Regulation of endothelial nitric oxide synthase (eNOS) in myocardium subjected to cardioplegic arrest.
BACKGROUND
Nitric oxide (NO) production by both coronary endothelial cells and cardiomyocytes is thought to play a significant role in myocardial pathophysiology following ischemia/reperfusion (I/R).


METHODS
In thirteen pigs subjected to 1 hour cardioplegic arrest (CA) on CPB, left ventricular (LV) biopsies were collected prior to CPB (baseline), at 60 min CPA, at 15 and 30 min reperfusion on CPB, and at 120 min post CPB. LV specimens were immunocytochemically stained against phospho-eNOS (Ser1177), phospho-eNOS (Thr495), phosphorylated ERK1/2, and AKT/PKB. Four additional pigs without CA served as controls. Cardiomyocytes were quantitatively investigated using TV densitometry (gray units: U).


RESULTS
After 60 min CA phosphorylation of eNOS (Ser1177) increased significantly and remained elevated until 30 min of reperfusion. In contrast, eNOS (Thr495) phosphorylation remained unchanged during CA and throughout reperfusion. In control animals, eNOS phosphorylation remained unchanged. Akt/PKB activity significantly increased after 60 min CA and decreased thereafter. ERK1/2 activity remained unchanged during ischemia but increased during reperfusion.


CONCLUSIONS
ENOS activation during ischemia occurs through phosphorylation at Ser1177 mediated by Akt/PKB. ERK1/2 does not seem to be involved in myocardial eNOS regulation especially not via phosphorylation at eNOS (Thr495).
Fischer
U M
UM
Pediatric Surgery, University of Texas Medical School at Houston, Houston, Texas, United States of America.
Klass
O
O
Cox
C S
CS
Stock
U
U
Antonyan
A
A
Brixius
K
K
Fischer
J H
JH
Mehlhorn
U
U
Bloch
W
W
D000818:Animals; D002315:Cardiopulmonary Bypass; D004789:Enzyme Activation; D005260:Female; D006324:Heart Arrest, Induced; D006352:Heart Ventricles / Q000201:enzymology; D007150:Immunohistochemistry; D008297:Male; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D023421:Models, Animal; D009200:Myocardial Contraction; D009206:Myocardium / Q000201:enzymology*; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D012694:Serine; D013552:Swine; D013912:Threonine; D013997:Time Factors; D016277:Ventricular Function, Left
D013912:Threonine; D012694:Serine; D052250:Nitric Oxide Synthase Type III; D051057:Proto-Oncogene Proteins c-akt; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3