Human hair follicle dermal sheath and papilla cells support keratinocyte growth in monolayer coculture.

Traditional skin grafting techniques are effective but limited methods of skin replacement. Autologous transplantation of rapidly cultured keratinocytes is successful for epidermal regeneration, but the current gold-standard technique requires mouse fibroblast feeders and serum-rich media, with serum-free systems and dermal fibroblast (DF) feeders performing relatively poorly. Here, we investigated the capacity of human hair follicle dermal cells to act as alternative supports for keratinocyte growth. Dermal papilla (DP) dermal sheath (DS), DF and 3T3 cells were used as inactivated feeder cells for human keratinocyte coculture. Under conditions favouring dermal cells, proliferation of keratinocytes in the presence of either DS or DP cells was significantly enhanced compared with DF cells, at levels comparable to keratinocytes cultured under gold-standard conditions. Secreted protein acidic and rich in cysteine (SPARC) expression increased DS and DP cells relative to DFs; however, further experiments did not demonstrate a role in keratinocyte support.

Chronological histological findings of cultured epidermal autograft over bilayer artificial dermis.

The application of cultured epidermal autograft (CEA) over bilayer artificial dermis theoretically should minimize surgical stress and donor site morbidity in severe burn patients. However, CEA over regenerated dermis is very fragile and easily detaches soon after application, because the very weak attachment. We performed chronological histological studies of the wounds of a 29 year-old patient, which was reconstructed using CEA (JACE(®)) and bilayer artificial dermis (Integra™). These studies included immunohistochemistry of anti-collagen (types III, IV, and VII) and anti-laminin, in addition to H&E and EVG staining. Reconstructed epidermis and dermis showed almost normal histological appearance with time, but formation of basement membrane proteins was delayed. Absent or immature basement membrane protein in the early phase after the CEA application was considered to be an important problem. In the late phase after the CEA application over the bilayer artificial dermis, the reconstructed skin was very durable and demonstrated no sign of skin stripping (although there was still a lack of basement membrane proteins).

Optimal use of negative pressure wound therapy for skin grafts.

Skin grafting is a technique used for transplanting human skin (i.e. epidermal and some dermal layers) from a harvest site to a recipient site. However, advancements in bioengineered matrices have also introduced alternatives to skin grafts. The method used to secure the graft, whether skin or biomatrix, is critical in reducing graft failure. During the past several years, negative pressure wound therapy using reticulated open-cell foam (NPWT/ROCF; V.A.C.® Therapy, KCI USA, Inc., San Antonio, TX) has become a well-established method for bolstering grafts to recipient beds and is being used more frequently over biomatrices to help improve graft outcomes. This review will combine expert opinion with scientific evidence for the use of NPWT/ROCF over grafts.

Bioelectrical impedance assessment of wound healing.

Objective assessment of wound healing is fundamental to evaluate therapeutic and nutritional interventions and to identify complications. Despite availability of many techniques to monitor wounds, there is a need for a safe, practical, accurate, and effective method. A new method is localized bioelectrical impedance analysis (BIA) that noninvasively provides information describing cellular changes that occur during healing and signal complications to wound healing. This article describes the theory and application of localized BIA and provides examples of its use among patients with lower leg wounds. This promising method may afford clinicians a novel technique for routine monitoring of interventions and surveillance of wounds.

Quality evaluation of meshed split-thickness skin grafts stored at 4°C in isotonic solutions and nutrient media by cell cultures.

OBJECTIVE: Excess split-skin autografts harvested and meshed during burn surgery are often stored at 4°C temporarily for later use. The quality of the stored skin is critical to clinical outcome and needs to be assured. METHODS: Meshed split-thickness skin graft (mSSG) stored in saline, Hartmann's solution and two cell culture media, Dulbecco's Modified Eagle Medium (DMEM) and DMEM/Ham F12 (DMEM/F12, 3:1 mixture) were analyzed by trypan blue staining, cell culture and microbiological testing through a 28-day time course for cell viability and microbial contamination. RESULTS: mSSG samples in all groups showed a progressive decrease of cell viability and colony forming efficiency through the time course of storage at 4°C. Cell culture media were better than saline and Hartmann's solution in maintaining the viability and growth capability of skin cells. The viability observed by trypan blue staining did not truly reflect the cell growth capacity after storage. mSSG in saline and Hartman's solution retained minimal keratinocyte growth potency after 7 days. mSSG in cell culture media had significant loss of keratinocyte colony growth potency after 7 days and minimal keratinocyte growth after 14 days. Dermal fibroblasts of all groups were less tolerant than keratinocytes to the storage. Microbial contaminations were common in mSSG harvested from burn surgery. CONCLUSIONS: Culture media instead of saline or Hartman's solution should be used for temporary storage of mSSG at 4°C. The stored mSSS should be used within seven days to have sufficient viable number and cell growth efficiency. After then, the efficacy of stored mSSG as a source of living cells for wound closure could be full of uncertainty due to significant decrease of keratinocyte colony forming efficiency. Precaution should be taken during skin harvest and storage to minimize the risk of sample contamination. Inclusion of antimicrobial agents in storage solution and microbiological testing are advisable to ensure the quality and clinical outcome.

Haptoglobin activates innate immunity to enhance acute transplant rejection in mice.

Immune tolerance to transplanted organs is impaired when the innate immune system is activated in response to the tissue necrosis that occurs during harvesting and implantation procedures. A key molecule in this immune pathway is the intracellular TLR signal adaptor known as myeloid differentiation primary response gene 88 (MyD88). After transplantation, MyD88 induces DC maturation as well as the production of inflammatory mediators, such as IL-6 and TNF-α. However, upstream activators of MyD88 function in response to transplantation have not been identified. Here, we show that haptoglobin, an acute phase protein, is an initiator of this MyD88-dependent inflammatory process in a mouse model of skin transplantation. Necrotic lysates from transplanted skin elicited higher inflammatory responses in DCs than did nontransplanted lysates, suggesting DC-mediated responses are triggered by factors released during transplantation. Analysis of transplanted lysates identified haptoglobin as one of the proteins upregulated during transplantation. Expression of donor haptoglobin enhanced the onset of acute skin transplant rejection, whereas haptoglobin-deficient skin grafts showed delayed acute rejection and antidonor T cell priming in a MyD88-dependent graft rejection model. Thus, our results show that haptoglobin release following skin necrosis contributes to accelerated transplant rejection, with potential implications for the development of localized immunosuppressive therapies.

Identification of the Fra-1 transcription factor in healing skin flaps transplants: a potential role as a negative regulator of VEGF release from keratinocytes.

The molecular mechanisms underlying successful myocutaneous skin flap integration, as well as the ischemic loss of transplanted tissue on surgery, remain largely unknown. In this study we used a mouse model of caudally based skin flaps to determine molecular patterns of acute transplant re-integration. Gene chip-based transcriptional analysis revealed an up-regulation of the transcription factor Fra-1 in murine skin flap tissue. Epidermal keratinocytes at the wound margins represented a dominant cellular source of Fra-1 in mice. Moreover, Fra-1 protein showed a clear nuclear localization. In addition, Fra-1 protein was also present in nuclei of wound margin keratinocytes located near the suture line in human skin flaps. In vitro studies using the human keratinocyte cell line HaCaT showed that epidermal growth factor (EGF) was a potent inducer of Fra-1 expression in keratinocytes. Ablation of Fral-1 protein using a specific Fra-1 small interfering (si)RNA markedly increased the EGF-induced vascular endothelial growth factor (VEGF) expression from keratinocytes. These data suggest the involvement of an injury-induced Fra-1 transcription factor as a regulator of keratinocyte gene expression, which might act as an antagonistic player to restrict epithelial-driven angiogenic responses during normal skin flap integration.

Smoking and diabetes mellitus type 2 reduce skin graft take; the use of fibrin glue might restore graft take to optimal levels.

Efficacy has been demonstrated in some uses of fibrin glue associated with graft loss. Smoking and hyperglycemia significantly decrease the success of skin graft survival in specific injuries. This retrospective study aimed to verify the association with decreased skin graft survival and whether fibrin glue is useful in reversing the influence of these factors. This bicentric, retrospective, cross sectional case control study was carried out on 1881 medical patients. Patients who met inclusion criteria were admitted to the Plastic Surgery Service of Reina Sofia University Hospital (Spain) and the Trauma/Burn intensive Care Unit of UAB Hospital at Birmingham (USA) between January 2000 and December 2009. The successful graft take for each group and its control were analyzed by a Chi-square test; the confidence interval was 95%. Smoking and DM type 2 decrease skin graft survival when compared with their control groups. There was a statistically significant improvement in skin graft take when fibrin glue was used. The percentage improvement in the control groups was approximately 10%, whereas in the study groups it was 2-3 times higher. We conclude that graft loss is associated with smoking and DM type 2, but fibrin glue might restore graft adherence to almost normal levels.

Improvement of skin-graft survival after autologous transplantation of adipose-derived stem cells in rats.

BACKGROUND: Skin grafts are frequently used for a variety of indications in plastic and reconstructive surgery. Their necrosis is a common complication, while different therapies have been proposed. Currently, adipose-derived stem cells (ASCs) hold great promise for their angiogenic potential and role during tissue repair. In this study, autologous transplantation of ASCs was used in skin grafts in rats to determine if it increases angiogenesis, skin-graft survival and wound healing. METHODS: ASCs were isolated, cultured, labelled with fluorescent dye and injected under full-thickness skin grafts in 10 rats (group 1), while 10 others served as controls (group 2). Skin grafts were analysed after 1 week. Collagen's framework was assessed with Masson's trichrome stain and angiogenesis with von Willebrand factor (vWF) immunohistochemistry. In addition, immunohistochemical staining intensity of vascular endothelial growth factor (VEGF) and transforming growth factor b3 (TGFb3) was assessed in all grafts. RESULTS: Mean area of graft necrosis was significantly less in group 1 than in group 2 (6.12% vs. 32.62%, p<0.01). Statistically significant increase of microvessel density, collagen density, VEGF and TGFb3 expression was noted in group 1 compared with group 2 (all: p<0.01). CONCLUSIONS: These findings suggest that autologous ASCs transplantation increases full-thickness skin-graft survival and shows promise for use in skin-graft surgery. This might be both due to in situ differentiation of ASCs into endothelial cells and increased secretion by ASCs of growth factors, such as VEGF and TGFb3 that enhance angiogenesis and wound healing.

Inflammatory and oxidative stress after surgery for the small area corrections of burn sequelae.

PURPOSE: To compare vitamin levels, inflammatory and oxidative stress markers before and after skin autograft surgery to correct burn scar areas. METHODS: This prospective study was conducted with 8 patients with a median age of 28 years (range, 16 to 40 years) that had burn sequelae and were admitted to a Burn Unit for correction of small burn scar areas [3.3 (1.0-5.0) % of the corporal surface]. The volunteers were evaluated before and 48 hours after excision of scar tissue and skin autograft. Routine laboratory data, along with a food questionnaire and anthropometry were collected in the preoperative period. Serum vitamin A, C, E, B(12) and folic acid levels, inflammatory markers (C-protein reactive, alpha-1-acid glycoprotein, ferritin) and oxidative stress markers (reduced glutathione - GSH and Thiobarbituric Acid Reactive Substances - TBARS) were determined at preoperative and postoperative phases. Data were analyzed with two-sample Wilcoxon test. RESULTS: All volunteers were clinically stable and had adequate nutritional status at admission. After surgery, C-reactive protein serum levels increased [0.4 (0.01-1.0) vs. 2.5 (0.6-4.7) mg/dL, p=0.01] and vitamin A levels decreased [3.4 (2.1-4.2) vs. 2.4 (1.6-4.1) µmol/L, p=0.01]. No changes occurred in other vitamins, ferrritin, alpha-1-acid glycoprotein, GSH and TBARS levels. CONCLUSION: Minimal metabolic changes were produced after skin autograft in small areas of well-nourished patients without active infection or inflammation.

Inflammatory and oxidative stress after surgery for the small area corrections of burn sequelae / Estresse inflamatório e oxidativo após cirurgia para correção de pequenas áreas de sequela de queimaduras

PURPOSE: To compare vitamin levels, inflammatory and oxidative stress markers before and after skin autograft surgery to correct burn scar areas. METHODS: This prospective study was conducted with 8 patients with a median age of 28 years (range, 16 to 40 years) that had burn sequelae and were admitted to a Burn Unit for correction of small burn scar areas [3.3 (1.0-5.0) percent of the corporal surface]. The volunteers were evaluated before and 48 hours after excision of scar tissue and skin autograft. Routine laboratory data, along with a food questionnaire and anthropometry were collected in the preoperative period. Serum vitamin A, C, E, B12 and folic acid levels, inflammatory markers (C-protein reactive, alpha-1-acid glycoprotein, ferritin) and oxidative stress markers (reduced glutathione - GSH and Thiobarbituric Acid Reactive Substances - TBARS) were determined at preoperative and postoperative phases. Data were analyzed with two-sample Wilcoxon test. RESULTS: All volunteers were clinically stable and had adequate nutritional status at admission. After surgery, C-reactive protein serum levels increased [0.4 (0.01-1.0) vs. 2.5 (0.6-4.7) mg/dL, p=0.01] and vitamin A levels decreased [3.4 (2.1-4.2) vs. 2.4 (1.6-4.1) µmol/L, p=0.01]. No changes occurred in other vitamins, ferrritin, alpha-1-acid glycoprotein, GSH and TBARS levels. CONCLUSION: Minimal metabolic changes were produced after skin autograft in small areas of well-nourished patients without active infection or inflammation.

OBJETIVO: Comparar os níveis séricos de vitaminas e dos marcadores de estresse oxidativo e inflamatório antes e após enxerto cutâneo para correção de pequenas áreas de cicatrizes hipertróficas de queimaduras. MÉTODOS: O estudo prospectivo foi conduzido com oito pacientes com mediana de idade de 28 anos (variação de 16 a 40 anos) que apresentavam cicatrizes fibróticas decorrentes de queimaduras. Todos os pacientes foram admitidos em Unidade de Queimados para serem submetidos a enxertos autólogos [3,3 (1,0 a 5,0) por cento da superfície corporal]. Os voluntários foram avaliados antes e 48 horas após a excisão do tecido cicatricial e do enxerto autólogo. Exames laboratoriais de rotina, além do questionário alimentar e da antropometria foram obtidos no período pré-operatório. Os níveis séricos das vitaminas A, C, E, B12 e ácido fólico, os marcadores inflamatórios (proteína C reativa, alfa-1-glicoproteína ácida e ferritina) e marcadores de estresse oxidativo (glutationa reduzida - GSH e Substâncias Reativas do Ácido Tiobarbitúrico - TBARS) foram determinados nas fases pré e pós-operatórias. Os dados foram analisados pelo teste de Wilcoxon pareado. RESULTADOS: Todos os voluntários estavam clinicamente estáveis e apresentavam estado nutricional adequado à admissão hospitalar. Após a cirurgia, houve aumento dos níveis séricos de proteína C reativa [0,4 (0,01-1,0) vs 2,5 (0,6-4,7) mg/dL, p=0,01], enquanto houve redução nos níveis de vitamina A [3,4 (2,1-4,2) vs 2,4 (1,6-4,1) µmol/L, p=0,01]. Não houve mudanças nos níveis séricos de outras vitaminas, ferritina, alfa-1-glicoproteína ácida, GSH e TBARS. CONCLUSÃO: Em pacientes com bom estado nutricional e sem evidência de atividade inflamatória ou infecciosa ocorrem mudanças metabólicas mínimas após enxerto autólogo de pequenas áreas de cicatrizes de queimadura.

DNA release and uptake associated with the development of pleomorphic cells in mammalian skin autotransplants / Liberación y absorción de ADN asociada con el desarrollo de células pleomórficas en autotrasplantes de piel en mamíferos

OBJECTIVE: Although several in vitro studies have demonstrated active release of DNA by living cells, there is still doubt. There are no such in vivo studies (1). The following experiment is an in vivo study to determine whether DNA release and uptake by cells and tissues occur and can be related to normal growth and differentiation, abnormal growth and cancer. METHODS: Epidermal and full-thickness ear-skin grafts were separately autotransplanted into two groups of mice. In a second group, host mice were labelled with tritiated thymidine and autografted separately, with unlabelled epidermal and full-thickness ear-skin grafts. Animals were sacrificed regularly in both cases. RESULTS: Full thickness grafts revealed cysts in 15 out of 16 grafts, with well-differentiated squamous epidermis, DNA labelling ofdermal fibroblasts and no DNA labelling ofepidermal cells. Epidermal grafts revealed cysts in six out of 20 grafts, with epidermal cells variable in shape and arrangement; some appeared normal but others were two to four times larger, forming solid nests ofcells. In some grafts, there were spindle-shaped pleomorphic cells loosely interconnected. DNA labelling was ob served in occasional epidermal cell. Two lung adenocarcinomas were found. CONCLUSION: These results suggest active release of DNA by host cells and DNA uptake by grafted cells. This phenomenon and the differential uptake of DNA labelling ofepidermal and dermal cells in the epidermal and full-thickness grafts suggest an association with abnormal, even pleomorphic epidermal cell behaviour due to the interference of dermal/epidermal interacting factors.

OBJETIVO: Aunque varios estudios in vitro han demostrado la liberación activa de DNA por las células vivas, todavía persisten las dudas. No existen tales estudios in vitro (1). El siguiente experimento constituye un estudio in vitro para determinar si hay liberación y absorción de ADN por parte de las células y los tejidos, y si estos procesos guardan relación con el crecimiento normal y la diferenciación, así como con el crecimiento anormal y el cáncer. MÉTODOS: Injertos de piel de la oreja, tanto de espesor total como epidérmicos fueron auto trasplantados por separado a dos grupos de ratones. En el segundo grupo, ratones huéspedes fueron etiquetados con timidina tritiada, y autoinjertados, por separado, con injertos de piel de la oreja no etiquetados, tanto de espesor total como epidérmicos. En ambos casos, fue necesario sacrificar animales de manera regular. RESULTADOS: Los injertos de espesor total revelaron quistes en 15 de cada 16 injertos, con epidermis escamosa bien diferenciada, etiquetado ADN de fibroblastos dérmicos, y no etiquetado ADN de células epidérmicas. Los injertos epidérmicos revelaron quistes en seis de 20 injertos, siendo las células epidérmicas variables en forma y ordenamiento. Algunas parecían normales, pero otras eran de dos a cuatro veces mayores, y formaban anidamientos celulares sólidos. En algunos de los injertos, se presentaron células pleomórficas en forma de huso, interconectadas con laxitud. Se observó etiquetado de DNA en células epidérmicas ocasionalmente. Se hallaron dos adenocarcinomas pulmonares. CONCLUSIÓN: Estos resultados sugieren la liberación activa de ADN por las células huéspedes y la absorción de ADN por las células injertadas. Este fenómeno y la absorción diferencial de etiquetado de ADN de células dérmicas y epidérmicas en los injertos epidérmicos y de espesor total, sugieren una asociación con el comportamiento celular anormal, e incluso pleomórfico epidérmico, debido a la interferencia de los factores dérmicos/epidérmicos interactuantes.

Cell-based vascularization strategies for skin tissue engineering.

Providing a blood-vascular network to promote survival and integration of cells in thick dermal substitutes for application in full-thickness wounds is essential for the successful outcome of skin tissue engineering. Nevertheless, promoting vascularization also represents a critical bottleneck in today's skin tissue engineering practice. Several cell types have been considered and tested, mostly in preclinical studies, to increase vascularization. When the clinical situation allows delayed reconstruction of the defect, an autologous approach is preferable, whereas in acute cases allogeneic therapy is needed. In both cases, the cells should be harvested with minimal donor-site morbidity and should be available in large amounts and safe in terms of tumor formation and transmission of animal diseases. Here, we outline the different mechanisms of cell-based vascularization and subsequently elaborate in more detail on the candidate cell types and their pros and cons in terms of clinical application and regulation of the wound healing process.

DNA release and uptake associated with the development of pleomorphic cells in mammalian skin autotransplants.

OBJECTIVE: Although several in vitro studies have demonstrated active release of DNA by living cells, there is still doubt. There are no such in vivo studies (1). The following experiment is an in vivo study to determine whether DNA release and uptake by cells and tissues occur and can be related to normal growth and differentiation, abnormal growth and cancer METHODS: Epidermal and full-thickness ear-skin grafts were separately autotransplanted into two groups of mice. In a second group, host mice were labelled with tritiated thymidine and autografted separately, with unlabelled epidermal and full-thickness ear-skin grafts. Animals were sacrificed regularly in both cases. RESULTS: Full thickness grafts revealed cysts in 15 out of 16 grafts, with well-differentiated squamous epidermis, DNA labelling of dermalfibroblasts and no DNA labelling of epidermal cells. Epidermal grafts revealed cysts in six out of 20 grafts, with epidermal cells variable in shape and arrangement; some appeared normal but others were two to four times larger, forming solid nests of cells. In some grafts, there were spindle-shaped pleomorphic cells loosely interconnected. DNA labelling was observed in occasional epidermal cell. Two lung adenocarcinomas were found. CONCLUSION: These results suggest active release of DNA by host cells and DNA uptake by grafted cells. This phenomenon and the differential uptake of DNA labelling of epidermal and dermal cells in the epidermal and full-thickness grafts suggest an association with abnormal, even pleomorphic epidermal cell behaviour due to the interference of dermal/epidermal interacting factors.

Temporary angiogenic transformation of the skin graft vasculature after reperfusion.

BACKGROUND: In the era of tissue engineering, the physiologic process of skin graft revascularization remains unclear, preventing the successful development of skin substitutes. Therefore, the authors developed a new in vivo model with which to visualize the process of engraftment and its microvascular architecture. The aim of this study was to specifically investigate the vascular transformations within the skin graft to gain applicable knowledge on how vascular processes during engraftment occur. METHODS: Microsurgical preparation of the modified dorsal skinfold chamber including autologous skin grafting was performed in male C57BL/6J mice (n = 10). In addition, immunohistochemistry of angiogenic factors, endothelial cells, and pericytes, and corrosion casting were performed to further characterize the specific mechanisms. RESULTS: The graft exhibited capillary widening starting at day 3, resulting in the temporary formation of spherical protrusions at the graft capillary divisions starting in the center of the graft 24 to 48 hours after revascularization. Confocal microscopy showed the simultaneous expression of CD31 and desmin. Corrosion casting and evaluation by light microscopy and scanning electron microscopy showed the three-dimensional formation of capillaries in the wound bed that connected to the preexisting capillary loops of the skin graft. CONCLUSIONS: The authors were able to show for the first time a temporary angiogenic response within the capillaries of the skin graft. This most likely represents a reaction to reperfusion allowing the supply of proangiogenic factors to the hypoxic skin graft. The detection of an angiogenic response within the graft capillaries is for the first time made possible in the newly developed model and is therefore completely novel.

IL-1 signalling determines the fate of skin grafts expressing non-self protein in keratinocytes.

Although IL-1 is a known inflammatory cytokine during pathogen infection, the role of IL-1 in skin graft rejection, particularly where foreign antigen is expressed exclusively in keratinocytes, is less understood. Here, we use a syngeneic skin graft system, where antigens are expressed in epithelial cells via either a keratin 14 or keratin 5 promoter, to explore the role of IL-1 in graft rejection and induction of epithelial antigen-specific effector CD8(+) T-cell function. Keratin 5 ovalbumin (K5mOVA) transgenic skin grafts destined for rejection demonstrated increased expression of IL-1beta and its receptors compared to K14 HPV16 E7 transgenic grafts that do not reject spontaneously. Rejection of OVA grafts lacking the IL-1 receptor (IL-1R1) was delayed and associated with decreased numbers of antigen-specific CD8 T cells. In contrast, K14E7 grafts survived on immunocompetent, syngeneic recipients with decreased graft levels of IL-1beta and IL-1R1 and 2. However, in the absence of the IL-1 receptor antagonist, IL-1Ra, skin grafts were spontaneously rejected and an E7-specific CD8 T-cell response was primed. Thus, expression of the HPV16E7 oncoprotein in epithelial cells prevents IL-1beta-associated skin graft rejection and induction of antigen-specific CD8 T-cell responses. Enhancing IL-1beta signalling, via blocking of the IL-1 receptor antagonist, may represent an alternative strategy for treatment of HPV16E7-associated cancers.

Photoacoustic monitoring of granulation tissue grown in a grafted artificial dermis on rat skin.

In this study, we investigated the validity of photoacoustic (PA) measurement for monitoring granulation tissue and hence adhesion of grafted artificial dermis (AD). A 2.5 cm x 2.5 cm, 3-mm-thick AD composed of an atelocollagen sponge sheet and a silicone film was grafted on a full-thickness open wound in rat dorsal skin. The grafted AD was irradiated with low-energy, 532-nm nanosecond laser pulses to photoacoustically excite blood in neovascularities, and the PA signals induced were measured using a piezoelectric transducer as a function of postgrafting time. The PA signals were compared with results of laser Doppler imaging and histological analysis. We found a significant correlation between the depths of the first or shallowest PA signal peaks and the depths of granulation tissues estimated from histology with hematoxylin & eosin staining (R=0.951, p<0.05). There was also a significant correlation between the amplitudes of the first PA signal peaks and densities of CD31-positive cells evaluated from histology with immunohistochemical staining (R=0.859, p<0.05). With laser Doppler imaging, no clear perfusion signals were observed, which is attributable to a high light scattering loss in ADs. These findings suggest the validity of PA measurement for monitoring the adhesion of grafted ADs.

Allograft mass as a possible contributing factor to the skin transplant outcome.

BACKGROUND: A composite tissue consists of tissues derived from ectoderm and mesoderm typically containing skin, fat, muscle, nerves, lymph nodes, bone, bone marrow or any combination of these. Solid organ transplants possessing larger allograft mass were reported to survive better. As for the vascularized composite tissues however, thus far no study has comparatively studied the survival of grafts possessing different tissue burdens. The purpose of this study was to explore the effect of transplanted tissue burden and tissue type on survival of skin element of composite tissues. MATERIALS AND METHODS: Forty-five transplantations were performed using four different vascularized composite tissue allotransplantation models. The survival periods and rejection severity of the skin parts of the transplants were compared by histological, immunological, and macroscopic evaluation: (a) under no immunosuppressive treatment (control group) (n = 21); (b) after 1 week of Cyclosporine (CsA) treatment (16 mg/kg) (experimental group) (n = 24). Total rejection was defined as necrosis of >90% of the skin flaps. Histopathologic evaluation and flow cytometric analysis to detect chimerism rates in the blood was performed in the CsA treated animals on day 18. RESULTS: The differences of mean survival times between and within the experimental and control groups were statistically significant (P < 0.05). Histopathological outcomes showed lower rejection grades in skin allografts transplanted with a higher tissue burden. Total CD4+ and CD8+ T-cell chimerism rates were less than 1% in isolated skin transplant groups and ranged from 6.1 to 33.5% in skin flaps transplanted with the entire hindlimb or a part of it. CONCLUSION: The transplanted tissue burden as well as the tissue type can be an important factor for the skin transplant outcome.

Meshed skin grafts placed upside down can take if desiccation is prevented.

BACKGROUND: The role of the wet environment in wound healing has been investigated in various studies. The current study explores the role of the wet wound environment in promoting healing of skin grafts. The authors hypothesized that the survival of skin grafts is dependent not only on the orientation of transplantation but also on the environment into which the skin is transplanted. METHODS: This study included 72 full-thickness (2.5 x 2.5-cm) wounds in six Yorkshire pigs. The wounds were grafted with autologous split-thickness skin grafts (meshed or sheet), placed either regularly (dermal side down) or inverted (dermal side up), and treated in a wet or a dry environment. Behavior of the skin grafts and healing were analyzed in histologic specimens collected on days 4, 6, 9, and 12 after wounding. Wound contraction was quantified by photoplanimetry. RESULTS: In the wet environment, not only did inverted meshed skin grafts survive, but also they proliferated to accelerate reepithelialization. In this environment, wounds transplanted with inverted and regular meshed grafts showed no significant difference in reepithelialization rate and contraction. In contrast, in the dry environment, wounds transplanted with inverted meshed grafts showed a significantly lower reepithelialization rate and a higher contraction rate than wounds transplanted with regular grafts. Inverted meshed grafts in a dry environment and inverted sheet grafts did not survive. CONCLUSION: The wound environment has an important role in the survival and proliferation of skin grafts, as demonstrated by survival of inverted meshed grafts in the wet environment and their contribution to accelerated reepithelialization, equal to the regularly placed grafts.