Adipose-derived stem cells (ADSC) are nowadays one of the most exploited cells in regenerative medicine. They are fast growing, capable of enhancing axonal elongation, support and locally stimulate Schwann cells (SCs), and protect de-innervated muscles from atrophy after a peripheral nerve injury. With the aim of developing a bio-safe, clinically translatable cell-therapy, we assessed the effect of ADSC pre-expanded with human platelet lysate in an in vivo rat model, delivering the cells into a 15 mm critical-size sciatic nerve defect embedded within a laminin-peptide-functionalized hydrogel (Biogelx-IKVAV) wrapped by a poly-ɛ-caprolactone (PCL) nerve conduit. ADSC retained their stemness, their immunophenotype and proliferative activity when tested in vitro. At 6 weeks post-implantation, robust regeneration was observed across the critical-size gap as evaluated by both the axonal elongation (anti-NF 200) and SC proliferation (anti-S100) within the human ADSC-IKVAV filled PCL conduit. All the other experimental groups manifested significantly lower levels of growth cone elongation. The histological gastrocnemius muscle analysis was comparable with no quantitative significant differences among the experimental groups. Taken together, these results suggest that ADSC encapsulated in Biogelx-IKVAV are a potential path to improve the efficacy of nerve regeneration. New perspectives can be pursued for the development of a fully synthetic bioengineered nerve graft for the treatment of peripheral nerve injury.