Bisphenol A (BPA) exposure in humans is widespread, and BPA has been detected in a variety of samples including follicular fluid. BPA levels have been found to negatively correlate with the developmental potential of oocytes in women undergoing in vitro fertilization and to induce meiotic abnormalities experimentally in human and mouse models. BPA may detrimentally affect oocyte maturation, and different concentrations of exposure can cause various outcomes. Because of the importance of oocyte maturation on developmental potential, disturbances during this time can significantly impact oocyte viability. Here, bovine oocytes were matured in vitro with and without BPA treatment of the media. The levels of BPA taken up by the oocytes were much lower than the initial exposure. Medium treatment with 30 ng/ml resulted in an average of 2.48 ng/ml BPA measured in mature oocytes. These oocytes exhibited decreased maturation and increased incidence of spindle abnormalities. Only 57.4% of oocytes exposed to 30 ng/ml BPA reached maturity compared to 72.4% of controls (p < 0.05). Mature oocytes following BPA exposure displayed increased abnormal spindle morphology (67.9%) and chromosome dispersal (60%) compared to all other groups analyzed (p < 0.05). Thus, exposure to BPA during in vitro oocyte maturation has the potential to decrease oocyte quality.

No abstract

Multi-copied gene families are prevalent in mammalian genomes, especially within the Y chromosome. Testis specific protein Y-encoded (TSPY) is present in variable copy number in many mammalian species. Previous studies have estimated that TSPY ranges from 50–200 copies in cattle. To examine TSPY localization on the Y chromosome we employed fluorescence in situ hybridization (FISH) and fiber-FISH. The results show a strong signal on the short arm of the Y chromosome (Yp). To investigate TSPY copy number we used relative real-time polymerase chain reaction (PCR) to analyze the DNA of 14 different cattle breeds. Variation both within and between breeds was observed. All breeds show significant variation in TSPY copy number among individual members. Brown Swiss (161 copies, CI = 133–195) had higher average levels of TSPY and Western Fjord Cattle (63 copies, CI = 45–86) had lower levels than some breeds. Overall, however, most breeds had a similar average TSPY copy number. The pooled average was 94 copies (CI = 88–100). The significance of the TSPY array remains uncertain, but as the function of TSPY is unraveled the purpose of the array may become clearer.

No abstract

High embryo loss occurs in the first week of bovine embryo development, with a high percentage of embryonic arrest. We hypothesized that arrested embryos enter a 'senescence-like state' and that both the cell cycle regulatory protein p53 and the stress-related protein p66(shc), which are involved in the onset of senescence in somatic cells, are responsible for this early embryonic arrest. In our in vitro production system, 13.5 +/- 0.5% of embryos arrest at the 2-4-cell stage. First cleavage occurs between 26 and 48 h post insemination (hpi), with early cleaving embryos showing only 0.6 +/- 0.3% arrest, with later cleaving embryos exhibiting up to 14.2 +/- 0.9% arrest. We compared 2-4-cell embryos collected at 28 hpi with those arrested at the 2-4-cell stage collected at day 8 post insemination. Quantification by real-time PCR and by semi-quantitative immunofluorescence showed significantly higher p66(shc) mRNA and protein levels in both arrested and late cleaving embryos versus 28 hpi embryos. By comparison, no significant changes in p53 mRNA, protein and phosphorylation levels were detected. Taken together, these results demonstrate that embryonic developmental potential is related to the time of first cleavage and that p66(shc), but not p53, is up-regulated in early arrested in vitro-produced bovine embryos.

In nondomestic and endangered species, the use of domestic animal oocytes as recipients for exotic donor nuclei causes the normal pattern of cytoplasmic inheritance to be disrupted, resulting in the production of nuclear-cytoplasmic hybrids. Evidence suggests that conflict between nuclear and cytoplasmic control elements leads to a disruption of normal cellular processes, including metabolic function and cell division. This study investigated the effects of nuclear-cytoplasmic interactions on the developmental potential of interspecies embryos produced by in vitro fertilization and somatic cell nuclear transfer: cattle x cattle, gaur x cattle, hybrid x cattle. Cattle control and hybrid embryos were examined for development to the blastocyst stage and blastocyst quality, as determined by cell number and allocation, apoptosis incidence, and expression patterns of mitochondria-related genes. These analyses demonstrated that a 100% gaur nucleus within a domestic cattle cytoplasmic environment was not properly capable of directing embryo development in the later preimplantation stages. Poor blastocyst development accompanied by developmental delay, decreased cell numbers, and aberrant apoptotic and related gene expression profiles, all signs of disrupted cellular processes associated with mitochondrial function, were observed. Developmental potential was improved when at least a portion of the nuclear genome corresponded to the inherited cytoplasm, indicating that recognition of cytoplasmic components by the nucleus is crucial for proper cellular function and embryo development. A better understanding of the influence of the cytoplasmic environment on embryonic processes is necessary before interspecies somatic cell nuclear transfer can be considered a viable alternative for endangered species conservation.

Background: Excessive developmental failure occurs during the first week of in vitro embryo development due to elevated levels of cell death and arrest. We hypothesize that permanently arrested embryos enter a stress-induced "senescence-like" state that is dependent on the oxidative stress-adaptor and lifespan determinant protein p66Shc. The aim of this study was to selectively diminish p66Shc gene expression in bovine oocytes and embryos using post-transcriptional gene silencing by RNA-mediated interference to study the effects of p66Shc knockdown on in vitro fertilized bovine embryos.

In this study we analyzed the pattern of polyadenylation changes that takes place between the resumption of meiosis and the first cleavage of bovine oocytes. Moreover, we investigated whether the delayed occurrence of the first cleavage division, which characterizes embryos of low developmental competence, is accompanied by an altered polyadenylation pattern of individual transcripts. We determined the polyadenylation status of a group of genes that characterize physiological processes, involved in early differentiation (Oct-4), compaction, and cavitation (beta-actin, plakophilin, connexin-32, connexin-43), energy metabolism (glucose transporter type 1, pyruvate dehydrogenase phosphatase), RNA processing (RNA poly(A) polymerase), and stress (heat shock protein 70). RNA was isolated from pools of 20 oocytes or embryos at the germinal vesicle (GV) stage, at the end of in vitro maturation, at the end of in vitro fertilization, and at the time of the first cleavage. Cleavage was assessed 27, 30, 36, 42 hr post insemination (hpi), and at the latter time the remaining uncleaved oocytes were retained as a group. Between oocyte isolation and first cleavage at 27 hpi (best quality embryos), the poly(A) tail of individual transcripts followed four patterns: no changes (beta-actin, PDP); gradual reduction (Cx-43, Oct-4, Plako); gradual elongation (Cx-32, TPA); reduction followed by elongation (PAP, HSP-70, Glut-1). If the interval between insemination and first cleavage was longer than 27 hpi (progressively lower quality embryos) further changes of polyadenylation were observed, which differed for each gene considered. These data indicated that specific changes in polyadenylation contribute to the modulation of gene expression in bovine embryos at this stage of development. Defective developmental competence is accompanied by abnormal polyadenylation levels of specific maternal mRNAs with synchrony between polyadenylation and cleavage emerging as an apparently important factor.