A Biomimetic Basement Membrane Substitute Based on Tri-Layered Nanofibrous Scaffold for Skin Reconstruction

Developing basement membranes (BMs) substitute remains major problem for constructing functional tissue engineered skin because of its complex structure and multifunction of regulating cellular behavior. Herein, a stable electrospinning method was employed to generate a biomimetic model of natural BMs based on novel scaffold electrospun from Poly(varepsilon-caprolactone) (PCL) and cellulose acetate (CA) incorporated with chitosan (CS). The morphology, structure, surface hydrophilicity, roughness and mechanical tensile strength of prepared monolayer and tri-layered scaffold were comprehensively compared. Besides, co-culture system via seeding keratinocytes (Kcs) and fibroblasts (Fbs) on opposite side of tri-layered scaffold revealed more effective segregation of both cell types within the central nanofibrous barrier together with enhanced cell attachment and proliferation than that on the monolayer scaffold. Moreover, the deposition of type VII collagen and laminin-5 was examined in comparison with normal skin BMs. Furthermore, the histological studies revealed characteristics of reconstructing BM zone at the junction of dermis-epidermis after in vivo implantation for 2 weeks, and wound healing while the seeded cells interacted with the endogenous cells. Additionally, the expression of active integrin beta1 and phosphorylated focal adhesion kinase (FAK) was promoted with treatment of tri-layered scaffold. This study stressed that this tri-layer scaffold might provoke biomimetic responses of Kcs and Fbs and thus be applied for future development of BMs containing tissues.