Objective. To understand the intracellular regulatory mechanisms in Fas-med
iated apoptosis of synoviocytes, we examined the involvement of caspases [c
aspase-1/ICE (interleukin-1 beta converting enzyme), caspase-3/CPP32, and c
aspase-8/FLICE] and Fas-associated death domain protein (FADD) forming a de
ath-inducing signalling complex (DISC) in Fas-mediated apoptosis of synovio
cytes.
Methods. Synoviocytes were obtained from rheumatoid arthritis (RA) and oste
oarthritis (OA) patients. The number of dead cells was counted after treatm
ent with anti-Fas monoclonal antibody in the presence of caspase-1-, -3-, o
r -8-specific inhibitors. The involvement of caspases and FADD in Fas-media
ted apoptosis of RA synoviocytes was examined by immunoblot and immunopreci
pitation 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 (P
ARP), corresponding to apoptosis of RA synoviocytes. Furthermore, specific
inhibitors for caspase-3 and caspase-8 but not caspase-1 suppressed Fas-ind
uced apoptosis of RA synoviocytes in a dose- and time-dependent manner. Cas
pase-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, con
sistent with sensitivity to the Fas-mediated apoptosis.
Conclusion. Our findings suggest that Fas-mediated apoptosis in synoviocyte
s may be regulated at the level of recruitment of FADD to the DISC, subsequ
ently leading to the activation of the FADD/caspase-8/caspase-3 signalling
pathway.