The demonstration that one of the mechanisms of action of
bisphosphonates (BPs) is the induction of osteoclast and macrophage apoptosis, suggests a potent therapeutic role for the BPs and other apoptosis-modulating agents in the management of periprosthetic
osteolysis. The purpose of this study was to improve our understanding of the basic underlying molecular events leading to the inhibitory effect of
pamidronate on the macrophage response to
ultra-high-molecular-weight polyethylene (
UHMWPE) particles. Murine J774 macrophages were incubated for 0-72 h in the presence of
UHMWPE particles and/or
pamidronate.
TNF-alpha release was measured by ELISA while
poly(ADP-ribose)polymerase (PARP) expression was measured by Western blot.
DNA was analyzed on
agarose. The appearance of PARP fragment and the fragmentation of
DNA were used as markers of apoptosis. We observed a dose-dependent response to
UHMWPE particles with
TNF-alpha release reaching 4, 10, and 19 times control with 10, 25, and 125 particles/macrophage, respectively.
UHMWPE particles (25 particles/macrophage) stimulate
TNF-alpha release by
a factor of 10, 7, and 6 after 24, 48, and 72 h, respectively, indicating a rapid stimulating effect of
UHMWPE particles on
TNF-alpha release. Our results also showed that
at 10 particles/macrophage,
pamidronate inhibits
UHMWPE-induced
TNF-alpha release by 12%, 14%, and 23% respectively after 24, 48, and 72 h (p<0.05 vs. 24 and 48 h). With 25 particles/macrophage, the inhibition of
TNF-alpha reached 9%, 12%, and 15% after 24, 48, and 72 h (p<0.05 vs. 24 h), respectively. There is no significant difference between the inhibition by
pamidronate of
TNF-alpha release induced by 125 particles/macrophages at 24, 48, and 72 h. When cells are pre-incubated for 48 h with
pamidronate prior to addition of
UHMWPE particles for 24 h, we observed an increased inhibition of
TNF-alpha compared to the co-incubation protocol. The inhibiting effect of
pamidronate reaches 56% when pre-incubated with macrophages prior to incubation with 10 particles of
UHMWPE/macrophage (p<0.05 vs. co-incubation).Co-incubation of
pamidronate with
UHMWPE particles also led to the appearance of the proteolytic PARP fragment after 24 h incubation. We also demonstrated the stimulation of DNA fragmentation (
DNA laddering) after 48-72 h with
pamidronate. The proteolytic cleavage of PARP, an early event in the induction of apoptosis, precedes the inhibition of
UHMWPE particle-induced
TNF-alpha release by
pamidronate whereas the fragmentation of
DNA, a late apoptotic event, parallels this inhibition. Our results suggest the induction of macrophage apoptosis is associated with the inhibitory effect of
pamidronate on
TNF-alpha release. There is a need for the development of medical management of periprosthetic
osteolysis. The demonstration that drugs such as
pamidronate induce specific apoptosis-related pathways in macrophages contributes data for a rational approach in the treatment and/or prevention of periprosthetic
osteolysis.