Background/Purpose: Rheumatoid arthritis (RA) synovial fibroblasts (SF) are central cells of cartilage destruction and neoangiogenesis. RASF show an increased migratory potential in the synovium towards sites of cartilage degradation. In the SCID mouse model, RASF were able to migrate through the vascular system. The interaction of RASF with endothelial cells (EC) is important for RASF-mediated angiogenesis and migration. Vessel sprouting and EC activation are two key mechanisms induced by interaction of RASF and EC. In this study, the kinetics of vessel growth by RASF in the SCID mouse model was analyzed as well as the E-selectin ligand (CD15s) expression in the synovial membrane, a mechanism potentially involved in RASF adhesion to EC.

Methods: Using 5µm frozen sections RA and osteoarthritis (OA) synovium fluorescence double staining was performed with FITC-labeled anti-CD15s and Cy3-labeled anti-vimentin (fibroblast) antibodies. Medium of cultured RA- and OASF was replaced with RA serum or serum from healthy donors for 48h. Then, cells were stained for CD15s. SCID mouse model (implants n=3-7/time point): the ipsilateral site contained healthy cartilage together with 1.5×10⁵RASF in a carrier matrix, the contralateral site contained cartilage without RASF. Neovascularization next to and into the implants was determined after 3 to 60 days. Implant images were taken, implants removed and angiogenesis analyzed by CD31 staining.

Results: CD15s signals were detectable in all RA tissues (n=12). In 67% CD15s signals were co-localized with vimentin, mainly located in sublining (50%) but also located in vessels (33%). After stimulation with RA serum, 80% of RASF (n=5) and 33% of OASF (n=3) showed an increased CD15s expression. Serum stimulation from healthy donors did not alter CD15s. Implant evaluation showed the early presence of truncated vessels (day 3-12) especially at the ipsilateral site which was reduced over time. Vessels in the murine skin close to ipsilateral implants increased in size and perfusion. Late during vascularization, very small vessels sprouted into the carrier matrix (day 9-60). Anti-CD31 staining showed that neoangiogenesis (<10 cell sizes) started at day 9 at both sites. Later (day 18), EC were detectable deep within the carrier matrix (>10 cell layers). Then (> day 27) the whole carrier matrix showed detectable EC signals. At this time the vessel lumen in the implants increased.

Conclusion: CD15s positive RASF were detectable in RA synovium and some of them located in vessels. RA serum increased CD15s expression by RASF which in turn allows RASF to adhere to EC. Implants co-implanted with RASF showed a stronger vessel formation at early time points then RASF-free implants. Vessel formation around and into implants showed an unexpected dynamics, starting with newly formed truncated vessels surrounding the implants especially at the ipsilateral site. Then these vessels disappear and EC invade into the implants, increasing over time. The perfusion of the surrounding vessels increases solely at the ipsilateral site. In summary, RASF are actively involved in neovascularization showing distinctive pattern in vessel formation. In turn, vascularization allows RASF long distance migration.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/neovascularization-and-cd15s-influence-long-distance-migration-of-synovial-fibroblasts-from-patients-with-rheumatoid-arthritis/