The present studies investigated whether the effect of high glucose levels on angiotensinogen (ANG) gene expression in kidney proximal tubular cells is mediated via reactive oxygen species (ROS) generation and p38 MAPK activation. Rat immortalized renal proximal tubular cells (IRPTCs) were cultured in monolayer. Cellular ROS generation and p38 MAPK phosphorylation were assessed by lucigenin assay and Western blot analysis, respectively. The levels of immunoreactive rat ANG secreted into the media and cellular ANG mRNA were determined by a specific RIA and RT-PCR, respectively. High glucose (25 mM) evoked ROS generation and p38 MAPK phosphorylation as well as stimulated immunoreactive rat ANG secretion and ANG mRNA expression in IRPTCs. These effects of high glucose were blocked by antioxidants (taurine and tiron), inhibitors of mitochondrial electron transport chain complex I (rotenone) and II (thenoyltrifluoroacetone), an inhibitor of glycolysis-derived pyruvate transport into mitochondria (alpha-cyano-4-hydroxycinnamic acid), an uncoupler of oxidative phosphorylation (carbonyl cyanide m-chlorophenylhydrazone), a manganese superoxide dismutase mimetic, catalase, and a specific inhibitor of p38 MAPK (SB 203580), but were not affected by an inhibitor of the malate-aspartate shuttle (aminooxyacetate acid). Hydrogen peroxide (>/=10(-5) M) also stimulated p38 MAPK phosphorylation, ANG secretion, and ANG mRNA gene expression, but its stimulatory effect was blocked by catalase and SB 203580. These studies demonstrate that the stimulatory action of high glucose on ANG gene expression in IRPTCs is mediated at least in part via ROS generation and subsequent p38 MAPK activation.