EXPRESS: Novel cholesterol efflux assay using immobilized liposome-bound gel beads: Confirmation and improvement for application in clinical laboratory

2021 ◽  
pp. 000456322110544
Author(s):  
Yuna Horiuchi ◽  
Shao-Jui Lai ◽  
Takahiro Kameda ◽  
Minoru Tozuka ◽  
Ryunosuke Ohkawa
Keyword(s):  
Reference Material ◽  
Cholesterol Efflux ◽  
Clinical Laboratory ◽  
Density Lipoprotein ◽  
Radioactive Materials ◽  
Simple Method ◽  
Cholesterol Efflux Capacity ◽  
Gel Beads ◽  
Potential Biomarker ◽  
High Bilirubin

Objectives Cholesterol efflux capacity (CEC), an atheroprotective function of high-density lipoprotein, is expected to be a potential biomarker for cardiovascular disease. However, CEC has not been widely introduced for application in clinical laboratories because of the complexity of the conventional CEC assay using cells and radioactive materials. Previously, we developed a novel CEC assay using immobilized liposome-bound gel beads (ILG), which solves these issues. We aimed to confirm the validation and further improve the ILG method for application in the clinical setting. Methods Cholesterol efflux capacity values by the ILG method assayed for shorter incubation time (4 h) were compared to those assayed for 16 h (our previous ILG method). To investigate a reference material that can correct the variation between ILG manufacturing lots, bovine serum albumin, human gamma-globulins, and globulin complexes were evaluated. CEC values were also estimated in plasmas obtained with different anticoagulants, serum treated with freeze-thaw cycles, and serum mixed with several interference substances. Results The CEC of 4- and 16-h incubation times were well correlated. Globulin complexes may be used as a reference material. Plasma can be used as the specimen. The serum and stored temperature of the specimen did not largely affect CEC. Hemoglobin and chyle did not have an effect on CEC, whereas high-bilirubin serum showed elevated CEC. The effect of bilirubin was nearly canceled by subtracting basal fluorescence intensity. Conclusions Present ILG method further fulfills some requirements for application in clinical laboratory. Using this reliable simple method, evaluation for clinical significance of CEC is expected.

scholarly journals Cholesterol efflux capacity assay using immobilized liposomes and apolipoprotein B-depleted serum

2019 ◽  
Vol 39 (6) ◽  
Author(s):  
Hima Rani Sapa
Keyword(s):  
Apolipoprotein B ◽  
Cholesterol Efflux ◽  
Ex Vivo ◽  
Density Lipoprotein ◽  
Poly Ethylene Glycol ◽  
Cholesterol Efflux Capacity ◽  
Gel Beads ◽  
Functional Step ◽  
Poly Ethylene

Abstract Cholesterol efflux capacity (CEC), an important functional step in reverse cholesterol transport, is the main anti-atherosclerotic function of high-density lipoprotein (HDL). Assays that improve the determination of CEC ex vivo for clinical applications are constantly explored. In the accompanying article, Horiuchi et al. (Biosci. Rep. (2019) 39(4), BSR20190213) evaluate the availability of apolipoprotein B-depleted serum for CEC assays. Using their recently developed immobilized liposome-bound gel beads (ILG) method, Horiuchi et al. demonstrate that apolipoprotein B-depleted serum obtained with poly ethylene glycol precipitation enables CEC assays to be easily and accurately introduced into laboratory medicine.

scholarly journals Comparison of a novel cholesterol efflux assay using immobilized liposome-bound gel beads with the conventional method

2020 ◽  
Vol 40 (8) ◽  
Author(s):  
Yuna Horiuchi ◽  
Shao-Jui Lai ◽  
Takahiro Kameda ◽  
Minoru Tozuka ◽  
Ryunosuke Ohkawa
Keyword(s):  
High Density Lipoprotein ◽  
Conventional Method ◽  
Strong Correlation ◽  
Cholesterol Efflux ◽  
Clinical Laboratory ◽  
Density Lipoprotein ◽  
Foam Cells ◽  
Atp Binding Cassette Transporter ◽  
Gel Beads ◽  
A Minor

Abstract Cholesterol efflux capacity (CEC) is an atheroprotective function of high-density lipoprotein (HDL). CEC is currently measured using artificially prepared foam cells composed of cultured macrophage and 3H-cholesterol. However, this conventional method is not suitable for clinical laboratory use due to poor repeatability, complexity, and low safety. Recently, we reported a novel CEC assay, called the immobilized liposome-bound gel beads (ILG) method. The ILG method is an alternative to foam cells, comprising gel beads and 4,4-diflioro-4-bora-3a,4a-s-indacene labeled cholesterol (BODIPY-cholesterol) instead of macrophage and 3H-cholesterol, respectively. The ILG method has shown adequate basic properties and strong correlation with the conventional method. Here, we aimed to compare this new ILG method with the conventional method in-depth. When apoB-depleted serum was used as the cholesterol acceptor (CA), the ILG method had far better reproducibility than the conventional method. The CEC of major HDL subclasses HDL2 and HDL3 had similar results in both the ILG and conventional method. However, the ILG method did not reflect the CEC of apolipoprotein (apo) A–I and a minor HDL subclass which uses ATP-binding cassette transporter A1 on foam cells. Superior reproducibility of the ILG method, which is a limitation of the conventional method, and similar CEC results for major HDL subclasses in the ILG and conventional methods, provide further evidence that the ILG method is promising for measuring CEC clinically. However, some HDL subclasses or apo might have poor CEC correlation between these methods. Further research is therefore needed to confirm the clinical significance of estimating CEC by the ILG method.

scholarly journals High Density Lipoprotein Cholesterol Efflux Capacity and Atherosclerosis in Cardiovascular Disease: Pathophysiological Aspects and Pharmacological Perspectives

Cells ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 574
Author(s):  
Maria Pia Adorni ◽  
Nicoletta Ronda ◽  
Franco Bernini ◽  
Francesca Zimetti
Keyword(s):  
High Density Lipoprotein ◽  
Cholesterol Efflux ◽  
Current Knowledge ◽  
Density Lipoprotein ◽  
Hdl Cholesterol ◽  
High Density ◽  
Current Evidence ◽  
Endocrine Disorders ◽  
Lifestyle Modifications ◽  
Cholesterol Efflux Capacity

Over the years, the relationship between high-density lipoprotein (HDL) and atherosclerosis, initially highlighted by the Framingham study, has been revealed to be extremely complex, due to the multiple HDL functions involved in atheroprotection. Among them, HDL cholesterol efflux capacity (CEC), the ability of HDL to promote cell cholesterol efflux from cells, has emerged as a better predictor of cardiovascular (CV) risk compared to merely plasma HDL-cholesterol (HDL-C) levels. HDL CEC is impaired in many genetic and pathological conditions associated to high CV risk such as dyslipidemia, chronic kidney disease, diabetes, inflammatory and autoimmune diseases, endocrine disorders, etc. The present review describes the current knowledge on HDL CEC modifications in these conditions, focusing on the most recent human studies and on genetic and pathophysiologic aspects. In addition, the most relevant strategies possibly modulating HDL CEC, including lifestyle modifications, as well as nutraceutical and pharmacological interventions, will be discussed. The objective of this review is to help understanding whether, from the current evidence, HDL CEC may be considered as a valid biomarker of CV risk and a potential pharmacological target for novel therapeutic approaches.

Abstract 97: Increased Plasma Level of ApoCIII-rich Electronegative HDL May Contribute to Cognitive Impairment in Alzheimer’s Disease

Stroke ◽  
2017 ◽  
Vol 48 (suppl_1) ◽  
Author(s):  
Hua-Chen Chan ◽  
Hsiao-Ting Lu ◽  
Mei-Chuan Chou ◽  
Ming-Hsien Tsai ◽  
Wei-Hsiang Chen ◽  
...  
Keyword(s):  
Cognitive Impairment ◽  
Cholesterol Efflux ◽  
Chemical Properties ◽  
Density Lipoprotein ◽  
Fold Increase ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Paraoxonase 1 ◽  
Cholesterol Efflux Capacity ◽  
Tnf Α

Background: High-density lipoprotein (HDL), the only lipoprotein class that can cross the blood brain barrier bidirectionally, is positively associated with cognitive functions. To delineate HDL’s role in Alzhenimer’s disease (AD), we analyzed the chemical properties of plasma HDL from AD and healthy normal adult (control) subjects. Methods and results: By using anion-exchange chromatography, we divided HDL into 5 increasingly electronegative subfractions, H1-H5. Compared to the control cohort (4.24±3.22%; n=20), HDL from AD patients (23.48±17.83%; n=30) had a 5.5-fold increase of H5 ( P <0.001; Figure ), accompanied by a decreased protein/lipid ratio attributed to a significant reduction of albumin essential for prevention of amyloid beta (Aβ) aggregation. As determined by LC/MS E and ProteinLynx Global SERVER (PLGS), AD-HDL was had a rich content of apolipoprotein (apo)CIII, but diminished amounts of sphingosine-1-phosphate (S1P)-associated apoM and antioxidative paraoxonase 1 (PON1). Exposure of murine RAW 264.7 macrophages to H5 induced vibrant expression of ganglioside GM1 in colocalization with apoCIII on lipid rafts, alongside a concomitant increase of TNF-α detectable in the cultured medium ( Figure ). LC/MS E examination localized posttranslational oxidation exclusively in ApoA1 residues of H5 in AD-HDL, which exhibited a compromised cholesterol efflux capacity. Conclusions: Plasma HDL from AD patients has a high proportion of H5, an apoCIII-rich electronegative HDL subfraction. The associated reduction in functional (albumin, S1P, apoM) and increase in proinflammatory (apoCIII, PON1, TNF-α) components may favor Aβ assembly and neuroinflammation. Additionally, a compromised cholesterol-efflux capacity of AD-HDL may also contribute to vascular cognitive impairment.

Abstract 540: Correlation of Improved Cholesterol Efflux Capacity of High Density Lipoprotein With Survival and Allograft Vasculopathy in Cardiac Transplant Recipients

2015 ◽  
Vol 35 (suppl_1) ◽  
Author(s):  
Ali Javaheri ◽  
Payman Zamani ◽  
Maria Molina ◽  
Amrith Rodrigues ◽  
Susan Chambers ◽  
...  
Keyword(s):  
Cohort Study ◽  
Cholesterol Efflux ◽  
Density Lipoprotein ◽  
Hdl Cholesterol ◽  
Cardiac Transplant ◽  
Transplant Recipients ◽  
Single Center ◽  
Allograft Vasculopathy ◽  
Cholesterol Efflux Capacity ◽  
Transplant Patients

Background: Coronary allograft vasculopathy (CAV) is an important cause of mortality after cardiac transplantation. High density lipoprotein (HDL) cholesterol efflux capacity has been inversely associated with coronary artery disease and is impaired in cardiac transplant recipients. We performed a single center case-cohort study to test the hypothesis that reduced efflux capacity is a risk factor for mortality and a second, multi-center retrospective study to test if efflux capacity is associated with CAV progression. Methods: We designed a single center case-cohort study in which we identified cardiac transplant patients who died between 2009-2012 (cases, n=34) and controls as cardiac transplant patients who were alive as of the fourth quarter of 2013 (n=57). Efflux capacity was measured by incubating apolipoprotein B-depleted serum with macrophages in a validated ex vivo system. In a second study, we utilized pre-transplant samples from the Clinical Trials in Organ Transplantation 5 (CTOT5) study to determine the association between ATP-binding-cassette (ABC) A1-dependent cholesterol efflux and CAV progression at 1 year. Results: In our single center study, the average time from transplant to study entry was well-matched between cases and controls (7.6±1.0 vs 7.7±0.8 years, respectively, p=0.48). Multivariable Cox proportional hazard ratios demonstrated that higher levels of HDL cholesterol efflux capacity were associated with survival (HR 0.61, 95% CI 0.43-0.85), even after adjustment for HDL cholesterol mass. To determine whether excess mortality observed in subjects with reduced efflux could be attributable to CAV progression, we tested the relationship between intravascular ultrasound (IVUS) progression of CAV and cholesterol efflux capacity using linear regression. ABCA1-dependent efflux and IVUS progression were significantly associated (β = -0.90, 95% CI [-1.73 - -0.07], p = 0.037, R2 = 0.37). Conclusion: Reduced efflux capacity is an important mediator of CAV progression and mortality in cardiac transplant recipients. This finding suggests that interventions to increase HDL cholesterol efflux capacity may provide clinical benefit in cardiac transplant recipients.

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