Mechanism of Siglec-8-mediated cell death in IL-5-activated eosinophils: Role for reactive oxygen species-enhanced MEK/ERK activation

Gen Kano, Maha Almanan, Bruce S. Bochner, Nives Zimmermann*

*Corresponding author for this work

Research output: Contribution to journalArticle

60 Scopus citations

Abstract

Background: Sialic acid-binding immunoglobulin-like lectin (Siglec)-8 is expressed on human eosinophils, where its ligation induces cell death. Paradoxically, Siglec-8-mediated cell death is markedly enhanced by the presence of the activation and survival factor IL-5 and becomes independent of caspase activity. Objective: In this report we investigate the mechanism of Siglec-8-mediated cell death in activated eosinophils. Methods: Human peripheral blood eosinophils were treated with agonistic anti-Siglec-8 antibody and IL-5, and cell death was determined by using flow cytometry and morphology. Phosphorylation of mitogen-activated protein kinase (MAPK) was determined by using phosphoLuminex, flow cytometry, and Western blotting. Reactive oxygen species (ROS) accumulation was determined by using dihydrorhodamine fluorescence. Results: Costimulation with anti-Siglec-8 and IL-5 significantly increased the rate and proportion of cell death by means of necrosis accompanied by granule release compared with that seen after stimulation with anti-Siglec-8 alone, in which apoptosis predominated. Together with the caspase-independent mode of cell death in costimulated cells, these findings suggest the activation of a specific and distinct biochemical pathway of cell death during anti-Siglec-8/IL-5 costimulation. Phosphorylation of extracellular signal-regulated kinase (ERK) 1/2 and MAPK-ERK kinase (MEK) 1 was significantly enhanced and sustained in costimulated cells compared with that seen in cells stimulated with IL-5 alone; anti-Siglec-8 alone did not cause ERK1/2 phosphorylation. MEK1 inhibitors blocked anti-Siglec-8/IL-5-induced cell death. ROS accumulation was induced by Siglec-8 ligation in a MEK-independent manner. In contrast, an ROS inhibitor prevented the anti-Siglec-8/IL-5-induced enhancement of ERK phosphorylation and cell death. Exogenous ROS mimicked stimulation by anti-Siglec-8 and was sufficient to induce enhanced cell death in IL-5-treated cells. Collectively, these data suggest that the enhancement of ERK phosphorylation is downstream of ROS generation. Conclusions: In activated eosinophils ligation of Siglec-8 leads to ROS-dependent enhancement of IL-5-induced ERK phosphorylation, which results in a novel mode of biochemically regulated eosinophil cell death.

Original languageEnglish (US)
Pages (from-to)437-445
Number of pages9
JournalJournal of Allergy and Clinical Immunology
Volume132
Issue number2
DOIs
StatePublished - Aug 1 2013

Keywords

  • Eosinophils
  • cell death
  • signaling

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

Fingerprint Dive into the research topics of 'Mechanism of Siglec-8-mediated cell death in IL-5-activated eosinophils: Role for reactive oxygen species-enhanced MEK/ERK activation'. Together they form a unique fingerprint.

  • Cite this