Effects of protamine on blood lipoproteins in the early period of hypercholesterolemia in rabbits

The content of protein, cholesterol and triglycerides in chylomicrons and different classes of blood serum lipoproteins was studied under protamine action in early (1 month) hypercholesterolemia. Except the low density lipoproteins the amount of triglycerides in the particles studied was practically unchanged. The content of proteins in all classes of lipoproteins was greatly increased and this may indicate that protamine activates the blood lipoprotein system. The enrichment of lipoproteins with cholesterol may be taken as being a result of protamine action in early hypercholesterolemia. The peculiar feature in the effect of protamine on mentioned lipoprotein components was that the protein/cholesterol ratio didn't change in chylomicrons, decreased in very low density lipoproteins and low density lipoproteins and increased in high density lipoproteins as compared with controls.     

Association Between Lipids, Lipoproteins Composition of HDL Particles and Triglyceride-Rich Lipoproteins, and LCAT and CETP Activity in Post-renal Transplant Patients

High-density lipoprotein (HDL) remodeling within the plasma compartment and the association between lecithin-cholesterol acyltransferase (LCAT) and cholesterol ester transfer protein (CETP) activity, and lipid, lipoprotein concentrations and composition were investigated. The aim was to examine the high sensitivity of C-reactive protein (hsCRP), lipid, apolipoprotein B (apoB), apoAI, total apoAII, apoAIInonB, apoB-containing apoAII (apoB:AII), total apoCIII, apoCIIInonB, apoB-containing apoCIII (apoB:CIII) concentration and LCAT and CETP activity to gain an insight into the association between them and LCAT and CETP, 57 post-renal transplant (Tx) patients with and without statin therapy and in 15 healthy subjects. Tx patients had moderate hypertriglyceridemia, hypercholesterolemia, and dyslipoproteinemia, disturbed triglyceride-rich lipoproteins (TRLs) and HDL composition, decreased LCAT, and slightly increased hsCRP but no CETP activity. Spearman’s correlation test showed the association between lipids and lipoproteins and LCAT or CETP, and multiple ridge stepwise forward regression showed that immunosuppressive therapy in Tx patients can disturb HDL and TRLs composition. The results suggest that inhibition or activation of LCAT is due, in part, to HDL-associated lipoprotein. Lipoprotein composition of apoAI, apoAIInonB, and apoCIIInonB in HDL particle and apoB:AII TRLs can contribute to decrease LCAT mass in Tx patients. Tx patients without statin and with lower triglycerides but higher HDL cholesterol concentration and disturbed lipoprotein composition of ApoAI and apoAII in HDL particle can decrease LCAT, increase LDL cholesterol, aggravate renal graft, and accelerate atherosclerosis and chronic heart diseases.     

Increased production of apolipoprotein B-containing lipoproteins in the absence of hyperlipidemia in transgenic mice expressing cholesterol 7alpha-hydroxylase

The finding that expression of a cholesterol 7alpha-hydroxylase (CYP7A1) transgene in cultured rat hepatoma cells caused a coordinate increase in lipogenesis and secretion of apoB-containing lipoproteins led to the hypothesis that hepatic production of apoB-containing lipoproteins may be linked to the expression of CYP7A1 (Wang, S.-L., Du, E., Martin, T. D., and Davis, R. A. (1997) J. Biol. Chem. 272, 19351-19358). To examine this hypothesis in vivo, a transgene encoding CYP7A1 driven by the constitutive liver-specific enhancer of the human apoE gene was expressed in C56BL/6 mice. The expression of CYP7A1 mRNA (20-fold), protein ( approximately 10-fold), and enzyme activity (5-fold) was markedly increased in transgenic mice compared with non-transgenic littermates. The bile acid pool of CYP7A1 transgenic mice was doubled mainly due to increased hydrophobic dihydroxy bile acids. In CYP7A1 transgenic mice, livers contained approximately 3-fold more sterol response element-binding protein-2 mRNA. Hepatic expression of mRNAs encoding lipogenic enzymes (i.e. fatty-acid synthase, acetyl-CoA carboxylase, stearoyl-CoA desaturase, squalene synthase, farnesyl-pyrophosphate synthase, 3-hydroxy-3-methylglutaryl-CoA reductase, and low density lipoprotein receptor) as well as microsomal triglyceride transfer protein were elevated approximately 3-5-fold in transgenic mice. CYP7A1 transgenic mice also displayed a >2-fold increase in hepatic production and secretion of triglyceride-rich apoB-containing lipoproteins. Despite the increased hepatic secretion of apoB-containing lipoproteins in CYP7A1 mice, plasma levels of triglycerides and cholesterol were not significantly increased. These data suggest that the 5-fold increased expression of the low density lipoprotein receptor displayed by the livers of CYP7A1 transgenic mice was sufficient to compensate for the 2-fold increase production of apoB-containing lipoproteins. These findings emphasize the important homeostatic role that CYP7A1 plays in balancing the anabolic lipoprotein assembly/secretion pathway with the cholesterol catabolic bile acid synthetic pathway.     

Concentration and composition of serum lipoproteins in the fetal rat]

1. Concentration and composition of the "very low density lipoproteins" (VLDL), "low density lipoproteins" (LDL) and "high density lipoproteins" (HDL) and of non-floatable lipids of fetal rat serum (day 22 of pregnancy) were determined by ultracentrifugation, thin-layer chromatographic separation of the floated lipids and quantitation of the lipid and protein moiety. 2. The concentration of VLDL is in the fetal rat by one order of magnitude lower, and that of LDL, 5fold higher than in the adult animal; the concentration of HDL in fetal serum amounts to 60% of the value of adult animals. 3. The composition of LDL and HDL of fetal serum does not differ from that in the serum of adult animals; in contrast, the fetal VLDL have a higher proportion of protein and cholesterol and a lower proportion of triglycerides than the VLDL of adult serum. The electrophoretic mobility of the fetal VLDL is lower than that of adult VLDL.     

Effect of ketoconazole on the activity of key enzymes of biosynthesis of cholesterol and its esters in the liver of rats maintained on a cholesterol-enriched diet]

Ketoconazole in vivo has been studied for its effect on the activity of key enzymes of the cholesterol and its esters' biosynthesis in the liver and on the cholesterol concentration in certain fractions of blood lipoproteins in normal and cholesterol-fed rats. It is established that ketoconazole decreases cholesterol concentration in low-density lipoproteins and in very low-density lipoproteins as well as decrease the acyl-CoA-cholesterol acyl-transferase activity and increases the 3-hydroxy-3-methyl-glutaryl-CoA-reductase activity in the liver microsomes of intact and test animals. It is supposed that the possible cause of the observed changes can be a disturbance in regulation of basic links of cholesterol metabolism in the liver.     

Comparative characterization of the lipoprotein spectrum and status of free radical processes in the blood of rabbits exposed to external gamma-irradiation and cholesterol administration]

Characteristic of lipoprotein spectrum and state of free-radical processes in the animals under action of ionizing radiation and cholesterin diet are comparatively studied. The content of lipids in the blood serum increases on the 5th day after irradiation and then decreases on the 15th and 30th days. Cholesterin diet increases the content of cholesterin under practically unchanged content of triglycerides in the blood serum in all the terms of investigations. Total decrease of the intensity of induced chemiluminescence of the blood serum and erythrocytes after irradiation is shown. Peroral introduction of cholesterin evokes differently directed changes in the light sum of chemiluminescence--a decrease in erythrocytes and increase in the blood serum.     

Dynamics of the lipoprotein distribution in early hypercholesterolemia characterizes the activation of cholesterol transport]

Using gradient gel electrophoresis, the levels of major classes of rabbit blood sera lipoproteins were studied over a period of 28 days. Hypercholesterolemia persisted up to the 4th day with a subsequent decrease. This was paralleled with an increase in the levels of triglyceride-rich and apoB-containing particles. The decrease of the electrophoretic mobility of low density lipoproteins correlated with an increased content of the intermediate fraction. On the 28th day after the beginning of experiment the concentration of total high density lipoproteins showed an increase. The subfractional redistribution of HDL3 and HDL2 subclasses pointed to the stimulation of the accepting process and the overall reverse cholesterol transport system. A comparison of experimental results with literary data allowed to conclude that the relative decrease of the serum cholesterol level typical of hypercholesterolemia of one month duration is due to the activation of specific and nonspecific preventive mechanisms.     

Characterization of subfractions of triglyceride-rich lipoproteins separated by gel chromatography from blood plasma of normolipemic and hyperlipemic humans

As judged from measurements of the diameters of particles fixed with osmium tetroxide and shadowed with platinum, gel chromatography on 2% agarose has been shown to be an effective quantitative method for separating triglyceride-rich lipoproteins according to particle size. Particles in the size range of chylomicrons, uncontaminated by lipoproteins smaller than about 700 A or by other serum proteins, emerged in the void volume of the column, and very low density lipoproteins with diameters between 400 and 700 A were separated into fractions with average standard deviation of 71 A from the mean. Systematic comparison of the relationship between diameter and chemical composition of fractions obtained from subjects with various hyperlipoproteinemic disorders demonstrated a precise correlation consistent with a spherical model for these lipoproteins in which phospholipids, free cholesterol, and protein occupy a surface monolayer with an invariant thickness of 21.5 A surrounding a liquid core of triglycerides and cholesteryl esters. The chemical composition of very low density lipoproteins of given particle size in most recognized types of hyperlipemia was similar to that of normolipemic subjects, but particles in the size range of chylomicrons sometimes had higher contents of cholesteryl esters and free cholesterol. Results obtained in subjects with dysbetalipoproteinemia were consistent with the presence of three populations of particles. Two of these, with mean diameters of about 850 and 350 A, had unusually high cholesteryl ester content and reduced triglyceride content and may represent "remnants" of the metabolism of structurally normal chylomicrons and very low density lipoproteins, respectively. The third, a heterogeneous group with intermediate range of particle size and pre-beta mobility, may represent a population of very low density lipoproteins with relatively normal composition.     

Characteristics of lipid composition of blood serum lipoproteins in insulin resistance

The purpose of this investigation was to study lipid composition of apoB- and apoA-containing lipoproteines in conditions of insulin resistance pathology. It was shown that hypertriglycerolemia and hypercholesterolemia were determined by triglycerole and cholesterol surplus in the composition of both apoB- and apoA-containing lipoproteins. Phospholipid deficiency in the composition of all lipoproteins fractions were established. Possible mechanisms of formation and metabolism in circulation of blood lipoproteins with altered lipid composition in the blood flow at insulin resistance is discussed.     

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.     

Comparative biochemical analysis of blood serum lipoproteins from human and various animal species

Lipid composition of blood serum and total lipids of low density lipoproteins (LDL) and high density lipoproteins (HDL2 and HDL3) were studied in human (donors, patients with ischemic heart disease, bronchial asthma, chronic obstructive bronchitis, as well as with a combined pathology), in mammals predisposed to atherosclerosis (pig, rabbit) and resistant to atherosclerosis (rat, mink, Arctic fox), in birds (hen, pigeon), in teleost fish (white fish, pikeperch, pike, bream, burbot) and cartilaginous fish (sturgeon, housen). It has been established that the most enriched in lipids is the blood serum of animals, particularly of cartilaginous fish. Twice lower is the lipid content in blood serum of donors than of animals. However, in the vascular, bronchial-pulmonary, and combined human pathologies the lipid level rises statistically significantly. In human and in animals predisposed to atherosclerosis the main mass of lipid is located in LDL, whereas in animals resistant to this disease--in HDL. The ratio of the human lipid content in LDL/HDL increases from 1.4 (in donors) to 2.7 in pathological states--in ischemic heart disease and its combination with chronic obstructive disease. In animals, a decrease of this ratio is noted from 1.0 to 0.2 in cartilaginous fish. By the example of one taxon (fish) there is established a regularity that indicates that evolution of lipoproteins occurred with an increase of the lipid amount in the "younger" LDL and with a decrease of concentration of the "colder" HDL.     

Effects of oleate and insulin on the production rates and cellular mRNA concentrations of apolipoproteins in HepG2 cells

We have reported previously that, in HepG2 cells, there is a lack of coordinate induction of triglyceride and apolipoprotein (apo) production by oleate and that insulin inhibits the production of triglyceride-rich, apoB-containing lipoproteins. The aim of the present study was to determine whether the effects of oleate and insulin on the net accumulation of apolipoproteins, specifically apoB, were related to their cellular mRNA concentrations. It was first established that the production of triglyceride-rich, apoB-containing lipoproteins and the concentration of mRNA for apolipoproteins A-I, A-II, B, and E were not affected by increasing the glucose concentration of medium from 5.5 to 20 mM. Oleate and insulin had no effect on either the accumulation in the medium or the cellular mRNA concentration of apolipoproteins A-I and A-II. On the other hand, the addition of oleate caused a two- to threefold increase in the accumulation of triglycerides in the medium without significantly affecting either the rates of accumulation or cellular mRNA levels of apolipoproteins B and E. In the presence of insulin, there was a dose-dependent decrease in the net accumulation of triglycerides and apoB and, to a lesser extent, cholesteryl esters and apoE. This inhibitory effect of insulin on the accumulation of triglycerides and apoB was partially abolished after a prolonged exposure of cells to insulin. Under these experimental conditions and at all concentrations tested, insulin had no effect on the cellular concentration of mRNA for either apoB or apoE.(ABSTRACT TRUNCATED AT 250 WORDS)     

Interaction of very low density lipoproteins (VLDL) with macrophages and their triboluminescence in hypercholesterolemia

Accumulation of cholesterol esters and triglycerides in peritoneal mice macrophages in the course of their interaction with lipoproteins of very low density (VLDL) is shown to grow considerably under conditions of hypercholesterolemia. A decrease of triboluminescence intensity characterizing the surface charge has been revealed at hypercholesterolemia both in VLDL and in the blood plasma. It is supposed that the triboluminescence method may be used for testing of the atherosclerotic process development.     

Isolation and characterization of an apoA-II-containing lipoprotein (LP-A-II:B complex) from plasma very low density lipoproteins of patients with Tangier disease and type V hyperlipoproteinemia

Previous studies have shown that very low density lipoproteins (VLDL) from patients with Tangier disease are less effective as a substrate for human milk lipoprotein lipase (LPL) than VLDL from normal controls as assessed by measuring the first order rate constant (k1) of triglyceride hydrolysis. Tangier VLDL also has a higher content of apolipoprotein (apo) A-II than normal VLDL. To explore the possible relationship between the relatively high concentration of apoA-II in VLDL and low k1 values, Tangier VLDL were fractionated on an anti-apoA-II immunosorber. The retained fraction contained a newly identified triglyceride-rich lipoprotein characterized by the presence of apolipoproteins A-II, B, C-I, C-II, C-III, D, and E (LP-A-II:B:C:D:E or LP-A-II:B complex), whereas the unretained fraction consisted of previously identified triglyceride-rich apoB-containing lipoproteins free of apoA-II. In VLDL from patients with Tangier disease or type V hyperlipoproteinemia, the LP-A-II:B complex accounted for 70-90% and 25-70% of the total apoB content, respectively. The LP-A-II:B complexes had similar lipid and apolipoprotein composition; they were poor substrates for LPL as indicated by their low k1 values (0.014-0.016 min-1). In contrast, the apoA-II-free lipoproteins present in unretained fractions were effective substrates for LPL with k1 values equal to or greater than 0.0313 min-1. These results indicate that triglyceride-rich lipoproteins consist of several apoB-containing lipoproteins, including the LP-A-II:B complex, and that lipoprotein particles of similar size and density but distinct apolipoprotein composition also possess distinct metabolic properties.     

Simvastatin causes the formation of cholesterol-rich remnants in mice lacking apoE

Mice that lack apolipoprotein E (apoE) display a severe hypercholesterolemia, caused by the accumulation of apolipoprotein B-48 (apoB-48)-carrying remnants of chylomicrons and very-low-density lipoproteins in the plasma. Statins are potent inhibitors of cholesterol synthesis that, when administered to mice lacking apoE, cause paradoxical further increases in plasma cholesterol levels. In the present study, we examined the mechanisms responsible for this phenomenon. ApoE-deficient mice fed a chow diet containing simvastatin developed, as anticipated, an enhanced increase in plasma cholesterol and a decrease in plasma triglycerides. Fractionation of the plasma lipoproteins by FPLC revealed that the lipid changes were confined to the lipoprotein remnants. Western blot analysis of the remnants from the untreated and simvastatin-treated mice showed no differences in their apoB-48 content, indicating that both groups of animals accumulated similar numbers of remnant particles in the plasma. Following the injection of Triton WR-1339, the simvastatin-treated mice accumulated in the plasma significantly more cholesterol and significantly less triglycerides than the untreated animals. These results indicate that the enhanced hypercholesterolemia observed in apoE-deficient mice treated with simvastatin is not the result of an increased number of remnant particles in circulation but is caused by synthesis and secretion into the plasma of lipoproteins that are enriched in cholesterol and depleted of triglycerides.     

Role of acyl-coenzyme A:cholesterol acyltransferase-1 in the control of hepatic very low density lipoprotein secretion and low density lipoprotein receptor expression in the mouse and hamster

Cholesteryl esters present in nascent very low density lipoproteins are generated in a reaction catalyzed by acyl CoA:cholesterol acyltransferase (ACAT). To examine the effect of cholesteryl esters on the secretion of apoB-containing lipoproteins, we transiently overexpressed human (h) ACAT-1 in the livers of low density lipoprotein (LDL) receptor(-/-) mice using adenovirus-mediated gene transfer. Overexpression of hACAT-1 increased hepatic total and esterified cholesterol but did not reduce hepatic free cholesterol due to a compensatory increase in the rate of de novo cholesterol synthesis. Overexpression of hACAT-1 markedly increased the plasma concentration and hepatic secretion of apoB-containing lipoproteins but had no effect on the clearance of very low density lipoprotein-apoB from plasma indicating that cholesteryl esters play an important role in regulating the assembly and secretion of apoB-containing lipoproteins. ACAT activity has been implicated in the regulation of the LDL receptor pathway by dietary fatty acids. It has been hypothesized that unsaturated fatty acids, by enhancing ACAT activity, reduce the amount of free cholesterol in a putative regulatory pool that feeds back on LDL receptor expression. We directly tested this hypothesis in hamsters by transiently overexpressing hACAT-1 in the liver. Enhanced cholesterol esterification in the liver resulted in a compensatory increase in de novo cholesterol synthesis but no induction of LDL receptor expression suggesting that fatty acids regulate LDL receptor expression via a mechanism independent of ACAT.     

Chinese hamster ovary cells require the coexpression of microsomal triglyceride transfer protein and cholesterol 7alpha-hydroxylase for the assembly and secretion of apolipoprotein B-containing lipoproteins

Due to the absence of microsomal triglyceride transfer protein (MTP), Chinese hamster ovary (CHO) cells lack the ability to translocate apoB into the lumen of the endoplasmic reticulum, causing apoB to be rapidly degraded by an N-acetyl-leucyl-leucyl-norleucinal-inhibitable process. The goal of this study was to examine if expression of MTP, whose genetic deletion is responsible for the human recessive disorder abetalipoproteinemia, would recapitulate the lipoprotein assembly pathway in CHO cells. Unexpectedly, expression of MTP mRNA and protein in CHO cells did not allow apoB-containing lipoproteins to be assembled and secreted by CHO cells expressing apoB53. Although expression of MTP in cells allowed apoB to completely enter the endoplasmic reticulum, it was degraded by a proteolytic process that was inhibited by dithiothreitol (1 mM) and chloroquine (100 microM), but resistant to N-acetyl-leucyl-leucyl-norleucinal. In marked contrast, coexpression of the liver-specific gene product cholesterol 7alpha-hydroxylase with MTP resulted in levels of MTP lipid transfer activity that were similar to those in mouse liver and allowed intact apoB53 to be secreted as a lipoprotein particle. These data suggest that, although MTP-facilitated lipid transport is not required for apoB translocation, it is required for the secretion of apoB-containing lipoproteins. We propose that, in CHO cells, MTP plays two roles in the assembly and secretion of apoB-containing lipoproteins: 1) it acts as a chaperone that facilitates apoB53 translocation, and 2) its lipid transfer activity allows apoB-containing lipoproteins to be assembled and secreted. Our results suggest that the phenotype of the cell (e.g. expression of cholesterol 7alpha-hydroxylase by the liver) may profoundly influence the metabolic relationships determining how apoB is processed into lipoproteins and/or degraded.     

Effect of a single and fractionated gamma-irradiation on lipid content in rat blood and tissues]

Contents of lipids, character of blood plasma chemiluminescence and separate classes of lipoproteins as well as the content of the lipids of tissues under single and fractionated gamma-irradiation of rats in a dose of 3 Gr. The development of hyperlipidemia and hypercholesterolemia for four weeks after irradiation which are more expressed under a single irradiation, a decrease of total content of cholesterol and triglycerides in the heart and liver tissues to the end of the experiment 24 hour after irradiation are registered in the both groups of animals.     

Plasma lipoproteins and cholesterol metabolism in Yoshida rats: an animal model of spontaneous hyperlipemia.

The purpose of this study was to characterize the lipoprotein profile and cholesterol metabolism in Yoshida rats, a strain of inbred genetically hyperlipemic animals. For comparison, Brown Norway rats were used as control animals. Plasma cholesterol and triglycerides were higher in Yoshida as compared to Brown Norway, the elevation of cholesterol being due to a rise in HDL fraction. Triglyceride distribution among lipoproteins showed an increase in VLDL fraction. Hyperlipemia was not related to diabetes, hypothyroidism or nephropathy. Plasma triglycerides production was increased in Yoshida rats, while lipoprotein and hepatic lipases were similar in the two groups. Hypercholesterolemia was associated with a defect of lipoprotein receptor activity and with elevated HMG-CoA reductase and cholesterol 7 alpha - hydroxylase; conversely ACAT activity was lower in Yoshida as compared to Brown Norway rats. Sterol fecal excretion was comparable in the two groups and hypercholesterolemia in Yoshida rats was not associated to an increase of cholesterol saturation of the bile. We suggest that lipoprotein overproduction is the main cause for hyperlipidemia in this strain of rats.