HDL Function Research Articles

HDL Structure.

HDL has various protein components, including enzymes, complement components, apolipoproteins, protease inhibitors, etc. In addition to proteins, lipids are also a significant component of HDL. These components and their structure determine the function of HDL. HDL is heavily involved in the acute response phase, complement regulation phase, hemostasis phase, immune response phase, and protease inhibition phase. Among the apolipoproteins, the predominant component is Apo A-I, which confers various atherogenic activities to HDL. Apo A-II, Apo-C, Apo-D, Apo-F, Apo-H, Apo-J, and Apo-O, which can bind free fatty acids, regulate the activity of many proteins involved in HDL metabolism, inhibit lipid transfer, and control the endogenous coagulation cascade. A major functional component is the enzyme LCAT, which helps catalyze the conversion of cholesterol to plasma-based lipoproteins and then to cholesteryl esters. Another enzyme associated with HDL is human paraoxonase, calcium-, PON1-, PON2-, and PON3-dependent lactone enzyme with catalytic activity, including reversible binding to substrates. PAF-AH is a phospholipase with lipoprotein properties, and HDL and LDL particles are commonly bound to plasma PAF-AH for circulation. As for lipid components, PC is an essential phospholipid subclass and may be a biomarker for constitutive inflammation. Sphingolipids, such as sphingomyelin and ceramide, also play an indispensable role in HDL function. In different physiological and pathological stages and plasma environments, HDL can exhibit different structural features, such as discoid HDL and spherical rHDL.

HDL and Lipid Metabolism.

Mediating reverse cholesterol transport (RCT) is the most classic function of HDL. HDL and HDL-C participate in the entire process of RCT, including cholesterol removal from cells, cholesterol transport in circulation, and cholesterol excretion. As cholesterol is a component of lipid rafts and lipid droplets in cells, HDL and RCT can influence cell activity. HDL has also been shown to be related to the metabolism of some other biological lipids, such as S1P and ox-PL. Here we will introduce in detail the molecular mechanism of HDL participation in RCT and its significance.

High Density Lipoprotein as a Biomarker, Potential Therapeutic Target and Therapy

HDL is a plasma lipoprotein with the function of reverse cholesterol transport and inflammation modulation. Studies suggest that HDL can function as both anti inflammatory and proinflammatory depending on absence or presence of inflammation and chronic disease. Functions of HDL may not always match its cholesterol levels. LDL on the other hand is one of the main lipoproteins in plasma which is in charge of extracellular lipid transportation.

Intestine-specific ablation of the Hepatocyte Nuclear Factor 4a (Hnf4a) gene in mice has minimal impact on serum lipids and ileum gene expression profile due to upregulation of its paralog Hnf4g

Ablation of the gene encoding the nuclear receptor Hepatocyte Nuclear Factor 4a (Hnf4a) in the liver strongly affects HDL concentration, structure and functionality but the role of this receptor in the intestine, the second organ contributing to serum HDL levels, has been overlooked. In the present study we show that mice with intestine-specific ablation of Hnf4a (H4IntKO) had undetectable levels of ΗΝF4A in ileum, proximal and distal colon but normal expression in liver. H4IntKO mice presented normal serum lipid levels, HDL-C and particle size (α1-α3). The expression of the major HDL biogenesis genes Apoa1, Abca1, Lcat was not affected but there was significant increase in Apoc3 as well as in Hnf4g, a paralog of Hnf4a. RNA-sequencing identified metabolic pathways significantly affected by Hnf4a ablation such as type II diabetes, glycolysis, gluconeogenesis and p53 signaling. Chromatin immunoprecipitation assays showed that HNF4G bound to various apolipoprotein gene promoters in control mice but its binding affinity was reduced in the ileum of H4IntKO mice suggesting a redundancy but also a cooperation between the two factors. In the distal colon of H4IntKO mice, where both HNF4A and HNF4G are absent and in a mouse model of DSS-induced colitis presenting decreased levels of HNF4A, most lipoprotein genes were strongly downregulated. In conclusion, Hnf4a ablation in mice does not significantly affect serum lipid levels or lipoprotein gene expression in ileum possibly due to compensatory effects by its paralog Hnf4g in this tissue.

Abstract 9976: New Risk Factors for Improved Identification of Coronary Artery Disease Risk in Type 2 Diabetes by Substituting Paraoxonase 1 Activity for HDL Cholesterol in Atherogenic Ratios

Introduction: HDL cholesterol (HDL-C) may not always represent the HDL function. This study was conducted to help predict the risk of coronary artery disease (CAD) in patients with type 2 diabetes (T2D) based on substituting paraoxonase 1 (PON1) as an important enzyme in HDL function for that of HDL-C in the atherogenic ratios LDL-C/HDL-C and log (TG/HDL-C). Methods: A total of 274 subjects undergoing diagnostic coronary angiography in this study; 92 without significant CAD and 182 with severe CAD. The hydrolysis rate of phenylacetate as a PON1 substrate was spectrophotometrically measured in kinetic mode. The area under the ROC curve (AUC) was calculated to quantify the overall discriminative capacity of the variables. Results: ROC analyses showed that LDL-C/PON1 (AUC=0.66, p=0.02) and log (TG/PON1) (AUC=0.65, p=0.019) had higher predictive powers compared with LDL-C/HDL-C (AUC=0.55, p=0.221) and log (TG/HDL-C) (AUC=0.57, p=0.121) for CAD characterization in T2D patients. Also, AUCs of LDL-C/PON1 (AUC=0.74, p=0.014) and log (TG/PON1) (AUC=0.67, p=0.057) were higher than those of LDL-C/HDL-C (AUC=0.51, p=0.469) and log (TG/HDL-C) (AUC=0.53, p=0.364) for predicting CAD in T2D patients with HbA1c ≥7%. The multiple regression analysis with adjustments for gender, age, BMI, duration of T2D, metformin and statin therapy indicated that LDL-C/PON1 [adjusted OR=1.1 (0.99,1.23), p=0.085] and log (TG/PON1) [adjusted OR=3.76 (0.94,14.89), p=0.058] were independently predictors of CAD in T2D patients. Conclusions: New cardiometabolic biomarkers using PON1 provide an incremental predictive value compared with those using HDL-C in identifying subjects at increased risk for CAD among patients with T2D and poor glycemic control. This study sought to exploit the lipoprotein-related biomarkers of CAD from a more effective perspective in T2D. Our findings may help develop clinically cost-effective and functional laboratory measurements to evaluate HDL function.

A lifestyle intervention with an energy-restricted Mediterranean diet and physical activity enhances HDL function: a substudy of the PREDIMED-Plus randomized controlled trial

Consumption of a Mediterranean diet, adequate levels of physical activity, and energy-restricted lifestyle interventions have been individually associated with improvements in HDL functions. Evidence of intensive interventions with calorie restriction and physical activity is, however, scarce. To determine whether an intensive lifestyle intervention with an energy-restricted Mediterranean diet plus physical activity enhanced HDL function compared to a non-hypocaloric Mediterranean eating pattern without physical activity. In 391 older adults with metabolic syndrome (mean age, 65 years; mean BMI, 33.3kg/m2) from 1 of the Prevención con Dieta Mediterránea-Plus trial centers, we evaluated the impact of a 6-month intervention with an energy-restricted Mediterranean diet plus physical activity (intensive lifestyle; n=190) relative to a nonrestrictive Mediterranean diet without physical activity (control; n=201) on a set of HDL functional traits. These included cholesterol efflux capacity, HDL oxidative/inflammatory index, HDL oxidation, and levels of complement component 3, serum amyloid A, sphingosine-1-phosphate, triglycerides, and apolipoproteins A-I, A-IV, C-III, and E in apoB-depleted plasma. The intensive-lifestyle intervention participants displayed greater 6-month weight reductions (-3.83kg; 95% CI: -4.57 to -3.09 kg) but no changes in HDL cholesterol compared with control-diet participants. Regarding HDL functional traits, the intensive lifestyle decreased triglyceride levels (-0.15mg/g protein; 95% CI: -0.29 to -0.014 mg/g protein) and apoC-III (-0.11mg/g protein; 95% CI: -0.18 to -0.026 mg/g protein) compared to the control diet, with weight loss being the essential mediator (proportions of mediation were 77.4% and 72.1% for triglycerides and apoC-III levels in HDL, respectively). In older adults with metabolic syndrome, an energy-restricted Mediterranean diet plus physical activity improved the HDL triglyceride metabolism compared with a nonrestrictive Mediterranean diet without physical activity. This trial is registered at isrctn.com as ISRCTN89898870.

Biological Basis and Proposed Mechanism of Action of CSL112 (Apolipoprotein A-I [Human]) for Prevention of Major Adverse Cardiovascular Events in Patients post Myocardial Infarction

Despite current standard of care treatment, the period shortly after acute myocardial infarction (AMI) is associated with particularly high residual cardiovascular (CV) risk, with high rates of recurrent AMI and CV death in the first 90 days following the index event. This represents an area of high unmet need which may be potentially addressed by the recent shift in focus away from raising levels of high-density lipoprotein cholesterol (HDL-C), to ones that optimize HDL function. Apolipoprotein A-I (apoA-I) is the major protein constituent of HDL and a key mediator of cholesterol efflux from macrophages within atherosclerotic plaque, a property which may convey therapeutic efficacy during the high-risk period. CSL112 is a novel formulation of human plasma-derived apolipoprotein A-I (apoA-I), reconstituted with phosphatidylcholine and stabilized with sucrose that is currently being evaluated in a phase 3 clinical trial (AEGIS-II) for the reduction of major adverse CV events (MACE) in the period of high-risk post AMI. Patients with diabetes mellitus are especially at high risk of recurrent MI, and are an enriched patient population in the AEGIS-II study. In this presentation, we provide an overview of the biological properties of CSL112 that contribute to its proposed mechanism of action for reducing rates of MACE post AMI. These properties include rapid and robust promotion of cholesterol efflux from cells abundant in atherosclerotic plaque, in addition to anti-inflammatory effects, which together, may have a stabilizing effect on atherosclerotic plaque. We provide a detailed overview of these mechanisms, in addition to information on the composition of CSL112 and how it is manufactured.

Children with familial hypercholesterolemia display changes in LDL and HDL function: A cross-sectional study.

The functional status of lipoprotein particles contributes to atherogenesis. The tendency of plasma low-density lipoprotein (LDL) particles to aggregate and the ability of igh-density lipoprotein (HDL) particles to induce and mediate reverse cholesterol transport associate with high and low risk for cardiovascular disease in adult patients, respectively. However, it is unknown whether children with familial hypercholesterolemia (FH) display lipoprotein function alterations. We hypothesized that FH children had disrupted lipoprotein functions. We analyzed LDL aggregation susceptibility and HDL-apoA-I exchange (HAE), and activity of four proteins that regulate lipoprotein metabolism (cholesteryl ester transfer protein, lecithin-cholesterol acyltransferase, phospholipid transfer protein, and paraoxonase-1) in plasma samples derived from children with FH (n = 47) and from normocholesterolemic children (n = 56). Variation in lipoprotein functions was further explored using an nuclear magnetic resonance-based metabolomics profiling approach. LDL aggregation was higher, and HAE was lower in FH children than in normocholesterolemic children. LDL aggregation associated positively with LDL cholesterol (LDL-C) and negatively with triglycerides, and HAE/apoA-I associated negatively with LDL-C. Generally, the metabolomic profile for LDL aggregation was opposite of that of HAE/apoA-I. FH children displayed increased atherogenicity of LDL and disrupted HDL function. These newly observed functional alterations in LDL and HDL add further understanding of the risk for atherosclerotic cardiovascular disease in FH children.

Cholesterol efflux capacity and its association with prevalent metabolic syndrome in a multi-ethnic population (Dallas Heart Study)

Metabolic syndrome (MetS) is characterized by adiposity and atherogenic dyslipidemia consisting of elevated triglyceride and decreased high density lipoprotein cholesterol (HDL-C) levels however, cholesterol concentration alone does not reflect HDL functionality. Cholesterol efflux capacity (CEC) captures a key anti-atherosclerotic function of HDL; studies linking CEC to MetS have yielded inconsistent findings and lacked racial/ethnic diversity. The aim of this study was to evaluate the association between CEC and MetS in a large multi-ethnic population utilizing two different CEC assays interrogating overlapping but distinct reverse cholesterol transport pathways. A cross-sectional study was performed using the Dallas Heart Study cohort and cholesterol efflux was measured with radiolabeled and fluorescent cholesterol assays. The relationship between CEC and MetS was assessed using multivariable regression analyses. A total of 2241 participants were included (mean age was 50 years; 38% men and 53% Blacks). CEC was independently and inversely associated with MetS irrespective of efflux assay (CEC-radiolabeled, adjusted OR 0·71 [95% CI 0·65–0·80]. CEC-fluorescent, adjusted OR 0·85 [95% CI 0·77–0·94]). Both CEC measures were inversely associated with waist circumference and directly associated with HDL-C but not with other MetS components. There was an interaction by sex but not by race such that the inverse associations between CEC and MetS were somewhat attenuated in men (OR 0·86, 95%CI 0·74–1·01). In this large multi-ethnic cohort, impaired CEC is linked to MetS irrespective of efflux assay and race/ethnicity but less so among men. Future studies are needed to assess whether CEC mediates the atherosclerotic cardiovascular disease risk of MetS.

The Effect of Curcumin on HDL Cholesterol Uptake Capacity in Obese Individuals: A Pilot Study

Introduction: Obesity is a common risk factor associated with cardiovascular disease (CVD) risk. Curcumin has been reported to exert beneficial effects on lipid metabolism, including HDL functionality. We have examined the effects of curcumin on HDL cholesterol uptake capacity in subjects with obesity.Materials and Methods: 30 obese individuals received curcumin and placebo 1 g per day for a period of 30 days. The subjects were crossed over to the alternative regimen after a 2-week washout period. A modified cholesterol uptake capacity (CUC) assay was used to determine serum HDL functionality.Results: The study groups had similar base line characteristics. We did not find significant effects (p>0.05) of curcumin on serum HDL CUC levels.Conclusion: Curcumin administration at a dose of 1 g per day for 30 days did not affect HDL CUC in subjects with obesity.

Breakfast partly restores the anti-inflammatory function of high-density lipoproteins from patients with type 2 diabetes mellitus

Background and aimsHigh-density lipoproteins (HDL) of patients with type 2 diabetes mellitus (T2DM) have impaired anti-inflammatory activities. The anti-inflammatory activity of HDL has been determined ex vivo after isolation by different methods from blood mostly obtained after overnight fasting. We first determined the effect of the HDL isolation method, and subsequently the effect of food intake on the anti-inflammatory function of HDL from T2DM patients. MethodsBlood was collected from healthy controls and T2DM patients after an overnight fast, and from T2DM patients 3 h after breakfast (n = 17 each). HDL was isolated by a two-step density gradient ultracentrifugation in iodixanol (HDLDGUC2), by sequential salt density flotation (HDLSEQ) or by PEG precipitation (HDLPEG). The anti-inflammatory function of HDL was determined by the reduction of the TNFα-induced expression of VCAM-1 in human coronary artery endothelial cells (HCAEC) and retinal endothelial cells (REC). ResultsHDL isolated by the three different methods from healthy controls inhibited TNFα-induced VCAM-1 expression in HCAEC. With apoA-I at 0.7 μM, HDLDGUC2 and HDLSEQ were similarly effective (16% versus 14% reduction; n = 3; p > 0.05) but less effective than HDLPEG (28%, p < 0.05). Since ultracentrifugation removes most of the unbound plasma proteins, we used HDLDGUC2 for further experiments. With apoA-I at 3.2 μM, HDL from fasting healthy controls and T2DM patients reduced TNFα-induced VCAM-1 expression in HCAEC by 58 ± 13% and 51 ± 20%, respectively (p = 0.35), and in REC by 42 ± 13% and 25 ± 18%, respectively (p < 0.05). Compared to preprandial HDL, postprandial HDL from T2DM patients reduced VCAM-1 expression by 56 ± 16% (paired test: p < 0.001) in HCAEC and by 34 ± 13% (paired test: p < 0.05) in REC. ConclusionsThe ex vivo anti-inflammatory activity of HDL is affected by the HDL isolation method. Two-step ultracentrifugation in an iodixanol gradient is a suitable method for HDL isolation when testing HDL anti-inflammatory function. The anti-inflammatory activity of HDL from overnight fasted T2DM patients is significantly impaired in REC but not in HCAEC. The anti-inflammatory function of HDL is partly restored by food intake.

HDL subfraction distribution in hypertensive patients with high plasma HDL-cholesterol levels

Background and Aims: Low HDL-Cholesterol (HDL-C) levels are associated with increased risk for atherosclerotic cardiovascular disease (ASCVD). However, many studies have indicated that a residual ASCVD risk exists even at high plasma HDL-C levels suggesting that other factors may affect the reverse association between HDL-C levels, including the HDL subfraction profile and HDL functionality and the existence of other ASCVD risk factors.We evaluated the plasma HDL subfraction profile in hypertensivepatients comparedtonormotensive subjects with high HDL-C levels.

A way to engage HDL functionality by substituting paraoxonase 1 activity for HDL cholesterol in the commonly used atherogenic ratios: new risk factors for improved identification of coronary artery disease risk

Background and Aims: HDL cholesterol (HDL-C) may not always represent the HDL function. The present study was conducted to help predict the risk of coronary artery disease (CAD) by investigating novel cardiometabolic biomarkers based on substituting paraoxonase 1 (PON1) as a crucial enzyme in the functionality of HDL for that of HDL-C.

Children with familial hypercholesterolemia display changes in LDL and HDL function: A Cross-sectional study

Background and Aims: The functional status of lipoprotein particles contributes to atherogenesis. The tendency of plasma LDL particles to aggregate and the ability of HDL particles to induce and mediate reverse cholesterol transport associate with high and low risk for cardiovascular disease in adult patients, respectively. However, it is unknown whether children with familial hypercholesterolemia (FH) display lipoprotein function alterations. We hypothesized that FH children had disrupted lipoprotein function.

Functional interaction between apolipoproteins A2 and E in the triglyceride rich lipoprotein metabolism: Is LDLR involved?

Background and Aims: APOA2 is the second most abundant protein of HDL and plays crucial role in HDL synthesis and function. Previous unpublished data from our laboratory show that hepatic APOA2 production, in the absence of functional APOE, leads to reduced cholesterol and triglycerides levels, mainly due to VLDL deposition and significantly reduced rate of VLDL triglyceride production. Since APOE is the main ligand of LDLr, we hypothesized that, APOA2 may lead to clearance of TRLs through direct binding with LDLr.

Cholesterol efflux capacity is associated with inflammation level and monocyte-platelet aggregates in healthy men.

Background and Aims: It is well established that HDL particles have antiatherogenic properties. However, there are conflicting evidence in regards to HDL particle function and its association with various CVD risk factors. The aim of our study was to analyze associations between the marker of HDL function - cholesterol efflux capacity (CEC) and traditional (age, serum lipids, CRP) and emerging (monocyte-platelet aggregates (MPAs)) CVD risk markers.

HDL in Atherosclerotic Cardiovascular Disease: In Search of a Role.

For a long time, high-density lipoprotein cholesterol (HDL-C) has been regarded as a cardiovascular disease (CVD) protective factor. Recently, several epidemiological studies, while confirming low plasma levels of HDL-C as an established predictive biomarker for atherosclerotic CVD, indicated that not only people at the lowest levels but also those with high HDL-C levels are at increased risk of cardiovascular (CV) mortality. This “U-shaped” association has further fueled the discussion on the pathophysiological role of HDL in CVD. In fact, genetic studies, Mendelian randomization approaches, and clinical trials have challenged the notion of HDL-C levels being causally linked to CVD protection, independent of the cholesterol content in low-density lipoproteins (LDL-C). These findings have prompted a reconsideration of the biological functions of HDL that can be summarized with the word “HDL functionality”, a term that embraces the many reported biological activities beyond the so-called reverse cholesterol transport, to explain this lack of correlation between HDL levels and CVD. All these aspects are summarized and critically discussed in this review, in an attempt to provide a background scenario for the “HDL story”, a lipoprotein still in search of a role.

Differential redox proteomic profiles of serum from severe asthma patients after one month of benralizumab and mepolizumab treatment

Mepolizumab and Benralizumab are biological drugs for severe asthma patients able to reduce moderate-to-severe exacerbation rate (peripheral eosinophilial % mepolizumab 1.6 ± 1.2; benralizumab 0; p < 0.0001), improving the quality of life and lung function parameters (FEV1%: mepolizumab 87.1 ± 21.5; benralizumab 89.7 ± 15, p < 0.04). Here we report a preliminary redox proteomic study highlighting the level of oxidative burst present in serum from patients before and after one month of both treatments.Our results highlighted apolipoprotein A1 oxidation after Mepolizumab treatment, that could be related to HDL functionality and could represent a potential biomarker for the treatment. On the other hand, after one month of Benralizumab we detected higher oxidation levels of ceruloplasmin and transthyretin, considered an important oxidative stress biomarker which action help to maintain redox homeostasis.

HDL and type 2 diabetes: the chicken or the egg?

HDL is a complex macromolecular cluster of various components, such as apolipoproteins, enzymes and lipids. Quality evidence from clinical and epidemiological studies led to the principle that HDL-cholesterol (HDL-C) levels are inversely correlated with the risk of CHD. Nevertheless, the failure of many cholesteryl ester transfer protein inhibitors to protect against CVD casts doubts on this principle and highlights the fact that HDL functionality, as dictated by its proteome and lipidome, also plays an important role in protecting against metabolic disorders. Recent data indicate that HDL-C levels and HDL particle functionality are correlated with the pathogenesis and prognosis of type 2 diabetes mellitus, a major risk factor for CVD. Hyperglycaemia leads to reduced HDL-C levels and deteriorated HDL functionality, via various alterations in HDL particles' proteome and lipidome. In turn, reduced HDL-C levels and impaired HDL functionality impact the performance of key organs related to glucose homeostasis, such as pancreas and skeletal muscles. Interestingly, different structural alterations in HDL correlate with distinct metabolic abnormalities, as indicated by recent data evaluating the role of apolipoprotein A1 and lecithin-cholesterol acyltransferase deficiency in glucose homeostasis. While it is becoming evident that not all HDL disturbances are causatively associated with the development and progression of type 2 diabetes, a bidirectional correlation between these two conditions exists, leading to a perpetual self-feeding cycle.

The role of abnormalities of lipoproteins and HDL functionality in small fibre dysfunction in people with severe obesity

Obesity and associated dyslipidemia may contribute to increased cardiovascular disease. Obesity has also been associated with neuropathy. We have investigated presence of peripheral nerve damage in patients with severe obesity without type 2 diabetes and the status of metabolic syndrome and lipoprotein abnormalities. 47participants with severe obesity and 30 age-matched healthy controls underwent detailed phenotyping of neuropathy and an assessment of lipoproteins and HDL-functionality. Participants with severe obesity had a higher neuropathy symptom profile, lower sural and peroneal nerve amplitudes, abnormal thermal thresholds, heart rate variability with deep breathing and corneal nerve parameters compared to healthy controls. Circulating apolipoprotein A1 (P = 0.009), HDL cholesterol (HDL-C) (P < 0.0001), cholesterol efflux (P = 0.002) and paroxonase-1 (PON-1) activity (P < 0.0001) were lower, and serum amyloid A (SAA) (P < 0.0001) was higher in participants with obesity compared to controls. Obese participants with small nerve fibre damage had higher serum triglycerides (P = 0.02), lower PON-1 activity (P = 0.002) and higher prevalence of metabolic syndrome (58% vs. 23%, P = 0.02) compared to those without. However, HDL-C (P = 0.8), cholesterol efflux (P = 0.08), apoA1 (P = 0.8) and SAA (P = 0.8) did not differ significantly between obese participants with and without small nerve fibre damage. Small nerve fibre damage occurs in people with severe obesity. Patients with obesity have deranged lipoproteins and compromised HDL functionality compared to controls. Obese patients with evidence of small nerve fibre damage, compared to those without, had significantly higher serum triglycerides, lower PON-1 activity and a higher prevalence of metabolic syndrome.