Density High-density Lipoprotein Research Articles

Associations between Endothelial Lipase, High-Density Lipoprotein, and Endothelial Function Differ in Healthy Volunteers and Metabolic Syndrome Patients.

Metabolic syndrome (MS) is characterized by endothelial- and high-density lipoprotein (HDL) dysfunction and increased endothelial lipase (EL) serum levels. We examined the associations between EL serum levels, HDL (serum levels, lipid content, and function), and endothelial function in healthy volunteers (HV) and MS patients. Flow-mediated dilation (FMD), nitroglycerin-mediated dilation (NMD), serum levels of HDL subclasses (measured by nuclear magnetic resonance (NMR) spectroscopy), and EL serum levels differed significantly between HV and MS patients. The serum levels of triglycerides in large HDL particles were significantly positively correlated with FMD and NMD in HV, but not in MS patients. Cholesterol (C) and phospholipid (PL) contents of large HDL particles, calculated as HDL1-C/HDL1-apoA-I and HDL1-PL/HDL1-apoA-I, respectively, were significantly negatively correlated with FMD in HV, but not in MS patients. Cholesterol efflux capacity and arylesterase activity of HDL, as well as EL, were correlated with neither FMD nor NMD. EL was significantly negatively correlated with HDL-PL/HDL-apoA-I in HV, but not in MS patients, and with serum levels of small dense HDL containing apolipoprotein A-II in MS patients, but not in HV. We conclude that MS modulates the association between HDL and endothelial function, as well as between EL and HDL. HDL cholesterol efflux capacity and arylesterase activity, as well as EL serum levels, are not associated with endothelial function in HV or MS patients.

Atherogenic lipidomics profile in healthy individuals with low cardiorespiratory fitness: The HUNT3 fitness study

Background and aimsLow cardiorespiratory fitness is a strong and independent risk factor for cardiovascular disease (CVD). Serum profiling of healthy individuals with large differences in cardiorespiratory fitness may therefore reveal early biomarkers of CVD development. Thus, we aimed to identify circulating lipoprotein subfractions differentially expressed between groups of individuals with large differences in cardiorespiratory fitness, measured as maximal oxygen uptake (VO2max). MethodsHealthy subjects (40–59 years) were selected from the third wave of the Trøndelag health study (HUNT3) based on having an age-dependent high VO2max (47.1 ± 7.7 mL kg−1·min−1, n = 103) or low VO2max (31.4 ± 4.9 mL kg−1·min−1, n = 108). The individuals were matched on gender, age, physical activity level and fasting status. Results99 lipoprotein subfractions were quantified in serum samples using nuclear magnetic resonance (NMR) lipidomics. Standard clinical analyses showed similar levels of total cholesterol, low-density lipoprotein (LDL)-cholesterol and high-density lipoprotein (HDL)-cholesterol between the groups, and slightly higher levels of triglycerides in participants with low VO2max. Thirteen lipoprotein subfractions were increased in the low VO2max group compared to the high VO2max group (p < 0.005), including mainly large very low-density lipoprotein (VLDL) subfractions. In addition, triglyceride levels in small-sized HDL and LDL particles were increased in participants with low VO2max. Correlation analyses between VO2max and lipoproteins subfractions displayed a negative correlation between VO2max and the levels of cholesterol and phospholipids in the small HDL particles. The lipoprotein profile of individuals with low VO2max is similar to the profile of insulin resistant individuals. ConclusionsLow VO2max was associated with enrichment of large VLDL particles, as well as an increased triglycerides content in the small and dense HDL and LDL particles. This unfavorable lipid profile is likely to be involved in the strong associations between VO2max and CVD.

Biological Variability of Small Dense LDL, HDL3, and Triglyceride-Rich Lipoprotein Cholesterol

Abstract The guideline-recommended lipid panel for cardiovascular disease (CVD) risk assessment measures total cholesterol, high-density lipoprotein (HDL) cholesterol, triglycerides, and calculated low-density lipoprotein (LDL) cholesterol. Measured cholesterol in subfractions of HDL and LDL purportedly improve CVD risk prediction. Homogenous enzymatic methods are now available for measurement of the cholesterol within small dense LDL (sLDL), small dense HDL (HDL3), and triglyceride-rich lipoproteins (TRL). For meaningful interpretation of these measurements, an understanding of the potential sources and extent of result variability is needed. The smallest difference between serial measurements within a patient that likely reflects a change in clinical status is called the reference change value (RCV). Biological variability and reference change values (RCV) are well-characterized for basic lipids but there is limited information for sLDL, HDL3 or TRL. The objective of this study was to determine intra- and inter-individual variability for sLDL, HDL3, and TRL in a healthy reference population. Serum samples were collected from 24 healthy subjects (n=14 female/10 male) daily for three days (non-fasting), daily for five days (fasting), weekly for four weeks (fasting), and monthly for 7 months (fasting). sLDL, HDL3, and TRL cholesterol were measured in duplicate by enzymatic colorimetric assays (Denka, Japan) on a Roche Cobas c501. Each source of variability (between subject, within subject, and analytical) was calculated using random-effects regression models to estimate each variance component including the overall variation, standard deviation (SD), coefficient of variation (CV), and proportion of total variance (between-subject, within-subject, and analytical). Using these analytical and biological variances, the reference change value (RCV), index of individuality (IoI), and intraclass correlation coefficient (ICC) were determined. Analytic variability (CVa) from monthly testing was 1.2%, 1.1%, and 1.5% for sLDL, HDL3, and TRL, respectively. Monthly within-subject variability (CVw) was 17.1% for sLDL, 7.4% for HDL3 and 25.7% for TRL. Monthly between-subject variability (CVb) was 32.2%, 13.93%, and 33.4% for sLDL, HDL3, and TRL, respectively. Most of the monthly variation was attributed to between-subject variation for all three tests. Within-subject variation accounted for 37% of TRL variation and 22% for both sLDL and HLD3. Within-subject RCVs for monthly measurements were 16.9mg/dL for sLDL, 5.3mg/dL for HDL3, and 15.1mg/dL for TRL. IoIs for monthly testing were 0.81 for TRL, 0.57 for sLDL, and 0.61 for HDL3. Our data demonstrate that sLDL, HDL3, and TRL show low analytical variability, moderate within-subject variability, but high between-subject variability when measured by homogenous assays in a healthy population. The IoI value (&amp;gt;0.6) for TRL suggests use of a reference interval is appropriate for result interpretation. Conversely, clinical cut-points may be more useful than reference intervals for sLDL and HDL3 which had IoIs ~0.6. These findings may be useful for clinical interpretation, particularly when comparing successive measurements of these analytes.

Structure dynamics of ApoA-I amyloidogenic variants in small HDL increase their ability to mediate cholesterol efflux

Apolipoprotein A-I (ApoA-I) of high density lipoproteins (HDLs) is essential for the transportation of cholesterol between peripheral tissues and the liver. However, specific mutations in ApoA-I of HDLs are responsible for a late-onset systemic amyloidosis, the pathological accumulation of protein fibrils in tissues and organs. Carriers of these mutations do not exhibit increased cardiovascular disease risk despite displaying reduced levels of ApoA-I/HDL cholesterol. To explain this paradox, we show that the HDL particle profiles of patients carrying either L75P or L174S ApoA-I amyloidogenic variants show a higher relative abundance of the 8.4-nm versus 9.6-nm particles and that serum from patients, as well as reconstituted 8.4- and 9.6-nm HDL particles (rHDL), possess increased capacity to catalyze cholesterol efflux from macrophages. Synchrotron radiation circular dichroism and hydrogen-deuterium exchange revealed that the variants in 8.4-nm rHDL have altered secondary structure composition and display a more flexible binding to lipids than their native counterpart. The reduced HDL cholesterol levels of patients carrying ApoA-I amyloidogenic variants are thus balanced by higher proportion of small, dense HDL particles, and better cholesterol efflux due to altered, region-specific protein structure dynamics.

Patients With Systemic Lupus Erythematosus Show an Increased Arterial Stiffness That is Predicted by IgM Anti-β2 -Glycoprotein I and Small Dense High-Density Lipoprotein Particles.

To investigate the metabolic and immunologic factors associated with the presence of central arterial stiffness as measured by the augmentation index (AIx). We conducted a cross-sectional study of 69 female patients with systemic lupus erythematosus (SLE) compared with a control group of 34 healthy women. The anthropometrical variables, the vascular studies, and the analytic data were obtained the same day. The AIx was assessed by peripheral arterial tonometry. The analysis of lipoprotein populations was performed using nuclear magnetic resonance (NMR) spectroscopy. Arterial stiffness was increased in patients with SLE compared with control subjects (mean ± SD 20.30 ± 21.54% versus 10.84 ± 11.51%; P = 0.0021). Values for the AIx were correlated with the Framingham risk score (r = 0.481, P < 0.001), carotid intima-media thickness (r = 0.503, P < 0.001), systolic blood pressure (r = 0.270, P < 0.001), and age (r = 0.365, P < 0.001). Patients receiving antimalarial drugs had a lower AIx (mean ± SD 11.74 ± 11.28% versus 24.97 ± 20.63%; P = 0.024). The AIx was correlated with the atherogenic lipoproteins analyzed by NMR. The immunologic variables associated with the AIx were C4 (r = 0.259, P = 0.046) and IgM anti-β2 -glycoprotein I (IgM anti-β2 GPI) (r = 0.284, P = 0.284). In the multivariate analysis, age (β = 0.347, 95% confidence interval [95% CI] 0.020-0.669, P = 0.035), IgM β2 GPI (β = 0.321, 95% CI 0.024-0.618, P = 0.035) and small dense high-density lipoprotein (HDL) particles (β = 1.288, 95% CI 0.246-2.329, P = 0.017) predicted the AIx. SLE patients had increased arterial stiffness compared with healthy control subjects. Arterial stiffness was decreased in patients treated with antimalarial drugs. Age, IgM β2 GPI, and the number of small dense HDL particles predicted the AIx.

Pathophysiology of Diabetic Dyslipidemia

Accumulating clinical evidence has suggested serum triglyceride (TG) is a leading predictor of atherosclerotic cardiovascular disease, comparable to low-density lipoprotein (LDL)-cholesterol (C) in populations with type 2 diabetes, which exceeds the predictive power of hemoglobinA1c. Atherogenic dyslipidemia in diabetes consists of elevated serum concentrations of TG-rich lipoproteins (TRLs), a high prevalence of small dense low-density lipoprotein (LDL), and low concentrations of cholesterol-rich high-density lipoprotein (HDL)2-C. A central lipoprotein abnormality is an increase in large TG-rich very-low-density lipoprotein (VLDL)1, and other lipoprotein abnormalities are metabolically linked to increased TRLs. Insulin critically regulates serum VLDL concentrations by suppressing hepatic VLDL production and stimulating VLDL removal by activation of lipoprotein lipase. It is still debated whether hyperinsulinemia compensatory for insulin resistance is causally associated with the overproduction of VLDL. This review introduces experimental and clinical observations revealing that insulin resistance, but not hyperinsulinemia stimulates hepatic VLDL production. LDL and HDL consist of heterogeneous particles with different size and density. Cholesterol-depleted small dense LDL and cholesterol-rich HDL2 subspecies are particularly affected by insulin resistance and can be named “Metabolic LDL and HDL,” respectively. We established the direct assays for quantifying small dense LDL-C and small dense HDL(HDL3)-C, respectively. Subtracting HDL3-C from HDL-C gives HDL2-C. I will explain clinical relevance of measurements of LDL and HDL subspecies determined by our assays. Diabetic kidney disease (DKD) substantially worsens plasma lipid profile thereby potentiated atherogenic risk. Finally, I briefly overview pathophysiology of dyslipidemia associated with DKD, which has not been so much taken up by other review articles.

Decreased serum PON1 arylesterase activity in familial hypercholesterolemia patients with a mutated LDLR gene.

Paraoxonase 1 (PON1) is a serum enzyme associated with high density lipoprotein (HDL) regulation through its paraoxonase and arylesterase activity. PON1 inhibits the oxidation of HDL and low density lipoprotein (LDL), and is involved in the pathogenesis of a variety of diseases including atherosclerosis. Conversely, mutations in the low density lipoprotein receptor (LDLR) result in failure of receptor mediated endocytosis of LDL leading to its elevated plasma levels and onset of familial hypercholesterolemia (FH). In the current study we investigated the role of PON1 polymorphisms rs662; c.575A > G (p.Gln192Arg) and rs854560; c.163T > A (p.Leu55Met) in a large family having FH patients harboring a functional mutation in LDLR. Genotypes were revealed by RFLP, followed by confirmation through Sanger sequencing. PON1 activity was measure by spectrophotometry. Our results show significantly reduced serum paraoxonase and arylesterase activities in FH patients compared with the healthy individuals of the family (p < 0.05). PON1 QQ192 genotype showed a significantly higher association with FH (p=0.0002). PON1 Q192 isoform was associated with reduced serum paraoxonase activity by in silico analysis and PON1 R192 exhibited higher serum paraoxonase and arylesterase activity than the other polymorphs. Our results highlight that the combination of LDLR mutations and PON1 MMQQ genotypes may lead to severe cardiac events.

PON1 L55M and Q192R gene polymorphisms and CAD risks in patients with hyperlipidemia : Clinical study of possible associations.

A decreased plasma high density lipoprotein (HDL) cholesterol level is a strong risk factor for coronary artery disease (CAD). Antioxidant activity of HDL mainly lies in the activity of paraoxonase (PON). This study aimed to investigate the relationships between PON1 L55M and Q192R polymorphisms, and the risks of CAD in patients with hyperlipidemia. From January 2014 to January 2016, 244 patients were divided into hyperlipidemia, hyperlipidemia+ CAD, and control groups. The hyperlipidemia and hyperlipidemia+ CAD groups were designated as the case group. Serum PON1 concentrations were measured using the enzyme-linked immunosorbent assay. After isolating genomic DNA, the PON1 L55M and Q192R genes were amplified by polymerase chain reaction and sequenced. In the case group, the genotypes LM and LL were detected significantly more often than in the control group, as were the alleles R (33.33%, 42.12%) and L (22.78%, 29.11%). The frequency of QR and RR genotypes was significantly higher in the hyperlipidemia+ CAD group than in the hyperlipidemia group; the allele R in the hyperlipidemia+ CAD group (42.77%) was more frequent than in the hyperlipidemia group (23.78%). The Q192R polymorphism was associated with low serum PON1 concentrations, and the lowest concentration was observed in the 192QR+ 192RR genotype (P= 0.03). Logistic regression analysis showed asignificant correlation between the 192R allele and smoking (P= 0.03), body mass index (P= 0.02), systolic blood pressure (P= 0.004), total cholesterol (P= 0.03), triglycerides (P= 0.01), HDL (P= 0.004), and low density lipoprotein (P= 0.02). The PON1 alleles 192R and 55L are associated with CAD, and the Q192R polymorphism may be arisk factor for CAD.

Leukocyte ABCA1 impedes progression of established atherosclerotic lesions after dietary cholesterol lowering in LDLr-/- mice

The ATP-binding cassette transporter A1 (ABCA1) facilitates the efflux of cholesterol and phospholipids to lipid-free apolipoprotein A1 and small dense high-density lipoproteins. Various studies have shown that leukocyte ABCA1 is an anti-atherogenic factor. Dietary cholesterol lowering stabilizes atherosclerotic lesions, however the role of ABCA1 in this process is unknown. Therefore, this study aims to investigate the effect of leukocyte ABCA1 on diet-induced atherosclerotic lesions after withdrawal of the atherogenic diet . Leukocyte ABCA1 was studied by transplanting bone marrow cells from donor mice that were either knock-out (ABCA1-/-) or wild-type for ABCA1 or that overexpressed ABCA1 (ABCA1Tg) into low-density lipoprotein receptor knock-out (LDLr-/-) mice. All three groups of chimeric mice were fed a Western-type diet (WTD: 0.25%cholesterol, 15Ecao butter) for 6 weeks to induce atherosclerotic lesion development. After this period, a baseline group was sacrificed (ABCA1-/- >LDLr-/- n=12; wild-type >LDLr-/- n=14; ABCA1Tg >LDLr-/- n=9) for lesion assessment. The remainder of chimeric mice was switched to a chow diet (low fat, no added cholesterol) for 3 weeks to lower plasma cholesterol levels (ABCA1-/- >LDLr-/- n=14; wild-type >LDLr-/- n=14; ABCA1Tg >LDLr-/- n=12).Withdrawal of the atherogenic diet normalized plasma cholesterol levels in all three groups. As a result, lesion development was stabilized in both the wild-type >LDLr-/- and ABCA1Tg >LDLr-/- mice, however not in the ABCA1-/- >LDLr-/- mice, which displayed a 1.5-fold increase (p Leukocyte ABCA1 is required to halt atherosclerotic lesion progression after dietary cholesterol lowering. Overexpression of ABCA1 did not result in any additional beneficial effects.

Technical note: Characterization of ceramide in bovine lipoproteins

The hepatic synthesis and export of ceramide is enhanced in diabetic monogastrics. Moreover, ceramide in lipoproteins can mediate the development of insulin resistance. We have previously demonstrated that circulating ceramide increases during the progression of insulin resistance in postpartum dairy cows. Considering that the origins of circulating ceramide required investigation, our objective was to develop a method to characterize the ceramide profile of lipoprotein fractions collected from dairy cows. Serum was collected from 4 nonpregnant and nonlactating Holstein dairy cows. Serum lipoproteins were isolated using size exclusion chromatography by fast protein liquid chromatography (SEC-FPLC). Measurement of triacylglycerol (TAG), phospholipid, total cholesterol, and protein was performed using standard colorimetry practices. Following lipid extraction, fractions were analyzed using electrospray ionization tandem mass spectrometry. Data were analyzed as repeated measures using a mixed model. Lipoprotein isolation using SEC-FPLC and subsequent colorimetric analyses confirmed the presence of 4 distinct fractions: TAG-rich, low density (LDL), and large (buoyant) and small (dense) high density lipoprotein (HDL) subclasses. As expected, the fraction representing mixed very low density lipoproteins and chylomicrons primarily contained TAG. Low density lipoprotein fractions were equally enriched with cholesterol and phospholipid. Buoyant HDL contained elevated levels of cholesterol, phospholipid, and protein. In contrast, the fraction containing dense HDL primarily contained protein. Our method revealed that LDL are enriched with ceramides. Ceramides were also compartmentalized to a lesser extent within both HDL subclasses and TAG-rich lipoproteins. Comparable to whole serum, C16:0-ceramide was the predominant ceramide quantified in all lipoprotein subclasses. Interestingly, the proportion of C24:0-ceramide to total ceramide was elevated in TAG-rich lipoproteins, relative to all other lipoprotein subclasses. We conclude that SEC-FPLC coupled with mass spectrometry is a means to quantify ceramides in lipoprotein fractions. Moreover, ceramides are enriched within bovine LDL, and lipoprotein ceramide profiles reflect levels observed in whole serum. Future investigation will need to determine the biological importance of lipoprotein ceramides with distinct C-chains at amide residues.

A Shift in ApoM/S1P Between HDL-Particles in Women With Type 1 Diabetes Mellitus Is Associated With Impaired Anti-Inflammatory Effects of the ApoM/S1P Complex.

Type 1 diabetes mellitus (T1D) patients have an increased risk of cardiovascular disease despite high levels of high-density lipoproteins (HDL). Apolipoprotein M (apoM) and its ligand sphingosine 1-phospate (S1P) exert many of the anti-inflammatory effects of HDL. We investigated whether apoM and S1P are altered in T1D and whether apoM and S1P are important for HDL functionality in T1D. ApoM and S1P were quantified in plasma from 42 healthy controls and 89 T1D patients. HDL was isolated from plasma and separated into dense, medium-dense, and light HDL by ultracentrifugation. Primary human aortic endothelial cells were challenged with tumor necrosis factor-α in the presence or absence of isolated HDL. Proinflammatory adhesion molecules E-selectin and vascular cellular adhesion molecule-1 were quantified by flow cytometry. Activation of the S1P1- receptor was evaluated by analyzing downstream signaling targets and receptor internalization. There were no differences in plasma levels of apoM and S1P between controls and T1D patients, but the apoM/S1P complexes were shifted from dense to light HDL particles in T1D. ApoM/S1P in light HDL particles from women were less efficient in inhibiting expression of vascular cellular adhesion molecule-1 than apoM/S1P in denser particles. The light HDL particles were unable to activate Akt, whereas all HDL subfractions were equally efficient in activating Erk and receptor internalization. ApoM/S1P in light HDL particles were inefficient in inhibiting tumor necrosis factor-α-induced vascular cellular adhesion molecule-1 expression in contrast to apoM/S1P in denser HDL particles. T1D patients have a higher proportion of light particles and hence more dysfunctional HDL, which could contribute to the increased cardiovascular disease risk associated with T1D.

Poor glycemic control in type 2 diabetes enhances functional and compositional alterations of small, dense HDL3c

High-density lipoprotein (HDL) possesses multiple biological activities; small, dense HDL3c particles displaying distinct lipidomic composition exert potent antiatherogenic activities which can be compromised in dyslipidemic, hyperglycemic insulin-resistant states. However, it remains indeterminate (i) whether such functional HDL deficiency is related to altered HDL composition, and (ii) whether it originates from atherogenic dyslipidemia, dysglycemia, or both.In the present work we analyzed compositional characteristics of HDL subpopulations and functional activity of small, dense HDL3c particles in treatment-naïve patients with well-controlled (n=10) and poorly-controlled (n=8) type 2 diabetes (T2D) and in normolipidemic age- and sex-matched controls (n=11).Our data reveal that patients with both well- and poorly-controlled T2D displayed dyslipidemia and low-grade inflammation associated with altered HDL composition. Such compositional alterations in small, dense HDL subfractions were specifically correlated with plasma HbA1c levels. Further analysis using a lipidomic approach revealed that small, dense HDL3c particles from T2D patients with poor glycemic control displayed additional modifications of their chemical composition. In parallel, antioxidative activity of HDL3c towards oxidation of low-density lipoprotein was diminished.These findings indicate that defective functionality of small, dense HDL particles in patients with T2D is not only affected by the presence of atherogenic dyslipidemia, but also by the level of glycemic control, reflecting compositional alterations of HDL.

Lifestyle intervention enhances high-density lipoprotein function among patients with metabolic syndrome only at normal low-density lipoprotein cholesterol plasma levels

Metabolic syndrome (MetS) is associated with altered lipoprotein metabolism and impairment in the functionality of small, dense high-density lipoprotein (HDL) particles secondary to compositional alterations. The objective of this study was to investigate the capacity of a lifestyle program to improve the composition and antioxidative function (AOX) of small dense HDL3c in MetS. Patients with MetS (n=33) not taking lipid-lowering drugs were recruited to follow a 12-week educational program to reduce caloric intake and to increase physical activity. HDL subfractions were preparatively isolated by isopycnic density-gradient ultracentrifugation. AOX of HDL3c was assessed as its capacity to inhibit low-density lipoprotein oxidation induced by an azoinitiator. AOX of HDL3c was significantly improved (mean reduction in the propagation rate of low-density lipoprotein oxidation by HDL3c, -6.8%, P=.03) and systemic oxidative stress, assessed as plasma levels of 8-isoprostanes, tended to decrease in normocholesterolemic MetS patients (low-density lipoprotein cholesterol [LDL-C] < 130mg/dL) but not in patients with elevated LDL-C levels and in the whole study population. In both the whole study population and the normocholesterolemic subgroup, lifestyle intervention resulted in a significant degree of normalization of HDL3c composition, (enrichment in apolipoprotein A-I and cholesteryl esters, depletion in triglycerides), which was more pronounced at LDL-C < 130mg/dL. In patients with MetS, a lifestyle program improves AOX of small, dense HDL in subjects with normal LDL-C levels. Correction of HDL composition, involving partial normalization of apoA-I content and core lipid composition, 2 central features of the lipid hydroperoxide-inactivating capacity of HDL, may account for this effect.

High-density lipoprotein, mitochondrial dysfunction and cell survival mechanisms

Ischemic injury is associated with acute myocardial infarction, percutaneous coronary intervention, coronary artery bypass grafting and open heart surgery. The timely re-establishment of blood flow is critical in order to minimize cardiac complications. Reperfusion after a prolonged ischemic period, however, can induce severe cardiomyocyte dysfunction with mitochondria serving as a major target of ischemia/reperfusion (I/R) injury. An increase in the formation of reactive oxygen species (ROS) induces damage to mitochondrial respiratory complexes leading to uncoupling of oxidative phosphorylation. Mitochondrial membrane perturbations also contribute to calcium overload, opening of the mitochondrial permeability transition pore (mPTP) and the release of apoptotic mediators into the cytoplasm. Clinical and experimental studies show that ischemic preconditioning (ICPRE) and postconditioning (ICPOST) attenuate mitochondrial injury and improve cardiac function in the context of I/R injury. This is achieved by the activation of two principal cell survival cascades: 1) the Reperfusion Injury Salvage Kinase (RISK) pathway; and 2) the Survivor Activating Factor Enhancement (SAFE) pathway. Recent data suggest that high density lipoprotein (HDL) mimics the effects of conditioning protocols and attenuates myocardial I/R injury via activation of the RISK and SAFE signaling cascades. In this review, we discuss the roles of apolipoproteinA-I (apoA-I), the major protein constituent of HDL, and sphingosine 1-phosphate (S1P), a lysosphingolipid associated with small, dense HDL particles as mediators of cardiomyocyte survival. Both apoA-I and S1P exert an infarct-sparing effect by preventing ROS-dependent injury and inhibiting the opening of the mPTP.

Small, dense high-density lipoprotein 3 particles exhibit defective antioxidative and anti-inflammatory function in familial hypercholesterolemia: Partial correction by low-density lipoprotein apheresis

Familial hypercholesterolemia (FH) features elevated oxidative stress and accelerated atherosclerosis driven by elevated levels of atherogenic lipoproteins relative to subnormal levels of atheroprotective high-density lipoprotein (HDL). Small, dense HDL3 potently protects low-density lipoprotein (LDL) against proinflammatory oxidative damage. To determine whether antioxidative and/or anti-inflammatory activities of HDL are defective in FH and whether such defects are corrected by LDL apheresis. Antioxidative and antiinflammatory activities of HDL were evaluated as protection of reference LDL from oxidative stress and capacity to prevent accumulation of proinflammatory oxidised lipids, respectively. Lipid surface rigidity of HDL was assessed using a fluorescent probe. HDL components were measured by analytical approaches. Systemic oxidative stress was characterized as plasma 8-isoprostanes. Pre-LDL-apheresis, FH patients (n = 10) exhibited elevated systemic oxidative stress (3.3-fold, P < 0.001) vs. sex- and age-matched normolipidemic controls (n = 10). Both antioxidative and antiinflammatory activity of HDL3 were impaired (up to -91%, P < 0.01) in FH. Sphingomyelin and saturated fatty acid contents were elevated in FH HDL3, resulting in enhanced lipid surface rigidity. The surface lipid content (phospholipids, free cholesterol) was reduced in FH (up to -15%, P < 0.001), whereas content of core lipids (cholesteryl esters, triglycerides) was elevated (up to +17%, P < 0.001). Molar apolipoprotein A-I content of HDL3 was subnormal in FH. A single LDL-apheresis session partially corrected (by up to 76%) deficient HDL antiatherogenic activities, attenuated systemic oxidative stress and partially normalised both the lipid composition and surface rigidity of HDL particles. FH features elevated oxidative stress and deficient antioxidative and anti-inflammatory activities of small, dense HDL3; such functional deficiency is intimately linked to anomalies in lipid and protein composition, which may impair the capacity of HDL to acquire and inactivate oxidized lipids.

Abstract 392: Effect of HDL on Platelet Apoptosis and Aggregation

Recent clinical trials have again questioned the cardioprotective role of high-density lipoprotein cholesterol (HDL-C). Attention has now focused on high-density lipoprotein (HDL) function. As HDL is highly heterogeneous in composition, and based upon some exciting preliminary results, we suspect that composition of HDL may play differential roles in platelet regulation, thus explaining the variable clinical results. Confirming a number of recent studies, we have found that HDL appears to inhibit platelet aggregation and activation in response to collagen in a time and dose response manner. This should be a clear beneficial effect of HDL, inhibiting thrombosis. However, in the same studies, we also observed that HDL induces substantial platelet mitochondrial membrane potential loss and damage, leading to platelet apoptosis (increased p53 phosphorylation, caspase 3 activation, and cytochrome c release), which has recently been shown to increase thrombosis. We were intrigued by how HDL can seemingly both inhibit and induce thrombosis. Based upon the literature and our further studies, we have developed a series of hypothesis to explain these seemingly opposing actions on platelets; 1) Apoptotic platelets do not aggregate and thus the inhibition of platelet aggregation, 2) With evidence that small dense HDL have more “platelet inhibitory effects” and are more prevalent in atherogenic dyslipidemia and diabetic patient, we hypothesize that the proapoptosis effect of HDL on platelet apoptosis is from small dense HDL and 3) Platelet apoptosis promotes thrombosis. Composition of HDL may have important differential roles on platelets.

Altered lipidome and antioxidative activity of small, dense HDL in normolipidemic rheumatoid arthritis: Relevance of inflammation

Objective: High-density lipoprotein (HDL) particles exert potent antiatherogenic activities, including antioxidative actions, which are relevant to attenuation of atherosclerosis progression. Such activities are enriched in small, dense HDL and can be compromised under conditions of chronic inflammation like rheumatoid arthritis (RA). However, structure–function relationships of HDL largely remain indeterminate. Methods: The relationships between HDL structure and function were evaluated in normolipidemic patients with active RA (DAS28 > 3.2; n = 12) and in normolipidemic age-matched controls (n = 10). Small, dense HDL3b and 3c particles were isolated from plasma or serum by density gradient ultracentrifugation and their physicochemical characteristics, lipidome (by LC/MS/MS) and antioxidative function (as protection of normolipidemic LDL from free radical-induced oxidation) were evaluated. Results: As expected, active RA patients featured significantly elevated plasma levels of high-sensitivity C-reactive protein (hsCRP; p < 0.001) and serum amyloid A (SAA; p < 0.01) relative to controls. Antioxidative activity and weight % chemical composition of small, dense HDL did not differ between RA patients and controls (p > 0.05), whereas HDL phosphosphingolipidome was significantly altered in RA. Subgroup analyses revealed that RA patients featuring high levels of inflammation (hsCRP>10 mg/l) possessed small, dense HDL with reduced antioxidative activities (p < 0.01). Furthermore, antioxidative activity of HDL was inversely correlated with plasma hsCRP (p < 0.01). Conclusions: These data revealed that (i) despite normolipidemic state, the lipidome of small, dense HDL was altered in RA and (ii) high levels of inflammation can be responsible for the functional deficiency of small, dense HDL in RA.

Small, Dense High-Density Lipoprotein-3 Particles Are Enriched in Negatively Charged Phospholipids

Objective— High-density lipoprotein (HDL) displays multiple atheroprotective activities and is highly heterogeneous in structure, composition, and function; the molecular determinants of atheroprotective functions of HDL are incompletely understood. Because phospholipids represent a major bioactive lipid component of HDL, we characterized the phosphosphingolipidome of major normolipidemic HDL subpopulations and related it to HDL functionality. Approach and Results— Using an original liquid chromatography–mass spectrometry/mass spectrometry methodology for phospholipid and sphingolipid profiling, 162 individual molecular lipid species were quantified across the 9 lipid subclasses, in the order of decreasing abundance, phosphatidylcholine&gt;sphingomyelin&gt;lysophosphatidylcholine&gt;phosphatidylethanolamine&gt;phosphatidylinositol&gt;ceramide&gt;phosphatidylserine&gt;phosphatidylglycerol&gt;phosphatidic acid. When data were expressed relative to total lipid, the contents of lysophosphatidylcholine and of negatively charged phosphatidylserine and phosphatidic acid increased progressively with increase in hydrated density of HDL, whereas the proportions of sphingomyelin and ceramide decreased. Key biological activities of HDL subpopulations, notably cholesterol efflux capacity from human THP-1 macrophages, antioxidative activity toward low-density lipoprotein oxidation, antithrombotic activity in human platelets, cell-free anti-inflammatory activity, and antiapoptotic activity in endothelial cells, were predominantly associated with small, dense, protein-rich HDL3. The biological activities of HDL particles were strongly intercorrelated, exhibiting significant correlations with multiple components of the HDL phosphosphingolipidome. Specifically, the content of phosphatidylserine revealed positive correlations with all metrics of HDL functionality, reflecting enrichment of phosphatidylserine in small, dense HDL3. Conclusions— Our structure–function analysis thereby reveals that the HDL lipidome may strongly affect atheroprotective functionality.

Lipoprotein lipase in aortic valve stenosis is associated with lipid retention and remodelling

Calcific aortic valve disease (CAVD) is a chronic disorder characterized by a fibrocalcific remodelling. It is suspected that lipid retention within the aortic valve may be one important mechanism participating to aortic valve remodelling. Lipoprotein lipase (LPL) is implicated in lipid metabolism and may play a role in lipid retention within the aortic valve. In 57 patients, CAVD were analysed for the expression of LPL by q-PCR and immunohistochemistry. Expression of oxidized-LDL (ox-LDL) and decorin was also documented. In addition, a complete blood profile, including the size of LDL and high-density lipoprotein (HDL) particles, were performed to find associations between the blood lipid profile and expression of ox-LDL and LPL within CAVD. Immunohistochemistry studies revealed that LPL was expressed in stenotic aortic valves as a diffuse staining and also in dense cellular areas where macrophages were abundant. Expression of LPL co-localized with decorin and ox-LDL. In turn, valves with higher amount of ox-LDL had elevated number of LPL transcripts. In addition, we documented that the small, dense HDL phenotype was associated with an elevated amount of ox-LDL and LPL transcripts within CAVD. Furthermore, expression of LPL was associated with several indices of fibrocalcific remodelling of the aortic valve. Expression of LPL within CAVD is related to the amount of ox-LDL, which is, in turn, associated with the small, dense HDL phenotype. Lipid retention associated with smaller HDL particles may participate in the expression of LPL, whereby a fibrocalcific remodelling of the aortic valve is promoted.

Associations between Small Dense LDL, HDL Subfractions (HDL2, HDL3) and Risk of Atherosclerosis in Japanese-Americans

Small dense low-density lipoprotein (sdLDL) has been suggested to be more atherogenic than large buoyant LDL. High-density lipoprotein (HDL) consists of two major subfractions (HDL2, HDL3), and just as controversy remains regarding which of the two is the more powerful negative risk factor for atherosclerosis, associations between sdLDL and these HDL subfractions are unclear. We measured sdLDL cholesterol (sdLDL-C), HDL2 cholesterol (HDL2-C) and HDL3 cholesterol (HDL3-C) by a newly developed method in 481 Japanese-Americans who were not using lipid-lowering medication, and examined the associations of these cholesterol concentrations with variables related to atherosclerosis. In multivariate analysis, sdLDL-C was positively correlated with the body mass index (BMI), fasting glucose and insulin, 2-h glucose, HOMA-IR, high sensitivity C-reactive protein (hsCRP), and carotid artery intima-media thickness (IMT) after adjustment for age and sex. In particular, sdLDL-C was positively correlated with IMT, even after adjustment for sex, age, smoking status, hypertension, diabetes mellitus and hsCRP. HDL2-C was more closely inversely correlated than total HDL-C with BMI, fasting glucose and insulin, 2-h glucose, HOMA-IR, and hsCRP, whereas HDL3-C was not correlated with these factors. Additionally, HDL2-C was more closely correlated than total HDL-C or HDL3-C with sdLDL-C, LDL-C, triglycerides (TG), and apolipoprotein B (apoB). SdLDL-C was closely associated with insulin resistance and glucose tolerance, lending credence to its potential as a useful risk marker in assessing carotid artery IMT and the present degree of atherosclerosis in Japanese-Americans. The findings also suggest that subjects with higher HDL2-C levels were better protected from atherosclerosis.