P-410 Repeated unsuccessful embryo transfers and inflammatory parameters in serum before IVF start and in follicle fluid at OPU
Authors: Nenonen H , Hjelmér I , Kondic A , Henic E 
Published: June 2022

Abstract

Study question
Is there a difference in cytokine Th1/Th2 ratio in women with repeated implantation failure (RIF) compared to women with a successful IVF treatment?

Summary answer
Proinflammatory activity in both serum and follicle fluid (FF) differs in RIF patients and patients with successful IVF treatment.

What is known already
Unexplained recurrent implantation failure (RIF) is generally used to describe patients who have not become pregnant after repeated embryo transfers (ETs) of good quality embryos (GQE) during IVF treatment. There is no unison definition. Implantation requires a receptive endometrium. Receptivity is modulated by cytokines amongst other factors. Cytokines are signaling molecules mainly produced by T-helper lymphocytes (Th; CD3+/CD4+) and macrophages. The guiding of the blastocyst to the endometrial lining, the trophoblast cell invasion and the subsequent remodeling of the damaged endometrial tissue all require proinflammatory cytokines (Th1 dominant), whereas the sustainment of pregnancy requires an anti-inflammatory state (Th2 dominant).

Study design, size, duration
Retrospective study based on material collected during a larger study including 384 women enrolled during 2007-2016. Inclusion criteria were: &lt;40 years of age, body mass index &lt;30kg/m2, non-smoking, regular menstruation cycle of 21-35 days and bilateral ovaries. Serum was obtained before treatment and FF was collected during OPU in the first IVF cycle.

Participants/materials, setting, methods
Patients with no implantation after ≥3 ETs with fresh or frozen GQEs and any indication were included in the RIF group (n = 29). Women with achieved pregnancy after ≤3 ETs, with an indication of male factor, were selected as controls (n = 26).
Cytokines were analysed with the MSD Proinflammatory Human Kit 1 (MSD, USA; IFN-γ, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13 and TNF-α). Th1/Th2 ratios were calculated and compared with the Mann-Whitney U test.

Main results and the role of chance
In serum there was a difference between RIF and non RIF in ratios for IFN-γ/IL10 (p = 0,01), IL-1β/IL10 (p = 0,04), IL2/IL10 (p = 0,00), TNF-α/IL10(p = 0,02), IFNγ/IL13(p = 0,01), IL12/IL13 (p = 0,02), IL2/IL13 (p = 0,00) and TNF-α/IL13 (p = 0,00) (p &lt; 0,05 for all), where the control group all had higher ratios, eg. a shift towards a Th1 profile before treatment start.
In FF there were differences in ratios between IL2/IL10 (p = 0,02), IL-8/IL-10 (0,02), TNF-α/IL10 (p = 0,02), IFNγ/IL13 (p = 0,01) and TNF-α/IL13 (p = 0,03). The RIF group had higher ratios of all these except for IFNγ/IL13, indicating that they had a shift towards a Th1 profile in their FF.
The RIF patients had a median age of 33 years, whilst the median age in the controls was 30 years (p = 0.01), there was no difference in BMI or cycle length.
Using inclusion and exclusion criteria regarding lifestyle factors and a strict definition of RIF and controls brings strength to the results and minimizes the role of chance. Also, there were no participants with chronic inflammatory disease.

Limitations, reasons for caution
The FF was pooled so the cytokine concentrations in the follicle containing the oocyte used for fertilization and ET was not determined. The age difference between the groups could have an effect. Yet, the median was below 35 years, where a decline in implantation and pregnancy rates are found.

Wider implications of the findings
Women at risk of immune related RIF could be identified proactively. Because it is taken at a timepoint closer to implantation, ratios in FF are specifically interesting as risk markers. Women with RIF and a Th1 shift in FF could be offered immune modulating therapy within a research setting.

Trial registration number
not applicable