Characterization of mRNA expression for key glycerophospholipid and sphingolipid enzymes within trophectoderm and embryonic disc during initiation of porcine conceptus elongation

Authors: Miles, Jeremy; Rempel, Lea; Snider, Alexandria - Alex; PANNIER, ANGELA - University Of Nebraska

Submitted to: Molecular Reproduction and Development
Publication Type: Abstract Only
Publication Acceptance Date: 6/1/2023
Publication Date: 8/31/2023
Citation: Miles, J.R., Rempel, L.A., Snider, A.P., Pannier, A.K. 2023. Characterization of mRNA expression for key glycerophospholipid and sphingolipid enzymes within trophectoderm and embryonic disc during initiation of porcine conceptus elongation [abstract]. Molecular Reproduction and Development. 90(7):736. https://doi.org/10.1002/mrd.23697.
DOI: https://doi.org/10.1002/mrd.23697

Interpretive Summary:

Technical Abstract: Adequate conceptus elongation plays a critical role for successful pregnancy outcomes in the pig. Recent RNA-Seq analysis demonstrated significant transcriptomic regulation of many key glycerophospholipid and sphingolipid enzymes as the conceptus undergoes the initial stages of elongation. However, it is not clear whether gene regulation of these enzymes is primarily influencing the trophectoderm (TD, cells which extensively expand during elongation and ultimately become the placenta) or the embryonic disc (ED, cells which become the fetal proper). Therefore, the objective of this study was to characterize mRNA expression patterns within TD or ED for glycerophospholipid and sphingolipid enzymes as blastocysts transform from small to large ovoid blastocysts. White crossbred gilts were bred at estrus and harvested at day 10 of gestation. Reproductive tracts were flushed with RPMI-1640 medium, and blastocyst morphology and size were evaluated. Pregnancies were selected and grouped as containing only small (3-4 mm) or large (8-10 mm) ovoid blastocysts (n = 5 gilts per size group). Within each litter of both sizes, ED was removed from TD using a stereomicroscope and surgical blade and corresponding ED and TD cells, as well as intact (INT) blastocysts were pooled (n = 3 blastocysts per pooled cell type). Gene expression for transcripts of glycerophospholipid (DGKH, LPCAT1, LPGAT1, MBOAT2, and PPAP2A/2C) and sphingolipid (SPTLC1/C2/C3, SPHK1, SGPL1, ACER2, CERS3/4/6, and CERK) enzymes were measured using real-time PCR. All data were analyzed for ANOVA with MIXED model analysis. Every glycerophospholipid transcript increased (P > 0.001) expression as blastocysts transitioned from small to large ovoids and most were not different (P < 0.10) among ED, TD, or INT cell types. There was a tendency (P = 0.07) for increased PAPP2A expression in ED from large ovoids. In contrast, PAPP2C was increased (P = 0.01) in TD and INT cells compared to ED. Similarly, nearly all the sphingolipid transcripts had increased (P > 0.05) expression in large ovoids, except there was an interaction (P > 0.001) for SPTLC1, which was decreased in ED cells from large ovoids. Finally, most sphingolipid transcripts were increased (P > 0.01) in TD and INT cells compared to ED, except CERS4 and CERK were increased in ED compared to TD and INT. These data confirm previously reported transcript regulation of glycerophospholipid and sphingolipid enzymes as the conceptus initiates elongation and illustrate that regulation of glycerophospholipid enzymes influence all cell types of the blastocyst, whereas sphingolipid enzymes are primarily regulated within the TD. In conclusion, this study further suggests the importance of these lipids to drive the initiation of conceptus elongation. USDA is an equal opportunity provider and employer. Funding provided by USDA-NIFA-AFRI Grants no. 2017-67015-26456 and 2021-67015-34416.