RESUMEN
Healing wounds represent a major public health problem, mainly when it is infected. Besides that, the antibiotics misuse and overuse favor the development of bacterial resistance. This study evaluated the effects of antimicrobial photodynamic therapy (aPDT) combined with artificial skin on disinfection of infected skin wound in rats. Twenty-four Wistar rats were randomly distributed into 4 groups (n = 6): (i) control-untreated; (ii) aPDT-treated with curcumin-mediated aPDT (blue light); (iii) artificial skin-treated with artificial skin alcohol-based; and (iv) aPDT plus artificial skin-treated with aPDT associated with artificial skin alcohol-based. For the in vivo model, a full-thickness biopsy with 0.80 cm was performed in order to inoculate the microorganism Staphylococcus aureus (ATCC 25923). The aPDT was performed with a curcumin gel and a blue LED light (450 nm, 80 mW/cm2) at the dose of 60 J/cm2 and the treatment with alcohol-based artificial skin was done with the topical application of 250 µL. Additional animals were submitted to aPDT combined with the artificial skin. After treatments, the number of colony-forming units (CFU) and the damage area were determined. Data were analyzed by two-way repeated measures ANOVA and Tukey tests. The highest reduction of the bacterial viability was observed in the PDT plus artificial skin group (4.14 log10), followed by artificial skin (2.38 log10) and PDT (2.22 log10) groups. In addition, all treated groups showed higher relative area of wound contraction (36.21% for the PDT, 38.41% for artificial skin, and 35.02% for PDT plus artificial) in comparison with the control group. These findings provide evidence for the positive benefits of aPDT with blue light and curcumin associated with artificial skin to decontaminate and accelerate the wound contraction.
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Curcumina/farmacología , Fotoquimioterapia , Piel Artificial/microbiología , Staphylococcus aureus/fisiología , Heridas y Lesiones/microbiología , Animales , Viabilidad Microbiana/efectos de los fármacos , Fármacos Fotosensibilizantes/farmacología , Ratas , Ratas Wistar , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/efectos de la radiaciónRESUMEN
Candida albicans is an opportunistic fungal infectious agent that can cause cutaneous candidiasis in humans. Biofilms formation of C. albicans is thought to be the major cause of antifungal drug resistance. Despite numerous studies conducted on C. albicans biofilms, a comprehensive understanding of how C. albicans biofilms induced cutaneous candidiasis in humans and the development of a more effective targeted therapy remain poorly investigated. Available animal models of cutaneous candidiasis and in vitro human skin cell cultures do not fully reflect the actual human skin microenvironment or the disease pathogenesis. We investigated the molecular pathology of C. albicans infection using an in vitro biofabricated 3D skin. This in vitro biofabricated 3D skin comprises a fully humanized three-dimensional (3D) skin equivalent, consisting of a stratified terminally differentiated epidermis and an underlying dermal compartment. Antifungal drug susceptibility testing, histological and electron microscopy study, biofilms study, and pro-inflammatory cytokines analysis were conducted in C. albicans infected skin. Histological results revealed that C. albicans covered and produced biofilm on the in vitro biofabricated 3D skin, invading the skin compartments including epidermis and dermis. Elevation of proinflammatory cytokines including MMP-9, IL-1ß, TNF-α, and IL-5 were examined in the C. albicans infected skin. However, treatment with itraconazole reduced the pathology of C. albicans infection. This study provides an alternative pathological model of cutaneous candidiasis, which can physiologically represent a close-up event during C. albicans. Moreover, it is rapid, cost-effective, and reproducible of the in vitro biofabricated 3D skin model, and may further highlight the importance of utilizing in vivo-like conditions to improve high-throughput screening for drug discovery against several antifungal drug resistant pathogens.
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Antifúngicos/farmacología , Biopelículas/efectos de los fármacos , Candida albicans/efectos de los fármacos , Técnicas In Vitro , Piel Artificial/microbiología , Candidiasis/tratamiento farmacológico , Farmacorresistencia Fúngica , Humanos , Pruebas de Sensibilidad MicrobianaRESUMEN
BACKGROUND: Biofilm formation represents a major microbial virulence attribute especially at epithelial surfaces such as the skin. Malassezia biofilm formation at the skin surface has not yet been addressed. OBJECTIVE: The present study aimed to evaluate Malassezia colonisation pattern on a reconstructed human epidermis (RhE) by imaging techniques. METHODS: Malassezia clinical isolates were previously isolated from volunteers with pityriasis versicolor and seborrhoeic dermatitis. Yeast of two strains of M furfur and M sympodialis were inoculated onto the SkinEthic™ RHE. The tissues were processed for light microscopy, wide-field fluorescence microscopy and scanning electron microscopy. RESULTS: Colonisation of the RhE surface with aggregates of Malassezia yeast entrapped in a multilayer sheet with variable amount of extracellular matrix was unveiled by imaging techniques following 24, 48, 72 and 96 hours of incubation. Whenever yeast were suspended in RPMI medium supplemented with lipids, the biofilm substantially increased with a dense extracellular matrix in which the yeast cells were embedded. Slight differences were found in the biofilm architectural structure between the two tested species with an apparently higher entrapment and viscosity in M furfur biofilm. CONCLUSION: Skin isolates of M furfur and M sympodialis were capable of forming biofilm in vitro at the epidermal surface simulating in vivo conditions. Following 24 hours of incubation, without added lipids, rudimental matrix was barely visible, conversely to the reported at plastic surfaces. The amount of biofilm apparently increased progressively from 48 to 96 hours. A structural heterogeneity of biofilm between species was found.
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Biopelículas , Epidermis/microbiología , Procesamiento de Imagen Asistido por Computador , Malassezia/aislamiento & purificación , Piel Artificial/microbiología , Dermatitis Seborreica/microbiología , Humanos , Malassezia/ultraestructura , Microscopía Electrónica de Rastreo , Tiña Versicolor/microbiologíaRESUMEN
Staphylococcal biofilms cause many infectious diseases and are highly tolerant to the effects of antimicrobials; this is partly due to the biofilm matrix, which acts as a physical barrier retarding the penetration and reducing susceptibility to antimicrobials, thereby decreasing successful treatment outcomes. In this study, both single and mixed micellar systems based on poly vinyl caprolactam (PCL)-polyethylene glycol (PEG) copolymers were optimised for delivery of chlorhexidine (CHX) to S. aureus, MRSA and S. epidermidis biofilms and evaluated for their toxicity using Caenorhabditis elegans. The respective polyethylene glycol (PEG) and poly vinyl caprolactam (PCL) structural components promoted stealth properties and enzymatic responsive release of CHX inside biofilms, leading to significantly enhanced penetration (56%) compared with free CHX and improving the efficacy against Staphylococcus aureus biofilms grown on an artificial dermis (2.4 log reduction of CFU). Mixing Soluplus-based micelles with Solutol further enhanced the CHX penetration (71%) and promoted maximum reduction in biofilm biomass (>60%). Nematodes-based toxicity assay showed micelles with no lethal effects as indicated by their high survival rate (100%) after 72â¯h exposure. This study thus demonstrated that bio-responsive carriers can be designed to deliver a poorly water-soluble antimicrobial agent and advance the control of biofilm associated infections.
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Antiinfecciosos/administración & dosificación , Clorhexidina/administración & dosificación , Óxido de Etileno/administración & dosificación , Lactonas/administración & dosificación , Micelas , Polietilenglicoles/administración & dosificación , Polivinilos/administración & dosificación , Staphylococcus aureus/efectos de los fármacos , Staphylococcus epidermidis/efectos de los fármacos , Animales , Antiinfecciosos/toxicidad , Biopelículas/efectos de los fármacos , Caenorhabditis elegans/efectos de los fármacos , Clorhexidina/toxicidad , Óxido de Etileno/toxicidad , Lactonas/toxicidad , Polietilenglicoles/toxicidad , Polivinilos/toxicidad , Piel Artificial/microbiología , Staphylococcus aureus/fisiología , Staphylococcus epidermidis/fisiologíaRESUMEN
We isolated and identified yeasts from burn wounds and evaluated the ability of Candida parapsilosis isolates from burn wounds to penetrate an acellular dermal matrix (ADM). A prospective study was conducted with patients from the burn treatment center of North Paraná University Hospital in Londrina, Brazil from February 2015 to January 2016. Yeast cultures were obtained from the tissue of burn wounds that had been debrided and cleansed with 2% chlorhexidine. After identification and confirmation of the purity of the culture, the yeasts were placed on ADM fragments and incubated for three or seven days. During the study period, 273 patients were treated, and 36 of these patients fulfilled the inclusion criteria and provided samples for culture. Yeasts were isolated in 19.44% (nâ¯=â¯7) of the cultures, and the following species were identified: C. parapsilosis (57.1%), C. albicans (28.6%), and C. glabrata (14.3%). C. parapsilosis, the most frequent species, was chosen for the ADM tests. We demonstrated active penetration of the ADM by the yeast isolates from burn wounds. C. parapsilosis grew on ADM and penetrated the matrix, indicating that this yeast, which is common in skin and cutaneous wounds, has the potential to colonize and pass through ADM, a medical device that is frequently used to dress and regenerate burn wounds.
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Dermis Acelular/microbiología , Quemaduras/microbiología , Candida parapsilosis/clasificación , Candida parapsilosis/aislamiento & purificación , Candidiasis/microbiología , Cicatrización de Heridas , Biopelículas/crecimiento & desarrollo , Brasil , Candida/clasificación , Candida/aislamiento & purificación , Candida albicans/aislamiento & purificación , Candida glabrata/aislamiento & purificación , Candida parapsilosis/patogenicidad , Candida parapsilosis/fisiología , Humanos , Estudios Prospectivos , Piel Artificial/microbiologíaRESUMEN
Hand hygiene, cleaning and disinfection are pre-requirements for hygiene management in hospital settings and the food industry. In order to facilitate risk management, different contamination scenarios and interventions need to be evaluated. In the present study data on transfer rates and reductions of Staphylococcus aureus were provided in an experimental set-up using artificial skin. Using this methodology, test persons were not exposed with pathogenic bacteria. An exposure assessment model was developed and applied to evaluate different contamination routes and hygiene interventions. The transfer rates of S. aureus from inoculated VITRO-SKIN® to fomites were calculated from blotting series. The VITRO-SKIN® was more prone to spread bacteria than fomites. When different surfaces were cleaned, the reduction of S. aureus varied between <1 and 7 log CFU. It could not be concluded that a certain coupon material, cleaning agent, cleaning wipe, soiling or humidity consistently resulted in a high or low reduction of S. aureus. The reduction of S. aureus and E. coli during hand washing was evaluated on artificial skin, VITRO-SKIN®. The reduction of E. coli on VITRO-SKIN® was similar to the log reduction obtained when washing human hands. The S. aureus count on a human hand was both calculated in different scenarios describing different contamination routes starting from a contaminated hand using the exposure assessment model, and measured on an experimental setup using VITRO-SKIN® for validation. A linear relationship was obtained between the analysed level of S. aureus and the calculated level. However, the calculated levels of S. aureus on the VITRO-SKIN® in the scenarios were 1-1.5 log lower than the analysed level. One of the scenarios was used to study the effect of interventions like hand washing and cleaning of surfaces.
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Descontaminación/métodos , Desinfección/métodos , Mano/microbiología , Infecciones Estafilocócicas/transmisión , Staphylococcus aureus/crecimiento & desarrollo , Desinfectantes/administración & dosificación , Fómites/microbiología , Desinfección de las Manos , Higiene de las Manos , Humanos , Piel Artificial/microbiología , Staphylococcus aureus/efectos de los fármacosRESUMEN
Children in community bodies like kindergartens are predisposed to suffer from impetigo. To consider important measures for infection prevention, direct and indirect transmission routes of pathogens must be revealed. Therefore, we studied the role of skin and fabrics in the spread of the impetigo pathogen Staphylococcus aureus and the strain Streptococcus equi (surrogate to Streptococcus pyogenes) in order to assess infection transfer in realistic scenarios. The transmission of test strains was studied with standardized fabric-skin models using a technical artificial skin and fabrics of different fiber types commonly occurring in German kindergartens. In synthetic pus, both test strains persisted on artificial skin and fabrics for at least 4 h. Friction enhanced transfer, depending on the fiber type or fabric construction. In a skin-to-skin setup, the total transfer was higher than via fabrics and no decrease in the transmission rates from donor to recipients could be observed after successive direct skin contacts. Children in kindergartens may be at risk of transmission for impetigo pathogens, especially via direct skin contact, but also by the joint use of fabrics, like towels or handicraft materials. Fabric-skin models used in this study enable further insight into the transmission factors for skin infections on the basis of a practical approach.
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Transmisión de Enfermedad Infecciosa , Impétigo/transmisión , Modelos Teóricos , Piel Artificial/microbiología , Staphylococcus aureus/aislamiento & purificación , Streptococcus equi/aislamiento & purificación , Textiles/microbiología , Guarderías Infantiles , Preescolar , Alemania , HumanosRESUMEN
Biofilm-protected microbial infections in skin are a serious health risk that remains to be adequately addressed. The lack of progress in developing effective treatment strategies is largely due to the transport barriers posed by the stratum corneum of the skin and the biofilm. In this work, we report on the use of Ionic Liquids (ILs) for biofilm disruption and enhanced antibiotic delivery across skin layers. We outline the syntheses of ILs, analysis of relevant physicochemical properties, and subsequent neutralization effects on two biofilm-forming pathogens: Pseudomonas aeruginosa and Salmonella enterica. Further, the ILs were also examined for cytotoxicity, skin irritation, delivery of antibiotics through the skin, and treatment of biofilms in a wound model. Of the materials examined, choline-geranate emerged as a multipurpose IL with excellent antimicrobial activity, minimal toxicity to epithelial cells as well as skin, and effective permeation enhancement for drug delivery. Specifically, choline-geranate was comparable with, or more effective than, bleach treatment against established biofilms of S. enterica and P. aeruginosa, respectively. In addition, choline-geranate increased delivery of cefadroxil, an antibiotic, by >16-fold into the deep tissue layers of the skin without inducing skin irritation. The in vivo efficacy of choline-geranate was validated using a biofilm-infected wound model (>95% bacterial death after 2-h treatment). This work establishes the use of ILs for simultaneous enhancement of topical drug delivery and antibiotic activity.
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Sistemas de Liberación de Medicamentos , Líquidos Iónicos/farmacología , Pseudomonas aeruginosa/fisiología , Salmonella enterica/fisiología , Administración Cutánea , Biopelículas/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Células Epiteliales/citología , Células Epiteliales/efectos de los fármacos , Humanos , Irritantes/toxicidad , Pruebas de Sensibilidad Microbiana , Pseudomonas aeruginosa/efectos de los fármacos , Reproducibilidad de los Resultados , Salmonella enterica/efectos de los fármacos , Piel/efectos de los fármacos , Piel Artificial/microbiología , Espectroscopía Infrarroja por Transformada de FourierRESUMEN
Burn wound infections are often difficult to treat due to the presence of multidrug-resistant bacterial strains and biofilms. Currently, mupirocin is used to eradicate methicillin-resistant Staphylococcus aureus (MRSA) from colonized persons; however, mupirocin resistance is also emerging. Since we consider antimicrobial peptides to be promising candidates for the development of novel anti-infective agents, we studied the antibacterial activities of a set of synthetic peptides against different strains of S. aureus, including mupirocin-resistant MRSA strains. The peptides were derived from P60.4Ac, a peptide based on the human cathelicidin LL-37. The results showed that peptide 10 (P10) was the only peptide more efficient than P60.4Ac, which is better than LL-37, in killing MRSA strain LUH14616. All three peptides displayed good antibiofilm activities. However, both P10 and P60.4Ac were more efficient than LL-37 in eliminating biofilm-associated bacteria. No toxic effects of these three peptides on human epidermal models were detected, as observed morphologically and by staining for mitochondrial activity. In addition, P60.4Ac and P10, but not LL-37, eradicated MRSA LUH14616 and the mupirocin-resistant MRSA strain LUH15051 from thermally wounded human skin equivalents (HSE). Interestingly, P60.4Ac and P10, but not mupirocin, eradicated LUH15051 from the HSEs. None of the peptides affected the excretion of interleukin 8 (IL-8) by thermally wounded HSEs upon MRSA exposure. In conclusion, the synthetic peptides P60.4Ac and P10 appear to be attractive candidates for the development of novel local therapies to treat patients with burn wounds infected with multidrug-resistant bacteria.
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Péptidos Catiónicos Antimicrobianos/farmacología , Biopelículas/efectos de los fármacos , Quemaduras por Electricidad/tratamiento farmacológico , Piel Artificial/microbiología , Heridas y Lesiones/tratamiento farmacológico , Secuencia de Aminoácidos , Antibacterianos/farmacología , Péptidos Catiónicos Antimicrobianos/síntesis química , Biopelículas/crecimiento & desarrollo , Quemaduras por Electricidad/microbiología , Epidermis/efectos de los fármacos , Epidermis/metabolismo , Epidermis/patología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Células Epiteliales/patología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Fibroblastos/patología , Humanos , Interleucina-8/biosíntesis , Interleucina-8/metabolismo , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Staphylococcus aureus Resistente a Meticilina/crecimiento & desarrollo , Pruebas de Sensibilidad Microbiana , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Mitocondrias/patología , Modelos Biológicos , Datos de Secuencia Molecular , Mupirocina/farmacología , Técnicas de Síntesis en Fase Sólida , Heridas y Lesiones/microbiología , CatelicidinasRESUMEN
Dermatophytes cause the majority of superficial mycoses in humans and animals. However, little is known about the pathogenicity of this specialized group of filamentous fungi, for which molecular research has been limited thus far. During experimental infection of guinea pigs by the human pathogenic dermatophyte Arthroderma benhamiae, we recently detected the activation of the fungal gene encoding malate synthase AcuE, a key enzyme of the glyoxylate cycle. By the establishment of the first genetic system for A. benhamiae, specific ΔacuE mutants were constructed in a wild-type strain and, in addition, in a derivative in which we inactivated the nonhomologous end-joining pathway by deletion of the A. benhamiae KU70 gene. The absence of AbenKU70 resulted in an increased frequency of the targeted insertion of linear DNA by homologous recombination, without notably altering the monitored in vitro growth abilities of the fungus or its virulence in a guinea pig infection model. Phenotypic analyses of ΔacuE mutants and complemented strains depicted that malate synthase is required for the growth of A. benhamiae on lipids, major constituents of the skin. However, mutant analysis did not reveal a pathogenic role of the A. benhamiae enzyme in guinea pig dermatophytosis or during epidermal invasion of the fungus in an in vitro model of reconstituted human epidermis. The presented efficient system for targeted genetic manipulation in A. benhamiae, paired with the analyzed infection models, will advance the functional characterization of putative virulence determinants in medically important dermatophytes.
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Arthrodermataceae/patogenicidad , Dermatomicosis/microbiología , Proteínas Fúngicas/genética , Eliminación de Gen , Recombinasas/genética , Factores de Virulencia/genética , Alopecia/microbiología , Animales , Arthrodermataceae/enzimología , Arthrodermataceae/genética , Eritema/microbiología , Femenino , Proteínas Fúngicas/metabolismo , Cobayas , Cabello/microbiología , Folículo Piloso/microbiología , Folículo Piloso/patología , Humanos , Malato Sintasa/genética , Malato Sintasa/metabolismo , Masculino , Recombinasas/metabolismo , Piel/microbiología , Piel/patología , Piel Artificial/microbiologíaRESUMEN
While infected skin wounds are on the increase because of ageing populations, rising incidence of diabetes, and antibiotic resistance, we lack relevant in vivo or in vitro models to study many aspects of bacterial interaction with skin. The aim of this study was to develop three-dimensional models of normal human skin to study bacterial infection. The common dermatological pathogens Staphylococcus aureus and Pseudomonas aeruginosa were used to infect tissue-engineered skin, and the course of infection in the skin was examined over several days. Two forms of model were developed-one in which bacteria were introduced directly to 10 mm wounds in the epidermis, and another in which wounds were created by burning a 4 mm hole in the center of the tissue before inoculation. The bacteria flourished within the engineered skin, and colonized the upper epidermal layers before invasion into the dermis. Infection with P. aeruginosa caused a loss of epidermis and de-keratinization of the skin constructs, as well as partial loss of basement membrane. These novel complex human skin infection models could be used to investigate microbial invasion of normal skin epithelium, basement membrane, and connective tissue, and as a model to study approaches to reduce bacterial burden in skin wounds.
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Fibroblastos/patología , Queratinocitos/patología , Enfermedades Cutáneas Bacterianas/microbiología , Enfermedades Cutáneas Bacterianas/patología , Piel Artificial/microbiología , Piel/lesiones , Piel/microbiología , Ingeniería de Tejidos/métodos , Células Cultivadas , Humanos , Piel/patologíaRESUMEN
BACKGROUND: Large wounds often require temporary allograft placement to optimize the wound bed and prevent infection until permanent closure is feasible. We developed and clinically tested a second-generation living human skin substitute (StrataGraft). StrataGraft provides both a dermis and a fully-stratified, biologically-functional epidermis generated from a pathogen-free, long-lived human keratinocyte progenitor cell line, Neonatal Immortalized KeratinocyteS (NIKS). METHODS: Histology, electron microscopy, quantitative polymerase chain reaction, and bacterial growth in vitro were used to analyze human skin substitutes generated from primary human keratinocytes or NIKS cells. A phase I/II, National Institute of Health-funded, randomized, safety, and dose escalation trial was performed to assess autograft take in 15 patients 2 weeks after coverage with StrataGraft skin substitute or cryopreserved cadaver allograft. RESULTS: StrataGraft skin substitute exhibited a fully stratified epidermis with multilamellar lipid sheets and barrier function as well as robust human beta defensin-3 mRNA levels. Analysis of the primary endpoint in the clinical study revealed no differences in autograft take between wound sites pretreated with StrataGraft skin substitute or cadaver allograft. No StrataGraft-related adverse events or serious adverse events were observed. CONCLUSIONS: The major finding of this phase I/II clinical study is that performance of StrataGraft skin substitute was comparable to cadaver allograft for the temporary management of complex skin defects. StrataGraft skin substitute may also eliminate the risk for disease transmission associated with allograft tissue and offer additional protection to the wound bed through inherent antimicrobial properties. StrataGraft is a pathogen-free human skin substitute that is ideal for the management of severe skin wounds before autografting.
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Trasplante de Piel , Piel Artificial , Traumatismos de los Tejidos Blandos/cirugía , Cicatrización de Heridas/fisiología , Adulto , Cadáver , Desbridamiento , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Piel Artificial/microbiología , StaphylococcusRESUMEN
BACKGROUND: Integra, a dermal replacement template consisting of bovine collagen, chondroitin-6-sulfate, and a silastic sheet is a postexcisional treatment for deep partial to full thickness burns where autograft is limited. This study correlates Integra histology and quantitative microbiology cultures with clinical outcomes after autografting. METHODS: Charts of 29 burn patients who underwent Integra treatment and neodermis biopsy at the time of ultra thin autografting were reviewed. We analyzed microbial contamination, inflammatory reaction, and autograft take. RESULTS: The mean burn size and age were 43% total body surface area and 39 years old, respectively. In quantitative neodermis cultures, 90% of samples had bacterial growth; nine samples (31%) had > 10(5) colony forming units per gram. The most common organism was Staphylococcus aureus (31%). Patients with quantitative bacterial counts >10(5) CFU/g received targeted systemic antibiotics. Integra take (83%) and autograft take (92%) were acceptable even in patients with high bacterial counts (78% Integra take; 86% autograft take). More than 50% of biopsies had dermal regeneration similar to normal dermis; foreign body reactions were unusual. Histologic evidence of inflammation, especially polymorphonuclear cells, was increased in biopsies with high bacterial counts. CONCLUSION: Integra and autograft take can be acceptable even with high bacterial counts if wounds are treated with appropriate targeted topical and systemic antibiotics in the presence of microbial contamination. Neodermis biopsies showed fibrous in-growth congruent with existing Integra fibers with minimal foreign body reaction. These data support Integra use as a safe and effective treatment modality in patients with major burns.
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Materiales Biocompatibles/uso terapéutico , Quemaduras/terapia , Sulfatos de Condroitina , Colágeno , Piel Artificial/microbiología , Infección de la Herida Quirúrgica/microbiología , Administración Tópica , Adulto , Bacitracina/uso terapéutico , Recuento de Colonia Microbiana , Combinación de Medicamentos , Femenino , Humanos , Masculino , Neomicina/uso terapéutico , Polimixina B/uso terapéutico , Estudios Retrospectivos , Trasplante de Piel , Infección de la Herida Quirúrgica/prevención & control , Resultado del TratamientoRESUMEN
AIMS: The aim of this study was to evaluate the respective influence of the physicochemical interactions and the roughness involved in the first part of the biological substrate biocontamination. METHODS AND RESULTS: Therefore we compared the bioadhesion results obtained on the biological model substrate (Episkin) and on a commonly employed inert substrate (AISI 304 stainless steel), frequently used either in dermatology or in development of medical devices. The two studied strains presented different characteristics, both physicochemical and microbiological. Staphylococcus epidermidis, a relatively hydrophobic bacteria capable of exchanging interactions which are principally of the van der Waals type, adhered more to 304 steel than to the surface of reconstituted skin. As for S. aureus, an essentially basic, hydrophilic bacteria, was more adherent to Episkin (a bipolar, hydrophilic substrate) than to stainless steel (a unipolar, basic, hydrophilic substrate). CONCLUSIONS: In the absence of electrostatic interactions, the adhesion of substrate-dependent bacteria to the surface of reconstituted skin was dependent upon the balance between gamma(LW), gamma(+) and gamma(-). SIGNIFICANCE AND IMPACT OF THE STUDY: Consequently, so as to restrict microbial adhesion and reduce adhesive binding between micro-organisms and the surface of the skin, it would be preferable to render this substrate hydrophobic and apolar through the use of appropriate surface treatment.
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Adhesión Bacteriana , Piel Artificial/microbiología , Acero Inoxidable , Staphylococcus/fisiología , Fenómenos Biomecánicos , Contaminación de Equipos , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Modelos Biológicos , Staphylococcus aureus/fisiologíaRESUMEN
Human living skin equivalents (LSEs) offer an alternative to the use of split-thickness autografts for the treatment of hard-to-heal wounds. LSEs consist of 4 active components: a well-differentiated stratum corneum derived from epidermal keratinocytes, dermal fibroblasts, and an extracellular collagen matrix. Neonatal foreskins are used as the source of keratinocytes and dermal fibroblasts for the manufacture of LSEs. Following isolation and expansion in vitro, the cells are cultured on a 3-dimensional scaffold to give an upper epidermal layer and supporting dermal layer. The resulting product has the appearance and handling characteristics of human skin. Safety evaluation of LSEs begins with insuring that foreskins are obtained only from healthy infants whose mothers are negative for a panel of adventitious agents. Keratinocyte and fibroblast cell banks are characterized using morphologic, biochemical, and histologic criteria; checked for the absence of contaminating cell types such as melanocytes, macrophages, lymphocytes, and Langerhans cells; subjected to rigorous microbiological testing (with any production materials of biological origin); and evaluated for in vivo tumorigenicity. The consistency of certain key morphologic and functional characteristics are regularly assessed. Because an LSE represents an allogeneic graft, preclinical safety studies include in vitro and in vivo determinations of its potential immunogenicity. Immunocompromised (SCID) mice reconstituted with human leukocytes or engrafted with human fetal hematolymphoid organs have been useful animal models for assessing possible immunologic responses to LSEs. Additional preclinical studies are being conducted to show that LSEs are noncytotoxic and lack allergenic, sensitizing, or irritation potential.
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Piel Artificial/efectos adversos , Animales , Humanos , Piel Artificial/microbiologíaRESUMEN
The effects of two commercially available econazole formulations (econazole nitrate cream, econazole liposome gel) on uninfected reconstructed human epidermis and on a model of human cutaneous candidosis were investigated. The morphological alterations of the reconstructed epidermis after infection and treatment were analysed with light and electron microscopy. The most important Candida albicans-specific alterations of the recently established in vitro model of human cutaneous candidosis were scaling, hyperkeratosis, parakeratosis, dyskeratosis and spongiosis. A single application of the cream to the uninfected reconstructed epidermis caused more epidermal barrier damage and irritative toxic effects than the liposome gel. Treatment of the modelled human cutaneous candidosis with the cream also resulted in increased toxic effects, e.g., enhancement of scaling with invasion of Candida albicans blastospores into the stratum corneum and intracellular vacuoles. After application of the liposomal preparation invasion of Candida albicans in the stratum corneum could not be detected and toxic effects were reduced. Some of the Candida albicans-specific alterations such as hyperkeratosis, focal thickening of the stratum corneum, dyskeratosis and parakeratosis were completely eliminated. The liposomal formulation increased slightly the morphological alterations of the blastospores. Remnants of the cream formulation could be detected only very rarely in the stratum corneum or the blastospores. The liposomal preparation showed a strong affinity for the Candida albicans cells and the stratum corneum. Intact liposomes could even be observed in the intercellular spaces of the upper stratum corneum. As successful treatment depends on the ability to target the liposomal agent to the wanted site of action, this might be useful for more effective treatment of cutaneous candidosis.