Normocholesterolemic Rats Research Articles

The diurnal rhythms of cholesterol metabolism and plasma clearance of model chylomicrons: comparison of normal and genetically hypercholesterolemic rats (RICO)

Previous studies showed a slower clearance of cholesterol-labeled lymph chylomicrons in genetically hypercholesterolemic rats (RICO) compared with normocholesterolemic rats. In this study, we compared rates of lipolysis and remnant clearance in RICO versus control normocholesterolemic rats of the same strain (RAIF) or with control Wistar rats, by injecting chylomicron-like lipid emulsions labeled with 14C-triolein to trace lipolysis, and 3H-cholesteryl ester to trace remnant clearance. Our findings showed slower clearance of chylomicron remnants in RICO compared with control RAIF or with control Wistar rats. During the light period, the clearance of lipids from chylomicron-like lipid emulsions injected intravenously was significantly slower in RICO rats compared with normocholesterolemic control rats of the same strain, RAIF. Within the RICO group, clearance of emulsion triolein (TO) was faster during the dark period compared with the light period. In contrast, however, the clearance of the emulsion remnants traced by cholesteryl oleate (CO) was slower during the dark period. This behaviour was not found within the Wistar group, where the clearances of TO and CO were similar in the light and dark period. Hepatic clearance of chylomicron remnants is mediated primarily by the low density lipoprotein (LDL) receptor, the expression of which shows diurnal variation. In both Wistar and RICO rats, the expression of LDL receptors was highest during the dark period. The LDL receptors in hepatic microsomal membranes from RICO rats migrated faster on SDS polyacrylamide gel electrophoresis when compared with normal Wistar and the RAIF. However in hepatic plasma membranes the LDL receptors from RICO and Wistar rats appeared identical after immunoblotting. Furthermore the LDL receptors from RICO and Wistar rats responded similarly to treatment with neuraminidase. An alteration in post-translational processing of the LDL receptor could possibly account for the slower clearance of chylomicron remnants in the RICO.

Consumption of enzyme resistant starch and cholesterol metabolism in normo- and hypercholesterolemic rats

The influence of raw and retrograded high amylose corn starch on cholesterol metabolism in normo- as well as hypercholesterolemic rats was studied. No effect on cholesterol metabolism was observed in normocholesterolemic rats. On the contrary, in hypercholesterolemic animals plasma free and total cholesterol concentrations were significantly lower when enzyme resistant starch was fed. Liver cholesterol concentrations were significantly decreased in rats given the hypercholesterolemic diet supplemented with retrograded high amylose corn starch. Total fecal bile acid excretion and bile acid spectrum were not changed by enzyme resistant starch consumption. Plasma total cholesterol concentrations showed a significantly negative correlation with fecal coprostanol excretion in hypercholesterolemic rats. However, there is no evidence that coprostanol is the only factor responsible for the observed hypocholesterolemic effect of enzyme resistant starch in cholesterol fed rats.

Neutrophil enhancement of reperfusion-induced capillary fluid filtration associated with hypercholesterolemia.

Fluid filtration rate (Jv/S) from individual mesenteric capillaries in normocholesterolemic and hypercholesterolemic rats was measured before and after 30 min each of ischemia and reperfusion (I/R). The median I/R-induced increase in Jv/S (I/R vs. baseline) was 44% in normocholesterolemic rats (n = 11) and 97% in hypercholesterolemic rats (n = 11). A positive correlation slope of 0.20% per mg/dl resulted when the percent Jv/S increase vs. plasma cholesterol concentration (P = 0.02) was plotted, demonstrating that hypercholesterolemia enhances the capillary response to I/R. Because microvascular pressure did not change significantly after I/R in either group of rats, the increments in Jv/S likely reflect increased capillary permeability. In hypercholesterolemic rats rendered neutropenic with antineutrophil serum, I/R did not elicit a significant increase in Jv/S, suggesting that activated neutrophils mediate the exaggerated endothelial barrier dysfunction associated with hypercholesterolemia.

Lymphatic transport of cholesterol in normocholesterolemic rats treated with pravastatin, an inhibitor of HMG-CoA reductase

Lymphatic absorption and transport of cholesterol and triacylglycerols were examined in rats treated with pravastatin, an inhibitor of 3-hydroxy-3-methyglutaryl-CoA (HMG-CoA) reductase. Pravastatin-treatment for 1, 7 and 28 days did not affect the recovery of cholesterol and triacylglycerols during 24 h after the lipid administration: the recovery was 52–59% and 82–93% for cholesterol and triacylglycerols, respectively. Rats treated with pravastatin for 28 days had a higher lymphatic recovery of the lipids during 3–6 h after the lipid administration than did control rats. Pravastatin treatment did not affect the ratio of phospholipid to cholesterol in the gut mucosa, the fatty acid composition of the lymph and mucosal lipids. We concluded that an inhibitor of HMG-CoA reductase would exert no adverse effect on absorption of fat-soluble nutrients by gut.

Effect of unprocessed and baked inulin on lipid metabolism in normo- and hypercholesterolemic rats

Male Wistar rats were fed a bread diet containing either corn starch or 6% raw or 6% baked inulin. Furthermore, the diets were either cholesterol-free or contained 1% cholesterol and 0.1% cholic acid. Adding unprocessed or baked inulin to the cholesterol-free diets resulted in significantly lower plasma cholesterol concentrations as well as in decreased liver cholesterol concentrations, while higher daily fecal excretions of bile acids were found. The normocholesterolemic rats showed a tendency to elevated fecal excretion of neutral steroids when inulin was fed, in spite of some decrease in the coprostanol excretion. In these groups there was a significantly inverse linear relationship between liver cholesterol concentrations and daily fecal bile acid output. The results suggest that the cholesterol lowering effect of inulin in normocholesterolemic rats may be due to higher fecal steroid excretion. Moreover, these rats tended to show higher HDL LDL cholesterol ratios when they received one of the inulin containing diets. In the hypercholesterolemic groups no significant effect from inulin intake on the plasma and liver cholesterol concentrations was found. In these rats, fecal cholesterol excretion was significantly increased by 40% when baked or unprocessed inulin was added to the diet, but fecal coprostanol excretion was significantly decreased by 95%. As a consequence, consumption of inulin did not change total neutral steroid excretion in the hypercholesterolemic groups. Moreover, fecal bile acid output was not significantly affected by dietary treatment. The fact that inulin intake did not change total steroid output might explain why plasma cholesterol was not reduced in the hypercholesterolemic rats receiving inulin, in contrast to the results obtained with the normocholesterolemic rats. Also no effect from inulin consumption was found on the HDL LDL cholesterol ratio in the hypercholesterolemic animals. Including inulin in bread did not alter its effect on lipid metabolism, neither in normo- nor in hypercholesterolemic rats.

Reduced smooth muscle cell regeneration in Yoshida (YOS) spontaneously hypercholesterolemic rats

Hypercholesterolemia is a predisposing factor for atherosclerosis. We studied the response to damage of vascular smooth muscle cells (SMC) from normocholesterolemic Brown Norway (BN) and from spontaneously hypercholesterolemic Yoshida (YOS) rats (16–24 month old). The regrowth rate of SMC from BN and YOS rats after freezeinduced damage was similar in the presence of fetal calf serum and of serum derived from normocholesterolemic rats, while it was reduced in the presence of serum from hypercholesterolemic rats. Freeze-injury of the abdominal aorta was followed by reduced neointima formation in YOS rats, as compared to BN rats, confirming the impaired response of vascular cells from hypercholesterolemic rats to injury. This defect may be due either to lipids or to unknown factors present in the hyperlypidemic serum.

The role of diet cholesterol changes on EEG.

Thirty three male albino rats, weight between 200 and 220 g were used in this experiment. Control animals consisting of 11 rats were fed with a normal lab diet for a period of 14 weeks and the others (22) were fed with a diet containing 1% cholesterol for the same period. At the end of the experimental period, plasma cholesterol level (Mean +/- SD) was 141.29 +/- 34.5 mg/dl in the cholesterol group and 70.66 +/- 10 mg/dl in the control group. Eleven of the rats from the cholesterol group were transferred to the normal diet for 14 weeks (normocholesterolemic group). Spectral analysis of EEG records from parietal lobes of animals showed that there was an obvious depression in the brain waves of hypercholesterolemic rats whereas no depression in normocholesterolemic rats.

Effects of oat-bran concentrate on rat serum lipids and liver fat infiltration

Oat bran concentrated with respect to both soluble and insoluble dietary fibre was fed to adult rats, and its effects on serum cholesterol and liver fat infiltration were studied. The feeds contained 15, 30 or 45 g beta-glucan/kg, except in control groups where the fibre of the feed was cellulose. Half the experimental groups received the feeds with an addition of 10 g cholesterol and 2 g cholic acid/kg in order to create an hypercholesterolaemic condition. In normocholesterolaemic rats the concentrated oat bran did not cause any significant changes in serum cholesterol concentrations, but reduced liver weight compared with control group rats fed on the cellulose-containing diet. In hypercholesterolaemic rats the concentrated oat bran reduced serum total cholesterol and increased high-density-lipoprotein cholesterol concentration. The effect was seen already at the 15 g/kg concentration of beta-glucan, the higher doses tested did not significantly improve the effect. Infiltration of fat into liver cells, especially in the periportal areas, was observed only in hypercholesterolaemic animals. The fat infiltration was accentuated in proportion to the amount of oat-bran concentrate fed.

The effects of tannic acid on serum and liver lipids of RAIF and RICO rats fed on high fat diet

1. 1. Male genetically hypercholesterolemic (RICO) and normocholesterolemic (RAIF) rats were divided into four groups and each group was fed one of the following diets for 10 weeks: a standard diet (STD); STD supplemented with tannic acid (STD + T); a high fat diet (HFD) and a high fat diet with tannic acid (HFD + T). 2. 2. The RAIF and RICO rats when fed on HFD + T showed beneficial changes in serum lipids compared to the rats fed HFD. The lipid parameters between the groups fed STD and STD + T in both strains were unchanged. 3. 3. The possible modes of hypocholesterolemic action of tannic acid in these rats is discussed.

Metabolism of intestinal triglyceride-rich lipoproteins in the genetically hypercholesterolemic rat (RICO)

Experiments were carried out to compare the catabolism of intestinal lipoproteins between genetically hypercholesterolemic (RICO) and normocholesterolemic (SW) rats. Kinetics of plasma cholesterol ester were studied after injection of cholesterol-labeled chylomicrons or VLDL. The chylomicron clearance is reduced in the RICO rat (rate constant, K = 7.2 ± 0.1 h −1 vs. 10.7 ± 0.1 h −1 in SW rat), while a much more minor alteration was observed in the catabolism of lymph VLDL ( K = 4.3 ± 0.6 h −1 in the RICO rat vs. 5.1 ± 0.4 h −1). The injection of chylomicrons from SW rats to RICO rats and from RICO rats to SW rats showed that the fall in the rate of catabolism of chylomicrons in RICO rats was not secondary to an increase in the production rate, but was related to the lipoprotein particle itself without any alteration of the catabolic system. The reduction in the rate of catabolism of chylomicrons in the RICO rat could be related to a change in their apolipoprotein composition (increase in the proportion of apolipoprotein E = 12 ± 2% vs. 3 ± 1% in the SW rat).

Cytomegalovirus infection induces vascular injury in the rat

The role of cytomegalovirus (CMV) in the early development of atherosclerosis was studied in a rat model. Arterial samples derived from virus-infected normo- and hypercholesterolaemic animals were investigated by light microscopy at 1, 4, 8 and 16 weeks post infection. Early atherogenic lesions comparable to those seen in non-infected hypercholesterolaemic rats were found in CMV-infected normocholesterolaemic and hypercholesterolaemic animals, starting at 1 week post infection. The changes consisted of minimal endothelial cell damage, as shown by the en face technique, and a more than 10-fold increase in the number of leukocytes adhering to the aortic intima. The increased adhesion of leukocytes was observed in infected normocholesterolaemic rats but only in the non-infected rats which were hypercholesterolaemic. The infection of hypercholesterolaemic rats did not enhance this effect although it resulted in increased migration of the leukocytes into the subendothelial space. CMV infection of normocholesterolaemic rats induced lipid accumulation in the endothelium. In these animals approximately 1% of the endothelial cells contained lipid at 1 week post infection. In the non-infected hypercholesterol-fed animals 10% of the cells contained lipid. CMV infection in these rats induced an extra increase of the lipid-containing endothelial area. The changes in the CMV infected animals largely corresponded with the intimal injury observed in the hypercholesterolaemic rats. These results support the hypothesis that CMV may be one of the factors involved in atherogenesis.

Turnover of [14C] sucrose HDL and uptake by organs in the normal or genetically hypercholesterolemic (RICO) rat using a constant infusion method

The turnover and tissular uptake of HDL (d 1.095-1.21) have been compared in normocholesterolemic or genetically hypercholesterolemic rats by a constant infusion method of [14C] sucrose labelled HDL for 8 h. The HDL clearance rate was not significantly smaller in the RICO than in the normocholesterolemic animal (320 +/- 22 microliters.h-1 versus 366 +/- 24 microliters.h-1 per 100 g of rat). It was the same case for the fractional catabolic rate, respectively equal to 7.8 and 9.4 +/- 0.6%.h-1. For both strains, liver and skeletal muscle were the main catabolic sites for HDL. The HDL uptake rates in intestine or kidney were 3-4-fold smaller than those in the liver. In the RICO rat, intestine, testis and adrenals showed a lesser HDL uptake capacity (expressed per g of organ) than the normocholesterolemic rat.

Identification of macrophages in intimal thickening of rat carotid arteries by cytochemical localization of purine nucleoside phosphorylase.

Complete desquamation of the endothelium of the rat carotid artery by balloon catheter stripping resulted within 2 weeks in the formation of a large intimal thickening. After an enzyme cytochemical technique was applied to localize cytosolic purine nucleoside phosphorylase (PNP), light microscopical evaluation indicated that this intimal thickening in normocholesterolemic rats was composed of 5.8% to 11.8% (mean 8.8%) PNP-positive cells. At the electron microscopic level, all these PNP-positive cells were identified as macrophages by the absence of a basement membrane and plasmalemmal vesicles and by the occurrence of specific intracytoplasmic granules. The nearly nonreactive intimal cells were classified as modified smooth muscle cells. Additional evidence of the macrophage nature of the PNP-stained intimal cells was obtained by differential immunogold labeling of these cells with a monoclonal antibody against rat macrophages. Moreover, in hypercholesterolemic rats, only the cells stained for PNP transformed into foam cells (between 8.5% and 11.4% of all nucleated intimal cells; mean 9.6%). This study shows that PNP cytochemistry discriminates macrophages from modified smooth muscle cells in the rat carotid intimal thickening. It further suggests that the intimal thickening in normocholesterolemic rats originates not only from migration and proliferation of smooth muscle cells, but also from a considerable number of leukocyte-derived macrophages. Whether the latter cells are actively involved in the establishment of the intimal thickening as has been suggested in dietary hypercholesterolemia, remains to be verified.

Evaluation of the hypolipidemic activity of xenalipin in experimental animals

Xenalipin (4'-trifluoromethyl-2-biphenylcarboxylic acid) is a chemically novel compound which has been found to be an effective hypolipidemic agent in two animal species. Significant reductions in serum cholesterol and triglycerides were observed in cholesterol-cholic acid-fed rats following oral doses of 10–30 mg/kg/day. Xenalipin was considerably more potent than clofibrate, nicotinic acid, and cholestyramine in the same model. Lipoprotein analysis showed that xenalipin reduced cholesterol and protein content in very low density lipoprotein (VLDL), intermediate density lipoprotein (IDL), and low density lipoprotein (LDL). Triglycerides were reduced in VLDL and IDL. Xenalipin was also effective in reducing serum cholesterol and triglyceride concentrations in normocholesterolemic rats. In diet-induced hypercholesterolemic African green monkeys, xenalipin at oral doses of 15–60 mg/kg b.i.d. reduced serum LDL-cholesterol concentrations. These results suggest that xenalipin has a profile of activity which would be beneficial in therapy for hyperlipidemia.

The use of three esterase kits to measure plasma cholesterol concentration in the rat and three other species

Three enzymatic methods for the determination of cholesterol and its esters were applied to plasma samples obtained from normocholesterolaemic rats, dogs, rabbits and man. A major difference was observed for rat plasma values, where the results obtained with a microbial esterase for the hydrolysis of cholesteryl esters were much lower than those obtained with mammalian esterases. This difference was greater for rats than for the other three species studied and is probably related to the plasma distribution of lipoproteins in the rat.

コレステロール食少量負荷ラットの脂質代謝に及ぼすγ-オリザノールの影響

コレステロール食少量負荷ラットの脂質代謝に及ぼすγ-オリザノールの影響

Comparative evaluation of some pharmacological properties and side effects of d-glucitol hexanicotinate (sorbinicate) and nicotinic acid correlated with the plasma concentration of nicotinic acid

In rabbits kept on a diet containing 1 g/day cholesterol for 12 weeks, the nicotinic acid derivative sorbinicate displayed greater hypolipemic and antiatherogenic activity than an equidose of plain nicotinic acid at much lower and more constant plasma nicotinic acid levels. In normocholesterolemic rats, nicotinic acid given at a level of 300 mg/kg per dose for 3 weeks induced plasma FFA and triglyceride rebound and triglyceride accumulation in the liver and possibly in the heart (all parameters determined 24 h after the last dosing), whereas an equidose of sorbinicate was free from these effects, potentially the two most dangerous side effects of nicotinic acid. By modulating the bioavailability of nicotinic acid, sorbinicate maintains and in some cases enhances the pharmacological activity of the acid, avoiding at least some of its major side effects.

Effect of diosgenin on lipid metabolism in rats.

The purpose of this study was to determine whether diosgenin suppresses cholesterol absorption in rats, and to examine relevant changes in cholesterol and bile acid metabolism. Diosgenin fed with the diet for 1 week inhibited cholesterol absorption as determined by the serum isotope ratio technique, as well as by measuring in the feces the amount of unabsorbed radioactivity from orally administered [3H]cholesterol. In addition, diosgenin suppressed the serum and liver uptake of radioactivity from co-administered [3H]cholesterol as well as the accumulation of liver cholesterol in the cholesterol-fed rat; diosgenin was substantially more active than cholestyramine or beta-sitosterol. In vitro, diosgenin had no effect on the activity of rat pancreatic esterase. Diosgenin decreased the elevated cholesterol in serum LDL and elevated cholesterol in the HDL fraction of cholesterol-fed rats; diosgenin had no effect on serum cholesterol in normocholesterolemic rats. In contrast to cholestyramine, diosgenin markedly increased neutral sterol excretion without altering bile acid excretion; in vitro, diosgenin had no effect on bile acid binding. Diosgenin treatment increased hepatic and intestinal cholesterol synthesis as well as the activity of hepatic HMG CoA reductase. This was accompanied by increased biliary concentration of cholesterol, but not of bile acids. Diosgenin had no effect on cholesterol synthesis when added to normal rat liver homogenates. It was concluded that diosgenin interferes with the absorption of cholesterol of both exogenous and endogenous origin; such interference is accompanied by derepressed, i.e., increased, rates of hepatic and intestinal cholesterol synthesis. The increased unabsorbed cholesterol together with enhanced secretion of cholesterol into bile resulted in increased excretion of neutral sterols without affecting the biliary and fecal excretion of bile acids.

Arylsulfonate esters of fatty alcohols as hypocholesterolemic agents. I: Oleyl and linoleyl p-toluenesulfonates.

Cholesterol levels of both plasma and liver of cholesterol-fed rats are lowered by feeding linoleyl and oleyl esters of p-toluenesulfonic acid. A structural specificity exists, since the effect is not observed with similar amounts of sodium p-toluenesulfonate nor with linoleyl methanesulfonate. Effective levels of these arylsulfonates do not alter liver to body weight ratios and maturation, nor reduce plasma cholesterol in normocholesterolemic rats.

Lipoprotein regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity in rat liver cell cultures.

A primary cell culture technique was used to study the effects of lipoproteins on rat hepatocyte 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity. In this system, lipoproteins prepared from normocholesterolemic rat and human plasma, including low density lipoproteins, did not inhibit hepatocyte HMG-CoA reductase activity whereas very low density lipoproteins and high density lipoproteins isolated from the same sources were stimulatory. A lipoprotein was isolated from the plasma of cholesterol-fed rats that did inhibit hepatocyte HMG-CoA reductase activity. An extensive chemical characterization of the inhibitory lipoprotein revealed that it was mainly d less than 1.019 g/ml and had beta mobility on lipoprotein electrophoresis. The lipoprotein was compared to the comparable density fraction in the normocholesterolemic rat plasma and there was no size difference appreciable by negatively stained electron micrographs. However, two important differences in chemical composition were evident: in the inhibitory lipoproteins the per cent of total apoprotein which was in the region of Mr = 35,000 was increased 1.5- to 2-fold, and there was a marked increase in cholesterol ester content. These chemical characteristics may be required for lipoproteins to regulate hepatocyte cholesterol synthesis. Primary cell culture of rat hepatocytes appears to be a useful system in which to study cholesterol metabolism in the liver.