Transcriptional profiling reveals upregulation of p53 signaling in porcine embryos produced in vitro

Authors: NOLAND, RYLIE - University Of Missouri; Redel, Bethany; LAMARTINA, MARISSA - University Of Missouri; PRATHER, RANDALL - University Of Missouri; Chen, Paula

Submitted to: Biology of Reproduction
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/23/2025
Publication Date: 5/14/2025
Citation: Noland, R.S., Redel, B.K., Lamartina, M.G., Prather, R.S., Chen, P.R. 2025. Transcriptional profiling reveals upregulation of p53 signaling in porcine embryos produced in vitro. Biology of Reproduction. 113(4): 777-786. https://doi.org/10.1093/biolre/ioaf113.
DOI: https://doi.org/10.1093/biolre/ioaf113

Interpretive Summary: Pig embryos produced in the laboratory setting are exposed to an artificial environment that induces a stress response. This can ultimately lead to poor development or death of the embryo. The goal of this study was to identify changes in gene expression in pig embryos due to culture in the artificial environment and ways to improve the culture system. Expression of genes related to cell stress and death were increased in the cultured pig embryos compared to those that developed inside of a pig. Therefore, a compound that inhibits this stress response was added to the pig embryo culture, and development of the embryos was improved. Embryos cultured with this compound were also able to establish pregnancies and resulted in live piglets, demonstrating that there was no negative impact on full-term development. As a result of the work, the embryo culture system in artificial environments has improved.

Technical Abstract: Although advances in the porcine embryo culture system have been achieved, the artificial environment continues to be stressful for the embryos which hinders development. To identify areas of improvement, transcriptional profiling was performed on in vivo-derived (IVV), in vivo-matured and in vitro-cultured (IVC), and in vitro-matured and cultured (IVMC) porcine blastocyst-stage embryos. Numerous differentially expressed genes were detected between IVC vs IVV (489 downregulated, 701 upregulated), IVMC vs IVV (435 downregulated, 1124 upregulated), and IVMC vs IVC (32 downregulated, 168 upregulated). Moreover, KEGG pathway analysis revealed upregulated pathways related to amino acid biosynthesis and metabolism in IVC and IVMC embryos compared to IVV. Interestingly, IVMC embryos demonstrated specific upregulation of the p53 signaling pathway compared to IVV embryos. Therefore, IVMC embryos were cultured with different p53 inhibitors, pifithrin-a (PFT-a), pifithrin-ß (PFT-ß), or pifithrin-µ (PFT-µ), to determine if the stress response could be suppressed to improve development to the blastocyst stage. Culture with 50 µM PFT-a improved development to the blastocyst stage (P'<'0.05), but total cell number and transcript abundance of p53 target genes remained unaltered. No difference in development was observed after culturing embryos with PFT-ß, and embryos cultured with pifithrin-µ (PFT-µ) demonstrated decreased development. Lastly, two embryo transfers of embryos cultured with PFT-a demonstrated that the inhibitor did not disrupt developmental competence of in vitro-produced embryos. Overall, addition of PFT-a in the porcine embryo culture medium was shown to have beneficial effects on development and is suitable for generating live pigs with this system.