Mechanism of cyclooxygenase-2 upregulation in late preconditioning.
Although the cardioprotection of late preconditioning (PC) is known to be mediated by both inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2), the signaling mechanism responsible for COX-2 upregulation and the interaction between iNOS and COX-2 remain unknown. A total of 122 mice were used to address this issue. In wild-type mice preconditioned with six cycles of 4-min coronary occlusion-4-min reperfusion, ischemic PC resulted in rapid activation of nuclear STAT1/3 through tyrosine phosphorylation (STAT1: 339 +/- 48% of control; STAT3: 389 +/- 46% of control) and increased STAT1/3-DNA binding activity (687 +/- 58% of control) at 30 min after PC, with subsequent upregulation of COX-2 protein (373 +/- 60% of control) and activity(increased myocardial levels of PGE2, PGF(2alpha), and 6-keto-PGF(1alpha)) at 24 h. However, COX-1 protein was not changed 24 h after ischemic PC. Pretreatment with the Janus tyrosine kinase (JAK) inhibitor AG-490 before the six occlusion-reperfusion cycles blocked both the tyrosine phosphorylation of STAT1/3 and the subsequent upregulation of COX-2 protein, demonstrating a necessary role of the JAK-STAT pathway in the induction of COX-2. Targeted disruption of the iNOS gene (iNOS-/-) did not block the increased expression of COX-2 protein 24 h after ischemic PC but completely blocked the increase in COX-2 activity, whereas targeted disruption of the COX-2 gene (COX-2-/-) did not alter ischemic PC-induced iNOS induction. Immunoprecipitation of preconditioned heart tissues with anti-COX-2 antibodies followed by immunoblotting with anti-iNOS antibodies revealed that the increased iNOS protein co-precipitated with COX-2. We conclude that (i) the upregulation of COX-2 protein expression after ischemic PC is mediated by a JAK1/2-STAT1/3-signaling cascade; (ii) COX-2 activity requires upregulated iNOS and iNOS-derived NO; and (iii) COX-2 forms complexes with iNOS, supporting a direct interaction between these two proteins. To our knowledge, this is the first evidence that myocardial COX-2 is upregulated via a JAK1/2-STAT1/3 pathway.
Experimental Research Laboratory, Division of Cardiology, University of Louisville and the Jewish Hospital Heart and Lung Institute, Louisville, KY 40292, USA.
Yu-Ting
YT
Xuan
Yiru
Y
Guo
Yanqing
Y
Zhu
Hui
H
Han
Robert
R
Langenbach
Buddhadeb
B
Dawn
Roberto
R
Bolli
D000818:Animals; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D019157:Ischemic Preconditioning, Myocardial; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins; D051379:Mice; D018345:Mice, Knockout; D019001:Nitric Oxide Synthase / Q000235:genetics / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D010766:Phosphorylation; D010770:Phosphotransferases; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics / Q000378:metabolism*; D050794:STAT1 Transcription Factor; D050796:STAT3 Transcription Factor; D015534:Trans-Activators / Q000378:metabolism; D015854:Up-Regulation / Q000502:physiology*
D004268:DNA-Binding Proteins; D007527:Isoenzymes; D008565:Membrane Proteins; D050794:STAT1 Transcription Factor; D050796:STAT3 Transcription Factor; C494080:Stat1 protein, mouse; C494087:Stat3 protein, mouse; D015534:Trans-Activators; D004247:DNA; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases; C496512:Ptgs1 protein, mouse; D010770:Phosphotransferases