Evidence for conserved DNA and histone H3 methylation reprogramming in mouse, bovine and rabbit zygotes

<p>Abstract</p> <p>Background</p> <p>In mammals the parental genomes are epigenetically reprogrammed after fertilization. This reprogramming includes a rapid demethylation of the paternal (sperm-derived) chromosomes prior to DNA replication in zygotes. Such active DNA demethylation in the zygote has been documented for several mammalian species, including mouse, rat, pig, human and cow, but questioned to occur in rabbit.</p> <p>Results</p> <p>When comparing immunohistochemical patterns of antibodies against 5-methyl-cytosine, H3K4me3 and H3K9me2 modifications we observe similar pronuclear distribution and dynamics in mouse, bovine and rabbit zygotes. In rabbit DNA demethylation of the paternal chromosomes occurs at slightly advanced pronuclear stages. We also show that the rabbit oocyte rapidly demethylates DNA of donor fibroblast after nuclear transfer.</p> <p>Conclusion</p> <p>Our data reveal that major events of epigenetic reprogramming during pronuclear maturation, including mechanisms of active DNA demethylation, are apparently conserved among mammalian species.</p

Transcriptome profiling of rabbit parthenogenetic blastocysts developed under in vivo conditions

Parthenogenetic embryos are one attractive alternative as a source of embryonic stem cells, although many aspects related to the biology of parthenogenetic embryos and parthenogenetically derived cell lines still need to be elucidated. The present work was conducted to investigate the gene expression profile of rabbit parthenote embryos cultured under in vivo conditions using microarray analysis. Transcriptomic profiles indicate 2541 differentially expressed genes between parthenotes and normal in vivo fertilised blastocysts, of which 76 genes were upregulated and 16 genes downregulated in in vivo cultured parthenote blastocyst, using 3 fold-changes as a cut-off. While differentially upregulated expressed genes are related to transport and protein metabolic process, downregulated expressed genes are related to DNA and RNA binding. Using microarray data, 6 imprinted genes were identified as conserved among rabbits, humans and mice: GRB10, ATP10A, ZNF215, NDN, IMPACT and SFMBT2. We also found that 26 putative genes have at least one member of that gene family imprinted in other species. These data strengthen the view that a large fraction of genes is differentially expressed between parthenogenetic and normal embryos cultured under the same conditions and offer a new approach to the identification of imprinted genes in rabbit. © 2012 Naturil-Alfonso et al.This work was supported by Generalitat Valenciana research programme (Prometeo 2009/125). Carmen Naturil was supported by Generalitat Valenciana research programme (Prometeo 2009/125). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Naturil Alfonso, C.; Saenz De Juano Ribes, MDLD.; Peñaranda, D.; Vicente Antón, JS.; Marco Jiménez, F. (2012). Transcriptome profiling of rabbit parthenogenetic blastocysts developed under in vivo conditions. PLoS ONE. 7(12):1-11. https://doi.org/10.1371/journal.pone.0051271S111712Harness, J. 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A Simple Approach for COnsumption and RElease (CORE) Analysis of Metabolic Activity in Single Mammalian Embryos

Non-invasive assay of the consumption and release of metabolites by individual human embryos could allow selection at the cleavage stage of development and facilitate Single Embryo Transfer in clinical IVF but will require simple, high throughput, sensitive methods applicable to small volume samples. A rapid, simple, non-invasive method has therefore been devised using a standard fluorescence plate reader, and used to measure the consumption of pyruvate and glucose, and release of lactate by single bovine embryos at all stages of preimplantation development in culture; amino acid profiles have been determined using HPLC. Early embryos with an ‘intermediate’ level (6.14±0.27 pmol/embryo/h) of pyruvate uptake were associated with the highest rate (68.3%) of blastocyst development indicating that a mid “optimum” range of pyruvate consumption correlates with high viability in this bovine model

Chronic Effects of Fusarium Mycotoxins in Rations with or without Increased Concentrate Proportion on the Insulin Sensitivity in Lactating Dairy Cows

The objective of this study was to investigate the effect of long-term exposure to a Fusarium toxin deoxynivalenol (DON, 5 mg/kg DM) on the energy metabolism in lactating cows fed diets with different amounts of concentrate. In Period 1 27 German Holstein cows were assigned to two groups and fed a control or mycotoxin-contaminated diet with 50% concentrate for 11 weeks. In Period 2 each group was further divided and fed either a diet containing 30% or 60% concentrate for 16 weeks. Blood samples were collected in week 0, 4, 8, 15, 21, and 27 for calculation of the Revised Quantitative Insulin Sensitivity Check Index and biopsy samples of skeletal muscle and the liver in w 0, 15, and 27 for analysis by real-time RT-qPCR. The DON-fed groups presented lower insulin sensitivities than controls at week 27. Concomitantly, muscular mRNA expression of insulin receptors and hepatic mRNA expression of glucose transporter 2 and key enzymes for gluconeogenesis and fatty acid metabolism were lower in DON-fed cows compared to the control. The study revealed no consistent evidence that DON effects were modified by dietary concentrate levels. In conclusion, long-term dietary DON intake appears to have mild effects on energy metabolism in lactating dairy cows

Vocalization as an indicator of estrus climax in Holstein heifers during natural estrus and superovulation

The reliable detection of estrus is an important scientific and practical challenge in dairy cattle farming. Female vocalization may indicate reproductive status, and preliminary evidence suggests that this information can be used to detect estrus in dairy cattle. The aim of this study was to associate the changes in the vocalization rate of dairy heifers with behavioral estrus indicators as well as test the influence of the type of estrus (natural estrus vs. superovulation-induced estrus). We analyzed 6 predefined estrus-related behavior patterns (standing to be mounted, head-side mounting, active mounting, chin resting, being mounted while not standing, and active sniffing in the anogenital region) and vocalization rates in the peri-estrus period (day of estrus ± 1 d) of 12 German Holstein heifers using audio-visual recordings. Each heifer was observed under natural estrus and a consecutive superovulation induced by FSH and cloprostenol. Estrus was determined by behavioral patterns and confirmed by clinical examination (vaginoscopy and ultrasound imaging of the ovaries) as well as by the concentration of peripheral progesterone. Estrus behavior and vocalization rates were analyzed in 3-h intervals (an average of 19 intervals for each heifer), and an estrus score was calculated based on the 6 behaviors. The interval with the highest estrus score (I0) was considered the estrus climax. We demonstrated similar time courses for the estrus score and vocalization rate independent of estrus type. However, in natural estrus, the maximum vocalization rate (±SE) occurred in the interval before estrus climax (I−1; 42.58 ± 21.89) and was significantly higher than that in any other interval except estrus climax (I0; 27.58 ± 9.76). During natural estrus, the vocalization rate was significantly higher within the interval before estrus climax (I−1; 42.58 ± 21.89 vs. 11.58 ± 5.51) than under superovulation. The results underscore the potential use of vocalization rate as a suitable indicator of estrus climax in automated estrus detection devices. Further studies and technical development are required to record and process individual vocalization rates