Revealing muscle stem cell heterogeneity in mice and humans through deep single-cell analysis

项目摘要/摘要 肌肉干细胞(MuSCs)，也被称为卫星细胞，是骨骼肌再生所必需的 在一生中。在老年人中，损伤后的肌肉质量和再生能力逐渐下降， 导致生活质量下降。我们最近已经证明了预期分离的MUSCs 衰老小鼠具有高度的异质性，作为一个群体，其再生能力显著降低。 相对于年轻的成年间充质干细胞，揭示了在老年间充质干细胞中先前未检测到的固有干细胞缺陷。这个 老年MSC功能障碍的特征是从可逆的静止转变为细胞衰老，驱动 由于p16INK4a细胞周期抑制因子的升高，以及细胞自主性异常导致的低效自我更新 P38丝裂原活化蛋白激酶和STAT3通路的激活。是什么导致了这些固有的 老年小鼠和人类的改变以及如何前瞻性地识别和治疗功能障碍的MSC仍然存在 悬而未决的问题。我们建议将高通量的深度单细胞RNA测序 成年小鼠和人类肌肉样本和干细胞群重建算法，以识别细胞- 明确区分健康和疾病小鼠和小鼠MUSCs的表面抗原图谱 人类的衰老。我们假设这种方法将使发现异质性表达的MUSC成为可能 在新发现的功能层次中区分不同干细胞能力的细胞表面抗原。我们会 预期分离和移植小鼠和人MUSCs亚群并进行有限稀释 利用生物工程培养微环境进行移植试验和自我更新试验。我们将表演 条件性缺失和瞬时敲除研究以调查已识别的抗原是否具有机械性 在体内和体外自我更新功能障碍中的作用。这些拟议的研究将提供一个深入剖析的 小鼠和人类衰老过程中肌肉干细胞和祖细胞的组织细胞图谱，应该能够 以失调干细胞为靶点促进老年人肌肉修复的合理治疗进展 在创伤之后。

PROJECT SUMMARY/ABSTRACT Muscle stem cells (MuSCs), also known as satellite cells, are essential to skeletal muscle regeneration throughout life. In aged individuals, muscle mass and regenerative capacity after injury progressively decline, leading to diminished quality of life. We have recently demonstrated that MuSCs prospectively isolated from aged mice are highly heterogeneous and, as a population, have a marked reduction in regenerative capacity relative to young adult MuSCs, revealing a previously undetected intrinsic stem cell defect in aged MuSCs. The dysfunction of aged MuSCs is characterized by a shift from reversible quiescence to cellular senescence, driven by elevation of the p16Ink4a cell-cycle inhibitor, and inefficient self-renewal, caused by aberrant cell-autonomous activation of the p38 mitogen-activated protein kinase and STAT3 pathways. What causes these inherent alterations and how to prospectively identify and treat dysfunctional MuSCs in aged mice and humans remain unanswered questions. We propose to combine high-throughput deep single-cell RNA-sequencing across varied adult mouse and human muscle samples and stem-cell population reconstruction algorithms to identify cell- surface antigen profiles that unambiguously distinguish between health and diseased MuSCs in mouse and human aging. We hypothesize that this approach will enable discovery of heterogeneously expressed MuSC cell-surface antigens that demark differing stem-cell capacities within a new-found functional hierarchy. We will prospectively isolate and transplant mouse and human MuSCs sub-populations and perform limiting dilution transplantation assays and self-renewal assays using bioengineered culture microenvironments. We will perform conditional deletion and transient knockdown studies to investigate if the identified antigens have mechanistic roles in self-renewal dysfunction in vivo and in vitro. These proposed studies will provide a deeply-profiled organized cellular atlas of muscle stem and progenitor cells in mouse and human aging that should enable rational therapeutic development targeting dysregulated stem cells for enhancing muscle repair in the elderly following trauma.