The design of multifunctional biomaterials that offer both structural support and bio-chemical cues is essential for enhancing tissue regeneration. In this study, hybrid nan-ofibrous scaffolds composed of poly(L-lactide-co-ε-caprolactone) (PLCL) and bioactive factors secreted by Wharton’s jelly-derived mesenchymal stem cells (WJ-MSCs) were fabricated via co-electrospinning. Nanofibers were produced in both aligned and ran-dom configurations following an optimized protocol developed at the Institute for Bi-oengineering of Catalonia (IBEC). Morphological and topographical features were characterized using light microscopy and atomic force microscopy (AFM), while fiber orientation was quantitatively assessed through Fast Fourier Transform (FFT) analysis. The controlled release kinetics of FITC-labeled bioactive compounds were evaluated, and human adipose-derived MSCs (AD-MSCs) were used as a cell model to evaluate scaffold biocompatibility, in respect to cell viability, adhesion, proliferation, and mi-gration. FFT analysis was also applied to quantify the orientation of AD-MSCs when cultured on the nanofibers, revealing higher cellular anisotropy and alignment on ori-ented scaffolds. We further show that aligned nanofibers supported cell viability and proliferation, as well as directed migration as wound closure assays showed signifi-cantly faster healing on aligned nanofibers compared to random ones. These results emphasize the synergistic effects of nanofiber alignment and biochemical functionali-zation in modulating cell behavior and promoting tissue regeneration, underscoring the potential of PLCL-based hybrid nanofibers for advanced wound healing applications.