A Multi-angled Approach to Discover and Improve Skeletal Muscle Stem Cell Therapies

Skeletal muscle plays an essential role in locomotion, metabolism, and thermoregulation. An intriguing characteristic of skeletal muscle is its remarkable regenerative capacity: a highly orchestrated cellular process, in which the resident muscle stem cells (MuSCs) play a central role. Upon tissue injury, quiescent MuSCs are activated and give rise to a population of primary myoblasts (pMBs). pMBs undergo several rounds of division and ultimately fuse with one another to form multi-nucleated myofibers and repair the muscle. As observed in a wide range of conditions such as muscular dystrophies and aging, disruptions in the repair process can lend to impaired regeneration and progressive muscle wasting. Strategies to restore strength and function to pathological muscle include cell-based therapies to replace defective myogenic cells, and treatments to restore the endogenous repair process. Despite substantial advances, these treatment options are still in the early stages of clinical translation. The specific goal of this thesis is to improve upon currently available strategies as well as to lay the groundwork for the emergence of new therapeutic entry-points for skeletal muscle treatments. In the area of cell transplantation therapy, challenges remain in producing clinically-relevant numbers of cells that, as a population, possess high regenerative potency to produce skeletal muscle and repopulate the stem cell niche. We demonstrate that by using a bioactive hydrogel (HAMC) as the cell delivery vehicle, it is possible to improve MuSC transplantation outcomes, and thereby reduce the overall number of required MuSCs. Next, we address the issue of producing sufficient numbers of highly regenerative myogenic cells. Using a high-throughput drug screen and in-vivo intramuscular transplantation assay validation, we identify epidermal growth factor receptor (Egfr) and vascular endothelial growth factor receptor 2 (Vegfr2, Kdr) as new druggable targets, that upon inhibition, produce a population of cultured MuSCs with greater regenerative potency than control treated. Finally, using single-cell RNA sequencing, we shed light on the diversity and intercommunication of cells present in skeletal muscle. This dataset serves as a valuable resource through which new regulators of MuSCs and other cells in skeletal muscle can be evaluated with an eye towards skeletal muscle regenerative medicine applications.Ph.D