Expression of CD95 and CD95L by OA chondrocytes. (a) Micrographs of immunohistochemcal analysis of CD95 expression in normal (bottom panels) and OA cartilage (top panels). Frozen sections from normal and OA articular cartilage were incubated with a monoclonal antibody CH 11 against CD95. Different (original) magnifications are indicated. Results are representative of two normal and four OA cartilage samples. (b) mRNA levels of CD95L in primary OA chondrocytes by real-time RT-PCR analysis. Total RNA was isolated from chondrocytes from OA cartilage following treatment as indicated below. *P < 0.05 versus treatment with Control Ab (a mouse isotype control antibody IgM). Each bar represents the mean ± standard deviation of three experiments. OA chondrocytes (donor age 63 years) stained with annexin V and PI. Results are representative of three OA cartilage samples. (c) Micrographs of immunocytochemical analysis of collagens type II and type I in OA chondrocytes following treatment as indicated below. Chondrocytes were reacted with rhodomaine mAb against type II or type I collagen, respectively. Chondrocyte nuclei were indicated by Hoechest blue dye staining. Anti-CD95: treatment with a mAb anti-Fas CH 11 (100 ng/ml) in serum-free medium for 17 hours. SB: treatment with SB203580 (10 μmol/l) for 17 hours. SB+Anti-CD95: treatment with SB203580 for 2 hours followed by anti-Fas treatment for 17 hours. Control: cells were treated with DMSO only. Results are representative of 3 OA cartilage samples. CD95L, CD95 ligand; DMSO, dimethyl sulfoxide; mAb, monoclonal antibody; OA, osteoarthritis; PI, propidium iodide.