Fig. 5.

Effect of ETC inhibitors and selective antioxidants on intracellular peroxide levels. ECs were preincubated with DCFH-DA (10 μM) and DAPI (1 μM) and with ebselen (5 μM), rotenone (2 μM), l-NAME (1 mM), or SS-31 or SS-20 (1, 10, or 100 nM) and sheared (30 min) in the presence of the respective drug. Digital images (62×) of DCF and DAPI fluorescence were obtained by confocal microscopy and overlaid. A, representative digital images from ECs exposed to different treatments. From all drugs tested on static ECs, only rotenone changed the DCF fluorescence signal (only rotenone is included). Because SS-20 had no effect on the fluorescence in sheared ECs at either concentration tested, only 10 nM is shown. B, using digital image processing, the mean DCF fluorescence intensity per image was calculated, averaged over three fields of view per experiment, and then averaged over three independent experiments. Normalized (to static controls) DCF fluorescence values are mean ± S.E.M. ^*, significantly different from static control group; P ≤ 0.05; n = 3. †, significantly different from shear group; P ≤ 0.05; n = 3.