Cholesteryl Ester Accumulation In Macrophages Research Articles

Low Doses of Innate Defence Regulator Peptide, IDR-1018, Enhances HDL-Mediated Cholesterol Efflux from Smooth Muscle Cells and Macrophages

Cardiovascular diseases due to atherosclerosis are the leading cause of death worldwide. In recent years, novel therapeutics designed to enhance high-density lipoprotein (HDL) activity as a treatment for atherosclerosis have been explored. Of particular interest is the use of amphipathic α-helical peptides to mimic the action of the main HDL protein, apolipoprotein (Apo)AI. Immunomodulatory peptides share many physical and functional
 properties with ApoAI and its mimetics; we therefore hypothesized that they too might be capable of enhancing cholesterol efflux. The aim of this study was to determine whether a potent immunomodulatory peptide, innate defence regulator (IDR)-1018, could promote cholesterol efflux from cells. Here, we report that IDR-1018 induced a dose-dependent increase in ApoAI on the cell surface and ATP-binding cassette transporter A1 (ABCA1)
 protein levels. Functional assays revealed that low doses of IDR-1018 improved HDL-mediated suppression of intracellular cholesteryl ester accumulation in macrophages and enhanced HDL-mediated cellular cholesterol efflux from smooth muscle cells and macrophages. Based on these results we propose that natural and synthetic immunomodulatory peptides like IDR-1018 represent a large new group of peptides that could be developed for
 the treatment of atherosclerosis.

Akt3 Deficiency in Macrophages Promotes Foam Cell Formation and Atherosclerosis in Mice

Akt, a serine-threonine protein kinase, exists as three isoforms. The Akt signaling pathway controls multiple cellular functions in the cardiovascular system, and the atheroprotective endothelial cell-dependent role of Akt1 has been recently demonstrated. The role of Akt3 isoform in cardiovascular pathophysiology is not known. We explored the role of Akt3 in atherosclerosis using mice with a genetic ablation of the Akt3 gene. Using hyperlipidemic ApoE(-/-) mice, we demonstrated a macrophage-dependent, atheroprotective role for Akt3. In vitro experiments demonstrated differential subcellular localization of Akt1 and Akt3 in macrophages and showed that Akt3 specifically inhibits macrophage cholesteryl ester accumulation and foam cell formation, a critical early event in atherogenesis. Mechanistically, Akt3 suppresses foam cell formation by reducing lipoprotein uptake and promoting ACAT-1 degradation via the ubiquitin-proteasome pathway. These studies demonstrate the nonredundant atheroprotective role for Akt3 exerted via the previously unknown link between the Akt signaling pathway and cholesterol metabolism.

Improved Insulin Sensitivity in High Fat- and High Cholesterol-fed Ldlr−/− Mice with Macrophage-specific Transgenic Expression of Cholesteryl Ester Hydrolase: ROLE OF MACROPHAGE INFLAMMATION AND INFILTRATION INTO ADIPOSE TISSUE

Cellular cholesterol balance induces changes in the inflammatory status of macrophages, and low grade chronic inflammation is increasingly being recognized as one of the key steps in the development of atherosclerosis as well as insulin resistance. Cholesteryl ester hydrolase (CEH) catalyzes the hydrolysis of intracellular stored cholesteryl esters (CEs) and thereby enhances free cholesterol efflux and reduces cellular CE content. We have earlier reported reduced atherosclerosis and lesion necrosis in macrophage-specific CEH transgenic mice on a Ldlr(-/-) background. In the present study, we tested the hypothesis that reduced intracellular accumulation of CE in macrophages from CEH transgenic mice will attenuate expression of proinflammatory mediators, thereby reducing infiltration into adipose tissue, alleviating inflammation, and resulting in improved insulin sensitivity. Western diet fed Ldlr(-/-)CEH transgenic mice showed improved insulin sensitivity as assessed by glucose and insulin tolerance tests. Macrophages from CEH transgenic mice expressed significantly lower levels of proinflammatory cytokines (interleukin-1beta and interleukin-6) and chemokine (MCP-1; monocyte chemoattractant protein). Attenuation of NF-kappaB- and AP-1-driven gene expression was determined to be the underlying mechanism. Infiltration of macrophages into the adipose tissue that increases inflammation and impairs insulin signaling was also significantly reduced in Ldlr(-/-)CEH transgenic mice. In the OP-9 adipocyte peritoneal macrophage co-culture system, macrophages from CEH transgenic mice had a significantly reduced effect on insulin signaling as measured by Akt phosphorylation compared with nontransgenic macrophages. Taken together, these studies demonstrate that macrophage-specific overexpression of CEH decreases expression of proinflammatory mediators and attenuates macrophage infiltration into the adipose tissue, resulting in decreased circulating cytokines and improved insulin sensitivity.

Reinioside C, a triterpene saponin of Polygala aureocauda Dunn, exerts hypolipidemic effect on hyperlipidemic mice

Reinioside C is a triterpene saponin from the the root of Polygala aureocauda Dunn (PAD). This study examined the effects of reinioside C on hyperlipidemic mice in vivo and endothelium cells, macrophages and smooth muscle cells in vitro. Mice were given a hyperlipidemic diet for 30 days, then administered reinioside C (4, 8, 16 mg/kg/day, p.o.) for 30 days. Then the serum lipid, superoxide dismutase (SOD), malonaldehyde (MDA), the total cholesterol (TC) and triglyceride (TG) in the liver extract were measured. Human umbilical vein endothelial (HUVECs), peritoneal macrophages and smooth muscle cells (SMCs) pre-treated with reinioside C were treated with oxidized low-density lipoprotein (OxLDL). The results showed that reinioside C decreased serum and liver tissue lipid profiles in hyperlipidemic mice. Moreover, reinioside C protected the HUVECs against the Ox-LDL induced LDH leakage and exerted a protective effect on oxidative lesions induced by OxLDL, inhibited cholesteryl ester accumulation in macrophages, and decreased [Ca2+](i) and SMC proliferation in vitro. Based on these results, it is suggested that reinioside C is a promising hypolipidemic candidate.

Estrogen prevents cholesteryl ester accumulation in macrophages induced by the HIV protease inhibitor ritonavir

Individuals with HIV can now live long lives with drug therapy that often includes protease inhibitors such as ritonavir. Many patients, however, develop negative long-term side effects such as premature atherosclerosis. We have previously demonstrated that ritonavir treatment increases atherosclerotic lesion formation in male mice to a greater extent than in female mice. Furthermore, peripheral blood monocytes isolated from ritonavir-treated females had less cholesteryl ester accumulation. In the present study, we have investigated the molecular mechanisms by which female hormones influence cholesterol metabolism in macrophages in response to the HIV protease inhibitor ritonavir. We have utilized the human monocyte cell line, THP-1 as a model to address this question. Briefly, cells were differentiated for 72 h with 100 nM PMA to obtain a macrophage-like phenotype in the presence or absence of 1 nM 17beta-estradiol (E2), 100 nM progesterone or vehicle (0.01% ethanol). Cells were then treated with 30 ng/ml ritonavir or vehicle in the presence of aggregated LDL for 24 h. Cell extracts were harvested, and lipid or total RNA was isolated. E2 decreased the accumulation of cholesteryl esters in macrophages following ritonavir treatment. Ritonavir increased the expression of the scavenger receptor, CD36 mRNA, responsible for the uptake of LDL. Additionally, ritonavir treatment selectively increased the relative levels of PPARgamma mRNA, a transcription factor responsible for the regulation of CD36 mRNA expression. Treatment with E2, however, failed to prevent these increases at the mRNA level. E2 did, however, significantly suppress CD36 protein levels as measured by fluorescent immunocytochemistry. This data suggests that E2 modifies the expression of CD36 at the level of protein expression in monocyte-derived macrophages resulting in reduced cholesteryl ester accumulation following ritonavir treatment.

Phagocytosis of Cholesteryl Ester Is Amplified in Diabetic Mouse Macrophages and Is Largely Mediated by CD36 and SR-A

Type 2 diabetes (T2D) is associated with accelerated atherosclerosis, which accounts for approximately 75% of all diabetes-related deaths. Here we investigate the link between diabetes and macrophage cholesteryl ester accumulation. When diabetic (db/db) mice are given cholesteryl ester intraperitoneally (IP), peritoneal macrophages (PerMΦs) recovered from these animals showed a 58% increase in intracellular cholesteryl ester accumulation over PerMΦs from heterozygote control (db/+) mice. Notably, PerMΦ fluid-phase endocytosis and large particle phagocytosis was equivalent in db/+and db/db mice. However, IP administration of CD36 and SR-A blocking antibodies led to 37% and 25% reductions in cholesteryl ester accumulation in PerMΦ. Finally, in order to determine if these scavenger receptors (SRs) were part of the mechanism responsible for the increased accumulation of cholesteryl esters observed in the diabetic mouse macrophages, receptor expression was quantified by flow cytometry. Importantly, db/db PerMΦs showed a 43% increase in CD36 expression and an 80% increase in SR-A expression. Taken together, these data indicate that direct cholesteryl ester accumulation in mouse macrophages is mediated by CD36 and SR-A, and the magnitude of accumulation is increased in db/db macrophages due to increased scavenger receptor expression.

β-Cryptoxanthin, a novel natural RAR ligand, induces ATP-binding cassette transporters in macrophages

Despite its serious adverse effects, recent accumulating evidence suggests that a physiological retinoic acid receptor (RAR) agonist, all- trans retinoic acid (atRA), exhibits preventive effects on atherogenesis. Therefore, the present study was designed to explore novel natural RAR ligands with anti-atherogenic effects in order to identify and develop a drug without severe side effects. Among xanthophylls and carotenoids studied, β-cryptoxanthin and lutein exhibited RAR ligand activity in yeast two-hybrid system that was found to be completely abolished by the RAR pan-antagonist LE540. Furthermore, these molecules can bind the RAR ligand-binding domain in the CoA-BAP system but not RXR ligand-binding domain. These results indicate that both β-cryptoxanthin and lutein serve as ligands for RAR, but not RXR, although their binding affinity was three orders of magnitude lower than that of atRA. Additionally, when applied to macrophages, β-cryptoxanthin indeed was found to induce the ATP-binding cassette transporter A1 (ABCA1) and ABCG1 mRNAs, which exert anti-atherosclerotic effects by preventing cholesteryl ester accumulation in macrophages. The induction of ABCA1 proteins by β-cryptoxanthin as well as atRA was abrogated by LE540. In summary, β-cryptoxanthin appears to be more an efficient provitamin A source than other carotenoids and xanthophylls including β-carotene, since β-cryptoxanthin can act not only as a RAR agonist but also a source of vitamin A. Taking into account that the pharmacodynamics difference between β-cryptoxanthin and atRA, β-cryptoxanthin appears to exert beneficial effects on atherogenesis through RAR activation in the manner different from atRA.

Hypoadiponectinemia: A common basis for diseases associated with overnutrition

Adiponectin is a plasma protein derived from adipose tissue, which we discovered from a human adipose cDNA project. Adiponectin exists in circulating plasma at concentrations ranging from 4 to 30 microg/mL, which is much higher than the concentrations of various other hormones and cytokines. Adiponectin has a sticky nature, binding to collagen I, III, and V, which are present in vascular intima. Adiponectin exhibits various antiatherogenic effects on vascular cells, suppressing the expression of adhesion molecules in vascular endothelial cells, proliferation of smooth muscle cells, and cholesteryl-ester accumulation in macrophages. However, its plasma levels are low in subjects with excess intra-abdominal fat. Adiponectin also has antidiabetic properties, and plasma adiponectin levels correlate positively with insulin sensitivity. Several clinical studies have demonstrated that hypoadiponectinemia is a risk factor for new-onset diabetes. Recent studies suggest that hypoadiponectinemia may partly contribute to the development of salt-sensitive hypertension and hypertensive heart failure, and can be also a risk factor for overnutrition-related cancers such as breast, colon, uterine, and prostate cancers. Hypoadiponectinemia might be at least in part the molecular basis of various diseases associated with overnutrition.

Lipoprotein(a) immune complexes induce cholesteryl ester accumulation in human macrophages

目的 探讨氧化、天然脂蛋白(a)[Lp(a)]循环免疫复合物(IC)致动脉粥样硬化作用.方法采用人源的氧化、天然Lp(a)与异源的抗载脂蛋白B(apoB)结合制备IC,观察不同浓度的Lp(a)-IC对小鼠腹腔巨噬细胞胆固醇酯的蓄积和泡沫细胞形成作用.结果氧化、天然Lp(a)-IC均引起巨噬细胞内胆固醇酯大量堆积,其效果强于Ox-Lp(a) (P<0.001),且随着剂量的增加而升高,并转化为泡沫细胞;而天然Lp(a)、抗apoB处理的巨噬细胞内未见胆固醇酯的堆积.结论 Lp(a)-IC导致巨噬细胞转化为泡沫细胞,参与动脉粥样硬化形成。

HDLs in apoA-I transgenic Abca1 knockout mice are remodeled normally in plasma but are hypercatabolized by the kidney

Patients homozygous for Tangier disease have a near absence of plasma HDL as a result of mutations in ABCA1 and hypercatabolize normal HDL particles. To determine the relationship between ABCA1 expression and HDL catabolism, we investigated intravascular remodeling, plasma clearance, and organ-specific uptake of HDL in mice expressing the human apolipoprotein A-I (apoA-I) transgene in the Abca1 knockout background. Small HDL particles (7.5 nm), radiolabeled with (125)I-tyramine cellobiose, were injected into recipient mice to quantify plasma turnover and the organ uptake of tracer. Small HDL tracer was remodeled to 8.2 nm diameter particles within 5 min in human apolipoprotein A-I transgenic (hA-I(Tg)) mice (control) and knockout mice. Decay of tracer from plasma was 1.6-fold more rapid in knockout mice (P < 0.05) and kidney uptake was twice that of controls, with no difference in liver uptake. We also observed 2-fold greater hepatic expression of ABCA1 protein in hA-I(Tg) mice compared with nontransgenic mice, suggesting that overexpression of human apoA-I stabilized hepatic ABCA1 protein in vivo. We conclude that ABCA1 is not required for in vivo remodeling of small HDLs to larger HDL subfractions and that the hypercatabolism of normal HDL particles in knockout mice is attributable to a selective catabolism of HDL apoA-I by the kidney.

Regulation of Macrophage Cholesterol Efflux through Hydroxymethylglutaryl-CoA Reductase Inhibition: A ROLE FOR RhoA IN ABCA1-MEDIATED CHOLESTEROL EFFLUX

The cholesterol biosynthetic pathway produces numerous signaling molecules. Oxysterols through liver X receptor (LXR) activation regulate cholesterol efflux, whereas the non-sterol mevalonate metabolite, geranylgeranyl pyrophosphate (GGPP), was recently demonstrated to inhibit ABCA1 expression directly, through antagonism of LXR and indirectly through enhanced RhoA geranylgeranylation. We used HMG-CoA reductase inhibitors (statins) to test the hypothesis that reduced synthesis of mevalonate metabolites would enhance cholesterol efflux and attenuate foam cell formation. Preincubation of THP-1 macrophages with atorvastatin, dose dependently (1-10 microm) stimulated cholesterol efflux to apolipoprotein AI (apoAI, 10-60%, p < 0.05) and high density lipoprotein (HDL(3)) (2-50%, p < 0.05), despite a significant decrease in cholesterol synthesis (2-90%). Atorvastatin also increased ABCA1 and ABCG1 mRNA abundance (30 and 35%, p < 0.05). Addition of mevalonate, GGPP or farnesyl pyrophosphate completely blocked the statin-induced increase in ABCA1 expression and apoAI-mediated cholesterol efflux. A role for RhoA was established, because two inhibitors of Rho protein activity, a geranylgeranyl transferase inhibitor and C3 exoenzyme, increased cholesterol efflux to apoAI (20-35%, p < 0.05), and macrophage expression of dominant-negative RhoA enhanced cholesterol efflux to apoAI (20%, p < 0.05). In addition, atorvastatin increased the RhoA levels in the cytosol fraction and decreased the membrane localization of RhoA. Atorvastatin treatment activated peroxisome proliferator activated receptor gamma and increased LXR-mediated gene expression suggesting that atorvastatin induces cholesterol efflux through a molecular cascade involving inhibition of RhoA signaling, leading to increased peroxisome proliferator activated receptor gamma activity, enhanced LXR activation, increased ABCA1 expression, and cholesterol efflux. Finally, statin treatment inhibited cholesteryl ester accumulation in macrophages challenged with atherogenic hypertriglyceridemic very low density lipoproteins indicating that statins can regulate foam cell formation.

Comparison of the effects of pioglitazone and rosiglitazone on macrophage foam cell formation

In order to elucidate the antiatherogenic effects of pioglitazone (a peroxisome proliferator-activated receptor [PPAR]γ agonist with PPARα agonistic activity) and rosiglitazone (a more selective PPARγ agonist), we examined gene expression and cholesteryl ester accumulation in THP-1-derived macrophages. Pioglitazone enhanced the mRNA expression of the proatherogenic factors CD36 and adipophilin, but was approximately 10 times less potent than rosiglitazone. The potencies of the two agents appeared to correspond to their PPARγ agonistic activities in this respect. However, both agents were similarly potent in enhancing the mRNA expression of the antiatherogenic factors liver X receptor α and ATP-binding cassette-transporter A1. Furthermore, both agents enhanced cholesteryl ester hydrolase mRNA expression and inhibited acyl-CoA cholesterol acyltransferase-1 mRNA expression and cholesteryl ester accumulation in macrophages. In this respect, their potencies appeared to correspond to their PPARα agonistic activities. These results suggest that pioglitazone has an equally beneficial effect on antiatherogenic events to rosiglitazone, despite being almost 10 times less potent than a PPARγ agonist.

Reduced atherosclerosis in hormone-sensitive lipase transgenic mice overexpressing cholesterol acceptors

Macrophage-specific overexpression of cholesteryl ester hydrolysis in hormone-sensitive lipase transgenic (HSL Tg) female mice paradoxically increases cholesterol esterification and cholesteryl ester accumulation in macrophages, and thus susceptibility to diet-induced atherosclerosis compared to nontransgenic C57BL/6 mice. The current studies suggest that whereas increased cholesterol uptake could contribute to transgenic foam cell formation, there are no differences in cholesterol synthesis and the expression of cholesterol efflux mediators (ABCA1, ABCG1, apoE, PPARγ, and LXRα) compared to wild-type macrophages. HSL Tg macrophages exhibit twofold greater efflux of cholesterol to apoA–I in vitro, suggesting the potential rate-limiting role of cholesteryl ester hydrolysis in efflux. However, macrophage cholesteryl ester levels appear to depend on the relative efficacy of alternate pathways for free cholesterol in either efflux or re-esterification. Thus, increased atherosclerosis in HSL Tg mice appears to be due to the coupling of the efficient re-esterification of excess free cholesterol to its limited removal mediated by the cholesterol acceptors in these mice. The overexpression of cholesterol acceptors in HSL-apoA-IV double-transgenic mice increases plasma HDL levels and decreases diet-induced atherosclerosis compared to HSL Tg mice, with aortic lesions reduced to sizes in nontransgenic littermates. The results in vivo are consistent with the effective efflux from HSL Tg macrophages supplemented with HDL and apoA-I in vitro, and highlight the importance of cholesterol acceptors in inhibiting atherosclerosis caused by imbalances in the cholesteryl ester cycle.

HIV protease inhibitors promote atherosclerotic lesion formation independent of dyslipidemia by increasing CD36-dependent cholesteryl ester accumulation in macrophages

Protease inhibitors decrease the viral load in HIV patients, however the patients develop hypertriglyceridemia, hypercholesterolemia, and atherosclerosis. It has been assumed that protease inhibitor-dependent increases in atherosclerosis are secondary to the dyslipidemia. Incubation of THP-1 cells or human PBMCs with protease inhibitors caused upregulation of CD36 and the accumulation of cholesteryl esters. The use of CD36-blocking antibodies, a CD36 morpholino, and monocytes isolated from CD36 null mice demonstrated that protease inhibitor-induced increases in cholesteryl esters were dependent on CD36 upregulation. These data led to the hypothesis that protease inhibitors induce foam cell formation and consequently atherosclerosis by upregulating CD36 and cholesteryl ester accumulation independent of dyslipidemia. Studies with LDL receptor null mice demonstrated that low doses of protease inhibitors induce an increase in the level of CD36 and cholesteryl ester in peritoneal macrophages and the development of atherosclerosis without altering plasma lipids. Furthermore, the lack of CD36 protected the animals from protease inhibitor-induced atherosclerosis. Finally, ritonavir increased PPAR-gamma and CD36 mRNA levels in a PKC- and PPAR-gamma-dependent manner. We conclude that protease inhibitors contribute to the formation of atherosclerosis by promoting the upregulation of CD36 and the subsequent accumulation of sterol in macrophages.

HIV protease inhibitors promote atherosclerotic lesion formation independent of dyslipidemia by increasing CD36-dependent cholesteryl ester accumulation in macrophages

Protease inhibitors decrease the viral load in HIV patients, however the patients develop hypertriglyceridemia, hypercholesterolemia, and atherosclerosis. It has been assumed that protease inhibitor–dependent increases in atherosclerosis are secondary to the dyslipidemia. Incubation of THP-1 cells or human PBMCs with protease inhibitors caused upregulation of CD36 and the accumulation of cholesteryl esters. The use of CD36-blocking antibodies, a CD36 morpholino, and monocytes isolated from CD36 null mice demonstrated that protease inhibitor–induced increases in cholesteryl esters were dependent on CD36 upregulation. These data led to the hypothesis that protease inhibitors induce foam cell formation and consequently atherosclerosis by upregulating CD36 and cholesteryl ester accumulation independent of dyslipidemia. Studies with LDL receptor null mice demonstrated that low doses of protease inhibitors induce an increase in the level of CD36 and cholesteryl ester in peritoneal macrophages and the development of atherosclerosis without altering plasma lipids. Furthermore, the lack of CD36 protected the animals from protease inhibitor–induced atherosclerosis. Finally, ritonavir increased PPAR-γ and CD36 mRNA levels in a PKC- and PPAR-γ–dependent manner. We conclude that protease inhibitors contribute to the formation of atherosclerosis by promoting the upregulation of CD36 and the subsequent accumulation of sterol in macrophages.

Transforming growth factor-beta1 inhibits macrophage cholesteryl ester accumulation induced by native and oxidized VLDL remnants.

Transforming growth factor beta1 (TGF-beta1) is secreted by various cells, including macrophages, smooth muscle cells, and endothelial cells. TGF-beta1 is present in atherosclerotic lesions, but its role in regulating macrophage foam cell formation is not understood. Hypertriglyceridemic very low density lipoprotein (VLDL) remnants (VLDL-REMs) in their native or oxidized form will induce cholesteryl ester (CE) and triglyceride (TG) accumulation in macrophages. Therefore, we examined whether TGF-beta1 can modulate the macrophage uptake of native or oxidized VLDL-REMs (oxVLDL-REMs). Incubation of J774A.1 macrophages with VLDL-REMs and oxVLDL-REMs compared with control cells increased cellular CE (13- and 21-fold, respectively) and TG mass (21-and 18-fold, respectively). Preincubation with TGF-beta1 before incubation with VLDL-REMs or oxVLDL-REMs significantly decreased CE (73% and 54%, respectively) and TG mass (42% and 41%, respectively). TGF-beta1 inhibited the activity and expression of 2 key components needed for VLDL-REM uptake: lipoprotein lipase and low density lipoprotein receptor. TGF-beta1 inhibited CE mass induced by oxVLDL-REMs in part by decreasing the expression of scavenger receptor type AI/II and CD36. Furthermore, TGF-beta1 enhanced cholesterol efflux through upregulation of the ATP-binding cassette (ABC) transporters ABCA1 and ABCG1. Thus, TGF-beta1 inhibits macrophage foam cell formation induced by VLDL-REMs or oxVLDL-REMs, which suggests an antiatherogenic role for this cytokine.

Effects of Fluvastatin and Its Major Metabolites on Low-Density Lipoprotein Oxidation and Cholesterol Esterification in Macrophages

We investigated effects of fluvastatin, a 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor, and its major metabolites, M2 and M4, on CuSO4-induced low-density lipoprotein (LDL) oxidation and cholesteryl ester accumulation in mouse peritoneal macrophages. All the test compounds inhibited LDL oxidation, and M2 had the most potent effect comparable to vitamin E. When LDL was previously incubated with the test compounds in the presence of CuSO4, the pre-treatment resulted in a marked reduction of facilitated cholesteryl ester accumulation in macrophages. Supplementation of mevalonate did not overcome the inhibitory effects of fluvastatin and its metabolites on both LDL oxidation and facilitated cholesterol esterification. Pravastatin, another HMG-CoA reductase inhibitor, did not show any inhibitory effect. Consequently, these effects of fluvastatin and its metabolites are considered to be derived from their own unique chemical structures. Moreover, fluvastatin and M2 directly inhibited cholesterol esterification induced by oxidized LDL in macrophages, but pravastatin was also found to have a weak effect. As their inhibitory effects were overcome by addition of mevalonate, the direct inhibitory effect on cholesterol esterification would be a common property of HMG-CoA reductase inhibitors. The inhibitory effects of fluvastatin and its metabolites on both LDL oxidation and cholesterol esterification in macrophages may contribute to the antiatherogenic action in vivo.

Effects of hypoxia on cholesterol metabolism in human monocyte-derived macrophages

We assessed the metabolism of low density lipoprotein (LDL) of human monocyte-derived macrophages under hypoxia. The specific binding and association of 125I-labeled LDL ( 125I-LDL) were not changed under hypoxia compared to normoxia. However, the degradation of 125I-LDL under hypoxia decreased to 60%. The rate of cholesterol esterification under hypoxia was 2-fold greater on incubation with LDL or 25-hydroxycholesterol. The cellular cholesteryl ester content was also greater under hypoxia on incubation with LDL. Secretion of apolipoprotein E into the medium was not altered under hypoxia, suggesting that apolipoprotein E independent cholesterol efflux may be reduced under hypoxia. Thus, hypoxia affects the intracellular metabolism of LDL, stimulates cholesterol esterification, and enhances cholesteryl ester accumulation in macrophages. Hypoxia is one of the important factors modifying the cellular lipid metabolism in arterial wall.

Functional evidence for anti-oxidant action of fluvastatin on low-density lipoprotein using isolated macrophages and aorta.

1. Fluvastatin has been reported to have not only a hypocholesterolaemic effect, but also a protective effect on low-density lipoprotein (LDL) from oxidation. We functionally evaluated the anti-oxidant effect of fluvastatin on oxidation of LDL by copper ions in vitro using mouse macrophages and rabbit aorta preparations. 2. After native LDL (N-LDL) from rabbit plasma had been pre-incubated in the presence or absence of fluvastatin (10 micromol/L) for 4 h, the N-LDL was mildly oxidized by incubation with 5 micromol/L CuCl2 for 5 h and two oxidized LDL, fluvastatin-pretreated (Flu-OxLDL) and -non-treated (OxLDL), were prepared. The level of thiobarbituric acid-reactive substances (TBARS) in Flu-OxLDL and OxLDL markedly increased compared with N-LDL. The degree of increment was significantly less in Flu-OxLDL than OxLDL. 3. When macrophages were incubated with Flu-OxLDL or OxLDL, the amount of cholesteryl ester that accumulated in the macrophages markedly increased compared with N-LDL. The degree of increment was significantly less in Flu-OxLDL than OxLDL. 4. Acetylcholine-induced endothelium-dependent relaxations in rabbit aortic rings were impaired in the presence of either Flu-OxLDL or OxLDL. The degree of impairment was significantly less in Flu-OxLDL. 5. The increased TBARS level, facilitated cholesteryl ester accumulation in macrophages and impaired endothelium-dependent relaxation elicited by OxLDL were not affected by simultaneous treatment with fluvastatin (10 micromol/L). 6. These findings indicate that fluvastatin can protect plasma LDL from oxidative modification and, thereby, prevent cholesterol accumulation in macrophages and endothelial dysfunction in blood vessels. This additional anti-oxidative effect of fluvastatin may be beneficial for preventing the progression of atherosclerosis.

Postprandial triglyceride metabolism in diabetes mellitus.

Individuals with diabetes have a two to four times higher risk of cardiovascular morbidity and mortality than nondiabetics. Patients with both type 1 and type 2 diabetes share a similar risk. Studies in individuals with type 1 diabetes have shown a decreased clearance of postprandial triglyceride-rich lipoprotein particles of abnormal composition. Particles isolated from diabetic individuals show abnormal composition and an increased tendency to cause cholesteryl ester accumulation in macrophages and are therefore potentially atherogenic. Various interventions may alter these abnormalities and improve the atherosclerotic risk. These include adopting a high-carbohydrate diet over a high monounsaturated diet, improving glycemic control, infusing insulin intraperitoneally, and using pharmacologic therapies such as the statins.