Figure 1

Increased ROS generation during reperfusion of OGD subjected cultures of retinoic acid differentiated SH-SY5Y involves NADPH oxidase activity. Representative fluorescent images from three independent experiments of reactive oxygen species production following OGD (A) with vehicle (1:1000 DMSO) or DPI (100 nM) detected using dihydroethidium (DHE). Spectrophotometric quantification of reactive oxygen species following OGD (B) with vehicle (1:1000 DMSO) or DPI (100 nM) utilizing nitrobluetetrazolium chloride (NBT). (C) Western blot shown is representative of three independent experiments comparing p67^phox, NR2A subunit protein expression in non- and differentiated SH-SY5Y cells. (D) Quantification of the relative densitometry of p67^phox immunoreactive band in non- and differentiated SH-SY5Y cells. Data represent fold change of OGD/R groups versus normoxic control group (arbitrary units) ± S.E.M from three separate experiments that consisted of at least 6 determinents (askteriks * indicates a p <0.05 from vehicle treated normoxic control; ANOVA with post hoc Bonferroni test). Phorbol 12-myristate 13-acetate (PMA; 1 μM for 15 minutes) was used as a positive control for NADPH oxidase activity in both the DHE and NBT assays. Quantitative data from p67^phox protein expression represent fold change of differentiated SH-SY5Y compared to non-differentiated SH-SY5Y cells (arbitrary units) ± S.E.M from three separate experiments (askteriks * indicates a p <0.01 from non-differentiated control; student t-Test).