Human Cholesterol Ester Transfer Protein Research Articles

Choline Supplementation Does Not Promote Atherosclerosis in CETP-Expressing Male Apolipoprotein E Knockout Mice

Dietary trimethylamines, such as choline, metabolized by intestinal microbiota to trimethylamine are absorbed by the gut and oxidized to trimethylamine N-oxide (TMAO). The objective of this study was to determine the effect of choline supplementation on atherosclerosis progression in Apoe−/− mice expressing human cholesterol ester transfer protein (hCETP) using the same diets as in previously reported studies. Mice expressing hCETP, after transfection with AAV2/8-hCETP, were fed an 18% protein diet with either 0.09% (standard chow), 0.5% or 1% choline for 16 weeks. Control mice not transfected with hCETP were fed 1% choline. Dietary choline supplementation increased plasma TMAO levels at 8 and 16 weeks. When atherosclerotic lesions were measured in the thoracic aorta and aortic root, there were no differences between any of the treatment groups in the amount of plaque development at either site. Throughout the study, no significant changes in plasma lipids or major classes of lipoproteins were observed in hCETP-expressing mice. Plasma-oxidized low density lipoprotein, myeloperoxidase and high density lipoprotein inflammatory index were measured at 16 weeks, with no significant changes in any of these inflammatory markers between the four treatment groups. Despite increasing plasma TMAO levels, dietary choline supplementation in Apoe−/− mice expressing hCETP did not promote atherosclerosis.

Atherosclerosis Regression and Cholesterol Efflux in Hypertriglyceridemic Mice.

[Figure: see text].

A15374 Genetic Variant of Human Cholesterol Ester Transfer Protein (CETP) Raises HDL levels and Is Associated with Increased Risk for Large Vessel and Small Vessel Stroke in Chinese Patients

Objectives: Cholesterol Ester Transfer Protein (CETP) mediates lipids transfer among lipoprotein particles, affects HDL levels in general population. The CETP deficiency mutation at CETP (442 D:G) raises HDL level. We set to access the association of raised HDL level and all stroke and major stroke subtypes in Chinese population. Methods: We carried out a case control study: INTERSTROKE-China study, with 3500 stroke cases and 3834 age/gender matched controls. Structured questionnaires were administered to each stroke cases and matched controls, and information of biometrics, socioeconomic status, lifestyle, biomarkers, etc, was obtained. Genotyping of the CETP deficiency mutation (CETP rs2303790) SNP was performed by the TaqMan Real Time PCR method. Results: We found that CETP G allele is associated with higher HDL levels (1.30 vs. 1.18 mmol, p = 7.8E-8 dominant), but not associated with all stroke OR 1.17 (95%CI; 0.95–1.46), even after adjustment for HDL level. In subtypes of stroke with CETP G allele, ischemic stroke showed positive association with OR 1.33 (1.05–1.69), but not hemorrhagic stroke with OR 0.99. We further test subtypes of ischemic stroke, and we found positive association of large vessel with OR 1.72 (1.06–2.62) and small vessel stroke with OR 1.38 (1.05–1.82) respectively. Borderline positive association was seen for subtypes of stroke with atrial fibrillation and cardiac cause. No hint of association was seen for encryptic stroke or cardioembolism. Conclusion: CETP genetic variant is association with raised HDL, but increased risk for large vessel and small vessel stroke, even after full adjustment for other stroke risk factors. We conclude that CETP variants raise HDL level also increase the risk for large and small vessel subtypes of ischemic stroke.

Harnessing a Physiologic Mechanism for siRNA Delivery With Mimetic Lipoprotein Particles

Therapeutics based on RNA interference (RNAi) have emerged as a potential new class of drugs for treating human disease by silencing the target messenger RNA (mRNA), thereby reducing levels of the corresponding pathogenic protein. The major challenge for RNAi therapeutics is the development of safe delivery vehicles for small interfering RNAs (siRNAs). We previously showed that cholesterol-conjugated siRNAs (chol-siRNA) associate with plasma lipoprotein particles and distribute primarily to the liver after systemic administration to mice. We further demonstrated enhancement of silencing by administration of chol-siRNA pre-associated with isolated high-density lipoprotein (HDL) or low-density lipoprotein (LDL). In this study, we investigated mimetic lipoprotein particle prepared from recombinant apolipoprotein A1 (apoA) and apolipoprotein E3 (apoE) as a delivery vehicle for chol-siRNAs. We show that apoE-containing particle (E-lip) is highly effective in functional delivery of chol-siRNA to mouse liver. E-lip delivery was found to be considerably more potent than apoA-containing particle (A-lip). Furthermore, E-lip–mediated delivery was not significantly affected by high endogenous levels of plasma LDL. These results demonstrate that E-lip has substantial potential as delivery vehicles for lipophilic conjugates of siRNAs.

Abstract 8817: Niacin Promotes Macrophage Reverse Cholesterol Transport in Hypercholesterolemic Mice In a CETP-Dependent Manner

Background- Niacin is the most effective drug currently available to increase HDL-cholesterol. The aim of the present study was to examine the effect of niacin on macrophage reverse cholesterol transport (RCT), a primary atheroprotective property of HDL, and to determine the mechanisms by which this occurs. Methods and Results- Niacin increased HDL-cholesterol in homozygous LDL receptor-deficient ( Ldlr -/- ) mice expressing human cholesterol ester transfer protein (CETP) ( Ldlr -/- . CETP ), but not in the mice without CETP. Niacin promoted macrophage RCT in the mice in a CETP-dependent manner. Treatment of Ldlr -/- . CETP mice with niacin decreased plasma CETP activity ex vivo , without the alteration of hepatic CETP expression or plasma CETP mass. Niacin also promoted macrophage RCT by reducing CETP activity even in the setting of partial lack of LDL receptor with CETP expression ( Ldlr +/- . CETP mice). Ldlr -/- . CETP mice and Ldlr +/- . CETP mice with niacin had the enhanced plasma capacity to promote macrophage cholesterol efflux. Moreover, niacin attenuated the transfer of cholesteryl esters from HDL to VLDL/LDL fractions in vivo , consistent with its inhibitory effect on CETP activity. Kinetic studies showed that niacin increased fecal excretion of HDL-derived cholesteryl esters in Ldlr -/- . CETP , but not in Ldlr -/- mice. Conclusions- Our observations demonstrate that niacin promotes macrophage RCT in vivo by decreasing plasma CETP activity in the setting of both complete and partial lack of LDL receptor. Niacin-mediated reduction in CETP activity leads to the enhanced direct pathway of RCT from HDL fractions into feces with the promotion of plasma efflux capacity from macrophages in the mice.

Peroxisome Proliferator‐Activated Receptor α Agonists Regulate Cholesterol Ester Transfer Protein

Peroxisome proliferator-activated receptor alpha (PPARalpha) belongs to the nuclear receptor superfamily that regulates multiple target genes involved in lipid metabolism. Cholesterol ester transfer protein (CETP) is a secreted glycoprotein that modifies high-density lipoprotein (HDL) particles. In humans, plasma CETP activity is inversely correlated with HDL cholesterol levels. We report here that PPARalpha agonists increase CETP mRNA, protein and accordingly its activity. In a human CETP transgenic animal model harboring the natural flanking regions (Jiang et al. in J Clin Investigat 90:1290-1295, 1992), both fenofibrate and a specific synthetic PPARalpha agonist LY970 elevated human CETP mRNA in liver, serum protein and CETP activity. In hamsters, the endogenous liver CETP mRNA level and the serum CETP activity were dose-dependently upregulated by fenofibrate. In addition Wy14643, a PPARalpha agonist, also significantly elevated CETP mRNA and activity. In a carcinoma cell line of hepatic origin, HepG2 cells, overexpression of PPARalpha resulted in increased CETP mRNA and agonist treatment further elevated CETP mRNA levels. We conclude that PPARalpha agonists upregulate CETP expression and activity and may play an important role in PPARalpha (agonist mediated HDL cholesterol homeostasis in humans.

Differential regulation of cholesterol homeostasis in transgenic mice expressing human cholesterol ester transfer proteinThis paper is one of a selection of papers published in this Special Issue, entitled The Cellular and Molecular Basis of Cardiovascular Dysfunction, Dhalla 70th Birthday Tribute.

We investigated whether expression of cholesterol ester transfer protein (CETP) in mice alters the regulation of cholesterol metabolism. Transgenic mice expressing human CETP (CETP-TG) and nontransgenic littermates (non-TG) were fed either a monounsaturated fatty acid (MUFA) or a saturated fatty acid (SFA)-rich diet in the presence or absence of cholesterol. Mice fed with MUFA diet had higher CETP activity compared with SFA-fed mice. Addition of cholesterol to the MUFA diet decreased CETP activity, whereas addition of cholesterol to the SFA diet had no effect. Cholesterol 7alpha-hydroxylase (Cyp7a) activity was higher in CETP-TG mice compared with non-TG mice when fed a MUFA diet, whereas SFA fed CETP-TG mice showed lower Cyp7a activity as compared with non-TG. Microsomal triglyceride transfer protein (MTTP) activity was higher in CETP-TG mice compared with non-TG mice when fed a MUFA diet. HMG-CoA reductase activity was lower in CETP-TG mice compared with non-TG mice when fed a MUFA or a SFA diet. These data demonstrate that the regulation of Cyp7a, HMG-CoA reductase, and MTTP is altered in CETP-TG mice as compared with non-TG mice and these alterations are further modulated by the quality of dietary fats. These findings highlight the importance of CETP in regulating cholesterol homeostasis.

Sterol upregulation of human CETP expression in vitro and in transgenic mice by an LXR element.

The cholesterol ester transfer protein (CETP) facilitates the transfer of HDL cholesterol esters from plasma to the liver. Transgenic mice expressing human CETP, controlled by its natural flanking region, increase expression of this gene in response to hypercholesterolemia. We established a CETP promoter-luciferase reporter assay in differentiated 3T3-L1 adipocytes to map the sterol upregulatory element. Promoter mutagenesis suggested that a direct repeat of a nuclear receptor binding sequence separated by 4 nucleotides (DR4 element, -384 to -399) was responsible for this activity. Using mice carrying normal or mutated promoter sequences, we confirmed the importance of this element for gene induction by dietary sterol. A gel retardation complex containing LXR/RXR was identified using the CETP DR4 element and adipocyte nuclear extracts. Both LXRalpha/RXRalpha and LXRbeta/RXRalpha transactivated the CETP promoter via its DR4 element in a sterol-responsive fashion. Thus, the positive sterol response of the CETP gene is mediated by a nuclear receptor binding site that is activated by LXRs. That Cyp7a, the rate-limiting enzyme for conversion of cholesterol into bile acids in the liver, is also regulated by LXRalpha suggests that this class of nuclear receptor coordinates the regulation of HDL cholesterol ester catabolism and bile acid synthesis in the liver.

Organization of Human Phospholipid Transfer Protein Gene

We have determined the exon/intron organization of the human phospholipid transfer protein gene. The gene, which spans approximately 13.3 kilobases, is comprised of 16 exons. The organization of the phospholipid transfer protein gene strikingly resembles that encoding another plasma lipid transfer protein, the human cholesterol ester transfer protein. The exon-intron junctions in these two genes are highly conserved, with eight out of fifteen junctions interrupting the same codons, while the remaining junctions lie within 5 residues of each other. The similarity in gene structure and homology in coding sequences suggests that these two genes most likely evolved from a common ancestral gene.

Interfacial properties of recombinant human cholesterol ester transfer protein.

We investigated the interfacial behavior of recombinant human cholesterol ester transfer protein (rCETP) using monolayer and surface balance techniques. rCETP bound to egg phosphatidylcholine monolayers spread at the air/water interface with a maximum surface pressure of 23 millinewtons (mN)/m at subphase concentrations between 3 and 5 x 10(-5) g/dl; the estimated dissociation constant was 7.5 x 10(-6) g/dl or 1 nM. The binding of rCETP to the lipid interface decreased linearly with increasing initial surface pressure; rCETP was excluded at pressures greater than 31 mN/m. rCETP catalyzed the desorption of [14C]cholesterol oleate from mixed lipid monolayers in a concentration dependent fashion. Similar studies with apolipoproteins A-I and A-IV established that cholesterol ester desorption was not caused by changes in surface pressure or cholesterol ester solubility. The desorption rate was proportional to subphase rCETP concentration, but at all concentrations surface radioactivity remained constant until surface pressure reached a plateau. The calculated binding stochiometry was one molecule of cholesterol ester desorbed for every 1000 molecules of rCETP in the subphase. We conclude that rCETP is surface active, binds to phospholipid monolayers with an affinity equivalent to that of the plasma apolipoproteins, and effects the desorption of cholesterol ester molecules from phospholipid monolayers by a carrier mechanism. Moreover, the relatively low equilibrium surface pressure of rCETP suggests that when bound to lipid the entire rCETP molecule may not penetrate the interface.