Occurrence of a RAGE-Mediated Inflammatory Response in Human Fetal Membranes.

Marilyne Lavergne,Régine Minet-Quinard,Loïc Blanchon,Héléna Choltus,Julie Durif,Corinne Belville,Vincent Sapin,Denis Gallot

doi:10.3389/fphys.2020.00581

Abstract

Context:Sterile inflammation has been shown to play a key role in the rupture of the fetal membranes (FMs). Moreover, an early and exacerbated runaway inflammation can evolve into a preterm premature rupture of membranes and lead to potential preterm birth. In this context, we investigated the receptor for advanced glycation end products (RAGE), an axis implied in physiological sterile inflammation, in conjunction with two major ligands: AGEs and High-Mobility Group Box 1 (HMGB1). Our first objective was to determine the spatiotemporal expression profiles of the different actors of the RAGE-signaling axis in human FMs, including its intracellular adaptors Diaphanous-1 and Myd88. Our second goal was to evaluate the functionality of RAGE signaling in terms of FMs inflammation.MethodsThe presence of the actors (RAGE, HMGB1, Myd88, and Diaphanous-1) at the mRNA level was investigated by reverse transcription quantitative polymerase chain reaction (RT-qPCR) in the human amnion and choriodecidua at the three trimesters and at term. Measurements were conducted at two distinct zones: the zone of intact morphology (ZIM) and the zone of altered morphology (ZAM). Then, proteins were quantified using Western blot analysis, and their localization was evaluated by immunofluorescence in term tissues. In addition, pro-inflammatory cytokine secretion was quantified using a Multiplex assay after the treatment of amnion and choriodecidua explants with two RAGE ligands (AGEs and HMGB1) in the absence or presence of a RAGE inhibitor (SAGEs).ResultsThe FMs expressed the RAGE-signaling actors throughout pregnancy. At term, RNA and protein overexpression of the RAGE, HMGB1, and Diaphanous-1 were found in the amnion when compared to the choriodecidua, and the RAGE was overexpressed in the ZAM when compared to the ZIM. The two RAGE ligands (AGEs and HMGB1) induced differential cytokine production (IL1β and TNFα) in the amnion and choriodecidua.ConclusionConsidered together, these results indicate that RAGE signaling is present and functional in human FMs. Our work opens the way to a better understanding of FMs weakening dependent on a RAGE-based sterile inflammation.

Highlights

Fetal membranes are an essential actor in human parturition; if they do not achieve their missions, the childbirth can be impacted (Naeye and Peters, 1980; Romero et al, 2006; Menon, 2016; Menon and Richardson, 2017)
We investigated the mRNA expression profile of the RAGE, its adaptors and one ligand (HMGB1) in FMs on amnion and choriodecidua samples throughout pregnancy (first trimester: 1 to 13 weeks of gestation (WG); second trimester: 14–26 WG; third trimester: 27–37 WG; at term: 38–40 WG, by cesarean or vaginal delivery)
No significant difference in RAGE expression was revealed by RT-qPCR between trimesters

Summary

Introduction

Fetal membranes are an essential actor in human parturition; if they do not achieve their missions, the childbirth can be impacted (Naeye and Peters, 1980; Romero et al, 2006; Menon, 2016; Menon and Richardson, 2017). These fetal tissues consist of two layers: the amnion, which is the innermost layer directly in contact with the amniotic fluid (AF), and the chorion, which adheres to the maternal decidua This 9-month organ participates in the correct development of the fetus by providing AF homeostasis as well as physical and microbial barriers during pregnancy; they play a role in parturition by their programmed rupture at term (after 37 weeks gestation) (Buhimschi et al, 2004; Moore et al, 2006; King et al, 2007; Prat et al, 2012). An increasing number of studies have shown the implication of one key phenomenon in the FMs weakening: sterile inflammation (Girard et al, 2014; Romero et al, 2014, 2015) This concept is dependent on specific molecules called alarmins or “damage-associated molecular patterns” (DAMPs), which are released and recognized by pattern recognition receptors (PRRs) leading to a microbial-free inflammatory response or a “sterile” inflammation. Lappas and colleagues demonstrated an induction of cytokine release (IL1β, IL6, IL8, TNFα) by FMs in response to AGEs (Lappas et al, 2007)

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