Upward migration of cultured autologous keratinocytes in IntegraÔ artificial skin: a preliminary report ISABEL JONES, FRCS; S. ELIZABETH JAMES, PhD; PHILIP RUBIN, FRCS; ROBIN MARTIN, PhD The combination of cultured autologous keratinocytes with the dermal regeneration template IntegraÔ could offer increased possibilities for reconstructive surgery and wound healing. A single-step application of cells, centrifuged deep into an IntegraÔ-like matrix at the silicone–matrix junction, has been described but might prove technically complex for clinical use. We have investigated the possibility of simplifying this procedure by applying cultured cells directly to the underside of the IntegraÔ or directly to the wound bed immediately prior to grafting. The objective was to see whether cells would migrate through the matrix in an upward direction. We tested the principle of this concept using a pig wound healing model. IntegraÔ was seeded directly with cultured cells and grafted onto fresh full- thickness wounds, or unseeded IntegraÔ was applied to freshly excised wound beds that had just been seeded with the same number of cells. Biopsies were taken at 3, 7, 11, and 14 days. Histological sections showed that the cells moved through the IntegraÔ to give a confluent surface epithelium. Direct seeding onto the IntegraÔ was the most efficient method. Transduction of cultured autologous keratinocytes in vitro with a MFGlacZnls retrovirus confirmed that the epidermis was derived from the cultured autologous keratinocytes. (WOUND REP REG 2003;11:132–138) Integra TM artificial skin is a commercially available dermal regeneration template for use in reconstructive surgery and in burns. 1 It is a bilayer product. The deeper layer is a collagen/chondroitin sulfate matrix that is replaced by new collagen synthesized by fibroblasts that grow in from the wound bed. The template matrix serves to regulate the in-growth of new tissue and hence minimize scarring. The purpose of the silicone sheet, which forms the superficial layer, is to protect the wound from infection and excessive loss of moisture and to limit granulation tissue formation. 2 Once the collagen layer has been integrated into the wound bed, the silicone layer begins to separate spontaneously. At this point it can be peeled away and replaced with an ultra- thin split-thickness skin graft (SSG) of approximately 0.1 mm. Whereas an average SSG measures 0.2–0.4 mm, due to the underlying IntegraÔ matrix, the ultra-thin SSG does not contract to the extent that would be expected if it were applied directly to the wound bed. This is supported by a long-term study. 3 Cultured autologous keratinocyte (CAK ) sheets have been proposed as an alternative form of definitive wound CAK Cultured autologous keratinocytes C-GAG Collagen-glycosaminoglycan FITC Fluorescein isothiocyanate H&E Hematoxylin & eosin SSG Spilt-thickness skin graft From the Blond McIndoe Research Center, Queen Vic- toria Hospital, East Grinstead, United Kingdom. This work was first reported, in part, at the 2000 ETRS meeting: Jones IT, James L, Rubin P, Martin R. Retroviral labeling as evidence of epidermal regeneration from autologous disaggregated cultured keratinocytes applied beneath Inte- graÔ artificial skin. Wound Rep Reg 2000;8:A420. Current addresses: Isabel Jones: Heatherwood Hospital, Ascot, Berkshire; P.R.: Department of Plastic Surgery, Nottingham City Hospital, Sheffield; R.M.: Smith & Nephew Group Research Center, Heslington, York, United Kingdom. Correspondence: Dr. Liz James, PhD, Blond McIndoe Center, Queen Victoria Hospital, East Grinstead, West Sussex, RH19 3DZ, United Kingdom. Fax: 44- 1342-301701; Email: liz.j@blondmcindoe.demon. co.uk. Copyright Ó 2003 by the Wound Healing Society. ISSN: 1067-1927 $15.00 + 0 132