Effects of deletion of macrophage ABCA7 on lipid metabolism and the development of atherosclerosis in the presence and absence of ABCA1

ABCA7, a close relative of ABCA1 which facilitates cholesterol efflux to lipid-poor apoproteins, has been implicated in macrophage lipid efflux and clearance of apoptotic cells in in vitro studies. In the current study, we investigated the in vivo effects of macrophage ABCA7 deficiency on lipid metabolism and atherosclerosis. Chimeras with dysfunctional ABCA7 in macrophages and other blood cells were generated by transplantation of bone marrow from ABCA7 knockout (KO) mice into irradiated low-density lipoprotein receptor (LDLr) KO mice. Unexpectedly, macrophage ABCA7 deficiency did not significantly affect atherosclerosis susceptibility of LDLr KO mice after 10 weeks Western-type diet feeding. However, ABCA7 deficiency was associated with 2-fold (p<0.05) higher macrophage ABCA1 mRNA expression levels. Combined disruption of ABCA1 and ABCA7 in bone-marrow-derived cells increased atherosclerotic lesion development (1.5-fold (p>0.05) as compared to wild type transplanted mice. However, single deletion of ABCA1 had a similar effect (1.8-fold, p<0.05). Macrophage foam cell accumulation in the peritoneal cavity was reduced in ABCA1/ABCA7 dKO transplanted animals as compared to single ABCA1 KO transplanted mice, which was associated with increased ABCG1 expression. Interestingly, spleens of ABCA1/ABCA7 double KO transplanted mice were significantly larger as compared to the other 3 groups and showed massive macrophage lipid accumulation, a reduction in CD3+ T-cells, and increased expression of key regulators of erythropoiesis. In conclusion, deletion of ABCA7 in bone marrow-derived cells does not affect atherogenesis in the arterial wall neither in the absence or presence of ABCA1. Interestingly, combined deletion of bone marrow ABCA1 and ABCA7 causes severe splenomegaly associated with cellular lipid accumulation, a reduction in splenic CD3+ T cells, and induced markers of erythropoeisis. Our data indicate that ABCA7 may play a role in T cell proliferation and erythropoeisis in spleen.     

Reactive oxygen species‐induced changes in glucose and lipid metabolism contribute to the accumulation of cholesterol in the liver during aging

Aging is a major risk factor for many chronic diseases due to increased vulnerability to external stress and susceptibility to disease. Aging is associated with metabolic liver disease such as nonalcoholic fatty liver. In this study, we investigated changes in lipid metabolism during aging in mice and the mechanisms involved. Lipid accumulation was increased in liver tissues of aged mice, particularly cholesterol. Increased uptake of both cholesterol and glucose was observed in hepatocytes of aged mice as compared with younger mice. The mRNA expression of GLUT2, GK, SREBP2, HMGCR, and HMGCS, genes for cholesterol synthesis, was gradually increased in liver tissues during aging. Reactive oxygen species (ROS) increase with aging and are closely related to various aging‐related diseases. When we treated HepG2 cells and primary hepatocytes with the ROS inducer, H₂O₂, lipid accumulation increased significantly compared to the case for untreated HepG2 cells. H₂O₂ treatment significantly increased glucose uptake and acetyl‐CoA production, which results in glycolysis and lipid synthesis. Treatment with H₂O₂ significantly increased the expression of mRNA for genes related to cholesterol synthesis and uptake. These results suggest that ROS play an important role in altering cholesterol metabolism and consequently contribute to the accumulation of cholesterol in the liver during the aging process.     

Diet-induced lipid accumulation in phospholipid transfer protein-deficient mice: its atherogenicity and potential mechanism

A high saturated fat diet induces free cholesterol and phospholipid accumulation in the plasma of phospholipid transfer protein (Pltp)-deficient mice. In this study, we examined the atherogenic consequence of this phenomenon and investigated the possible mechanism(s). Pltp KO/Apoe KO mice that were fed a coconut oil-enriched high-fat diet (COD) for 7 weeks had higher plasma free cholesterol (149%), phospholipids (15%), and sphingomyelin (54%) than Apoe KO controls. In contrast to chow-fed animals, COD-fed Pltp KO/Apoe KO mice had the same atherosclerotic lesion size as that of Apoe KO mice. Similar to Pltp KO mice, plasma from COD-fed Pltp KO/Apoe KO mice contained VLDL/LDL-sized lamellar particles. Bile measurement indicated that COD-fed Pltp KO mice have 33% less hepatic cholesterol output than controls. In conclusion, COD-fed, Pltp-deficient mice are no longer protected from atherosclerosis and have impaired biliary lipid secretion, which is associated with free cholesterol and phospholipid accumulation.     

Spontaneously hyperlipidemic (SHL) mice: Japanese wild mice with apolipoprotein E deficiency

During inbreeding of Japanese wild mice (Mus musculus molossinus), we established a strain of mice with severe cutaneous xanthomatous lesions. Since those mice showed high plasma cholesterol values, we named them spontaneously hyperlipidemic (SHL) mice; total cholesterol values of these mice (even when fed on conventional low-fat diet) are unusually high throughout the life span. The xanthomatous lesions appear in palms and distal extremities of forelimbs as early as 4 weeks after birth, and continue to expand to chest, abdomen, and face until the mice die before 14 months of age. Histological examination of these lesions revealed cholesterol crystal deposits, an infiltration of foam cells or macrophages, while that of the vascular system revealed atherosclerosis in the aortic sinus. Immunoblot and Northern blot analyses failed to detect apolipoprotein E (APOE) expression in these animals. Consistent with these findings, Southern blot analysis found disruption of the Apoe gene in SHL mice. Phenotypes of SHL mice, however, were distinct from those of Apoe ^tm1Unc (hereafter Apoe ^−/− ) mice, whose Apoe gene was disrupted by homologous recombination; hypercholesterolemia and xanthoma were more severe in SHL mice than in Apoe ^−/− mice, while atherosclerosis was milder in SHL mice. These distinctions suggest that there are modifier genes for the phenotypes. Alternatively, other gene(s), besides the Apoe gene, may be mutated in SHL mice. In either case, comparative genetic and molecular dissection of SHL mice will provide a good opportunity to understand the genetic basis for hyperlipidemia and atherosclerosis. Received: 2 October 1998 / Accepted: 2 December 1998     

Intestinal cholesterol absorption is substantially reduced in mice deficient in both ABCA1 and ACAT2

The process of cholesterol absorption has yet to be completely defined at the molecular level. Because of its ability to esterify cholesterol for packaging into nascent chylomicrons, ACAT2 plays an important role in cholesterol absorption. However, it has been found that cholesterol absorption is not completely inhibited in ACAT2-deficient (ACAT2 KO) mice. Because ABCA1 mRNA expression was increased 3-fold in the small intestine of ACAT2 KO mice, we hypothesized that ABCA1-dependent cholesterol efflux sustains cholesterol absorption in the absence of ACAT2. To test this hypothesis, cholesterol absorption was measured in mice deficient in both ABCA1 and ACAT2 (DKO). Compared with wild-type, ABCA1 KO, or ACAT2 KO mice, DKO mice displayed the lowest level of cholesterol absorption. The concentrations of hepatic free and esterified cholesterol and gallbladder bile cholesterol were significantly reduced in DKO compared with wild-type and ABCA1 KO mice, although these measures of hepatic cholesterol metabolism were very similar in DKO and ACAT2 KO mice. We conclude that ABCA1, especially in the absence of ACAT2, can have a significant effect on cholesterol absorption, although ACAT2 has a more substantial role in this process than ABCA1.     

Leukocyte ABCA1 Remains Atheroprotective in Splenectomized LDL Receptor Knockout Mice

Aim

ATP-binding cassette transporter A1 (ABCA1) is an important mediator of macrophage cholesterol efflux. It mediates the efflux of cellular cholesterol to lipid-poor apolipoprotein A-I. LDL receptor (LDLr) knockout (KO) mice deficient for leukocyte ABCA1 (ABCA1 KO→LDLr KO) show increased atherosclerosis and splenic lipid accumulation despite largely attenuated serum cholesterol levels. In the present study, we aimed to explore the importance of the spleen for the atheroprotective effects of leukocyte ABCA1.

Methods

LDLr KO mice were transplanted with bone marrow from ABCA1 KO mice or wild-type (WT) controls. After 8 weeks recovery, mice were either splenectomized (SP-x) or underwent a sham operation, and were subsequently challenged with a Western-type diet (WTD).

Results

In agreement with previous studies, the atherosclerotic lesion area in ABCA1 KO→LDLr KO sham animals (655±82×10³ µm²) was 1.4-fold (p = 0.03) larger compared to sham WT→LDLr KO mice (459±33×10³ µm²) after 8 weeks WTD feeding, despite 1.7-fold (p<0.001) lower serum cholesterol levels. Interestingly, deletion of ABCA1 in leukocytes led to 1.6-fold higher neutrophil content in the spleen in absence of differences in circulating neutrophils. Levels of KC, an important chemoattractant for neutrophils, in serum, however, were increased 2.9-fold (p = 0.07) in ABCA1 KO→LDLr KO mice. SP-x induced blood neutrophilia as compared to WT→LDLr KO mice (1.9-fold; p<0.05), but did not evoke differences in serum cholesterol and anti-oxLDL antibody levels. Atherosclerotic lesion development, however, was 1.3-fold induced both in the presence and absence of leukocyte ABCA1 (WT: 614±106×10³ µm², ABCA1 KO: 786±44×10³ µm²). Two-way ANOVA revealed independent effects on atherosclerosis for both leukocyte ABCA1 deficiency and SP-x (p<0.05).

Conclusions

The observed splenic alterations induced by leukocyte ABCA1 deficiency do not play a significant role in the anti-atherogenic effects of leukocyte ABCA1 on lesion development.     

Orp8 Deficiency in Bone Marrow-Derived Cells Reduces Atherosclerotic Lesion Progression in LDL Receptor Knockout Mice

Introduction

Oxysterol binding protein Related Proteins (ORPs) mediate intracellular lipid transport and homeostatic regulation. ORP8 downregulates ABCA1 expression in macrophages and cellular cholesterol efflux to apolipoprotein A-I. In line, ORP8 knockout mice display increased amounts of HDL cholesterol in blood. However, the role of macrophage ORP8 in atherosclerotic lesion development is unknown.

Methods and Results

LDL receptor knockout (KO) mice were transplanted with bone marrow (BM) from ORP8 KO mice and C57Bl/6 wild type mice. Subsequently, the animals were challenged with a high fat/high cholesterol Western-type diet to induce atherosclerosis. After 9 weeks of Western-Type diet feeding, serum levels of VLDL cholesterol were increased by 50% in ORP8 KO BM recipients compared to the wild-type recipients. However, no differences were observed in HDL cholesterol. Despite the increase in VLDL cholesterol, lesions in mice transplanted with ORP8 KO bone marrow were 20% smaller compared to WT transplanted controls. In addition, ORP8 KO transplanted mice displayed a modest increase in the percentage of macrophages in the lesion as compared to the wild-type transplanted group. ORP8 deficient macrophages displayed decreased production of pro-inflammatory factors IL-6 and TNFα, decreased expression of differentiation markers and showed a reduced capacity to form foam cells in the peritoneal cavity.

Conclusions

Deletion of ORP8 in bone marrow-derived cells, including macrophages, reduces lesion progression after 9 weeks of WTD challenge, despite increased amounts of circulating pro-atherogenic VLDL. Reduced macrophage foam cell formation and lower macrophage inflammatory potential are plausible mechanisms contributing to the observed reduction in atherosclerosis.     

Augmented atherogenesis in LDL receptor deficient mice lacking both macrophage ABCA1 and ApoE

Aim

ABCA1 protects against atherosclerosis by facilitating cholesterol efflux from macrophage foam cells in the arterial wall to extracellular apolipoprotein (apo) A-I. In contrast to apoA-I, apoE is secreted by macrophages and can, like apoA-I, induce ABCA1-mediated cholesterol efflux. Yet, the combined effect of macrophage ABCA1 and apoE on lesion development is unexplored.

Methods and Results

LDL receptor knockout (KO) mice were transplanted with bone marrow from ABCA1/apoE double KO (dKO) mice, their respective single KO''s, and wild-type (WT) controls and were challenged with a high-fat/high-cholesterol diet for 9 weeks. In vitro cholesterol efflux experiments showed no differences between ABCA1 KO and dKO macrophages. The serum non-HDL/HDL ratio in dKO transplanted mice was 1.7-fold and 2.4-fold (p<0.01) increased compared to WT and ABCA1 KO transplanted mice, respectively. The atherosclerotic lesion area in dKO transplanted animals (650±94×10³ µm²), however, was 1.9-fold (p<0.01) and 1.6-fold (p<0.01) increased compared to single knockouts (ABCA1 KO: 341±20×10³ µm²; apoE KO: 402±78×10³ µm², respectively) and 3.1-fold increased (p<0.001) compared to WT (211±20×10³ µm²). When normalized for serum cholesterol exposure, macrophage ABCA1 and apoE independently protected against atherosclerotic lesion development (p<0.001). Moreover, hepatic expression levels of TNFα and IL-6 were highly induced in dKO transplanted animals (3.0-fold; p<0.05, and 4.3-fold; p<0.001, respectively). In agreement, serum IL-6 levels were also enhanced in ABCA1 KO transplanted mice (p<0.05) and even further enhanced in dKO transplanted animals (3.1-fold as compared to ABCA1 KO transplanted animals; p<0.05).

Conclusions

Combined deletion of macrophage ABCA1 and apoE results in a defect in cholesterol efflux and, compared to ABCA1 KO transplanted mice, elevated serum total cholesterol levels. Importantly, these mice also suffer from enhanced systemic and hepatic inflammation, together resulting in the observed augmented atherosclerotic lesion development.     

Endothelial primary cilia inhibit atherosclerosis

Primary cilia are microtubule‐based structures present on most mammalian cells that are important for intercellular signaling. Cilia are present on a subset of endothelial cells where they project into the vessel lumen and are implicated as mechanical sensors of blood flow. To test the in vivo role of endothelial cilia, we conditionally deleted Ift88, a gene required for ciliogenesis, in endothelial cells of mice. We found that endothelial primary cilia were dispensable for mammalian vascular development. Cilia were not uniformly distributed in the mouse aorta, but were enriched at vascular branch points and sites of high curvature. These same sites are predisposed to the development of atherosclerotic plaques, prompting us to investigate whether cilia participate in atherosclerosis. Removing endothelial cilia increased atherosclerosis in Apoe^?/? mice fed a high‐fat, high‐cholesterol diet, indicating that cilia protect against atherosclerosis. Removing endothelial cilia increased inflammatory gene expression and decreased eNOS activity, indicating that endothelial cilia inhibit pro‐atherosclerotic signaling in the aorta.     

Serine palmitoyltransferase (SPT) deficient mice absorb less cholesterol

Serine palmitoyltransferase (SPT) is the key enzyme for the biosynthesis of sphingolipids. It has been reported that oral administration of myriocin (an SPT inhibitor) decreases plasma sphingomyelin (SM) and cholesterol levels, and reduces atherosclerosis in apoE knockout (KO) mice. We studied cholesterol absorption in myriocin-treated WT or apoE KO animals and found that, after myriocin treatment, the mice absorbed significantly less cholesterol than controls, with no observable pathological changes in the small intestine. More importantly, we found that heterozygous Sptlc1 (a subunit of SPT) KO mice also absorbed significantly less cholesterol than controls. To understand the mechanism, we measured protein levels of Niemann-Pick C1-like 1 (NPC1L1), ABCG5, and ABCA1, three key factors involved in intestinal cholesterol absorption. We found that NPC1L1 and ABCA1 were decreased, whereas ABCG5 was increased in the SPT deficient small intestine. SM levels on the apical membrane were also measured and they were significantly decreased in SPT deficient mice, compared with controls. In conclusion, SPT deficiency might reduce intestinal cholesterol absorption by altering NPC1L1 and ABCG5 protein levels in the apical membranes of enterocytes through lowering apical membrane SM levels. This may be also true for ABCA1 which locates on basal membrane of enterocytes. Manipulation of SPT activity could thus provide a novel alternative treatment for dyslipidemia.     

Deletion of p66Shc in mice increases the frequency of size‐change mutations in the lacZ transgene

Upon oxidative challenge the genome accumulates adducts and breaks that activate the DNA damage response to repair, arrest, or eliminate the damaged cell. Thus, reactive oxygen species (ROS) generated by endogenous oxygen metabolism are thought to affect mutation frequency. However, few studies determined the mutation frequency when oxidative stress is reduced. To test whether in vivo spontaneous mutation frequency is altered in mice with reduced oxidative stress and cell death rate, we crossed p66Shc knockout (p66KO) mice, characterized by reduced intracellular concentration of ROS and by impaired apoptosis, with a transgenic line harboring multiple copies of the lacZ mutation reporter gene as part of a plasmid that can be recovered from organs into Escherichia coli to measure mutation rate. Liver and small intestine from 2‐ to 24‐month‐old, lacZ (p66Shc+/+) and lacZp66KO mice, were investigated revealing no difference in overall mutation frequency but a significant increase in the frequency of size‐change mutations in the intestine of lacZp66KO mice. This difference was further increased upon irradiation of mice with X‐ray. In addition, we found that knocking down cyclophilin D, a gene that facilitates mitochondrial apoptosis acting downstream of p66Shc, increased the size‐change mutation frequency in small intestine. Size‐change mutations also accumulated in death‐resistant embryonic fibroblasts from lacZp66KO mice treated with H₂O₂. These results indicate that p66Shc plays a role in the accumulation of DNA rearrangements and suggest that p66Shc functions to clear damaged cells rather than affect DNA metabolism.     

Rutaecarpine suppresses atherosclerosis in ApoE−/− mice through upregulating ABCA1 and SR-BI within RCT

ABCA1 and scavenger receptor class B type I (SR-BI)/CD36 and lysosomal integral membrane protein II analogous 1 (CLA-1) are the key transporter and receptor in reverse cholesterol transport (RCT). Increasing the expression level of ABCA1 and SR-BI/CLA-1 is antiatherogenic. The aim of the study was to find novel antiatherosclerotic agents upregulating expression of ABCA1 and SR-BI/CLA-1 from natural compounds. Using the ABCA1p-LUC and CLA-1p-LUC HepG2 cell lines, we found that rutaecarpine (RUT) triggered promoters of ABCA1 and CLA-1 genes. RUT increased ABCA1 and SR-BI/CLA-1 expression in vitro related to liver X receptor alpha and liver X receptor beta. RUT induced cholesterol efflux in RAW264.7 cells. ApoE-deficient (ApoE^−/−) mice treated with RUT for 8 weeks showed ∼68.43, 70.23, and 85.56% less en face lesions for RUT (L), RUT (M), and RUT (H) groups, respectively, compared with the model group. Mouse macrophage-specific antibody and filipin staining indicated that RUT attenuated macrophages and cholesterol accumulations in atherosclerotic lesions, respectively. Additionally, ABCA1 and SR-BI expression was highly induced by RUT in livers of ApoE^−/− mice. Meanwhile, RUT treatment significantly increased the fecal ³H-cholesterol excretion, which demonstrated that RUT could promote RCT in vivo. RUT was identified to be a candidate that protected ApoE^−/− mice from developing atherosclerosis through preferentially promoting activities of ABCA1 and SR-BI within RCT.     

Proteomic analysis of HDL from inbred mouse strains implicates APOE associated with HDL in reduced cholesterol efflux capacity via the ABCA1 pathway

Cholesterol efflux capacity associates strongly and negatively with the incidence and prevalence of human CVD. We investigated the relationships of HDL’s size and protein cargo with its cholesterol efflux capacity using APOB-depleted serum and HDLs isolated from five inbred mouse strains with different susceptibilities to atherosclerosis. Like humans, mouse HDL carried >70 proteins linked to lipid metabolism, the acute-phase response, proteinase inhibition, and the immune system. HDL’s content of specific proteins strongly correlated with its size and cholesterol efflux capacity, suggesting that its protein cargo regulates its function. Cholesterol efflux capacity with macrophages strongly and positively correlated with retinol binding protein 4 (RBP4) and PLTP, but not APOA1. In contrast, ABCA1-specific cholesterol efflux correlated strongly with HDL’s content of APOA1, APOC3, and APOD, but not RBP4 and PLTP. Unexpectedly, APOE had a strong negative correlation with ABCA1-specific cholesterol efflux capacity. Moreover, the ABCA1-specific cholesterol efflux capacity of HDL isolated from APOE-deficient mice was significantly greater than that of HDL from wild-type mice. Our observations demonstrate that the HDL-associated APOE regulates HDL’s ABCA1-specific cholesterol efflux capacity. These findings may be clinically relevant because HDL’s APOE content associates with CVD risk and ABCA1 deficiency promotes unregulated cholesterol accumulation in human macrophages.     

Hyperglycemia accelerates ATP‐binding cassette transporter A1 degradation via an ERK‐dependent pathway in macrophages

An elevation in blood glucose concentration leads to increased risk of developing diabetes‐associated atherosclerotic cardiovascular disease due to an excessive accumulation of cholesterol in arterial macrophages. ATP‐binding cassette transporter A1 (ABCA1) is an atheroprotective protein that mediates the export of cholesterol from macrophages. The present study aims to investigate the effect of hyperglycemia on the regulation of ABCA1 expression and to explore its underlying mechanisms of regulation in macrophages. Our results show that high glucose activates the extracellular signal‐regulated kinases (ERK) signaling pathway via reactive oxygen species (ROS) production, which in turn down‐regulates ABCA1 mRNA and protein expression. This down‐regulation is mediated by accelerating ABCA1 mRNA and protein degradation in macrophages exposed to high concentrations of glucose. Our results provide evidence for the first time that hyperglycemia inhibits ABCA1 expression by ERK‐modulated ABCA1 mRNA and protein stability. Overall, these results provide a mechanism for hyperglycemia‐induced reduction in ABCA1 expression, which suggests a promising strategy for the treatment of diabetes‐associated atherosclerosis. J. Cell. Biochem. 114: 1364–1373, 2013. © 2012 Wiley Periodicals, Inc.     

Wogonin promotes cholesterol efflux by increasing protein phosphatase 2B-dependent dephosphorylation at ATP-binding cassette transporter-A1 in macrophages

Wogonin, one component in Scutellaria baicalensis Georgi extracts, has several beneficial properties for cancers and inflammatory diseases. However, the efficacy of wogonin in cholesterol metabolism of macrophages remains unknown. In macrophages, cholesterol uptake is controlled by scavenger receptors (SR-A and CD36) and cholesterol efflux by SR-BI, ATP-binding cassette transporter-A1 (ABCA1) and ABCG1. In the present study, we investigated the effect and underlying molecular mechanism of wogonin on the formation of macrophage foam cells by murine J774.A1 macrophages. Wogonin attenuated oxidized low-density lipoprotein (oxLDL)-induced cholesterol accumulation in macrophages. The binding of oxLDL to macrophages and protein expression of SR-A and CD36 were not affected by wogonin. Wogonin enhanced cholesterol efflux and increased the protein level of ABCA1 without affecting the protein expression of SR-BI or ABCG1. Inhibition of ABCA1 by pharmacological inhibitor 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid disodium salt or neutralizing antibody abolished this suppressive effect of wogonin on lipid accumulation. Moreover, the up-regulation of ABCA1 protein by wogonin resulted from a decrease in degradation rate of ABCA1 protein, with no effect on ABCA1 mRNA expression. This reduction in ABCA1 degradation was due to increased protein phosphatase 2B (PP2B)-mediated ABCA1 dephosphorylation, as evidenced by increased interaction between ABCA1 and PP2B; pharmacological inhibition of PP2B would prevent wogonin-induced ABCA1 protein expression, dephosphorylation and attenuation of lipid accumulation. Collectively, wogonin increases the protein stability of ABCA1 via PP2B-mediated dephosphorylation, thus leading to reduced cholesterol accumulation in macrophage foam cells.     

Lysophosphatidylcholine promotes cholesterol efflux from mouse macrophage foam cells via PPARgamma-LXRalpha-ABCA1-dependent pathway associated with apoE

Formation of macrophage-derived foam cells is a hallmark in earlier stages of atherosclerosis (AS). Increased cholesterol efflux from macrophage foam cells promote atherosclerotic regression. In the present study, lysophosphatidylcholine (LPC) promoting cholesterol efflux from macrophage foam cells was observed, and the mechanism underlying the action was investigated. Macrophage foam cells from mice were incubated with different concentrations of LPC (10, 20, 40, 80 microM), and the free cholesterol in medium increased but total intracellular cholesterol decreased. At the same time, the expression of PPARgamma, LXRalpha, ABCA1 was enhanced in a dose-dependent manner. The treatment of macrophage foam cells with 40 microM LPC for 12, 24 and 48 h promoted cellular cholesterol efflux in a time-dependent manner, meanwhile expression of PPARgamma, LXRalpha, ABCA1 was also raised respectively. Addition of different specific inhibitors of PPARgamma (GW9662), LXRalpha (GGPP), ABCA1 (DIDS) to the foam cells significantly suppressed LPC-induced cholesterol efflux. Also treatment with specific inhibitors of PPARgamma or LXRalpha decreased ABCA1 mRNA and protein expressions. LPC (40 microM)-induced cholesterol efflux was significantly lower in macrophage foam cells from apoE deficient mice than from normal C57BL/6J mice. In contrast, 10 microg apoAI-induced cholesterol efflux from foam cells remained in apoE deficient mice. The present results indicate that LPC promotes cholesterol efflux from macrophage foam cells via a PPARgamma-LXRalpha-ABCA1-dependent pathway. Furthermore, apoE may be involved in this process.     

PLK1 promotes cholesterol efflux and alleviates atherosclerosis by up-regulating ABCA1 and ABCG1 expression via the AMPK/PPARγ/LXRα pathway

Polo-like kinase 1 (PLK1) is a serine/threonine kinase involving lipid metabolism and cardiovascular disease. However, its role in atherogenesis has yet to be determined. The aim of this study was to observe the impact of PLK1 on macrophage lipid accumulation and atherosclerosis development and to explore the underlying mechanisms. We found a significant reduction of PLK1 expression in lipid-loaded macrophages and atherosclerosis model mice. Lentivirus-mediated overexpression of PLK1 promoted cholesterol efflux and inhibited lipid accumulation in THP-1 macrophage-derived foam cells. Mechanistic analysis revealed that PLK1 stimulated the phosphorylation of AMP-activated protein kinase (AMPK), leading to activation of the peroxisome proliferator-activated receptor γ (PPARγ)/liver X receptor α (LXRα) pathway and up-regulation of ATP binding cassette transporter A1 (ABCA1) and ABCG1 expression. Injection of lentiviral vector expressing PLK1 increased reverse cholesterol transport, improved plasma lipid profiles and decreased atherosclerotic lesion area in apoE-deficient mice fed a Western diet. PLK1 overexpression also facilitated AMPK and HSL phosphorylation and enhanced the expression of PPARγ, LXRα, ABCA1, ABCG1 and LPL in the aorta. In summary, these data suggest that PLK1 inhibits macrophage lipid accumulation and mitigates atherosclerosis by promoting ABCA1- and ABCG1-dependent cholesterol efflux via the AMPK/PPARγ/LXRα pathway.