Down‐regulation of microRNA‐34a* in rheumatoid arthritis synovial fibroblasts promotes apoptosis resistance

<jats:title>Abstract</jats:title><jats:sec><jats:title>Objective</jats:title><jats:p>To investigate the expression and effect of the microRNA‐34 (miR‐34) family on apoptosis in rheumatoid arthritis synovial fibroblasts (RASFs).</jats:p></jats:sec><jats:sec><jats:title>Methods</jats:title><jats:p>Expression of the miR‐34 family in synovial fibroblasts with or without stimulation with Toll‐like receptor (TLR) ligands, tumor necrosis factor α (TNFα), interleukin‐1β (IL‐1β), hypoxia, or 5‐azacytidine was analyzed by real‐time polymerase chain reaction (PCR). Promoter methylation was studied by combined bisulfite restriction analysis. The effects of overexpression and silencing of miR‐34a and miR‐34a* on apoptosis were analyzed by annexin V/propidium iodide staining. Production of X‐linked inhibitor of apoptosis protein (XIAP) was assessed by real‐time PCR and immunohistochemistry analysis. Reporter gene assay was used to study the signaling pathways of miR‐34a*.</jats:p></jats:sec><jats:sec><jats:title>Results</jats:title><jats:p>Basal expression levels of miR‐34a* were found to be reduced in synovial fibroblasts from RA patients compared to osteoarthritis patients, whereas levels of miR‐34a, miR‐34b/b*, and miR‐34c/c* did not differ. Neither TNFα, IL‐1β, TLR ligands, nor hypoxia altered miR‐34a* expression. However, we demonstrated that the promoter of miR‐34a/34a* was methylated and showed that transcription of the miR‐34a duplex was induced upon treatment with demethylating agents. Enforced expression of miR‐34a* led to an increased rate of FasL‐ and TRAIL‐mediated apoptosis in RASFs. Moreover, levels of miR‐34a* were highly correlated with expression of XIAP, which was found to be up‐regulated in RA synovial cells. Finally, we identified XIAP as a direct target of miR‐34a*.</jats:p></jats:sec><jats:sec><jats:title>Conclusion</jats:title><jats:p>Our data provide evidence of a methylation‐specific down‐regulation of proapoptotic miR‐34a* in RASFs. Decreased expression of miR‐ 34a* results in up‐regulation of its direct target XIAP, thereby contributing to resistance of RASFs to apoptosis.</jats:p></jats:sec>