Figure 3

Effect of H ₂ O ₂ on the R848-induced superoxide generation, elastase release and chemotaxis in human PMNs. (A) PMNs were preincubated for 30 min with or without 50 μM H₂O₂, and treated with vehicle or R848. Cells were then harvested and incubated with dihydro-rhodamine-123 (DHR-123) for 5 min. The amount of superoxide generation was indicated as the relative fluorescence intensity of DHR-123. (B) After incubation with or without 50 μM H₂O₂, PMNs were stimulated with various concentrations of R848 for 24 hrs. The media were assayed for elastase release by ELISA. (C) After one hour treatment with various concentrations of R848 with or without 50 μM H₂O₂, chemotactic capacity toward IL-8 was assessed by a modified boyden chamber method. Vertical axis: Relative ratio of the PMN counts (-fold increase). Relative ratio of the PMN counts was calculated as the ratio of the migrated cell count of each group to that of the control group. All values are mean values ± SEM of three to four separate experiments. *p < 0.05, compared with the values of vehicle-treated group; +p < 0.05, compared with the values of 50 μM H₂O₂-pretreated and vehicle-treated group; MFI = mean fluorescence intensity.