Juvenile vs Adult Skeletal Muscle Transplants in the Treatment of Volumetric Muscle Loss Injury

Background Volumetric muscle loss (VML) causes irreversible structural and functional deficits by removing myofibers, nerves, vasculature, extracellular matrix, and satellite cells, the resident muscle stem cells essential for regeneration. Skeletal muscle transplantation can restore tissue volume and reintroduce regenerative cells, yet functional outcomes remain incomplete. Age of the donor muscle has not been evaluated, despite evidence that juvenile muscle contains higher satellite cell density and greater myogenic plasticity than adult muscle. We hypothesized that these features would yield superior regenerative outcomes when juvenile muscle is used as a transplant source. Methods Tibialis anterior (TA) muscles from juvenile (21 d), adolescent (34 d), and adult (~ 120 d) male Lewis rats were compared for myofiber morphology, satellite cell density, and in-vitro myogenic behavior. GFP⁺ juvenile or adult muscle was then transplanted into standardized VML defects (~ 15–20% TA volume) in adult rats. Seven weeks post-surgery, in-vivo isometric strength, donor fiber integration, satellite cell distribution, and centralized nuclei were assessed. Results Juvenile muscle exhibited ~ 15× greater satellite cell density than adult (122.8 ± 28.4 vs. 8.4 ± 3.3 cells/mm², p < 0.0001) with enhanced in-vitro differentiation (fusion index + 73% vs. adult, p = 0.0067). In-vivo, both juvenile and adult transplants restored myofiber number to control levels (juvenile: 11,369 ± 1,511; adult: 9,115 ± 1,274; controls: 10,316 ± 685) and improved strength versus untreated VML (juvenile: +50%, p = 0.0016; adult: +36%, p = 0.0299). No significant functional differences were observed between donor ages. Donor fibers integrated but remained small, with localized satellite cell enrichment and increased centralized nuclei in transplant regions, consistent with ongoing regeneration. Conclusions Juvenile skeletal muscle displays cellular and structural attributes favorable for regeneration and superior in-vitro myogenic behavior compared to adult muscle. However, these advantages did not translate into greater short-term in-vivo recovery following VML transplantation. Enhancing donor fiber hypertrophy, neuromuscular integration, and satellite cell expansion beyond the transplant region, potentially through rehabilitation or pharmaceutical interventions, may be necessary to realize the full therapeutic potential of juvenile donor muscle for regenerative medicine applications.