Cellular Cholesterol Efflux Research Articles

Dipeptidyl Peptidase-4 Inhibitor Sitagliptin Phosphate Accelerates Cellular Cholesterol Efflux in THP-1 Cells.

Cholesterol efflux is a major atheroprotective function of high-density lipoproteins (HDLs) which removes cholesterol from the foam cells of lipid-rich plaques in Type 2 diabetes. The dipeptidyl peptidase-4 (DPP-4) inhibitor sitagliptin phosphate increases plasma glucagon-like peptide-1 (GLP-1) concentrations and is used to treat Type 2 diabetes. GLP-1 plays an important role in regulating insulin secretion and expression via the GLP-1 receptor (GLP-1R), which is expressed in pancreatic islets as well as freshly isolated human monocytes and THP-1 cells. Here, we identified a direct role of GLP-1 and DPP-4 inhibition in HDL function. Cholesterol efflux was measured in cultivated phorbol 12-myristate 13-acetate-treated THP-1 cells radiolabeled with 3H-cholesterol and stimulated with liver X receptor/retinoid X receptor agonists. Contrary to vildagliptin, sitagliptin phosphate together with GLP-1 significantly (p < 0.01) elevated apolipoprotein (apo)A1-mediated cholesterol efflux in a dose-dependent manner. The sitagliptin-induced increase in cholesterol efflux did not occur in the absence of GLP-1. In contrast, adenosine triphosphate-binding cassette transporter A1 (ABCA1) mRNA and protein expressions in the whole cell fraction were not changed by sitagliptin in the presence of GLP-1, although sitagliptin treatment significantly increased ABCA1 protein expression in the membrane fraction. Furthermore, the sitagliptin-induced, elevated efflux in the presence of GLP-1 was significantly decreased by a GLP-1R antagonist, an effect that was not observed with a protein kinase A inhibitor. To our knowledge, the present study reports for the first time that sitagliptin elevates cholesterol efflux in cultivated macrophages and may exert anti-atherosclerotic actions that are independent of improvements in glucose metabolism. Our results suggest that sitagliptin enhances HDL function by inducing a de novo HDL synthesis via cholesterol efflux.

Administration of an LXR agonist promotes atherosclerotic lesion remodelling in murine inflammatory arthritis.

The leading cause of mortality in patients with rheumatoid arthritis is atherosclerotic cardiovascular disease (CVD). We have shown that murine arthritis impairs atherosclerotic lesion regression, because of cellular cholesterol efflux defects in haematopoietic stem and progenitor cells (HSPCs), causing monocytosis and impaired atherosclerotic regression. Therefore, we hypothesised that improving cholesterol efflux using a Liver X Receptor (LXR) agonist would improve cholesterol efflux and improve atherosclerotic lesion regression in arthritis. Ldlr -/- mice were fed a western-type diet for 14 weeks to initiate atherogenesis, then switched to a chow diet to induce lesion regression and divided into three groups; (1) control, (2) K/BxN serum transfer inflammatory arthritis (K/BxN) or (3) K/BxN arthritis and LXR agonist T0901317 daily for 2 weeks. LXR activation during murine inflammatory arthritis completely restored atherosclerotic lesion regression in arthritic mice, evidenced by reduced lesion size, macrophage abundance and lipid content. Mechanistically, serum from arthritic mice promoted foam cell formation, demonstrated by increased cellular lipid accumulation in macrophages and paralleled by a reduction in mRNA of the cholesterol efflux transporters Abca1, Abcg1 and Apoe. T0901317 reduced lipid loading and increased Abca1 and Abcg1 expression in macrophages exposed to arthritic serum and increased ABCA1 levels in atherosclerotic lesions of arthritic mice. Moreover, arthritic clinical score was also attenuated with T0901317. Taken together, we show that the LXR agonist T0901317 rescues impaired atherosclerotic lesion regression in murine arthritis because of enhanced cholesterol efflux transporter expression and reduced foam cell development in atherosclerotic lesions.

ApoJ/Clusterin concentrations are determinants of cerebrospinal fluid cholesterol efflux capacity and reduced levels are associated with Alzheimer’s disease

BackgroundAlzheimer’s disease (AD) shares risk factors with cardiovascular disease (CVD) and dysregulated cholesterol metabolism is a mechanism common to both diseases. Cholesterol efflux capacity (CEC) is an ex vivo metric of plasma high-density lipoprotein (HDL) function and inversely predicts incident CVD independently of other risk factors. Cholesterol pools in the central nervous system (CNS) are largely separate from those in blood, and CNS cholesterol excess may promote neurodegeneration. CEC of cerebrospinal fluid (CSF) may be a useful measure of CNS cholesterol trafficking. We hypothesized that subjects with AD and mild cognitive impairment (MCI) would have reduced CSF CEC compared with Cognitively Normal (CN) and that CSF apolipoproteins apoA-I, apoJ, and apoE might have associations with CSF CEC.MethodsWe retrieved CSF and same-day ethylenediaminetetraacetic acid (EDTA) plasma from 108 subjects (40 AD; 18 MCI; and 50 CN) from the Center for Neurodegenerative Disease Research biobank at the Perelman School of Medicine, University of Pennsylvania. For CSF CEC assays, we used N9 mouse microglial cells and SH-SY5Y human neuroblastoma cells, and the corresponding plasma assay used J774 cells. Cells were labeled with [3H]-cholesterol for 24 h, had ABCA1 expression upregulated for 6 h, were exposed to 33 μl of CSF, and then were incubated for 2.5 h. CEC was quantified as percent [3H]-cholesterol counts in medium of total counts medium+cells, normalized to a pool sample. ApoA-I, ApoJ, ApoE, and cholesterol were also measured in CSF.ResultsWe found that CSF CEC was significantly lower in MCI compared with controls and was poorly correlated with plasma CEC. CSF levels of ApoJ/Clusterin were also significantly lower in MCI and were significantly associated with CSF CEC. While CSF ApoA-I was also associated with CSF CEC, CSF ApoE had no association with CSF CEC. CSF CEC is significantly and positively associated with CSF Aβ. Taken together, ApoJ/Clusterin may be an important determinant of CSF CEC, which in turn could mitigate risk of MCI and AD risk by promoting cellular efflux of cholesterol or other lipids. In contrast, CSF ApoE does not appear to play a role in determining CSF CEC.

2-Hydroxypropyl-beta-cyclodextrin Treatment Does Not Induce Atherosclerotic Lesion Regression in Western-Type Diet-Fed Apolipoprotein E Knockout Mice

2-Hydroxypropyl-beta-cyclodextrin (2HPβCD) is able to bind and solubilize unesterified cholesterol and may therefore be able to reverse the deposition of cholesterol in macrophages within the aortic vessel wall, a hallmark of atherosclerotic cardiovascular disease. However, conflicting results regarding the potential of 2HPβCD to induce regression of established atherosclerotic lesions have been described. In the current study, we therefore also investigated the ability of 2HPβCD to stimulate cholesterol removal from macrophage foam cells in vitro and induce the regression of established atherosclerotic lesions in apolipoprotein E knockout (APOE KO) mice. In vitro studies using murine thioglycollate-elicited peritoneal macrophages verified that 2HPβCD is able to induce cholesterol efflux from macrophages in an ATP-binding cassette transporter-independent manner. Switching Western-type-diet-fed APOE KO mice with established atherosclerotic lesions back to a chow diet was associated with a reduction in the hypercholesterolemia extent and an increase in the absolute lesion size and plaque collagen-to-macrophage ratio. Importantly, parallel subcutaneous administration of 2HPβCD was not able to prevent the diet-switch-associated lesion growth or induce atherosclerosis regression. Although in our hands, 2HPβCD does effectively stimulate cellular cholesterol efflux from macrophages, we do not consider it worthwhile to further pursue 2HPβCD as therapeutic moiety in the atherosclerosis regression context.

LXRβ Activation Inhibits the Proliferation of Small-cell Lung Cancer Cells by Depleting Cellular Cholesterol.

Liver X receptors (LXRs) are nuclear receptors with various functions, including the regulation of cholesterol metabolism, glucose homeostasis, and inflammation. We previously reported that LXR activation inhibits the growth of oral cancer cells by inducing cellular cholesterol efflux and that LXRβ is expressed mainly in small-cell lung cancer (SCLC) tissues. SCLC is one of the most aggressive cancers, and identifying an effective therapeutic target molecule is desirable. Therefore, we investigated whether LXRβ could be an effective target molecule for SCLC treatment through in vitro experiments. We evaluated the influence of treatment with the LXR agonist T0901317 on cell proliferation and apoptosis in SCLC cell lines using cell viability, BrdU-ELISA, FACS, and western blot analyses. Moreover, the mechanism by which T0901317 inhibits SCLC cell proliferation was elucidated using qRT-PCR, western blot, a cholesterol quantification assay, and a genome editing technique. We showed that cultivated SCLC cells expressed LXRβ and that an LXR agonist inhibited the proliferation of SCLC cells without toxicity to normal cells. Furthermore, the antitumoral effect of an LXR agonist on SCLC cells was attributed to the induction of ABCA1 by LXRβ activation, resulting in an increase in cellular cholesterol efflux via ABCA1. The activation of LXRβ up-regulates ABCA1 expression, causing cholesterol depletion in cancer cells. This mechanism could be a novel target strategy for SCLC.

Retinoic acid increases the cellular cholesterol predominantly in a mTOR-independent manner.

Retinoic acid (RA) plays a role in the mounting immune response and controls several functions of the human body, including cholesterol homeostasis. The synthesis, uptake, and efflux of cellular cholesterol are significantly linked to the mammalian target of rapamycin complex-1 (mTORC1). Activation of mTORC1 promotes the synthesis and uptake of the cholesterol and suppresses its efflux, thus causing accumulation of cellular cholesterol. It is intriguing to know the effect of a high dose of RA on cholesterol accumulation in macrophages (mφ) and whether it is via mTOR activation. It is important to note that the long-term treatment of RA in humans is safe. Therefore, we chose a high dose of RA to observe its effect, which may be implicated in diseases like visceral leishmaniasis, where cholesterol deficiency is established. In the present study, we found the increased expression of RAPTOR, a regulatory component of the mTORC1 complex, in mφ upon treatment with RA. We observed the increased expression of SREBP2, LDLR, and PCSK9 in RA-treated mφ under sufficient cholesterol conditions, which further increased cellular cholesterol levels. Notably, their expressions were decreased when the mTOR pathway was inhibited by rapamycin. However, treatment with rapamycin did not result in the loss of cellular cholesterol in RA-treated mφ. Comparison with rapamycin-treated mφ suggests that RA induces cellular cholesterol levels in a mTORC1-independent manner.

PIP2 and cholesterol interplay in inflammation and atherosclerosis

IntroductionCholesterol accumulation in arterial macrophages leads to foam cell formation, Nlrp3 inflammasome activation, and IL‐1b release. Phosphatidylinositol 4, 5‐bisphosphate (PIP2) plays a role in the cholesterol efflux pathway and IL‐1b release. The PIP2 phosphatase TMEM55b modulates cholesterol efflux but the mechanism of this effect is not clear. Miltefosine altered PIP2 localization and blunted Nlrp3 inflammasome in macrophages, but the mechanism of its anti‐atherosclerotic properties is not clear.ObjectivesTo determine the effect of cellular PIP2 levels on ABCA1 mediated cholesterol efflux. To determine the mechanism by which Miltefosine reduces atherosclerosis in mice.Methods and resultsThe correlation between cellular PIP2 levels and ABCA1 mediated cholesterol efflux was determined. PIP2 levels were markedly increased in cholesterol‐loaded cells vs. control macrophages (~ 22 % increase, N=3, mean±SD, *P=0.05). Expression of TMEM55b, a PIP2 phosphatase, in BHK cells resulted in ~ 70 % decrease in ABCA1 mediated cholesterol efflux (N=4, mean ± SD, P&lt;0.0001). Miltefosine treated mice showed blunted in‐vivo Nlrp3 inflammasome assembly and had significantly lowered IL‐1b levels in peritoneal lavage (mean +SD; N=5, **p&lt;0.001). Miltefosine‐fed mice showed significantly higher reverse cholesterol transport to plasma, liver, and feces. The hyperlipidemic apoE‐/‐ KO mice fed with Miltefosine (50 mg/kg/day) gained significantly less weight vs. controls and showed significantly decreased atherosclerotic lesions (≥50% smaller vs. controls; mean + SD, p&lt;0.001 in females with N=10, and p&lt;0.04 in males with N=12). Miltefosine altered gut microbiota in hyperlipidemic apoE‐/‐ KO mice.ConclusionPIP2 regulates ABCA1 mediated cellular cholesterol efflux. Miltefosine can dampen in‐vivo Nlrp3 inflammasome assembly, increase reverse cholesterol transport, reduce high fat diet‐induced weight gain, reduce atherosclerotic lesion area and alter gut microbiota.

Restoring cellular NAD(P)H levels by PPARα and LXRα stimulation to improve mitochondrial complex I deficiency

Mitochondrial complex I (CI), the first multiprotein enzyme complex of the oxidative phosphorylation system, plays a crucial role in cellular energy production. CI deficiency is associated with a variety of clinical phenotypes, including Leigh syndrome. At the cellular level, an increased NAD(P)H concentration is one of the hallmarks in CI-deficiency. AimsHere, we aimed to attenuate increased NAD(P)H levels by stimulation of ATP-dependent cassette (ABC)A1 and ABCG1-mediated cellular cholesterol efflux with various PPARα and LXRα agonists. Main methodsMitochondrial CI-deficient fibroblasts and chemically-induced CI-deficient HeLa cells were used to study the dose-dependent effects of various PPARα and LXRα agonists on cellular NAD(P)H levels and cholesterol efflux. Key findingsIn patient-derived mitochondrial CI-deficient fibroblasts, GW590735, astaxanthin, oleoylethanolamide, and GW3965 significantly reduced the enhanced NAD(P)H levels in CI-deficient fibroblasts. Similar effects were observed in chemically-induced CI-impaired HeLa cells, in which BMS-687453, Wy14643, GW7647, T0901317, DMHCA also demonstrated a beneficial effect. Surprisingly, no effect on ABCA1- and ABCG1-mediated cholesterol efflux in HeLa cells and fibroblasts was found after treatment with these compounds. The reduction in NAD(P)H levels by GW590735 could be partially reversed by inhibition of fatty acid synthase and β-oxidation, which suggests that its beneficial effects are possibly mediated via stimulation of fatty acid metabolism rather than cholesterol efflux. SignificanceCollectively, PPARα and LXRα stimulation resulted in attenuated cellular NAD(P)H levels in CI-impaired HeLa cells and patient-derived fibroblasts and could eventually have a therapeutic potential in CI deficiency.

Pathways and Mechanisms of Cellular Cholesterol Efflux-Insight From Imaging.

Cholesterol is an essential molecule in cellular membranes, but too much cholesterol can be toxic. Therefore, mammalian cells have developed complex mechanisms to remove excess cholesterol. In this review article, we discuss what is known about such efflux pathways including a discussion of reverse cholesterol transport and formation of high-density lipoprotein, the function of ABC transporters and other sterol efflux proteins, and we highlight their role in human diseases. Attention is paid to the biophysical principles governing efflux of sterols from cells. We also discuss recent evidence for cholesterol efflux by the release of exosomes, microvesicles, and migrasomes. The role of the endo-lysosomal network, lipophagy, and selected lysosomal transporters, such as Niemann Pick type C proteins in cholesterol export from cells is elucidated. Since oxysterols are important regulators of cellular cholesterol efflux, their formation, trafficking, and secretion are described briefly. In addition to discussing results obtained with traditional biochemical methods, focus is on studies that use established and novel bioimaging approaches to obtain insight into cholesterol efflux pathways, including fluorescence and electron microscopy, atomic force microscopy, X-ray tomography as well as mass spectrometry imaging.

In Vivo Cerebral Translocator Protein (TSPO) Binding and Its Relationship with Blood Adiponectin Levels in Treatment-Naïve Young Adults with Major Depression: A [11C]PK11195 PET Study.

Adiponectin is an adipokine that mediates cellular cholesterol efflux and plays important roles in neuroinflammatory processes. In this study, we undertook positron emission tomography (PET) with the translocator protein (TSPO) ligand [11C]PK11195 and measured serum adiponectin levels in groups of treatment-naïve young adult patients with major depressive disorder (MDD) and matched healthy controls. Thirty treatment-naïve MDD patients (median age: 24 years) and twenty-three healthy controls underwent [11C]PK11195 PET. We quantified TSPO availability in brain as the [11C]PK11195 binding potential (BPND) using a reference tissue model in conjunction with the supervised cluster analysis (SVCA4) algorithm. Age, sex distribution, body mass index, and serum adiponectin levels did not differ between the groups. Between-group analysis using a region-of-interest approach showed significantly higher [11C]PK11195 BPND in the left anterior and right posterior cingulate cortices in MDD patients than in controls. Serum adiponectin levels had significant negative correlations with [11C]PK11195 BPND in the bilateral hippocampus in MDD patients, but significant positive correlations in the bilateral hippocampus in the control group. Our results indicate significantly higher TSPO binding in the anterior and posterior cingulate cortices in treatment-naïve young MDD patients, suggesting microglial activation in these limbic regions, which are involved in cognitive and emotional processing. The opposite correlations between [11C]PK11195 BPND in the hippocampus with serum adiponectin levels in MDD and control groups suggest that microglial activation in the hippocampus may respond differentially to adiponectin signaling in MDD and healthy subjects, possibly with respect to microglial phenotype.

Abstract 10706: Alpha Aminoadipic Acid Down-Regulates Sr-bi Abca1 and Abcg1 Expression and May Regulate Plasma Lipid Profiles, Cellular Cholesterol Efflux, and Efferocytosis

Introduction: The metabolite α-aminoadipic acid (2-AAA) has been identified as a predictor of diabetes and atherosclerosis in large human prospective studies. However, the mechanisms linking this metabolite to disease pathophysiology remain unknown. Efferocytosis, a macrophage-mediated process for clearance of apoptotic cells (ACs), is critical in preventing necrosis of dead cells and triggers anti-inflammatory and pro-resolving responses in phagocytes. Cholesterol efflux from peripheral cells in atherosclerotic plaque is a critical step in Reverse Cholesterol Transport (RCT). We hypothesized that the 2-AAA pathway relates to atherosclerosis through modulation of plasma lipid profiles, cholesterol efflux, and macrophage efferocytosis. Results: We found a significant negative correlation between plasma 2-AAA and HDL cholesterol (r=-0.53, P&lt;0.0001), and a significant positive correlation with triglycerides (r=0.35, P=0.006), in two separate samples of individuals with and without cardiometabolic disease (N=98), suggesting a relationship between 2-AAA and cardiovascular risk. We investigated the consequences of 2-AAA stimulation in several primary cells and cell lines. We found that 2-AAA stimulation downregulates three key genes (SR-BI (up to 86% reduction, P=0.005), ABCA1 (up to 92% reduction, P=0.007), ABCG1 (up to 94% reduction, P=0.008)) that directly relate to cholesterol efflux in THP-1 macropohages, suggesting an important role of 2-AAA in cholesterol efflux. Next, we found that treatment with 2-AAA at physiologically-relevant doses (30μM and 100μM) for times ranging from 30 minutes to 6 hours led to consistent reductions of efferocytosis of up to 60% (P&lt;0.001) in mouse macrophages, J774 cells and Raw 264.7 cells, suggesting a pivotal role of 2-AAA in efferocytosis. Conclusions: Our data suggest 2-AAA is associated with plasma lipid profiles (HDL, triglycerides), cholesterol efflux, and macrophage efferocytosis. These novel data could provide a new target for the treatment of atherosclerosis.

MicroRNA-33-5p inhibits cholesterol efflux in vascular endothelial cells by regulating citrate synthase and ATP-binding cassette transporter A1

BackgroundA high level of total cholesterol is associated with several lipid metabolism disorders, including atherosclerosis and cardiovascular diseases. ATP-binding cassette (ABC) transporter A1 (ABCA1) and miR-33-5p play crucial roles in atherosclerosis by controlling cholesterol efflux. While citrate is a precursor metabolite for lipid and cholesterol synthesis, little is known about the association between citrate synthase (CS) and cholesterol efflux. This study investigated the role of the miR-33-5p/ABCA1/CS axis in regulating cholesterol efflux in vascular endothelial cells (VECs).Materials and methodsVECs were treated with oxidized low-density lipoprotein cholesterol (ox-LDL), or pretreated with plasmids overexpressing CS, ABCA1, siRNAs against CS and ABCA1, and an miR-33-5p inhibitor. Cell apoptosis, cellular senescence-associated β-galactosidase activity, inflammation, and cholesterol efflux were detected.ResultsTreatment with ox-LDL decreased ABCA1 and CS levels and increased miR-33-5p expression and apoptosis in dose-dependent manners. In contrast, treatment with the miR-33-5p inhibitor and ABCA1 and CS overexpression plasmids inhibited the above-mentioned ox-LDL-induced changes. In addition, treatment with ox-LDL decreased cholesterol efflux, induced aging, and promoted the production of inflammatory cytokines (i.e., IL-6 and tumor necrosis factor TNF-α), as well as the expression of Bax and Caspase 3 proteins in VECs. All these changes were rescued by miR-33-5p inhibition and ABCA1 and CS overexpression. The inhibition of ABCA1 and CS by siRNAs eliminated the effects mediated by the miR-33-5p inhibitor, and knockdown of CS eliminated the effects of ABCA1 on VECs.ConclusionsThis study demonstrated the crucial roles played by the miR-33-5p/ABCA1/CS axis in regulating cholesterol efflux, inflammation, apoptosis, and aging in VECs, and also suggested the axis as a target for managing lipid metabolism disorders.

Hormone-sensitive lipase deficiency affects the expression of SR-BI, LDLr, and ABCA1 receptors/transporters involved in cellular cholesterol uptake and efflux and disturbs fertility in mouse testis

Hormone-sensitive lipase (HSL) hydrolyse acylglycerols, cholesteryl and retinyl esters. HSL is a key lipase in mice testis, as HSL deficiency results in male sterility. The present work study the effects of the deficiency and lack of HSL on the localization and expression of SR-BI, LDLr, and ABCA1 receptors/transporters involved in uptake and efflux of cholesterol in mice testis, to determine the impact of HSL gene dosage on testis morphology, lipid homeostasis and fertility. The results of this work show that the lack of HSL in mice alters testis morphology and spermatogenesis, decreasing sperm counts, sperm motility and increasing the amount of Leydig cells and lipid droplets. They also show that there are differences in the localization of HSL, SR-BI, LDLr and ABCA1 in HSL+/+, HSL+/− and HSL−/− mice. The deficiency or lack of HSL has effects on protein and mRNA expression of genes involved in lipid metabolisms in mouse testis. HSL−/− testis have augmented expression of SR-BI, LDLr, ABCA1 and LXRβ, a critical sterol sensor that regulate multiple genes involved in lipid metabolism; whereas LDLr expression decreased in HSL+/− mice. Plin2, Abca1 and Ldlr mRNA levels increased; and LXRα (Nr1h3) and LXRβ (Nr1h2) decreased in testis from HSL−/− compared with HSL+/+; with no differences in Scarb1. Together these data suggest that HSL deficiency or lack in mice testis induces lipid homeostasis alterations that affect the cellular localization and expression of key receptors/transporter involved in cellular cholesterol uptake and efflux (SR-BI, LDRr, ABCA1); alters normal cellular function and impact fertility.

Compounds targeting OSBPL7 increase ABCA1-dependent cholesterol efflux preserving kidney function in two models of kidney disease

Impaired cellular cholesterol efflux is a key factor in the progression of renal, cardiovascular, and autoimmune diseases. Here we describe a class of 5-arylnicotinamide compounds, identified through phenotypic drug discovery, that upregulate ABCA1-dependent cholesterol efflux by targeting Oxysterol Binding Protein Like 7 (OSBPL7). OSBPL7 was identified as the molecular target of these compounds through a chemical biology approach, employing a photoactivatable 5-arylnicotinamide derivative in a cellular cross-linking/immunoprecipitation assay. Further evaluation of two compounds (Cpd A and Cpd G) showed that they induced ABCA1 and cholesterol efflux from podocytes in vitro and normalized proteinuria and prevented renal function decline in mouse models of proteinuric kidney disease: Adriamycin-induced nephropathy and Alport Syndrome. In conclusion, we show that small molecule drugs targeting OSBPL7 reveal an alternative mechanism to upregulate ABCA1, and may represent a promising new therapeutic strategy for the treatment of renal diseases and other disorders of cellular cholesterol homeostasis.

Recent developments in the regulation of cholesterol transport by natural molecules

The dysregulation of cholesterol metabolism is a high-risk factor for non-alcoholic fatty liver disease (NAFLD), dyslipidemia, and atherosclerosis (AS). Cholesterol transport maintains whole-body cholesterol homeostasis. Low-density apolipoprotein receptor (LDLR) mediates cholesterol uptake in cells and plays an important role in the primary route of circulatory cholesterol clearance in liver cells. Caveolins 1 is an integral membrane protein and shuttle between the cytoplasm and cell membrane. Caveolins 1 not only plays a role in promoting cholesterol absorption in cells but also in the transport of cellular cholesterol efflux by interacting with the ATP-binding cassette transporter A1 (ABCA1) and scavenger receptor class B type I (SR-BI). These proteins, which are associated with reverse cholesterol transport (RCT), are potential therapeutic targets for NAFLD and AS. Many studies have indicated that natural products have lipid-lowering effects. Moreover, natural molecules, derived from natural products, have the potential to be developed into novel drugs. However, the mechanisms underlying the regulation of cholesterol transport by natural molecules have not yet been adequately investigated. In this review, we briefly describe the process of cholesterol transport and summarize the mechanisms by which molecules regulate cholesterol transport. This article provides an overview of recent studies and focuses on the potential therapeutic effects of natural molecules; however, further high-quality studies are needed to firmly establish the clinical efficacies of natural molecules.

Editorial expression of concern: “MicroRNA-27a/b regulates cellular cholesterol efflux, influx and esterification/hydrolysis in THP-1 macrophages” [Atherosclerosis Volume 234, Issue 1, May 2014, Pages 54-64

Editorial expression of concern: “MicroRNA-27a/b regulates cellular cholesterol efflux, influx and esterification/hydrolysis in THP-1 macrophages” [Atherosclerosis Volume 234, Issue 1, May 2014, Pages 54-64

Apolipoprotein M promotes cholesterol uptake and efflux from mouse macrophages.

Apolipoprotein M (ApoM) exhibits various anti‐atherosclerotic functions as a component of high‐density lipoprotein (HDL) particles. Scavenger receptor class B type I (SR‐BI) is a classic HDL receptor that mediates selective cholesterol uptake and enhances the efflux of cellular cholesterol to HDL. However, the effect of ApoM on cholesterol transport in macrophages remains unclear. In this study, we identified for the first time that ApoM is expressed in mouse macrophages and is involved in cholesterol uptake, similar to SR‐BI. NBD‐cholesterol uptake and efflux in cells were characterized using fluorescence spectrophotometry. The uptake ratios of cholesterol by macrophages from ApoM −/− SR‐BI−/− mice were significantly lower than those from ApoM+/+SR‐BI −/− and ApoM −/− SR‐BI+/+ mice. Real‐time fluorescence quantitative PCR was used to analyze the expression of cholesterol transport‐related genes involved in cholesterol uptake. ApoM‐enriched HDL (ApoM+HDL) facilitated more cholesterol efflux from murine macrophage Ana‐1 cells than ApoM‐free HDL (ApoM−HDL). However, recombinant human ApoM protein inhibited the ability of ApoM−HDL to induce cholesterol efflux. In conclusion, ApoM promotes cholesterol uptake and efflux in mouse macrophages. A better understanding of ApoM function may lead to the development of novel therapeutic strategies for treating atherosclerotic diseases.

Myeloperoxidase Targets Apolipoprotein A-I for Site-Specific Tyrosine Chlorination in Atherosclerotic Lesions and Generates Dysfunctional High-Density Lipoprotein.

We previously demonstrated that apolipoprotein A-I (apoA-I), the major protein component of high-density lipoprotein (HDL), is an important target for myeloperoxidase (MPO)-catalyzed tyrosine chlorination in the circulation of subjects with cardiovascular diseases. Oxidation of apoA-I by MPO has been reported to deprive HDL of its protective properties. However, the potential effects of MPO-mediated site-specific tyrosine chlorination of apoA-I on dysfunctional HDL formation and atherosclerosis was unclear. Herein, Tyr192 in apoA-I was found to be the major chlorination site in both lesion and plasma HDL from humans with atherosclerosis, while MPO binding to apoA-I was demonstrated by immunoprecipitation studies in vivo. In vitro, MPO-mediated damage of lipid-free apoA-I impaired its ability to promote cellular cholesterol efflux by the ABCA1 pathway, whereas oxidation to lipid-associated apoA-I inhibited lecithin:cholesterol acyltransferase activation, two key steps in reverse cholesterol transport. Compared with native apoA-I, apoA-I containing a Tyr192 → Phe mutation was moderately resistant to oxidative inactivation by MPO. In high-fat-diet-fed apolipoprotein E-deficient mice, compared with native apoA-I, subcutaneous injection with oxidized apoA-I (MPO treated) failed to mediate the lipid content in aortic plaques while mutant apoA-I (Tyr192 → Phe) showed a slightly stronger ability to reduce the lipid content in vivo. Our observations suggest that oxidative damage of apoA-I and HDL involves MPO-dependent site-specific tyrosine chlorination, raising the feasibility of producing MPO-resistant forms of apoA-I that have stronger antiatherosclerotic activity in vivo.

The benzoate plant metabolite ethyl gallate prevents cellular- and vascular-lipid accumulation in experimental models of atherosclerosis

Ethyl gallate (EG) is a well-known constituent of medicinal plants, but its effects on atherosclerosis development are not clear. In the present study, the anti-atherosclerosis effects of EG and the underlying mechanisms were explored using macrophage cultures, zebrafish and apolipoprotein (apo) E deficient mice. Treatment of macrophages with EG (20 μM) enhanced cellular cholesterol efflux to HDL, and reduced net lipid accumulation in response to oxidized LDL. Secretion of monocyte chemotactic protein-1 (MCP-1) and interleukin-6 (IL-6) from activated macrophages was also blunted by EG. Fluorescence imaging techniques revealed EG feeding of zebrafish reduced vascular lipid accumulation and inflammatory responses in vivo. Similar results were obtained in apoE-/- mice 6.5 months of age, where plaque lesions and monocyte infiltration into the artery wall were reduced by 70% and 42%, respectively, after just 6 weeks of injections with EG (20 mg/kg). HDL-cholesterol increased 2-fold, serum cholesterol efflux capacity increased by ∼30%, and the levels of MCP-1 and IL-6 were reduced with EG treatment of mice. These results suggest EG impedes early atherosclerosis development by reducing the lipid and macrophage-content of plaque. Underlying mechanisms appeared to involve HDL cholesterol efflux mechanisms and suppression of pro-inflammatory cytokine secretion.

Effect of Tocilizumab on LDL and HDL Characteristics in Patients with Rheumatoid Arthritis. An Observational Study.

BackgroundIn patients with rheumatoid arthritis (RA), qualitative alterations of low and high-density lipoproteins (LDL and HDL, respectively) might partially explain their increased cardiovascular risk. Tocilizumab has been associated with an increase in lipids, including triglyceride (TG) and cholesterol levels. The aim of this study is to evaluate the effect of tocilizumab on certain LDL and HDL characteristics (oxidized LDL levels, HDL-associated enzymes, chemical composition of both total HDL and HDL3c subpopulation, and their capacity to promote cellular cholesterol efflux) at baseline and 3 months after the start of treatment in patients with RA.MethodsTwenty-eight RA patients (ACR/EULAR 2010 criteria) with indication of treatment with tocilizumab were included in the present study. Clinical assessment [Health assessment questionnaire (HAQ)], disease activity score 28 (DAS28), high-sensitivity C reactive protein (hsCRP) concentration, lipid profile, and lipoprotein (a) [Lp(a)] levels were evaluated in all patients at baseline and after 3 months of treatment with tocilizumab. Lipoprotein characteristics were evaluated through the levels of oxidized LDL (OxLDL), the activity of paraoxonase (PON) 1, the composition of total HDL and small, dense HDL3c subpopulation, and their ability to promote cellular cholesterol efflux.ResultsAfter 3 months of treatment with tocilizumab, HAQ (− 23%, p < 0.05), DAS28 (− 49%, p < 0.001), and hsCRP (− 94%, p < 0.01) levels decreased significantly. Total cholesterol (TC), LDL-C, non-HDL-C, and apo B levels showed a significant increase after treatment (TC: + 7.0%, p < 0.01; LDL-C: + 10%, p < 0.01; non-HDL-C: + 9.9%, p < 0.01; and apo B: + 9.6%, p < 0.05). Decreases in Lp(a) and OxLDL levels were also observed after treatment [Lp(a): − 50%, p < 0.01; and oxLDL: − 5.4%, p < 0.05]. The latter was in accordance with the increment detected in PON activity. No changes were observed in HDL capacity to promote cholesterol efflux (p > 0.05) in the whole group.ConclusionsTreatment with tocilizumab reduced hsCRP levels and displayed positive effects on certain lipoprotein-related parameters, such as a potent decrease inLp(a) and a reduction in OxLDL levels. Moreover, HDL capacity to promote cellular cholesterol efflux was maintained after 3 months of treatment.