Endothelin-1 (ET-1) Increases the Expression of Remodeling Genes in Vascular Smooth Muscle through Linked Calcium and cAMP Pathways: ROLE OF A PHOSPHOLIPASE A(2)(cPLA(2))/CYCLOOXYGENASE-2 (COX-2)/PROSTACYCLIN RECEPTOR-DEPENDENT AUTOCRINE LOOP

Several important genes that are involved in inflammation and tissue remodeling are switched on by virtue of CRE response elements in their promoters. The upstream signaling mechanisms that inflammatory mediators use to activate cAMP response elements (CREs) are poorly understood. Endothelin (ET) is an important vasoactive mediator that plays roles in inflammation, vascular remodeling, angiogenesis, and carcinogenesis by activating 7 transmembrane G protein-coupled receptors (GPCR). Here we characterized the mechanisms ET-1 uses to regulate CRE-dependent remodeling genes in pulmonary vascular smooth muscle cells. These studies revealed activation pathways involving a cyclooxygenase-2 (COX-2)/prostacyclin receptor (IP receptor) autocrine loop and an interlinked calcium-dependent pathway. We found that ET-1 activated several CRE response genes in vascular smooth muscle cells, particularly COX-2, amphiregulin, follistatin, inhibin-β-A, and CYR61. ET-1 also activated two other genes epiregulin and HB-EGF. Amphiregulin, follistatin, and inhibin-β-A and epiregulin were activated by an autocrine loop involving cPLA2, arachidonic acid release, COX-2-dependent PGI(2) synthesis, and IP receptor-linked elevation of cAMP leading to CRE transcription activation. In contrast COX-2, CYR61, and HB-EGF transcription were regulated in a calcium-dependent, COX-2 independent, manner. Observations with IP receptor antagonists and COX-2 inhibitors were confirmed with IP receptor or COX-2-specific small interfering RNAs. ET-1 increases in intracellular calcium and gene transcription were dependent upon ET(a) activation and calcium influx through T type voltage-dependent calcium channels. These studies give important insights into the upstream signaling mechanisms used by G protein-coupled receptor-linked mediators such as ET-1, to activate CRE response genes involved in angiogenesis, vascular remodeling, inflammation, and carcinogenesis.