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Objective

To examine the impact of Toll-like receptor 5 (TLR-5) on endothelial cell function in rheumatoid arthritis (RA) and vascularization in collagen-induced arthritis (CIA).

Methods

Endothelial cell migration and tube formation assays were used to demonstrate the direct role of TLR-5 ligation in angiogenesis. Mice with CIA were treated with the TLR-5 agonist flagellin to document the effect of TLR-5 ligation in RA pathology. Vascularization in CIA was determined by immunohistochemical analysis and determination of cytokine levels in ankle joints. Spleen Th17 cells and joint interleukin-17 (IL-17) were quantified by fluorescence-activated cell sorting analysis and enzyme-linked immunosorbent assay. The development of Th17 cells induced by TLR-5 ligation was validated in RA peripheral blood mononuclear cells.

Results

Ligation of TLR-5 to endogenous ligands expressed in RA synovial fluid contributed to endothelial cell infiltration and tube formation. Furthermore, treatment with flagellin after the onset of CIA exacerbated joint inflammation; in contrast, inflammation in control mice remained at a plateau phase. We showed that TLR-5–enhanced disease severity was attributable to Th17 cell differentiation and joint vascularization in CIA. Examination of the underlying mechanism using RA peripheral blood mononuclear cells documented that ligation of TLR-5 in myeloid cells and production of Th17–promoting cytokines were necessary for Th17 cell polarization. Additionally, we demonstrated that blockade of the IL-17 cascade markedly reduced endothelial cell migration activated by flagellin-conditioned medium, suggesting that TLR-5 ligation can mediate RA angiogenesis either directly by attracting endothelial cells or indirectly by fostering Th17 cell development.

Conclusion

Our data demonstrate a novel role for TLR-5 in RA angiogenesis; thus, TLR-5 may be a promising new target for RA treatment.