Activation of p38 and ERK1/2 MAPK by superoxide anion participates in Angiotensin II-induced COX-2 expression in smooth muscle cells from resistance arteries

Abstract

Introduction: Angiotensin II (Ang II) regulates vascular smooth muscle cell (VSMC) function by activating signalling cascades that promote vasoconstriction, growth and inflammation. The mechanisms implicated in Ang II-induced pro-inflammatory actions include activation of several mitogen-activated protein kinases (MAPKs), reactive oxygen species generation and the modulation of prostaglandins production by regulating cyclooxygenase-2 (COX-2) expression. Aim: To investigate the effect of Ang II on COX-2 expression in VSMC derived from small resistance arteries and the mechanisms involved. Methods: VSMC derived from rat mesenteric resistance arteries were used. Protein expression was determined by Western Blot, mRNA levels by Q-RT-PCR and superoxide anion (O2-) production by dihydroethidine fluorescence. Results: Ang II (0.1 ¿M) time-dependently increased COX-2 protein expression (30 min - 8 h) and mRNA levels (15 min - 4 h), while COX- 1 expression remained unmodified. In addition, Ang II did not modify AT1 receptor expression. The AT1 antagonist losartan (10 ¿M), but not the AT2 antagonist PD 122319 (10 ¿M), abolished the increase in COX-2 expression induced by Ang II (2 h). COX-2 expression was also reduced by the respective NADPHox and xanthine oxidase inhibitors, apocynin (30 mM) and allopurinol (10 mM). Furthermore, Ang II (3- 30 min) increased O2- production; this effect was reduced by losartan, allopurinol and apocynin but not by PD 123319. Ang II (2-30 min) induced the phosphorylation of p38 and ERK1/2 MAPK; this effect was reduced by losartan, allopurinol and apocynin. In addition, the respective inhibitors of p38 and ERK1/2, SB 203580 (10 mM) and PD 98059 (25 mM), reduced the Ang II-induced COX-2 expression. Conclusions: The present results provide evidences that angiotensin II increases COX-2 expression in VSMC from resistance arteries, at least in part, through mechanisms that include O2- production and the subsequent activation of p38 and ERK1/2 MAPK