海默病（AD）。本研究利用Tdp-43F/F条件性敲除小鼠模型，向其内侧前额叶皮层（mPFC）定点注射表达Cre重组酶（Cre recombinase）或GCaMP6f对照的腺相关病毒（AAV），构建短期与长期TDP-43功能耗竭模型，并与广泛应用的携带β淀粉样斑块病理特征的AD小鼠模型5xFAD小鼠进行对比，采用单细胞核RNA测序（single-nucleus RNA-sequencing）分析二者的趋同与趋异分子特征。研究结果显示，短期TDP-43功能耗竭后，小鼠内侧前额叶皮层内的γ-氨基丁酸能神经元（GABAergic neurons）出现显著丢失，并伴随明显的反应性胶质增生。本研究揭示了TDP-43功能丧失早期阶段中γ-氨基丁酸能系统与胶质细胞的显著参与，提示该系统对TDP-43病理具有易感性，可作为开发TDP-43相关AD型痴呆治疗策略与早期检测生物标志物的潜在靶点。整体实验设计：本研究通过向Tdp-43F/F小鼠内侧前额叶皮层（mPFC）定点注射表达Cre重组酶（Cre recombinase）或GCaMP6f对照的腺相关病毒（AAV），构建TDP-43功能丧失模型，并以5xFAD小鼠作为AD对照。实验分为短期（2个月）与长期（6个月）敲除组，同时设置相应的年龄匹配对照组。将新鲜解剖获取的内侧前额叶皮层（mPFC）组织混合后，采用10x Genomics 3'端测序平台进行单细胞核RNA测序（single-nucleus RNA-sequencing）。