Role of ERK-MAPK in endothelin receptor signaling in human aortic smooth muscle cellsReportar como inadecuado

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BMC Cell Biology

, 10:52

First Online: 03 July 2009Received: 27 February 2009Accepted: 03 July 2009


BackgroundEndothelin-1 ET-1 is a potent vasoactive peptide, which induces vasoconstriction and proliferation in vascular smooth muscle cells VSMCs through activation of endothelin type A ETA and type B ETB receptors. The extracellular signal-regulated kinase 1 and 2 ERK1-2 mitogen-activated protein kinases MAPK are involved in ET-1-induced VSMC contraction and proliferation. This study was designed to investigate the ETA and ETB receptor intracellular signaling in human VSMCs and used phosphorylation activation of ERK1-2 as a functional signal molecule for endothelin receptor activity.

ResultsSubconfluent human VSMCs were stimulated by ET-1 at different concentrations 1 nM-1 μM. The activation of ERK1-2 was examined by immunofluorescence, Western blot and phosphoELISA using specific antibody against phosphorylated ERK1-2 protein. ET-1 induced a concentration- and time- dependent activation of ERK1-2 with a maximal effect at 10 min. It declined to baseline level at 30 min. The ET-1-induced activation of ERK1-2 was completely abolished by MEK1-2 inhibitors U0126 and SL327, and partially inhibited by the MEK1 inhibitor PD98059. A dual endothelin receptor antagonist bosentan or the ETA antagonist BQ123 blocked the ET-1 effect, while the ETB antagonist BQ788 had no significant effect. However, a selective ETB receptor agonist, Sarafotoxin 6c S6c caused a time-dependent ERK1-2 activation with a maximal effect by less than 20% of the ET-1-induced activation of ERK1-2. Increase in bosentan concentration up to 10 μM further inhibited ET-1-induced activation of ERK1-2 and had a stronger inhibitory effect than BQ123 or the combined use of BQ123 and BQ788. To further explore ET-1 intracellular signaling, PKC inhibitors staurosporin and GF109203X, PKC-delta inhibitor rottlerin, PKA inhibitor H-89, and phosphatidylinositol 3-kinase PI3K inhibitor wortmannin were applied. The inhibitors showed significant inhibitory effects on ET-1-induced activation of ERK1-2. However, blockage of L-type Ca channels or calcium-calmodulin-dependent protein kinase II, chelating extracellular Ca or emptying internal Ca stores, did not affect ET-1-induced activation of ERK1-2.

ConclusionThe ETA receptors predominate in the ET-1-induced activation of ERK1-2 in human VSMCs, which associates with increments in intracellular PKC, PKA and PI3K activities, but not Ca signalling.

Electronic supplementary materialThe online version of this article doi:10.1186-1471-2121-10-52 contains supplementary material, which is available to authorized users.

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Autor: Qing-wen Chen - Lars Edvinsson - Cang-Bao Xu


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