CLINICAL STUDIES
Effects of angiotensin II on expression of the gap junction channel protein connexin43 in neonatal rat ventricular myocytes
Stephen M. Dodge, MD*,a,b,c,
Michael A. Beardslee, MDa,b,c,
Bruce J. Darrow, MD, PhD ,a,b,c,
Karen G. Green, BAa,b,c,
Eric C. Beyer, MD, PhD ,a,b,c and
Jeffrey E. Saffitz, MD, PhD, FACCa,b,c
a Department of Pathology, Washington University School of Medicine, Saint Louis, Missouri, USA
b Department of Medicine, Washington University School of Medicine, Saint Louis, Missouri, USA
c Department of Pediatrics, Washington University School of Medicine, Saint Louis, Missouri, USA
Manuscript received September 2, 1997;
revised manuscript received May 4, 1998,
accepted May 15, 1998.
Address for correspondence: Dr. Jeffrey E. Saffitz, Department of Pathology, Box 8118, Washington University School of Medicine, 660 S. Euclid Avenue, Saint Louis, Missouri 63110
saffitz{at}pathology.wustl.edu
Objectives. To elucidate signal transduction pathways regulating expression of myocardial gap junction channel proteins (connexins) and to determine whether mediators of cardiac hypertrophy might promote remodeling of gap junctions, we characterized the effects of angiotensin II on expression of the major cardiac gap junction protein connexin43 (Cx43) in cultured neonatal rat ventricular myocytes.
Background. Remodeling of the distribution of myocardial gap junctions appears to be an important feature of anatomic substrates of ventricular arrhythmias in patients with heart disease. Remodeling of intercellular connections may be initiated by changes in connexin expression caused by chemical mediators of the hypertrophic response.
Methods. Cultures were exposed to 0.1 µmol/liter angiotensin II for 6 or 24 h, and Cx43 expression was characterized by immunoblotting, confocal microscopy and electron microscopy.
Results. Immunoblot analysis revealed a twofold increase in Cx43 content in cells treated for 24 h with angiotensin II (n = 4, p < 0.05). This response was inhibited by the presence of 1.0 µmol/liter losartan, an AT1-receptor blocker. Confocal and electron microscopy demonstrated enhanced Cx43 immunoreactivity and increases in the number and size of gap junction profiles in cells exposed to angiotensin II for 24 h. These effects were also blocked by losartan. Immunoprecipitation of Cx43 from cells metabolically labeled with [35S]methionine demonstrated 2.4- and 2.9-fold increases in Cx43 radioactivity after 6 and 24 h exposure to angiotensin II, respectively (p < 0.03 at each time point).
Conclusions. Angiotensin II up-regulates gap junctions in cultured neonatal rat ventricular myocytes by increasing Cx43 synthesis. Signal transduction pathways activated by angiotensin II under pathophysiologic conditions could initiate remodeling of conduction pathways, leading to the development of anatomic substrates of arrhythmias.
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Abbreviations and Acronyms
| | cAMP | = cyclic adenosine 3'-5' monophosphate | | Cx43 | = connexin43 | | IgG | = immunoglobulin G | | PBS | = phosphate-buffered saline | | SDS–PAGE | = sodium dodecyl phosphate polyacrylamide gel electrophoresis | | TPBS | = phosphate-buffered saline containing 0.5% Triton X-100 |
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