Effects of discontinuous Percoll gradient centrifugation on the quality of bovine spermatozoa evaluated with computer-assisted semen analysis and fluorescent probes association

The objective of this study was to evaluate the quality of bovine frozen-thawed sperm cells after Percoll gradient centrifugation. Frozen semen doses were obtained from six bulls of different breeds, including three taurine and three Zebu animals. Four ejaculates per bull were evaluated before and after discontinuous Percoll gradient centrifugation. Sperm motility was assessed by computer-assisted semen analysis and the integrity of the plasma and acrosomal membranes, as well as mitochondrial function, were evaluated using a combination of fluorescent probes propidium iodide, fluorescein isothiocyanate-conjugated Pisum sativum agglutinin and 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine iodide. The procedure of Percoll gradient centrifugation increased the percentage of total and progressive sperm motility, beat frequency, rectilinear motility, linearity and rapidly moving cells. In addition, the percentage of cells with intact plasma membrane and mitochondrial membrane potential was increased in post-centrifugation samples. However, the percentage of sperm cells with intact acrosomal membrane was markedly reduced. The method used selected the motile cells with intact plasma membrane and higher mitochondrial functionality in frozen-thawed bull semen, but processing, centrifugation and/or the Percoll medium caused damage to the acrosomal membrane.

Computer game-based and traditional learning method: a comparison regarding students’ knowledge retention

Abstract Background Educational computer games are examples of computer-assisted learning objects, representing an educational strategy of growing interest. Given the changes in the digital world over the last decades, students of the current generation expect technology to be used in advancing their learning requiring a need to change traditional passive learning methodologies to an active multisensory experimental learning methodology. The objective of this study was to compare a computer game-based learning method with a traditional learning method, regarding learning gains and knowledge retention, as means of teaching head and neck Anatomy and Physiology to Speech-Language and Hearing pathology undergraduate students. Methods Students were randomized to participate to one of the learning methods and the data analyst was blinded to which method of learning the students had received. Students’ prior knowledge (i.e. before undergoing the learning method), short-term knowledge retention and long-term knowledge retention (i.e. six months after undergoing the learning method) were assessed with a multiple choice questionnaire. Students’ performance was compared considering the three moments of assessment for both for the mean total score and for separated mean scores for Anatomy questions and for Physiology questions. Results Students that received the game-based method performed better in the pos-test assessment only when considering the Anatomy questions section. Students that received the traditional lecture performed better in both post-test and long-term post-test when considering the Anatomy and Physiology questions. Conclusions The game-based learning method is comparable to the traditional learning method in general and in short-term gains, while the traditional lecture still seems to be more effective to improve students’ short and long-term knowledge retention.

The identification of the pillars of education in the class comprehensiveness in healthcare

The objective of the present study was to verify, based on the analysis of student portfolio narratives, if the four pillars of education were approached in the class "Comprehensiveness in health care", part of the integrated curriculum of the Baccalaureate in Nursing Program of the University of Sao Paulo at Ribeirao Preto College of Nursing. A qualitative, documental study was performed using 46 portfolios constructed during the classes. Data collection was performed using an assessment tool that contained items addressing cognitive and affective dimensions. The data were submitted to thematic categorical analysis using the pillars of education as predefined categories. The results show that the pillars of education were, apparently, included in the class. Despite the present study findings, no evidence was found that the expected competencies were actually discussed among students and faculty, according to the records regarding the evaluations of each pedagogical cycle of the studied class.

Quality characteristics and fertilizing ability of ram sperm subpopulations separated by partition in an aqueous two-phase system

Centrifugal countercurrent distribution (CCCD) in an aqueous two-phase system (TPS) is a resolute technique revealing sperm heterogeneity and for the estimation of the fertilizing potential of a given semen sample. However, separated sperm subpopulations have never been tested for their fertilizing ability yet. Here, we have compared sperm quality parameters and the fertilizing ability of sperm subpopulations separated by the CCCD process from ram semen samples maintained at 20 degrees C or cooled down to 5 degrees C. Total and progressive sperm motility was evaluated by computer-assisted analysis using a CASA system and membrane integrity was evaluated by flow cytometry by staining with CFDA/Pl. The capacitation state, staining with chlortetracycline, and apoptosis-related markers, such as phosphatidylserine (PS) translocation detected with Annexin V. and DNA damage detected by the TUNEL assay, were determined by fluorescence microscopy. Additionally, the fertilizing ability of the fractionated subpopulations was comparative assessed by zona binding assay (ZBA). CCCD analysis revealed that the number of spermatozoa displaying membrane and DNA alterations was higher in samples chilled at 5 degrees C than at 20 degrees C. which can be reflected in the displacement to the left of the CCCD profiles. The spermatozoa located in the central and right chambers (more hydrophobic) presented higher values (P<0.01) of membrane integrity, lower PS translocation (P<0.05) and DNA damage (P<0.001) than those in the left part of the profile, where apoptotic markers were significantly increased and the proportion of viable non-capacitated sperm was reduced. We have developed a new protocol to recover spermatozoa from the CCCD fractions and we proved that these differences were related with the fertilizing ability determined by ZBA, because we found that the number of spermatozoa attached per oocyte was significantly higher for spermatozoa recovered from the central and right chambers, in both types of samples. This is the first time, to our knowledge that sperm recovered from a two-phase partition procedure are used for fertilization assays. These results open up new possibilities for using specific subpopulations of sperm for artificial insemination or in vitro fertilization, not only regarding better sperm quality but also certain characteristics such as subpopulations enriched in spermatozoa bearing X or Y chromosome that we have already isolated or any other feature. (C) 2011 Elsevier B.V. All rights reserved.

Análise das distorções em leituras angulares de imagens fotográficas

CONTEXTUALIZAÇÃO: A biofotogrametria é uma técnica difundida na área da saúde e, apesar dos cuidados metodológicos, há distorções nas leituras angulares das imagens fotográficas. OBJETIVO: Mensurar o erro das medidas angulares em imagens fotográficas com diferentes resoluções digitais em um objeto com ângulos pré-demarcados. MÉTODOS: Utilizou-se uma esfera de borracha com 52 cm de circunferência. O objeto foi previamente demarcado com ângulos de 10º, 30º, 60º e 90º, e os registros fotográficos foram realizados com o eixo focal da câmera a três metros de distância e perpendicular ao objeto, sem utilização de zoom óptico e com resolução de 3, 5 e 10 Megapixels (Mp). Todos os registros fotográficos foram armazenados, e os valores angulares foram analisados por um experimentador previamente treinado, utilizando o programa ImageJ. As aferições das medidas foram realizadas duas vezes, com intervalo de 15 dias entre elas. Posteriormente, foram calculados os valores de acurácia, erro relativo e em graus, precisão e Coeficiente de Correlação Intraclasse (ICC). RESULTADOS: Quando analisado o ângulo de 10º, a média da acurácia das medidas foi maior para os registros com resolução de 3 Mp em relação às resoluções de 5 e 10 Mp. O ICC foi considerado excelente para as três resoluções de imagem analisadas e, em relação aos ângulos analisados nos registros fotográficos, pôde-se verificar maior acurácia, menor erro relativo e em graus e maior precisão para o ângulo de 90º, independentemente da resolução da imagem. CONCLUSÃO: Os registros fotográficos realizados com a resolução de 3 Mp proporcionaram medidas de maiores valores de acurácia e precisão e menores valores de erro, sugerindo ser a resolução mais adequada para gerar imagem de ângulos de 10º e 30º.