Abstract
Serum supplementation during bovine embryo culture has been demonstrated to promote cell proliferation and preimplantation embryo development. However, these desirable outcomes, have been associated with gene expression alterations of pathways involved in macroautophagy, growth, and development at the blastocyst stage, as well as with developmental anomalies such as fetal overgrowth and placental malformations. In order to start dissecting the molecular pathways by which serum supplementation of the culture medium during the preimplantation stage promotes developmental abnormalities, we examined blastocyst morphometry, inner cell mass and trophectoderm cell allocations, macroautophagy, and endoplasmic reticulum stress. On day 5 post-insemination, > 16 cells embryos were selected and cultured in medium containing 10% serum or left as controls. Embryo diameter, inner cell mass and trophectoderm cell number, and macroautophagy were measured on day 8 blastocysts (BL) and expanded blastocysts (XBL). On day 5 and day 8, we assessed transcript level of the ER stress markers HSPA5, ATF4, MTHFD2, and SHMT2 as well as XBP1 splicing (a marker of the unfolded protein response). Serum increased diameter and proliferation of embryos when compared to the no-serum group. In addition, serum increased macroautophagy of BL when compared to controls, while the opposite was true for XBL. None of the genes analyzed was differentially expressed at any stage, except that serum decreased HSPA5 in day 5 > 16 cells stage embryos. XBP1 splicing was decreased in BL when compared to XBL, but only in the serum group. Our data suggest that serum rescues delayed embryos by alleviating endoplasmic reticulum stress and promotes development of advanced embryos by decreasing macroautophagy.
Highlights
In vitro production (IVP) of bovine embryos is a common practice in the livestock industry, contributing to over one million transferrable embryos in 2019 [1]
As a result of these findings, we propose that the addition of serum to the culture medium on day 5 post insemination results in a higher number of embryos reaching the blastocyst stage on day 7 and 8 of development 1) by rescuing developmentally compromised embryos through alterations in autophagy- and ER stress-associated molecular mechanisms, and/or 2) by the dysregulation of molecular pathways involved in growth, such as the Mammalian Target of Rapamycin complex 1 (MTORC1) pathway, causing hasten embryonic growth which result in an embryo with altered development
We found that the presence of serum from day five through eight of bovine embryo culture increases embryo diameter and total cell number and causes a stage specific effect on autophagic activity and ER stress
Summary
In vitro production (IVP) of bovine embryos is a common practice in the livestock industry, contributing to over one million transferrable embryos in 2019 [1]. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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