František Kolář

Cardioprotective effects of chronic hypoxia and ischaemic preconditioning are not additive

Abstract The objective of the work was to examine whether adaptation to intermittent high altitude hypoxia and ischaemic preconditioning provide additive protection of the heart against subsequent acute ischaemic injury. Adult male rats were exposed to hypoxia (7000 m, 8 h/day, 24–30 exposures) in a hypobaric chamber. Susceptibility of their hearts to ischaemia-induced ventricular arrhythmias and infarction was evaluated in open-chest animals subjected to 30-min coronary artery occlusion and 4-h reperfusion. Preconditioning was induced by either two (PC1) or five (PC2) occlusions of the same artery for 5 min, each followed by 5-min reperfusion. Adaptation to hypoxia decreased the arrhythmia score from 2.75 ± 0.13 in normoxic controls to 2.17 ± 0.18. Both PC1 and PC2 reduced the arrhythmia score in the controls (0.15 ± 0.10 and 0.71 ± 0.24, respectively), as well as in the hypoxic groups (0.40 ± 0.15 and 0.27 ± 0.15, respectively). The infarct size was reduced from 66.6 ± 2.3% of the area at risk in the controls to 50.2 ± 1.9% in the adapted rats. PC1 conferred further protection in adapted animals (38.4 ± 2.8%) but this combined effect was of the same magnitude as that of preconditioning in the controls (37.5 ± 1.6%). Similar results were obtained using PC2. It is concluded that adaptation to hypoxia decreases the efficiency of ischaemic preconditioning; cardioprotective effects of these two phenomena are not additive. The results are consistent with the view that the mechanisms of protection conferred by chronic hypoxia and preconditioning may share the same signalling pathway.

Effects of mitochondrial KATP modulators on cardioprotection induced by chronic high altitude hypoxia in rats

OBJECTIVES Adaptation of rats to intermittent high altitude hypoxia increases the tolerance of their hearts to acute ischemia/reperfusion injury. Our aim was to examine the role of mitochondrial ATP-sensitive potassium channels (K(ATP)) in this form of protection. METHODS Adult male Wistar rats were exposed to hypoxia of 5000 m in a barochamber for 8 h/day, 5 days a week; the total number of exposures was 24-32. A control group was kept under normoxic conditions (200 m). Infarct size (tetrazolium staining) was measured in anesthetized open-chest animals subjected to 20-min regional ischemia (coronary artery occlusion) and 4-h reperfusion. Isolated perfused hearts were used to assess the recovery of contractile function following 20-min global ischemia and 40-min reperfusion. In the open-chest study, a selective mitochondrial K(ATP) blocker, 5-hydroxydecanoate (5 mg/kg), or openers, diazoxide (10 mg/kg) or BMS-191095 (10 mg/kg), were administered into the jugular vein 5 and 10 min before occlusion, respectively. In the isolated heart study, 5-hydroxydecanoate (250 micromol/l) or diazoxide (50 micromol/l) were added to the perfusion medium 5 or 10 min before ischemia, respectively. RESULTS In the control normoxic group, infarct size occupied 62.2+/-2.0% of the area at risk as compared with 52.7+/-2.5% in the chronically hypoxic group (P<0.05). Post-ischemic recovery of contractile function (dP/dt) reached 60.0+/-3.9% of the pre-ischemic value and it was improved to 72.4+/-1.2% by adaptation to hypoxia (P<0.05). While 5-hydroxydecanoate completely abolished these protective effects of chronic hypoxia, it had no appreciable influence in normoxic groups. In contrast, diazoxide significantly increased the recovery of contractile function and reduced infarct size in normoxic groups only. The later effect was also observed following treatment with BMS-191095. CONCLUSION The results suggest that opening of mitochondrial K(ATP) channels is involved in the cardioprotective mechanism conferred by long-term adaptation to intermittent high altitude hypoxia.

Structure of silicon oxycarbide glasses derived from poly(methylsiloxane) and poly[methyl(phenyl)siloxane] precursors

Abstract The preparation of silicon oxycarbide glasses and structure changes of polysiloxane precursors during heat-treatment in oxidative and nitrogen atmosphere are studied by 13 C and 29 Si magic angle spinning (MAS) NMR, Fourier-transform infrared (FTIR) spectroscopy and thermogravimetry analysis (TG). The reaction mechanisms in inert and oxidation conditions are discussed with respect to the chemical composition of the used precursors. We focused on the formation of new silicon oxycarbide units as well as a highly condensed aromatic carbon phase, the structure of which is discussed. A correlation of thermo-oxidative properties with chemical structure of prepared materials as well as with conditions of their preparation is given.

Regression of Chronic Hypoxia-Induced Pulmonary Hypertension, Right Ventricular Hypertrophy, and Fibrosis: Effect of Enalapril

Chronic hypoxia induces pulmonary hypertension and right ventricular hypertrophy. These changes are completely reversible, except for persistent myocardial fibrosis. The aim of the present study was to determine whether treatment with the angiotensin-converting enzyme (ACE) inhibitor enalapril can reduce the ventricular collagen content in animals recovering from chronic hypoxia. Adult male Wistar rats were exposed to intermittent high-altitude hypoxia simulated in a barochamber (7000 m, 8 hr/day, 5 days a week, 24 exposures), then transferred to normoxia and divided into two groups: (a) treated with enalapril (0.1 g/kg/day for 60 days) and (b) without treatment. The corresponding control groups were kept under normoxic conditions. Enalapril significantly decreased the heart rate, systemic arterial pressure, and absolute left and right ventricular weights in both hypoxic and control rats; on the other hand, the pulmonary blood pressure was unchanged. The content and concentration of collagen was reduced in both ventricles of enalapril-treated hypoxic and control animals by 10–26% compared with the corresponding untreated groups. These data suggest that the partial regression of cardiac fibrosis due to enalapril may be independent of the pressure load.

Inhibition of soluble epoxide hydrolase by cis-4-(4-(3-adamantan-1-ylureido)cyclohexyl- oxy)benzoic acid exhibits antihypertensive and cardioprotective actions in transgenic rats with angiotensin II-dependent hypertension

The present study was undertaken to evaluate the effects of chronic treatment with c-AUCB {cis-4-[4-(3-adamantan-1-ylureido)cyclohexyl-oxy]benzoic acid}, a novel inhibitor of sEH (soluble epoxide hydrolase), which is responsible for the conversion of biologically active EETs (epoxyeicosatrienoic acids) into biologically inactive DHETEs (dihydroxyeicosatrienoic acids), on BP (blood pressure) and myocardial infarct size in male heterozygous TGR (Ren-2 renin transgenic rats) with established hypertension. Normotensive HanSD (Hannover Sprague-Dawley) rats served as controls. Myocardial ischaemia was induced by coronary artery occlusion. Systolic BP was measured in conscious animals by tail plethysmography. c-AUCB was administrated in drinking water. Renal and myocardial concentrations of EETs and DHETEs served as markers of internal production of epoxygenase metabolites. Chronic treatment with c-AUCB, which resulted in significant increases in the availability of biologically active epoxygenase metabolites in TGR (assessed as the ratio of EETs to DHETEs), was accompanied by a significant reduction in BP and a significantly reduced infarct size in TGR as compared with untreated TGR. The cardioprotective action of c-AUCB treatment was completely prevented by acute administration of a selective EETs antagonist [14,15-epoxyeicosa-5(Z)-enoic acid], supporting the notion that the improved cardiac ischaemic tolerance conferred by sEH inhibition is mediated by EETs actions at the cellular level. These findings indicate that chronic inhibition of sEH exhibits antihypertensive and cardioprotective actions in this transgenic model of angiotensin II-dependent hypertension.

Gene expression profiling of sex differences in HIF1-dependent adaptive cardiac responses to chronic hypoxia

Although physiological responses to chronic hypoxia, including pulmonary hypertension and right ventricular hypertrophy, have been well described, the molecular mechanisms involved in cardiopulmonary adaptations are still not fully understood. We hypothesize that adaptive responses to chronic hypoxia are the result of altered transcriptional regulations in the right and left ventricles. Here we report results from the gene expression profiling of adaptive responses in a chronically hypoxic heart. Of 11 analyzed candidate genes, the expression of seven and four genes, respectively, was significantly altered in the right ventricle of hypoxic male and female mice. In the transcriptional profile of the left ventricle, we identified a single expression change in hypoxic males (Vegfa gene). To directly test the role of HIF1, we analyzed the expression profile in Hif1a partially deficient mice exposed to moderate hypoxia. Our data showed that Hif1a partial deficiency significantly altered transcriptional profiles of analyzed genes in hypoxic hearts. The expression changes were only detected in two genes in the right ventricle of Hif1a(+/-) males and in one gene in the right ventricle of Hif1a(+/-) females. First, our results suggest that hypoxia mainly affects adaptive expression profiles in the right ventricle and that each ventricle can respond independently. Second, our findings indicate that HIF1a plays an important role in adaptive cardiopulmonary responses and the dysfunction of HIF1 pathways considerably affects transcriptional regulation in the heart. Third, our data reveal significant differences between males and females in cardiac adaptive responses to hypoxia and indicate the necessity of optimizing diagnostic and therapeutic procedures in clinical practice, with respect to sex.

Chronic hypoxia alters fatty acid composition of phospholipids in right and left ventricular myocardium

Adult male Wistar rats were exposed to intermittent high altitude hypoxia of 7000 m simulated in a hypobaric chamber for 8 h/day, 5 days a week; the total number of exposures was 25. The concentration of individual phospholipids and their fatty acid (FA) profile was determined in right (RV) and left (LV) ventricles. Adaptation to hypoxia decreased the concentration of diphosphatidylglycerol (DPG) in hypertrophied RV by 19% and in non-hypertrophied LV by 12% in comparison with normoxic controls. Chronically hypoxic hearts exhibited lower phospholipid n-6 polyunsaturated FA (PUFA) content mainly due to decreased linoleic acid (18:2n-6), which was opposed by increased n-3 PUFA mainly due to docosahexaenoic acid (22:6n-3) in phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylinositol (PI). The content of arachidonic acid (20:4n-6) was unchanged in total phospholipids, but in PC it was increased in both ventricles (by 22%) and in PE decreased in LV only (by 20%). Chronic hypoxia increased the un-saturation index of PC and PE in both ventricles. The content of monounsaturated FA (MUFA) was increased and 18:2n-6 decreased in DPG. The proportion of saturated FA was increased in PC and PI of hypoxic RV but not LV. The FA composition of phosphatidylserine was not altered in hypoxic ventricles. It is concluded that chronic hypoxia led to only minor changes in individual phospholipid concentration in rat ventricular myocardium, but markedly altered their FA profile. These changes, in particular the greater incorporation of n-3 PUFA into phospholipids and increased un-saturation index, may lead to a better preservation of membrane integrity and thereby contribute to improved ischemic tolerance of chronically hypoxic hearts.

Mitochondrial BKCa channels contribute to protection of cardiomyocytes isolated from chronically hypoxic rats

Chronic hypoxia protects the heart against injury caused by acute oxygen deprivation, but its salutary mechanism is poorly understood. The aim was to find out whether cardiomyocytes isolated from chronically hypoxic hearts retain the improved resistance to injury and whether the mitochondrial large-conductance Ca2+-activated K+ (BKCa) channels contribute to the protective effect. Adult male rats were adapted to continuous normobaric hypoxia (inspired O2 fraction 0.10) for 3 wk or kept at room air (normoxic controls). Myocytes, isolated separately from the left ventricle (LVM), septum (SEPM), and right ventricle, were exposed to 25-min metabolic inhibition with sodium cyanide, followed by 30-min reenergization (MI/R). Some LVM were treated with either 30 μM NS-1619 (BKCa opener), or 2 μM paxilline (BKCa blocker), starting 25 min before metabolic inhibition. Cell injury was detected by Trypan blue exclusion and lactate dehydrogenase (LDH) release. Chronic hypoxia doubled the number of rod-shaped LVM and SEPM surviving the MI/R insult and reduced LDH release. While NS-1619 protected cells from normoxic rats, it had no additive salutary effect in the hypoxic group. Paxilline attenuated the improved resistance of cells from hypoxic animals without affecting normoxic controls; it also abolished the protective effect of NS-1619 on LDH release in the normoxic group. While chronic hypoxia did not affect protein abundance of the BKCa channel regulatory β1-subunit, it markedly decreased its glycosylation level. It is concluded that ventricular myocytes isolated from chronically hypoxic rats retain the improved resistance against injury caused by MI/R. Activation of the mitochondrial BKCa channel likely contributes to this protective effect.

Reperfusion-Induced Injury

In clinical therapy of evolving acute myocardial infarction, early coronary reperfusion has proved to be the only way to limit infarct size. However, there is also evidence from animal studies that reperfusion may contribute to further tissue damage, a phenomenon known as “reperfusion injury”.

Changes in calcium handling in atrophic heterotopically isotransplanted rat hearts

Atrophy of the rat heart induced by hemodynamic unloading after heterotopic transplantation is associated with impaired relaxation while systolic function remains normal when compared to the heart of the recipient animal. To identify possible underlying mechanisms for the above, we studied some aspects of membrane calcium handling using postextrasystolic potentiation of contractions in the isolated right ventricular papillary muscle and in the left ventricle of the Langendorff-perfused heart. We also compared the alterations of the unloaded heart with those of overloaded hypertrophic hearts of rats with suprarenal aortic constriction. In the atrophic heart the degree of potentiation after one extrasystole, considered to be proportional to the trans-sarcolemmal influx of Ca2+ during an action potential, was increased by 125% when compared with recipient hearts. The rate of decay of potentiation which reflects the fraction of activator Ca2+ recirculating in the cells via the sarcoplasmic reticulum, negatively correlated with the degree of potentiation, although its mean value was not significantly altered. In hypertrophic hearts the decay of potentiation was faster when compared with the hearts of sham-operated animals, indicating a decreased recirculating fraction of Ca2+. The data suggest that the relative importance of trans-sarcolemmal Ca2+ fluxes is increased both in cardiac atrophy and hypertrophy, but their quantitative role in the control of cardiac contraction might differ.