Veronika Fekete

Cholesterol diet-induced hyperlipidemia impairs the cardioprotective effect of postconditioning: role of peroxynitrite

The aim of the present study was to investigate if hyperlipidemia interferes with the infarct size-limiting effect of postconditioning and to study the involvement of peroxynitrite in this phenomenon. Rats were fed a 2% cholesterol-enriched or normal diet for 12 wk. Infarct size by triphenyltetrazolium chloride staining was measured in hearts isolated from both groups and subjected to 30 min coronary occlusion followed by 120 min reperfusion with or without the postconditioning protocol induced by six cycles of 10 s coronary occlusion and 10 s reperfusion at the onset of the reperfusion. Postconditioning significantly decreased infarct size in the normolipidemic but not in the hyperlipidemic group. Postconditioning increased cardiac 3-nitrotyrosine concentration (a marker for peroxynitrite formation) in the normal but not in the cholesterol-fed group when measured at the 5th min of reperfusion. Next, we tested if the postconditioning-induced acute increase in peroxynitrite is involved in the cardioprotection in normolipidemic animals in separate experiments. Postconditioning failed to decrease infarct size in the presence of the peroxynitrite decomposition catalyst 5,10,15,20-tetrakis-[4-sulfonatophenyl]-porphyrinato-iron [III] (20 mg/l) in normolipidemic animals. We conclude that an early increase in peroxynitrite after postconditioning plays a role in cardioprotection. Furthermore, hyperlipidemia blocks the cardioprotective effect of postconditioning at least in part via deterioration of the postconditioning-induced early increase in peroxynitrite formation.

Lovastatin interferes with the infarct size-limiting effect of ischemic preconditioning and postconditioning in rat hearts

Statins have been shown to be cardioprotective; however, their interaction with endogenous cardioprotection by ischemic preconditioning and postconditioning is not known. In the present study, we examined if acute and chronic administration of the 3-hydroxy-3-methylglutaryl CoA reductase inhibitor lovastatin affected the infarct size-limiting effect of ischemic preconditioning and postconditioning in rat hearts. Wistar rats were randomly assigned to the following three groups: 1) vehicle (1% methylcellulose per os for 12 days), 2) chronic lovastatin (15 mg.kg(-1).day(-1) per os for 12 days), and 3) acute lovastatin (1% methylcellulose per os for 12 days and 50 micromol/l lovastatin in the perfusate). Hearts isolated from the three groups were either subjected to a nonconditioning (aerobic perfusion followed by 30-min coronary occlusion and 120-min reperfusion, i.e., test ischemia-reperfusion), preconditioning (three intermittent periods of 5-min ischemia-reperfusion cycles before test ischemia-reperfusion), or postconditioning (six cycles of 10-s ischemia-reperfusion after test ischemia) perfusion protocol. Preconditioning and postconditioning significantly decreased infarct size in vehicle-treated hearts. However, preconditioning failed to decrease infarct size in acute lovastatin-treated hearts, but the effect of postconditioning remained unchanged. Chronic lovastatin treatment abolished postconditioning but not preconditioning; however, it decreased infarct size in the nonconditioned group. Myocardial levels of coenzyme Q9 were decreased in both acute and chronic lovastatin-treated rats. Western blot analysis revealed that both acute and chronic lovastatin treatment attenuated the phoshorylation of Akt; however, acute but not chronic lovastatin treatment increased the phosphorylation of p42 MAPK/ERK. We conclude that, although lovastatin may lead to cardioprotection, it interferes with the mechanisms of cardiac adaptation to ischemic stress.

Measurement of myocardial infarct size in preclinical studies.

Ischemic heart disease is a major cause of morbidity and mortality worldwide. Myocardial ischemia followed by reperfusion results in tissue injury termed ischemia/reperfusion injury which is characterized by decreased myocardial contractile function, occurrence of arrhythmias, and development of tissue necrosis (infarction). These pathologies are all relevant as clinical consequences of myocardial ischemia/reperfusion injury and they are also important as experimental correlates and endpoints. The most critical determinant of acute and long-term mortality after myocardial infarction is the volume of the infarcted tissue. Therefore, development of cardioprotective therapies aims at reducing the size of the infarct developing due to myocardial ischemia/reperfusion injury. Different techniques are available to measure myocardial infarct size in humans and in experimental settings, however, accurate determination of the extent of infarction is necessary to evaluate interventions that may delay the onset of necrosis and/or limit the total extent of infarct size during ischemia/reperfusion. This paper highlights recent advances of the different techniques to measure infarct size.

Metabolic syndrome influences cardiac gene expression pattern at the transcript level in male ZDF rats

BackgroundMetabolic syndrome (coexisting visceral obesity, dyslipidemia, hyperglycemia, and hypertension) is a prominent risk factor for cardiovascular morbidity and mortality, however, its effect on cardiac gene expression pattern is unclear. Therefore, we examined the possible alterations in cardiac gene expression pattern in male Zucker Diabetic Fatty (ZDF) rats, a model of metabolic syndrome.MethodsFasting blood glucose, serum insulin, cholesterol and triglyceride levels were measured at 6, 16, and 25 wk of age in male ZDF and lean control rats. Oral glucose tolerance test was performed at 16 and 25 wk of age. At week 25, total RNA was isolated from the myocardium and assayed by rat oligonucleotide microarray for 14921 genes. Expression of selected genes was confirmed by qRT-PCR.ResultsFasting blood glucose, serum insulin, cholesterol and triglyceride levels were significantly increased, glucose tolerance and insulin sensitivity were impaired in ZDF rats compared to leans. In hearts of ZDF rats, 36 genes showed significant up-regulation and 49 genes showed down-regulation as compared to lean controls. Genes with significantly altered expression in the heart due to metabolic syndrome includes functional clusters of metabolism (e.g. 3-hydroxy-3-methylglutaryl-Coenzyme A synthase 2; argininosuccinate synthetase; 2-amino-3-ketobutyrate-coenzyme A ligase), structural proteins (e.g. myosin IXA; aggrecan1), signal transduction (e.g. activating transcription factor 3; phospholipase A2; insulin responsive sequence DNA binding protein-1) stress response (e.g. heat shock 70kD protein 1A; heat shock protein 60; glutathione S-transferase Yc2 subunit), ion channels and receptors (e.g. ATPase, (Na+)/K+ transporting, beta 4 polypeptide; ATPase, H+/K+ transporting, nongastric, alpha polypeptide). Moreover some other genes with no definite functional clusters were also changed such as e.g. S100 calcium binding protein A3; ubiquitin carboxy-terminal hydrolase L1; interleukin 18. Gene ontology analysis revealed several significantly enriched functional inter-relationships between genes influenced by metabolic syndrome.ConclusionsMetabolic syndrome significantly alters cardiac gene expression profile which may be involved in development of cardiac pathologies in the presence of metabolic syndrome.

Myocardial postconditioning is lost in vascular nitrate tolerance

Abstract: Organic nitrates play an important role in the therapy of ischemic heart disease; however, their clinical application is limited by the development of vascular nitrate tolerance. We have previously shown attenuation of the cardioprotective effect of preconditioning in vascular nitrate tolerance. Here, we studied whether the development of vascular nitrate tolerance affects the infarct size, limiting effect of ischemic postconditioning (IPost) in the myocardium, and whether the activation of survival kinases plays a role in the molecular mechanism of postconditioning in the presence or absence of vascular nitrate tolerance. Male Wistar rats were treated with nitroglycerin/vehicle for 3 days to induce vascular nitrate tolerance. On the fourth day, isolated hearts were subjected to 30-minute coronary occlusion followed by 120-minute reperfusion with or without IPost. In nontolerant hearts, postconditioning significantly decreased infarct size as compared with ischemia/reperfusion; however, postconditioning failed to decrease infarct size in hearts of nitrate tolerant rats. Phosphorylation of ERK 1/2, Akt, or endothelial nitric oxide synthetase showed no significant differences between the groups at the 10th minute of reperfusion. Vascular nitrate tolerance interferes with the infarct size limiting effect of IPost. Activation of survival kinases is not crucial in the molecular mechanism of postconditioning, which remains unaffected in nitrate tolerance.

Tissue-specific Gene Expression in Rat Hearts and Aortas in a Model of Vascular Nitrate Tolerance.

Abstract: Nitroglycerin exerts a direct myocardial anti-ischemic effect even in the state of vascular nitrate tolerance. To examine the potentially diverse molecular responses in vascular and cardiac tissues, we investigated the gene expression profile of the heart and the aorta by DNA microarray in male Wistar rats that were previously made tolerant to the vascular effects of nitroglycerin. The blood pressure–lowering effect of nitroglycerin (1–100 &mgr;g/kg) was markedly attenuated in rats pretreated for 3 days with 3 × 100 mg/kg nitroglycerin. Nitric oxide content was significantly elevated in the heart but not in the aorta of nitrate-tolerant animals, which indicated tissue-specific differences in nitroglycerin bioconversion. Of 7742 genes analyzed by DNA microarray, we found that although the expression of 25 genes changed significantly in the heart (increased: Tas2r119, Map6, Cd59, Kcnh2, Kcnh3, Senp6, Mcpt1, Tshb, Haus1, Vipr1, Lrn3, Lifr; decreased: Ihh, Fgfr1, Cryge, Krt9, Agrn, C4bpb, Fcer1a, Csf3, Hsd17b11, Hsd11b2, Ctnnbl1, Prpg1, Hsf1), only 14 genes were altered in the aorta (increased: Tas2r119, Ihh, Rrad, Npm1, Snai1; decreased: Tubb2b, Usp15, Sema6c, Wfdc2, Rps21, Ramp2, Galr1, Atxn1, Lhx1) in vascular nitrate tolerance. Quantitative reverse transcription polymerase chain reaction analysis of genes related to oxidative/nitrative/nitrosative stress also showed differential expression pattern in the heart and aorta. This is the first pharmacogenomic analysis showing that nitroglycerin treatment leading to vascular nitrate tolerance differentially impacts gene expression in vascular and cardiac tissues, which indicates different tissue-specific downstream signaling pathways.