Skeletal muscle stem cells (MuSCs) are the major contributor to muscle regeneration. Upon injury, quiescent MuSCs activate, proliferate, and further differentiate to repair damaged muscle tissue or self-renew to replenish the stem cell pool. The goal of this study was to generate a toolbox to clear some technical roadblocks related to the molecular regulation of MuSC quiescence exit. In the first project, we established a label-free live imaging platform based on Stimulated Raman Scattering (SRS) microscopy, aiming to capture a dynamic template DNA strands co-segregation process during MuSC division. This would provide direct evidence supporting the "immortal strand hypothesis". In the second project, we generated a transgenic mouse that can serve as both a lineage tracer and cell divis...[ Read more ]

Skeletal muscle stem cells (MuSCs) are the major contributor to muscle regeneration. Upon injury, quiescent MuSCs activate, proliferate, and further differentiate to repair damaged muscle tissue or self-renew to replenish the stem cell pool. The goal of this study was to generate a toolbox to clear some technical roadblocks related to the molecular regulation of MuSC quiescence exit. In the first project, we established a label-free live imaging platform based on Stimulated Raman Scattering (SRS) microscopy, aiming to capture a dynamic template DNA strands co-segregation process during MuSC division. This would provide direct evidence supporting the "immortal strand hypothesis". In the second project, we generated a transgenic mouse that can serve as both a lineage tracer and cell division counter. This useful lineage tracer can identify, isolate, and trace somatic stem cell populations within adult tissues, as stem cells are only a small percentage of total cells within the tissue and heterogeneity exists within this population. Furthermore, lineage tracing of specific cells along with cell cycle history of the low turnover somatic stem cells will enable us to better understand how they behave during homeostasis, pathological conditions, and aging. In the third project, we developed a low-input ribosome profiling method to investigate translational regulations during muscle stem cell quiescence exit. Previously, it was technically challenging to apply ribosome profiling to quiescent MuSCs due to their low biological activity. Complementary to mRNA sequencing, our low-input ribosome profiling offers the possibility of a comprehensive understanding of how protein abundance is determined during stem cell lineage progression. Taken together, we generated a toolbox to assist the molecular regulation study of MuSCs quiescence exit, involving template stand co-segregation, cell cycle tracking as well as translation.