Dysfunctional HDL Research Articles

활동적 저항성 운동이 비만유도 노화 흰쥐 심장근의 apolipoprotein A-1 및 염증발현에 미치는 영향

The purpose of this study was to analyze the changes of dysfunctional HDL structure and inflammatory factors caused by toll like receptor 4 (TLR4) protein expression in order to enhance heart function for cardiovascular disease prevention. The study groups were consisted of 40 Wistar old male rats at 50 weeks, and were divided into 4 groups as a control group (Con; n=10), control exercise group (Con+Ex; n=10), high fat diet group (HFD; n=10), and high fat diet exercise group (HFD+Ex; n=10). Dynamic resistance exercise training program included of ladder-climbing exercise and treadmill exercise, 3 days per week, for 12 weeks. The expression of aplipoprotein A-1 (ApoA-I) and TLR4 p rotein was analyzed by western blot. Also, plasma ApoA-I and C-reaction protein (CRP) concentration was analyzed by ELISA. The body weight before sampling of HFD-Ex group was significantly (p.05) lower than HFD group. ApoA-I protein expression and plasma ApoA-I concentration in heart muscle were not significantly different between all groups. However, TLR4 protein expression and plasma CRP levels in heart muscle were significantly (p.05) decreased in the HFD-Ex group compared to the HFD group. Therefore, dynamic resistance exercise increases heart muscle function and reduces inflammatory cytokine secretion. However, additional protein analysis related to the function of dysfunctional HDL is needed and further studies will be needed.

Rare P376L variant in the SR-BI gene associates with HDL dysfunction and risk of cardiovascular disease

BackgroundScavenger receptor class B type 1 (SR-BI) encoded by SCARB1 gene serves as a multifunctional HDL receptor, facilitating the uptake of cholesteryl esters from HDL to the liver. Recent studies have identified the association between the P376L missense mutation of the SCARB1 gene with increased serum HDL-Cholesterol level. However, the contribution of this variant to the development of cardiovascular disease (CVD) remains unclear. ObjectiveWe have investigated the association between the P376L polymorphism with the properties of HDL and CVD outcomes in a population sample recruited as part of the Mashhad-Stroke and Heart-Atherosclerotic-Disorders (MASHAD) cohort. MethodsSix hundred and fifteen individuals who had a median follow-up period of 7 years were recruited as part of the MASHAD cohort. Anthropometric, biochemical parameters and HDL lipid peroxidation (HDLox) were assessed. Genotyping was performed using TaqMan-real-time-PCR based method. The association of P376L-rs74830766 with cardiovascular-risk-factors and CVD events were evaluated. ResultsCarriers of the P376L variant were significantly more likely than non-carriers to develop CVD using multivariate analyses adjusted for traditional CVD risk factors defined as: age, sex, BMI, presence of diabetes, or hypertension, positive smoking habit, and total cholesterol (OR: 3.75, 95%CI: 1.76–7.98, p = 0.001). In an adjusted model, there was a two fold increase in cardiovascular endpoints among individuals who were heterozygous for the P376L variant (hazard ratio, 2.08; 95% CI, 1.12-to 3.84, p = 0.02). Although there was no association between the presence of the P376L variant and HDL-C level, serum HDLox, measured as dysfunctional HDL, was 13% higher among carriers of the P376L variant than non-carriers. ConclusionWe have found that carriers of the P376L variant possessed higher HDLox and were at increased risk of CVD in a representative population-based cohort, as compared to non-carriers.

HDL Dysfunction Caused by Mutations in apoA-I and Other Genes that are Critical for HDL Biogenesis and Remodeling.

The "HDL hypothesis" which suggested that an elevation in HDL cholesterol (HDL-C) levels by drugs or by life style changes should be paralleled by a decrease in the risk for Cardiovascular Disease (CVD) has been challenged by recent epidemiological and clinical studies using HDL-raising drugs. HDL components such as proteins, lipids or small RNA molecules, but not cholesterol itself, possess various atheroprotective functions in different cell types and accumulating evidence supports the new hypothesis that HDL functionality is more important than HDL-C levels for CVD risk prediction. Thus, the detailed characterization of changes in HDL composition and functions in various pathogenic conditions is critically important in order to identify new biomarkers for diagnosis, prognosis and therapy monitoring of CVD. Here we provide an overview of how HDL composition, size and functionality are affected in patients with monogenic disorders of HDL metabolism due to mutations in genes that participate in the biogenesis and the remodeling of HDL. We also review the findings from various mouse models with genetic disturbances in the HDL biogenesis pathway that have been generated for the validation of the data obtained in human patients and how these models could be utilized for the evaluation of novel therapeutic strategies such as the use of adenovirus-mediated gene transfer technology that aim to correct HDL abnormalities.

Elevated high-density lipoprotein in adolescents with Type 1 diabetes is associated with endothelial dysfunction in the presence of systemic inflammation.

AimsHigh-density lipoprotein (HDL) function may be altered in patients with chronic disease, transforming the particle from a beneficial vasoprotective molecule to a noxious pro-inflammatory equivalent. Adolescents with Type 1 diabetes often have elevated HDL, but its vasoprotective properties and relationship to endothelial function have not been assessed.Methods and resultsSeventy adolescents with Type 1 diabetes (age 10–17 years) and 30 age-matched healthy controls supplied urine samples for the measurement of early renal dysfunction (albumin:creatinine ratio; ACR), blood samples for the assessment of cardiovascular risk factors (lipid profiles, HDL functionality, glycaemic control, and inflammatory risk score), and had their conduit artery endothelial function tested using flow-mediated dilation (FMD). HDL-c levels (1.69 ± 0.41 vs. 1.44 ± 0.29mmol/L; P < 0.001), and glycated haemoglobin (HbA1c) (8.4 ± 1.2 vs. 5.4 ± 0.2%; P < 0.001) were increased in all patients compared with controls. However, increased inflammation and HDL dysfunction were evident only in patients who also had evidence of early renal dysfunction (mean ± standard deviation for high-ACR vs. low-ACR and healthy controls: inflammatory risk score 11.3 ± 2.5 vs. 9.5 ± 2.4 and 9.2 ± 2.4, P < 0.01; HDL-mediated nitric-oxide bioavailability 38.0 ± 8.9 vs. 33.3 ± 7.3 and 25.0 ± 7.7%, P < 0.001; HDL-mediated superoxide production 3.71 ± 3.57 vs. 2.11 ± 3.49 and 1.91 ± 2.47nmol O2 per 250 000 cells, P < 0.05). Endothelial function (FMD) was impaired only in those who had both a high inflammatory risk score and high levels of HDL-c (P < 0.05).ConclusionIncreased levels of HDL-c commonly observed in individuals with Type 1 diabetes may be detrimental to endothelial function when accompanied by renal dysfunction and chronic inflammation.

Oxidized HDL, Adipokines, and Endothelial Dysfunction: A Potential Biomarker Profile for Cardiovascular Risk in Women with Obesity.

High BMI predicts adverse cardiovascular outcomes and positively correlates with increased levels of adipokines. The relationship among BMI, IL-6, TNFα, adiponectin, and oxidized high-density lipoprotein (Ox-HDL) with circulating endothelial cells (CECs) and endothelial progenitor cells (EPCs) has not been well studied. Elevated CEC levels have been described in both humans and mice with obesity and diabetes. Ox-HDL has been shown to be a potent driver of adipogenesis in vivo and in vitro. In this study, elevated BMI was examined in 2 groups of women studied in Brooklyn, New York, and Huntington, West Virginia, respectively. Twenty-six females with obesity and five lean controls without overt cardiovascular disease were enrolled, 13 from Huntington and 13 from Brooklyn. Cytokine levels, EPCs, and CECs were determined. Females with obesity had elevated levels of leptin, IL-6, and Ox-HDL, increased CEC levels, and decreased EPC and adiponectin levels (all P<0.01). The Ox-HDL levels were higher in women from Brooklyn versus Huntington (P<0.01), possibly from higher TNFα levels in Brooklyn or higher adiponectin levels in Huntington. Seventy-five percent of the variance in Ox-HDL levels could be predicted in this population (P<0.01). This study reveals a unique inflammatory biomarker profile in females with obesity.

Assessment of HDL Cholesterol Removal Capacity: Toward Clinical Application.

While there is a controversy regarding the causal relationship between high-density lipoprotein cholesterol (HDL-C) and cardiovascular disease (CVD), recent studies have demonstrated that the cholesterol efflux capacity (CEC) of HDL is associated with the incidence of CVD. However, there are several limitations to current assays of CEC. First, CEC measurements are not instantly applicable in clinical settings, because CEC assay methods require radiolabeled cholesterol and cultured cells, and these procedures are time consuming. Second, techniques to measure CEC are not standardized. Third, the condition of endogenous cholesterol donors would not be accounted for in the CEC assays. Recently, we established a simple, high-throughput, cell-free assay system to evaluate the capacity of HDL to accept additional cholesterol, which is herein referred to as “cholesterol uptake capacity (CUC)”. We demonstrated that CUC represents a residual cardiovascular risk in patients with optimal low-density lipoprotein cholesterol control independently of traditional risk factors, including HDL-C. Establishing reproducible approaches for the cholesterol removal capacity of HDL is required to validate the impact of dysfunctional HDL on cardiovascular risk stratification in the “real world”.

The combined utility of myeloperoxidase (MPO) and paraoxonase 1 (PON1) as two important HDL-associated enzymes in coronary artery disease: Which has a stronger predictive role?

Background and aimsSerum paraoxonase 1 (PON1) and myeloperoxidase (MPO) are HDL-associated enzymes that contribute significantly to the formation of dysfunctional HDL. The present study thus seeks to comparatively analyze the predictive role of PON1, MPO and the MPO/PON1 ratio and to also evaluate which one has a stronger predictive role in their combined utility as an MPO/PON1 ratio in coronary artery disease (CAD). MethodsPON1 activity and MPO concentrations were determined in patients with established CAD and those without significant CAD. Receiver operating characteristic (ROC) curves were drawn by plotting true positivity versus false positivity. ResultsThe ROC curve analyses showed that PON1 (AUC = 61%, p = 0.003) and MPO/PON1 (AUC = 60%, p = 0.01) have a better diagnostic performance than MPO (AUC = 50%, p = 0.42) in detecting patients with CAD. PON1 and MPO/PON1 were found to have a significantly stronger discriminatory power for the age range ≥52 and < 60 years (AUC = 69%, p = 0.008 for PON1; AUC = 66%, p = 0.022 for MPO/PON1). The multivariate analysis revealed PON1 as an independent variable that was significantly associated with the multi-vessel disease [odds ratio (OR) = 0.98; p = 0.017]. At the cutoff point of 30 μmol/mL/min for PON1 and 1.85 for MPO/PON1, specificities were 97% and 73% and sensitivities 30% and 54% for discriminating patients with single-vessel disease from non-CAD subjects. ConclusionsThe diagnostic performance of PON1 alone was comparable to that of the MPO/PON1 ratio for CAD risk assessment; however, MPO may increase the true positive rate. A larger number of blocked vessels seems to be associated with an increased predictive power for both PON1 and MPO/PON1. Recent data support the fact that PON1 and MPO may potentially be appropriate therapeutic targets for preventing CAD.

Potent anti-inflammatory properties of HDL in vascular smooth muscle cells mediated by HDL-S1P and their impairment in coronary artery disease due to lower HDL-S1P: a new aspect of HDL dysfunction and its therapy.

Dysfunctional HDL is associated with coronary artery disease (CAD), but its effect on inflammation in vascular smooth muscle cells (VSMCs) in atherosclerosis is unknown. We investigated the effect of healthy human HDL and CAD-HDL on TNF-α-driven inflammation in VSMCs and examined whether HDL-associated sphingosine-1-phosphate (HDL-S1P) could modulate inflammation with the aim of designing novel HDL-based anti-inflammatory strategies. Healthy human HDL, human CAD-HDL, and mouse HDL were isolated by ultracentrifugation, S1P was measured by liquid chromatography-tandem mass spectrometry, and TNF-α-induced inflammation was characterized by gene expression and analysis of NF-κB-dependent signaling. Mechanisms of S1P interference with TNF-α were assessed by S1P receptor antagonists, mouse knockouts, and short interfering RNA. We observed that healthy HDL potently inhibited the induction of TNF-α-stimulated inflammatory genes, such as iNOS (inducible NO synthase) and MMP9 (matrix metalloproteinase 9), a process that was entirely dependent on HDL-S1P, as evidenced by loss-of-function using S1P-less HDL and mimicked by genuine S1P. Inhibition was based on suppression of TNF-α-activated Akt signaling resulting in reduced IkBαSer32 and p65Ser534 NF-κB phosphorylation based on a persistent phosphatase and tensin homolog activation by S1P through the S1P receptor 2. Intriguingly, S1P suppressed inflammation even hours after initial TNF-α stimulation. The anti-inflammatory effect of healthy HDL correlated with HDL-S1P content and was superior to that of CAD-HDL featuring lower HDL-S1P. Nevertheless, therapeutic loading of HDL with S1P completely restored the anti-inflammatory capacity of CAD-HDL and greatly boosted that of both healthy and CAD-HDL. Suppression of inflammation by HDL-S1P defines a novel pathophysiologic characteristic that distinguishes functional from dysfunctional HDL. The anti-inflammatory HDL function can be boosted by S1P-loading and exploited by S1P receptor-targeting to prevent and even turn off ongoing inflammation.-Keul, P., Polzin, A., Kaiser, K., Gräler, M., Dannenberg, L., Daum, G., Heusch, G., Levkau, B. Potent anti-inflammatory properties of HDL in vascular smooth muscle cells mediated by HDL-S1P and their impairment in coronary artery disease due to lower HDL-S1P: a new aspect of HDL dysfunction and its therapy.

The High Density Lipoprotein Cholesterol Hypothesis Revisited

BACKGROUND: The strong inverse association of plasma levels of high-density lipoprotein cholesterol (HDL-C) with coronary heart disease (CHD) found in human epidemiological studies led to the development of the ‘HDL-C hypothesis’, which posits that intervention to raise HDL-C will result in reduced risk of CHD. However, recent evidence has raised doubts about the hypotheses that elevating HDL-C is necessarily therapeutic. Genetic variations that associate with altered HDL-C do not strongly associate with altered cardiovascular disease risk.CONTENT: HDL-mediated cholesterol efflux from macrophage foam cells or measurements of the flux of cholesterol from macrophages to the liver and feces seem to correlate better with atherosclerotic burden than with HDL-C levels. Thus, it may be time to modify the HDL-C hypothesis to the ‘HDL flux hypothesis’, where intervention to promote cholesterol efflux and reverse cholesterol transport will reduce CHD risk, regardless of whether it affects plasma HDL-C levels. A deeper understanding of the complex biology of HDL metabolism and its relationship to reverse cholesterol transport and atherothromobotic events is urgently needed. This might lead to biomarkers of HDL flux and functionality that are more informative than simple measurements of HDL-C levels.SUMMARY: It is now clear from recent clinical trial and genetic studies that some approaches to raising HDL-C levels may have no effect on CHD. This suggests the need to evaluate HDL-C-raising therapies in different clinical populations, as well as therapies targeted toward HDL flux and function rather than simply HDL-C elevation. Perhaps moving from a focus on the HDL-C hypothesis to a focus on the HDL flux hypothesis will permit a biologically based reassessment of the optimal therapeutic approach to targeting HDL for reduction in cardiovascular risk.KEYWORDS: reverse cholesterol transport, cholesterol efflux capacity, HDL dysfunction, HDL particle size, HDL lipidomics, HDL proteomics

Human ApoA-I overexpression ameliorates two main cardioprotective functions of HDL in db/db mice

Aim: The impact of ApoA-I, main protein of HDL, has been long examined. However, the impact of ApoA-I on atheroprotective action of dysfunctional HDL in diabetes is still limited. In this regard, we analyzed its consequences in important properties of HDL such as in vivo macrophage-specific reverse cholesterol transport (m-RCT) and ex vivo anti-oxidant protection in db/db mice

Glycation and Deamidation Result in HDL Dysfunction in Patients with Type 2 Diabetes

Diabetes is associated with HDL dysfunction and perturbed iron metabolism. ApoAI, transferrin (Trf) and ceruloplasmin (Cp) are the key HDL proteins involved in cholesterol efflux and iron metabolism. We tested the hypothesis that hyperglycemia-induced glycation contributes to HDL dysfunction and altered iron metabolism in diet-controlled patients with T2D. HDL from patients with T2D and matched healthy controls (n=9/group) was isolated and it’s anti-oxidant and cholesterol efflux properties were quantified. HDL proteome composition and post-translational modification of proteins were quantified by proteomics aproach. Metabolic 2H2O-labeling was applied to quantify HDL proteome dynamics. Patients with T2D and controls had similar lipid (triglycerides, total cholesterol, and HDL cholesterol) profile. HDL from T2D patients had reduced anti-oxidant (PON1 activity) and macrophage-cholesterol efflux capacity (P&amp;lt;0.05) and was enriched with glycated apoAI and transferrin (Tf), and deamidated ceruloplasmin (Cp), the key HDL proteins involved in cholesterol and iron transport. Both ApoAI and Trf glycation were directly correlated with HbA1c and the extent of glycations were associated with their increased degradation. ApoAI glycation was inversely correlated with cholesterol efflux activity of HDL. Trf glycations at Lys-2and Lys-534 sites involved in iron coordination, were 2-3 fold higher in T2D than in the controls. The PTM search on Cp revealed five asparagine deamidation sites in T2D patients with N-943 being located on one of the oxidation-prone motifs and may result in the loss of Cp’s ferroxidase activity. In vivo HDL flux study demonstrated that glycated apoAI and Tf, and deamidated Cp species were degraded 3, 10, and 2 fold faster than the respective non-modified native proteins. HDL dysfunction and oxidative stress in T2D is related to glycation- and deamidation-induced instability of HDL proteins, including ApoAI, Tf and Cp. Disclosure T. Kasumov: None. M. Golizeh: None. S. Kashyap: None.

Abstract 555: Investigation of 4-oxo-2-nonenal Mediated HDL Dysfunction and the Use of Lipid Dicarbonyl Scavengers in Preserving HDL Function

Background: Lipid peroxidation products such as hydroxy-2-nonenal (HNE) are elevated in atherosclerosis and cause HDL dysfunction. Generated in parallel to HNE is 4-oxo-2-nonenal (ONE), a less studied lipid dicarbonyl possibly due to its far greater reactivity. The consequences of ONE modification of HDL have not been studied. We have recently found that scavengers of lipid dicarbonyls such as salicylamine (SAM) and pentylpyridoxamine (PPM) prevent HDL dysfunction induced by malondialdehyde and isolevuglandin (IsoLG). In this study, we examine the impact of ONE adduct formation on HDL structure-function, and the scavenging abilities of various small molecules in preventing ONE modification. Methods and Results: By Western blot analysis, ONE crosslinked apoA-I on HDL at a concentration of 3 ONE molecules for every 10 apoA-I proteins (0.3 molar equivalence, eq.), which is 100 fold lower than HNE but comparable to IsoLG. ONE-mediated crosslinking of HDL proteins preferentially produced a 39 kDa band on SDS-PAGE, likely an apoA-I/apoA-II heterodimer. ONE-modified HDL partially inhibited the ability of HDL to protect against the inflammatory response of macrophages (as shown in TNFα and IL-1β mRNA expression), but did not render HDL pro-inflammatory. At 3 eq., ONE dramatically decreased the ability of apoA-I to exchange among HDLs, from ~46.5% to only ~18.4% (P&lt;0.001). HDL-mediated macrophage cholesterol efflux was decreased by ~70.6% (P&lt;0.005) and 56.1% (P&lt;0.001) by HDL modified by 0.3 and 3 eq. ONE, respectively. Investigation of various scavenger analogues in protecting HDL from ONE modification showed that while SAM and its fluoro- and chloro- analogues partially prevented ONE-mediated HDL protein crosslinking, PPM nearly completely blocked crosslinking. PPM pretreatment of HDL prior to 3 eq. ONE modification was also able to restore HDL-mediated macrophage cholesterol efflux, from 56.1~ to ~83.0% (P&lt;0.01) while the inactive analogue pentylpyridoxine had no effect. Conclusions: Our study is the first to show that ONE causes HDL dysfunction, and demonstrates that not all modified HDLs result in the same “dysfunction”. We also demonstrate the use of PPM in preferentially scavenging ONE in biological systems.

Abstract 551: Is Free Cholesterol Bioavailability a Determinant of Dysfunctional, Atherogenic High Density Lipoproteins? Refining the Model of Reverse Cholesterol Transport

Although cardiovascular disease (CVD) negatively correlates with high-density lipoprotein-cholesterol (HDL-C) levels, to date HDL-raising therapies have not reduced CVD; thus, in the context of reverse cholesterol transport (RCT), HDL quality, i.e., functionality, may be more important to atheroprotection than HDL quantity. The current RCT model comprises free cholesterol (FC) efflux from macrophages to apo AI to give nHDL, LCAT-mediated nHDL-FC esterification forming mature HDL, SR-B1-mediated hepatic uptake of HDL lipids, followed by sterol metabolism and excretion. Our recent studies challenge this model: We observed that nHDL apo AI, FC and phospholipid (PL) metabolically segregate. In mice, plasma nHDL FC and PL are hepatically cleared with t 1/2 ~3 min; nHDL-apo AI is cleared more slowly with t 1/2 = 460 min. FC esterification is 100X slower, and thus a minor RCT step. These results and the observation that nHDL is FC-rich (~64 mol%) led to a revised model of RCT, with a focus on FC bioavailabilty rather than CE uptake. HDL from SR-B1 -/- mice is also FC-rich and associated with atherosclerosis. Moreover, the magnitude of HDL-C content/particle is associated with carotid artery atheroprogression (Qi et al JACC 2015 65:355-363), and HDL from hyperlipidemic HIV patients, which are at increased CVD risk, also have a high mol% FC compared to controls (29 vs 16 mol%, p&lt;0.05). Thus, we hypothesize that high HDL-FC bioavailability, measured as the product of mol% HDL-FC and HDL particle number, is a metric for dysfunctional, atherogenic HDL. We studied SRB1 -/- mice, which are atherosusceptible and a model of dysfunctional HDL. Compared to WT mice, SR-B1 -/- mice have a higher HDL particle number and mol% FC (~58 vs. 15). Compared to WT HDL, SRB1 -/- HDL is more resistant to disruption by GdmHCl and serum opacity factor, indicating a resistance to remodeling. We plan to compare FC bioavailability of WT and SRB1 -/- HDL according to the kinetics of HDL-FC transfer to LDL. Our studies will determine if increased HDL-FC bioavailability in dysfunctional FC-rich HDL supports whole-body hypercholesterolemia that could increase CVD risk.

Abstract 394: Bacterial Serum Opacity Factor Rescues HDL Functionality in SR-B1 -/- Mice

Although plasma HDL-C levels negatively correlate with atherosclerotic cardiovascular disease (ACVD), attempts to reduce ACVD risk by raising plasma HDL have disappointed. Thus, hypotheses about salutary HDL effects have shifted from higher-is-better to function-is-more-important. The SR-B1 -/- mouse is an extreme model of HDL dsyfunctionality; compared to WT mice, SR-B1 -/- mice have higher plasma HDL levels and an HDL surface that is free cholesterol (FC)-rich (60 vs. 15 mol%). This would be expected to increase HDL-FC bioavailability to cytotoxic levels. HDL dysfunctionality among SR-B1 -/- mice is associated with multiple metabolic abnormalities—impaired cell membrane structure and function and atherosusceptibility, despite having high plasma HDL-C levels; moreover, female SR-B1 -/- mice are infertile. Liver-specific SR-B1 expression in SR-B1 -/- mice normalizes HDL size and FC content. Thus, the SR-B1 -/- mouse phenotype is due to lack of hepatic clearance of lipids from dysfunctional HDL. Serum opacity factor (SOF) is a bacterial protein that catalyzes the quantitative disproportionation of HDL into a cholesteryl ester-rich micro emulsion (CERM), neo HDL, and lipid-free apo AI. The CERM contains apo E and all HDL-CE. Injection of SOF (4 μg) into WT mice lowers plasma cholesterol by diverting the CERM to hepatic LDLR. Thus, we began testing whether adeno-viral delivery of SOF (AAV SOF ) to SR-B1 -/- mice rescues HDL functionality. A plasmid encoding the SOF gene was synthesized; SOF DNA was isolated by restriction enzyme digestion and cloned into a pAAV-TBG-mcs plasmid that was submitted to the PENN Vector Lab for virus production and isolation. AAV GFP plasmid (UPENN) was used as control. Good SOF production and secretion was confirmed by transfection of Huh7 hepatocytes. AAV SOF injection into SR-B1 -/- mice induced constitutive plasma SOF activity and reduced HDL-C levels to nil. Superposition of high plasma HDL levels and a high mol% FC in SR-B1 -/- is expected to increase HDL-FC bioavailablity that contributes to whole-body FC-toxicity and the observed metabolic abnormalities. Future tests will determine whether ablation of dysfunctional HDL in SR-B1 -/- mice rescues their pathological phenotype, especially atherosclerosis.

Abstract 035: Recombinant LCAT Restores Defective HDL mediated Endothelial Protection in Acute Coronary Syndrome

Recent evidence suggests that the vasoprotective effects of HDL are impaired in patients during an acute coronary syndrome (ACS). In particular, HDL from ACS patients are defective in promoting nitric oxide (NO) release from cultured endothelial cells (ECs). Lecithin:cholesterol acyltransferase (LCAT) plays a key role in HDL maturation and remodeling, and it is the enzyme responsible for cholesterol esterification in plasma. Very few studies have suggested that LCAT activity is decreased during an acute myocardial infarction, but the possible link between LCAT mass/activity and the HDL dysfunction observed in ACS has never been tested. To test this hypothesis, plasma from 30 STEMI patients was collected at admission, 48 and 72 hours and 30 days after event. LCAT concentration and activity significantly decreased 48 hours after event (-0.49±0.11 μg/ml, P&lt;0.001, and -3.15±1.05 nmol/ml/h, P=0.006) and remained reduced at 72 hours. HDL isolated from STEMI patients lose the capacity to promote NO production in ECs (-0.13 ± 0.06 fold, P=0.04) and the reduction is significantly correlated to decreased LCAT activity (R= 0.52, P=0.003). In vitro studies were performed in which STEMI patients’ plasma was added with rhLCAT and HDL vasoprotective activity assessed by measuring NO production in ECs. In vitro incubation of STEMI patients’ plasma with rhLCAT remodels phospholipids and protein content in HDL and restores HDL ability to promote endothelial NO production (+25%, P=0.03). Impairment of cholesterol esterification may be a major factor in the HDL dysfunction observed during ACS. rhLCAT is able to restore HDL-mediated NO production in vitro, suggesting LCAT as potential therapeutic target for restoring HDL functionality in ACS.

In vitro oxidized HDL and HDL from type 2 diabetes patients have reduced ability to efflux oxysterols from THP-1 macrophages

Oxidized LDL (OxLDL) that are enriched in products of lipid peroxidation including oxysterols have been shown to induce cellular oxidative stress and cytotoxicity therefore accelerating atheroma plaque formation. Upon oxLDL exposure of THP-1 macrophages, intracellular oxidation of LDL derived-cholesterol as well as endogenous cholesterol was increased. The oxysterols intracellularly produced were efficiently exported to HDL whereas apolipoprotein A1 was inefficient. These findings prompted us to investigate the consequences of modification of HDL by oxidation and glycation as observed in type 2 diabetes with respect to oxysterol and cholesterol efflux. We show that efflux of oxysterols was significantly impaired after in vitro oxidation and glycoxidation of HDL whereas glycation alone had no impact. Cholesterol efflux was only slightly decreased by oxHDL or glycoxidized HDL and not changed with glycated HDL. The defect of HDL towards oxysterol efflux was also observed with HDL isolated from diabetic subjects as compared to healthy controls. These findings support a deleterious cellular retention of oxysterols due to dysfunctional HDL in type 2 diabetes.

The impact of myeloperoxidase on HDL function in myocardial infarction.

The focus in cardiovascular research is shifting from determining mass HDL cholesterol levels toward investigating HDL functionalities as biomarker for cardiovascular disease. Myeloperoxidase (MPO), a main effector enzyme of the innate immune system, is increasingly implicated to negatively impact HDL function by various chemical modifications of HDL-associated proteins. This review summarizes recent insights how MPO affects HDL function in the setting of acute myocardial infarction (MI), mainly focusing on human data. First the mechanisms how MPO renders HDL particles dysfunctional and the usefulness of MPO as prospective biomarker for MI incidence and outcomes are described. Then the evidence for MPO causing specific HDL function impairments in MI and the clinical value of these observations is discussed in the context of the different HDL function assays employed. MPO modification of HDL in acute MI generates dysfunctional HDL. Features of HDL dysfunction can be used to stratify MI patients and seem associated with outcomes. More prospective studies are warranted to explore, if MPO-modified HDL is causally linked to severity and outcomes of MI. If this could be established, MPO would represent an attractive target to improve HDL dysfunction in MI and provide clinical benefit for patients.

High-Density Lipoprotein Function in Cardiovascular Disease and Diabetes Mellitus.

Cardiovascular disease (CVD) is the most common cause of death worldwide.1 Major CVD risk factors are hypertension, smoking, physical inactivity, abnormal glucose levels/diabetes mellitus, and dyslipidemia. Among these, dyslipidemia characterized by a low level of HDL-C (high-density lipoprotein cholesterol) is strongly and inversely correlated with CVD risk,2–4 and low HDL-C levels is part of the atherogenic dyslipidemia complex associated with diabetes mellitus.5 These observations triggered intense interest in increasing HDL-C levels for therapeutic intervention of CVD.6,7 However, several recent lines of evidence now suggest that the association between HDL-C levels and CVD status may be indirect or more complicated than previously recognized.7 Thus, human genetic studies demonstrate that genetically altered HDL-C levels do not necessarily translate to an altered risk of CVD.8–12 Phase III clinical trials of drugs that elevate HDL-C, such as niacin and CETP (cholesteryl ester transfer protein) inhibitors, also have largely failed to reduce CVD events in statin-treated subjects with established CVD.13–15 For several CETP inhibitors, improvement in CVD prevention was unsuccessful because of off-target effects (torcetrapib) or lack of efficacy (dalcetrapib and evacetrapib).14,16–18 The exception is the recent REVEAL trial (Randomized Evaluation of the Effects of Anacetrapib Through Lipid-Modification), which included more subjects and was performed for longer than previous CETP inhibitor trials. The REVEAL demonstrated that the CETP inhibitor anacetrapib exhibited beneficial effects on cardiovascular outcomes on top of those of statin therapy, although the risk reduction was moderate19,20 and might have been due, at least in part, to a reduction in non-HDL-C rather than elevated HDL-C.21 Considering these cumulative observations, it remains uncertain whether increased HDL-C directly impacts atherosclerosis and the …

Premature Coronary Heart Disease in South Asians: Burden and Determinants.

While the burden of cardiovascular disease (CVD) is on the decline globally, it is on the rise among South Asians. South Asians are also believed to present early with coronary artery disease (CAD) compared with other ethnicities. South Asians have demonstrated a higher burden of premature CAD (PCAD) compared with other ethnicities. These findings are not limited to non-immigrant South Asians but have also been found in immigrant South Asians settled around the world. In this article, we first discuss studies evaluating PCAD among South Asians residing in South Asia and among South Asian immigrants in other countries. We then discuss several traditional risk factors that could explain PCAD in South Asians (diabetes, hypertension, dietary factors, obesity) and lipoprotein-associated risk (low HDL-C levels, higher triglycerides, and elevated apolipoprotein B levels). We then discuss several emerging areas of research among South Asians including the role of dysfunctional HDL, elevated lipoprotein(a), genetics, and epigenetics. Although various risk markers and risk factors of CAD have been identified in South Asians, how they impact therapy is not well-known. PCAD is prevalent in the South Asian population. Large-scale studies are needed to identify how this information can be rationally utilized for early identification of risk among South Asians, and how currently available therapies can mitigate this increased risk.

Cell-free Biochemical Fluorometric Enzymatic Assay for High-throughput Measurement of Lipid Peroxidation in High Density Lipoprotein.

Low high-density lipoprotein cholesterol (HDL-C) levels are one of the most powerful independent negative predictors of atherosclerotic cardiovascular disease (CVD). The structure and function of HDL rather than HDL-C may more accurately predict atherosclerosis. Several HDL protein and lipid compositional changes that impair HDL function occur in inflammatory states such as atherosclerosis. HDL function is usually determined by cell based assays such as cholesterol efflux assay but these assays have numerous drawbacks lack of standardization. Cell-free assays may give more robust measures of HDL function compared to cell-based assays. HDL oxidation impairs HDL function. HDL has a major role in lipid peroxide transport and high amount of lipid peroxides is related to abnormal HDL function. Lipid-probe interactions should be considered when interpreting the results of non-enzymatic fluorescence assays for measuring the lipid oxidative state. This motivated us to develop a cell-free biochemical enzymatic method to assess HDL lipid peroxide content (HDLox) that contributes to HDL dysfunction. This method is based on the enzyme horseradish peroxidase (HRP) and the fluorochrome Amplex Red that can quantify (without cholesterol oxidase) the lipid peroxide content per mg of HDL-C. Here a protocol is describedfor determination of HDL-lipid peroxidation using the fluorochrome reagent. Assay variability can be reduced by strict standardization of experimental conditions. Higher HDLox values are associated with reduced HDL antioxidant function. The readout of this assay is associated with readouts of validated cell-based assays, surrogate measures of cardiovascular disease, systemic inflammation, immune dysfunction, and associated cardiovascular and metabolic risk phenotypes. This technical approach is a robust method to assess HDL function in human disease where systemic inflammation, oxidative stress and oxidized lipids have a key role (such as atherosclerosis).