Human chemerin induces eryptosis at concentrations exceeding circulating levels

Abstract

Introduction: Human chemerin is an adipokine that regulates chemotaxis, inflammation, and glucose metabolism. In addition, accumulating evidence suggests that chemerin promotes apoptosis, autophagy, and pyroptosis. However, there are no data on its impact on eryptosis. The current study aimed to analyze the effects of human active Glu21-Ser157 chemerin on eryptosis in vitro. Materials and Methods: Human chemerin 0-2-10-50 µg/mL was incubated for 24 h with human erythrocytes (hematocrit 0.4%) obtained from eight healthy individuals. Flow cytometry-based determination of phospholipid scrambling, reactive oxygen species (ROS) production, and intracellular Ca2+ levels was performed. To supplement data on ROS and Ca2+ signaling in chemerin-mediated eryptosis, incubation in the presence or absence of antioxidants vitamin C and N-acetylcysteine and Ca2+-binding agent EGTA was carried out, respectively. Confocal microscopy-based techniques were used to detect reactive nitrogen species (RNS) generation, involvement of caspase-3 and caspase-8, as well as the state of lipid order in cell membranes of erythrocytes exposed to human Glu21-Ser157 chemerin. Results: Our observations suggest that human Glu21-Ser157 chemerin had no impact on eryptosis parameters at 2 µg/mL. However, chemerin stimulated phosphatidylserine externalization, ROS production, and Ca2+ accumulation at higher concentrations suggesting activation of eryptosis. Ca2+ uptake turned out to be at least partly required for chemerin-mediated eryptosis. Chemerin-mediated erythrotoxicity was additionally mediated by RNS, caspase-3, and caspase-8. Moreover, Glu21-Ser157 chemerin promoted reduction in the liquid-ordered phase of cell membranes in erythrocytes. Conclusions: The present study first discloses that human chemerin can induce eryptosis via Ca2+-dependent mechanisms at concentrations noticeably exceeding circulating levels. Thus, chemerin-induced eryptosis can hardly contribute to eryptosis-mediated anemia in diseases associated with enhanced levels of chemerin in blood.