Takashi Yanaga

The effects of renin-angiotensin system inhibition on aortic cholesterol content in cholesterol-fed rabbits

To investigate how the renin-angiotensin system (RAS) might be involved in cholesterol-induced atherosclerosis, we studied the effects of a nonsulhydryl angiotensin converting enzyme (ACE) inhibitor, enalapril, and an angiotensin II receptor antagonist, E-4177, in cholesterol fed rabbits. Japanese white rabbits were randomly divided into four groups with the following dietary regimens: group A (n = 8) received a standard diet; group B (n = 8) had a 0.5% cholesterol diet; group C (n = 8) had a 0.5% cholesterol diet plus enalapril (10 mg/kg/day, p.o.); group D (n = 8) received a 0.5% cholesterol diet plus E-4177 (20 mg/kg/day, p.o.) and were fed these diets for 5 weeks. Enalapril or E-4177 had no significant effect on either the total plasma or the high density lipoprotein (HDL) cholesterol concentrations. However, the aortic cholesterol content in groups C and D was equally significantly less than that in group B. The plasma and aortic ACE activities were significantly reduced only in group C compared with those in the other groups. The aortic ACE mRNA and AT1 mRNA levels were assessed by a reverse transcription polymerase chain reaction (RT-PCR). The aortic ACE mRNA level was only significantly less in group C than in any of the other groups. The aortic AT1 mRNA level increased significantly in group B compared with that in group A and was significantly and equally reduced in both groups C and D compared with that in group B. These data indicate that angiotensin II rather than ACE may therefore be related to aortic cholesterol content. It follows therefore that the inhibition of angiotensin II by either ACE inhibitor or angiotensin II (type 1) receptor antagonist may play a role in prevention of atherosclerosis.

Interleukin 1α-induced expression of manganous superoxide dismutase reduces myocardial reperfusion injury in the rat

We investigated the effects of pretreatment with interleukin(IL)-1α on the expression of manganous (Mn) superoxide dismutase (DOD) mRNA and reperfusion-induced arrhythmias and the size of myocardial infarct in rats. Male Wistar rats received 10 mg intraperitoneal injetions of human recombinant IL-1α. Their hearts were thereafter isolated at 6, 12, 24, 36 h. A Northern analysis showed that Mn-SOD mRNA was mainly expressed in the heart and slightly in kidney, but not in any other organs. The expression of Mn-SOD mRNA peaked at 6 h after the injection of IL-1α. The Mn-SOD protein content was most increased 12 h after injection. In the isolated heart model, the rats were pretreated with Il-1α 24 h earlier and their hearts were perfused by the Langendorff method. After 20 min of ischemia which was induced by a ligation of a coronary artery, reperfusion-induced arrhythmias were observed. There we no significant differences in the incidence of ventricular arrhythmias between the IL-1α pretreated and the untreated hearts. IL-1α pretreatment significantly reduced the mean duration of the ventricular arrhythmias and also delayed the onset of arrhythmias. The effect of IL-1α pretreatment was also investigated in a 30-min model of ischemia followed by a 3-min reperfusion in anesthetized rats. The infarct size expressed as as percentage of the area at risk was significantly reduced in the IL-1α pretreated hearts compared with the untreated hearts. The left ventricular systolic pressure increased significantly in rat hearts pretreated with IL-1α. Our results therefore showed that the pretreatment with IL-1α induced the overexpression of Mn-SOD mRNA in the rat hearts and also suggested that pretreatment with IL-1α 24 h before ischemia reduced the risk of ischemia-reperfusion injury.

Studies of arrhythmias by 24-hour polygraphic recordings: Relationship between atrioventricular block and sleep states

The relationship between AV conduction disturbances and sleep states was investigated using continuous polygraphic recordings for one full night. The results were as follows: (1) In the case of first-degree AV block, the phasic shortening of the PQ interval was observed during rapid eye movement (REM) sleep. (2) In the case of second-degree AV block (Wenckebach type), the conduction ratio increased transiently during REM sleep and significant relationships were observed between the number of nonconducted P waves and the mean heart rate in each sleep stage. (3) In the case of advanced AV block, complete AV block was observed less frequently during REM sleep and the period of falling asleep. Since the reproducibility of these results from night to night has not yet been investigated, additional evaluation is required.

Frequency characteristics of the heart rate variability produced by Cheyne-Stokes respiration during 24-hr ambulatory electrocardiographic monitoring

Spectral analysis of heart rates during 24-hr ambulatory electrocardiographic monitoring has been carried out to characterize the heart rate spectral components of Cheyne-Stokes respiration (CSR) by using fast Fourier transformation (FFT). Eight patients with congestive heart failure were selected for the study. FFT analyses have been performed for 614.4 sec. Out of the power spectrum, five parameters were extracted to characterize the CSR. The low peak frequencies in eight subjects were between 0.0179 Hz (56 sec) and 0.0081 Hz (123 sec). The algorithms used to detect CSR are the followings: (i) if the LFPA/ULFA ratios were above the absolute value of 1.0, and (ii) the LFPP/MLFP ratios were above the absolute values of 4.0, then the power spectrum is suggestive of CSR. We conclude that the automatic detection of CSR by heart rate spectral analysis during ambulatory ECG monitoring may afford a tool for the evaluation of the patients with congestive heart failure.

Effects of oral diltiazem on ventricular premature contractions.

The effects of oral diltiazem (90-180 mg/day for four weeks) on ventricular premature contractions (VPCs) were studied in 16 patients with frequent VPCs using 24-hour ambulatory ECG recordings. VPC frequency was evaluated as a function of underlying heart rate. Plots of VPC frequency vs. heart rate were made at 1-beat/min intervals for all heart rates recorded for at least five minutes during 24 hours. Patterns of correlation between VPC frequency and heart rate observed before diltiazem therapy included: 1) a relatively linear increase in VPCs with heart rate (positive correlation) in ten patients, 2) a linear decrease (negative correlation) in one patient, and 3) an increase at low heart rates and a decrease at high heart rates (bidirectional correlation) in five patients. Diltiazem significantly reduced the mean VPC frequency per 24 hours for patients with a positive correlation, but induced no significant change for patients with a negative or a bidirectional correlation. At the 65% level of VPC reduction, diltiazem was effective in eight of ten patients with a positive correlation but was not effective in the six patients with other correlations (p less than 0.01). These results suggest that an evaluation of VPC frequency as a function of heart rate predicts the response of VPCs to diltiazem.

Effects of amiloride on the mechanical, electrical and biochemical aspects of ischemia-reperfusion injury

Although many causal factors have been proposed for the ischemia-reperfusion injury, the exact mechanisms for interdependent derangements of mechanical, electrical and metabolic events remains unclear. For this purpose, the Langendorff-perfused rat hearts were subjected to regional brief ischemia followed by reperfusion to study the protective effects of amiloride, an inhibitor of Na+−H+ exchange. Amiloride (0.1 mM) attenuated the rise in tissue Na+ and Ca2+, both duration and incidence of arrhythmias (p<0.05 vs. control), sarcolemmal injury (assessed by Na−K ATPase) and lipid peroxidation (assessed by malonedialdehyde formation) during reperfusion. Treatment of hearts with monensin, a sodium inophore, reversed the protective effects of amiloride. Reduction in transsarcolemmal Na+ and pH gradients during ischemia exhibited protective effects similar to those seen with amiloride. These results suggest that cardiac dysfunction, sarcolemmal injury and triggered arrhythmias during ischemia-reperfusion are due to the occurrence of intracellular Ca2+ overload caused by the activation of Na+−H+ exchange and Na+−Ca2+ exchange systems in the myocardium.

Modulation of Na+-Ca2+ exchange in cardiac sarcolemmal vesicles by Ca2+ antagonists

SummaryThe purpose of this study was to examine the effect of three classes of Ca2+ antagonists, diltiazem, verapamil and nifedipine on Na+-Ca2+ exchange mechanism in the sarcolemmal vesicles isolated from canine heart. Na+-Ca2+ exchange and Ca2+ pump (ATP-dependent Ca2+ uptake) activities were assessed using the Millipore filtration technique. sarcolemmal vesicles used in this study are estimated to consist of several subpopulations wherein 23% are inside-out and 55% are right side-out sealed vesicles in orientation. The affect of each Ca2+ antagonist on the Na+-dependent Ca2+ uptake was studied in the total population of sarcolemmal vesicles, in which none of the agents depressed the initial rate of Ca2+ uptake until concentrations of 10 μM were incubated in the incubation medium. However, when sarcolemmal vesicles were preloaded with Ca2+ via ATP-dependent Ca2+ uptake, cellular Ca2+ influx was depressed only by verapamil (28%) at 1 μM in the efflux medium with 8 mM Na+. Furthermore, inhibition of Ca2+ efflux by verapamil was more pronounced in the presence of 16 mM Na+ in the efflux medium. The order of inhibition was; verapamil > diltiazem > nifedipine. These results indicate that same forms of Ca2+-antagonist drugs may affect the Na+-Ca2+ exchange mechanism in the cardiac sarcolemmal vesicles and therefore we suggest this site of action may contribute to their effects on the myocardium.

Role of the suprachiasmatic nuclei of the hypothalamus on diurnal rhythm in cardiac arrhythmias

SummaryAmbulatory EGG and EEG recordings were recorded under a 14/10-h light-dark illumination schedule using rats. The rats consisted of two groups: a suprachiasmatic (Sch) lesioned group (n=5) and a normal control group (n=5). Bilateral Sch nuclei were lesioned electrically (DC, 2.5 mA, 30 s for each) using a pair of platinum electrodes 0.3 mm in diameter. After recovery from surgery, recordings of ECGs (leads I, II, and III) and EEGs from the cortex and the left dorsal hippocampus were continued for 6 days. Diurnal periodicity in bradyarrhythmia (sinoatrial block, atrioventricular block) and heart rate was analyzed by the least square fit of 24-h cosines. Significant diurnal rhythm was observed in control rats, whereas Sch-lesioned rats showed no significant diurnal rhythm. The integrity of the Sch nuclei, therefore, is necessary for the generation and/or the expression of diurnal periodicity in bradyarrhythmia in rats.

Effect of dietary omega-3 eicosapentaenoic acid supplements on cholesteryl ester transfer from HDL in cholesterol-fed rabbits.

We investigated the reactivities of cholesteryl ester transfer protein (CETP) in Japanese white rabbits fed either a low-cholesterol diet containing 0.1% cholesterol (Control group) or a diet containing 0.1% cholesterol plus 17.5% omega-3 eicosapentaenoic acid (omega-3 20:5, EPA) of 4.5% (w/w) total lipid (EPA group) for 6 weeks. The plasma total and LDL cholesterol levels and aortic cholesterol content were all significantly higher in the EPA group than in the control group. The aortic cholesterol content significantly correlated with LDL cholesterol (r = 0.81). HDL cholesterol levels tended to be lower in the EPA group compared with control group, which was not statistically significant. The plasma VLDL cholesterol levels did not differ significantly between the groups. In addition, no significant differences were observed in the plasma CETP activity or lecithin:cholesterol acyltransferase (LCAT) activity between the groups. However, the cholesteryl ester (CE) mass transfer from fractionated HDL in the EPA group to excess VLDL and/or LDL as acceptors by purified CETP increased significantly compared with the control group, even if the acceptors were fractionated from either the EPA or the control group. Fatty acid analyses of CE showed that the omega-3 18:3, 20:4 or omega-3 20:5 fatty acid acyl groups in CE of HDL were significantly more transferred to apo B-containing lipoproteins compared with the 14:0,16:0, 18:0, 18:1 or 18:2 fatty acid acyl groups in CE of HDL during the incubation period. The amount of CE in HDL containing omega-3 18:3 and omega-3 20:5 fatty acid acyl groups was greater, while the amount of CE containing 18:2 fatty acid acyl groups was smaller in the EPA group than in the control group. These results show that although CETP itself did not change, the transfer of CE in HDL to apo B-containing lipoproteins by CETP increased in the rabbits fed a diet containing EPA as the HDL is modified by the diet, which may partly explain why atherogenicity was thus found to progress in the rabbits fed a cholesterol plus EPA diet.

Modulation of adrenergic receptors during regression of cardiac hypertrophy.

Objective To determine whether α1- or β-adrenergic receptors are altered during regression of cardiac hypertrophy produced by antihypertensive agents. Design and methods Cardiac hypertrophy was induced in rats by aortic banding. After 6 weeks banding the rats were treated with an angiotensin converting enzyme (ACE) inhibitor (enalapril), an α1-adrenergic antagonist (bunazosin) or a β-adrenergic antagonist (propranolol) for 6 weeks to induce regression. The numbers of α1- and β-adrenergic receptors, haemodynamics, tissue noradrenaline content and tissue ACE activity were measured. Results Regression of cardiac hypertrophy occurred after treatment of aortic banded rats with a high dose of enalapril, bunazosin or propranolol, and was accompanied by a reduction in systolic blood pressure. The number of α1- or β-adrenergic receptors was unchanged by propranolol treatment, but the number of α1-adrenergic receptors was increased in the hearts of rats treated with bunazosin. A low dose of enalapril (3 mg/kg body weight) caused regression of hypertrophy without a concomitant reduction in blood pressure, and decreased the number of α1-adrenergic receptors. The dissociation constants for α1- and β-adrenergic receptors were not different among the experimental groups, and the positive derivatives of left ventricular pressure was unaltered in rats treated with a low dose of enalapril but was reduced by the other drugs. Conclusion Of the three drugs tested, only the low dose of enalapril affected adrenergic receptors during regression of cardiac hypertrophy, causing a decrease in α1-adrenergic receptor number without a reduction in blood pressure. This effect may be explained by non-haemodynamic actions of the ACE inhibitor enalapril, probably by modulation of peripheral sympathetic activity.