Skeletal muscles provide necessary contractile forces to enable locomotion and breathing. Healthy skeletal muscle can recover from minor injuries, through a regeneration process involving skeletal muscle stem cells. However, there are several pathologic conditions (e.g. myopathies, aging-related sarcopenia) in which skeletal muscle is unable to sufficiently regenerate and ultimately atrophies. There is currently no cell-based therapy available to alleviate these disorders. Our objective is to develop innovative culture environments and investigate the mechanisms that could permit in vitro expansion of aged or diseased muscle stem cells.
In the first and second aims, we have developed a new stiffness-adjustable artificial support material that allows preservation of muscle stem cell viability in vitro (Aim 1) and, more importantly, permits maintenance of muscle stem cell potential as observed after in vivo transplantation of the cultured muscle stem cells (Aim 2). We have demonstrated that the rigidity of the support material on which the cells are expanded is critical for maintaining their transplantation potential. We observed that standard rigid culture surfaces negatively affect muscle stem cell viability and are unfavorable for preserving the muscle stem cell function. We noticed that a soft, very elastic culture material can dramatically enhance the survival of muscle stem cells. We have analyzed the genes expressed by the muscle stem cells isolated from young and aged mouse skeletal muscles as we reasoned that the poor stem cell potential of older stem cells could be resulting from the lack of key stem cell genes. Here we show that gene expression patterns differ between the aged and young muscle stem cells, and that identification of these differentially expressed genes could help us to investigate their potential for rejuvenating the aged cells. We observed that aged muscle stem cells seeded in culture and observed by time lapse videos have diminished behaviors in vitro. Our goal is to exploit this altered behavior to test our ten selected regulators for expanding and rejuvenating aged muscle stem cells.
In the third aim, we have developed a method to isolate muscle stem cells from human samples. Until recently, there has been a lack of information regarding the characterization of human stem muscle from skeletal tissue. By using flow cytometry, we have managed to identify a family of cell surface molecules that allows enrichment of the cells from a very heterogeneous population. These cells are highly myogenic, which means that in vitro they proliferate and generate muscle fibers, and we are now in the process of testing their behavior in culture on the artificial platform described above. In parallel, we are analyzing the genes expressed by these cells isolated from young or aged patients and we seek to compare these analyses with the mouse data that we have acquired.