Human Cholesteryl Ester Transfer Protein Research Articles

SiRNA-induced liver ApoB knockdown lowers serum LDL-cholesterol in a mouse model with human-like serum lipids

Increased serum apolipoprotein (apo)B and associated LDL levels are well-correlated with an increased risk of coronary disease. ApoE⁻/⁻ and low density lipoprotein receptor (LDLr)⁻/⁻ mice have been extensively used for studies of coronary atherosclerosis. These animals show atherosclerotic lesions similar to those in humans, but their serum lipids are low in apoB-containing LDL particles. We describe the development of a new mouse model with a human-like lipid profile. Ldlr CETP⁺/⁻ hemizygous mice carry a single copy of the human CETP transgene and a single copy of a LDL receptor mutation. To evaluate the apoB pathways in this mouse model, we used novel short-interfering RNAs (siRNA) formulated in lipid nanoparticles (LNP). ApoB siRNAs induced up to 95% reduction of liver ApoB mRNA and serum apoB protein, and a significant lowering of serum LDL in Ldlr CETP⁺/⁻ mice. ApoB targeting is specific and dose-dependent, and it shows lipid-lowering effects for over three weeks. Although specific triglycerides (TG) were affected by ApoB mRNA knockdown (KD) and the total plasma lipid levels were decreased by 70%, the overall lipid distribution did not change. Results presented here demonstrate a new mouse model for investigating additional targets within the ApoB pathways using the siRNA modality.

The conformation and CETP inhibitory activity of [10]-dehydrogingerdione isolated from Zingiber officinale.

In the course of searching for cholesteryl ester transfer protein (CETP) inhibitors from natural sources, a new type of CETP inhibitor, [10]-dehydrogingerdione (1), was isolated from the extract of rhizomes of Zingiber officinale Roscoe. By NMR spectroscopic analysis of its (1)HNMR, (13)C-NMR, and (1)H-(1)H COSY, HMBC, HMQC and NOESY, more precise structure, compared with its originally proposed structures, of [10]-dehydrogingerdione has been elucidated. This active compound inhibited human plasma CETP with IC(50) values of 35 μM.

Aerobic Exercise Improves Reverse Cholesterol Transport in Cholesteryl Ester Transfer Protein Transgenic Mice

We analyzed the effect of a 6-week aerobic exercise training program on the in vivo macrophage reverse cholesterol transport (RCT) in human cholesteryl ester transfer protein (CETP) transgenic (CETP-tg) mice. Male CETP-tg mice were randomly assigned to a sedentary group or a carefully supervised exercise training group (treadmill 15 m/min, 30 min sessions, five sessions per week). The levels of plasma lipids were determined by enzymatic methods, and the lipoprotein profile was determined by fast protein liquid chromatography (FPLC). CETP activity was determined by measuring the transfer rate of ¹⁴C-cholesterol from HDL to apo-B containing lipoproteins, using plasma from CETP-tg mice as a source of CETP. The reverse cholesterol transport was determined in vivo by measuring the [³H]-cholesterol recovery in plasma and feces (24 and 48 h) and in the liver (48 h) following a peritoneal injection of [³H]-cholesterol labeled J774-macrophages into both sedentary and exercise trained mice. The protein levels of liver receptors were determined by immunoblot, and the mRNA levels for liver enzymes were measured using RT-PCR. Exercise training did not significantly affect the levels of plasma lipids or CETP activity. The HDL fraction assessed by FPLC was higher in exercise-trained compared to sedentary mice. In comparison to the sedentary group, a greater recovery of [³H]-cholesterol from the injected macrophages was found in the plasma, liver and feces of exercise-trained animals. The latter occurred even with a reduction in the liver CYP7A1 mRNA level in exercised trained animals. Exercise training increased the liver LDL receptor and ABCA-1 protein levels, although the SR-BI protein content was unchanged. The RCT benefit in CETP-tg mice elicited by exercise training helps to elucidate the role of exercise in the prevention of atherosclerosis in humans.

Structure and Function of Cholesteryl Ester Transfer Protein in the Tree Shrew

Cholesteryl ester transfer protein (CETP) plays an important role in reverse cholesterol transport (RCT). To study on the structure and function of CETP in the tree shrew, a kind of animal resistant to atherosclerosis, we completed the cloning of the full-length tree-shrew CETP cDNA sequence based on the reported partial sequence. The full-length cDNA of tree shrew CETP was 1,704bp and the deduced protein of the cDNA showed a sequence identity of 81, 80 and 74%, respectively, with the human, monkey and rabbit CETP. The level of CETP mRNA in the liver was much more abundant than that in the other tissues. A mutant protein with a substitution of Asn at position 110 by Gln was found to possess an impaired secretion property compared with the wild-type tree shrew CETP. The mutant proteins, respectively, with a substitution of Pro at position 344 by Ser and a substitution of Gln at position 452 by Arg displayed similar secretion ability, but a decreased cholesteryl ester transfer capability compared with the wild type (48 and 26% lower, respectively). These findings demonstrate that liver is the main tissue synthesizing CETP in the tree shrew. Asn at position 110 plays an important role in the secretion of tree shrew CETP. The residues at position 344 and 452 play essential roles in cholesteryl ester transferring process.

A PCSK9-binding antibody that structurally mimics the EGF(A) domain of LDL-receptor reduces LDL cholesterol in vivo1[S

Proprotein convertase subtilisin-like/kexin type 9 (PCSK9) regulates LDL cholesterol levels by inhibiting LDL receptor (LDLr)-mediated cellular LDL uptake. We have identified a fragment antigen-binding (Fab) 1D05 which binds PCSK9 with nanomolar affinity. The fully human antibody 1D05-IgG2 completely blocks the inhibitory effects of wild-type PCSK9 and two gain-of-function human PCSK9 mutants, S127R and D374Y. The crystal structure of 1D05-Fab bound to PCSK9 reveals that 1D05-Fab binds to an epitope on the PCSK9 catalytic domain which includes the entire LDLr EGF(A) binding site. Notably, the 1D05-Fab CDR-H3 and CDR-H2 loops structurally mimic the EGF(A) domain of LDLr. In a transgenic mouse model (CETP/LDLr-hemi), in which plasma lipid and PCSK9 profiles are comparable to those of humans, 1D05-IgG2 reduces plasma LDL cholesterol to 40% and raises hepatic LDLr protein levels approximately fivefold. Similarly, in healthy rhesus monkeys, 1D05-IgG2 effectively reduced LDL cholesterol 20%-50% for over 2 weeks, despite its relatively short terminal half-life (t(1/2) = 3.2 days). Importantly, the decrease in circulating LDL cholesterol corresponds closely to the reduction in free PCSK9 levels. Together these results clearly demonstrate that the LDL-lowering effect of the neutralizing anti-PCSK9 1D05-IgG2 antibody is mediated by reducing the amount of PCSK9 that can bind to the LDLr.

Point: The Relationship between Cholesteryl Ester Transfer Protein and Cardiovascular Risk

There is overwhelming epidemiologic evidence that increased concentrations of LDL cholesterol (LDL-C)2) and low concentrations of HDL cholesterol (HDL-C) are independently associated with increased cardiovascular risk. Furthermore, the risk associated with low HDL-C concentrations persists even when LDL-C has been reduced to concentrations <1.8 mmol/L (<70 mg/dL) by treatment with statins (1). It is logical, therefore, to investigate strategies designed to redistribute cholesterol away from atherogenic LDL particles in favor of potentially protective HDL particles. One such approach is to inhibit the cholesteryl ester transfer protein (CETP), a protein in human plasma that transfers cholesterol from the HDL to non-HDL fractions (2). CETP activity is prominent in humans, nonhuman primates, and rabbits (2). In contrast, there is little or no CETP activity in the plasma of most other species, including rodents (2). In general, species with CETP activity in plasma are susceptible to the development of atherosclerosis, whereas those lacking the protein tend to be naturally resistant. Furthermore, inhibiting CETP in rabbits markedly reduces the development of atherosclerosis, regardless of whether the CETP inhibition is achieved by genetic manipulation (3), by vaccination against CETP (4), or by chemical inhibitors of CETP (5). ### CETP POLYMORPHISMS AND SUSCEPTIBILITY TO ATHEROSCLEROSIS IN HUMANS Several polymorphisms of the human CETP (cholesteryl ester transfer protein, plasma) gene have been reported (2). Although many of these studies have demonstrated associations between CETP single-nucleotide polymorphisms and small changes in plasma CETP and HDL concentrations, the relationship between these polymorphisms and susceptibility to atherosclerosis is variable (2). A recent metaanalysis of 92 studies involving 113 833 participants (6) concluded, however, that those CETP genotypes associated with inhibition of CETP activity show an inverse association with coronary risk and that the magnitude of the reduction of coronary events was consistent with the reductions in risk for equivalent increases in HDL-C concentration as predicted from …

P260 CETP EXPRESSION REVERSES THE RECONSTITUTED HDL-INDUCED INCREASE IN VLDL

Human data suggest that reconstituted HDL (rHDL) infusion can induce atherosclerosis regression. Studies in mice indicated that rHDL infusion adversely affects VLDL levels, but this effect is less apparent in humans. This discrepancy may be explained by the fact that humans, in contrast to mice, express cholesteryl ester transfer protein (CETP). The aim of this study was to investigate the role of CETP in the effects of rHDL on VLDL metabolism by using APOE*3-Leiden (E3L) mice, a well-established model for human-like lipoprotein metabolism. At 1 h after injection, rHDL increased plasma VLDL-C and TG in E3L mice, but not in E3L mice cross-bred onto a human CETP background (E3L.CETP mice). This initial raise in VLDL, caused by competition between rHDL and VLDL for LPLmediated TG hydrolysis, was thus prevented by CETP. At 24 h after injection, rHDL caused a second increase in VLDL-C and TG in E3L mice, whereas rHDL had even decreased VLDL in E3L.CETP mice. This secondary raise in VLDL was due to increased hepatic VLDL-TG production. Collectively, we conclude that CETP protects against the rHDL-induced increase in VLDL. We anticipate that studies evaluating the anti-atherosclerotic efficacy of rHDL in mice that are naturally deficient for CETP should be interpreted with caution, and that treatment of atherogenic dyslipidemia by rHDL should not be combined with agents that aggressively reduce CETP activity. Copyright © 2011 by the American Society for Biochemistry and Molecular Biology, Inc.

Restoration of High-Density Lipoprotein Levels by Cholesteryl Ester Transfer Protein Expression in Scavenger Receptor Class B Type I (SR-BI) Knockout Mice Does Not Normalize Pathologies Associated With SR-BI Deficiency

Disruption of scavenger receptor class B type I (SR-BI) in mice impairs high-density lipoprotein (HDL)-cholesterol (HDL-C) delivery to the liver and induces susceptibility to atherosclerosis. In this study, it was investigated whether introduction of cholesteryl ester transfer protein (CETP) can normalize HDL-C transport to the liver and reduce atherosclerosis in SR-BI knockout (KO) mice. Expression of human CETP in SR-BI(KO) mice resulted in decreased plasma HDL-C levels, both on chow diet (1.8-fold, P<0.001) and on challenge with Western-type diet (1.6-fold, P<0.01). Furthermore, the presence of CETP partially normalized the abnormally large HDL particles observed in SR-BI(KO) mice. Unexpectedly, expression of CETP in SR-BI(KO) mice did not reduce atherosclerotic lesion development, probably because of consequences of SR-BI deficiency, including the persistence of higher VLDL-cholesterol (VLDL-C) levels, unchanged elevated free cholesterol/total cholesterol ratio, and the increased oxidative status of the animals. In addition, CETP expression did not normalize other characteristics of SR-BI deficiency, including female infertility, reticulocytosis, thrombocytopenia, and impaired platelet aggregation. CETP restores HDL-C levels in SR-BI(KO) mice, but it does not change the susceptibility to atherosclerosis and other typical characteristics that are associated with SR-BI disruption. This may indicate that the pathophysiology of SR-BI deficiency is not a direct consequence of changes in the HDL pool.

Selective CETP Inhibition and PPARα Agonism Increase HDL Cholesterol and Reduce LDL Cholesterol in Human ApoB100/Human CETP Transgenic Mice

Cholesteryl ester transfer protein (CETP) plays a key role in high-density lipoprotein (HDL) cholesterol metabolism, but normal mice are deficient in CETP. In this study, transgenic mice expressing both human apolipoprotein B 100 (ApoB-100) and human CETP (hApoB100/hCETP) were used to characterize the effects of CETP inhibition and peroxisome proliferator-activated receptor alpha (PPARalpha) agonism on lipid profiles. Torcetrapib (3, 10, and 30 mg/kg), a CETP inhibitor, fenofibrate (30 mg/kg), a weak PPARalpha agonist, and GW590735 (3 and 10 mg/kg), a potent and selective PPARalpha agonist were given orally for 14 days to hApoB100/hCETP mice and lipid profiles were assessed. The average percentages of HDL, low-density lipoprotein (LDL), and very-low-density lipoprotein (VLDL) cholesterol fractions in hApoB100/hCETP mice were 34.8%, 61.6%, and 3.6%, respectively, which is similar to those of normolipidemic humans. Both torcetrapib and fenofibrate significantly increased HDL cholesterol and reduced LDL cholesterol, and there was a tendency for torcetrapib to reduce VLDL cholesterol and triglycerides. GW590735 significantly increased HDL cholesterol, decreased LDL and VLDL cholesterol, and significantly reduced triglycerides. Maximal increases in HDL cholesterol were 37%, 53%, and 84% with fenofibrate, torcetrapib, and GW590735, respectively. These results, in mice that exhibit a more human-like lipid profile, demonstrate an improved lipid profile with torcetrapib, fenofibrate, and GW590735, and support the use of selective PPARalpha agonism for the treatment of lipid disorders. In addition, these data demonstrate the use of hApoB100/hCETP transgenic mice to identify, characterize, and screen compounds that increase HDL cholesterol.

Cholesteryl Ester Transfer Protein Penetrates Lipoproteins For Cholesteryl Ester Transfer

Human cholesteryl ester transfer protein (CETP), a hydrophobic glycoprotein (476 amino acids, ∼74 kDa), mediates the transfer of cholesteryl ester (CE) from high-density lipoproteins (HDL) to apo-B containing lipoproteins (low-density lipoproteins - LDL; very low density lipoproteins - VLDL). The crystal structure of CETP reveals a banana-shaped protein consisting of N- and C-terminal β-barrel domains, a central β-sheet, and a ∼60-A-long hydrophobic cavity. Since CETP deficiency is associated with elevated ‘good-cholesterol’ (HDL-associated) levels via delayed catabolism, as a result, CETP has become a high interest pharmacological drug target.Here, we investigated the mechanism of CETP-mediated neutral lipid transfer by electron microscopy (EM), image-processing, and molecular dynamic simulations (MDS). We discovered that: 1) CETP bridges HDL and LDL/VLDL to form a ternary complex; 2) the CETP C-terminal beta-barrel domain penetrates partially into the LDL/VLDL surface; 3) the N-terminal beta-barrel domain penetrates through the HDL surface and interacts with the CE core; and 4) MDS analysis suggested that the penetrating regions of CETP are highly mobile and form pores that connect to cavities running through the central axis of the CETP molecule to generate a tunnel. The tunnel was mainly hydrophobic and connected the distal ends through its central cavities. Thus, we proposed CETP resembles a Chinese finger trap, in which both beta-barrel domains penetrate the lipoprotein surface, resulting in pressure from both ends. The pressure may result in the anti-twisting of the two domains, leading to opening of the tunnel for CE transfer between HDL and LDL. The distal end of N-terminal beta-barrel domain could be particularly important for the design of pharmacological inhibitors, as blockage of the N-terminal beta-barrel domain would prevent CE transfer with a low probability of off target effects on normal HDL metabolism.

Different Binding Modes of Compounds Affecting CETP Activity: Dalcetrapib and Torcetrapib

Cholesteryl ester transfer protein (CETP) is a hydrophobic glycoprotein in plasma and facilitates the exchange of neutral lipids between various lipoproteins resulting in a net transport of cholesteryl ester from HDL to LDL. Decrease in CETP activity leads to an increase of HDL cholesterol which potentially reduces cardiovascular risk. Human recombinant CETP (rhCETP) was purified to homogeneity by hydrophobic interaction chromatography and size exclusion chromatography (SEC). The protein showed an extraordinarily high thermodynamic stability against thermal as well as denaturant-induced unfolding. The effects of CETP on lipoprotein profiles were characterized after in-vitro incubation of plasma with rhCETP followed by SEC and reverse phase protein arrays. Monitoring the distribution of apolipoproteins and the cholesterol profiles confirmed the transfer of cholesteryl ester from HDL to LDL induced by CETP. In the presence of the potent CETP inhibitor torcetrapib (1 µM) added CETP is bound to lipoproteins while free CETP can still be detected in presence of dalcetrapib (3 µM), a compound from a different chemical class. 14C-torcetrapib and 14C-dalcetrapib were bound to CETP immobilized on sepharose and incubated with excess of unlabelled compounds. 14C-dalcetrapib bound as Dalcetrapib-thiol could only be displaced in the presence of reducing agent by an excess of unlabeled dalcetrapib but not by torcetrapib. Dalcetrapib did not compete for binding of 14C-torcetrapib. Interaction studies with Surface Plasmon Resonance confirmed reversible covalent binding of dalcetrapib-thiol and different binding sites for dalcetrapib and torcetrapib. In agreement with literature cysteine 13 of CETP was identified as attachment point of dalcetrapib-thiol. All data suggested that torcetrapib and dalcetrapib bind to different sites on CETP which may be related to differences observed in their pharmacological profiles.

CETP does not affect triglyceride production or clearance in APOE*3-Leiden mice

The cholesteryl ester transfer protein (CETP) facilitates the bidirectional transfer of cholesteryl esters and triglycerides (TG) between HDL and (V)LDL. By shifting cholesterol in plasma from HDL to (V)LDL in exchange for VLDL-TG, CETP aggravates atherosclerosis in hyperlipidemic APOE*3-Leiden (E3L) mice. The aim of this study was to investigate the role of CETP in TG metabolism and high-fat diet-induced obesity by using E3L mice with and without the expression of the human CETP gene. On chow, plasma lipid levels were comparable between both male and female E3L and E3L.CETP mice. Further mechanistic studies were performed using male mice. CETP expression increased the level of TG in HDL. CETP did not affect the postprandial plasma TG response or the hepatic VLDL-TG and VLDL-apolipoprotein B production rate. Moreover, CETP did not affect the plasma TG clearance rate or organ-specific TG uptake after infusion of VLDL-like emulsion particles. In line with the absence of an effect of CETP on tissue-specific TG uptake, CETP also did not affect weight gain in response to a high-fat diet. In conclusion, the CETP-induced increase of TG in the HDL fraction of E3L mice is not associated with changes in the production of TG or with tissue-specific clearance of TG from the plasma.

Pharmacological inhibition of ABCA1 degradation increases HDL biogenesis and exhibits antiatherogenesis

Expression of ABCA1 is regulated by transcription of the gene and calpain-mediated proteolytic degradation, and inhibition ABCA1 degradation results in increased ABCA1 and HDL biogenesis in vitro. We examined whether this approach could be a potential antiatherogenic treatment. Although probucol inhibits both the activity and degradation of ABCA1, its oxidized products, spiroquinone and diphenoquinone, reduce degradation of ABCA1 without inhibiting its activity or altering transcription of the ABCA1 gene. Accordingly, both compounds enhanced apolipoprotein A-I/ABCA1-dependent generation of HDL in vitro, and increased hepatic ABCA1 and plasma HDL without increasing antioxidant activity in plasma when given to rabbits. Both compounds also decreased vascular lipid deposition in cholesterol-fed rabbits. We therefore conclude that stabilization of ABCA1 against calpain-mediated degradation is a novel and potentially important strategy to increase HDL formation and prevent atherosclerosis. Spiroquinone and diphenoquinone are potential seeds for development of such drugs.

Liver X Receptor–Mediated Induction of Cholesteryl Ester Transfer Protein Expression Is Selectively Impaired in Inflammatory Macrophages

Cholesteryl ester transfer protein (CETP) is a target gene for the liver X receptor (LXR). The aim of this study was to further explore this regulation in the monocyte-macrophage lineage and its modulation by lipid loading and inflammation, which are key steps in the process of atherogenesis. Exposure of bone marrow-derived macrophages from human CETP transgenic mice to the T0901317 LXR agonist increased CETP, PLTP, and ABCA1 mRNA levels. T0901317 also markedly increased CETP mRNA levels and CETP production in human differentiated macrophages, whereas it had no effect on CETP expression in human peripheral blood monocytes. In inflammatory mouse and human macrophages, LXR-mediated CETP gene upregulation was inhibited, even though ABCA1, ABCG1, and SREBP1c inductions were maintained. The inhibition of CETP gene response to LXR agonists in inflammatory cells was independent of lipid loading (ie, oxidized LDL increased CETP production in noninflammatory macrophages with a synergistic effect of synthetic LXR agonists). LXR-mediated induction of human CETP expression is switched on during monocyte-to-macrophage differentiation, is magnified by lipid loading, and is selectively lost in inflammatory macrophages, which suggests that inflammatory cells may not increase the circulating CETP pool on LXR agonist treatment.

Effects of cholesteryl ester transfer protein inhibition on apolipoprotein A-II-containing HDL subspecies and apolipoprotein A-II metabolism

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.

Scavenger receptor class B type I-mediated uptake of serum cholesterol is essential for optimal adrenal glucocorticoid production

Impaired scavenger receptor class B type I (SR-BI)-mediated uptake of HDL-cholesterol esters (HDL-CE) induces adrenal insufficiency in mice. Humans contain an alternative route of HDL-CE clearance, namely through the transfer by cholesteryl ester transfer protein (CETP) to apolipoprotein B lipoproteins for subsequent uptake via the LDL receptor. In this study, we determined whether CETP can compensate for loss of adrenal SR-BI. Transgenic expression of human CETP (CETP Tg) in SR-BI knockout (KO) mice increased adrenal HDL-CE clearance from 33-58% of the control value. SR-BI KO/CETP Tg and SR-BI KO mice displayed adrenal hypertrophy due to equally high plasma adrenocorticotropic hormone levels. Adrenal cholesterol levels and plasma corticosterone levels were 38-52% decreased in SR-BI KO mice with and without CETP expression. SR-BI KO/CETP Tg mice also failed to increase their corticosterone level after lipopolysaccharide challenge, leading to an identical >4-fold increased tumor necrosis factor-alpha response compared with controls. These data indicate that uptake of CE via other routes than SR-BI is not sufficient to generate the cholesterol pool needed for optimal adrenal steroidogenesis. In conclusion, we have shown that CETP-mediated transfer of HDL-CE is not able to reverse adrenal insufficiency in SR-BI knockout mice. Thus, SR-BI-mediated uptake of serum cholesterol is essential for optimal adrenal function.

Cholesteryl ester transfer protein (CETP) increases postprandial triglyceridaemia and delays triacylglycerol plasma clearance in transgenic mice

The CETP (cholesteryl ester transfer protein) is a plasma protein synthesized in several tissues, mainly in the liver; CETP reduces plasma HDL (high-density lipoprotein) cholesterol and increases the risk of atherosclerosis. The effect of CETP levels on postprandial intravascular metabolism of TAGs (triacylglycerols) is an often-overlooked aspect of the relationship between CETP and lipoprotein metabolism. Here, we tested the hypothesis that CETP delays the plasma clearance of TAG-rich lipoprotein by comparing human CETP expressing Tg (transgenic) and non-Tg mice. After an oral fat load, the postprandial triglyceridaemia curve was markedly increased in CETP-Tg compared with non-Tg mice (280+/-30 versus 190+/-20 mg/dl per 6 h respectively, P<0.02). No differences in intestinal fat absorption and VLDL (very-low-density lipoprotein) secretion rates were observed. Kinetic studies of double-labelled chylomicron-like EMs (emulsions) showed that both [(3)H]triolein and [(14)C]cholesteryl oleate FCRs (fractional clearance rates) were significantly reduced ( approximately 20%) in CETP-Tg mice. Furthermore, TAG from lipid EM pre-incubated with CETP-Tg plasma had plasma clearance and liver uptake significantly lower than the non-Tg plasma-treated lipid EM. In addition, reductions in post-heparin plasma LPL (lipoprotein lipase) activity (50%) and adipose tissue mRNA abundance (39%) were verified in CETP-Tg mice. Therefore we conclude that CETP expression in Tg mice delays plasma clearance and liver uptake of TAG-rich lipoproteins by two mechanisms: (i) transferring TAG to HDLs and increasing CE content of the remnant particles and (ii) by diminishing LPL expression. These findings show that the level of CETP expression can influence the responsiveness to dietary fat and may lead to fat intolerance.

Blood Lipid-Lowering and Antioxidant Effects of a Structured Lipid Containing Monoacylglyceride Enriched with Monounsaturated Fatty Acids in C57BL/6 Mice

We recently reported that a synthetic edible oil-containing monoacylglyceride (MAG) and diacylglyceride (DAG) exerted anti-atherosclerotic effects. In order to further investigate the activities and individual effects of MAG and DAG on the atherosclerotic process, we prepared a structured oil with various MAG and DAG contents and tested them both in vitro and in vivo, using C57BL/6 mice. The structured oil to be tested was mixed (final concentration 5%, wt/wt) with a high-cholesterol high-fat diet (1.2% cholesterol/15% fat/0.5% sodium cholate) and provided to the mice for 7 weeks. After administration, the mice consuming MAG97%-oil and DAG50%/MAG10%-oil evidenced 17% and 24% decreases in plasma total cholesterol (TC) level, respectively, as compared to a group of mice fed on lard. The experimental mice also had reduced plasma triglyceride concentrations and elevated high-density lipoprotein-cholesterol to TC ratios, by up to 31% in the case of the DAG50%/MAG10%-oil fed mice. The mice fed on MAG97%-oil exhibited elevated plasma antioxidant activity and lecithin:cholesterol acyltransferase activity. Histological assessments of the livers of the mice showed that the consumption of MAG-containing oil attenuated the adhesion of inflammatory cells and also ameliorated fatty liver changes, as compared to what was observed in the case of DAG85%-oil consumption. In conclusion, the MAG-containing oil exhibited anti-inflammatory and antioxidant activities in vivo, as well as in vitro inhibitory activity against human cholesteryl ester transfer protein. These results provide us with new insights into MAG-containing oil in terms of hypocholesterolemic effects and antioxidant activities.

Cholesteryl Ester Transfer Protein (CETP) Inhibitors

Epidemiological studies have demonstrated an inverse correlation between plasma concentrations of high-density lipoprotein cholesterol (HDL-C) and incidence of coronary heart disease (CHD); thus new therapies for raising HDL-C levels have been the focus of significant efforts by the cardiovascular medicine community. Inhibition of cholesteryl ester transfer protein (CETP) is one approach to increasing HDL-C concentrations. CETP is a plasma glycoprotein that mediates the transfer of cholesteryl esters from HDL to the apoB-containing lipoproteins, with a balanced transfer of triglycerides. Inhibition of CETP results in an accumulation of cholesteryl esters in HDL, thus resulting in increased HDL-C. Pharmacological inhibition of CETP in humans has been shown to result in increased levels of HDL-C, although any beneficial effect of this inhibition on CHD has yet to be established. This review article will discuss the complex role of CETP in lipid metabolism, recent developments for small-molecule inhibitors of CETP, and future prospects for CETP inhibitors in the treatment of CHD.

The role of plasma lipid transfer proteins in lipoprotein metabolism and atherogenesis

The plasma lipid transfer proteins promote the exchange of neutral lipids and phospholipids between the plasma lipoproteins. Cholesteryl ester transfer protein (CETP) facilitates the removal of cholesteryl esters from HDL and thus reduces HDL levels, while phospholipid transfer protein (PLTP) promotes the transfer of phospholipids from triglyceride-rich lipoproteins into HDL and increases HDL levels. Studies in transgenic mouse models and in humans with rare genetic deficiencies (CETP) or common genetic variants (CETP and PLTP) highlight the central role of these molecules in regulating HDL levels. Human CETP deficiency is associated with dramatic elevations of HDL cholesterol and apolipoprotein A-I levels, while PLTP variants with increased expression are associated with higher HDL levels. A recent meta-analysis suggests that common CETP alleles causing reduced CETP and increased HDL levels are associated with reduced coronary heart disease. The failure of a clinical trial with the CETP inhibitor torcetrapib may have been related in part to off-target toxicity. Ongoing phase 3 clinical trials with other CETP inhibitors may help to clarify if this strategy can ultimately be successful in the treatment of atherosclerosis.