Cytokine-induced nitric-oxide-dependent apoptosis in mouse osteoblastic cells: involvement of p38MAP kinase.

The apoptotic signalling induced by pro-inflammatory cytokines was examined in mouse osteoblastic MC3T3-E1 cells. Annexin-V/propidium iodine double-staining analysis demonstrated that the combination of tumour necrosis factor-alpha, interleukin-1beta and interferon-gamma caused cell death in osteoblastic cells mediated by apoptosis, not necrosis. Treatment with these cytokines resulted in potent enhancement of inducible nitric-oxide synthase (iNOS) mRNA and nitric-oxide (NO) in the cells. A specific inhibitor of p38 mitogen-activated protein (MAP) kinase, i.e. SB203580, dose dependently inhibited the induction of iNOS mRNA, its enzyme product, NO and DNA fragmentation (as an apoptosis index) in the cytokine-treated cells (P<0.05). In contrast, PD98059, a specific inhibitor of MEK that acts immediately upstream of classic MAP kinase, had no effect on the induction of iNOS, NO or DNA fragmentation in the cells. These results demonstrate that this cytokine-induced apoptosis in mouse osteoblastic cells was mediated by a p38MAP-kinase-dependent iNOS system.