Antiinflammatory Actions of HDL

The fact that a low level of high-density lipoprotein (HDL) cholesterol is highly predictive of future cardiovascular events has been established in population studies beyond all reasonable doubt. Furthermore, the evidence is overwhelming that the relationship is one of cause and effect rather than an epiphenomenon, with numerous studies in animals demonstrating that HDL-raising interventions translate into profound reductions in atherosclerosis. Such interventions have included the infusion of both native and reconstituted HDLs into rabbit models of atherosclerosis, the overexpression of apolipoprotein (apo) A-I (the major HDL protein) in transgenic mice and rabbits, and the inhibition of cholesteryl ester transfer protein (CETP) in rabbits. See pages 1325 and 1426 Evidence that raising the level of HDLs is also antiatherogenic in humans is mounting, although there are still relatively few human studies that have directly tested the phenomenon. Fibrates, statins, and niacin all raise the level of HDL cholesterol, and treatment with each of these agents has been shown to be associated with a reduction in future cardiovascular events. Fibrates are especially effective in event reduction in people with insulin resistance or with other features of the metabolic syndrome, although the benefits cannot be explained in terms of the observed HDL raising.1 Statins are effective in reducing events in all subjects, but the benefits can be explained almost completely in terms of the achieved LDL lowering2; the contribution of a statin-induced elevation of HDL is difficult to evaluate. Niacin, the most effective of the currently available HDL-raising agents, has also been shown to reduce cardiovascular events, especially when coadministered with a statin3; but again, other properties of niacin may …

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It is generally thought that the large heterogeneity of human HDL confers antiatherogenic properties; however, the mechanisms governing HDL biogenesis and speciation are complex and poorly understood. Here, we show that incubation of exogenous apolipoprotein A-I (apoA-I) with fibroblasts, CaCo-2, or CHO-overexpressing ABCA1 cells generates only alpha-nascent apolipoprotein A-I-containing particles (alpha-LpA-I) with diameters of 8-20 nm, whereas human umbilical vein endothelial cells and ABCA1 mutant (Q597R) cells were unable to form such particles. Interestingly, incubation of exogenous apoA-I with either HepG2 or macrophages generates both alpha-LpA-I and prebeta1-LpA-I. Furthermore, glyburide inhibits almost completely the formation of alpha-LpA-I but not prebeta1-LpA-I. Similarly, endogenously secreted HepG2 apoA-I was found to be associated with both prebeta1-LpA-I and alpha-LpA-I; by contrast, CaCo-2 cells secreted only alpha-LpA-I. To determine whether alpha-LpA-I generated by fibroblasts is a good substrate for LCAT, isolated alpha-LpA-I as well as reconstituted HDL [r(HDL)] was reacted with LCAT. Although both particles had similar V(max) (8.4 vs. 8.2 nmol cholesteryl ester/h/microg LCAT, respectively), the K(m) value was increased 2-fold for alpha-LpA-I compared with r(HDL) (1.2 vs. 0.7 microM apoA-I). These results demonstrate that 1) ABCA1 is required for the formation of alpha-LpA-I but not prebeta1-LpA-I; and 2) alpha-LpA-I interacts efficiently with LCAT. Thus, our study provides direct evidence for a new link between specific cell lines and the speciation of nascent HDL that occurs by both ABCA1-dependent and -independent pathways.

Epidemiological evidence strongly favors the notion that the risk of cardiovascular disease (CVD) is inversely related to the plasma high-density lipoprotein (HDL) cholesterol concentration.1 Low HDL cholesterol is still predictive of high CVD risk in subjects with low LDL cholesterol,2 as well as during statin treatment.3 These observational data and other studies, which show that HDL particles contain a large number of antioxidative, antiinflammatory, and antiproliferative proteins, underlie the generally held view that HDL particles have atheroprotective properties.1–5 However, evidence is accumulating supporting the concept that high HDL cholesterol levels do not always predict reduced CVD risk. The Incremental Decrease in End Points through Aggressive Lipid Lowering (IDEAL) trial and the European Prospective Investigation into Cancer and Nutrition (EPIC)-Norfolk case-control study revealed that (recurrent) CVD risk is not decreased in subjects with the highest HDL cholesterol and the greatest mean HDL particle size.6 More recently, a high HDL cholesterol, high C-reactive protein (CRP) subgroup of individuals at increased risk for a first cardiovascular event was identified in the community-dwelling Prevention of Renal and Vascular End-Stage Disease (PREVEND) cohort using the “outcome event mapping approach,” a graphical exploratory data analysis tool that has been originally developed by Corsetti et al.7 Applying this analytic method to the Thrombogenic Factors and Recurrent Coronary Events (THROMBO) postinfarction cohort, the presence of a subgroup …

Macrophages in the arterial intima take up modified LDL and become cholesterol-ester laden foam cells, which are the primary cell type in newly formed fatty streak lesions, and which play an important role throughout lesion progression and plaque vulnerability. Macrophages can unload their excess cellular cholesterol stores via lipid efflux, the first step in the protective reverse cholesterol transport pathway. In this pathway, efflux of cellular cholesterol to extracellular acceptors, such as HDL and lipid-poor apolipoproteins, targets this cholesterol for delivery to the liver for metabolism and direct excretion into the bile. The mechanisms of cellular cholesterol efflux are the focus of intensive research, and this field was advanced greatly by the discovery of ABCA1 as the Tangier disease gene in 1999.1 Tangier disease subjects have almost no plasma HDL, and their cells have a complete deficiency in cholesterol and phospholipid efflux to apolipoprotein A-I (apoAI) and a partial defect in lipid efflux to HDL.2 Thus, it was apparent that there is more than one pathway for lipid efflux to HDL. In 2001, Schmitz and colleagues demonstrated that ABCG1, another member of the ABC gene superfamily, was upregulated by cholesterol in human macrophages, and that an ABCG1 antisense oligonucleotide that reduced ABCG1 expression also decreased lipid efflux to HDL.3 The role of ABCG1 in lipid efflux was confirmed in 2004 when Wang and colleagues and Edwards and colleagues showed that ABCG1 transfected cells had increased cholesterol efflux to HDL.4,5 See page 1310 In the current issue of Arteriosclerosis, Thrombosis, and Vascular Biology , Wang et al provide novel insights both into the regulation of ABCG1 activity and into aspects of the mechanism of ABCG1-mediated lipid efflux,6 which differentiate it from the mechanism of ABCA1-mediated lipid efflux. Their essential discovery is that treatment of cells with …

Endothelial lipase (EL) is a triglyceride lipase gene family member that has high phospholipase and low triglyceride lipase activity. The aim of this study was to determine whether the phospholipase activity of EL is sufficient to remodel HDLs into small particles and mediate the dissociation of apolipoprotein A-I (apoA-I). Spherical, reconstituted HDLs (rHDLs) containing apoA-I only [(A-I)rHDLs], apoA-II only [(A-II)rHDLs], or both apoA-I and apoA-II [(A-I/A-II) rHDLs] were prepared. The rHDLs, which contained only cholesteryl esters in their core and POPC on the surface, were incubated with EL. As the rHDLs did not contain triacylglycerol, only the POPC was hydrolyzed. Hydrolysis was greater in the (A-I/A-II)rHDLs than in the (A-I)rHDLs. The (A-II)rHDL phospholipids were not hydrolyzed by EL. EL remodeled the (A-I)rHDLs and (A-I/A-II)rHDLs, but not the (A-II)rHDLs, into smaller particles. The reduction in particle size was related to the amount of phospholipid hydrolysis, with the diameter of the (A-I/A-II)rHDLs decreasing more than that of the (A-I)rHDLs. These changes did not affect the conformation of apoA-I, and neither apoA-I nor apoA-II dissociated from the rHDLs. Comparable results were obtained when human plasma HDLs were incubated with EL. These results establish that the phospholipase activity of EL remodels plasma HDLs and rHDLs into smaller particles without mediating the dissociation of apolipoproteins.

The last decade has witnessed an explosion in studies of the role of lipoproteins in brain function. Neurons require a continuous supply of lipids for membrane synthesis and acetylcholine production. Indeed, the brain is a site of intense lipid turnover—even though the central nervous system (CNS) accounts for only 2.1% of body weight, it contains 23% of total body cholesterol.1 Lipid metabolism in the brain is tightly controlled locally, as plasma lipoproteins are shielded from the brain by the blood-brain barrier. Although neuronal cells are capable of de novo synthesis of a wide spectrum of molecular species of lipids, they rely heavily on exogenous sources and readily bind and internalize lipoproteins of the extracellular fluid.2 Equally, neurons need to dispose of excess lipids; lipoprotein-mediated lipid transport is therefore bidirectional and includes cellular efflux of cholesterol.3 See accompanying article on page 1556 Human cerebrospinal fluid (CSF) primarily contains spherical lipoproteins of approximately 10 to 12 nm in diameter with hydrated density in the range 1.063 to 1.25 g/mL, thereby resembling HDL in human plasma.3,4⇓ Lipid concentrations in CSF are however much lower (eg, 300- to 400-fold for total cholesterol and phospholipids) as compared to the plasma compartment.4 Apolipoproteins (apo) E and A-I are the major apolipoproteins in human CSF (typical concentration range: 0.1 to 0.4 mg/dL3,4⇓), with apoA-II, A-IV, J, D, C-II, C-III, C-IV, and H equally present.3,4⇓ Importantly, CSF lipoproteins carry amyloid-β (Aβ), a 39- to 43-aa peptide produced in neuronal cells, which is the major component of senile amyloid plaques.5 The metabolism of CSF lipoproteins remains poorly characterized but seems to be distinct from that of plasma lipoproteins. ApoE-rich HDL are synthesized locally in CNS and secreted by astrocytes as discoidal complexes enriched in free cholesterol.6 High …

Human plasma HDLs are classified on the basis of apolipoprotein composition into those that contain apolipoprotein A-I (apoA-I) without apoA-II [(A-I)HDL] and those containing apoA-I and apoA-II [(A-I/A-II)HDL]. ApoA-I enters the plasma as a component of discoidal particles, which are remodeled into spherical (A-I)HDL by LCAT. ApoA-II is secreted into the plasma either in the lipid-free form or as a component of discoidal high density lipoproteins containing apoA-II without apoA-I [(A-II)HDL]. As discoidal (A-II)HDL are poor substrates for LCAT, they are not converted into spherical (A-II)HDL. This study investigates the fate of apoA-II when it enters the plasma. Lipid-free apoA-II and apoA-II-containing discoidal reconstituted HDL [(A-II)rHDL] were injected intravenously into New Zealand White rabbits, a species that is deficient in apoA-II. In both cases, the apoA-II was rapidly and quantitatively incorporated into spherical (A-I)HDL to form spherical (A-I/A-II)HDL. These particles were comparable in size and composition to the (A-I/A-II)HDL in human plasma. Injection of lipid-free apoA-II and discoidal (A-II)rHDL was also accompanied by triglyceride enrichment of the endogenous (A-I)HDL and VLDL as well as the newly formed (A-I/A-II)HDL. We conclude that, irrespective of the form in which apoA-II enters the plasma, it is rapidly incorporated into spherical HDLs that also contain apoA-I to form (A-I/A-II)HDL.

See related article, pages 598–606 An elevated plasma level of homocysteine has long been known as an independent predictor of cardiovascular disease.1,2 However, in the absence of a clear mechanism linking homocysteine to cardiovascular disease, there has been an ongoing debate about whether this relationship is one of cause and effect or whether an elevated level of plasma homocysteine is an epiphenomenon, reflecting the presence of some other proatherogenic factor that is actually responsible for the cardiovascular disease. In the current issue of this journal, Liao et al3 suggest that the mechanistic link may be a homocysteine-induced reduction in the concentration of high density lipoproteins (HDLs). Liao et al3 report that homocysteine reduces the concentration of HDL cholesterol in plasma by inhibiting the hepatic synthesis of apoA-I, the main HDL apolipoprotein. This conclusion supports the findings from another recent study by Mikael et al4 who also reported that homocysteine inhibits apoA-I synthesis. The results of these 2 studies not only explain the documented inverse correlation between the plasma concentrations of HDL cholesterol and homocysteine5,6 but also raise the real possibility that a homocysteine-induced inhibition of apoA-I synthesis is the mechanism linking homocysteine to the development of atherosclerosis. A low concentration of HDL cholesterol has been shown in numerous human population studies to be highly predictive of premature cardiovascular disease.7,8 Furthermore, treatments that increase the level of HDL cholesterol in plasma in both animals9,10 and humans11,12 reduce the progression or even promote …

This study was designed to establish the mechanism responsible for the increased apolipoprotein (apo) A-II levels caused by the cholesteryl ester transfer protein inhibitor torcetrapib. Nineteen subjects with low HDL cholesterol (<40 mg/dl), nine of whom were also treated with 20 mg of atorvastatin daily, received placebo for 4 weeks, followed by 120 mg of torcetrapib daily for the next 4 weeks. Six subjects in the nonatorvastatin cohort participated in a third phase, in which they received 120 mg of torcetrapib twice daily for 4 weeks. At the end of each phase, subjects underwent a primed-constant infusion of [5,5,5-(2)H(3)]L-leucine to determine the kinetics of HDL apoA-II. Relative to placebo, torcetrapib significantly increased apoA-II concentrations by reducing HDL apoA-II catabolism in the atorvastatin (-9.4%, P < 0.003) and nonatorvastatin once- (-9.9%, P = 0.02) and twice- (-13.2%, P = 0.02) daily cohorts. Torcetrapib significantly increased the amount of apoA-II in the alpha-2-migrating subpopulation of HDL when given as monotherapy (27%, P < 0.02; 57%, P < 0.003) or on a background of atorvastatin (28%, P < 0.01). In contrast, torcetrapib reduced concentrations of apoA-II in alpha-3-migrating HDL, with mean reductions of -14% (P = 0.23), -18% (P < 0.02), and -18% (P < 0.01) noted during the atorvastatin and nonatorvastatin 120 mg once- and twice-daily phases, respectively. Our findings indicate that CETP inhibition increases plasma concentrations of apoA-II by delaying HDL apoA-II catabolism and significantly alters the remodeling of apoA-II-containing HDL subpopulations.

Low serum levels of high-density lipoprotein (HDL) are commonly encountered in patients with coronary artery disease (CAD). An example of this type of patient is a 42-year-old white man with a history of sudden-onset angina secondary to a 90% obstructive lesion along the proximal left anterior descending coronary artery. The family history was significant for his father, who died of a myocardial infarction (MI) at age 44 years. The patient underwent percutaneous transluminal angioplasty with stenting but developed in-stent restenosis. He underwent cutting balloon angioplasty and brachytherapy and was asymptomatic for approximately 6 months. The stent then developed a high-grade occlusion with recurrence of angina, and the patient required single-vessel bypass surgery. The patient’s baseline serum lipid profile revealed low-density lipoprotein (LDL) 128 mg/dL, HDL 27 mg/dL, and triglycerides 92 mg/dL. His lipoprotein(a), C-reactive protein, and homocysteine levels were normal. He was not hypertensive, had no impairment of glycemic control, and did not smoke. With a combination of simvastatin 40 mg and niacin (Niaspan; Kos Pharmaceuticals) 1000 mg daily, the patient’s lipid profile improved, with LDL 78 mg/dL, HDL 43 mg/dL, and triglycerides 60 mg/dL. Follow-up stress testing demonstrated normal myocardial perfusion, and the patient has been asymptomatic for 2 years. With few exceptions, low HDL is an independent risk factor for CAD in case-control and prospective observational studies. In contrast, high HDL levels are associated with longevity and are protective against the development of atherosclerotic disease. In the Framingham Study, risk for CAD increases sharply as HDL levels fall progressively below 40 mg/dL.1 In the Quebec Cardiovascular Study, for every 10% reduction in HDL, risk for CAD increased 13%.2 Many clinicians believe that low HDL is associated with increased CAD risk because it is a marker for hypertriglyceridemia and elevated remnant particle concentrations. The Prospective Cardiovascular Munster …

The prevailing concept of mechanisms responsible for the development of atherosclerotic lesions largely focuses on the accumulation and retention of low-density lipoproteins in the arterial intima and their subsequent oxidative modification. This oxidation leads to activation of the endothelium, and particularly, expression of adhesion molecules that mediate leukocyte adherence and chemokines which initiate the inflammation reaction that is widely accepted as being responsible for the development and progression of atherosclerotic lesions.1,2 There is also a strong body of evidence to indicate that elevated triacylglycerides (triglycerides) are an independent risk factor for atherosclerosis.3–5 Article p 731 One mechanism that can contribute to elevated triglycerides involves apolipoprotein CIII (apoCIII). apoCIII is a small protein that resides on the surface of very-low-density lipoproteins (VLDLs), low-density lipoproteins, chylomicrons, and high-density lipoproteins (Figure). It exists as multiple species, as either a nonglycosylated isoform (apoCIIIo) or a glycosylated isoform (apoCIII1 or apoCIII2); all three isoforms have similar plasma half-lives and probably have very similar physiological functions. Increased apoCIII production is a characteristic feature of patients with hypertriglyceridemia,6 and plasma apoCIII levels have been positively correlated with plasma triacylglycerol concentrations and also have been associated with severity of hypertriglyceridemia.7 Elevated plasma apoCIII concentration and, specifically, accumulation of apoCIII in triacylglycerol-rich lipoproteins is casually related to hypertriglyceridemia in patients with metabolic syndrome and has also been associated with insulin resistance.8 apoCIII is a major regulator of lipolysis, as it noncompetitively inhibits endothelial-bound lipoprotein lipase, the enzyme that hydrolyzes triacylglycerols in …

Plasma high density lipoproteins (HDLs) from humans, from transgenic mice to human apolipoprotein A-I (HuAITg mice), from transgenic mice to human apolipoprotein A-II (HuAIITg mice), from transgenic mice to human apolipoproteins A-I and A-II (HuAIAIITg mice), and from C57BL/6 control mice were isolated, and their ability to interact with the human cholesteryl ester transfer protein (CETP) was studied. Whereas cholesteryl ester transfer rates were gradually enhanced by the addition of moderate amounts of HDL from the different sources, striking differences appeared when HDL levels kept increasing beyond a maximal transfer value. Indeed, while a plateau value corresponding to maximal CETP activity was maintained when raising the concentration of HuAITg HDL and HuAIAIITg HDL, inhibitions could be observed with the highest levels of human, control mouse, and HuAIITg mouse HDL. The concentration-dependent inhibition of CETP activity could be reproduced by the addition of delipidated HDL apolipoproteins from control mice, but it was abolished by a 1-h preheating treatment at 56 degrees C. In contrast, no significant inhibition of CETP activity was observed with the delipidated protein moiety of HuAITg HDL, and cholesteryl ester transfer rates remained unchanged before and after a 1-h, 56 degrees C preheating step. Finally, the CETP-mediated transfer of radiolabeled cholesteryl esters from human low density lipoprotein to human HDL was significantly higher in the presence of lipoprotein-deficient plasma from HuAITg mice than in the presence of lipoprotein-deficient plasma from control mice. Interestingly, cholesteryl ester transfer rates measured with both control and HuAITg lipoprotein-deficient plasmas became remarkably similar following a 1-h, 56 degrees C preheating treatment. It is concluded that human, control mouse, and HuAIITg mouse HDL contain a heat-labile lipid transfer inhibitory activity that is absent from HDL of HuAITg and HuAIAIITg mice. Alterations in CETP-lipoprotein binding did not account for differential lipid transfer inhibitory activities.

We characterized the molecular mechanisms by which high density lipoprotein (HDL) inhibits the expression of adhesion molecules, including vascular cell adhesion molecule-1 and intercellular adhesion molecule-1, induced by sphingosine 1-phosphate (S1P) and tumor necrosis factor (TNF) alpha in endothelial cells. HDL inhibited S1P-induced nuclear factor kappaB activation and adhesion molecule expression in human umbilical vein endothelial cells. The inhibitory HDL actions were associated with nitric-oxide synthase (NOS) activation and were reversed by inhibitors for phosphatidylinositol 3-kinase and NOS. The HDL-induced inhibitory actions were also attenuated by the down-regulation of scavenger receptor class B type I (SR-BI) and its associated protein PDZK1. When TNFalpha was used as a stimulant, the HDL-induced NOS activation and the inhibitory action on adhesion molecule expression were, in part, attenuated by the down-regulation of the expression of S1P receptors, especially S1P(1), in addition to SR-BI. Reconstituted HDL composed mainly of apolipoprotein A-I and phosphatidylcholine mimicked the SR-BI-sensitive part of HDL-induced actions. Down-regulation of S1P(3) receptors severely suppressed the stimulatory actions of S1P. Although G(i/o) proteins may play roles in either stimulatory or inhibitory S1P actions, as judged from pertussis toxin sensitivity, the coupling of S1P(3) receptors to G(12/13) proteins may be critical to distinguish the stimulatory pathways from the inhibitory ones. In conclusion, even though S1P alone stimulates adhesion molecule expression, HDL overcomes S1P(3) receptor-mediated stimulatory actions through SR-BI/PDZK1-mediated signaling pathways involving phosphatidylinositol 3-kinase and NOS. In addition, the S1P component of HDL plays a role in the inhibition of TNFalpha-induced actions through S1P receptors, especially S1P(1).

Endothelial nitric oxide synthase (eNOS) is one of three NOS isoforms that catalyze the formation of nitric oxide (NO) and L-citrulline by the oxidation of the guanido-nitrogen group of L-arginine. The cardiovascular importance of this reaction relies on the formation of NO, a signaling molecule that regulates vascular tone, platelet aggregation, oxidative stress, leukocyte adherence, and smooth muscle cell mitogenesis.1 Peroxisome proliferator-activated receptors (PPARs) are a subfamily of the nuclear receptor family of transcription factors that control the expression of key genes involved in the regulation of metabolism, inflammation, and thrombosis.2 Transcriptional control involves ligand activation followed by either heterodimerization with a retinoid X receptor and binding to the promoter region of target genes, or a DNA-binding independent mechanism that interferes negatively with proinflammatory factor pathways. Of the three PPAR isoforms (α, β/δ, and γ), PPAR-α is expressed chiefly in fatty acid-oxidizing tissues including liver, skeletal muscle, and heart, but also in endothelial and vascular smooth muscle cells and macrophages within the arterial wall. Despite a plethora of basic research demonstrating that PPAR-α activation by synthetic ligands (eg, fibrates) has favorable antiatherogenic properties,2 the corresponding effects on eNOS and the biology of NO has surprisingly not yet been explored. See page 658 In this issue of Arteriosclerosis, Thrombosis, and Vascular Biology , Goya et al3 demonstrate for the first time that specific PPAR-α agonists, such as fenofibrate, regulate eNOS in cultured endothelial cells. Using classical molecular biology techniques and bovine aortic endothelial cells as a model, fenofibrate was shown to increase the mRNA expression, protein level, and enzyme activity of eNOS in a dose-dependent manner at concentrations within the range of its EC50 value for human PPARα. …