Interchange of apoprotein components between the human plasma high density lipoprotein subclasses HDL2 and HDL3 in vitro.

The major apoproteins of human high density lipoproteins (HDL) labeled with 125I have been shown to exchange between the two major HDL subclasses HDL2 and HDL3 in vitro. This bidirectional exchange process is inhibited by cross-linking with bifunctional reagents and is apparently dependent upon the formation of collision complexes. This exchange has been demonstrated both when the subclasses of HDL are free in solution and also when one of them is covalently bound to Sepharose. Using system involving Sepharose-bound HDL, it could be shown that not only free apoprotein molecules but subunits consisting of lipid-apoprotein combinations were exchanged between HDL2 and HDL3. The rate of exchange in these processes is significant in the lifetime of the protein particles in vivo equalling approximately 2.5% per h for apoprotein exchange. These experiments suggest that there is a dynamic relationship between HDL2 and HDL3 even though each of them exists alone in vitro as stable separate entities; when they are placed together in solution significant interaction occurs between the particles. Apoprotein exchange occurs between HDL2:HDL2 and HDL3:HDL3 as well as between HDL2 and HDL3 molecules. These data also suggest that the interconversion of HDL2 and HDL3 may be affected by the availability of lipids.     

HDL-induced cardiac prostacyclin synthesis: relative contribution of HDL apoprotein and lipid

The objective of this study was to determine if the apoprotein or lipid constituents of high density lipoproteins (HDL) mediate HDL-induced prostacyclin synthesis in the Langendorff-perfused rabbit heart. Acetylation, acetoacetylation, or partial removal by trypsin digestion of HDL apoprotein did not reduce the ability of the lipoprotein to stimulate cardiac prostacyclin synthesis. Delipidated apoproteins were less effective in stimulating cardiac prostacyclin synthesis in comparison to intact HDL. In contrast, protein-free lipid vesicles, made from HDL lipids, caused a pronounced stimulation of cardiac prostacyclin synthesis. These results suggest that HDL apoproteins, in their native state, are not essential for HDL-induced cardiac prostacyclin synthesis. The stimulation of cardiac prostacyclin synthesis by HDL may depend on the lipoprotein's lipid rather than on its apoprotein constituents.     

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.     

A rapid electrophoretic technique for identification of subunit species of apoproteins in serum lipoproteins

The denaturing solvent tetramethylurea (TMU) delipidates and quantitatively liberates the apoproteins of human serum high-density lipoprotein (HDL) in soluble form while virtually the whole apoprotein of human lowdensity lipoprotein (LDL) is precipitated. A fraction of the apoprotein of very low density lipoprotein (VLDL) which appears to represent its content of LDL-like protein (apo B) is precipitated by this reagent, while the remaining apoprotein species are liberated in soluble form.The dissociation of the soluble apoproteins from lipid by TMU obviates the need for time-consuming delipidation by organic solvents, permitting immediate electrophoretic analysis in polyacrylamide gels. Bands are observed with mobilities corresponding to those of all the major soluble polypeptide species isolated from serum lipoproteins by ion-exchange chromatography. The apparent distribution of these elements in the different classes of lipoproteins is in agreement with findings of studies employing chromatographic methods. The predominant apoprotein of HDL, which has been identified immunochemically in VLDL, appears to comprise less than 1% of the apoprotein of VLDL from normal serum.     

Characterization of baboon plasma high-density lipoproteins and of their major apoproteins.

Baboon high-density lipoproteins (HDL) were isolated by preparative ultracentrifugation between d = 1.063 and 1.215 g/mL. The HDL contains 48.8% protein and a lipid distribution similar to human HDL. The phospholipid distribution shows a low sphingomyelin value (5.9%), and the fatty acid composition of HDL is comparable to the human data except for the 18:1/18:2 ratio as a result of a higher 18:1 content in the CE and a lower 18:2 concentration in the PL. The major HDL apoproteins isolated on diethylaminoethyl-cellulose had a mobility on sodium dodecyl sulfate--polyacrylamide gel electrophoresis and a molecular weight and an amino acid composition similar to human apoA-I. However, the amino acid sequence of the first 30 residues of baboon apoA-I differed from the human apoprotein in residues 15 and 21. Treatment of apoA-I with carboxypeptidase A indicated a carboxyl-terminal sequence of Leu-Ser-Thr-Gln. Baboon apoHDL contained monomeric apoA-II with the mobility of monomeric human apoA-II and a molecular weight of 8500. The amino acid composition differed from the human apoA-II by the presence of arginine and by the absence of half-cystine and isoleucine. The circular dichroic spectra of apoA-I and apoA-II demonstrated a higher helicity compared to the human apoproteins. Recombination studies by microcalorimetry of apoHDL with dimyristoylphosphatidylcholine (DMPC) indicated similarities in the thermodynamic binding properties of the HDL apoproteins from man and baboon. The maximal-binding enthalpies of DMPC to apoHDL, apoA-I, and apoA-II were lower for the baboon than for the human apoprotein.     

Biosynthesis of high density lipoprotein by chicken liver: nature of nascent intracellular high density lipoprotein

Young chickens were administered L-[(3)H]leucine and after 10 or 30 min the livers were removed and fractioned into rough (RER) and smooth (SER) endoplasmic reticulum fractions and into light, intermediate, and heavy golgo cell fractions. The labeled high density lipoprotein (HDL), contained within these intracellular organelles was isolated either by immunoprecipitation using rabbit antiserum to rooster HDL, or by ultracentrifugal glotation between densities 1.063 and 1.21 g/ml. The radioactive apoproteins of nascent HDL were analyzed by SDS PAGE and detected by fluorography. Analyses of radioactive apoproteins obtained by immunoprecipitation from the contents of the RER, the SER, and the three golgi complex fractions revealed only one apoprotein, A1. The C peptide present in serum HDL was not detected intracellularly. The radioactive apoprotein A1 which is present within the cisternae of the RER and the SER fractions failed to float, whereas apoprotein A1, present within the golgi apparatus, readily floated between densities 1.063 and 1.21 g/ml. The HDL particles, isolated by flotation from the golgi apparatus content, were further characterized by lipid and protein analyses and by electron microscopy. Golgi HDL particles have the same density as serum HDL. On a percentage basis, golgi HDL contains less protein and more phospholipids than does serum HDL. Morphologically, golgi HDL is different in appearance from serum HDL. It is more heterogeneous in size, with most of the particles ranging 8.3-25 nm in diameter. The spherical particles contain small membrane tails. Occasionally, a few disk-shaped bilayer structures are also found within the golgi apparatus. These studies show that the newly synthesized apoprotein A1, present within the RER and the SER cell fractions, is not fully complexed with lipid and that apoprotein A1 does not acquire sufficient lipid to float at the proper HDL density until it enters the golgi apparatus. The difference in chemical composition and the heterogeneous size of golgi HDL may be attributed to the different stages of HDL maturation.     

Proteolysis of apoprotein B during the transfer of very low density lipoprotein from hens' blood to egg yolk

We report an example of the enzymic cleavage of an apoprotein B (apoB), the main apoprotein in the very low density lipoprotein (VLDL) of laying hens' blood, in a normal biological process, the formation of egg yolk. Plasma VLDL was labeled in vivo with 3H-amino acids, isolated by centrifuging, and injected into another laying hen. Yolk VLDL was isolated and its apoproteins were separated. ApoB was not detected in this lipoprotein. Most of the label originally in apoB was distributed among four smaller yolk apoproteins, apovitellenins III to VI, which are a large proportion of the apoproteins of VLDL in yolk. This distribution of 3H suggested that 80% of apoB was cleaved at three places. One yolk apoprotein, apovitellenin II, was not labeled, indicating that it did not originate from an apoprotein in plasma VLDL. The site for cleavage of apoB in the ovarian tissue has not been determined, but cleavage may occur during receptor-mediated endocytosis. The pattern of cleavage of apoB during transfer to yolk was not imitated by some known proteolytic enzymes.     

Characterization of high density lipoprotein binding activity in rat adrenocortical cells

Rat adrenocortical cells take up high density lipoprotein cholesterol for use as steroidogenic substrate. To better understand this unique uptake process, we have first characterized HDL binding. Infusion of human 125I-labeled HDL into rats pretreated with 4-APP demonstrated that the adrenal and ovary accumulate HDL in a saturable fashion in vivo. Subsequent studies using isolated rat adrenocortical cells demonstrated that cellular uptake of HDL is comprised of two events. One event is characterized by reversible membrane binding and is complete by 60 min (t1/2 = 20 min). The second event is marked by irreversible apoprotein accumulation which continues for at least 3 hr. Reversibly bound material exhibits the same apoprotein distribution as unincubated HDL. Irreversible accumulation could not be attributed to internalization or lysosomal accumulation inasmuch as it also occurred with partially purified plasma membranes and was not enhanced by addition of chloroquine. Reversible binding of human HDL3 exhibited a saturable dependence on concentration (Kd = 27 micrograms protein/ml; N = 3.0 X 10(6) sites/cell) similar to that previously reported for rat liver, ovary, and testis. Cell accumulation of HDL decreased by over 80% at 4 degrees C compared to 37 degrees C, did not require calcium, and was not diminished by prior cell treatment with trypsin or pronase. These results indicate that rat adrenocortical cells possess plasma membrane recognition sites for HDL with different properties than those of the LDL receptor. Moreover, adrenal accumulation of HDL apoproteins does not lead to secondary lysosome formation.     

Plasma levels of lipids and apoprotein in a group of middle-school students 11-14 years of age: preliminary data]

Plasma lipids and apoprotein A and B levels were measured in 63 children, of both sexes, in the age range 11-14 years. The children have been subjected to a blood drawing after a 12 hour fast at least. Statistical analysis proves that total cholesterol (TC) is positively correlated with triglycerides (TG), HDL cholesterol (HDL) with apolipoproteins A (Apo A), apolipoproteins A (Apo A) with apoproteins B (Apo B). In the end we confirm the utility of determining plasma lipids and apoproteins to estimate lipidic risk for atherosclerosis in pediatric age.     

Apoprotein E-rich high density lipoproteins inhibit ovarian androgen synthesis

The influence of high density lipoproteins (HDL) on luteinizing hormone-stimulated rat ovarian theca/interstitial cell steroidogenesis was studied. Without HDL the cells produced primarily androgens from progestin precursors. In the presence of rat or human HDL steroid output increased 3-5-fold, but the type of steroid produced was dependent on the source of the HDL. Human HDL nonselectively amplified luteinizing hormone-stimulated steroid production, whereas rat HDL promoted progestin production without a concomitant increase in androgen output. Comparisons of the activities of apoprotein E-rich HDL (e.g. HDL from intact mature rats) with apoprotein E-poor HDL (e.g. human HDL or rat HDL from hypophysectomized immature rats) suggested that apoprotein E was responsible for the inhibition of androgen production. Furthermore, the inhibitory activity of rat HDL was abolished by depleting apoprotein E-containing lipoproteins with heparin affinity chromatography. Direct evidence that apoprotein E was the inhibitory constituent of rat HDL was obtained by showing that isolated lipid-free rat apoprotein E inhibited androgen production, whereas isolated rat apoproteins A-I and A-IV did not. The possible paracrine function of apoprotein E in ovarian follicular maturation of the ovary is discussed.     

Interaction of rat plasma very low density lipoprotein with lipoprotein lipase-rich (postheparin) plasma.

Incubation of 125I-labeled very low density lipoprotein (VLDL) with lipoprotein lipase-rich (postheparin) plasma obtained from intact or supradiaphragmatic rats resulted in the transfer of more than 80% of apoprotein C from VLDL to high density lipoprotein (HDL), whereas apoprotein B was associated with lipoprotein of density less than 1.019 g/ml (intermediate lipoprotein). The transfer of 125I-labeled apoprotein C from VLDL to HDL increased with time and decreased in proportion to the amount of VLDL in the incubation system. A relationship was established between the content of triglycerides and apoprotein C in VLDL, whereas the amount of apoprotein C in VLDL was independent of that of other apoproteins, especially apoprotein B. The injection of heparin to rats preinjected with 125I-labeled VLDL caused apoprotein interconversions similar to those observed in vitro. The intermediate lipoprotein was relatively rich in apoprotein B, apoprotein VS-2, cholesterol, and phospholipids and poor in triglycerides and apoprotein C. The mean diameter of intermediate lipoprotein was 269 A (compared with 427 A, the mean Sf rate was 30.5 (compared with 115), and the mean weight was 7.0 X 10(6) daltons (compared with 23.1 X 10(6)). From these data it was possible to calculate the mass of lipids and apoproteins in single lipoprotein particles. The content of apoprotein B in both particles was virtually identical, 0.7 X 10(6) daltons. The relative amount of all other constituents in intermediate lipoprotein was lower than in VLDL: triglycerides, 22%; free cholesterol, 37%; esterified cholesterol, 68%; phospholipids, 41%; apoprotein C, 7%, and VS-2 apoprotein, 60%. The data indicate that (a) one and only one intermediate lipoprotein is formed from each VLDL particle, and (b) during the formation of the intermediate lipoprotein all lipid and apoprotein components other than apoprotein B leave the density range of VLDL to a varying degree. Whether these same changes occur during the clearance of VLDL in vivo is yet to be established.     

Endurance training increased HDL cholesteryl ester metabolism in rats

We studied the effects of endurance training on the metabolism of high-density lipoprotein (HDL, 1.063 less than density less than 1.15 kg/l) cholesteryl ester and proteins in rats fed a cholesterol-rich (1%) semipurified diet. The HDL were labeled with 131I in the apoproteins and with cholesteryl-[1-14C]oleate in the esters. The HDL were intravenously administered to endurance-trained (n = 10) and cage-sedentary (n = 10) rats. Blood samples were taken over the next 36 h while the rats were conscious and feeding. The trained rats had higher plasma HDL cholesterol (0.72 vs. 0.28 mM) and HDL apoprotein (461 vs. 267 mg/l) concentrations than the sedentary rats. The production or disposal rate of HDL cholesteryl ester was higher in the trained rats (1.36 mumol/h) than in the sedentary rats (0.72 mumol/h), whereas the production or disposal rate of HDL apoproteins was similar in the trained (0.64 mg/h) and sedentary (0.60 mg/h) rats. The residence time of the HDL cholesteryl esters (4.72 +/- 0.22 vs. 3.37 +/- 0.21 h) and HDL apoprotein (7.65 +/- 0.36 vs. 4.55 +/- 0.28 h) was longer for the trained than for the sedentary rats. These data indicate that endurance training resulted in a significant change in the metabolism of HDL cholesteryl esters and apoproteins as well as an increase in their concentrations.     

Abnormal interaction of the human apolipoprotein A-I variant [Lys107----0] with high density lipoproteins

Several isoforms of apoprotein A-I [apoA-I], the major apoprotein of high density lipoproteins [HDL], have been described. We compared the in vivo and in vitro properties of normal human apoA-I with those of apoA-I [Lys107----0]. Fluorescence and circular dichroic spectra showed that deletion of Lys107 decreases apoprotein self-association. In vivo metabolic studies in the rat indicated that the interaction of apoA-I [Lys107----0] with HDL was lower than normal. We conclude that deletion of Lys107 results in a reorganization of the apoprotein structure that decreases its potential to form hydrophobic associations.     

An alternate apoprotein conformation in high density apolipoprotein discoidal complexes. A Fourier transform infra-red study.

Discoidal complexes have been prepared from 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) and the apoproteins of HDL3 (apo HDL3) or purified apo A-I. Gel electrophoresis established that apo HDL3 contained 74% apo A-I. Deconvolution and curve-fitting of the infra-red amide I band of the apoprotein in the lipid-protein complex revealed a secondary structure containing approximately 40% alpha-helix and 50% beta-structure. This contrasted with the results from circular dichroism studies (Surewicz et al. (1986) J. Biol. Chem., 261, 16191) of apo A-I/DMPC complexes which predicted 68% alpha-helix and 7% beta-structure. The discrepancy between the two methods and limitations of the two techniques for lipoproteins is discussed.     

Regulation of apoprotein synthesis and secretion in the human hepatoma Hep G2. The effect of exogenous lipoprotein

The regulation of lipoprotein assembly and secretion at a molecular level is incompletely understood. To begin to identify the determinants of apoprotein synthesis and distribution among lipoprotein classes, we have examined the effects of chylomicron remnants which deliver triglyceride and cholesterol, and beta very low density lipoprotein (beta VLDL), which deliver primarily cholesterol, on apolipoprotein synthesis and secretion by the human hepatoma Hep G2. Hep G2 cells were incubated with remnants or beta VLDL for 24 h, the medium was changed and the cells then incubated with [35S]methionine. The secreted lipoproteins were separated by gradient ultracentrifugation and the radiolabeled apoproteins were isolated by immunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis and counted. Remnants caused a 14-fold, and beta VLDL a 7-fold, increase in VLDL apoprotein (apo) secretion; the apoB/apoE ratio in this class was unchanged. Preincubation with either of the lipoproteins also stimulated low density lipoprotein apoB secretion. Preincubation with beta VLDL, but not with remnants, significantly increased apoE and apoA-I secreted in high density lipoprotein (HDL). In addition, the apoE/apoA-I ratio precipitated from the HDL of beta VLDL-treated cells by anti-apoE was 2.2-fold higher than that precipitated by anti-apoA-I. There was no difference in the ratios precipitated from control HDL. This was due to the secretion of a lipoprotein, subsequently isolated by immunoaffinity chromatography, that contained predominantly apoE. When Hep G2 cells were preincubated with oleic acid alone, total apoprotein secretion was not altered. However, cholesterol-rich liposomes stimulated secretion of newly synthesized apoE, but not apoB, while apoA-I secretion was variably affected. Cholesterol-poor liposomes had no effect. Thus, lipid supply is a determinant of apoprotein synthesis and secretion, and cholesterol may be of particular importance in initiating apoprotein synthesis.     

Metabolism of apoE-free high density lipoproteins in rat hepatoma cells: evidence for a retroendocytic pathway

The cellular metabolism of apoE-free HDL (HDL) was studied in rat hepatoma cells (FU5AH). Cells incubated with HDL showed a dose-dependent decreased incorporation of [14C]acetate into cell sterol, indicating a net cholesterol delivery to the cells. HDL was localized both at the cell surface and inside the cell. This conclusion was drawn from both the association of 125I-labeled HDL with the cells under different experimental conditions and morphological evidence based on the association of colloidal gold-labeled HDL with the cells. Up to 63% of the 125I-labeled HDL protein initially inside the cell was subsequently recovered in the media as trichloroacetic acid precipitable (TCA-ppt) protein after a 30-min, 37 degrees C chase with a 100-fold concentration of unlabeled HDL. About 27% of the TCA-ppt apoprotein originally inside the cell was recovered as TCA-soluble material. Thus, we conclude that of the HDL apoprotein taken up by the cells, the majority is resecreted by a retroendocytosis pathway. The quantity of HDL apoprotein reappearing in the media was stimulated by the presence of unlabeled HDL in the media, while the amount of TCA-soluble material produced was not. Retroendocytosis of HDL was inhibited at 0 degree C and by the presence of 10 mM NaCN, 20 mM 2-deoxy-D-glucose in the media. Thus, the pathway appears to be both temperature- and energy-sensitive. HDL resecreted by the cell were depleted of cholesteryl ester and showed an altered size distribution, indicative of lipoprotein catabolism and remodeling. This study provides evidence for the existence of an endocytosis-retroendocytosis pathway for HDL apoproteins in a rat hepatoma cell and for the possibility that the endocytosis-retroendocytosis pathway may be involved in lipid delivery to the cell.     

Isolation and characterization of a larval lipoprotein from the hemolymph of Manduca sexta.

1. The larval high density lipoprotein (HDL) from the hemolymph of Manduca sexta, isolated by density gradient centrifugation, contains 61% protein, 37% lipid and 2% carbohydrate. 2. The molecular weight of HDL is 6 x 10(5), with two apoproteins of 2.85 x 10(5) and 8.1 x 10(4) daltons. 3. The large apoprotein is destroyed by trypsin treatment of the particle, while the small one is not. 4. Calculations based upon size and composition show that this particle is very different in structure from mammalian lipoproteins. It is proposed that a portion of the apoprotein occupies the central core region.     

Alterations of the plasma lipoproteins and apoproteins following cholesterol feeding in the rat.

The feeding of cholesterol to rats resulted in marked alterations in the type and distribution of the plasma lipoproteins and their apoproteins. The hyperlipoproteinemia was characterized by an increase in the d < 1.006 lipoproteins (B-VLDL and VLDL), an increase in the intermediate and low density lipoproteins (LDL), and the appearance of HDL(c). Associated with these lipoproteins was a prominence of the arginine-rich apoprotein. The high density lipoproteins (HDL) were decreased. A two-dimensional immunoelectrophoretic procedure was adapted to quantitate the changes in distribution of the arginine-rich apoprotein in the plasma and various ultracentrifugal fractions obtained from control and cholesterol-fed rats. In rats fed the cholesterol diet, the total plasma arginine-rich apoprotein increased from a control value of approximately 29 mg/dl to 47 mg/dl. The method of ultracentrifugation, however, was found to markedly alter the quantitative results. When the 60 Ti rotor was used at maximum speed to isolate the ultracentrifugal fractions, less than 50% of the total plasma arginine-rich apoprotein was associated with the lipoproteins in the d < 1.006 or the d 1.006-1.02, 1.02-1.063, or 1.063-1.21 ultracentrifugal fractions. By contrast, after limited ultracentrifugation with the 40 rotor, much less arginine-rich apoprotein was lost, with approximately 20% of the arginine-rich apoprotein in control rats and 10% in cholesterol-fed rats found in the d > 1.21 fraction. Significant alterations in the arginine-rich apoprotein quantitation notwithstanding, the observations of increased arginine-rich apoprotein in the B-VLDL, intermediate fraction, and HDL(c) following cholesterol feeding remained valid. However, precise quantitation awaits refinements in lipoprotein isolation techniques.     

Cross-linking of apoproteins in high density lipoprotein by dimethylsuberimidate inhibits specific lipoprotein binding to membranes

Apoprotein E-free high density lipoproteins (HDL) bind to various cells and cell membrane preparations with properties typical of ligand-receptor interactions. This specific binding can be inhibited by treatment of HDL with tetranitromethane (TNM). During treatment of HDL with TNM, in addition to the expected nitration of tyrosine residues, cross-linking of lipids to apoproteins and of apoproteins to each other occurs. We have recently shown that cross-linking of phospholipids to apoproteins is not responsible for the inhibition of binding (1987. Chacko, G. K., et al. J. Lipid Res. 28: 332-337). To determine the role of cross-linking of apoproteins to each other in the inhibition, we used the bifunctional reagent dimethylsuberimidate (DMS) to cross-link the apoproteins in HDL3. Over 80% of apoproteins in DMS-HDL3 were cross-linked, as analyzed by SDS-polyacrylamide gel electrophoresis. DMS-HDL3 was similar to control HDL3 in its lipid composition. Gel filtration chromatography did not reveal any significant difference in size between DMS-HDL3 and control HDL3. As determined by competitive binding with 125I-labeled HDL3, DMS-HDL3 was almost completely unable to bind specifically to rat liver plasma membranes and human skin fibroblasts. It is concluded from these results that TNM inhibits the specific binding of HDL3 to membranes by a mechanism that involves cross-linking of apoproteins to each other in HDL3 particles. This observation implies that the specific binding of HDL3 to cells may depend on the native quaternary structure of apoproteins in the HDL particle. Because of its reduced ability to bind to the specific binding sites, DMS-HDL3 may be useful for studies related to the functional aspects of HDL binding sites.     

Lipoprotein metabolism in the suckling rat: characterization of plasma and lymphatic lipoproteins

Suckling rat plasma contains (in mg/dl): chylomicrons (85 +/- 12); VLDL (50 +/- 6); LDL (200 +/- 23); HDL1 (125 +/- 20); and HDL2 (220 +/- 10), while lymph contains (in mg/dl): chylomicrons (9650 +/- 850) and VLDL (4570 +/- 435) and smaller amounts of LDL and HDL. The lipid composition of plasma and lymph lipoproteins are similar to those reported for adults, except that LDL and HDL1 have a somewhat higher lipid content. The apoprotein compositions of plasma lipoproteins are similar to those of adult lipoproteins except for the LDL fraction, which contains appreciable quantities of apoproteins other than apoB. Although the LDL fraction was homogeneous by analytical ultracentrifugation and electrophoresis, the apoprotein composition suggests the presence of another class of lipoproteins, perhaps a lipid-rich HDL1. The lipoproteins of lymph showed low levels of apoproteins E and C. The triacylglycerols in chylomicrons and VLDL of both lymph and plasma are rich in medium-chain-length fatty acids, whereas those in LDL and HDL have little or none. Phospholipids in all lipoproteins lack medium-chain-length fatty acids. The cholesteryl esters of the high density lipoproteins are enriched in arachidonic acid, whereas those in chylomicrons, VLDL, and LDL are enriched in linoleic acid, suggesting little or no exchange of cholesteryl esters between these classes of lipoproteins. The fatty acid composition of phosphatidylcholine, sphingomyelin, and lysophosphatidylcholine were relatively constant in all lipoprotein fractions, suggesting ready exchange of these phospholipids. However, the fatty acid composition of phosphatidylethanolamine in plasma chylomicrons and VLDL differed from that in plasma LDL, HDL1, and HDL2. LDL, HDL1, and HDL2 were characterized by analytical ultracentrifugation and shown to have properties similar to that reported for adult lipoproteins. The much higher concentration of triacylglycerol-rich lipoproteins in lymph, compared to plasma, suggests rapid clearance of these lipoproteins from the circulation.