肌肉干细胞负责出生后骨骼肌的生长和再生，在损伤或疾病诱导的肌肉再生过程中，肌肉干细胞由静息状态被激活，经过增殖、分化后形成新的肌纤维，部分增殖后的肌肉干细胞重新回到静息状态，回归干细胞库。在上述过程中，肌肉干细胞如何在激活后继续保持干性，同时大量扩增，目前所知甚少。由于激活的肌肉干细胞无法在体外长期培养条件下保持干性，而体内系统具有瞬时性和复杂性，难以进行系统的机制研究。申请人建立了能够长期维持激活的肌肉干细胞干性的体外培养系统，并在此过程中鉴定出两类具有不同整合能力的肌肉干细胞，为研究肌肉干细胞的干性保持提供了优良的实验系统。我们将利用此实验系统，通过RNA-seq、Mass-spec等方法鉴定与干性保持和活跃扩增有关的蛋白和非编码RNA，在肌肉干细胞中和体内验证这些相关因子的功能，并阐明其作用的分子机制。

Muscle stem cells are the major source for postnatal muscle growth and muscle regeneration. These cells are activated from quiescent status upon injury or disease, then further go through proliferation and differentiation processes to form new functional myofibers. Part of the activated muscle stem cells can go back to the quiescent status and replenish the stem cell reservoir. How activated stem cells maintain their stemness and the abilities of rapid expansion remains to be elusive. Investigaton on the above questions has been hamperred by the lack of proper experimental system. The in vivo muscle stem cell activation and expansion are transient and difficult to trace; while muscle stem cells lose most of their stemness once cultured in vitro. We rescently established a long term expansion system to culture the activated muscle stem cells and maintain their stemness in vitro. In this system, we identified two groups of muscle stem cells with high or low ability to repair muscle injury in vivo, respectively, though they appear similarly in vitro. These findings provide us an exerllent system to investigate the stemness maintenance of muscle stem cells. We will perform RNA-seq, mass-spec, and other high throughput assays to identify factors (proteins and non-coding RNAs) critical for stemness maintenance, the functions of these factors will be further examined in cultured muscle stem cells and in vivo. The mechanism of how these factors regulating stemness maintenance and long term expansion will also be investigated.