Expression of C/EBPβ in rheumatoid arthritis (RA) synovium.
(A) Expression and distribution of C/EBPβ and RANKL in RA synovium by immunofluorescence staining. RANKL: anti-RANKL antibodies; C/EBPβ: anti-C/EBPβ antibodies; IgG: normal IgG antibodies. The appropriate species Alexa Fluor 488 and 568-conjugated antibodies were used as secondary antibodies and 4′,6-diamidino-2-phenylindole dihydrochloride (DAPI) was applied as a nuclear stain. Original magnification, 200×. Magnified views (600×) at the synovial lining layer are also shown. (B,
C) Western blotting for C/EBPβ protein consisting of 38 kD liver-enriched activator protein (LAP*), 36 kD LAP and 20 kD liver-enriched inhibitory protein (LIP) (B) Experiments with whole extracts of primary cultured passage-1 RA fibroblast-like synoviocytes (RA-FLS) from three patients (RA1, RA2 and RA3) and normal human fibroblast-like synoviocytes (HFLS). A band on 30 kD in RA2 might be a non-specific band. (C) RA-FLS were treated for 48 hours with IL-1β (2 ng/ml), TNF-α (10 ng/ml), IL-6 (5 ng/ml), or IL-17 (100 ng/ml), respectively. Nuclear extracts and cytoplasmic protein from RA-FLS treated with cytokines were analyzed. Representative data from three independent experiments are shown.