NITRIC-OXIDE ACTS IN CONJUNCTION WITH PROINFLAMMATORY CYTOKINES TO PROMOTE CELL-DEATH IN OSTEOBLASTS

Proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alp ha), interferon-gamma (IFN-gamma), and interleukin-1 beta are known mo dulators of bone remodeling in vitro and in vivo, The same cytokines i nduce the production of nitric oxide (NO) in various cell types, inclu ding osteoblasts and osteoclasts, and NO has recently been implicated in the regulation of bone resorption, We investigated the relationship between NO levels and, cell viability in MC3T3-E1, a well-characteriz ed osteoblastic cell line. NO donors at high concentrations (greater t han or equal to 0.5 mM) produce a significant cytotoxic effect over a 48 h period, Various combinations of the three cytokines strongly prom ote endogenous NO production, and high NO levels are correlated with t he loss of cell viability. Although TNF-alpha produces NO-independent cytotoxicity, NO greatly enhances this cytotoxic effect, Human and mou se TNF-alpha differ in their cytotoxic effects, and human TNF-alpha in duces lower levels of NO production, In cocultures of RAW 264.7 mouse macrophages stimulated with lipopolysaccharide and IFN-gamma, and untr eated MC3T3-E1 osteoblasts, addition of anti-TNF-alpha antibody and in hibition of NO synthesis have additive, protective effects on osteobla st viability, NO cytotoxicity involves an apoptotic mechanism, Our res ults underline the importance of NO and TNF-alpha as cytotoxic mediato rs in the osseous microenvironment and might explain the observed defi ciency of bone formation in inflammatory sites.