P-789 Trophoblast organoids as a 3D model for the study of oxygen concentration during in vitro preimplantation embryo culture
Authors: Barry F , Brouillet S , Hamamah S 
Published: July 2024

Abstract

Study question
To determine if trophoblast organoids could represent a relevant robust model to study the impact of oxygen concentration on human preimplantation embryo development.

Summary answer
Transcriptomic analysis of trophoblast organoids cultured in monophasic (5%) or biphasic (5-2%) O2 concentration identified 1,184 RNAs differentially expressed.

What is known already
Our team has previously demonstrated that a biphasic (5-2%) O2 concentration strategy during preimplantation embryo culture (from Day 0 to Day 6) significantly improved blastulation and live birth rates in in vitro fertilization (IVF). We also identified differentially expressed RNAs between monophasic and biphasic O2 strategies through whole transcriptome analysis on human embryos donated for research. However, the number of embryos donated to research is limited, they are all genetically different, and their use iethically controversial, representing a significative drawback to experimentation. Latest studies on trophoblast organoids indicate that they could represent a relevant model for studying preimplantation embryo development.

Study design, size, duration
A prospective monocentric study approved by the Institutional Review Board (I.R.B.) was conducted between February 2022 and September 2023: four early first-trimester human placentas were collected after surgical abortions and used for derivation of organoids in a 3D culture system. The protocol for organoid culture is described in the study of Sheridan and al, 2020. Organoids were cultured with either a monophasic or a biphasic oxygen concentration strategy.

Participants/materials, setting, methods
Trophoblast organoids were either exposed to 5% oxygen concentration during 5 days or to 5% O2 for 3 days and to 2% O2 for 2 days. Total RNA extraction of 21 organoids for each condition was performed. After whole transcriptome analysis using human Clariom D arrays (ThermoFisher Scientific®), microarray data were normalized and differentially expressed RNAs were identified using the Transcriptome Analysis Console software; the genetic network was generated with Ingenuity Pathway Analysis (Qiagen®).

Main results and the role of chance
Transcriptomic analysis of trophoblast organoids cultured in monophasic (5%) or biphasic (5-2%) O2 concentration identified 1,184 RNAs that were differentially expressed depending on the O2 strategy (fold change &gt;2 or &lt;-2, p &lt; 0.05), with most of them (75.1%, n = 889/1,184) being downregulated in the biphasic (5-2%) O2 group. IPA analysis highlighted downregulation of transcripts involved in cell survival and differentiation as well as upregulation of transcripts involved in embryo angiogenesis and vascularization of extraembryonic tissue. The transcript HIF1a was upregulated in organoids cultured in biphasic strategy compared to monophasic strategy (fold change 2, p = 0.0129). Comparison between differentially expressed transcripts in monophasic and biphasic strategies highlighted 22 transcripts in common between organoids and embryos donated to research from our previous study.

Limitations, reasons for caution
Our study was limited by a low sample size.

Wider implications of the findings
This innovative model of trophoblast organoids could help us to overcome the rarity as well as the genetic/epigenetic heterogeneity associated with IVF human embryos donated to the research, thus leading to the realization of large-scale experimentations and promoting the identification of new non-invasive predictive biomarkers for IVF.

Trial registration number
not applicable