Inverse regulation of preproendothelin-1 and endothelin-converting enzyme-1beta genes in cardiac cells by mechanical load.
Mechanical stretch and para- and/or autocrine factors, including endothelin-1, induce hypertrophy of cardiac myocytes and proliferation of fibroblasts. To investigate the effect of mechanical load on endothelin-1 production and endothelin system gene expression in neonatal rat ventricular myocytes and fibroblasts, we exposed cells to cyclic mechanical stretch in vitro (0.5 Hz, 10-25% elongation, from 1 min to 24 h). Endothelin-1 peptide levels were measured from culture media of myocytes and fibroblasts and human umbilical vein endothelial cells (positive control) by specific radioimmunoassay. Preproendothelin-1 promoter activity was measured via transfection of reporter plasmids and mRNA levels with Northern blot analysis or quantitative RT-PCR. Activity of extracellular signal-regulated kinase was quantified with specific kinase assay. We found that stretching of myocytes activated preproendothelin-1 gene expression, including promoter activation, transient mRNA level increases, and augmented endothelin-1 secretion. In contrast, preproendothelin-1 gene expression was inhibited in stretched fibroblasts. Endothelin-converting enzyme-1beta mRNA levels elevated in stretched fibroblasts but decreased in stretched myocytes. Endothelin receptor type A mRNA levels declined in stretched myocytes, whereas levels were below detection in fibroblasts. Stretch activated extracellular signal-regulated kinase in myocytes, and when the kinase activity was pharmacologically inhibited, the preproendothelin-1 induction was suppressed. Transient overexpression of mitogen-activated ERK-activating kinase-1 induced preproendothelin-1 promoter in myocytes. In summary, mechanical stretch distinctly regulates endothelin system gene expression in cardiac myocytes and fibroblasts. The inhibition of the endothelin system may affect cardiac mechanotransduction and therefore provides an approach in treatment of load-induced cardiac pathology.
Pikkarainen
Sampsa
S
Department of Pharmacology and Toxicology, Biocenter Oulu, University of Oulu, Finland.
Tokola
Heikki
H
Kerkelä
Risto
R
Ilves
Mika
M
Mäkinen
Markus
M
Orzechowski
Hans-Dieter
HD
Paul
Martin
M
Vuolteenaho
Olli
O
Ruskoaho
Heikki
H
D000818:Animals; D016282:Aspartic Acid Endopeptidases / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism; D019332:Endothelin-1 / Q000235:genetics* / Q000378:metabolism; D000072396:Endothelin-Converting Enzymes; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005419:Flavonoids / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics*; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D048369:MAP Kinase Kinase 1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D008666:Metalloendopeptidases / Q000235:genetics* / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D016760:Proto-Oncogene Proteins c-fos / Q000235:genetics; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D044022:Receptor, Endothelin A / Q000235:genetics; D013314:Stress, Mechanical; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
D019332:Endothelin-1; D005419:Flavonoids; D007093:Imidazoles; D047428:Protein Kinase Inhibitors; D016760:Proto-Oncogene Proteins c-fos; D011725:Pyridines; D044022:Receptor, Endothelin A; D048049:Extracellular Signal-Regulated MAP Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D048369:MAP Kinase Kinase 1; D016282:Aspartic Acid Endopeptidases; D008666:Metalloendopeptidases; D000072396:Endothelin-Converting Enzymes; D013755:Tetradecanoylphorbol Acetate; C093642:SB 203580; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one