High levels of circulating extracellular vesicles with altered expression and function during pregnancy.

Extracellular vesicles (EVs) are widely considered important modulators of cell-cell communication and may interact with target cells locally and on a systemic level. Several studies had shown that circulating EVs' levels are increased during pregnancy. However, EVs characteristics, composition and biological functions in pregnancy still need to be clarified. This study aims to determine if circulating EVs during pregnancy are modified regarding levels, markers and cytokine profile as well as their reactivity towards peripheral blood cells. 26 pregnant women (PW) being in the second gestational trimester and 59 non-pregnant women (NPW) were investigated. EVs enrichment was performed by ExoQuick™ or ultracentrifugation; nanoparticle tracking analysis, SDS-PAGE followed by Western Blotting and densitometry, and IFN-γ, IL-10 and TGF-β1 ELISA for EVs characterization; imaging flow cytometry to analyze EVs' uptake by peripheral blood cells and flow cytometry were performed to analyze EVs function regarding induction of caspase-3 activity. Circulating EVs' levels were increased during pregnancy [26.9×10(6)EVs/ml (range: 6.4-46.3); p=0.003] vs NPW [18.9×10(6)EVs/ml (range: 2.5-61.3)]. Importantly, the immunosuppressive TGF-β1 and IL-10 cytokine cargo were increased in EVs of PW even after normalization to 1 million EVs [TGF-β1: 0.25pg/10(6)EVs (range: 0.0-2.0); p<0.0001] and [IL-10: 0.21pg/10(6)EVs (range: 0.0-16.8); p=0.006] vs NPW. Although EVs derived from non-pregnant and pregnant women were taken up by NK cells, the latter exclusively enhanced the caspase-3 activity in CD56(dim) NK cells (8.2±0.9; p=0.02). The qualitative and quantitative pregnancy-related alterations of circulating EVs provide first hints for an immune modulating role of circulating EVs during pregnancy.