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AB0175 Elimination of Granulocytic Myeloid-Derived Suppressor Cells in Lupus-Prone Mice Due to Ros-Dependent Extracellular Trap Formation
  1. K. Vlachou1,2,3,
  2. K. Mintzas3,
  3. M. Glymenaki2,
  4. M. Ioannou1,2,
  5. G. Papadaki1,2,
  6. Z. Vlata1,
  7. G. Bertsias2,
  8. P. Sidiropoulos2,
  9. D.T. Boumpas1,3,4,
  10. P. Verginis3,5
  1. 1Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology
  2. 2Laboratory of Autoimmunity and Inflammation, Medical School, University of Crete, Heraklion
  3. 3Biomedical Research Foundation of the Academy of Athens
  4. 44th Department of Medicine, Medical School, University of Athens, Athens, Greece
  5. 5Department of Clinical Pathobiochemistry, Medical Faculty, Technische Universität Dresden, Dresden, Germany

Abstract

Background Systemic lupus erythematosus (SLE) is characterized by a breakdown of self-tolerance against nuclear Ags leading to inflammation and multi-organ damage1-2. The mechanisms that participate in the breakdown of self-tolerance as well as the role of regulatory cells and molecules towards the re-establishment of tolerance remain elusive. Emerging evidence supports a crucial role of myeloid-derived suppressor cells (MDSCs) in the regulation of autoimmune diseases3; however their role in SLE remains unknown.

Objectives Thus, we sought to assess MDSC role in SLE inflammatory environment.

Methods The F1 hybrid of New Zealand Black x New Zealand White mice (NZB/W F1) was used as animal model for SLE. C57BL/6 (B6) mice were used as control mice. Experimental Autoimmune Disease (EAE) model was induced by injection of B6 mice with MOG35-55 peptide emulsified in CFA. Cell frequencies were analyzed by flow cytometry (FACS). FACS sorting was used for the isolation of MDSCs and T cells to perform T cell proliferation assay. Ly6G+ cells were isolated by the bone marrow of B6 mice with a discontinuous Percoll gradient (52%, 67%, 75%). Extracellular Trap formation (ETosis) was assessed by fluorescent microscopy; ETs were characterized as structures positively stained for neutrophil elastase (NE), myeloperoxidase (MPO) and DNA. Total ROS production levels were quantified by 2', 7' Dichlorofluorescin diacetate (DCF) reagent, whereas mitochondrial ROS production by MitoSox; results were obtained by FACS analysis. Gene expression analysis was performed by real time quantitative PCR and protein levels were assessed by Western blot.

Results Our results revealed an impaired expansion and defective function of MDSCs in the lymphoid organ of NZB/W F1 lupus-prone mice with established disease that involved predominantly the granulocytic MDSC cell subset (G-MDSCs). More specifically, we found increased elimination of G-MDSCs due to ET formation driven by the inflammatory milieu of lupus and we demonstrate a role of cytokines such as IFN-α, IFN-γ and IL-6 in this process. Induction of ET release by G-MDSCs was mediated by ROS production since inhibition of ROS generation significantly reduced ET release.

Conclusions Collectively, our findings reveal the elimination of a crucial regulatory immune cell subset in SLE microenvironment and provide new insights into the pathogenetic mechanisms of the disease.

References

  1. G. C. Tsokos. 2011. Systemic lupus erythematosus. N. Engl. J. Med. 365: 2110-2121.

  2. Bertsias, G. K., Salmon, J. E., and D. T. Boumpas. 2010. Therapeutic opportunities in systemic lupus erythematosus: state of the art and prospects for the new decade. Ann. Rheum. Dis. 69: 1603-1611.

  3. Crook, K. R. and P. Liu. 2014. Role of myeloi-derived suppressor cells in autoimmune disease. World J. Immunol. 4(1): 26-33.

Acknowledgements This work was supported by grants from the Greek General Secretariat of Research and Technology (Synergasia 09SYN-12-1074 to P.V.), (Aristeia I 2344 to D.B.), and European Union project Innovative Medicine Initiative 6 (“BeTheCure”, contract number 115142-2, to P.V and D.B.).

Disclosure of Interest None declared

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