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Diet plays an important role in the development of chronic metabolic diseases (diabetes, obesity, cardiovascular disease) and as dietary fat consumption has increased, so has the incidence of these disorders. Metabolic syndrome, a clustering of risk factors that includes central obesity, increased plasma triacylglycerol (TG), elevated fasting glucose and glucose intolerance is perhaps the most notorious and aggressive. Animal and human studies indicate that bioactive fatty acids can influence cellular energy metabolism. Using susceptible rodent models (apoE-/- and LDLr-/- mice and Syrian Golden hamsters) this project investigated whether supplementation of a western type diet (WD) with bioactive fatty acids could improve hepatic lipid metabolism, plasma lipoprotein profiles or liver markers of lipogenesis. In mice, dietary supplementation with t-10, c-12 conjugated linoleic acid (CLA) decreased the weight gain induced by high fat diet compared with WD (p<0.01) and was accompanied by hyperinsulinemia (p<0.05) in the ApoE-/- and hypoadiponectinemia (p<0.01) in both mice strains. Although t-10, c-12 CLA supplementation increased plasma lipids and was associated with profound liver steatosis there was a reduction in atherosclerotic lesions in both mouse models (p<0.05). Analysis of mRNA and protein levels in the liver suggested that the differences in liver and plasma lipids may reflect inappropriate lipogenic response to t-10,c-12 CLA. In the high fat and fructose-fed hamster, the modulating role of fish fatty acids was investigated. The addition of DHA increased weight gain and adiposity compared to EPA and c-9, t-11 CLA supplementation. However, glucose tolerance was improved after 6 weeks of DHA supplementation (p? 0.01). Using [35S]methionine radiolabelling, DHA supplementation decreased apolipoprotein B100 synthesis and secretion. Newly synthesized cellular and secreted TG, as measured by [3H]glycerol incorporation, were also decreased with DHA supplementation. Although the effects of EPA were similar to those with DHA, the magnitude was generally lower. These results suggest that supplementation with fish fatty acids can improve several of the risk factors of the metabolic syndrome. Taken together, these observations indicate that some, but not all, bioactive fatty acids may be useful supplements for mediating cardiovascular risk factors.

The effect of acute exercise on blood lipids and lipoproteins has been examined and their response is believed to reflect the changes that occur as a result of chronic exercise training. The cross sectional differences between trained and untrained individuals and the results of longitudinal investigations suggest LDL particle size and distribution may be altered after acute exercise. Moreover, the response of the lipid and lipoprotein profile of Black men may be different than that of White men, owing to initial concentration differences or a genetic predisposition. PURPOSE: To examine the effects of expending 300 and 600 calories through treadmill walking at a moderate intensity (65% VO2max) on blood lipids and lipoproteins as compared to a non-exercise, control trial over the course of 72 hours. METHODS: Ten White and ten Black men participated in this study after being screened for strict inclusion criteria (including: parents/grandparents either Black or White and from the United States; sedentary, but otherwise healthy; VO2max 30-45ml⋅kg⋅min-1; and body fatness ¡Ü25%). Analysis of variance with repeated measures design was used (Group x Trial x Time). Blood was collected at Baseline (0hr), 12hr, 24hr, 48hr and 72hr for determination of blood lipid concentrations, LDL particle size, LDL distribution and CETPa. RESULTS: Black men of this investigation had higher HDL-C (49.4 vs 41.8 mg⋅dL-1), HDL3-C (35.1 vs 30.8 mg⋅dL-1) and CETPa (82.1 vs 52.8 pmol⋅ml-1⋅3hr-1) and lower TC (148.9 vs 177.1 mg⋅dL- 1), LDL-C (83.4 vs 116.3 mg⋅dL-1) and TC to HDL ratios (3.06 vs 4.46) than the White men (p<0.05) over the control period. There were no group differences detected in LDL particle size (White 25.63nm vs Black 25.79nm), LDL distribution (Zone 1, White 58.19% vs Black 64.71%; Zone 2, White 20.16% vs Black 16.31%; Zone 3, White 21.65% vs Black 18.98%), HDL2-C (White 11.5 vs Black 14.2 mg⋅dL-1), or TG (White 95.0 vs Black 79.4 mg⋅dL-1). Black and White men responded to acute exercise in similar fashion for these variables and therefore our statistical model collapsed the data into a single group by trial and over time. Exercise did not statistically alter TC, LDL-C, LDL particle size, LDL distribution, HDL2-C or CETPa in the 72 hours following each exercise bout. However, HDL-C was increased 6.2%, 10.1% and 5.8%, HDL3-C was increased 9.8%, 13.8% and 9.8%, while TG were reduced 24.8%, 27.3% and 22.4% at 12hr, 24hr and 48hr, respectively from the Base value of the 600 Kcal Trial. Additionally, TG were reduced 18.0% at 12hr from the baseline value of the 300 Kcal Trial. CONCLUSION: These results are the first to indicate that acute treadmill walking at moderate intensity of sufficient caloric expenditure modified HDL-C, HDL3-C and TG in Black men. No differences were determined in the manner in which Black and White American men statistically increased HDL-C and HDL3-C and reduced TG after acute exercise. In addition, there was no impact of acute exercise on TC, LDL-C, HDL2-C, CETPa, LDL particle size or LDL distribution in either group. However, these data suggest that Black/White differences exist in CETPa, where Black men have greater activity of this enzyme / Dissertation / PhD

Mortality from cardiovascular disease in patients on chronic hemodialysis (HD) is 10 to 20 times greater than in the general population. One major risk factor is renal dyslipidemia, characterised by an impaired catabolism of triglyceride (TG)-rich lipoproteins with accumulation of atherogenic remnant particles. A contributing factor may be derangement of the lipoprotein lipase (LPL) system, the major lipase in the catabolism of TG-rich lipoproteins. The functional pool of LPL is located at vascular surfaces, and is released by heparin into the circulating blood and extracted and degraded by the liver. Unfractionated heparin (UFH) is commonly used during dialysis to avoid clotting in the extracorporeal devices, but is increasingly replaced by various low molecular weight heparin (LMWH) preparations. Plasma LPL activity is usually lower after injection of LMWH which is therefore said to release less LPL and cause less disturbance of lipoprotein metabolism than UFH. However, animal studies have revealed that LMWH is as efficient as UFH in releasing LPL but is less efficient in retarding hepatic uptake. The aim of this study was to explore the effects of UFH and a LMWH (dalteparin) on LPL activity and TG concentrations in HD-patients compared with healthy controls, matched for age and gender. A disturbed LPL system might contribute to an impaired lipoprotein metabolism, and hence, an aggravated cardiovascular condition. An 8-hour primed infusion of UFH to controls gave rise to an initial peak of LPL activity within 30 minutes. The activity then dropped by almost 80% over the next two hours and levelled off to a plateau that corresponded to 15% of the peak level. When UFH was infused to HD-patients the curve for LPL activity resembled that for controls, but was reduced by 50% during the peak, while the plateau activities were comparable. The interpretation was that the functional pool, represented by the initial peak, was impaired in HD-patients, while the production of lipase molecules, reflected by the plateau, was only marginally reduced. During the peak of LPL activity TG decreased in both groups, but less in HD-patients, as was expected from the lower circulating lipase activity. During the plateau phase with low lipase activity, TG increased towards and beyond baseline values. When dalteparin was infused, the same pattern of plasma LPL activity was observed, although remarkably reduced. In controls the peak was only 30% and the subsequent plateau 40% compared with the activities during the UFH infusion. A bolus of UFH given when the LPL activity had levelled off to a plateau brought out about the same amount of activity, regardless of whether dalteparin or UFH had been infused. The conclusion was that both heparin preparations had reduced endothelial LPL to a similar extent, but that dalteparin less efficiently retarded the hepatic uptake of the enzyme. As a consequence to this, TG tended to reach higher levels after the dalteparin infusion. The LPL activities were further reduced in HD-patients during infusion with dalteparin, the peak was only 27% and the plateau 35% compared with the activities when UFH was infused. There was no decrease in TG, but rather a continuous increase, suggesting a profound depletion of functional LPL. In another study in HD-patients, two anticoagulation regimes based on present clinical practice were compared, and the doses were adjusted to the respective manufacturers recommendation. UFH was administered as a primed infusion, whereas dalteparin was given only as a single bolus pre-dialysis, not followed by an infusion. The results were in line with those in the experimental studies and indicate that also in the clinical setting LMWH interferes with the LPL system as least as much as an infusion of UFH does, and temporarily impairs lipolysis of TG. This interference might, in consequence, contribute to an aggravated cardiovascular condition in HD-patients.

Emerging evidence has shown that an abnormal postprandial accumulation of dietary tat IS atherogenic. The aim of this study is to measure triacylglycerol (TAG) kinetics in endogenous and exogenous lipoproteins in both fed and fasted states using stable isotopes.

In recent years, studies have revealed a central role for the intestine in regulation of lipid homeostasis and development of insulin resistance and type-2 diabetes. The function of intestinal Scavenger Receptor Class-B type-I remains unknown, however it is believed to play a role in dietary lipid uptake. Recently, our laboratory demonstrated a correlation between intestinal SR-BI expression and chylomicron secretion. We hypothesized that intestinal SR-BI is involved in chylomicron secretion and contributes to chylomicron oversecretion in insulin resistance. I first characterized chylomicron production in healthy and insulin resistant Syrian golden hamsters. Inhibition of SR-BI resulted in reduced postprandial chylomicron accumulation in plasma, and resistance to diet-induced hyperlipidemia and weight-gain. Lower postprandial triglyceride levels were also observed in SR-BI-/- mice. In summary, these data demonstrate a key role for intestinal SR-BI in chylomicron secretion and control of lipid homeostasis, implicating intestinal SR-BI in chylomicron overproduction in insulin resistant states.

In recent years, studies have revealed a central role for the intestine in regulation of lipid homeostasis and development of insulin resistance and type-2 diabetes. The function of intestinal Scavenger Receptor Class-B type-I remains unknown, however it is believed to play a role in dietary lipid uptake. Recently, our laboratory demonstrated a correlation between intestinal SR-BI expression and chylomicron secretion. We hypothesized that intestinal SR-BI is involved in chylomicron secretion and contributes to chylomicron oversecretion in insulin resistance. I first characterized chylomicron production in healthy and insulin resistant Syrian golden hamsters. Inhibition of SR-BI resulted in reduced postprandial chylomicron accumulation in plasma, and resistance to diet-induced hyperlipidemia and weight-gain. Lower postprandial triglyceride levels were also observed in SR-BI-/- mice. In summary, these data demonstrate a key role for intestinal SR-BI in chylomicron secretion and control of lipid homeostasis, implicating intestinal SR-BI in chylomicron overproduction in insulin resistant states.

Individuals with high levels of plasma triglycerides are at high risk to develop cardiovascular disease (CVD), currently one of the major causes of death worldwide. Recent epidemiological studies show that loss-of-function mutations in the APOC3 gene lower plasma triglyceride levels and reduce the incidence of coronary artery disease. The APOC3 gene encodes for apolipoprotein (APO) C3, known as an inhibitor of lipoprotein lipase (LPL) activity. Similarly, a common gain-of-function mutation in the LPL gene is associated with reduced risk for CVD. LPL is central for the metabolism of lipids in blood. The enzyme acts at the endothelial surface of the capillary bed where it hydrolyzes triglycerides in circulating triglyceride-rich lipoproteins (TRLs) and thereby allows uptake of fatty acids in adjacent tissues. LPL activity has to be rapidly modulated to adapt to the metabolic demands of different tissues. The current view is that LPL is constitutively expressed and that the rapid modulation of the enzymatic activity occurs by some different controller proteins. Angiopoietin-like protein 4 (ANGPTL4) is one of the main candidates for control of LPL activity. ANGPTL4 causes irreversible inactivation through dissociation of the active LPL dimer to inactive monomers. Other proteins that have effects on LPL activity are the APOCs which are surface components of the substrate TRLs. APOC2 is a well-known LPL co-factor, whereas APOC1 and APOC3 independently inhibit LPL activity. Given the important role of LPL for triglyceride homeostasis in blood, the aim of this thesis was to find small molecules that could increase LPL activity and serve as lead compounds in future drug discovery efforts. Another aim was to investigate the molecular mechanisms for how APOC1 and APOC3 inhibit LPL activity. Using a small molecule screening library we have identified small molecules that can protect LPL from inactivation by ANGPTL4 during incubations in vitro. Following a structure-activity relationship study we have synthesized lead compounds that more efficiently protect LPL from inactivation by ANGPTL4 in vitro and also have dramatic triglyceride-lowering properties in vivo. In a separate study we show that low concentrations of fatty acids possess the ability to prevent inactivation of LPL by ANGPTL4 under in vitro conditions. With regard to APOC1 and APOC3 we demonstrate that when bound to TRLs, these apolipoproteins prevent binding of LPL to the lipid/water interface. This results in decreased lipolysis and in an increased susceptibility of LPL to inactivation by ANGPTL4. We demonstrate that hydrophobic amino acid residues that are centrally located in the APOC3 molecule are critical for attachment of this protein to lipid emulsion particles and consequently for inhibition of LPL activity. In summary, this work has identified a lead compound that protects LPL from inactivation by ANGPTL4 in vitro and lowers triglycerides in vivo. In addition, we propose a molecular mechanism for inhibition of LPL activity by APOC1 and APOC3.

L’hypertriglycéridémie (hyperTG) est une dyslipidémie fréquente, caractérisée par une augmentation de la concentration plasmatique en triglycérides (TG). L’hyperTG est considérée comme un facteur de risque indépendant de la maladie cardiovasculaire, particulièrement de la maladie coronarienne athérosclérotique. Plusieurs facteurs environnementaux et génétiques ont été associés avec l’hyperTG. Cependant, près de 90% des cas d’hyperTG primaire sont encore incomplètement caractérisés au niveau moléculaire. Dernièrement, la protéine GPIHBP1 (glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1), qui a un rôle clef dans le métabolisme des TG, a été associée à l’expression d’hyperTG sévère et rare chez l’humain. Ce mémoire présente les résultats de nos travaux qui ont été effectués afin d’identifier de nouvelles bases moléculaires associées à l’expression de l’hyperTG dans le locus du gène GPIHBP1. Nous avons observé que le polymorphisme GPIHBP1 g.-469G>A (rs72691625), dont la fréquence de l’allèle mineure a été évaluée à 19,6% dans notre échantillon, serait associé à l’expression d’hyperTG (TG ≥ 2mmol/L) dans une population canadienne-française. Ce polymorphisme est associé à un risque 1,67 fois plus grand d’exprimer une triglycéridémie ≥ 2mmol/L chez les porteurs hétérozygotes et 5,7 fois plus grand chez les porteurs homozygotes, comparativement aux non-porteurs. Ce risque d’hyperTG serait exacerbé par la présence concomitante d’une mutation hypertriglycéridémiante dans le gène codant pour la lipoprotéine lipase. La présence de ce polymorphisme serait particulièrement associée à l’expression de la dysbêtalipoprotéinémie familiale et de l’hypertriglycéridémie familiale endogène. GPIHBP1 g.-469G>A est le premier polymorphisme fréquent identifié dans le promoteur du gène à être associé avec l’expression d’hyperTG. GPIHBP1 émerge de plus en plus comme un gène candidat intéressant pour la recherche de nouvelles bases moléculaires pouvant expliquer certaines formes d’hyperTG primaire fréquente. / Hypertriglyceridemia (hyperTG) is a frequent dyslipidemia referring to an increased fasting plasma triglyceride (TG) level ≥ 2 mmol/L. HyperTG is an independent risk factor for cardiovascular disease, such as coronary artery diseases. Several environmental and genetic factors have been associated with hyperTG. Although several gene factors were associated with hyperTG, nearly 90% of cases of primary hyperTG are still incompletely characterized at the molecular level. Recently, few cases of rare and severe hyperTG have been associated with some rare polymorphisms in the gene coding for GPIHBP1 (glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1). This manuscript resumes our research regarding the identification of new molecular bases associated with the expression of frequent hyperTG subtypes in the gene locus GPIHBP1. Our results show that the GPIHBP1 g.-469G>A polymorphism (rs72691625), whose the minor allele frequency was estimated to 19.6% in our sample, was associated with the expression of hyperTG (TG ≥ 2 mmol/L) in a French-Canadian population. Subjects heterozygous and homozygous for this polymorphism respectively had a 1.67-fold and 5.70-fold increased risk to exhibit plasma TG levels ≥ 2mmol/L as compared to non-carriers. This increased risk of hyperTG observed in g.-469A carriers seems to be exacerbated by the concomitant presence of a frequent loss-of-function lipoprotein lipase gene variant. This polymorphism seems also particularly associated with dysbetalipoproteinemia and familial hypertriglyceridemia. The g.-469G>A polymorphism is the first common polymorphism in the GPIHBP1 gene promoter to be associated with the expression of hyperTG. GPIHBP1 emerges as a significant candidate for the molecular based of primary hyperTG.

Wichtige Marker in der klinischen Routine für die Risikoabschätzung von kardiovaskulären Erkrankungen (CVD) sind Blutcholesterinwerte auf Basis von Lipoproteinklassen wie ''schlechtes'' LDL oder ''gutes'' HDL. Dies vernachlässigt, dass jede Lipoproteinklasse eine nicht-homogene Population von Lipoproteinpartikeln unterschiedlicher Zusammensetzung aus Lipiden und Proteinen bildet. Studien zeigen zudem, dass solche Sub-populationen von Lipoproteinen im Stoffwechsel als auch im Beitrag zu CVD unterschiedlich sind. Mehrwert und routinemäßiger Einsatz einer detaillierteren Auftrennung von Lipoproteinen sind jedoch umstritten, da die experimentelle Fraktionierung und Analyse aufwendig, zeit- und kostenintensiv sind. Die vorliegende Arbeit ''Computational Lipidology'' präsentiert einen neuartigen Modellierungsansatz für die Berechnung von Lipoproteinverteilungen (Lipoproteinprofil) im Blutplasma, wobei erstmals individuelle Lipoproteinpartikel anstelle von Lipoproteinklassen betrachtet werden. Das Modell berücksichtigt elementare Bestandteile (Lipide, Proteine) und Prozesse des Stoffwechsel von Lipoproteinen. Stochastische wie deterministische Simulationen errechnen auf Basis aller Lipoproteinpartikel im System deren Dichteverteilung. Die Modellberechnungen reproduzieren erfolgreich klinisch gemessene Lipoproteinprofile von gesunden Patienten und zeigen Hauptmerkmale von pathologischen Situationen, die durch Störung eines der zugrundeliegenden molekularen Prozesse verursacht werden. Hochaufgelöste Lipoproteinprofile zeigen die Verteilung von sogenannten ''high-resolution density sub-fractions'' (hrDS) innerhalb von Hauptlipoproteinklassen. Die Ergebnisse stimmen mit klinischen Beobachtungen sehr gut überein, was die Arbeit als einen signifikanten Schritt in Richtung Analyse von individuellen Unterschieden, patienten-orientierte Diagnose von Fettstoffwechselstörungen und Identifikation neuer Sub-populationen von potentiell klinischer Relevanz qualifiziert. / Monitoring the major lipoprotein classes, particularly low-density lipoproteins (''bad'' LDL) and high-density lipoproteins (''good'' HDL) for characterizing risk of cardiovascular disease (CVD) is well-accepted and routine in clinical practice. However, it is only one-half of the truth as lipoprotein classes comprise non-homogeneous populations of lipoprotein particles varying significantly in their composition of lipids and apolipoproteins. Various studies have shown differing metabolic behavior and contribution to CVD of individual lipoprotein sub-populations. Nevertheless, the superiority of more detailed lipoprotein fractionation is still a matter of debate because experimental separation and analysis is an elaborate, time-consuming and expensive venture and not yet worthwhile for routine measurements. The present work ''Computational Lipidology'' aims at establishing a novel modeling approach to calculate the distribution of lipoproteins (lipoprotein profile) in blood plasma being the first that settles on individual lipoprotein complexes instead of common lipoprotein classes. Essential lipoprotein constituents and processes involved in the lipoprotein metabolism are taken into account. Stochastic as well as deterministic simulations yield the distribution of lipoproteins over density based on the set of individual lipoprotein complexes in the system. The model calculations successfully reproduce lipoprotein profiles measured in healthy subjects and show main characteristics of pathological situations elicited by disorder in one of the underlying molecular processes. Moreover, the model reveals the distribution of high-resolution lipoprotein sub-fractions (hrDS) within major density classes. The results show satisfactory agreement with clinical observations which qualifies the work as a significant step towards analyzing inter-individual variability, patient-oriented diagnosis of lipid disorders and identifying new sub-fractions of potential clinical relevance.