4.0 Article

Down-Regulation of Myeloid Cell Leukemia 1 by Epigallocatechin-3-Gallate Sensitizes Rheumatoid Arthritis Synovial Fibroblasts to Tumor Necrosis Factor α-Induced Apoptosis

Journal

ARTHRITIS AND RHEUMATISM
Volume 60, Issue 5, Pages 1282-1293

Publisher

WILEY
DOI: 10.1002/art.24488

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Funding

  1. NIH [AT-003633, AR-055741, AI-40987, AR-48267]
  2. American Heart Association Postdoctoral Fellowship [0425749Z]
  3. French Society of Rheumatology, Lavoisier, and the Philippe Foundation
  4. Frederick G. L. Huetwell and William D. Robinson, MD, Professorship in Rheumatology
  5. Office of Research and Development, Medical Research Service, Department of Veterans Affairs

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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 a (TNF alpha)-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 TNF alpha-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-kappa B. Results. In RA synovial fibroblasts, EGCG (5-50 mu M) inhibited constitutive and TNF alpha-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 TNF alpha-induced Akt and NF-kappa B pathways. Caspase 3 activation by EGCG also suppressed RA synovial fibroblast growth, and this effect was mimicked by Akt and NF-kappa B inhibitors. Interestingly, Mcl-1 degradation by EGCG sensitized RA synovial fibroblasts to TNF alpha-induced PARP cleavage and apoptotic cell death. Conclusion. Our findings indicate that EGCG itself induces apoptosis and further sensitizes RA synovial fibroblasts to TNF alpha-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.

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