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INTRODUCTION: Deep facial burns are often combined with inhalation injury that could lead to patient destabilization. Accurate timing of surgical debridement of deep burns in a critical patient is the real medical art. Especially in patients with deep burned face and hands, in whom early debridement promises better functional and aesthetic results. CASE PRESENTATION: A fifty-three-year-old woman sustained burns of 16% TBSA including face area. The treatment of the burn injury was complicated by severe inhalation trauma, which led to patient destabilization shortly after admission. Standard surgical debridement was risky at the time. We used a new enzymatic agent for early burn eschar necrolysis instead. All the debrided areas were temporarily covered with porcine xenografts. The facial burns healed spontaneously without the need for a skin transplant. Definitive surgery treatment of full-thickness burns was postponed until the patient´s stabilization. DISCUSSION: The new enzymatic debridement is minimally invasive and can be applied bedside without the need for general anesthesia. All advantages of the new enzymatic debridement had led to extend its use at the face area, although it was not tested in this area during pre-registration studies. Especially in facial area high selectivity and significant reduction of skin grafting expect a better aesthetic and functional outcome. CONCLUSION: Bromelain-based enzymatic debridement proved to be safe and effective on the face in a very high-risk patient with unstable circulation and severe inhalation injury as an alternative to tangential excision.

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Amniotic membrane is a biological material widely used in plastic and reconstructive surgery and in ophthalmology. Due to its excellent biocompatibility and strength we tried to use it as a scaffold for the in vitro cultivation of different cell types, especially keratinocytes and limbal stem cells. It was possible to cultivate limbal stem cells and keratinocytes without using 3T3 mouse fibroblast feeder cells on deep frozen amniotic membranes. The amniotic membrane can also be used as a carrier for suspensions of different types of cells, allowing a substantial reduction of the cultivation time needed to prepare cell cultures for clinical application to burn patients. Our results show that the amniotic membrane seems not only to be an excellent carrier for human keratinocytes and corneal limbal stem cells, but also for other cell types, including dermal fibroblasts, adipose tissue-derived mesenchymal stem cells and chondrocytes.

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Acellular matrices are used for various purposes and they have been studied extensively for their potential roles in regenerating tissues or organs. The acellular matrix generates physiological cues that mimic the native tissue microenvironment. Acellular dermal matrix (ADM) is a soft connective tissue graft generated by a decellularization process that preserves the intact extracellular skin matrix. Upon implantation, this structure serves as a scaffold for donor-side cells to facilitate subsequent incorporation and revascularization. In breast reconstruction, ADM is used mainly for lower pole coverage and the shaping of a new breast. It helps control the positioning of the implant in the inframammary fold, and prevent the formation of contractile pseudocapsule around the breast implant. In this study, we provide a comprehensive histological description of ADM used for human breast reconstruction over the course of several months following implementation. Using immunohistochemical methods (a panel of 12 antibodies) coupled with optical and transmission electron microscopy, we confirmed that the original acellular dermal matrix became recolonized by fibroblasts and myofibroblasts, and also by various other free cells of the connective tissue (lymphocytes, macrophages and multinucleated giant cells, granulocytes, mast cells) after implantation into the patient's body. Within the implanted ADM, there was a relatively rapid ingrowth of blood vessels. Lymphatic vessels were only detected in one case 9 months after the implantation of the ADM. These results suggest that lymphangiogenesis is a longer process than angiogenesis.

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Acellular dermal matrix (ADM) is a tissue graft of allogeneic origin from post-mortem tissue donors prepared by an innovative decellularization process. The newly developed non-toxic and low cost decellularization process of cadaver origin dermis included ADM in breast reconstruction procedures proved to help coverage of the lower-pole of breast expanders or implants. As the results have shown, it did help to eliminate autologous dermis donor site morbidity along with shortening the operation time by avoiding elevation of additional muscle or fascia during the operation. Main aims of this article include histology evaluation of allogeneic acellular dermal matrix prepared by a new decellularization method and presentation of clinical results of its use. A total of 22 patients underwent 26 ADM based breast reconstructions. The mean patient's follow up was 12.6 months. Average total size of ADM used for one breast was 273 cm2. Post-operative complications occurred in 3 patients including one expander infection, one expander extrusion and one expander pocket disfiguration. Microscopic analysis of tissue samples has confirmed incorporation of the acellular dermal matrices into the surrounding connective tissue without any noticeable immune reaction. In a majority of the ADM samples we found pseudocapsullar formation on implant side of samples without acute or chronic inflammatory cells. The use of ADM prepared by new preparation method in expansive post mastectomy breast reconstruction was associated by a relatively low complication rate resulting in good outcomes.

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In this study we evaluated the biocompatibility of a modified polyurethane nanofiber membrane on a polypropylene spunbond substrate. This material was treated with plasma using diffuse coplanar surface barrier discharge, and subsequent modification was done by continuous spraying of a biologically active chitosan solution (CHIT) containing an inclusion complex of ß-cyclodextrin (ß-CD) encapsulating berberine (BRB). Biocompatibility was evaluated using several in vitro assays. Human dermal fibroblasts (HDFs) and 3T3 murine fibroblasts were used as biological models. The results of these assays showed that a polyurethane nanofiber membrane modified by CHIT/ß-CD/BRB appears to be non-toxic and biocompatible; potentially, it could be used as a wound dressing after further testing.

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The preparation and study of three-dimensional functional skin substitutes has been the focus of intense research for several decades. Dermal substitutes are now commonly used in medical practice for a variety of applications. Here, we assess the toxicity of seven selected acellular dermal matrix materials to establish their potential for use in future three-dimensional skin substitute studies. The cytotoxicity of acellular dermis (of Allo- and Xenograft origin) prepared in our lab and biomaterials based on collagen and hyaluronic acid (Coladerm H and Coladerm H-L) were compared to that seen in three commercially available products (Xe-Derma, AlloDerm and Xeno-Impl). Murine fibroblasts NIH-3T3 and human dermal fibroblasts were used in cytotoxicity tests, with any resultant cytotoxic effects caused by the seven tested dermal scaffolds visualised using an inverted microscope system and confirmed in parallel using colorimetric MTT cell proliferation assays. While most of the dermal substitutes did not demonstrate a cytotoxic effect on our two cell types, Xeno and Xeno-Impl scaffolds clearly did. The cytotoxic effect of acellular Xeno dermal matrix could essentially be removed through a regime of multiple washes, but we were unable to remove the cytotoxic effect of Xeno-Impl. Thus, Xeno-Impl alone has been excluded from our future work on preparation of 3D skin substitutes.

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Substitution of skin, particularly in extensive burns, is one of the key points for patients mortality reduction. In addition to the use of allogeneic and autologous skin substitutes, new developments in tissue engineering would enable the use biosynthetic and combined skin substitutes, which could mimic the structure and functions of normal skin. Several such types of substitutes like cultured allogeneic and autologous keratinocytes, allogeneic/autologous composites, acellular matrices, matrices based on biological substances such as collagen/hyaluronic acid, and matrices seeded by different cell types (keratinocytes, dermal fibroblasts, stem cells) already exist. Recent development in skin substitutes research aims gradually to establish a fully functional skin substitute which could mimic skin not only by its structure, but which could be capable to assure also its revascularization, reinnervations, and replacement of skin appendages (hair follicles, sebaceous glands etc.) as well. Creation of such a skin substitute will require collaboration of a wide range of research specialists including molecular biology, material sciences, genetic and tissue engineering, computer sciences, and, of course, clinical specialists in the field of plastic surgery and burn medicine. Recent advances in this field are promising and give hope that in the near future such a fully functional skin substitute would become a reality. This article aims to give information on the available skin substitutes at the present time.

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Both allogenic and autologous cultured skin cells have been used clinically on burn patients. In vitro cultivation of human keratinocytes has been routinely provided by the Central Tissue Bank in Bratislava since 1996, with an average annual production of around 7,000 cm(2). Keratinocytes have been cultivated using a version of the original by Rheinwald and Green (Cell 6:317-330, 1975) methodology which has been modified over time in our laboratory as we gained more experience with this serial passage system. We have observed that the growth of cultured keratinocytes depends on several important factors, including the timing of skin sample procurement, the method of skin sample procurement, the general condition of the patient, the quality and composition of the culture media and, to a lesser extent, the age of the patient. We aim to share our experience with other cell cultivation facilities.

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Glucan preparations, primarily modified water-soluble glucans, are involved in the activation of the body's natural defense mechanisms and in the acceleration of the skin's wound-healing processes. Pleuran, an insoluble beta-D-glucan in hydrogel form, offers a natural alternative to more common chemically derivated soluble beta-D-glucans. Pleuran was applied to human keratinocyte primary cultures, and after 24 h of incubation the release of matrix metalloproteinase 9 (MMP-9) and metalloproteinase 2 (MMP-2) by stimulated keratinocytes was detected using gelatine zymography. There was a concentration-dependent increase in pro-MMP-9 release after treatment with pleuran over the concentration range of 2 to 200 microg/ml, but pro-MMP-2 was detected at a constant level. Moreover, the active forms of both MMPs were not detectable, indicating that in vitro autoactivation of these zymogens did not occur. The results indicate that pleuran is a potent keratinocyte stimulator of proMMP-9 release, which implies its application in dermatological therapies.

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The worldwide growing interest to biomaterials over the last years results from their irreplaceable role in medical clinic. Hydroxyapatite is used in bone reconstruction because of its similar chemical structure compared to the inorganic composition of human bone and it is basic building component of many newly prepared biomaterials. In this study, we evaluated cytotoxic/antiproliferative activity of hydroxyapatite extract using murine fibroblast cell line NIH-3T3 and two in vitro different cytotoxic assays: growth inhibition assay and MTT assay. Hydroxyapatite extract after 72 h of incubation manifested the significant in vitro cytotoxic/antiproliferative effect only at the highest concentration tested (100 %). The antiproliferative effect of hydroxyapatite extract at the other concentrations tested (75 %, 50 %, 25 %, 10 %, 5 % and 1 %) was directly proportional to the concentration and the time of influence. The inhibition of cell proliferation was 86.8 - 0 %. The sensitivity of cell growth inhibition assay (direct counting of viable cells) to the extract influence was higher than that of MTT test.

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