Microencapsulated Pomegranate Modifies the Composition and Function of High-Density Lipoproteins (HDL) in New Zealand Rabbits.

Previous studies demonstrated that pomegranate, which is a source of several bioactive molecules, induces modifications of high-density lipoproteins (HDL) lipid composition and functionality. However, it remains unclear whether the beneficial effects of pomegranate are related to improvement in the lipid components of HDL. Therefore, in this placebo-controlled study, we characterized the size and lipid composition of HDL subclasses and assessed the functionality of these lipoproteins after 30 days of supplementation with a pomegranate microencapsulated (MiPo) in New Zealand white rabbits. We observed a significant decrease in plasma cholesterol, triglycerides, and non-HDL sphingomyelin, as well as increases in HDL cholesterol and HDL phospholipids after supplementation with MiPo. Concomitantly, the triglycerides of the five HDL subclasses isolated by electrophoresis significantly decreased, whereas phospholipids, cholesterol, and sphingomyelin of HDL subclasses, as well as the HDL size distribution remained unchanged. Of particular interest, the triglycerides content of HDL, estimated by the triglycerides-to-phospholipids ratio, decreased significantly after MiPo supplementation. The modification on the lipid content after the supplementation was associated with an increased resistance of HDL to oxidation as determined by the conjugated dienes formation catalyzed by Cu2+. Accordingly, paraoxonase-1 (PON1) activity determined with phenylacetate as substrate increased after MiPo. The effect of HDL on endothelial function was analyzed by the response to increasing doses of acetylcholine of aorta rings co-incubated with the lipoproteins in an isolated organ bath. The HDL from rabbits that received placebo partially inhibited the endothelium-dependent vasodilation. In contrast, the negative effect of HDL on endothelial function was reverted by MiPo supplementation. These results show that the beneficial effects of pomegranate are mediated at least in part by improving the functionality of HDL, probably via the reduction of the content of triglycerides in these lipoproteins.

Bone Morphogenetic Protein-2 and Osteopontin Gene Expression in Epicardial Adipose Tissue from Patients with Coronary Artery Disease Is Associated with the Presence of Calcified Atherosclerotic Plaques.

PURPOSE: It has been proposed that the cardiovascular effects of obesity are related to epicardial adipose tissue (EAT), which seems to play an active role on the development and calcification of atherosclerotic plaques, but the mechanisms are still unknown. Therefore, the aim of this study was to determine whether the EAT expresses the genes of calcifying factors and whether such expression is associated with the body mass index (BMI) and with the presence of coronary artery calcium (CAC) in patients with coronary artery disease (CAD). PATIENTS AND METHODS: Forty-three patients with CAD were enrolled specifically for this study, and their CAC score and EAT volume were determined by computed tomography. As the group of comparison, 41 patients with aortic valve stenosis and CAC = 0 were included (control group). A representative subgroup of 16 CAD patients and 23 controls were selected to obtain EAT biopsies during the chirurgical procedure from the atrio-interventricular groove. The mRNA expression of bone morphogenetic protein-2 and -4 (BMP-2, BMP-4), osteopontin (OPN), osteonectin (ON), and osteoprotegerin (OPG) in EAT was determined by qPCR. RESULTS: The gene expression of OPN and BMP-2 was 70% and 52% higher in the EAT from CAD patients than that in controls, respectively, whereas the expression of OPG, ON, and BMP-4 was similar in both groups. The EAT volume positively correlated with OPG and with the BMI, suggesting a relationship of obesity with local higher expression of calcifying genes in the coronary territory. The logistic regression analysis showed that high levels of both OPN and BMP-2 increased about 6 and 8 times the odds of coronary calcification (CAC score > 0), respectively. CONCLUSION: EAT correlated with BMI and expressed the mRNA of calcifying genes but only OPN and BMP-2 expression was higher in CAD patients. Higher levels of both OPN and BMP-2 statistically determined the presence of calcium in coronary arteries of CAD patients.

Atorvastatin and Fenofibrate Increase the Content of Unsaturated Acyl Chains in HDL and Modify In Vivo Kinetics of HDL-Cholesteryl Esters in New Zealand White Rabbits.

Previous studies demonstrated modifications of high-density lipoproteins (HDL) structure and apolipoprotein (apo) A-I catabolism induced by the atorvastatin and fenofibrate combination. However, it remains unknown whether such structural and metabolic changes of HDL were related to an improvement of the HDL-cholesteryl esters (HDL-CE) metabolism. Therefore, we determined the structure of HDL and performed kinetic studies of HDL-CE radiolabeled with tritium in rabbits treated with atorvastatin, fenofibrate, and a combination of both drugs. The atorvastatin and fenofibrate combination increased the HDL size and the cholesterol and phospholipid plasma concentrations of the largest HDL subclasses. Moreover, the relative amount of unsaturated fatty acids contained in HDL increased, in detriment of saturated fatty acids as determined by gas chromatography-mass spectrometry. The transfers of cholesteryl esters (CE) from HDL to very low-density lipoproteins/low-density lipoproteins (VLDL/LDL) and vice versa were enhanced with atorvastatin, alone or in combination. Moreover, the direct elimination of CE from plasma via VLDL/LDL decreased with fenofibrate, whereas the direct elimination of CE via HDL augmented with the combination treatment. Taken together, the rise of unsaturated fatty acid content and the size increase of HDL, suggest that atorvastatin and fenofibrate induce more fluid HDL particles, which in turn favor an enhanced CE exchange between HDL and VLDL/LDL. Our results contribute to a better understanding of the relationship between the structure and function of HDL during the use of anti-dyslipidemic drugs.

Atorvastatin and Fenofibrate Exert Opposite Effects on the Vascularization and Characteristics of Visceral Adipose Tissue in New Zealand White Rabbits.

Statins may precipitate the onset of type 2 diabetes (T2D) in high-risk patients. In contrast, only the subset of individuals with insulin resistance and/or diabetes receives cardiovascular benefits with fibrates. In this context, previous observations from our laboratory suggested that atorvastatin induced an increase in visceral adipose tissue (VAT), whereas fenofibrate had the opposite effects in rabbits. Therefore, we determined the mass, morphology, and vascularization of VAT in New Zealand white rabbits (n = 6/group) that received 0.33 or 2.6 mg/kg/d of atorvastatin or fenofibrate, respectively, during 2 months. As expected, the cholesterol from the atorvastatin group was lower after treatment, while triglycerides decreased in the fenofibrate group. The mass of VAT from the fenofibrate group was 46% lower compared to the controls, meanwhile atorvastatin was associated with a larger diameter of adipocytes (+65%) than that of the control and fenofibrate groups. Fibroblast growth factor 2 (FGF2) gene expression was lower in the fenofibrate group than in the control group (-54%). By contrast, vascular endothelial growth factor A (VEGF-A) gene expression in fenofibrate-treated rabbits was 110% higher than in the control group. In agreement with the gene expression, the marker of angiogenesis platelet endothelial cell adhesion molecule 1 was slightly but significantly higher (+10%) in rabbits treated with fenofibrate than in controls, as determined by immunohistochemistry. These results suggest that fenofibrate is associated with a favorable remodeling of VAT, that is, reduced mass and increased vascularization in normolipemic rabbits; in contrast, atorvastatin induced a nonfavorable remodeling of VAT. These results may be related to the cardiovascular benefits of fenofibrate and the increased risk of T2D in high-risk patients induced by atorvastatin.

Hyperuricemia is Associated with Increased Apo AI Fractional Catabolic Rates and Dysfunctional HDL in New Zealand Rabbits.

The potential cause-effect relationship between uric acid plasma concentrations and HDL functionality remains elusive. Therefore, this study aimed to explore the effect of oxonic acid (OA)-induced hyperuricemia on the HDL size distribution, lipid content of HDL subclasses, and apo AI turnover, as well as HDL functionality in New Zealand white rabbits. Experimental animals received OA 750 mg/kg/day by oral gavage during 21 days. The HDL-apo AI fractional catabolic rate (FCR) was determined by exogenous labeling with 125I, and HDL subclasses were determined by sequential ultracentrifugation and PAGE. Paraoxonase-1 activity (PON-1) and the effect of HDL on relaxation of aorta rings in vitro were determined as an indication of HDL functionality. Oxonic acid induced a sixfold increase of uricemia (0.84 ± 0.06 vs. 5.24 ± 0.12 mg/dL, P < 0.001), and significant decreases of triglycerides and phospholipids of HDL subclasses, whereas HDL size distribution and HDL-cholesterol remained unchanged. In addition, HDL-apo AI FCR was significantly higher in hyperuricemic rabbits than in the control group (0.03697 ± 0.0038 vs. 0.02605 ± 0.0017 h-1 respectively, P < 0.05). Such structural and metabolic changes were associated with lower levels of PON-1 activities and deleterious effects of HDL particles on endothelium-mediated vasodilation. In conclusion, hyperuricemia is associated with structural and metabolic modifications of HDL that result in impaired functionality of these lipoproteins. Our data strongly suggest that uric acid per se exerts deleterious effects on HDL that contribute to increase the risk of atherosclerosis.

Small HDL subclasses become cholesterol-poor during postprandial period after a fat diet intake in subjects with high triglyceridemia increases.

BACKGROUND: Postprandial triglyceridemia may transitory affect the structure of HDL subclasses and probably their antiatherogenic properties but little is known in this field. We analyzed the HDL subclasses lipid content along postprandial period. METHODS: Fifteen metabolic syndrome (MS) patients and 15 healthy controls were enrolled. HDL were isolated from plasma samples obtained at fasting and every 2-h up to 8-h, after a 75-g fat meal. Cholesterol (C), triglycerides (TAG), and phospholipid (Ph) plasma concentrations of five HDL subclasses were determined by densitometry of electrophoresis gels enzymatically stained. RESULTS: The increase of postprandial triglyceridemia expressed as the incremental area under the curve (iAUC) was twice in MS patients than in controls. Only large HDL2b-TAG were higher in MS than controls at 4, 6 and 8h after meal intake, whereas cholesterol of HDL2a, 3a and 3b were lower at 8h. HDL size distribution shifted towards large HDL and HDL3a-, 3b- and 3c-subclasses had a lower content of cholesterol (estimated by the C-to-Ph ratio) in subjects whose iAUC>289.5mgh/dl (n=15) in comparison with those subjects with iAUC below this cutoff point (n=15), independently of the MS status and fasting TAG. Triglycerides content of HDL subclasses changed only discreetly along the postprandial period, whereas paraoxonase-1 remained unchanged. CONCLUSIONS: A high postprandial triglyceridemia conditions the shift of HDL size distribution towards large particles and the decrease of cholesterol in HDL3 subclasses. These data demonstrate that postprandial hypertriglyceridemia contributes to a transitory hypoalphalipoproteinemia that may increase the risk of cardiovascular disease.

Increased HDL Size and Enhanced Apo A-I Catabolic Rates Are Associated With Doxorubicin-Induced Proteinuria in New Zealand White Rabbits.

The catabolism and structure of high-density lipoproteins (HDL) may be the determining factor of their atheroprotective properties. To better understand the role of the kidney in HDL catabolism, here we characterized HDL subclasses and the catabolic rates of apo A-I in a rabbit model of proteinuria. Proteinuria was induced by intravenous administration of doxorubicin in New Zealand white rabbits (n = 10). HDL size and HDL subclass lipids were assessed by electrophoresis of the isolated lipoproteins. The catabolic rate of HDL-apo A-I was evaluated by exogenous radiolabelling with iodine-131. Doxorubicin induced significant proteinuria after 4 weeks (4.47 ± 0.55 vs. 0.30 ± 0.02 g/L of protein in urine, P < 0.001) associated with increased uremia, creatininemia, and cardiotoxicity. Large HDL2b augmented significantly during proteinuria, whereas small HDL3b and HDL3c decreased compared to basal conditions. HDL2b, HDL2a, and HDL3a subclasses were enriched with triacylglycerols in proteinuric animals as determined by the triacylglycerol-to-phospholipid ratio; the cholesterol content in HDL subclasses remained unchanged. The fractional catabolic rate (FCR) of [(131)I]-apo A-I in the proteinuric rabbits was faster (FCR = 0.036 h(-1)) compared to control rabbits group (FCR = 0.026 h(-1), P < 0.05). Apo E increased and apo A-I decreased in HDL, whereas PON-1 activity increased in proteinuric rabbits. Proteinuria was associated with an increased number of large HDL2b particles and a decreased number of small HDL3b and 3c. Proteinuria was also connected to an alteration in HDL subclass lipids, apolipoprotein content of HDL, high paraoxonase-1 activity, and a rise in the fractional catabolic rate of the [(131)I]-apo A-I.

Atorvastatin and fenofibrate combination induces the predominance of the large HDL subclasses and increased apo AI fractional catabolic rates in New Zealand white rabbits with exogenous hypercholesterolemia.

The anti-atherogenic properties of high-density lipoproteins (HDLs) may be related to their structure and metabolism. The HDL physicochemical characteristics that determine their plasma clearance during treatment with statins and fibrates are not well understood. In this study, we analyzed HDL-apo AI fractional catabolic rates (FCRs), size distributions, and the lipid composition of the HDL subclasses in New Zealand white rabbits with exogenous dyslipidemia that received low doses of atorvastatin and fenofibrate. Hypercholesterolemia decreased only partially with the combination of both drugs. HDL size distribution shifted toward larger particles among the groups of rabbits that received atorvastatin, fenofibrate, or their combination, compared with both the control group and the dyslipidemic group. The HDL subclasses were significantly rich in cholesterol in each of the groups compared with controls. The structural changes noted in the HDL subclasses were not associated with impaired plasma paraoxonase-1 (PON1) activity. The groups receiving monotherapy and the drug combination group were all associated with a higher apo AI FCR value compared with both the dyslipidemic rabbits and the control group. In conclusion, the combination of atorvastatin and fenofibrate induced a more favorable HDL subclass profile than did the individual use of these drugs. Similarly, the apo AI FCR values were augmented in every group receiving drug treatment (either monotherapy or combination therapy) in the setting of hypercholesterolemia. The anti-atherogenic properties of HDLs, excluding their capacity to bind PON1, may be enhanced by the structural and metabolic modifications induced by the combination of atorvastatin and fenofibrate.