Triacylglycerol and very low density lipoprotein secretion into plasma of squirrel monkeys.

We determined the effects of varying the types and level of dietary fat and cholesterol on the increase in plasma total triacylglycerol concentrations after injection of Triton WR-1339, an inhibitor of lipoprotein lipase, into monkeys that had been subjected to an overnight fast. The monkeys that had been treated with Triton WR-1339 were then given a test meal by intragastric intubation. Dietary cholesterol, high levels of fat and saturated fat in the habitual diet reduced the rate of release of triacylglycerol to plasma in the fasted monkey. We also determined the changes in protein and lipid concentrations of the different lipoprotein fractions. The injection of Triton WR-1339 resulted in a linear increase with time in the concentration of protein and triacylglycerol in the very low density (chylomicron-free and d less than 1.006) lipoproteins, but there was an increase in the ratio of traicylglycerol to protein in that fraction. Most of the increase (96%) in very low density protein was in the B protein. Regardless of the habitual diet, a test meal accentuated the rate of triacylglycerol appearance in whole plasma and in the very low density lipoproteins of Triton WR-1339-treated monkeys, and the rate of increase of the protein component after feeding was slightly higher. Thus the administration of a meal to the fasted Triton WR-1339-treated squirrel monkey further increased the proportion of triacylglycerol in very low density lipoproteins. Although dietary cholesterol and saturated fat in the habitual diet depressed the rate of increase in very low density triacylglycerol during fasting, the rate of protein synthesis was not significantly affected. After administration of a test meal the rates of increase in triacylglycerol and protein in the very low density lipoproteins were similar for monkeys from the different diet groups. Triton WR-1339 administration caused a slight and progressive increase in the intermediate density (d 1.006-1.019) lipoproteins and a marked and progressive decrease in the low density (d 1.019-1.063) lipoproteins. There was an immediate (by 5 min) drop of 70% or more in high density (d 1.063-1.21) lipoprotein protein, but the lipids except triacylglycerol remained unchanged. There was a decrease in both the A (the major fraction) and C proteins. The rates of very low density B protein secretion were comparable to the rates of low density lipoprotein catabolism that had been previously demonstrated for this species.     

The effects of hyperlipidemia on lipoprotein metabolism in squirrel monkeys and rabbits.

We studied the metabolism of different classes of lipoprotein in squirrel monkeys and rabbits. Lipoproteins were labeled in vivo in donor animals with (3H)leucine and (3H)cholesterol. The rate of disappearance from plasma of recipient squirrel monkeys of the protein moiety of the very low density lipoproteins was rapid, that of high density lipoproteins slow, and the rate for low density lipoproteins was intermediate. The fractional turnover of the apoprotein of low density lipoproteins was slightly reduced in hyperlipidemic monkeys, but the absolute rates of synthesis and catabolism were increased. Hyperdipidemia in rabbits resulted in a dramatic reduction in the fractional catabolic rate of low density lipoprotein apoprotein. Hyperlipidemia in the donors of biosynthetic low density lipoproteins also influenced the rates of catabolism in rabbits. We showed the cycloheximide that although there was recycling of (3H)leucine into other proteins, the reutilization of leucine from low density lipoproteins for nascent low density lipoproteins was not significant. In most tissues the ratio of cholesterol:protein radioactivity was much greater than that for plasma 24 h after administration of labeled low density lipoproteins, but the ratios for aortic intima plus inner media and for plasma low density lipoproteins were similar. The presence of atherosclerosis resulted in a large increase in the apparent uptake of low density lipoproteins by the aortas of rabbits and monkeys.     

Changes in the concentration of plasma lipoproteins and apoproteins following the administration of Triton WR 1339 to rats.

Changes in whole plasma and lipoprotien apoprotein concentrations were determined after a single injection of Triton WR 1339 into rats. Concentrations of apoproteins A-I (an activator of lecithin:cholesterol acyl transferase), arginine-rich apoprotein (ARP), and B apoprotein were measured by electroimmunoassay. The content of C-II apoprotein (an activaor of lipoprotein lipase) was estimated by the ability of plasma and lipoprotein fractions to promote hydrolysis of triglyceride in the presence of cow's milk lipase and also by isoelectric focusing on polyacrylamide gels. Apoproteins C-II and A-I were rapidly removed from high density lipoprotein (HDL) after Triton treatment and were recovered in the d 1.21 g/ml infranate fraction. A-I was then totally cleared from the plasma within 10--20 hr after injection. Arginine-rich apoprotein was removed from HDL and also partially cleared from the plasma. The rise in very low density lipoprotein (vldl) apoprotein that followed the removal of apoproteins from HDL was mostly antributed to the B apoprotein, although corresponding smaller increases were observed in VLDL ARP and C apoproteins. The triglyceride:cholesterol, triglyceride:protein, and B:C apoprotein ratios of VLDL more closely resembled nascent rather than plasma VLDL 10 hr after Triton injection. These studies suggest that the detergent may achieve its hyperlipidemic effct by disrupting HDL and thus removing the A-I and C-II proteins from a normal activating environment compirsing VLDL, HDL, and the enzymes. The possible involvement of intact HDL in VLDL catabolism is discussed in relation to other recent reports which also suggest that abnormalities of the VLDL-LDL system may be due to the absence of normal HDL.     

Swine lipoproteins and atherosclerosis. Changes in the plasma lipoproteins and apoproteins induced by cholesterol feeding.

Cholesterol feeding in miniature swine resulted in a hypercholesterolemia with a distinctive hyperlipoproteinemia and the subsequent development of atherosclerosis. Alterations in the type and distribution of plasma lipoproteins induced by cholesterol feeding were as follows: (a) the occurrence of beta-migrating lipoproteins (B-VLDL) as well as very low density lipoproteins in the d less than 1.006 ultracentrifugal fraction; (b) an increased prominence of the intermediate lipoproteins (d = 1.006-1.02); (c) an increased prominence of low density lipoproteins; and (d) the occurrence of a distinctive lipoprotein with alpha mobility which was referred to as HDLc (cholesterol induced). Characterization of the various plasma lipoproteins included chemical composition, size by electron microscopy, and apoprotein content. The B-VLDL resembled the beta-migrating lipoproteins of human Type III hyperlipoproteinemia and contained a prominent protein equivalent to the arginine-rich apoprotein in addition to the B apoprotein, apo-A-I, and the fast-migrating apoproteins (apo-C). The HDLc were rich in cholesterol, ranged in size from 100 to 240 A in diameter, and contained the arginine-rich apoprotein and apo-A0I but lacked the B apoprotein. The arginine-rich apoproteins isolated from B-VLDL and HDLc by gel chromatography were similar in amino acid analyses, with glutamic acid as their amino-terminal residue. The occurrence of a spectrum of cholesterol-rich lipoproteins which contained the arginine-rich apoprotein with the occurrence of accelerated atherosclerosis suggested an interesting, although speculative, association.     

The effects of dietary fat and cholesterol on the metabolism of plasma low density lipoprotein apoproteins in squirrel monkeys.

Low density lipoprotein apoproteins from squirrel monkeys (Saimiri sciureus) had characteristic 2-phase die-away curves in plasma. The kinetic constants were similar with three methods of labeling: in vitro with 125I by the iodine monochloride or the Bolton-Hunter methods or in vivo by the injection of [3H]-leucine into a donor animal. Dietary cholesterol and the type of dietary fat influenced the concentration of plasma cholesterol and low density lipoproteins. The fractional turnover of low density lipoprotein apoprotein was greaterin monkeys fed semipurified diets with safflower oil than in those on butter but was not influenced by dietary cholesterol. The total low density lipoprotein apoprotein turnover (the product of fractional turnover and plasma lipoprotein concentration) was highest in monkeys fed butter plus added cholesterol and lowest in those on safflower oil without cholesterol. Dietary safflower oil resulted in a smaller proportion of the total low density lipoprotein pool in the intravascular compartment than did butter, regardless of whether cholesterol was added.     

Ultrastructural and physiological evidence for corticosteroid-induced alterations in hepatic production of very low density lipoprotein particles

The cause of corticosteroid-induced hyperlipoproteinemia was studied in rats and mice. An ultrastructural morphometric method was utilized to demonstrate alterations in hepatocyte very low density lipoprotein content, and Triton WR 1339-treated rats were used to identify changes in the removal of very low density lipoproteins from plasma. The results show that corticosteroid treatment results in (1) an increase in both plasma triglyceride and cholesterol levels, (2) an increase in rate of accumulation of triglyceride after inhibition of very low density lipoprotein removal by Triton, and (3) an increase in the number and size of Golgi-associated very low density lipoprotein particles in hepatocytes. These combined results suggest that corticosteroids induce hyperlipoproteinemia through increased hepatic production of very low density lipoproteins.     

Intestinal lipoprotein synthesis. Comparison of nascent Golgi lipoproteins from chow-fed and hypercholesterolemic rats

Hypercholesterolemia, induced by a cholesterol-enriched diet, is associated with distinctive modifications in the serum lipoproteins of a variety of species. Present in the serum of these animals are several classes of lipoproteins enriched in cholesteryl esters and apolipoprotein E. To investigate the role of intestinal lipoprotein synthesis in diet-induced hypercholesterolemia, we characterized nascent lipoproteins retrieved from Golgi apparatus-rich fractions of intestinal epithelial cells from chow-fed control and hypercholesterolemic rats. To eliminate chylomicrons from the preparations, rats were fasted overnight prior to the experiments. Golgi very low density lipoproteins (d less than 1.006 g/ml) from control rats were triglyceride-rich lipoproteins that migrated slightly slower than pre-beta migrating serum very low density lipoproteins. These particles contained apoproteins B-240, A-IV, and A-I. Golgi very low density lipoproteins from hypercholesterolemic rats were likewise triglyceride-rich lipoproteins migrating electrophoretically like control Golgi very low density lipoproteins and they contained apoproteins B-240, A-IV, and A-I. However, these latter particles contained less triglyceride and more cholesterol compared to control Golgi very low density lipoproteins. In addition, by radioisotope incorporation studies, Golgi very low density lipoproteins from hypercholesterolemic rats contained relatively more apoprotein A-IV (21.6 vs. 11.0%) and less apoprotein B-240 (17.0 vs. 27.0%) than found in control Golgi very low density lipoproteins. Approximately 60% of the total apoprotein radioactivity was found in apoprotein A-I in both preparations. We conclude that intestinal lipoprotein synthesis is modified by diet-induced hypercholesterolemia. The significance of these modifications with respect to the marked hypercholesterolemia observed in these animals remains to be determined.     

Apoproteins of the lipoproteins in a nonrecirculating perfusate of rat liver.

The apoproteins of serum lipoproteins and of lipoproteins present in a nonrecirculating perfusate of rat liver were compared by immunochemical, gel electrophoretic, and solubility techniques. Serum and perfusate very low density lipoprotein apoprotein composition were not different. No evidence for the presence of a lipoprotein resembling serum low density lipoprotein was obtained. However, the apoprotein composition of circulatory high density lipoprotein was quantitatively different from the secretory product in the density 1.06-1.21 range. As measured by stained sodium dodecyl sulfate gel electrophoretic patterns, the arginine-rich protein was the major secretory apoprotein while the A-I protein was the major apoprotein in circulating high density lipoprotein. A very similar pattern was seen in perfusates of orotic acid-fatty livers. It was concluded that although the liver secrets lipoproteins in the high density class, circulatory high density lipoprotein is largely a product of catabolic processes.     

Characterization of the plasma lipoproteins and apoproteins of the Erythrocebus patas monkey.

Patas monkey lipoproteins were fractionated into four distinct classes by a combination of ultracentrifugation and Geon-Pevikon block electrophoresis and characterized with respect to their chemical and physical properties. Very low density lipoproteins (VLDL) were isolated at d is less than 1.006, were triglyceride rich, and were in the size range 300-850 A. They were similar in apoprotein content to the VLDL of man, dog, and swine. The Patas monkey low density lipoprotein referred to as LDL-I had beta mobility and a size which ranged from 190 to 240 A in diameter. Their chemical composition and apoprotein content were similar to those of human LDL. A second low density lipoprotein referred to as LDL-II occurred at a density of 1.05-1.085, ranged in size from 190 to 300 A, and contained the B, arginine-rich, and A-I apoproteins. Differences between LDL-I and LDL-II included a higher sialic acid content for LDL-II and lipid to protein ratios of 3.7 and 3.0 for LDL-I and LDL-II, respectively. In addition, the LDL-II, but not LDL-I, reacted immunochemically with antisera prepared to human Lp(a). The physical, chemical, and immunochemical properties indicated that monkey LDL-II were equivalent to the human Lp(a). Patas monkey HDL, equivalent to human HDL, were protein and phospholipid rich and ranged in size from 70 to 100 A in diameter. The two major HDL apoproteins, A-I and A-II, were isolated from apo-HDL by column chromatography. The amino-terminal sequence of Patas A-I showed striking homology to that reported for human, dog, and swing A-I. The amino acid composition of monkey A-II was very similar to that of human A-II; however, unlike human A-II, the monkey apoprotein was shown to exist as a monomer similar to that reported for Rhesus monkey A-II. The similarities between the plasma lipoproteins of the monkey and of man suggest that the Patas monkey would serve as a suitable model for metabolic studies.     

Liver-specific inactivation of the abetalipoproteinemia gene completely abrogates very low density lipoprotein/low density lipoprotein production in a viable conditional knockout mouse

Conventional knockout of the microsomal triglyceride transfer protein large subunit (lMTP) gene is embryonic lethal in the homozygous state in mice. We have produced a conditional lMTP knockout mouse by inserting loxP sequences flanking exons 5 and 6 by gene targeting. Homozygous floxed mice were born live with normal plasma lipids. Intravenous injection of an adenovirus harboring Cre recombinase (AdCre1) produced deletion of exons 5 and 6 and disappearance of lMTP mRNA and immunoreactive protein in a liver-specific manner. There was also disappearance of plasma apolipoprotein (apo) B-100 and marked reduction in apoB-48 levels. Wild-type mice showed no response, and heterozygous mice, an intermediate response, to AdCre1. Wild-type mice doubled their plasma cholesterol level following a high cholesterol diet. This hypercholesterolemia was abolished in AdCre1-treated lMTP-/- mice, the result of a complete absence of very low/intermediate/low density lipoproteins and a slight reduction in high density lipoprotein. Heterozygous mice showed an intermediate lipoprotein phenotype. The rate of accumulation of plasma triglyceride following Triton WR1339 treatment in lMTP-/- mice was <10% that in wild-type animals, indicating a failure of triglyceride-rich lipoprotein production. Pulse-chase experiments using hepatocytes isolated from wild-type and lMTP-/- mice revealed a failure of apoB secretion in lMTP-/- animals. Therefore, the liver-specific inactivation of the lMTP gene completely abrogates apoB-100 and very low/intermediate/low density lipoprotein production. These conditional knockout mice are a useful in vivo model for studying the role of MTP in apoB biosynthesis and the biogenesis of apoB-containing lipoproteins.     

Influx of cholesterol into plasma in rabbits with fasting hyperbetalipoproteinemia

New Zealand white rabbits exhibited as much as a threefold increase in plasma cholesterol but no change in hepatic cholesterol when fasted for 7-9 days. Agarose electrophoresis and ultracentrifugation of plasma samples showed that only low density lipoprotein increased during fasting. Fasting changed the composition of the low density lipoprotein by increasing the percentage of cholesterol and decreasing the percentage of triglyceride while protein and phospholipid remained the same. Rates of cholesterol secretion into plasma, measured by Triton WR 1339 injection, and rates of plasma cholesteryl ester synthesis, determined by [2-(14)C]mevalonate injection, were similar for fed and fasted rabbits. These findings suggest that fasting hypercholesterolemia in rabbits did not result from increased production of low density lipoproteins. Triton WR 1339 was shown to inhibit plasma cholesterol esterification in vitro.     

Studies on the composition of pig serum lipoproteins. Isolation and characterization of different apoproteins.

1. Different lipoprotein density fractions from pig serum were isolated by phosphotungstate precipitation followed by purification in the preparative ultra-centrifuge. 2. The protein part of very low density lipoproteins was composed of approximately 52 percent lipoprotein B apoprotein and the rest of lipoprotein C II apoprotein and other as yet unidentified peptides. 3. The protein moiety of low density lipoproteins consisted primarily of lipoprotein B apoprotein (over 95 percent); the amino acid compositions of lipoprotein B apoprotein of very low and low density lipoproteins were practically identical. 4. The predominant polypeptide of pig serum high density lipoproteins exhibited an amino acid composition and a molecular weight very similar to human liprotein A I apoprotein. In contrast to human lipoprotein A I apoprotein, the apoprotein from pigs was found to release leucine first followed by alanine, threonine, and lysine upon incubation with carboxypeptidase A. 5. In pig serum the major lipoprotein C apoprotein was found to be a polypeptide similar in amino acid composition to lipoprotein C II apoprotein from human serum. The molecular weight of this polypeptide is approximately 8000. Incubation experiments with carboxypeptidase A indicate serine to be the most likely C-terminal amino acid.     

Interaction of plasma apolipoproteins with lipid monolayers.

The monolayer technique has been used to study the interaction of lipids with plasma apolipoproteins. Apolipoprotein C-II and C-III from human very low density lipoproteins, apolipoprotein A-I from human high density lipoproteins and arginine-rich protein from swine very low density lipoproteins were studied. The injection of each apoprotein underneath a monolayer of egg phosphatidy[14C]choline at 20 mN/m caused an increase in surface pressure to approximately 30 mN/m. With apolipoprotein C-II and apolipoprotein C-III there was a decrease in surface radioactivity indicating that the apoproteins were removing phospholipid from the interface; the removal of phospholipid was specific for apolipoprotein C-II and apolipoprotein C-III. Although there was a removal of phospholipid from the monolayer, the surface pressure remained constant and was due to the accumulation of apoprotein at the interface. The rate of surface radioactivity decrease was a function of protein concentration, required lipid in a fluid state and, of the lipids tested, was specific for phosphatidylcholine. Cholesterol and phosphatidylinositol were not removed from the interface. The addition of 33 mol% cholesterol to the phosphatidylcholine monolayer did not affect the removal of phospholipids by apolipoprotein C-III. The addition of phospholipid liposomes to the subphase greatly facilitated the apolipoprotein C-II-mediated removal of phospholipid from the interface. Although apolipoprotein A-I and arginine-rich protein gave surface pressure increases, phospholipid was only slightly removed fromthe interface by the addition of liposomes. Based on these findings, we conclude that the apolipoproteins C interact specifically with phosphatidylcholine at the interface. This interaction is important as it relates to the transfer of the apolipoproteins C and phospholipids from very low density lipoproteins to other plasma lipoproteins. The addition of human plasma high density lipoproteins or very low density lipoproteins to the subphase increased the apolipoprotein C-mediated removal of phosphatidyl[14C]choline from the interface 3--4 fold. Low density lipoproteins did not affect the rate of decrease. During lipolysis of very low density lipoproteins to the subphase increased the apolipoprotein C-mediated removal of with the lipid monolayer. Lipolysis experiments were performed in a monolayer trough containing a surface film of egg phosphatidyl[14C]choline and a subphase of very low density lipoproteins and bovine serum albumin. Lipolysis was initiated by the addition of purified milk lipoprotein lipase to the subphase. As a result of lipolysis, there was a decrease in surface radioactivity of phosphatidylcholine. The pre-addition of high density lipoproteins decreased the rate of decrease in surface radioactivity...     

Blockade of intestinal lipoprotein clearance in rabbits injected with Triton WR 1339-ethyl oleate

Although Triton WR 1339 has been used to block triglyceride or cholesterol removal from plasma, no data are available on the extent to which Triton WR 1339 administered to rabbits blocks clearance of newly absorbed dietary lipids. In the present study, we have measured the efficiency of this blockade during a 24-hr interval. After the Triton WR 1339 administration, plasma Sf greater than 400 and d less than 1.019 g/ml lipoprotein lipid concentrations increased greatly, but the concentration of d greater than 1.019 g/ml lipids decreased. In the rabbits fed 0.5% cholesterol for 1 week, the increase in d less than 1.019 g/ml and the decrease in 1.019 less than d less than 1.063 g/ml lipoprotein fractions 24 hr after the Triton WR 1339 injection were much greater than in the chow-fed Tritonized rabbits. After the Triton treatment, 50% of intravenously injected LDL-125I-labeled apoB disappeared in 24 hr, but little or no apoB appeared in other lipoprotein fractions and no VLDL apoB was converted to LDL. Labeled cholesterol and retinol were fed to rabbits and 24-hr increments in plasma cholesteryl- and retinyl-ester label and mass were measured. In chow-fed Tritonized rabbits about one-half of the absorbed oral doses of both labeled lipids was recovered in plasma, indicating that Triton WR 1339 does not completely inhibit the clearance of intestinal lipoproteins. When rabbits were injected with Triton and an ethyl oleate emulsion, the blockade of dietary lipid removal from plasma was substantially improved and chylomicron cholesterol uptake by extra-hepatic tissues was completely abolished.     

The turnover of human plasma very low density lipoprotein protein.

The plasma decay of three groups of iodinated apoproteins on human very low density lipoproteins were evauluated in two normals, two subjects with endogenous hypertriglyceridemia and another two with dysbetalipoproteinemia. The apo beta decay was more rapid than that of the C apoproteins in all patients. The apo beta decay was more rapid for the normals than for either the subjects with hypertriglyceridemia or dysbetalipoprotenemia. The apo C protein had an irregular decay in the normals but decayed less irregularly for the hypertriglyceridemics. The arginine rich apoprotein had a decay somewhat similar to apo C protein in the normals. The apo beta protein of the alpha2 very low density lipoprotein of a dysbetalipoproteinemic was consistent with a precursor relationship to the apo beta of beta very low density lipoprotein of this subject, but the arginine rich apoprotein was not.     

Hypolipidemic effect of pregnancy in the rabbit

New Zealand white rabbits showed large decreases in plasma cholesterol and phospholipid concentrations during the second half of pregnancy. All lipoproteins (very low density, low density, and high density) participated in the decrease. Very large decreases in plasma cholesterol concentrations were observed even when the animals were maintained on high cholesterol diets. Increases in plasma cholesterol concentrations, after the intravenous administration of Triton WR 1339, were at least as great in pregnant as in nonpregnant animals. It is concluded that the decrease in plasma cholesterol concentrations is not the result of impaired plasma lipoprotein production.     

The distribution and partial characterization of the serum apolipoproteins in the guinea pig.

1. Very-low-density (VLD), low-density (LD) and high-density (HD) lipoproteins were isolated by sequential ultracentrifugation from the serum of male guinea pigs fed on a diet containing 3--4% fat. The apoproteins of these lipoproteins (apo-VLD, apo-LD and apo-HD lipoproteins) were studied after delipidation with organic solvents or extraction with tetramethylurea. 2. The major apolipoprotein of LD lipoprotein isolated by gel filtration was found to closely resemble apolipoprotein B of human serum in its chemical and physical properties. Electrophoresis in sodium dodecyl sulphate-polyacrylamide gel showed that this apoprotein consisted of a number of polypeptides. 3. Tetramethylurea precipitated an apoprotein from guinea-pig serum lipoproteins that is probably the apolipoprotein B-like component. This apoprotein accounted for about 80% of the apo-LD lipoprotein, about 55% of the apo-VLD lipoprotein and about 50% of the apo-HD lipoprotein. 4. The distribution of apolipoproteins soluble in tetramethylurea was determined by densitometric scanning of stained polyacrylamide disc gels. 5. A glycine-rich component of high electrophoretic mobility (band I) and a triplet of soluble apolipoproteins (bands II-IV) were present in both VLD and LD lipoprotein classes. These components constituted a higher proportion of the tetramethylurea-soluble apoproteins of VLD lipoprotein (60--80%) than of LD lipoprotein (40--55%). 6. Small amounts (10--15%) of a component of intermediate mobility, which contained traces of half-cystine, were also present in both VLD and LD lipoproteins. 7. A group of soluble components of basic character (bands VI-X), present as minor components of VLD lipoprotein (10--20%), constituted a major proportion (30--45%) of the soluble apoproteins of LD lipoprotein. Two of these apoproteins were rich in lysine, and two of lower electrophoretic mobility were rich in arginine. 8. The pattern of tetramethylurea-soluble apoproteins in HD lipoprotein was distinguished by the presence of two polypeptides of low electrophoretic mobility as its predominant components. One of these components, band VI, resembled the A-I apolipoprotein of man in both its amino acid profile and in its electrophoretic mobility. The second major component, band VI-B, was rich in lysine and resembled the C-I apolipoprotein of man in amino acid composition. 9. The soluble components of bands I and IX were analogous in physicochemical properties to the R-X1 and R-X2 (high-arginine polypeptide) peptides of human serum lipoproteins respectively.     

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.     

Separation and characterization of plasma lipoproteins of rhesus monkeys (Macaca mulatta).

A group of 14 adult male rhesus monkeys was maintained on a low cholesterol-high fat diet. Periodically, animals were fasted and blood samples were taken for characterization of the plasma lipoproteins. Complete separation of individual plasma lipoprotein classes was not achieved by traditional sequential ultracentrifugation techniques. Rather, initial separation of lipoprotein classes according to size was effected and density centrifugation was used subsequently for further separation. At least six lipoprotein fractions were identified, each of which was unique as defined by the properties of size, density (d), and electrophoretic mobility. These lipoprotein fractions were characterized by determination of chemical compositions and apoprotein patterns. The lipoproteins present in highest concentration in these monkeys were designated as region IV lipoproteins. This fraction had alpha-migration on agarose electrophoresis, 1.063 < d < 1.225, and the size, composition, and apoprotein pattern characteristic of HDL. No fewer than three fractions were identified with densities that overlapped the 1.019 < d < 1.063 range. Of these, the fraction designated as region III lipoproteins was present in highest concentration, had beta-migration by agarose electrophoresis, a predominant B apoprotein, and a chemical composition and size characteristic of LDL. Two larger subfractions, identified as region II lipoproteins, were separated from each other at a density of 1.050 g/ml. Agarose electrophoresis showed that the fraction with d < 1.050 had a migration intermediate between beta and pre-beta. The chemical composition and apoprotein pattern were consistent with the possibility that these lipoproteins were remnants of VLDL catabolism. The fraction with d > 1.050, had pre-beta mobility and a size and composition similar to the Lp(a) lipoprotein in plasma of human beings. At least two VLDL subfractions, identified as region I and IIa lipoproteins, were found although both were present in very low concentrations. Region I lipoproteins were larger and contained relatively more cholesteryl ester and more of the apoproteins that migrated with the mobility of apo-B and arg-rich apoprotein in SDS-polyacrylamide gel electrophoresis. Some of the region I lipoproteins were beta-migrating by agarose electrophoresis. These results suggested the possibility that a beta-migrating VLDL was present in these normal animals.     

Cortisol, adrenocorticotropic hormone and apolipoprotein synthesis in rat hepatocytes]

The influence of cortisol (5 mg/kg body wt administered daily for 5 and 10 days) on biosynthesis of apoproteins of lipoproteins of very low density in the liver and on the synthesis of apolipoproteins of very low, low, and high density (VLDL, LDL, and HDL apoproteins, respectively) in the blood serum of adrenalectomized animals, and after replacement cortisol therapy was studied. Cortisol treatment during these periods resulted in the VLDL apoproteins biosynthesis inhibition in the rat liver. The synthesis of apolipoproteins was increased by adrenalectomy; this effect was eliminated after replacement cortisol treatment. The apoprotein synthesis was stimulated within 5 hours by single injection of cortisol or ACTH. Study of the blood serum apolipoproteins specific radioactivity indicated metabolic change of lipoproteins, such as disturbed conversion from VLDL to LDL. Single and prolonged cortisol administration led to the opposite results. The authors believe that the metabolic disturbances of lipoproteins in the blood play a more important role in the pathogenesis of cortisol-induced hyperlipidemia than lipoprotein syntesis stimulation in the liver.