IL-12- and IL-18-Mediated, Nitric Oxide-Induced Apoptosis in TNF-α-Mediated Osteoclastogenesis of Bone Marrow Cells

TNF-α has been recognized as an important factor for osteoclastogenesis and plays an important role in bone resorption under pathological conditions. IL-12 and IL-18, which are T-cell mediators, are also important inflammatory cytokines. We have reported that IL-12 and IL-18 induce apoptosis in bone marrow cells treated with TNF-α in vitro and that osteoclastogenesis is inhibited by the interaction of TNF-α-induced Fas and the IL-12-induced Fas ligand (FasL). However, the anti-FasL antibody could not completely inhibit apoptosis. Therefore, it is possible that IL-12 and IL-18 may also trigger some other apoptotic mechanisms. Nitric oxide (NO) may act as a mediator of the apoptotic effect. In this study, we examined whether NO causes the IL-12- and IL-18-induced apoptosis of bone marrow cells in TNF-α-mediated osteoclast formation. We found that NO production was induced in bone marrow cells cultured with IL-12 and IL-18 in the presence of TNF-α. When bone marrow cells were cultured with TNF-α, osteoclasts were formed. In contrast, when bone marrow cells were cultured with both TNF-α and IL-12 or IL-18, the adherent cells were induced to undergo apoptosis. Apoptosis was partially inhibited when bone marrow cells were treated with NO synthase inhibitors. Furthermore, IL-12 and IL-18 synergistically induced cell death and upregulated NO production in the presence of TNF-α. These results indicate that the simultaneous effects of TNF-α and IL-12 or IL-18 on bone marrow cells induce apoptosis and that apoptosis is induced by the production of NO.