The alpha(1D)-adrenergic receptor induces vascular smooth muscle apoptosis via a p53-dependent mechanism

Activation of the endogenous alpha(1)-adrenergic receptor (AR) associated with human aortic smooth muscle cells resulted in a dose- and time-dependent increase in the levels of mitochondrial reactive oxygen species (ROS). ROS increases were apparent within 10 min and maximal after 45 min. Prolonged activation (> 4 h) of the alpha(1)-AR resulted in smooth muscle cell apoptosis. Both the increase in ROS and apoptotic cell death were blocked by the nonselective alpha(1)-AR antagonist prazosin as well as the selective alpha(1D)-AR antagonist 8-[2-[4-(2-methoxyphenyl)-1- piperazinyl] ethyl]-8-azaspiro[4.5]decane-7,9-dione( BMY 7378). Increases in ROS and apoptosis produced by alpha(1)-AR activation were also blocked by the p38 mitogen-activated protein kinase inhibitor 4-(4-fluorophenyl)-2-(4-hydroxyphenyl)5-(4-pyridyl)-1H-imidazole (SB 202190) and the NAPDH oxidase inhibitor apocynin. The extracellular signal-regulated kinase 1/2 inhibitor 2 '-amino-3 '-methoxyflavone (PD 98059) or the c-Jun NH2-terminal kinase inhibitor 1,9-pyrazoloanthrone anthra(1,9-cd)pyrazol-6(2H)-one (SP 600125) was without effect on increases in ROS levels or apoptosis. Pifithrin-alpha, an inhibitor of the tumor suppressor protein p53, had no effect on ROS generation but did block alpha(1D)-AR-induced apoptosis. Activation of the alpha(1D)-AR resulted in translocation of p53 to the mitochondria. The mitochondrial translocation of p53 was blocked by prazosin, BMY 7378, apocynin, SB 202190, and pifithrin-alpha. Apoptosis was also blocked by small interfering RNA directed against p53. These data show that the alpha(1D)-AR is coupled to the generation of mitochondrial ROS by a pathway involving p38 and NADPH oxidase. Sustained activation of the alpha(1D)-AR results in smooth muscle cell apoptosis in a pathway that involves the tumor suppressor protein p53 and the mitochondrial translocation of p53. The data also provide evidence of a linkage between the alpha(1D)-AR and p53.