John F Hardesty, MD Department of Ophthalmology and Visual Sciences, Washington University School of Medicine, St. Louis, MO, USA.
Department of Developmental Biology, Washington University School of Medicine, St. Louis, MO, USA.
Methods Mol Biol. 2025;2848:117-134. doi: 10.1007/978-1-0716-4087-6_8.
Retinal degenerative diseases including age-related macular degeneration and glaucoma are estimated to currently affect more than 14 million people in the United States, with an increased prevalence of retinal degenerations in aged individuals. An expanding aged population who are living longer forecasts an increased prevalence and economic burden of visual impairments. Improvements to visual health and treatment paradigms for progressive retinal degenerations slow vision loss. However, current treatments fail to remedy the root cause of visual impairments caused by retinal degenerations-loss of retinal neurons. Stimulation of retinal regeneration from endogenous cellular sources presents an exciting treatment avenue for replacement of lost retinal cells. In multiple species including zebrafish and Xenopus, Müller glial cells maintain a highly efficient regenerative ability to reconstitute lost cells throughout the organism's lifespan, highlighting potential therapeutic avenues for stimulation of retinal regeneration in humans. Here, we describe how the application of single-cell RNA-sequencing (scRNA-seq) has enhanced our understanding of Müller glial cell-derived retinal regeneration, including the characterization of gene regulatory networks that facilitate/inhibit regenerative responses. Additionally, we provide a validated experimental framework for cellular preparation of mouse retinal cells as input into scRNA-seq experiments, including insights into experimental design and analyses of resulting data.
视网膜退行性疾病包括年龄相关性黄斑变性和青光眼，据估计目前在美国影响超过 1400 万人，随着年龄的增长，视网膜退行性疾病的患病率也在增加。人口老龄化和寿命延长预计会导致视力障碍的患病率和经济负担增加。改善视觉健康和进行视网膜退行性病变的治疗方案可以减缓视力丧失。然而，目前的治疗方法无法根治视网膜退行性病变导致的视力损害的根本原因——视网膜神经元的丧失。内源性细胞来源的视网膜再生刺激为替代丧失的视网膜细胞提供了一个令人兴奋的治疗途径。在包括斑马鱼和非洲爪蟾在内的多种物种中，Müller 胶质细胞具有高度有效的再生能力，可以在整个生物体的生命周期中重建丢失的细胞，这突出了刺激人类视网膜再生的潜在治疗途径。在这里，我们描述了单细胞 RNA 测序 (scRNA-seq) 的应用如何增强我们对 Müller 胶质细胞衍生的视网膜再生的理解，包括鉴定促进/抑制再生反应的基因调控网络。此外，我们还提供了一个经过验证的小鼠视网膜细胞的细胞制备实验框架，作为 scRNA-seq 实验的输入，包括对实验设计和结果数据分析的见解。
