Integridade e funcionalidade dos espermatozóides ovinos submetidos à criopreservação após a incorporação de colesterol, desmosterol, ácido oléico-linoléico e alfa-lactoalbumina

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Qualidade espermática de sêmen de cães naturalmente infectados por Leishmania sp:

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Efeitos do processo de seleção do sexo por centrifugação em gradiente de densidade na qualidade de espermatozódes bovinos

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Physiological basis of salt stress tolerance in rice expressing the antiapoptotic gene SfIAP

Programmed cell death-associated genes, especially antiapoptosis-related genes have been reported to confer tolerance to a wide range of biotic and abiotic stresses in dicotyledonous plants such as tobacco (Nicotiana tabacum L.) and tomato (Solanum lycopersicum L.). This is the first time the antiapoptotic gene SfIAP was transformed into a monocotyledonous representative: rice (Oryza sativa L.). Transgenic rice strains expressing SfIAP were generated by the Agrobacterium-mediated transformation method and rice embryogenic calli, and assessed for their ability to confer tolerance to salt stress at both the seedling and reproductive stages using a combination of molecular, agronomical, physiological and biochemical techniques. The results show that plants expressing SfIAP have higher salt tolerance levels in comparison to the wild-type and vector controls. By preventing cell death at the onset of salt stress and maintaining the cell membrane’s integrity, SfIAP transgenic rice plants can retain plant water status, ion homeostasis, photosynthetic efficiency and growth to combat salinity successfully.

Efeito da centrifugação sobre a qualidade do sêmen canino

Avaliou-se o efeito da centrifugação sobre a viabilidade do sêmen canino e compararam-se três meios de diluição pré-centrifugação. Utilizaram-se 10 ejaculados completos de 10 cães que, após a avaliação inicial, foram divididos em quatro porções (grupos). Uma das amostras, não centrifugada, formou o grupo-controle; as outras foram diluídas em três diferentes meios e centrifugadas a 800 x g por 15 minutos, formando os grupos: CPSA - constituído por sêmen centrifugado em plasma seminal autólogo; CLG - sêmen centrifugado em meio à base de leite desnatado e glicose (LG); e CPer- sêmen centrifugado em gradientes de Percoll (45% e 90%). Após a centrifugação e a eliminação do sobrenadante, procedeu-se à ressuspensão de todas as porções do ejaculado em LG e à imediata avaliação quanto à motilidade, vigor, aglutinação espermática e integridade das membranas espermáticas. Todas as suspensões foram, então, incubadas a 37ºC por 30 minutos e reavaliadas. O processo de centrifugação não causou danos aos espermatozoides e a centrifugação em meio LG melhorou a viabilidade espermática.

Viabilidade e fertilidade de sêmen ovino criopreservado com glicerol e dimetilacetamida acrescido de trealose EDTA

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Perfil lipídico da membrana plasmática e alterações morfofuncionais de esperamatozóides de garanhões resistentes e sensíveis à criopreservação

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Efeito da adição de butil-hidroxitolueno nos meios de refrigeração e congelação sobre a viabilidade espermática de equinos

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Evaluación de tolerancia al NaCl en cultivares viníferas y portainjertos híbridos del género Vitis

El objetivo fue evaluar la supervivencia al riego con soluciones de NaCl en 28 genotipos del género Vitis para identificar su grado de tolerancia salina. Se estudiaron 11 híbridos de Vitis americanos y 17 variedades de Vitis vinifera. Plantas de 1 y 2 años se regaron durante 66 días con una solución 100 mM de NaCl. Para clasificar las variedades de acuerdo con su grado de tolerancia a la salinidad, se calculó el día en que el 25% de la población había muerto (1er cuartil de la muestra). Se consideraron como genotipos sensibles aquellos en que al menos 25% había muerto antes del día 30 (3309 Couderc, 161-49 Couderc, Fercal, Freedom, 1103 Paulsen, 99 Richter, SO4, Torrontés Sanjuanino), como poco tolerantes cuando al menos 25% murió entre los días 30 y 60 (Cereza, Colombard, Criollas Blanca y Ballista, Palomino, 110 Richter, 140 Ruggeri, Syrah, Torrontés Riojano) y como tolerantes cuando el 75% o menos sobrevivió más de 60 días (101-14 Millardet-Grasset, 196-17 Castel, Criollas Chica y Sanjuanina, Moscatel de Alejandría, Pedro Giménez). Aun en los genotipos más tolerantes cuando las plantas crecieron bajo salinidad, la integridad de membranas celulares se redujo un 17% y el contenido de clorofila total disminuyó un 52%.

The influence of matrix integrity on stress-fiber remodeling in 3D.

Matrix anisotropy is important for long term in vivo functionality. However, it is not fully understood how to guide matrix anisotropy in vitro. Experiments suggest actin-mediated cell traction contributes. Although F-actin in 2D displays a stretch-avoidance response, 3D data are lacking. We questioned how cyclic stretch influences F-actin and collagen orientation in 3D. Small-scale cell-populated fibrous tissues were statically constrained and/or cyclically stretched with or without biochemical agents. A rectangular array of silicone posts attached to flexible membranes constrained a mixture of cells, collagen I and matrigel. F-actin orientation was quantified using fiber-tracking software, fitted using a bi-model distribution function. F-actin was biaxially distributed with static constraint. Surprisingly, uniaxial cyclic stretch, only induced a strong stretch-avoidance response (alignment perpendicular to stretching) at tissue surfaces and not in the core. Surface alignment was absent when a ROCK-inhibitor was added, but also when tissues were only statically constrained. Stretch-avoidance was also observed in the tissue core upon MMP1-induced matrix perturbation. Further, a strong stretch-avoidance response was obtained for F-actin and collagen, for immediate cyclic stretching, i.e. stretching before polymerization of the collagen. Results suggest that F-actin stress-fibers avoid cyclic stretch in 3D, unless collagen contact guidance dictates otherwise.

The influence of matrix integrity on stress-fiber remodeling in 3D

Matrix anisotropy is important for long term in vivo functionality. However, it is not fully understood how to guide matrix anisotropy in vitro. Experiments suggest actin-mediated cell traction contributes. Although F-actin in 2D displays a stretch-avoidance response, 3D data are lacking. We questioned how cyclic stretch influences F-actin and collagen orientation in 3D. Small-scale cell-populated fibrous tissues were statically constrained and/or cyclically stretched with or without biochemical agents. A rectangular array of silicone posts attached to flexible membranes constrained a mixture of cells, collagen I and matrigel. F-actin orientation was quantified using fiber-tracking software, fitted using a bi-model distribution function. F-actin was biaxially distributed with static constraint. Surprisingly, uniaxial cyclic stretch, only induced a strong stretch-avoidance response (alignment perpendicular to stretching) at tissue surfaces and not in the core. Surface alignment was absent when a ROCK-inhibitor was added, but also when tissues were only statically constrained. Stretch-avoidance was also observed in the tissue core upon MMP1-induced matrix perturbation. Further, a strong stretch-avoidance response was obtained for F-actin and collagen, for immediate cyclic stretching, i.e. stretching before polymerization of the collagen. Results suggest that F-actin stress-fibers avoid cyclic stretch in 3D, unless collagen contact guidance dictates otherwise. © 2012 Elsevier Ltd.

Towards integrated anti-microbial capabilities: Novel bio-fouling resistant membranes by high velocity embedment of silver particles

Biofilm formation on membranes during water desalination operation and pre-treatments limits performance and causes premature membrane degradation. Here, we apply a novel surface modification technique to incorporate anti-microbial metal particles into the outer layer of four types of commercial polymeric membranes by cold spray. The particles are anchored on the membrane surface by partial embedment within the polymer matrix. Although clear differences in particle surface loadings and response to the cold spray were shown by SEM, the hybrid micro-filtration and ultra-filtration membranes were found to exhibit excellent anti-bacterial properties. Poly(sulfone) ultra-filtration membranes were used as for cross-flow filtration of Escherichia coli bacteria solutions to investigate the impact of the cold spray on the material[U+05F3]s integrity. The membranes were characterized by SEM-EDS, FT-IR and TGA and challenged in filtration tests. No bacteria passed through the membrane and filtrate water quality was good, indicating the membranes remained intact. No intact bacteria were found on hybrid membranes, loaded with up to 15. wt% silver, indicating the treatment was lysing bacteria on contact. However, permeation of the hybrid membranes was found to be reduced compared to control non-modified poly(sulfone) membranes due to the presence of the particles across the membrane material. The implementation of cold spray technology for the modification of commercial membrane products could lead to significant operational savings in the field of desalination and water pre-treatments.

Evaluation of plasma membrane integrity of frozen-thawed goat spermatozoa with or without seminal plasma

The aim of the present work was to evaluate plasma membrane integrity, motility, vigor and morphology of fresh and frozen goat spermatozoa with or without seminal plasma. Semen samples were diluted in Tris solution, before and after thawing, with a combination of carboxifluorescein diacetate and propidium iodide. The results showed differences (P < 0.01) for motility and minor defects in the presence or absence of seminal plasma, for both fresh and frozen samples. Periods of collection had a significant effect on motility, probably due to changes in the photoperiod. Plasma membrane integrity was significantly reduced by the freezing process, whether seminal plasma was present or absent. In conclusion, removal of seminal plasma decreased motility and vigor rates in frozen samples. The photoperiod probably decreased the testosterone level, contributing negatively to the high percentage of sperm abnormalities, mainly damaged membranes. The use of fluorescent probes allowed a better estimation of the percentage of functional cells, instead of only estimating the percentage of motile cells or morphology defects. © 2001 Elsevier Science B.V. All rights reserved.

Effect of manganese supplementation on the membrane integrity and the mitochondrial potential of the sperm of grazing Nelore bulls

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Principais marcadores utilizados na avaliação da qualidade do sêmen de animais domésticos

Methods of semen cryopreservation allow changes in spermatic cells, such as damage in plasma and acrossomal membrane and modifications in mitochondrial function due to a disorder in the lipidic bilayer. For effective oocyte fertilization, spermatozoa require functional competent membranes, and intact organelles, acrosome and DNA. However, most laboratory methods used to evaluate semen quality are not highly correlated with fertilizing capacity. The discovery of a variety of fluorochromes and compounds conjugated to fluorescent probes has enabled an accurate assessment of the viability, integrity and function of spermatozoa. Among the most used probes that label the various compartments of the sperm cell there are the membrane impermeable fluorescent dyes to test the membrane integrity, as well as acylated dyes that pass the intact membrane. For the acrossomal integrity the most commonly used method is lectins labeled by a fluorescent probe. The acrosome reaction and spermatic capacitation is detected by the evaluation of membrane architecture and disorder of lipids in plasma membrane. Mitochondrial function can be determined using markers for their aerobic activity. The DNA status of spermatozoa has been determined using the metachromatic properties of Acridine Orange, and the DNA fragmentation can also be assessed by TUNEL assay. Finally, DNA condensation is analyzed using a single cell DNA gel electrophoresis assay that indicates DNA compactation. This monograph aims to compile the various tests used to detect damaged spermatozoa under cryopreservation methods, searching for improve the predictive value of semen analysis with the intention of a successful conception