Pronounced Segregation of Donor Mitochondria Introduced by Bovine Ooplasmic Transfer to the Female Germ-Line

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

Aberrant expression of E-cadherin and beta-catenin proteins in placenta of bovine embryos derived from somatic cell nuclear transfer

Abnormal placental development is common in the bovine somatic cell nuclear transfer (SCNT)-derived fetus. In the present study, we characterised the expression of E-cadherin and beta-catenin, structural proteins of adherens junctions, in SCNT gestations as a model for impaired placentation. Cotyledonary tissues were separated from pregnant uteri of SCNT (n - 6) and control pregnancies (n - 8) obtained by artificial insemination. Samples were analysed by western blot, quantitative RT-PCR (qRT-PCR) and immunohistochemistry. Bovine trophectoderm cell lines derived from SCNT and control embryos were analysed to compare with the in utero condition. Although no differences in E-cadherin or beta-catenin mRNA abundance were observed in fetal tissues between the two groups, proteins encoded by these genes were markedly under-expressed in SCNT trophoblast cells. Immunohistochemistry revealed a different pattern of E-cadherin and total beta-catenin localisation in SCNT placentas compared with controls. No difference was observed in subcellular localisation of dephosphorylated active-beta-catenin protein in SCNT tissues compared with controls. However, qRT-PCR confirmed that the wingless (WNT)/beta-catenin signalling pathway target genes CCND1, CLDN1 and MSX1 were downregulated in SCNT placentas. No differences were detected between two groups of bovine trophectoderm cell lines. Our results suggest that impaired expression of E-cadherin and beta-catenin proteins, along with defective beta-catenin signalling during embryo attachment, specifically during placentation, is a molecular mechanism explaining insufficient placentation in the bovine SCNT-derived fetus.

Development of Y-chromosome-Specific SCAR Markers Conserved in Taurine, Zebu and Bubaline Cattle

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

Ooplast-mediated developmental rescue of bovine oocytes exposed to ethidium bromide

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

Xenooplasmic Transfer between Buffalo and Bovine Enables Development of Homoplasmic Offspring

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

Chemically Assisted Enucleation Results in Higher G6PD Expression in Early Bovine Female Embryos Obtained by Somatic Cell Nuclear Transfer

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

Sexagem de embriões bovinos fecundados in vitro pela técnica de PCR multiplex

Neste trabalho, a técnica de PCR (polymerase chain reaction) foi utilizada para a sexagem de 92 embriões bovinos fertilizados in vitro. Os embriões originaram-se de fertilização in vitro de oócitos aspirados de ovários de fêmeas bovinas, provenientes de abatedouros comerciais. Os oócitos foram maturados, fertilizados e cultivados até o estádio de blastocisto. Os embriões foram lavados em solução de PBS, transferidos para tubos de polipropileno contendo água ultrapura, e imediatamente congelados a -196ºC. Os embriões foram descongelados sobre isopor contendo gelo picado e tratados com proteinase K. Para a reação de PCR, utilizaram-se alíquotas de 34 µl de cada tudo, onde foram acrescidos dois pares de primers, seqüência BC1.2 e seqüência satélite 1.715, desoxinucleotídeos, MgCl2, tampão PCR 10X, TaqDNA polimerase e água, em um volume final de 50 µl. As amostras foram amplificadas e a eletroforese realizada em gel de poliacrilamida a 8%. Os géis foram corados com solução de brometo de etídio e analisados em transiluminador de luz ultravioleta. Um índice de 93,47% de amplificação foi atingido, com 41 embriões (47,67%) machos e 45 (52,32%) embriões fêmeas. O uso de gel de poliacrilamida a 8% foi eficaz na separação de fragmentos de DNA muito próximos.

The effects of ovalbumin as a protein source during the in vitro production of bovine embryos

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

The effect of interaction between macromolecule supplement and oxygen tension on bovine oocytes and embryos cultured in vitro

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

Meiotic inhibition of bovine oocytes in medium supplemented with a serum replacer and hormones: effects on meiosis progression and developmental capacity

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

Effect of moderate and severe heat stress on avian embryonic Hsp70 gene expression

Stress response is a universal mechanism developed by all organisms to deal with adverse changes in the environment, which lead to the synthesis of heat shock proteins (Hsps). In this study, the effect of moderate (41degreesC) and severe (44degreesC) heat stress on Hsp70 transcript expression pattern was investigated during chicken embryogenesis. Acute exposure to severe heat stress for one hour resulted in a fifteen-fold increase in Hsp70 mRNA levels. The return of stressed embryos to normal incubation temperature resulted in Hsp70 mRNA levels five-fold higher than control after three hours and normal levels after six hours. Moderate heat stress did not induce enhancements on Hsp70 mRNA levels. The spatial expression of Hsp70 transcripts was detected in embryos under normal incubation conditions. Whole-mount in situ hybridization analysis showed that Hsp70 transcripts were constitutively present in somite and in distinct encephalic domains (predominantly in prosencephalon and mesencephalon areas) of the chicken embryo. These results showed that Hsp70 induction is dependent on incubation temperature conditions, suggesting that early chicken embryos may induce a quick emergence response to cope with severe heat stress by increasing Hsp70 mRNA levels.

Heat or cold chronic stress affects organ weights and Hsp70 levels in chicken embryos

This study was carried out with the objective of evaluating the effect of heat (38.8 degreesC) or cold (35.8 degreesC) stress on chicken embryo development and tissues Hsp70 levels, after the 13th day of incubation. Embryo weight (percent egg weight), organ weight (percent embryo weight) and Hsp70 levels (ng Hsp70 mug(-1) total protein) in different tissues (liver, breast muscle, heart, lungs, brain and kidney) were studied at the end of incubation. Cold stress induced a lower embryo weight and lower kidney and lungs weights, whereas heart and liver were lighter in heat-stressed embryos. An interaction between temperature and age was obtained only for Hsp70 levels in kidney and heart. Cold-stressed embryos showed higher Hsp70 levels in the brain, lungs and liver; a decrease in brain and breast muscle Hsp70 levels was seen from the 19th to 20th days in control embryos. Hsp70 levels increased with age in kidneys of control embryos and in heart of heat- and cold-stressed embryos. In conclusion, this study showed that chicken embryo organ weights are affected by incubation temperature, and that Hsp70 expression is tissue dependent (higher levels being seen in the brain) being cold-stress more effective in increasing Hsp70 levels in most studied tissues.

Hematological values and body, heart and liver weights of male and female broiler embryos of young and old breeder eggs

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

Breeder age and bone development in broiler chicken embryos

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Effect of breeder age on eggshell thickness, surface temperature, hatchability and chick weigh

Two experiments were carried out to study the effect of breeder age on incubation parameters (hatchability, eggshell thickness, egg surface temperature and chick weight). In Exp. 1, fertile eggs (30- and 60-wk-old breeders) were incubated at three different temperatures (36.8, 37.8 and 38.8 ºC). Eggshell surface temperature was measured by attaching a thermocouple to the shell and data were collected in a datalogger every ten minutes. This study was conducted according to a 3 x 2 factorial design (three temperatures and two breeder ages). Data revealed that eggshell surface temperature changed according to incubation temperature, with the main increase occurring between 10 and 13 days of incubation, and that the maximum increase in eggshell surface temperature was not higher than +0.6 ºC, irrespective of incubator temperature. The incubator temperature affected total incubation period and hatchability (%) at 38.8 ºC, independent of breeder age. Heavier eggs resulted in heavier chicks, irrespective of incubator temperature. In Exp 2, the eggs (30- and 60-wk-old breeders) were incubated at 37.8 ºC and eggs characteristics (weight, specific gravity, total hatchability and chicks weight) were evaluated according to a randomized experimental design. The data showed that breeder age affected eggshell thickness and chick weight (heavier eggs resulted in heavier chicks), but not specific gravity, eggshell surface temperature or hatchability. The findings of this study revealed that hatchability can be influenced by incubation temperature, but not by the breeder age. Breeder age can affect eggshell thickness, egg weight and eggshell surface temperature, but not specific gravity.