Effect of protamine on the composition of blood lipoproteins at various periods of hypercholesterolemia in rabbits

Protamine has been studied for its effect on changes in the protein-lipid composition of main fractions of blood lipoproteins in different periods (1- and 7-months) of hypercholesterolemia in rabbits. Daily introduction of protamine to intact animals for 30 days did not change the studied parameters. Results of analysis of the protamine action against the background of the 1-month hypercholesterolemia testify to an increase of the protein and cholesterin concentration in apoB-containing lipoproteins. A moderate increase of the cholesterin concentration in the HDL composition has been observed, the protein concentration remaining at high level. The atherogenic diet for 7 month has induced in animals subjected to its dyslipoproteinemia and atherosclerosis of vessels. Under these conditions protamine has inhibited "proatherogenic" alterations in the composition of lipoproteins. The concentration of protein, cholesterin and triglycerides in the apoB-containing fractions decreased and the concentration of total cholesterin and triglycerides in the blood serum fell considerably.     

Preparation and investigation of 99m technetium-labeled low-density lipoproteins in rabbits with experimentally induced hypercholesterolemia

Low-density lipoproteins (LDL) were radiolabeled in atherosclerosis studies. The aim was to investigate the biodistribution and uptake of ^99mTc-labeled LDL by atherosclerotic plaques in experimentally induced hyperlipidemia. Rabbits were fed a diet containing 2% cholesterol for 60 days to develop hyperlipidemia and atheromatous aortic plaques. A combination of preparative and analytical ultracentrifugation was used to investigate human LDL aliquots, to prepare radioactive-labeled lipoproteins and in rabbits with induced hyperlipidemia. Preparative density gradient centrifugation was applied for the simultaneous isolation of the major lipoprotein density classes, which form discrete bands of lipoproteins in the preparative tubes. The cholesterol and protein levels in the lipoprotein fractions were determined. LDL was subsequently dialysed against physiological solution and sterilized and apolipoprotein fragments and aggregates were eliminated by passage through a 0.22-micron filter. LDL was radiolabeled with ^99mTc by using sodium dithionite as a reducing agent. Radiochemical purity and in vitro stability were controlled by paper chromatography in acetone. The labelling efficiency was 85–90% for human LDL. Two months after the start of cholesterol feeding, the total cholesterol in the blood serum had increased approximately 33-fold in comparison with the basal cholesterol content of hypercholesterolemic rabbits. Investigation of LDL was performed by Schlieren analysis after adjustment of the density of serum and underlayering by salt solution in a spinning ultracentrifugation capillary band-forming cell. Quantitative results were obtained by measuring the Schlieren areas between the sample curves and the reference baseline curve by means of computerized numerical and graphic techniques. In this manner we measured the concentrations of human LDL and analyzed rabbit LDL levels in induced hyperlipidemia. Gamma scintillation camera scanning of the rabbits was performed. Overnight fasted rabbits were injected in the marginal ear vein with ^99mTc-labeled human LDL (4–10 mCi, 0.5–1.5 mg protein). The initial scintigram showing a typical blood-pool scan, gradually changing with time to an image of specific organ uptake of radioactivity by the liver, kidneys and brain and in the bladder. Gamma camera in vivo scintigraphy on rabbits revealed visible signals corresponding to atherosclerotic plaques in the aorta and carotid arteries. Our results show that ^99mTc-LDL can be used to assess the organ distribution pattern of LDL in the rabbit, and to detect and localize areas of arterial atherosclerotic lesions.     

Toxic activity of the blood plasma in the early postresuscitation period

The time course of blood plasma toxicity was studied in dogs in the early postresuscitation period 10 minutes after clinical death because of acute hemorrhage. The duration of dying, the rate of the recovery of the main vitally important body functions were discovered to affect the rate of accumulation in the blood of substances with different molecular weights. There were 3 phases in blood plasma toxicity accumulation. The first phase is marked by the overflow of blood vessels with low-molecular substances, the second one by relative lessening of the content of low-molecular metabolites and an increase in the content of medium-molecular substances. The second phase is in agreement with the maximal rise of blood plasma toxicity. The third phase that develops by the 60th minute of the postresuscitation period is characterized by reduction of the peak activity of all substances under study and by overt toxicity with relative normalization of the main body functions.     

Effects of dietary flaxseed on vascular contractile function and atherosclerosis during prolonged hypercholesterolemia in rabbits

Dietary flaxseed has significant anti-atherogenic effects. However, the limits of this action and its effects on vascular contractile function are not known. We evaluated the effects of flaxseed supplementation on atherosclerosis and vascular function under prolonged hypercholesterolemic conditions in New Zealand White rabbits assigned to one of four groups for 6, 8, or 16 wk of feeding: regular diet (RG), 10% flaxseed-supplemented diet (FX), 0.5% cholesterol-supplemented diet (CH), and 0.5% cholesterol- and 10% flaxseed-supplemented diet (CF). Cholesterol feeding resulted in elevated plasma cholesterol levels and the development of atherosclerosis. The CF group had significantly less atherosclerotic lesions in the aorta and carotid arteries after 6 and 8 wk than the CH animals. However, the anti-atherogenic effect of flaxseed supplementation was completely attenuated by 16 wk. Maximal tension induced in aortic rings either by KCl or norepinephrine was not impaired by dietary cholesterol until 16 wk. This functional impairment was not prevented by including flaxseed in the high-cholesterol diet. Aortic rings from the cholesterol-fed rabbits exhibited an impaired relaxation response to acetylcholine at all time points examined. Including flaxseed in the high-cholesterol diet completely normalized the relaxation response at 6 and 8 wk and partially restored it at 16 wk. No significant changes in the relaxation response induced by sodium nitroprusside were observed in any of the groups. In summary, dietary flaxseed is a valuable strategy to limit cholesterol-induced atherogenesis as well as abnormalities in endothelial-dependent vasorelaxation. However, these beneficial effects were attenuated during prolonged hypercholesterolemic conditions.     

Blood lipoproteins in modeling of atherosclerosis and administration of protamine]

The influence of protamine on the HDL, LDL, VLDL species concentration variation as well as on their cholesterol and protein content in respect to total blood serum cholesterol in different periods (1 and 7 months) of hypercholesteremia in rabbits is investigated. Daily administration of protamine (10 mg/kg) during 30 days to intact animals did not change the investigated parameters. In the early period of hypercholesteremia protamine considerably increased the content of all lipoprotein fractions. Under the conditions of prolonged hypercholesteremia the LDL and VLDL concentrations decreased by 23 and 60 per cent, respectively; HDL increased by 101 per cent.     

Effects of hypercholesterolemia on the electrical and contractile properties of the blood vessel wall

The investigations were performed on the ring strips of rabbit aorta. The electrical activity of the vessels smooth muscle cells was registered by the "sucrose gap" method; the contractile activity of the strips was determined simultaneously. Experimental atherosclerosis was induced by keeping rabbits on a special diet enriched by cholesterol for 2 and 4 months. Strips were prepared with intact or mechanically removed endothelium. Hypercholesterolemia was shown to inhibit the reactivity of the vessel's wall to the weakening action of acetylcholine due to endothelial stimulation. The cause of these changes was the inhibition of the endothelial functional activity and inactivation of mechanisms by which endothelium influences smooth muscle cells.     

Influence of mevinolin on metabolism of low density lipoproteins in primary moderate hypercholesterolemia

Mevinolin, a competitive inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase, was used for treatment of 12 patients with moderate hypercholesterolemia, but not classical familial hypercholesterolemia. For most patients, measurements of turnover of low density lipoprotein-apolipoprotein B (LDL-apoB) were made on placebo and during treatment with two doses of mevinolin. LDL turnover was determined after injection of autologous 125I-labeled radioiodinated LDL. Compared to placebo, a low dose of mevinolin (10 mg, twice daily (BID] caused reductions of plasma total cholesterol and LDL-cholesterol averaging 15% and 20%, respectively; corresponding reductions on high doses of mevinolin (20 mg BID) were 22% and 31%, respectively. Triglyceride levels were unchanged by the drug. High density lipoprotein cholesterol levels rose significantly on the high dose, but not on the low dose. Neither dose produced a stastistically significant change in fractional catabolic rate (FCR) for LDL-apoB for the whole group, although several patients had increases in FCR on both doses. In contrast, both doses of mevinolin caused decreases in production rates of LDL-apoB. Thus, the fall in LDL levels in patients with moderate hypercholesterolemia can be explained more by a reduction in the input rate of LDL-apoB than by enhanced fractional removal of LDL from the circulation.     

Change in the rheologic properties of blood during the early postreanimation period]

Experiments were conducted on nembutal-anesthetized cats; a study was made of the dynamic viscosity of the blood, and also of an aggregation capacity of the blood formed elements for two hours of the postresuscitative period. There was established an increase in the viscosity both in the zone of a low (1.82 dynes/cm2) and high (10.94 dynes/cm2) sear stress. A definite role in increase of the blood viscosity is played by a rise in the hematocrite index and by enhanced aggregation capacity of the blood formed elements.     

Effect of hypercholesterolemia on myocardial function in New Zealand white rabbits

Although hypercholesterolemia is a well-known risk factor for atherosclerosis, little is known about the effect of hypercholesterolemia on cardiac contractile function. The objective of this study was to examine the effect of hypercholesterolemia on myocardial contractility. Fifteen New Zealand white rabbits were fed standard chow (control group) and another 15 were fed a cholesterolenriched diet (HC group) for 12 weeks. The contractile response of ventricular muscle strips was measured in various extracellular calcium concentrations and at different pacing rates. The whole-cell calcium current recording, and mRNA and protein levels of cellular calcium-handling proteins were also analyzed. With 2 mM Ca²⁺ and stimulation at 3 Hz, the contractile force of HC strips was less than that of the controls (3.63±0.20 vs. 4.61±0.50 mN, p<0.05). The time to peak tension was longer for HC strips (93.3±2.16 vs. 82.2±2.81 ms, p < 0.05). The peak L-type calcium inward current density was slightly higher in HC myocytes but did not reach statistical significance (–14.90±0.94 vs. –12.44±0.84 pA/pF, p=0.15). The mRNA level of sarcoplasmic reticulum Ca²⁺-ATPase (SERCA), normalized to GAPDH, was significantly lower in the HC than that in the control group (2.85±0.14 vs. 7.67±0.67, p<0.05), as was the ryanodine receptor (RyR; 0.42±0.06 vs. 0.71±0.13, p<0.05). The mRNA of the Na⁺/Ca²⁺ exchanger (NCX) was statistically higher in the HC group (0.90±0.12 vs. 0.48±0.05, p<0.05). Western blot experiments revealed that protein expression of SERCA in the HC strips decreased, but that of the NCX increased. The protein expression of the dihydropyridine receptor was similar between these two groups. We concluded that hypercholesterolemia results in suppression of the maximal contractile function and in a longer systolic contractile time course. These changes may partially be mediated through a decrease in SERCA and RyR but an increase in NCX expression.