Lipid peroxidation--the factor promoting cholesterol accumulation in cells in atherogenesis

The cholesterol transfer between human erythrocytes and main classes of serum lipoproteins (LP) from healthy donors and artery-coronary disease patients was studied (artery-coronary disease is the main manifestation of atherosclerosis). It is shown that low-density lipoproteins (LDL) are capable of transporting cholesterol to erythrocytes, which lack the specific receptors for LDL. The cell cholesterol content in comparison with erythrocytes incubated without LDL was increased by 11.4%. The effect was even higher in case of LDL, isolated from serum of artery-coronary subjects (the cell cholesterol content was increased by 33.8%). High-density lipoproteins (HDL) accept cholesterol from cell membranes. However, cholesterol-accepting properties of HDL from artery-coronary disease patients were suppressed as compared with normal HDL. Both discovered events must promote the cholesterol accumulation in cell membranes in atherosclerosis. As it is shown by the spin probe method, lipid peroxidation (LPO) causes the disturbance of the structural organization of LP and as the consequence of that--the increase of LDL cholesterol-donating ability and the decrease of HDL cholesterol-accepting ability. The greater LDL are oxidized, the more cholesterol they transport to erythrocytes during incubation. The greater is the level of HDL peroxidation, the stronger their cholesterol-accepting function is suppressed. These results suggest that LPO can play an important role in LP modification, the disturbance of their interaction with cell surface and the cholesterol accumulation in cells in atherosclerosis.     

Uptake of Endoneurial Lipoprotein into Schwann Cells and Sensory Neurons Is Mediated by Low Density Lipoprotein Receptors and Stimulated After Axonal Injury

During Wallerian degeneration of rat sciatic nerve, the expression of apolipoprotein E increases and apolipoprotein E-containing endoneurial lipoproteins accumulate in the distal nerve segment. In established primary cultures dissociated from dorsal root ganglia, Schwann cells and sensory neurons internalized rhodamine-labeled lipoproteins isolated from crushed rat sciatic nerve as well as low density lipoprotein (LDL) from human serum. The uptake of endoneurial lipoproteins could be inhibited by an excess of LDL or at low temperature (4 degrees C). After transection of nerve fibers in dorsal root ganglia explant cultures, the uptake of lipoproteins was markedly stimulated in Schwann cells that were in close proximity to degenerating neurites. A specific monoclonal antibody directed to the bovine LDL receptor (clone C7) was shown to cross-react with LDL receptor preparations of rat endoneurial cells. LDL receptor immunoreactivity was expressed by cell bodies and processes of cultured Schwann cells, sensory neurons, and fibroblasts from dorsal root ganglia. Incubation of Schwann cells and neurons with the LDL receptor antibody strongly inhibited the uptake of endoneurial lipoproteins. Our results provide direct evidence for the important role of the LDL receptor-mediated pathway to internalize endoneurial lipoproteins into Schwann cells and peripheral neurons required for reuse of cholesterol and other lipids in myelin and plasma membrane biogenesis during nerve repair.     

Free radical modification of lipoproteins and cholesterol accumulation in cells upon atherosclerosis.

An electron spin probe study was made of the effect of lipid peroxidation (LPO) on the structure of surface proteolipid layer of human serum low-density lipoproteins (LDL). The results obtained with a positively charged spin label and stearic acid spin probes with doxyl labels at positions 5, 12, and 16 revealed that LPO caused a decrease in phospholipid molecule mobility both in the region of polar heads and in the region of acyl chains till the depth of at least 1.7 mm from water-lipid interface. Under relatively high levels of oxidation (more than 6 mumol MDA/g LDL phospholipid) the polarity of lipid phase increased. The decrease in efficiency of tryptophan fluorescence quenching by nitroxide fragments incorporated in hydrophobic regions at the depth of approximately 2 nm from water-lipid interface indicated that lipid-protein interaction was disturbed as a result of oxidation of LDL lipids. In addition, the LPO-induced modification of apo-B, the main protein of LDL, was examined with maleimide spin label. LPO led to increase in mobility of strongly immobilized maleimide labels and in the number of weakly immobilized ones. Oxidized LDL revealed decreased ability to incorporate spin-labeled steroid (androstane) as compared to native ones. LPO-induced structural changes of LDL surface are supposed to be a reason of enhanced accumulation of cholesterol in human monocytes during their incubation with oxidized LDL. The cholesterol content in red cells was shown to be directly correlated to MDA content in apo-B containing lipoproteins but not in whole serum. Our findings suggest that free radical modification of serum lipoproteins but not solely an increased level of LPO products in blood is one important cause for cholesterol accumulation in cells and, apparently, for their transformation into foam cells during atherosclerosis.     

Transport of lipids from high and low density lipoproteins via scavenger receptor-BI.

The scavenger receptor-BI (SR-BI) delivers sterols from circulating lipoproteins to tissues, but the relative potency of individual lipoproteins and the transported cholesterol has not been studied in detail. In this study, we used Chinese hamster ovary cells that express recombinant mouse SR-BI but have no functional low density lipoprotein (LDL) receptors (ldlA7-SRBI cells) to compare the fate of lipids transferred from high or low density lipoproteins to cells by SR-BI. HDL and LDL were equally effective in mediating the transfer of [(3)H]cholesterol to cells. Only 5% of the free cholesterol transferred to cells was esterified, in direct contrast to the findings in the cells that express LDL receptors in which 50% of the transported cholesterol was esterified. Almost all the free cholesterol transferred from lipoproteins to cells was rapidly excreted when the ldlA7-SRBI cells were switched to media containing unlabeled lipoproteins. SR-BI expression was associated with an increase in selective cholesteryl ester uptake from both lipoproteins, but HDL was a more effective donor. HDL and LDL were equally effective in delivering cholesterol to the intracellular regulatory pool via SR-BI. These data indicate that SR-BI is able to exchange cholesterol rapidly between lipoproteins and cell membranes and can mediate the uptake of cholesteryl esters from both classes of lipoproteins.     

The effect of serum lipoproteins on cholesterol content and sterol exchange in cultured human endothelial cells.

Cultured human endothelial cells preincubated with the infranatant of human serum increased their content of cholesterol when subsequently exposed to low density lipoproteins (LDL) as compared to control cultures further incubated in the presence of infranatant only. Replacing LDL with high density lipoproteins (HDL) resulted in no change in the cellular cholesterol content compared to the control. The addition of HDL did not influence the increase in cellular cholesterol content mediated by LDL. HDL stimulated the efflux of endogenously synthesized 14C-labelled sterols compared to the infranatant fraction, whereas LDL had only a slight effect. Cells preincubated with whole serum did not change their cholesterol content when subsequently exposed to LDL, compared to cultures further incubated in presence of whole serum. Replacing whole serum (during the final incubation) with infranatant, resulted in a decrease of the cellular cholesterol content, which was not influenced by further addition of HDL.     

Characterization of plasma lipoproteins of grain- and cholesterol-fed White Carneau and Show Racer pigeons

Plasma cholesterol concentrations from White Carneau (WC) and Show Racer (SR) pigeons consuming a cholesterol-free grain diet averaged about 300 mg/dl, approximately 200 mg/dl as high density lipoproteins (HDL) and the remainder as low density lipoproteins (LDL). Consumption of a cholesterol-containing diet increased plasma cholesterol concentrations in both breeds to greater than 2000 mg/dl. Approximately one-half of this increase was as LDL with the remainder as beta-migrating very low density lipoproteins (beta-VLDL). There was little change in HDL concentration. LDL from cholesterol-fed animals had a greater net negative charge than control LDL, and was larger (Mr = 10 X 10(6) vs 3.2 X 10(60)) due to an increase in the number of cholesteryl ester molecules per particle. The principal apoprotein of LDL was apoB-100 with smaller amounts of apoA-I and several minor unidentified apoproteins. beta-VLDL was cholesteryl ester-rich, could be separated into two size populations by gel chromatography, and contained apoB-100 as its principal apoprotein. Apoprotein E was not detected in any of the plasma lipoproteins. HDL from control and cholesterol-fed animals was composed of a single class of particles with virtually identical composition resembling HDL2. The major apoprotein of HDL was apoA-I. There were no consistent quantitative or qualitative differences in the lipoproteins of the two breeds of pigeons that could help to explain the susceptibility to atherosclerosis of the WC or the resistance of the SR.     

Sterol-modulated glycolipid sorting occurs in niemann-pick C1 late endosomes

The Niemann-Pick C1 (NPC1) protein and endocytosed low density lipoprotein (LDL)-derived cholesterol were shown to enrich separate subsets of vesicles containing lysosomal associated membrane protein 2. Localization of Rab7 in the NPC1-containing vesicles and enrichment of lysosomal hydrolases in the cholesterol-containing vesicles confirmed that these organelles were late endosomes and lysosomes, respectively. Lysobisphosphatidic acid, a lipid marker of the late endosomal pathway, was found in the cholesterol-enriched lysosomes. Recruitment of NPC1 to Rab7 compartments was stimulated by cellular uptake of cholesterol. The NPC1 compartment was shown to be enriched in glycolipids, and internalization of GalNAcbeta1-4[NeuAcalpha2-3]Galbeta1-4Glcbeta1-1'-ceramide (G(M2)) into endocytic vesicles depends on the presence of NPC1 protein. The glycolipid profiles of the NPC1 compartment could be modulated by LDL uptake and accumulation of lysosomal cholesterol. Expression in cells of biologically active NPC1 protein fused to green fluorescent protein revealed rapidly moving and flexible tubular extensions emanating from the NPC1-containing vesicles. We conclude that the NPC1 compartment is a dynamic, sterol-modulated sorting organelle involved in the trafficking of plasma membrane-derived glycolipids as well as plasma membrane and endocytosed LDL cholesterol.     

The lipid composition of serum lipoproteins in the harbour seal, Phoca vitulina

The density profile of serum lipoproteins and their lipid composition was studied in 12 adult, female harbour seals. The animals were sampled after an approximate 20 hr fast. The density profile of lipoproteins showed that the harbour seals displayed a distinct VLDL (density less than 1.006 g/ml) and HDL band (density about 1.125 g/ml), but no clear LDL band. There was a rather diffuse population of lipoproteins in the density range of 1.019-1.100 g/ml. Mean serum total cholesterol concentration was 5.7 mmol/l; about 60% of this cholesterol was located in the HDL fraction (density greater than 1.063 g/ml). The fasted seals were found to carry 4% of serum total lipids in chylomicrons. These lipoproteins consisted of 51% of triaclyglycerols (on the basis of total chylomicron lipids). The LDL (defined as heparin-manganese precipitable lipoproteins in VLDL and chylomicron-deficient serum) contained 49% of cholesterol and 43% of phospholipids (on the basis of total LDL lipids). The HDL (defined as heparin-manganese soluble lipoproteins in VLDL and chylomicron-deficient serum) contained 36% of cholesterol and 58% of phospholipids (on the basis of total HDL lipids).     

Differential effects of isolated lipoproteins from normal and hypercholesterolemic rhesus monkeys on cholesterol esterification and accumulation in arterial smooth muscle cells in culture.

Whole serum obtained from hypercholesterolemic rhesus monkeys was found to stimulate cholesterol esterification and cholesteryl ester accumulation in rhesus monkey arterial smooth muscle cells in culture to a significantly greater extent than normocholesterolemic serum. This was true even when the cholesterol concentration of the culture medium was equalized. Isolation and characterzation of the low density lipoproteins (LDL) from rhesus monkeys indicated that the LDL from hypercholesterolemic animals was 33% larger than LDL from normocholesterolemic animals due principally to an increase in the amount of cholesteryl ester per molecule. As a result, LDL from hypercholesterolemic animals transported over 50% more cholesterol per molecule than did normal LDL. The LDL of altered composition from hypercholesterolemic animals, when added to smooth muscle cells in culture, was nearly twice as effective in stimulating cholesterol esterification and cholesteryl ester accumulation than was LDL of normal composition. Results suggest that at least part of the exaggerated ability of whole hypercholesterolemic serum to stimulate the esterification and accumulation of cholesterol in cells in culture is due to the presence of LDL of altered composition.     

Lipid composition of cells and low-density lipoproteins in blood serum of human and some vertebrate species

To establish interaction of atherogenic low-density lipoproteins (LDL) with the erythrocyte membrane, the content of lipid components in blood cells and serum LDL was studied in healthy people (donors) and in 12 species of vertebrates (the mammals non-predisposed to atherosclerosis — birds and fish). Lipid composition of blood cells and LDL was also analyzed in patients with pathologies: ischemic heart disease (IHD), bronchial asthma (BA), and chronic obstructive bronchitis (COB), as well as in 2 species of mammals predisposed to atherosclerosis, in whose blood LDL predominated. The content of lipids in the blood cells and LDL of the studied vertebrates has been found to depend on their taxonomy and on the clear trends either for an increase in the cholesterol content and a decrease in the phosphatidylcholine level in patients, particularly with IHD, or for a rise of the ratio of the content of the more saturated sphingomyelin and cholesterol to the less saturated phosphatidylcholine from the lower to the higher organisms, including humans (donors). The highest levels of free cholesterol in blood cells of total cholesterol in LDL, as well as of parameters of ratio of the cholesterol/phosphatidylcholine content have been revealed in patients, especially with IHD, and in the mammals predisposed to atherosclerosis, i.e. in representatives with predominance of blood LDL, in contrast to donors and the mammals resistant to atherosclerosis. The highest parameters of lipid components were determined in blood cells and LDL in patients with IHD. The lipid LDL composition affects directly the composition and ratio of lipids in blood cells.     

Multiple effects of serum low-density lipoprotein on cultured human fibroblasts.

The effects of serum low-density lipoproteins (LDL) were studied in cultures of human skin fibroblasts grown in medium supplemented with human serum deficient in lipoproteins and in platelet factor. The LDL led to a temporary increase in the rate of cell replication, to increases in the cell content of protein and cholesterol, to an increase in average cell size, and to an increased secretion of glycosaminoglycans. The increases in cholesterol and protein were proportional to the increase in cell size, suggesting that the additional protein and cholesterol were of a structural, rather than a storage, nature. The increase in cell protein during the first few days of exposure to LDL was due to a decrease in the rate of protein degradation. Ultrafiltration of the serum to remove substances of molecular weight less than 30,000 did not reduce the basal rate of cell proliferation but did prevent the stimulation of proliferation by LDL; it did not alter the effect of LDL on cell protein and cholesterol, indicating that the latter responses are independent of the mitogenic action. The response of cells from diabetic donors did not differ from that of normal cells.     

Lipoprotein-dependent synthesis of prostacyclin in smooth muscle cells

Low (LDL) and high density lipoproteins (HDL) stimulated prostacycline (PGI2) synthesis in rabbit and human aorta smooth muscle cells growing in culture. The lipoproteins were added to the cells in concentrations equal to that of cholesterol. It was shown that HDL exerted a stronger stimulating effect as compared to LDL. The maximal effect was observed with HDL3. HDL3 isolated from blood serum of healthy volunteers appeared to be more active in PGI2 synthesis promotion than those of CDH patients with documented coronary atherosclerosis. Purified Apo A-1 stimulated the transformation of [14C]arachidonic acid into the products of its metabolism with increased accumulation of 6-keto-PGF1 alpha among labeled metabolites. Estradiol (1.10(-7) M) showed a stimulating effect; norepinephrine (1.10(-6) M) and progesterone (1.10(-7) M) showed an inhibiting effect, whereas corticosterone (1.10(-6) M) and deoxycorticosterone (1.10(-6) M) did not influence the rate of LDL-dependent PGI2 synthesis.     

Quantitation of the in vitro free cholesterol exchange of human red cells and lipoproteins

The exchange of free cholesterol in vitro between human red blood cells and low density lipoproteins (LDL) was quantified. The flux of sterol between LDL and red cells was relatively constant over a wide range of concentrations of free cholesterol in lipoproteins. In a system containing a suspension of red blood cells in a mixed solution of high density lipoproteins (HDL) and LDL, the fractional rate of exchange of HDL cholesterol was most rapid followed by LDL and lastly, by red cells. Increasing the ionic strength of the incubation media had no effect on the exchange of cholesterol between LDL and red cells. However, when both HDL and LDL were incubated with red cells in a buffer of increased ionic strength, total red cell cholesterol exchange was unaltered, but proportionately more exchange occurred with HDL and less with LDL. Addition of acetone to the buffer increased the exchange of cholesterol between LDL and red cells but produced no increment in red cell-HDL exchange.     

The interaction of prostaglandins with serum low-density lipoproteins

The interaction of human serum low-density lipoproteins (LDL) with various types of prostaglandins (PG) was studied using equilibrium dialysis, steady-state fluorescence polarization spectroscopy and photolabeling methods. Low concentrations (10(-13)-10(-9) M) of PGE1 and PGF2 alpha were shown to induce specific rearrangements of the lipids on the LDL surface, whereas the closely related PGE2 and PGF1 alpha had no effect. With fluorescent labeled LDL, the PGE1-induced changes of the steady-state fluorescence polarization (P) were shown to be time- and concentration-dependent, saturable and reversible. However, equilibrium dialysis revealed a very low binding capacity of LDL for PGE1 (approx. 1 prostaglandin molecule per 600 LDL particles). Approximately the same PGE1 concentration was sufficient to cause maximal changes of P, to enhance the binding to apolipoprotein B of a photoreactive sphingomyelin analogue inserted into the LDL surface and to alter the thermal phase behavior of the LDL surface lipids. It is proposed that the LDL surface rearrangement caused by prostaglandins is due to the interaction of prostaglandins with apolipoprotein B, resulting in formation of short-lived complexes. The mechanism of this interaction is discussed in terms of the non-equilibrium ligand-receptor interaction model proposed earlier to explain the interaction of prostaglandins with high-density lipoproteins (Bergelson, L.D. et al. (1987) Biochim. Biophys. Acta 921, 182-190). It is suggested that direct prostaglandin-lipoprotein interactions may play a role in the homeostasis of cholesterol.     

Characterization of plasma low density lipoproteins on nonhuman primates fed dietary cholesterol.

LDL from animals of three nonhuman primate species, Macaca mulatta, Macaca fascicularis, and Cercopithecus aethiops, were studied. A standard preparation of 125I-LDL was added to isolated lipoprotein mixtures just prior to separation of plasma lipoproteins by agarose gel chromatography. A relative size index, rI, was determined by dividing the elution volume of the iodinated LDL by the elution volume of the sample LDL, both volumes being determined simultaneously during chromatographic elution. Comparison of rI with molecular weights measured by flotation equilibrium analysis in the analytical ultracentrifuge showed a linear relationship across a molecular weight range of 2.5-8.0 X 10(6), r = 0.985. A regression equation describing this relationship was used to calculate molecular weights of LDL from a group of M. fascicularis that were fed cholesterol-containing diets. In these animals, plasma cholesterol concentration ranged from 100 to over 700 mg/dl and was highly correlated with LDL molecular weight and with the micromolar concentration of the LDL. Using multiple regression analyses, the two variables of plasma LDL could be shown to account for 94% of the variation in plasma cholesterol concentration in the M. fascicularis of this study. Micromolar concentration and molecular weight of LDL were not correlated with each other, suggesting that in M. fascicularis at least two independent types of controls are operative in the response of plasma LDL to dietary cholesterol. The increase in LDL molecular weight was associated with a large increase in cholesteryl ester content and concomitant smaller increases in protein, phospholipid, and free cholesterol. As molecular weight increased, these components appeared to be added to the LDL particles together as discrete increments of fixed composition. The data are consistent with a spherical model of LDL structure with a core of cholesteryl ester and triglyceride and a 21.3 A-thick coat of phospholipid, free cholesterol, and protein.     

Measurement of cholesterol bidirectional flux between cells and lipoproteins

We developed an assay that quantitates bidirectional cholesterol flux between cells and lipoproteins. Incubating Fu5AH cells with increasing concentrations of human serum resulted in increased influx and efflux; however, influx was 2- to 3-fold greater at all serum concentrations. With apolipoprotein B (apoB)-depleted serum, the ratio of influx to efflux (I/E) was close to 1, indicating cholesterol exchange. The apoB fraction of serum induced influx and little efflux, with I/E > 1. Using block lipid transport-1 to block scavenger receptor class B type I (SR-BI)-mediated flux with different acceptors, we determined that 50% to 70% of efflux was via SR-BI. With HDL, 90% of influx was via SR-BI, whereas with LDL or serum, 20% of influx was SR-BI-mediated. Cholesterol-enriched hepatoma cells produced increased efflux without a change in influx, resulting in reduced I/E. The assay was applied to cholesterol-normal and -enriched mouse peritoneal macrophages exposed to serum or LDL. The enrichment enhanced efflux without shifts in influx. With cholesterol-enriched macrophages, HDL efflux was enhanced and influx was greatly reduced. With all lipoproteins, cholesterol enrichment of murine peritoneal macrophages led to a reduced I/E. We conclude that this assay can simultaneously and accurately quantitate cholesterol bidirectional flux and can be applied to a variety of cells exposed to isolated lipoproteins or serum.     

Regulation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase activity in mouse peritoneal macrophages.

The lipoprotein-mediated regulation of 3-hydroxy-3-methylglutaryl-(HMG-) CoA reductase in cultured mouse peritoneal macrophages has been investigated. In contrast to what has been reported for other cells, HMG-CoA reductase activity is not suppressed by normal serum or by normal low density lipoproteins (LDL) from humans or dogs. Suppression of reductase activity occurred when cells were cultured in the presence of beta-migrating very low density lipoproteins (beta-VLDL) or LDL from hypercholesterolaemic dogs, or LDL modified by acetoacetylation. Human beta-VLDL from an atypical type III hyperlipoproteinaemic patient was also effective, as was apolipoprotein (apo) E-containing high density lipoproteins (HDL) from cholesterol-fed dogs (apo-E HDLc). The results indicate that cholesterol biosynthesis in mouse peritoneal macrophages is regulated by lipoprotein cholesterol entering via receptor-mediated endocytosis. Normal LDL were not effective because of the poor binding and uptake of these lipoproteins by the apo-B, E (LDL) receptor. Only beta-VLDL, apo-E HDLc, and hypercholesterolaemic LDL were avidly taken up by this receptor and were able to suppress HMG-CoA reductase. Acetoacetylated LDL were internalized via the acetyl-LDL (scavenger) receptor. Thus, mouse macrophages differ from human fibroblasts and smooth muscle cells in their physiological regulation of cholesterogenesis.     

Specific targeting of high density lipoproteins to liver hepatocytes by incorporation of a tris-galactoside-terminated cholesterol derivative

A triantennary galactose-terminated cholesterol derivative, N-(tris(beta-D-galactopyranosyloxymethyl) methyl)-N alpha-(4(5-cholesten-3 beta-yloxy)succinyl)glycinamide (Tris-Gal-Chol), which dissolves easily in water, was added to human apolipoprotein E-free high density lipoproteins (HDL) in varying quantities. Incorporation of 5 or 13 micrograms of Tris-Gal-Chol into HDL (20 micrograms of protein) stimulates the liver association of the HDL apoprotein radioactivity 24- and 55-fold, respectively, at 10 min after intravenous injection into rats. The increased interaction of Tris-Gal-Chol HDL with the liver is blocked by preinjection of asialofetuin or N-acetylgalactosamine but not influenced by N-acetylglucosamine. The parenchymal liver cell uptake of HDL is stimulated 42- or 105-fold, respectively, by incorporation of 5 or 13 micrograms of Tris-Gal-Chol into HDL (20 micrograms of protein), while the association with nonparenchymal cells is stimulated only 1.7- or 5-fold. It can be calculated that 98.0% of the Tris-Gal-Chol HDL is associated with parenchymal cells. In contrast, incorporation of 13 micrograms of Tris-Gal-Chol into LDL (20 micrograms of protein) leads to a selective association of LDL with nonparenchymal cells (92.3% of the total liver uptake). It is concluded that Tris-Gal-Chol incorporation into HDL leads to a specific interaction of HDL with the asialoglycoprotein (galactose) receptor on parenchymal cells whereas Tris-Gal-Chol incorporation into LDL leads mainly to an interaction with a galactose receptor from Kupffer cells. Probably this highly selective cellular targeting of LDL and HDL by Tris-Gal-Chol is caused by the difference in size between these lipoproteins. The increased interaction of HDL with the parenchymal cells upon Tris-Gal-Chol incorporation is followed by degradation of the apolipoprotein in the lysosomes. It is concluded that Tris-Gal-Chol incorporation into LDL or HDL leads to a markedly increased catabolism of LDL by way of the Kupffer cells and HDL by parenchymal cells which might be used for lowering serum cholesterol levels. The use of Tris-Gal-Chol might also find application for targeting drugs or other compounds of interest to either Kupffer or parenchymal liver cells.     

Evaluation of rat and rabbit sera lipoproteins in experimentally induced hyperlipidemia by analytical ultracentrifugation

Animals of various species are widely used as models with which to study atherosclerosis and the lipoprotein metabolism. The objective of this study was to investigate the lipoprotein profiles in Wistar rats and New Zealand white rabbits with experimentally induced hyperlipidemia by means of ultracentrifugation. The Schlieren curves were utilized to compare suckling and adult rat sera to determine whether aging causes alterations in lipoprotein profiles. A striking feature of the data is the high concentration of low-density lipoproteins (LDL), (>5.2 mmol/l cholesterol) in the 2-week old rat serum pool which was greatly decreased in the 3-weeks rat serum pool (<1.3 mmol/l cholesterol). Additional experiments were performed to permit a direct comparison of the amounts of lipoprotein present in rat sera in experimental hyperlipidemia post-Triton WR 1339 administration. Rapid changes in concentrations in very low-density lipoproteins (VLDL), LDL and high-density lipoproteins (HDL) were observed after Triton injection. The administration of Triton WR 1339 to fasted rats resulted in an elevation of serum cholesterol levels. Triton physically alters VLDL, rendering them refractive to the action of lipolytic enzymes in the blood and tissues, preventing or delaying their removal from the blood. Whereas the VLDL concentration was increased markedly, those of LDL and HDL were decreased at 20 h after Triton treatment. 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 of LDL aliquots, to prepare radioactive-labeled lipoproteins and to study induced hyperlipidemia in rabbits. Analytical ultracentrifugation was applied to investigate the LDL flotation peaks before and after cholesterol feeding of rabbits. Modified forms of LDL were detected in the plasma of rabbits with experimentally induced atherosclerosis. ApoB-containing particles, migrating as LDL, intermediate density lipoproteins and VLDL were the most abundant lipoproteins. Gamma camera in vivo scintigraphy on rabbits with radiolabeled lipoproteins revealed visible signals corresponding to atherosclerotic plaques of the aorta and carotid arteries.