High Density Lipoprotein Formation Research Articles

High-density lipoproteins and non-alcoholic fatty liver disease

Background and aimsNon-alcoholic fatty liver disease (NAFLD), a high incidence liver pathology, is associated with a ∼1.5-fold higher cardiovascular disease risk. This phenomenon is generally attributed to the NAFLD-associated increase in circulating levels of pro-atherogenic apolipoprotein B100-containing small dense low-density lipoprotein and plasma hypertriglyceridemia. However, also a significant reduction in cholesterol transported by anti-atherogenic high-density lipoproteins (HDL) is frequently observed in subjects suffering from NAFLD as compared to unaffected people. In this review, we summarize data regarding the relationship between NAFLD and plasma HDL-cholesterol levels, with a special focus on highlighting potential causality between the NAFLD pathology and changes in HDL metabolism. Methods and resultsPublications in PUBMED describing the relationship between HDL levels and NAFLD susceptibility and/or disease severity, either in human clinical settings or genetically-modified mouse models, were critically reviewed for subsequent inclusion in this manuscript. Furthermore, relevant literature describing effects on lipid loading in cultured hepatocytes of models with genetic alterations related to HDL metabolism have been summarized. ConclusionsAlthough in vitro observations suggest causality between HDL formation by hepatocytes and protection against NAFLD-like lipid accumulation, current literature remains inconclusive on whether relative HDL deficiency is actually driving the development of fatty liver disease in humans. In light of the current obesity pandemic and the associated marked rise in NAFLD incidence, it is of clear scientific and societal interest to gain further insight into the relationship between HDL-cholesterol levels and fatty liver development to potentially uncover the therapeutic potential of pharmacological HDL level and/or function modulation.

Hepatocyte ABCA1 deficiency is associated with reduced HDL sphingolipids.

ATP binding cassette transporter A1 (ABCA1) limits the formation of high density lipoproteins (HDL) as genetic loss of ABCA1 function causes virtual HDL deficiency in patients with Tangier disease. Mice with a hepatocyte-specific ABCA1 knockout (Abca1 HSKO) have 20% of wild type (WT) plasma HDL-cholesterol levels, suggesting a major contribution of hepatic ABCA1 to the HDL phenotype. Whether plasma sphingolipids are reduced in Tangier disease and to what extent hepatic ABCA1 contributes to plasma sphingolipid (SL) levels is unknown. Here, we report a drastic reduction of total SL levels in plasma of a Tangier patient with compound heterozygosity for mutations in ABCA1. Compared to mutation-free controls, heterozygous mutations in ABCA1 had no significant effect on total SLs in plasma; however, apoB-depleted plasma showed a reduction in total SL also in het carriers. Similarly, liver specific Abca1 KO mice (Abca1 HSKO) showed reduced total sphingolipids in plasma and liver. In parallel, apoM and sphingosine-1-phosphate (S1P) levels were reduced in plasma of Abca1 HSKO mice. Primary hepatocytes from Abca1 HSKO mice showed a modest, but significant reduction in total SLs concentration compared to WT hepatocytes, although SL de novo synthesis and secretion were slightly increased in Abca1 HSKO hepatocytes. We conclude that hepatic ABCA1 is a signficant contributor to maintaining total plasma pool of HDL sphingolipids, including sphingomyelins and S1P.

#5668 PERITONEAL DIALYSIS ADDITIVE REDUCES TRANSPERITONEAL LOSS OF BLOOD MICROPARTICLE AND EXTRACELLULAR MATRIX PROTEINS IN A CONTROLLED CROSSOVER TRIAL

Abstract Background and Aims About 11% of patients requiring dialysis worldwide are treated with peritoneal dialysis (PD). With each PD exchange high amounts of protein are removed from the peritoneal cavity stemming from the systemic circulation (reflecting toxin removal, loss of serum proteins, inflammatory processes) and local inflammatory processes. Thereby, the clinically measured peritoneal protein loss (PPL) is the result of transperitoneal serum loss, lymphatic drainage, transmesothelial reabsorption, local production, and cellular components. Although high PPL has been found to be predictive of mortality and cardiovascular morbidity, the pathophysiological concept is still poorly understood. Method Peritoneal dialysis effluent (PDE) samples were obtained from peritoneal equilibrium tests during a prospective, multicenter, double-blinded, controlled, randomized, dual-period, 2-treatment, crossover, phase II, proof-of-concept study in Austria. PDE samples were submitted to proteomic analysis by tandem mass tag (TMT) labeling and 2D reversed phase liquid chromatography mass spectrometry. To disentangle transmembrane serum protein loss from local protein production we developed a novel and unique analytical approach. Briefly, partitioning around medoids and proximity in the Euclidean space for between visit (non-treatment visits only) single-protein kinetics was utilized to achieve this separation. A multivariate mixed-effects model was calculated to identify treatment (intraperitoneal alanyl-glutamine administration) effects. Results We identified 2,624 different proteins within the PDE of 12 patients across two time-points. A large cluster of proteins with stable abundances in collinearity with well-established clinical characteristics of transperitoneal transport (e.g., dialysate-to-plasma-albumin) in accordance with the three-pore-model was identified. After clustering and separation, the proteins resembling similar time-course kinetics to clinical transperitoneal membrane transport characteristics (one of five clusters) were all identified to be able to theoretically traverse through the small peritoneal pore (< 4-6 nm). Of these 549 proteins lost from the systemic circulation via transperitoneal small pore diffusion, the appearance of 51 in the PD effluent was significantly reduced by intraperitoneal administration of alanyl-glutamine. The affected proteins are mainly involved in extracellular matrix organization, part of extracellular exosome signaling and blood microparticles, e.g. high density lipoprotein (HDL) formation (Apolipoprotein A1). Conclusion PPL is an important but poorly understood undesired effect of PD associated with cardiovascular morbidity and mortality. Our novel analytical approach enables us to disentangle the complexity of transperitoneal serum protein loss and local protein production on single-protein level across the peritoneal proteome. Furthermore, we were able to delineate the potential beneficial effects of intraperitoneal alanyl-glutamine treatment on PPL and peritoneal health which might even suggest potential beneficial effects on cardiovascular morbidity (e.g., HDL formation).

Activity and Structural Dynamics of Human ABCA1 in a Lipid Membrane

The human ATP-binding cassette (ABC) transporter ABCA1 plays a critical role in lipid homeostasis as it extracts sterols and phospholipids from the plasma membrane for excretion to the extracellular apolipoprotein A-I and subsequent formation of high-density lipoprotein (HDL) particles. Deleterious mutations of ABCA1 lead to sterol accumulation and are associated with atherosclerosis, poor cardiovascular outcomes, cancer, and Alzheimer’s disease. The mechanism by which ABCA1 drives lipid movement is poorly understood, and a unified platform to produce active ABCA1 protein for both functional and structural studies has been missing. In this work, we established a stable expression system for both a human cell-based sterol export assay and protein purification for in vitro biochemical and structural studies. ABCA1 produced in this system was active in sterol export and displayed enhanced ATPase activity after reconstitution into a lipid bilayer. Our single-particle cryo-EM study of ABCA1 in nanodiscs showed protein induced membrane curvature, revealed multiple distinct conformations, and generated a structure of nanodisc-embedded ABCA1 at 4.0-Å resolution representing a previously unknown conformation. Comparison of different ABCA1 structures and molecular dynamics simulations demonstrates both concerted domain movements and conformational variations within each domain. Taken together, our platform for producing and characterizing ABCA1 in a lipid membrane enabled us to gain important mechanistic and structural insights and paves the way for investigating modulators that target the functions of ABCA1.

Endocytosis of LXRs: Signaling in liver and disease.

Nuclear receptors are among one of the major transcriptional factors that induces gene regulation in the nucleus. Liver X receptor (LXR) is a transcription factor which regulates essential lipid homeostasis in the body including fatty acid, cholesterol and phospholipid synthesis. Liver X receptor-retinoid X receptor (LXR-RXR) heterodimer is activated by either of the ligand binding on LXR or RXR. The promoter region of the gene which is targeted by LXR is bound to the response element of LXR. The activators bind to the heterodimer once the corepressor is dissociated. The cellular process such as endocytosis aids in intracellular trafficking and endosomal formation in transportation of molecules for essential signaling within the cell. LXR isotypes play a crucial role in maintaining lipid homeostasis by regulating the level of cholesterol. In the liver, the deficiency of LXRα can alter the normal physiological conditions depicting the symptoms of various cardiovascular and liver diseases. LXR can degrade low density lipoprotein receptors (LDLR) by the signaling of LXR-IDOL through endocytic trafficking in lipoprotein uptake. Various gene expressions associated with cholesterol level and lipid synthesis are regulated by LXR transcription factor. With its known diversified ligand binding, LXR is capable of regulating expression of various specific genes responsible for the progression of autoimmune diseases. The agonists and antagonists of LXR stand to be an important factor in transcription of the ABC family, essential for high density lipoprotein (HDL) formation. Endocytosis and signaling mechanism of the LXR family is broad and complex despite their involvement in cellular growth and proliferation. Here in this chapter, we aimed to emphasize the master regulation of LXR activation, regulators, and their implications in various metabolic activities especially in lipid homeostasis. Furthermore, we also briefed the significant role of LXR endocytosis in T cell immune regulation and a variety of human diseases including cardiovascular and neuroadaptive.

Association of methylation status of ABCA1/G1 genes with the risk of coronary artery disease

BackgroundATP-binding cassette transporters A1/G1 (ABCA1/G1) is a main regulator of HDL (high-density lipoprotein) formation and reverse cholesterol transport. Impaired ABCA1/G1 genes function may seriously affect cholesterol homeostasis, leading to increased risk of cardiovascular disease. In the present study, the association of ABCA1/G1 genes methylation status with the risk of coronary artery disease (CAD), risk factors of CAD, and serum level of lipid parameters was investigated.This study was conducted on 70 CAD patients and 40 control subjects. All CAD subjects with diabetes mellitus were excluded. The promoter methylation status of ABCA1/G1 genes was determined by the methylation-specific polymerase chain reaction (MS-PCR) method and serum lipid parameters were assessed using commercial kits.ResultsABCA1 promoter methylation was higher in CAD group compared to the control participants (80% vs. 60%). Hypermethylation of the ABCA1 gene significantly increases the risk of CAD in the total population (OR 3.886, 95% CI (1.181–12.791), p = 0.026). ABCG1 methylation status showed no difference between CAD and control subjects. In addition, no significant association was noted between methylation status of ABCA1/G1 and serum level of lipid profile.ConclusionsAltogether, our study shows that ABCA1 gene promoter hypermethylation may increase the risk of CAD, which may help identify people at risk of developing CAD.

Specificity of ABCA7-mediated cell lipid efflux

Adenosine triphosphate-binding cassette transporter subfamily A member 7 (ABCA7) performs incompletely understood biochemical functions that affect pathogenesis of Alzheimer's disease. ABCA7 is most similar in primary structure to ABCA1, the protein that mediates cell lipid efflux and formation of high-density lipoprotein (HDL). Lipid metabolic labeling/tracer efflux assays were employed to investigate lipid efflux in BHK-ABCA7(low expression), BHK-ABCA7(high expression) and BHK-ABCA1 cells. Shotgun lipid mass spectrometry was used to determine lipid composition of HDL synthesized by BHK-ABCA7 and BHK-ABCA1 cells. BHK-ABCA7(low) cells exhibited significant efflux only of choline-phospholipid and phosphatidylinositol. BHK-ABCA7(high) cells had significant cholesterol and choline-phospholipid efflux to apolipoprotein (apo) A-I, apo E, the 18A peptide, HDL, plasma and cerebrospinal fluid and significant efflux of sphingosine-lipid, serine-lipid (which is composed of phosphatidylserine and phosphatidylethanolamine in BHK cells) and phosphatidylinositol to apo A-I. In efflux assays to apo A-I, after adjustment to choline-phospholipid, ABCA7-mediated efflux removed ~4 times more serine-lipid and phosphatidylinositol than ABCA1-mediated efflux, while ABCA1-mediated efflux removed ~3 times more cholesterol than ABCA7-mediated efflux. Shotgun lipidomic analysis revealed that ABCA7-HDL had ~20 mol% less phosphatidylcholine and 3–5 times more serine-lipid and phosphatidylinositol than ABCA1-HDL, while ABCA1-HDL contained only ~6 mol% (or ~1.1 times) more cholesterol than ABCA7-HDL. The discrepancy between the tracer efflux assays and shotgun lipidomics with respect to cholesterol may be explained by an underestimate of ABCA7-mediated cholesterol efflux in the former approach. Overall, these results suggest that ABCA7 lacks specificity for phosphatidylcholine and releases significantly but not dramatically less cholesterol in comparison with ABCA1.

Rationale for the use of hypo-hyperoxic training in patients with metabolic syndrome

Recently, the interest of scientists around the world in studying the relationship of metabolic disorders and obesity with the growth of cardiovascular diseases has increased significantly. Pharmacological medicine does not yet have effective drugs with which it is possible to solve the problem of metabolic syndrome in a complex, therefore, drugs are most often used to reduce cholesterol levels in the blood, blood pressure, appetite suppression to reduce body weight, etc. In recent years, the method of interval hypoxic and hypoxic-hyperoxic training has been very intensively researched and applied in clinical practice as a modern highly effective physical method of prevention and treatment of various diseases. It was found that with the course use of IGT, a complex of compensatory responses develops: optimization of the functioning of the sympatho-adrenal system; changes in the metabolism of lipids and lipoproteins due to the activation of key enzymes that catalyze the esterification of cholesterol and regulate the formation of high-density lipoproteins. Such a systemic response of the body is very important in people with insulin resistance, metabolic syndrome and impaired carbohydrate tolerance.

Pathways and Mechanisms of Cellular Cholesterol Efflux-Insight From Imaging.

Cholesterol is an essential molecule in cellular membranes, but too much cholesterol can be toxic. Therefore, mammalian cells have developed complex mechanisms to remove excess cholesterol. In this review article, we discuss what is known about such efflux pathways including a discussion of reverse cholesterol transport and formation of high-density lipoprotein, the function of ABC transporters and other sterol efflux proteins, and we highlight their role in human diseases. Attention is paid to the biophysical principles governing efflux of sterols from cells. We also discuss recent evidence for cholesterol efflux by the release of exosomes, microvesicles, and migrasomes. The role of the endo-lysosomal network, lipophagy, and selected lysosomal transporters, such as Niemann Pick type C proteins in cholesterol export from cells is elucidated. Since oxysterols are important regulators of cellular cholesterol efflux, their formation, trafficking, and secretion are described briefly. In addition to discussing results obtained with traditional biochemical methods, focus is on studies that use established and novel bioimaging approaches to obtain insight into cholesterol efflux pathways, including fluorescence and electron microscopy, atomic force microscopy, X-ray tomography as well as mass spectrometry imaging.

Apolipoprotein A-I Inhibits Transendothelial Transport of Apolipoprotein B-Carrying Lipoproteins and Enhances Its Associated High-Density Lipoprotein Formation.

Caveola-located scavenger receptor type B class I (SR-BI) and activin receptor-like kinase-1 (ALK1) are involved in transendothelial transport of apolipoprotein B-carrying lipoproteins (apoB-LPs). Transport of apoB-LPs though mouse aortic endothelial cells (MAECs) is associated with apoE-carrying high-density lipoprotein (HDL)-like particle formation and apoAI induces raft-located proteins to shift to non-raft membranes by upregulation of ATP-binding cassette transporter A1 (ABCA1). To investigate apoAI's effect on transendothelial transport of apoB-LPs, MAECs and human coronary artery endothelial cells (HCAECs) were treated with apoB-LPs ± apoAI. Our data demonstrated that apoAI neither altered SR-BI and ALK1 expression nor affected apoB-LP binding to MAECs. ApoAI inhibited MAEC uptake, transcellular transport, and intracellular accumulation of apoB-LPs and accelerated their resecretion in MAECs. ApoAI enhanced transendothelial apoB-LP transport-associated HDL-like particle formation, upregulated ABCA1 expression, shifted SR-BI and ALK1 to the non-raft membrane in MAECs, inhibited transcellular transport of apoB-LPs, and enhanced associated HDL-like particle formation in HCAECs. ABCA1 knockdown attenuated apoAI-induced membrane SR-BI and ALK1 relocation and diminished apoAI's effect on transendothelial apoB-LP transport and HDL-like particle formation in MAECs. This suggests that upregulation of ABCA1 expression is a mechanism, whereby apoAI provokes caveola-located receptor relocation, inhibits transendothelial apoB-LP transport, and promotes associated HDL-like particle formation.

AICAR upregulates ABCA1/ABCG1 expression in the retinal pigment epithelium and reduces Bruch's membrane lipid deposit in ApoE deficient mice

The etiology of age-related macular degeneration (AMD) is diverse; however, recent evidence suggests that the lipid metabolism-cholesterol pathway might be associated with the pathophysiology of AMD. The ATP-binding cassette (ABC) transporters, ABCA1 and ABCG1, are essential for the formation of high-density lipoprotein (HDL) and the regulation of macrophage cholesterol efflux. The failure of retinal or retinal pigment epithelium (RPE) cholesterol efflux to remove excess intracellular lipids causes morphological and functional damage to the retina. In this study, we investigated whether treatment with 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR), an AMP-activated protein kinase (AMPK) activator, improves RPE cholesterol efflux and Bruch's membrane (BM) lipid deposits. The protein and mRNA levels of ABCA1 and ABCG1 in ARPE-19 cells and retinal and RPE/choroid tissue from apolipoprotein E-deficient (ApoE-/-) mice were evaluated after 24 weeks of AICAR treatment. The cholesterol efflux capacity of ARPE-19 cells and the cholesterol-accepting capacity of apoB-depleted serum from mice were measured. The thickness of the BM and the degree of lipid deposition were evaluated using electron microscopy. AICAR treatment increased the phosphorylation of AMPK and the protein and mRNA expression of ABCA1 and ABCG1 in vitro. It promoted cholesterol efflux from ARPE-19 cells and upregulated the protein and mRNA levels of ABCA1 and ABCG1 in the retina and RPE in vivo. ApoB-depleted serum from the AICAR-treated group showed enhanced cholesterol-accepting capacity. Long-term treatment with AICAR reduced BM thickening and lipid deposition in ApoE-/- mice. In conclusion, AICAR treatment increased the expression of lipid transporters in the retina and RPE in vivo, facilitated intracellular cholesterol efflux from the RPE in vitro, and improved the functionality of HDL to accept cholesterol effluxed from the cell, possibly via AMPK activation. Collectively, these effects might contribute to the improvement of early age-related pathologic changes in the BM. Pharmacological improvement of RPE cholesterol efflux via AMPK activation may be a potential treatment strategy for AMD.

Current models of apolipoprotein A-I lipidation by adenosine triphosphate binding cassette transporter A1.

The primary cardioprotective function of high-density lipoprotein (HDL) is to remove excess cellular free cholesterol (FC) from peripheral tissues and deliver it to the liver. Here, we summarize recent research that examines apolipoprotein A-I (apoA-I) lipidation models by adenosine triphosphate binding cassette transporter A1 (ABCA1) and discuss its relevance in atherosclerotic cardiovascular disease (ASCVD). The first step in HDL formation involves the interaction between apoA-I and ABCA1, where ABCA1 mediates the removal of FC and phospholipids from lipid-laden macrophages to form discoidal nascent HDL (nHDL). However, there are currently no clear-cut systematic models that characterize HDL formation. A number of recent studies have investigated the importance of apoA-I C- and N-terminal domains required for optimal cholesterol efflux and nHDL production. Furthermore, functional ABCA1 is required for direct or indirect binding to apoA-I where ABCA1 dimer-monomer interconversion facilitates apoA-I lipidation from plasma membrane microdomains. Microparticles are also another lipid source for apoA-I solubilization into nHDL. ApoA-I and ABCA1 are key factors in macrophage-mediated cholesterol efflux and nHDL production. Understanding of the key steps in HDL formation may unlock the therapeutic potential of HDL and improve clinical management of ASCVD.

Circadian regulation of apolipoprotein gene expression affects testosterone production in mouse testis

The circadian clock system plays an important role in regulating testosterone synthesis in mammals. Male Bmal1−/− mice are infertile with low serum testosterone levels and decreased expression of testicular steroidogenic genes, suggesting that circadian clock genes regulate testosterone biosynthesis by activating steroidogenic gene transcription. However, whether the circadian clock regulates testosterone production via other genes remains unknown. Using Bmal1−/− mice and their wild-type (WT) siblings, we aimed to identify additional genes by which the circadian clock regulates testosterone synthesis. WT and Bmal1−/− mouse testes sections had similar normal morphologies, although there was a decrease in testicular spermatozoa in the Bmal1−/− mice. Low serum testosterone levels were detected in the Bmal1−/− mice. RNA sequencing identified 37 and 48 genes that were differentially expressed between WT and Bmal1−/− mouse testes at circadian time (CT2 and CT14), respectively. The cholesterol metabolism pathway was significantly enriched in the KEGG pathway analysis, and there was lower expression of three apolipoprotein genes (Apoa1, Apoa2, and Apoc3) at CT2 in the testes of Bmal1−/− mice than in those of WT mice. These decreases in Apoa1, Apoa2, and Apoc3 expression were verified by quantitative polymerase chain reaction analysis, which also revealed downregulation of the expression of the circadian clock (Per2, Dbp, and Nr1d1) and steroidogenic (StAR, Cyp11a1, and Hsd17b3) genes. The expression of circadian clock genes was relatively stable in WT mice over a 20-h period, whereas there was clear circadian rhythmic expression of Apoa1, Apoa2, Apoc3, StAR, Cyp11a1, Hsd3b2, and Hsd17b3. Bmal1−/− mice showed severely reduced expression of testicular circadian clock genes at three time points (CT4, CT12, and CT20), and a reduction in mRNA expression levels of Apo (Apoa1, Apoa2, and Apoc3) and steroidogenic (StAR, Cyp11a1, Hsd3b2, and Hsd17b3) genes. Oil Red O staining showed decreased lipid aggregation in the Leydig cells of Bmal1−/− mouse testes. Considering the vital role of Apo genes in high-density lipoprotein formation and cholesterol transport, the present data suggest that the circadian clock system regulates testosterone production by orchestrating the rhythmic expression of Apo genes. These data extend our understanding of the role of the circadian clock in regulating testosterone production in mammals.

Myeloperoxidase Targets Apolipoprotein A-I for Site-Specific Tyrosine Chlorination in Atherosclerotic Lesions and Generates Dysfunctional High-Density Lipoprotein.

We previously demonstrated that apolipoprotein A-I (apoA-I), the major protein component of high-density lipoprotein (HDL), is an important target for myeloperoxidase (MPO)-catalyzed tyrosine chlorination in the circulation of subjects with cardiovascular diseases. Oxidation of apoA-I by MPO has been reported to deprive HDL of its protective properties. However, the potential effects of MPO-mediated site-specific tyrosine chlorination of apoA-I on dysfunctional HDL formation and atherosclerosis was unclear. Herein, Tyr192 in apoA-I was found to be the major chlorination site in both lesion and plasma HDL from humans with atherosclerosis, while MPO binding to apoA-I was demonstrated by immunoprecipitation studies in vivo. In vitro, MPO-mediated damage of lipid-free apoA-I impaired its ability to promote cellular cholesterol efflux by the ABCA1 pathway, whereas oxidation to lipid-associated apoA-I inhibited lecithin:cholesterol acyltransferase activation, two key steps in reverse cholesterol transport. Compared with native apoA-I, apoA-I containing a Tyr192 → Phe mutation was moderately resistant to oxidative inactivation by MPO. In high-fat-diet-fed apolipoprotein E-deficient mice, compared with native apoA-I, subcutaneous injection with oxidized apoA-I (MPO treated) failed to mediate the lipid content in aortic plaques while mutant apoA-I (Tyr192 → Phe) showed a slightly stronger ability to reduce the lipid content in vivo. Our observations suggest that oxidative damage of apoA-I and HDL involves MPO-dependent site-specific tyrosine chlorination, raising the feasibility of producing MPO-resistant forms of apoA-I that have stronger antiatherosclerotic activity in vivo.

The effect of 10 days of energy-deficit diet and high-intensity exercise training on the plasma high-density-lipoprotein (HDL) level among healthy collegiate males

ABSTRACT An energy-deficit (ED) diet increases lipid mobilisation, while endurance exercise improves lipid profile by promoting formation of high-density lipoproteins (HDLs) among moderately active population. However, it is not clear whether ED with high-intensity exercise training can improve lipid profiles. Therefore, 20 recreationally active males (20.3 ± 2.7 years old with peak oxygen consumption (O2peak) of 2.8 ± 0.2 L min−1) with HDL concentration of 1.44 ± 0.32 mmol L−1, were evenly allocated to either an ED group (1590 ± 79 kcal day−1: 40% of total caloric omitted at lunchtime) or a control (CON) group (2570 ± 139 kcal day−1). Participants in both groups performed high-intensity cycling at 90% O2peak at a constant workload for 8 min, 2 h after breakfast on day D0, D2, D4, D6, D8 and D10 (Dn is day number), after which blood samples were collected. In ED, compared to D0, triglycerides (TG) decreased on D6-10 (p < .01) while total low-density lipoprotein (LDL), total cholesterol (TC) increased on D2-10, while HDL progressively increased on D6-10 (p = .01) with a final value of 1.67 ± 0.24 mmol L−1. In CON, there were no changes in TG, TC and HDL while LDL was reduced on D8-10 (p = .01). Moreover, the proportions of TC/HDL and LDL/HDL increased in ED on D2-10 and D2-8 (p = .05), respectively, while LDL/HDL in CON was reduced on D8-10 (p = .01). In brief, 10 days of ED and a series of high-intensity exercise sessions show progressive elevation of HDL which suggest longer period is required to observe changes in lipid ratios.

A key point mutation (V156E) affects the structure and functions of human apolipoprotein A-I.

A naturally occurring point mutant of human apolipoprotein A-I (apoA-I), V156E, which is associated with extremely low plasma apoA-I and high density lipoprotein (HDL) levels, and coronary artery disease (Huang, W., Sasaki, J., Matsunaga, A., Nanimatsu, H., Moriyama, K., Han, H. Kugi, M., Koga, T., Yamaguchi, K., and Arakawa, K. (1998) Arterioscler. Throm. Vasc. Biol. 18, 389-396), was produced in an Escherichia coli expression system. The purified recombinant proapoA-I V156E mutant was examined in its structural and functional properties, both, in the lipid-free and lipid-bound states. In the lipid-free form the mutant protein exhibited small changes in conformation, but was more stable, and quite resistant to self-association, compared with control apoA-I. The V156E mutant was able to interact with phospholipid (PL) at high PL:protein ratios (95:1, mol/mol), but was inefficient in forming reconstituted HDL (rHDL) complexes at lower PL:protein ratios (40:1). In the lipid-bound, rHDL state, the mutant protein was somewhat more alpha-helical and formed a larger complex (110 A) than control apoA-I (97 A). Furthermore, the rHDL particles containing the V156E mutant did not rearrange to smaller particles in the presence of low density lipoproteins, and had minimal reactivity with lecithin-cholesterol acyltransferase (LCAT), compared with rHDL particles made with control apoA-I. These results suggest a key role for Val-156, or the adjacent central region of apoA-I in the modulation of apoA-I conformation, stability, and self-association in solution, and in the formation of small HDL, the conformational adaptability of apoA-I leading to structural rearrangements of HDL, and the activation of LCAT.

High-density lipoprotein-related cholesterol metabolism in Alzheimer's disease.

High-density lipoproteins (HDL) are a heterogeneous class of molecules whose main function is to remove excess cholesterol through a mechanism called reverse transport, in which cholesterol is transported from peripheral organs and from arterial foam cells to the liver, where it is subsequently eliminated with bile. While its ability to eliminate excess cholesterol has always been viewed as its main feature, its beneficial effects go beyond this single effect. Many of the proteins that are associated with HDL are responsible for anti-oxidant and anti-inflammatory properties. These proteins that are associated with HDL during its generation and remodelling, are referred to as 'protein cargo', which has been extensively analysed by mass spectrometry analysis in healthy and diseased individuals. In this review, we discuss the pathway that leads to HDL formation and its subsequent remodelling and catabolism with regards to the possible involvement of HDL 'protein cargo' in Alzheimer's disease.

Adiponectin's mechanisms in high-density lipoprotein biogenesis and cholesterol efflux

AimAmong adiponectin's beneficial properties is its ability to promote cellular cholesterol efflux, thereby generating high-density lipoprotein (HDL) particles. However, adiponectin's role in the regulation of macrophage lipid metabolism, a crucial process in atherogenesis, remains poorly investigated. The aim of this study was to characterize the adiponectin's role in HDL biogenesis. Methods and resultsWe perform kinetics studies in baby hamster kidney (BHK) and Tamm-Horsfall protein 1 (THP-1) cell lines to elucidate adiponectin's role in HDL biogenesis. In cholesterol-enriched cells, specific molar doses of adiponectin stimulated cholesterol efflux with high efficiency to apoA-I. In the presence of adiponectin, BHK cells expressing ATP binding cassette transporter A1 (ABCA1) or ABCG1 generated lipidated particles having α electrophoretic mobility (α-HDL) and a molecular size of 7.5–20 nm. Interestingly, in THP-1 macrophages, cholesterol efflux was associated with more lipidated preβ1-HDL particles. Direct molecular interaction of adiponectin with apoA-I enhanced the affinity of apoA-I to free cholesterol and resulted in an increase in preβ1-HDL particles from plasma ex vivo. Adiponectin increased ABCA1 and ABCG1 protein expression and activated the formation of ABCA1-linked cholesterol oxidase sensitive plasma membrane domains. ConclusionAdiponectin upregulated ABCA1 and ABCG1 protein expression, reduced lipid accumulation, and efficiently promoted nascent HDL formation. These results highlight that these cellular processes are interconnected through adiponectin and ABCA1- and ABCG1-dependent. In this pathway, adiponectin increased the affinity of apoA-I to cholesterol and effectively accelerated cholesterol removal from the plasma membrane to HDL particles. Thus, by accelerating HDL biogenesis, adiponectin may have therapeutic potential for atherosclerotic cardiovascular disease prevention and management.

Apolipoprotein M: Research Progress and Clinical Perspective.

Apolipoprotein M (apoM) was first identified and characterized to the apolipoprotein family in 1999. Human apoM gene is located in a highly conserved segment in the major histocompatibility complex (MHC) class III locus on chromosome 6 and codes for an about 23kDa protein that structurally belongs to the lipocalin superfamily. ApoM is selectively expressed in hepatocytes and in the tubular epithelium of kidney. In human plasma, apoM is mainly confined to the high-density lipoprotein (HDL) particles, but it may also occur in other lipoprotein classes, such as in the triglyceride-rich particles after fat intake. It has been demonstrated that apoM is critical for the formation of HDL, notably pre-beta HDL1. The antiatherogenic function of HDL is well established, and its ability to promote cholesterol efflux from foam cells in the atherosclerotic lesions is generally regarded as one of the key mechanisms behind this protective function. However, HDL could also display a variety of properties that may affect the complex atherosclerotic processes by other mechanisms, thus being involved in processes related to antioxidant defense, immune system, and systemic effects in septicemia, which may be partly contributed via its apolipoproteins and/or phospholipids. Moreover, it has been demonstrated that apoM functions as a natural carrier of sphingosin-1-phosphate (S1P) in vivo which may be related to its antiatherosclerotic and protective effects on endothelial cell barrier and anti-inflammatory properties. These may also provide a link between the diverse effects of HDL.

Phosphorylation by protein kinase C stabilizes ABCG1 and increases cholesterol efflux.

ATP-binding cassette protein G1 (ABCG1) plays an important role in eliminating excess cholesterol from macrophages and in the formation of high-density lipoprotein (HDL), which contribute to the prevention and regression of atherosclerosis. The post-translational regulation of ABCG1 remains elusive, although phosphorylation by protein kinase A destabilizes ABCG1 proteins. We examined the phosphorylation of ABCG1 using HEK293 and Raw264.7 cells. ABCG1 phosphorylation was enhanced by treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA), a protein kinase C (PKC) activator. PKC activation by TPA increased ABCG1 protein levels and promoted ABCG1-dependent cholesterol efflux to HDL. This activity was suppressed by Go6976, a PKCα/βI inhibitor, suggesting that PKC activation stabilizes ABCG1. To confirm this, the degradation rate of ABCG1 was analysed; ABCG1 degradation was suppressed upon PKC activation, suggesting that PKC phosphorylation regulates ABCG1 levels. To confirm this involvement, we co-expressed ABCG1 and a constitutively active form of PKCα in HEK cells. ABCG1 was increased upon co-expression. These results suggest that PKC-mediated phosphorylation, probably PKCα, stabilizes ABCG1, consequently increasing ABCG1-mediated cholesterol efflux, by suppressing ABCG1 degradation. PKC activation could thus be a therapeutic target to suppress the development of atherosclerosis.