Fig. 2

Intracellular formation of ROS. Fluorescence distribution of DCFH-DA oxidation or DHR oxidation was estimated by flow cytometry in NB9 exposed to 2 mM L-Arg. Lane 1, control (50 µM L-Arg); lane 2, 2 mM L-Arg; lane 3, 2 mM L-Arg and 5 mM L-NAME; lane 4, 50 µM L-Arg and 250 µM L-Cit. (A) NB9 was incubated for 48 hr as described above, then with 5 µM DCFH-DA for 30 min at 37°C, and the formation of DCFH-DA oxidation was estimated as fluorescence intensity. (B) NB9 was incubated for 48 hr as described above, then with 2 µM DHR for 30 min at 37°C and the formation of rhodamine 123 was estimated flow cytometrically as for DCFH-DA. The histograms represent the number of cells as a function of relative fluorescence intensity. Data are representative of three experiments with similar results.