to dihydroxyeicosatrienoic acids. EETs are formed from arachidonic acid during myocardial ischemia and play a protective role against ischemic cell death. Deletion of sEH has been shown to be protective against myocardial ischemia in the isolated heart preparation. We tested the hypothesis that sEH inactivation by targeted gene deletion or pharmacological inhibition reduces infarct size (I) after regional myocardial ischemia-reperfusion injury in vivo. Male C57BL‫6گ‬J wildtype or sEH knockout mice were subjected to 40 min of left coronary artery (LCA) occlusion and 2 h of reperfusion. Wild-type mice were injected intraperitoneally with 12-(3-adamantan-1-yl-ureido)-dodecanoic acid butyl ester (AUDA-BE), a sEH inhibitor, 30 min before LCA occlusion or during ischemia 10 min before reperfusion. 14,15-EET, the main substrate for sEH, was administered intravenously 15 min before LCA occlusion or during ischemia 5 min before reperfusion. The EET antagonist 14,15-epoxyeicosa-5(Z)-enoic acid (EEZE) was given intravenously 15 min before reperfusion. Area at risk (AAR) and I were assessed using fluorescent microspheres and triphenyltetrazolium chloride, and I was expressed as I/AAR. I was significantly reduced in animals treated with AUDA-BE or 14,15-EET, independent of the time of administration. The cardioprotective effect of AUDA-BE was abolished by the EET antagonist 14,15-EEZE. Immunohistochemistry revealed abundant sEH protein expression in left ventricular tissue. Strategies to increase 14,15-EET, including sEH inactivation, may represent a novel therapeutic approach for cardioprotection against myocardial ischemia-reperfusion injury.14,15-epoxyeicosatrienoic acids; 12-(3-adamantan-1-yl-ureido)-dodecanoic acid butyl ester THE P-450 EPOXYGENASE PATHWAY metabolizes arachidonic acid into four biologically active eicosanoids, referred to as epoxyeicosatrienoic acids (5,6-, 8,9-, 11,12-, and 14,15-EET) (10). EETs play an important role in regulating tissue perfusion in both cardiac and extracardiac organs. The actions of EETs are terminated by conversion to dihydroxyeicosatrienoic acids (DHETs) by epoxide hydrolases (11). Two major epoxide hydrolases are found in mammalian tissues, the microsomal (mEH) and soluble epoxide hydrolases (sEH) (2). However, sEH is the primary enzyme involved in the in vivo metabolism of EETs (19). In addition to their vascular effects, EETs exhibit a cardioprotective effect, which has been linked to activation of the reperfusion injury salvage kinase pathway (18) and mediated in part through activation of the phosphatidylinositol 3-kinase/Akt pathway and the mitochondrial ATP-sensitive K ϩ channels (5, 15). Augmenting endogenously released EETs by inhibiting the converting enzyme sEH represents an attractive strategy to increase ischemic tolerance. Our laboratory has recently used this strategy to show that pharmacological inhibition (20) and gene deletion (21) are protective against experimental stroke in vivo. Similarly, using the isolated heart preparation, Seubert et al. (16) demonstrat...

Acute MI stimulates a spectrum of changes in remote vessels, including up-regulation of endothelial inflammatory adhesion molecules and platelet-endothelial adhesion from endothelial-associated VWF multimers. These remote arterial alterations persist longer in the presence of hyperlipidemia, are associated with accelerated plaque growth and inflammation, and are attenuated by Nox inhibition.

The third-generation tyrosine kinase inhibitor (TKI) ponatinib has been associated with high rates of acute ischemic events. The pathophysiology responsible for these events is unknown. We hypothesized that ponatinib produces an endothelial angiopathy involving excessive endothelial-associated von Willebrand factor (VWF) and secondary platelet adhesion. In wild-type mice and ApoE−/− mice on a Western diet, ultrasound molecular imaging of the thoracic aorta for VWF A1-domain and glycoprotein-Ibα was performed to quantify endothelial-associated VWF and platelet adhesion. After treatment of wild-type mice for 7 days, aortic molecular signal for endothelial-associated VWF and platelet adhesion were five- to sixfold higher in ponatinib vs sham therapy (P < .001), whereas dasatinib had no effect. In ApoE−/− mice, aortic VWF and platelet signals were two- to fourfold higher for ponatinib-treated compared with sham-treated mice (P < .05) and were significantly higher than in treated wild-type mice (P < .05). Platelet and VWF signals in ponatinib-treated mice were significantly reduced by N-acetylcysteine and completely eliminated by recombinant ADAMTS13. Ponatinib produced segmental left ventricular wall motion abnormalities in 33% of wild-type and 45% of ApoE−/− mice and corresponding patchy perfusion defects, yet coronary arteries were normal on angiography. Instead, a global microvascular angiopathy was detected by immunohistochemistry and by intravital microscopy observation of platelet aggregates and nets associated with endothelial cells and leukocytes. Our findings reveal a new form of vascular toxicity for the TKI ponatinib that involves VWF-mediated platelet adhesion and a secondary microvascular angiopathy that produces ischemic wall motion abnormalities. These processes can be mitigated by interventions known to reduce VWF multimer size.

A previous study demonstrated that the threshold dose of intra-arterial angiotensin II required to induce a pressor response in the rat was significantly lower when the drug was administered into the carotid artery than when administered into the abdominal aorta. This result was interpreted to indicate that part of the increase in arterial pressure produced by low concentrations of blood-borne angiotensin in this species was the result of an effect on structures in the central nervous system selectively accessible via the carotid vascular bed. The purpose of the present study was to establish more precisely the site of the pressor action of angiotensin within the central nervous system. The central component of the pressor effect of angiotensin was quantified as the difference in pressor responses to intracarotid and intra-aortic infusions of angiotensin II (delta c-a). In conscious rats, delta c-a was attenuated by administration of the angiotensin antagonist, saralasin, into the third cerebral ventricle. In rats with chronic electrolytic lesions of the anteroventral third ventricle (AV3V), delta c-a was abolished. Periventricular structures surrounding the third ventricle appear to mediate the central component of the pressor action of blood-borne angiotensin in the rat.

Soluble epoxide hydrolase (sEH) metabolizes epoxyeicosatrienoic acids (EETs), primarily 14,15-EET. EETs are derived from arachidonic acid via P-450 epoxygenases and are cardioprotective. We tested the hypothesis that sEH deficiency and pharmacological inhibition elicit tolerance to ischemia via EET-mediated STAT3 signaling in vitro and in vivo. In addition, the relevance of single nucleotide polymorphisms (SNPs) of EPHX2 (the gene encoding sEH) on tolerance to oxygen and glucose deprivation and reoxygenation and glucose repletion (OGD/RGR) was assessed in male C57BL\6J (WT) or sEH knockout (sEHKO) cardiomyocytes by using transactivator of transcription (TAT)-mediated transduction with sEH mutant proteins. Cell death and hydrolase activity was lower in Arg287Gln EPHX2 mutants vs. nontransduced controls. Excess 14,15-EET and SEH inhibition did not improve cell survival in Arg287Gln mutants. In WT cells, the putative EET receptor antagonist, 14,15-EEZE, abolished the effect of 14,15-EET and sEH inhibition. Cotreatment with 14,15-EET and SEH inhibition did not provide increased protection. In vitro, STAT3 inhibition blocked 14,15-EET cytoprotection, but not the effect of SEH inhibition. However, STAT3 small interfering RNA (siRNA) abolished cytoprotection by 14,15-EET and sEH inhibition, but cells pretreated with JAK2 siRNA remained protected. In vivo, STAT3 inhibition abolished 14,15-EET-mediated infarct size reduction. In summary, the Arg287Gln mutation is associated with improved tolerance against ischemia in vitro, and inhibition of sEH preserves cardiomyocyte viability following OGD/RGR via an EET-dependent mechanism. In vivo and in vitro, 14,15-EET-mediated protection is mediated in part by STAT3.