Carboxyl ester lipase expression in macrophages increases cholesteryl ester accumulation and promotes atherosclerosis

Carboxyl ester lipase (CEL, also called cholesterol esterase or bile salt-dependent lipase) is a lipolytic enzyme capable of hydrolyzing cholesteryl esters, triacylglycerols, and phospholipids in a trihydroxy bile salt-dependent manner but hydrolyzes ceramides and lysophospholipids via bile salt-independent mechanisms. Although CEL is synthesized predominantly in the pancreas, a low level of CEL expression was reported in human macrophages. This study used transgenic mice with macrophage CEL expression at levels comparable with that observed in human macrophages to explore the functional role and physiological significance of macrophage CEL expression. Peritoneal macrophages from CEL transgenic mice displayed a 4-fold increase in [(3)H]oleate incorporation into cholesteryl [(3)H]oleate compared with CEL-negative macrophages when the cells were incubated under basal conditions in vitro. When challenged with acetylated low density lipoprotein, cholesteryl ester accumulation was 2.5-fold higher in macrophages expressing the CEL transgene. The differences in cholesteryl ester accumulation were attributed to the lower levels of ceramide and lysophosphatidylcholine in CEL-expressing cells than in CEL-negative cells. CEL transgenic mice bred to an atherosclerosis susceptible apoE(-/-) background displayed an approximate 4-fold higher atherosclerotic lesion area than apoE(-/-) mice without the CEL transgene when both were fed a high fat/cholesterol diet. Plasma level of the atherogenic lysophosphatidylcholine was lower in the CEL transgenic mice, but plasma cholesterol level and lipoprotein profile were similar between the two groups. These studies documented that CEL expression in macrophages is pro-atherogenic and that the mechanism is because of its hydrolysis of ceramide and lysophosphatidylcholine in promoting cholesterol esterification and decreasing cholesterol efflux.     

Effect of pantethine on post-heparin plasma lipolytic activities and adipose tissue lipoprotein lipase in rats

The lipid-lowering effect of pantethine, a new drug affecting lipid metabolism, had been evaluated in carbohydrate-induced hyperlipidemic rats. Administration of the drug raised post-heparin lipolytic activities, the change being due to an increase in lipoprotein lipase activity, whereas hepatic lipase activity remained virtually unchanged. Total lipoprotein lipase activity per g of adipose tissue increased in pantethine-treated rats compared with controls. Furthermore, the soluble lipoprotein lipase of fat-pads was fractionated by heparin-Sepharose affinity chromatography. The first active peak, originated from the microsomal fractions, significantly increased after the drug treatment, while the second one, originated from the plasma membranes, remained unchanged. The increase in the microsomal lipoprotein lipase activity may be due to an increase in intracellular synthesis of lipoprotein lipase enzyme proteins. The heterogeneity of lipoprotein lipase of rat adipose tissues was ensured using affinity chromatography on heparin-Sepharose.     

Acid Lipase of Castor Bean Lipid Bodies : Isolation and Characterisation

The acid lipase of castor endosperm lipid bodies has been studied using colorimetric assay based on the measure of the hydrolytic activity of p-nitrophenyl ester of palmitate and other acyl derivatives. These substrates are compatible with the natural triacylglycerols for the measure of lipolytic activities. The subcellularly-surveyed acid lipolytic activity in the germinated castor bean endospermal tissue was found to be enhanced in the lipid bodies. The lipase, which is partially latent and tightly associated with lipid bodies, is an exceptionally stable enzyme with an optimum activity at pH 4.5 and displays an inverse relationship between its activity and the acyl chain length of its substrate. To facilitate isolation of the acid lipase, a procedure has been developed to solubilise the membrane-bound enzyme in an active form. The detergent-solubilised acid lipase after two chromatographic steps yielded an eight-fold active preparation which after gel permeation resolved as heterogeneous aggregate in excess of 500 kD. Lipase-enriched preparations showed consistent presence of 14 and 60 kD proteins which constituted the most abundant species of the lipid bodies. Although it has not been possible to obtain an active lipase preparation in a state free of either the 14 or 60 kD protein, the lipase activity in the detergent extracts of lipid bodies was immunoprecipitable with antibodies raised against the 60 kD component.     

Human lipoprotein lipase: the loop covering the catalytic site is essential for interaction with lipid substrates.

Lipoprotein lipase (LPL), a key enzyme which initiates the hydrolysis of triglycerides present in chylomicrons and very low density lipoproteins, consists of multiple functional domains which are necessary for normal activity. The catalytic domain of LPL mediates the esterase function of the enzyme but separate lipid binding sites have been proposed to be involved in the interaction of LPL with emulsified lipid substrates at the water-lipid interface. Like pancreatic lipase (PL), LPL contains a surface loop covering the catalytic pocket that may modulate access of the substrate to the active site of the enzyme. Secondary structural analysis of this loop reveals a helix-turn-helix motif with two short amphipathic helices that have hydrophobic moments of 0.64 and 0.68. In order to investigate the role of the loop in the initial interaction of LPL with its substrate, we utilized site-directed mutagenesis to generate eight constructs in which the amphipathic properties of the loop were altered and expressed them in human embryonal kidney-293 cells. Reducing the amphiphilicity without changing the predicted secondary structure of the loop abolished the ability of the lipase to hydrolyze emulsified, long chain fatty acid triglycerides (triolein) but not the water soluble substrate tributyrin. Replacing the loop of LPL with the loop of hepatic lipase, which differs in 15 of 22 amino acids but is also amphiphilic, led to the expression of an enzyme that retained both triolein and tributyrin hydrolyzing activity. Substitution of the LPL loop by a short four amino acid peptide, which may allow more direct access to the active site than the 22 amino acid loop, enhanced hydrolysis of short chain fatty acid triglycerides by more than 2-fold, while the ability to hydrolyze emulsified substrates was abolished. Thus, disruption of the amphipathic structure of the LPL loop selectively decreases the hydrolysis of emulsified lipid substrate without affecting the esterase or catalytic function of the enzyme. These studies establish that the loop with its two amphipathic helices is essential for hydrolysis of long chain fatty acid substrate by LPL providing new insight into the role of the LPL loop in lipid-substrate interactions. We propose that the interaction between the lipoprotein substrates and the amphipathic helices within this loop may in part determine lipase substrate specificity.     

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.     

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.     

Concerted action of human carboxyl ester lipase and pancreatic lipase during lipid digestion in vitro: importance of the physicochemical state of the substrate

The pancreatic enzyme carboxyl ester lipase (CEL) has been shown to hydrolyse a large number of different esters, including triacylglycerols, cholesteryl esters and retinyl esters with an absolute requirement for bile salts. Some of the lipids that are substrates for CEL can also be hydrolysed by pancreatic lipase. In order to investigate the relative roles of human CEL and pancreatic lipase, the two enzymes were incubated on a pH-stat with isotope-labelled lipid substrate mixtures in physicochemical forms resembling the state of the dietary lipids in human intestinal contents. In the first set of experiments, cholesteryl oleate (CO) and retinyl palmitate (RP) were solubilised in an emulsion of triolein (TO) stabilised by egg phosphatidylcholine and bile salts. Lipase (always added together with its cofactor, colipase) hydrolysed TO, with monoolein and oleic acid as end-products, whereas CEL alone could not hydrolyse TO in the presence of phosphatidylcholine (PC). Lipase alone did not hydrolyse CO or RP, but CEL did hydrolyse these esters if lipase was present. Release of [3H]glycerol from labelled TO increased only slightly if CEL was added compared to lipase alone, suggesting that monoolein hydrolysis was slow under these conditions. In the second set of experiments, CO and RP were dissolved in bile salt/monoolein/oleic acid dispersions with varying bile salt concentrations. CEL hydrolysed CO and RP more rapidly in a system with a high bile salt concentration containing mixed micelles than in a system with a low bile salt concentration, where the lipids were dispersed in the form of mixed micellar and non-micellar aggregates; both types of aggregate have been reported to exist in human intestinal contents. In conclusion, these data suggest that the main function of CEL under physiological conditions is to hydrolyse cholesteryl and retinyl esters, provided that the triacylglycerol oil phase is hydrolysed by pancreatic lipase, which probably causes a transfer of the substrate lipids of CEL from the oil emulsion phase to an aqueous bile salt/lipolytic product phase. Depending on the bile salt/lipolytic product ratio, the substrate will reside in either micellar or non-micellar lipid aggregates, of which the micellar state is preferred by CEL.     

Post-heparin lipolytic activity with no hepatic triacylglycerol lipase involved in a mammalian species

It was found that lipolytic activity in bovine post-heparin plasma differed from that of other mammalian species by the fact that intravenous heparin induced the release of lipoprotein lipase but not hepatic triacylglycerol lipase.Initially, this fact was strongly suspected when no remaining lipolytic activity could be found after whole bovine post-heparin plasma had been tested with either 1 M NaCl or antiserum against lipoprotein lipase. This was further confirmed by using heparin-Sepharose affinity chromatography when the entire lipolytic activity was eluted with 1.5 M NaCl but none with 0.4 or 0.7 M NaCl. The active fraction had lipoprotein lipase characteristics i.e it required serum activators to produce optimum activity and was fully inhibited by NaCl of high molarity and by anti-lipoprotein lipase antiserum. Neither the different doses of heparin nor the various times of sampling altered the results. This raises the question whether hepatic triacylglycerol lipase is absent from the bovine liver or whether this enzyme is present but cannot be released by heparin.     

Bile salt-stimulated carboxyl ester lipase influences lipoprotein assembly and secretion in intestine: a process mediated via ceramide hydrolysis.

Bile salt-stimulated carboxyl ester lipase (CEL), also called cholesterol esterase, is one of the major proteins secreted by the pancreas. The physiological role of CEL was originally thought to be its mediation of dietary cholesterol absorption. However, recent studies showed no difference between wild type and CEL knockout mice in the total amount of cholesterol absorbed in a single meal. The current study tests the hypothesis that CEL in the intestinal lumen may influence the type of lipoproteins produced. A lipid emulsion containing 4 mm phospholipid, 13.33 mm [(3)H]triolein, and 2.6 mm [(14)C]cholesterol in 19 mm taurocholate was infused into the duodenum of lymph fistula CEL(+/+) and CEL(-/-) mice at a rate of 0.3 ml/h. Results showed no difference between CEL(+/+) and CEL(-/-) mice in the rate of cholesterol and triglyceride transport from the intestinal lumen to the lymph. However, CEL(-/-) mice produced predominantly smaller lipoproteins, whereas the CEL(+/+) mice produced primarily large chylomicrons and very low density lipoprotein. The proximal intestine of CEL(-/-) mice was also found to possess significantly less ceramide hydrolytic activity than that present in CEL(+/+) mice. By using Caco2 cells grown on Transwell membranes as a model, sphingomyelinase treatment inhibited the secretion of larger chylomicron-like lipoproteins without affecting total cholesterol secretion. In contrast, the addition of CEL to the apical medium increased the amount of large lipoproteins produced and alleviated the inhibition induced by sphingomyelinase. Taken together, this study identified a novel and physiologically significant role for CEL, namely the promotion of large chylomicron production in the intestine. The mechanism appears to be mediated through CEL hydrolysis of ceramide generated during the lipid absorption process.     

The lipoprotein lipase of white adipose tissue. Changes in the adipocyte cell-surface content of enzyme in response to extracellular effectors in vitro.

An indirect labelled-second-antibody cellular immunoassay for adipocyte surface lipoprotein lipase was used to assess the changes that occurred during the incubation of cells in the presence and absence of effectors. In the absence of any specific effectors, the amount of immunodetectable lipoprotein lipase present at the surface of adipocytes remained constant throughout the 4 h incubation period at 37 degrees C. Under such conditions total cellular enzyme activity also remained constant, with no activity appearing in the medium. In the presence of heparin, cell-surface immunodetectable lipoprotein lipase increased by up to 20%, whereas in the presence of cycloheximide they decreased by up to 60%. Thus the obvious turnover of enzyme from this cell-surface site was found to be relatively rapid and dependent for its replenishment, at least in part, on protein synthesis. In the presence of insulin alone, a substantial increase in cell-surface lipoprotein lipase protein occurred, only part of which was dependent on protein synthesis. The total cellular activity of lipoprotein lipase was unaffected by the presence of insulin. The insulin-dependent increase in cell-surface enzyme was potentiated somewhat in the presence of dexamethasone, which was not shown to exert any independent effect. Glucagon, adrenaline and theophylline all produced a significant decline in the cell-surface immunodetectable lipoprotein lipase, which in the case examined (adrenaline) was partially additive with regard to the independent effect of cycloheximide. Cell-surface immunodetectable lipoprotein lipase amounts were decreased significantly when cells were incubated in the presence of either colchicine or tunicamycin. The concerted way in which cell-surface lipoprotein lipase altered during the incubations of adipocytes in the presence of effectors suggested that the translocation of enzyme to and from this cellular site was dependent on hormonal action and the integrity of intracellular protein-transport mechanisms.     

Functional topology of a surface loop shielding the catalytic center in lipoprotein lipase.

Lipoprotein lipase (LPL), hepatic lipase, and pancreatic lipase show high sequence homology to one another. The crystal structure of pancreatic lipase suggests that it contains a trypsin-like Asp-His-Ser catalytic triad at the active center, which is shielded by a disulfide bridge-bounded surface loop that must be repositioned before the substrate can gain access to the catalytic residues. By sequence alignment, the homologous catalytic triad in LPL corresponds to Asp156-His241-Ser132, absolutely conserved residues, and the homologous surface loop to residues 217-238, a poorly conserved region. To verify these assignments, we expressed in vitro wild-type LPL and mutant LPLs having single amino acid mutations involving residue Asp156 (to His, Ser, Asn, Ala, Glu, or Gly), His241 (to Asn, Ala, Arg, Gln, or Trp), or Ser132 (to Gly, Ala, Thu, or Asp) individually. All 15 mutant LPLs were totally devoid of enzyme activity, while wild-type LPL and other mutant LPLs containing substitutions in other positions were fully active. We further replaced the 22-residue LPL loop which shields the catalytic center either partially (replacing 6 of 22 residues) or completely with the corresponding hepatic lipase loop. The partial loop-replacement chimeric LPL was found to be fully active, and the complete loop-replacement mutant had approximately 60% activity, although the primary sequence of the hepatic lipase loop is quite different. In contrast, replacement with the pancreatic lipase loop completely inactivated the enzyme. Our results are consistent with Asp156-His241-Ser132 being the catalytic triad in lipoprotein lipase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hepatic lipase and HDL metabolism

Hepatic lipase is a lipolytic enzyme that has been suggested to have a role in HDL metabolism. Evidence suggests that HDL-cholesterol level is at least partly regulated by hepatic lipase level. Recent studies have shown that hepatic lipase not only hydrolyzes triglyceride and phospholipid in HDL, but also stimulates HDL cholesterol ester uptake by hepatocytes. Therefore, hepatic lipase, together with lipid transfer proteins, determines both HDL-cholesterol level and its function in reverse cholesterol transport. These conclusions are based on observations from in-vitro model substrate studies, cell culture studies, transgenic animal studies, and clinical studies. At present time, it is not known whether hepatic lipase action increases or decreases risk of developing atherosclerosis.     

Interaction of synthetic peptides of apolipoprotein C-II and lipoprotein lipase at monomolecular lipid films

The triacylglycerol hydrolyase and phospholipase A1 activities of bovine milk lipoprotein lipase toward long-chain fatty acyl ester substrates were investigated with monomolecular lipid films containing trioleoylglycerol and phosphatidylcholine. In a monolayer of egg phosphatidylcholine containing 3 mol% [14C]trioleoylglycerol, and in the presence of apolipoprotein C-II, a 79 amino acid activator protein for lipoprotein lipase, enzyme activity was maximal at a surface pressure of 21-22 mN X m-1 (37 mumol oleic acid released/h per mg enzyme); enzyme activity was enhanced 9-fold by apolipoprotein C-II. At surface pressures between 22 and 30 mN X m-1, lipoprotein lipase activity decreased over a broad range and was nearly zero at 30 mN X m-1. Apolipoprotein C-II and the synthetic fragments of the activator protein containing residues 56-79, 51-79 and 44-79 were equally effective at 20 mN X m-1 in enhancing lipoprotein lipase catalysis. However, at surface pressures between 25 and 29 mN X m-1, only apolipoprotein C-II and the phospholipid-associating fragment containing residues 44-79 enhanced enzyme catalysis. The effect of apolipoprotein C-II and synthetic peptides on the phospholipase A1 activity of lipoprotein lipase was examined in sphingomyelin:cholesterol (2:1) monolayers containing 5 mol% di[14C]myristoylphosphatidylcholine. At 22 mN X m-1, apolipoprotein C-II and the synthetic fragments containing residues 44-79 or 56-79 enhanced lipoprotein lipase activity (70-80 nmol/h per mg enzyme). In contrast to trioleoylglycerol hydrolysis, the synthetic fragments were not as effective as apolipoprotein C-II enhancing enzyme activity towards di[14C]myristoylphosphatidylcholine at higher surface pressures. We conclude that the minimal amino acid sequence of apolipoprotein C-II required for activation of lipoprotein lipase is dependent both on the lipid substrate and the packing density of the monolayer.     

Digestive lipases: from three-dimensional structure to physiology

Human gastric lipase (HGL) is a lipolytic enzyme that is secreted by the chief cells located in the fundic part of the stomach. HGL plays an important role in lipid digestion, since it promotes the subsequent hydrolytic action of pancreatic lipase in duodenal lumen. Physiological studies have shown that HGL is able of acting not only in the highly acid stomach environment but also in the duodenum in synergy with human pancreatic lipase (HPL). Recombinant HGL (r-HGL) was expressed in the baculovirus/insect cell system in the form of an active protein with a molecular mass of 45 kDa. The specific activities of r-HGL were found to be similar to that of the native enzyme when tested on various triacylglycerol (TG) substrates. The 3-D structure of r-HGL was the first solved within the mammalian acid lipase family. This globular enzyme (379 residues) shows a new feature, different from the other known lipases structures, which consists of a core domain having the alpha/beta hydrolase fold and a cap domain including a putative 'lid' of 30 residues covering the active site of the lipase (closed conformation). HPL is the major lipolytic enzyme involved in the digestion of dietary TG. HPL is a 50 kDa glycoprotein which is directly secreted as an active enzyme. HPL was the first mammalian lipase to be solved structurally, and it revealed the presence of two structural domains: a large N-terminal domain (residues 1-336) and a smaller C-terminal domain (residues 337-449). The large N-terminal domain belongs to the alpha/beta hydrolase fold and contains the active site. A surface loop called the lid domain (C237-C261) covers the active site in the closed conformation of the lipase. The 3-D structure of the lipase-procolipase complex illustrates how the procolipase might anchor the lipase at the interface in the presence of bile salts: procolipase binds to the C-terminal domain of HPL and exposes the hydrophobic tips of its fingers at the opposite site of its lipase-binding domain. These hydrophobic tips help to bring N-terminal domain into close conformation with the interface where the opening of the lid domain probably occurs. As a result of all these conformational changes, the open lid and the extremities of the procolipase form an impressive continuous hydrophobic plateau, extending over more than 50 A. This surface might able to interact strongly with a lipid-water interface. The biochemical, histochemical and clinical studies as well as the 3-D structures obtained will be a great help for a better understanding of the structure-function relationships of digestive lipases.     

Hepatic lipase: a marker for cardiovascular disease risk and response to therapy

PURPOSE OF REVIEW: Hepatic lipase plays a key role in the metabolism of pro-atherogenic and anti-atherogenic lipoproteins affecting their plasma level as well as their physico-chemical properties. However, controversial evidence exists concerning whether hepatic lipase is pro or anti-atherogenic. The goal of this review is to summarize recent evidence that connects the enzyme to cardiovascular disease. The potential impact of genetic determinants of hepatic lipase activity in modulating both the development of coronary and carotid atherosclerosis will be discussed based on hepatic lipase proposed roles in lipoprotein metabolism. RECENT FINDINGS: Twenty to 30% of individual variation of hepatic lipase activity is accounted for by the presence of a common polymorphism in the promoter region (-514 C to T) of the hepatic lipase gene (LIPC). This polymorphism, via its impact on hepatic lipase synthesis and activity, appears to contribute to (1) individual susceptibility to cardiovascular disease: the presence of the T allele (low hepatic lipase activity) may carry a marginally increased risk of atherosclerosis; (2) carotid plaque composition and individual susceptibility to cerebrovascular events: the presence of the C allele (high hepatic lipase activity) is associated with increased carotid intima-media thickness and abundance of macrophages in the carotid plaque (unstable plaque); and (3) response of cardiovascular disease patients to lipid-lowering therapy: patients with the CC genotype have the greatest clinical benefit from intensive lipid-lowering therapy. SUMMARY: Convincing evidence shows that hepatic lipase plays a key role in remnant lipoprotein catabolism as well as in remodeling of LDL and HDL particles. The anti or pro-atherogenic role of hepatic lipase is likely to be modulated by the concurrent presence of other lipid abnormalities (i.e. increased LDL cholesterol levels) as well as by the genetic regulation of other enzymes involved in lipoprotein metabolism. Characterization of patients by their LIPC genotype will contribute to a better definition of individual risk of coronary and cerebrovascular events, specifically in patients with qualitative (small, atherogenic LDL and low HDL2 cholesterol) rather than quantitative lipid abnormalities for whom the routine lipid profile may underestimate the risk of coronary and cerebrovascular disease.     

Hepatic lipase: structure/function relationship,synthesis, and regulation

Hepatic lipase (HL) is a lipolytic enzyme, synthesized by hepatocytes and found localized at the surface of liver sinusoid capillaries. In humans, the enzyme is mostly bound onto heparan-sulfate proteoglycans at the surface of hepatocytes and also of sinusoid endothelial cells. HL shares a number of functional domains with lipoprotein lipase and with other members of the lipase gene family. It is a secreted glycoprotein, and remodelling of the N-linked oligosaccharides appears to be crucial for the secretion process, rather than for the acquisition of the catalytic activity. HL is also present in adrenals and ovaries, where it might promote delivery of lipoprotein cholesterol for steroidogenesis. However, evidence of a local synthesis is still controversial. HL activity is fairly regulated according to the cell cholesterol content and to the hormonal status. Coordinate regulations have been reported for both HL and the scavenger-receptor B-I, suggesting complementary roles in cholesterol metabolism. However, genetic variants largely contribute to HL variability and their possible impact in the development of a dyslipidemic phenotype, or in a context of insulin-resistance, is discussed.     

Lipoprotein lipase activity in bovine aorta.

A lipoprotein lipase in the bovine arterial wall has been identified and partially characterized. The enzyme has a Km apparent of 1 mM for triolein in a phosphatidylcholine stabilized emulsion. The lipase was stimulated 20- to 30-fold by the addition of heated rat plasma to the assay medium. The activity exhibited a pH optimum at 8.6. Protamine sulfate (1.0 mg/ml) inhibited the activity by 50%, whereas 1.4 M sodium chloride inhibited by 85%. Sodium fluoride, an inhibitor of the hormone-sensitive lipase, had no effect on the activity. Additions of low concentrations of heparin or Ca-2+ to the enzyme caused a slight stimulation of the lipolytic activity. A crude sectioning of the aorta revealed specific activity of lipoprotein lipase to be highest at the endothelial side of the artery.     

Lipoprotein lipases and vitellogenins in relation to the known three-dimensional structure of pancreatic lipase

A 106-residue region of high similarity between lipoprotein/pancreatic/hepatic lipases and Drosophila vitellogenins encompasses four beta-strands with all residues but one strictly conserved or conservatively replaced between the structures, and enclosing the putative active site Ser-152. The properties suggest a common folding pattern but the region probably does not function as an 'interface recognition site' in the lipases, although it might well bind fatty acid esters of ecdysteroids or single lipid molecules in the vitellogenins. C-terminally of this 106-residue region, a surface loop ('flap') covers the active site. No residue within this loop is conserved through all lipases, but adjacent segments exhibit 60-70% residue identity. Hepatic and lipoprotein lipases probably hydrolyze both soluble and emulsified substrates at the same site. They lack residues corresponding to a second active site postulated in pancreatic lipase to account for hydrolysis of soluble substrates. In addition, due to structural differences the flap could prevent entry of soluble substrate molecules into the active site of pancreatic lipase.     

In vivo evidence of a role for hepatic lipase in human apoB-containing lipoprotein metabolism, independent of its lipolytic activity

Hepatic lipase (HL) is a key player in lipoprotein metabolism by modulating, through its lipolytic activity, the triglyceride (TG) and phospholipid content of apolipoprotein B (apoB)-containing lipoproteins and of high density lipoproteins (HDL), thereby affecting their size and density. A new and separate role has been suggested for HL in cellular lipoprotein metabolism, in which it serves as a ligand promoting cellular uptake of apoB-containing remnant lipoproteins and HDL. We tested the hypothesis that HL has both a lipolytic and a nonlipolytic role in human lipoprotein metabolism, by measuring lipid plasma concentrations, lipoprotein density distribution by density gradient ultracentrifugation, and lipoprotein composition, in three subjects with HL deficiency: two of the patients (S-1 and S-3) were characterized as having neither plasma HL activity nor detectable HL protein; the third subject (S-2) had no plasma HL activity but a detectable amount (35.5 ng/ml) of HL protein. All HL-deficient subjects showed a severalfold increase in lipoprotein TG content across the lipoprotein density spectrum [very low density lipoprotein (VLDL), intermediate density lipoprotein (IDL), low density lipoprotein (LDL), and HDL] as compared with control subjects. They also had remarkably more buoyant LDL particles (LDL-R(f) = 0.342;-0.394) as compared with the control subjects (LDL-R(f) = 0.303). Subjects S-1 and S-3 (no HL activity or protein) presented with a distinct increase in cholesterol and apoB levels in the IDL and VLDL density range as compared with patient S-2, with detectable HL protein, and the control subjects.This study provides evidence in humans that HL indeed plays an important role in lipoprotein metabolism independent of its enzymatic activity: in particular, inactive HL protein appears to affect VLDL and IDL particle concentration, whereas HL enzymatic activity seems to influence VLDL-, IDL-, LDL-, and HDL-TG content and their physical properties.