Physical Training Improves Cardiac Structure and Function of Rats After Doxorubicin-Induced Cardiomyopathy

Abstract Background: The use of doxorubicin in chemotherapy has been associated with cardiotoxicity and heart failure. Physical exercise produces favorable morphofunctional adaptations in the cardiovascular system and may reverse cardiac dysfunction in patients undergoing chemotherapy. Objective: To assess the effects of physical training on myocardial structure, cardiac function, and exercise tolerance in Wistar rats initiated after the onset of cardiotoxicity-induced cardiotoxicity. Methods: This study investigated 30 adult male Wistar rats randomly divided into four groups: control (C), exercise (EX), doxorubicin (DX), and doxorubicin and exercise (DXEX). The DX and DXEX groups received six doses of doxorubincin from 1.25 mg/kg body weight up to a cumulative dose of 7.5 mg/kg. Injections were administered intraperitoneally three times a week for two weeks; after this stage, the EX and DXEX groups started physical training (swimming) sessions three times a week with a load of 5% of their body weight. Echocardiography and exercise tolerance tests were performed. Generalized linear models were used in statistical analysis, and a p<0.05 was set as statistically significant. Results: Left ventricular shortening fraction and ejection fraction were reduced in the DX group compared to C, EX, and DXEX. The DXEX group showed greater tolerance to effort when compared to the DX and C groups. Conclusion: Physical training, initiated after the onset of doxorubicin-induced cardiotoxicity, improved cardiac function and exercise tolerance in rats.

Classification of the sperm chromatin compaction alterations in bulls (Bos taurus) and its correlation with the efficiency of the in vitro production of embryos

This paper proposes to classify the sperm chromatin compaction alterations in bulls, according to the affected area location and its objective is evaluating the correlation of the intensity, the heterogeneity and these kinds of chromatin decompaction with the rates of cleavage and the formation of blastocysts of in vitro production of embryos (IVPE). It was used several subfertile animals sperm samples, which were evaluated using the toluidine blue staining and computer image analysis, making possible the categorization of the chromatin decompaction according to their location. The percentages of chromatin decompaction and heterogeneity were also evaluated. IVPEs were done and the rates of cleavage and of blastocysts were correlated with the chromatin characteristics. It made possible the classification of the chromatin decompaction according to the head affected part in at least four types: base decompaction, basal half decompaction, central axis decompaction, total decompaction. Based on the correlation, it can be implicated that each type of classification has different influences on the bull fertility. It made possible understanding that sperms amount with 5% or more of chromatin decompaction intensity interferes in the bull fertility and this condition can be featured as an uncompensable defect, while the heterogeneity of chromatin is not an important factor in the IVPE results.

Gene expression of FOXP3, indoleamine 2,3-dioxygenase (IDO), IL10 and CSF1 in the uterus of cows that received an intrauterine infusion of maternal and paternal antigens / Expressão gênica de FOXP3, indoleamina 2,3-dioxigenase (IDO), IL10 e CSF1 em útero de vacas que receberam infusão intrauterina de antígenos paternos e maternos

Sensitization with conceptus antigens has been shown to be useful for improving reproductive performance facilitating maternal acceptance of an allogeneic embryo through the induction of cytokines and immunoregulatory cells in the uterine microenvironment. As FOXP3, IDO, IL10 and CSF1 in the uterus are important on the recognition and development of embryos during early pregnancy, this study aimed to determine whether simultaneous or isolated administration of paternal (semen) and maternal (PBMCs) antigens in the uterus of cow, on the day of estrus, influence the gene expression of these cytokines. Forty crossbred cows were divided into four treatments: T0: Control; T1: Semen; T2: PBMCs (peripheral blood mononuclear cells) from another cow and T3: PBMCs+Semen. Antigens were administered into the uterine body on the estrus day (D0). Uterine biopsies designed for molecular analysis of gene expression were collected in vivo seven (D7) and fourteen (D14) days after immunostimulation. Transcripts from FOXP3, IDO, IL-10 and CSF-1 were detected in all RNA samples extracted from uterine biopsies. The semiquantitative analysis showed that none of the treatments caused significant increase in the expression of these genes. Furthermore, on D14 all treatments led to a decline in the number of CSF-1 transcripts; moreover, treatment with PBMCs+Semen also led to a drop in the abundance of IL-10 transcripts. Such results suggest that isolated or simultaneous administration of both antigens would not increase maternal tolerance to embryo alloantigens, nor would it create favorable conditions to its growth and pre-implantation development, at least regarding the effects mediated by these genes on D7 and D14 of the estrous cycle.

A sensibilização intrauterina com antígenos do concepto tem-se mostrado útil para melhorar o desempenho reprodutivo, facilitando a aceitação materna de um embrião alogênico por meio da indução de citocinas e células imunorreguladoras no microambiente uterino. Como FOXP3, IDO, IL10 e CSF1 no útero são importantes no reconhecimento e desenvolvimento de embriões durante a gestação inicial, este estudo teve como objetivo determinar se a administração simultânea ou isolada de antígenos paterno (sêmen) e materno (PBMCs) no útero de vacas, no dia do estro, influenciam a expressão gênica dessas citocinas. Quarenta vacas mestiças foram divididas em quatro tratamentos: T0: Controle; T1: Sêmen; T2: PBMCs (células mononucleares do sangue periférico) e T3: PBMCs + Sêmen. Os antígenos foram administrados no corpo do útero no dia do estro (D0). Biópsias uterinas projetadas para análise molecular da expressão gênica foram coletadas in vivo sete (D7) e catorze (D14) dias após imunoestimulação. Transcritos de FOXP3, IDO, IL-10 e CSF-1 foram detetados em todas as amostras de RNA extraídas de biópsias uterinas. A análise semiquantitativa mostrou que nenhum dos tratamentos causou um aumento significativo na expressão desses genes. Além disso, no D14, todos os tratamentos levaram a um declínio na quantidade de transcritos do CSF-1; Além disso, o tratamento com PBMCs + sêmen também levou a uma queda na abundância de transcritos de IL-10. Estes resultados sugerem que a administração isolada ou simultânea de ambos os antígenos não deve aumentar a tolerância materna aos aloantígenos do embrião, nem deve criar condições favoráveis ao seu crescimento e desenvolvimento pré-implantação, pelo menos em relação aos efeitos mediados por esses genes nos dia sete e catorze do ciclo estral.

Micro-RNAs differentially expressed in comparative analysis of sperm samples with high and low efficiency in the in vitro production of bovine (Bos Taurus) embryos / Micro-rnas diferencialmente expressos em análise comparativa de amostras de espermatozoides com alta e baixa efi ciência na produção in vitro de embriões bovinos (bos taurus )

Infertility or subfertility in bovine males may be related to spermatic microRNAs (miRNAs), whose function seems to be associated with the regulation of gene expression, degradation orstorage of messenger RNAs (mRNAs) for later translation into early embryonic development. Thus, the purpose of this study was to identify differentially expressed miRNAs in semen samples from bulls (Bos taurus) with low and high efficiency in the in vitro embryo production (IVEP) and to evaluate if they can be used as markers of semen efficiency for IVEPs. In order to identify miRNA markers of semen efficiency in thein vitro embryo production, eight semen samples from each animal, one bull with high and two bulls with low efficiency in IVEPs were used to perform the RNAseq technique for miRNAs. Initially the samples were washed with PBS to remove the extender semen and subsequently were submitted to RNA extraction protocols performed according to procedures described by mirVana™ miRNA Isolation Kit. Then, the amplification of the miRNAs was carried out, not to mention the preparation of the library (Ion Total RNA-Seq Kit v2), the PCR emulsion reaction, enrichment, as well as the injection of the sample on the chip by the Ion Chef equipment. The sequencing was done on Ion Proton equipment. The comparison between the samples was established using two methodologies for searching for targets to increase the robustness of the analytical procedure: the miRanda program using as cutoff minimum free energy of the hybridization -20 kcal/Mol, 100% of identity between nucleotides 2 and 8 of the miRNA, and the RNAhybrid program, using as cutoff minimum free energy of hybridization -20 kcal/mol. In sum, 1306 miRNAs were identified in the samples. The bta-miR-380-5p, bta-miR-155, bta-miR-30c and bta-miR-34a genes were identified by the Bioinformatics as being strongly differentially expressed between the groups, indicating that these genes may present themselves as possible efficiency markers. However, it has become clear that there is no single miRNA that marks different types and causes of fertility problems.

A infertilidade ou subfertilidade em machos bovinos pode estar relacionada a microRNAs espermáticos (miRNAs), cuja função parece estar associada à regulação da expressão gênica, degradação ou armazenamento de RNAs mensageiros (mRNAs), para posterior tradução no desenvolvimento embrionário inicial. Assim, o objetivo deste estudo foi identificar miRNAs diferencialmente expressos em amostras de sêmen de touros (Bos taurus) com baixa e alta eficiência na produção in vitro de embriões (PIVE) e avaliar se eles podem ser utilizados como marcadores de eficiência do sêmen em PIVEs. Para identificar miRNA marcadores da eficiência de sêmen em PIVE, oito amostras de sêmen de cada animal, sendo um touro com alto e dois touros com baixa eficiência, foram utilizados para realizar a técnica de RNAseq para miRNAs. Inicialmente as amostras foram lavadas com PBS para remover o diluente do sêmen e, posteriormente, foram submetidas a protocolos de extração de RNA realizados de acordo com os procedimentos descritos pelo Kit de isolamento de miRNA mirVana ™. Em seguida, foi realizada a amplificação dos miRNAs, a preparação da biblioteca (Ion RNA-Seq Kit v2), a reação de emulsão de PCR, enriquecimento e a injeção das amostras no chip apropriado utilizando o equipamento Ion. Chef. O sequenciamento foi realizado no equipamento Ion Proton. A comparação entre as amostras foi estabelecida utilizando duas metodologias de busca de alvos para aumentar a robustez do procedimento analítico: o programa miRanda utilizando como valor de corte a energia mínima livre de hibridização -20 kcal / Mol e 100% de identidade entre os nucleotídeos 2 e 8 do miRNA, e o programa RNAhybrid, utilizando como valor de corte a energia mínima livre de hibridização -20 kcal / mol. Em suma, 1306 miRNAs foram identificados nas amostras. Os genes bta-miR-380-5p, bta-miR-155, bta-miR-30c e bta-miR-34a foram identificados pela bioinformática como sendo fortemente diferencialmente expressos entre os grupos, indicando que esses genes podem se apresentar como possíveis marcadores de eficiência. No entanto, ficou claro que não existe um único miRNA que marque diferentes tipos e causas de problemas de fertilidade.

Ambiente web para citomorfometria usando imagens 3D / Web environment for histomorphometry using 3D images

Softwares de bioimage são utilizados para a análise de imagens microscópicas e auxiliam os usuários em suas tomadas de decisões, a usabilidade pode limitar o seu uso. OBJETIVOS: Desenvolver um software de bioimage que pode ser acessado em um navegador web. Auxiliar patologistas e outros usuários na tomada de decisão, minimizando a subjetividade de suas avaliações. MÉTODOS: Foram realizados estudos nos softwares de bioimage open sources, o qual foram identificadas as características positivas e negativas, permitindo a escolha das tecnologias apropriadas para desenvolver o ambiente. RESULTADOS: O trabalho proveu um software de bioimage para análise citomorfométrica usando imagens tridimensionais que pode ser acessado por meio de um navegador web. CONCLUSÃO: O ambiente proposto é capaz de subsidiar os usuários com informações sobre as estruturas das células para tomada de decisão, fornecendo dados quantitativos e permitindo a exploração por meio de uma cena tridimensional.

Bioimage softwares are used for the analysis of microscopic images and assist users in their decision making,usability can limit its use. PURPOSES: Develop a bioimage software that can be accessed through a web browser. Assist pathologists and other users in decision making, minimizing the subjectivity of its evaluations. METHODS: Studies were carried out in the bioimage open sources softwares, which have been identified the positive and negative characteristics, allowing the choice of the appropriate technologies to developing the environment. RESULTS: The work provided abioimage software for histomorphometric analysis using three-dimensional images that can be accessed through a webbrowser. CONCLUSION: The proposed environment can subsidize the users with information on the structures of cells to decision making, providing quantitative data and allowing the exploration of a three-dimensional scene.

Changes in cortical bone channels network and osteocyte organization after the use of zoledronic acid

Objective The aim of this study was to evaluate the effects of zoledronic acid (ZA) on the cortical bone channels network (CBCN) and osteocyte organization in relation to the bone channels. Materials and methods Eighteen male Wistar rats were divided into control (CG) and test groups (TG). Twelve animals from TG received 3 ZA doses (7.5 µg/kg), and 6 animals from CG did not receive any medication. TG animals were euthanized at 14 (n = 6) and 75 (n = 6) dadys after drug injection. CBCN was analyzed in mandibles and tibias using computational routines. The osteocyte organization was qualitatively evaluated in tibias using a three-dimensional reconstruction of images from serial histological sections. Results Significant differences in CBCN of tibia were found between the treated and untreated rats, with a wider range of sizes and shapes of the channels after the use of ZA (channels area p = 0.0063, channels area SD p = 0.0276) and less bone matrix (bone volume p = 0.0388). The alterations in the channels’ morphology were more evident at 75 days after the drug injection (channels perimeter p = 0.0286). No differences were found in mandibles CBCN. The osteocyte distribution revealed more variable patterns of cell distribution in ZA groups, with non-homogeneous distribution of cells in relation to the bone channels. Conclusion Zoledronic acid induces structural changes in CBCN and modifies the osteocyte arrangement in cortical bone in the tibia; also, the variability in the morphology of bone channels became more evident after a certain time of the use of the drug.

Plasma rico em plaquetas autólogas na cicatrização do tendão do músculo gastrocnêmio de coelhos / Plasma platelet-rich autogenous healing tendon of the gastrocnemius muscle in rabbits

As lesões tendíneas podem envolver a secção parcial ou total do tendão calcâneo comum e ocasionar alterações posturais do membro locomotor. O presente trabalho avaliou o efeito da aplicação tópica de concentrado de plaquetas autólogas na reparação do tendão do músculo gastrocnêmio de coelhos, decorridos 45 e 90 dias de período pós-operatório (PO). Doze coelhos adultos foram separados em dois grupos (n = 6) e submetidos à punção cardíaca e colheita de 10 mL de sangue para obtenção do plasma rico em plaquetas (PRP). Nos animais de ambos os grupos, foi realizada a tenotomia transversal total no terço médio do ventre lateral do tendão do músculo gastrocnêmio e a sua aproximação com a sutura de Kessler modificada e fio de náilon. Nos animais do grupo tratado, foi aplicado sobre a síntese dos tendões em média 490.644 mil plaquetas/μL do PRP e notou-se maior quantidade de fibras colágenas em relação ao controle, sendo que aos 90 dias de PO a intensidade de colágeno observado foi maior que nos animais com 45 dias, com maior quantidade de fibroblastos nos controles em relação aos tratados. A administração de concentrado de plasma de plaquetas autógenas no reparo do tendão gastrocnêmio de coelhos estimula e organiza o processo de reparação e ocasiona produção precoce de fibras colágenas.

Tendon lesions may involve the partial or total section of the common calcaneal tendon and cause postural changes of the member. This study evaluated, after 45 and 90 postoperative days (PO), the repair of the tendon of gastrocnemius muscle of rabbits with topical application of autologous platelet concentrate. Twelve adult rabbits were divided into two groups (n = 6) undergoing cardiac puncture and collection of 10 ml of blood to obtain platelet rich-plasma (PRP). Animals of both groups had a transverse tenotomy in the middle third of the lateral belly of the gastrocnemius tendon and muscle that was approximated with modified Kessler suture and nylon thread. In the animals of the treated group it was applied the average of 490.644 platelets / uL of PRP, per animal over the tendon synthesis. The treated group showed a higher amount of collagen fibers than the control one, and at 90 PO days the intensity of collagen was higher than at 45 days with more fibroblasts in the control than in treated one. The administration of plasma autogenous platelet concentrate in the repair of the gastrocnemius tendon of rabbits stimulates and organizes the repair process and causes early production of collagen fibers.

Campylobacter jejuni in commercial eggs

This study evaluated the ability of Campylobacter jejuni to penetrate through the pores of the shells of commercial eggs and colonize the interior of these eggs, which may become a risk factor for human infection. Furthermore, this study assessed the survival and viability of the bacteria in commercial eggs. The eggs were placed in contact with wood shavings infected with C. jejuni to check the passage of the bacteria. In parallel, the bacteria were inoculated directly into the air chamber to assess the viability in the egg yolk. To determine whether the albumen and egg fertility interferes with the entry and survival of bacteria, we used varying concentrations of albumen and SPF and commercial eggs. C. jejuni was recovered in SPF eggs (fertile) after three hours in contact with contaminated wood shavings but not in infertile commercial eggs. The colonies isolated in the SPF eggs were identified by multiplex PCR and the similarity between strains verified by RAPD-PCR. The bacteria grew in different concentrations of albumen in commercial and SPF eggs. We did not find C. jejuni in commercial eggs inoculated directly into the air chamber, but the bacteria were viable during all periods tested in the wood shavings. This study shows that consumption of commercial eggs infected with C. jejuni does not represent a potential risk to human health.

Morphological aspects of the liver of the Podocnemis expansa(Testudines, Podocnemididae)

The liver of P. expansa was characterized morphohistologically. To this end, twenty livers from clinicallyhealthy male and female Podocnemis expansa, weighing from 2.0 to 4,5 kg, supplied by the commercial breederFazenda Moenda da Serra, in Araguapaz, state of Goiás, Brazil, were analyzed macro - and microscopically.The coelomatic cavity was opened and the topography of the fresh organs was examined visually. After thehistological preparation, the slides were stained with Hematoxylin and Eosin (HE), Periodic Acid-Schiff(PAS), Gomori Trichrome, Reticulin and Picrosirius. The liver of P. expansa is a voluminous organ with anapproximately rectangular shape and brown coloration, varying from light to dark shades, and is divided into aright lobe, left lobe, and a central portion. The right lobe is the largest of the three portions. The gall bladderis located in a depression in the caudal portion of the right lobe, where the gall duct begins and emptiesinto the duodenum. Histologically, the hepatocytes are arranged in the form of double cords surroundedby winding sinusoidal capillaries. In cross section, they resemble acini containing approximately two to fivehepatocytes surrounding a probable central biliary canaliculus. The hepatocytes are polyhedral or pyramidalin shape, of uniform size, with a few central nuclei and others displaced peripherally, and the cytoplasm islittle eosinophilic when analyzed by the HE staining technique. The parenchyma is supported by delicatereticular fibers surrounding hepatocytes and sinusoids. The parenchyma and perisinusoidal spaces containlarge quantities of melanomacrophages, mainly close to the portal spaces.

Transmission of Campylobacter coli in chicken embryos

Campylobacter coli is an important species involved in human cases of enteritis, and chickens are carriers of the pathogen mainly in developing country. The current study aimed to evaluate the transmission of C. coli and its pathogenic effects in chicken embryos. Breeder hens were inoculated intra-esophageally with C. coli isolated from chickens, and their eggs and embryos were analyzed for the presence of bacteria using real-time PCR and plate culture. The viability of embryos was verified. In parallel, SPF eggs were inoculated with C. coli in the air sac; after incubation, the embryos were submitted to the same analysis as the embryos from breeder hens. In embryos and fertile eggs from breeder hens, the bacterium was only identified by molecular methods; in the SPF eggs, however, the bacterium was detected by both techniques. The results showed no relationship between embryo mortality and positivity for C. coli in the embryos from breeder hens. However, the presence of bacteria is a cause of precocious mortality for SPF embryos. This study revealed that although the vertical transmission is a possible event, the bacteria can not grow in embryonic field samples.

Ponte miocárdica associada a lesões cardiovasculares em bovinos adultos da raça Canchim / Myocardial bridge associated with cardiovascular injuries in bovines adult of Canchim race / Puente miocárdico asociado a lesiones cardiovasculares en bovinos adultos de la raza canchim

FUNDAMENTO: A influência que a ponte miocárdica exerce sobre a corrente sanguínea no curso do segmento arterial sob a ponte tem sido objeto de discussão pela comunidade científica. OBJETIVO: Comparar o tecido muscular ultraestrutural da ponte miocárdica e a parede ventricular; analisar o grau de lesão da camada íntima dos segmentos arteriais e investigar possíveis mudanças que podem preceder ou iniciar o processo de lesões ateroscleróticas. MÉTODOS: Quarenta corações bovinos da raça Canchim foram estudados em relação às alterações da camada íntima das artérias coronarianas nos diferentes segmentos de ponte miocárdica. Para o exame microscópico, foram feitas colorações por hematoxilina-eosina e fucsina-resorcina seguindo técnicas microscópicas convencionais. Para o exame de microscopia eletrônica, os segmentos da ponte miocárdica de doze corações bovinos Canchim foram coletados a partir da parede ventricular e da artéria coronariana e foram processados de acordo com técnicas convencionais. RESULTADOS: Na microscopia de luz, foi observada maior frequência de lesões em segmentos pré-ponte e pós-ponte da camada íntima, em comparação ao segmento ponte. Espessamentos da camada íntima foram seguidos por um desarranjo na lâmina limitante elástica interna. Essas células frequentemente apresentaram seus citoplasmas ingurgitados por gotas lipídicas, compondo as chamadas células de espuma. A microscopia eletrônica revelou que as fibras musculares da ponte miocárdica geralmente se unem de forma reta e lisa apresentando ramos laterais com um número maior de mitocôndrias no músculo ventricular do que na ponte. CONCLUSÃO: Há poucas diferenças entre os tecidos musculares estudados; lesões da camada íntima são menos frequentes em regiões da ponte em comparação com as regiões pré e pós-ponte.

BACKGROUND: The influence that myocardial bridge exercises over blood stream in the course of arterial segment under the bridge has been discussed by the scientific community. OBJECTIVE: To compare ultrastructural muscle tissue of myocardial bridge and the ventricular wall; to analyze the degree of injury to the tunica intima of the arterial segments, and look for possible changes that may precede or initiate the process of atherosclerotic lesions. METHODS: Forty Canchim bovine hearts were studied regarding alterations of the tunica intima from coronary arteries on the different myocardial bridge segments. For the microscopic examination, hematoxylin-eosin and fuchsin-resorcin staining following conventional microscope techniques were made. For the electronic microscopic examination, myocardial Bridge segments from twelve Canchim bovine hearts were collected from the ventricle wall and coronary artery and were processed according to conventional techniques. RESULTS: In the light microscopy, a higher frequency of lesions on prepontine and postpontine segments of the tunica intima was observed, compared to the pontine segment. Tunica intima thickenings were followed by a disarrangement on the internal elastic limitant lamina. These cells often presented their cytoplasmas engorged by lipidic drops, making up the so-called foam cells. Electronic microscopy revealed that the muscular fibers of the myocardial bridge are usually joined in a straight and smooth way presenting lateral branches with a greater number of mitochondria in the ventricular muscle than in the bridge. CONCLUSION: There are few differences between the muscle tissues studied; intimae lesions are less frequent in pontine regions compared to pre and post-pontine regions.

FUNDAMENTO: La influencia que el puente miocárdico ejerce sobre la corriente sanguínea en el transcurso del segmento arterial bajo el puente, ha sido objeto de discusión por parte de la comunidad científica. OBJECTIVO: Comparar el tejido muscular ultra estructural del puente miocárdico y la pared ventricular; analizar el grado de lesión de la capa íntima de los segmentos arteriales e investigar posibles cambios que pueden preceder o iniciar el proceso de lesiones ateroscleróticas. MÉTODO: Cuarenta corazones bovinos de la raza Canchim fueron estudiados con respecto a las alteraciones de la capa íntima de las arterias coronarias en los diferentes segmentos del puente miocárdico. Para el examen microscópico, se hicieron coloraciones por hematoxilina-eosina y fucsina-resorcina secundando las técnicas microscópicas convencionales. Para el examen de microscopía electrónica, los segmentos del puente miocárdico de doce corazones de bovinos de la raza Canchim, fueron recolectados así como de la pared ventricular y de la arteria coronaria y fueron procesados de acuerdo con las técnicas convencionales. RESULTADOS: En la microscopía de luz, observamos una mayor frecuencia de lesiones en segmentos de pre puente y pos-puente de la capa íntima, en comparación con el segmento puente. Los espesamientos de la capa íntima vinieron acompañados por un desarreglo en la lámina limitante elástica interna. Esas células a menudo presentan sus citoplasmas ingurgitados por gotas lipídicas, lo que compone las llamadas células de espuma. La microscopía electrónica reveló que las fibras musculares del puente miocárdico generalmente se unen de forma recta y lisa, presentando ramas laterales con un número mayor de mitocondrias en el músculo ventricular que en el puente. CONCLUSIONES: Existen pocas diferencias entre los tejidos musculares estudiados. Las lesiones de la capa íntima son menos frecuentes en las regiones del puente en comparación con las regiones pre y pos puente.

Metabolic activity of Streptococcus mutans biofilms after treatment with different mouthwash formulations

Aim: The aim of this study was to investigate the metabolic activity of Streptococcus mutans biofilms after treatment with mouthwashes with different compositions. Methods: S. mutans biofilms were growth on polystyrene plates during 18 h, washed with sterile saline and treated with the following mouthwashes during 1 min: Listerine®, Oral B®, Parodontax® and Periogard® with and without alcohol. After the treatment, the biofilms were incubated with complete medium containing sucrose during 60, 120 or 180 min, and then samples were collected for pH measurements. In addition, biofilms were grown in microscope coverslips treated as described above, followed by staining with Propidium Iodide and Fluoresceine for visualization with a confocal laser scanning microscopy. Results: For all mouthwashes evaluated, treatment was deleterious to cell metabolism, since little or no acidification was observed at least 60 min after treatment. Mouthwashes containing 0.2% chlorhexidine (Parodontax®) or essential oils (Listerine®) induced a significant reduction in the metabolic activity of biofilms during the tested time points (120 and 180 min after treatment), being thus more effective than the mouthwashes containing 0.12% chlorhexidine (Periogard®) or cetylpyridinium plus fluoride (Oral B®). The confocal analysis overall confirmed the results observed in the analysis of metabolic activity. Conclusions: The treatment of biofilms with mouthwashes containing 0.2% chlorhexidine or essential oils induced significant reduction in S. mutans metabolism.

Inibição de metaloproteinases por extratos aquosos de Aloe vera, Annona muricata e chá preto / Inhibition of metalloproteinases by Aloe vera, Annona muricata, and black tea aqueous extracts

As Metaloproteinases de Matriz (MMPs) podem exercer importantes funções na remodelagem fisiológica e patológica da matriz extracelular. Elas são implicadas na progressão tumoral de muitas malignidades humanas interferindo em eventos angiogênicos, invasivos e metastáticos. Produtos obtidos de algumas plantas têm sido consistentemente associados a funções neoplásicas, possivelmente por reproduzir a atividade neutralizadora dos inibidores teciduais (TIMPs) das MMPs. No presente trabalho, investigou-se a inibição da atividade gelatinolítica das MMPs -2 e -9 por extratos aquosos (EA) de Aloe vera, chá preto e Annona muricata diluídos em tampão de ativação (TA, com cálcio). A atividade proteolítica das MMPs foi avaliada em ensaios zimográficos. Para avaliar o efeito inibitório dos produtos testados, géis foram seccionados e incubados em soluções contendo o TA e os EA em três diluições seriadas. Associação indireta foi observada entre o EA de A. vera e chá preto e a inibição da atividade de MMP-2. EA de A. muricata exerceu pequeno efeito inibitório na MMP-2 nas duas menores diluições. A atividade de MMP-9 foi consistentemente neutralizada por todos os EA testados, de forma dose-dependente. Estes resultados sugerem que os efeitos antitumorais verificados paraos EA de A. vera, A. muricaca, e chá preto podem ser explicados parcialmente por sua atividade inibitória sobre as MMP-2 e -9.

Matrix metalloproteinases may exert important roles in both physiologic and pathologicextracellular matrix remodeling. They have been implicated in tumoral progression of many human malignancies interfering on angiogenic, invasive, and metastatic events. Products obtained from some plants have been consistently associated to anti-neoplastic roles, possibly by reproducing the neutralizing activity of tissue inhibitors over MMPs. In the present work, we investigate the inhibition of the MMP-2 and -9 gelatinolytic activity by Aloe vera, black tea, and Annona muricata aqueous extracts (AE) diluted in activation buffer (AB, with calcium). The proteolytic activity of MMP was evaluated on zymographic assays. To evaluate the inhibitory effect of the tested products, gels were sectioned and incubated in the solutions containing the AB and AE from three serial dilutions. Indirect association was observed between the dilution of A. vera and black tea and MMP-2 activity inhibition. A. muricata exerted only small inhibition on MMP-2 effect in the two smallest dilutions. The MMP-9 activity was consistently neutralized by all of the natural solutions in a dose-dependent way. The present results suggest that the antitumoral effects verified for A. vera, A. muricaca, and black tea AE may be partially explained by their inhibitory activity on MMP.

Alterações de cromatina em espermatozóides de ovinos e caprinos avaliadas por azul de toluidina e alaranjado de acridina / Chromatin alterations in ram and goat spermatozoa evaluated by toluidine blue and acridine orange

Reprodutores com espermograma normal podem se comportar como subférteis ou passarem por períodos de subfertilidade. As alterações na cromatina dos espermatozóides são possíveis explicações encontradas para tais comportamentos. Conduziu-se este trabalho, com o objetivo de testar a eficiência de azul de toluidina (AT) e do alaranjado de acridina (AA) na identificação de alterações na compactação de cromatina em espermatozóides de ovinos e caprinos, além de avaliar a correlação entre essas alterações e as de morfologia espermática. Para tal, foram avaliadas amostras de sêmen de 15 ovinos e de 15 caprinos, com dez repetições para cada método por animal. Calcularam-se a média, o desvio padrão (DP) e o coeficiente de variação (CV) para cada técnica e animal. Utilizou-se o teste t-Student para avaliar diferença entre as médias obtidas nos dois métodos. Também foram calculados a correlação de Pearson e os coeficientes kappa ponderado e não ponderado para avaliar a concordância entre os métodos com AT e AA. Foi verificado que nem sempre as anomalias morfológicas de cabeça são acompanhadas por alterações na cromatina identificáveis pelos métodos utilizados neste trabalho. O método AT é mais estável e possui maior sensibilidade do que AA para ambas as espécies, sendo o mais indicado para caprinos. Contudo, em razão de apresentar repetibilidade muito baixa, ambos os métodos não são indicados para avaliação espermática em ovinos.

Males with normal spermogram can behave as subfertile or pass for periods of subfertility. Chromatin alterations of spermatozoa can account for such behavior. The objective of the present work was to test the efficacy of toluidine blue (TB) and acridine orange (AO) in the identification of alterations in chromatin compaction in spermatozoa from rams and goats, in addition to evaluate the correlation between those alterations and the ones of spermatic morphology. In order to do that, samples of semen from 15 rams and 15 goats were evaluated with 10 replications for each method. Mean, standard deviation and coefficient of variation were calculated for each animal and technique. Student's t-test was used to evaluate differences between the averages of two methods. Pearson correlation coefficient, weighted and non weighted kappa coefficient were also calculated to evaluate the agreement between methods with TB and AO. It was verified that morphological alterations of head not always are accompanied by alterations in chromatin identified by the methods used in this work. The TB method is more stable and sensitive than AO method for both species and this is most appropriate for goats. However, due to the very low repeatability, both methods are not indicated for evaluating ram spermatozoa.

Neurorrafia do ramo bucal dorsal do nervo facial em coelhos com proteção de segmento intestinal alógeno / Neurorrhaphy of the dorsal buccal branch of facial nerve in rabbits with protection o f allograft intestinal segment

Foram utilizados 18 coelhos, Nova Zelândia, machos, adultos, para avaliação clínica e histológica do reparo do ramo bucal dorsal do nervo facial, decorridos 15, 30 e 60 dias de pós-operatório (PO). Os animais foram distribuídos em dois grupos para secção e aproximação epineural do ramo bucal com fio náilon monofilamentoso 10-0. Nos animais do grupo I, o nervo foi revestido com proteção de segmento de jejuno alógeno conservado em glicerina a 98% e o grupo II apenas aplicação de sutura epineural. Nos coelhos dos dois grupos ocorreu retorno da movimentação do lábio superior a partir da oitava semana.Verificou-se infiltrados celulares e células gigantes com fibrose desorganizada e fibras colágenas do envoltório alógeno entremeadas ao tecido conjuntivo. Aos 15 e 30 dias de PO, os cotos distais deambos os grupos encontravam-se com degeneração walleriana e aos 60 dias, com fibras regeneradas. A reparação do ramo bucal dorsal do nervo facial com o segmento intestinal não foi significativamente diferente nos coelhos do controle, quanto à avaliação de recuperação funcional e histológica.

18 rabbits, New Zealand, males, adults were used for clinical and histological evaluation of repair dorsal buccal branch of facial nerve after 15, 30 and 60 days postoperatively (PO). The animals were divided into two groups for transection and 10-0 nylon monofilament epineural suture of buccal branch. In animals in Group I, the nerve was coated with protection of jejunum allograft preserved in glycerin 98% and in group II was applied epineural suture. Both groups occurred the return of movement of the upper lip from the eighth week. There was infiltrated cellular and giant cells with fibrosis unsystematic and collagen fibers of the allograft jejunum joing to the connective tissue. At 15 and 30 days of PO, the distal nerve stumps of both groups were found with degeneration wallerian and in 60 days, regenerated fibers. The repair of the dorsal buccal branch offacial nerve with the allograft wasn’t significantly different between the control rabbits as to the assessment of histological and functional recovery.

Spironolactone effects on myocardium changes induced by thyroid hormone in rats / Efeitos da espironolactona sobre as alterações miocárdicas induzidas pelo hormônio tireoideano em ratos

FUNDAMENTO: Estudar o papel do sistema renina-angiotensina-aldosterona na hipertrofia miocárdica induzida pelo hormônio tireoideano, utilizando-se a espironolactona. OBJETIVO: Analisar as alterações morfológicas no miocárdio induzidas pelo hormônio tireoideano e os efeitos da espironolactona nesse processo. MÉTODOS: Foram estudados 40 ratos Wistar, divididos em quatro grupos, que receberam: veículo utilizado para a diluição do hormônio tireoideano (C); levotiroxina sódica (50 µg/rato/dia) (H); espironolactona (0,3 mg/kg/dia) (E) e hormônio tireoideano + espironolactona (HE), nas mesmas doses citadas, durante 28 dias consecutivos. Todos os animais foram submetidos a pesagem, coleta de sangue para dosagens hormonais e realização de ECG no início e no final do experimento. Ao final do período de estudo, os animais foram sacrificados para determinação do peso do ventrículo esquerdo (VE) e obtenção de cortes de VE para análise morfológica. RESULTADOS: Houve aumento dos níveis de T3 no plasma, perda de peso corporal e aumento da freqüência cardíaca nos animais que receberam o hormônio. O peso do VE foi maior nos grupos H e HE. A análise histométrica mostrou maiores diâmetros dos miócitos no grupo H, com os valores decrescentes nos grupos HE, E e C, sendo as diferenças estatisticamente significantes entre todos os grupos. A espironolactona associada ao hormônio tireoideano (HT) diminuiu em 14,6 por cento a hipertrofia transversal dos miócitos. CONCLUSÃO: Em ratos tratados com hormônio tireoideano ocorre hipertrofia cardíaca com aumento do peso do VE e do diâmetro do miócito. A associação de espironolactona ao hormônio tireoideano previne parcialmente essa hipertrofia por mecanismos ainda desconhecidos.

BACKGROUND: To study the possible role of aldosterone on thyroid hormone-induced myocardium hypertrophy, using spironolactone. OBJECTIVE: To evaluate morphological changes in the myocardium induced by thyroid hormone and the possible effects of spironolactone use on these alterations. METHODS: Forty Wistar rats were studied. The animals were allocated to four groups and were given: the vehicle used for dilution of the thyroid hormone (C); sodium levothyroxin at 50 µg/rat/day (H); spironolactone, 0.3 mg/kg/day (S); or thyroid hormone plus spironolactone (HS), at the same doses mentioned above, for 28 consecutive days. All the animals were weighed, had blood drawn for hormonal measurements and underwent ECG at the start and the end of the experiment. At the end of experiment all animals were euthanized, the weight of the left ventricle (LV) was determined and LV slices were obtained for morphological analysis. RESULTS: There was an increase in T3 levels, decrease of body weight and higher heart rate in the animals from group H. The LV weight was significantly higher in the H e HS groups. The histometric analyses that measured the diameter of the myocytes showed higher values in group H and a progressive decrease in groups HS, S and C, with a significant difference among all the groups. The addition of spironolactone decreased the transversal myocyte hypertrophy by 14.6 percent. CONCLUSION: Rats treated with thyroid hormone present cardiac hypertrophy with increased LV weight and greater myocyte diameter. Spironolactone, when associated with thyroid hormone, can partially prevent this hypertrophy through mechanisms that are yet to be determined.

Morfogênese do testículo de embriões e fetos de vacas da raça nelore (Bos taurus indicus) / Morfogenesis of Testis From Embryos and Fetuses of Nelore Breed Cows (Bos taurus Indicus)

Este estudo teve como objetivo acompanhar o processo de desenvolvimento testicular desde a fase indiferenciada até sua completa formação. Embriões e fetos de vacas da raça nelore (Bos taurus indicus) foram obtidos em frigoríficos próximos à cidade de Uberlândia, Minas Gerais. As gônadas dos fetos e os embriões foram fixados em bouin e processados para microscópica óptica convencional. A gônada foi observada primeiramente em um embrião de 1,0 cm de comprimento. Em embriões com 2,5 cm a presença da albugínea permite a identificação do sexo. A espessura média da albugínea variou de 29,08 a 558,46 mm. Gradativamente, observou-se aumento da vascularização da albugínea e do parênquima. O mediastino encontrava-se localizado centralmente. Houve uma diminuição no espaço ocupado pelos cordões testiculares de 63,71 para 41,99% do volume total dos testículos. O seu diâmetro variou de 31,68 a 48,80 mm. O diâmetro das células germinativas (e dos seus núcleos) foi de 12,27(6,65) a 16,95 (14,21) mm, respectivamente. A quantidade de células germinativas por corte transversal de cordão diminuiu de um máximo de 2,80 para 0,76. O número total de células germinativas foi de 16 no princípio da colonização da gônada para 18,32 x 106 no final do estudo. O número de células de Sertoli por corte transversal de cordão variou de 10,00 a 16,25. Os resultados obtidos mostram que a origem e a formação dos testículos nos embriões e fetos de vacas da raça Nelore (Bos taurus indicus) ocorre de forma muito semelhante ao do que é descrito para Bos taurus taurus.

The aim of this study was to accompany the process of testicular development from the non-differentiable phase to its complete formation. Embryos and fetuses of Nelore breed cows (Bos Taurus indicus) were obtained in slaughterhouses near the Uberlandia city, Minas Gerais. The gonads and the embryos were fixed in Bouin’s fixative and afterwards processed for conventional optical microscopy. The gonadal was observed firstly in a 1.0 cm long embryo. In 2.5 cm long embryos the presence of the albuginea allows the sex identification. The mean thickness of the albuginea ranged from 29.08 to 558.45 mm. Gradually increase of vascularization of the albuginea and parenchyma is observed. The mediastinum is located centrally. There was a decrease in the space occupied by the testicular cords, from 63.71 to 41.99% of the total testes volume. Its diameter ranged from 31.68 to 48.80 mm. The diameter of germinal cells (and their nuclei) was from 12.27 (6.65) to 16.95 914.21) mm. The quantity of germinal cells by cross section of cord decreased from a maximum of 2.80 to 0.76. The total number of germinal cells was from 16 at the beginning of colonization of the gonad to 18.32 x 106 at the end of the study. The number of Sertoli’s cells by cross section of cord ranged from 10.00 to 16.25. The results obtained show that the origin and formation of testes in embryos and fetuses from Nelore breed cows (Bos taurus indicus) does occur in a very similar way to what is described for Bos taurus taurus.

Avaliação da morfologia e da compactação cromatínica em espermatozóides de galo (Gallus gallus, Linnaeus, 1758) através de microscopia eletrônica de transmissão / Morphology and chromatin condensation evaluation in fowl spermatozoa (Gallus gallus, Linnaeus, 1758) through the transmission electron microscopy

O espermograma é um dos principais métodos de avaliação da fertilidade em mamíferos, contudo, algumas alterações do sêmen como a compactação da cromatina dos espermatozóides, não são identificadas nessa rotina, podendo interferir significativamente na fertilidade de reprodutores. Na avicultura, geralmente a avaliação de reprodutores é feita por amostragem e os parâmetros avaliados são em menor número que em mamíferos, sendo que a compactação da cromatina dos espermatozóides de galo nunca foi explorada. Este trabalho teve como objetivo verificar através da microscopia eletrônica de transmissão, se existe diferentes intensidades de compactação de cromatina em espermatozóides de galo. Para isto, foram coletadas 20 amostras de sêmen de galo, que foram fixadas por 48 horas em glutaraldeído 4% tamponado em cacodilato de sódio 0,1M a pH 7,2, após centrifugado e lavado em tampão cacodilato o sêmen foi pós-fixados em tetróxido de ósmio a 1% mais ferrocianeto de potássio a 1,25%. O sedimento foi incluído, cortado e posteriormente contrastado. Os cortes foram examinados e documentados em microscópio eletrônico Zeiss EM-109. Geralmente, os espermatozóides de galo possuem acrossoma com material homogêneo ou levemente granular e de densidade moderada, o núcleo com cromatina densa e levemente granular, varia as tonalidades de cinza, ou seja, os graus de compactação. Observou-se também o “perforatorium”, estrutura que une o núcleo ao acrossoma. A inserção da cauda é por meio de dois centríolos, sendo um transversal e o outro longitudinal ao eixo. A peça intermediária possui axonema típico envolto por mitocôndrias aparentemente dispostas longitudinalmente. A transição entre as peças intermediária e principal da cauda é marcada pela ausência das mitocôndrias. A peça principal é formada pelo axonema envolto por uma camada de citoplasma granular. Na avaliação das patologias morfológicas observaram-se cabeças arredondadas, dobradas, envoltas por conteúdo da peça intermediária...

The spermiogram is one of the principal methods of evaluation of the fertility in mammals; however some semen modifications as the spermatozoa chromatin condensation, are not identified in this routine, and can expressively interfere in the fertility of reproducers. In poultry breeding, generally the evaluation of reproducers is made by sampling and the evaluated parameters are in fewer quantities than in mammals, considering the spermatozoa chromatin condensation of the fowl has never been exploited. The objective of this work is verifying through the transmission electron microscopy, whether there are different intensities of spermatozoa chromatin condensation of fowl. So that, have been collected 20 samples of fowl semen, which were placed in 4% glutaraldehyde buffered in 0.1M sodium cacodilate to pH 7.2 for 48 hours, after being centrifugated and rinsed in cacodilate tampon, the semen was post-fixed in 1 % osmium tetroxide plus 1.25% potassium ferrocyanide. The sediment was embedded, cut and contrasted. The thin sections were examined and documented in transmission electronic microscopy Zeiss EM -109. Generally the fowl spermatozoon has acrosome with homogenic or slightly granular and of moderate density material, the nucleus with dense or slightly granular chromatin, varies the gray scales, that is, the grades of condensation. It was also observed the "perforatorium", structure that links the nucleus to the acrosome. The insertion of the tail is made through two cetriules, being one transversal and the other longitudinal to the axle. The intermediate piece has typical axoneme involved by mitochondrias, apparently displayed longitudinally. The transition between the principal and the intermediate pieces of the tail is marked by the absence of the mitochondrias. The principal piece is formed by axoneme involved by a granular cytoplasm layer. In the evaluation of the morphological pathologies, it was observed rounded heads, folded, involved by the...

Avaliação da integridade cromatínica de espermatozóides de galos (Gallus gallus, Linnaeus, 1758) de linhagem pesada em duas idades / Evaluation of chromatin integrity of fowl spermatozoa (Gallus gallus, Linnaeus, 1758) from meat chicken in two ages

Na avicultura, a avaliação de fertilidade em machos é de extrema importância, para garantir uma melhor produção de ovos férteis. As técnicas para avaliação de fertilidade em galos são pouco exploradas, sendo que na maioria das vezes, a avaliação é feita por amostragem e levando em consideração, apenas fatores morfofisiológicos, diretamente relacionados com o espermatozóide. Mas é sabido que, em outras espécies, além dos fatores morfofisiológicos, existem problemas intrínsecos ao espermatozóide, como a baixa compactação da cromatina, que pode levar a distúrbios de fertilidade, que na maioria das vezes não são diagnosticados. O objetivo desse trabalho foi a adaptação de técnicas de avaliação da cromatina, já descritas em outras espécies, para aves de linhagem pesada (Gallus gallus, Linnaeus, 1758), correlacionando as alterações cromatínicas com as alterações morfológicas e com a fertilidade. Para tanto, sêmen de galo com diferentes níveis de fertilidade, foram utilizados em diferentes métodos para identificação de alterações na cromatina, utilizando os corantes azul de toluidina e alaranjado de acridina. As avaliações demonstraram que esfregaços de sêmen fresco de galo com posterior fixação geram artefatos que levam a alterações na forma da cabeça e na integridade da cromatina, não sendo indicados em métodos de avaliação de fertilidade. Apesar de todos os métodos testados apresentarem falhas metodológicas e um certo grau de subjetividade, o método que gerou melhores resultados foi a mistura de uma gota de sêmen conservado em formol salina e uma gota do alaranjado de acridina sobre lâmina de microscopia, com posterior secagem e observação em microscopia de fluorescência com filtro de excitação azul. Pelo método verificou-se que alterações na compactação da cromatina de espermatozóides de galo geralmente não são acompanhadas por alterações morfológicas e que geralmente as alterações morfológicas de espermatozóides de galo são acompanhadas por alterações na ...

In poultry breeding, male fertility has extreme importance to assure a better production of fertile eggs. Evaluation techniques for fertility of fowls are not well explored and in most cases, the evaluation is done through sampling considering only morphophysiological factors directly related with the spermatozoon. It's known that in other species, besides morphophysiological factors, there are intrinsic problems to the spermatozoon such as less chromatin condensation that can lead to fertility problems that in most cases are not diagnosised. The purpose of this work was the adaptation of techniques of chromatin evaluation for chicken (Gallus Gallus, Linnaeus, 1758) which have already been described in other species, correlating the chromatin alterations with the morphological alterations and the fertility. Fowl semen samples with different levels of fertility were used for test different methods to identify chromatin alterations using toluidine blue and acridine orange die. Evaluations have shown that smears of fresh semen with posterior fixation generate artfacts that change the head shape and the chromatin integrity not being indicated in methods of fertility evaluation. Although all the tested methods have presented methodology imperfections and some certain degree of subjectivity, the method that has generated best results was the mixture of one semen drop conserved in buffered phormol and a drop of acridine orange on microscopy slide with posterior staining and observation in fluorescence microscopy with blue excitation filter. Through this method it was verified that alterations in the chromatin condensation of fowl spermatozoa are generally not followed by morphological alterations and that generally the morphologic alterations of fowl spermatozoa are generally followed by alterations in chromatin condensation. This is an important factor in the determination of the fowl fertility.

Morphometric features and chromatin condensation abnormalities evaluated by toluidine blue staining in bull spermatozoa

In this article, we investigated the correlation between the morphometric features of sperm heads and abnormalities in chromatin condensation using computational image analysis of sperm smears stained with toluidine blue. The chromatin structure was characterized in terms of the average gray-level intensity and the corresponding coefficient variation based on images of the head. The geometry of the sperm heads was determined based on several morphometric measurements, including the area, perimeter, width, length, width: length ratio, ellipticity, shape factor, Fourier descriptor and symmetry. The correlation between the chromatin structure and the geometrical properties was evaluated. The correlation coefficients between the morphometric features and chromatin alterations varied substantially from sample to sample, although a normal morphology was found in heads with or without abnormalities in chromatin condensation. Variations in the abnormalities in chromatin condensation influenced the head morphology in different ways. The finding that spermatozoa with chromatin abnormalities did not necessarily show morphological alterations demonstrates the need for the morphological analysis of sperm to be followed by an evaluation of chromatin structure.