Evaluation of a biodegradable matrix containing cultured human fibroblasts as a dermal replacement beneath meshed skin grafts on athymic mice.

Meshed, expanded split-thickness skin grafts (MSTSG) frequently achieve poor results when used to cover full-thickness wounds. Poor cosmetic and functional results occur in part because the epithelium that grows across the skin graft interstices lacks a dermis. We used a living dermal replacement composed of either polyglycolic acid (PGA) or polyglactin-910 (PGL) mesh containing confluent, cultured human fibroblasts. These grafts were applied to full-thickness wounds on athymic mice; widely expanded, 3:1 ratio human MSTSG was then placed over the dermal graft. Histologic examination of wounds during a 99-day period after graft placement showed that PGA/PGL-fibroblast grafts vascularized to the wound, and the MSTSG simultaneously vascularized to the PGA/PGL-fibroblast graft. Epithelialization from the MSTSG bridges proceeded rapidly across the surface of the PGA/PGL-fibroblast grafts, resulting in an epithelialized layer that covered a densely cellular substratum that resembled dermis. Basement membrane formation at the dermal-epidermal junction of the epithelialized interstices was confirmed by immunohistochemical microscopy. Minimal inflammatory reaction to the PGA/PGL-fibroblast grafts was seen. Grafts composed of PGA or PGL biodegradable meshes combined with cultured fibroblasts vascularize in full-thickness wounds, resulting in formation of organized tissue beneath the epithelialized surface that resembles dermis.     

在胶原海绵膜上构建人表皮细胞和成纤维细胞复合移植物的实验研究

目的将原代培养的人表皮细胞和成纤维细胞，接种到一种新的载体-胶原海绵膜上，构建新的复合皮肤替代物。方法来源于人包皮的表皮细胞和成纤维细胞分别原代培养，成纤维细胞经消化后接种在胶原海绵膜上(1×10     

表皮细胞、成纤维细胞和几丁质-胶原蛋白复合人工皮的体外构建

目的 探讨几丁质—胶原蛋白膜作为表皮细胞、成纤维细胞培养支架的可能性，体外构建包含双层细胞的复合皮。方法 取健康成人环切包皮，分离成纤维细胞、表皮细胞，分别制成单细胞悬液，将几丁质-胶原蛋白膜(有孔面向上)平铺于60mm培养皿中，首先接种成纤维细胞，培养2d后，将含10％小牛血清的DMEM更换成完全型DMEM，翻转膜，使光滑面朝上，再种植表皮细胞，每日换液。定期观察细胞与材料的粘附、细胞贴壁及其生长增殖情况。结果 几丁质—胶原蛋白膜对成纤维细胞、表皮细胞无明显毒性作用，成纤维细胞培养1d后，细胞贴附于材料支架，细胞胞体较大，呈典型的梭形。加入表皮细胞复合培养2d示表皮细胞贴壁生长，并分化、增殖。复合皮构建2周，网格支架及孔内均有大量细胞生长，膜表面细胞融合成片，表皮细胞分化形成复层。结论 几丁质—胶原蛋白膜对细胞无毒性，有利于培养细胞的粘附、生长。在体外可以构建成功类似生理性皮肤的人工皮肤。     

Genetic modification of cultured skin substitutes by transduction of human keratinocytes and fibroblasts with platelet-derived growth factor-A

Gene therapy promises the potential for improved treatment of cutaneous wounds. This study evaluated whether genetically modified cultured skin substitutes can act as vehicles for gene therapy in an athymic mouse model of wound healing. Human keratinocytes and fibroblasts were genetically engineered by retroviral transduction to overexpress human platelet-derived growth factor-A chain. Three types of skin substitutes were prepared from collagen-glycosaminoglycan substrates populated with fibroblasts and keratinocytes: HF-/HK-, containing both unmodified fibroblasts and keratinocytes; HF-/HK+, containing unmodified fibroblasts and modified keratinocytes; and HF+/HK-, containing modified fibroblasts and unmodified keratinocytes. Skin substitutes were cultured for two weeks before grafting to full-thickness wounds on athymic mice. The modified skin substitutes secreted significantly elevated levels of platelet-derived growth factor throughout the culture period. Expression of retroviral platelet-derived growth factor-A mRNA was maintained after grafting to mice, and was detected in all HF-/HK+ grafts and one HF+/HK- graft at two weeks after surgery. Although no differences were seen between control and modified grafts, the results suggest that genetically modified cultured skin substitutes can be a feasible mechanism for cutaneous gene therapy. The cultured skin model used for these studies has advantages over other skin analogs containing only epidermal cells; because it contains both fibroblasts and keratinocytes, it therefore offers greater opportunities for genetic modification and potential modulation of wound healing.     

One-step grafting procedure using artificial dermis and split-thickness skin in burn patients

The study aimed to achieve a one-step grafting procedure using artificial dermis and split-thickness skin. We performed simultaneous grafting of artificial dermis and skin in two severely burned patients. Artificial dermis was treated with fresh autogenous platelet-derived wound-healing factors (PDWHF), cryopreserved allogeneic cultured endothelial cells, and fibroblasts. Dermal microvascular endothelial cells and fibroblasts were obtained from a single human donor’s skin. The cultured cells were cryopreserved until use in grafting. The PDWHF was prepared from autogenous blood from each patient prior to the surgery. In two patients, the artificial dermis treated with this method was grafted to a full-thickness burn wound. Immediately after artificial dermis grafting, meshed split-thickness skin was grafted. In each case, the skin graft took well, and the skin texture was acceptable. Histological examination revealed that bovine collagen tissue remained in the dermis after surgery, indicating the success of the simultaneous grafting of the artificial dermis and the skin. The present study indicates that one-step grafting of artificial dermis and split-skin is possible when the artificial dermis is treated with PDWHF and cultured endothelial cells and fibroblasts. Level of Evidence: Level V, therapeutic study.     

The efficacy of collagen dermis membrane and fibrin on cultured epidermal graft using an athymic mouse model

A human skin substitute consisting of human cultured keratinocytes, collagen dermis, and fibrin was evaluated in athymic mice. Eighty athymic mice were divided randomly into four groups. A 1.5x1.5-cm full-thickness wound defect was created on the back of each athymic mouse under anesthesia. These wounds were covered by sheets of cultured epidermal graft (group A), cultured epidermal graft with collagen dermis and fibrin (group B), cultured epidermal graft with collagen dermis (group C), or cultured epidermal graft with fibrin (group D). The grafts were secured and kept moist by specially designed saline gauze chambers. The take rates of the cultured graft with more than 50% of the wound covered were 65%, 15%, 50%, and 45% respectively. Group B had a significantly lower graft take rate, however the difference was not significant among groups A, C, and D. Light microscopy of biopsies of the grafted sites at 12 days showed complete epithelialization. The incidence of discharge from wound beds in groups A, B, C, and D was 0%, 15%, 15%, and 10% respectively. The results suggest that cultured cells are best grafted directly onto the wound bed or in combination with either a thin layer of collagen or fibrin but not both because the collagen dermal membrane and the fibrin together may impose too great a diffusion barrier for the cultured cell graft to become vascularized.     

用含活性细胞的胶原支架复合皮修复小鼠全层皮肤缺损的研究

目的构建具有活性细胞成分的复合皮，观察其在修复小鼠全层皮肤缺损中的作用。方法将异种成纤维细胞种植于脱软骨细胞胶原支架上培养3d，形成真皮替代物。再将其放置于培养的上皮细胞膜上共同培养10d，组成含活性细胞成分的复合皮。将复合皮移植于小鼠的全层皮肤缺损处，记录其生长情况并于术后定期活检，进行组织学观察。结果胶原支架中可见生长良好的成纤维细胞和复层表皮细胞。移植术后1周，复合皮与小鼠缺损创面粘连紧密，可见明显的血管化。术后6周，创面愈合良好，移植物与创缘融合，未见明显排斥反应。结论以脱软骨细胞胶原为支架构建的复合皮，可作为皮肤替代物修复全层皮肤组织缺损。     

In vivo cultured skin composed of two-layer collagen sponges with preconfluent cells.

Although various kinds of cultured skin substitutes have been developed, it takes several weeks to produce them before grafting. In their previous study, the authors succeeded in producing cultured skin easily in a short period of time by layering two collagen sponges. In the current study, to shorten this period even further, they grafted the cell-preconfluent artificial skin immediately after seeding the cells. They used two collagen sponges with different pore sizes and crosslink densities. They seeded 1,000,000 cells per square centimeter of fibroblasts and 1,000,000 cells per square centimeter of keratinocytes on the respective collagen sponges and grafted them on a full-thickness, excised wound on the back of severe combined immunodeficient mice. Two weeks after grafting, epithelium and dermislike tissue were formed. They then decreased the number of keratinocytes and grafted them. Four weeks after grafting, at seeding densities of 50,000 to 1,000,000 cells per square centimeter of keratinocytes, the preconfluent artificial skin took histologically, and human type IV and type VII collagen were stained immunohistochemically. This cell-preconfluent artificial skin composed of two-layer collagen sponges seems promising for widespread clinical use.     

An experimental study on the repair of full skin loss of nude mice with composite graft of epidermal stem cells

This study is to constitute a composite skin substitute with epidermal stem cells (ESCs) and fibroblasts on collagen sponge. ESCs were selected by rapid attachment to collagen IV for 10 min. Collagen was extracted from rat's tail. The matrix lattice was fabricated by freeze-dryer and cross-linked with glutaraldehyde. Fibroblasts were inoculated on collagen sponge and cultured for 1 week prior to inoculation of ESCs. Having cultured for 2 weeks in submerged culture, the bioengineered tissue was raised to the air-liquid interface and cultured for 2 weeks. The artificial skin was then grafted onto full skin loss wounds of nude mice. Collagen sponge membrane lacking cell inoculation and an artificial skin with epidermal cells (ECs) and fibroblasts were used as controls. The wounds were observed daily. Tissue samples were harvested and examined by means of histology, immunohistochemistry and electron microscopy. The wounds in the test group healed at a significantly faster rate than controls, with good skin appearance and minimal scar formation. The control group showed delayed wound healing and intensive wound contraction as compared to the test group. Thus the skin substitute with ESCs seemed to be a good equivalent.     

The effects of combined application of autogenous fibroblast cell culture and full-tissue skin graft (FTSG) on wound healing and contraction in full-thickness tissue defects

IntroductionThis study investigates the effects of culture grown fibroblasts on contraction and dermal regeneration when used concurrently with full-thickness skin graft (FTSG) in full-thickness wounds.Materials and methodsFourteen Sprague-Dawley male rats were divided into two groups. In the first group, wound contraction was evaluated visually. Two full thickness tissue defects were produced on the back of the seven rats. The skin harvested from these areas was prepared as a full-thickness graft and sutured back to their original beds. Just before the last suture, autogenous fibroblast suspension was applied between the graft and the bed in area 1, and area 2 served as control. The surface area of grafts were calculated and compared with “Image J” program. In the second group, contraction and dermal regeneration were evaluated histologically. Three full-thickness tissue defects were produced on the back of seven rats. Area 1 and 2 were prepared as described above and area 3 was left to secondary healing. On the 14th and 30th days, punch biopsies were harvested from the center of the areas 1–3. Preparations were examined under light microscopy.ResultsWound contraction was significantly less in area 1 on day 14 (p < 0.01). Histologically neovascularization, fibroblast density and collagen synthesis were more evident in cultured fibroblast applied areas on day 14. However epithelialization did not show any difference between areas both on days 14 and 30. On day 30, area 1 still a higher degree of fibroblast intensity than the other areas but neovascularization and collagen synthesis were not different than the other areas.ConclusionAccording to the data obtained from the study, cultured fibroblasts, particularly with a dermal support, do not regress when transplanted to a living tissue. They contribute to the wound healing process; reduce the contraction of the wound; and support collagen synthesis and neovascularization.     

Reconstruction of a full-thickness alar wound using an auricular conchal composite graft

Nasogastric intubation has become a frequently used method for alleviating gastrointestinal symptoms. Necrosis from alar pressure during prolonged nasogastric and nasotracheal intubation is common, and can result in considerable deformity if it is unrecognized. The reconstruction of full-thickness alar wounds often requires multiple challenging surgical procedures. Difficult full-thickness alar defects often require nasal mucosal replacement for lining, cartilage batten graft support for the preservation of nasal function, and skin coverage for the restoration of an aesthetically correct appearance. Free composite conchal grafting can offer a single-staged, one-step repair of difficult full-thickness alar wounds that are no larger than 1.5 cm in size. A thorough explanation of the graft design and execution is presented, as well as a case report and literature review. Free composite conchal grafting can produce aesthetic and functional results that rival the most sophisticated flap reconstructions of the lateral ala.     

Evaluation of Permacol as a cultured skin equivalent

Skin loss following severe burn requires prompt wound closure to avoid such complications as fluid and electrolyte imbalance, infection, immune suppression, and pain. In clinical situations in which insufficient donor skin is available, the development of cultured skin equivalents (dermal matrices seeded with keratinocytes and fibroblasts) may provide a useful alternative. The aim of this study was to assess the suitability of a porcine-derived dermal collagen matrix (Permacol™) to function as a cultured skin equivalent in supporting the growth of keratinocytes in vitro and providing cover to full thickness wounds in the BALB C/nude mouse model. A histological comparison was against Glycerol treated-Ethylene Oxide Sterilised Porcine Dermis (Gly-EO Dermis) which has successfully been used as a cultured skin equivalent in previous studies. Both Gly-EO Dermis and to a lesser extent Permacol™ were able to support the growth of cultured keratinocytes following a 16-day period of cell culture, however, this study was only able to demonstrate the presence of an epidermal layer on Gly-EO dermis 2 weeks after grafting onto full-thickness wounds in the BALB C/nude mouse model.     

Viability and function of autologous and allogeneic fibroblasts seeded in dermal substitutes after implantation

BACKGROUND: Fibroblast-seeded collagen sponges have been used for the treatment of skin defects and skin ulcers. However, the viability of the fibroblasts after implantation is still unknown. The objective of this study was to investigate the viability and distribution of autologous and allogeneic fibroblasts after implantation and to clarify which type is more effective for wound healing. MATERIALS AND METHODS: Skin samples of Hartley guinea pigs were retrieved and autologous fibroblasts were isolated and cultured. Fibroblasts isolated from the skin of a Strain2 guinea pig were used as allogeneic fibroblasts. Three full-thickness wounds were created on the backs of guinea pigs and an acellular collagen sponge, a collagen sponge seeded with autologous fibroblasts, and a collagen sponge seeded with allogeneic fibroblasts were transplanted. Before implantation, fibroblasts were labeled with PKH26. The guinea pigs were sacrificed 1, 2, and 3 weeks after implantation. The epithelization and contraction of the wounds were assessed, and the viability and distribution of the seeded fibroblasts were observed in cross sections. RESULTS: Three weeks after implantation, the PKH26-labeled autologous and allogeneic fibroblasts remained viable. In the wounds covered with the autologous fibroblast-seeded collagen sponge, the epithelization was fastest, and the percent wound contraction was smallest. In contrast, in the wounds covered with allogeneic fibroblasts, the epithelization was slowest and the percent contraction was largest. CONCLUSION: The allogeneic fibroblasts seeded in the collagen sponge survived and remained viable on the grafted area, but did not accelerate wound healing.     

Development and evaluation of a new composite Laserskin graft

BACKGROUND: Tremendous effort has been made to improve the graft take rate of cultured epidermal autograph. The purpose of this study is to develop and evaluate a new composite Laserskin graft (CLSG) as a human skin substitute for wound resurfacing. METHODS: The seeding efficacy of cultured keratinocytes on plain Laserskin was compared with the 3T3 cell-seeded Laserskin and allogenic fibroblast-populated Laserskin. Three different types of CLSG, 2 cm in diameter each, were prepared and tested in rats. Type A CLSG consisted of proliferative allogenic rat fibroblasts on both sides of the Laserskin with autologous keratinocytes also on the upper side. Fibroblasts and keratinocytes were seeded only on the upper side of the Laserskin in type B CLSG. Keratinocytes alone were seeded on plain Laserskin in type C CLSG. Type B CLSG consisting of autologous keratinocytes and autologous dermal fibroblasts was tested on five selected wounds (5x5 cm each) of a patient with full-thickness burn. In another burn patient, type B CLSG consisting of autologous keratinocytes and allogenic dermal fibroblasts was grafted onto three wounds (5x5 cm each). RESULTS: The seeding efficacy of human keratinocytes on plain Laserskin increased from 75% to 95% when proliferative allogenic fibroblasts were grown as a feeder layer on the Laserskin. The seeding efficacy of rat keratinocytes increased from 36% to 88% in the presence of a proliferative allogenic fibroblast feeder layer, whereas human/rat keratinocytes had respective seeding efficacy of 98%/91% on Laserskin preseeded with mitomycin C-treated 3T3 cells. Skin biopsies of grafted type A CLSG on day 14 after grafting showed complete epithelialization without severe inflammation in 16 of 20 (80%) grafted surgical wounds in rats. There were eight (40%) and seven (35%) "takes" of the CLSG in types B and C, respectively. The infection rate in type B CLSG was two (10%). There was one (5%) infection in types A and C. The respective take rates on the two patients grafted with type B CLSG were 60% and 100%. CONCLUSION: The animal experiment and the preliminary clinical data showed that the CSLGs consisting of autologous keratinocytes and of autologous/allogenic fibroblasts are good human skin substitutes in terms of durability, biocompatibility, high seeding efficacy for keratinocytes, high graft take rate, and low infection rate.     

The application of new biosynthetic artificial skin for long-term temporary wound coverage

Temporary dressings protect wounds from desiccation and infection. In our previous study, we used meshed acellular porcine dermis (APD) to enhance wound healing and decrease wound contraction; however, the wounds showed meshed scar [Wang HJ, Chen TM, Cheng TY. Use of a porcine dermis template to enhance widely expanded mesh autologous split-thickness skin graft growth: preliminary report. J Trauma 1997;42(2):177–82]. In this study, we produced an artificial skin composed of a cross-linked silicon sheet on the surface of APD which we have called silicone acellular porcine dermis (SAPD). This new artificial skin can protect the wound long enough to promote wound healing either by second intention or covered long enough until cultured epithelium autograft (CEA) or autologous skin graft can be harvested for permanent coverage.

We delivered 4 cm × 5 cm full-thickness wound on the back of 350 g Sprague–Dawley rats. Thirty-six rats were divided into two groups. Eighteen rats had SAPD and the other 18 were covered with Biobrane. The wounds were first examined 2 weeks after grafting and followed weekly for an additional 4 weeks to evaluate the wound and study pathological changes by using H.E. and Masson's stains. Wound size was calculated by ruler and analyzed by Student's t-test.

At the 2-week inspection, both SAPD and Biobrane showed tight adherence to the wound with no change of wound size. Both the SAPD and Biobrane dermal templates were pink. In the Biobrane-covered group, the wounds contracted soon after the tie-over dressing was removed. Its dermal layer is a layer of thin porcine dermal substance, which was promptly digested by tissue hyaluronidase and provides no real dermal template. In the SAPD-covered group however, the wound size was maintained significantly from third to sixth week after grafting (p < 0.001). SAPD was designed with thick epidermal silicone and a well-organized porcine dermis so that it incorporates into the recipient wound. Clinically the silicone layer of SAPD dislodged from APD about 6–7 weeks after grafting and was followed by dermal matrix exposure and infection. In pathological examination, much like a human skin graft, new vessels were found in APD about 1 week after grafting with minimal inflammatory cells infiltrated in the graft and wound. Six weeks after grafting, the collagen of APD incorporated into the wound, showing palisade arrangement and no sign of rejection. In the Biobrane group however, the wounds showed severe inflammation, the porcine dermal matrix was digested and disappeared 3 weeks after coverage.

In conclusion, SAPD is a thick biosynthetic artificial skin, which protects the rat wound significantly longer than Biobrane and prevents contraction. We expect that using of SAPD for temporary wound coverage will provide enough time to grow autologous-cultured epithelium or to reharvest skin grafts.     

Tissue-engineered product: allogeneic cultured dermal substitute composed of spongy collagen with fibroblasts

Recently, various types of allogeneic skin substitutes including cultured epidermal substitute (CES), cultured dermal substitute (CDS), and cultured skin substitute (CSS), which are composed of keratinocytes and/or fibroblasts as the cellular component(s), have been used as biological wound dressings. In our study, the allogeneic CDS was prepared by plating fibroblasts on a spongy collagen. The clinical evaluation was conducted using fresh or cryopreserved allogeneic CDS. In 145 of our clinical cases, 95% (138/145) of various wounds were evaluated as achieving good or excellent results, including 96% (22/23) of deep dermal burns (DDB) and dermal burns (DB), 100% (53/53) of partial-thickness donor wounds, 91% (21/23) of traumatic skin defects, 100% (5/5) of pressure ulcers, 82% (9/11) of chronic skin ulcers, 100% (6/6) of coverage for debrided DB, and 92% (22/24) of coverage for autologous meshed graft. The results obtained in our study suggest that the allogeneic CDS is able to provide an effective therapy for patients with partial and/or full-thickness skin defects.     

Fibroblasts improve performance of cultured composite skin substitutes on athymic mice

BACKGROUND: This study investigated the impact of adding human fibroblasts to a cultured composite skin substitute model of cultured human keratinocytes and acellular human dermis. METHODS: Skin substitutes were prepared by seeding human keratinocytes on the papillary side of acellular dermis with or without seeding fibroblasts on the reticular side. Performance of the grafts was compared both in vitro by histology and in vivo on surgically created full-thickness wounds on athymic mice. Graft size and contraction were measured and immunohistochemical stains were done to reveal vascularization. RESULTS: Skin substitutes with fibroblasts formed thicker epidermis than skin substitutes without fibroblasts. When transplanted onto athymic mice, skin substitutes with fibroblasts maintained their original size with only 2% contraction. In contrast, skin substitutes without fibroblasts showed 29% contraction. Vascular basement membrane specific mouse CD31staining and endothelial cell specific mouse collagen type IV staining revealed vascularization as early as 1 week posttransplant in grafts with fibroblasts, and was significantly higher than grafts without fibroblasts at 2 weeks. CONCLUSIONS: Addition of fibroblasts to keratinocyte based composite skin substitutes improves epidermis formation, enhances vascularization and reduces contraction.     

Cultured human keratinocytes as a single cell suspension in fibrin glue combined with preserved dermal grafts enhance skin reconstitution in athymic mice full-thickness wounds

Cultured human keratinocytes as a single cell suspension in fibrin glue combined with preserved dermal grafts enhance skin reconstitution in athymic mice full-thickness wounds. The technique of transplanting cultured human keratinocytes suspended as single cells in a fibrin-glue matrix (KFGS) has been recently developed to overcome common disadvantages of standard cultured epidermal sheet grafts. The combination of this method with glycerolized (nonvital) xenograft overlays in standardized nude mice full-thickness wounds, as compared to KFGS alone or controls with no grafts, showed enhancement of epithelial regeneration in terms of epithelial thickness and diminished wound contraction during the 6-week follow-up. Total scar thickness was increased after the combined KFGS/xenograft technique. The time taken to complete wound reepithelialization was similar in the two groups. Reconstitution of the dermo-epidermal junction zone as shown by electron microscopy and immunohistochemistry was enhanced by the KFGS+xenograft technique, showing structures resembling rete ridges 6 weeks postoperatively. The combined KFGS/xenograft technique is able to transfer proliferative single keratinocytes. The method simplifies the application when compared to conventional epithelial sheet grafting and reduces wound contraction when compared to pure keratinocyte grafting. Received: 15 October 1998 / / Accepted: 10 March 1999     

Long-term remodeling of a bilayered living human skin equivalent (Apligraf®) grafted onto nude mice: immunolocalization of human cells and characterization of extracellular matrix

Type I collagen is a clinically approved biomaterial largely used in tissue engineering. It acts as a regenerative template in which the implanted collagen is progressively degraded and replaced by new cell-synthesized tissue. Apligraf, a bioengineered living skin, is composed of a bovine collagen lattice containing living human fibroblasts overlaid with a fully differentiated epithelium made of human keratinocytes. To investigate its progressive remodeling, athymic mice were grafted and the cellular and the extracellular matrix components were studied from 0 to 365 days after grafting. Biopsies were analyzed using immunohistochemistry with species-specific antibodies and electron microscopy techniques. We observed that this bioengineered tissue provided living and bioactive cells to the wound site up to 1 year after grafting. The graft was rapidly incorporated within the host tissue and the bovine collagen present in the graft was progressively replaced by human and mouse collagens. A normal healing process was observed, i.e., type III collagen appeared transiently with type I collagen, the major collagen isoform present at later stages. New molecules, such as elastin, were produced by the living human cells contained within the graft. This animal model combined with species-specific immunohistochemistry tools is thus very useful for studying long-term tissue remodeling of bioengineered living tissues.     

Preliminary clinical studies of a biological skin equivalent in burned patients

The possibility of covering large areas of full thickness skin loss with 'living skin equivalent' produced by a modification of Bell's method was studied. Living skin equivalents, composed of a dermal equivalent (fibroblasts plus collagen) covered by epithelial cells were grafted, meshed or non-meshed, onto granulation tissue and, in one patient, onto fascia. Eight patients with full skin thickness burn wounds covering over 15 per cent of the body surface area were thus partially covered. The graft 'take' was evaluated every 48 h. In every patient grafted, an extensive lysis (60-90 per cent) of the skin equivalent graft was observed at the first dressing (48 h). In one patient only, a significant percentage of 'take' (40 per cent) was observed 14 days after grafting. These disappointing results were probably related to the presence of collagenases or proteases produced on the wound bed either by bacteria or by surrounding human cells. It appears that at the present time the biochemical nature of the dermal equivalent used is not yet completely appropriate to serve routinely as a substitute for human skin.