Figure 1. LPS induced greater activation of NADPH oxidase in Nrf2^−/− macrophages by modulating PKC.

(A–B) Intracellular levels of GSH (A) and ROS (B) in Nrf2^−/− and Nrf2^+/+ macrophages treated with or without BSO (200 µM) for 16 h followed by LPS stimulation (1 h). (C) Levels of phosphorylated-serine and p47^phox in Nrf2^−/− and Nrf2^+/+ macrophages after LPS stimulation. After LPS activation, cell lysates were prepared and immunoprecipatated using anti-p47^phox antibody. The immune complex was resolved on SDS-PAGE and levels of phosphorylated-serine and p47^phox were analyzed using anti-phosphorylated serine and anti-p47^phox antibody by Western blot analysis. (D) Densitometry analysis of phosphorylated–P47^phox immunoblot normalized to total P47^phox using ImageJ software. Data are represented as mean ± SD, arbitrary units (AU) from three independent experiments. (E) PKC activity in Nrf2^−/− and Nrf2^+/+ macrophages treated with or without BSO for 16 h followed by LPS stimulation (30 min). (F) Flow cytometric analysis of ROS in Nrf2^−/− and Nrf2^+/+ macrophages treated with or without starosporine (STS) for 30 min followed by LPS stimulation (1 h). Analysis of ROS was performed within 3 h after macrophage isolation. Data are represented mean channel florescence (MCF) from two independent experiments (n=3). *P<0.05