Muscle stem cell therapy is being studied for the treatment of skeletal muscle pathologies, such as severe muscle injury or muscular dystrophies, to enhance muscle regeneration. In vivo, the pool of skeletal muscle stem cells (MuSCs) is the main cell population that allows skeletal muscle regeneration. Therefore, MuSCs are considered as a good cell source for cell therapy. However, it is known that MuSCs lose some of their regenerative potential when expanded in vitro. To date, there has been no identification of the ideal cell product.

In the laboratory, it has been demonstrated that human muscle reserve cells (MuRCs) generated in vitro are highly comparable to human MuSCs. Human MuRCs are quiescent Pax7+/MyoD- cells with enhanced survival and a higher potency to generate Pax7+ cells in vivo compared to human myoblasts. Furthermore, it is possible to generate human MuRCs in vitro in large numbers, making them suitable for future therapeutic applications.

Based on these promising results obtained earlier in the laboratory, I then proposed to evaluate the molecular signature, metabolic profile, and therapeutic potential of human MuRCs. In a first study, I assessed the expression of Pax7 in human MuRCs, a known marker of MuSCs. I showed by flow cytometry that human MuRCs are heterogeneous for Pax7 expression with Pax7^High and Pax7^Low subpopulations. Using bulk-RNA sequencing, we showed that Pax7High cells exhibited a deeper quiescent state and a stronger shift towards fatty acid oxidation (FAO) compared to Pax7^Low -MuRCs. However, it is not possible to obtain viable cells to study their functional properties in vitro or in vivo with the procedure (fixation and permeabilization) used to isolate human Pax7^High and Pax7^Low MuRCs populations. In a second study, I investigated whether autofluorescence could be used as a tool to isolate viable and functional human MuRCs. I showed that compared to human MB, human MuRCs are highly autofluorescent (AF) cells. We can distinguish two subpopulations of MuRCs based on their autofluorescence AFHigh and AFLow. After sorting, these cells were shown to be viable and functional in vitro. In addition, AF^High and AF^Low were shown to be functional cells in vivo as they participate in muscle regeneration and in MuSCs pool formation after their intramuscular injection in immunodeficient mice.

Overall, my PhD project has the potential to improve our knowledge concerning the molecular signature, metabolic profile and therapeutic potential of human MuRCs. Taken together, these results suggest that human MuRCs AF^High/Pax7^High may be a suitable stem cell source for potential therapeutic applications in skeletal muscle diseases.