THP-1 Macrophage-derived Foam Cells Research Articles

Polyunsaturated Fatty Acids and Modulation of Cholesterol Homeostasis in THP-1 Macrophage-Derived Foam Cells

Transformation of macrophages to foam cells is determined by the rates of cholesterol uptake and efflux. This study uses a real time RT-PCR technique to investigate the role of conjugated linoleic acid (CLA), α-linolenic acid (ALA) and eicosapentaenoic acid (EPA) in the regulation of the ATP-binding cassette A1 (ABCA1) and liver X receptor α (LXR) genes, which are involved in cholesterol homeostasis. Accordingly, these fatty acids significantly reduced the total, free and esterified cholesterols within the foam cells. While the expression of the ABCA1 and LXRα genes was increased in the presence of the pharmacological LXRα ligand, T0901317, their mRNA expression was not significantly affected by CLA, ALA and EPA. These results suggest that although polyunsaturated fatty acids have an effect on cholesterol homeostasis, they cannot change the expression of the ABCA1 and LXRα genes. Alternatively, several other genes and proteins may be involved.

Ibrolipim increases ABCA1/G1 expression by the LXRα signaling pathway in THP-1 macrophage-derived foam cells

To determine the effects and potential mechanisms of ibrolipim on ATP-binding membrane cassette transporter A-1 (ABCA1) and ATP-binding membrane cassette transporter G-1 (ABCG1) expression from human macrophage foam cells, which may play a critical role in atherogenesis. Human THP-1 cells pre-incubated with ox-LDL served as foam cell models. Specific mRNA was quantified using real-time RT-PCR and protein expression using Western blotting. Cellular cholesterol handling was studied using cholesterol efflux experiments and high performance liquid chromatography assays. Ibrolipim 5 and 50 μmol/L significantly increased cholesterol efflux from THP-1 macrophage-derived foam cells to apoA-I or HDL. Moreover, it upregulated the expression of ABCA1 and ABCG1. In addition, LXRα was also upregulated by the ibrolipim treatment. In addition, LXRα small interfering RNA completely abolished the promotion effect that was induced by ibrolipim. Ibrolipim increased ABCA1 and ABCG1 expression and promoted cholesterol efflux, which was mediated by the LXRα signaling pathway.

Contribution of D4-F to ABCA1 Expression and Cholesterol Efflux in THP-1 Macrophage-derived Foam Cells

Adenosine triphosphate-binding cassette transporter A1 (ABCA1) plays a crucial role in apolipoprotein A-I (apoA-I) binding activity and promotes cellular cholesterol efflux. ApoA-I mimetic peptide D4-F has reported to have the similar ability as apoA-I. However, the detailed mechanisms of ABCA1 regulation by D4-F are not understood. In the present study, we investigated the effects of D4-F on ABCA1 expression and ABCA1-dependent cholesterol efflux and examined the role of Cdc42/cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA) pathway on the regulation of ABCA1 by D4-F in THP-1 macrophage-derived foam cells. Results showed that D4-F stabilized ABCA1 protein and enhanced ABCA1-dependent cholesterol efflux but had no effect on ABCA1 messenger RNA expression. We also revealed that D4-F enhanced cAMP level and PKA activity and ABCA1 serine phosphorylation. Short interfering RNA of PKA led to reduction of ABCA1 serine phosphorylation and ABCA1-mediated cholesterol efflux compensated by D4-F. PKA-specific activation by PKA agonist enhanced the upregulation of ABCA1 serine phosphorylation and ABCA1-mediated cholesterol efflux by D4-F. However, ABCA1 expression did not change by treatment with PKA agonist or PKA-short interfering RNA. We found that secramine B of Cdc42 inhibitor reduced the cAMP level compensated by D4-F. These results provide evidence that D4-F enhances ABCA1 serine phosphorylation and ABCA1-dependent cholesterol efflux through Cdc42/cAMP/PKA pathway in THP-1 macrophage-derived foam cells.

Lipopolysaccharide Down-regulates ABCA1 Expression in Foam Cells in a Nucleus Factor-κB Pathway-dependent Manner*

The immnue-inflammatory response mediated by LPS is firmly related to the development of atherosclerosis. ABCA1 has been identified to play a key role in the cellular cholesterol efflux which is regarded to anti-atherosclerosis. To investigate the changes of cholesterol efflux, ATP-binding cassette transporter A1 (ABCA1) mRNA and protein expression in THP-1 macrophage derived foam cells treated with LPS, and to discover the role of TLR4/NF-κB pathway and LXRs in this process. The foam cells were exposed to different concentration of LPS for 24 h or exposed to LPS for different times, and more, the cells were treated with LPS along or together with N-p-Tosyl-L-phenylalanine chloromethyl ketone (TPCK) for 24 h. The mRNA levels of ABCA1, TLR4 and LXRα were measured by reverse transcriptase-polymerase chain reaction,and the protein levels of ABCA1, LXRα and intranuclear NF-κB p65 were measured by Western blotting. Cellular lipid accumulation was determined by high performance liquid chromatography analysis. Cholesterol efflux was determined by FJ-2107P type liquid scintillator. The results show that the expression of ABCA1 were decreased in a dose- and time-dependent manner after treated with LPS, while TLR4 expression and the intranuclear NF-κB p65 protein level were increased by LPS, and these changes can be reversed partly by pretreatment with TPCK. LPS and TPCK can not effect LXRα expression. The results also show that cellular lipid accumulation was increased, while the cellular cholesterol efflux was decreased in THP-1 macrophage derived foam cells after exposured to LPS for 24 h. It was concluded that LPS can down-regulate the expression of ABCA1, promote the accumulation of lipid and decrease cellular cholesterol efflux in THP-1 macrophage derived foam cells, which may be related to the TLR4/NF-κB dependent and LXRα-independent pathway.

Lipopolysaccharide Down-regulates ABCA1 Expression in Foam Cells in a Nucleus Factor-κB Pathway-dependent Manner*

The immnue-inflammatory response mediated by LPS is firmly related to the development of atherosclerosis. ABCA1 has been identified to play a key role in the cellular cholesterol efflux which is regarded to anti-atherosclerosis. To investigate the changes of cholesterol efflux, ATP-binding cassette transporter A1 (ABCA1) mRNA and protein expression in THP-1 macrophage derived foam cells treated with LPS, and to discover the role of TLR4/NF-κB pathway and LXRs in this process. The foam cells were exposed to different concentration of LPS for 24 h or exposed to LPS for different times, and more, the cells were treated with LPS along or together with N-p-Tosyl-L-phenylalanine chloromethyl ketone (TPCK) for 24 h. The mRNA levels of ABCA1, TLR4 and LXRα were measured by reverse transcriptase-polymerase chain reaction,and the protein levels of ABCA1, LXRα and intranuclear NF-κB p65 were measured by Western blotting. Cellular lipid accumulation was determined by high performance liquid chromatography analysis. Cholesterol efflux was determined by FJ-2107P type liquid scintillator. The results show that the expression of ABCA1 were decreased in a dose- and time-dependent manner after treated with LPS, while TLR4 expression and the intranuclear NF-κB p65 protein level were increased by LPS, and these changes can be reversed partly by pretreatment with TPCK. LPS and TPCK can not effect LXRα expression. The results also show that cellular lipid accumulation was increased, while the cellular cholesterol efflux was decreased in THP-1 macrophage derived foam cells after exposured to LPS for 24 h. It was concluded that LPS can down-regulate the expression of ABCA1, promote the accumulation of lipid and decrease cellular cholesterol efflux in THP-1 macrophage derived foam cells, which may be related to the TLR4/NF-κB dependent and LXRα-independent pathway.

TGF-β1 Up-Regulates Expression of ABCA1, ABCG1 and SR-BI through Liver X Receptor α Signaling Pathway in THP-1 Macrophage-Derived Foam Cells

High density lipoprotein (HDL) and its apolipoproteins can promote cholesterol efflux from macrophage foam cells via the ATP-binding cassette transporter A1 (ABCA1), ABCG1, and scavenger receptor class B type I (SR-BI). Liver X receptors (LXRs) operate as cholesterol sensors which may protect from cholesterol overload by stimulating cholesterol efflux from cells to HDL through ABCA1, ABCG1 and SR-BI. The regulation of ABCA1, ABCG1 and SR-BI expression by cytokines present within the microenvironment of the atheroma may play an important role in determining the impact of reverse cholesterol transport on the atherosclerotic lesion. In the current study, we examined the effect of transforming growth factor-beta1 (TGF-beta1) on expressions of ABCA1, ABCG1 and SR-BI and explored the role of LXR alpha in the regulation of ABCA1, ABCG1 and SR-BI in THP-1 macrophage-derived foam cells. TGF-beta1 significantly increased expressions of ABCA1, ABCG1 and SR-BI at both transcriptional and translational levels in a dose-dependent and time-dependent manner. Cellular cholesterol content was decreased while cholesterol efflux was increased by TGF-beta1 treatment. Moreover, LXR alpha was up-regulated by TGF-beta1 treatment. In addition, LXR alpha small interfering RNA completely abolished the promotion effect induced by TGF-beta1. These results provide evidence that TGF-beta1 up-regulates expressions of ABCA1, ABCG1 and SR-BI through the LXR alpha pathway in THP-1 macrophage-derived foam cells.

Modulation Peroxisome Proliferators Activated Receptor alpha (PPAR α) and Acyl Coenzyme A: Cholesterol Acyltransferase1 (ACAT1) Gene expression by Fatty Acids in Foam cell

BackgroundOne of the most important factors in the initiation and progression of atherosclerosis is the default in macrophage cholesterol homeostasis. Many genes and transcription factors such as Peroxisome Proliferators Activated Receptors (PPARs) and Acyl Coenzyme A: Cholesterol Acyltransferase1 (ACAT1) are involved in cholesterol homeostasis. Fatty Acids are important ligands of PPARα and the concentration of them can effect expression of ACAT1. So this study designed to clarified on the role of these genes and fatty acids on the lipid metabolism in foam cells.MethodsThis study examined effects of c9, t11-Conjugated Linoleic Acid(c9, t11-CLA), Alpha Linolenic Acid (LA), Eicosapentaenoic Acid (EPA) on the PPARα and ACAT1 genes expression by using Real time PCR and cholesterol homeostasis in THP-1 macrophages derived foam cells.ResultsIncubation of c9, t11-CLA, LA cause a significant reduction in intracellular Total Cholesterol, Free Cholesterol, cellular and Estrified Cholesterol concentrations (P ≤ 0.05). CLA and LA had no significant effect on the mRNA levels of ACAT1, but EPA increased ACAT1 mRNA expression (P = 0.003). Treatment with EPA increased PPARα mRNA levels (P ≤ 0.001), although CLA, LA had no significant effect on PPARα mRNA expression.ConclusionIn conclusion, it seems that different fatty acids have different effects on gene expression and lipid metabolism and for complete conception study of the genes involved in lipid metabolism in foam cell all at once maybe is benefit.

NO-1886 Up-regulates Niemann–Pick C1 Protein (NPC1) Expression Through Liver X Receptor α Signaling Pathway in THP-1 Macrophage-Derived Foam Cells

The Niemann-Pick C1 (NPC1) protein regulates the transport of cholesterol from late endosomes/lysosomes to other compartments responsible for maintaining intracellular cholesterol homeostasis. Liver X receptors (LXRs) operate as cholesterol sensors which may protect from cholesterol overload by increasing the amount of free cholesterol in the plasma membrane through inducing NPC1 expression. NO-1886 has been proven to be highly effective at increasing liver X receptor alpha expression and promoting cellular cholesterol efflux. In this study, the effects of NO-1886 on NPC1 expression were investigated in THP-1 macrophage-derived foam cells. Results showed that NO-1886 markedly increased expression of NPC1 at both mRNA level and protein level in a dose-dependent and time-dependent manner. Cellular cholesterol content was decreased while cholesterol efflux was increased by NO-1886 treatment. In addition, LXR alpha was also up-regulated by NO-1886 treatment. And LXR alpha small interfering RNA completely abolished the promotion effect which was induced by NO-1886. These results provide evidence that NO-1886 up-regulates expression of NPC1 through LXR alpha pathway in THP-1 macrophage- derived foam cells.

Eicosapentaenoic acid reduces ABCA1 serine phosphorylation and impairs ABCA1-dependent cholesterol efflux through cyclic AMP/protein kinase A signaling pathway in THP-1 macrophage-derived foam cells

ABCA1 is a key mediator of cholesterol efflux to apoA-I in cholesterol loaded macrophages, a first step of RCT in vivo. Unsaturated fatty acids can inhibit cholesterol efflux from macrophages by increasing degradation of ABCA1. However, the detailed mechanisms of ABCA1 regulation by unsaturated fatty acids are not fully understood. In the present study, we investigated the effects of EPA on ABCA1 expression and ABCA1-dependent cholesterol efflux and examined the role of cAMP/PKA pathway on the regulation of ABCA1 by EPA in THP-1 macrophage-derived foam cells. Results showed that EPA significantly destabilized ABCA1 protein and reduced ABCA1-dependent cholesterol efflux but had no effect on ABCA1 mRNA expression. We also revealed that EPA markedly reduced cAMP level and PKA activity and ABCA1 serine phosphorylation. PKA-specific activation by PKA agonist markedly compensated the down-regulation of ABCA1 serine phosphorylation and ABCA1-mediated cholesterol efflux by EPA, while, siRNA of PKA leaded to reduce of ABCA1 serine phosphorylation and ABCA1-mediated cholesterol efflux more significantly than EPA. However, EPA-Induced enhancement of degradation rate of ABCA1 protein did not change by treatment with PKA agonist or PKA-siRNA. These results provide evidence that EPA may have dual negative effects on ABCA1 activity by decreasing ABCA1 protein level and by reducing PKA-mediated ABCA1 serine phosphorylation in THP-1 macrophage-derived foam cells.

IFN-γ down-regulates ABCA1 expression by inhibiting LXRα in a JAK/STAT signaling pathway-dependent manner

Interferon gamma (IFN-γ) is an immunomodulatory and anti-microbial cytokine, which has a variety of proatherogenic effects. It has been reported that IFN-γ can down-regulate ABCA1 expression. However, its mechanism is elusive. In the present study, we have investigated the effect of IFN-γ on ABCA1 expression and cholesterol efflux in THP-1 macrophage-derived foam cells. IFN-γ decreased ABCA1 expression at both transcriptional and translational levels in a dose-dependent manner. Cellular cholesterol content was increased while cholesterol efflux was decreased by IFN-γ treatment. Liver X receptor α (LXRα), which can regulate the expression of ABCA1, was also down-regulated by IFN-γ treatment. LXRα-specific activation by LXRα agonist almost compensated the down-regulation of ABCA1 expression by IFN-γ, while siRNA of LXRα led to down-regulation of ABCA1 expression more significantly than IFN-γ. IFN-γ induced phosphorylation of STAT1 and expression of STAT1α in the nucleus, which was inhibited by a JAK inhibitor AG-490. Treatment with STAT1 siRNA further enhanced down-regulation of LXRα mRNA by IFN-γ. Furthermore, AG-490 and STAT1 siRNA almost compensated the effect of IFN-γ on ABCA1 expression and cholesterol efflux. In conclusion, IFN-γ may first down-regulate expression of LXRα through the JAK/STAT1 signaling pathway and then decrease expression of ABCA1 and cholesterol efflux in THP-1 macrophage-derived foam cells. Therefore, our study may be useful in understanding the critical effect of IFN-γ in pathogenesis of atherosclerosis.

Liver X receptor agonist T0901317 reduces atherosclerotic lesions in apoE-/- mice by up-regulating NPC1 expression

In this study, we studied the effect of liver X receptor (LXR) agonist T0901317 on Niemann-Pick C1 protein (NPC1) expression in apoE-/- mice. Male apoE-/- mice were randomized into 4 groups, baseline group (n=10), control group (n = 14), treatment group (n = 14) and prevention group (n = 14). All of the mice were fed with a high-fat/high-cholesterol (HFHC) diet containing 15% fat and 0.25% cholesterol. The baseline group treated with vehicle was sacrificed after 8 weeks of the diet. The control group and the prevention group were treated with either vehicle or T0901317 daily by oral gavage for 14 weeks. The treatment group was treated with vehicle for 8 weeks, and then was treated with the agonist T0901317 for additional 6 weeks. Gene and protein expression was analyzed by real-time quantitative PCR, immunohistochemistry and Western blotting, respectively. Plasma lipid concentrations were measured by commercially enzymatic methods. We used RNA interference technology to silence NPC1 gene expression in THP-1 macrophage-derived foam cells and then detected the effect of LXR agonist T0901317 on cholesterol efflux. Plasma triglyceride (TG), total cholesterol (TC), high density lipoprotein cholesterol (HDL-C) and apoA-I concentrations were markedly increased in T0901317-treated groups. T0901317 treatment reduced the aortic atherosclerotic lesion area by 64.2% in the prevention group and 58.3% in the treatment group. LXR agonist treatment increased NPC1 mRNA expression and protein levels in the small intestine, liver and aorta of apoE-/- mice. Compared with the normal cells, cholesterol efflux of siRNA THP-1 macrophage-derived foam cells was significantly decreased, whereas cholesterol efflux of LXR agonist T0901317-treated THP-1 macrophage-derived foam cells was significantly increased. Our results suggest that LXR agonist T0901317 inhibits atherosclerosis development in apoE-/- mice, which is related to up-regulating NPC1 expression.

Interleukin‐10 inhibits the down‐regulation of ATP binding cassette transporter A1 by tumour necrosis factor‐alpha in THP‐1 macrophage‐derived foam cells

It is suggested that cholesterol efflux mediated by ATP binding cassette transporter A1 (ABCA1) plays an important role in anti-atherogenesis. However, the effects of inflammatory cytokines on ABCA1 expression and cholesterol accumulation in foam cells are little known. This study investigates the effects of tumour necrosis factor-alpha (TNF-alpha) and interleukin-10 (IL-10) on ABCA1 expression and cholesterol content in THP-1 macrophage-derived foam cells. ABCA1mRNA and protein levels were determined by RT-PCR and Western blot, respectively. The total cholesterol content in THP-1 macrophage-derived foam cells was detected by the zymochemistry method. Results revealed that TNF-alpha could increase cholesterol content by down-regulating ABCA1 expression in a time-dependent manner in THP-1 macrophage-derived foam cells, which may contribute to its pro-atherosclerotic effect. In addition IL-10 time-dependently decreased cholesterol accumulation by up-regulating ABCA1 expression and inhibited the down-regulation of ABCA1 by TNF-alpha in THP-1 macrophage-derived foam cells, which may be one of the mechanisms of IL-10 contributing to its anti-atherosclerotic action.

Th-P15:209 Mast cells change cellular cholesterol contents and inhibit cholesterol efflux from THP-1 macrophage-derived foam cells

Th-P15:209 Mast cells change cellular cholesterol contents and inhibit cholesterol efflux from THP-1 macrophage-derived foam cells

W08.193 Effect of rolipram on ATP binding cassette transporter 1 and cholesterol efflux in THP-1 macrophage-derived foam cells

W08.193 Effect of rolipram on ATP binding cassette transporter 1 and cholesterol efflux in THP-1 macrophage-derived foam cells

W08.193 Effect of rolipram on ATP binding cassette transporter 1 and cholesterol efflux in THP-1 macrophage-derived foam cells

W08.193 Effect of rolipram on ATP binding cassette transporter 1 and cholesterol efflux in THP-1 macrophage-derived foam cells

Effect of apolipoprotein A-I on ATP binding cassette transporter A1 degradation and cholesterol efflux in THP-1 macrophage-derived foam cells.

Cholesterol-loaded macrophage foam cells are a central component of atherosclerotic lesions. ATP binding cassette transporter A1 (ABCA1), the defective molecule in Tangier disease, mediates the efflux of phospholipid and cholesterol from cells to apolipoprotein A-I (apoA-I), reversing foam cell formation. This study investigated the effect of apoA-I on ABCA1 degradation and cholesterol efflux in THP-1 macrophage-derived foam cells. After exposure of the cultured THP-1 macrophage-derived foam cells to apoA-I for different time, cholesterol efflux, ABCA1 mRNA and protein levels were determined by FJ-2107P type liquid scintillator, RT-PCR and Western blot, respectively. The mean ABCA1 fluorescence intensity on THP-1 macrophage-derived foam cells was detected by flow cytometry. Results showed that apoA-I markedly increased ABCA1-mediated cholesterol efflux from THP-1 macrophage-derived foam cells. This was accompanied by an increase in the content of ABCA1. ApoA-I did not alter ABCA1 mRNA abundance. Significantly, thiol protease inhibitors increased the level of ABCA1 protein and slowed its decay in THP-1 macrophage-derived foam cells, whereas none of the proteosome-specific inhibitor lactacystin, other protease inhibitors, or the lysosomal inhibitor NH4Cl showed such effects. The apoA-I-mediated cellular cholesterol efflux was enhanced by thiol protease inhibitors. Our results suggested that thiol protease inhibitors might provide an alternative way to upregulate ABCA1 protein. This strategy is especially appealing since it may mimic the stabilizing effect of the natural ligands apoA-I.