ROS generated by pollen NADPH oxidase provide a signal that augments antigen-induced allergic airway inflammation
J. Clin. Invest. Istvan Boldogh, et al. 115:2169 doi:10.1172/JCI24422 [Go to this article.]

Figure 1
Pollen grain extracts show NADPH oxidase activity. (A) Reduction of NBT to formazan by allergenic extracts using NBT assay in the presence (+) or absence (–) of NADPH. RWE, RWE^H, and Amb a 1 were tested. X+XO were used as a positive control. ^#P < 0.001; ^##P < 0.0001. (B) pRWE^OX+-induced NBT reduction is inhibited by NADPH oxidase inhibitors DPI, QA, and SOD. The presence or absence of NADPH in the reaction mixture is indicated. (C) NBT reduction by allergenic extracts in situ after nondenaturing PAGE. RWE^H, heat-inactivated oak extract (Oak^H), and heat-inactivated timothy grass extract (Timothy^H) are shown. (D) Detection of p67^phox by Western blot analysis using a rabbit anti-human p67^phox antibody. (E and F) Immunolocalization of p67^phox in ragweed pollens detected by fluorescence microscopy using anti-p67^phox antibody (E) or normal rabbit IgG control (F). Right panels show differential interference contrast images of the same pollens. Magnification, ×600. (G) Kinetics of O₂^•– generation determined by cytochrome c assay. Shown are cytochrome c (filled diamonds); cytochrome c plus pRWE^OX+ (filled squares); cytochrome c plus NADPH (open triangles); cytochrome c plus pRWE^OX+ and NADPH (open squares); and cytochrome c plus pRWE^OX+, NADPH, and SOD (filled triangles).