Cholesteryl Ester Transfer Protein Mice Research Articles

CETP inhibition enhances monocyte activation and bacterial clearance and reduces streptococcus pneumonia-associated mortality in mice.

Sepsis is a leading cause of mortality worldwide, and pneumonia is the most common cause of sepsis in humans. Low levels of high-density lipoprotein cholesterol (HDL-C) levels are associated with an increased risk of death from sepsis, and increasing levels of HDL-C by inhibition of cholesteryl ester transfer protein (CETP) decreases mortality from intraabdominal polymicrobial sepsis in APOE*3-Leiden.CETP mice. Here, we show that treatment with the CETP inhibitor (CETPi) anacetrapib reduced mortality from Streptococcus pneumoniae-induced sepsis in APOE*3-Leiden.CETP and APOA1.CETP mice. Mechanistically, CETP inhibition reduced the host proinflammatory response via attenuation of proinflammatory cytokine transcription and release. This effect was dependent on the presence of HDL, leading to attenuation of immune-mediated organ damage. In addition, CETP inhibition promoted monocyte activation in the blood prior to the onset of sepsis, resulting in accelerated macrophage recruitment to the lung and liver. In vitro experiments demonstrated that CETP inhibition significantly promoted the activation of proinflammatory signaling in peripheral blood mononuclear cells and THP1 cells in the absence of HDL; this may represent a mechanism responsible for improved bacterial clearance during sepsis. These findings provide evidence that CETP inhibition represents a potential approach to reduce mortality from pneumosepsis.

Atherosclerosis: an overview of mouse models and a detailed methodology to quantify lesions in the aortic root.

Cardiovascular disease, the primary cause of human mortality globally, is predominantly caused by a progressive disorder known as atherosclerosis. Atherosclerosis refers to the process of accumulation of cholesterol-enriched lipoproteins and the concomitant initiation of inflammatory processes in the arterial wall, including the recruitment of immune cells. This leads to the formation of atherosclerotic plaques, initially causing a thickening of the arterial wall and narrowing of arteries. However, as plaque formation progresses, atherosclerotic plaques may become unstable and rupture, leading to a blood clot that blocks the affected artery or travels through the blood to block blood flow elsewhere. In the early 1990s, emerging gene editing methods enabled the development of apolipoprotein E knockout (Apoe-/- ) and low-density lipoprotein receptor knockout (Ldlr-/- ) mice. These mice have been instrumental in unraveling the complex pathogenesis of atherosclerosis. Around the same time, human APOE*3-Leiden transgenic mice were generated, which were more recently cross-bred with human cholesteryl ester transfer protein (CETP) transgenic mice to generate APOE*3-Leiden.CETP mice. This model appears to closely mimic human lipoprotein metabolism and responds to classic lipid-lowering interventions due to an intact ApoE-LDLR pathway of lipoprotein remnant clearance. In this review, we describe the role of lipid metabolism and inflammation in atherosclerosis development and highlight the characteristics of the frequently used animal models to study atherosclerosis, with a focus on mouse models, discussing their advantages and limitations. Moreover, we present a detailed methodology to quantify atherosclerotic lesion area within the aortic root region of the murine heart, as well as details required for scoring atherosclerotic lesion severity based on guidelines of the American Heart Association adapted for mice.

The impact of cholesteryl ester transfer protein on the progression of cutaneous leishmaniasis.

Pathogenesis of cutaneous leishmaniases involves parasite growth, persistent inflammation, and likely participation of lipoproteins (LP). The cholesteryl ester transfer protein (CETP), involved in LP remodeling, has been shown to participate in the inflammatory response and the evolution of infectious conditions. We evaluated the impact of the presence of CETP on infection by Leishmania (L.) amazonensis in an experimental model of cutaneous leishmaniasis using C57BL6/J mice transgenic for human CETP (CETP), having as control their littermates that do not express the protein, wild-type (WT) mice. The progression of the lesion after infection in the footpad was monitored for 12 weeks. Two groups of animals were formed to collect the plantar pad in the 4th and 12th week post-infection. The lesion increased from the 3rd week onwards, in both groups, with a gradual decrease from the 10th week onwards in the CETP group compared to the WT group, showing a reduction in parasitism and an improvement in the healing process, a reduction in CD68+ cells, and an increase in CD163+ and CD206, characterizing a population of M2 macrophages. A reduction in ARG1+ cells and an increase in INOS+ cells were observed. During infection, the LP profile showed an increase in triglycerides in the VLDL fraction in the CETP group at 12 weeks. Gene expression revealed a decrease in the CD36 receptor in the CETP group at 12 weeks, correlating with healing and parasite reduction. In vitro, macrophages derived from bone marrow cells from CETP mice showed lower parasite load at 48 h and, a reduction in arginase activity at 4 h accompanied by increased NO production at 4 and 24 h compared to WT macrophages, corroborating the in vivo findings. The data indicate that the presence of CETP plays an important role in resolving Leishmania (L.) amazonensis infection, reducing parasitism, and modulating the inflammatory response in controlling infection and tissue repair.

Human cholesteryl ester transfer protein lacks lipopolysaccharide transfer activity, but worsens inflammation and sepsis outcomes in mice

Bacterial lipopolysaccharides (LPSs or endotoxins) can bind most proteins of the lipid transfer/LPS-binding protein (LT/LBP) family in host organisms. The LPS-bound LT/LBP proteins then trigger either an LPS-induced proinflammatory cascade or LPS binding to lipoproteins that are involved in endotoxin inactivation and detoxification. Cholesteryl ester transfer protein (CETP) is an LT/LBP member, but its impact on LPS metabolism and sepsis outcome is unclear. Here, we performed fluorescent LPS transfer assays to assess the ability of CETP to bind and transfer LPS. The effects of intravenous (iv) infusion of purified LPS or polymicrobial infection (cecal ligation and puncture [CLP]) were compared in transgenic mice expressing human CETP and wild-type mice naturally having no CETP activity. CETP displayed no LPS transfer activity in vitro, but it tended to reduce biliary excretion of LPS in vivo. The CETP expression in mice was associated with significantly lower basal plasma lipid levels and with higher mortality rates in both models of endotoxemia and sepsis. Furthermore, CETPTg plasma modified cytokine production of macrophages in vitro. In conclusion, despite having no direct LPS binding and transfer property, human CETP worsens sepsis outcomes in mice by altering the protective effects of plasma lipoproteins against endotoxemia, inflammation, and infection.

Inhibition of Cholesteryl Ester Transfer Protein Preserves High-Density Lipoprotein Cholesterol and Improves Survival in Sepsis.

The high-density lipoprotein hypothesis of atherosclerosis has been challenged by clinical trials of cholesteryl ester transfer protein (CETP) inhibitors, which failed to show significant reductions in cardiovascular events. Plasma levels of high-density lipoprotein cholesterol (HDL-C) decline drastically during sepsis, and this phenomenon is explained, in part, by the activity of CETP, a major determinant of plasma HDL-C levels. We tested the hypothesis that genetic or pharmacological inhibition of CETP would preserve high-density lipoprotein levels and decrease mortality in clinical cohorts and animal models of sepsis. We examined the effect of a gain-of-function variant in CETP (rs1800777, p.Arg468Gln) and a genetic score for decreased CETP function on 28-day sepsis survival using Cox proportional hazard models adjusted for age and sex in the UK Biobank (n=5949), iSPAAR (Identification of SNPs Predisposing to Altered Acute Lung Injury Risk; n=882), Copenhagen General Population Study (n=2068), Copenhagen City Heart Study (n=493), Early Infection (n=200), St Paul's Intensive Care Unit 2 (n=203), and Vasopressin Versus Norepinephrine Infusion in Patients With Septic Shock studies (n=632). We then studied the effect of the CETP inhibitor, anacetrapib, in adult female APOE*3-Leiden mice with or without human CETP expression using the cecal-ligation and puncture model of sepsis. A fixed-effect meta-analysis of all 7 cohorts found that the CETP gain-of-function variant was significantly associated with increased risk of acute sepsis mortality (hazard ratio, 1.44 [95% CI, 1.22-1.70]; P<0.0001). In addition, a genetic score for decreased CETP function was associated with significantly decreased sepsis mortality in the UK Biobank (hazard ratio, 0.77 [95% CI, 0.59-1.00] per 1 mmol/L increase in HDL-C) and iSPAAR cohorts (hazard ratio, 0.60 [95% CI, 0.37-0.98] per 1 mmol/L increase in HDL-C). APOE*3-Leiden.CETP mice treated with anacetrapib had preserved levels of HDL-C and apolipoprotein-AI and increased survival relative to placebo treatment (70.6% versus 35.3%, Log-rank P=0.03), whereas there was no effect of anacetrapib on the survival of APOE*3-Leiden mice that did not express CETP (50.0% versus 42.9%, Log-rank P=0.87). Clinical genetics and humanized mouse models suggest that inhibiting CETP may preserve high-density lipoprotein levels and improve outcomes for individuals with sepsis.

Novel role of cholesteryl ester transfer protein (CETP): attenuation of adiposity by enhancing lipolysis and brown adipose tissue activity

ObjectiveThe systemic function of CETP has been well characterized. CETP plasma activity reduces HDL cholesterol and thus increases the risk of atherosclerosis. Here, we investigated whether CETP expression modulate adiposity. MethodsBody adiposity and energy metabolism related assays and gene/protein expression were compared in CETP transgenic and non-transgenic mice and in hamsters treated with CETP neutralizing antibody. ResultsWe found that transgenic mice expressing human CETP present less white adipose tissue mass and lower leptinemia than nontransgenic (NTg) littermates. No differences were found in physical activity, food intake, fat fecal excretion, lipogenesis or exogenous lipid accumulation in adipose depots. Nonetheless, adipose lipolysis rates and whole-body energy expenditure were elevated in CETP mice. In accordance, lipolysis-related gene expression and protein content were increased in visceral and brown adipose tissue (BAT). In addition, we verified increased BAT temperature and oxygen consumption. These results were confirmed in two other animal models: 1) hamsters treated with CETP neutralizing antibody and 2) an independent line of transgenic mice expressing simian CETP. ConclusionsThese findings reveal a novel anti-adipogenic role for CETP.

Cholesteryl ester transfer protein alters liver and plasma triglyceride metabolism through two liver networks in female mice

Elevated plasma TGs increase risk of cardiovascular disease in women. Estrogen treatment raises plasma TGs in women, but molecular mechanisms remain poorly understood. Here we explore the role of cholesteryl ester transfer protein (CETP) in the regulation of TG metabolism in female mice, which naturally lack CETP. In transgenic CETP females, acute estrogen treatment raised plasma TGs 50%, increased TG production, and increased expression of genes involved in VLDL synthesis, but not in nontransgenic littermate females. In CETP females, estrogen enhanced expression of small heterodimer partner (SHP), a nuclear receptor regulating VLDL production. Deletion of liver SHP prevented increases in TG production and expression of genes involved in VLDL synthesis in CETP mice with estrogen treatment. We also examined whether CETP expression had effects on TG metabolism independent of estrogen treatment. CETP increased liver β-oxidation and reduced liver TG content by 60%. Liver estrogen receptor α (ERα) was required for CETP expression to enhance β-oxidation and reduce liver TG content. Thus, CETP alters at least two networks governing TG metabolism, one involving SHP to increase VLDL-TG production in response to estrogen, and another involving ERα to enhance β-oxidation and lower liver TG content. These findings demonstrate a novel role for CETP in estrogen-mediated increases in TG production and a broader role for CETP in TG metabolism.

Abstract 397: Transgenic Expression of Cholesteryl Ester Transfer Protein in Female Mice Disrupts Liver and Plasma Triglyceride Metabolism With Estrogen Treatment in a Liver Network Involving Estrogen Receptor Alpha and Small Heterodimer Partner

Background: Triglycerides are an important risk factor for cardiovascular disease, especially in women. Hormone replacement therapy raises triglycerides (TG) in postmenopausal women, but mechanisms responsible remain poorly understood. Cholesteryl Ester Transfer Protein (CETP) is a serum protein that shuttles TG and cholesteryl ester between lipoproteins in humans and plays an important role in lipoprotein metabolism. The role of CETP in regulating whole body triglyceride homeostasis remains unclear. Mice (which naturally lack CETP) expressing a transgenic copy of CETP or their wild-type (WT) littermates were used to determine the role of CETP in regulating plasma and liver TG metabolism in response to estrogen. Methods and Results: Female CETP and WT mice were ovariectomized and given vehicle or estrogen treatment. Estrogen raised plasma TG in CETP females, but not in WT females. CETP expression impaired TG clearance independent of estrogen. Estrogen treatment raised very-low density lipoprotein (VLDL) TG production in CETP females, but not in WT females. Estrogen treatment augmented mRNA expression and protein activity of VLDL assembly and secretion targets (Apob, Mttp, PDI) in CETP females, but not in WT females. Additionally, CETP expression lowered liver TG content and increased mRNA expression of TG oxidation targets (Ppara, Acox1, Acadm, Cpt2). Deletion of liver estrogen receptor alpha prevented CETP-mediated decreases in liver TG content and increases in mRNA expression of TG oxidation targets. Liver-specific deletion of estrogen receptor alpha did not attenuate estrogen-mediated increases in plasma TG and VLDL-TG production in female mice expressing CETP. Deletion of liver small heterodimer partner attenuated CETP-mediated increases in plasma TG and VLDL-TG production with estrogen treatment. Deletion of small heterodimer partner also attenuated CETP-mediated increases in mRNA expression and protein activity of VLDL synthesis and assembly targets with estrogen treatment. Conclusion: These studies indicate several novel functions of CETP in regulating plasma and liver TG homeostasis. We also discovered that CETP expression in female mice also augments several aspects of estrogen action in liver that impact liver TG biology.

Cholesteryl Ester Transfer Protein (CETP) expression does not affect glucose homeostasis and insulin secretion: studies in human CETP transgenic mice.

BackgroundCholesteryl ester transfer protein (CETP) is a plasma protein that mediates the exchange of triglycerides for esterified cholesterol between HDL and apoB-lipoproteins. Previous studies suggest that CETP may modify glucose metabolism in patients or cultured cells. In this study, we tested if stable CETP expression would impair glucose metabolism.MethodsWe used human CETP transgenic mice and non-transgenic littermate controls (NTg), fed with control or high fat diet, as well as in dyslipidemic background and aging conditions. Assays included glucose and insulin tolerance tests, isolated islets insulin secretion, tissue glucose uptake and adipose tissue GLUT mRNA expression.ResultsCETP expression did not modify glucose or insulin tolerance in all tested conditions such as chow and high fat diet, adult and aged mice, normo and dyslipidemic backgrounds. Fasting and fed state plasma levels of insulin were not differ in CETP and NTg mice. Direct measurements of isolated pancreatic islet insulin secretion rates induced by glucose (11, 16.7 or 22 mM), KCl (40 mM), and leucine (10 mM) were similar in NTg and CETP mice, indicating that CETP expression did not affect β-cell function in vivo and ex vivo. Glucose uptake by insulin target tissues, measured in vivo using 3H-2-deoxyglucose, showed that CETP expression had no effect on the glucose uptake in liver, muscle, perigonadal, perirenal, subcutaneous and brown adipose tissues. Accordingly, GLUT1 and GLUT4 mRNA in adipose tissue were not affected by CETP.ConclusionsIn summary, by comparing the in vivo all-or-nothing CETP expressing mouse models, we demonstrated that CETP per se has no impact on the glucose tolerance and tissue uptake, global insulin sensitivity and beta cell insulin secretion rates.

CETP Lowers TLR4 Expression Which Attenuates the Inflammatory Response Induced by LPS and Polymicrobial Sepsis.

Sepsis is a systemic inflammatory response to infection eliciting high mortality rate which is a serious health problem. Despite numerous studies seeking for therapeutic alternatives, the mechanisms involved in this disease remain elusive. In this study we evaluated the influence of cholesteryl ester transfer protein (CETP), a glycoprotein that promotes the transfer of lipids between lipoproteins, on the inflammatory response in mice. Human CETP transgenic mice were compared to control mice (wild type, WT) after polymicrobial sepsis induced by cecal ligation and puncture (CLP), aiming at investigating their survival rate and inflammatory profiles. Macrophages from the peritoneal cavity were stimulated with LPS in the presence or absence of recombinant CETP for phenotypic and functional studies. In comparison to WT mice, CETP mice showed higher survival rate, lower IL-6 plasma concentration, and decreased liver toll-like receptor 4 (TLR4) and acyloxyacyl hydrolase (AOAH) protein. Moreover, macrophages from WT mice to which recombinant human CETP was added decreased LPS uptake, TLR4 expression, NF-κB activation and IL-6 secretion. This raises the possibility for new therapeutic tools in sepsis while suggesting that lowering CETP by pharmacological inhibitors should be inconvenient in the context of sepsis and infectious diseases.

CETP Expression Protects Female Mice from Obesity-Induced Decline in Exercise Capacity.

Pharmacological approaches to reduce obesity have not resulted in dramatic reductions in the risk of coronary heart disease (CHD). Exercise, in contrast, reduces CHD risk even in the setting of obesity. Cholesteryl Ester Transfer Protein (CETP) is a lipid transfer protein that shuttles lipids between serum lipoproteins and tissues. There are sexual-dimorphisms in the effects of CETP in humans. Mice naturally lack CETP, but we previously reported that transgenic expression of CETP increases muscle glycolysis in fasting and protects against insulin resistance with high-fat diet (HFD) feeding in female but not male mice. Since glycolysis provides an important energy source for working muscle, we aimed to define if CETP expression protects against the decline in exercise capacity associated with obesity. We measured exercise capacity in female mice that were fed a chow diet and then switched to a HFD. There was no difference in exercise capacity between lean, chow-fed CETP female mice and their non-transgenic littermates. Female CETP transgenic mice were relatively protected against the decline in exercise capacity caused by obesity compared to WT. Despite gaining similar fat mass after 6 weeks of HFD-feeding, female CETP mice showed a nearly two-fold increase in run distance compared to WT. After an additional 6 weeks of HFD-feeding, mice were subjected to a final exercise bout and muscle mitochondria were isolated. We found that improved exercise capacity in CETP mice corresponded with increased muscle mitochondrial oxidative capacity, and increased expression of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α). These results suggest that CETP can protect against the obesity-induced impairment in exercise capacity and may be a target to improve exercise capacity in the context of obesity.

Prolonged niacin treatment leads to increased adipose tissue PUFA synthesis and anti-inflammatory lipid and oxylipin plasma profile

Prolonged niacin treatment elicits beneficial effects on the plasma lipid and lipoprotein profile that is associated with a protective CVD risk profile. Acute niacin treatment inhibits nonesterified fatty acid release from adipocytes and stimulates prostaglandin release from skin Langerhans cells, but the acute effects diminish upon prolonged treatment, while the beneficial effects remain. To gain insight in the prolonged effects of niacin on lipid metabolism in adipocytes, we used a mouse model with a human-like lipoprotein metabolism and drug response [female APOE*3-Leiden.CETP (apoE3 Leiden cholesteryl ester transfer protein) mice] treated with and without niacin for 15 weeks. The gene expression profile of gonadal white adipose tissue (gWAT) from niacin-treated mice showed an upregulation of the "biosynthesis of unsaturated fatty acids" pathway, which was corroborated by quantitative PCR and analysis of the FA ratios in gWAT. Also, adipocytes from niacin-treated mice secreted more of the PUFA DHA ex vivo. This resulted in an increased DHA/arachidonic acid (AA) ratio in the adipocyte FA secretion profile and in plasma of niacin-treated mice. Interestingly, the DHA metabolite 19,20-dihydroxy docosapentaenoic acid (19,20-diHDPA) was increased in plasma of niacin-treated mice. Both an increased DHA/AA ratio and increased 19,20-diHDPA are indicative for an anti-inflammatory profile and may indirectly contribute to the atheroprotective lipid and lipoprotein profile associated with prolonged niacin treatment.

PCSK9 inhibition fails to alter hepatic LDLR, circulating cholesterol, and atherosclerosis in the absence of ApoE

LDL cholesterol (LDL-C) contributes to coronary heart disease. Proprotein convertase subtilisin/kexin type 9 (PCSK9) increases LDL-C by inhibiting LDL-C clearance. The therapeutic potential for PCSK9 inhibitors is highlighted by the fact that PCSK9 loss-of-function carriers exhibit 15-30% lower circulating LDL-C and a disproportionately lower risk (47-88%) of experiencing a cardiovascular event. Here, we utilized pcsk9(-/-) mice and an anti-PCSK9 antibody to study the role of the LDL receptor (LDLR) and ApoE in PCSK9-mediated regulation of plasma cholesterol and atherosclerotic lesion development. We found that circulating cholesterol and atherosclerotic lesions were minimally modified in pcsk9(-/-) mice on either an LDLR- or ApoE-deficient background. Acute administration of an anti-PCSK9 antibody did not reduce circulating cholesterol in an ApoE-deficient background, but did reduce circulating cholesterol (-45%) and TGs (-36%) in APOE*3Leiden.cholesteryl ester transfer protein (CETP) mice, which contain mouse ApoE, human mutant APOE3*Leiden, and a functional LDLR. Chronic anti-PCSK9 antibody treatment in APOE*3Leiden.CETP mice resulted in a significant reduction in atherosclerotic lesion area (-91%) and reduced lesion complexity. Taken together, these results indicate that both LDLR and ApoE are required for PCSK9 inhibitor-mediated reductions in atherosclerosis, as both are needed to increase hepatic LDLR expression.

Anacetrapib reduces progression of atherosclerosis, mainly by reducing non-HDL-cholesterol, improves lesion stability and adds to the beneficial effects of atorvastatin.

The residual risk that remains after statin treatment supports the addition of other LDL-C-lowering agents and has stimulated the search for secondary treatment targets. Epidemiological studies propose HDL-C as a possible candidate. Cholesteryl ester transfer protein (CETP) transfers cholesteryl esters from atheroprotective HDL to atherogenic (V)LDL. The CETP inhibitor anacetrapib decreases (V)LDL-C by ∼15-40% and increases HDL-C by ∼40-140% in clinical trials. We evaluated the effects of a broad dose range of anacetrapib on atherosclerosis and HDL function, and examined possible additive/synergistic effects of anacetrapib on top of atorvastatin in APOE*3Leiden.CETP mice. Mice were fed a diet without or with ascending dosages of anacetrapib (0.03; 0.3; 3; 30 mg/kg/day), atorvastatin (2.4 mg/kg/day) alone or in combination with anacetrapib (0.3 mg/kg/day) for 21 weeks. Anacetrapib dose-dependently reduced CETP activity (-59 to -100%, P < 0.001), thereby decreasing non-HDL-C (-24 to -45%, P < 0.001) and increasing HDL-C (+30 to +86%, P < 0.001). Anacetrapib dose-dependently reduced the atherosclerotic lesion area (-41 to -92%, P < 0.01) and severity, increased plaque stability index and added to the effects of atorvastatin by further decreasing lesion size (-95%, P < 0.001) and severity. Analysis of covariance showed that both anacetrapib (P < 0.05) and non-HDL-C (P < 0.001), but not HDL-C (P = 0.76), independently determined lesion size. Anacetrapib dose-dependently reduces atherosclerosis, and adds to the anti-atherogenic effects of atorvastatin, which is mainly ascribed to a reduction in non-HDL-C. In addition, anacetrapib improves lesion stability.

Abstract 246: Anacetrapib Adds to the Antiatherogenic Effect of Atorvastatin, Mainly by Increasing the Clearance of Non-HDL Cholesterol

Introduction: The residual risk that remains after statin treatment has stimulated the search for secondary treatment targets. Epidemiological studies propose HDL-C as a possible candidate. Cholesteryl ester transfer protein (CETP) transfers cholesteryl esters from atheroprotective HDL to atherogenic (V)LDL. The CETP inhibitor anacetrapib decreases (V)LDL-C by ~15-40% and increases HDL-C by ~40-140% in clinical trials. We evaluated the effects of anacetrapib on lipid metabolism, HDL function and atherosclerosis, and examined possible additive/synergistic effects of anacetrapib on top of atorvastatin in APOE*3Leiden.CETP mice. Methods and results: Mice were fed a diet without or with ascending dosages of anacetrapib (0.03; 0.3; 3; 30 mg/kg/d), atorvastatin (2.4 mg/kg/d) alone or in combination with anacetrapib (0.3 mg/kg/d) for 21 weeks. Anacetrapib dose-dependently reduced CETP activity (-59% to -100%, P&lt;0.01), thereby decreasing nonHDL-C (-24% to -45%, P&lt;0.001) and increasing HDL-C (+30% to +86%, P&lt;0.001). Anacetrapib did not affect HDL function. Additionally, we showed that anacetrapib reduced nonHDL-C mainly by increasing the clearance by the liver. Anacetrapib dose-dependently reduced atherosclerotic lesion area (-41% to -92%, P&lt;0.01) and severity, increased the plaque stability index and added to the effects of atorvastatin by further decreasing lesion size (-95%, P&lt;0.001) and severity. Analysis of covariance showed that both anacetrapib (P&lt;0.05) and nonHDL-C (P&lt;0.001), but not HDL-C (P=0.76), independently decreased lesion size. Conclusions: Anacetrapib reduces atherosclerosis, and adds to the anti-atherogenic effect of atorvastatin. The anti-atherogenic effect is mainly ascribed to a reduction in nonHDL-C, as a consequence of an increased clearance. In addition, anacetrapib improves lesion stability.

Cholesteryl ester transfer protein protects against insulin resistance in obese female mice

Cholesteryl ester transfer protein (CETP) shuttles lipids between lipoproteins, culminating in cholesteryl ester delivery to liver and increased secretion of cholesterol as bile. Since gut bile acids promote insulin sensitivity, we aimed to define if CETP improves insulin sensitivity with high-fat feeding. CETP and nontransgenic mice of both sexes became obese. Female but not male CETP mice had increased ileal bile acid levels versus nontransgenic littermates. CETP expression protected female mice from insulin resistance but had a minimal effect in males. In liver, female CETP mice showed activation of bile acid-sensitive pathways including Erk1/2 phosphorylation and Fxr and Shp gene expression. In muscle, CETP females showed increased glycolysis, increased mRNA for Dio2, and increased Akt phosphorylation, known effects of bile acid signaling. These results suggest that CETP can ameliorate insulin resistance associated with obesity in female mice, an effect that correlates with increased gut bile acids and known bile-signaling pathways.

Abstract 69: Anacetrapib Dose-dependently Decreases Atherosclerosis Development and Adds to the Beneficial Effects of Atorvastatin in APOE*3Leiden.CETP Mice

Introduction The residual risk of cardiovascular disease that remains after statin treatment has triggered the search for a secondary treatment target. Epidemiological studies propose HDL-cholesterol (HDL-C) as a possible candidate. Cholesteryl ester transfer protein (CETP) transfers cholesteryl esters from atheroprotective HDL to atherogenic (V)LDL. In human intervention trials, the CETP inhibitor anacetrapib decreases (V)LDL-C by 30-40% and increases HDL-C by 40-140%. Hypothesis Complete inhibition of CETP activity may result in adverse effects as compared to partial inhibition due to the appearance of a dysfunctional HDL. We, therefore, evaluated the effect of a broad treatment window of anacetrapib-induced CETP inhibition with partial to full inhibition, as well as the combination of atorvastatin and anacetrapib on atherosclerosis development in APOE*3Leiden.CETP mice. Methods Female mice were fed a Western-type diet containing 0.1% cholesterol without or with incremental dosages of anacetrapib (0.03; 0.3; 3; 30 mg/kg/d), atorvastatin (2.4 mg/kg/d) or a combination of anacetrapib (0.3 mg/kg/d) and atorvastatin (2.4 mg/kg/d) for 20 weeks. Effects on plasma lipids, CETP activity and levels, as well as atherosclerotic lesion size and severity were assessed. Results Anacetrapib dose-dependently reduced CETP activity (-60% to -100%, P&lt;0.001) and increased CETP levels (+13% to +31%, P&lt;0.05), thereby decreasing nonHDL-C (-23% to -44%, P&lt;0.001) and increasing HDL-C (+32% to +88%, P&lt;0.001). Atorvastatin decreased CETP activity (-29%, P&lt;0.001) and nonHDL-C (-36%, P&lt;0.001). Anacetrapib dose-dependently decreased atherosclerotic lesion size (-36%, P&lt;0.05 to -92%, P&lt;0.001) and the percentage severe lesions. Anacetrapib added to the effects of atorvastatin by further reducing nonHDL-C (-36%, P&lt;0.001), increasing HDL-C (+72%, P&lt;0.001) and decreasing lesion size (-86%, P&lt;0.001) and severity. Results on lesion composition are pending. Conclusions Anacetrapib dose-dependently decreases atherosclerosis development and adds to the beneficial effects of atorvastatin in APOE*3Leiden.CETP mice. Total blockage of CETP activity does not reveal adverse effects as compared to partial blockage.

Osteoarthritis development is induced by increased dietary cholesterol and can be inhibited by atorvastatin in APOE*3Leiden.CETP mice—a translational model for atherosclerosis

ObjectiveHypercholesterolaemia, a risk factor for atherosclerosis (ATH), has been suggested to have a role in the development of osteoarthritis (OA). To test this hypothesis, the effect of cholesterol and different...

Torcetrapib Does Not Reduce Atherosclerosis Beyond Atorvastatin and Induces More Proinflammatory Lesions Than Atorvastatin

Although cholesteryl ester transfer protein (CETP) inhibition is regarded as a promising strategy to reduce atherosclerosis by increasing high-density lipoprotein cholesterol, the CETP inhibitor torcetrapib given in addition to atorvastatin had no effect on atherosclerosis and even increased cardiovascular death in the recent Investigation of Lipid Level Management to Understand its Impact in Atherosclerotic Events trial. Therefore, we evaluated the antiatherogenic potential and adverse effects of torcetrapib in humanized APOE*3-Leiden.CETP (E3L.CETP) mice. E3L.CETP mice were fed a cholesterol-rich diet without drugs or with torcetrapib (12 mg x kg(-1) x d(-1)), atorvastatin (2.8 mg x kg(-1) x d(-1)), or both for 14 weeks. Torcetrapib decreased CETP activity in both the absence and presence of atorvastatin (-74% and -73%, respectively; P<0.001). Torcetrapib decreased plasma cholesterol (-20%; P<0.01), albeit to a lesser extent than atorvastatin (-42%; P<0.001) or the combination of torcetrapib and atorvastatin (-40%; P<0.001). Torcetrapib increased high-density lipoprotein cholesterol in the absence (30%) and presence (34%) of atorvastatin. Torcetrapib and atorvastatin alone reduced atherosclerotic lesion size (-43% and -46%; P<0.05), but combination therapy did not reduce atherosclerosis compared with atorvastatin alone. Remarkably, compared with atorvastatin, torcetrapib enhanced monocyte recruitment and expression of monocyte chemoattractant protein-1 and resulted in lesions of a more inflammatory phenotype, as reflected by an increased macrophage content and reduced collagen content. CETP inhibition by torcetrapib per se reduces atherosclerotic lesion size but does not enhance the antiatherogenic potential of atorvastatin. However, compared with atorvastatin, torcetrapib introduces lesions of a less stable phenotype.

Cholesteryl ester transfer protein expression prevents diet-induced atherosclerotic lesions in male db/db mice.

Accompanying more atherogenic lipoprotein profiles and an increased incidence of atherosclerosis, plasma cholesteryl ester transfer protein (CETP) is depressed in diabetic obese patients compared with nondiabetic obese counterparts. The depressed levels of CETP in the plasma of diabetic obese individuals may contribute to the development of an atherogenic lipoprotein profile and atherogenesis. We have examined the effect of CETP expression on vascular health in the db/db model of diabetic obesity. Transgenic mice expressing the human CETP minigene were crossed with db/db strain, and 3 groups of offspring (CETP, db, and db/CETP) were placed on an atherogenic diet for 16 weeks. The proximal aorta was then excised and examined for the presence of atherosclerotic plaques. In db mice, 9 of 11 had intimal lesions with a mean area of 26 098+/-7486 microm2. No lesions greater than 1000 microm2 were observed in db/CETP or CETP mice. CETP-expressing mice had lower circulating cholesterol concentrations than db mice. Fractionating plasma lipids by FPLC indicated that the difference in total cholesterol was primarily attributable to differences in VLDL and LDL. The expression of human CETP in db/db mice prevented the formation of diet-induced lesions, suggesting an antiatherogenic effect of CETP in the context of diabetic obesity.