Arthritis & Rheumatism, Volume 63,
November 2011 Abstract Supplement

Abstracts of the American College of
Rheumatology/Association of Rheumatology Health Professionals
Annual Scientific Meeting
Chicago, Illinois November 4-9, 2011.


Downregulation of Mcl-1 by Ursolic Acid, a Pentacyclic Triterpenoid, Sensitizes Rheumatoid Arthritis Synovial Fibroblasts for TRAIL-Induced Apoptosis.

Zielinski,  Carolyn, Beamer,  Maria, Ahmed,  Salahuddin

Background/Purpose:

In rheumatoid arthritis (RA), enhanced expression of Mcl-1 contributes significantly to the resistance of synovial fibroblasts to apoptosis. In the present study, we evaluated the effect of ursolic acid (UA), a potent anti-inflammatory pentacyclic triterpenoid, on TRAIL-induced apoptosis in RA synovial fibroblasts.

Methods:

Effects of UA (2.5–20 mM) and TRAIL (100 ng/ml), alone or in combination, on the cultured RA synovial fibroblast morphology and cell viability were determined through 72 hours of observation by microscopy and a colorimetric MTT cell viability assay. Caspase-3 activity was determined by a colorimetric assay. Apoptosis was measured by the cleavage of poly-ADP-ribose polymerase (PARP). Western blotting was used to evaluate the apoptosis mediators, Bcl-2, Mcl-1, Noxa, and Bax; the cell survival protein Akt; and nuclear translocation of nuclear factor-kB (NF-kB).

Results:

UA (2.5–20 mM) decreased the cell viability of RA synovial fibroblasts in a dose-dependent manner and synergistically enhanced TRAIL-induced cell death by ~15% as compared to the UA alone treated group. Importantly, UA (5–20 mM) selectively induced RA synovial fibroblast Mcl-1 degradation within 24 hours of treatment, with no marked effect on the expression levels of Bcl-2 or Bax (p<0.05; n=3–5). In addition, UA treatment dose-dependently induced the expression of BH3-only Noxa, which may partly contribute to the inhibition of Mcl-1 expression by UA (p<0.05; n=3). Inhibition of Mcl-1 by UA resulted in the sensitization of RA synovial fibroblasts to TRAIL-induced PARP cleavage and apoptotic cell death. The addition of MG132 (a proteasome inhibitor; 10 mM) blocked the inhibitory effect of UA on Mcl-1 expression and downstream apoptotic events, implicating the proteasomal degradation of Mcl-1 as a fundamental mechanism of RA synovial fibroblast sensitization to apoptosis by UA. Furthermore, evaluation of the signaling events showed that UA specifically blocked the constitutive and TRAIL-activated phospho-Akt expression and the nuclear translocation of NF-kB to induce apoptosis in RA synovial fibroblasts.

Conclusion:

Our novel findings indicate that UA itself induces apoptosis and further sensitizes RA synovial fibroblasts to TRAIL-induced apoptosis by specifically blocking Mcl-1 expression partly through Noxa expression. Thus, UA may be a promising adjunct therapeutic option in regulating the invasive growth of synovial fibroblasts in RA.

To cite this abstract, please use the following information:
Zielinski, Carolyn, Beamer, Maria, Ahmed, Salahuddin; Downregulation of Mcl-1 by Ursolic Acid, a Pentacyclic Triterpenoid, Sensitizes Rheumatoid Arthritis Synovial Fibroblasts for TRAIL-Induced Apoptosis. [abstract]. Arthritis Rheum 2011;63 Suppl 10 :370
DOI:

Abstract Supplement

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