Deciphering the dynamic heterogeneity of regulatory T-cells in muscle injury using single-cell RNA sequencing

由于遗传或年龄相关的病理导致的肌肉再生能力差代表了一个越来越大的健康挑战，要求扩大该领域的研究。免疫细胞在调节肌肉修复和再生中的作用越来越受到重视。最近，Diane Mathis博士和Christophe Benoist博士实验室的研究人员描述了一种独特的调节性t细胞（Tregs）群体，它们在受伤的肌肉中积累，并在组织修复中发挥基本作用。来自这些研究的几条证据表明，肌肉treg包括表型和功能异质性的细胞亚群。这推动了一种假设，即不同的Treg种群被动态地招募到受伤的肌肉中，以便在时间上准时的方式对修复的不同方面做出贡献。然而，目前传统的批量分析方法只能对这种复杂系统的异质性提供有限的见解，并且在研究罕见细胞群方面表现出很大的不足。因此，本项目旨在利用新颖的单细胞RNA测序技术inDrop来破译损伤后肌肉Tregs的动态变化。此外，鉴定的转录组将与t细胞受体序列连接，以推断不同Treg簇之间的克隆关系。此后，新发现的Treg群体的关键转录调节因子将被发现并验证其在肌肉营养不良和慢性损伤模型中的功能相关性。该项目的预期结果将对我们对Treg在组织损伤中的专门化的理解产生直接影响。此外，它将为旨在揭示各种炎症过程中免疫细胞动态异质性的后续研究提供方法学框架。最终，将这些发现转化为人类环境将对肌肉萎缩症患者和老年个体具有临床潜力。

Poor muscle regenerative ability due to genetic or age-related pathologies represents an increasing health challenge that mandates expanding research in that field. Immune cells have been increasingly appreciated for their role in regulating muscle repair and regeneration. Recently, researchers in the lab of Dr. Diane Mathis and Dr. Christophe Benoist have described a unique population of regulatory T-cells (Tregs) that accumulate in injured muscles and play a fundamental role in orchestrating tissue repair. Several lines of evidence from these studies suggested that muscle Tregs comprise phenotypically and functionally heterogeneous cell subsets. This drove the hypothesis that distinct Treg populations are dynamically recruited to injured muscle in order to contribute to different aspects of repair in a temporally punctual manner. However, the current conventional bulk analysis methods offer only a limited insight into the heterogeneity of such complex system and are rendered largely insufficient for studying rare cell populations. Therefore, the proposed project aims at using the novel single-cell RNA sequencing technology, inDrop, to decipher the dynamic changes in muscle Tregs after injury. Moreover, the identified transcriptomes will be linked to T-cell receptor sequences to infer the clonal relationship between different Treg clusters. Thereafter, key transcriptional regulators of the newly-identified Treg populations will be uncovered and validated for their functional relevance in models of muscle dystrophy and chronic injury. The expected findings of this project will have immediate impact on our understanding of Treg specialization in tissue injury. Furthermore, it will provide a methodological framework for subsequent studies aiming to uncover immune cell dynamic heterogeneity in various inflammatory processes. Ultimately, translation of these findings to the human context would have clinical potential for both muscular dystrophy patients and geriatric individuals.