Abstract
We demonstrated previously that local, intra-articular injection of an adenoviral vector expressing human tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in a rabbit knee model of inflammatory arthritis stimulated synovial apoptosis and reduced inflammation. To examine whether intra-articular injection of recombinant chimeric human TRAIL protein (rTRAIL) also induces apoptosis of proliferating rabbit synovium and reduces inflammation, we used an experimental rabbit arthritis model of rheumatoid arthritis, induced by intra-articular introduction of allogeneic fibroblasts genetically engineered to secrete human IL-1β. Analysis of synovium isolated from the rabbits treated with intra-articular injection of rTRAIL, relative to saline control, showed areas of extensive acellular debris and large fibrous regions devoid of intact cells, similar to adenoviral mediated TRAIL gene transfer. Extensive apoptosis of the synovial lining was demonstrated using TUNEL analysis of the sections, corresponding to the microscopic findings in hematoxylin and eosin staining. In addition, leukocyte infiltration into the synovial fluid of the inflamed knee joints following rTRAIL treatment was reduced more than 50% compared with the saline control. Analysis of the glycosaminoglycan synthetic rate by cultured cartilage using radiolabeled sulfur and cartilage histology demonstrated that rTRAIL did not adversely affect cartilage metabolism and structure. Analysis of serum alanine aminotransferase showed that intra-articular injection of rTRAIL did not have adverse effects on hepatic function. These results demonstrate that intra-articular injection of rTRAIL could be therapeutic for treating pathologies associated with rheumatoid arthritis.
Highlights
Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a type II transmembrane protein that was initially identified according to the homology of its extracellular domain with CD95L (FasL), TNF-α and lymphotoxin-α [1,2]
Results recombinant chimeric human TRAIL protein (rTRAIL) induces synovial apoptosis following intraarticular injection To determine whether rTRAIL can induce apoptosis of rabbit synoviocytes, synovial fibroblasts from inflamed knee joints of rabbit were cultured in 24-well culture plates and co-incubated with various doses (5, 20, and 40 μg/ml) of rTRAIL for 48 h. rTRAIL exhibited a small but insignificant effect on rabbit synovial fibroblast viability relative to saline control as determined using an MTT assay (Figure 2)
Addition of increasing doses of rTRAIL to two different cultures of human Rheumatoid arthritis (RA) synovial fibroblasts showed marginal effects on cell viability. These results suggest that rTRAIL is not effective in inducing apoptosis of synovial cells in culture
Summary
Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a type II transmembrane protein that was initially identified according to the homology of its extracellular domain with CD95L (FasL), TNF-α and lymphotoxin-α [1,2]. TRAIL is able to induce apoptosis of a wide variety of human tumor cells, but generally appears not to affect normal cells [5]. The removal of the synovial pannus by either surgery [8] or radioactive isotopes [9] has proven to be useful in treating RA in certain cases, resulting in pain relief and better outcome. These methodologies, have inherent limitations in the treatment
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