Regulation of apoptosis in fibroblast‐like synoviocytes by the hypoxia‐induced Bcl‐2 family member Bcl‐2/adenovirus E1B 19‐kd protein–interacting protein 3

Abstract

Objective. Rheumatoid arthritis (RA) synovial hyperplasia is related in part to a resistance to apoptosis exhibited by fibroblast-like synoviocytes (FLS). Since hypoxia is a regulator of apoptosis, and since RA synovium is hypoxic, we conducted this study to examine the effects of hypoxia on the Bcl-2 pathway and the role this may play in regulating apoptosis in FLS.Methods. Synovium samples from RA patients, osteoarthritis (OA) patients, and normal subjects were used for immunohistologic assessments and for generating FLS lines in vitro. FLS were stimulated under conditions of hypoxia (1% O 2 ) and using 100 M CoCl 2 to simulate the effects of severe hypoxia. Changes in the gene expression profile of FLS were evaluated using microarrays and were confirmed by quantitative polymerase chain reaction (PCR). Changes in protein expression were detected by Western blotting. The effect of transient transfection with a BNIP3 plasmid on the apoptosis of FLS was evaluated in the presence and absence of cytokines.Results. Gene expression profiling demonstrated that BNIP3 was unique among the BCL2 family, in that it was induced by hypoxia in FLS. Quantitative PCR indicated a 2-3-fold induction of BNIP3 messenger RNA, and Western blotting showed a 3-5-fold increase in the 30-kd Bcl-2/adenovirus E1B 19-kd proteininteracting protein 3 (BNIP-3) monomer. BNIP-3 was widely expressed in RA synovium and was prominent in FLS from the lining layer. Overexpression of BNIP3 increased FLS apoptosis under hypoxic conditions, an effect that was inhibited by tumor necrosis factor ␣ and interleukin-1␤.Conclusion. The proapoptotic protein BNIP-3 is induced in FLS by hypoxia and is widely expressed in RA synovium, but its proapoptotic effects may be inhibited in vivo by proinflammatory cytokines. Since overexpression of BNIP3 in FLS increases apoptosis, this may provide a novel approach for controlling synovial hyperplasia in RA.

Cell death in rheumatoid arthritis

“…In addition, they provide a link between cytokine-mediated stimulation of these cells and their resistance to apoptotic cell-death. The latter notion is at least in part supported also by most recent observations that systematically analysed the Bcl-2 pathway in RA-FLS [20]. In this study, the apoptotic Bcl-2 family member Bcl-2/adenovirus E1B 19-kd protein-interacting protein 3 (BNIP3) was identified as being uniquely up-regulated by hypoxia but inhibited in its pro-apoptotic action by TNFalpha and IL-1.…”

Synovial tissue hypoxia and inflammation in vivo

“…Lining layer SFCs undergo phenotypic changes as they invade adjacent cartilage and bone, while RA SFCs can significantly promote endothelial cell activation and leucocyte recruitment compared to normal or osteoarthritis (OA) SFCs 10 11. The precise mechanisms involved in regulation of persistent infiltration and invasion are unclear, however high levels of cytokines and impaired apoptotic pathways in the microenvironment are implicated 12–14…”

Effect of Bone Marrow Mesenchymal Stem Cell Transplant on Synovial Proliferation in Rats With Type II Collagen-Induced Arthritis

“…Most hyperplasia synovial cells are fibroblastlike synoviocytes. 11 Serum vascular endothelial growth factor, TNF-α, and IL-17 are important factors in RA synovitis.…”