P-168 Does the concentration of Interleukin-6 (IL-6) in spent embryo culture medium tell us anything? Association with implantation potential after euploid embryo transfer
Authors: Albert Rodriguez C , Bori L , Valera M.A , Conversa L , Delgado A , Meseguer M 
Published: June 2022

Abstract

Study question
Could Interleukin-6 (IL-6) secretion by euploid embryos provide extra information in the prediction of clinical outcomes after human blastocyst transfer?

Summary answer
The concentration of IL-6 in the spent embryo culture medium provides additional and non-redundant information associated with the success of a euploid embryo transfer.

What is known already
Previous studies revealed that the concentration of IL-6 in the culture medium could be useful in selecting the embryo for implantation (Dominguez et al., 2015) and higher concentrations of IL-6 was reported in embryos that reach blastocyst stage than in arrested ones (Lindgren et al., 2018). Recently, proteomic information gained from the analysis of the embryonic secretome have been used to predict the potential of a euploid embryo to lead to a live birth (Bori et al., 2021). Out of 92 proteins measured, IL-6 was one of the most relevant for the prediction of live birth along with blastocyst morphology.

Study design, size, duration
This prospective study included 157 euploid embryos from the preimplantation genetic testing for aneuploidies (PGT-A) program. Embryos were cultured until the blastocyst stage in EmbryoScope systems with single-step medium (Gems, Genea). All of them were routinely assessed by senior embryologists. Morphological quality was based on ASEBIR criteria and morphokinetic parameters were automatically annotated by guided annotations tool. The spent culture medium was collected on day 5/6 of embryo development (day of trophectoderm biopsy).

Participants/materials, setting, methods
Chromosome analysis was performed using next-generation sequence technology. In parallel, 20 µL of spent culture medium from each blastocyst was analyzed with Human IL-6 ELISA Kit. Only euploid embryos were used in the following analysis. We examined the association between IL-6 levels (pg/ul) and embryo morphology and morphokinetics. Out of the total, 77 embryos were selected and transferred according to ASEBIR morphological quality. Pregnancy, implantation and live birth rate were correlated to IL-6 concentration.

Main results and the role of chance
The average concentration of IL-6 in spent embryo culture medium was 1.76±0.93 pg/ul. The protein value was not related to embryo morphology (p = 0.332): 2.35±1.60 pg/ul for embryos graded as A; 1.72±0.95 pg/ul for embryos graded as B and 1.79±0.76 pg/ul for embryos graded as C. Likewise, IL-6 level was not related to morphokinetic parameters. Division time to 2 cells, 3 cells, 4 cells, 5 cells and time to blastulation were not statistically different among the following quartiles of IL-6: ≤ 1.06 pg/ul (n = 41), from 1.07 to 1.66 pg/ul (n = 39), from 1.67 to 2.21 pg/ul (n = 40) and &gt;2.21 pg/ul (n = 37). However, the concentration of IL-6 was higher in those euploid embryos that achieved a clinical pregnancy than in those embryos that failed after transfer. For spent culture media with less than 1.66 pg/ul of IL-6, pregnancy rate was 50%*, implantation rate was 50%* and live birth rate 43.80%. For spent culture media with more than 1.66 pg/ul of IL-6, pregnancy rate was 77.80% *, implantation rate was 75,60%* and live birth rate was 64.40%. *p&lt;0.05

Limitations, reasons for caution
The detection limit in protein quantification and the specific culture medium used that may affect IL-6 concentrations are the main limitations of our study.

Wider implications of the findings
Our findings demonstrated that the concentration of IL-6 in spent culture medium is independent of embryo morphology and morphokinetics, but it is related to the success of a PGT-A treatment. This may provide a promising approach to predict implantation after euploid embryo transfer by measuring the IL-6 protein.

Trial registration number
CDTI (IDI-20191102)