Chrissta X Maracle1,2,3, Paulina Kucharzewska4, Boy Helder1,2,3, Corine van der Horst1,2,3, Pedro Correa de Sampaio5, Ae-Ri Noort1,2,3, Katinka van Zoest1,2,3, Arjan W Griffioen6, Henric Olsson4, Sander W Tas7,2,3. 1. Amsterdam Rheumatology and immunology Center. 2. Department of Clinical Immunology & Rheumatology. 3. Department of Experimental Immunology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands. 4. AstraZeneca R&D, Gothenburg, Sweden. 5. MD Anderson Cancer Centre, University of Texas, Houston, TX, USA. 6. Angiogenesis Laboratory, Department of Medical Oncology, VU University Medical Center, Amsterdam, The Netherlands. 7. Amsterdam Rheumatology and immunology Center s.w.tas@amc.uva.nl.
Abstract
OBJECTIVE: Angiogenesis is crucial in RA disease progression. Lymphotoxin β receptor (LTβR)-induced activation of the non-canonical nuclear factor-κB (NF-κB) pathway via NF-κB-inducing kinase (NIK) has been implicated in this process. Consequently, inhibition of this pathway may hold therapeutic potential in RA. We describe a novel three-dimensional (3D) model of synovial angiogenesis incorporating endothelial cells (ECs), RA fibroblast-like synoviocytes (RAFLSs) and RA synovial fluid (RASF) to further investigate the contributions of NF-κB in this process. METHODS: Spheroids consisting of RAFLSs and ECs were stimulated with RASF, the LTβR ligands LTβ and LIGHT, or growth factor bFGF and VEGF, followed by quantification of EC sprouting using confocal microscopy and digital image analysis. Next, the effects of anginex, NIK-targeting siRNA (siNIK), LTβR-Ig fusion protein (baminercept) and a novel pharmacological NIK inhibitor were investigated. RESULTS: RASF significantly promoted sprout formation, which was blocked by the established angiogenesis inhibitor anginex (P < 0.05). LTβ and LIGHT induced significant sprouting (P < 0.05), as did bFGF/VEGF (P < 0.01). siNIK pre-treatment of ECs led to reductions in LTβR-induced vessel formation (P < 0.05). LTβR-Ig not only blocked LTβ- or LIGHT-induced sprouting, but also RASF-induced sprouting (P < 0.05). The NIK inhibitor blocked angiogenesis induced by LTβ, LIGHT, growth factors (P < 0.05) and RASF (P < 0.01). CONCLUSION: We present a novel 3D model of synovial angiogenesis incorporating RAFLSs, ECs and RASF that mimics the in vivo situation. Using this system, we demonstrate that non-canonical NF-κB signalling promotes neovascularization and show that this model is useful for dissecting relative contributions of signalling pathways in specific cell types to angiogenic responses and for testing pharmacological inhibitors of angiogenesis.
OBJECTIVE: Angiogenesis is crucial in RA disease progression. Lymphotoxin β receptor (LTβR)-induced activation of the non-canonical nuclear factor-κB (NF-κB) pathway via NF-κB-inducing kinase (NIK) has been implicated in this process. Consequently, inhibition of this pathway may hold therapeutic potential in RA. We describe a novel three-dimensional (3D) model of synovial angiogenesis incorporating endothelial cells (ECs), RA fibroblast-like synoviocytes (RAFLSs) and RA synovial fluid (RASF) to further investigate the contributions of NF-κB in this process. METHODS: Spheroids consisting of RAFLSs and ECs were stimulated with RASF, the LTβR ligands LTβ and LIGHT, or growth factor bFGF and VEGF, followed by quantification of EC sprouting using confocal microscopy and digital image analysis. Next, the effects of anginex, NIK-targeting siRNA (siNIK), LTβR-Ig fusion protein (baminercept) and a novel pharmacological NIK inhibitor were investigated. RESULTS: RASF significantly promoted sprout formation, which was blocked by the established angiogenesis inhibitor anginex (P < 0.05). LTβ and LIGHT induced significant sprouting (P < 0.05), as did bFGF/VEGF (P < 0.01). siNIK pre-treatment of ECs led to reductions in LTβR-induced vessel formation (P < 0.05). LTβR-Ig not only blocked LTβ- or LIGHT-induced sprouting, but also RASF-induced sprouting (P < 0.05). The NIK inhibitor blocked angiogenesis induced by LTβ, LIGHT, growth factors (P < 0.05) and RASF (P < 0.01). CONCLUSION: We present a novel 3D model of synovial angiogenesis incorporating RAFLSs, ECs and RASF that mimics the in vivo situation. Using this system, we demonstrate that non-canonical NF-κB signalling promotes neovascularization and show that this model is useful for dissecting relative contributions of signalling pathways in specific cell types to angiogenic responses and for testing pharmacological inhibitors of angiogenesis.
Authors: Muhammad Farooq Rai; Hua Pan; Huimin Yan; Linda J Sandell; Christine T N Pham; Samuel A Wickline Journal: Transl Res Date: 2019-07-10 Impact factor: 7.012
Authors: Mario Rothbauer; Eva I Reihs; Anita Fischer; Reinhard Windhager; Florien Jenner; Stefan Toegel Journal: Front Bioeng Biotechnol Date: 2022-06-15
Authors: Kim C M Jeucken; Charlotte C N van Rooijen; Yik Y Kan; Lotte A Kocken; Aldo Jongejan; Abraham C I van Steen; Jaap D van Buul; Henric K Olsson; Jan Piet van Hamburg; Sander W Tas Journal: Front Immunol Date: 2022-06-13 Impact factor: 8.786
Authors: Chrissta X Maracle; Kim C M Jeucken; Boy Helder; Thomas M van Gulik; Anne Steins; Hanneke W M van Laarhoven; Sander W Tas Journal: Oncotarget Date: 2018-06-19