The main goal in burn management has always been increasing the survival of severely burned patients by rapid debridement and early closure of burn wounds, consequently reducing the risk of infection, sepsis and multi-organ failure. However, in the last decennia, surgical emphasis has shifted from survival to ‘quality of survival’, especially by improving the residual scars and preventing secondary contractures. Research and development of dermal skin substitutes has been directed towards improvement of scar quality by a bi-layered reconstruction of dermis and epidermis.
Allogeneic skin recovered from deceased donors is often used in the management of burns and other full thickness skin defects as a temporary coverage before definitive closure with autologous skin. Donor skin is used to improve the quality of the wound bed before grafting with autologous skin, as a biological dressing for partial thickness wounds (Hermans, 1989), or as overlay on widely expanded autograft (sandwich technique, Kreis et al., 1989). Excised full thickness defects treated with thin and widely expanded split thickness autografts often heal with an un-aesthetic, hypertrophic scar as final result as shown in figure 1.
In patients with extensive burns it is not possible to use thicker autografts with limited expansion because of increased donor site morbidity. These results may improve if a dermal subsitute is placed underneath the split thickness skin graft. This substitute can replace the dermal tissue that is lost and provide a scaffold to the cells infiltrating in the wound bed during the healing process. The scaffold must be designed in such a way the cells will produce the new collagen fibers in a random organized structure, replacing slowly the scaffold material instead of making the typical parallel oriented fibers of scars (Van der Veen et al., 2010). In the past decades, several dermal substitutes products have become available but the clinical evidence on the effectiveness is limited until now (Brusselaers et al., 2010). These commercially available products are based on animal derived collagen or human skin and associated with high costs (Jones et al., 2002).
2. Development of dermal substitute
Dermal substitutes can be obtained from animal or human tissue as well as synthetic materials. The advantage of human skin is the presence of the natural collagen- elastin fibre network, providing the optimal three dimensional fibre structure for ingrowing fibroblast and blood vessels. Antigenic structures must be absent from the dermal substitute to avoid the induction of inflammatory processes that can worsen final scar quality. Human allogeneic skin has been applied to the wound as dermal substitute in the past as described by Cuono et al., 1987. In this method, the more immunogenic epidermis was removed several days after grafting the human donor skin and replaced by autogous cultured epithelial sheets. Technically, this method is difficult and in the allogeneic dermis cells of the donor remain present that can induce inflammation after wound closure. Therefore, several methods have been developed to remove the donor cells and hairs from the donor skin. This resulted in products known as Alloderm, Surederm, Graft jacket using enzymes and freeze drying techniques.
We have developed a cost-effective method to eliminate the antigenic structures by using low concentrations of sodium hydroxide (Richters et al. 2008). De-cellularization of glycerol 85% preserved skin starts by rinsing out the glycerol, thereafter the skin is incubated in 0.06M NaOH for 6 weeks. The NaOH solution is refreshed every week and finally neutralized. The acellular dermis is again preserved in glycerol 85%. This method avoids freezing which may damage the collagen and elastin fibers and, at the same time, it is very effective in washing out the donor cells and hairs. The prototypes, consisting of an intact, native collagen-elastin matrix obtained using the NaOH method were tested in a porcine model. Full thickness wounds of 4x4 cm were transplanted with the prototype and an autologous split skin graft meshed 1:3. Contraction of the wounds treated with this prototype was significantly lower compared to the control wounds, transplanted with only autologous skin (Richters et al., 2008, Pirayesh et al., 2007 and 2008). The six weeks treatment with NaOH showed the best results. Further experiments showed a two stage procedure was more optimal with the take rate of the autologous skin > 90% if the interval between application of the prototype and the autologous skin was 7 days. Final result with respect to contraction was comparable to Integra in this porcine model but the scar was more smooth (Pirayesh et al., 2011, manuscript in preparation). This encouraging results let to the first pilot of clinical experiments on burn patients.
3. Clinical application of Glyaderm
This novel dermal substitute is called Glyaderm, which is an abbreviation for Glycerol-preserved Acellular Dermis. Before application of Glyaderm, the wound bed must show viable granulation tissue. In most cases, woundbed preparation with allogeneic donor skin is needed; in burns this takes about 7 days after debridement. Glyaderm is preserved in
85% glycerol and must be rinsed in sterile 0.9% NaCl before use on the patient. The Glyaderm is meshed 1:1 to allow drainage of wound fluid. Glyaderm was used on the patients also in a two stage procedure in clinical studies, approved by the local ethics committee of the University of Ghent, Belgium. A pilot study and a comparative study were performed sofar.
4. Pilot study
In the first non-randomized pilot experiments 15 patients with full thickness burns (TBSA < 40%) were selected to study the optimal wound dressing, to avoid dehydration and bacterial contamination of the Glyaderm during the first days before the autologous skin was applied. Then a group of 10 patients with full thickness wounds was selected (5 with burns and 5 with reconstructive wounds) and was treated to define the optimal time between the two operations using Laser Doppler Imaging. The depth of the burn wounds was first assessed using laser Doppler imaging (Monstrey et al., 2007) to assure the wounds were deep dermal. The patients were recruited and treated with Glyaderm and split thickness skin graft after wound bed preparation with glycerol preserved allografts. Patient demographics were recorded and Laser Doppler imaging was used to monitor the vascularization of glyaderm. We also discovered that Laser Doppler Imaging allows us to measure wound tissue perfusion and monitor daily ingrowth of blood vessels into the dermal substitute. This allows us to know the exact timeframe in which the dermal bed is vascularised enough and skin grafting can be carried out. This is typically after 5-7 days, much shorter than the period necessary for ingrowth of the currently most widely used commercial dermal substitute Integra (Dantzer et al., 2001)
4.1. Randomised, controlled, paired, intra-individual Comparative study in 30 patients
After completing the pilot studies, a comparative study was performed. Patients with full thickness burns or full thickness lower arm defects after free flap harvesting were recruited. Part of the wounds were treated with Glyadem and split thickness skin and the other part with split thickness skin only. This was done in anatomically related areas to allow paired, intra-individual comparison of the wound healing. Preferably, a left/right comparison was made and the experimental treatment was randomized. Patient demographics were recorded and at regular follow up time points of 1, 3, 6 and 12 months after wound closure biopsies were taken and objective and subjective scar assessment was performed. The dermatospectrometer was used to measure scar colour, Dermalab cutometer for pliability and the modified Vancouver scar scale as well as a contour scale (Brusselaers et al 2010) were used. All data were recorded in a purpose designed database and subjected to statistical analysis with SPSS software package.
5. Results of the pilot study
All recruited patients responded well to a dressing regimen of surfasoft, a semipermeable membrane, with betadine gel and paraffin gauze in terms of bacterial control and prevention from desiccation of Glyaderm with capital. Laser Doppler Imaging proved to show enhanced vascularization from day 1 to day 5 allowing to exactly delineate the optimal engraftment interval. This was found to be 6 days with a 1 day standard deviation. The increase in blood vessel growth can be observed clearly in Figure 3, the areas with a red color represent blood vessel flow and these are larger on day 7 in this patient. The take of the Glyaderm was > 90% and the take of the autologous skin > 95%. The results with respect to scar formation in this pilot study were encouraging. Figure 4 shows pictures of the wound and final scar of the same patient as in Figure 3. The color of the scar is not red and the patient is able to move the head normally, indicating good elasticity.
6. Results of the comparative study
The take rates in the STSG on Glyaderm were comparable to the take rates in the wounds with a STSG alone (Wilcoxon Singed Ranks Test Z = -0.823, p =.41). Pliability and Visual Scores and contour scale assessment indicate a trend of improvement with Glyaderm treatments; however, follow-up times are currently too short with small numbers to be able to draw conclusions, we expect a necessary follow-up interval of two years. Comparing the Dermalab measurements for elasticity “Glyaderm + STSG” versus “STSG alone” statistics indicate that: Glyaderm + STSG” has significantly more elasticity when compared to “STSG alone” 1 month and 12 months after wound closure.
The results of the comparative, controlled, intra-individual clinical study show beneficial effects of a dermal substitute on the elasticity of the scar. The natural structure of the collagen and elastin fibers in glyaderm may have contributed to this effect. Glyaderm is the first non-commercial dermal substitute that can compete with all currently available dermal equivalents. Laser Doppler imaging allows monitoring of vascular ingrowth in dermal substitutes such as Glyaderm. Although most burn experts advocate the use of dermal substitutes, the challenge remains to objectively show the perceived benefit over split thickness skin grafting alone. The evolving evaluation with objective scar assessment tools within these studies may help to demonstrate this benefit in the near future.
Our next study with Glyaderm has started recently; a multi-centre clinical study in major European Burn centers. To allow also quality of life and cost-effectiveness studies, 60 patients will be treated with glyaderm + STSG and 60 patients with STSG alone.
Researchers continue their quest for the ideal skin substitute, and in the future it should be possible to create such an advanced skin substitute, containing melanocytes, hair follicles and sebaceous glands. The available products remain rather expensive, due to commercial incentives, high manufacturing costs, special shipment and storage conditions. Nevertheless, accelerated healing and closure of the wound will reduce the labour intensive dressing changes, hospital stay and the need for reconstructive surgery. Until the optimal off the shelve skin substitute becomes available, the burn surgeon can improve aesthetic and functional outcome by choosing from the gamut of currently available scaffolds for bilayered skin restoration. Glyaderm may well be an optimal cost-effective solution to bridge this gap in the near future.
Brusselaers N. Pirayesh A. Hoeksema H. Richters C. D. Verbelen J. Beele H. Blot S. I. Monstrey S. 2010J Trauma; 68 490 501
Brusselaers N. Pirayesh A. Hoeksema H. Verbelen J. Blot S. Monstrey S. 2010Burn scar assessment: a systematic review of objective scar assessment tools. Burns, 36 1157 1164
Cuono C. B. Langdon R. Birchall N. Barttelbart S. Mc Guire J. 1987Composite autologous-allogeneic skin replacement: development and clinical application. Plast Recon Surg, 80 626 637
Dantzer E. Braye F. M. 2001Reconstructive surgery using an artificial dermis (Integra): results with 39 grafts. Br J Plast Surg., 54 659 664
Hermans M. H. E. 1989Clinical experience with glycerol preserved donor skin treatment in partial thickness burns. Burns; 15 57 60
Jones I. Currie L. Martin P. 2002A guide to biological skin substitutes. Br J Plast Surg, 55 185 193
Kreis R. W. Vloemans A. F. P. M. Hoekstra M. J. Mackie D. P. Hermans R. P. 1989The use of non-viable glycerol-preserved cadaver skin combined with widely expanded autografts in the treatment of extensive third-degree burns. J Trauma; 29 51 55
Monstrey S. Hoeksema H. Verbelen J. Pirayesh A. Blondeel P. 2008Assessment of burn depth and burn wound healing potential. Burns; 34 761 769
Pirayesh A. Monstrey S. Blondeel P. Hoekstra H. Vanoorbeek J. Hoeksema H. Richters C. D. 2007Development of a novel dermal substitute based on glycerinized allograft: Clinical (Phase I) and experimental evaluation. Burns; 33:S13
Pirayesh A. Dur A. F. M. Paauw N. J. Monstrey S. Kreis R. W. Hoekstra M. J. Richters C. D. 2008Evaluation of acellular dermis for closure of abdominal wall defects in a rat model. Eur Surg res; 41 345 352
Pirayesh A. Hoeksema H. Richters C. D. Monstrey S. 2011Determination of the optimal time interval between application of a dermal substitute and grafting with autologous skin using Laser Doppler Imaging. Manuscript in preparation.
Richters C. D. Pirayesh A. Hoeksema H. Kamperdijk E. W. A. Kreis R. W. Dutrieux R. P. Monstrey S. Hoekstra M. J. 2008Development of a dermal matrix from glycerol preserved allogeneic skin. Cell and Tissue Banking; 9 309 315
Midelkoop E. Van der Veen V. Van der Wal M. B. Van Leeuwen M. C. MM Ulrich 2010Biological background of dermal substitutes. Burns; 36 305 321