Analysis Tools
cellular
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• blastocysts exhibit normal reactive oxygen species (ROS) levels relative to wild-type and heterozygous controls, indicating normal oxidative homeostasis
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• blastocysts exhibit hyperacetylated histones, with a marked increase in histone H4 acetylation on multiple residues (H4K5 + K8 + K12 + K16) and in histone H3 acetylation on lysine 56 (H3K56ac) relative to controls
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• percentage of active TRP53-positive apoptotic cells (phosphorylation at Ser15) is significantly increased relative to control blastocysts, indicating increased apoptosis
• percentage of TUNEL-positive nuclei is significantly higher than in control blastocysts, indicating increased DNA breaks and cell death
• however, percentage of EdU-positive nuclei is similar to that in control blastocysts, indicating normal cell proliferation
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embryo
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• percentage of active TRP53-positive apoptotic cells (phosphorylation at Ser15) is significantly increased relative to control blastocysts, indicating increased apoptosis
• percentage of TUNEL-positive nuclei is significantly higher than in control blastocysts, indicating increased DNA breaks and cell death
• however, percentage of EdU-positive nuclei is similar to that in control blastocysts, indicating normal cell proliferation
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• although blastocysts appear morphologically normal at E3.5, in vitro outgrowth assays show that 3 of 6 blastocysts exhibit arrested growth and differentiation immediately after hatching
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• in vitro outgrowth assays show 3 of 6 blastocysts fail to hatch with some blastomeres still trapped in the zona pellucida
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• percentage of NANOG-positive epiblast cells is significantly reduced relative to control blastocysts, indicating a defective epiblast lineage
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• percentage of SOX17-positive primitive endoderm (PE) cells is significantly reduced relative to control blastocysts, indicating a defective PE lineage
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• percentage of POU5F1/OCT4-positive inner cell mass (ICM) cells is severely decreased relative to control blastocysts; only ~30% of the appropriate number of ICM cells are present
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• percentage of CDX2-positive trophectoderm (TE) cells is significantly increased relative to control blastocysts
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mortality/aging
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IMPC - UCD
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IMPC - UCD
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• only a few homozygous embryos are recovered at E3.5 (3% versus expected 25%, n=32); no homozygous embryos or excess resorption sites are observed at E7.5
(J:279207)
• homozygous embryos cannot be recovered in vivo after E3.5
(J:290528)
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IMPC - UCD
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reproductive system
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• blastocysts hatch from the zona pellucida but fail to form typical outgrowth colonies after 3 days in culture
(J:279207)
• no embryos or increased numbers of empty decidua are found at E7.5, suggesting failure of embryo implantation
(J:290528)
• in vitro outgrowth assays show that none of the embryos form a successful outgrowth after 3 days in culture
(J:290528)
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