Background Interleukin-34 (IL-34) is an inflammatory cytokine mainly involved in macrophage differentiation, angiogenesis and osteoclastogenesis in inflammatory arthritis (IA)1. IA is characterized by synovial hypoxia, increased oxidative stress and altered angiogenesis2.
Objectives To investigate the role of IL-34 in regulation of angiogenesis in IA.
Methods Twenty-three patients with active inflammatory arthritis (RA n=18; PsA n=5) were recruited and underwent knee arthroscopy. A subgroup of patients (n=6) pre-posttherapy were also recruited. Synovial tissue (ST) was collected and in vivo tissue oxygen levels (tpO2) were measured using combined Licox oxygen probe. IL-34 expression in ST and RA synovial fibroblasts (RASFC) was assessed by immunohistology (IHC) and RT-PCR. Synovial expression of angiogenic markers (VEGF, Ang2, Tie2) and marker of proliferation (Ki67) were also assesed by IHC. Synovial primary fibroblasts from RA (RASFC) and PsA (PsASFC) patients (n=8) and Human Microvascular Endothelial Cells (HMVEC) were stimulated with IL-34 (10 ng/ml and 100 ng/ml) under normoxia and 3% hypoxia, proliferation, angiogenesis and VEGF were assessed using a crystal violet proliferation assay, matrigel tube formation assay and ELISA. The effect of tumor necrosis factor α ( TNFα) and acute serum amyloid A (A-SAA) on IL-34 mRNA expression in RASFCs was measured by RT-PCR.
Results At baseline the mean synovial tissue pO2 level was profoundly hypoxic at 25.94mmHg (3.3%). IL-34 expression was observed throughout the synovium, with higher expression observed in the perivascular/vascular regions compared to the lining layer and sub-lining[DUF1]. Synovial IL-34 expression correlated with VEGF (r=0.60, p=0.011), Tie2 (r=0.50, p=0.021), Ang2 (r=0.70, p=0.013) and Ki67 (r=0.56, p=0.025) and with macroscopic vascularity (r=0.47, p=0.043). For those individuals who measured >20 mmHg ST tpO2, synovial IL-34 expression was also higher in any layers of the ST, however it did not reach significance. Posttherapeuthic synovial IL-34 expression significantly decreased in SL and VC layers (p=0.039, p=0.026) with a simultaneous tpO2 increase from 20.9 to 23.2 mmHg. At functional level, IL-34 significantly induced RASFC/HMVEC proliferation and HMVEC tube formation (all p<0.05), an effect that was potentiated under 3% hypoxic conditions (p<0.05). Furthermore IL-34 induced VEGF expression in RASFC[g3] s. . Preliminary results also show that upstream triggers of TNFα and A-SAA induced IL-34 mRNA expression in RASFCs that was potenciated by hypoxia.
Conclusions IL-34 is strongly associated with synovial inflammation and promotes synovial angiogenesis and cell proliferation, an effect that is potentiated by hypoxia.
Interleukin 34 expression is associated with synovitis severity in rheumatoid arthritis patients. Chemel M, Le Goff B, Brion R, Cozic C, Berreur M, Amiaud J, Bougras G, Touchais S, Blanchard F, Heymann MF, Berthelot JM, Verrecchia F, Heymann D. Ann Rheum Dis. 2012 Jan;71(1):150-4.
Redox balance dynamically regulates vascular growth and remodeling. Shyamal C. Bir, Gopi K Kolluru, Kai Fang, Christopher G. Kevil. Semin Cell Dev Biol. 2012 Sep;23(7):745-57.
Acknowledgements Translational Research Group, DAMC, SVUH
Disclosure of Interest E. Balogh Grant/research support from: MSD, M. Biniecka: None Declared, M. Connolly: None Declared, J. McCormick: None Declared, D. Veale Grant/research support from: Abbott, Opsona, Pfizer, Roche, Consultant for: MSD, Pfizer, Roche, Speakers bureau: Janssen, MSD, Pfizer, UCB, U. Fearon: None Declared
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