Human Skin Allografts Research Articles

Bacterial contamination of human skin allografts and antimicrobial resistance: a skin bank problem

BackgroundBacterial contamination remains the major problem in skin banks, even after antimicrobial treatment, and results in high rates of tissue discarding. This study aimed to analyze bacterial contamination in 32 human skin allografts from the skin bank of Dr. Roberto Corrêa Chem from the Hospital Complex Santa Casa de Misericórdia de Porto Alegre. These samples were already discarded due to microbial contamination. The identification of the bacteria isolated from skin allografts was performed by matrix assisted laser desorption ionization–time of flight. The antimicrobial susceptibility of the isolates to six different classes of antimicrobials was determined using the disk-diffusion agar method, and the evaluation of the inhibitory potential was determined by the minimal inhibitory concentration (50/90) of antimicrobials already used in the skin bank and those that most isolates were susceptible to.ResultsA total of 21 (65.6%) skin samples were contaminated with Gram-positive bacteria: 1 (4.7%) with Paenibacillus sp., 12 (61.9%) with Bacillus sp., 6 (28.5%) with Staphylococcus sp., and 2 (9.5%) with Bacillus sp. and Staphylococcus sp. Several resistance profiles, including multiresistance, were found among the isolates. Most of the isolates were susceptible to at least one of the antimicrobials used in the skin bank. All isolates were susceptible to amikacin, gentamicin, and tetracycline, which demonstrated the best inhibitory activities against the isolates and were considered as potential candidates for new antimicrobial treatments.ConclusionsBacillus, Paenibacillus, and Staphylococcus were isolated from the skin allografts, thus demonstrating the predominance of Gram-positive bacteria contamination. Other factors not related to the resistance phenotype may also be involved in the persistence of bacterial isolates in the skin allografts after antibiotic treatment. Gentamicin, amikacin, and tetracycline can be considered as an option for a more effective treatment cocktail.

Regulation of human T cell responses by dNP2-ctCTLA-4 inhibits human skin and microvessel graft rejection

Manipulation of human T cell functioning by delivery of macromolecules such as DNA, RNA, or protein is limited, unless the human T cells have been stimulated or electropermeabilized. To achieve successful adaptation and survival of a grafted organ, the alloreactive T cells that induce graft rejection must be regulated. Corticosteroids, calcineurin inhibitors, and mTOR inhibitors, which are systemic immunosuppressants, are currently used for transplantation, with significant side effects. In this study, we demonstrated that a cell-permeable peptide (CPP), dNP2, could efficiently deliver proteins into human CD4 and CD8 T cells. We confirmed regulatory functioning of the cytoplasmic domain of CTLA-4 conjugated with dNP2 (dNP2-ctCTLA-4) in human T cell activation, proliferation, and chemokine receptor expression. We utilized a human skin allograft system in SCID/beige mice to examine whether dNP2-ctCTLA-4 could inhibit allograft rejection by controlling T cell responses. The grafted skin tissue inflammation, allogeneic T cell infiltration, and blood cytokine level was markedly reduced by dNP2-ctCTLA-4, resulting in successful transplantation. In addition, it also inhibited T cell alloresponses against microvessels formed form Bcl-2-transduced human umbilical vein endothelial cells implanted into Balb/c Rag1−/−/IL-2Rγ−/- double knockout (DKO) mice, assessed as reduced T cell infiltration and granzyme B expression. These results collectively suggest that dNP2 peptide conjugation offers a valuable tool for delivering macromolecules like proteins into human T cells, and dNP2-ctCTLA-4 is a novel agent that shows potential in controlling human T cell responses to allow successful adaptation of grafted tissues.

Skin banking at a regional burns centre—The way forward

In India approximately 1 million people get burnt every year and most of them are from the lower or middle income strata. Therefore it is obligatory to find out an economic way of treatment for the affected populace. Since use of human skin allograft is the gold standard for the treatment of burn wound, in-house skin banking for a burn unit hospital is prerequisite to make the treatment procedure affordable. Although, there was one skin bank at India till 2009, but it was difficult for a single bank to cover the entire country’s need. Looking at the necessities, National Burns Centre (a tertiary burn care centre) along with Rotary International and Euro Skin Bank collaborated and developed an effective cadaveric skin banking model in Mumbai, Maharashtra in 2009. Initial two to three years were formation phase; by the year 2013 the entire system was organized and started running full fledged. The model has also been replicated in other states of India to accommodate the large burn population of the country. This paper therefore, gives a step by step account of how the bank evolved and its present status.

93 The Impact of Skin Allograft on Inpatient Outcomes for Major Burns 20–50%: a Propensity Score Matched Analysis Using NIS

Human cadaveric skin (allograft) is used in treating major burns both as temporizing wound coverage and a means of testing wound bed viability following burn excision. There is limited information on outcomes, and clinicians disagree on indications for application in intermediate-sized burns. This study aims to improve understanding of allograft use in 20–50% total body surface burns by assessing current utilization and evaluating inpatient outcomes. Discharge data from the Nationwide Inpatient Sample (NIS), Healthcare Cost and Utilization Project (HCUP), Agency for Healthcare Research and Quality assessed 3,557 major burn patients (>20% total body surface area and ≥ second degree depth, 20–50% TBSA) undergoing operative treatment. Patient demographics and hospital characteristics were evaluated with multivariate logistic regression. Outcomes were evaluated with propensity score matching. The primary outcome was mortality with secondary outcomes including complications, length of stay, total burn operations, and charges. After matching, 771 allografted patients were paired with 2,786 controls. Covariate mean standard differences were all <10% after matching. The average treatment effect (ATE) of allograft on inpatient mortality was an increase of 3.2% (95% CI 0.9–5.5%, p=0.006). Allograft ATEs were all significantly higher for secondary outcomes: composite complication index increased 0.11 (95% CI 0.04–0.17, p<0.001), length of stay increased 9.5 days (95% CI 7.4–11.6 days, p<0.001), total burn operations increased by 1.6 (95% CI 1.4–1.8, p<0.001), and total charges increased $148,578 (95% CI $111,166–185,991, p<0.001). After adjusting for covariates, allograft use significantly increased inpatient mortality, and was associated with more complications, longer length of stay, more burn operations, and greater total charges. This analysis suggests that better studies are needed to justify the use of this costly and limited resource in intermediate-sized major burns. Inform providers on judicious use of allograft and negative repercussions in TBSA burns 20–50%.

Selective blockade of CD28 on human T cells facilitates regulation of alloimmune responses.

T cells are central to the detrimental alloresponses that develop in autoimmunity and transplantation, with CD28 costimulatory signals being key to T cell activation and proliferation. CTLA4-Ig molecules that bind CD80/86 and inhibit CD28 costimulation offer an alternative immunosuppressive treatment, free from some of the chronic toxicities associated with calcineurin inhibition. However, CD80/86 blockade by CTLA4-Ig also results in the loss of coinhibitory CTLA4 signals that are critical to the regulation of T cell activation. Here, we show that a nonactivating monovalent anti-CD28 that spares CTLA4 signaling is an effective immunosuppressant in a clinically relevant humanized mouse transplant model. We demonstrate that selective CD28 blockade prolongs human skin allograft survival through a mechanism that includes a reduction in the cellular graft infiltrate. Critically, selective CD28 blockade promotes Treg function in vivo and synergizes with adoptive Treg therapy to promote transplant survival. In contrast to CTLA4-Ig treatment, selective CD28 blockade promotes regulation of alloimmune responses and facilitates Treg-based cellular therapy.

Non-uraemic calciphylaxis: a diagnostic and management challenge for the burns team

Calciphylaxis is a rare systemic condition usually seen in patients with end-stage renal disease. It is characterised by pathological calcification of dermal blood vessels, causing ischaemia and necrosis in the skin and subcutaneous fat. The lesions are usually small but, in extreme cases, may be very extensive and may mimic necrotising fasciitis, prompting extensive surgical debridement. We describe the challenges faced in managing such a case. A 38-year-old female patient was admitted to another hospital with widespread progressive truncal skin necrosis. A provisional diagnosis of necrotising fasciitis was made, and she underwent debridement of 32% of her total body surface area (TBSA). However, her condition rapidly deteriorated and she was transferred to our burn centre for further management. Diagnosis of calciphylaxis was based on clinical features and confirmed by tissue biopsy. Characteristic findings of progressive skin and fat necrosis at the wound margins with healthy underlying fascia and muscle were recognised from previously treated cases of calciphylaxis; however, the absence of renal disease proved misleading. Non-uraemic calciphylaxis is very rare but can, as in this case, be associated with alcoholic liver disease. At our centre, the patient required prolonged ventilation and supportive treatment in burns critical care, combined with intensive wound management, repeated episodes of debridement, temporary wound cover using human skin allografts and reconstruction using split-thickness autograft. We report this case to alert others to this rare condition as a possible diagnosis in patients presenting with extensive panniculitis. We discuss associations, diagnostic difficulties, novel therapies and the importance of multidisciplinary care in managing these complex cases. Level of evidence: Level V, diagnostic study.

Transient Anti-CD45RC mAb Treatment Induces Specific Transplant Tolerance Through Potentiation of Tregs

Introduction: The main goal in transplantation is to induce specific rather than general immunosuppression. The CD45 protein is an essential regulator of T and B cell antigen receptor signaling. We have previously shown that T cells expressing low levels of CD45RC have strong immunoregulatory properties while T cells expressing high levels of CD45RC are essentially effector cells. Thus, we investigated the effect of depleting CD45RC+ cells on graft survival. Methods: Tregs were analyzed for the expression of CD45 isoforms by flow cytometry. LEW-1A rats were grafted in a heterotopic position with a LEW-1 W heart. NSG mice were 2Gy-irradiated or grafted with human skin and injected with 5x106 or 1,5x107 human PBMCs respectively. Animals were treated with 0.8 mg/kg/2.5d for 20d of anti-CD45RC mAb (OX22 or MT2 clones) or isotype. Humoral responses were assessed after KLH + CFA or exogenous red blood cells immunization. FACS Aria-sorted Tregs were i.v. transferred in 4,5Gy-irradiated recipients. CD45RClow/- Tregs were analyzed by RNA sequencing. Results: First, we have shown that CD45RC is the only CD45-isoform not expressed by Foxp3+ Tregs both in human and rat. Interestingly, transient treatment with anti-CD45RC Mabs induced transplant tolerance in a fully mismatch cardiac allograft model in rat. Importantly, this treatment prevented specifically anti-donor humoral responses while preserved primary and memory responses against cognate antigens. As expected, anti-CD45RC Mabs induced rapid cell death of CD45RC+ T cells through intrinsic cell signaling while preserved CD45RClow/- Tregs. Furthermore, the short-term treatment induced a transient homeostatic proliferation and sustained transcriptional changes in long-term CD8+ and CD4+ CD45RClow/- Tregs that maintained an indefinite, dominant and donor-specific tolerance, as adoptive transfer of Tregs succeeded in this allograft combination only. Finally, we translated these findings to human, in humanized mice models of xenogeneic GVHD and human skin allograft rejection. We demonstrated that ex vivo depletion of CD45RC+ PBMCs prevented development of GVHD, and that transient treatment with anti-human CD45RC Mabs delayed GVHD development and allograft rejection. Conclusion: This is the first proof of concept that targeting CD45RC with short-term Mabs treatment is an efficient innovative strategy to induce a donor-specific tolerance in transplantation without any other treatment.

An update and review of cell-based wound dressings and their integration into clinical practice

Chronic wounds affect over 4 million individuals and pose a significant burden to the US healthcare system. Diabetes, venous stasis, radiation or paralysis are common risk factors for chronic wounds. Unfortunately, the current standard of care (SOC) has a high relapse rate and these wounds continue to adversely affect patients' quality of life. Fortunately, advances in tissue engineering have allowed for the development of cell-based wound dressings that promote wound healing by improving cell migration and differentiation. As the available options continue to increase in quantity and quality, physicians should have a user-friendly guide to reference when deciding which dressing to use. The objective of this review is to identify the currently available biologic dressings, describe their indications, and provide a framework for integration into clinical practice. This review included 53 studies consisting of prospective and retrospective cohorts as well as several randomized control trials. Three general categories of cell-based biologic dressings were identified and nine brands were included. Cell-based biologic dressings have shown efficacy in a broad range of scenarios, and studies examining their efficacy have improved our understanding of the pathophysiology of chronic wounds. Amniotic and placental membranes have the widest scope and can be used to treat all subtypes of chronic wounds. Human skin allografts and bioengineered skin substitutes can be used for chronic ulcers but generally require a vascularized wound bed. Autologous platelet rich plasma (PRP) has shown promise in venous stasis ulcers and decubitus ulcers that have failed conventional treatment. Overall, more research is necessary to determine if these novel therapeutic options will change the current SOC, but current studies demonstrate encouraging results in the treatment of chronic wounds.

Retrospective observational analysis of the use of an architecturally unique dermal regeneration template (Derma Pure®) for the treatment of hard-to-heal wounds.

The purpose of this analysis was to evaluate the use of DermaPure, a decellularised human skin allograft, in the treatment of a variety of challenging wounds. This retrospective observational analysis reviewed a total of 37 patients from 29 different wound clinics across the USA. Each patient received one application of DermaPure which was followed until complete closure. A statistical analysis was performed with the end point being complete healing. All wounds on average, had a duration of 56 weeks and healed in an average time of 10·58 weeks. Individual wound categories included diabetic foot ulcers, which healed in 8·21 weeks; venous leg ulcers, which healed in 11·29 weeks; and surgical/traumatic wounds, which healed in 11·8 weeks.

Characterization of a Cryopreserved Split-Thickness Human Skin Allograft–TheraSkin

The purpose of this study was to examine the characteristics of a cryopreserved split-thickness skin allograft produced from donated human skin and compare it with fresh, unprocessed human split-thickness skin. Cutaneous wound healing is a complex and organized process, where the body re-establishes the integrity of the injured tissue. However, chronic wounds, such as diabetic or venous stasis ulcers, are difficult to manage and often require advanced biologics to facilitate healing. An ideal wound care product is able to directly influence wound healing by introducing biocompatible extracellular matrices, growth factors, and viable cells to the wound bed. TheraSkin (processed by LifeNet Health, Virginia Beach, Virginia, and distributed by Soluble Systems, Newport News, Virginia) is a minimally manipulated, cryopreserved split-thickness human skin allograft, which contains natural extracellular matrices, native growth factors, and viable cells. The authors characterized TheraSkin in terms of the collagen and growth factor composition using ELISA, percentage of apoptotic cells using TUNEL analysis, and cellular viability using alamarBlue assay (Thermo Fisher Scientific, Waltham, Massachusetts), and compared these characteristics with fresh, unprocessed human split-thickness skin. It was found that the amount of the type I and type III collagen, as well as the ratio of type I to type III collagen in TheraSkin, is equivalent to fresh unprocessed human split-thickness skin. Similar quantities of vascular endothelial growth factor, insulinlike growth factor 1, fibroblast growth factor 2, and transforming growth factor β1 were detected in TheraSkin and fresh human skin. The average percent of apoptotic cells was 34.3% and 3.1% for TheraSkin and fresh skin, respectively. Cellular viability was demonstrated in both TheraSkin and fresh skin.

Disinfection of human skin allografts in tissue banking: a systematic review report.

The use of skin allografts to temporarily replace lost or damaged skin is practiced worldwide. Naturally occurring contamination can be present on skin or can be introduced at recovery or during processing. This contamination can pose a threat to allograft recipients. Bacterial culture and disinfection of allografts are mandated, but the specific practices and methodologies are not dictated by standards. A systematic review of literature from three databases found 12 research articles that evaluated bioburden reduction processes of skin grafts. The use of broad spectrum antibiotics and antifungal agents was the most frequently identified disinfection method reported demonstrating reductions in contamination rates. It was determined that the greatest reduction in the skin allograft contamination rates utilized 0.1 % peracetic acid or 25 kGy of gamma irradiation at lower temperatures.Electronic supplementary materialThe online version of this article (doi:10.1007/s10561-016-9569-2) contains supplementary material, which is available to authorized users.

Sometimes patience pays off: When amputation is the alternative

Extended lower extremities defects due to systemic diseases can lead to amputation. This study aims to illustrate the successful treatment of two patients with chronic and therapy-resistant soft tissue defects of the lower leg, which healed with the use of gamma-irradiated human skin allograft (Gammagraft™, Promethean LifeSciences Inc, Pittsbourgh, PA/USA) and artificial dermis (Integra™, Integra LifeSciences Corporation, Plainsboro, NJ/USA) respectively. Both patients presented at admission with extended lower leg necrotic ulcers and both had a positive history of vasculopathy. The patient A was a 42 years old, male, with Scleromyxedema (SM) and the patient B was a 52 years old, female, diabetic and thrombophilic, which also suffered from Systemic Lupus Erythematosus (SLE). Serial wound debridements, amputation of single toes and skin grafting, were paired with modern reconstructive tools, like hydrosurgery system (Versajet◊, Smith &amp; Nephew, London/UK), Vacuum Assisted Closure system (VAC®, KCI, San Antonio, Tx/USA) and tissue engineered products: Gammagraft™ on patient A and Integra™ on patient B. The follow up revealed a long-term durable coverage of the defects. While there is no evidence to support a gold standard for extended chronic lower extremities defects of such origin, whose other alternative seems to be amputation, we believe that patience combined with the use of different reconstructive methods can add another valuable tool in our armamentarium and should considered as a valid reconstructive option.

The clinical use of cryopreserved human skin allografts for transplantation

Abstract Background The biological recovery of human skin allografts is the gold standard for preservation in Skin Banks. However, there is no worldwide consensus about specific allocation criteria for preserved human skin allografts with living cells. A report is presented on the results of 5 years of experience of using human skin allografts in burned patient in the Skin and Tissue Bank at Instituto Nacional de Rehabilitacion . Material and methods The human skin allografts were obtained from multi-organ donors, processed and preserved at −80 °C for 12 months. Allocation criteria were performed according to blood type match, clinical history, and burned body surface. Results Up to now, the Skin and Tissue Bank at Instituto Nacional de Rehabilitacion has processed and recovered 125,000 cm 2 of human skin allografts. It has performed 34 surgical implants on 21 burned patients. The average of burn body surface was 59.2%. More than two-thirds (67.7%) of recipients of skin allografts were matched of the same to type blood of the donor, and 66.6% survived after 126 days hospital stay. Conclusion It is proposed to consider recipient's blood group as allocation criteria to assign tissue; and use human skin allografts on patiens affected with burns over 30% of body surface (according the “rule of the 9”).

Skin donors and human skin allografts: evaluation of an 11-year practice and discard in a referral tissue bank.

The Saint Louis hospital tissue bank provides skin allografts to pediatric and adult burn units in the Paris area. The aim of this study was to analyze our activity during the last 11 years focusing on the reasons for skin discard. Skin is procured solely from the back of the body, which is divided into 10 zones that are harvested and processed separately. This retrospective study included all skin donors harvested between June 2002 and June 2013, representing a total of 336 donors and 2770 zones. The donors were multiorgan heart-beating donors in 91 % of cases (n = 307). The main reason for discarding harvested skin was microbial contamination, detected in 99 donors (29 %). Most contaminants were of low pathogenicity. Other reasons for discard included positive serologic tests for 2 donors [17 zones (0.61 %)], unsuitable physical skin characteristics for 3 zones (0.11 %), the donor's medical history for 53 zones (1.91 %), and technical issues with processing or distribution for 61 zones (2.2 %). In our experience, microbial contamination continues to be the main reason for discarding potential skin allografts. However, discards are limited by separate harvesting and processing of multiple zones in each donor.

El uso clínico de aloinjertos de piel humana criopreservados con fines de trasplante

BackgroundThe biological recovery of human skin allografts is the gold standard for preservation in Skin Banks. However, there is no worldwide consensus about specific allocation criteria for preserved human skin allografts with living cells.A report is presented on the results of 5 years of experience of using human skin allografts in burned patient in the Skin and Tissue Bank at the “Instituto Nacional de Rehabilitacion” Material and methodsThe human skin allografts were obtained from multi-organ donors. processed and preserved at -80°C for 12 months. Allocation criteria were performed according to blood type match, clinical history, and burned body surface. ResultsUp to now, the Skin and Tissue Bank at ‘Instituto Nacional de Rehabilitacion” has processed and recovered 125,000cm2 of human skin allografts. It has performed 34 surgical implants on 21 burned patients. The average of burn body surface was 59.2%. More than two-thirds (67.7%) of recipients of skin allografts were matched of the same to type blood of the donor, and 66.6% survived after 126 days hospital stay. ConclusionIt is proposed to consider recipient's blood group as allocation criteria to assign tissue; and use human skin allografts on patiens affected with burns over 30% of body surface (according the “rule of the 9”).

Recellularizing of human acellular dermal matrices imaged by high-definition optical coherence tomography.

High-definition optical coherence tomography (HD-OCT) permits real-time 3D imaging of the impact of selected agents on human skin allografts. The real-time 3D HD-OCT assessment of (i) the impact on morphological and cellular characteristics of the processing of human acellular dermal matrices (HADMs) and (ii) repopulation of HADMs in vitro by human fibroblasts and remodelling of the extracellular matrix by these cells. Four different skin decellularization methods, Dispase II/Triton X-100, Dispase II/SDS (sodium dodecyl sulphate), NaCl/Triton X-100 and NaCl/SDS, were analysed by HD-OCT. HD-OCT features of epidermal removal, dermo-epidermal junction (DEJ) integrity, cellularity and dermal architecture were correlated with reflectance confocal microscopy (RCM), histopathology and immunohistochemistry. Human adult dermal fibroblasts were in vitro seeded on the NaCl/Triton X-100 processed HADMs, cultured up to 19days and evaluated by HD-OCT in comparison with MTT proliferation test and histology. Epidermis was effectively removed by all treatments. DEJ was best preserved after NaCl/Triton X-100 treatment. Dispase II/SDS treatment seemed to remove all cellular debris in comparison with NaCl/Triton X-100 but disturbed the DEJ severely. The dermal micro-architectural structure and vascular spaces of (sub)papillary dermis were best preserved with the NaCl/Triton X-100. The impact on the 3D structure and vascular holes was detrimental with Dispase II/SDS. Elastic fibre fragmentation was only observed after Dispase II incubation. HD-OCT showed that NaCl/Triton X-100 processed matrices permitted in vitro repopulation by human dermal fibroblasts (confirmed by MTT test and histology) and underwent remodelling upon increasing incubation time. Care must be taken in choosing the appropriate processing steps to maintain selected properties of the extracellular matrix in HADMs. Processing HADMs with NaCl/Triton X-100 permits in vitro the proliferation and remodelling activity of human dermal fibroblasts. HD-OCT provides unique real-time and non-invasive 3D imaging of tissue-engineered skin constructs and complementary morphological and cytological information.

Anti-CD28 Antagonism Using a Monovalent Fab Antibody for the Prolongation of Human Skin Allograft Survival.

Background. T cells play a principal detrimental role in alloresponses after transplantation. Costimulation through CD28 induces T cells activation, cytokine production, proliferation, and differentiation, whereas the absence of CD28 signalling leads to T cell apoptosis or anergy. Importantly, the development of effector or regulatory responses after initial antigen exposure may be influenced by the balance between CD28 and cytotoxic T lymphocyte antigen 4 (CTLA-4) costimulatory signalling. Influencing this balance by blocking costimulation through CD28 is theoretically advantageous to agonistic signaling through CTLA4 (for example using CTLA4-Ig) as this would not interfere with CTLA4 and PDL-1 co-inhibitory signaling. In the current study we have investigated the hypothesis that a non-activating monovalent anti-CD28 antibody would prevent rejection of human skin allografts in vivo. Methods. Using a humanised mouse system, we treated mice that had received human skin transplants with FR104 (5 mg/kg), CTLA4-Ig (10mg/kg) or vehicle control intravenously twice a week for three weeks. All mice were started on the treatment regimen three weeks post-adoptive transfer of allogeneic human peripheral blood mononuclear cells (PBMCs) to ensure adequate levels of human leukocyte chimerism. Results. FR104 significantly prolonged skin allograft survival in humanised mice compared with CTLA4-Ig (median survival time, MST=63 vs. 31 (p=0.002)) or Saline (MST=31). While FR104 treatment significantly impacted the level of human leukocyte chimerism in the peripheral blood, it remained above the required reconstitution level known to induce skin rejection. The number of graft infiltrating CD4+ (47.8±24.8 vs 83.3±5.1 cells/HPF, P<0.02) and CD8+ (50.0±11.9 vs 111±34.0 cells/HPF, P<0.04) cells was significantly reduced by FR104 treatment as compared to CTLA-4 Ig treatment. No differences were noted in the number of graft infiltrating Foxp3+ cells between FR104 and CTLA4-Ig groups. FR104 significantly suppressed the serum cytokine production of IL-2, IFNγ, TNFα and IL-10. Conclusion. FR104 is an effective immunosuppressant in vivo. By sparing CTLA-4 and PDL-1 coinhibitory signals, FR104 may theoretically lead to improved therapeutic responses compared with CTLA-4-Ig.

Total scalp reconstruction following a dog bite in a pediatric patient.

The purpose of this case report is to present the reconstruction of a full-thickness scalp defect in a 2-year-old Hispanic male mauled by a dog, which was performed by a novel technique. After the scalp defect was debrided multiple times and treated with greater than 600 cm2 of cryopreserved human skin allograft, Integra was applied to the scalp defect following debridement and burring-out the outer bony cortex to promote bleeding and integration, followed by the application of a VAC dressing. Four weeks later, the mature Integra was grafted with split-thickness grafts. One year after the original injury, the patient demonstrated 98% take of the skin grafts to the calvarium. The resultant soft-tissue cover was supple, pliable, uniform in texture, and stable. The proposed technique of reconstruction of large full-thickness scalp defects in pediatric patients is easily reproducible, involves a short operative time, and produces satisfying cosmetic results.

The biomechanical and histological sequelae of common skin banking methods

Human skin allografts are used worldwide as an adjunct for the healing of burns when autograft skin is not available or not indicated. Allograft skin comes from human cadaveric donors, and so must be preserved until use. This study forms the first investigation to compare the mechanical and histological integrity of human split-thickness skin grafts preserved by either glycerolisation or cryopreservation (with or without the cryoprotectant DMSO). Stress relaxation was used to assess mechanical properties, whilst histological analysis allowed for evaluation of structural integrity.Preservation of tissue, whether by freezing or glycerolisation, altered the relaxation behaviours of skin. Young's modulus upon initial loading significantly decreased for skin frozen without cryoprotectant, but remained unchanged for skin frozen with cryoprotectant and skin preserved with glycerol. After 1.5h of stress relaxation, both fresh skin and skin frozen without DMSO displayed similar relaxation rates. Samples frozen with DMSO or preserved with glycerol had increased relaxation rate and had not reached load equilibrium within this time.To understand the structural basis for the biomechanical changes, samples were histologically assessed. All preservation protocols resulted in a similar degree of visible damage, but cryopreservation appeared particularly damaging to the extracellular matrix, whereas glycerolisation caused dramatic separation of the epidermis from the underlying dermis.The mechanical property alterations reveal that preservation results in laxity, which clinically could hinder contact dependent healing properties, but alternatively may increase capacity for coverage. The structural changes confirm that preservation techniques do not conserve grafts in an in vivo state.

An outcome analysis and long-term viability of cryopreserved cultured epidermal allografts: assessment of the conservation of transplantable human skin allografts

To assess the viability of cultured epithelium and preserved by freezing for periods varying from one month to one year. Samples of cultured epithelium were incubated in cryoprotectant medium (Group A), packed in aluminum envelopes and packed in polystyrene boxes. The boxes were subjected to a temperature of-70 ºC. After freezing for a period of time ranging from one to 12 months, cultured epithelial samples were assessed for their viability by vital staining (Trypan blue) and metabolic analysis based on glucose consumption and lactate production. Samples of not frozen cultured epithelium (Group B) were also tested for viability and the results obtained were used as comparison parameter for the variation of viability. Statistical analysis between the group A and B indicate that the mean age of the donors (p=0.51) and the culture time (p=1.18) showed no statistical difference. In 30 days we obtained 37% of the original viability of cultured epithelium, 25% at six months and one year, less than 15%. This trend was confirmed statistically with a reduction of approximately 1.8% of the original viability epithelium cultured every 30 days of storage. In the analysis by lactate production, similar results were observed. In the analysis by the glucose consumption results were not significant. The viability indices show statistically significant difference between the group A and B (p<0.0001). Although cryopreserved cultured epithelium showed significant reduction of viability, all samples remained viable. It was also found that the viability of cryopreserved cultured epithelial decreased as a function of storage time.