Abstract 17

The Expression of TRAIL and TRAIL Receptors in Cultured Synovial Fibroblasts from Rheumatoid and Osteoarthritis

Anak Agung Sagung Sri Kencana Dharmapatni^1, David M. Findlay ^2, Andreas Evdokiou^2, Holding C^1, Wallwork, M^3, David R.Haynes¹

¹ Dept. of Pathology, University of Adelaide, South Australia, Australia
² Dept. of Orthopaedics and Trauma, University of Adelaide, and Hanson Institute South Australia, Australia
³ Adelaide Microscopy, University of Adelaide, South Australia, Australia

In this study, we investigated TRAIL (Tumor necrosis factor related apoptosis inducing ligand) and TRAIL receptor expression on cultured synovial fibroblasts and correlate this with their ability to undergo TRAIL and disease modifying anti rheumatic drugs (DMARDs)- induced apoptosis.

Synovial tissues were obtained from patients undergoing arthroscopy biopsy. Synovial fibroblasts were isolated by digestion with collagenase and dispase for 11/2- 2 hours. After repeated passaging, the cells were used at passage 3 to 6. Immunocytochemistry and immunoflurescence were used to investigate the expression of TRAIL and TRAIL receptors on these cells. Apoptosis was induced with TRAIL, Methotrexate and Actinomycin D or combination of these reagents. DAPI (4'-6-Diamidino-2-phenylindole) staining were used to examine the morphology of cells that undergoing apoptosis. Cell viability was quantitavely examined by crystal violet staining.

We found that both synovial fibroblasts from RA And OA constitutively express TRAIL, TRAIL Receptor 4 and Osteoprotegerin (OPG). Induction of apoptosis with soluble TRAIL (100 to 1000 ng/ml) for 24 hours did not induce cell death in rheumatoid synovial fibroblasts from both rheumatoid (RA) and osteoarthritis (OA). These cells were sensitive to Actinomycin D-induced apoptosis. Combination with soluble TRAIL and pretreatment with TNF _ and IL-1 _ enhanced the sensitivity of RASF to Actinomycin D-induced apoptosis.

In conclusion, synovial fibroblasts from RA and OA are relatively resistent to TRAIL induced apoptosis but sensitive to Actinomycin D induced apoptosis. Expression of TRAIL decoy receptors (TRAIL R4 and OPG ) may be involved in the resistance of these cells to TRAIL induced apoptosis.

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