5-Azacytidine incorporated skeletal muscle-derived hydrogel promotes rat skeletal muscle regeneration

Decellularized skeletal muscle is a promising biomaterial for muscle regeneration due to the mimicking of the natural microenvironment. Previously, it has been reported that 5-Azacytidine (5-Aza), a DNA methyltransferase inhibitor, induces myogenesis in different types of stem cells. In the current study, we investigated the effect of 5-Aza incorporated muscle-derived hydrogel on the viability and proliferation of muscle-derived stem cells (MDSCs) in vitro and muscle regeneration in vivo. Wistar rat skeletal muscles were decellularized using a physico-chemical protocol. The decellularized tissue was analyzed using SEM, histological staining and evaluation of DNA content. Then, muscle-derived hydrogel was made from Pepsin-digested decellularized muscle tissues. 5-Aza was physically adsorbed in prepared hydrogels. Then, MDSCs were cultured on hydrogels with/without 5-Aza, and their proliferation and cell viability were determined using LIVE/DEAD and DAPI staining. Moreover, myectomy lesions were done in rat femoris muscles, muscle-derived hydroges with/without 5-Aza were injected to the myectomy sites, and histological evaluation was performed after three weeks. The analysis of decellu-larized muscle tissues showed that they maintained extracellular matrix components of native muscles, while they lacked DNA. LIVE/DEAD and DAPI staining showed that the hydrogel containing 5-Aza supported MDSCs viability. Histological analysis of myectomy sites showed an improvement in muscle regeneration after admin-istration of 5-Aza incorporated hydrogel. These findings suggest that the combination of 5-Aza with skeletal muscle hydrogel may serve as an alternative treatment option to improve the regeneration of injured muscle tissue.