BioProject

The uterus is transiently receptive for embryo implantation. It remains to be understood why the uterus does not reject semi-allogeneic or allogeneic embryos for implantation. Uterine early response genes at the time approaching embryo attachment may provide insights. Uteri from C57BL/6 pseudo-pregnant (PP) mice and pregnant (P) mice approaching embryo attachment on day 3 post-coitum (D3) @22 h were analyzed for early response genes by microarray. SAM algorithm retained 21,858 unique probesets. Principal component analysis (PCA) indicated a clear separation between the PP and P groups. There were 105 upregulated and 5 downregulated protein-coding genes in the pregnant uterus (fc>1.5 & q-value<5%). Gene ontology (GO) analysis of these upregulated genes revealed that 84% of the GO terms were related to immune responses, with a dominant natural killer (NK) cell activation signature. Among the top 8 upregulated protein-coding genes, Cyp26a1 inactivates retinoic acid (RA) while Lrat promotes vitamin A storage, both of which attenuate RA bioavailability; Atp6v0d2 and Gjb2 play roles in ion transport and transmembrane transport; Gzmb, Gzmc, and Il2rb are involved in immune responses; and Tdo2 is important for kynurenine pathway. Most of these genes or their related pathways have been implicated in immune regulations. RA signaling has been implicated in tolerance and immunity and uterine NK cells have been implicated in immunotolerance at the maternal-fetal interface in the placenta. The mechanisms of immune responses approaching embryo attachment remain to be elucidated. The coordinated effects of the early response genes may hold the keys to the question of why the uterus does not reject an implanting embryo. Overall design: Embryo attachment can be detected by faint blue dye reaction in mice and it normally occurs ~midnight 4 days post-coitum (D4 @0h or D3 @24h). The earliest time for faint blue dye reaction to be detectable in some C57BL/6 mice is D3 @22h (DOI: 10.1016/j.fertnstert.2011.01.160). Young virgin C57BL/6 mice were purchased from The Jackson Laboratory. They were randomly splitted into two sets. One set of mice were mated with stud males. Mating night was defined as D0 and mating was verified by a vaginal plug the next morning. The pregnant uterine tissues were collected on D3 @22h. Only the ones without blue dye reaction (to verify no embryo attachment yet) but with healthy-looking blastocysts (to verify pregnancy status) flushed from the uterine horns were included. The other set of mice were mated with vasectomized males to produce pseudopregnant mice. The pseudopregnant uterine tissues were also collected on D3 @22h. The uterine tissues were processed for total RNA isolation. Four samples each from the pregnant and pseudopregnant groups with the best RNA qualities (260/280 ratio: 1.84~1.99; RIN: 6.9~8.7) were selected from microarry.