P-447 Dynamic changes in uterine natural killer cell (uNK) function across menstrual phases: Implications for implantation
Authors: Sheth S , Woon E  V , Male V 
Published: July 2024

Abstract

Study question
Does the function of uNK cells change from the proliferative stage of the menstrual cycle to the secretory phase in readiness for implantation?

Summary answer
During the secretory phase, uNK cells demonstrate an enhanced ability to produce cytokines indicating their importance for implantation.

What is known already
The abundance of uNK cells during implantation, and their secretion of trophoblast chemoattractant factors, suggests their pivotal role in placentation. uNK cells express Killer Immunoglobulin-like Receptors (KIR), which interact with Human Leukocyte Antigen-C (HLA-C) on trophoblasts, inducing cytokine production that aids trophoblast migration. It has recently been discovered that uNK cells consist of three distinct subsets, with different propensities to produce cytokines. However, the cytokine production abilities of each subset, and how this changes over the menstrual cycle, remains unexplored.

Study design, size, duration
A prospective cross-sectional study was conducted to compare uNK cell function between the proliferative and secretory phases of the menstrual cycle. The proliferative phase was defined as before day 14 of the menstrual cycle, and the secretory phase after this, in those with a regular 28-32 day menstrual cycle. The current data analysed is from 6 proliferative phase samples and 10 secretory phase samples however recruitment is ongoing.

Participants/materials, setting, methods
Endometrial samples were collected with a Pipelle biopsy from healthy fertile women attending the sexual health clinic for contraception. In the laboratory, endometrial lymphocytes were isolated and cultured with Anti-CD107a PerCP-eFlour710 (100ul/ml), Brefeldin (3ug/ml) and Monensin (2uM/ml). Cells were then stimulated with phorbol 12-myristate 13-acetate (PMA) and ionomycin (2ul/ml) and incubated for 4 hours at 37 °C. Flow cytometry was employed to analyse the samples after staining with viability dye, cell surface antibodies, and intracellular markers.

Main results and the role of chance
In initial experiments, degranulation declined during the proliferative phase in both uNK2 and uNK3 compared to the other two phases of the menstrual cycle. There was a significant increase in the production of TNFa by uNK1 and IL-8 by uNK3 in the secretory phase compared to the proliferative phase. There were also increases in IFNg and GM-CSF production across all subsets although this did not reach significance (Statistical testing was done using Kruskal Wallis with a post-hoc Dunn test, p &lt; 0.05). Current work explores the production of more recently identified trophoblast chemoattractants, including XCL1 and CSF1, which we would expect to increase in a similar manner.

Limitations, reasons for caution
Limitations include the cross-sectional nature of the study, limiting the ability to establish causation. However another study, tracking pregnancy outcomes in those with recurrent implantation failure aims to address this. Additionally, variations within menstrual phases may exist. The sample size is also a limitation which I am working to expand.

Wider implications of the findings
A comprehensive understanding of uNK throughout various menstrual phases holds profound implications for reproductive health. The heightened cytokine production observed during the secretory phase indicates their pivotal role in the process of implantation. This finding not only underscores their significance but also serves as a catalyst for further research.

Trial registration number
not applicable