Marie Milerová

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.

Neonatal cardiac mitochondria and ischemia/reperfusion injury

Postnatal maturation of the heart is characterized by decreasing tolerance to ischemia/reperfusion (I/R) injury associated with significant changes in mitochondrial function. The aim of this study is to test the hypothesis that the role of the mitochondrial membrane permeability transition pore (MPTP) in the I/R injury differs in the neonatal and in the adult heart. For this purpose, the effect of blockade of MPTP on the degree of I/R injury and the sensitivity of MPTP to swelling-inducing agents was compared in hearts from neonatal (7 days old) and adult (90 days old) Wistar rats. It was found that the release of NAD+ from the perfused heart induced by I/R can be prevented by sanglifehrin A (SfA) only in the adult myocardium; SfA had no protective effect in the neonatal heart. Furthermore, the extent of Ca-induced swelling of mitochondria from neonatal rats was significantly lower than that from the adult animals; mitochondria from neonatal rats were more resistant at higher concentrations of calcium. In addition, not only the extent but also the rate of calcium-induced swelling was about twice higher in adult than in neonatal mitochondria. The results support the idea that lower sensitivity of the neonatal MPTP to opening may be involved in the mechanism of the higher tolerance of the neonatal heart to I/R injury.

Differences between atrial and ventricular protein profiling in children with congenital heart disease

The purpose of the present study was to compare protein profiling of atria and ventricles in children operated for congenital heart disease. Tissue samples were obtained during surgery from patients with normoxemic (ventricular and atrial septal defects) and hypoxemic (tetralogy of Fallot) diseases. Protein fractions were isolated by stepwise extraction from both fight ventricular and atrial musculature. The concentration of total atrial protein in the normoxemic patients exceeded the ventricular value (110±2.1 vs 99.9±4.0mg.g−1 wet weight, respectively); in the hypoxemic group this atrio-ventricular difference disappeared. The concentration of contractile proteins in all cardiac samples was significantly higher in the ventricles as compared with atria, while the concentration of collagenous proteins was significantly higher in the atria (due to a higher amount of the insoluble collagenous fraction). The concentration of sarcoplasmic proteins (containing predominantly enzyme systems for aerobic and anaerobic substrate utilization), however did not differ between ventricles and atria. Furthermore, ventricular contractile fractions obtained from both normoxemic and hypoxemic patients were contaminated with the myosin light chain of atrial origin. Soluble collagenous fractions (containing newly synthesized collagenous proteins, predominantly collagen I and III), derived from all ventricular samples, were contaminated by low molecular weight fragments (mol. weight 29–35 kDa). The proportion of the soluble collagenous fraction was significantly higher in atrial but not in ventricular myocardium of hypoxemic children as compared with the normoxemic group. It seems, therefore, that lower oxygen saturation affects the svnthesis of collagen preferentially in atrial tissue.

Activity of Cytochrome c Oxidase in the Right and Left Ventricular Myocardium of Male and Female Rats Exposed To Intermittent High Altitude Hypoxiaa

Abstract: The aim of the present study was to compare the capacity of the oxidative metabolism (total activity of cytochrome c oxidase, COX) in the right and left ventricular myocardium of adult rats exposed to intermittent high altitude (IHA) hypoxia simulated in a barochamber (5,000 m, 8 h/day, 5 days/wk, for a total of 32 exposures). In male and female rats, IHA induced significant increases of the right ventricular (RV) weight and protein content, whereas left ventricular (LV) weight and protein content remained unaffected. Consequently, the RV/LV ratio in both sexes markedly increased. Similarly, IHA induce an increase of the total activity of COX in RV in both sexes. The specific activities of COX in homogenate as well as in isolated mitochondria were not changed in IHA‐exposed animals, which indicates that the increase of total activity of COX is proportional to the increase of total protein content and RV weight.

Cardiac phosphocreatine deficiency induced by GPA during postnatal development in rat

The effect of chronic administration of β-guanidinopropionic acid (GPA) on the protein profiling, energy metabolism and right ventricular (RV) function was studied in the rat heart during the weaning and adolescence period. GPA was given in tap water (1–1.5%) using pair drink controls. The feeding of animals with GPA solution for a six week period resulted in elevation of heart to body weight ratio due to body growth retardation. GPA accumulated in the myocardium up to 67.37 ± 5.3 μmoles.g dry weight and the tissue content of total creatine, phosphocreatine and ATP was significantly decreased to 15%, 9% and 65% of control values respectively. Total activity of creatine kinase (CK) was not changed, but the proportion of mitochondrial (Mi) CK isoenzyme was decreased; the percentage of MB isoenzyme of CK was significantly higher. GPA treatment resulted in an elevation of the content of cardiac collagenous proteins and decrease of non-collagenous proteins in the heart; in parallel, a decrease of the collagen I to collagen III ratio was detected. The function of the RV was assessed using an isolated perfused heart with RV performing pressure-volume work. As compared to pair-drink controls, RV function was significantly impaired the GPA group: at any given right atrial filling pressure, the RV systolic pressure and the rate of pressure development were decreased by almost a factor of two. Elevation of the RV diastolic pressure with increasing pulmonary artery diastolic pressure was also significantly steeper in the GPA group which also showed decrease of cardiac output, especially at high outflow resistance. It may be assumed that chronic administration of GPA deeply influenced metabolic parameters, protein profiles and contractile function of the developing heart. On the other hand, concentrations of glucose, total lipids and triglycerides in blood plasma were not affected. All these data confirm the concept that the CK system is of central importance both for heart function and for the regulation of normal growth of cardiac myocytes.

Effect of the preweaning nutritional state on the cardiac protein profile and functional performance of the rat heart

The aim of the study was to find out whether the changes in nutritional status induced by different litter size during early postnatal development can influence quantitative and qualitative protein remodeling and contractile performance of the myocardium. Male Wistar rats born at the same day were pooled together at 2 days postbirth and assigned by random selection to dams in groups of 4, 8 or 16 rats/litter. The animals were investigated at the age of 4 and 16 weeks. The results revealed that the early postnatal nutritional modification altered weight parameters: whereas lower heart weight persisted in slow-growing rats until 16 weeks, higher body weight of fast-growing rats returned to the control level at the age of 16 weeks. Altered nutritional status influenced also protein remodeling of the myocardium: the concentration of all noncollagenous proteins (fractions of metabolic and contractile proteins) significantly increased in slow-growing rats, on the other hand, the concentration of collagenous proteins (pepsin-soluble and -insoluble fractions) was higher in fast-growing animals. The changes were, however, only transitional: three months after the end of the weaning period most protein changes returned to the control level. However, higher concentration of total blood lipids and triglycerides in fast-growing rats persisted until adulthood. Nutritional changes had, however, only minor effect on ventricular performance. No differences among groups were observed in basal values of the left ventricular pressure, while the maximum pressure attained after an acute ventricular loading and the contractile reserve were significantly decreased in slowgrowing 4 week old rats. The functional consequence of altered nutritional status during weaning was only transitional, in agreement with the transient character of most structural and biochemical markers of myocardial remodeling.

Development of cytochrome-c oxidase activity in rat heart : Downregulation in newborn rats

Cytochrome-c oxidase (COX) activity of the rat heart was two- to sevenfold activated when the membrane integrity was disrupted by hypotonic swelling, freezing-thawing, or a detergent, indicating that a large portion of COX capacity in intact mitochondria is not active. The effect of detergent was tested in heart mitochondria isolated from 1-, 5-, 15-, 29-, and 60-d-old rats; activation by detergent was up to 20-fold in 1-d-old animals, whereas in mitochondria from 60-d-old rats it was only 7-fold. Our data indicate that the rat heart exhibits significant developmental changes dependent on downregulation of COX activity in the neonatal period.

Tissue Specific Sensitivity of Mitochondrial Permeability Transition Pore to Ca2+ Ions.

Ca2+-induced opening of the mitochondrial permeability transition pore (MPTP) is involved in induction of apoptotic and necrotic processes. We studied sensitivity of MPTP to calcium using the model of Ca2+-induced, cyclosporine A-sensitive mitochondrial swelling. Presented data indicate that the extent of mitochondrial swelling (dA520/4 min) induced by addition of 25 μM Ca2+ is seven-fold higher in liver than in heart mitochondria (0.564 ± 0.08/0.077± 0.01). The extent of swelling induced by 100 μM Ca2+ was in liver tree times higher than in heart mitochondria (0.508±0.05/ 0.173±0.02). Cyclosporine A sensitivity showed that opening of the MPTP is involved. We may thus conclude that especially at low Ca2+ concentration heart mitochondria are more resistant to damaging effect of Ca2+ than liver mitochondria. These finding thus support hypothesis that there exist tissue specific strategies of cell protection against induction of the apoptotic and necrotic processes.

Protein and Phospholipids Composition of Human Myocardium in Children with Congenital Heart Disease

Congenital heart disease is caused by abnormalities in the first 6–8 weeks of fetal life; incidence of heart malformations is approximately eight out of 1000; they resulted from aberrant organogenesis due to dysfunction of genes and the encoded proteins. We, therefore, analyzed protein and phospholipids profiles of human myocardium (tissue samples were obtained during surgery of children with normoxemic and hypoxemic congenital heart disease). Protein profile of the atrial and ventricular myocardium differs significantly; higher concentration of non-collagenous proteins due to contractile ones and lower concentration of extracellular matrix proteins (ECM) due to pepsin-insoluble collagenous proteins were in ventricles. Furthermore, there were significant differences in activities of metabolic enzymes from both glycolytic and oxidative pathways; it correlated with higher amount of cardiolipin in ventricle (mitochondrial phospholipids). Moreover, electrophoretic pattern of both heavy and light chains of myosin in ventricular and atrial musculature of damage myocardium were different; however, hypoxemia decreased ATPase activity of myosin in ventricle only. Compartmentation of troponin T between myofibrillar and cytosolic pools was not affected by hypoxemia. Synthesis of newly formed collagenous proteins depend on balance between synthetic and degradation processes; they are mediated by metaloproteinases (major band in atria and ventricle has mol..weight of 95kDa). Hypoxemia induced in atria both higher amount of collagen (predominantly collagen III) and activation of metaloproteinases. Significantly higher concentration of phospholipids was in ventricles; hypoxemia did not affected this atrio/ventricular difference. However, composition of phospholipids species in both parts of diseased myocardium did not differ (phosphatidylcholine and phosphatidylethanolamine occupied about 70% from all phosholipids). Surgeons, who are reconstructing cardiac anatomy, should be aware that they are dealing with biochemical abnormal tissue and they should respect atrio/ventricular differences.