Optimization of the Timing of Induction for the Assessment of Nitric Oxide Production in Leishmania major Infected Macrophage Cells

BACKGROUND: The present study was conducted to investigate the optimized timing for macrophages induction and nitric oxide (NO) production against invading Leishmania parasite. METHODS: The present study examined the murine macrophage cell line, B10R, in three different states. In the first state, the cells were first infected with L. major and then treated with IFN-γ and LPS as stimulants. In the second state, the cells were infected after stimulation with IFN-γ and LPS. In the third state, the cells were only exposed to stimulants as controls. In all the three states, cell culture supernatants were collected at three points in time (6, 24 and 48 h) and the amount of NO production was measured using Griess assay. RESULTS: The treatment of macrophages with inducers prior to infection with stationary phase parasite led to the secretion of significant amounts of NO, particularly at early time points quit contrary to the cells infected with parasites prior to induction. The amount of NO produced by cells induced after infection was detected significantly lower. CONCLUSION: The induction of macrophages prior to infection with parasites leads to the production and secretion of greater amounts of NO, resulting in an increased ability to suppress and inhibit parasite proliferation even in the early stages of infection.