Figure 4

Robo and Slit expression in RASF in three different passages. (a) Robo1, Robo2, Robo3, Slit1, Slit2, and Slit3 mRNA expression in rheumatoid arthritis synovial fibroblasts (RASF; n = 5) was quantified by real-time (RT)-PCR starting from passage 3 (P3) over passage 5 (P5) to passage 10 (P10; logarithmic scale). Robo1 and Robo2 expression decrease slightly in RASF over the passages. Robo3 mRNA levels dropped dramatically in P5 and P10 compared with P3. No considerable differences in mRNA expression were measured with Slit1, Slit2, and Slit3 over the passages. (b) Protein expression of Robo3 in RASF of P3 and P5 was analyzed by FACS using a Robo3 specific antibody. RASF in early passages showed a shift (I) compared with negative control, whereas cells of later passages (II) did not reveal Robo3 expression. (c) In vitro assays on cartilage destruction revealed significant reduction of free glycosaminoglycans (GAGs) after coculture with RASF of late passages compared with early passages.