Preparation and characterization of skin cell sheets for treatment of burn wounds

Wound closure is a crucial step in the treatment of burns, often performed using autologous skin graft. Unfortunately, in extensive burn injuries, only limited area of normal skin can be used to make a graft which is not enough to cover the entire wound surface. To overcome the shortage of the skin donor site, a skin graft constructed from a sheet of co-cultured human dermal fibroblast and keratinocyte has been proposed. In this study, human dermal fibroblasts were isolated from skin biopsies obtained from patients, hospitalized for burn injuries. To construct cell sheets, fibroblasts were grown on the commercial temperature-responsive culture surface (UpCell™) and on our UV-polymerized poly(N-isopropy-lacrylamide-co-acrylamide) (PNIAM-CO-AM), which also exhibited a phase transition with temperature. Fibroblasts grew well on both surfaces at 37°c. To detach the cells as an intact cell sheet from UpCell™ surface, the incubation temperature was lowered to 20°C for 30 minutes. On the other hand, a two-step incubation (10°C for 30 minutes and 20°C for 60 minutes) was required to detach the fibroblasts cell sheet from PNIAM-CO-AM coated surface. The qualities of the multilayered cell sheets, as assessed by histochemical staining show that both surfaces yielded high quality tissue-like sheet consisting of healthy fibroblasts.