Background Inflammatory arthritides (IA) are associated with both local and systemic inflammation. Hypoxia and angiogenesis are recognized as important events in the perpetuation of joint destruction in IA. Oxidative stress, which arises from an imbalance between Reactive Oxygen Species (ROS) and antioxidant defences, also play important role in the pathogenesis of IA1,2.
Objectives To examine if hypoxia-induced oxidative stress regulates angiogenic mechanisms in IA.
Methods Patients with inflammatory arthritis (IA; n=44) underwent arthroscopy and synovial tissue oxygen (tpO2) measurements, and synovial tissue biopsies and clinical assessment were obtained. Twenty patients pre/post-TNFi therapy were also recruited. Immunohistology (IHC) and dual-immunofluorescence (IF) were performed to quantify vascular factors (VEGF, Ang2 and Tie2), cell specific markers (CD3, CD68) and markers of oxidative DNA damage (8-oxo-dG) and lipid peroxidation (4-HNE). Rheumatoid arthritis (RA) and psoriatic arthritis (PsA) synovial primary fibroblasts (SFC) and Human Microvascular Endothelial Cells (HMVEC) were stimulated with 4-HNE (5µM) under normoxia and 3% hypoxia conditions, and proliferation, tube formation and angiogenic growth factors were assessed using crystal violet assay, matrigel tube formation assays and ELISA.
Results VEGF, Ang2 and Tie-2 were expressed in the inflamed synovium localised in the lining layer (LL), sub-lining (SL) and vascular regions (VC) with the highest expression of VEGF, Ang2 and Tie2 scored in the VC layer, compared to LL and SL (all p<0.005). VEGF expression correlated with macroscopic synovitis (r=0.41, p=0.031) and vascularity (r=0.40, p=0.034), Ang2 expression correlated with synovitis (r=0.56, p=0.013) and CD68+ cells (r=0.53, p=0.013) and Tie2 correlated with CD3+ cells (r=0.41, p=0.023) and CD68+ cells (r=0.46, p=0.010). Treatment with TNFi showed a significant reduction in expression of Tie2 (p=0.034) and Ang2 (p=0.021), this was paralleled by an increase in mean tpO2 levels (22.8 mmHg-30.3 mmHg). A significant association between increased ΔVEGF, ΔAng2 and ΔTie2 with high DNA damage (Δ8-oxo-dG) and lipid peroxidation (Δ4-HNE) was demonstrated (all p<0.05), suggesting an interplay between oxidative stress and angiogenesis in a progression of chronic inflammatory arthritis. Colocalisation of angiogenic factors VEGF, Ang2 and Tie2 with oxidative damage markers 4-HNE and 8-oxo-dG in inflamed synovium was demonstrated. Functionally, 4-HNE induced RASFC proliferation (p<0.01), HMVEC tube formation (p<0.001) and the effect was significantly potentiated under hypoxic conditions (p<0.001).
Conclusions Angiogenesis is closely linked to hypoxia and oxidative stress; both of which promote angiogenic inflammatory cell influx and proliferation in inflammatory athritis.
Hypoxia--a key regulator of angiogenesis and inflammation in rheumatoid arthritis.
Konisti S, Kiriakidis S, Paleolog EM. Nat Rev Rheumatol. 2012 Jan 31;8(3):153-62.
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, D. Veale Grant/research support from: Abbott, Opsona, Pfizer, Roche, Consultant for: MSD, Pfizer, Roche, Speakers bureau: Janssen, MSD, Pfizer, UCB, J. McCormick: None Declared, Z. Szekanecz: None Declared, C. NG: None Declared, U. Fearon: None Declared, M. Biniecka: None Declared