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.

A New Genus and Species of a Minute Suckermouth Armored Catfish (Siluriformes: Loricariidae) from the Rio Tocantins Drainage, Central Brazil: The Smallest Known Loricariid Catfish

Nannoplecostomus eleonorae, a new genus and species of a miniature suckermouth armored catfish, is described based on specimens collected from the karst region of Sao Domingos, upper Rio Tocantins basin, Goias State, central Brazil. The new genus and species can be diagnosed among loricariids by presenting a unique reductive pattern of lateral dermal plates, with most of the body covered by only three series of plates (viz., dorsal, mid-ventral, and ventral). Based on the available published phylogenetic studies for the family, we provisionally consider Nannoplecostomus eleonorae as being an incertae sedis taxon within Loricariidae. Achieving a maximum standard length of 22.2 mm SL, Nannoplecostomus eleonorae is the smallest known loricariid catfish, and a list of the remaining smallest loricariids is provided.

Ciprofloxacin has antifibrotic effects in scleroderma fibroblasts via downregulation of Dnmt1 and upregulation of Fli1

Systemic sclerosis (SSc) is characterized by fibrosis of the skin and internal organs. The present study was undertaken to examine the effects of ciprofloxacin, a fluoroquinolone antibiotic implicated in matrix remodeling, on dermal and lung fibroblasts obtained from SSc patients. Dermal and lung fibroblasts from SSc patients and healthy subjects were treated with ciprofloxacin. Western blotting was used to analyze protein levels and RT-PCR was used to measure in RNA expression. The pharmacologic inhibitor UO126 was used to block Erk1/2 signaling. SSc dermal fibroblasts demonstrated a significant decrease in collagen type I mRNA and protein levels after antibiotic treatment, while healthy dermal fibroblasts were less sensitive to ciprofloxacin, downregulating collagen only at the protein levels. Connective tissue growth factor (CCN2) gene expression was significantly reduced and matrix metalloproteinase (MMPI) levels were enhanced after ciprofloxacin treatment to a similar extent in healthy and SSc fibroblasts. Ciprofloxacin induced Erk1/2 phosphorylation, and Erk1/2 blockade completely prevented MMP1 upregulation. However. Smad1 and Smad3 activation in response to TGF beta was not affected. The expression of friend leukemia integration factor 1 (Fli1). a transcriptional repressor of collagen, was increased after treatment with ciprofloxacin only in SSc fibroblasts, and this was accompanied by a decrease in the levels of DNA methyltransferase 1 (Dnmt1). Similar effects were observed in SSc-interstitial lung disease (ILD) lung fibroblasts. In summary, our study demonstrates that ciprofloxacin has antifibrotic actions in SSc dermal and lung fibroblasts via the downregulation of Dnmt1, the upregulation of Fli1 and induction of MMPI gene expression via an Erk1/2-dependent mechanism. Thus, our data suggest that ciprofloxacin may he an attractive therapy for SSc skin and lung fibrosis.

Fluorescence spectroscopy as a tool to detect and evaluate glucocorticoid-induced skin atrophy

Topical glucocorticoid (GC) therapy has been successfully used in the treatment of several common cutaneous diseases in clinical practice for a long time, and skin atrophy is one of the most typical cutaneous side effects of this therapy. The aim of this study was to evaluate the potential of noninvasive fluorescence spectroscopy (FS) technique in the detection and classification of GC-induced skin atrophy. A total of 20 male Wistar rats were used in the experimental protocol under controlled environmental conditions and with free access to food. One group received topical application of clobetasol propionate 0.05% for 14 days to induce cutaneous atrophy (atrophic group) and the other (control) group received only vehicle application following the same protocol and schedule. Histological analyses and FS measurements with laser excitation at both 532 nm and 408 nm were obtained on days 1 and 15. The FS results were classified as "normal" or "atrophic" according by histological analysis. Fluorescence spectra obtained with excitation at 408 nm allowed a clear distinction between the control and atrophic groups, and were more informative than the those obtained at 532 nm. Our results reveal that, if correctly applied, FS allows noninvasive evaluation of corticosteroid-induced skin atrophy, and thus represents an important step towards better monitoring of undesirable side effects of cutaneous therapy.

A New Diminutive Species of Loricaria (Siluriformes: Loricariidae) from the Rio Paraguay System, Mato Grosso do Sul, Brazil

Loricaria coximensis. new species, is described from the Rio Paraguay basin, in Mato Grosso do Sul State, Brazil. It is distinguished from all other species of Loricaria, except L. holmbergi, L. lundbergi, L. parnahybae, and L. pumila by having abdominal plates confined to the pre-anal shield and posterior median abdominal area, usually loosely joined or separated by naked areas, pectoral girdle mostly naked, with isolated plates near base of pectoral fins and posterior to gill opening vs. abdominal plates well developed and tightly arranged across the entire median abdominal area, including the pectoral girdle. It is distinguished from these other four species by meristic counts and several morphometric proportions, particularly a broader head (19.4-21.3% SL vs. 14.1-19.1% SL). Loricaria coximensis becomes the third described paedomorphic species of Loricaria, along with L. nickeriensis and L. pumila, based on its small adult size (<100 mm SL), reduced number of lateral dermal plates, and incomplete abdominal plate development. The new species is presently known only from a limited section of the Rio Coxim, which has been severely impacted by a hydroelectric dam and widespread conversion of land for agriculture.

Occupational safety and health practices among flower greenhouses workers from Alto Tiete region (Brazil)

In this preliminary study the occupational safety and health practices among flower greenhouses workers were evaluated. The study was carried out in the alto Tiete region, located at the Sao Paulo State, Brazil. Inadequate welfare facilities; poor pesticide storage, use and disposal conditions; use of highly toxic pesticides; lack of adequate data regarding pesticide use; and incorrect use and maintenance of PPE were observed in most of the visited greenhouses. These results suggest that, in greenhouses, workers may be at higher risk of pesticide exposure, due to many factors that can intensify the exposure such as the lack of control on reentry intervals after pesticide application. Specific regulations are needed to ensure better OSH practices on pesticide use and to improve working conditions in greenhouses, in order to deal with the peculiarities of greenhouse working environment. Some of the special requirements for greenhouses workers' protection are the establishment of ventilation criteria for restricted entry interval; clear reentry restrictions; and EPI for workers other than applicators that need to enter the greenhouse before expiring REI interval. Another important way to improve OSH practices among workers includes the distribution of simple guidelines on the dos and don'ts regarding OSH practices in greenhouses and extensively training interventions to change the perception of hazards and the behavior towards risk. (C) 2011 Elsevier B.V. All rights reserved.

No evidence of selenosis from a selenium-rich diet in the Brazilian Amazon

Selenium (Se) is an essential element and a well-known anti-oxidant. In the Lower Tapajos River region of the Brazilian Amazon, biomarkers of Se range from normal to very high. The local traditional diet includes important Se sources such as Brazil nuts, chicken, game meat and certain fish species. Some studies have reported alterations in keratin structure, gastrointestinal problems and paresthesia in populations with high Se intake. The objective of the present study was to evaluate cutaneous and garlic odor of the breath signs and sentinel symptoms of Se toxicity (selenosis) in relation to Se status in communities along the Tapajos River. Participants (N = 448), aged 15-87 years, were recruited from 12 communities. Se concentrations were measured in blood (B-Se) and plasma (P-Se) by ICP-MS. A nurse performed an examination of the hair, nails, skin and breath for signs of Se toxicity. Interview-administered questionnaires were used to collect information on socio-demographics, medical history and possible symptoms of Se toxicity. In this population, the median levels of B-Se and P-Se were 228.4 mu g/L (range 103.3-1500.2 mu g/L) and 134.8 mu g/L (range 53.6-913.2 mu g/L) respectively. Although B-Se and P-Se surpassed concentrations considered toxic (B-Se: 1000 mu g/L (U.S. EPA, 2002)), no dermal or breath signs or symptoms of Se toxicity were associated with the biomarkers of Se status. In the present study population, where Se intake is mostly from traditional diet, there is no evidence of selenosis. These findings support the need to re-assess Se toxicity considering factors such as the chemical form of Se exposure, route of exposure (inhaled versus ingested), co-exposures to toxic elements such as mercury. Considering the current food transition towards a western diet in the Amazon, further studies should address the possible association between high Se status and cardiometabolic health in this study population. (C) 2011 Elsevier Ltd. All rights reserved.

The occurrence of defensive alkaloids in non-integumentary tissues of the Brazilian red-belly toad Melanophryniscus simplex (Bufonidae)

The red-belly toads (Melanophryniscus) of southern South America secrete defensive alkaloids from dermal granular glands. To date, all information on Melanophryniscus alkaloids has been obtained by extraction from either skins or whole organisms; however, in other amphibians, tetrodotoxins, samandarines, and bufadienolides have been detected in both skin and other organs, which raise the possibility that lipophilic alkaloids may occur in non-integumentary tissues in Melanophryniscus as well. To test this hypothesis, we studied the distribution of alkaloids in the skin, skeletal muscle, liver, and mature oocytes of the red-belly toad M. simplex from three localities in southern Brazil. Gas chromatography and mass spectrometry of skin extracts from 11 individuals of M. simplex resulted in the detection of 47 alkaloids (including isomers), 9 unclassified and 38 from 12 known structural classes. Each alkaloid that was present in the skin of an individual was also present in the same relative proportion in that individual's skeletal muscle, liver, and oocytes. The most abundant and widely distributed alkaloids were the pumiliotoxins 251D, 267C, and 323A, 5,8-disubstituted indolizidines 207A and 223D, 5,6,8-trisubstituted indolizidine 231B, 3,5-disubstituted pyrrolizidines cis-223B and cis- and trans-251K, and izidine 211C. We report the first record of piperidines in Melanophryniscus, bringing the total number of alkaloid classes detected in this genus to 16. Alkaloid composition differed significantly among the three study sites. The functional significance of defensive chemicals in non-integumentary tissues is unknown.

The expression of melanopsin and clock genes in Xenopus laevis melanophores and their modulation by melatonin

Vertebrates have a central clock and also several peripheral clocks. Light responses might result from the integration of light signals by these clocks. The dermal melanophores of Xenopus laevis have a photoreceptor molecule denominated melanopsin (OPN4x). The mechanisms of the circadian clock involve positive and negative feedback. We hypothesize that these dermal melanophores also present peripheral clock characteristics. Using quantitative PCR, we analyzed the pattern of temporal expression of Opn4x and the clock genes Per1, Per2, Bmal1, and Clock in these cells, subjected to a 14-h light:10-h dark (14L:10D) regime or constant darkness (DD). Also, in view of the physiological role of melatonin in the dermal melanophores of X. laevis, we determined whether melatonin modulates the expression of these clock genes. These genes show a time-dependent expression pattern when these cells are exposed to 14L:10D, which differs from the pattern observed under DD. Cells kept in DD for 5 days exhibited overall increased mRNA expression for Opn4x and Clock, and a lower expression for Per1, Per2, and Bmal1. When the cells were kept in DD for 5 days and treated with melatonin for 1 h, 24 h before extraction, the mRNA levels tended to decrease for Opn4x and Clock, did not change for Bmal1, and increased for Per1 and Per2 at different Zeitgeber times (ZT). Although these data are limited to one-day data collection, and therefore preliminary, we suggest that the dermal melanophores of X. laevis might have some characteristics of a peripheral clock, and that melatonin modulates, to a certain extent, melanopsin and clock gene expression.

In vitro cultivation of canine multipotent mesenchymal stromal cells on collagen membranes treated with hyaluronic acid for cell therapy and tissue regeneration

Support structures for dermal regeneration are composed of biodegradable and bioresorbable polymers, animal skin or tendons, or are bacteria products. The use of such materials is controversial due to their low efficiency. An important area within tissue engineering is the application of multipotent mesenchymal stromal cells (MSCs) to reparative surgery. The combined use of biodegradable membranes with stem cell therapy may lead to promising results for patients undergoing unsuccessful conventional treatments. Thus, the aim of this study was to test the efficacy of using membranes composed of anionic collagen with or without the addition of hyaluronic acid (HA) as a substrate for adhesion and in vitro differentiation of bone marrow-derived canine MSCs. The benefit of basic fibroblast growth factor (bFGF) on the differentiation of cells in culture was also tested. MSCs were collected from dog bone marrow, isolated and grown on collagen scaffolds with or without HA. Cell viability, proliferation rate, and cellular toxicity were analyzed after 7 days. The cultured cells showed uniform growth and morphological characteristics of undifferentiated MSCs, which demonstrated that MSCs successfully adapted to the culture conditions established by collagen scaffolds with or without HA. This demonstrates that such scaffolds are promising for applications to tissue regeneration. bFGF significantly increased the proliferative rate of MSCs by 63% when compared to groups without the addition of the growth factor. However, the addition of bFGF becomes limiting, since it has an inhibitory effect at high concentrations in culture medium.

Localized scleroderma: assessment of the therapeutic response to phototherapy

BACKGROUND: Scleroderma is a chronic autoimmune disease characterized by progressive connective tissue sclerosis and microcirculatory changes. Localized scleroderma is considered a limited disease. However, in some cases atrophic and deforming lesions may be observed that hinder the normal development. Literature reports indicate phototherapy as a therapeutic modality with favorable response in cutaneous forms of scleroderma. OBJECTIVES: This study had the purpose of assessing the phototherapy treatment for localized scleroderma. METHODS: Patients with localized scleroderma were selected for phototherapy treatment. They were classified according to the type of localized scleroderma and evolutive stage of the lesions. Clinical examination and skin ultrasound were used to demonstrate the results thus obtained. RESULTS: Some clinical improvement was observed after an average of 10 phototherapeutic sessions. All skin lesions were softer at clinical palpation with scores reduction upon pre and post treatment comparison. The ultrasound showed that most of the assessed lesions presented a decrease in dermal thickness, and only five maintained their previous measure. Treatment response was similar regardless of the type of phototherapeutic treatment employed. CONCLUSIONS: The proposed treatment was effective for all lesions, regardless of the phototherapeutic modality employed. The improvement was observed in all treated skin lesions and confirmed by clinical evaluation and skin ultrasound.

Mastopexia após perda ponderal maciça: suspensão dérmica, remodelação do parênquima e aumento com tecido autógeno

INTRODUÇÃO: Pacientes com perda ponderal significativa podem apresentar mamas com ptose acentuada, perda de projeção do polo superior e excesso de tecido na porção toracolateral. Rubin & Khachi descreveram técnica de mastopexia com suspensão dérmica e remodelação do parênquima associada a aumento com tecido autógeno, tratando a deformidade mamária e o excesso toracolateral em um só estágio. Neste trabalho, é ilustrada essa técnica cirúrgica e demonstradas sua reprodutibilidade e suas complicações. MÉTODO: Foram operadas 14 pacientes com deformidade graus 2 e 3 pela Escala de Pittsburgh, no Hospital Estadual de Sapopemba (São Paulo, SP, Brasil), no período de dezembro de 2008 a dezembro de 2009, utilizando a técnica referida. Foram analisados os seguintes dados: tipos de deformidade das mamas, translocação do complexo areolopapilar (CAP), dimensões dos retalhos, tempo cirúrgico, tempo de permanência do dreno e incidência de complicações. RESULTADOS: A média de idade das pacientes foi de 41,21 + 7,67 anos e o índice de massa corporal médio foi de 29,30 + 2,77. O tempo de seguimento das pacientes variou de 3 meses a 18 meses, com média de 8 meses. Dentre as 14 pacientes operadas, 4 (28,6%) apresentavam deformidade grau 3 e 10 (71,4%), grau 2. A média de translocação do CAP foi de 6,38 cm. As dimensões médias do retalho foram de 25,21 cm x 6,92 cm. O tempo cirúrgico médio foi de 188,57 minutos. Os drenos permaneceram, em média, por 6,21 dias. Foram observadas as seguintes complicações: epiteliose de CAP, deiscência na junção do T, hematoma pequeno e linfedema toracolateral. CONCLUSÕES: A mastopexia com suspensão dérmica, remodelação do parênquima e aumento com tecido autólogo é uma técnica reprodutível, rápida e com baixo índice de complicações.

Uso de matriz dérmica associado ao curativo por pressão negativa na abordagem da contratura em pacientes queimados

INTRODUÇÃO: O aperfeiçoamento no tratamento inicial do paciente queimado, por meio da reposição volêmica e, principalmente, com excisão e enxertia precoce das lesões, resultou em profundo impacto na evolução dos indivíduos queimados, ocorrendo aumento da taxa de sobrevida. Com a maior sobrevida desses pacientes surgiu um novo desafio para a cirurgia reparadora, o tratamento das sequelas de queimaduras, principalmente compostas pelas contraturas. O objetivo deste estudo é demonstrar o uso da matriz dérmica artificial associado a terapia por pressão negativa no tratamento de sequelas de queimaduras. MÉTODO: Foram selecionados 10 pacientes com contratura por queimadura. Os pacientes selecionados foram submetidos a liberação da contratura cicatricial e colocação de matriz dérmica artificial (Integra®), conforme técnica padrão, e curativo por pressão negativa para cobertura. A cada 5 dias, foram realizadas trocas do curativo até completar período de 3 semanas a 4 semanas. Após esse período, o leito foi submetido a enxertia de pele. Os pacientes foram questionados, no pré e pós-operatório, quanto a sua satisfação com os aspectos estético e funcional da região abordada. RESULTADOS: Houve integração de cerca de 98% da matriz de regeneração dérmica na área em que a contratura foi ressecada. Também ocorreu integração de aproximadamente 85% dos enxertos utilizados. Todos os pacientes abordados referiram significativa melhora estética e, principalmente, funcional da região abordada. CONCLUSÕES: O uso da matriz de regeneração dérmica associado a terapia por pressão negativa promove maior taxa de sucesso na abordagem da contratura cicatricial, proporcionando melhor resultado tanto funcional como estético nos pacientes com sequelas graves de queimadura.

Construção de substituto da pele composto por matriz de colágeno porcino povoada por fibroblastos dérmicos e queratinócitos humanos: avaliação histológica

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.

The effect of bone allografts combined with bone marrow stromal cells on the healing of segmental bone defects in a sheep model

Background: The repair of large bone defects is a major orthopedic challenge because autologous bone grafts are not available in large amounts and because harvesting is often associated with donor-site morbidity. Considering that bone marrow stromal cells (BMSC) are responsible for the maintenance of bone turnover throughout life, we investigated bone repair at a site of a critically sized segmental defect in sheep tibia treated with BMSCs loaded onto allografts. The defect was created in the mid-portion of the tibial diaphysis of eight adult sheep, and the sheep were treated with ex-vivo expanded autologous BMSCs isolated from marrow aspirates and loaded onto cortical allografts (n = 4). The treated sheep were compared with control sheep that had been treated with cell-free allografts (n = 4) obtained from donors of the same breed as the receptor sheep. Results: The healing response was monitored by radiographs monthly and by computed tomography and histology at six, ten, fourteen, and eighteen weeks after surgery. For the cell-loaded allografts, union was established more rapidly at the interface between the host bone and the allograft, and the healing process was more conspicuous. Remodeling of the allograft was complete at 18 weeks in the cell-treated animals. Histologically, the marrow cavity was reestablished, with intertrabecular spaces being filled with adipose marrow and with evidence of focal hematopoiesis. Conclusions: Allografts cellularized with AOCs (allografts of osteoprogenitor cells) can generate great clinical outcomes to noncellularized allografts to consolidate, reshape, structurally and morphologically reconstruct bone and bone marrow in a relatively short period of time. These features make this strategy very attractive for clinical use in orthopedic bioengineering