Immunomodulatory effects of Simvastatin in Rheumatoid arthritis synoviocytes are mediated via BNIP3 (119.1)

Abstract

Abstract We investigated the anti-inflammatory effects of Simvastatin (SMV) in vitro, in fibroblast-like synoviocyte (FLS) cell lines from control and RA subjects, in the presence of an activated TLR2 signaling pathway. Cells (10^5 cells/well) were grown in 12-well culture plates and treated with SMV in triplicates for 4 hours and then treated with TLR2 ligand (Pam3CSK4) for 24-hours. Cytotoxicity was assessed by Alamar-blue assay, interleukin-6 (IL-6) by ELISA, reactive oxygen species (ROS) and apoptosis by flow cytometry. Inositol phosphates (IPs) were measured by labeling IPs with tritium in cell culture, followed by anion exchange chromatography and analyzed in a beta scintillation counter. Gene expression analysis of 84-ROS associated genes was performed using a real time PCR array profiling kit from SABiosciences. SMV caused dose-dependent cytotoxicity in both cell lines and this was due to apoptosis. Basal apoptosis was higher in normal than RA-FLS (14% vs 4.3%). ROS was increased modestly (5% basal to 12%). IL-6 increased and IP-1 and IP-2 decreased with SMV treatment. Gene expression of GSR, TXNRD1, PRDX1, SOD2, MT3 and PTGS2 were significantly upregulated. BNIP3 expression was increased 2200-fold in RA-FLS after SMV/TLR2 treatment. These findings indicate that SMV induce FLS apoptosis in RA by upregulating BNIP3 and FLS react to survive by increasing IL-6, decreasing IP-1 and IP-2 and upregulating anti-oxidant genes.