Perfusable adipose decellularized extracellular matrix biological scaffoldco-recellularizedwith adipose-derived stem cellsand L6 promotes functional skeletal muscle regeneration following volumetric muscle loss
In this study, we used a recellularization method of perfusing cells into biological scaffolds through vascular pedicles. ASCs and L6 cells were co-seeded into adipose decellularized extracellular matrix (adECM) biological scaffolds. On one hand, this strategy ensures uniform distribution of seeded cells in the scaffold and avoids the formation of non-functional "muscle islands" later. On the other hand, co-seeding of L6 cells and ASCs addresses the issue of low myogenic differentiation potential of ASCs. Subsequently, the recellularized biological scaffold treatment of VML animal experiments showed that the combined recellularization strategy had significantly better muscle regeneration and angiogenesis than the single ASCs recellularization strategy, and the former had better functional recovery of tibialis anterior muscle (TA). Further single cell sequencing analysis found that compared to singly seeded ASCs in biological scaffolds, L6 cells in the combined recellularization induced ASCs to transform into a new subpopulation of cells highly expressing Mki67, CD34 and CDK1 genes, which had stronger ability of oriented myogenic differentiation and formed more mature and larger muscle fibers, significantly restoring TA muscle function. Overall design: Healthy Sprague Dawley male rats aged 8 weeks were used to establish the tibialis anterior (TA) muscle VML model (n = 10). The right hind leg skin of the rat was incised, and the fascia and muscle were separated. A muscle defect (7 × 10× 3 mm) was excised from TA. Simultaneously, the tissue constructs could be implanted at this point.adECM + ASCs group: The adECM was implanted in the VML defect with ASCs recellularization; adECM+ASCs+L6 group: The adECM was implanted in the VML defect with ASCs+L6 co-recellularization. Both side of the muscle of each rat were removed from the 7day of modeling for snRNA-seq.