Cholesterol feeding alters the metabolism of thoracic-duct lymph lipoprotein cholesterol in rabbits but not in rats.
ABSTRACT: 1. Labelled thoracic-duct lymph was collected from rats and rabbits after test meals containing [(14)C]cholesterol and [2-(3)H]glyceryl trioleate. 2. The metabolism of labelled cholesterol and triglyceride was studied in normally fed and cholesterol-fed rats and rabbits injected with radioactive lymph from the same species. 3. In normally fed animals of both species, 10min after intravenous administration, about 80% of lymph cholesteryl ester but only about 10% of triglyceride was recovered in the liver after clearance from the plasma. This distribution is consistent with participation of ;remnant' particles in the metabolism of dietary lymph particles. 4. The metabolism of cleared lymph lipoprotein constituents was unchanged in cholesterol-fed rats, but the recovery of cholesteryl ester in the livers of the cholesterol-fed rabbits was decreased to 30% of the cleared dose. 5. The low recovery in cholesterol-fed rabbits was accounted for mainly by increased hydrolysis of cholesteryl ester. 6. It is proposed that differences between rats and rabbits in metabolism of dietary cholesterol might be partly due to the observed enhancement of hydrolysis of lymph lipoprotein cholesteryl ester in rabbits.
Project description:1. The appearance of exogenous cholesterol in free cholesterol and ester cholesterol of plasma chylomicra, very-low-density (VLD), low-density (LD) and high-density (HD) lipoproteins was studied in unanaesthetized rabbits after ingestion of a meal containing 5% fat and 0.08% [(3)H]cholesterol. 2. The specific radioactivity of ester cholesterol of VLD lipoproteins reached the highest value of any lipoprotein fraction and for each lipoprotein it increased at a faster rate and reached a higher maximum than that of free cholesterol; the maximum in VLD lipoproteins occurred later than in chylomicra. 3. The pattern of appearance of exogenous cholesterol in chylomicra and VLD lipoproteins of plasma was similar to the pattern previously observed in lymph. The specific radioactivity of ester cholesterol in plasma VLD lipoproteins was higher than that in chylomicra in spite of a larger pool size and dilution of cholesteryl esters from VLD lipoproteins produced by the liver. These results support the concept that during absorption the major portion of exogenous cholesterol is transported in VLD lipoproteins as ester cholesterol. 4. The specific radioactivity of ester cholesterol of chylomicra and VLD lipoproteins increased at a faster rate than that of LD and HD lipoproteins. However, the rate of increase and the absolute values of the specific radioactivity in LD and HD lipoproteins were identical. Since cholesteryl esters are thought not to exchange between lipoproteins, this observation supports the hypothesis that a result of VLD lipoprotein and chylomicron metabolism is the formation of LD and HD lipoproteins. 5. Results in vivo showed that the free cholesterol of individual plasma lipoproteins does not equilibrate within a period of 24h.
Project description:1. Ezetimibe potently inhibits the transport of cholesterol across the intestinal wall, thereby reducing plasma cholesterol in preclinical animal models of hypercholesterolemia. The effect of ezetimibe on known absorptive processes was determined in the present studies. 2. Experiments were conducted in the hamster and/or rat to determine whether ezetimibe would affect the absorption of molecules other than free cholesterol, namely cholesteryl ester, triglyceride, ethinylestradiol, progesterone, vitamins A and D, and taurocholic acid. In addition, to determine whether exocrine pancreatic function is involved in the mechanism of action of ezetimibe, a biliary anastomosis model, which eliminates exocrine pancreatic function from the intestine while maintaining bile flow, was established in the rat. 3. Ezetimibe reduced plasma cholesterol and hepatic cholesterol accumulation in cholesterol-fed hamsters with an ED(50) of 0.04 mg kg(-1). Utilizing cholesteryl esters labelled on either the cholesterol or the fatty acid moiety, we demonstrated that ezetimibe did not affect cholesteryl ester hydrolysis and the absorption of fatty acid thus generated in both hamsters and rats. The free cholesterol from this hydrolysis, however, was not absorbed (92 - 96% inhibition) in the presence of ezetimibe. Eliminating pancreatic function in rats abolished hydrolysis of cholesteryl esters, but did not affect the ability of ezetimibe to block absorption of free cholesterol (-94%). Ezetimibe did not affect the absorption of triglyceride, ethinylestradiol, progesterone, vitamins A and D, and taurocholic acid in rats. 4. Ezetimibe is a potent inhibitor of intestinal free cholesterol absorption that does not require exocrine pancreatic function for activity. Ezetimibe does not affect the absorption of triglyceride as a pancreatic lipase inhibitor (Orlistat) would, nor does it affect the absorption of vitamin A, D or taurocholate, as a bile acid sequestrant (cholestyramine) would.
Project description:The effects of atorvastatin (3 mg kg(-1)) and simvastatin (3 mg kg(-1)) on hepatic enzyme activities involved in very low density lipoprotein metabolism were studied in coconut oil/cholesterol fed rabbits. Plasma cholesterol and triglyceride levels increased 19 and 4 fold, respectively, after 7 weeks of feeding. Treatment with statins during the last 4 weeks of feeding abolished the progression of hypercholesterolaemia and reduced plasma triglyceride levels. 3-Hydroxy-3-methyl-glutaryl Coenzyme A reductase, acylcoenzyme A:cholesterol acyltransferase, phosphatidate phosphohydrolase and diacylglycerol acyltransferase activities were not affected by drug treatment. Accordingly, hepatic free cholesterol, cholesteryl ester and triglyceride content were not modified. Simvastatin treatment caused an increase (72%) in lipoprotein lipase activity without affecting hepatic lipase activity. Atorvastatin caused a reduction in hepatic phospholipid content and a compensatory increase in CTP:phosphocholine cytidylyl transferase activity. The results presented in this study suggest that, besides the inhibitory effect on 3-hydroxy-3-methyl-glutaryl Coenzyme A reductase, simvastatin and atorvastatin may have additional effects that contribute to their triglyceride-lowering ability.
Project description:Proteins from different fish species and different raw materials such as fish fillets and by-products have shown promising cardioprotective effects in rodents and humans, including effects on cholesterol metabolism. Blue whiting is used mainly to produce fish meal for the feed industry and during this production, a water-soluble protein fraction, containing small peptides that are easily absorbed and may hold bioactive properties, is isolated. The effects of water-soluble fish protein on cholesterol metabolism were investigated in twelve male obese Zucker fa/fa rats. Rats were fed diets with water-soluble protein from blue whiting (BWW) as 1/3 of the total protein and the remaining 2/3 as casein (BWW group) or with casein as the sole protein source (control group). After 5 weeks intervention, the BWW group had lower serum total, high-density lipoprotein (HDL), and low-density lipoprotein (LDL) cholesterol concentrations and lower cholesteryl ester concentration compared to controls. Hepatic concentrations of cholesterol, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, and LDL receptors were also lower in the BWW group. The groups had a similar concentration of serum total bile acids and similar fecal excretions of cholesterol and bile acids. To conclude, the BWW diet led to lower concentrations of serum and liver cholesterol in obese Zucker fa/fa rats, probably due to lower hepatic cholesterol synthesis.
Project description:Acyl-CoA:cholesterol O-acyl transferase 2 (ACAT2) promotes cholesterol absorption by the intestine and the secretion of cholesteryl ester-enriched very low density lipoproteins by the liver. Paradoxically, mice lacking ACAT2 also exhibit mild hypertriglyceridemia. The present study addresses the unexpected role of ACAT2 in regulation of hepatic triglyceride (TG) metabolism. Mouse models of either complete genetic deficiency or pharmacological inhibition of ACAT2 were fed low fat diets containing various amounts of cholesterol to induce hepatic steatosis. Mice genetically lacking ACAT2 in both the intestine and the liver were dramatically protected against hepatic neutral lipid (TG and cholesteryl ester) accumulation, with the greatest differences occurring in situations where dietary cholesterol was elevated. Further studies demonstrated that liver-specific depletion of ACAT2 with antisense oligonucleotides prevents dietary cholesterol-associated hepatic steatosis both in an inbred mouse model of non-alcoholic fatty liver disease (SJL/J) and in a humanized hyperlipidemic mouse model (LDLr(-/-), apoB(100/100)). All mouse models of diminished ACAT2 function showed lowered hepatic triglyceride concentrations and higher plasma triglycerides secondary to increased hepatic secretion of TG into nascent very low density lipoproteins. This work demonstrates that inhibition of hepatic ACAT2 can prevent dietary cholesterol-driven hepatic steatosis in mice. These data provide the first evidence to suggest that ACAT2-specific inhibitors may hold unexpected therapeutic potential to treat both atherosclerosis and non-alcoholic fatty liver disease.
Project description:OBJECTIVE: Scavenger receptor-class B type I (SR-BI), the receptor for HDL-cholesterol, plays a key role in HDL metabolism, whole body cholesterol homeostasis, and reverse cholesterol transport. We investigated the in vivo impact of hepatic SR-BI inhibition on lipoprotein metabolism and the development of atherosclerosis employing RNA interference. METHODS: Small hairpin RNA plasmid specific for rabbit SR-BI was complexed with galactosylated poly-l-lysine, allowing an organ-selective, receptor-mediated gene transfer. Rabbits were fed a cholesterol-rich diet, and were injected with plasmid-complexes once a week. RESULTS: After 2 weeks of treatment hepatic SR-BI mRNA levels were reduced by 80% accompanied by reduced SR-BI protein levels and a modulation of the lipoprotein profile. Rabbits treated with SR-BI-specific plasmid-complexes displayed higher cholesteryl ester transfer from HDL to apoB-containing lipoproteins, lower HDL-cholesterol, and higher VLDL-cholesterol levels, when compared to controls. In a long-term study, this gene therapeutic intervention led to a similar modulation of the lipoprotein profile, to lower total cholesterol levels, and most importantly to a 50% reduction of the relative atherosclerotic lesion area. CONCLUSION: Our results are another indication that the role of SR-BI in lipoprotein metabolism and atherogenesis in rabbits--a CETP-expressing animal model displaying a manlike lipoprotein profile may be different from the one found in rodents.
Project description:To study potential effects of hepatic cholesterol concentration on secretion of very-low-density lipoprotein (VLDL) by the liver, male rats were fed on unsupplemented chow, chow with lovastatin (0.1%), or chow with lovastatin (0.1%) and cholesterol (0.1%) for 1 week. Livers were isolated from these animals and perfused in vitro, with a medium containing [2-14C]acetate, bovine serum albumin and glucose in Krebs-Henseleit buffer, and with an oleate-albumin complex. With lovastatin feeding, the hepatic concentrations of cholesteryl esters and triacylglycerols before perfusion were decreased, although free cholesterol was unchanged. However, hepatic secretion of all the VLDL lipids was decreased dramatically by treatment with lovastatin. Although total secretion of VLDL triacylglycerol, phospholipid, cholesterol and cholesteryl esters was decreased, the decrease in triacylglycerol was greater than that in free cholesterol or cholesteryl esters, resulting in secretion of a VLDL particle enriched in sterols relative to triacylglycerol. In separate studies, the uptake of VLDL by livers from control animals or animals treated with lovastatin was measured. Uptake of VLDL was estimated by disappearance of VLDL labelled with [1-14C]oleate in the triacylglycerol moiety, and was observed to be similar in both groups. During perfusion, triacylglycerol accumulated to a greater extent in livers from lovastatin-fed rats than in control animals. The depressed output of VLDL triacylglycerols and the increase in triacylglycerol in the livers from lovastatin-treated animals was indicative of a limitation in the rate of VLDL secretion. Addition of cholesterol (either free cholesterol or human low-density lipoprotein) to the medium perfusing livers from lovastatin-fed rats, or addition of cholesterol to the diet of lovastatin-fed rats, increased the hepatic concentration of cholesteryl esters and the output of VLDL lipids. The concentration of cholesteryl esters in the liver was correlated with the secretion of VLDL by the liver. These data suggest that cholesterol is an obligate component of the VLDL required for its secretion. It is additionally suggested that cholesteryl esters are in rapid equilibrium with a small pool of free cholesterol which comprises a putative metabolic pool available and necessary for the formation and secretion of the VLDL. Furthermore, the specific radioactivity (d.p.m./mumol) of the secreted VLDL free cholesterol was much greater than that of hepatic free cholesterol, suggesting that the putative hepatic metabolic pool is only a minor fraction of total hepatic free cholesterol.
Project description:Plasma lipoproteins, plasma activities of lecithin:cholesterol acyltransferase (LCAT), phospholipid transfer protein (PLTP), cholesteryl ester transfer protein (CETP) and post-heparin lipases were measured before and after cholesterol challenge in two inbred strains of rabbits with either a high (hyper-responders) or a low (hyporesponders) response of plasma cholesterol to dietary cholesterol. The purpose of this study was to provide clues about the mechanisms underlying the effect of dietary cholesterol on lipoprotein levels and composition, and particularly those underlying the strain difference of this effect. Cholesterol feeding (0.15 g of cholesterol/100 g of diet) caused increased plasma total cholesterol concentrations and an increased ratio of cholesteryl esters:triacylglycerol in all lipoprotein particles in both strains; these effects were significantly greater in hyper- than hypo-responsive rabbits. Feeding on the high-cholesterol diet lowered plasma triacylglycerols in hyper-responders, but caused increased plasma triacylglycerol levels in hyporesponders. This was accompanied by significantly greater increases in the activities of hepatic triacylglycerol lipase and lipoprotein lipase in hyper- than in hypo-responders. Both strains showed a dietary-cholesterol-induced rise in plasma CETP as well as in PLTP activity. The increase in PLTP activity was greater in the hyper-responders, but that of CETP was less. There was no effect of dietary cholesterol on LCAT activity. It is hypothesized that the lipases are involved in the removal of cholesterol-rich lipoproteins.
Project description:Dysregulated free cholesterol (FC) metabolism has been implicated in nearly all stages of atherosclerosis, the underlying cause of most cardiovascular disease. According to a widely cited model, the burden of macrophage FC in the arterial wall is relieved by transhepatic reverse cholesterol transport (RCT), which comprises three successive steps: (1) macrophage FC efflux to high-density lipoprotein (HDL) and/or its major protein, apolipoprotein AI; (2) FC esterification by lecithin:cholesterol acyltransferase (LCAT); and (3) HDL-cholesteryl ester (CE) uptake via the hepatic HDL-receptor, scavenger receptor class B type 1 (SR-B1). Recent studies have challenged the validity of this model, most notably the role of LCAT, which appears to be of minor importance. In mice, most macrophage-derived FC is rapidly cleared from plasma (t1/2 < 5 min) without esterification by hepatic uptake; the remainder is taken up by multiple tissue and cell types, especially erythrocytes. Further, some FC is cleared by the nonhepatic transintestinal pathway. Lastly, FC movement among lipid surfaces is reversible, so that a higher-than-normal level of HDL-FC bioavailability-defined by high plasma HDL levels concurrent with a high mol% HDL-FC-leads to the transfer of excess FC to cells in vivo. SR-B1-/- mice provide an animal model to study the mechanistic consequences of high HDL-FC bioavailability that provokes atherosclerosis and other metabolic abnormalities. Future efforts should aim to reduce HDL-FC bioavailability, thereby reducing FC accretion by tissues and the attendant atherosclerosis.
Project description:The hypothesis tested in this study was that cholesterol esterification by ACAT2 would increase cholesterol absorption efficiency by providing cholesteryl ester (CE) for incorporation into chylomicrons. The assumption was that absorption would be proportional to Acat2 gene dosage. Male ACAT2?/?, ACAT2?/?, and ACAT2?/? mice were fed a diet containing 20% of energy as palm oil with 0.2% (w/w) cholesterol. Cholesterol absorption efficiency was measured by fecal dual-isotope and thoracic lymph duct cannulation (TLDC) methods using [³H]sitosterol and [¹?C]cholesterol tracers. Excellent agreement among individual mice was found for cholesterol absorption measured by both techniques. Cholesterol absorption efficiency in ACAT2?/? mice was 16% compared with 46-47% in ACAT2?/? and ACAT2?/? mice. Chylomicrons from ACAT2?/? and ACAT2?/? mice carried ?80% of total sterol mass as CE, whereas ACAT2?/? chylomicrons carried >90% of sterol mass in the unesterified form. The total percentage of chylomicron mass as CE was reduced from 12% in the presence of ACAT2 to ?1% in ACAT2?/? mice. Altogether, the data demonstrate that ACAT2 increases cholesterol absorption efficiency by providing CE for chylomicron transport, but one copy of the Acat2 gene, providing ?50% of ACAT2 mRNA and enzyme activity, was as effective as two copies in promoting cholesterol absorption.