74 resultados para Embryo Culture Techniques


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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

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

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Pós-graduação em Zootecnia - FCAV

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The acceptance of biotechnology for the most equine breeders association had a significant effect in the horse industry, gaining popularity around the world, because the increasing on the genetic gain, allowing the use of sub fertile mares and stallions with high genetics value on reproduction. The embryos in vitro production of human and cattle has been used with success, however in vitro embryo production is not efficient in the horse, as oocyte transfer (OT) and intracytoplasmatic sperm injection (ICSI). The oocyte transfer has been used especially in subfertile old mares presenting reproductive pathologies as: endometrite, cervical and uterine adhesions, blocked oviduct, perineal laceration and ovulation failures. During oocyte recovery process, the oocytes must be collected from immature follicles that need be matured in vitro or in vivo matured oocytes from pre-ovulatory follicles through the transvaginal aspiration guided by ultrasound. The recovered oocyte is transferred to a previously inseminated recipient mare, through the flank laparotomy. The intracytoplasmatic sperm injection (ICSI) is a procedure of in vitro fertilization that needs only one sperm that is aspirated and injected inside the oocyte. The oocytes used, can be from mature and immature follicles. Fresh, cooled and frozen semen can be used, because the procedure not requires a functional sperm. The use of Piezo drill resulted in a breakthrough the pellucid zone, allowing the vibration per minute provided in the sperm injection pipette, a major result of cleaved oocytes, due to a better sperm injection in the oocyte. The embryo transfer can be straight inside the oviduct, as also transcervical transferred after embryo culture produced in vitro. In conclusion both procedures (OT and ICSI) are effective to be used on equine assisted reproduction, getting results even lower than expected, but satisfactory from animal genetically superior

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

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

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This clinical study was conducted to correlate the levels of endotoxins and bacterial counts found in primary endodontic infection with the volume of periapical bone destruction determined by cone-beam computed tomography (CBCT) analysis. Moreover, the levels of bacteria and endotoxins were correlated with the development of clinical features. Twenty-four root canals with primary endodontic disease and apical periodontitis were selected. Clinical features such as pain on palpation, pain on percussion, and previous episode of pain were recorded. The volume (cubic millimeters) of periapical bone destruction was determined by CBCT analysis. Endotoxins and bacterial samplings were collected by using sterile/apyrogenic paper points. Endotoxins were quantified by using limulus amebocyte lysate assay (KQCL test), and bacterial count (colony-forming units [CFU]/mL) was determined by using anaerobic culture techniques. Data were analyzed by Pearson correlation and multiple logistic regression (P < .05). Endotoxins and bacteria were detected in 100% of the root canal samples (24 of 24), with median values of 10.92 endotoxin units (EU)/mL (1.75-128 EU/mL) and 7.5 × 10(5) CFU/mL (3.20 × 10(5)-8.16 × 10(6) CFU/mL), respectively. The median volume of bone destruction determined by CBCT analysis was 100 mm(3) (10-450 mm(3)). The multiple regression analysis revealed a positive correlation between higher levels of endotoxins present in root canal infection and larger volume of bone destruction (P < .05). Moreover, higher levels of endotoxins were also correlated with the presence of previous pain (P < .05). Our findings revealed that the levels of endotoxins found in root canal infection are related to the volume of periapical bone destruction determined by CBCT analysis. Moreover, the levels of endotoxin are related to the presence of previous pain.

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This study was carried out to assess the influence of bovine embryo culture medium Beltsville Agriculture Research Center (BARC), supplemented with FCS, BSA or PVA, on the in vitro oocyte maturation, evidenced by cleavage rate and blastocysts production at different developmental stages. Three experiments were performed, as follows: exp.1: addition of FCS to BARC medium at concentrations of 0, 5 and 10%; exp. 2: addition of BSA to BARC medium at concentrations of 0, 4 and 8 mg/ml; exp. 3: addition of PVA to BARC medium at concentrations of 0, 0.5 and 1.0 mg/ml. TCM 199 supplemented with bicarbonate, pyruvate, gentamicin sulfate, FSH, LH and FCS was used as control group. Oocytes obtained from cow ovaries at slaughterhouse were selected in PBS, and then matured in BARC medium supplemented with FSH, LH and gentamicin sulfate, according to the experimental design. Percoll gradient was used for sperm selection and TALP medium for IVF. In vitro embryo culture was in SOF-m medium; a humidified atmosphere with 5% CO2, in air, at 38.7oC was used for all steps. The number of oocytes reaching blastocyst, expanded blastocyst, and hatched blastocyt stages was recorded, respectively at 72 and 168 h post-insemination. ANOVA and Bonferroni t test were used to determine differences among groups. Differences of P<0.05 were taken as significant. Higher percentage (P<0.05) of cleaved oocytes was observed in group TCM + FCS than for the other groups matured in BARC supplemented with FCS or BSA, regardless the concentration used. However, the cleavage rate was similar between groups BARC plus PVA with 1 mg/ml (85.7%) and TCM + FCS (90.8%). Significant difference was found among groups for the production of blastocysts, with the control group yielding a higher number of blastocysts (results ranging from 47.4 to 51.4%, in comparison with groups using BARC + FCS (4.1 to 19.7%), BSA (1.4 to 5.6%) and PVA (5.7 to 10.6%). In conclusion, BARC medium supplemented with different macromolecules did not promote a beneficial effect on in vitro oocyte maturation, resulting in lower rate of cleavage and blastocyst production when compared with TCM + FCS medium.