Protocols for obtainment and isolation of two mesenchymal stem cell sources in sheep

PURPOSE: To evaluate different protocols to isolate stem cells from ovine umbilical cord blood and adipose tissue. METHODS: There were used 5 samples of umbilical blood and 5 samples of perirenal adipose tissue from 10 female sheep. All the samples were obtained through surgery, to harvest aseptic samples. There were used 3 protocols for obtainment and culture of umbilical cord blood stem cells and 4 protocols for ovine adipose tissue stem cells. RESULTS: It was possible to observe only one successful protocol for the obtainment of umbilical cord blood stem cells. When analyzing the techniques used to obtain adipose tissue stem cells, only one of the methods was effective as well. Through colony forming unit assay, there were obtained 58 colonies of cells after seven days in culture. Flow citometry tests revealed the cells were positive to CD44 and exhibited negative reaction to CD38, CD45, CD41/61. These cells showed a growth curve with very well defined phases LOG, LAG and PLATEAU. This phases are typically seem in mesenchymal stem cells growth curves. CONCLUSIONS: The isolation and culture of mesenchymal stem cells from ovine umbilical cord blood are complex and request more detailed assays. Stem cells from fat tissue sheep showed mesenchymal characteristics, according to their cell growth curve, ability to origin colonies of fibroblastoid cells and positive reactivity with the antibody CD44 by flow citometry.

Mechanisms of pigmentation loss in subterranean fishes

Troglobitic (exclusively subterranean) organisms usually present, among their apomorphies related to the subterranean life (troglomorphisms), the regression of eyes and melanic pigmentation. The degree of regression varies among species, from a slight reduction to the complete loss of eyes and dark pigmentation, without a taxonomic correlation. While mechanisms of eye reduction have been intensively investigated in some troglobites such as the Mexican blind tetra characins, genus Astyanax, and the European salamander, Proteus anguinus, few studies have focused on pigmentation. The Brazilian subterranean ichthyofauna distinguishes not only by the species richness (23 troglobitic fishes so far known) but also by the variation in the degree of reduction of eyes and pigmentation. This study focused on Brazilian fishes completely devoid of melanic pigmentation: the characiform Stygichthys typhlops (Characidae) and the siluriforms Ancistrus formoso (Loricariidae), Rhamdiopsis sp.1 (Heptapteridae; from caves in the Chapada Diamantina, Bahia) and Rhamdiopsis sp. 2 (cave in Campo Formoso, Bahia). In order to investigate if such depigmentation is the result of blockage in some step in the melanogenesis, in vitro tests of administration of L-DOPA were done, using caudal-fin fragments extracted from living fish. Except for Rhamdiopsis sp. 2, all the studied species were DOPA(+), i.e., melanin was synthesized after L-DOPA administration. This indicates these fish do have melanophores but they are unable to convert L-tyrosine to L-DOPA. On the other hand, Rhamdiopsis sp. 2, like the albino specimens of Trichomycterus itacarambiensis previously studied (which correspond to one third of the population), are DOPA(-), either because the block of melanin synthesis occurs downstream in melanogenesis, which is probably the case with T. itacarambiensis (monogenic system in view of the phenotypic discontinuity), or because the so-called albinos do no possess melanophores. The physiological loss in the ability to synthesize melanin, apparently caused by different genetic processes in DOPA(+) and in DOPA(-) fishes, may co-exist in subterranean populations with a decrease in the density of melanophores, as observed in the pigmented two thirds of T. itacarambiensis population, a morphological reduction apparently controlled by polygenic systems producing a continuous phenotypic variation.

Etiology of cutaneous leishmaniasis and anthropophilic vectors in Juruti, Pará State, Brazil

In a preliminary study in Juruti, a mining municipality in western Pará State, Brazil, 12 out of 21 patients suspected of presenting cutaneous leishmaniasis showed positive PCR (SSUrDNA and G6PD): Leishmania (Viannia) braziliensis (9/12; 75%) and L. (V.) sp. (3/12; 25%). Entomological studies in the same location revealed the presence of 12 different phlebotomine species (n =105). One of the most common species was Lutzomyia (Psychodopygus) complexa (17%) which is both highly anthropophilic and a known vector of L. (V.) braziliensis in other regions of Pará. These preliminary findings should serve to guide future epidemiological surveillance in Juruti.

Ituglanis mambai, a new subterranean catfish from a karst area of Central Brazil, rio Tocantins basin (Siluriformes: Trichomycteridae)

Ituglanis mambai, new species, is described from a cave in the Mambaí karst area, State of Goiás, Central Brazil. The new species distinguishes from epigean and cave congeners by the combination of the following characteristics: posterior supraoccipital fontanel absent; pectoral-fin rays usually i,7; six pleural ribs; total vertebrae 37-38 behind Weberian apparatus; shorter predorsal length (65.1-70.8% SL); shorter caudal peduncle length (8.4-11.9% SL); shorter dorsal-fin base length (7.7-11.3% SL); wider interobital width (29.2-36.5% HL); larger mouth width (43.4-64.0% HL); intermediate between epigean and other cave Ituglanis species as regards to both eyes (diameter varying from 0.5 to1.0 mm in adults, 7.8-10.1 % HL) and pigmentation, composed by irregular light brown spots along the body. The latter indicate the troglobitic status for I. mambai. In addition, this species has the maxillae with a discrete medial-posterior projection; fronto-lacrimal one half-length of the maxillae and pointed backwards; posterior process of palatine half its length, with a tenuous medial concavity; 14 dorsal and 12 ventral procurrent rays. In the natural habitat, I. mambai displayed cryptobiotic habits, trying to hide in the graveled bottom or under boulders when disturbed, apparently showing a negative response to light. It was observed a preference to slow-moving waters. Recent flood marks were observed in the stream conduit in March/April 2007 (end of the rainy season) when less individuals were observed on 300 m of the subterranean stream compared to September 2004 (end of the dry season).

Reaction time assessment in children with ADHD

Attention deficit, impulsivity and hyperactivity are the cardinal features of attention deficit hyperactivity disorder (ADHD) but executive function (EF) disorders, as problems with inhibitory control, working memory and reaction time, besides others EFs, may underlie many of the disturbs associated with the disorder. OBJECTIVE: To examine the reaction time in a computerized test in children with ADHD and normal controls. METHOD: Twenty-three boys (aged 9 to 12) with ADHD diagnosis according to Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition, 2000 (DSM-IV) criteria clinical, without comorbidities, Intelligence Quotient (IQ) >89, never treated with stimulant and fifteen normal controls, age matched were investigated during performance on a voluntary attention psychophysical test. RESULTS: Children with ADHD showed reaction time higher than normal controls. CONCLUSION: A slower reaction time occurred in our patients with ADHD. This findings may be related to problems with the attentional system, that could not maintain an adequate capacity of perceptual input processes and/or in motor output processes, to respond consistently during continuous or repetitive activity.

Leishmania spp. parasite isolation through inoculation of patient biopsy macerates in interferon gamma knockout mice

Isolation of Leishmania parasite and species identification are important for confirmation and to help define the epidemiology of the leishmaniasis. Mice are often used to isolate pathogens, but the most common mouse strains are resistant to infection with parasites from the Leishmania (Viannia) subgenus. In this study we tested the inoculation of interferon gamma knockout (IFNγ KO) mice with biopsy macerates from Leishmania-infected patients to increase the possibility of isolating parasites. Biopsies from twenty five patients with clinical signs of leishmaniasis were taken and tested for the presence of parasites. Immunohistochemical assay (IHC) and conventional histopathology detected the parasite in 88% and 83% of the patients, respectively. Leishmania sp. were isolated in biopsy macerates from 52% of the patients by culture in Grace's insect medium, but 13% of isolates were lost due to contamination. Inoculation of macerates in IFNγ KO mice provides isolation of parasites in 31.8% of the biopsies. Most isolates belong to L. (Viannia) subgenus, as confirmed by PCR, except one that belongs to L. (Leishmania) subgenus. Our preliminary results support the use of IFNγ KO mice to improve the possibility to isolate New World Leishmania species.

Boosting the suppressive effects of Ascaris suum components in IFN-γ-deficient mice

High molecular weight components from Ascaris suum extract suppress ovalbumin-specific immunity in mice. In IFN-γ-deficient mice, ovalbumin-specific delayed-type hypersensitivity reactions are more strongly downregulated by these suppressive components. Here, the cellularity of the delayed-type hypersensitivity reaction in IFN-γ-deficient mice and the increased downregulation induced by Ascaris suum components were analyzed. IL-12p40-dependent neutrophilic influx was predominant. Suboptimal doses of the suppressive fraction from this nematode completely inhibited the hypersensitivity reaction, thus indicating intensification of the immunosuppression under conditions of intense recruitment of IFN-γ-independent neutrophils.

Primeiro registro de Trypanosoma vivax em bovinos no Estado do Rio Grande do Sul, Brasil

Este estudo teve o objetivo de relatar a ocorrência de Trypanosoma vivax no Sul do Brasil. O protozoário foi diagnosticado em esfregaço sanguíneo de um bovino e a identificação baseada na morfologia das formas tripomastigotas e confirmada pela técnica de reação em cadeia de polimerases (PCR). O animal infectado apresentou sintomatologia compatível com a forma nervosa da infecção por T. vivax. Outros bovinos que compartilhavam o mesmo ambiente apresentaram resultados negativos para T. vivax por PCR. Este é o primeiro registro de T. vivax no Estado do Rio Grande do Sul e na região Sul do Brasil.

Avaliação da cicatrização de feridas em dorso de ratos tratados com mitomicina C tópica e posterior reforço com injetável

Estudo experimental em animais. A mitomicina C vem sendo usada como inibidor de fibroblastos, acarretando, com isso, diminuição do processo cicatricial em feridas cirúrgicas. OBJETIVO: Este trabalho visa avaliar o uso de Mitomicina C para diminuir o processo cicatricial, através de seu uso tópico com reforços posteriores injetáveis. MATERIAL E MÉTODOS: Foi usado um modelo de feridas em dorso de ratos, com retirada circular da pele e cicatrização por segunda intenção. Foram usados 18 ratos, divididos em três grupos: controle; com uso tópico; e com reforço de mitomicina C injetável, mensalmente e por 2 meses. Após 3 meses os animais foram sacrificados e as cicatrizes retiradas cirurgicamente e submetidas a estudo histológico. RESULTADOS: Notou-se sob vários critérios que a cicatrização com o uso tópico é menos intensa, mas ao se usar o reforço injetável os parâmetros voltam a ser comparados ao do grupo controle. DISCUSSÃO: Acreditamos que a administração injetável de mitomicina C nas cicatrizes, pela sua elevada característica tóxica, acarreta destruição tecidual e neoformação cicatricial. CONCLUSÕES: A mitomicina C diminui o processo cicatricial quando usada topicamente, mas acarreta aumento da cicatrização quando nestas feridas são feitos reforços injetáveis.

Fatores preditivos do maior número de estádios na cirurgia micrográfica de Mohs para o tratamento do carcinoma espinocelular da cabeça

FUNDAMENTOS: Os carcinomas espinocelulares da pele da cabeça têm como opção terapêutica mais segura a cirurgia micrográfica de Mohs, que apresenta os menores índices de recidiva e a máxima preservação tecidual. Características dos carcinomas espinocelulares podem estar relacionadas a maior número de estádios cirúrgicos. OBJETIVO: Definir características dos carcinomas espinocelulares que sejam preditoras de maior número de estádios na cirurgia de Mohs. MÉTODOS: Análise retrospectiva de 51 carcinomas espinocelulares da cabeça tratados pela cirurgia de Mohs para determinar fatores de risco de maior número de estádios. Foram analisados limites clínicos, morfologia, recidiva, histologia e tamanho, relacionando-os ao número de estádios cirúrgicos. A análise estatística foi realizada pelo teste exato de Fisher e regressão logística multivariada. RESULTADOS: Os carcinomas recidivados tiveram tendência a maior número de estádios (p=0,081). Os tumores com limites imprecisos apresentaram três vezes mais possibilidades de maior número de fases na análise da razão de chances. Esse achado foi compatível com dados da literatura, apesar de não ter sido estatisticamente significante. CONCLUSÃO: Características pré-operatórias dos carcinomas espinocelulares, como recidiva e limites imprecisos, apesar de não preditivas, indicaram tendência a maior número de estádios na cirurgia micrográfica de Mohs.

Synthesis, screening for antiacetylcholinesterase activity and binding mode prediction of a new series of [3-(disubstituted-phosphate)-4,4,4-trifluoro-butyl]-carbamic acid ethyl esters

A series of nine new [3-(disubstituted-phosphate)-4,4,4-trifluoro-butyl]-carbamic acid ethyl esters (phosphate-carbamate compounds) was obtained through the reaction of (4,4,4-trifluoro-3-hydroxybut-1-yl)-carbamic acid ethyl esters with phosphorus oxychloride followed by the addition of alcohols. The products were characterized by ¹H, 13C, 31P, and 19F NMR spectroscopy, GC-MS, and elemental analysis. All the synthesized compounds were screened for acetylcholinesterase (AChE) inhibitory activity using the Ellman method. All compounds containing phosphate and carbamate pharmacophores in their structures showed enzyme inhibition, being the compound bearing the diethoxy phosphate group (2b) the most active compound. Molecular modeling studies were performed to investigate the detailed interactions between AChE active site and small-molecule inhibitor candidates, providing valuable structural insights into AChE inhibition.

The optical analysis of the abdominal wall using the biospeckle after implants of polypropylene mesh in rats

PURPOSE: To evaluate an experimental animal model to study the abdominal tissue activity considering its interaction with a polypropylene mesh, through the use of one of the optical phenomena of light Laser, the biospeckle. METHODS: Fifty Wistar male rats were divided into four groups: Group 1: ten animals not submitted to surgery; Group 2: ten animals submitted to surgery without polypropylene mesh; Group 3: 20 animals submitted to surgery followed by the mesh placement; Group 4: (sham) with ten animals. None of the animals presented post surgical complications being submitted to the optical tests at the 20th postoperative day. RESULTS: The analysis from the biospeckle tests, comparing the medians and standard deviations with T Student test, indicated that no significative difference was observed on the abdominal wall tissue activity in the four groups considered, with and without polypropylene mesh prosthesis implantation. CONCLUSION: The animal model is viable and the biospeckle open ways for a great number of experiments to be developed in evaluating tissue activity.

Relação conjugal, violência psicológica e complementaridade fusional

Neste artigo abordamos um tipo específico de dinâmica conjugal marcada pela violência psicológica. Na clínica com casais, temos observado situações nas quais são utilizadas agressões verbais e humilhações, que afetam a auto-estima e a capacidade de reação e decisão da pessoa agredida. Forma-se um laço perverso caracterizado por um movimento recíproco no sentido do domínio do parceiro, com condições e exigências que submetem a ambos, levando a um tipo de relação na qual impera a complementaridade fusional. O discurso característico da contemporaneidade, de obtenção de prazer e busca da perfeição nos relacionamentos amorosos e da presença do modelo igualitário nas relações homem/mulher, põe em destaque esse tipo de dinâmica relacional. Observa-se, em alguns casos, uma fixação do modelo "homem-ativo-fálico/mulher-passiva-castrada" que permanece subjacente à mudança dos tempos.

Avaliação da memória de trabalho em crianças com transtorno do déficit de atenção e hiperatividade

Esse projeto investigou as habilidades cognitivas da Memória de Trabalho de crianças e adolescentes com o “Transtorno do Déficit de Atenção e Hiperatividade” (TDAH). Foi aplicado teste TIHC - Teste Infantil de Habilidades Cognitivas informatizado, avaliando principalmente cinco habilidades cognitivas Raciocínio Indutivas (RI), Memória de Armazenamento Auditiva (MAA) e Visual e Memória de Trabalho Auditiva (MTA) e Visual (MTV) usando o modelo de Cattell-Horn-Carroll. O teste foi aplicado em 62 sujeitos sendo 32 com TDAH do Hospital das Clínicas-SEPIA-USP e 30 sujeitos controle “normal” do Ensino Fundamental da rede municipal de 1ª à 8ª séries com idades entre 7 e 15 anos, de ambos os sexos. As análises executadas indicaram a existência de diferenças em diversos aspectos relacionados à memória nos tipos peculiares de crianças com TDAH. Verificou-se que as crianças com TDAH obtiveram bom desempenho nas provas de memória visual em detrimento as provas de memória auditiva. A correlação dos resultados do TIHC verificara que o tempo de reação da prova de memória de armazenamento visual manteve uma correlação significativa com a prova de cálculo.

Thermal decomposition of mercerized linter cellulose and its acetates obtained from a homogeneous reaction

Cellulose acetates with different degrees of substitution (DS, from 0.6 to 1.9) were prepared from previously mercerized linter cellulose, in a homogeneous medium, using N,N-dimethylacetamide/lithium chloride as a solvent system. The influence of different degrees of substitution on the properties of cellulose acetates was investigated using thermogravimetric analyses (TGA). Quantitative methods were applied to the thermogravimetric curves in order to determine the apparent activation energy (Ea) related to the thermal decomposition of untreated and mercerized celluloses and cellulose acetates. Ea values were calculated using Broido's method and considering dynamic conditions. Ea values of 158 and 187 kJ mol-1 were obtained for untreated and mercerized cellulose, respectively. A previous study showed that C6OH is the most reactive site for acetylation, probably due to the steric hindrance of C2 and C3. The C6OH takes part in the first step of cellulose decomposition, leading to the formation of levoglucosan and, when it is changed to C6OCOCH3, the results indicate that the mechanism of thermal decomposition changes to one with a lower Ea. A linear correlation between Ea and the DS of the acetates prepared in the present work was identified.