Spent embryo culture medium metabolites are related to the in vitro attachment ability of blastocysts

Abstract

The metabolomic profile of an embryo culture medium can aid in the advanced prediction of embryonic developmental potential and genetic integrity. But it is not known if this technology can be used to determine the in vitro potential of inner cell mass (ICM) in adherence and proliferation. Here, we investigated the developmental potential of mouse 2-cell embryos carrying cisplatin-induced DNA lesions (IDL), beyond blastocyst stage using ICM outgrowth assay. The genetic integrity of ICM cells was determined by comet assay. The metabolic signatures of spent medium were recorded 84 hours post injection of hCG (hpi-hCG), and after 96 hours of extended in vitro culture (Ex 96) by NMR spectroscopy. We observed that blastocysts that lack the ability to adhere in vitro had an increased requirement of pyruvate (p < 0.01), lactate (p < 0.01), and were accompanied by a significant reduction of pyruvate-alanine ratio in the culture medium. We propose that the aforementioned metabolites from 84 hpi-hCG spent medium be further explored using appropriate experimental models, to prove their potential as biomarkers in the prediction of implantation ability of in vitro derived human embryos in clinical settings.

Early prepubertal cyclophosphamide exposure in mice results in long-term loss of ovarian reserve, and impaired embryonic development and blastocyst quality

“…Ability of the blastocyst to attach and proliferate post-implantation in vitro was assessed using ICM outgrowth assay. In vitro derived blastocysts appearing morphologically normal with expanded blastocoel at 120 hour-post insemination (hpi) were selected for ICM outgrowth assay [21,22]. Briefly, multi-well dishes (D7039, Sigma Aldrich, USA) were pre-coated with gelatin (0.1%) for 30 min at room temperature.…”

Advantages of the outgrowth model for evaluating the implantation competence of blastocysts

“…The regulation of blastocyst development and outgrowth adhesion independently of gene activation enables cells to adapt to alter-ations in the conditions of in vitro culture. It was recently suggested that spent embryo culture medium metabolites might be related to the ability of blastocysts to undergo outgrowth [58]. In a metabolite analysis of embryo culture medium, non-outgrowth blastocysts that lacked the ability to adhere in vitro had increased requirements for lactate and pyruvate, and showed a significant reduction of the pyruvate-alanine ratio.…”

“…In a metabolite analysis of embryo culture medium, non-outgrowth blastocysts that lacked the ability to adhere in vitro had increased requirements for lactate and pyruvate, and showed a significant reduction of the pyruvate-alanine ratio. Thus, it was proposed that the aforementioned metabolites from the spent medium should be further analyzed using proper experimental models to substantiate their potential as biomarkers for predicting the implantation competence of embryos in clinical IVF-ET programs [58].…”

Embryo competence and cryosurvival: Molecular and cellular features

“…When analyzing the energy metabolism during early embryonic development, energy consumption is relatively low until pre-compaction stage and the main source of ATP is derived from pyruvate through oxidative phosphorylation, whereas with blastocyst formation and cavitation process a significant increase on energy demand and consumption of glucose, pyruvate and oxygen are expected. Additionally, after formation of the blastocyst, other sources of energy are required in greater proportion, including: glucose, amino acids and lipids (D'Souza et al, 2018;Hu and Yu, 2017).…”