Exosome Production in C2C12 Myoblasts Improves Proliferation and Myogenic Differentiation following Exercise

Introduction: Mesenchymal stem cells (MSCs) influence tissue regeneration and healing primarily through paracrine secretion of proregenerative factors. Many of these factors are contained within exosomes suggesting that exosome suspensions may be an effective cell-free therapy for tissue regeneration and repair. We have previously shown that mechanical strain (exercise) rescues the myogenic potential of muscle derived stem cells from an aged mouse model (progeria mice). We hypothesized that in vitro “exercise” can improve the regenerative potential of exosomes derived from myoblastic precursors.

Materials and Methods: C2C12 myoblast cells were subjected to two different ex vivo exercise routines: low-intensity endurance and high-intensity interval using biaxial or uniaxial strain on a Flexcell FX-5000TT bioreactor. Exosomes were quantified from each experimental condition and 20 µg of exosomes (normalized to protein content) from each condition were added to myogenic differentiation media. After 3 days, cell proliferation and myotube quantification was performed.

Results: All exercised cells secreted significantly higher concentrations of exosomes than unexercised controls. Low-intensity exercise resulted in lower exosome concentrations than high-intensity exercise. Exosomes isolated from all exercised conditions resulted in significantly greater proliferation and myotube formation than controls. Exosome number was not significantly impacted by storage at −80°C for up to 4 months.

Discussion/Conclusion: Parent cell conditions influence exosome payload and subsequent regenerative effects on progenitor cells. Here we show that exercising progenitor cells ex vivo can improve the proliferation and differentiation potential of whole exosome suspensions. Exosomes represent a powerful new platform for cell-free regenerative therapies

Acknowledgment: This project was funded by The Borgen Family Foundation.