Hyperoxia and transforming growth factor β1 signaling in the post-ischemic mouse heart.
OBJECTIVE
Following ischemic injury, myocardial healing and remodeling occur with characteristic myofibroblast trans-differentiation and scar formation. The current study tests the hypothesis that hyperoxia and nitric oxide (NO) regulate TGF-β1 signaling in the post-ischemic myocardium.


METHODS
C57BL/6 wild-type (WT), endothelial and inducible nitric oxide synthase knockout (eNOS(-/-) and iNOS(-/-)) mice were subjected to 30-min left anterior descending coronary artery occlusion followed by reperfusion. Myocardial tissue oxygenation was monitored with electron paramagnetic resonance oximetry. Protein expressions of TGF-β1, receptor-activated small mothers against decapentaplegic homolog (Smad), p21 and α-smooth muscle actin (α-SMA) were measured with enzyme-linked immunosorbent assay (ELISA), Western immunoblotting, and immunohistochemical staining.


RESULTS
There was a hyperoxic state in the post-ischemic myocardial tissue. Protein expressions of total and active TGF-β1, p-Smad2/3 over t-Smad2/3 ratio, p21, and α-SMA were significantly increased in WT mice compared to Sham control. Knockout of eNOS or iNOS further increased protein expression of these signals. The expression of α-SMA was more abundant in the infarct of eNOS(-/-) and iNOS(-/-) mice than WT mice. A protein band indicating nitration of TGF-β type-II receptor (TGFβRII) was observed from WT heart. Carbogen (95% O2 plus 5% CO2) treatment increased the ratio of p-Smad2/t-Smad2, which was inhibited by 10006329 EUK (EUK134) and sodium nitroprusside (SNP). In conclusion, hyperoxia up-regulated and NO/ONOO(-) inhibited cardiac TGF-β1 signaling and myofibroblast trans-differentiation.


CONCLUSIONS
These findings may provide new insights in myocardial infarct healing and repair.
Davis Heart and Lung Research Institute and Division of Cardiovascular Medicine, Department of Internal Medicine, The Ohio State University Wexner Medical Center, Columbus, OH 43210, USA.
Yuanjing
Y
Li
Ming
M
Cai
Qinghua
Q
Sun
Zhenguo
Z
Liu
Arturo J
AJ
Cardounel
Harold M
HM
Swartz
Guanglong
G
He
D000199:Actins / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects; D004578:Electron Spin Resonance Spectroscopy; D004797:Enzyme-Linked Immunosorbent Assay; D018496:Hyperoxia / Q000209:etiology / Q000378:metabolism*; D007150:Immunohistochemistry; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D017202:Myocardial Ischemia / Q000150:complications*; D058628:Myofibroblasts / Q000166:cytology; D009569:Nitric Oxide / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II / Q000235:genetics; D052250:Nitric Oxide Synthase Type III / Q000235:genetics; D010092:Oximetry; D015398:Signal Transduction / Q000502:physiology*; D051899:Smad2 Protein / Q000378:metabolism; D051900:Smad3 Protein / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000378:metabolism*
C558396:Acta2 protein, mouse; D000199:Actins; D051899:Smad2 Protein; C497783:Smad2 protein, mouse; D051900:Smad3 Protein; C497791:Smad3 protein, mouse; D053773:Transforming Growth Factor beta1; D009569:Nitric Oxide; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; C496317:Nos2 protein, mouse; C496315:Nos3 protein, mouse