Regulatory effects of bio-intensity electric field on transformation of human skin fibroblasts / 中华烧伤杂志

Objective: To investigate the regulatory effects of bio-intensity electric field on the transformation of human skin fibroblasts (HSFs). Methods: The experimental research methods were used. HSFs were collected and divided into 200 mV/mm electric field group treated with 200 mV/mm electric field for 6 h and simulated electric field group placed in the electric field device without electricity for 6 h. Changes in morphology and arrangement of cells were observed in the living cell workstation; the number of cells at 0 and 6 h of treatment was recorded, and the rate of change in cell number was calculated; the direction of cell movement, movement velocity, and trajectory velocity within 3 h were observed and calculated (the number of samples was 34 in the simulated electric field group and 30 in 200 mV/mm electric field group in the aforementioned experiments); the protein expression of α-smooth muscle actin (α-SMA) in cells after 3 h of treatment was detected by immunofluorescence method (the number of sample was 3). HSFs were collected and divided into simulated electric field group placed in the electric field device without electricity for 3 h, and 100 mV/mm electric field group, 200 mV/mm electric field group, and 400 mV/mm electric field group which were treated with electric fields of corresponding intensities for 3 h. Besides, HSFs were divided into simulated electric field group placed in the electric field device without electricity for 6 h, and electric field treatment 1 h group, electric field treatment 3 h group, and electric field treatment 6 h group treated with 200 mV/mm electric field for corresponding time. The protein expressions of α-SMA and proliferating cell nuclear antigen (PCNA) were detected by Western blotting (the number of sample was 3). Data were statistically analyzed with Mann-Whitney U test, one-way analysis of variance, independent sample t test, and least significant difference test. Results: After 6 h of treatment, compared with that in simulated electric field group, the cells in 200 mV/mm electric field group were elongated in shape and locally adhered; the cells in simulated electric field group were randomly arranged, while the cells in 200 mV/mm electric field group were arranged in a regular longitudinal direction; the change rates in the number of cells in the two groups were similar (P>0.05). Within 3 h of treatment, the cells in 200 mV/mm electric field group had an obvious tendency to move toward the positive electrode, and the cells in simulated electric field group moved around the origin; compared with those in simulated electric field group, the movement velocity and trajectory velocity of the cells in 200 mV/mm electric field group were increased significantly (with Z values of -5.33 and -5.41, respectively, P<0.01), and the directionality was significantly enhanced (Z=-4.39, P<0.01). After 3 h of treatment, the protein expression of α-SMA of cells in 200 mV/mm electric field group was significantly higher than that in simulated electric field group (t=-9.81, P<0.01). After 3 h of treatment, the protein expressions of α-SMA of cells in 100 mV/mm electric field group, 200 mV/mm electric field group, and 400 mV/mm electric field group were 1.195±0.057, 1.606±0.041, and 1.616±0.039, respectively, which were significantly more than 0.649±0.028 in simulated electric field group (P<0.01). Compared with that in 100 mV/mm electric field group, the protein expressions of α-SMA of cells in 200 mV/mm electric field group and 400 mV/mm electric field group were significantly increased (P<0.01). The protein expressions of α-SMA of cells in electric field treatment 1 h group, electric field treatment 3 h group, and electric field treatment 6 h group were 0.730±0.032, 1.561±0.031, and 1.553±0.045, respectively, significantly more than 0.464±0.020 in simulated electric field group (P<0.01). Compared with that in electric field treatment 1 h group, the protein expressions of α-SMA in electric field treatment 3 h group and electric field treatment 6 h group were significantly increased (P<0.01). After 3 h of treatment, compared with that in simulated electric field group, the protein expressions of PCNA of cells in 100 mV/mm electric field group, 200 mV/mm electric field group, and 400 mV/mm electric field group were significantly decreased (P<0.05 or P<0.01); compared with that in 100 mV/mm electric field group, the protein expressions of PCNA of cells in 200 mV/mm electric field group and 400 mV/mm electric field group were significantly decreased (P<0.05 or P<0.01); compared with that in 200 mV/mm electric field group, the protein expression of PCNA of cells in 400 mV/mm electric field group was significantly decreased (P<0.01). Compared with that in simulated electric field group, the protein expressions of PCNA of cells in electric field treatment 1 h group, electric field treatment 3 h group, and electric field treatment 6 h group were significantly decreased (P<0.01); compared with that in electric field treatment 1 h group, the protein expressions of PCNA of cells in electric field treatment 3 h group and electric field treatment 6 h group were significantly decreased (P<0.05 or P<0.01); compared with that in electric field treatment 3 h group, the protein expression of PCNA of cells in electric field treatment 6 h group was significantly decreased (P<0.01). Conclusions: The bio-intensity electric field can induce the migration of HSFs and promote the transformation of fibroblasts to myofibroblasts, and the transformation displays certain dependence on the time and intensity of electric field.

Evaluation of dermal collagen stained with picrosirius red and examined under polarized light microscopy

Abstract: The special picrosirius red staining highlights the natural birefringence of collagen fibers when exposed to polarized light. The results from birefringence allow to evaluate the organization of the collagen fibers in the tissues. The authors intend to elucidate all steps to obtain and capture images of histological sections stained with picrosirius red and evaluated under polarized light microscopy, as well as possible artefacts that may occur.

Métodos avançados para tratamento de queimaduras: uma revisão / Advanced methods for the treatment of burns: a review / Métodos avanzados para el tratamiento de quemaduras: una revisión

Introdução: As lesões cutâneas por queimadura são consideradas uma das causas frequentes de mortalidade e grave incapacidade em longo prazo. Visando a reabilitação desses indivíduos, materiais biocompatíveis destinam-se a mimetizar a matriz extracelular em um microambiente para o crescimento de células in vitro. Objetivo: Apresentar metodologias desenvolvidas no tratamento de lesões de pele. Método: Trata-se de uma revisão narrativa na qual o levantamento bibliográfico deu-se por fontes de evidência primária e secundária, tais como: Biblioteca Nacional de Medicina dos Estados Unidos da América (MEDLINE) via PubMed e SciVerse Scopus. Como complementaridade na pesquisa, livros e endereço eletrônico de associações médicas-científicas e governamentais correspondentes a publicações dos últimos cinco anos foram utilizados. Resultados: Verificaram-se diferentes possibilidades para o tratamento de lesões de pele. Dentre as quais, o desenvolvimento de "Materiais Inteligentes" e sua interação com o tecido biológico numa leitura simultânea de biomarcadores capazes de replicar funções de órgãos seria auspicioso, em um sistema de cultura dinâmico e tecnologia futurista. Conclusão: Técnicas promissoras avançam no desenvolvimento de substitutos de pele funcionais em sua organização multiescalar

Introduction: Cutaneous burn injuries are considered one of the frequent causes of mortality and severe long-term disability. Aiming at the rehabilitation of these individuals, biocompatible materials are intended to mimic the extracellular matrix through a microenvironment for cell growth in vitro. Objective: To present methodologies developed in the treatment of skin lesions. Methods: This is a narrative review which was based on primary and secondary evidence sources, such as: National Library of Medicine of the United States of America (MEDLINE) through PubMed and SciVerse Scopus. As a complement to the research, books and electronic address of medical-scientific and governmental associations corresponding to publications of the last five years were used. Results: There were different possibilities for the treatment of skin lesions. Among them, the development of "Intelligent Materials" and their interaction with biological tissue in a simultaneous reading of biomarkers capable of replicating organ functions would be auspicious, in a dynamic culture system and futuristic technology. Conclusion: Promising techniques advance towards the development of functional skin substitutes in their multiscalar organization

Introducción: Las lesiones cutáneas resueltas de quemaduras son consideradas una de las causas frecuentes de mortalidad y grave incapacidad a largo plazo. Visando la rehabilitación clínica de estos individuos, los materiales biocompatibles se destinan a mimetizar la matriz extracelular a través de un microambiente para el crecimiento de células in vitro. Objetivo: Exponer metodologías desarrolladas en el tratamiento de lesiones de piel. Método: Se trata de una revisión narrativa donde el levantamiento bibliográfico se dio por fuentes de evidencia primaria y secundaria, tales como: la Biblioteca Nacional de Medicina de los Estados Unidos de América (MEDLINE) vía PubMed y SciVerse Scopus. Como complementariedad en la investigación, libros y los sitios en internet de asociaciones médicas-científicas y gubernamentales correspondientes a publicaciones de los últimos cinco años se utilizaron. Resultados: Se verificaron diferentes posibilidades para el tratamiento de lesiones de piel. Entre las cuales, el desarrollo de "Materiales Inteligentes" y su interacción con el tejido biológico en una lectura simultánea de biomarcadores capaces de replicar funciones de órganos sería auspicioso, en un sistema de cultura dinámica y tecnología futurista. Conclusión: Técnicas prometedoras avanzan en el desarrollo de sustitutos de piel funcionales en su organización multiescalar.

Aldefluor protocol to sort keratinocytes stem cells from skin

Abstract Purpose: To investigate the use Aldefluor® and N, N - Dimethylaminobenzaldehyde (DEAB) to design a protocol to sort keratinocyte stem cells from cultured keratinocytes from burned patients. Methods: Activated Aldefluor® aliquots were prepared and maintained at temperature between 2 to 8°C, or stored at -20°C. Next, the cells were collected following the standard protocol of sample preparation. Results: Best results were obtained with Aldefluor® 1.5µl and DEAB 15 µl for 1 x 106 cells, incubated at 37°C for 15 minutes. Flow cytometer range for keratinocyte stem cells separation was evaluated. There were 14.8% of stem cells separated in one sample of keratinocyte culture used to pattern the protocol. After being defined the ideal concentration, the same test pattern was performed in other keratinocyte samples. We observed a final mean of 10.8%. Conclusion: Aldefluor® has been shown as a favorable marking of epidermal keratinocyte stem cells for subsequent separation on a flow cytometer, with detection of 10.8% of epidermal keratinocyte stem cells, in this protocol.

Keratinocyte growth factor and the expression of wound-healing-related genes in primary human keratinocytes from burn patients

ABSTRACT PURPOSE: To evaluate the effect of keratinocyte growth factor (KGF) treatment on the expression of wound-healing-related genes in cultured keratinocytes from burn patients. METHODS: Keratinocytes were cultured and divided into 4 groups (n=4 in each group): TKB (KGF-treated keratinocytes from burn patients), UKB (untreated keratinocytes from burn patients), TKC (KGF-treated keratinocytes from controls), and UKC (untreated keratinocytes from controls). Gene expression analysis using quantitative polymerase chain reaction (qPCR) array was performed to compare (1) TKC versus UKC, (2) UKB versus UKC, (3) TKB versus UKC, (4) TKB versus UKB, (5) TKB versus TKC, and (6) UKB versus TKC. RESULTS: Comparison 1 showed one down-regulated and one up-regulated gene; comparisons 2 and 3 resulted in the same five down-regulated genes; comparison 4 had no significant difference in relative gene expression; comparison 5 showed 26 down-regulated and 7 up-regulated genes; and comparison 6 showed 25 down-regulated and 11 up-regulated genes. CONCLUSION: There was no differential expression of wound-healing-related genes in cultured primary keratinocytes from burn patients treated with keratinocyte growth factor.

Platelet-Rich Fibrin Lysate Can Ameliorate Dysfunction of Chronically UVA-Irradiated Human Dermal Fibroblasts

To determine whether platelet-rich fibrin lysate (PRF-L) could restore the function of chronically ultraviolet-A (UVA)-irradiated human dermal fibroblasts (HDFs), we isolated and sub-cultured HDFs from six different human foreskins. HDFs were divided into two groups: those that received chronic UVA irradiation (total dosages of 10 J cm-2) and those that were not irradiated. We compared the proliferation rates, collagen deposition, and migration rates between the groups and between chronically UVA-irradiated HDFs in control and PRF-L-treated media. Our experiment showed that chronic UVA irradiation significantly decreased (p<0.05) the proliferation rates, migration rates, and collagen deposition of HDFs, compared to controls. Compared to control media, chronically UVA-irradiated HDFs in 50% PRF-L had significantly increased proliferation rates, migration rates, and collagen deposition (p<0.05), and the migration rates and collagen deposition of chronically UVA-irradiated HDFs in 50% PRF-L were equal to those of normal fibroblasts. Based on this experiment, we concluded that PRF-L is a good candidate material for treating UVA-induced photoaging of skin, although the best method for its clinical application remains to be determined.

Prolonged viability of human organotypic skin explant in culture method (hOSEC)

Abstract BACKGROUND: Currently, the cosmetic industry is overwhelmed in keeping up with the safety assessment of the increasing number of new products entering the market. To meet such demand, research centers have explored alternative methods to animal testing and also the large number of volunteers necessary for preclinical and clinical tests. OBJECTIVES: This work describes the human skin ex-vivo model (hOSEC: Human Organotypic Skin Explant Culture) as an alternative to test the effectiveness of cosmetics and demonstrate its viability through cutaneous keratinocytes' proliferative capacity up to 75 days in culture. METHODS: The skin explants obtained from surgeries were cultured in CO2-humid incubator. After 1, 7, 30 and 75 days in culture, skin fragments were harvested for analysis with histomorphological exam (HE staining) on all days of follow-up and immunohistochemistry for Ck5/6, Ck10 and Ki-67 only on the 75th day. RESULTS: On the 7th day, the epidermis was perfect in the dermoepidermal junction, showing the viability of the model. On the 30th day, the epidermis was thicker, with fewer layers on the stratum corneum, although the cutaneous structure was unaltered. On the 75th day, the skin became thinner but the dermoepidermal junctions were preserved and epidermal proliferation was maintained. After the 75th day on culture, the skin was similar to normal skin, expressing keratinocytes with Ck5/6 on supra-basal layers; Ck10 on differentiated layers; and viability could be assessed by the positivity of basal cells by Ki-67. CONCLUSION: The hOSEC model seems a good alternative to animal testing; it can be used as a preclinical test analogous to clinical human skin test with similar effectiveness and viability proven by immunohistological analyses. .

Characterization of victims of aggression and transportation accidents treated at the Forensic Medicine and Dentistry Institute - Campina Grande, Paraíba, Brazil - 2010 / Caracterização de vítimas de agressão e de acidentes de transporte atendidas no Instituto de Medicina e Odontologia Forense - Campina Grande, Paraíba, Brasil - 2010

The objective of this cross-sectional census study was to characterize agression and land-based transport accidents in a city in the Northeast of Brazil. Data was analyzed from live victims who were treated at a forensic service (N = 2.379). In the descriptive analysis, the majority of events were represented by aggression (71.6%); which occurred on weekdays (65%), with 35.1% at night. Trauma occurred to the whole body (63.6%) and to soft tissue (74.2%). On the basis of multiple correspondence analysis, two dimensions were formed: the first dimension (internal reliability = 0.654) was formed by the cause of the event, the trauma and the age group and the second dimension (reliability = 0.514), by age group, occupation and civil status. Three groups with distinct profiles were formed for accidents and aggression: young women who suffered aggression, with trauma to the face and soft tissues during the evening and at weekends; adult men who suffered car accidents, in the morning and on work days; and retired elderly widowers, who were run over.

O objetivo deste estudo tranversal censitário foi caracterizar a agressão e os acidentes de transporte terrestre em uma cidade do nordeste do Brasil. Foram analisados os dados de vítimas vivas que foram atendidas em um serviço forense (N = 2.379). Na análise descritiva, a maioria dos eventos foi a agressão (71,6%); que ocorreu nos dias úteis (65%), sendo 35,1% no período noturno. Os traumas ocorreram no corpo todo (63,6%) e envolveram o tecido mole (74,2%). A partir da análise de correspondência múltipla formaram-se duas dimensões: a primeira dimensão (confiabilidade interna = 0,654) foi formada pela causa do evento, o trauma e a faixa etária e, a segunda dimensão (confiabilidade interna = 0,514), pela faixa etária, a ocupação e o estado civil. Formaram-se três grupos com perfis distintos para os acidentes e agressão. Mulheres jovens que sofreram agressões com traumas faciais, em tecidos moles, no período da tarde e durante os finais de semana. Homens, adultos que sofreram acidentes automobilísticos, pela manhã e em dias úteis, e idosos, viú vos, aposentados e que sofreram atropelamento. Há um elevado número de vítimas de agressão interpessoal, seguido por acidentes de moto e os tipos de acidentes estão associados a grupos populacionais.

Medicamentos esenciales y Acuerdo sobre los ADPIC: colisión entre el derecho a la salud y el derecho de propiedad intelectual / Essential medicines and the TRIPS Agreement: collision between the right to health and intellectual property rights

El fortalecimiento mundial de la patente farmacéutica tensiona el acceso a los medicamentos esenciales. De acuerdo a este trabajo ello ha derivado en la colisión del derecho de propiedad intelectual que impulsa el Acuerdo sobre los Aspectos de los Derechos de Propiedad Intelectual relacionados con el Comercio (ADPIC) de la Organización Mundial del Comercio (OMC), con el derecho a la salud previsto en el Pacto Internacional de Derechos Económicos, Sociales y Culturales (PIDESC). Diversas controversias ventiladas en la OMC ilustran el enfrentamiento entre países con una poderosa industria farmacéutica y los intereses de países en desarrollo. Se concluye que las normas ADPIC-plus suscritas en tratados de libre comercio por países en desarrollo, que confieren al titular de la patente farmacéutica más derechos que los previstos en el propio Acuerdo sobre los ADPIC, son incompatibles con las obligaciones que asumen respecto del acceso a medicamentos esenciales en el marco del derecho a la salud del PIDESC.

The strengthening of pharmaceutical patent protection globally puts strains on access to essential medicines. According to the present paper, this process has led to the collision of the intellectual property rights adopted in the World Trade Organization (WTO) Trade-Related Aspects of Intellectual Property Rights (TRIPS) Agreement and the right to health stated in the International Covenant on Economic, Social and Cultural Rights (ICESCR). Several controversies disputed in the WTO illustrate the confrontation between countries with a powerful pharmaceutical industry and the interests of developing countries. It is concluded that the TRIPS-plus rules subscribed to by developing countries in free trade agreements which give the pharmaceutical patent holder more rights than those stipulated in the original TRIPS Agreement are incompatible with the obligations to provide access to essential medicines under the right to health of the ICESCR.

Detección de anticuerpos circulantes en biopsia de tejido para el diagnóstico de dermatitis herpetiforme / Detection of circulating antibodies in tissue biopsy for the diagnosis of dermatitis herpetiformis

Introducción: la dermatitis herpertiforme (DH), es una enfermedad crónica inflamatoria de la piel. Se caracteriza por la presencia de lesiones pruriginosas de diversas formas y de hallazgos histopatológicos típicos. Se ha considerado como la expresión cutánea de la enteropatía sensible al gluten (ESG), indistinguible de la enfermedad celiaca. Es una entidad con baja prevalencia y se presenta con más frecuencia en la tercera década de la vida. Afecta principalmente a individuos caucásicos. El diagnóstico de realiza a través de la detección de un depósito granular de Inmunoglobulina A (IgA) en la unión dermoepidérmica, a través de un estudio de inmunofluorescencia directa (IFD) de una muestra de piel sana perilesional. Estos depósitos con frecuencia se localizan en las puntas de las papilas dérmicas. La negatividad del estudio de inmunofluorescencia directa debe hacer dudar del diagnóstico. El tratamiento consiste en mantener una dieta estricta libre de gluten, el manejo de las lesiones y la prescripción de dapsona para el manejo de las erupciones cutáneas. Objetivo: evaluar la utilidad de la detección de anticuerpos circulantes en biopsia de tejido para el diagnóstico de la dermatitis herpetiforme. \r\nResultados: se identificaron 174 publicaciones. Con los resultados obtenidos, no fue posible identificar revisiones sistemáticas de la literatura ni estudios de validez diagnóstica de la IFD. Se hizo una preselección de 18 estudios observacionales descriptivos. Fueron incluidos nueve series de casos. Se presentan los datos descriptivos sobre la positividad de la IFD para el diagnóstico de la DH. Conclusiones: \r\nactualmente se considera que la IFD es el patrón de oro para el diagnóstico de la DH.(AU)

Stem cells in dermatology

Preclinical and clinical research have shown that stem cell therapy could be a promising therapeutic option for many diseases in which current medical treatments do not achieve satisfying results or cure. This article describes stem cells sources and their therapeutic applications in dermatology today.

Touch DNA of shed skin cells from the deployed airbag to address drunken driving crimes / 法医学杂志

In the criminal cases of driving under the influence (DUI), DNA evidence can be collected from the deployed airbag of the motor vehicle and submitted to the crime lab for touch DNA analysis. The evidence can be acquired when the skin cells are observed on the surface of the airbag in a traffic accident. However, the low quantity or quality of the evidence collected from a crime scene prevents further identification analysis in many cases. In the current study, we reported a case of identifying touch DNA extraction from the shed skin cells from the deployed airbag of a motor vehicle. We managed to collect DNA evidence from the shed skin cells in an airbag using a proper approach of collection and extraction. The 5.87 ng of extracted DNA was sufficient for genotyping and forensic identification, which helped to identify the driver of the car in collision with a pier in the street. In DUI cases and other traffic accidents, therefore, the amount of touch DNA extracted from the deployed airbag can be sufficient for DNA marker genotyping and further analysis.

Human beta defensin-4 and keratinocyte growth factor gene expression in cultured keratinocyte and fibroblasts of burned patients

PURPOSE: To evaluate KGF and human beta defensin-4 (HBD-4) levels produced by dermic fibroblasts and keratinocytes cultivated from burned patients' skin samples. METHODS: Keratinocytes and fibroblasts of 10 patients (four major burns, four minor burns and two controls) were primarily cultivated according to standard methods. HBD-4 and KGF genes were analyzed by quantitative PCR. RESULTS: In fibroblasts, KGF gene expression was 220±80 and 33.33±6.67 (M±SD; N=4), respectively for major and minor burn groups. In keratinocytes, KGF gene expression was 11.2±1.9 and 3.45±0.37 (M±SD; N=4), respectively for major and minor burn groups. In fibroblasts, HBD-4 gene expression was 15.0±4.0 and 11.5±0.5 (M±SD; N=4), respectively for major and minor burn. In keratinocyte, HBD-4 gene expression was 0.0±0.0 and 13.4±4.8 (M±SD; N=4), respectively for major and minor burn. CONCLUSIONS: KGF expression was increased in burn patient fibroblasts compared to control group. In keratinocytes culture, KGF suppression is inversely proportional to burn extension; it is active and increased in major burn but decreased in minor burn. HBD-4 expression was increased in fibroblasts and decreased in keratinocytes from all burned patients. .

TGF-β1 on induced osteogenic differentiation of human dermal fibroblast

PURPOSE: To evaluate the role of transforming growth factor beta 1 (TGF-β1) on the induced osteogenic differentiation of human dermal fibroblasts. METHODS: We performed four groups with cultured dermal fibroblasts according to the culture medium: CONTROL (DMEM culture medium); TGF-β1 (DMEM culture medium with 10 ng/ml of TGF-β1); OSTEOG (DMEM culture medium with 0.5 µg/ml of ascorbic acid, 10 mmol/l of β-glycerophosphate and 10 nmol/L of dexamethasone); and OSTEOG/TGF-β1 (osteogenic medium with 10 ng/ml of TGF-β1). Alkaline phosphatase (ALP) activity and the amount of osteocalcin (OC) in the supernatant, as well as the capability to form calcium phosphate deposits, were analysed for 28 days RESULTS: There were significant differences (p<0.05) between CONTROL and TGF-β1 groups in comparison with OSTEOG and OSTEOG/TGF-β1 groups in the ALP activity and OC amount. Although, both osteogenic groups had the same behavior with regard the expression curve during the experimental time, the OSTEOG/TGF-β1 group achieved significantly higher ALP and OC levels and showed no significant difference in the levels of mineralized deposits and in comparison with the levels found in the OSTEOG group. CONCLUSION: The addition of transforming growth factor beta 1 to the osteogenic culture medium increased the activity of alkaline phosphatase and the amount of osteocalcin, but TGF-β1 did not alter the presence of mineralized calcium phosphate deposits. .

Keratinocyte growth factor, tumor necrosis factor-alpha and interleukin-1 beta gene expression in cultured fibroblasts and keratinocytes from burned patients

PURPOSE: To evaluate the gene expression of KGF, TNF-alpha and IL-1 beta in skin fibroblasts and keratinocytes cultured from burned patients. METHODS: Three patients with large burns and three patients with small burns, as well as two controls, were included. The cell culture was initiated by the enzymatic method. After extraction and purification of mRNA, qPCR was used to assess the gene expression of KGF, TNF-alpha and IL-1 beta. RESULTS: The expression of KGF was increased on average 220-fold in large burns and 33.33-fold in small burns in fibroblasts, and 11.2-fold in large burns and 3.45-fold in small burns in keratinocytes compared to healthy patients (p<0.05). Expression of TNF-alpha was not observed. IL-1 beta is down-regulated in fibroblasts of burned patients, and much more repressed in small burns (687-fold, p<0.05). In keratinocytes, the repression of IL-1 beta expression occurs in patients with small burns (28-fold), while patients with large burns express this gene intensively (15-fold). CONCLUSIONS: The study showed a quantitative pattern in the expression of KGF gene, which is more expressed according to the size of the burn. TNF-alpha was not expressed. A qualitative pattern in the expression of IL-1 beta gene was demonstrated.

Ocorre alta incidência de células da pele na primeira e terceira gotas do líquido cefalospinal em raquianestesia / There is high incidence of skin cells in the first and third drops of cerebrospinal fluid in spinal anesthesia / Ocurre una alta incidencia de células de la piel en la primera y tercera gotas del líquido cefaloespinal en la raquianestesia

JUSTIFICATIVA E OBJETIVOS: Fragmentos de pele durante punções subaracnóideas podem desenvolver tumores epidermoides intraespinais. O objetivo deste estudo foi verificar a incidência de células epiteliais que refluem junto com a primeira e terceira gotas de líquor de pacientes submetido a raquianestesia. MÉTODO: Foram obtidas amostras da primeira e terceira gotas de líquor em 39 pacientes adultos submetidos à raquianestesia com agulha 25G Quincke, sendo confeccionadas quatro lâminas: da primeira gota, da terceira gota, da agulha e uma quarta lâmina controle com uma gota de soro fisiológico. As lâminas foram examinadas de forma randomizada pelo patologista. RESULTADOS: Foram identificadas células epiteliais escamosas em 35 (89,7%) das amostras da primeira gota, em 34 (87,2%) da terceira gota e em 24 (61,5%) das agulhas espinais. A terceira gota apresentou em média maior número de células que a primeira gota (p = 0,046). Células epiteliais nucleadas foram encontradas em uma (2,56%) das amostras da primeira gota, em quatro (10,25%) da terceira gota e em uma (2,56%) das agulhas espinais. A terceira gota apresentou em média maior número de células nucleadas que a primeira gota sem diferença estatística (p = 0,257). CONCLUSÕES: Encontramos uma alta porcentagem de células epiteliais que refluem na primeira (89,7%) e na terceira (87,2%) gotas do líquor e nas agulhas utilizadas (61,5%). Mesmo utilizando agulhas de pequeno calibre, descartáveis e com mandril bem adaptado, foram encontradas células da pele.

BACKGROUND AND OBJECTIVES: Skin fragments during lumbar punctures may develop intraspinal epidermoid tumors. The aim of this study was to determine the incidence of epithelial cells that reflow along with the first and third drops of CSF of patients undergoing spinal anesthesia. METHODS: Samples of the first and third drops of cerebrospinal fluid were collected from 39 adult patients undergoing spinal anesthesia with a 25G Quincke needle. Four microscope slides were prepared: one for the first drop, one for third drop, one for the needle, and one with a drop of saline for control. A pathologist examined the slides randomly. RESULTS: Squamous epithelial cells were identified in 35 (89.7%) samples from the first drop, 34 (87.2%) from the third drop, and 24 (61.5%) from spinal needle. The third drop showed a mean number of cells larger than the first drop (p = 0.046). Nucleated epithelial cells were found in a sample of the first drop (2.56%), in four samples of third drop (10.25%), and in one spinal needle (2.56%). Third drop showed a mean number of nucleated cells higher than first drop with no statistical difference (p = 0.257). CONCLUSIONS: High percentage of epithelial cells was found in the first (89.7%) and third (87.2%) drops of CSF reflow and in used needles (61.5%). Skin cells were found even using small gauge disposable needles with well-adapted mandrel,.

JUSTIFICATIVA E OBJETIVOS: Algunos fragmentos de piel durante las punciones subaracnoideas pueden desarrollar tumores epidermoides intraespinales. El objetivo de este estudio fue verificar la incidencia de células epiteliales que refluyen junto con la primera y tercera gotas de líquido cefalorraquídeo de los pacientes sometidos a la raquianestesia. MÉTODO: Se obtuvieron muestras de la primera y tercera gotas de líquido cefalorraquídeo en 39 pacientes adultos sometidos a la raquianestesia con una aguja 25G Quincke, siendo confeccionadas cuatro láminas: de la primera gota, de la tercera gota, de la aguja y una cuarta lámina control con una gota de suero fisiológico. Las láminas fueron examinadas de forma aleatoria por el patólogo. RESULTADOS: Se identificaron células epiteliales escamosas en 35 (89,7%) de las muestras de la primera gota, en 34 (87,2%) de la tercera gota y en 24 (61,5%) de las agujas espinales. La tercera gota tuvo como promedio un mayor número de células que la primera gota (p = 0,046). Las células epiteliales nucleadas fueron encontradas en una (2,56%) de las muestras de la primera gota, en cuatro (10,25%) de la tercera gota y en una (2,56%) de las agujas espinales. La tercera gota presentó como promedio, un mayor número de células nucleadas que la primera gota sin diferencias estadísticas (p = 0,257). CONCLUSIONES: Encontramos un alto porcentaje de células epiteliales que refluyen en la primera (89,7%) y en la tercera (87,2%) gotas del líquido cefalorraquídeo y en las agujas utilizadas (61,5%). Y aunque hayamos usado agujas de pequeño calibre, desechables y con un mandril bien adaptado, se encontraron células de la piel.

Influência do envelhecimento na qualidade da pele de mulheres brancas: o papel do colágeno, da densidade de material elástico e da vascularização / Influence of aging on the skin quality of white-skinned women: the role of collagen, elastic material density, and vascularization

INTRODUÇÃO: O objetivo deste estudo é avaliar a influência do envelhecimento na qualidade da pele de mulheres brancas, analisando o colágeno, as fibras elásticas e a vascularização. MÉTODO: Foi realizada análise histológica e morfométrica de 218 retalhos pré-auriculares de mulheres brancas, que se submeteram a cirurgia estética facial. Foram utilizados o imunomarcador AntiCD 34, que evidencia os vasos sanguíneos, a coloração de Weigert, para visibilização das fibras elásticas, e a coloração de Picrosirius Ultrared, para analisar e quantificar os colágenos I, III e total. Os dados foram analisados de acordo com a faixa etárias das doadoras: < 40 anos, 40 anos a 49 anos, 50 anos a 59 anos, 60 anos a 69 anos, e > 70 anos. RESULTADOS: Foi observada fragmentação e desorganização das fibras de colágeno, especialmente acima de 60 anos. Não houve diferenças significantes entre a idade e a espessura da derme e da epiderme, porém foi identificada relação com as porcentagens de colágeno I, III e total (P < 0,001). Houve aumento da densidade de fibras elásticas com a progressão da idade (P < 0,001). Comparando-se as peles das pacientes de faixas etárias vizinhas, com diferença de uma década entre elas, não houve diferença significativa na quantidade de material elástico dessas peles; porém, ao se comparar aquelas com diferença de 2 ou mais décadas nas faixas etárias, o aumento foi significante (P < 0,05). A diferença do número de vasos não foi significante (P = 0,112). CONCLUSÕES: O envelhecimento promoveu redução do colágeno, degradação e fragmentação das fibras, e aumento da densidade de material elástico desorganizado, e não influenciou no número de vasos sanguíneos da derme.

BACKGROUND: In the present study, we aimed to evaluate the influence of aging on the skin quality of white-skinned women by assessing collagen levels, elastic material density, and vascularization. METHODS: Histological and morphometric analyses were performed on 218 preauricular skin fragments from white-skinned women who underwent facial cosmetic surgery. Anti-CD34 was used to identify the blood vessels, Weigert's staining was used to visualize elastic fibers, and Picro-sirius Ultra Red staining was employed for analyzing and quantifying the expression of type I, III, and total collagen. Data were analyzed according to the following donor age groups: < 40, 40-49, 50-59, 60-69, and > 70 years. RESULTS: Fragmentation and disorganization of collagen fibers were observed in certain samples, particularly in samples from patients aged > 60 years. Significant differences between age and the thickness of the dermis and epidermis were not detected. However, a relationship was identified between age and the percentages of type I, III, and total collagen, and an increase of elastic fibers density was associated with age progression (P < 0.001). The comparison of the skin of patients with a decade difference in age did not reveal a significant difference in the elastic material quality; however, when the age difference was of 2 decades or more, there was a significant difference in elastic fibers (P < 0.05). The difference in the number of blood vessels between the groups was not significant (P = 0.112). CONCLUSIONS: Aging promoted collagen reduction, fiber degradation and fragmentation, and increased disorganized elastic material density; however, it did not affect the number of dermal blood vessels.

Pluripotent lineage of CD133 stem cells isolated from human skin samples.

Skin stem cells are very important in cosmetics, pharmacological and regenerative medicine and burn cases. Foreskin samples surgically removed after circumcision from boys below 7 years were collected and primary epidermal cells were prepared by enzymatic and mechanical tituration method. Selecting CD133 (prominin-1) multipotent stem cell marker, enriched stem cells were analyzed by MACS using CD133 antibodies conjugated with magnetic beads. CD133 positive and negative cells with specific skin stem cells markers like - CD34 (Universal stem cells marker), CD29 (integrin beta-1) and CD49f (integrin alpha-6) immunophenotypes were screened and sorted in flowcytometer. Further the expression of four embryonic genes or transcription factors of pluripotent stem cells were analyzed for pluripotent character of sorted cells. It was found that skin stem cell markers associated with CD133 cells, differentially expressed CD34, CD29 and CD49f immunophenotyes on both positive and negative CD133 cells in FACS analysis. The embryonic stem cell markers (induced pluripotent stem cell markers) like Oct4, SOX2, Notch-2 and K19 genes were expressed in CD133 positive epidermal cells. It is therefore evident that foreskin derived epidermal stem cells showed pluripotent or multipotent nature. This finding opens up avenues for new uses of these stem cells for direct cell seeding in wound healing, surgical suturing and drug screening.

Construção de substituto da pele composto por matriz de colágeno porcino povoada por fibroblastos dérmicos e queratinócitos humanos: avaliação histológica / Construction of a skin substitute composed of porcine collagen matrix populated with human dermal fibroblasts and keratinocytes: histological evaluation", "_e": "n

INTRODUÇÃO: O uso de enxertos autólogos é limitado pela extensão da área doadora e pelo estado clínico dos pacientes, no caso de lesões extensas. Alotransplantes coletados de cadáveres ou voluntários são rejeitados após uma ou duas semanas, servindo apenas como cobertura temporária para essas lesões. O tratamento de grandes lesões cutâneas com tegumento autólogo reconstruído constitui alternativa atraente, já que, a partir de um pequeno fragmento de pele do paciente, pode-se obter culturas de células que se multiplicam rapidamente e podem ser criopreservadas, permitindo, assim, sua utilização em novos tratamentos por tempo indeterminado. Este estudo pretendeu avaliar o comportamento histológico de queratinócitos e fibroblastos humanos cultivados sobre uma matriz de colágeno porcino derivada da submucosa intestinal. MÉTODO: Células da epiderme e derme humana foram cultivadas separadamente e semeadas sobre matriz de colágeno porcino, onde permaneceram em ambiente controlado por 21 dias, antes de serem submetidas a análise histológica. RESULTADOS: Observou-se que os fibroblastos invadem e colonizam a matriz de colágeno, enquanto os queratinócitos se organizam de forma laminar e estratificada sobre a superfície em que foram semeados. CONCLUSÕES: A utilização da matriz de colágeno porcino como carreador de células da pele humana é possível e a organização dessas células se assemelha à arquitetura da pele humana.

BACKGROUND: In the case of extensive lesions, the use of autologous grafts is limited by the extent of the donor area and the clinical condition of patients. Allografts collected from cadavers or volunteers are usually rejected after 1 to 2 weeks, thus serving only as temporary cover for these lesions. Treating major cutaneous lesions with reconstructed autologous skin is an attractive alternative, because it is possible to obtain cultures of cells that multiply rapidly and can be cryopreserved from a small fragment of the patient's skin, thereby facilitating its indefinite use in new treatments. This study evaluated the histological behavior of cultured human keratinocytes and fibroblasts on a collagen matrix derived from porcine small intestinal submucosa. METHODS: Cells from human epidermis and dermis were grown separately and seeded on porcine collagen matrix, which was maintained in a controlled environment for 21 days before being subjected to histological analysis. RESULTS: Fibroblasts invaded and colonized the collagen matrix, whereas keratinocytes were organized in laminated and stratified layers on the surface on which they were seeded. CONCLUSIONS: The use of porcine collagen matrix as a support for human skin cells is feasible, and the organization of these cells resembles the architecture of human skin.

Stem cells in embryonic skin development

The skin is a complex stratified organ which acts not only as a permeability barrier and defense against external agents, but also has essential thermoregulatory, sensory and metabolic functions. Due to its high versatility and activity, the skin undergoes continuous self-renewal to repair damaged tissue and replace old cells. Consequently, the skin is a reservoir for adult stem cells of different embryonic origins. Skin stem cell populations reside in the adult hair follicle, sebaceous gland, dermis and epidermis. However, the origin of most of the stem cell populations found in the adult epidermis is still unknown. Far more unknown is the embryonic origin of other stem cells that populate the other layers of this tissue. In this review we attempt to clarify the emergence, structure, markers and embryonic development of diverse populations of stem cells from the epidermis, dermis and related appendages such as the sebaceous gland and hair follicle.