Method for measuring the activities of cholesteryl ester transfer protein (lipid transfer protein)

Epps, Dennis E. Van; Harris, Jessica; Greenlee, K.A.; Fisher, Jed F.; Marschke, Charles K.; Castle, Christine K.; Ulrich, Roger G.; Moll, Th.; Melchior, George W.; Kézdy, Ferenc J.

doi:10.1016/0009-3084(95)02454-q

Cited by 7 publications

(6 citation statements)

References 17 publications

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“…Gates were set to exclude nonviable cells as determined by propidium iodide staining. Cholesteryl ester uptake was quantified as previously described [30]; in brief, samples were excited at 490 nm and fluorescence was measured at 530 nm (554 and 571 for Dil-AcLDL), uptake was quantified by subtracting median fluorescence of control cells from median fluorescence of cells treated with bodipy-cholesteryl ester or Dil-AcLDL.…”

Section: Methodsmentioning

confidence: 99%

Phagocytosis of Cholesteryl Ester Is Amplified in Diabetic Mouse Macrophages and Is Largely Mediated by CD36 and SR-A

et al. 2007

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Type 2 diabetes (T2D) is associated with accelerated atherosclerosis, which accounts for approximately 75% of all diabetes-related deaths. Here we investigate the link between diabetes and macrophage cholesteryl ester accumulation. When diabetic (db/db) mice are given cholesteryl ester intraperitoneally (IP), peritoneal macrophages (PerMΦs) recovered from these animals showed a 58% increase in intracellular cholesteryl ester accumulation over PerMΦs from heterozygote control (db/+) mice. Notably, PerMΦ fluid-phase endocytosis and large particle phagocytosis was equivalent in db/+and db/db mice. However, IP administration of CD36 and SR-A blocking antibodies led to 37% and 25% reductions in cholesteryl ester accumulation in PerMΦ. Finally, in order to determine if these scavenger receptors (SRs) were part of the mechanism responsible for the increased accumulation of cholesteryl esters observed in the diabetic mouse macrophages, receptor expression was quantified by flow cytometry. Importantly, db/db PerMΦs showed a 43% increase in CD36 expression and an 80% increase in SR-A expression. Taken together, these data indicate that direct cholesteryl ester accumulation in mouse macrophages is mediated by CD36 and SR-A, and the magnitude of accumulation is increased in db/db macrophages due to increased scavenger receptor expression.

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Section: Methodsmentioning

confidence: 99%

Phagocytosis of Cholesteryl Ester Is Amplified in Diabetic Mouse Macrophages and Is Largely Mediated by CD36 and SR-A

et al. 2007

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“…Powdered hen egg L-a-phosphatidylcholine was purchased from Avanti Polar Lipids. Triolein, 1,3-diolein, and oleic acid were from Sigma Chemical Co. Hexabromotriolein and Bodipy-TG were synthesized as described previously (Epps et al, 1995). [cholesteryl-4-I4C]Cholesteryl oleate, 45-60 mCi/mmol, and triolein, [9,10-3H(N)]-[CH3(CH2)7CH=CH-(CH2)7C0]303C3H5, 10-30 Ci/mmol, were purchased from New England Nuclear.…”

Section: Methodsmentioning

confidence: 99%

“…CETP activity was measured both by a fluorescence assay (Epps et al, 1995) and by a radioactivity assay (Pape et al, 1991). Inhibitors were added to the reaction mixtures from stock concentrated DMSO solutions.…”

Section: Methodsmentioning

confidence: 99%

“…We have utilized assays for measuring both inhibition of the transfer reaction and for identifying agents that are other than competitive. A continuous recording fluorescence assay (Epps et al, 1995) was used to monitor lipid transfer mediated by CETP from cynomolgus monkeys (Sarcich et al, 1995). Kinetic equations were derived for analyzing the donor and acceptor dependencies of lipid transfer.…”

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confidence: 99%

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Kinetics and Inhibition of Lipid Exchange Catalyzed by Plasma Cholesteryl Ester Transfer Protein (Lipid transfer protein)

Epps

Greenlee²,

Harris³

et al. 1995

Biochemistry

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The cholesteryl ester transfer protein-catalyzed cholesteryl ester transfer is inhibited by two compounds identified by a large-scale screening of cholesterol backbone-containing molecules. Kinetic analysis shows that U-95,594, an amino steroid, inhibits competitively the cholesteryl ester transfer protein-catalyzed transfer of both cholesteryl esters and triglycerides, as well from high-density lipoproteins as from synthetic microemulsions. In contrast, U-617, an organomercurial derivative of cholesterol, inhibits competitively the transfer of cholesteryl ester from either donor but is without any effect on triglyceride transfer. In addition to the rapid, competitive inhibition of cholesteryl ester transfer, U-617 also slowly and reversibly reacts with cholesteryl ester transfer protein to produce an additional 10-fold decrease in cholesteryl ester transfer activity but, again, without effect on triglyceride transfer.

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“…In order to precisely model these processes, detailed knowledge of their main characteristics, including rate constants, diffusion coefficients, viscosity, thermal conductivity and others, is required; these data are however largely absent from the literature. For this reason, we modelled the exchange and transfer processes as first-order concentrationdependent kinetics, consistent with established first-order kinetics of FC and PL transfer across lipoproteins [37][38][39]. When rate constants were unknown, they were modelled as time-or concentration-dependent parameters as long as such dependences were established elsewhere.…”

Section: Metabolic Processesmentioning

confidence: 97%

Mathematical Modelling of Material Transfer to High-Density Lipoprotein (HDL) upon Triglyceride Lipolysis by Lipoprotein Lipase: Relevance to Cardioprotective Role of HDL

Schekatolina¹,

Lahovska²,

Bekshaev

et al. 2022

Metabolites

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High-density lipoprotein (HDL) contributes to lipolysis of triglyceride-rich lipoprotein (TGRL) by lipoprotein lipase (LPL) via acquirement of surface lipids, including free cholesterol (FC), released upon lipolysis. According to the reverse remnant-cholesterol transport (RRT) hypothesis recently developed by us, acquirement of FC by HDL is reduced at both low and extremely high HDL concentrations, potentially underlying the U-shaped relationship between HDL-cholesterol and cardiovascular disease. Mechanisms underlying impaired FC transfer however remain indeterminate. We developed a mathematical model of material transfer to HDL upon TGRL lipolysis by LPL. Consistent with experimental observations, mathematical modelling showed that surface components of TGRL, including FC, were accumulated in HDL upon lipolysis. The modelling successfully reproduced major features of cholesterol accumulation in HDL observed experimentally, notably saturation of this process over time and appearance of a maximum as a function of HDL concentration. The calculations suggested that the both phenomena resulted from competitive fluxes of FC through the HDL pool, including primarily those driven by FC concentration gradient between TGRL and HDL on the one hand and mediated by lecithin-cholesterol acyltransferase (LCAT) and cholesteryl ester transfer protein (CETP) on the other hand. These findings provide novel opportunities to revisit our view of HDL in the framework of RRT.

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Method for measuring the activities of cholesteryl ester transfer protein (lipid transfer protein)

Cited by 7 publications

References 17 publications

Phagocytosis of Cholesteryl Ester Is Amplified in Diabetic Mouse Macrophages and Is Largely Mediated by CD36 and SR-A

Phagocytosis of Cholesteryl Ester Is Amplified in Diabetic Mouse Macrophages and Is Largely Mediated by CD36 and SR-A

Kinetics and Inhibition of Lipid Exchange Catalyzed by Plasma Cholesteryl Ester Transfer Protein (Lipid transfer protein)

Mathematical Modelling of Material Transfer to High-Density Lipoprotein (HDL) upon Triglyceride Lipolysis by Lipoprotein Lipase: Relevance to Cardioprotective Role of HDL

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