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Introduction: The aim was to investigate syncytiotrophoblast extracellular vesicle (STBEV) uptake mechanisms by primary endothelial cells, the effects on gene expression, cell activation as well as the effect of aspirin., Methods: The STBEVs were derived using the placental perfusion system, from normal or preeclamptic placentas. Endothelial uptake was analysed with flow cytometry. To elucidate uptake, different inhibitors were tested; Cytochalasin D, Chlorpromazine hydrochloride, Methyl-B-cyclodextrin, Dynasore and Wortmannin. Endothelial gene expression was evaluated using an endothelial cell biology qPCR array. Cell activation was studied by ICAM-1 surface expression after STBEV exposure, with and without aspirin treatment., Results: Normal and preeclamptic STBEV uptake was blocked in similar ways. Chlorpromazine, Dynasore and Wortmannin almost completely blocked STBEV uptake. Methyl-B-cyclodextrin blocked 45-60% of the uptake while Cytochalasin D did not block uptake at all. Neither normal nor preeclamptic STBEVs had any significant effects on endothelial gene expression. Normal STBEVs down-regulated cell surface protein ICAM-1 expression, with and without aspirin treatment. Aspirin had no effect on STBEV uptake or cellular gene expression on its own, however it down regulated ICAM-1 protein expression in combination with preeclamptic STBEV exposure., Discussion: STBEV uptake primarily occurred through clathrin-mediated endocytosis. The STBEVs had no significant effect on gene expression but did have effects on ICAM-1 surface expression. The prophylactic mechanisms of aspirin may be by preventing the endothelium from being activated by the preeclamptic STBEVs., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

During the pregnancy associated syndrome preeclampsia (PE), there is increased release of placental syncytiotrophoblast extracellular vesicles (STBEVs) and free foetal haemoglobin (HbF) into the maternal circulation. In the present study we investigated the uptake of normal and PE STBEVs by primary human coronary artery endothelial cells (HCAEC) and the effects of free HbF on this uptake. Our results show internalization of STBEVs into primary HCAEC, and transfer of placenta specific miRNAs from STBEVs into the endoplasmic reticulum and mitochondria of these recipient cells. Further, the transferred miRNAs were functional, causing a down regulation of specific target genes, including the PE associated gene fms related tyrosine kinase 1 (FLT1). When co-treating normal STBEVs with HbF, the miRNA deposition is altered from the mitochondria to the ER and the cell membrane becomes ruffled, as was also seen with PE STBEVs. These findings suggest that STBEVs may cause endothelial damage and contribute to the endothelial dysfunction typical for PE. The miRNA mediated effects on gene expression may contribute to the oxidative and endoplasmic reticulum stress described in PE, as well as endothelial reprogramming that may underlay the increased risk of cardiovascular disease reported for women with PE later in life.

Preeclampsia (PE) is a pregnancy syndrome characterized by hypertension and organ damage manifesting after 20 gestational weeks. The etiology is of multifactorial origin, where placental stress causes increased levels of placenta-derived extracellular vesicles (STBEVs) in the maternal circulation, shown to cause inflammation, endothelial activation, vasoconstriction, and anti-angiogenic activity. General endothelial dysfunction is believed to be initiated by endothelial insult during pregnancy that alters vascular function resulting in increased arterial stiffness, cardiac dysfunction, and increased risk of cardiovascular disease later in life. We compared the effect of normal and PE derived STBEVs in vitro on vascular contractility of human subcutaneous arteries using wire myography. Cellular structures of exposed vessels were investigated by transmission electron microscopy. We explored strategies to pharmacologically block the effects of the STBEVs on human vessels. The PE STBEVs caused significantly stronger angiotensin II-mediated contractions and extended structural damage to human subcutaneous arteries compared to normal STBEVs. These negative effects could be reduced by blocking vesicle uptake by endothelial cells, using chlorpromazine or specific antibodies towards the LOX-1 receptor. The therapeutic potential of blocking vesicle uptake should be further explored, to reduce the permanent damage caused on the vasculature during PE pregnancy to prevent future cardiovascular risk. [ABSTRACT FROM AUTHOR]

The literature on extracellular vesicles consists of rapidly expanding and often contradictory information. In this paper we attempt to review what is currently known regarding extracellular vesicles released specifically from human placental syncytiotrophoblast cells with a focus on the common but complex pregnancy-associated syndrome pre-eclampsia, where the level of syncytiotrophoblast extracellular vesicle release is significantly increased. We review common methods for syncytiotrophoblast extracellular vesicle derivation and isolation and we discuss the cargo of syncytiotrophoblast extracellular vesicles including proteins, RNA and lipids and their possible functions. A metaanalysis of available trophoblast-derived extracellular vesicle proteomic datasets revealed only three proteins in common: albumin, fibronectin-1 and plasminogen activator inhibitor-1, suggesting some variability in vesicle cargo, most likely reflecting stage and cell type of origin. We discuss the possible sources of variability that may have led to the low number of common markers, which has led us to speculate that markers and density in common use may not be strict criteria for identifying and isolating placenta-derived exosomes. [ABSTRACT FROM AUTHOR]