Neutrophils Clear Circulating Apoptotic Cell Derived Extracellular Vesicles and Secrete Pro-Regenerative Factors after Liver Resection

Abstract

Background: Inflammatory processes relying on intrahepatic neutrophil accumulation predominate the priming phase of liver regeneration. During later stages the relevance of neutrophils remains incompletely understood. Here we aimed to investigate dynamic phenotypical changes of circulating neutrophils upon stimulation with apoptotic cell derived extracellular vesicles (aEV) and their potential role in human liver regeneration. Methods: 93 patients undergoing liver resection were included in this study. Dynamic perioperative changes of aEV, caspase cleaved cytokeratin 18 (M30), M65 and HGF were assessed. In vitro efferocytosis of aEV by neutrophils was assessed by image flow cytometry. Secreted proteins were measured using a multiplex bead array assay. Results: M30 and aEV increased transiently at POD1 in patients with full recovery of liver function. Patients who developed liver dysfunction failed to increase M30. Neutrophils engulf aEV efficiently, which was associated with increased expression of CD11b, CD16, CD66b, and CD62L shedding, but neither induced NETosis, respiratory burst, or degranulation. While bacterial stimulation of neutrophils lead to an inflammatory neutrophil phenotype, engulfment of aEV lead to reduced secretion of pro-inflammatory cytokines but could not inhibit the activation of co-cultured T cells. Circulating HGF levels correlated with the M30 levels. Conclusion: While bacterial translocation, occurring during the priming phase of liver regeneration, shifts neutrophils towards an inflammatory phenotype, liver derived aEV, shed during later stages of liver regeneration are engulfed by neutrophils leading to a non-inflammatory, non-immunosuppressive polarization state and a tissue repair phenotype indicating a dynamic phenotypical switch during in human liver regeneration.