Functional integrity of human neutrophils following 24 hour incubation with hydroxyapatite and fluoride

We have previously shown that hydroxyapatite (HA) priming of human neutrophils to a second stimulus of formyl-methionyl-leucyl-phenylalanine (fMPL) is influenced by a bisphosphonate and fluoride. The purpose of this study was to investigate the long-term effects of low concentrations of NaF (10(-3)-10(-11) mol/LF) on HA-mediated neutrophil chemiluminescence (CL) as a measure of oxidative function. CL assays were conducted following extended time periods of incubation (30 min, 3 h, 18 h and 24 h). Results were calculated as integrals of total energy output and expressed as the difference between the experimental (NaF/HA) and control (cells alone) assays. Transmission electron microscopy (TEM) was used to estimate cellular integrity and confirm HA phagocytosis. CL inhibition was observed at all fluoride concentrations at 30 min incubation. No significant difference compared to the control was observed in the CL output at 3 or 18 h. However, at 24 h the response showed a significant increase in activity at all NaF concentrations. The TEM results confirmed the functional integrity of the neutrophils, particularly those phagocytosing HA particles up to 24 h. Based on these results we demonstrate that human peripheral blood neutrophils can be maintained in a fully functional state with respect to the respiratory burst and morphology for at least 24 h.