Down‐regulation of myeloid cell leukemia 1 by epigallocatechin‐3‐gallate sensitizes rheumatoid arthritis synovial fibroblasts to tumor necrosis factor α–induced apoptosis

• Published in: Arthritis and rheumatism 2009-05, Vol.60 (5), p.1282-1293
• Main Authors: Ahmed, Salahuddin, Silverman, Matthew D., Marotte, Hubert, Kwan, Kevin, Matuszczak, Natalie, Koch, Alisa E.
• Format: Article
• Language: English
• Subjects: Anticarcinogenic Agents - pharmacology; Apoptosis - drug effects; Apoptosis - physiology; Arthritis, Rheumatoid - metabolism; Arthritis, Rheumatoid - physiopathology; Biological and medical sciences; Blotting, Western; Caspase 3 - analysis; Catechin - analogs & derivatives; Catechin - pharmacology; Cell Survival; Cells, Cultured; Diseases of the osteoarticular system; Down-Regulation - physiology; Fibroblasts - physiology; Hematologic and hematopoietic diseases; Inflammatory joint diseases; Leukemias. Malignant lymphomas. Malignant reticulosis. Myelofibrosis; Medical sciences; Myeloid Cell Leukemia Sequence 1 Protein; Osteoarthritis - metabolism; Proto-Oncogene Proteins c-bcl-2 - analysis; Synovial Membrane - cytology; Synovial Membrane - physiopathology; Tumor Necrosis Factor-alpha - pharmacology
• ISSN: 0004-3591; 1529-0131
• DOI: 10.1002/art.24488

Objective Overexpression of the antiapoptotic protein myeloid cell leukemia 1 (Mcl‐1) in rheumatoid arthritis (RA) synovial fibroblasts is a major cause of their resistance to tumor necrosis factor α (TNFα)–induced apoptosis. This study was undertaken to evaluate the efficacy of epigallocatechin‐3‐gallate (EGCG) in down‐regulating Mcl‐1 expression and its mechanism of RA synovial fibroblast sensitization to TNFα‐induced apoptosis. Methods EGCG effects on cultured RA synovial fibroblast cell morphology, proliferation, and viability over 72 hours were determined by microscopy and a fluorescent cell enumeration assay. Caspase 3 activity was determined by a colorimetric assay. Western blotting was used to evaluate the apoptosis mediators poly(ADP‐ribose) polymerase (PARP), Mcl‐1, Bcl‐2, Akt, and nuclear translocation of NF‐κB. Results In RA synovial fibroblasts, EGCG (5–50 μM) inhibited constitutive and TNFα‐induced Mcl‐1 protein expression in a concentration‐ and time‐dependent manner (P < 0.05). Importantly, EGCG specifically abrogated Mcl‐1 expression in RA synovial fibroblasts and affected Mcl‐1 expression to a lesser extent in osteoarthritis and normal synovial fibroblasts or endothelial cells. Inhibition of Mcl‐1 by EGCG triggered caspase 3 activity in RA synovial fibroblasts, which was mediated via down‐regulation of the TNFα‐induced Akt and NF‐κB pathways. Caspase 3 activation by EGCG also suppressed RA synovial fibroblast growth, and this effect was mimicked by Akt and NF‐κB inhibitors. Interestingly, Mcl‐1 degradation by EGCG sensitized RA synovial fibroblasts to TNFα‐induced PARP cleavage and apoptotic cell death. Conclusion Our findings indicate that EGCG itself induces apoptosis and further sensitizes RA synovial fibroblasts to TNFα‐induced apoptosis by specifically blocking Mcl‐1 expression and, hence, may be of promising adjunct therapeutic value in regulating the invasive growth of synovial fibroblasts in RA.