PLASMA EXTRACELLULAR VESICLES DAMPEN ACUTE INFLAMMATORY RESPONSES IN NEUTROPHILS STIMULATED WITH BACTERIAL PAMPs

FABIANO, MARTINA; LEICAJ, LUZ; ADAMCZYK, ALAN; MAZZITELLI, IGNACIO; ERRA DIAZ, FERNANDO; PEREZ, PAULA; OSTROWSKI, MATÍAS

Congreso; REUNIÓN CONJUNTA SAIC SAI & FAIC SAFIS 2022; 2022

Extracellular vesicles are heterogeneous membrane structures that mediate intercellular communication both in physio and pathological conditions. We have previously shown that healthy donors’ plasma EVs (pEVs) have anti-inflammatory and pro-resolution effects on monocyte derived macrophages simultaneously treated with a PAMP. Herein we aimed at studying the pEV-mediated modulation of neutrophil activation by diverse PAMPs. We analyzed neutrophils’ oxidative burst, degranulation, cytokine production and cell viability. pEVs were purified from healthy donor plasma by size-exclusion chromatography followed by centrifugation, and characterized by western blotting (WB). Neutrophils were isolated from blood from healthy donors by centrifugation on Ficoll-Paque followed by dextran sedimentation and hypotonic lysis of erythrocytes. N-Formyl-Met-Leu-Phe (fMLP)-induced oxidative burst was analyzed by determining dihydrorhodamine 123 (DHR) oxidation; neutrophils’ degranulation was generated also with fMLP and assessed by CD11b and CD66b surface staining; cell viability was evaluated with Annexin V and propidium iodide. Measurements were obtained by flow cytometry (FC). Cytokines in cell culture supernatants were evaluated by ELISA. We observed that pEV treatment induced a dose-dependent reduction of both oxidative burst and degranulation of neutrophils following fMLP stimulation. However, IL-8 concentrations were higher in simultaneously LPS and pEVs treated neutrophils. Neutrophils stimulated with LPS 150 ng/mL and pEVs for 18 hs showed similar live cells percentages than only LPS treated ones. In conclusion, pEVs may contribute to controlling inflammation by diminishing neutrophils’ respiratory burst and degranulation while contributing to wound healing by promoting the secretion of the angiogenic cytokine IL-8.