Anacetrapib promotes reverse cholesterol transport and bulk cholesterol excretion in Syrian golden hamsters

Cholesteryl ester transfer protein (CETP) transfers cholesteryl ester (CE) and triglyceride between HDL and apoB-containing lipoproteins. Anacetrapib (ANA), a reversible inhibitor of CETP, raises HDL cholesterol (HDL-C) and lowers LDL cholesterol in dyslipidemic patients; however, the effects of ANA on cholesterol/lipoprotein metabolism in a dyslipidemic hamster model have not been demonstrated. To test whether ANA (60 mg/kg/day, 2 weeks) promoted reverse cholesterol transport (RCT), 3H-cholesterol-loaded macrophages were injected and (3)H-tracer levels were measured in HDL, liver, and feces. Compared to controls, ANA inhibited CETP (94%) and increased HDL-C (47%). 3H-tracer in HDL increased by 69% in hamsters treated with ANA, suggesting increased cholesterol efflux from macrophages to HDL. 3H-tracer in fecal cholesterol and bile acids increased by 90% and 57%, respectively, indicating increased macrophage-to-feces RCT. Mass spectrometry analysis of HDL from ANA-treated hamsters revealed an increase in free unlabeled cholesterol and CE. Furthermore, bulk cholesterol and cholic acid were increased in feces from ANA-treated hamsters. Using two independent approaches to assess cholesterol metabolism, the current study demonstrates that CETP inhibition with ANA promotes macrophage-to-feces RCT and results in increased fecal cholesterol/bile acid excretion, further supporting its development as a novel lipid therapy for the treatment of dyslipidemia and atherosclerotic vascular disease.     

Postprandial lipemia enhances the capacity of large HDL2 particles to mediate free cholesterol efflux via SR-BI and ABCG1 pathways in type IIB hyperlipidemia

Lipid and cholesterol metabolism in the postprandial phase is associated with both quantitative and qualitative remodeling of HDL particle subspecies that may influence their anti-atherogenic functions in the reverse cholesterol transport pathway. We evaluated the capacity of whole plasma or isolated HDL particles to mediate cellular free cholesterol (FC) efflux, cholesteryl ester transfer protein (CETP)-mediated cholesteryl ester (CE) transfer, and selective hepatic CE uptake during the postprandial phase in subjects displaying type IIB hyperlipidemia (n = 16). Postprandial, large HDL2 displayed an enhanced capacity to mediate FC efflux via both scavenger receptor class B type I (SR-BI)-dependent (+12%; P < 0.02) and ATP binding cassette transporter G1 (ABCG1)-dependent (+31%; P < 0.008) pathways in in vitro cell systems. In addition, the capacity of whole postprandial plasma (4 h and 8 h postprandially) to mediate cellular FC efflux via the ABCA1-dependent pathway was significantly increased (+19%; P < 0.0003). Concomitantly, postprandial lipemia was associated with elevated endogenous CE transfer rates from HDL2 to apoB lipoproteins and with attenuated capacity (−17%; P < 0.02) of total HDL to deliver CE to hepatic cells. Postprandial lipemia enhanced SR-BI and ABCG1-dependent efflux to large HDL2 particles. However, postprandial lipemia is equally associated with deleterious features by enhancing formation of CE-enriched, triglyceride-rich lipoprotein particles through the action of CETP and by reducing the direct return of HDL-CE to the liver.     

HDL-apolipoprotein A-I exchange is independently associated with cholesterol efflux capacity

HDL is the primary mediator of cholesterol mobilization from the periphery to the liver via reverse cholesterol transport (RCT). A critical first step in this process is the uptake of cholesterol from lipid-loaded macrophages by HDL, a function of HDL inversely associated with prevalent and incident cardiovascular disease. We hypothesized that the dynamic ability of HDL to undergo remodeling and exchange of apoA-I is an important and potentially rate-limiting aspect of RCT. In this study, we investigated the relationship between HDL-apoA-I exchange (HAE) and serum HDL cholesterol (HDL-C) efflux capacity. We compared HAE to the total and ABCA1-specific cholesterol efflux capacity of 77 subjects. We found that HAE was highly correlated with both total (r = 0.69, P < 0.0001) and ABCA1-specific (r = 0.47, P < 0.0001) efflux, and this relationship remained significant after adjustment for HDL-C or apoA-I. Multivariate models of sterol efflux capacity indicated that HAE accounted for approximately 25% of the model variance for both total and ABCA1-specific efflux. We conclude that the ability of HDL to exchange apoA-I and remodel, as measured by HAE, is a significant contributor to serum HDL efflux capacity, independent of HDL-C and apoA-I, indicating that HDL dynamics are an important factor in cholesterol efflux capacity and likely RCT.     

Cellular SR-BI and ABCA1-mediated cholesterol efflux are gender-specific in healthy subjects

We evaluated the impact of gender differences in both the quantitative and qualitative features of HDL subspecies on cellular free cholesterol efflux through the scavenger receptor class B type I (SR-BI), ABCA1, and ABCG1 pathways. For that purpose, healthy subjects (30 men and 26 women) matched for age, body mass index, triglyceride, apolipoprotein A-I, and high density lipoprotein-cholesterol (HDL-C) levels were recruited. We observed a significant increase (+14%; P < 0.03) in the capacity of whole sera from women to mediate cellular free cholesterol efflux via the SR-BI-dependent pathway compared with sera from men. Such enhanced efflux capacity resulted from a significant increase in plasma levels of large cholesteryl ester-rich HDL2 particles (+20%; P < 0.04) as well as from an enhanced capacity (+14%; P < 0.03) of these particles to mediate cellular free cholesterol efflux via SR-BI. By contrast, plasma from men displayed an enhanced free cholesterol efflux capacity (+31%; P < 0.001) via the ABCA1 transporter pathway compared with that from women, which resulted from a 2.4-fold increase in the plasma level of prebeta particles (P < 0.008). Moreover, in women, SR-BI-mediated cellular free cholesterol efflux was significantly correlated with plasma HDL-C (r = 0.72, P < 0.0001), whereas this relationship was not observed in men. In conclusion, HDL-C level may not represent the absolute indicator of the efficiency of the initial step of the reverse cholesterol transport.     

LDL-apheresis depletes apoE-HDL and pre-β1-HDL in familial hypercholesterolemia: relevance to atheroprotection

Subnormal HDL-cholesterol (HDL-C) and apolipoprotein (apo)AI levels are characteristic of familial hypercholesterolemia (FH), reflecting perturbed intravascular metabolism with compositional anomalies in HDL particles, including apoE enrichment. Does LDL-apheresis, which reduces HDL-cholesterol, apoAI, and apoE by adsorption, induce selective changes in HDL subpopulations, with relevance to atheroprotection? Five HDL subpopulations were fractionated from pre- and post-LDL-apheresis plasmas of normotriglyceridemic FH subjects (n = 11) on regular LDL-apheresis (>2 years). Apheresis lowered both plasma apoE (−62%) and apoAI (−16%) levels, with preferential, genotype-independent reduction in apoE. The mass ratio of HDL2:HDL3 was lowered from ∼1:1 to 0.72:1 by apheresis, reflecting selective removal of HDL2 mass (80% of total HDL adsorbed). Pre-LDL-apheresis, HDL2 subpopulations were markedly enriched in apoE, consistent with ∼1 copy of apoE per 4 HDL particles. Large amounts (50-66%) of apoE-HDL were removed by apheresis, preferentially in the HDL2b subfraction (−50%); minor absolute amounts of apoE-HDL were removed from HDL3 subfractions. Furthermore, pre-β1-HDL particle levels were subnormal following removal (−53%) upon apheresis, suggesting that cellular cholesterol efflux may be defective in the immediate postapheresis period. In LDL-receptor (LDL-R) deficiency, LDL-apheresis may enhance flux through the reverse cholesterol transport pathway and equally attenuate potential biglycan-mediated deposition of apoE-HDL in the arterial matrix.     

Zymosan-mediated inflammation impairs in vivo reverse cholesterol transport

Inflammation has been proposed to impair HDL function and reverse cholesterol transport (RCT). We investigated the effects of inflammation mediated by zymosan, a yeast glucan, on multiple steps along the RCT pathway in vivo and ex vivo. Acute inflammation with 70 mg/kg zymosan impaired RCT to plasma, liver, and feces similarly by 17-22% (P < 0.05), with no additional block at the liver. Hepatic gene expression further demonstrated no change in ABCG5, ABCB4, and ABCB11 expression but a decline in ABCG8 mRNA (32% P < 0.05). Plasma from zymosan-treated mice had a 21% decrease in cholesterol acceptor ability (P < 0.01) and a 35% decrease in ABCA1-specific efflux capacity (P < 0.01) in vitro. Zymosan treatment also decreased HDL levels and led to HDL remodeling with increased incorporation of serum amyloid A. In addition, cholesterol efflux from cultured macrophages declined with zymosan treatment in a dose dependent manner. Taken together, our results suggest that zymosan impairs in vivo RCT primarily by decreasing macrophage-derived cholesterol entering the plasma, with minimal additional blocks downstream. Our study supports the notion that RCT impairment is one of the mechanisms for the increased atherosclerotic burden observed in inflammatory conditions.     

Selective uptake of HDL cholesteryl esters and cholesterol efflux from mouse peritoneal macrophages independent of SR-BI

Scavenger receptor class B type I (SR-BI) mediates the selective uptake of HDL cholesteryl esters (CEs) and facilitates the efflux of unesterified cholesterol. SR-BI expression in macrophages presumably plays a role in atherosclerosis. The role of SR-BI for selective CE uptake and cholesterol efflux in macrophages was explored. Macrophages and HDL originated from wild-type (WT) or SR-BI knockout (KO; homozygous) mice. For uptake, macrophages were incubated in medium containing 125I-/3H-labeled HDL. For lipid removal, [3H]cholesterol efflux was analyzed using HDL as acceptor. Selective uptake of HDL CE ([3H]cholesteryl oleyl ether - 125I-tyramine cellobiose) was similar in WT and SR-BI KO macrophages. Radiolabeled SR-BI KO-HDL yielded a lower rate of selective uptake compared with WT-HDL in WT and SR-BI KO macrophages. Cholesterol efflux was similar in WT and SR-BI KO cells using HDL as acceptor. SR-BI KO-HDL more efficiently promoted cholesterol removal compared with WT-HDL from both types of macrophages. Macrophages selectively take up HDL CE independently of SR-BI. Additionally, in macrophages, there is substantial cholesterol efflux that is not mediated by SR-BI. Therefore, SR-BI-independent mechanisms mediate selective CE uptake and cholesterol removal. SR-BI KO-HDL is an inferior donor for selective CE uptake compared with WT-HDL, whereas SR-BI KO-HDL more efficiently promotes cholesterol efflux.     

ApoE promotes hepatic selective uptake but not RCT due to increased ABCA1-mediated cholesterol efflux to plasma

ApoE plays an important role in lipoprotein metabolism. This study investigated the effects of adenovirus-mediated human apoE overexpression (AdhApoE3) on sterol metabolism and in vivo reverse cholesterol transport (RCT). In wild-type mice, AdhApoE3 resulted in decreased HDL cholesterol levels and a shift toward larger HDL in plasma, whereas hepatic cholesterol content increased (P < 0.05). These effects were dependent on scavenger receptor class B type I (SR-BI) as confirmed using SR-BI-deficient mice. Kinetic studies demonstrated increased plasma HDL cholesteryl ester catabolic rates (P < 0.05) and higher hepatic selective uptake of HDL cholesteryl esters in AdhApoE3-injected wild-type mice (P < 0.01). However, biliary and fecal sterol output as well as in vivo macrophage-to-feces RCT studied with (3)H-cholesterol-loaded mouse macrophage foam cells remained unchanged upon human apoE overexpression. Similar results were obtained using hApoE3 overexpression in human CETP transgenic mice. However, blocking ABCA1-mediated cholesterol efflux from hepatocytes in AdhApoE3-injected mice using probucol increased biliary cholesterol secretion (P < 0.05), fecal neutral sterol excretion (P < 0.05), and in vivo RCT (P < 0.01), specifically within neutral sterols. These combined data demonstrate that systemic apoE overexpression increases i) SR-BI-mediated selective uptake into the liver and ii) ABCA1-mediated efflux of RCT-relevant cholesterol from hepatocytes back to the plasma compartment, thereby resulting in unchanged fecal mass sterol excretion and overall in vivo RCT.     

Physiological importance of SR-BI in the in vivo metabolism of human HDL and LDL in male and female mice

The physiological role of murine scavenger receptor class B type I (SR-BI) was evaluated by in vivo clearances of human HDL3 and LDL in normal and SR-BI knockout (KO) mice. In normal mice, cholesteryl esters (CEs) were removed faster than proteins, indicating a selective uptake process from both HDL3 and LDL. SR-BI KO mice showed 80% losses of HDL-CE selective uptake and the complete loss of LDL-CE selective uptake in the first phase of clearance. However, the second phase was characterized by an acceleration of CE disappearance in SR-BI KO mice. Thus, SR-BI is the only murine receptor mediating HDL-CE selective uptake, whereas a SR-BI-independent pathway specific to LDL can rescue SR-BI deficiency. The analysis of LDL recovered 3 h after injection in mice from different genotypes revealed that LDLs are significantly depleted in CE (reduction from 19% to 50% of the CE/protein ratios). A smaller LDL size in comparison with that of noninjected LDL was also detectable but was more evident for LDL recovered from normal mice. All LDL preparations migrate faster than noninjected LDL on agarose-barbital gels. Thus, both SR-BI-dependent and -independent pathways lead to substantial changes in LDL.     

SR-BI inhibits ABCG1-stimulated net cholesterol efflux from cells to plasma HDL

This study compares the roles of ABCG1 and scavenger receptor class B type I (SR-BI) singly or together in promoting net cellular cholesterol efflux to plasma HDL containing active LCAT. In transfected cells, SR-BI promoted free cholesterol efflux to HDL, but this was offset by an increased uptake of HDL cholesteryl ester (CE) into cells, resulting in no net efflux. Coexpression of SR-BI with ABCG1 inhibited the ABCG1-mediated net cholesterol efflux to HDL, apparently by promoting the reuptake of CE from medium. However, ABCG1-mediated cholesterol efflux was not altered in cholesterol-loaded, SR-BI-deficient (SR-BI(-/-)) macrophages. Briefly cultured macrophages collected from SR-BI(-/-) mice loaded with acetylated LDL in the peritoneal cavity did exhibit reduced efflux to HDL. However, this was attributable to reduced expression of ABCG1 and ABCA1, likely reflecting increased macrophage cholesterol efflux to apolipoprotein E-enriched HDL during loading in SR-BI(-/-) mice. In conclusion, cellular SR-BI does not promote net cholesterol efflux from cells to plasma HDL containing active LCAT as a result of the reuptake of HDL-CE into cells. Previous findings of increased atherosclerosis in mice transplanted with SR-BI(-/-) bone marrow probably cannot be explained by a defect in macrophage cholesterol efflux.     

Hypertriglyceridemia is associated with prebeta-HDL concentrations in subjects with familial low HDL

Prebeta-HDL particles act as the primary acceptors of cellular cholesterol in reverse cholesterol transport (RCT). An impairment of RCT may be the reason for the increased risk of coronary heart disease (CHD) in subjects with familial low HDL. We studied the levels of serum prebeta-HDL and the major regulating factors of HDL metabolism in 67 subjects with familial low HDL and in 64 normolipidemic subjects. We report that the subjects with familial low HDL had markedly reduced prebeta-HDL concentrations compared with the normolipidemic subjects (17.4 +/- 7.2 vs. 23.4 +/- 7.8 mg apolipoprotein A-I/dl; P < 0.001). A positive correlation was observed between prebeta-HDL concentration and serum triglyceride (TG) level (r = 0.334, P = 0.006). In addition, serum TG level was found to be the strongest predictor of prebeta-HDL concentration in subjects with familial low HDL. The activities of cholesteryl ester transfer protein and hepatic lipase were markedly increased in subjects with familial low HDL without a significant correlation to prebeta-HDL concentration. Our results support the hypothesis that impaired RCT is one mechanism behind the increased risk for CHD in subjects with familial low HDL.     

In vivo effects of anacetrapib on preβ HDL: improvement in HDL remodeling without effects on cholesterol absorption

Cholesteryl ester transfer protein (CETP) transfers cholesteryl ester and triglyceride between HDL and apoB-containing lipoproteins. Anacetrapib (ANA), a reversible inhibitor of CETP, raises HDL cholesterol and lowers LDL cholesterol in dyslipidemic patients. We previously demonstrated that ANA increases macrophage-to-feces reverse cholesterol transport and fecal cholesterol excretion in hamsters, and increased preβ HDL-dependent cholesterol efflux via ABCA1 in vitro. However, the effects of ANA on in vivo preβ HDL have not been characterized. In vitro, ANA inhibited the formation of preβ, however in ANA-treated dyslipidemic hamsters, preβ HDL levels (measured by two-dimensional gel electrophoresis) were increased, in contrast to in vitro findings. Because changes in plasma preβ HDL have been proposed to potentially affect markers of cholesterol absorption with other CETP inhibitors, a dual stable isotope method was used to directly measure cholesterol absorption in hamsters. ANA treatment of hamsters (on either dyslipidemic or normal diet) had no effect on cholesterol absorption, while dalcetrapib-treated hamsters displayed an increase in cholesterol absorption. Taken together, these data support the notion that ANA promotes preβ HDL functionality in vivo, with no effects on cholesterol absorption.     

Plasma cholesterol efflux capacity from human THP-1 macrophages is reduced in HIV-infected patients: impact of HAART

The capacity of HDL to remove cholesterol from macrophages is inversely associated with the severity of angiographic coronary artery disease. The effect of human immunodeficiency virus (HIV) infection or its treatment on the ability of HDL particles to stimulate cholesterol efflux from human macrophages has never been studied. We evaluated the capacity of whole plasma and isolated HDL particles from HIV-infected subjects (n = 231) and uninfected controls (n = 200), as well as in a subset of 41 HIV subjects receiving highly active antiretroviral therapy (HAART) to mediate cholesterol efflux from human macrophages. Plasma cholesterol efflux capacity was reduced (−12%; P = 0.001) in HIV patients as compared with controls. HIV infection reduced by 27% (P < 0.05) the capacity of HDL subfractions to promote cholesterol efflux from macrophages. We observed a reduced ABCA1-dependent efflux capacity of plasma (−27%; P < 0.0001) from HIV-infected subjects as a result of a reduction in the efflux capacity of HDL3 particles. HAART administration restored the capacity of plasma from HIV patients to stimulate cholesterol efflux from human macrophages (9.4%; P = 0.04). During HIV infection, the capacity of whole plasma to remove cholesterol from macrophages is reduced, thus potentially contributing to the increased coronary heart disease in the HIV population. HAART administration restored the removal of cholesterol from macrophages by increasing HDL functionality.     

Effects of amino acid substitutions at glycine 420 on SR-BI cholesterol transport function

Scavenger receptor class B type I (SR-BI) facilitates the uptake of HDL cholesteryl esters (CEs) in a two-step process involving binding of HDL to its extracellular domain and transfer of HDL core CEs to a metabolically active membrane pool, where they are subsequently hydrolyzed by a neutral CE hydrolase. Recently, we characterized a mutant, G420H, which replaced glycine 420 in the extracellular domain of SR-BI with a histidine residue and had a profound effect on SR-BI function. The G420H mutant receptor exhibited a reduced ability to mediate selective HDL CE uptake and was unable to deliver HDL CE for hydrolysis, despite the fact that it retained the ability to bind HDL. This did not hold true if glycine 420 was replaced with an alanine residue; G420A maintained wild-type HDL binding and cholesterol transport activity. To further understand the role that glycine 420 plays in SR-BI function and why there was a disparity between replacing glycine 420 with a histidine versus an alanine, we generated a battery of point mutants by substituting glycine 420 with amino acids possessing side chains that were charged, hydrophobic, polar, or bulky and tested the resulting mutants for their ability to support HDL binding, HDL cholesterol transport, and delivery for hydrolysis. The results indicated that substitution with a negatively charged residue or a proline impaired cell surface expression of SR-BI or its interaction with HDL, respectively. Furthermore, substitution of glycine 420 with a positively charged residue reduced HDL CE uptake as well as its subsequent hydrolysis.     

Changes in lipoprotein subfraction concentration and composition in healthy individuals treated with the CETP inhibitor anacetrapib

We investigated the effects of the cholesteryl ester (CE) transfer protein inhibitor anacetrapib (ANA) on plasma lipids, lipoprotein subfraction concentrations, and lipoprotein composition in 30 healthy individuals. Participants (n = 30) were randomized to ANA 20 mg/day, 150 mg/day, or placebo for 2 weeks. Changes in concentration of lipoprotein subfractions were assessed using ion mobility, and compositional analyses were performed on fractions separated by density gradient ultracentrifugation. ANA 150 mg/day versus placebo resulted in significant decreases in LDL-cholesterol (26%) and apo B (29%) and increases in HDL-cholesterol (82%). Concentrations of medium and small VLDL, large intermediate density lipoprotein (IDL), and medium and small LDL (LDL2a, 2b, and 3a) decreased whereas levels of very small and dense LDL4b were increased. There was enrichment of triglycerides and reduction of CE in VLDL, IDL, and the densest LDL fraction. Levels of large buoyant HDL particles were substantially increased, and there was enrichment of CE, apo AI, and apoCIII, but not apoAII or apoE, in the mid-HDL density range. Changes in lipoprotein subfraction concentrations and composition with ANA 20 mg/day were similar to those for ANA 150 mg/day but were generally smaller in magnitude. The impact of these changes on cardiovascular risk remains to be determined.     

Evaluation of HDL-modulating interventions for cardiovascular risk reduction using a systems pharmacology approach

The recent failures of cholesteryl ester transport protein inhibitor drugs to decrease CVD risk, despite raising HDL cholesterol (HDL-C) levels, suggest that pharmacologic increases in HDL-C may not always reflect elevations in reverse cholesterol transport (RCT), the process by which HDL is believed to exert its beneficial effects. HDL-modulating therapies can affect HDL properties beyond total HDL-C, including particle numbers, size, and composition, and may contribute differently to RCT and CVD risk. The lack of validated easily measurable pharmacodynamic markers to link drug effects to RCT, and ultimately to CVD risk, complicates target and compound selection and evaluation. In this work, we use a systems pharmacology model to contextualize the roles of different HDL targets in cholesterol metabolism and provide quantitative links between HDL-related measurements and the associated changes in RCT rate to support target and compound evaluation in drug development. By quantifying the amount of cholesterol removed from the periphery over the short-term, our simulations show the potential for infused HDL to treat acute CVD. For the primary prevention of CVD, our analysis suggests that the induction of ApoA-I synthesis may be a more viable approach, due to the long-term increase in RCT rate.     

Cholesteryl ester handling by RAW264 macrophages: response to native and acetylated low density lipoprotein

Summary The effects of LDL and Ac-LDL on the growth properties, morphology, and cholesteryl ester (CE) metabolism of the RAW264 macrophage cell line have been characterized. Cells were grown in media supplemented by a defined media (DM) mixture or fetal bovine serum (FBS). The addition of LDL or Ac-LDL to the culture media did not significantly alter cell growth properties. Cytoplasmic deposition of CE was observed by fluorescence microscopy in macrophages treated with LDL or Ac-LDL but not in untreated controls. Dose-response studies have shown that cholesteryl ester (CE) can accumulate in RAW264 treated with LDL. Cellular cholesterol content saturated at 4 hours with 50 g/ml LDL; this effect may be associated with receptor saturation. Dose-response studies conducted with Ac-LDL in DM have shown dramatic increases in total cell cholesterol content. However, deposition of CE was not observed below Ac-LDL concentrations of 100 g/ml. This indicates that a critical concentration of Ac-LDL must be reached to trigger deposition in DM. In contrast, no critical concentration of Ac-LDL was observed in macrophages grown in medium supplemented with 10% FBS. Cholesterol esterification in response to LDL and Ac-LDL was examined by ¹⁴C-oleic acid incorporation into CE. These results confirmed the mass cellular cholesterol and CE measurements. Kinetic studies conducted with RAW264 cells treated with 50 or 100 g/ml Ac-LDL resulted in a cholesterol efflux from the cells at 6–12 hours of incubation. Therefore, these studies show that (1) the nature of CE deposition is highly dependent upon the incubation media and (2) CE deposition is very sensitive to Ac-LDL concentration under certain conditions.Abbreviations LDL Low Density Lipoprotein - Ac-LDL Acetylated Low Density Lipoprotein - FBS Fetal Bovine Serum - DM Defined Medium - PBS Phosphate Buffered Saline - CE Cholesteryl Ester     

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-α 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.     

In vivo tissue cholesterol efflux is reduced in carriers of a mutation in APOA1

Atheroprotection by high density lipoprotein (HDL) is considered to be mediated through reverse cholesterol transport (RCT) from peripheral tissues. We investigated in vivo cholesterol fluxes through the RCT pathway in patients with low plasma high density lipoprotein cholesterol (HDL-c) due to mutations in APOA1. Seven carriers of the L202P mutation in APOA1 (mean HDL-c: 20 ± 19 mg/dl) and seven unaffected controls (mean HDL-c: 54 ± 11 mg/dl, P < 0.0001) received a 20 h infusion of ¹³C₂-cholesterol (¹³C-C). Enrichment of plasma and erythrocyte free cholesterol and plasma cholesterol esters was measured. With a three-compartment SAAM-II model, tissue cholesterol efflux (TCE) was calculated. TCE was reduced by 19% in carriers (4.6 ± 0.8 mg/kg/h versus 5.7 ± 0.7 mg/kg/h in controls, P = 0.02). Fecal ¹³C recovery and sterol excretion 7 days postinfusion did not differ significantly between carriers and controls: 21.3 ± 20% versus 13.3 ± 6.3% (P = 0.33), and 2,015 ± 1,431 mg/day versus 1456 ± 404 mg/day (P = 0.43), respectively. TCE is reduced in carriers of mutations in APOA1, suggesting that HDL contributes to efflux of tissue cholesterol in humans. The residual TCE and unaffected fecal sterol excretion in our severely affected carriers suggest, however, that non-HDL pathways contribute to RCT significantly.     

The effect of hepatic lipase on coronary artery disease in humans is influenced by the underlying lipoprotein phenotype

Increased or decreased hepatic lipase (HL) activity has been associated with coronary artery disease (CAD). This is consistent with the findings that gene variants that influence HL activity were associated with increased CAD risk in some population studies but not in others. In this review, we will explain the conditions that influence the effects of HL on CAD. Increased HL is associated with smaller and denser LDL (sdLDL) and HDL (HDL₃) particles, while decreased HL is associated with larger and more buoyant LDL and HDL particles. The effect of HL activity on CAD risk is dependent on the underlying lipoprotein phenotype or disorder. Central obesity with hypertriglyceridemia (HTG) is associated with high HL activity that leads to the formation of sdLDL that is pro-atherogenic. In the absence of HTG, where large buoyant cholesteryl ester-enriched LDL is prominent, elevation of HL does not raise the risk for CAD. In HTG patients, drug therapy that decreases HL activity selectively decreases sdLDL particles, an anti-atherogenic effect. Drug therapy that raises HDL₂ cholesterol has not decreased the risk for CAD. In trials where inhibition of cholesterol ester transfer protein (CETP) or HL occurs, the increase in HDL₂ most likely is due to inhibition of catabolism of HDL₂ and impairment of reverse cholesterol transport (RCT). In patients with isolated hypercholesterolemia, but with normal triglyceride levels and big-buoyant LDL particles, an increase in HL activity is beneficial; possibly because it increases RCT. Drugs that lower HL activity might decrease the risk for CAD only in hypertriglyceridemic patients with sdLDL by selectively clearing sdLDL particles from plasma, which would override the potentially pro-atherogenic effect on RCT. This article is part of a Special Issue entitled Advances in High Density Lipoprotein Formation and Metabolism: A Tribute to John F. Oram (1945-2010).