Comparison of different cellular models measuring in vitro the whole human serum cholesterol efflux capacity

2006 ◽  
Vol 36 (8) ◽  
pp. 552-559 ◽  
Author(s):  
S. Mweva ◽  
J. L. Paul ◽  
M. Cambillau ◽  
D. Goudouneche ◽  
P. Beaune ◽  
...  
Keyword(s):  
Human Serum ◽  
Serum Cholesterol ◽  
Cholesterol Efflux ◽  
Cholesterol Efflux Capacity ◽  
Cellular Models

scholarly journals 260Identification of differential roles of microRNA-33a and -33b during atherosclerosis progression with genetically modified mice

2019 ◽  
Vol 40 (Supplement_1) ◽  
Author(s):  
T Horie ◽  
S Koyama ◽  
T Kimura ◽  
K Ono
Keyword(s):  
Bone Marrow ◽  
Serum Cholesterol ◽  
Cholesterol Efflux ◽  
Target Genes ◽  
Genetically Modified ◽  
Expression Levels ◽  
Cholesterol Efflux Capacity ◽  
Predominant Expression ◽  
Progression Of Atherosclerosis

Abstract Background MicroRNAs (miRs) are small non-protein-coding RNAs that bind to specific mRNAs and inhibit translation or promote mRNA degradation. Recent reports, including ours, indicated that miR-33a located within the intron of sterol regulatory element-binding factor (SREBF) 2 targets cholesterol transporter ATP-binding cassette protein A1 (ABCA1) or other anti-atherogenic targets and contributes to atherogenesis. Its inhibition or deletion is known to result in the amelioration of atherosclerosis. However, mice lack the other member of miR-33 family, miR-33b, which exists in humans. Precise evaluation and comparison of the responsibilities of these two miRs during the progression of atherosclerosis are essential and need to be investigated. Methods and results The difference between miR-33a and miR-33b in vitro and in vivo were analyzed from multiple directions using genetically modified miR-33a knock-out (KO) and miR-33b knock-in (KI) humanized mice. At first, we performed transcriptomic analysis of primary cultured hepatocytes transfected with synthetic miR-33a, miR-33b, and a control miR and found similar potential target repression and targeting motif of miR-33a and miR-33b in vitro. However, we noticed distinct expression patterns of miR-33a and miR-33b in several organs. By crossing miR-33a KO and miR-33b KI mice, we established four strains with or without miR-33a and miR-33b. Comparison of these strains showed distinct distribution and regulation of miR-33 family. In particular, comparison between mice with only miR-33a (wild-type mice) and mice with only miR-33b (miR-33a−/− miR-33b+/+) revealed 4-fold predominant expression of miR-33b in the liver. Such differential expression resulted in a reduced expression of target genes such as ABCA1 and worsened serum cholesterol profile in mice with only miR-33b. On the contrary, in macrophages the expression levels of miR-33 family genes were similar and their effects on target genes and cholesterol efflux capacity to ApoA-I or HDL cholesterol (HDL-C) were almost comparable. To evaluate the whole body atherogenic potency, we developed ApoE−/− miR-33a+/+ miR-33b−/− mice and ApoE−/− miR-33a−/− miR-33b+/+ mice. ApoE−/− miR-33a−/− miR-33b+/+ mice developed increased atherosclerotic plaque compared with ApoE−/− miR-33a+/+ miR-33b−/− mice, in line with the predominant expression of miR-33b in the liver and decreased serum HDL-C levels whose lower cholesterol efflux capacity were confirmed in 3H-labeled macrophages. On the contrary, a bone marrow transplantation study showed no significant difference in atherosclerosis and serum cholesterol profile, and this was consistent with the relevant expression levels of miR-33a and miR-33b in bone marrow cells. Conclusions miR-33 family exhibited differences in distribution and regulation, and particularly in the progression of atherosclerosis, miR-33b would be more potent than miR-33a.

Abstract 238: Incubation of MDCO216 With Human Serum or Plasma Potentiates ABCA1-Mediated Cholesterol Efflux Capacity, Increases Prebeta-1 HDL and Causes a Shift From Small to Large Alpha HDL Particles

2014 ◽  
Vol 34 (suppl_1) ◽  
Author(s):  
Herman J Kempen ◽  
Dorota B Schranz ◽  
Bela Asztalos ◽  
Elias Jeyarajah ◽  
James Otvos ◽  
...  
Keyword(s):  
Human Serum ◽  
Human Plasma ◽  
Cholesterol Efflux ◽  
Time Dependent ◽  
Plaque Burden ◽  
Strong Increase ◽  
2D Electrophoresis ◽  
Cholesterol Efflux Capacity ◽  
J774 Cells ◽  
Synergistic Increase

MDCO216 is a complex of dimeric apoA-IMilano and POPC, shown to reduce atherosclerotic plaque burden. Here we studied the effect of incubation of human plasma or serum with MDCO216 on cholesterol efflux capacity from J774 cells, on prebeta-1 HDL and various HDL subfractions. At clinically relevant concentrations MDCO216 by itself markedly stimulates global and ABCA1-mediated cholesterol efflux. When incubated with human serum a time-dependent synergistic increase of the ABCA1 mediated efflux capacity is observed. Using the Sekisui ELISA for prebeta-1 HDL, MDCO216 itself is poorly detected. Prebeta-1 HDL is rapidly lost when human plasma alone is incubated at 37 o C. However, incubation of human plasma with MDCO216 at 37 o C causes a robust generation of new prebeta-1 HDL. 2D-Electrophoresis followed by immunoblotting with a general apoA-I antibody that also detects apoA-IM confirms the increase in prebeta-1 HDL (having a different mobility than pure MDCO216 particles), and shows a concomitant disappearance of alpha-3 HDL, alpha-4 HDL and MDCO216, and an increase in alpha-1 and alpha-2 HDL. NMR analysis of plasma incubated with MDCO216 at 47 o C confirms very rapid disappearance of small HDL and increase of medium and large HDL particles. In conclusion, incubation of human plasma or serum with MDCO216 strongly enhances ABCA1-mediated cholesterol efflux capacity, causes a strong increase of prebeta-1 HDL and drastically changes HDL subfraction distribution, consistent with anti-atherosclerotic activity.

Abstract 17232: Comparison of Methods for the Measurement of Cholesterol Efflux Capacity of Plasma HDL With J774 Mouse Macrophages Reveals Superiority of the Radioactive Cholesterol Method over the BODIPY-Cholesterol-2 Method

Circulation ◽  
2015 ◽  
Vol 132 (suppl_3) ◽  
Author(s):  
David Rhainds ◽  
Renaud Julien ◽  
Boulé Marie ◽  
Denis Maxime ◽  
Rhéaume Eric ◽  
...  
Keyword(s):  
Cholesterol Efflux ◽  
Hdl Cholesterol ◽  
Fold Increase ◽  
Strongly Correlated ◽  
Response Curves ◽  
Complex Samples ◽  
Cholesterol Efflux Capacity ◽  
Mouse Macrophages ◽  
Efflux Time

Recent clinical studies suggest that raising HDL-cholesterol (HDL-C) concentration is insufficient to lower cardiovascular risk and that protection derives from other characteristics of HDL particles. One such characteristic that can be measured in vitro is the cholesterol efflux capacity of plasma, an assay where HDL accepts labelled cholesterol from macrophages. Recently, a method for the measurement of cholesterol efflux with a fluorescent tracer, BODIPY-cholesterol-2 (BC2), has been proposed as an alternative to the radioactive method. We undertook a systematic comparison of BC2 and 3H-cholesterol methods with J774 macrophages in basal and cAMP-stimulated conditions with purified acceptors, plasma from healthy volunteers and patients with myocardial infarction of the Montreal Heart Institute Biobank. Dose-response curves show higher affinity of BC2 vs. 3H cholesterol with apoA-I, HDL and apoB-depleted plasma (all p<0.01) and a higher maximal efflux for HDL and depleted plasma (both p<0.001) in stimulated conditions. This was reflected in a faster kinetics of BC2 efflux compared to 3H-cholesterol efflux (time for half-maximal efflux 4.1h vs. 10.2 h) and resulted in a 2.5-fold increase of BC2 maximal efflux (p<0.01). With 50 normolipidemic plasmas (2.8%, 4h efflux), the two methods were not correlated in basal conditions (r2=0.079) and strongly correlated in stimulated conditions (r2=0.830). BC2 values did not correlate with HDL-C in all conditions, contrary to 3H cholesterol values (p<0.01). Using more complex samples from patients with MI (n=115), the correlations were modest in basal (r2=0.219) and stimulated (r2=0.400) conditions. Differences between controls and MI cases showed reduced significance with BC-2 compared to 3H cholesterol. Lastly, in macrophages examined under the confocal microscope, BC2 labels vesicular structures colocalized with filipin, a free cholesterol marker, suggesting endosomal loading of BC2. Thus, the BC2 method is not equivalent to the classic 3H cholesterol method for cholesterol efflux measurement.

Abstract 319: Dalcetrapib-Mediated Modulation of Cholesteryl Ester Transfer Protein Activity Increases HDL-Cholesterol, Apolipoprotein A-I Levels and Cholesterol Efflux Capacity in Chow-Fed Rabbits

2012 ◽  
Vol 32 (suppl_1) ◽  
Author(s):  
Mathieu R Brodeur ◽  
David Rhainds ◽  
Daniel Charpentier ◽  
Téodora Mihalache-Avram ◽  
Cyrille Maugeais ◽  
...  
Keyword(s):  
Cholesteryl Ester ◽  
Cholesteryl Ester Transfer Protein ◽  
Cholesterol Efflux ◽  
Hdl Cholesterol ◽  
Lipid Transfer ◽  
Protein Activity ◽  
Cholesterol Efflux Capacity ◽  
Transfer Protein

Introduction: A potential approach to reduce CV risk is to increase HDL-C levels. This could be achieved by reducing cholesteryl ester transfer protein (CETP) activity. Dalcetrapib, which modulates CETP activity by changing its conformation and raises HDL-C without inhibiting CETP-induced pre-β-HDL formation in humans, was shown to decrease progression of atherosclerosis in rabbits. Hypothesis: Investigate the modifications of HDL particle size distribution and cholesterol efflux capacity of serum produced by dalcetrapib in normocholesterolemic rabbits. Methods: New Zealand white rabbits were treated with dalcetrapib (300 mg/kg as food admix) or placebo for 14 days. We evaluated CETP conformation and mass by ELISAs (including antibodies sensitive to conformational change), CETP activity by fluorescent lipid transfer, lipid profile and apoA-I distribution in HDL subclasses by 2D-non denaturing gradient gels (2D-NDGGE). Cholesterol efflux capacity of rabbit sera was determined after loading cells with 3 H-free cholesterol, using HepG2 hepatocytes to measure SR-BI-dependent efflux and by inducing ABCA1 or ABCG1 expression in BHK cells. Results: Dalcetrapib modified the conformation of rabbit CETP in vitro and in vivo and, after 14 days, this was associated with increased CETP mass (+50%, p<0.001) and reduced CETP activity (-86%, p<0.001). Total cholesterol was increased with dalcetrapib (+178%, p<0.001), due to a higher HDL-C level. In contrast, dalcetrapib reduced LDL-C and triglycerides by 41% (p<0.01) and 48% (p<0.001). Serum analysis by 2D-NDGGE showed that total rabbit apoA-I was increased 1.7- fold in animals treated with dalcetrapib. This was associated with an increase in large HDL but also in small α-migrating HDL with pre-β-HDL size. Cholesterol efflux assays showed that ABCA1-, ABCG1- and SR-BI-dependent efflux were all increased in dalcetrapib-treated rabbits (+24%, p=0.038; +21%, p=0.021; +44%, p<0.001). Conclusion: Modulation of CETP activity and conformation by dalcetrapib increases HDL-C and apoA-I levels and affects apoA-I distribution in HDL subclasses. These changes are associated with increased cholesterol efflux capacity, suggesting that HDL functionality is preserved in dalcetrapib-treated chow-fed rabbits.

scholarly journals In Vivo Inflammation Does Not Impair ABCA1-Mediated Cholesterol Efflux Capacity of HDL

Cholesterol ◽  
2012 ◽  
Vol 2012 ◽  
pp. 1-8 ◽  
Author(s):  
Remco Franssen ◽  
Alinda W. M. Schimmel ◽  
Sander I. van Leuven ◽  
Simone C. S. Wolfkamp ◽  
Erik S. G. Stroes ◽  
...  
Keyword(s):  
Severe Sepsis ◽  
Inflammatory Disease ◽  
Cholesterol Efflux ◽  
Chronic Inflammatory Disease ◽  
Cholesterol Efflux Capacity ◽  
Hdl Function ◽  
In Vivo Inflammation ◽  
The Impact

HDL provides atheroprotection by facilitating cholesterol efflex from lipid-laden macrophages in the vessel wall. In vitro studies have suggested impaired efflux capacity of HDL following inflammatory changes. We assessed the impact of acute severe sepsis and mild chronic inflammatory disease on the efflux capacity of HDL. We hypothesize that a more severe inflammatory state leads to stronger impaired cholesterol efflux capacity. Using lipid-laden THP1 cells and fibroblasts we were able to show that efflux capacity of HDL from both patients with severe sepsis or with Crohn's disease (active or in remission), either isolated using density gradient ultracentrifugation or using apoB precipitation, was not impaired. Yet plasma levels of HDL cholesterol and apoA-I were markedly lower in patients with sepsis. Based on the current observations we conclude that inflammatory disease does not interfere with the capacity of HDL to mediate cholesterol efflux. Our findings do not lend support to the biological relevance of HDL function changes in vitro.

scholarly journals ABCA1-dependent serum cholesterol efflux capacity inversely correlates with pulse wave velocity in healthy subjects

2012 ◽  
Vol 54 (1) ◽  
pp. 238-243 ◽  
Author(s):  
Elda Favari ◽  
Nicoletta Ronda ◽  
Maria Pia Adorni ◽  
Francesca Zimetti ◽  
Paolo Salvi ◽  
...  
Keyword(s):  
Pulse Wave Velocity ◽  
Wave Velocity ◽  
Serum Cholesterol ◽  
Healthy Subjects ◽  
Cholesterol Efflux ◽  
Pulse Wave ◽  
Cholesterol Efflux Capacity

scholarly journals HDL Phospholipid Content and Composition as a Major Factor Determining Cholesterol Efflux Capacity From Fu5AH Cells to Human Serum

1997 ◽  
Vol 17 (11) ◽  
pp. 2685-2691 ◽  
Author(s):  
Natalie Fournier ◽  
Jean-Louis Paul ◽  
Véronique Atger ◽  
Anne Cogny ◽  
Théophile Soni ◽  
...  
Keyword(s):  
Human Serum ◽  
Cholesterol Efflux ◽  
Phospholipid Content ◽  
Cholesterol Efflux Capacity

scholarly journals Relationship between HDL Cholesterol Efflux Capacity, Calcium Coronary Artery Content, and Antibodies against ApolipoproteinA-1 in Obese and Healthy Subjects

2019 ◽  
Vol 8 (8) ◽  
pp. 1225 ◽  
Author(s):  
Nicolas Vuilleumier ◽  
Sabrina Pagano ◽  
Fabrizio Montecucco ◽  
Alessandra Quercioli ◽  
Thomas H. Schindler ◽  
...  
Keyword(s):  
Coronary Artery ◽  
Cholesterol Efflux ◽  
Positive Association ◽  
Hdl Cholesterol ◽  
Cholesterol Content ◽  
Cholesterol Efflux Capacity ◽  
Clinical Observations ◽  
Framingham Risk ◽  
Obese Subjects

Aims: To explore the associations between cholesterol efflux capacity (CEC), coronary artery calcium (CAC) score, Framingham risk score (FRS), and antibodies against apolipoproteinA-1 (anti-apoA-1 IgG) in healthy and obese subjects (OS). Methods and Results: ABCA1-, ABCG1-, passive diffusion (PD)-CEC and anti-apoA-1 IgG were measured in sera from 34 controls and 35 OS who underwent CAC score determination by chest computed tomography. Anti-apoA-1 IgG ability to modulate CEC and macrophage cholesterol content (MCC) was tested in vitro. Controls and OS displayed similar ABCG1-, ABCA1-, PD-CEC, CAC and FRS scores. Logistic regression analyses indicated that FRS was the only significant predictor of CAC lesion. Overall, anti-apoA-1 IgG were significantly correlated with ABCA1-CEC (r = 0.48, p < 0.0001), PD-CEC (r = −0.33, p = 0.004), and the CAC score (r = 0.37, p = 0.03). ABCA1-CEC was correlated with CAC score (r = 0.47, p = 0.004) and FRS (r = 0.18, p = 0.29), while PD-CEC was inversely associated with the same parameters (CAC: r = −0.46, p = 0.006; FRS: score r = −0.40, p = 0.01). None of these associations was replicated in healthy controls or after excluding anti-apoA-1 IgG seropositive subjects. In vitro, anti-apoA-1 IgG inhibited PD-CEC (p < 0.0001), increased ABCA1-CEC (p < 0.0001), and increased MCC (p < 0.0001). Conclusions: We report a paradoxical positive association between ABCA1-CEC and the CAC score, with the latter being inversely associated with PD in OS. Corroborating our clinical observations, anti-apoA-1 IgG enhanced ABCA1 while repressing PD-CEC, leading to MCC increase in vitro. These results indicate that anti-apoA-1 IgG have the potential to interfere with CEC and macrophage lipid metabolism, and may underpin paradoxical associations between ABCA1-CEC and cardiovascular risk.

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