Confirmation of ED in ovine embryos.

<p>(<i>a</i>) Proportion of BrdU- and TUNEL-positive cells in diapausing ovine and murine blastocysts flushed from ovariectomised mouse uteri and in controls (<i>b</i>) qRT-PCR analysis of genes involved in ED control. Genes that positively regulate cell proliferation (<i>PCNA</i>) and signaling (<i>HB-EGF</i>) were not expressed in diapausing ovine blastocysts, while the anti-proliferative gene <i>BTG1</i> was significantly over-expressed. <i>IGF2R</i> mRNA expression did not differ statistically between diapausing and control blastocysts. (c) Immunolocalization of CB1 (green) in diapausing (middle panel) and control ovine blastocysts (upper panel). Nuclei (red) were visualized with propidium iodide. CB1 expression is higher in diapausing ovine blastocysts. Lower panel: ovine blastocysts incubated with neutralized anti-CB1 antibody showing no positive signal. For each experiment ≥5 blastocysts were used and it was repeated 3–5 times. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033027#s2" target="_blank">Results</a> are mean ± S.E.M. *** p<0.0001, ** p<0.003, *p<0.03.