Morphine inhibits VEGF expression in myocardial ischemia.
BACKGROUND
Vascular endothelial growth factor (VEGF) may contribute to the resolution of myocardial ischemia by stimulating collateral circulation. Morphine analgesia after myocardial ischemia is thought to increase infarct size. We hypothesize that morphine inhibits myocardial VEGF expression by inhibiting hypoxia-induced factor 1alpha (HIF-1alpha) and the signal transduction mechanisms involving Erk-1,2 MAP kinase (p42/p44), and PI3 kinase activity (phospho-Akt).


METHODS
(1) In vitro: primary cultures of rat cardiac myocytes; (2) in vivo: rat coronary ligation model; (3) mRNA measurement: real-time reverse transcriptase-polymerase chain reaction; (4) protein measurements: enzyme-linked immunosorbent assay, Western immunoblot, electromobility shift assay (EMSA), and immunohistochemistry.


RESULTS
Using rat cardiac myocytes in vitro, we show that morphine: (1) decreases hypoxia-induced VEGF(121) and VEGF(165) mRNA expression and VEGF protein concentration through an opioid receptor mechanism; (2) decreases HIF-1alpha protein expression (immunoblot) and nuclear protein binding to the VEGF HIF-1alpha DNA response element (EMSA); and (3) inhibits phospho-Erk-1,2 MAP kinase (immunoblot) and phospho-Akt kinase activity (immunoblot). Using a rat coronary ligation model, we show that morphine treatment: (1) decreases myocardial VEGF protein expression (immunohistochemistry); (2) decreases HIF-1alpha protein expression (immunoblot); and (3) decreases phospho-Erk-1,2 and phospho-Akt expression.


CONCLUSIONS
(1) Morphine inhibits hypoxia-induced VEGF transcription, in part, through an HIF-1alpha-mediated mechanism and (2) morphine inhibition of hypoxia-induced HIF-1alpha may be mediated by inhibition of ERK 1,2 MAP kinase activity and PI3 kinase activity.
Roy
S
S
Department of Pharmacology, University of Minnesota, Minneapolis Veterans Affairs Medical Center, Minneapois, MN 55417, USA.
Balasubramanian
S
S
Wang
Jinghua
J
Chandrashekhar
Y
Y
Charboneau
R
R
Barke
Roderick
R
D000818:Animals; D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism; D016228:Endothelial Growth Factors / Q000037:antagonists & inhibitors* / Q000235:genetics; D000860:Hypoxia / Q000378:metabolism; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics; D008222:Lymphokines / Q000037:antagonists & inhibitors* / Q000235:genetics; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors; D009020:Morphine / Q000494:pharmacology*; D017202:Myocardial Ischemia / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism; D009294:Narcotics / Q000494:pharmacology*; D009687:Nuclear Proteins / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010750:Phosphoproteins / Q000037:antagonists & inhibitors; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000037:antagonists & inhibitors; D051057:Proto-Oncogene Proteins c-akt; D012333:RNA, Messenger / Q000037:antagonists & inhibitors; D051381:Rats; D017207:Rats, Sprague-Dawley; D011957:Receptors, Opioid / Q000378:metabolism; D020218:Response Elements; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D016228:Endothelial Growth Factors; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D036341:Intercellular Signaling Peptides and Proteins; D008222:Lymphokines; D009294:Narcotics; D009687:Nuclear Proteins; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D011957:Receptors, Opioid; D014157:Transcription Factors; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D009020:Morphine; D004247:DNA; C494920:Akt1 protein, rat; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases