Circulating exosomal and Argonaute-bound microRNAs in preeclampsia

Abstract

Introduction: microRNAs (miRNAs) play important role in the regulation of placental development, and abnormal miRNA expression is associated with preeclampsia (PE). miRNAs are released from trophoblast cells to maternal blood flow, where they are highly stable, being encapsulated inside extracellular vesicles, like exosomes or bound to Argonaute proteins. In PE, placental dysfunction leads to aberrant extracellular miRNA secretion. hsa-miR-210 is a hypoxia-sensitive miRNA found to be upregulated in PE, however, it is unknown whether it is the cause or the consequence of the disease. Objective: Our aim was to analyze the expression of several miRNAs, including hsa-miR-210 in placenta, exosome and Ago-bound fractions comparing normal (N) and PE pregnancies. We performed in vitro analyses of extracellular hsa-miR-210 secretion of trophoblast cell cultures (of villous and extravillous origin) under hypoxic condition. Methods: PE and N placenta samples were collected from C-sections, and blood samples were drawn from each pregnant woman in the third trimester. Htr-8 and Jar cell lines were cultured in exosome-free media and treated with hypoxia-mimetic agents. Exosome and Agobound fractions were isolated by membrane affinity spin column method from plasma and cell media. Short RNAs were extracted from exosomes and vesicle-free fractions, and total-RNA was isolated from the placenta samples. The RNA purity and concentration were measured by spectrophotometry. Expression analysis was carried out by qPCR with specific primers to target and reference miRNAs. Results: The level of hsa-miR-210 was significantly higher in PE placentas, which could cause a minor increase of exosomal and a high elevation of Ago-bound miR-210 in circulation. Hypoxia leads to intracellular hsa-miR-210 upregulation in trophoblast cell lines. In extravillous cell (HTR8) media, only the level of exosomal hsa-miR-210 was increased but no change in Ago-bound hsa-miR-210 level was observed. In contrast, in villous cell (JAR) media, the level of exosomal hsa-miR-210 was increased and enhanced release of Ago-bound hsa-miR-210 was also observed. Conclusion: Based on our data, we postulate that in PE, exosomal hsa-miR-210 are secreted actively from the trophoblast, and by intercellular communication, it may have a role in disease etiology. In addition, there is a passive release of Ago-bound hsa-miR-210 into the circulation, which may represent by-products of cell-death and is thereby a possible consequence of the disease.

The role of epigenetic changes in preeclampsia

“…The level of placenta has‐miR‐517c in PE group was upregulated but in exosome and Ago‐bound plasma no significant difference in the level of has‐miR‐517c between PE and normal group was detected. It was also shown that the level of has‐miR‐16 in placenta, exosome, and Ago‐bound plasma had no significant difference between two groups . Exosomal has‐miR‐210 may have impact in intracellular communication and etiology of PE disease and also release of Ago‐bound has‐miR‐210 could be the by‐product of cell death and several manifestations of PE diseases .…”

“…Has‐miR‐210 is one of the main markers of hypoxic responses in endothelial and trophoblast cells . Biro et al have reported that the level of has‐miR‐210 in exosome, Ago‐bound plasma, and placental was significantly upregulated in PE compared with normal group. Has‐miR‐517c is part of the primate‐specific C19MC miRNA cluster and expressed in placenta .…”

“…It was also shown that the level of has-miR-16 in placenta, exosome, and Ago-bound plasma had no significant difference between two groups. 93 Exosomal has-miR-210 may have impact in intracellular communication and etiology of PE disease and also release of Ago-bound has-miR-210 could be the by-product of cell death and several manifestations of PE diseases. 93 The total miRNA concentration in exosome samples was six times higher in sever PE and three times higher in moderate PE groups compared with normotensive group.…”

“…93 Exosomal has-miR-210 may have impact in intracellular communication and etiology of PE disease and also release of Ago-bound has-miR-210 could be the by-product of cell death and several manifestations of PE diseases. 93 The total miRNA concentration in exosome samples was six times higher in sever PE and three times higher in moderate PE groups compared with normotensive group. Moreover, exosomal has-miR-210-3p expression was higher in both sever PE and moderate PE groups and in sever PE higher than moderate PE group.…”

Effects of extracellular vesicles on placentation and pregnancy disorders

“…Moreover, miR-136, -494 and -495 in the EVs secreted by umbilical cord mesenchymal stem cells (UCMSCs) were also highly expressed in PE patients when compared to healthy controls. The expression of miR-210 was found to be higher in PE placenta when compared to normal placenta; coincidentally, hypoxic conditions induce a high level of miR-210 release via EVs (Biro et al 2019).…”