Permanent Skin Replacement Research Articles

Report of outcomes in burn patients enrolled in the Cultured epidermal autograft prospective Registry

Cultured epidermal autograft (CEA) is a permanent skin replacement indicated for use in adult and pediatric patients with deep dermal or full thickness burns comprising a total body surface area (TBSA) ≥ 30 %. CEA (Epicel®) was approved for use in adults in the United States in 2007 as a Humanitarian Use Device (HUD) under a Humanitarian Device Exemption (HDE) and was approved for pediatric use in 2016. In 2019, a CEA Registry was established with an objective of prospective data collection and analysis of demographic, treatment, and outcome data for the real-world use of CEA. At the time of data cut–off (June 2022), 68 patients (50 adults and 18children) had completed data in the registry, up through hospital discharge, for this analysis. Mean total body surface area (TBSA) of the burn was 58 % in adults and 56 % in pediatric patients, and almost half had inhalation injury in each group. Approximately 74 % of adults and 67 % of pediatric patients had TBSA ≥ 50 %. Overall survival was 87 % (84 % adults, 94 % pediatric). Mean % graft take (engraftment) of CEA was 81 % in adults and 84 % in pediatric patients. Outcomes from this prospective collection of data in severely burned patients treated with CEA demonstrate favorable engraftment and survival rates and are in general agreement with recent literature.

770 Outcome Results of the Prospective CEA Registry

Abstract Introduction Cultured epidermal autograft (CEA) is a permanent skin replacement indicated for use in adult and pediatric patients with deep dermal or full thickness burns comprising a total body surface area (TBSA) ≥ 30%. CEA may be used in conjunction with split-thickness autografts or alone, in patients for whom split-thickness autografts may not be an option due to the severity and extent of their burns. CEA was approved for use in adults in the United States in 2007 as a Humanitarian Use Device (HUD) under a Humanitarian Device Exemption (HDE) and was approved for pediatric use in 2016. Methods The objective of the CEA Registry is to allow prospective collection and analysis of demographic, treatment, and outcome data for the real-world use of CEA. Prior to initiation of the Registry, a central Institutional Review Board reviewed and approved written procedures and informed consent from each participating burn center. As of the date of this analysis, 5 centers were enrolling patients in the Registry. The data are collected and stored in a 21 Code of Federal Regulations (CFR) Part 11 and Health Insurance Portability and Accountability Act (HIPAA) compliant environment. First patient enrollment in the Registry was 13 June 2019 and data cutoff for this analysis was 28 July 2022. Results Sixtyeight patients (50 adults and 18 children) had completed data in the registry, up through hospital discharge, for this analysis. Except for age and revised Baux score, demographics and burn characteristics were similar between the adult and pediatric populations. The majority of patients were male in both the adult (76%) and pediatric (67%) groups, with burn caused by flame the most common etiology (86% and 78%, respectively). The total body surface area (TBSA) of burn was 58% in adults and 56% in pediatric patients (range 30% to 96%), and almost half had inhalation injury in each group. Approximately 74% of adults and 67% of pediatric patients had TBSA ≥50%. Median age in the adult group was 42 years, and in the pediatric group was 15 years. Since age is a component of the calculation of Baux score, pediatric patients had lower mean revised Baux scores (75) compared to adults (107.5). Overall, 59 of the 68 patients (87%) survived until hospital discharge. The mean % graft take (engraftment) in both adult (81%) and pediatric (84%) groups is notable when compared to mean final graft take in 453 Epiceltreated patients reported in the HDE database (67% in adults and 70% in pediatric). By age group, adults had 84% survival rate and pediatric patients had 94% survival. Conclusions Outcomes from this prospective collection of data in severely burned patients treated with CEA demonstrate favorable engraftment and survival rates and are in general agreement with recent literature. Applicability of Research to Practice Reported outcomes from the CEA Registry can be used as a benchmark by centers using CEA for treatment of burns.

Skin Immunity and Tolerance: Focus on Epidermal Keratinocytes Expressing HLA-G.

Although the role of epidermal cells in skin regeneration has been extensively documented, their functions in immunity and tolerance mechanisms are largely underestimated. The aim of the present review was to outline the state of knowledge on resident immune cells of hematopoietic origin hosted in the epidermis, and then to focus on the involvement of keratinocytes in the complex skin immune networks acting in homeostasis and regeneration conditions. Based on this knowledge, the mechanisms of immune tolerance are reviewed. In particular, strategies based on immunosuppression mediated by HLA-G are highlighted, as recent advances in this field open up perspectives in epidermis-substitute bioengineering for temporary and permanent skin replacement strategies.

3D Printing of Strontium Silicate Microcylinder-Containing Multicellular Biomaterial Inks for Vascularized Skin Regeneration.

The reconstruction of dermal blood vessels is essential for skin regeneration process. However, the lack of vascular structure, insufficient angiogenesis induction, and ineffective graft-host anastomosis of the existing skin substitutes are major bottle-necks for permanent skin replacement in tissue engineering. In this study, the uniform strontium silicate (SS) microcylinders are successfully synthesized and integrated into the biomaterial ink to serve as stable cell-induced factors for angiogenesis, and then a functional skin substitute based on a vascularization-induced biomimetic multicellular system is prepared via a "cell-writing" bioprinting technology. With an unprecedented combination of vascularized skin-mimicking structure and vascularization-induced function, the SS-containing multicellular system exhibits outstanding angiogenic activity both in vitro and in vivo. As a result, the bioprinted skin substitutes significantly accelerate the healing of both acute and chronic wounds by promoting the graft-host integration and vascularized skin regeneration in three animal models. Therefore, the study provides a referable strategy to fabricate biomimetic multicellular constructs with angiogenesis-induced function for regeneration of vascularized complex and hierarchical tissues.

Therapeutic Strategies for Full‐Thickness Wounds: Skin and Corneal Tissue‐Engineered Substitutes with Stem Cells

The LOEX centre of Laval University is specialized in stem cells and tissue engineering. In addition to fundamental research, LOEX focuses on translational medicine to transfer the discoveries into better treatment for the patients. The LOEX self‐assembly approach of tissue engineering was designed to elaborate complex human living tissue substitutes. Autologous self‐assembled skin substitutes (SASSs) are produced to treat wounds of severely burned patients. The procedure is based on primary culturing dermal fibroblasts and keratinocytes from a few cm2 skin biopsy taken from an unaffected site. Then, dermal fibroblasts are cultured in the presence of ascorbic acid. They secrete and organize their own extracellular matrix (ECM) on which keratinocytes rebuild the epithelium. SASSs exhibit a well‐developed stratified cornified epithelium. The basement membrane is structured and favors long‐term preservation of stem cells in the basal layer of the tissue‐engineered epidermis. The dense collagen network of the tissue‐engineered dermis provides resistance and allow SASS manipulation. After transplantation of autologous SASSs on burn wounds, the graft take was excellent (90–100%). No significant contraction was observed in vivo after grafting. The presence of the ECM promoted a good healing and skin suppleness. SASS presented a well‐organized epidermis and stem cells were settled and maintained in the basal layer after grafting. A complete basement membrane with numerous hemidesmosomes was observed before and after grafting, supporting a strong cohesion between the dermis and epidermis. The integrity of the transplanted SASS persisted over time (1‐ to 8‐year follow‐up) with no defect in epidermal regeneration and no significant contracture. Minimal hypertrophic scars were only observed between SASS. Thus, SASSs provide permanent skin replacement for severe burns. These living substitutes comprising cells and extracellular matrix possess the capability of expanding with the child’s growth. We conclude that the TES produced by this approach is a promising skin substitute for resurfacing full‐thickness skin injury given its functional characteristics: minimal contraction after grafting and promotion of long‐term tissue regeneration. LOEX also used cultured epithelial corneal autografts (CECA), produced from a small biopsy of the limbal area containing stem cells, for the treatment of limbal stem cell deficiency. The first clinical trial is completed, 14 of the 15 patients treated improved their visual conditions. These regenerative medicine technologies pave the way for better treatment of rare diseases.Support or Funding InformationSupported by the Canadian Institutes for Health Research (CIHR), The Stem Cell Network, Fonds de Recherche du Québec en Santé (FRQS), Fondation des Pompiers du Québec pour les Grands Brûlés and the Cell and Tissue Therapy Network of the FRQS. LG is holder of the Canada Research Chair on Stem Cells and Tissue Engineering of CIHR.

On skin substitutes for wound healing: Current products, limitations, and future perspectives

Cutaneous wound healing is a highly coordinated process involving numerous molecular pathways that regulate the function of specific skin cell types. One of the key decisions in acute and/or chronic wound healing management is to choose the skin substitute that, based on its composition and/or properties, may act as permanent skin replacement or temporary wound cover.The current products however are limited in their action, especially in the context of large chronic wounds or extensive second-/third-degree burns, due to the risk of developing infection, the limited vascularization, the inability to integrate to host tissue, and the lack in the healed area of skin apparatus responsible for temperature control, pigmentation, immune regulation, and nerve supply. In addition, the high cost of skin substitutes precludes their use in small- and moderate-size burns because they are economically impractical and protract the time to definitive wound closure. Thus, the need to develop cost-effective substitutes of high quality is imperative. The purpose of this review is to discuss current available products, their limitations and to provide some perspectives on future research toward generation of cost-effective, high-quality substitutes.

338 - First-in-human application of the self-assembled skin substitute as permanent graft to treat burn patients: A case series of 14 patients indicating clinical effectiveness

There is a need for diseased or fatally wounded tissue replacement. The aim of this study was to assess whether autologous engineered tissues produced by the self-assembly approach of tissue engineering could provide a permanent tissue replacement. The clinical outcome of autologous self-assembled skin substitutes (SASSs) as permanent coverage of full-thickness burn wounds was monitored. SASSs were used to treat patients suffering from severe burn wounds over more than 50% of their total body surface area. The SASSs were grafted after surgical debridement of full-thickness wounds in fourteen severely burned patients. The mean percentage of SASS graft take was 98% (SD 5). The integrity of the SASSs persisted over time (average follow-up time: 3.2 years) without noticeable deficiency in epidermal regeneration. Assessment of scar quality (skin elasticity, erythema, thickness) was performed on a subset of patients. Minimal scarring was observed. However, non-homogeneous pigmentation was noticed in several patients. We conclude that SASSs allow permanent skin replacement. This first-in-human application of a self-assembled skin substitute could lead to the treatment of various epithelial/stroma defects. SASSs could be combined with gene therapy to develop a treatment for genetic cutaneous diseases. Moreover, the self-assembly approach of tissue engineering can be used to produce other tissues or even organs and could eventually lead to new therapies in the cardiovascular or ophthalmology domains.

Adult Burn Patients With More Than 60% TBSA Involved–Meek and Other Techniques to Overcome Restricted Skin Harvest Availability–The Viennese Concept

Despite the fact that early excision and grafting has significantly improved outcome over the last decades, the management of severely burned adult patients with >/=60% total body surface area (% TBSA) burned still represents a challenging task for burn care specialists all over the world. In this article, we present our current treatment concept for this entity of severely burned patients and analyze its effect in a comparative cohort study. Surgical strategy comprised the use of split-thickness skin grafts (Meek, mesh) for permanent coverage, fluidized microsphere bead-beds for wound conditioning, temporary coverage (polyurethane sheets, Epigard; nanocrystalline silver dressings, Acticoat; synthetic copolymer sheets based on lactic acid, Suprathel; acellular bovine derived collagen matrices, Matriderm; allogeneic cultured keratinocyte sheets; and allogeneic split-thickness skin grafts), and negative-pressure wound therapy (vacuum-assisted closure). The autologous split-thickness skin graft expansion using the Meek technique for full-thickness burns and the delayed approach for treating dorsal burn wounds is discussed in detail. To demonstrate differences before and after the introduction of the Meek technique, we have compared patients of 2007 with >/=60% TBSA (n = 10) to those in a matched observation period (n = 7). In the first part of the comparative analysis, all patients of the two samples were analyzed with regard to age, abbreviated burn severity index, Baux, different entities of % TBSA, and survival. In the second step, only the survivors of both years were separated in two groups as follows: patients receiving skin grafts, using the Meek technique (n = 6), were compared with those without Meek grafting (n = 4). When comparing the severely burned patients of 2007 with a cohort of 2006, there were no differences for age (2007: 46.4 +/- 13.4 vs. 2006: 39.1 +/- 14.8 years), abbreviated burn severity index score (2007: 12.2 +/- 1.0 vs. 2006: 12.1 +/- 1.2) or % TBSA (2007: 72.1 +/- 11.7 vs. 2006: 69.3 +/- 8.7% TBSA). In these two rather small groups of severely burned patients with >/=60% TBSA, the overall survival rate of patients was 70.0% (7/10) in 2007 and 42.9% (3/7) in 2006, respectively. Almost all nonsurvivors in both years died within the first 5 days after admission. If assessing the different treatment modalities of the survivors, we found that although the Meek group patients were older (Meek 48.8 +/- 13.3 vs. non-Meek 26.8 +/- 11.5 years, P = .0381) and had consequently higher Baux scores (Meek 124.0 +/- 2.9 vs. non-Meek 93.8 +/- 8.5, P = .0095) than the non-Meek patients, this seemed to have no effect on length-of-stay (80.5 +/- 9.7 vs. non-Meek 79.8 +/- 33.0 days), hospital length-of-stay (85.7 +/- 14.8 vs. non-meek 84.3 +/- 26.1 days) or number of operations (6.5 +/- 1.0 vs. non-Meek 7.0 +/- 4.1 operations). The achieved results represent a combination of various treatment changes and, therefore, cannot be attributed to a single modality. The Meek technique is one of the technical options to choose from, to achieve permanent skin replacement; we think that it has its place if integrated in a whole treatment concept for management of severely burned patients.

High Yields of Autologous Living Dermal Equivalents Using Porcine Gelatin Microbeads as Microcarriers for Autologous Fibroblasts

Permanent skin replacement requires a dermal component to ensure adequate long-term graft stability and to prevent wound contraction. This study was to construct a bioreactor microcarrier cell culture system (Bio-MCCS) to produce autologous living dermal equivalents on a large scale. Autologous fibroblasts were isolated from split-thickness skin biopsy from a leg ulcer patient, inoculated onto macroporous porcine gelatin microbeads, and incubated in a bioreactor (Cellspin) in serum-free fibroblast growth medium or in DMEM medium containing 10% fetal calf serum (FCS). Fibroblasts rapidly adhered to and actively proliferated on the microbeads in the bioreactor in both serum-free and serum-containing medium. MTT assay showed the number of fibroblasts on the microbeads reached up to 5.3- or 4.0-fold the cells seeded in DMEM medium containing 10% FCS or serum-free medium, respectively. When removed from Bio-MCCS and cultured under static conditions, fibroblasts were able to leave the microbeads and proliferate to confluence on the bottom of tissue culture flasks. When stored at room temperature in DMEM containing 10% FBS, fibroblast cultured on the microbeads retained highest viabilities for at least 3 weeks, up to 82% of originals. This Bio-MCCS using porcine gelatin microbeads as carriers for fibroblasts offers a new option of mass production of autologous living dermal equivalents.

Allogenic skin in the treatment of burns

Allogenic skin has had a major role in acute burns care for over 100 years. The principle source of allogenic skin is from cadavers. Allogenic skin provides the gold standard for temporary skin substitutes. The main drawbacks to its wider use are availability and disease transmission. The major obstacle to prolonged use is its immunogenicity. As more effective means are developed to ensure the supply and safety of allogenic skin and novel ways of circumventing the immunologic problems are developed, it is possible that allogenic skin may find a new role as a permanent skin replacement in future burns care.

Initial experience with a composite autologous skin substitute

Patients with large burns are surviving in increasing numbers, but there remains no durable and reliable permanent skin replacement. After initial favorable small animal experiments, a pilot trial of a composite skin replacement was performed in patients with massive burns. A composite skin replacement (CSR) was developed by culturing autologous keratinocytes on acellular allogenic dermis. This material was engrafted in patients with massive burns and compared to a matched wound covered with split thickness autograft. With human studies committee approval, 12 wounds in 7 patients were grafted with CSR while a matched control wound was covered with split thickness autograft. These 7 children had an average age of 6.4±1.4 yr and burn size of 75.9±5.0% of the body surface. Nine wounds were acute burns and three were reconstructive releases. Successful vascularization at 14 days averaged 45.7±14.2% (range 0–100%) in the study wounds and 98±1% (range 90–100%) in the control sites ( P<0.05). Reduced CSR take seemed to correlate with wound colonization. All children survived. While CSR did not engraft with the reliability of standard autograft, this pilot experience is encouraging in that successful wound closure with this material is possible, if not yet dependable. It is hoped that a more mature epidermal layer may facilitate engraftment, and trials to explore this possibility are in progress.

Artificial skin.

The skin is a complex organ that is difficult to replace when it is irreversibly damaged by burns, trauma, or disease. Although autologous skin transplantation remains the most common form of treatment in patients with significant skin loss, there are now a number of commercially available products that can be used to replace the skin temporarily or permanently. Here we describe several such products under the rubric "artificial skin," focusing on two types of technology that have been applied to the problem of permanent skin replacement.

Dendritic ice morphology in unidirectionally solidified collagen suspensions

Collagen is a fibrillar protein which is commonly used as a biodegradable biomaterial. A typical application of such a biomaterial is a freeze-dried collagen sponge which is primarily used as a permanent skin replacement for the treatment of deep dermal burns. Especially to diminish scar formation caused by a severe loss of skin, it is necessary to induce a cell migration into the sponges and the regeneration of endogenous tissue within the sponges. Thereby, the three-dimensional structure and the pore sizes of the collagen sponges strongly influence the wound healing. In order to study this influence, the development of a process to manufacture collagen sponges with an adjustable and homogeneous three-dimensional structure is necessary. The porous structure of freeze-dried sponges corresponds to the ice crystal morphology after freezing. In order to form an adjustable and homogeneous ice morphology, the unidirectional solidification of collagen suspensions was investigated. All experiments were performed in a cryomicroscope according to the Bridgman technique. To induce a constitutional supercooling leading to the breakdown of the planar ice front and the formation of a stable dendritic ice crystal morphology (without side branches), the addition of solutes to the basic collagen suspension is required. We used acetic acid and ethanol as soluble additives, because they are suitable for medical application. The effects of the temperature gradient, the ice front velocity, and the solute concentration on the primary spacing and the dendrite sizes were examined. In order to find a simplified experimental system, acetic acid solutions without collagen were solidified under the same freezing conditions. Although the primary spacings and the dendrite sizes almost varied in the same range for solutions and suspensions, they were influenced in a different manner by the freezing parameters and the solute concentrations. Varying the aforementioned parameters the primary spacing could be adjusted in the range from 40 to 60 μm. Furthermore, the ice crystal sizes, which correspond to the pore size after freeze-drying, could be varied between 30 and 50 μm.

Cultured epithelial autografts: diving from surgery into matrix biology.

Cultured epithelial autografts offer an exciting approach to cover extensive skin wounds. The main problem of this method is mechanical instability during the first weeks after grafting. There is evidence that the shortcomings of autografting cultured keratinocytes result from the lack of a mature and functional dermo-epidermal junction. This article summarizes the current knowledge regarding the de novo formation of the dermo-epidermal junction and the dynamics of "take" and stabilization of cultured epithelial autografts. Future strategies are discussed of how to improve and accelerate the process conferring definitive stabilization of cultured epithelial autografts including the potential therapeutic use of transglutaminase as well as cocultivation of a dermo-epidermal equivalent in order to facilitate a permanent skin replacement.

Permanent skin replacement using chimeric epithelial cultured sheets comprising xenogeneic and syngeneic keratinocytes.

The present study was undertaken to evaluate the possibility of permanent skin replacement using chimeric xenogeneic-syngeneic graftable sheets previously obtained in vitro. Newborn (<3 days old) BALB/c and human keratinocytes were isolated and cocultured in different ratios as follows: 50% BALB/c to 50% human and 25% BALB/c to 75% human keratinocytes. Four to 5 days after culture and prior to their grafting, all chimeric sheets contained both cell types in ratios similar to those used to seed the initial chimeric cultures. Fourteen and 30 days after chimeric sheet grafting onto BALB/c mice dorsum, the newly generated cutaneous tissue showed a histologically well-organized epidermis presenting basal and suprabasal cell layers. Cutaneous cells in these structures secreted laminin and type IV collagen in blood vessels, and at ground level of the dermoepidermal junction there were signs of physiologically active skin. Cell phenotyping revealed the presence of only syngeneic keratinocytes, whereas xenogeneic cells were passively eliminated without a total rejection of the chimeric implant. This selective and passive elimination of xenogeneic keratinocytes went through cellular and humoral immunity activation. Data suggest that this chimeric culture method can be used for cutaneous therapies such as large congenital nevi, skin ulcers, and extensively burned skin. Indeed, for large third-degree wounded skin treatment, this culture method may shorten the time (4-5 weeks) needed for cell growth and graftable sheet production. Moreover, since the ultimate aim in allogeneic and xenogeneic transplantation is to achieve an immunological acceptance and tolerance to these foreign tissues, the chimeric culture approach may provide ways to lighten tolerance phenomena on cutaneous tissue.

Permanent grafting of living skin substitutes: surgical parameters to control for successful results.

Autologous mesh grafting, widely used in the treatment of severe burns, remains the most conventional approach for permanent skin replacement. However, during the last decade several types of skin substitutes were reported as suitable alternatives for full-thickness burn wound coverage. The clinical use of such dressings requires new surgical skills to maintain the integrity of the grafts and favor their permanent implantation in vivo. This article reports observations made on nude mice grafted with cultured human skin equivalents. Some parameters such as the quality of adhesion between the implant and the graft bed, the size, the stability and the thickness of the graft, the humidity of the chamber, and the protocol of antibiotic administration were identified as crucial for the success of the surgery. The grafting procedures are described in this paper. These results should be taken into consideration in all transplantations of skin grafts in vivo.

Cultured epidermal autografts and allodermis combination for permanent burn wound coverage

Cultured epidermal autografts (CEA) have been shown to be an effective permanent skin replacement for major burn injuries, but are more sensitive to adverse conditions than split thickness grafts (Clarke et al., 1988). Cuono et al. (1986, 1987) have described the successful use of engrafted allodermis as a wound bed for cultured grafts. We report on a method of preparing allodermis and grafting CEA in five patients with major burns (48–70 per cent TBSA, average 59.6 per cent). The average age was 38.8 years (20–60 years). All full thickness wounds were excised down to fat within 7 days of admission, and covered with meshed split thickness cryopreserved homograft. Over the ensuing 2–3 weeks, the homograft became engrafted. At surgery, the allo-epidermis was removed, leaving the dermal components as a viable bed for the CEA. Keratinocytes derived from a full thickness biopsy were. grown to confluence by the method of Rheinwald and Green (1975), and 25 cm 2 sheets were stapled to Vaseline gauze backings and applied to freshly excised wounds. Seven to 10 days after surgery, the gauze backings were removed. The average take ranged from 87–100 per cent (average 93.6 per cent). Follow-up for up to 4 years shows supple skin that has been durable, and resistant to trauma and infection.

Burn wound closure using permanent skin replacement materials.

Over the past decade, very significant advances in the development of clinically useful, permanent skin replacement materials have taken place. The most prominent and successful approaches to the physiological closure of an open wound have been either by creating a totally artificial dermal matrix material, by using culture techniques to expand cell populations for autologous transplantation, or by using a combination of these methods. As a result of substantial early progress in this field, permanent skin replacement materials as a treatment modality promise significant contributions to improved wound management and increased survival rates for patients with devastating soft tissue destruction such as massive burn injuries.

Epidermis generated in vitro: practical considerations and applications

The technology for culture of epidermis is one of the most advanced to date for generation of a tissue in vitro. Cultured epidermis is already used for a number of applications ranging from use as a permanent skin replacement to use as an organotypic culture model for toxicity testing and basic research. While simple epidermal sheets have been grafted successfully, more advanced models for skin replacement consisting of both dermal and epidermal components are in development and being tested in a number of laboratories. One of the most advanced in vitro models is the living skin equivalent, an organotypic model consisting of a collagen lattice contracted and nourished by dermal fibroblasts overlaid with a fully formed epidermis.

Skin Substitutes in Upper Extremity Burns

Four techniques for permanent skin replacement with skin substitutes are described. A claim of superiority to conventional skin grafting on upper extremity and hand burns is not made, but some clinical observations and histologic evidence of different healing characteristics are shown. The composite grafts described appear to effectively replace the bilayered structure of skin and seem to have good subjective resistance to shear forces. If the skin replacement is durable and heals with less scarring than conventional skin grafts, the inexorable course to stiffness and contracture may be altered. Further basic science and clinical investigation may provide us with a better way of managing these difficult problems.