Cell-Signaling Evidence for Adenosine Stimulation of Coronary Smooth Muscle Proliferation via the A 1 Adenosine Receptor

Abstract

For decades, it has been thought that adenosine is exclusively antimitogenic on vascular smooth muscles via the A 2 -type adenosine receptor. Recently, we have demonstrated that adenosine stimulates proliferation of porcine coronary artery smooth muscle cells (CASMC) through the A 1 adenosine receptor. However, the cell-signaling mechanisms underlying A 1 receptor–mediated CASMC proliferation in response to adenosine have not been defined. Here, we show that in cultured CASMC, adenosine stimulates phosphorylation of extracellular signal–regulated kinase (ERK), Jun N-terminal kinase (JNK), and AKT in a concentration- and time-dependent manner. This effect is fully mimicked by NECA (nonselective agonist), largely mimicked by CCPA (A 1 -selective agonist), weakly mimicked by 2-Cl-IB-MECA (A 3 -selective agonist), but not by CGS21680 (A 2A -selective agonist), indicating that adenosine signals strongly via the A 1 receptor to these mitogenic signaling pathways. This interpretation is supported by the finding that adenosine- and CCPA-induced phosphorylation of ERK, JNK, and AKT are inhibited by pertussis toxin (inactivator of Gi proteins) and by DPCPX (A 1 -selective antagonist), but not by SCH58261, MRS1706, and VUF5574 (A 2A -, A 2B -, and A 3 -selective antagonists, respectively). In addition, adenosine- and CCPA-induced phosphorylation of ERK, JNK, and AKT is inhibited, respectively, by U0126, PD98059 (mitogen-activated protein kinase kinase inhibitors), SP600125 (JNK kinase inhibitor), and wortmannin (phosphatidylinositol 3-kinase inhibitor). Furthermore, these kinase inhibitors abolish or diminish adenosine- and CCPA-induced increases in the rate of cellular DNA synthesis, bromodeoxyuridine incorporation, protein synthesis, and cell number. We conclude that adenosine activates the ERK, JNK, and phosphatidylinositol 3-kinase/AKT pathways primarily through the A 1 receptor, leading to CASMC mitogenesis.