Fig. 2: Synovial fibroblasts (SF) are the major cell type displaying GALA in arthritic synovium.

a Co-staining of VVL, mouse fibroblast marker Vimentin (VIM), immune cell marker CD45 and nuclei in the synovium from 7-day CAIA mice joint section, relative enrichment areas for immune cells (circle) and fibroblasts (rectangle) in the pannus tissues is indicated. Scale bar, 50 μm. S synovium, B bone. b Representative HE image of synovial tissues obtained from RA patients. *: Infiltration of immune cells in the sub-lining of RA synovium; SL synovial lining. Scale bar, 100 μm. c Representative IF images of human RA synovium from (b); lining synovial fibroblasts (SF) identified by FAPα (arrowheads) and immune cells identified by CD45 in the sub-lining (limit of lining indicated by the dashed line). Scale bar, 50 μm. d Quantification of helix pomatia lectin (HPL) levels per cell in patient purified HCSF, OASF and RASF under basal conditions and upon presence of cytokines IL1β and TNFα (CYTO) plated on plastic (No coating) or plated on cartilage ECM or collagen I enriched ECM. Data are the mean ± SEM of three independent experiments. **, p < 0.01 (p = 0.0014; No coating, UT HCSF vs RASF) ***, p < 0.001 (p = 0.0001; No coating, CYTO, HCSF. vs OASF), ∗∗∗∗, p < 0.0001, NS not significant (p = 0.0958; No coating, CYTO, HCSF vs OASF) (Two-way ANOVA test). e Representative immunofluorescence images of HCSF, OASF, and RASF stained with HPL, Golgi marker Giantin and Hoechst on plastic wells or cartilage ECM treated wells. Scale bar, 20 μm. f Representative images of wildtype SW982 cells stimulated with or without CYTO and cartilage ECM stained with GALNT2, ER marker PDIA4 and Golgi marker TGN46. Images were acquired at 100× magnification. Scale bar, 10 μm. g Quantification of Mander’s coefficient of GALNT1 and ER marker PDIA4 in (f). M1 represents the fraction of GALNT1 staining coincident with the ER and M2 represents the fraction of the ER marker coincident with GALNT1 staining. 23 and 36 cells from unstimulated and CYTO-cartilage ECM stimulated respectively. Data present are the mean ± SD of cells quantified from two experiments. ****, p < 0.0001 (Two sided Student’s t-test, two-tailed p-value).