Simple SummaryThe use of combined estrus synchronization/superovulation (SS) treatments alters ovarian and endometrial gene expression patterns, impairing follicle and oocyte growth, fertilization, and embryo development. Since the impact of SS treatments on the transcriptome of the surviving embryos remains unidentified, we examined gene expression changes in day 6 blastocysts that survived a brief regimen of synchronization treatment combined with superovulation. Embryos were surgically collected on day 6 after AI, and transcriptome analysis was performed on blastocyst-stage embryos with good morphology to identify differentially expressed genes (DEGs) between groups atp-value < 0.05 and </>1.5-fold change. Compared with the blastocysts from control (untreated) sows, the blastocysts from SS-treated sows had moderate gene expression changes, with 7 pathways disrupted and a total of 10 transcripts affected, including up-regulation of metabolicRDH10andSPTLC2genes and down-regulation of oxidation-relatedGSTK1andGSTO1genes. These gene expression alterations may suggest suboptimal embryo quality and could depress the embryos’ response to oxidative stress, thereby impairing subsequent embryo development. These and previous findings call for the avoidance of SS treatments in embryo transfer programs.AbstractThe combination of estrus synchronization and superovulation (SS) treatments causes alterations in ovarian and endometrial gene expression patterns, resulting in abnormal follicle and oocyte growth, fertilization, and embryo development. However, the impact of combined SS treatments on the transcriptome of the surviving embryos remains unidentified. In this study, we examined gene expression changes in day 6 blastocysts that survived a brief regimen of synchronization treatment combined with superovulation. The sows were included in one of three groups: SS7 group (n= 6), sows were administered Altrenogest (ALT) 7 days from the day of weaning and superovulated with eCG 24 h after the end of ALT treatment and hCG at the onset of estrus; SO group (n= 6), ALT nontreated sows were superovulated with eCG 24 h postweaning and hCG at the onset of estrus; control group (n= 6), weaned sows displaying natural estrus. Six days after insemination, the sows underwent a surgical intervention for embryo collection. Transcriptome analysis was performed on blastocyst-stage embryos with good morphology. Differentially expressed genes (DEGs) between groups were detected using one-way ANOVA with an un-adjustedp-value < 0.05 and a fold change </> 1.5. The effect of SO treatment on the number of altered pathways and DEGs within each pathway was minimal. Only four pathways were disrupted comprising only a total of four altered transcripts, which were not related to reproductive functions or embryonic development. On the other hand, the surviving blastocysts subjected to SS7 treatments exhibited moderate gene expression changes in terms of DEGs and fold changes, with seven pathways disrupted containing a total of 10 transcripts affected. In this case, the up-regulation of certain pathways, such as the metabolic pathway, with two up-regulated genes associated with reproductive functions, namelyRDH10andSPTLC2, may suggest suboptimal embryo quality, while the down-regulation of others, such as the glutathione metabolism pathway, with down-regulated genes related to cellular detoxification of reactive oxygen species, namelyGSTK1andGSTO1, could depress the embryos’ response to oxidative stress, thereby impairing subsequent embryo development. The gene expression changes observed in the present study in SS7 embryos, along with previous reports indicating SS7 can negatively affect fertilization, embryo production, and reproductive tract gene expression, make its use in embryo transfer programs unrecommendable.