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Fas-associated death domain protein is a Fas-mediated apoptosis modulator in synoviocytes
K Okamoto1, T Kobayashi, T Kobata
1Rheumatology, Immunology, and Genetics Program, Institute of Medical Science, St. Marianna University School of Medicine, 2-16-1 Sugao, Miyamae-ku, Kawasaki 216-8512, Japan.
Objective:
To understand the intracellular regulatory mechanisms in Fas-mediated apoptosis of synoviocytes, we examined the involvement of caspases [caspase-1/ICE (interleukin-1beta converting enzyme), caspase-3/CPP32, and caspase-8/FLICE] and Fas-associated death domain protein (FADD) forming a death-inducing signalling complex (DISC) in Fas-mediated apoptosis of synoviocytes.
Methods:
Synoviocytes were obtained from rheumatoid arthritis (RA) and osteoarthritis (OA) patients. The number of dead cells was counted after treatment with anti-Fas monoclonal antibody in the presence of caspase-1-, -3-, or -8-specific inhibitors. The involvement of caspases and FADD in Fas-mediated apoptosis of RA synoviocytes was examined by immunoblot and immunoprecipitation analyses.
Results:
RA synoviocytes expressed high levels of caspase-3, caspase-8, and FADD compared with OA synoviocytes. Interestingly, Fas ligation activated caspase-8 and caspase-3 with the cleavage of poly(ADP-ribose) polymerase (PARP), corresponding to apoptosis of RA synoviocytes. Furthermore, specific inhibitors for caspase-3 and caspase-8 but not caspase-1 suppressed Fas-induced apoptosis of RA synoviocytes in a dose- and time-dependent manner. Caspase-8-specific inhibitor suppressed the activation of caspase-3 after Fas ligation on RA synoviocytes. Importantly, FADD was selectively recruited to the Fas death domain during Fas-mediated apoptosis of RA synoviocytes, consistent with sensitivity to the Fas-mediated apoptosis.
Conclusion:
Our findings suggest that Fas-mediated apoptosis in synoviocytes may be regulated at the level of recruitment of FADD to the DISC, subsequently leading to the activation of the FADD/caspase-8/caspase-3 signalling pathway.
Insights
Fas-mediated apoptosis in rheumatoid arthritis synoviocytes involves the FADD/caspase-8/caspase-3 pathway. Recruitment of Fas-associated death domain protein (FADD) to the death-inducing signaling complex (DISC) regulates this process.
Area of Science:
- Immunology
- Cell Biology
- Molecular Biology
Background:
- Synoviocytes play a crucial role in joint inflammation, particularly in rheumatoid arthritis (RA).
- Fas-mediated apoptosis is a key mechanism for regulating cell numbers in tissues.
Purpose of the Study:
- To elucidate the intracellular mechanisms regulating Fas-mediated apoptosis in synoviocytes.
- To investigate the roles of caspases and Fas-associated death domain protein (FADD) in this process.
Main Methods:
- Synoviocytes from RA and osteoarthritis (OA) patients were treated with anti-Fas antibody and caspase inhibitors.
- Immunoblot and immunoprecipitation analyses were used to examine protein involvement.
Main Results:
- RA synoviocytes showed higher levels of caspase-3, caspase-8, and FADD than OA synoviocytes.
- Fas ligation activated caspase-8 and caspase-3, leading to apoptosis, which was suppressed by caspase-3 and -8 inhibitors.
- FADD recruitment to the Fas death domain was observed during Fas-mediated apoptosis.
Conclusions:
- Fas-mediated apoptosis in synoviocytes is regulated by FADD recruitment to the DISC.
- This initiates the FADD/caspase-8/caspase-3 signaling cascade, driving apoptosis.