P-516 Defective decidualization secretome in severe preeclampsia
Authors: Castillo Marco N , Muñoz-Blat I , Cordero T , Ebner S , Meissner F , Monfort R , Satorres E , Simon C , Garrido-Gómez T 
Published: June 2023

Abstract

Study question
How defective decidualization (DD) induces a differential secretome compromising embryo invasion in severe preeclampsia (sPE).

Summary answer
DD endometrial stromal cells isolated from sPE patients secrete a differential secretome in response to hormonal stimuli.

What is known already
Endometrial decidualization is a primary driver of embryo invasion and placentation. This process is highly coordinated by estradiol and progesterone, which transform the endometrial stromal cells (ESCs) into decidualized cells specialized in the secretion of specific molecules. The secretome of ESCs regulates the endometrial differentiation into the decidua, including immune system and endothelial function regulation. Previously, we have demonstrated the existence of in vitro and in vivo defective decidualization (DD) in women who suffered sPE due to an impaired response to hormonal stimuli. However, the molecular mechanism that connects DD with a hostile maternal environment for embryo invasion remains unexplored.

Study design, size, duration
Human ESCs from women who suffered sPE (n = 3) and women with normal pregnancies as controls (n = 3) were isolated from endometrial biopsies collected at the time of late-secretory phase. ESCs were cultured using an established in vitro decidualization model, including three technical replicates per sample and condition. Culture media (n = 36) were analysed by mass spectrometry to characterize their global secretome, unveiling the differential secreted factors in DD underlaying sPE. The study duration was 12 months.

Participants/materials, setting, methods
For in vitro decidualization, isolated human ESCs were cultured for three days in the absence or presence of hormonal stimuli (0.5mM cAMP + 1μM MPA) in serum-free media. Culture media from decidualized and non-decidualized conditions were collected to analyse the secreted proteins by high-resolution mass spectrometry-based proteomics. Protein quantification was performed in the MaxQuant and Perseus software. This approach enables the characterization of the global secretory program as well as the discovery of proteins.

Main results and the role of chance
We identified and quantified a total of 1.117 proteins secreted by ESCs. In the transition from non-decidualized to decidualized status, the secretome of the control group showed significant differences in the abundance of 83 proteins (p-value&lt;0.05), including two classical markers of decidualization (IGFBP1 and PRL), as well as other hormonally regulated proteins (e.g. VEGFA, WNT4 and MFAP2), supporting the validity of our experimental method. Then, we compared the secretome of decidualized ESCs from controls versus sPE patients, unveiling the altered differential secretome linked to DD. We detected 160 proteins significantly deregulated in sPE compared controls (p-value&lt;0.05) during DD, including proteins reported as risk factors of sPE (e.g. LPA and TF), decidualization modulators (e.g. MFAP2 and TRAP1), mediators of maternal-fetal communication (NPTX1) and immune response regulators (e.g. C3, C5, C7, and SERPIN). Downregulated proteins were associated with cytoskeleton organization and small GTPase mediated signal transduction, consistent with defective decidualization. In contrast, upregulated proteins were implicated in complement activation, regulation of humoral and inflammatory response. A heatmap based on the 160 deregulated proteins successfully classified samples into sPE and controls, supporting the existence of a differential secretome in DD linked to sPE.

Limitations, reasons for caution
Sample size for in vitro decidualization was based on six patients, analysing two conditions and three technical replicates per patient (N total samples=36) to ensure robustness and reproducibility. Samples clustering in the heatmap support the biological variance was higher than the technical variance, supporting the effectiveness of the hallmark identified.

Wider implications of the findings
ESCs isolated from women who suffered sPE show an aberrant differential secretome linked to DD. These secreted factors into the decidual microenvironment might impact in embryo invasion, supporting the maternal contribution to sPE. Understanding the mechanisms underlying DD might provide new biomarkers for sPE risk prediction.

Trial registration number
Not applicable