ERK phosphorylation mediates sildenafil-induced myocardial protection against ischemia-reperfusion injury in mice.
Sildenafil, a selective inhibitor of phosphodiesterase type 5, induces powerful protection against myocardial ischemia-reperfusion injury through activation of cGMP-dependent protein kinase (PKG). We further hypothesized that PKG-dependent activation of survival kinase ERK may play a causative role in sildenafil-induced cardioprotection via induction of endothelial nitric oxide synthase (eNOS)/inducible nitric oxide synthase (iNOS) and Bcl-2. Our results show that acute intracoronary infusion of sildenafil in Langendorff isolated mouse hearts before global ischemia-reperfusion significantly reduced myocardial infarct size (from 29.4 +/- 2.4% to 15.9 +/- 3.0%; P < 0.05). Cotreatment with ERK inhibitor PD98059 abrogated sildenafil-induced protection (31.8 +/- 4.4%). To further evaluate the role of ERK in delayed cardioprotection, mice were treated with sildenafil (ip) 24 h before global ischemia-reperfusion. PD98059 was administered (ip) 30 min before sildenafil treatment. Infarct size was reduced from 27.6 +/- 3.3% in controls to 7.1 +/- 1.5% in sildenafil-treated mice (P < 0.05). The delayed protective effect of sildenafil was also abolished by PD98059 (22.5 +/- 2.3%). Western blots revealed that sildenafil significantly increased phosphorylation of ERK1/2 and GSK-3beta and induced iNOS, eNOS, Bcl-2, and PKG activity in the heart 24 h after treatment. PD98059 inhibited the enhanced expression of iNOS, eNOS, and Bcl-2 and the phosphorylation of GSK-3beta. PD98059 had no effect on the sildenafil-induced activation of PKG. We conclude that these studies provide first direct evidence that PKG-dependent ERK phosphorylation is indispensable for the induction of eNOS/iNOS and Bcl-2 and the resulting cardioprotection by sildenafil.
Das
Anindita
A
Division of Cardiology, Department of Internal Medicine, Box 980281, Virginia Commonwealth Univ. Medical Center, 1101 East Marshall St., Rm. 7-040, Richmond, VA 23298-0281, USA. adas2@vcu.edu
Salloum
Fadi N
FN
Xi
Lei
L
Rao
Yuan J
YJ
Kukreja
Rakesh C
RC
D000818:Animals; D002317:Cardiovascular Agents / Q000494:pharmacology*; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism; D054706:Cyclic Nucleotide Phosphodiesterases, Type 5 / Q000378:metabolism; D004195:Disease Models, Animal; D004789:Enzyme Activation; D005419:Flavonoids / Q000494:pharmacology; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D006439:Hemodynamics / Q000187:drug effects; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009200:Myocardial Contraction / Q000187:drug effects; D009203:Myocardial Infarction / Q000201:enzymology / Q000503:physiopathology / Q000517:prevention & control*; D015428:Myocardial Reperfusion Injury / Q000201:enzymology / Q000503:physiopathology / Q000517:prevention & control*; D009206:Myocardium / Q000201:enzymology* / Q000473:pathology; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism; D058986:Phosphodiesterase 5 Inhibitors; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology*; D010766:Phosphorylation; D010879:Piperazines / Q000494:pharmacology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011687:Purines / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D000068677:Sildenafil Citrate; D013450:Sulfones / Q000494:pharmacology*; D013997:Time Factors; D051028:bcl-2-Associated X Protein / Q000378:metabolism
C494827:Bax protein, mouse; D002317:Cardiovascular Agents; D005419:Flavonoids; D058986:Phosphodiesterase 5 Inhibitors; D010726:Phosphodiesterase Inhibitors; D010879:Piperazines; D047428:Protein Kinase Inhibitors; D019253:Proto-Oncogene Proteins c-bcl-2; D011687:Purines; D013450:Sulfones; D051028:bcl-2-Associated X Protein; D000068677:Sildenafil Citrate; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; C496317:Nos2 protein, mouse; C496315:Nos3 protein, mouse; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; D017869:Cyclic GMP-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D038362:Glycogen Synthase Kinase 3; D054706:Cyclic Nucleotide Phosphodiesterases, Type 5; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one