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G Olivecrona

Publications and source records attributed to G Olivecrona.

At least 37 records · Page 2Linked to original sources
PubMed2000

Nutritional regulation of binding sites for lipoprotein lipase in rat heart.

Several laboratories have shown that when rats are fasted, the amount of lipoprotein lipase (LPL) at the vascular endothelium in heart (monitored as the amount released by heparin) increases severalfold without corresponding changes in the production of LPL. This suggests that there is a change in endothelial binding of LPL. To study this, (125)I-labeled bovine LPL was injected. The fraction that bound in the heart was more than twice as high in fasted than in fed rats, 4.3% compared with 1.9% of the injected dose. Refeeding reversed this in 5 h. When unlabeled LPL was injected before the tracer, the fraction of (125)I-LPL that bound in heart decreased, indicating that the binding was saturable. When isolated hearts were perfused at 4 degrees C with a single pass of labeled LPL, twice as much bound in hearts of fasted rats. We conclude that fasting causes a change in the vascular endothelium in heart such that its ability to bind LPL increases.

Adipose Tissue↗
PubMed2000

Functional characterization of 4 polymorphisms in promoter region of hepatic lipase gene.

Hepatic lipase (HL) is a lipolytic enzyme involved in the metabolism of plasma lipoproteins, especially high density lipoproteins. Association studies have provided strong evidence for relations of common mutations in the promoter region of the HL gene to postheparin plasma HL activity and the plasma high density lipoprotein cholesterol concentration, but the functional relevance of these polymorphisms has not been evaluated to date. We analyzed the physiological significance of 4 common polymorphisms (-250G/A, -514C/T, -710T/C, and -763A/G, all in strong linkage disequilibrium) in the promoter of the HL gene by use of electrophoretic mobility shift assays and transient transfection studies in HepG2 cells. No consistent evidence was found for a significant contribution of any of these polymorphisms to the basal rate of transcription of the HL gene. These data suggest that the 4 polymorphisms in the promoter region of the HL gene are in linkage disequilibrium with >/=1 as-yet-unknown functional polymorphisms in the HL gene locus with a significant effect on HL metabolism and/or enzymatic activity.

Adult↗
PubMed1999

The second and fourth cluster of class A cysteine-rich repeats of the low density lipoprotein receptor-related protein share ligand-binding properties.

The low density lipoprotein receptor-related protein (LRP) is a multifunctional endocytic cell-surface receptor that binds and internalizes a diverse array of ligands. The receptor contains four putative ligand-binding domains, generally referred to as clusters I, II, III, and IV. In this study, soluble recombinant receptor fragments, representing each of the four individual clusters, were used to map the binding sites of a set of structurally and functionally distinct ligands. Using surface plasmon resonance, we studied the binding of these fragments to methylamine-activated alpha(2)-macroglobulin, pro-urokinase-type plasminogen activator, tissue-type plasminogen activator (t-PA), plasminogen activator inhibitor-1, t-PA.plasminogen activator inhibitor-1 complexes, lipoprotein lipase, apolipoprotein E, tissue factor pathway inhibitor, lactoferrin, the light chain of blood coagulation factor VIII, and the intracellular chaperone receptor-associated protein (RAP). No binding of the cluster I fragment to any of the tested ligands was observed. The cluster III fragment only bound to the anti-LRP monoclonal antibody alpha(2)MRalpha3 and weakly to RAP. Except for t-PA, we found that each of the ligands tested binds both to cluster II and to cluster IV. The affinity rate constants of ligand binding to clusters II and IV and to LRP were measured, showing that clusters II and IV display only minor differences in ligand-binding kinetics. Furthermore, we demonstrate that the subdomains C3-C7 of cluster II are essential for binding of ligands and that this segment partially overlaps with a RAP-binding site on cluster II. Finally, we show that one RAP molecule can bind to different clusters simultaneously, supporting a model in which RAP binding to LRP induces a conformational change in the receptor that is incompatible with ligand binding.

Binding Sites↗
PubMed1999

Sortilin/neurotensin receptor-3 binds and mediates degradation of lipoprotein lipase.

Lipoprotein lipase and the receptor-associated protein (RAP) bind to overlapping sites on the low density lipoprotein receptor-related protein/alpha2-macroglobulin receptor (LRP). We have investigated if lipoprotein lipase interacts with the RAP binding but structurally distinct receptor sortilin/neurotensin receptor-3. We show, by chemical cross-linking and surface plasmon resonance analysis, that soluble sortilin binds lipoprotein lipase with an affinity similar to that of LRP. The binding was inhibited by heparin and RAP and by the newly discovered sortilin ligand neurotensin. In 35S-labeled 3T3-L1 adipocytes treated with the cross-linker dithiobis(succinimidyl propionate), lipoprotein lipase-containing complexes were isolated by anti-sortilin antibodies. To elucidate function in cells, sortilin-negative Chinese hamster ovary cells were transfected with full-length sortilin and shown to express about 8% of the receptors on the cell surface. These cells degraded 125I-labeled lipoprotein lipase much faster than the wild-type cells. The degradation was inhibited by unlabeled lipoprotein lipase, indicating a saturable pathway, and by RAP and heparin. Moreover, inhibition by the weak base chloroquine suggested that degradation occurs in an acidic vesicle compartment. The results demonstrate that sortilin is a multifunctional receptor that binds lipoprotein lipase and, when expressed on the cell surface, mediates its endocytosis and degradation.

Adaptor Proteins, Vesicular Transport↗
PubMed1999

Apolipoprotein CIII from guinea pig (Cavia porcellus) is shorter and less homologous than apolipoprotein CIII from other mammals.

Apolipoprotein (apo) CIII plays an important role in metabolism of triglyceride-rich lipoproteins as a regulator of lipolysis and/or lipoprotein-receptor interaction. With the method of RT-PCR, the cDNA of guinea pig apo CIII was cloned and sequenced. The deduced amino acid sequence of 91 amino acids residues consists of a highly conserved signal peptide of 20 residues and a mature protein of 71 residues. Compared to mouse, rat, dog, bovine and human apo CIII, guinea pig apo CIII has a deletion of eight or nine amino acids at its C-terminus and it shows the lowest degree of homology to the presently known apo CIII sequences. Interestingly, the most conserved areas of guinea pig apo CIII are found in two regions, residues 16-33 and residues 50-69. Corresponding regions in human and dog apo CIII were previously predicted to form amphipathic helices, which are assumed to play important roles in the inhibition of lipoprotein lipase (LPL) and binding to lipid. Our present study could be helpful for the future elucidation of the structure-function relationships and evolution of apo CIII.

Amino Acid Sequence↗
PubMed1999

Chylomicron metabolism in an animal model for hyperlipoproteinemia type I.

Mink homozygous for the mutation Pro214Leu in lipoprotein lipase (LPL) had only traces of LPL activity but amounts of LPL protein in their tissues similar to those of normal mink. In normal mink, lymph chylomicrons from rats given [3H]retinol (incorporated into retinyl esters, providing a core label) and [14C]oleic acid (incorporated mainly in triglycerides (TG)) were rapidly cleared from the circulation. In the homozygous mink, clearance was much retarded. The ratio of TG to core label in plasma did not decrease and much less [14C]oleic acid appeared in plasma. Still, half of the labeled material disappeared from the circulating blood within 30;-40 min and the calculated total turnover of TG in the hypertriglyceridemic mink was almost as large as in normal mink. The core label was distributed to the same tissues in hypertriglyceridemic mink as in normal mink. Half to two-thirds of the cleared core label was in the liver. The large difference was that in the hypertriglyceridemic mink, TG label (about 40% of the total amount removed) followed the core label to the liver and there was no preferential uptake of TG over core label in adipose or muscle tissue. In normal mink, only small amounts of TG label (<10%) appeared in the liver, while most was in adipose and muscle tissues. Apolipoprotein B-48 dominated in the accumulated TG-rich lipoproteins in blood of hypertriglyceridemic mink, even in fasted animals.

Amino Acid Substitution↗
PubMed1998

Mild oxidation of lipoproteins increases their affinity for surfaces covered by heparan sulfate and lipoprotein lipase.

Lipoprotein lipase (LPL) is present in cells involved in development of atherosclerosis (endothelial cells, smooth muscle cells, and macrophages). A direct involvement of LPL in atherogenesis has been suggested. Previously we used the surface plasmon resonance technique to study the interaction of lipoproteins with surfaces covered by heparan sulfate proteoglycans (HSPG) and LPL [A. Lookene et al. (1997) Biochemistry 36, 5267-5275]. The binding was much increased by the presence of LPL. Here we demonstrate that mild oxidation of low-density-lipoprotein (LDL) and very-low-density lipoprotein (VLDL) in vitro increases their binding to surfaces covered by HSPG and LPL, while extensive oxidation decreases it. Similar results were obtained with a lipid emulsion (Intralipid), indicating that oxidation-induced changes of the lipid part could explain the effects. LPL increased binding and uptake of the mildly oxidized (compared to nonoxidized) LDL by THP-I monocyte-derived macrophages. Our studies indicate that LPL has the highest affinity for mildly oxidized LDL and support its involvement in development of atherosclerosis.

Animals↗
PubMed1998

Clearance of artificial triacylglycerol particles.

In this review we discuss the metabolism of parenteral emulsions in relation to their natural counterpart, the chylomicrons. A major reaction is lipoprotein lipase-mediated hydrolysis of triglycerides at the vascular endothelium in extrahepatic tissues. The lipase is retained at the cell surface by interactions with heparan sulfate proteoglycans but can move along the surface. Lipoproteins and emulsion particles are initially steered to the endothelium by electrostatic forces. These weak interactions are reinforced by recruitment of lipase molecules. Small particles, whether injected as such or formed as remnants of larger particles, are catabolized mainly through receptor-mediated endocytosis in the liver. In contrast, many of the larger particles are removed by other, less well defined, mechanisms.

Animals↗
PubMed1998

A mutation in the lipoprotein lipase gene associated with hyperlipoproteinemia type I in mink: studies on lipid and lipase levels in heterozygotes.

Severe hypertriglyceridemia was previously observed in mink. Affected animals had no detectable lipoprotein lipase activity, but normal amounts of lipoprotein lipase protein in post-heparin plasma. We have now cloned cDNA for lipoprotein lipase from normal mink and identified a single point mutation in the affected animals which most likely explains the deficiency of active lipase. The mutation is located in exon 6 and results in a Pro214Leu substitution. In heterozygote mink the levels of lipoprotein lipase activity and mass in post-heparin plasma were lower than in normal mink, but could not be used to identify carriers of the mutation. In some tissues (heart, muscle, kidney and lung), lipoprotein lipase activity was decreased to about 50%. In adipose tissue there seemed to be a mechanism to compensate for the mutation, resulting in increased mass and approximately the same activity of lipoprotein lipase as in animals not carrying the mutation. Mink had high lipoprotein lipase activity and mass in kidneys, although the levels of mRNA in kidney were many fold lower than in adipose tissue. Mink had very low levels of cholesteryl ester transfer protein activity in plasma. This may contribute to the high levels of HDL in this animal species.

Adipose Tissue↗
PubMed1998

Binding and intracellular trafficking of lipoprotein lipase and triacylglycerol-rich lipoproteins by liver cells.

The cellular mechanisms and pathways by which lipoprotein lipase (LPL) enhances the binding and uptake of lipoproteins remains unknown. Confocal and immunoelectron microscopy demonstrated that primary binding of bovine LPL (bLPL) occurs at the microvilli surface of HepG2 cells and hepatocytes. Internalized bLPL was associated with endocytic vesicles and multivesicular bodies. Quantitative immunofluorescence indicated that the presence of bLPL caused a marked increase in the cell-surface binding of DiI-conjugated triacylglycerol-rich lipoproteins (DiI-TRL). Confocal microscopy showed that when DiI-TRL was incubated with bLPL at 4 degrees C, the distributions of bound LPL and DiI-TRL were totally coincident, and covered the apical surface of both HepG2 cells and hepatocytes. When incubated separately, the time-courses of the internalization of fluorescence associated with DiI-TRL and bLPL were different: DiI-TRL was quickly internalized by both HepG2 cells and hepatocytes, and reached a plateau at 30 min, whereas intracellular LPL increased continuously, but more slowly in the same period. In the presence of bLPL, DiI-TRL was internalized progressively by HepG2 and by cultured hepatocytes for up to 1 h and no saturation was reached. At this time the intensity of labeling of bLPL was lower than of DiI-TRL and a higher number of DiI spots did not colocalize with bLPL immunofluorescence, suggesting that the ligands follow a different pathway after internalization. The data suggest that when lipoprotein lipase (LPL) is associated with the lipoproteins it directs them to specific endocytic pathways. A hypothetical model of the intracellular pathways followed by triacylglycerol-rich lipoproteins and LPL after internalization is proposed.

Animals↗
PubMed1998

Lipoprotein lipase transport in plasma: role of muscle and adipose tissues in regulation of plasma lipoprotein lipase concentrations.

Lipoprotein lipase (LPL) is synthesized in tissues involved in fatty acid metabolism such as muscle and adipose tissue. LPL is also found in the circulation, but is mostly lipolytically inactive. The proportion of active circulating LPL increases after a fatty meal. We investigated the release of active and inactive LPL from adipose tissue and muscle in the fasting and postprandial states. Arteriovenous concentration gradients of LPL across adipose tissue and forearm muscle were measured in male subjects before and after a fat-rich meal (n = 7) and before and during infusion of a triacylglycerol emulsion (Intralipid) (n = 6). Plasma LPL activity rose after the meal and more so during Intralipid infusion. Plasma LPL mass (>95% inactive LPL) increased after the meal but decreased after Intralipid infusion. In the fasting state (n = 13) muscle efflux of LPL activity was 0.263 +/- 0.098 mU/min per 100 ml of muscle tissue whereas there was an influx of LPL activity to adipose tissue of 0.085 +/- 0.100 mU/min per 100 g of adipose tissue (P < 0. 02 muscle vs. adipose tissue). Similarly in the postprandial state only muscle released LPL activity. Both tissues released LPL mass. In the fasting state efflux was 17.8 +/- 8.8 ng/min per 100 ml muscle and 55.2 +/- 21.3 ng/min per 100 g of adipose tissue (P < 0. 05 muscle vs. adipose tissue). Release of LPL, either active or inactive, was not correlated with levels of non-esterified fatty acids or plasma triacylglycerol. In conclusion, there is a substantial release of LPL from adipose tissue and muscle, most of which is inactive. A small proportion of active LPL seems to be redistributed from muscle to adipose tissue.

Adipose Tissue↗
PubMed1997

Interaction of lipoproteins with heparan sulfate proteoglycans and with lipoprotein lipase. Studies by surface plasmon resonance technique.

Interaction of different classes of lipoproteins with heparan sulfate, heparin, and lipoprotein lipase was studied by a surface plasmon resonance based technique on a BIAcore. The proteoglycans were covalently attached to sensor chips as previously described [Lookene, A., Chevreuil, O., Ostergaard, P., & Olivecrona, G. (1996) Biochemistry 35, 12155-12163]. Binding of all lipoproteins, except for beta-VLDL, to endothelial heparan sulfate was low. Binding of chylomicrons (from rat lymph) and of human VLDL was much increased by the presence of lipoprotein lipase. With human LDL, binding was low in the absence of lipase or at low lipase concentrations. For efficient binding, 2-4 lipase dimers per LDL particle were necessary, indicating cooperativity in the interaction. In contrast, HDL did not bind under any conditions. Heparin had higher binding capacity for lipoproteins than heparan sulfate. This was due to a higher number of binding sites on the heparin chains. Binding of LDL, VLDL, and chylomicrons to heparan sulfate-covered surfaces, both in the presence and in the absence of lipoprotein lipase, was characterized by high values for association rate constants (10(4)-10(5) M(-1) s(-1)) and low values for dissociation rate constants (10(-4)-10(-5) M(-1) s(-1)). In some experiments, rabbit beta-VLDL were directly immobilized to the sensor chips. Binding of lipoprotein lipase to these surfaces was characterized by a very high association rate constant (10(6) M(-1) s(-1)). The dissociation of triacylglycerol-rich lipoproteins was more rapid with catalytically active lipase than with active site-inhibited lipase. It was also markedly increased in the presence of free heparin, suggesting fast exchange kinetics at the surface. Based on that, we propose that lipoproteins are relatively mobile at heparan sulfate covered surfaces. Our study emphasizes the important role of lipoprotein lipase, or molecules with similar properties (apolipoprotein E, hepatic lipase), as mediators for binding of lipoproteins to proteoglycans. It also demonstrates the great potential for the use of biosensors for studies of lipoprotein interactions.

Animals↗
PubMed1997

Mutation of tryptophan residues in lipoprotein lipase. Effects on stability, immunoreactivity, and catalytic properties.

Previous studies had pointed to an important function of a putative exposed loop in the C-terminal domain of lipoprotein lipase for activity against emulsified lipid substrates. This loop contains 3 tryptophan residues (Trp390, Trp393, and Trp394). We have expressed and characterized lipase mutants with tryptophan to alanine substitutions at positions 55, 114, 382, 390, 393, and 394 and a double mutant at residues 393 and 394. The substitutions in the N-terminal domain (W55A and W114A) led to poor expression of completely inactive lipase variants. Heparin-Sepharose chromatography showed that mutant W114A eluted at the same salt concentration as inactive wild-type monomers, indicating that this substitution prevented subunit interaction or led to an unstable dimer. In contrast, all mutants in the C-terminal domain were expressed as mixtures of monomers and dimers similarly to the wild-type. The dimers displayed at least some catalytic activity and had the same apparent heparin affinity as the active wild-type dimers. The mutants W390A, W393A, W394A, and W393A/W394A had decreased reactivity with the monoclonal antibody 5D2, indicating that the 5D2 epitope is longer than was reported earlier, or that conformational changes affecting the epitope had occurred. The mutants W390A, W393A, W394A, and W393A/W394A had decreased catalytic activity against a synthetic lipid emulsion of long-chain triacylglycerols (IntralipidR) and in particular against rat lymph chylomicrons. The most pronounced decrease of activity was found for the double mutant W393A/W394A which retained only 6% of the activity of the wild-type lipase, while 70% of the activity against water-soluble tributyrylglycerol was retained. In the case of chylomicrons also the affinity for the substrate particles was lowered, as indicated by severalfold higher apparent Km values. This effect was less prominent with the synthetic lipid emulsion. We conclude that the tryptophan cluster Trp390-Trp393-Trp394 contributes to binding of lipoprotein lipase to lipid/water interfaces. Utilizing different lipid substrates in different physical states, we have demonstrated that the tryptophan residues in the C-terminal domain may have a role also in the productive orientation of the enzyme at the lipid/water interface.

Amino Acid Sequence↗
PubMed1997

Regulation of adipose tissue lipoprotein lipase in young and old rats.

OBJECTIVE: Changes in tissue lipoprotein lipase have been reported in several of the metabolic disorders commonly associated with ageing, like insulin resistance, obesity and impaired hormonal balance. We have investigated the effect of normal ageing on the nutritional regulation of lipoprotein lipase (LPL) in rat tissues. MEASUREMENTS: In the first experiment, LPL activity and immunoreactive mass were measured in epididymal and perirenal adipose tissue, and soleus and heart muscle tissue. In the following experiments we focused on epididymal adipose tissue. RESULTS: In young rats (aged 29 d, 87 +/- 5 g), fasting for 24 h decreased LPL activity in epididymal and perirenal adipose tissue to 31% and 51% of fed control, respectively, while LPL mass increased to 146% and 261%, respectively. Consequently, LPL specific activity (activity/mass ratio) decreased to 20% of control. Other tissues studied did not show any large changes in LPL specific activity with the nutritional state. This suggests that the mechanism responsible for the down-regulation of LPL specific activity is specific for adipose tissue. The down-regulation was gradually blunted with increasing age and was non-existent in the old rats (aged 265 d, 564 +/- 14 g). LPL in soleus muscle from young rats was regulated by another mechanism, and was associated with a large increase in LPL activity and mass during fasting (297% and 458% of fed control). Also, this mechanism did not exist in soleus muscle from old rats. Prolonging the fasting period of the old rats to 96 h did not induce the changes in adipose tissue or soleus muscle LPL seen in the young rats. CONCLUSION: The results indicate that the nutritional regulation of LPL in adipose tissue and soleus muscle changes during normal ageing.

Adipose Tissue↗
PubMed1997

Lipid binding of apolipoprotein CII is required for stimulation of lipoprotein lipase activity against apolipoprotein CII-deficient chylomicrons.

Human apolipoprotein CII (apo CII) consists of 79 amino acid residues. The amino-terminal two thirds of the molecule binds to lipid through the formation of amphipathic helixes, while the carboxy-terminal third is engaged in activation of lipoprotein lipase (LPL). On the basis of studies in model systems, it was previously concluded that fragments of apo CII spanning residues 51-79 were sufficient for activation, although they do not bind to lipid. In the present study, we used chylomicrons from an apo CII-deficient patient to reinvestigate this possibility, with a physiologically relevant substrate. Human LPL expressed very low activity against these chylomicrons. Addition of apo CII caused an immediate > 100-fold increase in lipase activity. The apo CII fragment 50-79 caused very little stimulation, though with some synthetic lipid substrates, this fragment was fully effective. LPL bound to the chylomicrons even in the absence of apo CII but apparently in a nonproductive manner. In accord with this finding, the main effect of apo CII was on the VMAX for the reaction, with little or no change in the apparent K(M). We conclude that the lipid-binding part of apo CII is needed for activity of LPL against chylomicrons. This idea is in accord with previous studies with lipid monolayers, which showed that the lipid-binding part is necessary for activation of the enzyme at high surface pressures.

Apolipoprotein C-II↗
PubMed1997

Lipoprotein lipase deficiency with pancreatitis in mink: biochemical characterization and pathology.

A severe hyperlipemia in mink, with a pattern that suggested recessive inheritance, was observed at a farm in Norway. On a normal mink diet, affected animals had grossly elevated levels of plasma triglycerides which decreased towards normal on a low-fat diet. Normal minks had the main part of their plasma cholesterol in the HDL fraction. Affected minks, although severely hypertriglyceridaemic, had almost normal levels of both LDL and HDL. Affected minks frequently had lipogranulomas in the mesentery and the pancreas. The lipogranulomatous tissue contained spaces filled with an amorphous, sudanophilic substance with many foamy macrophages in the fibrous tissue between the lesions. Separation of postheparin plasma on heparin-agarose revealed that the affected minks had no detectable lipoprotein lipase activity but normal activity of hepatic lipase. Both normal and affected minks had inactive lipoprotein lipase protein in pre- and post-heparin plasma. This protein, which eluted before the active lipase from heparin-agarose, probably corresponds to lipase monomers. The presence of lipoprotein lipase mass in the affected minks, but no activity, indicates that there might be a point mutation in the lipase gene. The minks provide a new animal model for studies on pancreatitis induced by hypertriglyceridemia and on lipoprotein metabolism in the lipoprotein lipase-deficient state and show features similar to those found in human hyperlipoproteinemia type I.

Animals↗