The hepatic uptake of VLDL in lrp-ldlr-/-vldlr-/- mice is regulated by LPL activity and involves proteoglycans and SR-BI

LPL activity plays an important role in preceding the VLDL remnant clearance via the three major apolipoprotein E (apoE)-recognizing receptors: the LDL receptor (LDLr), LDL receptor-related protein (LRP), and VLDL receptor (VLDLr). The aim of this study was to determine whether LPL activity is also important for VLDL remnant clearance irrespective of these receptors and to determine the mechanisms involved in the hepatic remnant uptake. Administration of an adenovirus expressing LPL (AdLPL) into lrp(-)ldlr(-/-)vldlr(-/-) mice reduced both VLDL-triglyceride (TG) and VLDL-total cholesterol (TC) levels. Conversely, inhibition of LPL by AdAPOC1 increased plasma VLDL-TG and VLDL-TC levels. Metabolic studies with radiolabeled VLDL-like emulsion particles showed that the clearance and hepatic association of their remnants positively correlated with LPL activity. This hepatic association was independent of the bridging function of LPL and HL, since heparin did not reduce the liver association. In vitro studies demonstrated that VLDL-like emulsion particles avidly bound to the cell surface of primary hepatocytes from lrp(-)ldlr(-/-)vldlr(-/-) mice, followed by slow internalization, and involved heparin-releaseable cell surface proteins as well as scavenger receptor class B type I (SR-BI). Collectively, we conclude that hepatic VLDL remnant uptake in the absence of the three classical apoE-recognizing receptors is regulated by LPL activity and involves heparan sulfate proteoglycans and SR-BI.     

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.     

The effect of added monoacylglycerols on the removal from plasma of chylomicron-like emulsions injected intravenously in rats

Lipid emulsions were prepared with compositions similar to the triacylglycerol-rich plasma lipoproteins, but also incorporating added small amounts of monoacylglycerols. Control emulsions without monoacylglycerol were metabolized similarly to natural chylomicrons or very-low-density lipoproteins when injected intravenously in rats. The emulsion triacylglycerols and cholesteryl esters were both removed rapidly from the bloodstream, with the removal rates of triacylglycerols faster than those of cholesteryl esters. Much of the removed cholesteryl ester was found in the liver, but only a small fraction of the triacylglycerol, consistent with hepatic uptake of the triacylglycerol-depleted remnants of the injected emulsion. Emulsions incorporating added monooleoylglycerol or stearic acid were metabolized similarly. Added 1- or 2-monostearoylglycerol had no effect on triacylglycerol removal from plasma, but the removal rate of cholesteryl esters was decreased and less cholesteryl ester was found in the liver. These effects are similar to those recently described when emulsions and chylomicrons contained triacylglycerols with a saturated acyl chain at the glycerol 2-position, suggesting that saturated monoacylglycerol produced by the action of lipoprotein lipase may cause triacylglycerol-depleted remnant particles to remain in the plasma instead of being rapidly taken up by the liver.     

Removal of chylomicron remnants in transgenic mice overexpressing normal and membrane-anchored hepatic lipase

The LDL receptor and the LDL receptor-related protein (LRP) mediate the removal of chylomicron remnants. The LRP pathway involves sequestration of particles in the space of Disse. It has been proposed that either alone or in combination with other factors, such as apolipoprotein E and proteoglycans, hepatic lipase (HL) may contribute to the sequestration of chylomicron remnants. To test this hypothesis, we generated two lines of transgenic mice producing rat HL as a native or as a membrane-anchored form. These animals express HL at levels similar to normal rat. Chylomicron remnants were perfused in a single nonrecirculating pass into the livers of the rat HL transgenic, HL-deficient, and wild-type (WT) mice for 20 min, and the rate of chylomicron remnant removal was measured. Chylomicron remnants were removed at a rate of approximately 50% per pass in WT mice. It was slightly increased in both transgenic mice and reduced in HL-deficient mice compared with the WT mice. Confocal microscopy of liver sections showed that a modest amount of HL colocalized with chylomicron remnant clusters in the transgenic mice, suggesting that HL is a component of the LRP-proteoglycan clusters. These data suggest that HL helps to direct cholesterol to the tissues in which it is localized by a nonenzymatic mechanism.     

Inhibitory effects of C apolipoproteins from rats and humans on the uptake of triglyceride-rich lipoproteins and their remnants by the perfused rat liver

Like rat C apolipoproteins, each of the C apolipoproteins from human blood plasma (C-I, C-II, C-III-1, and C-III-2) bound to small chylomicrons from mesenteric lymph of estradiol-treated rats and inhibited their uptake by the isolated perfused rat liver. This inhibitory effect of the C apolipoproteins was independent of apolipoprotein E, which is present only in trace amounts in these chylomicrons. Addition of rat apolipoprotein E to small chylomicrons from mesenteric lymph of normal rats did not displace C apolipoproteins and had no effect on the uptake of these particles by the perfused liver, indicating that an increased ratio of E apolipoproteins to C apolipoproteins on chylomicron particles, unaccompanied by depletion of the latter, may not promote recognition by the chylomicron remnant receptor. The hepatic uptake of remnants of rat hepatic very low density lipoproteins (VLDL) and small chylomicrons, which had been produced in functionally eviscerated rats, was also inhibited by addition of C apolipoproteins. These observations are consistent with the hypothesis that the addition of all of the C apolipoproteins to newly secreted chylomicrons and VLDL inhibits premature uptake of these particles by the liver and that depletion of all of these apolipoproteins from remnant particles facilitates their hepatic uptake. Remnants of chylomicrons and VLDL incubated with rat C apolipoproteins efficiently took up C-III apolipoproteins, but not apolipoprotein C-II (the activator protein for lipoprotein lipase). Preferential loss of apolipoprotein C-II during remnant formation may regulate the termination of triglyceride hydrolysis prior to complete removal of triglycerides from chylomicrons and VLDL.     

Plasma clearance and liver uptake of chylomicron remnants generated by hepatic lipase lipolysis: evidence for a lactoferrin-sensitive and apolipoprotein E-independent pathway

Chylomicrons labeled with [3H]cholesterol and [14C]triglyceride fatty acids were lipolyzed by hepatic lipase (HL) in vitro and then injected intravenously into normal mice fed low- or high-fat diets, and into apolipoprotein (apo) E-deficient mice. In normal mice fed the high-fat diet and injected with non-lipolyzed chylomicrons, the plasma clearance and hepatic uptake of the resulting [3H]cholesterol-labeled remnants was markedly inhibited. In contrast, chylomicrons lipolyzed by HL were taken up equally rapidly by the livers of mice fed the low- and high-fat diets. The removal of non-lipolyzed chylomicrons lacking apoE from the plasma of apoE-deficient mice was inhibited, but not the removal of chylomicrons lipolyzed by HL. Pre-injection of lactoferrin into normal mice inhibited the plasma clearance of both non-lipolyzed chylomicrons and chylomicrons lipolyzed by HL. The removal of HL from the surface of the lipolyzed particles by proteolytic digestion did not affect their rapid uptake, indicating that the hepatic recognition of the lipoproteins was not mediated by HL. These observations support previous findings that phospholipolysis of chylomicrons by hepatic lipase generates remnant particles that are rapidly cleared from circulation by the liver. They also support the concept that chylomicron remnants can be taken up by the liver by an apolipoprotein E-independent mechanism. We hypothesize that this mechanism is modulated by the remnant phospholipids and that it may involve their interaction with a phospholipid-binding receptor on the surface of hepatocytes such as the class B scavenger receptor BI.     

Heparan sulfate proteoglycans and triglyceride-rich lipoprotein metabolism

PURPOSE OF REVIEW: Clearance of triglyceride-rich lipoprotein remnants by the liver is a key step in preventing hypertriglyceridemia, an independent risk factor for cardiovascular disease. We review recent genetic evidence that heparan sulfate proteoglycans work in concert with the LDL receptor in the liver to facilitate binding and clearance of both triglyceride and cholesterol-rich lipoproteins from the circulation. RECENT FINDINGS: Partial reduction of sulfation of liver heparan sulfate using the Cre-loxP system caused accumulation of hepatic and dietary triglyceride-rich lipoprotein particles due to delayed clearance. Compounding the mutation with LDL receptor deficiency caused enhanced accumulation of both cholesterol and triglyceride-rich particles compared with mice lacking only LDL receptors. These findings provide the first genetic evidence that hepatic heparan sulfate proteoglycans play a central role in the clearance of lipoproteins by the liver and work independently of LDL receptors. SUMMARY: A role for hepatocyte heparan sulfate in lipoprotein metabolism has now been genetically established in mice. Given this finding, mild, but clinically relevant, hyperlipidemias in human patients may be a result of alterations in heparan sulfate structure or possible genetic polymorphisms in the relevant biosynthetic genes.     

Effects of threonine-poor apolipoproteins on post-heparin plasma hepatic triacylglycerol lipase and lipoprotein lipase

Whole-irradiated rabbit pre-heparin plasma had an important inhibitory effect on hepatic triacylglycerol lipase and lipoprotein lipase activities, whereas control rabbit pre-heparin plasma slightly inhibited hepatic triacylglycerol lipase activity at a high concentration and enhanced lipoprotein lipase activity. As some apolipoproteins were known to modulate these two lipolytic enzymes, the inhibitory effects of irradiated rabbit plasma were investigated in apolipoproteins. Three apolipoproteins, with isoelectric points of about 6.58, 6.44 and 6.12, characterized by their low content in threonine (threonine-poor apolipoproteins) were produced in high concentrations in rabbit VLDL and HDL after irradiation. The effects of these apolipoproteins on control rabbit post-heparin plasma hepatic triacylglycerol lipase and extrahepatic lipoprotein lipase were studied. Threonine-poor apolipoproteins substantially inhibited the hepatic triacylglycerol lipase activity and enhanced the apolipoprotein C-II-stimulated activity of lipoprotein lipase. The amounts of these apolipoproteins in triacylglycerol-rich lipoprotein particles may determine the lipolytic activity of lipoprotein lipase and hepatic triacylglycerol lipase in triacylglycerol hydrolysis. The existence of another inhibitor of lipoprotein lipase remains to be determined.     

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.     

Mouse preheparin plasma contains high levels of hepatic lipase with low affinity for heparin

It was recently noted that newborn mice have much higher lipase activity in plasma than rats or humans, and that most of the activity is due to an enzyme related to the hepatic (heparin-releasable) lipase. Here we report that this lipase is present in plasma of adult mice also. In contrast to the high activity of hepatic lipase, the activity of lipoprotein lipase in plasma was low and similar to that in rats. The source of the plasma lipase was probably the liver, since we could not demonstrate hepatic lipase-like activity in any other organ. When human hepatic lipase was injected into mice, it rapidly disappeared from plasma. Most of the injected lipase located in the liver, and could be released back into circulation by injection of heparin. These results indicate that there are binding sites for hepatic lipase in mouse liver, and suggest that mouse hepatic lipase has an affinity for these sites which is lower than usual. It is currently believed that the endothelial acceptors are heparan-sulfate or similar molecules. Mouse hepatic lipase eluted from heparin-Sepharose at lower salt concentration than rat or human hepatic lipase, demonstrating that it has a relatively low affinity for heparin-like polysaccharides.     

Distribution of lipoprotein lipase and hepatic lipase between plasma and tissues: effect of hypertriglyceridemia

Lipoprotein lipase and hepatic lipase were measured in rat plasma using specific antisera. Mean values for lipoprotein lipase in adult rats were 1.8-3.6 mU/ml, depending on sex and nutritional state. Values for hepatic lipase were about three times higher. Lipoprotein lipase activity in plasma of newborn rats was 2-4-times higher than in adults. In contrast, hepatic lipase activity was lower in newborn than in adult rats. Following functional hepatectomy there was a progressive increase in lipoprotein lipase activity in plasma, indicating that transport of the enzyme from peripheral tissues to the liver normally takes place. Lipoprotein lipase, but not hepatic lipase, increased in plasma after a fat meal. An even more marked increase, up to 30 mU/ml, was seen after intravenous injection of Intralipid. Plasma lipase activity decreased in parallel with clearing of the injected triacylglycerol. 125I-labeled lipoprotein lipase injected intravenously during the hyperlipemia disappeared somewhat slower from the circulation than in fasted rats, but the uptake was still primarily in the liver. Hyperlipemia, or injection of heparin, led to increased lipoprotein lipase activity in the liver. This was seen even when the animals had been pretreated with cycloheximide to inhibit synthesis of new enzyme protein. These results suggest that during hypertriglyceridemia lipoprotein lipase binds to circulating lipoproteins/lipid droplets which results in increased plasma levels of the enzyme and increased transport to the liver.     

Phosphatidylcholine biosynthesis and lipoprotein metabolism

Phosphatidylcholine (PC) is the major phospholipid component of all plasma lipoprotein classes. PC is the only phospholipid which is currently known to be required for lipoprotein assembly and secretion. Impaired hepatic PC biosynthesis significantly reduces the levels of circulating very low density lipoproteins (VLDLs) and high density lipoproteins (HDLs). The reduction in plasma VLDLs is due in part to impaired hepatic secretion of VLDLs. Less PC within the hepatic secretory pathway results in nascent VLDL particles with reduced levels of PC. These particles are recognized as being defective and are degraded within the secretory system by an incompletely defined process that occurs in a post-endoplasmic reticulum compartment, consistent with degradation directed by the low-density lipoprotein receptor and/or autophagy. Moreover, VLDL particles are taken up more readily from the circulation when the PC content of the VLDLs is reduced, likely due to a preference of cell surface receptors and/or enzymes for lipoproteins that contain less PC. Impaired PC biosynthesis also reduces plasma HDLs by inhibiting hepatic HDL formation and by increasing HDL uptake from the circulation. These effects are mediated by elevated expression of ATP-binding cassette transporter A1 and hepatic scavenger receptor class B type 1, respectively. Hepatic PC availability has recently been linked to the progression of liver and heart disease. These findings demonstrate that hepatic PC biosynthesis can regulate the amount of circulating lipoproteins and suggest that hepatic PC biosynthesis may represent an important pharmaceutical target. This article is part of a Special Issue entitled Triglyceride Metabolism and Disease.     

Absorption and lipoprotein transport of sphingomyelin

Dietary sphingomyelin (SM) is hydrolyzed by intestinal alkaline sphingomyelinase and neutral ceramidase to sphingosine, which is absorbed and converted to palmitic acid and acylated into chylomicron triglycerides (TGs). SM digestion is slow and is affected by luminal factors such as bile salt, cholesterol, and other lipids. In the gut, SM and its metabolites may influence TG hydrolysis, cholesterol absorption, lipoprotein formation, and mucosal growth. SM accounts for approximately 20% of the phospholipids in human plasma lipoproteins, of which two-thirds are in LDL and VLDL. It is secreted in chylomicrons and VLDL and transferred into HDL via the ABCA1 transporter. Plasma SM increases after periods of large lipid loads, during suckling, and in type II hypercholesterolemia, cholesterol-fed animals, and apolipoprotein E-deficient mice. SM is thus an important amphiphilic component when plasma lipoprotein pools expand in response to large lipid loads or metabolic abnormalities. It inhibits lipoprotein lipase and LCAT as well as the interaction of lipoproteins with receptors and counteracts LDL oxidation. The turnover of plasma SM is greater than can be accounted for by the turnover of LDL and HDL particles. Some SM must be degraded via receptor-mediated catabolism of chylomicron and VLDL remnants and by scavenger receptor class B type I receptor-mediated transfer into cells.     

Uptake of chylomicron remnants by the liver: further evidence for the modulating role of phospholipids

Rat lymph chylomicrons were treated with rat heparin-releasable hepatic lipase (HL) or with bovine milk lipoprotein lipase (LPL). The ability of the resulting particles to be taken up by the liver in vivo was assessed following their infusion into the portal vein of partially hepatectomized animals. The following observations were made: a) the rate of phospholipid depletion, relative to the rate of triglyceride hydrolysis, induced by HL was two- to threefold higher than that observed for LPL; b) the depletion of at least 57% of phospholipids from the surface of HL-treated chylomicrons caused no major alterations in the apoprotein profile of the particles; c) for the same extent of triglyceride hydrolysis, HL-treated chylomicrons were taken up by liver at a rate significantly higher (P less than 0.005) than LPL-treated particles; d) the liver uptake of HL-treated chylomicrons was competitively inhibited by endogenously generated chylomicron remnants, indicating that these two types of lipoproteins share the same process of recognition and uptake by liver cells. It is concluded that the in vivo changes in phospholipid content, or composition, on the surface of chylomicrons during their transformation into remnants, modulate the differentiation of these two particles by the hepatic remnant receptor.     

Mechanism of the hypertriglyceridemia induced by tumor necrosis factor administration to rats

4 h after intravenous injection of recombinant HuTNF-alpha to fed rats, an increase in heart, diaphragm, and plasma lipoprotein lipase activity was observed. At the same time, a 40-60% decrease in enzymic activity in epididymal fat pad and kidney and 40% decrease in hepatic lipase activity in liver had occurred. Similar results were obtained 20 h after injection of recombinant HuTNF-alpha into fasted rats. Pretreatment with Indomethacin did not affect the changes in tissue lipoprotein lipase activity observed following recombinant HuTNF-alpha administration. Serum triacylglycerol concentration increased by 2- and 6-fold; 4 and 20 h after recombinant HuTNF-alpha administration. Disappearance of 14C-labeled triacylglycerol from the circulation after injection of small chylomicrons, biosynthetically labeled in their triacylglycerol and cholesterol moieties, was lower in TNF-treated than in control rats. However, the clearance rate of triacylglycerol was the same or even higher in recombinant HuTNF-alpha treated rats (assuming that 14C-labeled chylomicron triacylglycerol represents the serum triacylglycerol pool). The livers of recombinant HuTNF-alpha-treated rats and controls contained similar amounts of 14C-labeled lipids, but less [3H]cholesterol, suggesting that in recombinant HuTNF-alpha-treated rats, the liver took up chylomicron remnant particles enriched with triacylglycerol. Separation of the d less than 1.04 g/ml fraction of serum obtained from control and recombinant HuTNF-alpha treated rats by zonal ultracentrifugation revealed that in recombinant HuTNF-alpha-treated rats the lipoprotein particles were less lipolyzed than in controls. The secretion rate of [3H]triacylglycerol into the serum was determined 90 min after injection of [3H]palmitate albumin complex and Triton WR 1339. In recombinant HuTNF-alpha-treated rats, the secretion of [3H]triacylglycerol into plasma was 48% higher than in controls. It is suggested that the increase in lipoprotein lipase activity of heart and diaphragm resulted from an indirect effect of TNF. It is concluded that the increase in serum triacylglycerol in the recombinant HuTNF-alpha-treated rats is due mainly to an increased secretion of triacylglycerol by the liver. Impaired lipolysis, probably due to a fall in hepatic lipase could also contribute to the rise in plasma triacylglycerol.     

Recombinant human bone morphogenetic protein-2 stimulates differentiation in primary cultures of fetal rat calvarial osteoblasts

Molecular mechanisms of lipid synthesis and their controls in hepatic stellate cells are not known. We have previously proposed that, in contrast to other fat storing cells, hepatic stellate cells are not involved in energy storage, but they represent a particular cell population specialized in storage of lipid-soluble substances, the major one being probably retinol. In agreement with this hypothesis, induction of the lipocyte phenotype in stellate cells is not under the control of insulin, but responds to retinoids and other molecules that modify the gene expression program in these cells. In the present study we have monitored the activity of the two major enzymes involved in lipid synthesis during the induction of the lipocyte phenotype in hepatic stellate cells: glycerol-3-phosphate dehydrogenase (GPDH) that mediates the de novo lipid synthesis, and lipoprotein lipase that mediates incorporation of plasma lipids. In early stages of lipocyte induction, both pathways of lipid synthesis are activated. When lipocytes have already constituted the lipid droplets, lipoprotein lipase pathway is downregulated, while GPDH activity remains high. Adult liver has been reported to lack lipoprotein lipase, but under stress, lipase activity was detected around and at the surface of the intrahepatic vasculature. We have now shown that the lipase activity can be induced in the hepatic stellate cells, located in the Disse's space. The high lipoprotein lipase activity under acute induction of lipocyte phenotype, followed by the low activity under conditions of metabolic equilibrium, are in compass with the increased activity of this enzyme under stress, and its low activity in adult liver parenchyma under normal conditions.     

The uptake of chylomicron remnants and very low density lipoprotein remnants by the perfused rat liver

The regulation of the hepatic uptake of chylomicron remnants and very-low-density lipoprotein (VLDL) remnants was studied in the rat using a nonrecirculating liver perfusion system. The hepatic removal of remnant lipoproteins was shown to be by receptor-mediated processes since the concentration-dependent uptake was saturable and reductive methylation of the particles reduced the uptake of each lipoprotein by two-thirds. Treatment of liver donor rats with 17 alpha-ethinyl estradiol resulted in a 2-fold increase in the hepatic uptake of VLDL remnants, while cholesterol feeding of liver donor rats caused complete suppression of the receptor-mediated uptake of VLDL remnants. Chylomicron remnant removal was unaffected by estradiol administration and only slightly diminished by cholesterol feeding. The results of competition studies also indicated that a specific chylomicron remnant receptor exists in the liver. Apoprotein E was shown to be required for the receptor-mediated uptake of both remnant lipoproteins. Chylomicron remnants which contained no apoprotein E and VLDL remnants which contained reductively methylated apoprotein E were removed by the liver to about one-third of the extent of native particles. Thus the hepatic uptake of remnant lipoproteins occurs by receptor-mediated processes and the specific removal of both particles is mediated by apoprotein E. In addition, the uptake of VLDL remnants is regulated by the same factors that control hepatic low-density lipoprotein removal, while chylomicron remnant removal is unaffected by these factors.     

Low density lipoprotein receptor-related protein and gp330 bind similar ligands, including plasminogen activator-inhibitor complexes and lactoferrin, an inhibitor of chylomicron remnant clearance.

The low density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor (LRP) and gp330, two members of the low density lipoprotein receptor gene family, share a multitude of cysteine-rich repeats. LRP has been shown to act as an endocytosis-mediating receptor for several ligands, including protease-antiprotease complexes and plasma lipoproteins. The former include alpha 2-macroglobulin-protease complexes and plasminogen activator inhibitor-activator complexes. The latter include chylomicron remnant-like particles designated beta-very low density lipoproteins (beta-VLDL) complexed with apoprotein E or lipoprotein lipase. The binding specificity of gp330 is unknown. In the current studies we show that gp330 from rat kidney membranes binds several of these ligands on nitrocellulose blots. We also show that both LRP and gp330 bind an additional ligand, bovine lactoferrin, which is known to inhibit the hepatic clearance of chylomicron remnants. Lactoferrin blocked the LRP-dependent stimulation of cholesteryl ester synthesis in cultured human fibroblasts elicited by apoprotein E-beta-VLDL or lipoprotein lipase-beta-VLDL complexes. Cross-competition experiments in fibroblasts showed that the multiple ligands recognize at least three distinct, but partially overlapping sites on the LRP molecule. Binding of all ligands to LRP and gp330 was inhibited by the 39-kDa protein, which co-purifies with the two receptors, suggesting that the 39-kDa protein is a universal regulator of ligand binding to both receptors. The correlation of the inhibitory effects of lactoferrin in vivo and in vitro support the notion that LRP functions as a chylomicron remnant receptor in liver. LRP and gp330 share a multiplicity of binding sites, and both may function as endocytosis-mediating receptors for a large number of ligands in different organs.     

Hydrolysis of chylomicron polyenoic fatty acid esters with lipoprotein lipase and hepatic lipase

The lipolysis of rat chylomicron polyenoic fatty acid esters with bovine milk lipoprotein lipase and human hepatic lipase was examined in vitro. Chylomicrons obtained after feeding fish oil or soy bean oil emulsions were used as substrates. The lipolysis was followed by gas chromatography or by using chylomicrons containing radioactive fatty acids. Lipoprotein lipase hydrolyzed eicosapentaenoic (20:5) and arachidonic acid (20:4) esters at a slower rate than the C14-C18 acid esters. More 20:5 and 20:4 thus accumulated in remaining tri- and diacylglycerols. Eicosatrienoic, docosatrienoic and docosahexanoic acids exhibited an intermediate lipolysis pattern. When added together with lipoprotein lipase, hepatic lipase increased the rate of lipolysis of 20:5 and 20:4 esters of both tri- and diacylglycerols. Addition of NaCl (final concentration 1 M) during the course of lipolysis inhibited lipoprotein lipase as well as the enhancing effect of hepatic lipase on triacylglycerol lipolysis. Hepatic lipase however, hydrolyzed diacylglycerol that had already been formed. Chylomicron 20:4 and 20:5 esters thus exhibit a relative resistance to lipoprotein lipase. It is suggested that the tri- and diacylglycerol species containing these fatty acids may accumulate at the surface of the remnant particles and act as substrate for hepatic lipase during a concerted action of this enzyme and lipoprotein lipase.     

Fatty acid-induced atherogenic changes in extracellular matrix proteoglycans

PURPOSE OF REVIEW: Nonesterified fatty acids change the expression and properties of the extracellular matrix proteoglycans of arterial and hepatic cells. We review how this may contribute to arterial disease in insulin resistance and type 2 diabetes. RECENT FINDINGS: Elevated nonesterified fatty acids characterize the dyslipidemia of insulin resistance and type 2 diabetes. In hepatocytes high levels of fatty acids cause changes in proteoglycans leading to a matrix with decreased affinity for VLDL remnants. Furthermore, liver proteoglycans from insulin resistant hyperlipidemic Zucker rats showed alterations also associated with decreased remnant affinity. In arterial smooth muscle cells overexposure to fatty acids augmented expression of matrix proteoglycans for which LDL showed increased affinity. Fatty acids appeared to compromise insulin signaling by protein kinase C activation. The observed fatty acid-induced changes in matrix proteoglycans in liver and arteries can be an important component of the atherogenicity of the dyslipidemia of insulin resistance and type 2 diabetes. SUMMARY: Overexposure to fatty acids can contribute to generate a remnant-rich dyslipidemia and to precondition the arterial intima for lipoprotein deposition via changes in expression of matrix proteoglycans. Normalizing fatty acid should be a key target in treatment of the atherogenic dyslipidemia of insulin resistance.