The role of AT1 receptor, Ras and NAD(P)H oxidase on p38 MAPK phosphorylation by angiotensin II stimulation in primary cultured vascular smooth muscle cells

Abstract

Aim: Ang II stimulates vascular smooth muscle cells (VSMCs) and activates mitogen protein kinases (MAPKs) in culture media. p38 MAPK, a member of MAPK family, is stimulated by both Ang II and reactive oxygen species. This study was carried out to show Ang II stimulated p38 MAPK activation through Ras and subsequent NADPH oxidase activation in VSMCs. Material and Method: In our study, primary VSMCs isolated from rat aorta were stimulated by Ang II and with different inhibitors, and western blot was used to measure p38 MAPK phosphorylation. Results: The optimum p38 MAPK phosphorylation was observed at 100 nM Ang II concentration for 5 minutes and the next experiments were carried out under this conditions. To analyze the effect of Ras, which is an upstream mediator of p38 MAPK phosphorylation, VSMCs were incubated with a Ras spesific inhibitor FTS (Farnesyl thiosalicylic acid). After the stimulation of VSMCs by Ang II, p38 MAPK phosphorylation was inhibited. p38 MAPK phosphorylation was also completely inhibited in the presence of DPI (Diphenyl Iodinoum), a spesific NAD(P)H oxidase inhibitor, after Ang II stimulation. Conclusion: These findings demonstrate that Ang II stimulated p38 MAPK phosphorylation is through AT1 receptor and Ras-NAD(P)H oxidase dependent pathway in VSMCs. Understanding this pathway may contribute to the cellular mechanisms underlying in cardiovascular diseases.