PubMed Health⌕ Search

Biomedical subjects

D B Zilversmit

Publications and source records attributed to D B Zilversmit.

At least 37 records · Page 2Linked to original sources
PubMed1986

Parallel changes in plasma cholesterol and lipid transfer activity in pregnant rabbits.

The relationship between the concentration of plasma cholesterol and the lipid transfer activity (LTA) of lipoprotein-deficient plasma (d greater than 1.21) was studied in two models of pregnancy in the rabbit. Plasma cholesterol and the protein-mediated transfer of cholesteryl ester and triglyceride were monitored throughout gestation, 48 hr after parturition, and during lactation in New Zealand white (NZW) and heterozygous WHHL rabbits. Lipoprotein cholesterol was determined prior to and 48 hr after parturition. For both NZW and heterozygous WHHL rabbits, the progressive hypocholesterolemia of gestation was associated with parallel changes in LTA. Similarly, the rapid postpartum increase in plasma cholesterol was paralleled by increased LTA for both strains. In relation to basal values, the relative changes in plasma cholesterol and LTA were virtually identical. These data provide further evidence that in the rabbit plasma cholesterol and LTA are closely related.

Animals↗
PubMed1985

Purification and characterization of microsomal triglyceride and cholesteryl ester transfer protein from bovine liver microsomes.

A lipid transfer protein was isolated from bovine liver. Following the release of soluble proteins from liver microsomes, the transfer protein was purified 75-fold to near homogeneity by a combination of DEAE-cellulose ion exchange, Sephadex G-200 gel permeation, and hydroxylapatite chromatography. About 7% of the original activity was recovered. The purified fraction promoted the transfer of triglyceride, cholesteryl ester, phosphatidylcholine, and phosphatidylethanolamine. When the fractional rates of lipid transfer were compared, the transfer of apolar lipids was over 10 times faster than that of phospholipid. The purified transfer complex contained less than 5% lipid. No carbohydrate was detected. Electrophoresis of the purified protein on polyacrylamide gels under non-denaturing conditions showed a single band. Elution of protein from slices of unstained gels showed that lipid transfer activities coincided with the position of the protein band on the stained gel. When the purified protein was electrophoresed in the presence of SDS, two bands, accounting for more than 95% of the staining density, were observed with molecular weights at 58 000 and 88 000. The purified transfer protein eluted from a Sephadex G-200 column at a position corresponding to a protein with a molecular weight of 220 000, which probably represents a complex of two or more polypeptides. The purified transfer protein was activated by increasing NaCl concentrations up to about 100 mM. At higher NaCl concentrations the transfer activity decreased. Maximal transfer activities were observed at pH 7. The protein was inactivated by heating above 50 degrees C. The transfer rates were not greatly increased by changing the assay temperatures between 20 degrees C and 50 degrees C. These activity characteristics of the transfer protein were the same whether triglyceride or cholesteryl ester transfer activities were measured.

Animals↗
PubMed1984

Direct determination of human and rabbit apolipoprotein B selectively precipitated with butanol-isopropyl ether.

A method is described for the rapid, selective, and quantitative precipitation of apolipoprotein B from isolated hypercholesterolemic rabbit and human very low density lipoproteins (VLDL), intermediate density lipoproteins (IDL), and low density lipoproteins (LDL). Lipoprotein samples are heat-treated at 100 degrees C in 1% SDS. The denatured apoprotein solutions are then mixed briefly with two volumes of butanol-isopropyl ether 45:55 (v/v) to precipitate the apoB. The supernatant solutions, containing the non-apoB proteins and lipids, are removed and the apoB pellet is washed once with water. To determine apoB specific activity, the apoB pellet is resolubilized in 0.5 M NaOH by heating for 30 min at 120 degrees C. The hydrolyzed apoB protein is quantitated by fluorescence of a fluorescamine derivative. The precipitation of apoB is quantitative and selective: 99.5% of rabbit 125I-labeled LDL-apoB and 97.5% of human 125I-labeled LDL-apoB is precipitated and less than 5% of 125I-labeled HDL added to unlabeled VLDL, IDL, or LDL is precipitated. Triglyceride and cholesteryl ester contamination of the apoB pellet is less than 2% of their original radioactivities.

Animals↗
PubMed1984

Purification and characterization of human plasma proteins that inhibit lipid transfer activities.

A protein which inhibits cholesteryl ester and triacylglycerol transfer activities was purified from human lipoprotein-deficient plasma by chromatography on phenyl-Sepharose CL-4B, chromatofocusing, Bio-Gel A-0.5m and hydroxylapatite. The inhibitor is a sialoglycoprotein with molecular weight 32 000 and a relatively broad isoelectric region of 3.9-4.3. The inhibitor suppressed triacylglycerol and cholesteryl ester transfer activities to a similar extent. Apolipoprotein A-I, which was separated from the inhibitor by chromatofocusing chromatography, suppressed triacyglycerol transfer more than cholesteryl ester transfer. The percentage reduction of lipid transfer between lipoproteins by the inhibitor was independent of the concentration of transfer protein but was decreased at higher lipoprotein concentrations. The inhibition was not observed during lipid transfer between liposomes. These results indicate that the inhibitor interacts with substrates rather than with the transfer protein.

Blood Proteins↗
PubMed1984

A triglyceride and cholesteryl ester transfer protein associated with liver microsomes.

An intracellular protein accelerates the transfer of triglyceride and cholesteryl ester. The fraction of phospholipid transferred was much less than for the less polar lipids. A rich source of this activity was obtained from low ionic strength washes of liver microsomes. The protein was partially purified by column chromatography on Bio-Gel A-5m and hydroxylapatite. The elution position of the transfer protein on gel filtration corresponds to a protein with a molecular weight of about 200,000. The isoelectric point of the partially purified protein is between pH 5.2 and 5.6. At each step of the purification the stimulation of triglyceride transfer was greater than that of cholesteryl ester.

Animals↗
PubMed1984

Facilitated transfer of cholesteryl ester between rough and smooth microsomal membranes by plasma lipid transfer protein.

The accessibility of intracellular membrane cholesteryl esters to removal was tested with plasma lipid transfer protein as a tool. Incubation of a mixture of non-radioactive smooth microsomes + rough microsomes prelabeled with cholesteryl ester resulted in slight movement (2-4%) of radioactive cholesteryl ester into smooth microsomes. With the addition of increasing amounts of plasma lipid transfer protein to the mixture, the % transfer of cholesteryl ester into smooth microsomes progressively increased until a plateau was reached at 14%. Movement of cholesteryl ester in the reverse direction was examined with non-radioactive rough microsomes as an acceptor and smooth microsomes prelabeled with cholesteryl ester as a donor. The pattern of the % cholesteryl ester transferred in the reverse and forward direction was almost identical in the presence of plasma lipid transfer protein, showing bidirectional movement of cholesteryl ester between membranes.

Animals↗
PubMed1984

Comparison of various methods for in vitro cholesteryl ester labeling of lipoproteins from hypercholesterolemic rabbits.

Little or no information is available on biologically valid labeling of hypercholesterolemic plasma lipoproteins with cholesteryl ester. The esterification of labeled unesterified cholesterol in hypercholesterolemic rabbit plasma by the lecithin: cholesterol acyltransferase reaction is inefficient. The use of the d greater than 1.063 plasma fraction for this reaction greatly improves the efficiency, but some labeled unesterified cholesterol remains in the end products. The latter disadvantage can be avoided by the addition to whole plasma of labeled cholesteryl ester dissolved in DMSO or acetone. However, in hypercholesterolemic rabbit plasma only a small fraction of the added cholesteryl ester was associated with lipoproteins. When phosphatidylcholine/cholesteryl ester liposomes were incubated with hypercholesterolemic rabbit plasma for 18-24 h at 37 degrees C the labeled cholesteryl ester was quantitatively incorporated into lipoproteins. Chylomicron-like, cholesteryl ester-rich particles were removed by centrifugation (10(6) g X min) and the subsequently isolated d less than 1.019 and d = 1.019-1.063 (LDL) fractions were injected intravenously into normal and hypercholesterolemic rabbits. The disappearance of d less than 1.019 and LDL cholesteryl ester and the appearance of cholesteryl ester in other lipoprotein fractions was indistinguishable from that of in vivo-labeled lipoproteins. In vivo and in vitro cholesteryl ester-labeled lipoproteins were also compared by measuring the exchangeability of their cholesteryl ester with HDL cholesteryl ester in vitro. Equal exchangeability of the two labels was observed in the d less than 1.019 fraction from which the chylomicron-like particles had been removed. These findings demonstrate that when cholesteryl ester is incorporated by the liposome procedure, the distribution of labeled cholesteryl ester within the lipoprotein complex corresponds closely to that of the in vivo-incorporated labeled cholesteryl ester.

Animals↗
PubMed1983

Inhibition of hormone-stimulated adenylate cyclase activity after altering turkey erythrocyte phospholipid composition with a nonspecific lipid transfer protein. Phosphatidylinositol uncouples catecholamine binding from adenylate cyclase activation.

The nonspecific lipid transfer protein from beef liver was used to modify the phospholipid composition of intact turkey erythrocytes in order to study the dependence of isoproterenol-stimulated adenylate cyclase activity on membrane phospholipid composition. Incorporation of phosphatidylinositol into turkey erythrocytes inhibited isoproterenol-stimulated cyclic AMP accumulation in a linear, concentration-dependent manner. Inhibition was relatively specific for phosphatidylinositol; phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylglycerol and phosphatidic acid were from 3 to 7 times less effective as inhibitors of hormone-stimulated cyclase activity. Inhibition by phosphatidylinositol was not reversible when up to 90% of the incorporated phosphatidylinositol was removed, either by incubation with phosphatidylinositol-specific phospholipase C or a second incubation with transfer protein; possibly adenylate cyclase activity depends on a small pool of phosphatidylinositol that is inaccessible to either phospholipase C hydrolysis or removal by lipid transfer protein. Phosphatidylinositol incorporation inhibits adenylate cyclase activity by uncoupling beta-adrenergic receptors from the remainder of the cyclase complex. Phosphatidylinositol incorporation had no effect on stimulation of cAMP accumulation by either cholera toxin or forskolin, indicating that inhibition occurs only at the level of receptor. Phosphodiesterase activity was not altered in phosphatidylinositol-modified cells. Inhibition of cAMP accumulation was not the result of changes in either membrane fluidity or in cAMP transport out of modified turkey erythrocytes. Phosphatidylinositol inhibition of isoproterenol-stimulated cyclase activity may serve as a useful model system for hormone-induced desensitization.

Adenylyl Cyclase Inhibitors↗
PubMed1983

Inter-relationship of lipids transferred by the lipid-transfer protein isolated from human lipoprotein-deficient plasma.

In a previous study we demonstrated that highly purified lipid-transfer protein facilitated the transfer of triglyceride, cholesteryl ester, and phosphatidylcholine between plasma lipoproteins. It remained unclear, however, whether these lipids were transferred by independent sites on the lipid-transfer protein. To address this point, we have studied the protein-mediated transfer of triglyceride, cholesteryl ester, and phosphatidylcholine as a function of the concentration and lipid composition of donor and acceptor lipoproteins. Lipoproteins labeled in vitro, reconstituted lipoproteins of defined lipid composition, and phosphatidylcholine liposomes with or without triglyceride and/or cholesteryl ester have been used to investigate the inter-relationships of lipids transferred by the lipid-transfer protein. In studies of initial (less than or equal to 10-13%) transfer, we found that, although absolute transfer rates were affected, the ratio of cholesteryl ester to triglyceride transferred was independent of donor and acceptor lipoprotein concentrations and acceptor lipoprotein lipid composition. With reconstituted lipoproteins as donor, we demonstrated that this ratio was linearly related to the ratio of cholesteryl ester to triglyceride in the donor particle; the sum of triglyceride and cholesteryl ester transferred remained constant and independent of the lipid composition of the donor. Experiments with intact lipoproteins labeled in vitro and with small unilamellar vesicles in the presence and absence of p-chloromercuriphenylsulfonate, confirmed the interdependence of triglyceride and cholesteryl ester transfer. In contrast, under all assay conditions, no correlation was found between the amount of phosphatidylcholine transferred and the transfer of triglyceride and/or cholesteryl ester. We conclude that triglyceride and cholesteryl ester compete for transfer and that the extent of transfer for each lipid is determined by its relative concentration in the donor particle, whereas phosphatidylcholine transfer is independent of triglyceride and cholesteryl ester transfer. The data also strongly support the conclusion that lipid transfer protein promotes both the exchange and net transfer of triglyceride and cholesteryl ester and that the net transfer process proceeds by a reciprocal exchange of triglyceride and cholesteryl ester without net transfer of core lipid between lipoproteins.

Carrier Proteins↗
PubMed1983

Lack of secretion of retinyl ester by livers of normal and cholesterol-fed rabbits.

The use of retinyl ester as a tracer for chylomicrons and chylomicron remnants depends on the observation that newly absorbed dietary retinol is transported as retinyl ester in plasma chylomicrons or chylomicron remnants and also on the assumption that organs other than intestine do not contribute retinyl ester to plasma. To measure the secretion of retinyl ester by rabbit liver, the liver was labeled by injecting labeled retinol intravenously 1) as a colloidal dose, 2) incorporated into liposomes or 3) dispersed in a solution of Tween 20. Depending on the dose, between 63 and 80% of the labeled retinol in the liver was esterified and was found in both parenchymal and nonparenchymal cells. For all types of doses in both normal and cholesterol-fed rabbits, less than 1% of the injected dose was present in the plasma as retinyl ester during the 24-hour time period after injection. The secretion of retinyl ester by liver in response to the uptake of retinyl ester-enriched chylomicrons was also measured. This was done by feeding a diet enriched in retinol followed by a retinol-free diet. Only an insignificant quantity of retinyl ester accumulated in plasma during a 24-hour period after blocking the removal of triglyceride-rich lipoproteins with Triton WR 1339. Apparently, there is little, if any, secretion of retinyl ester by the liver of normal or cholesterol-fed rabbits.

Animals↗
PubMed1983

Plasma very low density lipoprotein (VLDL) in cholesterol-fed rabbits: chylomicron remnants or liver lipoproteins?

When iodinated hypercholesterolemic plasma very low density lipoprotein (VLDL), chylomicrons or chylomicron remnants were injected intravenously, the apoB of chylomicrons and chylomicron remnants was removed more rapidly than apoB of hypercholesterolemic VLDL. In perfused livers from normal or cholesterol-fed rabbits, chylomicron remnants were removed 69% and 125% more rapidly, respectively, than hypercholesterolemic VLDL. Chylomicron remnants were removed equally well by perfused livers from normal and cholesterol-fed rabbits, but hypercholesterolemic plasma VLDL was removed more slowly by perfused livers of cholesterol-fed than by those of normal rabbits. Chylomicron remnant removal by livers from normal and cholesterol-fed rabbits was inhibited by high levels of hypercholesterolemic plasma VLDL in the perfusate. Twenty-four hours after a single dose of retinyl ester was fed to cholesterol-fed rabbits, less than 1% of the absorbed retinol remained in the plasma as retinyl ester. Thus, increases in plasma VLDL cholesterol levels in cholesterol-fed rabbits cannot be accounted for by the accumulation of chylomicron remnants. Apparently, a cholesterol-rich VLDL, probably of hepatic origin, accumulates in the postabsorptive plasma of the cholesterol-fed rabbit.

Animals↗
PubMed1983

A model for cholesterol absorption: isotope vs. mass; single dose vs. constant infusion.

A study is presented to evaluate the relative merits of isotope and cholesterol mass measurements for cholesterol absorption. In this study, cholesterol absorption is simulated as a sequence of 10 two-pool segments in which a concentration gradient of cholesterol mass and/or label exists between the site of exogenous cholesterol entry and that of fecal loss. The model is governed by first order rate constants both for label and for mass. The appearance of labeled cholesterol in lymph and feces provides a reliable measure of the cholesterol mass increment in lymph due to exogenous cholesterol absorption. Net cholesterol absorption, calculated from constant infusion experiments, differs numerically from this mass increment. A dual isotope fecal ratio method agrees with other labeling techniques, but gives reliable information only when feces are collected for a sufficiently long time.

Animals↗
PubMed1982

High de novo synthesis of glycerolipids compared to deacylation-reacylation in rat liver microsomes.

A microsomal system characterized by high flux through the entire de novo pathway from glycerol phosphate to phosphatidylcholine and triacylglycerol has been developed. Optimum synthesis of phosphatidylcholine requires CDPcholine, Mg2+, KCl and a palmitoyl-CoA-generating system containing palmitic acid, ATP and CoA. Incorporation of [14C]glycerol phosphate into phosphatidylcholine/triacylglycerol synthesis ratio decreases as palmitate is increased. Phosphatidylcholine synthesis from glycerol phosphate is stimulated more by palmitate than by other saturated fatty acids; phosphatidylcholine synthesis increases with increasing unsaturation of the added fatty acids. The ratio of incorporation of [3H]palmitate to [14C]glycerol phosphate was determined for phosphatidic acid, diacylglycerol, phosphatidylcholine and triacylglycerol. This ratio is approximately 2 for all diacylglycerolipids and 3 for triacylglycerol. In our system, incorporation of palmitate into microsomal glycerolipid proceeds primarily by the de novo pathway, with minimal fatty acid recycling via deacylation-reacylation.

Animals↗
PubMed1982

Exchange of retinyl and cholesteryl esters between lipoproteins of rabbit plasma.

Normal or hypercholesterolemic rabbit plasma stimulates the transfer of retinyl ester as well as cholesteryl ester from rabbit lymph chylomicrons, chylomicron remnants or from cholesteryl ester-rich plasma VLDL to the d greater than 1.019 lipoprotein fractions. The presence of p-chloromercuriphenylsulfonate does not inhibit the transfer of these esters. Partially purified lipid transfer protein from rabbit or from human plasma also accelerates the transfer of the above esters. Whereas the rabbit plasma transfer protein preferentially accelerates the transfer of retinyl ester, the human plasma transfer protein appears to have a somewhat greater stimulating effect on the transfer of cholesteryl ester from low- to high-density lipoproteins.

Animals↗