ERK and p38 MAPK, but not NF-kappa B, are critically involved in reactive oxygen species-mediated induction of IL-6 by angiotensin II in cardiac fibroblasts

We recently reported that angiotensin II (Ang II) induced IL-6 mRNA express ion in cardiac fibroblasts, which played an important role in Ang II-induce d cardiac hypertrophy in paracrine fashion. The present study investigated the regulatory mechanism of Ang II-induced IL-6 gene expression, focusing e specially on reactive oxygen species (ROS)-mediated signaling in cardiac fi broblasts. Ang II increased intracellular ROS in cardiac fibroblasts, and t he increase was completely inhibited by the AT-I blocker candesartan and th e NADH/NADPH oxidase inhibitor diphenyleneiodonium (DPI). We first confirme d that antioxidant N-acetylcysteine, superoxide scavenger Tiron, and DPI su ppressed Ang II-induced IL-6 expression. Because we observed that exogenous H2O2 also increased IL-6 mRNA, the signaling pathways downstream of Ang II and exogenous H2O2 were compared. Ang II, as well as exogenous H2O2, activ ated ERK, p38 MAPK, and JNK, which were significantly inhibited by N-acetyl cysteine and DPI. In contrast with exogenous H2O2, however, Ang II did not influence phosphorylation and degradation of I kappaB-alpha/beta or nuclear translocation of p65, nor did it increase NF-KB promoter activity. PD98059 and SB203580 inhibited Ang II-induced IL-6 expression. Truncation and muta tional analysis of the IL-6 gene promoter showed that CRE was an important cis-element in Ang II-induced IL-6 gene expression. NF-KB-binding site was important for the basal expression of IL-6, but was not activated by Ang II . Ang II phosphorylated CREB through the ERK and p38 MAPK pathway in a ROS- sensitive manner. Collectively, these data indicated that Ang II stimulated ROS production via the ATI receptor and NADH/NADPH oxidase, and that these ROS mediated activation of MAPKs, which culminated in IL-6 gene expression through a CRE-dependent, but not NF-kappaB-dependent, pathway in cardiac f ibroblasts.