Blastomere Removal from Cleavage-Stage Mouse Embryos Alters Steroid Metabolism During Pregnancy1

Abstract

Preimplantation genetic diagnosis (PGD) is a genetic screening of embryos conceived with assisted reproduction technologies (ART). A single blastomere from an early-stage embryo is removed and molecular analyses follow to identify embryos carrying genetic defects. PGD is considered highly successful for detecting genetic anomalies, but the effects of blastomere biopsy on fetal development are understudied. We aimed to determine whether single blastomere removal affects steroid homeostasis in the maternal-placental-fetal unit during mouse pregnancy. Embryos generated by in vitro fertilization (IVF) were biopsied at the four-cell stage, cultured to morula/early blastocyst, and transplanted into the oviducts of surrogate mothers. Nonbiopsied embryos from the same IVF cohorts served as controls. Cesarean section was performed at term, and maternal and fetal tissues were collected. Embryo biopsy affected the levels of steroids (estradiol, estrone, and progesterone) in fetal and placental compartments but not in maternal tissues. Steroidogenic enzyme activities (3beta-hydroxysteroid dehydrogenase, cytochrome P450 17alpha-hydroxylase, and cytochrome P450 19) were unaffected but decreased activities of steroid clearance enzymes (uridine diphosphate-glucuronosyltransferase and sulfotransferase) were observed in placentas and fetal livers. Although maternal body, ovarian, and placental weights did not differ, the weights of fetuses derived from biopsied embryos were lower than those of their nonbiopsied counterparts. The data demonstrate that blastomere biopsy deregulates steroid metabolism during pregnancy. This may have profound effects on several aspects of fetal development, of which low birth weight is only one. If a similar phenomenon occurs in humans, it may explain low birth weights associated with PGD/ART and provide a plausible target for improving PGD outcomes.

Embryo biopsy and development: the known and the unknown

“…Of relevance, a low fetal/placental weight ratio was observed in BB conceptuses, suggesting increased risk of cardiovascular diseases later in life (Thornburg et al 2010). Although placental gross morphology and weight seem not to be affected by BB, impaired placental functions have recently been described (Sugawara et al 2012, Sato et al 2014, Yao et al 2016). Sugawara and coworkers showed alterations in steroid metabolism, such as increased progesterone levels and impaired steroid clearance/elimination, in placentae and fetal livers (Sugawara et al 2012).…”

“…Although placental gross morphology and weight seem not to be affected by BB, impaired placental functions have recently been described (Sugawara et al 2012, Sato et al 2014, Yao et al 2016). Sugawara and coworkers showed alterations in steroid metabolism, such as increased progesterone levels and impaired steroid clearance/elimination, in placentae and fetal livers (Sugawara et al 2012). Compromised steroid metabolism has been associated with (i) defective placental vasculogenesis, which may affect nutrient supply to the fetus; (ii) defective signaling for the induction of labor and (iii) delayed fetal development, resulting in low birth weight and indicative of a higher predisposition to cardiovascular disease later in life.…”

“…Though the majority of studies did not show an increased incidence of pregnancy complications following embryo biopsy, some reported that BB is associated with reduced implantation rates, increased risks of low birth weight or preterm birth (Banerjee et al 2008, Thomaidis et al 2012, Cimadomo et al 2016 and, in early childhood, with impaired cognitive and/or motor development (Banerjee et al 2008, Middelburg et al 2011, Thomaidis et al 2012, Cimadomo et al 2016, while no indication on TB pregnancy/babies are available. Research on mouse model has shown that BB is not only associated with increased pregnancy loss and reduced/delayed fetal development, but also with impaired placental functions and epigenetic defects in both fetuses and placentae (Sugawara et al 2012, Yang et al 2012, Sato et al 2014, Wu et al 2014, Yao et al 2016. Regarding TB, research on mouse model is very limited so far.…”

“…Studies to assess epigenetic, behavioral, reproductive and/or metabolic health in the pre-or postnatal life following TB have not been published so far, while few research groups have performed studies to assess BB risk(s) and safety (Yu et al 2009, 2013, Sugawara et al 2012, Yang et al 2012, Zeng et al 2013, Sampino et al 2014, Sato et al 2014, Wu et al 2014. Thus, in the following sections, we will focus our attention on BB, rather than TB.…”

“…As widely described in human and animal models, a suboptimal preimplantation environment (such as in vitro culture) can perturb proper epigenetic remodeling or critical transcriptional events during early development (Thompson et al 2006, Lucas 2013, resulting in impaired fetoplacental and postnatal development. With respect to BB, there is limited knowledge of epigenetic or transcriptional perturbation in early embryos (Wu et al 2014); however, due to defects observed in BB conceptuses and offspring (Sugawara et al 2012, Wu et al 2014, it is plausible that the removal of one blastomere from the developing embryo perturbs the proper sequence of developmental programming events.…”