The toxic effect of fluoride on MG-63 osteoblast cells is also dependent on the production of nitric oxide

Some soda-lime-phospho-silicate glasses, such as Hench’s Bioglass® 45S5, form bone-like apatite ontheir surface when bound to living bone. To improve their osteointegration for clinical purposes, thefluoride insertion in their structure has been proposed, but we recently showed that fluoride causesoxidative damage in human MG-63 osteoblasts, via inhibition of pentose phosphate oxidative path-way (PPP) and its key enzyme glucose 6-phosphate dehydrogenase (G6PD). In the same cells we havenow investigated the role of nitric oxide (NO) in these effects. Fluoride-containing bioactive glasses andNaF caused, as expected, release of lactate dehydrogenase in the extracellular medium, accumulation ofintracellular malonyldialdehyde, inhibition of PPP and G6PD: we have now observed that these effectswere significantly reverted not only by superoxide dismutase (SOD) plus catalase (scavengers of reactiveoxygen species), but also by N-monomethyl l-arginine (l-NMMA, a NOS inhibitor) and 2-phenyl-4,4,5,5,-tetramethylimidazoline-1oxyl 3-oxide (PTIO, a NO scavenger). Moreover the two highest concentrationsof both fluoride-containing bioglasses and NaF caused increase of nitrite (a stable derivative of NO) levelsin the culture supernatant, which was inhibited by l-NMMA, erythrocytes, PTIO and SOD/catalase, andincrease of intracellular NO synthase (NOS) activity. The incubation with bioglasses or NaF increasedalso the phosphorylation of Ser1177 in the endothelial NOS isoform. Furthermore, the NO donor spermineNONOate was able to inhibit G6PD activity in vitro, and this effect was partly reverted by PTIO. Thereforeour results suggest that most cytotoxic effects of fluoride are mediated by the production of NO: reactiveoxygen species are important, causing NOS phosphorylation. We also observed, for the first time, thatTempol, but not SOD/catalase, besides inhibiting the oxidative stress induced by fluoride, also scavengesfluoride ions. For this reason it is not a selective inhibitor of the oxidative effects of fluoride.