ECTS2016 Poster Presentations Arthritis and other joint diseases: translational and clinical (11 abstracts)
Chemotactic signals mediating osteoclast progenitor migration in collagen induced arthritis
Darja Flegar 1, , Alan Sucur 1, , Antonio Markotic 1, , Tomislav Kelava 1, , Natasa Kovacic 2, , Sanja Ivcevic 1, , Katerina Zrinski-Petrovic 2, , Vedran Katavic 2, & Danka Grcevic 1,
1Department of Physiology and Immunology, University of Zagreb School of Medicine, Zagreb, Croatia; 2Laboratory for Molecular Immunology, Croatian Institute for Brain Research, Zagreb, Croatia; 3Department of Anatomy, University of Zagreb School of Medicine, Zagreb, Croatia.
Introduction: Collagen induced arthritis (CIA) is a mouse model of rheumatoid arthritis marked by persistent inflammation and enhanced osteoresorption at the affected joints as well as systemic osteopenia due to increased activation of osteoclast progenitors (OCP). Abundantly secreted chemokines presumably attract OCPs to the inflamed sites. Our goal was to identify chemotactic signals important for OCPs recruitment in CIA.
Methods: After obtaining approval from the Ethical Committee, C57BL/6 mice were immunized with chicken type II collagen in Freund’s adjuvant. Arthritis development was confirmed by clinical scoring and anti-collagen antibody detection. To determine OCP phenotype, peripheral blood mononuclear cells (PBMCs), bone marrow and collagenase digested tarsometatarseal joints were assessed by series of markers (CD3, B220, NK1.1, CD45, CD11b, CD115, CCR2, CCR5, CCR9, CXCR4) using flow-cytometry. For in vitro migration assay, PBMCs were stimulated with M-CSF and RANKL, seeded into transwell inserts and analyzed for the migration potential toward CCL2 or CCL5 contained in the lower chambers.
Results: Immunized mice developed arthritis with the incidence of about 60%. Frequency of OCPs contained among CD3-B220-NK1.1-CD45+CD11b+CD115+ population was increased in CIA (3.4±0.13% in control vs 7.02±2.59% in CIA, p<0.05). Chemokine receptors were expressed by certain proportion of OCPs in both groups, with substantial expression of CCR2 (27.95±5.93%) and CCR5 (24.45±5.35%), and variable expression of CCR9 (9.69±8.30%) and CXCR4 (1.13±1.04%). OCPs from mice with CIA showed increased migration potential toward chemoattractant gradients (median 11.5 (IQR 11 to 13.25) migrated cells in control vs 20.5 (IQR 20–25) in CIA for CCL2, P<0.05, and 12.5 (IQR 9–18.5) in control vs 33.5 (IQR 23–35.75) in CIA for CCL5, P<0.05).
Conclusions: Our results indicate that CCL2/CCR2 and CCL5/CCR5 signaling possibly contribute to increased OCP migration. Therapeutic blocking of such chemotactic signals represents a promising approach to antagonize enhanced osteoresorption in inflammatory diseases.
Volume 5
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