Hypoxic induction of receptor activity-modifying protein 2 alters regulation of pulmonary endothelin-1 by adrenomedullin: induction under normoxia versus inhibition under hypoxia.
The vasodilator adrenomedullin (AM) is up-regulated in pulmonary hypertension, and inhaled AM is beneficial in patients. Therefore, we investigated the effects of AM on pulmonary endothelin-1 (ET-1). In normoxic isolated rat lungs (IRL) and rat pulmonary artery endothelial cells (RPAEC), the calcitonin gene-related peptide type-1 receptor (CGRP1R) antagonist human (h)CGRP(8-37) decreased ET-1 secretion, and the AM receptor antagonist hAM(22-52) had no effect. Exogenous AM (1 and 10 pM) increased ET-1 levels, which was abolished by hCGRP(8-37) and protein kinase A (PKA) inhibition. At 50 and 100 pM, AM decreased ET-1, an effect sensitive to hAM(22-52), NO inhibition, and protein kinase G (PKG) inhibition. In RPAEC, these results were attributed to altered ET-1 gene expression; low exogenous AM also promoted activity of endothelin-converting enzyme, and high AM increased the number of endothelin type-B (ETB) receptor sites. Hypoxia significantly elevated AM and ET-1 levels in IRL and RPAEC, and hAM(22-52), NO inhibition, or PKG inhibitors caused a further ET-1 rise. These interventions also prevented the hypoxia-related increase in ETB sites in RPAEC. In RPAEC, both high AM and hypoxia down-regulated receptor activity-modifying protein (RAMP)1, but they up-regulated RAMP2 protein and AM receptor sites, and RAMP2 silencing by small interference RNA proved its pivotal role for signal switching. In conclusion, endogenous pulmonary AM up-regulates ET-1 and endothelin-converting enzyme activity under physiological conditions, via CGRP1R and PKA. In contrast, hypoxia-induced high AM levels, via AM1 receptor and NO/PKG, down-regulate ET-1 gene expression and promote expression of ETB receptors. This hypoxia-related switch of AM signaling can be attributed to up-regulation of the RAMP2/AM1 receptor system.
Dschietzig
Thomas
T
Department of Cardiology and Angiology, University Medical Centre Charité, Campus Mitte, Schumannstr. 20/21, 10117 Berlin, Germany. thomas.dschietzig@charite.de
Richter
Christoph
C
Asswad
Louay
L
Baumann
Gert
G
Stangl
Karl
K
D053607:Adrenomedullin / Q000494:pharmacology*; D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D015153:Blotting, Western; D015740:Calcitonin Gene-Related Peptide / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism; D019332:Endothelin-1 / Q000378:metabolism*; D000860:Hypoxia / Q000378:metabolism*; D066298:In Vitro Techniques; D047908:Intracellular Signaling Peptides and Proteins; D008168:Lung / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D008565:Membrane Proteins / Q000096:biosynthesis*; D009569:Nitric Oxide / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D034741:RNA, Small Interfering / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D058263:Receptor Activity-Modifying Protein 2; D058261:Receptor Activity-Modifying Proteins; D044023:Receptor, Endothelin B / Q000187:drug effects / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D012997:Solvents
D000894:Anti-Inflammatory Agents, Non-Steroidal; D019332:Endothelin-1; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; C545503:RAMP2 protein, human; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C545509:Ramp2 protein, rat; D058263:Receptor Activity-Modifying Protein 2; D058261:Receptor Activity-Modifying Proteins; D044023:Receptor, Endothelin B; D012997:Solvents; D053607:Adrenomedullin; D009569:Nitric Oxide; D000242:Cyclic AMP; D006152:Cyclic GMP; D015740:Calcitonin Gene-Related Peptide