RESUMEN
The goal of the present study was to determine the effect of sole horn thickness (SHT) and sole horn hardness (SHD) on ultrasonographic visualization of sole structures in the inner and outer claws of 150 Holstein-Friesian cows, and to evaluate different ultrasound frequencies for this purpose. Ultrasonographic views of the sole structure were considered complete when 3 echogenic lines, representing the ventral surface of the sole horn, the borders of the sole horn and soft-tissue layer, and the ventral surface of the distal phalanx, were seen. The proportion of complete ultrasonographic views of the sole structures, designated as the ultrasonographic visualization proportion (UVP), and the measurement errors of SHT were evaluated by comparing images from computed tomography (CT) and ultrasonography. The latter images were generated using 3 different probes, frequencies of 6.5 and 5.0 MHz, and 2 different ultrasound machines (#1 and #2) to assess the apex, middle, and heel regions of the claws. The UVP were 60.8 to 77.9% for the 6.5-MHz probe in ultrasound machine #1 (probe A), which were lower than those (>90%) for both the 5.0-MHz probe in ultrasound machine #1 (probe B) and the 5.0-MHz probe in ultrasound machine #2 (probe C). The UVP was significantly lower in claws with an SHD ≥50 units than in claws with an SHD <40 or 40 to <50 units (UVP: 77.1% compared with 93.7 and 91.4%, respectively) when measured with probe B. In claws with an SHT <10 mm, the UVP was significantly lower when SHD was ≥50 units compared with <40 or 40 to >50 units; the values were 69.0% versus 91.3 and 85.9%, respectively, for probe A, and 89.7% versus 100 and 100%, respectively, for probe B. When SHT were measured by either probes A or B in ultrasound machine #1, the proportions of claws in which ultrasonographic values were within a ±1 mm range compared with the values obtained by CT were 84.9 to 91.3% for CT-determined SHT <5 mm, 66.7 to 71.9% for CT-determined SHT 5 to <7 mm, 28.9 to 51.2% for CT-determined SHT 7 to <10 mm, and 6.2 to 19.7% for CT-determined SHT ≥10 mm. The data indicated that increased SHT was associated with a decrease in ultrasonographic measurement accuracy. In claws with an SHT <5 mm, the high proportion of ultrasonographic values that were accurate within a ±1 mm range suggests that this imaging modality would be useful in cows with thin soles.
Asunto(s)
Bovinos/anatomía & histología , Pezuñas y Garras/diagnóstico por imagen , Tomografía Computarizada por Rayos X/veterinaria , Ultrasonografía/veterinaria , Animales , Femenino , Dureza , Reproducibilidad de los ResultadosRESUMEN
Nuclear factor kappa B (NF-κB) signaling plays critical roles in many physiological and pathological processes, including regulating organogenesis. Down-regulation of NF-κB signaling during development results in hypohidrotic ectodermal dysplasia. The roles of NF-κB signaling in tooth development, however, are not fully understood. We examined mice overexpressing IKKß, an essential component of the NF-κB pathway, under keratin 5 promoter (K5-Ikkß). K5-Ikkß mice showed supernumerary incisors whose formation was accompanied by up-regulation of canonical Wnt signaling. Apoptosis that is normally observed in wild-type incisor epithelium was reduced in K5-Ikkß mice. The supernumerary incisors in K5-Ikkß mice were found to phenocopy extra incisors in mice with mutations of Wnt inhibitor, Wise. Excess NF-κB activity thus induces an ectopic odontogenesis program that is usually suppressed under physiological conditions.
Asunto(s)
Incisivo/embriología , FN-kappa B/fisiología , Odontogénesis/fisiología , Germen Dentario/embriología , Proteínas Adaptadoras Transductoras de Señales , Ameloblastos/citología , Amelogenina/análisis , Animales , Apoptosis/fisiología , Proteínas Morfogenéticas Óseas/genética , Esmalte Dental/citología , Epitelio/embriología , Proteínas Hedgehog/fisiología , Quinasa I-kappa B/fisiología , Imagenología Tridimensional/métodos , Incisivo/anomalías , Queratina-15/genética , Ratones , Ratones Mutantes , Microrradiografía/métodos , Mutación/genética , Receptores Patched , Fenotipo , Regiones Promotoras Genéticas/genética , Receptores de Superficie Celular/fisiología , Germen Dentario/anomalías , Diente Supernumerario/etiología , Diente Supernumerario/genética , Vía de Señalización Wnt/genética , Vía de Señalización Wnt/fisiología , Microtomografía por Rayos X/métodosRESUMEN
Our study was conducted to assess the follicular development and availability of sound ovarian oocytes for in vitro production (IVP) of embryos in pre-pubertal cats. The relationship between body and ovarian weight was examined in 93 cats. The results revealed that ovarian weight rapidly increased until 100 days of estimated age. By histological evaluation of ovaries obtained from 11 pre-pubertal cats with estimated age of <20, 20-40 and 100-120 days, it was clarified that the increase in ovarian weight during kitten growth accompanied the increase in the number and size of antral follicles. The follicular diameter and percentage of normal oocytes in secondary/antral follicles also increased as estimated age (body weight) increased. The oocytes obtained from pre-pubertal cats with 100-120 days of estimated age were used for IVP of embryos. The results showed that the success rates of in vitro maturation, in vitro fertilization and development to blastocysts after in vitro culture in pre-pubertal cats were lower than in sexually mature cats. However, the percentage of blastocysts based on the cleaved embryos and cell number of blastocysts in pre-pubertal cats were comparable to those in mature cats. In conclusion, these results suggest that the ovaries of pre-pubertal cats with ≥100 days of age contain oocytes with in vitro developmental competence to blastocysts.
Asunto(s)
Gatos/fisiología , Oocitos/citología , Oocitos/fisiología , Folículo Ovárico/fisiología , Maduración Sexual/fisiología , Animales , Peso Corporal , Femenino , Fertilización In Vitro , MeiosisRESUMEN
The objective of this study was to develop a preservation method for canine sperm using microencapsulation. Pooled ejaculates from three beagles (Canis familiaris) were extended in egg yolk Tris extender and were encapsulated in gel (alginate only) or polycation (poly-L-lysine membrane bound) microcapsules at 0.75% and 1.0% alginate concentration. In Experiment 1, characteristics of microcapsule and microencapsulated sperm were evaluated during chilling storage for 48 h. Gel microcapsules at 0.75% alginate concentration had a teardrop-like structure with fragility, whereas those at 1.0% alginate had a solid spherical structure. In all groups, diameter of the microcapsules increased with duration of storage (P<0.05). Alginate concentration did not affect the sperm recovery rate from microcapsules. Total average recovery rate of sperm from polycation microcapsules was lower than that of gel microcapsules (P<0.05). Progressive motility of polycation microencapsulated sperm and unencapsulated sperm (control) was higher than that of the gel microencapsulated sperm, both at 0.75% and 1.0% alginate concentration (P<0.05), although viability of sperm was similar among the three groups. In Experiment 2, to evaluate the sperm longevity after chilling storage, sperm were microencapsulated in polycation microcapsules at 1.0% alginate concentration, stored at 4 degrees C for 0, 1, 4, and 7 d, and then cultured at 38.5 degrees C for 0, 6, and 24h. Progressive motility and viability of microencapsulated sperm were higher than those of unencapsulated spermatozoa at 0 to 24h of culture after 4 and 7 d of chilling storage (P<0.05). In conclusion, polycation microencapsulation at 1.0% alginate concentration can be successfully applied for chilling storage of canine sperm by maintaining motility and viability for up to 7 d.
Asunto(s)
Frío , Perros , Composición de Medicamentos/métodos , Preservación de Semen/métodos , Espermatozoides , Animales , Cápsulas/química , Supervivencia Celular/fisiología , Composición de Medicamentos/efectos adversos , Masculino , Tamaño de la Partícula , Poliaminas/química , Polielectrolitos , Refrigeración/métodos , Preservación de Semen/instrumentación , Recuento de Espermatozoides , Motilidad Espermática/fisiología , Espermatozoides/citologíaRESUMEN
This study was conducted to improve in vitro production of embryos from domestic cats using TCM-199 as an IVM medium. The time sequence of nuclear maturation and the optimal timing of in vitro insemination were examined. Most oocytes were at the germinal vesicle stage immediately after collection; however, 8.3% had already resumed meiosis before IVM culture. After 30 h of IVM culture, the percentage of oocytes at metaphase II (MII) reached a peak (75.5%) and did not change (P>0.05) from 30 to 48 h after IVM culture. The percentage of oocytes with two pronuclei was higher (P<0.05) for oocytes matured for 30 and 36 h (38.2 and 33.0%, respectively) than for those after IVM culture for only 24 h (18.5%). Total sperm penetration rate was highest (P<0.05) for oocytes that had been matured for 30 h (46.1%). After 30 h of IVM and 18 h of IVF culture, 66.3 and 24.8% of inseminated oocytes had cleaved and developed to the blastocyst stage, respectively. We concluded that IVM of feline oocytes for 30 h in TCM-199 resulted in optimal nuclear maturation and sperm penetration.
Asunto(s)
Blastocisto/fisiología , Gatos/fisiología , Fertilización In Vitro/veterinaria , Oocitos/fisiología , Animales , Técnicas de Cultivo de Célula/métodos , Técnicas de Cultivo de Célula/veterinaria , Femenino , Fertilización In Vitro/métodos , Masculino , Embarazo , Factores de TiempoRESUMEN
AIMS: Glypican 3 (GPC3) is a cell surface heparan sulphate proteoglycan expressed specifically in the fetal liver and malignant neoplasms of hepatocyte lineage. The aim was to evaluate the significance of GPC3 in alpha-fetoprotein (AFP)-producing gastric carcinoma (GC) and other forms of GC. METHODS AND RESULTS: We immunohistochemically evaluated GPC3 expression in representative cases of AFP-producing GC and in a tissue microarray of a consecutive series of GCs with other markers of hepatocyte lineage (AFP, PIVKA-II and hepatocyte antigen, HEP). In a series of 10 cases of AFP-producing GC, we observed immunohistochemical positivity for GPC3, PIVKA-II and HEP in 10, three and three cases in components with a hepatoid pattern and in nine, two and five cases in components with a non-hepatoid pattern, respectively. In a series of 118 cases of GC, we observed positivity for AFP, GPC3, PIVKA-II and HEP in one (0.8%), four (3.4%), six (5.1%) and 26 cases (22%), respectively. GPC3 was observed concurrently with AFP and discordantly with PIVKA-II and HEP. GPC3 positivity was clearly stronger in a larger area compared with immunoreactivity for AFP. CONCLUSIONS: GPC3 is a sensitive marker for AFP-producing GC and its hepatoid component and is therefore useful to identify this aggressive subgroup of GC.
Asunto(s)
Adenocarcinoma/metabolismo , Glipicanos/metabolismo , Neoplasias Gástricas/metabolismo , alfa-Fetoproteínas/metabolismo , Adenocarcinoma/patología , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/metabolismo , Femenino , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Neoplasias Gástricas/patología , Análisis de Matrices TisularesRESUMEN
The objective of this study was to collect oocytes from ovaries of bitches with pyometra and to characterize the quality of the oocytes recovered. In 10 of 12 cases of pyometra, follicles with a diameter of 500 microm to 1mm were observed in the ovaries. A total of 710 oocytes were collected from 10 bitches by puncturing individual follicles after slicing the ovarian tissues. Oocyte recovery was successful from a bitch with severe clinical signs of pyometra. Of the oocytes collected, 53.5% were surrounded by > or =2 layers of cumulus cells, and 55.0% of these cumulus-oocyte complexes (COCs) had a darkly pigmented ooplasm >110 microm in diameter (large-dark COCs). The number of large-dark COCs per bitch varied from 1 to 72. A germinal vesicle with fine filaments of chromatin (Type A) was observed in 51.8% (range 21.1-100%) of the oocytes of large-dark COCs. Out of 50 oocytes cultured for 72 h, 6.0% developed to Metaphase II. In conclusion, there were many follicles with a diameter of 500 microm to 1mm in ovaries of bitches with pyometra, and many oocytes recovered from these follicles underwent meiotic maturation in vitro. The number of oocytes and COCs, and the morphological quality of the germinal vesicles varied among individual bitches.
Asunto(s)
Enfermedades de los Perros/patología , Enfermedades de los Perros/fisiopatología , Oocitos/fisiología , Folículo Ovárico/citología , Enfermedades Uterinas/veterinaria , Animales , Núcleo Celular/ultraestructura , Células Cultivadas , Perros , Femenino , Meiosis , Oocitos/ultraestructura , Supuración , Enfermedades Uterinas/patología , Enfermedades Uterinas/fisiopatologíaRESUMEN
The objective was to characterize the separation of canine epididymal spermatozoa on a Percoll gradient. Epididymal spermatozoa were overlaid on a 45 and 90% discontinuous Percoll gradient and centrifuged at 700 x g for 20 min. The Percoll column was separated into six fractions (top to bottom, A-F) after centrifugation. Fractions A-C contained few spermatozoa. Spermatozoa with bent or folded tails and a large amount of granular debris were observed in Fraction B. Fraction D contained many nonmotile spermatozoa, erythrocytes and round epithelial cells. Spermatozoa in Fraction E had significantly lower motility than those in the initial layer. Spermatozoa in Fraction F had motility similar to those before separation. Fraction F contained 40.6% of the motile spermatozoa layered and 67.5% of all motile spermatozoa recovered. There was no significant difference between Fraction F and the initial layer in sperm membrane integrity. In the sperm-oocyte penetration assay, spermatozoa from Fraction F had a significantly higher penetration rate into the immature homologous oocytes than those from Fraction E. Although the recovery rate of the motile spermatozoa was low, the canine epididymal spermatozoa with motility, membrane integrity and penetrating capability could be separated by two-layer discontinuous Percoll gradient centrifugation.
Asunto(s)
Separación Celular/veterinaria , Centrifugación por Gradiente de Densidad , Perros , Epidídimo/citología , Espermatozoides/citología , Animales , Separación Celular/métodos , Femenino , Masculino , Oocitos , Povidona , Dióxido de Silicio , Motilidad Espermática , Interacciones Espermatozoide-Óvulo , Espermatozoides/anomalíasRESUMEN
The aim of this study is to clarify influence of cold storage of deer epididymides on sperm quality and suitability for cryopreservation. The epididymides were obtained postmortem from sika deer during the breeding season. When epididymides were removed 8-12h postmortem and stored at 4 degrees C for 1-4 days, the collected spermatozoa showed low motility (6.4%). When spermatozoa were collected from epididymides removed within 4h postmortem, sperm motility and viability were 71.8 and 82.4%, respectively. Sperm motility decreased as prolongation of the storage period of the epididymides continued up to 7 days, but sperm viability was not affected. Pyknosis of the epithelial cells and their release into the lumen were observed in the stored epididymides. Epididymal spermatozoa frozen on Day 0 showed 58.1% motility and 83.2% viability. Motility of the frozen-thawed spermatozoa from epididymides stored at 4 degrees C for 1 day (41.9%) was similar to that of nonfrozen spermatozoa from epididymides stored for 4 days (41.8%). These results suggest that refrigeration of deer epididymides or cryopreservation of spermatozoa from refrigerated epididymides can be used for assisted reproductive techniques when epididymal spermatozoa cannot be collected immediately.
Asunto(s)
Criopreservación/veterinaria , Ciervos/fisiología , Epidídimo/citología , Preservación de Semen/veterinaria , Espermatozoides/fisiología , Animales , Frío , Criopreservación/métodos , Criopreservación/normas , Masculino , Preservación de Semen/métodos , Preservación de Semen/normas , Motilidad Espermática , Factores de TiempoRESUMEN
Facultatively psychrophilic alkaliphilic strains were isolated from seawater obtained off the coast of Rumoi, Hokkaido, Japan. They were Gram-negative, aerobic straight rods with polar flagella. The isolates were catalase- and oxidase-positive and able to grow at 4 degrees C, but not at 40 degrees C. They produced acid from D-glucose under aerobic conditions. The isolates reduced nitrate to nitrite and hydrolysed casein and gelatin, but not starch or DNA. NaCl was required for growth at pH 10 but was not required at neutral pH. The major isoprenoid quinone was ubiquinone-9 (Q-9) and the DNA G+C content was 62.3-63.2 mol%. The whole-cell fatty acids mainly consisted of C16:0, C16:1(9c) and C18:1(9c), with 3-OH C10:0 and 3-OH C12:0 as the hydroxyl fatty acids. A larger amount of trans-unsaturated fatty acid, C16:1(9t) was observed when the cells were grown at pH 7 compared to when cells were grown at pH 10. Phylogenetic analysis based on 16S rRNA gene sequencing indicated that the bacteria are members of the genus Pseudomonas. Analysis of DNA-DNA relatedness data with several close phylogenetic neighbours revealed a low level of hybridization (less than 61%). On the basis of phenotypic characteristics, phylogenetic analysis and DNA-DNA relatedness data, it is concluded that these isolates represent a separate new species. Accordingly, the name Pseudomonas alcaliphila is proposed. The type strain is AL15-21T (= JCM 10630T = IAM 14884T).
Asunto(s)
Pseudomonas/clasificación , Microbiología del Agua , Álcalis , Técnicas de Tipificación Bacteriana , Carbohidratos , ADN Ribosómico , Ácidos Grasos/análisis , Datos de Secuencia Molecular , Hibridación de Ácido Nucleico , Pseudomonas/citología , Pseudomonas/genética , Pseudomonas/aislamiento & purificación , ARN Ribosómico 16S , Agua de Mar , Terminología como AsuntoRESUMEN
Facultatively alkaliphilic Bacillus sp. strain YN-2000 was isolated from an indigo ball. Although the strain has been extensively investigated as a representative strain of alkaliphilic bacillus, its taxonomic position is not yet known. Morphological, biochemical, and physiological characteristics and chemotaxonomic properties indicated that the strain was closely related to Bacillus cohnii; this was confirmed by the high homology of the 16S rRNA sequence and the construction of a phylogenetic tree on the basis of the 16S rRNA sequence and DNA-DNA relatedness data. Strain YN-2000 contained a larger amount of unsaturated fatty acids compared with Bacillus subtilis and the obligate alkaliphile, Bacillus alcalophilus, regardless of its culture pH. When the cells were grown at pH 10, the unsaturated fatty acid content and anteiso-/iso-branched fatty acid ratio became lower than those at pH 7. This result suggests that membrane fluidity decreases when the cells are grown at pH 10 compared to those of pH 7. In the cells of strain YN-2000 grown at pH 10, the cell-surface aspect was rougher, the cell shape was longer, and the cell-surface layer was thicker compared with those of the cells grown at pH 7. The cell-surface structural change might be related to adaptation to an alkaline environment.
Asunto(s)
Bacillus/química , Bacillus/ultraestructura , Pared Celular/química , Ácidos Grasos/análisis , Álcalis , Bacillus/clasificación , Bacillus/crecimiento & desarrollo , Pared Celular/ultraestructura , Clasificación , Medios de Cultivo/química , ADN Bacteriano/genética , Ácidos Grasos/química , Concentración de Iones de Hidrógeno , Microscopía Electrónica de Rastreo , Datos de Secuencia Molecular , Hibridación de Ácido Nucleico , Fenotipo , Filogenia , ARN Ribosómico 16S/genética , Análisis de Secuencia de ARNRESUMEN
Effects of the embryo retrieval stages and addition of glutathione (GSH) on post-thaw development of mouse morula were evaluated in 2 consecutive experiments. In the first experiment, 1-, 2-, 3- to 4- and 5- to 8-cell stage embryos were collected and cultured to the morula stage in Whitten's medium containing 0.1 mM ethylenediaminetetraacetic acid (EDTA). The development rate of 1-cell embryos to the morula stage was lower than that of the other stages (P<0.01). The post-thaw development rate of the morulae obtained from in vitro culture of 1-, 2-, 3- to 4-, and 5- to 8-cell embryos and from in vivo embryos (control) to the blastocyst stage was 55.5, 84.9, 87.4, 90.1 and 90.8%, respectively. The post-thaw development rate of morula obtained from in vitro produced 1-cell embryos was significantly lower than from the other stages or from the in vivo counterparts (P<0.0001). In Experiment 2, the impact of GSH supplementation of the culture medium in the presence or absence of EDTA was evaluated for embryo development to the morula stage and post-thaw survival, using in the 2 x 2 factorial design. Although EDTA supplementation increased development rates to the morulae (P<0.01) stage, GSH did not have an influence on morula development. However, the presence of either GSH or EDTA in the culture medium supported development to the blastocyst stage (P<0.01) of in vitro produced morulae. These data demonstrate that 1-cell embryos from a blocking-strain mouse cultured in vitro to the morula stage have a lower development rate following freezing and thawing than embryos collected at the 2-cell or later stages. Addition of EDTA or GSH, individually or in combination, to the culture medium may improve the development rate of morula to blastocyst stage following cryopreservation.
Asunto(s)
Transferencia de Embrión/veterinaria , Glutatión/farmacología , Mórula , Animales , Frío , Técnicas de Cultivo , Ácido Edético/farmacología , Ratones , Conservación de Tejido/veterinariaRESUMEN
Pronuclear stage embryos with intact (ZI), slit (ZS) or completely removed (ZF) zona pellucida were encapsulated with an artificial zona pellucida (AZP) made of 1.5% sodium alginate. Embryos were cultured in KSOM medium with or without protein and their development in vitro to the blastocyst stage was recorded. AZP significantly (P < 0.05) improved the development of embryos to the blastocyst stage regardless of the presence of the natural zona pellucida. The encapsulated embryos developed at a higher rate (P < 0.05) in the absence of protein as compared with non-encapsulated embryos. Furthermore, the cell contacts at the 4-cell stage were significantly improved (P < 0.05) with encapsulation. AZP improved (P < 0.05) the development of pronuclear stage embryos with a slit zona pellucida to morula and blastocyst stages as compared with ZS embryos. It is concluded that AZP improves the in vitro development of pronuclear stage embryos with intact or completely removed zona pellucida as well as micromanipulated embryos to the blastocyst stage.
Asunto(s)
Alginatos , Materiales Biocompatibles , Desarrollo Embrionario y Fetal/fisiología , Zona Pelúcida , Transferencia Intrafalopiana del Cigoto/veterinaria , Análisis de Varianza , Animales , Medios de Cultivo/química , Femenino , Ácido Glucurónico , Ácidos Hexurónicos , Ratones , Ratones Endogámicos ICR , Proteínas/farmacología , Transferencia Intrafalopiana del Cigoto/métodosRESUMEN
In Japan, very hot moist compresses applied to the lumbar region have been used to promote intestinal movement and flatus or defecation. This skill was based on empirical knowledge. One of the purposes of this study is to make clear the effect of very hot compresses on intestinal movement. The other purpose is to prove the safety of the skin where very hot compresses applied. The sample consisted of 8 healthy female volunteers, ranging in age from 27 to 47 with a mean age of 37.4. A very hot compress was applied to the lumbar region for 10 minutes. The results were as follows: 1) Bowel sounds immediately after hot compresses increased 1.7 times compared with bowel sounds before the hot compresses. 2) Skin temperature of the back increased to 41.1-43.1 degrees centigrade under the hot compresses, then decreased rapidly. 3) Blood flow to the back during hot compresses, increased to 156%. Blood flow to the upper arm also increased. 4) Body temperature, pulse rate and blood pressure were not remarkably changed by the hot compresses. The results of the experiments show that very hot compresses applied to the lumbar region affect intestinal movement. And the results suggest that a very hot compress is a useful nursing skill to promote flatus or defecation.
Asunto(s)
Auscultación , Vendajes , Calor/uso terapéutico , Región Lumbosacra , Peristaltismo , Agua , Adulto , Temperatura Corporal , Investigación en Enfermería Clínica , Defecación , Femenino , Flatulencia , Humanos , Región Lumbosacra/irrigación sanguíneaRESUMEN
The effects of different concentrations of etoposide and cycloheximide (ETO-CHXM), used for chemical enucleation of mouse oocytes, on polar body extrusion and chromatin expulsion were tested. The developmental ability of blastomeres of late 2-cell stage embryos fused to chemically enucleated oocytes of different ages or cytoplasts from different sources was also examined in vitro. Metaphase I oocytes cultured in different concentrations of ETO-CHXM (10-50 micrograms/ml/each) extruded polar bodies at rates similar to those cultured without ETO-CHXM (58.5-65.9% and 64.6%, respectively). However, low percent of the oocytes (1.7-6.2%) expressed signs of meiotic perturbation, which was manifested by blebbing of the cytoplasmic membrane and extrusion of two or more polar body-like fragments. Twenty-three percent of the chemically enucleated oocytes cultured in ETO-CHXM-free medium spontaneously fused to their polar bodies. The rates of total chromatin expulsion were similar when ETO-CHXM concentrations were 36 and 50 micrograms/ml (93.5 and 98%, respectively). The results also showed that the cleavage rates of reconstituted embryos were significantly (P < 0.001) affected by the age of the chemically enucleated oocytes. Cytoplasts of bisected oocytes that matured in vivo supported the development of 31.7% of the reconstituted embryos to the blastocyst stage. However, both cytoplasts of chemically enucleated oocytes and in vitro matured oocytes did not support the development to the blastocyst stage. A high percentage (85.5%) of the reconstituted embryos with chemically enucleated recipients displayed abnormality of the metaphase plate. These results suggest that concentrations of etoposide between 36 and 50 micrograms/ml are optimum for enucleation of mouse oocytes. Furthermore, increasing the age or reducing the cytoplasmic volume of the chemically enucleated oocytes did not improve the development of the reconstituted embryos to the blastocyst stage.
Asunto(s)
Blastómeros/citología , Blastómeros/fisiología , Desarrollo Embrionario y Fetal/fisiología , Oocitos/citología , Oocitos/fisiología , Animales , Antineoplásicos Fitogénicos/farmacología , Fusión Celular , Núcleo Celular/efectos de los fármacos , Núcleo Celular/ultraestructura , Células Cultivadas , Cicloheximida/farmacología , Relación Dosis-Respuesta a Droga , Combinación de Medicamentos , Etopósido/farmacología , Femenino , Ratones , Ratones Endogámicos C57BL/embriología , Ratones Endogámicos CBA/embriología , Ratones Endogámicos ICR/embriología , Oocitos/ultraestructura , Inhibidores de la Síntesis de la Proteína/farmacologíaRESUMEN
To study the effects of insulin and insulin-like growth factor-I (IGF-I) on the development of bovine embryos, fertilized bovine embryos in vitro were cultured in a chemically defined, protein-free medium: modified synthetic oviduct fluid (mSOF) supplemented with 1 mg/ml polyvinyl alcohol. Dose-response studies showed that insulin (0.5 to 10 microg/ml) and IGF-I (2 to 200 ng/ml) stimulated the development of bovine embryos to the morula stage 5 d after in vitro fertilization. The addition of 0.5 microg/ml insulin or 2 ng/ml IGF-I to the mSOF had beneficial effects on embryonic development to the morula stage in the presence of amino acids, but insulin and IGF-I did not affect the development of bovine embryos to the morula stage in the absence of amino acids. The antiIGF-I receptor antibody (alphaIR-3) completely blocked the stimulation of development to the morula stage by insulin and IGF-I. These findings suggest that the stimulation of embryonic development by insulin and IGF-I is mediated through the IGF-I receptor.
RESUMEN
The influence of oxygen concentrations in the gas atmosphere on the development of IVM/IVF bovine embryos was determined by culturing them in a microdrop of modified synthetic oviduct fluid medium supplemented with amino acids, insulin and PVA (mSOFai). After removing the cumulus cells at 18 hr post-insemination, presumptive zygotes were cultured in mSOFai for 104-106 hr under 5% CO2 with various O2 concentrations (2.5 to 20%). Reduced O2 (5-10%) improved the development to the morula stage, and 5% O2 gave the highest development. In the next experiment, morulae obtained after 102-104 hr of culture, were further cultured for 50 hr in mSOFai with 2mM glucose under 5 and 20% O2. An increase in the mean cell number in blastocysts, but not in the frequency of blastocysts, was observed under 5% O2. In the third experiment, zygotes were cultured for 152-154 hr in mSOFai under 5 and 20% O2, or cocultured with bovine oviduct epithelial cells in TCM199 + 10% FCS under 5% CO2 in air. Percentage of blastocysts for mSOFai in 5% O2 doubled to that for 20% O2, and was similar to that for coculture. Moreover, mean cell number in the blastocysts for mSOFai in 5% O2 was significantly higher than that for coculture. Results demonstrate that oxygen concentration critically affects embryonic development through zygotes to blastocysts, and suggest that around 5% 02 is optimal. It also indicates that bovine zygotes can be cultured up to the blastocyst stage using a chemically defined medium with rates similar to those of a conventional coculture system.
Asunto(s)
Bovinos/embriología , Medios de Cultivo/farmacología , Desarrollo Embrionario y Fetal/efectos de los fármacos , Fertilización In Vitro/veterinaria , Oxígeno/farmacología , Aminoácidos/análisis , Aminoácidos/farmacología , Animales , Medios de Cultivo/análisis , Relación Dosis-Respuesta a Droga , Desarrollo Embrionario y Fetal/fisiología , Femenino , Fertilización In Vitro/métodos , Insulina/análisis , Insulina/farmacología , Embarazo , Cigoto/efectos de los fármacos , Cigoto/fisiologíaRESUMEN
Mouse demi-embryos that developed from bisected morulae were transferred to recipients. The eu-blastocysts (distinct inner cell mass and well-developed trophectoderm) contained cells equal to 51% of the controls that developed from zona-free morulae. The rate of decidual cell reaction induced by the eu-blastocysts was not significantly different from that of the controls, but the size of the deciduum containing the egg cylinder was significantly smaller on Day 5.5 of pregnancy (P < 0.001). A significant increase in embryonic loss was observed from Day 7.5 to Day 9.5 in the eu-blastocysts (P < 0.05), while the controls exhibited no significant difference. Although the embryos from the eu-blastocysts showed retardation of developmental stages and decreased size, they attained normal stages and size regulation up to 90% of that of the control on Day 10.5. The pseudo-blastocysts (poorly developed inner cell mass enclosed by trophectoderm) contained cells equal to 25% of those of the controls and showed less than a 10% developmental rate to the egg cylinder stage. The trophectodermal vesicles (no enclosed cells) and nonintegrated forms (disorganized clusters of cells) contained cells less than 18% of those of the controls. They showed lower rates of decidual cell reaction than those in the controls (P < 0.05), and no egg cylinder was found in the deciduum. The results indicate that a severe decrease in the number of embryonic cells affects the regulation of embryonic development and decidual cell reaction in the uterus.
RESUMEN
The concept of care/caring has been recognized as the central component of nursing in the United States of America since the 1980s. Although caring is the essence and the central focus of clinical nursing, the term has several meanings, and there is no consensus regarding its definition. The concept of care/caring should be examined not only from the nurse's perspective, but also from the patient's perspective. The purpose of this research is to identify the common characteristics of the concept of care/caring from both the patient's and nurse's perspectives, and to clarify the structure of the concept of care/caring in the English articles. Thirty qualitative research papers written by 21 authors and 17 quantitative research papers written by 9 authors were sampled. From the analysis of the 47 articles, 5 categories and 13 sub-categories were identified as attributes of care/caring. The 5 categories were 1) Characteristics of Nurse, 2) Nursing Activities, 3) Patient-Nurse Relationship, 4) Caring Outcomes, and 5) Others. As the next stage of the research, the concept of care/caring in Japanese nursing should be analyzed. It must be determined whether the concept of care/caring is also the central component of nursing in Japan and if so can these identified characteristics of the concept be applied to the concept of care/caring in Japanese nursing.