Antioxidant actions contribute to the antihypertrophic effects of atrial natriuretic peptide in neonatal rat cardiomyocytes.
OBJECTIVE
Reactive oxygen species (ROS) such as superoxide have been linked to the hypertrophic response of the heart to stimuli including angiotensin II (AngII), mechanical stretch, and pressure overload. We have previously demonstrated that cGMP and protein kinase G mediate the antihypertrophic actions of the natriuretic peptides in rat cardiomyocytes and isolated whole hearts. The impact of natriuretic peptides on cardiac ROS generation, however, has not been investigated. We tested the hypothesis that reduced superoxide accumulation contributes to the antihypertrophic action of atrial natriuretic peptide (ANP).


METHODS
Neonatal rat cardiomyocytes were cultured in serum-free medium with and without AngII (1 micromol/L) or endothelin-1 (ET(1), 60 nmol/L) in the presence and absence of ANP (1 micromol/L) or tempol (100 micromol/L). Hypertrophic responses, cardiomyocyte superoxide generation, and cardiomyocyte expression of NADPH oxidase were determined.


RESULTS
AngII induced increases in cardiomyocyte size (to 176 +/- 9% n = 8 p < 0.001, at 48 h), beta-myosin heavy chain expression (to 4.0 +/- 1.6-fold n = 6 p < 0.05, at 48 h), c-fos expression (to 1.9 +/- 0.5-fold n = 7 p < 0.01, at 6 h), superoxide generation (to 181+/-21% n = 8 p < 0.005, at 24 h), and expression of the gp91phox subunit of NADPH oxidase (to 2.4 +/- 0.5-fold n = 7 p < 0.05, at 48 h). These effects were all significantly inhibited by ANP: cardiomyocyte size, beta-myosin heavy chain expression, c-fos expression, superoxide generation and gp91phox expression were reduced to 107 +/- 5% (n = 5 p < 0.05), 1.2 +/- 0.2-fold (n = 6 p < 0.05), 0.9 +/- 0.2-fold (n = 7 p < 0.05), 141 +/- 21% (n = 8 p < 0.05), and to 1.0 +/- 0.5-fold (n = 7 p < 0.05), respectively. These effects were mimicked by tempol. ANP and tempol also significantly inhibited ET1-induced increases in cardiomyocyte size and superoxide generation, but had no effect on markers of hypertrophy when studied alone.


CONCLUSIONS
This data indicates that the antihypertrophic actions of ANP are accompanied by reduced levels of superoxide, suggesting an antioxidant action contributes to the antihypertrophic actions of ANP.
Laskowski
Adrienne
A
Wynn Department of Metabolic Cardiology, Baker Heart Research Institute, Melbourne 8008, Australia.
Woodman
Owen L
OL
Cao
Anh H
AH
Drummond
Grant R
GR
Marshall
Tanneale
T
Kaye
David M
DM
Ritchie
Rebecca H
RH
D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D000975:Antioxidants / Q000494:pharmacology*; D009320:Atrial Natriuretic Factor / Q000494:pharmacology*; D049108:Cell Enlargement / Q000187:drug effects; D002478:Cells, Cultured; D003497:Cyclic N-Oxides / Q000494:pharmacology; D019332:Endothelin-1 / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D016762:Genes, fos; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism*; D018995:Myosin Heavy Chains / Q000235:genetics; D019255:NADPH Oxidases / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D013113:Spin Labels; D013481:Superoxides / Q000032:analysis
D000975:Antioxidants; D003497:Cyclic N-Oxides; D019332:Endothelin-1; C000594638:MYH7 protein, rat; D013113:Spin Labels; D013481:Superoxides; D000804:Angiotensin II; D009320:Atrial Natriuretic Factor; D019255:NADPH Oxidases; D018995:Myosin Heavy Chains; C001803:tempol