Abstract Background: Volumetric muscle loss (VML) is a traumatic massive loss of muscular tissue which frequently leads to amputation, limb loss, or lifetime disability. The current medical intervention is limited to autologous tissue transfer, which usually leads to non-functional tissue recovery. Tissue engineering holds a huge promise for functional recovery. 
Methods: In this work, we evaluated the potential of human adipose-derived mesenchymal stem cells (hASCs) pre-cultured in Gellan Gum based spongy-like hydrogels (SLH). 
Results: In vitro, hASCs were spreading, proliferating, and releasing growth factors and cytokines (i.e. FGF, HGF, IGF-1, IL6, IL8, IL-10, VEGF) important for muscular regeneration. After implantation into a volumetric muscle loss (VML) mouse model, implants were degrading overtime, entirely integrating into the host between 4 and 8 weeks. In both SLH and SLH+hASCs defects, infiltrated cells were observed inside constructs associated with matrix deposition. Also, minimal collagen deposition was marginally observed around the constructs along both time-points. Neovascularization (CD31+ vessels) and neoinnervation (β-III tubulin+ bundles) were significantly detected in the SLH+hASCs group, in relation to the SHAM (empty lesion). A higher density of α-SA+ and MYH7+ cells were found in the injury site among all different experimental groups, at both time-points, in relation to the SHAM. The levels of α-SA, MyoD1, and MHC proteins were moderately increased in the SLH+hASCs group after 4 weeks, and in the hASCs group after 8 weeks, in relation to the SHAM. 
Conclusions: Taken together, defects treated with hASCs-laden SLH promoted angiogenesis, neoinnervation, and the expression of myogenic proteins.