Both the AT|1 and AT|2 receptor subtypes mediate angiotensin II-stimulated growth in vascular smooth muscle cells
Angiotensin II (AngII), a key growth factor for smooth muscle cells (SMCs), contributes to the pathophysiology of cardiovascular diseases. Although at least two distinct receptor subtypes have been identified, the AT1 receptor is regarded as the principal mediator of AngII's actions and the biological role of the AT2 receptor remains unclear. The hypothesis that activation of both AT1 and AT2 receptors contribute to SMC proliferation through distinct signalling pathways was investigated. A comprehensive study of the relative contribution of (i) the AT 1 and AT2 receptor subtypes to AngII-mediated SMC growth and (ii) receptor-specific intracellular signalling pathways was undertaken in primary cultures of porcine coronary artery (PCA) SMCs. Both AT1 and AT2 receptors were coupled to the mitogenic response based on the inhibition of AngII-stimulated RNA and DNA synthesis, PCNA expression and hyperplasia by the non-peptide receptor antagonists losartan (AT 1) or PD123319 (AT2). A novel role for phosphatidylinositol 3-kinase (PI3K) in AT1-mediated signalling was established with assays for P13K activity and the formation of a p85 signalling complex with IRS1, paxillin, FAK and the AT1 receptor. AngII regulated P13K through transient tyrosine phosphorylation of p85 and translocation of the p85 and p 110 subunits to different intracellular compartments. A role for P13K in AngII-stimulated growth was indicated by the inhibitor LY294002. A key signalling pathway for the AT2 receptor was identified as prostaglandin synthesis, based on the ability of indomethacin to inhibit AngII-stimulated growth and PD123319 to block AngII-stimulated arachidonic acid release. In support of these data, AngII-stimulated growth of fetal rat A10 SMCs was mediated by AT2 receptors, via a prostaglandin synthesis pathway, that involved a novel requirement for insulin pretreatment. A cellular mechanism for crosstalk between the RAS and insulin system was demonstrated by insulin regulation of AT2 receptor mRNA levels. In conclusion, these data support the hypothesis and challenge the accepted view of the AT1 receptor as the sole mediator of the mitogenic actions of AngII in SMCs.