Facilitation of fas mediated apoptosis of human chondrocytes by the
proteasome inhibitor and actinomycin d.
J Rheumatol 2003 Mar;30(3):550-8
Kim HA, Kim YH, Song YW.
Department of Internal Medicine, Hallym University College of Medicine, Seoul,
Korea.
OBJECTIVE: We investigated the susceptibility of chondrocytes to apoptosis
induced by anti-Fas and various potentiators, and the relevant signaling
pathway. METHODS: Chondrocytes were cultured from cartilages obtained at the
time of joint replacement surgery for knee osteoarthritis (OA) or femur neck
fracture. Fas receptor ligation was performed with agonistic anti-Fas antibody
(clone CH-11) at concentrations ranging from 0.5 to 1.0 micro g/ml. Mitogen
activated protein kinase inhibitors SB203580 and PD98059, cycloheximide,
bisindolylmaleimide, actinomycin D, or MG132 were added with anti-Fas to
facilitate cell death. Chondrocyte surface expression of Fas was analyzed by
FACS, and the expression of apoptosis related proteins analyzed by Western blot.
RESULTS: Cell death increased upon coculture with 0.5 micro g/ml of anti-Fas and
0.2 micro g/ml of actinomycin D or 20 micro M MG132. Apoptosis potentiated by
actinomycin D or MG132 was effectively inhibited by caspase inhibitors,
implicating the involvement of the caspase cascade in chondrocyte apoptosis.
Compared with untreated cells or actinomycin D treated cells, cells treated with
MG132 showed distinct shifts in the distribution of surface Fas fluorescence.
Although concentrations of Bcl-2, Bax, FLICE inhibitory protein (FLIP), and Fas
ligand were unaffected by MG132 or actinomycin D, both treatments led to a
significant increase of p53. The expression of the p53 response proteins, MDM2
and p21, was elevated in MG132 treated chondrocytes. CONCLUSION: Our results
suggest that chondrocytes can be rendered sensitive to anti-Fas mediated
apoptosis by the proteasome inhibitor MG132 and the transcription inhibitor
actinomycin D. MG132 and actinomycin D show different characteristics in terms
of apoptosis signaling.
