Analysis Tools
mortality/aging
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• although normal looking blastocysts are present in expected Mendelian ratios at E3.5, no homozygous embryos or excess resorption sites are observed at E7.5
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embryo
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• although blastocysts hatch from the zona pellucida with apparent trophectoderm (TE) and primitive endoderm (PrE) cells, inner cell mass (ICM) cells fail to form a typical outgrowth colony after 3 days in culture, suggesting impaired signaling from or proliferation of the ICM
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• blastocysts flushed at E3.5 and cultured overnight contain a significantly decreased total number and percentage of NANOG+ epiblast cells
• 4 of 7 blastocysts exhibit very few epiblast cells while the remaining 3 contain no NANOG+ cells within the ICM, indicating defective epiblast allocation
• however, the number and percentage of SOX17+ primitive endoderm (PE) cells is normal
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• ICM/epiblast cells fail to form a typical outgrowth colony after 3 days in culture
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• immunofluorescence analysis of POU5F1 (aka OCT4) shows that the percentage of POU5F1+ ICM cells is severely decreased, with only ~45% of the appropriate number of ICM cells present
• however, the total cell number per blastocyst is similar to that in controls, and no increase in Trp53-dependent apoptosis or changes in histone H4 acetylation (H4K5 + K8 + K12 + K16) are noted in the ICM
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• immunofluorescence analysis of the trophectoderm (TE) marker CDX2 shows that the percentage of CDX2+ TE cells is significantly increased
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cellular
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• although blastocysts hatch from the zona pellucida with apparent trophectoderm (TE) and primitive endoderm (PrE) cells, inner cell mass (ICM) cells fail to form a typical outgrowth colony after 3 days in culture, suggesting impaired signaling from or proliferation of the ICM
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• ICM/epiblast cells fail to form a typical outgrowth colony after 3 days in culture
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reproductive system
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• no embryos or increased numbers of empty decidua are found at E7.5, suggesting failure of embryo implantation
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