Insulin regulation of lipoprotein lipase in cultured 3T3-L1 cells

Lipoprotein lipase activity is produced by the 3T3-L1 cell an established mouse fibroblast line which resembles an adipocyte after reaching a confluent stage of growth. Since insulin has been shown to be an important regulator of lipoprotein lipase in other mammalian systems, a two hour incubation period was utilized to determine if insulin could enhance an acute response of enzyme activity. Over the range of concentrations tested (0.4, 4.0 and 40 ng/ml), insulin increased lipoprotein lipase activity in acetone ether powders of cells (intracellular enzyme) and the activity secreted into the culture medium. A simultaneous decrease in lipoprotein lipase activity releasable with heparin in a subsequent incubation (membrane bound activity) indicates two distinct effects of insulin on the enzyme in this system.     

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.     

Monoacylglycerol accumulation in low and high density lipoproteins during the hydrolysis of very low density lipoprotein triacylglycerol by lipoprotein lipase.

We have demonstrated that low and high density lipoproteins from monkey plasma are capable of accepting and accumulating monoacylglycerol that is formed by the action of lipoprotein lipase on monkey lymph very low density lipoproteins. Furthermore, the monoacylglycerol that accumulates in both low and high density lipoproteins is not susceptible to further hydrolysis by lipoprotein lipase but is readily degraded by the monoacylglycerol acyltransferase of monkey liver plasma membranes. These observations suggest a new mechanism for monoacylglycerol transfer from triacylglycerol rich lipoproteins to other lipoproteins. In addition, the finding that monoacylglycerol bound to low and high density lipoprotein is degraded by the liver enzyme but not lipoprotein lipase lends support to the hypothesis that there are distinct and consecutive extrahepatic and hepatic stages in the metabolism of triacylglycerol in plasma lipoproteins.     

Heparin-binding apoproteins. Effects on lipoprotein lipase and hepatic uptake of remnants.

Apoprotein-free heparin-binding and non-binding chylomicrons were used as substrates to test the effects on lipoprotein lipase activity of (a) chylomicron protein I; (b) the mixture of proteins I, II and apoprotein E and (c) human beta 2-glycoprotein I. No activation of the enzyme was observed with any of those apoproteins. When rats were injected simultaneously with [3H]cholesterol-labelled heparin-binding chylomicrons (containing proteins I and II) and [14C]cholesterol-labelled non-binding chylomicrons, no differences were detected between the rates of removal from circulation of those two types of particles. Clearance of chylomicrons from circulation was accompanied by the incorporation of 3H and 14C labels into the livers at similar rates. It is concluded that proteins I, II and apoprotein E have no effect on the degradation of chylomicrons by lipoprotein lipase and that the hepatic recognition of remnants does not appear to be affected by proteins I and II.     

Lipoprotein lipase activity in the flight muscle of Locusta migratoria and its specificity for haemolymph lipoproteins

Lipoprotein lipase activity in flight muscle homogenates of Locusta migratoria was measured, using natural radiolabelled lipoproteins as substrates. The flight specific lipoprotein A⁺ (or low density lipophorin) stimulated lipoprotein lipase activity several-fold compared to the resting lipoprotein A_y (or high density lipophorin). However, with the high mol. wt lipoprotein fraction O_AKH as a substrate, lipase activity was even doubled compared to lipoprotein A⁺. Lipase activity was not increased in flight muscle homogenates of insects which had flown. Neither adipokinetic hormone, nor octopamine had any direct effect on lipoprotein lipase activity. Aspects of hormonal regulation and apoprotein activation of the locust flight muscle lipoprotein lipase are discussed and compared with the model for vertebrate lipoprotein lipase.     

Synthesis of lipoprotein lipase in cultured avian granulosa cells

Avian granulosa cells cultured as a homogeneous parenchymal population contain lipolytic activity. This activity is stimulated 2--5-fold by serum, inhibited 90% by 1 M NaCl and inhibited 80% by specific anti-lipoprotein lipase immunoglobulins. 85% of the activity binds to heparin-Sepharose 4B, and 70% of bound activity is eluted with 1.5 M NaCl. Thus, the lipolytic activity of cultured granulosa cells is lipoprotein lipase. Granulosa cells were shown to synthesize lipoprotein lipase in culture by incorporating [3H]leucine into the enzyme protein, as measured with an immunoadsorption technique. Finally, colchicine was shown to increase intracellular lipolytic activity, suggesting an inhibition of secretion of this enzyme by cultured granulosa cells.     

CL-proteins and the regulation of lipoprotein lipase activity in locust flight muscle

Lipoprotein lipases in the flight muscles of Locusta migratoria show a marked substrate specificity: diacylglycerols associated with the adipokinetic hormone (AKH)-induced lipoprotein, A+, are hydrolysed at 4 to 5 times the rate of those associated with the lipoprotein in resting (non-hormone-stimulated) locusts, Ayellow. To determine the basis for this discrimination, the effect on the activity of flight muscle lipoprotein lipase of CL-proteins, a major constituent of lipoprotein A+, but not of Ayellow, has been investigated; they inhibit the flight muscle enzyme in a competitive manner whether activity is measured with a natural lipoprotein substrate, a lipid emulsion or a water soluble substrate. Experiments in vivo suggest that the flight muscle enzyme is normally inhibited in resting (non-AKH-stimulated) locusts but, interestingly, injection of synthetic AKH-I relieves the inhibition and increases the activity by 30 to 40%. This is not a direct effect of the hormone on the enzyme, but appears to be related to the hormone-induced formation of lipoprotein A+, so that the majority of CL-proteins in the haemolymph become bound to this lipoprotein and the concentration of free CL-proteins is markedly reduced. We suggest that CL-proteins play a major role in the regulation of lipoprotein lipase in locust flight muscle.     

The mechanism of heparin stimulation of rat adipocyte lipoprotein lipase

Free fat cells and stromal-vascular cells were prepared from rat adipose tissue by incubation with collagenase. NH(4)OH-NH(4)Cl extracts of acetone-ether powders prepared from fat cells contained lipoprotein lipase activity but extracts of stromal-vascular cells did not. Intact fat cells released lipoprotein lipase activity into incubation medium, but intact stromal-vascular cells did not. The lipoprotein lipase activity of the medium was increased when fat cells were incubated with heparin, and this was accompanied by a corresponding decrease in the activity of subsequently prepared fat cell extracts. Heparin did not release lipoprotein lipase activity from stromal-vascular cells. The lipoprotein lipase activity of NH(4)OH-NH(4)Cl extracts of fat cell acetone powders is increased by the presence of heparin during the assay. This increase is not due to preservation of enzyme activity, but to increased binding of lipoprotein lipase to chylomicrons. Protamine sulfate and sodium chloride have little effect on the binding of lipoprotein lipase to chylomicrons, but they inhibit enzyme activity after binding to substrate has occurred. These inhibitors do, however, inhibit the stimulatory effect of heparin on enzyme-substrate binding.     

Synthesis and secretion of lipoprotein lipase in 3T3-L1 adipocytes. Demonstration of inactive forms of lipase in cells

3T3-L1 adipocytes in culture incorporated [35S]methionine into a protein which could be immunoprecipitated with chicken antiserum to bovine lipoprotein lipase. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed this protein had an Mr of 55,000, similar to that of bovine lipoprotein lipase, and accounted for 0.1-0.5% of total protein synthesis in the adipocytes. Lipoprotein lipase protein was present in small amounts in confluent 3T3-L1 fibroblasts, and the amount increased many-fold as the cells differentiated into adipocytes. This increase was accompanied by parallel increases in cellular lipase activity and secretion. When cells were grown with [35S]methionine, the amount of label incorporated into lipoprotein lipase increased for 2 h and then leveled off. Pulse-chase experiments showed that half-life of newly synthesized lipase was about 1 h. Turnover of lipoprotein lipase in control cells involved both release to the medium and intracellular degradation. When N-linked glycosylation was blocked by tunicamycin, the cells synthesized a form of lipase that had a smaller Mr (48,000), was catalytically inactive, and was not released to the medium. Radioimmunoassay demonstrated that 3T3-L1 adipocytes contained an unexpectedly large amount of lipoprotein lipase protein. 55% of the enzyme protein in acetone/ether powder of the cells was insoluble in 50 mM NH3/NH4Cl at pH 8.1, a solution commonly used to extract lipoprotein lipase; 27% of the lipase protein was soluble but did not bind to heparin-Sepharose and had very low lipase activity; and the remaining 13% was soluble, bound to heparin-Sepharose, and had high lipolytic activity. About one-half of the lipase released spontaneously to the medium was inactive, and lipase inactivation proceeded in the medium with little loss of enzyme protein. Lipoprotein lipase released heparin, in contrast, was fully active and more stable. When protein synthesis was blocked by cycloheximide, the level of lipoprotein lipase activity in adipocytes decreased more rapidly than the amount of lipase protein in the cells. Most of the inactive lipoprotein lipase in adipocytes probably results from dissociation of active dimeric lipase, but some could be a precursor of active enzyme.     

Endogenous plasma lipoprotein lipase activity in fed and fasting rats may reflect the functional pool of endothelial lipoprotein lipase

In this study, a correlation was sought between the circulating lipoprotein lipase activity and nutritional state in the rat. In fed rats, the plasma lipoprotein lipase activity was between 30 and 120 munits/ml, whereas after an overnight fast in restraining cages, the lipoprotein lipase plasma levels were between 280 and 500 munits/ml. The plasma lipoprotein lipase activity was inhibited by a specific high titre goat antiserum to rat lipoprotein lipase. No effect of fasting was seen on the plasma hepatic triacylglycerol lipase. 6 h after fasting, adipose tissue lipoprotein lipase decreased maximally, but plasma lipoprotein lipase was not changed and rose only after 16 h. Thus, it seems that most of the lipoprotein lipase activity in the fasting plasma was related to the 3-fold rise in lipoprotein lipase activity in the heart, which may represent total muscle lipoprotein lipase. The increase in heart lipoprotein lipase was due in part to an increase in the t1/2 of the enzyme from 1.2 to 2.9 h. To determine whether the high plasma levels in the fasting rats might result from impaired clearance of the enzyme by the liver, functional hepatectomy was carried out. 15 min after hepatectomy, plasma lipoprotein lipase rose up to 20-fold in fed and about 6-fold in fasting rats. Lipoprotein lipase activity extracted by the liver was calculated to be 30-60 munits/ml in the fed and 171-247 munits/ml plasma per min in fasting rats. An increase in lipoprotein lipase activity in extrahepatic tissues (heart, lung, kidney, diaphragm and adrenal) occurred 30 min after hepatectomy in fed rats. The increase in heart lipoprotein lipase was due to an increase in heparin-releasable fraction. Since no impairment of hepatic clearance of circulating plasma lipoprotein lipase was found, the high fasting plasma lipoprotein lipase activity may be related to an increase in enzyme synthesis, decreased enzyme turnover and an expansion of the functional pool in tissues such as the heart and probably muscle. The present findings indicate that measurement of endogenous plasma lipoprotein lipase can provide information with respect to the size of the functional pool under normal and pathological conditions.     

Analysis and lipoprotein lipase activation capacity of plasma lipoproteins isolated from atherosclerosis-susceptible White Carneau pigeon and atherosclerosis-resistant Show Racer pigeon

The lipoprotein composition and apoprotein composition of the major lipoprotein fraction (high density lipoprotein) were compared in White Carneau and Show Racer plasma. The capacity of the plasma and lipoproteins to activate the triacylglycerol hydrolyzing activity of lipoprotein lipase in vitro was compared in the two strains of birds and found to be identical in each case. It appears unlikely that differences in lipoprotein composition or tissue lipoprotein lipase activity will be reflected in the flux rates of lipoproteins in the two strains which have different susceptibilities to atherosclerosis.     

Metabolism of lipoprotein acylglycerols by liver parenchymal cells.

We investigated the metabolism by hepatocyte suspensions of the acylglycerols in lipoprotein remnants as well as those associated with albumin and low or high density lipoproteins. Remnants, albumin and plasma lipoproteins, rich in monoacylglycerol were prepared by short-term incubations of radio-labeled chylomicra or very low density lipoproteins with extrahepatic lipoprotein lipase in the presence of albumin and low and high density lipoproteins. We demonstrated that liver parenchymal cells contain an active monoacylglycerol acyltransferase that is located on the extracellular surface of the cell plasma membrane. Further, the enzyme is capable of degrading the monoacylglycerol in all the above forms. Triacylglycerol in intact chylomicra and very low density lipoproteins were not metabolized by the cells to any appreciable degree. The degradation of the remnant triacylglycerol appeared to depend solely on the activity of the lipoprotein lipase bound to the lipoprotein remnants. Little uptake of intact lipoprotein acylglycerols by the hepatocytes was observed; instead, hydrolysis of the substrates in the medium always preceded the uptake of the products. The products were then utilized for the synthesis of triacylglycerol and phospholipid within the cells.     

Serum lipoprotein composition, lecithin cholesterol acyltransferase and tissue lipase activities in pregnant diabetic rats and their offspring receiving enriched n-3 PUFA diet

The effects of dietary n-3 polyunsaturated fatty acids on lipoprotein concentrations and on lipoprotein lipase (LPL), hepatic triglyceride lipase (HTGL) and lecithin cholesterol acyltransferase (LCAT) activities were studied in streptozotocin-induced diabetic rats during pregnancy and in their macrosomic offspring from birth to adulthood. Pregnant diabetic and control rats were fed Isio-4 diet (vegetable oil) or EPAX diet (concentrated marine omega-3 EPA/DHA oil), the same diets were consumed by pups at weaning. Compared with control rats, diabetic rats showed, during pregnancy, a significant elevation in very low density lipoprotein (VLDL) and low and high density lipoprotein (LDL-HDL(1))-triglyceride, cholesterol and apoprotein B100 concentrations and a reduction in apoprotein A-I levels. HTGL activity was high while LPL and LCAT activities were low in these rats. The macrosomic pups of Isio-4-fed diabetic rats showed a significant enhancement in triglyceride and cholesterol levels at birth and during adulthood with a concomitant increase in lipase and LCAT activities. EPAX diet induces a significant diminution of VLDL and LDL-HDL(1) in mothers and in their macrosomic pups, accompanied by an increase in cholesterol and apoprotein A-I levels in HDL(2-3) fraction. It also restores LPL, HTGL and LCAT activities to normal range. EPAX diet ameliorates considerably lipoprotein disorders in diabetic mothers and in their macrosomic offspring.     

Multiple effects of tumor necrosis factor on lipoprotein lipase in vivo

A single dose of recombinant murine tumor necrosis factor (TNF) suppressed lipoprotein lipase activity in adipose tissue of fed rats, mice, and guinea pigs for 48 h, even though TNF itself is rapidly metabolized in vivo. Immunoprecipitation of [35S]lipoprotein lipase from fat pads pulse-labeled with [35S]methionine showed a decrease in relative synthesis of the enzyme, which correlated to the decrease in activity. There was no decrease in general protein synthesis and no change in distribution of the enzyme between adipocytes and extracellular locations in the tissue. This is in contrast to fasting in which case there is redistribution of the enzyme within the tissue, decrease in general protein synthesis, but no change in relative synthesis of lipoprotein lipase. TNF did not decrease lipoprotein lipase activity in any tissue other than the adipose but increased the activity in several cases, most markedly in the liver. No [35S]methionine was incorporated into lipoprotein lipase by liver slices from normal or TNF-treated animals. Thus, the increased activity can not be ascribed to enhanced hepatic synthesis of the enzyme. There was an increase in lipoprotein lipase activity in plasma, which correlated to the increase in liver. Thus, TNF suppresses lipoprotein lipase synthesis in adipocytes, but not in other tissues, and has some as yet undefined effect on lipoprotein lipase turnover in extrahepatic tissues, which results in increased transport of active lipase through plasma to the liver.     

Influence of nutritional state on lipoprotein lipase activities in the hypothyroid rat

We have investigated the effects of nutritional state on the lipoprotein lipase activities of the experimentally hypothyroid rat. Both short-term effects (i.e., those of a 24 h fast with and without re-feeding) and long-term effects (due to decreased food intake in hypothyroidism) have been studied. The hypothyroid rats had significantly higher lipoprotein lipase activities of adipose tissue and heart muscle. The effect of hypothyroidism on adipose tissue lipoprotein lipase activities was modified by the nutritional state. In rats studied after 24 h fasting, the hypothyroid group had significantly higher lipoprotein lipase activities than weight-matched, age-matched and pair-fed (i.e., semi-starved) control groups. In rats studied in the re-fed state, the effects of hypothyroidism as such were less evident, since the pair-fed group also demonstrated significantly higher enzyme activities than did the other control groups. We have also studied the lipoprotein lipase activities of different enzyme preparations from adipose tissue. The effects of hypothyroidism were most clearly reflected in an increase of heparin-elutable enzyme activity from adipose tissue, whereas adipocyte lipoprotein lipase activity and the lipoprotein lipase secretion rate from adipocytes were affected to a lesser extent. We conclude that alterations in food intake strongly influence the lipoprotein lipase activities in the hypothyroidism. Our data also imply that the increased lipoprotein lipase activity in the hypothyroid state is due to a decreased degradation of the enzyme, both intra- and extracellularly.     

Actin in young and senescent fibroblasts.

Long-term exposure of guinea pigs to a diet rich in maize oil caused an increase in adipose-tissue lipoprotein lipase activity. A similar diet rich in beef tallow had no such effect, and neither diet affected the enzyme activity in heart, lung, diaphragm or skeletal muscle.     

Intra- and extracellular forms of lipoprotein lipase in adipose tissue.

The location of lipoprotein lipase activity in rat adipose tissue was studied using intact epididymal fat pads, isolated adipocytes, and lipoprotein lipase activity secreted from adipocytes as enzyme sources. The enzyme activities of these preparations were characterized by gel filtration. The method used for isolation of adipocytes had been modified to minimize activation of lipoprotein lipase during the procedures. Extracts of intact adipose tissue separated into two major lipoprotein lipase activity peaks, designated "a" and "b", the "a" fraction representing about 30 (fasted rats) to 50% (fed rats) of the total enzyme activity. An intermediate fraction (designated "i") was frequently observed. Extracts of isolated adipocytes from fed rats contained about 35% and those from fasted rats about 65% of the lipoprotein lipase activity present in intact tissue. The "b" fraction constituted 80--97% of the adipocyte lipoprotein lipase activity. In contrast, the enzyme activity secreted from the adipocytes contained only the "a" and "i" fractions. These data implicate the existance of one intracellular form of lipoprotein lipase (corresponding to the "b" fraction), different from extracellular forms of the enzyme (corresponding to fractions "a" and "i"). A transformation of the intracellular to the extracellular forms appears to occur in conjunction with secretion of enzyme from the fat cell.     

Importance of the different steps of glycosylation for the activity and secretion of lipoprotein lipase in rat preadipocytes studied with monensin and tunicamycin

Lipoprotein lipase synthesized by cultured rat preadipocytes is present in three compartments: an intracellular, a surface-related 3-min heparin-releasable, and that secreted into the culture medium. 30 min after addition of 6 microM monensin, the lipoprotein lipase activity in the heparin-releasable compartment starts to decrease; by 4 h of monensin treatment the lipoprotein lipase activity in the heparin-releasable pool and in the culture medium is about 10% of that found in control dishes. The intracellular activity, which had been identified as lipoprotein lipase by an antiserum to lipoprotein lipase, increases slowly and doubles by 24 h. However, since the cellular compartment accounts for 10-25% of total activity, this increase does not account for the missing enzyme activity. To determine whether this enzyme molecule is synthesized but is not active, incorporation of labeled leucine, mannose and galactose into immunoadsorbable lipoprotein lipase was studied in control, monensin- or tunicamycin-treated cells. Addition of tunicamycin (5 micrograms/ml) for 24 h caused a 30-50% reduction in immunoadsorbable lipoprotein lipase, but the enzyme activity was reduced by 90%. On the other hand, 4 h monensin treatment reduced both incorporation of [3H]leucine into immunoadsorbable lipoprotein lipase and heparin-releasable and medium lipoprotein lipase activity by 57 to 77%. The immunoadsorbable lipoprotein lipase in the intracellular compartment has a [14C]mannose to [3H]galactose ratio of 0.15 and this ratio increased 6-fold in monensin-treated cells. The intracellular lipoprotein lipase in monensin-treated cells had the same affinity for both the native and synthetic substrate as the lipoprotein lipase in control cells, yet its spontaneous secretion into the culture medium and its release by 3 min heparin treatment was markedly decreased. The present results indicate that: the presence of asparagine-linked oligosaccharide (formation of which is inhibited by tunicamycin) is mandatory for the expression of lipoprotein lipase activity; lipoprotein lipase is active also in a high mannose form; and terminal glycosylation and oligosaccharide processing, which is inhibited by monensin, may be important for the appearance of heparin-releasable lipoprotein lipase and secretion of lipoprotein lipase into the medium.     

Interaction of lipoprotein lipase with native and modified heparin-like polysaccharides

1. Lipoprotein lipase (EC 3.1.1.34), which was previously shown to bind to immobilized heparin, was now found to bind also to heparan sulphate and dermatan sulphate and to some extent to chondroitin sulphate. 2. The relative binding affinities were compared by determining (a) the concentration of NaCl required to release the enzyme from polysaccharide-substituted Sepharose; (b) the concentration of free polysaccharides required to displace the enzyme from immobilized polysaccharides; and (c) the total amounts of enzyme bound after saturation of immobilized polysaccharides. By each of these criteria heparin bound the enzyme most efficiently, followed by heparan sulphate and dermatan sulphate, which were more efficient than chondroitin sulphate. 3. Heparin fractions with high and low affinity for antithrombin, respectively, did not differ with regard to affinity for lipoprotein lipase. 4. Partially N-desulphated heparin (40–50% of N-unsubstituted glucosamine residues) was unable to displace lipoprotein lipase from immobilized heparin. This ability was restored by re-N-sulphation or by N-acetylation; the N-acetylated product was essentially devoid of anticoagulant activity. 5. Partial depolymerization of heparin led to a decrease in ability to displace lipoprotein lipase from heparin–Sepharose; however, even fragments of less than decasaccharide size showed definite enzyme-releasing activity. 6. Studies with hepatic lipase (purified from rat post-heparin plasma) gave results similar to those obtained with milk lipoprotein lipase. However, the interaction between the hepatic lipase and the glycosaminoglycans was weaker and was abolished at lower concentrations of NaCl. 7. The ability of the polysaccharides to release lipoprotein lipase to the circulating blood after intravenous injection into rats essentially conformed to their affinity for the enzyme as evaluated by the experiments in vitro.     

Binding of chylomicron remnants and beta-very low density lipoproteins to hepatic and extrahepatic lipoprotein receptors. A process independent of apolipoprotein B48

The ability of apolipoprotein (apo-) B48 to interact with lipoprotein receptors was investigated using three different types of lipoproteins. First, canine chylomicron remnants, which contained apo-B48 as their primary apoprotein constituent, were generated by the hydrolysis of chylomicrons with milk lipoprotein lipase. These apo-B48-containing chylomicron remnants are deficient in apo-E and reacted very poorly with apo-E receptors on adult dog liver membranes and the low density lipoprotein (apo-B,E) receptors on human fibroblasts. Addition of normal human apo-E3 restored the receptor binding activity of these lipoproteins. Second, beta-very low density lipoproteins (beta-VLDL) from cholesterol-fed dogs were subfractionated into distinct classes containing apo-E along with either apo-B48 or apo-B100. Both classes bound to the apo-B,E and apo-E receptors. Their binding was almost completely mediated by apo-E, as evidenced by the ability of the anti-apo-E to inhibit the receptor interaction. Third, beta-VLDL from type III hyperlipoproteinemic patients were subfractionated by immunoaffinity chromatography into lipoproteins containing apo-E plus either apo-B48 or apo-B100. Both subfractions bound poorly to apo-B,E and apo-E receptors due to the presence of defective apo-E2. However, the residual binding of the apo-B48-containing and apo-B100-containing human beta-VLDL was inhibited by the anti-apo-E. After lipase hydrolysis, apo-B100 became a more prominant determinant responsible for mediating receptor binding to the apo-B,E receptor. By contrast, lipase hydrolysis did not increase the binding activity of the apo-B48-containing beta-VLDL. These results indicate that apo-B48 does not play a direct role in mediating the interaction of lipoproteins with receptors on fibroblasts or liver membranes.