Postheparin Plasma Lipoprotein Lipase Activity Research Articles

Corn Oil, Palm Oil and Butterfat Fractions Affect Postprandial Lipemia and Lipoprotein Lipase in Meal-Fed Rats

Our objective was to investigate the time course of postprandial lipemia and lipolytic activity in male Sprague-Dawley rats trained to eat meals containing butterfat fractions, palm oil or corn oil. Baseline and postprandial blood samples were obtained via a carotid cannula in rats fed the experimental diets for 4 wk. Rats fed saturated fats compared with corn oil showed a significantly greater peak increase in postprandial triacylglycerol concentrations. Corn oil ingestion resulted in significantly lower concentrations of cholesterol and triacylglycerol in plasma and significantly less triacylglycerol accumulation (millimoles per liter per 24 h) compared with ingestion of saturated fats. Postheparin plasma lipoprotein lipase activity and plasma insulin concentration were generally greater with ingestion of corn oil compared with palm oil or butterfat. Palm oil ingestion resulted in a biphasic plasma triacylglycerol response curve and greater postheparin plasma lipoprotein lipase activity compared with butterfat ingestion, suggesting differential effects of saturated fats on postprandial lipemia. Our results indicate that greater postprandial lipemia with ingestion of saturated fats compared with corn oil may be due in part to slower plasma triacylglycerol clearance.

Correlation between chylomicronemia diagnosis scores and post-heparin lipoprotein lipase activity

IntroductionSustained chylomicronemia is a defect in post-prandial triglyceride management characterized by severe hypertriglyceridemia (triglyceride > 10 mmol/L) due to functional or genetic defects in lipoprotein lipase (LPL)-mediated triglyceride-rich lipoprotein lipolysis. Familial chylomicronemia syndrome (FCS) is a rare mendelian form of chylomicronemia caused by loss-of–function variants in LPL or LPL-related genes. Most individuals with chylomicronemia however present multifactorial chylomicronemia (MCS), in which LPL bio-availability and activity are variable. FCS and MCS differ in terms of clinical characteristics and risk of disease, and diagnosis scoring systems have been proposed to accurately distinguish FCS from MCS. ObjectiveThe aim of this study was to assess the strength of the relationship between plasma post-heparin LPL activity and two published chylomicronemia diagnosis scoring systems. Design and MethodsPost-heparin plasma LPL activity was measured using colorimetric assays in a sample of 29 subjects with sustained chylomicronemia (20 FCS and 9 MCS).Chylomicronemia diagnosis scores were obtained for all subjects using the scoring system A (model A), which integrates apolipoprotein B and free glycerol, a surrogate marker of triglyceride hydrolysis, and the scoring system B (model B). Correlation analyses were conducted to estimate the linear relationship between LPL activity and the two diagnosis scoring systems. ResultsThere was a significant (p < 0.001) difference in post-heparin LPL activity between FCS and MCS. Both scoring systems significantly correlated with post-heparin LPL activity (model A: rs = −0.64, p < 0.001; model B: rs = −0.54, p = 0.002). ConclusionsThese result suggest that chylomicronemia diagnosis scoring systems correlate with LPL activity and adequately contribute to distinguish FCS from MCS.

Role of lipoprotein lipase activity measurement in the diagnosis of familial chylomicronemia syndrome

Activity assays for lipoprotein lipase (LPL) are not standardised for use in clinical settings. This study sought to define and validate a cut-off points based on a ROC curve for the diagnosis of patients with familial chylomicronemia syndrome (FCS). We also evaluated the role of LPL activity in a comprehensive FCS diagnostic workflow. A derivation cohort (including an FCS group (n=9), a multifactorial chylomicronemia syndrome (MCS) group (n=11)), and an external validation cohort (including an FCS group (n=5), a MCS group (n=23) and a normo-triglyceridemic (NTG) group (n=14)), were studied. FCS patients were previously diagnosed by the presence of biallelic pathogenic genetic variants in the LPL and GPIHBP1 genes. LPL activity was also measured. Clinical and anthropometric data were recorded, and serum lipids and lipoproteins were measured. Sensitivity, specificity and cut-offs for LPL activity were obtained from a ROC curve and externally validated. All post-heparin plasma LPL activity in the FCS patients were below 25.1 mU/mL, that was cut-off with best performance. There was no overlap in the LPL activity distributions between the FCS and MCS groups, conversely to the FCS and NTG groups. We conclude that, in addition to genetic testing, LPL activity in subjects with severe hypertriglyceridemia is a reliable criterium in the diagnosis of FCS when using a cut-off of 25.1 mU/mL (25% of the mean LPL activity in the validation MCS group). We do not recommend the NTG patient based cut-off values due to low sensitivity.

Using Synthetic ApoC-II Peptides and nAngptl4 Fragments to Measure Lipoprotein Lipase Activity in Radiometric and Fluorescent Assays.

Lipoprotein lipase (LPL) plays a crucial role in preventing dyslipidemia by hydrolyzing triglycerides (TGs) in packaged lipoproteins. Since hypertriglyceridemia (HTG) is a major risk factor for cardiovascular disease (CVD), the leading cause of death worldwide, methods that accurately quantify the hydrolytic activity of LPL in clinical and pre-clinical samples are much needed. To date, the methods used to determine LPL activity vary considerably in their approach, in the LPL substrates used, and in the source of LPL activators and inhibitors used to quantify LPL-specific activity, rather than other lipases, e.g., hepatic lipase (HL) or endothelial lipase (EL) activity. Here, we describe methods recently optimized in our laboratory, using a synthetic ApoC-II peptide to activate LPL, and an n-terminal Angiopoietin-Like 4 fragment (nAngptl4) to inhibit LPL, presenting a cost-effective and reproducible method to measure LPL activity in human post-heparin plasma (PHP) and in LPL-enriched heparin released (HR) fractions from LPL secreting cells. We also describe a modified version of the triolein-based assay using human serum as a source of endogenous activators and inhibitors and to determine the relative abundance of circulating factors that regulate LPL activity. Finally, we describe how an ApoC-II peptide and nAngptl4 can be applied to high-throughput measurements of LPL activity using the EnzChek™ fluorescent TG analog substrate with PHP, bovine LPL, and HR LPL enriched fractions. In summary, this manuscript assesses the current methods of measuring LPL activity and makes new recommendations for measuring LPL-mediated hydrolysis in pre-clinical and clinical samples.

The zinc finger and BTB domain containing protein ZBTB20 regulates plasma triglyceride metabolism by repressing lipoprotein lipase gene transcription in hepatocytes.

Lipoprotein lipase (LPL) is responsible for the lipolytic processing of triglyceride-rich lipoproteins, the deficiency of which causes severe hypertriglyceridemia. Liver LPL expression is high in suckling rodents but relatively low at adulthood. However, the regulatory mechanism and functional significance of liver LPL expression are incompletely understood. We have established the zinc finger protein ZBTB20 as a critical factor for hepatic lipogenesis. Here, we evaluated the role of ZBTB20 in regulating liver Lpl gene transcription and plasma triglyceride metabolism. Hepatocyte-specific inactivation of ZBTB20 in mice led to a remarkable increase in LPL expression at the mRNA and protein levels in adult liver, in which LPL protein was mainly localized onto sinusoidal epithelial cells and Kupffer cells. As a result, the LPL activity in postheparin plasma was substantially increased, and postprandial plasma triglyceride clearance was significantly enhanced, whereas plasma triglyceride levels were decreased. The dysregulated liver LPL expression and low plasma triglyceride levels in ZBTB20-deficient mice were normalized by inactivating hepatic LPL expression. ZBTB20 deficiency protected the mice against high-fat diet-induced hyperlipidemia without causing excessive triglyceride accumulation in the liver. Chromatin immunoprecipitation and gel-shift assay studies revealed that ZBTB20 binds to the LPL promoter in the liver. A luciferase reporter assay revealed that ZBTB20 inhibits the transcriptional activity of LPL promoter. The regulation of LPL expression by ZBTB20 is liver-specific under physiological conditions. Liver ZBTB20 serves as a key regulator of LPL expression and plasma triglyceride metabolism and could be a therapeutic target for hypertriglyceridemia.

Molecular basis of the familial chylomicronemia syndrome in patients from the National Dyslipidemia Registry of the Spanish Atherosclerosis Society

Familial chylomicronemia syndrome (FCS) is an extremely rare lipoprotein disorder caused by mutations in at least 5 genes of the lipoprotein lipase (LPL) complex. This work shows the molecular analysis of patients diagnosed with FCS, who attended the Spanish Arteriosclerosis Society lipid units and were included in the National Dyslipidemia Registry. Among the 238 patients registered with severe hypertriglyceridemia (fasting triglycerides>1000mg/dL), 26 were diagnosed with FCS as they had confirmed postheparin plasma LPL activity deficiency and/or homozygosity for loss-of-function mutations in LPL, GPIHBP1, APOC2, LMF1, or Apolipoprotein A5 (APOA5). Among the 26 FCS cases, 23 had mutations in the homozygous state: 19 in LPL and 4 in the GPIHBP1 gene. The molecular analysis revealed 3 novel mutations: 2 in LPL, in 2 unrelated patients (c.312delA; p.Asp105Thrfs*66 and c.629A>G; p.His210Arg), and 1 in GPHIBP1 in a third patient (c.502delC; p.Leu168Serfs*83). These 3 patients had confirmed lack of LPL activity. Three additional patients with confirmed LPL activity deficiency were heterozygous carriers of mutations in the genes analyzed. Among these, we found 2 novel mutations in APOA5 (c.50-1G>A and c.326_327insC; p.Tyr110Leufs*158). We have identified 5 novel pathogenic mutations: 2 in LPL, 1 in GPIHBP1, and 2 in the APOA5 gene. The genetic defaults accounting for the LPL activity deficiency of 23 of them have been clearly identified and 3 patients, who harbored mutations in heterozygosity, were diagnosed based on LPL activity deficiency, which raises the question of the involvement of new genes in the manifestation of FCS.

Inulin Improves Postprandial Hypertriglyceridemia by Modulating Gene Expression in the Small Intestine.

Postprandial hyperlipidemia is an important risk factor for cardiovascular diseases in the context of obesity. Inulin is a non-digestible carbohydrate, known for its beneficial properties in metabolic disorders. We investigated the impact of inulin on postprandial hypertriglyceridemia and on lipid metabolism in a mouse model of diet-induced obesity. Mice received a control or a western diet for 4 weeks and were further supplemented or not with inulin for 2 weeks (0.2 g/day per mouse). We performed a lipid tolerance test, measured mRNA expression of genes involved in postprandial lipid metabolism, assessed post-heparin plasma and muscle lipoprotein lipase activity and measured lipid accumulation in the enterocytes and fecal lipid excretion. Inulin supplementation in western diet-fed mice decreases postprandial serum triglycerides concentration, decreases the mRNA expression levels of Cd36 (fatty acid receptor involved in lipid uptake and sensing) and apolipoprotein C3 (Apoc3, inhibitor of lipoprotein lipase) in the jejunum and increases fecal lipid excretion. In conclusion, inulin improves postprandial hypertriglyceridemia by targeting intestinal lipid metabolism. This work confirms the interest of using inulin supplementation in the management of dyslipidemia linked to obesity and cardiometabolic risk.

Striated muscle gene therapy for the treatment of lipoprotein lipase deficiency.

Excessive circulating triglycerides due to reduction or loss of lipoprotein lipase activity contribute to hypertriglyceridemia and increased risk for pancreatitis. The only gene therapy treatment for lipoprotein lipase deficiency decreases pancreatitis but minimally reduces hypertriglyceridemia. Synthesized in multiple tissues including striated muscle and adipose tissue, lipoprotein lipase is trafficked to blood vessel endothelial cells where it is anchored at the plasma membrane and hydrolyzes triglycerides into free fatty acids. We conditionally knocked out lipoprotein lipase in differentiated striated muscle tissue lowering striated muscle lipoprotein lipase activity causing hypertriglyceridemia. We then crossed lipoprotein lipase striated muscle knockout mice with mice possessing a conditional avian retroviral receptor gene and injected mice with either a human lipoprotein lipase retrovirus or an mCherry control retrovirus. Post-heparin plasma lipoprotein lipase activity increased for three weeks following human lipoprotein lipase retroviral infection compared to mCherry infected mice. Human lipoprotein lipase infected mice had significantly lower blood triglycerides compared to mCherry controls and were comparable to wild-type blood triglyceride levels. Thus, targeted delivery of human lipoprotein lipase into striated muscle tissue identifies a potential therapeutic target for lipoprotein lipase deficiency.

Streptozotocin-Treated High Fat Fed Mice: A New Type 2 Diabetes Model Used to Study Canagliflozin-Induced Alterations in Lipids and Lipoproteins.

The pharmacological effects of type 2 diabetes (T2DM) medications on lipoprotein metabolism are difficult to assess in preclinical models because those created failure to replicate the human condition in which insulin deficiency is superimposed on obesity-related insulin resistance. To create a better model, we fed mice with high fat (HF) diet and treated the animals with low dose streptozotocin (STZ) to mimic T2DM. We used this model to evaluate the effects of canagliflozin (CANA), a drug that reduces plasma glucose by inhibiting the sodium-glucose transporter 2 (SGLT2), which mediates ~90% of renal glucose reabsorption] on lipid and lipoprotein metabolism. After 6 weeks of CANA (30 mg/kg/day) treatment, the increase in total plasma cholesterol in HF-STZ diabetic mice was reversed, but plasma triglycerides were not affected. Lipoprotein fractionation and cholesterol distribution analysis showed that CANA kept HDL-Cholesterol, LDL-Cholesterol, and IDL-Cholesterol levels steady while these lipoprotein species were increased in placebo- and insulin-treated control groups. CANA treatment of HF-STZ mice reduced post-heparin plasma lipoprotein lipase (LPL) activity at 2 (-40%) and 5 (-30%) weeks compared to placebo. Tissue-specific LPL activity following CANA treatment showed similar reduction. In summary, CANA prevented the total cholesterol increase in HF-STZ mice without effects on plasma lipids or lipoproteins, but did decrease LPL, implying a potential role of LPL-dependent lipoprotein metabolism in CANA action. These effects did not recapitulate the effect of SGLT2 inhibitors on lipids and lipoproteins in human, suggesting that a better murine T2DM model (such as the ApoB100 humanized CETP-overexpressing mouse) is needed next.

Novel mutations in the GPIHBP1 gene identified in 2 patients with recurrent acute pancreatitis

Glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1 (GPIHBP1) has been demonstrated to be essential for the invivo function of lipoprotein lipase (LPL), the major triglyceride (TG)-hydrolyzing enzyme involved in the intravascular lipolysis of TG-rich lipoproteins. Recently, loss-of-function mutations of GPIHBP1 have been reported as the cause of type I hyperlipoproteinemia in several patients. Two unrelated patients were referred to our Lipid Units because of a severe hypertriglyceridemia and recurrent pancreatitis. We measured LPL activity in postheparin plasma and serum ApoCII and sequenced LPL, APOC2, and GPIHBP1. The 2 patients exhibited very low LPL activity not associated with mutations in LPL gene or with ApoCII deficiency. The sequence of GPIHBP1 revealed 2 novel point mutations. One patient (proband 1) was found to be homozygous for a C>A transversion in exon 3 resulting in the conversion of threonine to lysine at position 80 (p.Thr80Lys). The other patient (proband 2) was found to be homozygous for a G>T transversion in the third base of the ATG translation initiation codon in exon 1, resulting in the conversion of methionine to isoleucine (p.Met1Ile). In conclusion, we have identified 2 novel GPIHBP1 missense mutations in 2 unrelated patients as the cause of their severe hypertriglyceridemia.

Acute alcohol consumption downregulates lipoprotein lipase activity in vivo

ObjectiveAcute alcohol consumption can induce hypertriglyceridemia. Such an effect could be explained in part by the influence of alcohol on lipoprotein lipase (LPL) – the key enzyme responsible for triglyceride hydrolysis in circulation. Therefore, we have studied the effects of acute moderate alcohol consumption on LPL activity and on the concentrations of angiopoietin-like proteins 3 and 4 (ANGPTL3 and ANGPTL4), which are known to inhibit LPL. MethodsTwo experiments were carried out in 8 healthy volunteers. They received 25g of alcohol (vodka) in one experiment and water in the other (control). The in vivo function of LPL was estimated using intravenous fat tolerance tests (IVFTT) carried out before, 2 and 4hours after alcohol administration. At the end of each experiment, LPL activity and mass were measured in post-heparin plasma (PHP). The concentrations of ANGPTL3 and ANGPTL4 in blood were measured before alcohol consumption and at the end of the experiments. ResultsLPL activity, as estimated using the IVFTT, was reduced by 25% and 24% two and four hours after the administration of alcohol, respectively, and was not affected in the control experiment. At the end of the experiment, LPL activity in PHP was 23% lower after alcohol consumption than in the controls. The concentrations of ANGPTL3 and ANGPTL4 had dropped to 67% and 86% of baseline values, respectively, at 280min after alcohol consumption. These levels were not affected in the control experiment. The levels of ANGPTL4 but not those of ANGPTL3 were increased in PHP compared to both baseline values and values at 280min. ConclusionThe capacity for triglyceride clearance seemed to be acutely reduced by alcohol consumption and the effect persisted for several hours. The levels of LPL activity in PHP were reduced to a similar extent. This reduction in LPL activity could not be explained by the changes in the levels of ANGPTL3 or ANGPTL4, which both decreased.

Homozygosity for a partial deletion of apoprotein A-V signal peptide results in intracellular missorting of the protein and chylomicronemia in a breast-fed infant

Deficiency of apoprotein A-V (apoA-V) can cause hypertriglyceridemia. In an 11 months old boy presenting with a severe hypertriglyceridemia, a formerly unknown 24 nucleotide deletion in exon 2 of the APOA5 gene was detected. The homozygous mutation results in an eight amino acid loss in the signal peptide sequence (c.16_39del; p.Ala6_Ala13del). Screening of control persons proved that this deletion is a rare mutation. Hypertriglyceridemia in the patient was only found at the time when he was breast fed, while after weaning, triglyceride levels were close to normal. Under both dietary conditions, apoA-V protein was undetectable in plasma while post-heparin plasma lipoprotein lipase activity was normal.Expression analysis of normal and mutated protein by Western blot and immunofluorescence in apoA-V deficient primary hepatocytes revealed that, due to changes in the signal peptide, mutated apoA-V was intracellularly missorted to lipid droplets and not secreted. Wild type apoA-V, instead, was not targeted to lipid droplets but transported via endosomal compartments to the plasma membrane for secretion.It is concluded that the c.16_39del mutation in the APOA5 gene leads to hepatic missorting and impaired secretion, which consequently results in undetectable apoA-V plasma levels. The absence of apoA-V in plasma leads under conditions of fat-rich diets to severe chylomicronemia, suggestive for a modulatory role of apoA-V for lipoprotein lipase mediated intravascular triglyceride lipolysis.

Hepatic lipase activity is decreased in Japanese patients with type III hyperlipoproteinemia

BackgroundType III hyperlipoproteinemia (HLP), a disorder associated with a high incidence of premature cardiovascular diseases, is characterized by the accumulation of remnant lipoproteins in the plasma. The primary genetic defect in patients with type III HLP is the presence of apolipoprotein E2 (apoE2), an isoform of apoE, and accumulation of remnant lipoproteins in the plasma has been thought to be attributable to the presence of apoE2, which bind poorly to low density lipoprotein receptors, resulting in defective remnant lipoprotein clearance. On the other hand, the activity of hepatic lipase (HL), the enzyme that plays a pivotal role in the removal of remnant lipoproteins, in type III HLP has not been investigated. MethodsWe examined post-heparin plasma lipoprotein lipase (LPL) and HL activities in 7 patients with type III HLP. The activities of HL and LPL in post-heparin plasma were measured separately by an immunochemical method using antiserum specifically directed against HL. ResultsThe post-heparin plasma HL activity was significantly reduced, while the LPL activity was normal. ConclusionsReduced HL activity may account, at least in part, for the accumulation of remnant lipoproteins in the plasma, a characteristic feature of type III HLP.

Effects of NO-1886, a Lipoprotein Lipase Promoting Agent, on Homozygous and Heterozygous Watanabe Heritable Hyperlipidaemic Rabbits

The novel compound NO-1886 ([4-(4-bromo-2-cyano-phenylcarbamoyl)-benzyl]-phosphonic acid diethyl ester, CAS 133208-93-2) is a lipoprotein lipase (LPL) activator, and long term administration of NO-1886 protects against the development of experimental atherosclerosis in rats and rabbits. In the present experiments, the effects of this compound were examined in Watanabe heritable hyperlipidaemic (WHHL) rabbits, an animal model for familial hypercholesterolemia lacking low density lipoprotein (LDL) receptors. NO-1886 increased postheparin plasma LPL activity, resulting in a reduction of plasma triglycerides with concomitant elevation of HDL cholesterol in heterozygous WHHL rabbits. However, the compound did not cause any changes in plasma lipids and postheparin plasma LPL activity in homozygous WHHL rabbits. The different responses suggest that the effects of NO-1886 may be either mediated by LDL receptors, or that persistent exposure to extreme hypercholesterolemia might affect the cellular response to this particular compound in homozygous WHHL rabbits.

The lipid-lowering effect of dietary proanthocyanidins in rats involves both chylomicron-rich and VLDL-rich fractions

Proanthocyanidins have been shown to improve postprandial hypertriacylglycerolaemia. The present study aims to determine the actual contribution of chylomicrons (CM) and VLDL in the hypotriacylglycerolaemic action of grape seed proanthocyanidin extract (GSPE) in the postprandial state and to characterise the mechanisms by which the GSPE treatment reduces TAG-rich lipoproteins in vivo. A plasma lipid tolerance test was performed on rats fasted for 14 h and orally loaded with lard containing either GSPE or not. GSPE (250 mg/kg body weight) markedly blocked the increase in plasma TAG induced by lard, with a statistically significant reduction of 22 % in the area under the curve. The VLDL-rich fraction was the major contributor (72 %) after 1 h, whereas the CM-rich fraction was the major contributor (85 %) after 3 h. At 5 and 7 h after treatment, CM-rich and VLDL-rich fractions showed a similar influence. Plasma post-heparin lipoprotein lipase (LPL) activity and LPL mRNA levels in white adipose tissue and muscle were not affected by GSPE. On the contrary, GSPE treatment significantly repressed (30 %) the secretion of VLDL-TAG. In the liver, GSPE treatment induced different effects on the expression of acyl-coenzyme A synthetase long-chain family member 1, Apoc3 and 3-hydroxy-3-methylglutaryl-coenzyme A reductase at 1 h and Cd36 at 5 h, compared to those induced by lard. Furthermore, GSPE treatment significantly increased the activity of carnitine palmitoyltransferase 1a at 1 h. In conclusion, both CM-rich and VLDL-rich fractions contributed to the hypotriacylglycerolaemic action of GSPE, but their influence depended on time. GSPE induces hypotriacylglycerolaemic actions by repressing lipoprotein secretion and not by increasing LPL activity.

ATP-binding cassette G5/G8 deficiency causes hypertriglyceridemia by affecting multiple metabolic pathways

Mutations in ABCG5 or ABCG8 transporters are responsible for sitosterolemia, an autosomal recessive disease characterized by the accumulation of plant sterols. The aim of this study was to investigate the effects of ABCG5 and ABCG8 deficiency on TG metabolism in mice. Experiments were carried out in wild-type (G5/G8+/+) mice, mice heterozygous for ABCG5 and ABCG8 deficiency (G5/G8+/−) and ABCG5/G8-deficient (G5/G8−/−) mice fed a chow diet. Plasma TG were 2.6 and 4.3-fold higher in fasted G5/G8+/− and G5/G8−/− mice, respectively, than in G5/G8+/+ mice. Postprandial TG were 5-fold higher in G5/G8−/− mice. TG metabolism studies indicate that: first, the fractional catabolic rate was significantly lower in G5/G8+/− (1.3-fold) and G5/G8−/− mice (1.5-fold) compared to G5/G8+/+ and postheparin plasma lipoprotein lipase activities were significantly lower in G5/G8+/− (1.8-fold) and G5/G8−/− mice (5.4-fold) than in G5/G8+/+. Second, liver TG secretion was 1.3-fold higher in G5/G8+/− and G5/G8−/− than in G5/G8+/+ mice and this was associated with an increase in liver LXR, FAS, ACAC and CD36 gene expression. Third, TG intestinal secretion, determined after an oral fat gavage of glycerol tri[9,10(n)-3H] oleate, was 5.8-fold higher in G5/G8−/− than in G5/G8+/+ mice. Also, the HOMA index was 2.6-fold higher in G5/G8−/− than in G5/G8+/+ mice, reflecting a degree of insulin resistance. In conclusion, ABCG5/G8 deficiency in mice fed a chow diet markedly raises TG levels by impairing TG catabolism and by increasing liver and intestinal TG secretion.

Activation of lipoprotein lipase increases serum high density lipoprotein 2 cholesterol and enlarges high density lipoprotein 2 particles in rats

It is known that postheparin plasma lipoprotein lipase (LPL) activity correlates with serum high density lipoprotein cholesterol (HDL-C) levels in humans and animals. Furthermore, LPL has been reported to cause enlargement of HDL particle size in vitro. However, these effects have not yet been experimentally proven. The aim of this study was to determine whether LPL has a role in increase in HDL-C and enlargement of HDL particle by activating the LPL function with NO-1886, the LPL promoting agent. NO-1886 administration increased postheparin plasma LPL activity without influencing hepatic triglyceride lipase activity. NO-1886 increased serum HDL2-cholesterol (HDL2-C) concentration and enlarged HDL2 particle size, but did not increase serum HDL3-cholesterol concentration or enlarge HDL3 particle size. Also, serum HDL2-C concentrations were positively correlated with HDL2 particle size (r=0.910). Our study demonstrates that the LPL activation induced with NO-1886 may cause production of HDL2-C by catabolism of triglyceride-rich lipoproteins and enlarges HDL2 particle size in rats.

Determination of lipoprotein lipase activity using a novel fluorescent lipase assay

A novel, real-time, homogeneous fluorogenic lipoprotein lipase (LPL) assay was developed using a commercially available substrate, the EnzChek lipase substrate, which is solubilized in Zwittergent. The triglyceride analog substrate does not fluoresce, owing to apposition of fluorescent and fluorescent quenching groups at the sn-1 and sn-2 positions, respectively, fluorescence becoming unquenched upon release of the sn-1 BODIPY FA derivative following hydrolysis. Increase in fluorescence intensity at 37°C was proportional to LPL concentration. The assay was more sensitive than a similar assay using 1,2-O-dilauryl-rac-glycero-3-glutaric acid-(6-methylresorufin ester) and was validated in biological samples, including determination of LPL-specific activity in postheparin mouse plasma. The simplicity and reproducibility of the assay make it ideal for in vitro, high-throughput screening for inhibitors and activators of LPL, thus expediting discovery of drugs of potential clinical value.

Hypertriglyceridemia and delayed clearance of fat load in transgenic rabbits expressing human apolipoprotein CIII

Apolipoprotein CIII (apoCIII) has been implicated in hypertriglyceridemia and plasma apoCIII independently predicts risk for coronary heart disease. While hypertriglyceridemia in patients has been demonstrated to correlate with elevated plasma apoCIII levels and reduced lipoprotein lipase (LPL) activity, apoCIII transgenic mice show elevated LPL activity. In this study, we generated transgenic (Tg) rabbits expressing human apoCIII gene exclusively in liver and investigated the effect of apoCIII overexpression on lipid metabolism of rabbits. In comparison with non-Tg littermates, Tg rabbits had 3- and 3.2-fold increases in fed and fasted plasma triglycerides, respectively. In contrast, no significant differences were observed in plasma total cholesterol and high density lipoprotein cholesterol levels between Tg and non-Tg rabbits. Analysis of lipoprotein fractions revealed that elevated plasma triglyceride levels in Tg rabbits were mainly attributed to an increase in very low density lipoprotein/chylomicron-sized particles. Furthermore, Tg rabbits showed markedly delayed clearance of plasma triglycerides accompanied with significantly reduced LPL activity in post-heparin plasma compared to that in non-Tg controls. In conclusion, these results indicate apoCIII transgenic rabbits develop hypertriglyceridemia with similar mechanism in hypertriglyceridemic patients via delayed clearance of plasma triglycerides, and could be used as a valuable tool for the study of human hyperlipidemia in relation with atherosclerotic disorders.

P285 FIRST DEMONSTRATION OF THE LOCALIZATION OF HEPATIC apoB100- AND INTESTINAL apoB48-CONTAINING LIPOPROTEIN PARTICLES IN HUMAN ATHEROSCLEROTIC PLAQUES USING DUAL-STAINING IMMUNOFLUORESCENCE

Mutations in ABCG5 or ABCG8 transporters are responsible for sitosterolemia, an autosomal recessive disease characterized by the accumulation of plant sterols. The aim of this study was to investigate the effects of ABCG5 and ABCG8 deficiency on TG metabolism in mice. Experiments were carried out in wild-type (G5/G8+/+) mice, mice heterozygous for ABCG5 and ABCG8 deficiency (G5/G8+/−) and ABCG5/G8-deficient (G5/G8−/−) mice fed a chow diet. Plasma TG were 2.6 and 4.3-fold higher in fasted G5/G8+/− and G5/G8−/− mice, respectively, than in G5/G8+/+ mice. Postprandial TG were 5-fold higher in G5/G8−/− mice. TG metabolism studies indicate that: first, the fractional catabolic rate was significantly lower in G5/G8+/− (1.3-fold) and G5/G8−/− mice (1.5-fold) compared to G5/G8+/+ and postheparin plasma lipoprotein lipase activities were significantly lower in G5/G8+/− (1.8-fold) and G5/G8−/− mice (5.4-fold) than in G5/G8+/+. Second, liver TG secretion was 1.3-fold higher in G5/G8+/− and G5/G8−/− than in G5/G8+/+ mice and this was associated with an increase in liver LXR, FAS, ACAC and CD36 gene expression. Third, TG intestinal secretion, determined after an oral fat gavage of glycerol tri[9,10(n)-3H] oleate, was 5.8-fold higher in G5/G8−/− than in G5/G8+/+ mice. Also, the HOMA index was 2.6-fold higher in G5/G8−/− than in G5/G8+/+ mice, reflecting a degree of insulin resistance. In conclusion, ABCG5/G8 deficiency in mice fed a chow diet markedly raises TG levels by impairing TG catabolism and by increasing liver and intestinal TG secretion.