Elsevier

Molecular Immunology

Volume 45, Issue 1, January 2008, Pages 160-168
Molecular Immunology

IL-17 attenuates the anti-apoptotic effects of GM-CSF in human neutrophils

https://doi.org/10.1016/j.molimm.2007.04.027Get rights and content

Abstract

Interleukin (IL)-17A is a pleiotropic, pro-inflammatory cytokine that is implicated in chronic inflammatory and degenerative disorders. IL-17 has been demonstrated to link activated T-lymphocyte with the recruitment of neutrophils at sites of inflammation, however whether IL-17 can mediate neutrophil survival and subsequently affect inflammatory responses has not fully been elucidated. In our study, we demonstrate that human peripheral blood and HL-60 differentiated neutrophils express mRNA and cell surface IL-17A receptor. IL-17A does not affect the rate of spontaneous neutrophil apoptosis, however significantly decreased granulocyte macrophage-colony stimulating factor (GM-CSF)-mediated survival by antagonizing the signal transduction pathways of p38, Erk1/2 and signal transducer and activator of transcription (STAT) 5B. These events were associated with reduced myeloid cell lymphoma-1 (Mcl-1) protein levels, increased translocation and aggregation of Bax to mitochondria, decreased mitochondrial transmembrane potential and in an increase in caspase-3/7 activity. These events were independent of increased Fas or soluble Fas ligand expression levels. Taken together, our findings suggest that IL-17 may regulate neutrophil homeostasis and favor the resolution of inflamed tissues by attenuating the delay in neutrophil apoptosis induced by inflammatory cytokines.

Introduction

Human neutrophils are the most abundant granulocyte accounting for 60% of the total circulating leukocytes found in blood (Houwen, 2001, Walker and Willemze, 1980). Although short lived, polymorphonuclear leukocytes are actively recruited to sites of inflammation by a multitude of chemotatic and infectious agents which can boost their survival. Endothelial transmigration (Watson et al., 1997), cytokines (Lee et al., 1993) and bacterial agents such as LPS (Hachiya et al., 1995) have been identified as neutrophilic survival factors. These factors can contribute to an acute or chronic state of inflammation by prolonging neutrophil activation and delaying apoptosis. Removal of apoptotic neutrophils is an essential factor in the normal resolution of the inflammatory response as it prevents damage to healthy tissues that would otherwise occur by necrotic cell lysis (Marshall and Watson, 1997).

Since neutrophils undergo spontaneous cell death without the need of a positive death signal, anti-apoptotic signals generated by pro-survival factors appear to coordinate the fate of these cells (Edwards et al., 2003). Of note, myeloid cell lymphoma-1 (Mcl-1) retains a principal function in the prolonged survival of these inflammatory cells (Edwards et al., 2004, Moulding et al., 2001). Cellular levels of Mcl-1 in human neutrophils are collectively associated with increased cell survival and its role as a primary modulator of apoptosis is further accentuated by the absence of Bcl-2 and Bcl-XL (Harter et al., 2003, Moulding et al., 1998).

Interleukin (IL)-17 is a pleiotropic, pro-inflammatory cytokine released by activated CD4+ and CD8+ T-lymphocytes. IL-17 has been established as a neutrophilic chemoattractant intermediate and links antigen specific T-cell with the recruitment and development of neutrophils through the induction of chemokines and myeloid growth factors from macrophages, fibroblasts, epithelial and endothelial cells (Aggarwal and Gurney, 2002, Fossiez et al., 1996, Jovanovic et al., 1998, Shin et al., 1999, Yao et al., 1995). Elevated levels of IL-17 have been associated with acute and chronic inflammatory disorders and have been detected in inflamed tissue from bacterial infections (Johnson et al., 2004, Luzza et al., 2000), synovial fluids from arthritic patients (Raza et al., 2005) and from sera or bronchoalveolar lavage fluid from asthmatic patients (Molet et al., 2001, Wong et al., 2001). Nevertheless, whether IL-17 has the ability to mediate the activity or survival of neutrophils and contribute to heighten inflammatory responses remains to be fully investigated.

Our present study aimed to determine the functional implication of IL-17A in relation to the survival outcomes of human neutrophils. Our results demonstrate that human peripheral blood and differentiated HL-60 neutrophils express cell surface IL-17RA. Stimulation with IL-17 did not induce apoptosis however attenuated survival effects of GM-CSF when added in combination. IL-17 antagonized GM-CSF-induced p38, Erk1/2 (p42/p44) and signal transducer and activator of transcription (STAT) 5B (p93) signal transduction pathways. This effect was associated with reduced Mcl-1 protein levels, increased Bax translocation, decreased mitochondrial transmembrane potential and increased caspase-3/7 activity. Collectively, our results demonstrate that IL-17 disrupts GM-CSF-induced anti-apoptotic signaling pathways regulating Bcl-2 family members and apoptosis.

Section snippets

Isolation and cell culture conditions

This study was approved by the Ethics Committee of the Faculty of Medicine at the University of Manitoba. Blood was collected into sterile heparin tubes from the peripheral vein of healthy volunteer donors. Neutrophils were separated from whole blood by Dextran, Ficoll-Paque histopaque sedimentation (Amersham Pharmacia Biotech) and hypotonic lysis method (Gounni et al., 2001). Cytological examination of stained neutrophils by the Wright–Giemsa method (Fisher Scientific) accounted for 95–98%

Circulating human neutrophils express mRNA and cell surface IL-17A receptor

To determine whether human neutrophils expressed the IL-17A receptor (IL-17RA), mRNA and cell surface receptor expression were examined. IL-17RA mRNA was detected in neutrophils from two healthy donors (lanes 1 and 2) and in differentiated HL-60 neutrophil-like cells (lane 4) but not in undifferentiated HL-60 promyelocytes (lane 3). Purified CD16+ human neutrophils expressed cell surface IL-17RA by a geometric mean of 3.5 ± 1.2 compared to isotype control (representative data shown in Fig. 1B, n = 

Discussion

To date, the focal point of IL-17s function on the myeloid lineage has generally been limited to its pro-inflammatory properties and to its ability to induce granulopoiesis or recruit neutrophils. New evidence from our study and others suggest that IL-17 may also possess anti-inflammatory properties. Andoh and colleagues have demonstrated that IL-17 can specifically inhibit TNF-α mediated RANTES secretion in human colonic subepithelial myofibroblasts through an auto-regulatory mechanism

Acknowledgements

We would like to acknowledge the helpful discussions brought forward by Drs. Aaron Marshall and Spencer Gibson. We also thank Dr. Keith Fowke and John Rutherford (University of Manitoba) for technical assistance in collecting human blood samples.

References (44)

  • R.B. Yang et al.

    A novel interleukin-17 receptor-like protein identified in human umbilical vein endothelial cells antagonizes basic fibroblast growth factor-induced signaling

    J. Biol. Chem.

    (2003)
  • S. Aggarwal et al.

    IL-17: a prototype member of an emerging cytokine family

    J. Leukoc. Biol.

    (2002)
  • M. Alvarado-Kristensson et al.

    p38-MAPK signals survival by phosphorylation of caspase-8 and caspase-3 in human neutrophils

    J. Exp. Med.

    (2004)
  • A. Andoh et al.

    IL-17 selectively down-regulates TNF-alpha-induced RANTES gene expression in human colonic subepithelial myofibroblasts

    J. Immunol.

    (2002)
  • K. Aoshiba et al.

    Role of p38-mitogen-activated protein kinase in spontaneous apoptosis of human neutrophils

    J. Immunol.

    (1999)
  • J.G. Clohessy et al.

    Characterisation of Mcl-1 cleavage during apoptosis of haematopoietic cells

    Br. J. Haematol.

    (2004)
  • S.W. Edwards et al.

    Regulation of neutrophil apoptosis by Mcl-1

    Biochem. Soc. Trans.

    (2004)
  • S.W. Edwards et al.

    Regulation of neutrophil apoptosis

    Chem. Immunol. Allergy

    (2003)
  • P.K. Epling-Burnette et al.

    Cooperative regulation of Mcl-1 by Janus kinase/stat and phosphatidylinositol 3-kinase contribute to granulocyte–macrophage colony-stimulating factor-delayed apoptosis in human neutrophils

    J. Immunol.

    (2001)
  • G. Fossati et al.

    The mitochondrial network of human neutrophils: role in chemotaxis, phagocytosis, respiratory burst activation, and commitment to apoptosis

    J. Immunol.

    (2003)
  • F. Fossiez et al.

    T cell interleukin-17 induces stromal cells to produce proinflammatory and hematopoietic cytokines

    J. Exp. Med.

    (1996)
  • A.S. Gounni et al.

    Human neutrophils express the high-affinity receptor for immunoglobulin E (Fc epsilon RI): role in asthma

    FASEB J

    (2001)
  • Cited by (62)

    • Effect of aging on sputum inflammation and asthma control

      2017, Journal of Allergy and Clinical Immunology
      Citation Excerpt :

      Furthermore, IL-27 has been reported to inhibit Treg cells.51,52 TH17 cells promote neutrophilia through multiple mechanisms,53,54 including secretion of the neutrophil chemoattractant CXCL8 (IL-8),55 and are associated with asthma in younger adults that is less responsive to ICSs.56,57 Although we did not measure clinical responsiveness to ICSs in our asthmatic patients, aged patients, despite similar ICS treatments, had significantly decreased asthma control compared with younger patients, which was associated with increased levels of IL-6 and the TH17 cytokine MIP-3α/CCL20.

    View all citing articles on Scopus

    Supported by grants from the Canadian Institute of Health Research (CIHR, MOP-53104) and the Manitoba Medical Service Foundation to ASG. ASG is supported by a CIHR New Investigator Award. SD is supported by a studentship from the Health Sciences Centre Foundation, the Manitoba Health Research Council and by the CIHR National Training Program in Allergy and Asthma.

    View full text