Cultured Skin Grafts

Abstract

ages. The use of autologous skin grafts has been the "gold standard" for wound closure, but in patients with wounds of large surface area, such as extensive burns, this strategy cannot be realized because of the lack of adequate donor sites for grafting. Because the ultimate outcome of many burn patients is determined by wound coverage, temporary coverage of wounds with materials other than the patient's own skin has been performed in these situations using fresh or cadaver skin allografts from related or unrelated donors. However, such allografts are rejected, usually after two weeks, and regrafting is necessary.1,2 See also JAMA 1988;259:2566-2571. DEVELOPMENT OF KERATINOCYTE CULTURE TECHNIQUESAs techniques for organ culture and tissue culture were developing in the 1970s, attempts were made to cover skin wounds with epidermal sheets derived from in vitro cultivation of skin. In explant cultures, a skin sample was placed with the dermal surface in contact with the culture dish to allow attachment and then incubated in culture medium. The epider¬ mis grew down the side of the expiant, attached to the dish, then migrated outward to produce epitheli¬ al layers that could be used as allografts in animal models.35 However, proliferation of epidermal cells using this method was insufficient to produce the large amounts of epidermis required for grafting, and fibroblast overgrowth in the cultures was a significant problem. By six to eight weeks, the grafted epithelium began to deteriorate, leaving only a small residue of grafted epithelium on the con¬ tracted wound. The development of techniques of in vitro cultivation and serial subculture of epidermal cells to produce viable epithelial sheets changed the situation.6 In 1975, Rheinwald and Green6 described a method allowing the successful serial subcultivation of human keratinocytes. Whole skin was trypsinized to disaggregate epidermal cells, which were then cultivated on a monolayer of lethally irradiated 3T3 mouse fibroblasts. Since that time, there have been many reported modifications in the technique, and it is now possible, beginning with a skin sample as small as 1 cm2, to expand the amount of cultured epithelium to an area comparable to body surface. Using the enzyme Dispase (a neutral protease from Bacillus polymyxa; Boehringer-Mannheim GmbH, Mannheim, West Germany), a confluent layer of epithelial cells can be detached as an intact sheet and transferred to the graft bed.7 In addition, new culture systems have been developed,8" but their relevance to large-scale grafting needs to be further docu¬ mented. CLINICAL APPLICATIONS: CULTURED AUTOGRAFTS AND CULTURED ALLOGRAFTSThe first successful clinical application of this technique was reported by O'Connor et al12 in 1981, when small skin biopsy specimens from burned patients were cultured in vitro. These cultured autografts were placed on full-thickness wounds on the arms of two adult burn patients and successfully grew to cover the wounds in six weeks. The epitheli¬ um on these wounds appeared stable for the f...

Tissue Engineering of Skin

“…It has also been reported that cultured sheet grafts eventually reform a dermal-like subepithelial region, but only after extended periods, as long as 4-5 years (Gallico and O'Connor, 1985;Tittlekow and Scott, 1986;Phillips, 1988). A characterization of this "neo-dermis" after the transplantation of keratinocyte autografts on the muscle fascia in burn patients has been reported (Woodley et al, 1990).…”

A Prospective Multicenter Study of the Efficacy and Tolerability of Cryopreserved Allogenic Human Keratinocytes to Treat Venous Leg Ulcers

“…Freshly cultured autologous and allogeneic keratinocytes are reported to be useful to treat wounds resulting from burns to leg ulcers. [11][12][13][14][15][16][17][18] Allogeneic cultured keratinocytes offer the advantages of availability and obviate the need for harvesting skin. They are reported to enhance wound healing, [19][20][21] offering a reduced mean time to complete closure.…”

Sheets of embryonic epidermis can be isolated and cultured with or without the basal lamina

“…In contrast, separation by the more traditional enzyme, trypsin, causes dispersion of the cells (Green et al, 1979). This neutral protease (DII) has also been used to obtain confluent sheets of cultured human epidermal cells to use for autografting (Green et al, 1979;Phillips 1988). In our experiments, embryonic avian epidermis could be isolated as a continuous sheet with DII for either immediate examination or organ culture experiments.…”