Olga Nováková

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.

Seasonal changes in the phospholipid composition in thoracic muscles of a heteropteran, Pyrrhocoris apterus

Abstract The fatty acid composition of phospholipids in thoracic muscles of Pyrrhocoris apterus was related to acclimatization temperature and diapause. Two unsaturated fatty acids, linoleic (18:2n-6) and oleic (18:1n-9), and two saturated, palmitic (16:0) and stearic (18:0), dominated at all temperatures. In contrast to most other reports, the proportion of unsaturated fatty acids did not increase with decreasing temperature; there was a positive correlation between the unsaturation ratio and temperature in total phospholipids ( r =0.67). The most prominent response to cold acclimatization was an increase in the proportion of 16:0 fatty acid and a corresponding decrease in the proportion of fatty acids with 18 carbons. The negative correlation between the proportion of 16:0 and temperature was stronger in phospholipids with phosphatidylethanolamine (PE) head group ( r =−0.85) than in phospholipids with phosphatidylcholine (PC) head group ( r =−0.58). Changes in fatty acid profiles associated with photoperiodic induction of diapause had the same trend as changes related to cold acclimatization. Similar to most other reports, the proportion of PE increased, while the proportion of PC decreased with decreasing temperature. In contrast to a general rule, the PE-phospholipids were less unsaturated than PC-phospholipids.

Deep brain stimulation of the subthalamic nucleus affects resting EEG and visual evoked potentials in Parkinson's disease

OBJECTIVE We studied changes of the EEG spectral power induced by deep brain stimulation (DBS) of the subthalamic nucleus (STN) in patients with Parkinsons disease (PD). Also analyzed were changes of visual evoked potentials (VEP) with DBS on and off. METHODS Eleven patients with advanced PD treated with bilateral DBS STN were examined after an overnight withdrawal of L-DOPA and 2 h after switching off the neurostimulators. All underwent clinical examination followed by resting EEG and VEP recordings, a procedure repeated after DBS STN was switched on. RESULTS With DBS switched on, the dominant EEG frequency increased from 9.44+/-1.3 to 9.71+/-1.3 Hz (P<0.01) while its relative spectral power dropped by 11% on average (P<0.05). Switching on the neurostimulators caused a decrease in the N70/P100 amplitude of the VEP (P<0.01), which inversely correlated with the intensity of DBS (black-and-white pattern: P<0.01; color pattern: P<0.05). CONCLUSIONS Despite artifacts generated by neurostimulators, the VEP and resting EEG were suitable for the detection of effects related to DBS STN. The acceleration of dominant frequency in the alpha band may be evidence of DBS STN influence on speeding up of intracortical oscillations. The spectral power decrease, seen mainly in the fronto-central region, might reflect a desynchronization in the premotor and motor circuits, though no movement was executed. Similarly, desynchronization of the cortical activity recorded posteriorly may by responsible for the VEP amplitude decrease implying DBS STN-related influence even on the visual system. SIGNIFICANCE Changes in idling EEG activity observed diffusely over scalp together with involvement of the VEP suggest that the effects of DBS STN reach far beyond the motor system influencing the basic mechanisms of rhythmic cortical oscillations.

Role of time and concentration on carbon tetrachloride toxicity in rats

SummaryThe hepatotoxic effect of different exposure schemes to carbon tetrachloride (CCl4) was studied in inhalation experiments in rats. The duration of exposures at different concentrations of (CCl4) vapours in air was changed in such a way as to give a constant product of concentration and time (CT). The animals were exposed for 4 successive days a week.A concentration of 1,625 mg/m3 (250 ppm) (CCl4) for 72 min (CT = 300 ppm × h) caused a higher increase in SGPT activity than the exposure to 325 mg/m3 (50 ppm) for 6h (CT = 300); the effect of 6,500 mg/m3 (1,000 ppm) for 3 min six times at 1-h intervals (CT = 300) had a much smaller effect than the exposure to 6,500 mg/m3 (1,000 ppm) for 18 min (again CT = 300) (1 ppm (CCl4) = 6.5 mg/m3). Similar results were obtained at other concentrations and by increasing the number of exposures up to 18; the effects were also confirmed by other biochemical changes in blood serum and liver and by histological examination of the liver.The results indicate that the severity of liver lesions is more influenced by the concentration of (CCl4) in the inhaled air (and accordingly in the blood entering the liver) than by the total inhaled (and absorbed) amount of (CCl4). This also explains the differences between the two types of exposure in the concentration of 6,500 mg/m3 (1,000 ppm): blood cannot be saturated with (CCl4) to the same level within 3 min as within 18 min of exposure.

Structural and biochemical remodelling in catecholamine-induced cardiomyopathy: comparative and ontogenetic aspects

Excessive release or administration of beta-mimetic catecholamines may induce cardiomegaly, necrotic lesions and accumulation of connective tissue in the heart of adult homoiotherms. It was examined here whether similar changes can also be observed at different stages of evolution of the cardiovascular system, i.e. in poikilotherms and in homoiotherms during embryonic life.Sensitivity of the poikilothermic hearts (carp, frog, turtle) to isoproterenol (IPRO) was significantly lower than in the homoiotherms. Necrotic lesions, if present, were localized in the inner spongious musculature which has no vascular supply but which exhibits higher activities of enzymes connected with aerobic oxidation. Moreover, the IPRO-induced decrease of the phospholipid content was also significantly more expressed in the spongious layer. IPRO treatment did not influence the total weight of the fish heart but the proportion of the outer compact layer was significantly higher. These changes were accompanied by an increase of collagen, higher water content and an increase of isomyosin with a lower ATPase activity. The response of the poikilothermic heart to IPRO-induced overload thus differs significantly from that in the homoiotherms.The administration of IPRO during embryonic life of homoiotherms (chick) induces serious cardiovascular disturbances, including cardiomegaly and cellular oedema. Necroses of myofibrils, characteristic of IPRO-induced lesions of adults, were, however, rather exceptional. IPRO did not elevate the concentration of85Sr (as a calcium homologue) in the immature myocardium; it seems, therefore, that IPRO-induced changes of the embryonic heart are not necessarily due to an intracellular calcium overload.It may be concluded that the character of catecholamine-induced cardiomyopathy is not uniform and depends strictly on the stage of cardiac development.

Corticosteroid effect on Caco-2 cell lipids depends on cell differentiation

Previous studies from our laboratory have indicated that secondary hyperaldosteronism affects phospholipids of rat colonic enterocytes. To assess whether this represents a direct effect of mineralocorticoids on enterocytes, the role of aldosterone and dexamethasone in the regulation of lipid metabolism was examined in Caco-2 cells during development of their enterocyte phenotype. Differentiation of Caco-2 cells was associated with increased levels of triglycerides (TG) and cholesteryl esters (CE), a decreased content of cholesterol and phospholipids and changes in individual phospholipid classes. The phospholipids of differentiated cells had a higher content of n-6 polyunsaturated fatty acids (PUFA) and lower amounts of monounsaturated (MUFA) and saturated fatty acids than subconfluent undifferentiated cells. Differentiated cells exhibited a higher ability to incorporate [3H]arachidonic acid (AA) into cellular phospholipids and a lower ability for incorporation into TG and CE. Incubation of subconfluent undifferentiated cells with aldosterone or dexamethasone was without effect on the content of lipids, their fatty acids and [3H]AA incorporation. In contrast, aldosterone treatment of differentiated cells diminished the content of TG, increased the content of phospholipids and modulated their fatty acid composition. The percentage of n-6 and n-3 PUFA in phospholipids was increased and that of MUFA decreased, whereas no changes in TG were observed. The incorporation of [3H]AA into phospholipids was increased and into TG decreased and these changes were blocked by spironolactone. Treatment of differentiated cells with dexamethasone increased their CE content but no effect was identified upon other lipids, their fatty acid composition and on the incorporation of [3H]AA. As expected for the involvement of corticosteroid hormones the mineralocorticoid and glucocorticoid receptors were identified in Caco-2 cells by RT-PCR. The results suggest that aldosterone had a profound influence on lipid metabolism in enterocytes and that its effect depends on the stage of differentiation. The aldosterone-dependent changes occurring in phospholipids and their fatty acid composition may reflect a physiologically important phenomenon with long-term consequences for membrane structure and function.

Postnatal development of phospholipids and their fatty acid profile in rat heart

The aim of this study was to determine the concentration of phospholipids (PL), plasmalogen components of choline (PC) and ethanolamine (PE) phosphoglycerides (PLPC, PLPE) and fatty acid profile of PL and triacylglycerols (TAG) in developing rat left ventricular myocardium between postnatal day (d) 2 and 100. The steepest increase of total PL (TPL) concentration occurs between d2 and d5, followed by a further slower increase between d20 and d40. Similar developmental changes were observed in PC and PE. The PLPE concentration rises by d10, whereas PLPC does not change during the whole period investigated, except for the transient decline on d5. The concentration of diphosphatidylglycerol (DPG) increases by d60; the steepest rise occurs between d20 and d40. Phosphatidylinositol (PI) concentration rises only by d5. The concentration of phosphatidylserine (PS) decreases between d5 and d10 and then it does not change. Sphingomyelin (SM) concentration is maintained till d10, it declines on d20 and does not change thereafter. The proportion of saturated fatty acids (SFA) increases by d5 in PC, PE, PS and TAG, and by d10 in DPG and PI. After d20 the SFA proportion gradually decline in all lipids. Monounsaturated FA (MUFA) proportion decreases in PC, PE, PI and PS from d2 till d10, and in the weaning period it tends to rise again. In contrast, in DPG and TAG the proportion of MUFA declines during the whole postnatal period. N-6 polyunsaturated FA (PUFA) decrease in all PL by d20 and rise again thereafter; in TAG they decline between d2 and d10 and return to the initial level by d100. N-3 PUFA increase in all PL during the suckling period and decline after weaning; in TAG they increase only by d5 and then they decline. This remodeling of myocardial PL and TAG composition during postnatal development may affect membrane properties and contribute to developmental changes in the function of membrane proteins and cell signaling.

Dietary polyunsaturated fatty acids and adaptation to chronic hypoxia alter acyl composition of serum and heart lipids

The effects of dietary supplementation with fat of different fatty acid profile and chronic intermittent hypoxia (CIH) on the fatty acid composition of serum and heart lipids were analysed. Adult male Wistar rats were fed a standard non-fat diet enriched with 10 % of lard, fish oil (n-3 PUFA) or maize oil (n-6 PUFA) for 10 weeks. After 4 weeks on the diets, each group was divided in two subgroups, either exposed to CIH in a barochamber (7000 m, twenty-five exposures) or kept at normoxia. In normoxic rats, the fish oil diet increased the level of conjugated dienes. The n-6:n-3 PUFA ratio in serum TAG, phospholipids (PL), cholesteryl esters (CE) and heart TAG, PL and diacylglycerols (DAG) followed the ratio in the fed diet (in the sequence maize oil>lard>fish oil). In heart TAG, PL and DAG, 20 : 4n-6 and 18 : 2n-6 were replaced by 22 : 6n-3 in the fish oil group. The main fatty acid in CE was 20 : 4n-6 in the lard and maize oil groups whereas in the fish oil group, half of 20 : 4n-6 was replaced by 20 : 5n-3. CIH further increased 20 : 5n-3 in CE in the fish oil group. CIH decreased the n-6:n-3 PUFA ratio in serum CE, heart TAG, PL and DAG in all dietary groups and stimulated the activity of catalase in the maize and fish oil groups. In conclusion, PUFA diets and CIH, both interventions considered to be cardioprotective, distinctly modified the fatty acid profile in serum and heart lipids with specific effects on conjugated diene production and catalase activity.

Protein kinase C activity and isoform expression during early postnatal development of rat myocardium

Total protein kinase C (PKC) activity, its isoform expression, and concentration and fatty acid (FA) composition of diacylglycerol (DAG) were determined in the left ventricular myocardium of the rat during early postnatal development (d 2, 3, 5, 7, and 10). PKC activity measured by the incorporation of 32P into histone IIIS decreased between d 2 and 10 in the homogenate as well as in cytosolic, membrane (100,000g), and nuclear-cytoskeletal-myofilament fractions (1000g). Likewise, the expression of PKC isoforms (α, δ, and ε) determined by immunoblotting generally declined during the period analyzed, although with a variable pattern. In the membrane and nuclear cytoskeletal myofilament fractions, PKCδ and PKCε expression decreased markedly by d 3, returning to or close to the d 2 level immediately on d 5. PKCα expression in the membrane fraction remained almost unchanged by d 7, declining thereafter. PKCδ and PKCε were associated predominantly with particulate fractions, whereas PKCα was more abundant in the cytosolic fraction. DAG concentration exhibited a significant decline by d 5, consistent with the decrease in maximal PKC activity. The unsaturation index of FA in DAG tended to decrease on d 3 owing to the lowered proportion of all polyunsaturated FA of n−6 and n−3 series. These results demonstrate that the developmental decrease in PKC activity and expression in the rat myocardium is not linear and that subcellular localization of the enzyme exhibits isoform-specific day-by-day changes during the early postnatal period. These changes are compatible with the view that PKC signaling may be involved in the control of a rapid switch of myocardial growth pattern during the first week of life.

Aldosterone alters the phospholipid composition of rat colonocytes

Previous studies have shown that aldosterone treatment of amphibian epithelial cells results not only in stimulation of Na(+) absorption but also in changes in phospholipid composition which are necessary for the mineralocorticoid action of aldosterone. The present study was designed to investigate the effect of aldosterone on phospholipids of mammalian epithelia. Phospholipid and fatty acid composition was examined in colonic epithelium (mineralocorticoid target tissue) and thymus (non-mineralocorticoid but glucocorticoid target tissue) of rats which had received aldosterone or vehicle by a miniosmotic pump for 7 days. Aldosterone increased the mass of colonic phospholipids relative to cellular proteins with concomitant changes in the percentage distribution of fatty acids, whereas the relative distribution of membrane phospholipds was not changed. Phosphatidylcholine increased the content of polyunsaturated and decreased that of monounsaturated fatty acids, which predominantly reflected the accretion of arachidonic and a decrease in oleic and palmitoleic acids. Within the phosphatidylethanolamine subclass, pretreatment of rats with aldosterone decreased the content of monounsaturated fatty acids (predominantly oleic and palmitoleic acid) and of n-3 fatty acids, and increased the content of saturated fatty acids (palmitic acid). The saturated-to-nonsaturated fatty acid ratio also significantly increased after aldosterone treatment. No changes in thymic phospholipids were seen. The results are consistent with the contention that aldosterone specifically modulates phospholipid concentration and metabolism in mineralocorticoid target tissue. The changes in phospholipid content and its fatty acid composition during the fully developed effect of aldosterone may reflect a physiologically important phenomenon with long-term consequences for membrane structure and function.