Trophoblast cells inhibit neutrophil extracellular trap formation and enhance apoptosis through vasoactive intestinal peptide-mediated pathways

G CALO; F SABBIONE; D VOTA; D PAPARINI; R RAMHORST; A TREVANI; C PÉREZ LEIRÓS

HUMAN REPRODUCTION

OXFORD UNIV PRESS

Lugar: Oxford; Año: 2017 vol. 32 p. 55 - 64

0268-1161

STUDY QUESTION: Do human trophoblast cells modulate neutrophil extracellular trap (NET) formation, reactive oxygen species (ROS)synthesis and neutrophil apoptosis through mechanisms involving vasoactive intestinal peptide (VIP)?SUMMARY ANSWER: Trophoblast cells inhibited NET formation and ROS synthesis and enhanced neutrophil apoptosis through VIPmediatedpathways in a model of maternal?placental interaction.WHAT IS KNOWN ALREADY: Immune homeostasis maintenance at the maternal?placental interface is mostly coordinated by trophoblastcells. Neutrophil activation and NET formation increases in pregnancies complicated by exacerbated pro-inflammatory responses. VIPhas anti-inflammatory and immunosuppressant effects and is synthesized by trophoblast cells.STUDY DESIGN, SIZE, DURATION: This is a laboratory-based observational study that sampled circulating neutrophils from 50 healthyvolunteers to explore their response in vitro to factors derived from human trophoblast cells.PARTICIPANTS/MATERIALS, SETTING, METHODS: Peripheral blood neutrophils were isolated from healthy volunteers andtested in vitro with first trimester trophoblast cell line (Swan-71 and HTR8) conditioned media (CM) or with VIP. The effect of VIP andtrophoblast CM on NET formation was assessed by co-localization of elastase and DNA by confocal microscopy, DNA release and elastaseactivity measurement. Neutrophil apoptosis was determined by flow cytometry or fluorescence microscopy. ROS formation was assessed byflow cytometry with a fluorescent probe. VIP silencing was performed by siRNA transfection. For phagocytosis of apoptotic neutrophils,autologous monocytes were sampled, and engulfment and cytokines were assessed by flow cytometry and ELISA.MAIN RESULTS AND THE ROLE OF CHANCE: Trophoblast CM and 10 nM VIP promoted neutrophil deactivation by preventingphorbol myristate acetate?induced NET formation and ROS synthesis while they increased neutrophil spontaneous apoptosis and reversedthe anti-apoptotic effect of lipopolysaccharide (all P < 0.05 versus control). The effects of trophoblast CM were prevented by a VIP antagonistor when VIP knocked-down trophoblast cells were used (P < 0.05 versus control). Neutrophils driven to apoptosis by trophoblast CM couldbe rapidly engulfed by monocytes without increasing IL-12 production.LARGE SCALE DATA: Not applicable.LIMITATIONS, REASONS FOR CAUTION: The mechanisms of neutrophil deactivation by trophoblast VIP are based on the resultsobtained with neutrophils drawn from peripheral blood of healthy individuals interacting with trophoblast cell lines in vitro. These studies weredesigned to investigate biological processes at the cellular and molecular level; therefore, they have the limitations of studies in vitro and it is not possible to ascertain if these mechanisms operate similarly in vivo. We tested 50 neutrophil samples from healthy volunteers that have a normal variability in their responses. Cell lines derived from human trophoblast were used, and we cannot rule out a differential behavior of trophoblast cells in contact with neutrophils in vivo.WIDER IMPLICATIONS OF THE FINDINGS: Results presented here are consistent with an active mechanism through which neutrophilsin contact with trophoblast cells would be deactivated and silently cleared by decidual macrophages throughout pregnancy. They supporta novel immunomodulatory role of trophoblast VIP on neutrophils at the placenta, providing new clues for pharmacological targeting ofimmune and trophoblast cells in pregnancy complications associated with exacerbated inflammation.