Extracellular Vesicles from iPSC derived Muscle Progenitor Cells Rejuvenate the Dysfunctional Muscle Stem Cells during Aging

• Published in: The FASEB journal Vol. 36; no. S1
• Main Authors: Xuan, Wanling, Tipparaju, Srinivas M., Ashraf, Muhammad
• Format: Journal Article
• Language: English
• Published: United States The Federation of American Societies for Experimental Biology 01-05-2022
• ISSN: 0892-6638; 1530-6860
• DOI: 10.1096/fasebj.2022.36.S1.L7524

Background and Objective
Sarcopenia is defined as the loss of skeletal muscle mass and function due to age, which is a major concern for life long health and wellbeing in elderly population. Age‐associated decline in the performance of muscle stem cells (MuSC) contributes to the deterioration of skeletal muscle during aging. Using small molecule Givinostat and CHIR99021 we previously demonstrated the generation of muscle progenitor cells (Givi‐MPC) of human Induced pluripotent stem cells (iPSC) from multiple cell lines. We found that extracellular vesicles (EVs) from these MPC have a specific cargo compared with human myoblasts. Here, we investigated whether these EVs can rejuvenate the aging dysfunctional muscle stem cells.
Methods and Results
Using size exclusion chromatography EVs were isolated and purified from iPSC derived Givi‐MPC and primary human myoblasts. The EVs were visualized by transmission electron microscopy. Next, we isolated primary MuSC from 25 months old aged C57/B6 mice, which were confirmed by the expression of myogenic transcription factor, Pax7. We found the aged MuSC pretreated with EVs from MPC promoted clonal expansion and restored myogenic potential in vitro with more myotubes formed after 72h differentiation in medium with 2% horse serum compared with PBS pretreated MuSC (MuSC (Fusion index: 40.3±5.2% vs.15.0± 2.1%, n=3).). To test the regenerative potential of the EVs on aged MuSC, 8 weeks old mdx/scid mice (dystrophin deficient mice) were irradiated to destroy the host MuSC and then injured their tibialis anterior (TA) muscle with cardiotoxin (CTX). 24 hours after injury, we transplanted the aging MuSC (5000 cells) either with EV or PBS pretreatment into TA muscles, respectively. 5 days after transplantation, the EVs pretreated MuSC increased muscle regeneration with muscle fibers positive for embryonic myosin heavy chain compared to PBS pretreated MuSC. One month later, we found transplantation of EVs pretreated MuSC restored dystrophin expression in these mdx mice. We further tested the effects of EVs pretreatment on MuSC in 25 months old mice following CTX injury. Consistent with the results in mdx mice, pretreatment of mice with EVs from Givi‐MPC showed enhanced regeneration (6‐fold increase in number of new muscle cells with central nuclei) in CTX injured aged muscle accompanied by an increase of Pax7 positive MuSC compared with PBS treated mice or treatment with EVs from human myoblasts (number of Pax7 positive MuSC: EVs‐Givi‐MPC 10±5/mm2; PBS 2±1/mm2; EVs‐human myoblast 6±2/mm2, n=3).
Conclusion
The regenerative potential of aging MuSC is restored with the treatment of EVs from Givi‐MPC allowing stem cell renewal and differentiation upon activation.