Differential apoptotic response of J774 macrophages to alumina and ultra-high-molecular-weight polyethylene particles

Abstract

We recently identified apoptosis in in vitro wear particle-stimulated macrophages. The recent explosion of interest in apoptosis lies in the fact that it is under positive and negative regulation through evolutionary conserved biochemical pathways. It may also be possible to modulate macrophage apoptosis in the treatment of periprosthetic osteolysis. The purpose of this study was to compare the macrophage response to identically sized particles of alumina ceramic (Al 2O 3) and ultra-high-molecular-weight polyethylene (UHMWPE) in terms of TNF-α release and induction of apoptosis. J774 mouse macrophages were incubated for 0–24 h in the presence of Al 2O 3 and UHMWPE particles. TNF-α release was measured by ELISA; Poly(ADP-ribose)polymerase (PARP) and caspase-3 expression was analyzed by Western blot; DNA fragmentation (DNA laddering) was visualized on agarose gel containing ethidium bromide. Al 2O 3 particles induced TNF-α release after 4 h incubation with concentrations reaching 483 and 800 pg/ml after 24 h with 125 and 250 particles/macrophage, respectively (control=161 pg/ml) ( P<0.05 vs. control). The same concentrations of UHMWPE particles induced a much larger and significant TNF-α release after only 1 h incubation, increasing up to 6250 pg/ml after 24 h ( P<0.05 vs. control). Western blot analysis demonstrated that the active caspase-3 fragment (17 kDa) and the proteolytic PARP fragment (85 kDa) were expressed after 2 h incubation with 125 and 250 Al 2O 3 particles/macrophage. The active caspase-3 and the PARP fragment had lower expression and appeared after a longer incubation time (8 h) with 125 and 250 UHMWPE particles/macrophage. Finally, DNA fragmentation (DNA laddering) was observed after 16 h with 125 and 250 particles of Al 2O 3 per macrophage whereas no laddering was induced by UHMWPE particles even after 24 h incubation. This study shows that although both Al 2O 3 and UHMWPE particles induce TNF-α release, this stimulation was much greater (8–10 times higher) with UHMWPE than Al 2O 3 ( P<0.05 vs. control). As well, the induction of apoptosis, as measured by activation of caspase-3, PARP cleavage and DNA laddering, is different for these two particles, being faster and more important with Al 2O 3 than UHMWPE. We hypothesize that the ability of Al 2O 3 to induce macrophage apoptosis may explain the lower TNF-α release observed with these particles and explain the differences seen in osteolysis patterns of ceramic–ceramic (CC) vs. metal–polyethylene (Mpe) articulations. In conclusion, apoptosis may be a major internal mechanism to decrease macrophage activity and may be a desired therapeutic endpoint. The identification of an apoptosis-related pathway in the macrophage response to ceramic particles provides crucial data for a rational approach in the treatment and/or prevention of periprosthetic osteolysis.