Further characterization of the changes occurring in the plasma lipoprotein spectrum in the European badger (Meles meles L.) during winter.

The plasma lipoprotein pattern in the European badger has been shown previously to undergo marked and complex quantitative and qualitative seasonal modifications (Laplaud, P.M. et al., 1980, J. Lipid Res., 21, 724-738). However, the conventional ultracentrifugal techniques then in use in our laboratory were of insufficient discriminating power with regard to the numerous lipoprotein fractions whose presence was suggested by our analyses. In the present study, a new density gradient ultracentrifugation procedure was applied to the more detailed determination of the distribution of plasma lipoproteins. The first series of analyses was performed in early December and the second in March, i.e. at the dates when the maximum and minimum, respectively, of lipidemia occur in this species. The fractions thus obtained, each of which corresponded to a narrow density interval, were analyzed subsequently for chemical composition, appearance upon polyacrylamide gel electrophoresis, and for their content of tetramethylurea-soluble apolipoproteins in alkaline-urea gels. Changes occurring from December to March included a large decrease in the plasma concentration of the 1.015-1.065 g/ml lipoproteins, chemical analysis of this material being compatible with the presence of at least two lipoprotein populations. On the other hand, high-density lipoproteins (1.065-1.162 g/ml) appeared less variable in chemical composition, although the proportion of those with lower density decreased considerably in early spring. Polyacrylamide gel electrophoresis of the native fractions showed multiple bands in most of them; the tetramethylurea-soluble apoprotein profile remained similar at the two dates considered with an apolipoprotein A-I-like component present in large amounts throughout the entire low- and high-density ranges.

Distribution and partial characterization of the plasma lipoproteins in the hedgehog (Erinaceus europaeus L.), a hibernator with potential use in the study of the hormonal regulation of lipoprotein metabolism.

The hedgehog is a hibernator in which yearly cycles of several endocrine activities and seasonal variations of plasma lipids have already been demonstrated. We have consequently undertaken a study of plasma lipids and lipoproteins in this animal, bled during late April and May. Plasma cholesterol levels (178 +/- 30 mg/100 ml) were comparable with those in normal humans, while triacylglycerol was lower (46 +/- 17 mg/100 ml) and phospholipids higher (252 +/- 35 mg/100 ml). The main characteristics of the plasma lipoprotein spectrum, as determined by sequential and density gradient preparative ultracentrifugation, analytical ultracentrifugation and gel filtration chromatography, were (1) a low concentration of very-low-density components (d less than 1.006 g/ml, about 20 mg/100 ml); (2) a continuity between the low (1.006-1.063 g/ml) and high (1.063-1.21 g/ml) density components, the former (about 150 mg/100 ml) exhibiting a considerable heterogeneity upon polyacrylamide gel electrophoresis while the latter were largely predominating (570 mg/100 ml); (3) the presence, at a density of 1.087 g/ml, of a band migrating electrophoretically like human low-density lipoproteins, a finding consistent with the results of apolipoprotein electrophoresis, which showed the presence of a high-molecular-weight counterpart to apolipoprotein B, in both the low- and high-density ranges defined above; (4) the presence, throughout the entire density spectrum, of an apolipoprotein with molecular weight and mobility in polyacrylamide/urea gels similar to human apolipoprotein A-I, and (5) the presence of very-high-density lipoproteins (d 1.178-1.259 g/ml) responsible for the transport of approximately 15% of plasma cholesterol and 20% of phospholipids.

Seasonal variations of plasma lipids and lipoproteins in the hedgehog, an animal model for lipoprotein (a) metabolism: relation to plasma thyroxine and testosterone levels.

We describe a study of the seasonal variations of hedgehog plasma lipids and lipoproteins and their correlation with changes in the activities of the thyroid and testis. In ten male hedgehogs, plasma concentrations of lipids, thyroxine and testosterone were assayed each month for 1 year beginning in September, while plasma lipoproteins from five of these animals were analyzed at the same dates using density gradient ultracentrifugation. All classes of plasma lipids (cholesterol, total glycerol and phospholipids) exhibited statistically significant seasonal variations in their respective concentrations, with simultaneous maxima (cholesterol: 207 +/- 39 mg/100 ml; total glycerol: 50 +/- 9 mg/100 ml; phospholipids: 266 +/- 25 mg/100 ml) during late fall-early winter, i.e., during the period of the year when plasma levels of both thyroxine and testosterone were minimal. Plasma lipids subsequently decreased to minimal levels either in early summer (cholesterol: 129 +/- 18 mg/100 ml; phospholipids: 178 +/- 20 mg/100 ml) or in late winter (total glycerol: 22 +/- 9 mg/100 ml). Very low density lipoproteins (d less than 1.015 g/ml) were found at low levels (less than 15 mg/100 ml) during the cold months, and then became detectable as trace components only. The total concentration of the mixed lipoprotein population (i.e., low density lipoproteins, Lp(a), and high density lipoprotein (HDL)-like particles) in the d 1.015-1.065 g/ml interval decreased by almost 50% from January to February (from 164.3 to 89.2 mg/100 ml), i.e., following a 10-fold increase in the level of plasma testosterone, and immediately before the rapid doubling in plasma thyroxine concentration. The staining intensity of the electrophoretic band with migration characteristics corresponding to those of Lp(a) decreased considerably during winter. At the same period of the year, lower density (1.032-1.055 g/ml) HDL-like particles disappeared. The concentration of lipoproteins with d 1.065-1.162 g/ml, which included Lp(a) particles in addition to typical HDL, equally underwent seasonal variations. These variations consisted of two successive maxima in late fall (426.4 mg/100 ml) and late winter (458.3 mg/100 ml) with two subsequent decreases leading to minima in February (327.8 mg/100 ml) and August (257.1 mg/100 ml). Finally, very high density lipoproteins (d 1.162-1.259 g/ml) were heterogeneous, containing both cholesterol-rich (d 1.162-1.227 g/ml) and phospholipid-rich (d 1.194-1.259 g/ml) subpopulations.(ABSTRACT TRUNCATED AT 400 WORDS)

Plasma lipid transport in the preruminant calf, Bos spp: primary structure of bovine apolipoprotein A-I.

The preruminant calf (Bos spp.) is a model of considerable interest with regard to hepatic and intestinal lipoprotein metabolism (Bauchart et al., J. Lipid Res. (1989) 30, 1499-1514 and Laplaud et al., J. Lipid Res. (1990) 31, 1781-1792). As a preliminary step towards future experiments dealing with HDL metabolism in the calf, we have purified apoA-I from this animal and determined its complete amino acid sequence. Thus, approx. 10% of calf apoA-I was shown to contain a propeptide, with the sequence Arg-His-Phe-Trp-Gln-Gln. Enzymatic cleavage of apoA-I resulted in 10 proteolytic peptides. The complete apoA-I sequence was obtained after alignment of peptides on the basis of their homologies with those from rabbit apoA-I. Thus calf apoA-I consists of 241 amino acid residues, and exhibits high sequence homology with all mammalian apoA-I's studied to date. The bovine protein contained 10 hydrophobic amphipathic helical regions, occurring between residues 43-64, 65-86, 87-97, 98-119, 120-141, 142-163, 164-184, 185-206, 207-217 and 218-241. A computer-constructed phylogenetic tree showed that bovine apoA-I was more closely related to its dog counterpart, including the presence of a single methionine, than to the corresponding macaque and human proteins. Comparative predictions of the respective antigenic structures of human and bovine apoA-I's using the Hopp-Woods algorithm indicated similar positions for all 13 detectable antigenic sites, among which 7 were of identical, or closely related, amino acid composition. This finding was confirmed by demonstration of partial immunological identity between the two proteins upon immunodiffusion analysis, a result obtained using a monospecific rabbit antiserum against bovine apoA-I. Finally, comparison of sequence homology between bovine apoA-I and the lecithin:cholesterol acyl transferase (LCAT) activating region of human apoC-I suggests that several LCAT activating domains may be present in calf apoA-I.

Adenovirus-mediated overexpression of tissue inhibitor of metalloproteinase-1 reduces atherosclerotic lesions in apolipoprotein E-deficient mice.

BACKGROUND: To define the role of metalloproteinases (MMPs) in the development of lipid-rich atherosclerotic lesions in relation to the balance between proteolytic and antiproteolytic activities, we investigated the impact of adenovirus-mediated elevation in the circulating levels of human tissue inhibitor of MMP (TIMP-1) in atherosclerosis-susceptible apolipoprotein E-deficient (apoE(-/-)) mice. METHODS AND RESULTS: Infusion of apoE(-/-) mice fed a lipid-rich diet with rAd.RSV.TIMP-1 (1x10(11) viral particles) resulted in high hepatic expression of TIMP-1. At 2 weeks after injection, plasma TIMP-1 levels ranged from 7 to 24 micrograms/mL (mean 14.8+/-6.8). Marked overexpression of TIMP-1 was transient, with levels of TIMP-1 decreasing to 2.5 to 8 micrograms/mL (mean 4.3+/-2.1) at 4 weeks. Plasma lipid and lipoprotein levels in mice treated with rAd.RSV.TIMP-1 were similar to those treated with rAd.RSV.betaGal. However, rAd.RSV.TIMP-1-infused mice displayed a marked reduction (approximately 32%; P<0.05) in mean lesion area per section (512+/-121 micrometers(2)x10(3); n=12 sections from 4 animals) as compared with rAd.RSV.betaGal-infused mice (750+/-182 micrometers(2)x10(3); n=12 sections from 4 animals). Similarly, marked reduction in macrophage deposition as well as MMP-2, MMP-3, and MMP-13 antigens was observed. CONCLUSIONS: Histological and immunohistologic analyses of atherosclerotic lesions revealed increases in collagen, elastin, and smooth muscle alpha-actin content in mice treated with rAd.RSV.TIMP-1. These qualitative and quantitative features were the consequence of TIMP-1 infiltration from plasma to arterial intima, as immunohistochemical analyses revealed an abundance of TIMP-1 specifically in lesions of rAd.RSV. TIMP-1-treated mice.

The complete cDNA sequence encoding plasminogen from the European hedgehog (Erinaceus europaeus).

Using a PCR-based strategy, we have determined the complete cDNA sequence encoding hedgehog plasminogen (Plg). The 2700-nucleotide cDNA (corresponding to a 2.9-kb liver-derived transcript) encodes an open reading frame of 811 amino acids which shares 74-76% identity with Plg characterized from mouse, human and rhesus monkey. Residues corresponding to the catalytic triad, tPA-cleavage site, as well as seven of the eight lysine-binding residues in kringle IV are conserved in the hedgehog. However, potential N-linked glycosylation sites which have been reported in human and rhesus Plg are not present in analogous positions in the hedgehog Plg sequence.

A year-long study of changes induced by thyroidectomy in the plasma lipid and lipoprotein spectrum in the European badger.

Hypothyroidism is associated with hypercholesterolemia and increased risk for atherosclerotic disease. The European badger exhibits large seasonal changes in thyroid activity and the annual minimum of plasma thyroxine level in this species occurs at the same period of the year (i.e. late fall) as a pronounced hypercholesterolemia. We examined the plasma lipid and lipoprotein spectrum in a group of thyroidectomized male badgers every month for a year. Non-operated animals were used as controls. Our analyses included measurement of plasma lipid levels, density gradient ultracentrifugation of lipoproteins, electrophoresis of lipoproteins and apolipoproteins, and histological studies. Maximal differences between the two groups of animals were observed during spring, occurring concomitantly with the annual maximum of plasma thyroxine concentration in control badgers. Comparison with the latter animals revealed a permanent hypercholesterolemia and hyperphospholipidemia in thyroidectomized badgers, while their lipoprotein spectrum was characterized by the continual presence of elevated concentrations of cholesterol-rich lipoproteins of d congruent to 1.015 - 1.027 g/ml. The ratio of triglyceride/cholesteryl ester content in such lipoproteins remained constant throughout the year, resembling that noted in intact animals during late fall. Other features distinguishing the lipoprotein spectrum in thyroidectomized badgers were: (1) higher levels of lipoproteins with d 1.027 - 1.065 g/ml and d 1.065 - 1.100 g/ml, and (2) a cholesteryl ester enrichment of both these lipoprotein subclasses. The two groups of animals shared a heterogeneity of low density lipoprotein subfractions isolated on density gradients, together with the presence of apolipoproteins with molecular weights respectively typical of human apolipoproteins A-I and B throughout the low density range. Arterial walls and heart tissues from intact and thyroidectomized animals were free of atherosclerotic lesions at the end of the experimental period.

A year-long study of changes induced by castration in the plasma lipid and lipoprotein spectrum in the European badger.

In man, an influence of male sex hormones on plasma lipid transport is well established; however, recent data on this subject in the literature are both relatively lacking and occasionally conflicting. The male European badger exhibits seasonal variations of large amplitude in its gonadic function. We have therefore attempted to establish the influence of male sex steroids on plasma lipids and lipoproteins in this species. For this purpose, we have examined the plasma lipid and lipoprotein spectrum in a group of castrated male badgers every month for a year, non-operated animals being used as controls. Our analyses included measurement of plasma lipid levels, density gradient ultracentrifugation of lipoproteins, electrophoresis of lipoproteins and apolipoproteins, and evaluation of plasma testosterone and thyroxine levels. The differences observed between the 2 groups of animals were maximal during the months when plasma testosterone was elevated in intact badgers (January to July). For this period, castration resulted in higher plasma concentrations of cholesterol, phospholipids and triglycerides, while the latter alone remained significantly more elevated in operated animals until the end of our experiments. With regard to lipoproteins, the main effect of castration consisted of a large augmentation in the concentration of lipoproteins with d approximately equal to 1.027-1.065 g/ml which were responsible for the transport of most of the increased amounts of triglycerides present in the plasma of castrated badgers. The proportion of apoprotein B in the protein moiety of these lipoprotein components was enhanced after castration. Other changes in the lipoprotein spectrum included (1) a moderate increase in the concentration of lipoproteins with d less than 1.015 g/ml and 1.019-1.027 g/ml, and (2) a modification of the respective proportions of high density lipoproteins with d 1.065-1.100 g/ml and d 1.100-1.162 g/ml. Finally, no considerable differences between the 2 groups of animals were noted in the respective percentages of the various chemical constituents in each lipoprotein subfraction assayed, except for those with d 1.023-1.027 g/ml, which, in castrated badgers, did not exhibit the enrichment in triglycerides usually noted during late winter and spring in intact animals.

Effects of soy protein and casein in low cholesterol diets on plasma lipoproteins in normolipidemic subjects.

Dietary plant proteins may lower plasma cholesterol and LDL concentrations in hypercholesterolemic patients when substituted for animal proteins, particularly in diets with low cholesterol and saturated fat content. Plant protein diets appear, however, to be without effect on plasma lipoprotein levels in normal subjects. In the present study, we have examined whether the origin of the dietary protein, i.e. plant (soy) or animal (casein), affects the plasma lipoproteins in normolipidemic subjects when these proteins are presented as components of diets low in cholesterol and saturated fat. The study followed a crossover design. Five men and 5 women consumed liquid formula diets containing 20% of calories as casein or soy protein, 28% as fat (mainly monounsaturated), and 52% as carbohydrate; the intake of cholesterol was less than 100 mg per day. The two dietary periods, each of 1 month duration, were separated by an interim period of 1 month on self-chosen food. Following an initial 30% reduction of cholesterol and LDL plasma levels on both diets, the concentrations of each of the major lipoprotein classes (VLDL, IDL, LDL, HDL2 and HDL3) were similar during the two experimental dietary periods. Body weights were essentially constant. Dietary soy protein and casein could not be distinguished in their effects on the plasma concentrations and chemical composition of the major lipoprotein classes in normolipidemic subjects.

Antioxidant action of Vaccinium myrtillus extract on human low density lipoproteins in vitro: initial observations.

Oxidative modifications of low density lipoproteins (LDL) are now recognised as one of the major processes in atherogenesis. Various drugs, as well as a number of natural products, have been proposed to inhibit such processes. Among the naturally-occurring constituents of plants which appear to possess antioxidant activity are polyphenolic compounds such as flavonoids. The aqueous extract of Vaccinium myrtillus is rich in such molecules. In this report, we describe the in vitro antioxidative potential of this extract on human LDL. The copper-induced oxidative modification of these lipoproteins was assessed using 1) measurement of oxidative resistance as determined by the lag-phase preceding conjugated diene formation; 2) quantification of the amount of lipoperoxides and thiobarbituric acid-reactive substances generated, and measurement of the modification in the net negative electrical charge of the lipoproteins, over a 7-hour time course experiment. Trace amounts of V myrtillus extract (15 to 20 micrograms/mL) induce statistically significant changes in the oxidation behaviour of LDL, which include 1) prolongation of the lag-phase of conjugated diene production (P < 0.01); 2) reduction in the formation of lipoperoxides and of thiobarbituric acid-reactive substances up to 7 hours and especially between 1 and 5 hours (P < 0.01); and 3) inhibition of modification in the net negative charge of LDL. These results demonstrate that V myrtillus extract exerts potent protective action on LDL particles during in vitro copper-mediated oxidation. Calculation of IC50 values indicates that, on a molar basis, this extract may indeed be more potent than either ascorbic acid or butylated hydroxytoluene in the protection of LDL particles from oxidative stress.

Effects of dietary n-6 or n-3 polyunsaturated fatty acids protected or not against ruminal hydrogenation on plasma lipids and their susceptibility to peroxidation in fattening steers.

Two experiments were conducted using crossbred Salers x Charolais fattening steers fed diets enriched with no supplemental oilseeds or oils rich in either n-6 PUFA (from sunflower seeds) or n-3 PUFA (from linseeds) provided either as seeds incorporated in the diet (i.e., not protected from ruminal bacterial hydrogenation) or by chronic infusion into the duodenum (protected form). In the Sunflower experiment, animals (initial age = 454 +/- 20 d; initial BW = 528 +/- 36 kg) received a control diet for 70 d (CS, n = six) consisting of hay and concentrate, or the same basal diet supplemented with sunflower oil (4% of dietary DM), either fed as seeds (SS, n = six) or infused into the duodenum (ISO, n = six). The same experimental design was applied to animals (initial age = 412 +/- 33 d; initial BW = 536 +/- 33 kg) used in the Linseed experiment (CL, LS, and ILO; n = 8 per group). For all animals, blood was sampled every 15 d during 70 d. In both trials, a significant diet x time interaction (P < 0.001) was detected for plasma concentrations of apolipoprotein A-I, phospholipids, and free and esterified cholesterol, with values increasing with time during administration of the PUFA-rich diets being more evident with ISO and ILO diets. Plasma fatty acids were altered with oil infusions, with increased concentrations of n-6 (1.6-fold; P < 0.05) and n-3 PUFA (4.5-fold; P < 0.05) and of their respective indicies of peroxidizability (1.2- and 1.5-fold with Diets ISO and ILO, respectively; P < 0.05). In vitro copper-induced peroxidation of lipids revealed a decreased length of the lag phase in the process of conjugated diene generation by 48% (P < 0.005) with the ILO diet, indicating less resistance against peroxidation than in control steers. Compared with CS, the ISO treatment increased plasma alpha-tocopherol (x2.5; P < 0.05) leading to similar resistance against peroxidation. After depletion of this vitamin, the rates of peroxidation and production of conjugated dienes were greater (twofold; P < 0.05) with the ISO and ILO diets than with the others. In conclusion, infusion of sunflower or linseed oil into the duodenum altered the composition and distribution of plasma lipids and increased the plasma concentration of PUFA. The sensitivity of plasma PUFA to peroxidation depends on the plasma level of antioxidants, especially vitamin E, a nutrient important both for the health of animals and for the stability of the blood lipids until their tissue deposit.

A spontaneously seasonal hypercholesterolemic animal: plasma lipids and lipoproteins in the European badger (Meles meles L.).

The European badger has previously been shown to exhibit yearly cycles of locomotor activity, endocrine secretions, and body weight, as well as seasonal variations in plasma cholesterol. Over a period of 2 years, we have followed the plasma levels of free and esterified cholesterol, triglycerides and phospholipids, and of plasma lipoproteins (by means of polyacrylamide gel electrophoresis, agarose column chromatography and preparative and analytical ultracentrifugation). Some preliminary observations on the qualitative characteristics of the plasma apoproteins, obtained by application of electrophoretic techniques, are also described. Our results provide evidence for considerable synchronous and spontaneous variations of each of the plasma lipid components studied, all of them reaching a maximum in late autumn/early winter, then decreasing to a minimum in early spring. In some animals, the amplitude of observed variations was as large as 650% for total cholesterol, 420% for phospholipids and 180% for triglycerides. While the plasma concentration of very low density lipoproteins (d < 1.006 g/ml) remained at low or moderate levels, major changes in the lipoprotein spectrum occurred in the low density (1.006--1.063 g/ml) and high density (1.063--1.21 g/ml) lipoproteins, these two classes exhibiting marked heterogeneity. This led to an autumn/winter prominence of the 1,006--1.063 g/ml components and of those in the lower part of the high density range, with an enrichment in cholesterol in lipoproteins in the low density region. These phenomena occur simultaneously and/or immediately after the annual minimum of plasma thyroxine concentration in the species considered. In contrast, early spring patterns displayed more classical features with higher density lipoproteins predominating. Our findings thus suggest that the badger may provide a useful model for future experiments regarding the hormonal regulation of plasma lipid transport as well as the metabolism and physiopathological implications of some cholesterol-rich lipoproteins.

Diet-induced and physiologically occurring hypercholesterolemias in the spontaneous hypothyroid European badger (Meles meles L.): a density gradient study of lipoprotein profile.

As previously shown in this laboratory (Laplaud, P. M. et al. J. Lipid Res. 1980. 21: 724-738), the European badger is, with regard to its plasma lipid transport system, an original and complex animal of great potential interest to lipoprotein research. In an effort to study the response of this animal to cholesterol feeding, we gave a diet supplemented with 1% cholesterol to six male badgers (group H) during the late fall period when spontaneous hypercholesterolemia and hypothyroidism occur. Six more male animals of similar age received the standard diet (group C) and were simultaneously used as controls. Plasma lipids were measured using enzymatic methodologies, while the use of a recently described density gradient ultracentrifugation technique allowed detailed examination of lipoprotein composition and polyacrylamide gel electrophoresis of lipoproteins and tetramethylurea-soluble apoproteins in the fractions. The results suggest the superimposition, in H badgers, of the spontaneous and diet-induced hypercholesterolemias, maximum levels being reached in December in both C and H groups. While the two groups were very similar at the beginning of the experiment, highly significant differences (P < 0.01) were subsequently observed between C and H animals in plasma cholesterol and phospholipid concentrations. Density gradient ultracentrifugation provided evidence for the following diet-induced changes in lipoprotein profile: 1) a twofold increase in cholesteryl esters in particles of d < 1.006 g/ml; 2) the occurrence of large amounts of supplementary cholesterol-rich low density lipoproteins, mainly in the 1.019-1.027 g/ml region; 3) an increase in the 1.039-1.055 g/ml low density lipoproteins; and 4) a change in the ratio of the concentrations of high density lipoproteins of d 1.065-1.100 g/ml and d 1.100-1.162 g/ml, to the benefit of the former. Electrophoresis of the density gradient fractions revealed marked heterogeneity, especially in the low density part of the spectrum. Electrophoresis of the low molecular weight, tetramethylurea-soluble apoproteins failed to show marked differences between C and H badgers. However, chromatographic determination of the proportion of apoB in the protein moiety of the two main low density components showed that 1) it was consistently low, 2) its contribution to the higher density fraction (d 1.039-1.046 g/ml) was unaffected by the hypercholesterolemic diet (being about 25% in both C and H animals), and 3) its contribution to the lower density fraction (d 1.019-1.027 g/ml) decreased under the same nutritional conditions, representing about 20% in C as compared to about 10% in H badgers.-Laplaud, P. M., Beaubatie, and D. Maurel. Dietinduced and physiologically occurring hypercholesterolemias in the spontaneous hypothyroid European badger (Meles meles L.): a density gradient study of lipoprotein profile.

Paraoxonase as a risk marker for cardiovascular disease: facts and hypotheses.

Paraoxonase (PON1) is a Ca2+-dependent enzyme whose mechanism of action is incompletely elucidated. PON1 was originally found to be responsible for the hydrolysis of paraoxon, a catabolite of the insecticide parathion, but this enzyme is equally able to hydrolyze other substrates such as phenyl acetate. PON1 exhibits two sequence polymorphisms, Arg-->Gln 192 and Met-->Leu 55, respectively, of which the former is responsible for the distinct catalytic activity of the two corresponding allozymes against paraoxon. The PON1 gene is a member of a family of at least three related genes. Although the physiologic substrate of PON1 is unknown, a protective role against the oxidative degradation of serum lipoproteins has been attributed to this enzyme. Indeed, PON1 is a component of a spectrum of circulating high density lipoprotein particles and can hydrolyze oxidized phospholipids and cholesteryl ester hydroperoxides. Studies have been conducted to evaluate the possible "protective" role of PON, and especially the influence of the Arg-->Gln 192 polymorphism, in coronary artery disease. Results from these investigations are conflicting, and recent data suggest a complex pattern with influences from other polymorphisms in either the PON1 and/or the PON2 and PON3 genes, or even another region of the gene cluster. A number of related factors, which include the heterogeneity of the high density lipoprotein particles incorporating PON(s), the metabolism of associated apolipoproteins such as apoJ/clusterin, the respective roles of PON(s) and other high density lipoprotein-associated enzymes such as platelet-activating-factor acetyl-hydrolase and lecithin-cholesterol acyltransferase, modifications of high density lipoprotein composition and activity under acute-phase conditions, the dietary and environmental regulation of PON(s), and the actual in situ availability of PON in the atherosclerotic artery wall, must equally be taken into account.

Lipoprotein[a] is the major apoB-containing lipoprotein in the plasma of a hibernator, the hedgehog (Erinaceus europaeus).

We have undertaken studies aimed at elucidating the interrelationships existing between the seasonal modifications in endocrine status (already demonstrated by Saboureau, M., and J. Boissin. 1978. C.R. Acad. Sci. (Paris) 286D: 1479-1482) and plasma lipoprotein metabolism in the male hedgehog. During the course of these studies, we discovered that a lipoprotein comparable to human Lp[a] was a prominent component of the plasma lipoprotein spectrum in the hedgehog. This lipoprotein was present in the 1.040-1.100 g/ml density range (approximately), exhibited pre beta mobility upon agarose gel electrophoresis, and its Stokes diameter was 275 A. Its apolipoprotein moiety consisted of two proteins with molecular weights and amino acid compositions similar to those of human apoB-100 and apo[a], respectively. These two apolipoproteins were present in hedgehog Lp[a] as a complex that could be dissociated using dithiothreitol and whose stoichiometry could be 1:1. Lp[a] polymorphism due to size heterogeneity of apo[a] appeared to be present in the hedgehog as in man. The chemical composition of hedgehog Lp[a], obtained from animals bled during spring and summer, differed from that of its human counterpart in that the proportion of triglycerides was approximately three times higher in the hedgehog particle (13% vs. 4%), to the detriment of cholesteryl esters. Dissociation of the apoB:apo[a] complex has allowed us to obtain Lp[a] devoid of its specific polypeptide (Lp[a-]), a particle that retained the characteristics of Lp[a] as regards its lipid composition but whose Stokes diameter decreased by 30 to 40 A. The plasma concentration of LDL particles, defined as lipoproteins containing apoB-100 as their sole apolipoprotein constituent, was considerably lower than that of Lp[a]. These findings suggest that the hedgehog could be a unique animal model for studies regarding Lp[a] metabolism.

Density distribution and physicochemical properties of plasma lipoproteins and apolipoproteins in the goose, Anser anser, a potential model of liver steatosis.

The fractionation and physicochemical characterization of the complex molecular components composing the plasma lipoprotein spectrum in the goose, a potential model of liver steatosis, are described. Twenty lipoprotein subfractions (d less than 1.222 g/ml) were separated by isopycnic density gradient ultracentrifugation, and characterized according to their chemical composition, particle size and particle heterogeneity, electrophoretic mobility, and apolipoprotein content. Analytical ultracentrifugal analyses showed high density lipoproteins (HDL) to predominate (approximately 450 mg/dl plasma), the peak of its distribution occurring at d approximately 1.090 g/ml (F1.21 approximately 2.5). The HDL class displayed marked density heterogeneity, HDL1-like particles being detected up to a lower density limit of approximately 1.020 g/ml, particle size decreasing progressively from 17-19 nm at d 1.024-1.028 g/ml to 10.5-12 nm (d 1.055-1.065 g/ml), and then remaining constant (approximately 9 nm) at densities greater than 1.065 g/ml. HDL subfractions displayed multiple size species; five subspecies were present over the range d 1.103-1.183 g/ml with diameters of 10.5, 9.9, 9.0, 8.2, and 7.5 nm, four in the range d 1.090-1.103 g/ml (diameters 10.5, 9.9, 9.0, and 8.2 nm) and three over the range d 1.076-1.090 g/ml (diameters 10.5, 9.9, and 9.0 nm). ApoA-I (Mr 25,000-27,000) was the major apolipoprotein in all goose HDL subfractions, while the minor components (apparent Mr 100,000, 91,000, 64,000, 58,000, approximately 42,000, 18,000 and apoC-like proteins) showed marked quantitative and qualitative variation across this density range (i.e., 1.055-1.165 g/ml). The d 1.063 g/ml boundary for separation of goose low density lipoproteins (LDL) from HDL was inappropriate, since HDL-like particles were present in the density interval 1.024-1.063 g/ml, while particles enriched in apoB (Mr approximately 540,000) and resembling LDL in size (approximately 20.5 nm) were detected up to a density of approximately 1.076 g/ml. Goose LDL itself was a major component of the profile (90-172 mg/dl) with a single peak of high flotation rate (Sf approximately 10.5). The physicochemical properties and apolipoprotein content of intermediate density lipoproteins (IDL) and LDL varied but little over the range d 1.013-1.040 g/ml, presenting as two particle species (diameters 20.5 and 21 nm) of essentially constant chemical composition; LDL (d 1.019-1.040 g/ml) were separated from HDL1 by gel filtration chromatography and appeared to contain primarily apoB with lesser amounts of apoA-I.(ABSTRACT TRUNCATED AT 400 WORDS)

Plasma lipoproteins and apolipoproteins in the preruminant calf, Bos spp: density distribution, physicochemical properties, and the in vivo evaluation of the contribution of the liver to lipoprotein homeostasis.

The in vivo role of the liver in lipoprotein homeostasis in the preruminant calf, a functional monogastric, has been evaluated. To this end, the hydrodynamic and physicochemical properties, density distribution, apolipoprotein content, and flow rates of the various lipoprotein particle species were determined in the hepatic afferent (portal vein and hepatic artery) and efferent (hepatic vein) vessels in fasting, 3-week-old male preruminant calves. Plasma lipoprotein profiles were established by physicochemical analyses of a series of subfractions isolated by isopycnic density gradient ultracentrifugation. Triglyceride-rich very low density lipoproteins (VLDL) (d less than 1.018 g/ml) were minor plasma constituents (approximately 1% or less of total d less than 1.180 g/ml lipoproteins). The major apolipoproteins of VLDL were apoB-like species, while the complement of minor components included bovine apoA-I and apoC-like peptides. Particles with diameters (193-207 A) typical of low density lipoproteins (LDL) were present over the density interval 1.026-1.076 g/ml; however, only LDL of d 1.026-1.046 g/ml were present as a unique and homogeneous size subspecies, containing the two apoB-like species as major protein components in addition to elevated cholesteryl ester contents. LDL represented approximately 10% of total d less than 1.180 g/ml lipoproteins in fasting plasma from all three hepatic vessels. Overlap in the density distribution of particles with the diameters of LDL and of high density lipoproteins (HDL) occurred in the density range from 1.046 to 1.076 g/ml; these HDL particles were 130-150 A in diameter. HDL were the major plasma particles (approximately 90% of total d less than 1.180 g/ml substances) and presented as two distinct populations which we have termed light (HDLL) and heavy (HDLH) HDL. Light HDL (d 1.060-1.091 g/ml) ranged in size from 120 to 140 A, and were distinguished by their high cholesteryl ester (29-33%) and low triglyceride (1-3%) contents; apoA-I was the principal apolipoprotein. Small amounts of apolipoproteins with Mr less than 60,000, including apoC-like peptides, were also present. Heavy HDL (d 1.091-1.180 g/ml) accounted for almost half (47%) of total calf HDL, and like HDLL, were also enriched in cholesteryl ester and apoA-I; they ranged in size from 93 to 120 A. The protein moiety of HDLH was distinct in its possession of an apoA-IV-like protein (Mr 42,000). Blood flow rates were determined by electromagnetic flowmetry, thereby permitting determination of net lipoprotein balance across the liver. VLDL were efficiently removed during passage through the liver (net uptake 1.06 mg/min per kg body weight).(ABSTRACT TRUNCATED AT 400 WORDS)

Lipoproteins and apolipoproteins in intestinal lymph of the preruminant calf, Bos spp., at peak lipid absorption.

We have recently evaluated the in vivo role of the liver in lipoprotein homeostasis in the preruminant calf (Bauchart, D., D. Durand, P. M. Laplaud, P. Forgez, S. Goulinet, and M. J. Chapman, 1989. J. Lipid Res. 30: 1499-1514). We now present the partial characterization of lipoprotein particles in postprandial intestinal lymph at peak lipid absorption (i.e., 10 h after a meal) in the preruminant calf fed a curdled milk replacer. Intestinal lymph from four male preruminant calves was analyzed for its content of lipids and fractionated by sequential and density gradient ultracentrifugation into chylomicrons (Sf greater than 400), very low density lipoproteins (VLDL) (Sf less than 400; d less than 1.006 g/ml), and a series of lipoprotein subfractions with d greater than 1.006 g/ml. Postprandial lymph contained predominantly triglycerides (1099 +/- 611 mg/100 ml), with lesser amounts of phospholipids (197 +/- 107 mg/100 ml) and cholesterol (52 +/- 30 mg/100 ml). The most abundant particles were triglyceride-rich chylomicrons and VLDL which accounted for approximately 76% and approximately 19%, respectively, of total d less than 1.21 g/ml lipoproteins. As judged by negative stain electron microscopy, chylomicron particle diameters ranged from 650 to 2400 A, while VLDL were smaller and distributed over a distinct size range (340-860 A). These two lipoprotein classes each presented protein components with Mr comparable to those of human apoB-48, apoA-I, and C apoproteins, together with an Mr 52,000 protein resembling human beta 2-glycoprotein-I. In addition, VLDL exhibited a polypeptide with Mr approximately 61,000. Lymph lipoproteins with d greater than 1.006 g/ml consisted primarily (approximately 81% of total) of particles distributed over the 1.053-1.119 g/ml density range. Electrophoretic analysis of the latter lipoprotein fraction showed it to be heterogeneous, including particles with the migration characteristics of low and of high density lipoproteins, respectively. Subfractions in the d 1.053-1.076 g/ml range were dominated by particles with Stokes diameters typical of high density lipoproteins (HDL), but also contained three different populations of low density lipoprotein-like particles. The high molecular weight apolipoproteins in these same cholesteryl ester-rich (greater than 30% of lipoprotein mass) subfractions comprised components with Mr resembling those of human apoB-100 and apoB-48, respectively, and with the latter protein predominating to a varying degree. A counterpart to human apoA-I was the major protein component over the entire density range from d 1.053 to 1.119 g/ml.(ABSTRACT TRUNCATED AT 400 WORDS)