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. Author manuscript; available in PMC: 2009 Nov 1.
Published in final edited form as: Free Radic Biol Med. 2008 Aug 16;45(9):1340–1351. doi: 10.1016/j.freeradbiomed.2008.08.013

Figure 6.

Figure 6

Analysis of O2∸ and H2O2 in Nox4-depleted RASMCs. (A) Production of intracellular O2∸ was measured by DHE/HPLC following accumulation of 2-hydroxyethidium using HPLC as described in Materials and Methods. RASMCs treated with scrambled (Scr) or Nox4 siRNA (sinox4) were stimulated with 100 nM AngII for 4-hours; (B) Cellular H2O2 was measured using the fluorescent probe Amplex Red and normalized by cellular protein. RASMCs were stimulated with AngII and incubated with Amplex Red for 2-hours. Accumulation of the fluorescent signal was blocked by supplementation with catalase (20 µg/ml) (not shown). (C) ESR measurements of NADPH oxidase activity in membrane preparations of siRNA-treated RASMC by analysis of NADPH-dependent O2∸ production; (D) Measurements of NADPH oxidase activity in membrane preparations of siRNA-treated RASMC by analysis of NADPH-dependent H2O2 production using ESR spectroscopy as described in Methods. Data are from 4 to 6 separate experiments (*P<0.05 AngII vs non-stimulated).