Morphometry of bovine blastocysts produced in vitro in culture media with antioxidants cysteamine or oily extract of Lippia origanoides / [Morfometria de blastocistos bovinos produzidos in vitro em meio de cultura com antioxidantes cisteamina ou extrato oleoso de Lippia origanoides]

This study aimed to evaluate the ultrastructural morphometry of bovine embryos produced in vitro grown at different concentrations of antioxidants. After in vitro maturation and fertilization, the presumptive zygotes were assigned into five treatments. T1) without the addition of any antioxidants (negative control); T2) addition of 50µM/mL cysteamine; and T3, T4 and T5) adding 2.5µg/mL, 5.0µg/mL or 10.0µg/mL of the antioxidants derived from the oily extract from Lippia origanoides, respectively. On D7 of culture, the embryos in the blastocyst stage were fixed and prepared for electron transmission microscopy. These were evaluated for the proportion of cytoplasm-to-nucleus, cytoplasm-to-mitochondria, cytoplasm-to-vacuoles, cytoplasm-to-autophagic vacuoles and cytoplasm-to-lipid droplets. Blastocysts cultured in media containing oily extract of Lippia origanoides presented morphological characteristics such as high cell:mitochondria ratio and low cell:vacuoles and cell:autophagic vacuole ratio, possibly been morphological indicators of embryonic quality. Inner cell mass (ICM) from blastocysts cultured in media without any antioxidants had the highest cell:vacuole ratio. Similar results were found in the trophectoderm (TE) cells of blastocysts from treatment 2. Embryo culture media supplemented with antioxidants derived from Lippia origanoides oil produced embryos with a higher cytoplasmic proportion of organelles, such as mitochondria. Also, treatments without any antioxidants or with the addition of cysteamine presented cytoplasmic vacuolization, a characteristic related to production of poor-quality embryos.(AU)

Este estudo teve como objetivo avaliar a morfometria ultraestrutural de embriões bovinos produzidos in vitro e cultivados em diferentes concentrações de antioxidantes. Após a maturação e a fertilização in vitro, os possíveis zigotos foram divididos em cinco tratamentos: T1) sem adição de antioxidantes (controle negativo); T2) adição de 50µM/mL de cisteamina; e T3, T4 e T5) adição de 2,5µg/mL, 5,0µg/mL ou 10,0µg/mL dos antioxidantes derivados do extrato oleoso de Lippia origanoides, respectivamente. No D7 de cultivo, os embriões em estágio de blastocisto foram fixados e preparados para microscopia eletrônica de transmissão. Estes foram avaliados para a proporção entre citoplasma e núcleo, citoplasma e mitocôndria, citoplasma e vacúolos, citoplasma e vacúolos autofágicos e citoplasma e gotículas lipídicas. Blastocistos cultivados em meio contendo extrato oleoso de Lippia origanoides apresentaram características morfológicas como alta relação célula:mitocôndria e baixa relação célula:vacúolos e célula:vacúolo autofágico, possíveis indicadores morfológicos de qualidade embrionária. A massa celular interna (MCI) de blastocistos cultivados em meio sem quaisquer antioxidantes teve a maior razão célula:vacúolo. Resultados semelhantes foram encontrados nas células do trofectoderma (TE) de blastocistos do tratamento 2. Portanto, o meio de cultivo embrionário suplementado com antioxidantes derivados do óleo de Lippia origanoides produziu embriões com maior proporção citoplasmática de organelas, como mitocôndrias. Além disso, tratamentos sem antioxidantes ou com adição de cisteamina apresentaram vacuolização citoplasmática, característica relacionada à produção de embriões de baixa qualidade.(AU)

Análise ultraestrutural do colágeno de feridas cutâneas de coelhos tratadas com plasma rico em plaquetas de equino / Ultrastructural analysis of the collagen of rabbit skin wounds treated with platelet-rich equine plasma

O colágeno é sintetizado e segregado no espaço extracelular e organizados em fibrilas estriadas de acordo com o tipo de tecido. Utilizaram-se 24 coelhos brancos da raça Nova Zelândia, com idade de 12 meses e com 3,0kg de peso corporal, para avaliar a porcentagem de colágeno das feridas cutâneas tratadas com plasma rico em plaquetas de equino e pomada contendo gentamicina, sulfanilamida, sulfadiazina, ureia e vitamina A. Os animais foram separados em quatro grupos de igual número e submetidos à remoção de pele na região das linhas médias dorsal torácica (feridas tratadas) e lombar (feridas controle). As feridas torácicas foram tratadas com plasma rico em plaqueta de equino e pomada contendo gentamicina, sulfanilamida, sulfadiazina, ureia e vitamina A, e as do grupo controle somente com a pomada. Dos animais do grupo I, foi coletado tecido cutâneo, para a avaliação histológica e a ultraestrutural, com três dias de pós-operatório; dos animais do grupo II, com sete dias; do grupo III, com 14 dias; e do grupo IV, com 21 dias. Decorrido o período de avaliação de cada grupo, foi coletado fragmento de pele para avaliação da porcentagem de colágeno, bem como do diâmetro e da densidade da fibrila de colágeno por microscopia eletrônica de transmissão. O tratamento com PRP de equino associado à aplicação tópica da pomada mostrou-se eficaz na maturação das fibrilas colágenas e na antecipação do processo cicatricial.(AU)

Collagen is synthesized and secreted into the extracellular space and organized into striated fibrils according to the tissue type. This study evaluated the concentration of collagen in rabbit skin wounds treated with equine platelet-rich plasma (PRP) and ointment containing gentamicin, sulfanilamide, sulfadiazine, urea, and vitamin A. Twenty-four New Zealand white rabbits aged 2 to 12 months and weighing 3.0kg were included. The animals were allocated equally into four groups and the skin was removed from the thoracic dorsal midline (treated wound) and lumbar (control wound) regions. The thoracic wounds were treated with equine PRP and ointment containing gentamicin, sulfanilamide, sulfadiazine, urea, and vitamin A, and the control group was treated with the ointment alone. For histological and ultrastructural assessment, cutaneous tissue was collected on postoperative days 3 (group I), 7 (group II), 14 (group III), and 21 (group IV). After the evaluation period, in each group, a skin fragment was collected for analysis of the collagen concentration, as well as the collagen fibril diameter and density by transmission electron microscopy. The results indicated that treatment with equine PRP combined with topical application of the ointment was effective in facilitating the maturation of collagen fibrils and the wound healing process.(AU)

Duplication of Dio3 genes in teleost fish and their divergent expression in skin during flatfish metamorphosis.

Deiodinase 3 (Dio3) plays an essential role during early development in vertebrates by controlling tissue thyroid hormone (TH) availability. The Atlantic halibut (Hippoglossus hippoglossus) possesses duplicate dio3 genes (dio3a and dio3b). Expression analysis indicates that dio3b levels change in abocular skin during metamorphosis and this suggests that this enzyme is associated with the divergent development of larval skin to the juvenile phenotype. In larvae exposed to MMI, a chemical that inhibits TH production, expression of dio3b in ocular skin is significantly up-regulated suggesting that THs normally modulate this genes expression during this developmental event. The molecular basis for divergent dio3a and dio3b expression and responsiveness to MMI treatment is explained by the multiple conserved TREs in the proximal promoter region of teleost dio3b and their absence from the promoter of dio3a. We propose that the divergent expression of dio3 in ocular and abocular skin during halibut metamorphosis contributes to the asymmetric pigment development in response to THs.

Orchestrating change: The thyroid hormones and GI-tract development in flatfish metamorphosis.

Metamorphosis in flatfish (Pleuronectiformes) is a late post-embryonic developmental event that prepares the organism for the larval-to-juvenile transition. Thyroid hormones (THs) play a central role in flatfish metamorphosis and the basic elements that constitute the thyroid axis in vertebrates are all present at this stage. The advantage of using flatfish to study the larval-to-juvenile transition is the profound change in external morphology that accompanies metamorphosis making it easy to track progression to climax. This important lifecycle transition is underpinned by molecular, cellular, structural and functional modifications of organs and tissues that prepare larvae for a successful transition to the adult habitat and lifestyle. Understanding the role of THs in the maturation of organs and tissues with diverse functions during metamorphosis is a major challenge. The change in diet that accompanies the transition from a pelagic larvae to a benthic juvenile in flatfish is associated with structural and functional modifications in the gastrointestinal tract (GI-tract). The present review will focus on the maturation of the GI-tract during metamorphosis giving particular attention to organogenesis of the stomach a TH triggered event. Gene transcripts and biological processes that are associated with GI-tract maturation during Atlantic halibut metamorphosis are identified. Gene ontology analysis reveals core biological functions and putative TH-responsive genes that underpin TH-driven metamorphosis of the GI-tract in Atlantic halibut. Deciphering the specific role remains a challenge. Recent advances in characterizing the molecular, structural and functional modifications that accompany the appearance of a functional stomach in Atlantic halibut are considered and future research challenges identified.

Transcriptome of the Atlantic halibut (Hippoglossus hippoglossus).

Although the Atlantic halibut (Hippoglossus hippoglossus) is an important commercial species, there is still a deficit with regard to the number of transcripts in the databases, which can be accessed and exploited for targeted candidate gene and pathway studies. In this study, the RNAs from head, skin and GI tract from different developmental stages were sequenced to generate 22,272 contigs of 500 base pairs or greater as a molecular resource for this species.

The spreading process of Ehrlichia canis in macrophages is dependent on actin cytoskeleton, calcium and iron influx and lysosomal evasion.

Ehrlichia canis is an obligate intracellular microorganism and the etiologic agent of canine monocytic ehrlichiosis. The invasion process has already been described for some bacteria in this genus, such as E. muris and E. chaffeensis, and consists of four stages: adhesion, internalisation, intracellular proliferation and intercellular spreading. However, little is known about the spreading process of E. canis. The aim of this study was to analyse the role of the actin cytoskeleton, calcium, iron and lysosomes from the host cell in the spreading of E. canis in dog macrophages in vitro. Different inhibitory drugs were used: cytochalasin D (actin polymerisation inhibitor), verapamil (calcium channel blocker) and deferoxamine (iron chelator). Our results showed a decrease in the number of bacteria in infected cells treated with all drugs when compared to controls. Lysosomes in infected cells were cytochemically labelled with acid phosphatase to allow the visualisation of phagosome-lysosome fusion and were further analysed by transmission electron microscopy. Phagosome-lysosome fusion was rarely observed in vacuoles containing viable E. canis. These data suggest that the spreading process of E. canis in vitro is dependent on cellular components analysed and lysosomal evasion.

Molecular and cellular changes in skin and muscle during metamorphosis of Atlantic halibut (Hippoglossus hippoglossus) are accompanied by changes in deiodinases expression.

Flatfish metamorphosis is the most dramatic post-natal developmental event in teleosts. Thyroid hormones (TH), thyroxine (T4) and 3,3'-5'-triiodothyronine (T3) are the necessary and sufficient factors that induce and regulate flatfish metamorphosis. Most of the cellular and molecular action of TH is directed through the binding of T3 to thyroid nuclear receptors bound to promoters with consequent changes in the expression of target genes. The conversion of T4 to T3 and nuclear availability of T3 depends on the expression and activity of a family of 3 selenocysteine deiodinases that activate T4 into T3 or degrade T4 and T3. We have investigated the role of deiodinases in skin and muscle metamorphic changes in halibut. We show that, both at the whole body level and at the cellular level in muscle and skin of the Atlantic halibut (Hippoglossus hippoglossus) during metamorphosis, the coordination between activating (D2) and deactivating (D3) deiodinases expression is strongly correlated with the developmental TH-driven changes. The expression pattern of D2 and D3 in cells of both skin and muscle indicate that TH are necessary for the maintenance of larval metamorphic development and juvenile cell types in these tissues. No break in symmetry occurs in the expression of deiodinases and in metamorphic developmental changes occurring both in trunk skin and muscle. The findings that two of the major tissues in both larvae and juveniles maintain their symmetry throughout metamorphosis suggest that the asymmetric changes occurring during flatfish metamorphosis are restricted to the eye and head region.

The influence of urbanization on natural radiation levels in anomalous areas.

This paper verifies the effects of urbanization on the original levels of environmental gamma radiation in an anomalous area. The fieldwork was done in the city of Guarapari, on the seacoast of the state of Espírito Santo, Brazil. This place was chosen because the whole region is naturally rich in monazite sand, which contains thorium and uranium. A similar study was also carried out in the 1960s. The measurements were done using an inorganic NaI scintillometer detector with Cu+Pb filters coupled to a scaler. The methodology of measurements used was similar to the one applied in the fieldwork carried out in the 1960s. In addition, a simulation was performed in the laboratory, to determine the attenuation of the materials used in the urbanization of the region. The results of this work clearly show that there was a reduction in the levels of external radiation in the streets and squares of Guarapari. It was concluded that the reduction was due to attenuation by the materials used in the urbanization.

[Reactions and feelings of the nursing professional towards death]. / Reações e sentimentos do profissional de enfermagem diante da morte.

Through this paper which self explanatory and description we attempted to understand the reaction and feelings of nurses facing patients who are out of therapeutic possibility. W conducted individual, semi guided interviews with nine nurses from a general hospital in Maringá-PR about their reactions and feelings regarding forthcoming death on a daily base can be an extremely painful experience, therefore, they normally use self defense mechanics such as denial and reasoning death in order to live with daily losses. As a result these professional nurses may fail to notice the limits and their anxiety thus preventing them from providing a good quality of work in assisting their patients. Based on this fact we would like to suggest a prearranged space at the hospital where these health professionals could meet to share, reflect upon and express their anxieties, their fears, their helplessness and pain in face of death, enabling them to be better prepared for their daily losses.