(Verbatim from the application): The long-term goals are to identify at the biochemical and cellular levels the signal transduction pathway(s) which link Angiotensin II (Ang II) to the production of nitric oxide (NO), a potent vasodilator, in endothelial cells. This effect of Ang II is in sharp contrast to its well-known role in the cardiovascular system as a potent vasoconstrictor. The underlying hypothesis is that these apparently contradictory effects of Ang II reflect signaling events coupled to the AT1 and AT2 receptors, respectively, and that the vasodilatory effects of Ang II are mediated by the AT2 receptors via currently unidentified signaling pathways. This emerges from the novel findings that Ang II stimulates NO production in pulmonary endothelial cells, via increased expression of endothelial nitric oxide synthase (eNOS). Preliminary data indicate that this is mediated via the AT2 receptor, and support a mechanism by which Ang II binding to a G protein-coupled AT2 receptor leads to an increase in tyrosine phosphorylation, which ultimately results in increased eNOS protein expression. Furthermore, signaling via the AT1 receptor appears to negatively regulate eNOS protein expression.
Aim I is directed at the characterization of AT1/AT2 receptor subtype distribution on endothelial cells, and further investigation of the mechanism of Ang II-stimulated eNOS mRNA and protein expression. The role of the Ang II receptor subtypes in mediating Ang II dependent enhancement of hypoxic vasoconstriction in pulmonary endothelium will be determined. The goal of the second aim is to identify the pertussis toxin-sensitive G protein that is linked to AT2 receptor.
In Aim 3 the tyrosine kinase(s) that are activated, or protein tyrosine phosphatase(s) that are inhibited, and that provide the signaling linkage between Ang II and NO production, will be identified. Receptor antagonists, binding studies, and analysis of mRNA and protein levels will be used to identify the receptor subtypes. Pharmacological inhibitors, antisense oligonucleotides and antibody immunoneutralization experiments will be used to investigate the role of various signaling components (Mg II receptors, GTP binding proteins and protein kinases). These studies could provide a molecular basis for a novel hypothesis that would rationalize the physiologically important but opposing effects of Ang II on smooth muscle cells versus endothelial cells. These studies are of significant medical importance to heart and pulmonary disease, which involve pathological changes in blood pressure.
