Akt isoform-dependent regulation of ATP-Binding cassette A1 expression by apolipoprotein E.
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ABSTRACT:
SUBMITTER: Okoro EU
PROVIDER: S-EPMC4935621 | biostudies-other | 2016 Aug
REPOSITORIES: biostudies-other
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Project description:Apolipoprotein M (apoM) is a novel apolipoprotein that is reportedly necessary for pre beta HDL formation; however, its detailed function remains unknown. We investigated the biogenesis and properties of apoM and its effects on the initial steps of nascent pre beta HDL assembly by ABCA1 in HEK293 cells. Transiently transfected apoM was localized primarily in the endomembrane compartment. Pulse-chase analyses demonstrated that apoM is inefficiently secreted, relative to human serum albumin, and that approximately 50% remains membrane-associated after extraction with sodium carbonate, pH 11.5. To investigate the role of apoM in nascent pre beta HDL formation, ABCA1-expressing or control cells, transfected with empty vector, apoM, or C-terminal epitope-tagged apoM (apoM-C-FLAG), were incubated with (125)I-apoA-I for 24 h. Conditioned media were harvested and fractionated by fast-protein liquid chromatography (FPLC) to monitor HDL particle size. Pre beta HDL particles were formed effectively in the absence of apoM expression; however, increased apoM expression stimulated the formation of larger-sized nascent pre beta HDLs. Immunoprecipitation with anti-apoA-I antibody followed by apoM Western blot analysis revealed that little secreted apoM was physically associated with pre beta HDL. Our results suggest that apoM is an atypical secretory protein that is not necessary for ABCA1-dependent pre beta HDL formation but does stimulate the formation of larger-sized pre beta HDL. We propose that apoM may function catalytically at an intracellular site to transfer lipid onto pre beta HDL during or after their formation by ABCA1.
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Project description:The contribution of soluble epoxide hydrolase (sEH) to atherosclerosis has been well defined. However, less is understood about the role of sEH and its underlying mechanism in the cholesterol metabolism of macrophages. The expression of sEH protein was increased in atherosclerotic aortas of apolipoprotein E-deficient mice, primarily in macrophage foam cells. Oxidized low-density lipoprotein (oxLDL) increased sEH expression in macrophages. Genetic deletion of sEH (sEH-/- ) in macrophages markedly exacerbated oxLDL-induced lipid accumulation and decreased the expression of ATP-binding cassette transporters-A1 (ABCA1) and apolipoprotein AI-dependent cholesterol efflux following oxLDL treatment. The down-regulation of ABCA1 in sEH-/- macrophages was due to an increase in the turnover rate of ABCA1 protein but not in mRNA transcription. Inhibition of phosphatase activity, but not hydrolase activity, of sEH decreased ABCA1 expression and cholesterol efflux following oxLDL challenge, which resulted in increased cholesterol accumulation. Additionally, oxLDL increased the phosphatase activity, promoted the sEH-ABCA1 complex formation and decreased the phosphorylated level of ABCA1 at threonine residues. Overexpression of phosphatase domain of sEH abrogated the oxLDL-induced ABCA1 phosphorylation and further increased ABCA1 expression and cholesterol efflux, leading to the attenuation of oxLDL-induced cholesterol accumulation. Our findings suggest that the phosphatase domain of sEH plays a crucial role in the cholesterol metabolism of macrophages.
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Project description:Atherosclerosis is a chronic inflammatory disease related to a massive accumulation of cholesterol in the artery wall. Photobiomodulation therapy (PBMT) has been reported to possess cardioprotective effects but has no consensus on the underlying mechanisms. Here, we aimed to investigate whether PBMT could ameliorate atherosclerosis and explore the potential molecular mechanisms. The Apolipoprotein E (ApoE)-/- mice were fed with western diet (WD) for 18 weeks and treated with PBMT once a day in the last 10 weeks. Quantification based on Oil red O-stained aortas showed that the average plaque area decreased 8.306 ± 2.012% after PBMT (P < .05). Meanwhile, we observed that high-density lipoprotein cholesterol level in WD + PBMT mice increased from 0.309 ± 0.037 to 0.472 ± 0.038 nmol/L (P < .05) compared with WD mice. The further results suggested that PBMT could promote cholesterol efflux from lipid-loaded primary peritoneal macrophages and inhibit foam cells formation via up-regulating the ATP-binding cassette transporters A1 expression. A contributing mechanism involved in activating the phosphatidylinositol 3-kinases/protein kinase C zeta/specificity protein 1 signalling cascade. Our study outlines that PBMT has a protective role on atherosclerosis by promoting macrophages cholesterol efflux and provides a new strategy for treating atherosclerosis.
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Project description:ATP-binding cassette transporter (ABC) A1 is required for the lipidation of apolipoprotein A-I to generate high density lipoprotein (HDL). This process is proposed to occur through a retro-endocytosis pathway in which apoA-I internalizes with ABCA1 and generates HDL from the endosomal compartments before resecretion. The aim of this study was to determine the route of apoA-I endocytosis and whether endocytosis contributes to HDL biogenesis. Using confocal microscopy, we found that internalized apoA-I only transiently colocalized with transferrin, a retro-endocytosis marker. Instead, apoA-I perfectly colocalized with a bulk phase uptake marker (fluorescein isothiocyanate-dextran) and, at later time points, with LysoTracker in several cell models including macrophages, fibroblasts, and baby hamster kidney cells. ABCA1 colocalized poorly with internalized apoA-I. To determine the contribution of internalized apoA-I to HDL biogenesis, we specifically removed apoA-I from the cell surface and analyzed the fate of internalized apoA-I. We found that 23% of cell-associated apoA-I was internalized at steady state. Of internalized apoA-I, only 20% was converted to HDL, and the rest was degraded, consistent with a lysosomal destination. We also found that apoA-I was released approximately five times faster from the plasma membrane than from the intracellular compartments. From these kinetic parameters, we estimated that approximately 5.6% of apoA-I that interacts with cells is degraded and that internalized apoA-I contributes to approximately 1.4% of total HDL production. We also found that blocking endocytosis with sucrose or cytochalasin D did not decrease cholesterol efflux or HDL biogenesis. We therefore conclude that the plasma membrane is the main platform where ABCA1-mediated lipidation of apoA-I occurs.
| S-EPMC3259641 | biostudies-literature
Project description:Cholesterol homeostasis is essential in normal physiology of all cells. One of several proteins involved in cholesterol homeostasis is the ATP-binding cassette transporter A1 (ABCA1), a transmembrane protein widely expressed in many tissues. One of its main functions is the efflux of intracellular free cholesterol and phospholipids across the plasma membrane to combine with apolipoproteins, mainly apolipoprotein A-I (Apo A-I), forming nascent high-density lipoprotein-cholesterol (HDL-C) particles, the first step of reverse cholesterol transport (RCT). In addition, ABCA1 regulates cholesterol and phospholipid content in the plasma membrane affecting lipid rafts, microparticle (MP) formation and cell signaling. Thus, it is not surprising that impaired ABCA1 function and altered cholesterol homeostasis may affect many different organs and is involved in the pathophysiology of a broad array of diseases. This review describes evidence obtained from animal models, human studies and genetic variation explaining how ABCA1 is involved in dyslipidemia, coronary heart disease (CHD), type 2 diabetes (T2D), thrombosis, neurological disorders, age-related macular degeneration (AMD), glaucoma, viral infections and in cancer progression.
| S-EPMC7915494 | biostudies-literature
Project description:Fibroblasts from patients with Tangier disease carrying ATP-binding cassette A1 (ABCA1) loss-of-function mutations are characterized by cardiolipin accumulation, a mitochondrial-specific phospholipid. Suppression of ABCA1 expression occurs in glomeruli from patients with diabetic kidney disease (DKD) and in human podocytes exposed to DKD sera collected prior to the development of DKD. We demonstrated that siRNA ABCA1 knockdown in podocytes led to reduced oxygen consumption capabilities associated with alterations in the oxidative phosphorylation (OXPHOS) complexes and with cardiolipin accumulation. Podocyte-specific deletion of Abca1 (Abca1fl/fl) rendered mice susceptible to DKD, and pharmacological induction of ABCA1 improved established DKD. This was not mediated by free cholesterol, as genetic deletion of sterol-o-acyltransferase-1 (SOAT1) in Abca1fl/fl mice was sufficient to cause free cholesterol accumulation but did not cause glomerular injury. Instead, cardiolipin mediates ABCA1-dependent susceptibility to podocyte injury, as inhibition of cardiolipin peroxidation with elamipretide improved DKD in vivo and prevented ABCA1-dependent podocyte injury in vitro and in vivo. Collectively, we describe a pathway definitively linking ABCA1 deficiency to cardiolipin-driven mitochondrial dysfunction. We demonstrated that this pathway is relevant to DKD and that ABCA1 inducers or inhibitors of cardiolipin peroxidation may each represent therapeutic strategies for the treatment of established DKD.
| S-EPMC6668702 | biostudies-literature
Project description:ObjectiveDysregulation of cholesterol metabolism in the brain is responsible for many lipid storage disorders, including Niemann-Pick disease type C (NPC). Here, we have investigated whether cyclodextrin (CD) and apolipoprotein A-I (apoA-I) induce the same signal to inhibit cell cholesterol accumulation by focusing on the main proteins involved in cholesterol homeostasis in response to CD and apoA-I treatment.Materials and methodsIn this experimental study, astrocytes were treated with apoA-I or CD and then lysed in RIPA buffer. We used Western blot to detect protein levels of 3-hydroxy-3-methyl-glutaryl coenzyme A reductase (HMGCR) and ATP-binding cassette transporter A1 (ABCA1). Cell cholesterol content and cholesterol release in the medium were also measured.ResultsApoA-I induced a significant increase in ABCA1 and a mild increase in HMGCR protein level, whereas CD caused a significant increase in HMGCR with a significant decrease in ABCA1. Both apoA-I and CD increased cholesterol release in the medium. A mild, but not significant increase, in cell cholesterol content was seen by apoA-I; however, a significant increase in cell cholesterol was detected when the astrocytes were treated with CD.ConclusionCD, like apoA-I, depletes cellular cholesterol. This depletion occurs in a different way from apoA-I that is through cholesterol efflux. Depletion of cell cholesterol with CDs led to reduced protein levels of ABCA1 along with increased HMGCR and accumulation of cell cholesterol. This suggested that CDs, unlike apoA-I, could impair the balance between cholesterol synthesis and release, and interfere with cellular function that depends on ABCA1.
| S-EPMC7944131 | biostudies-literature
Project description:ObjectiveIncreasing plasma glucose levels are associated with increasing risk of vascular disease. We tested the hypothesis that there is a glycaemia-mediated impairment of reverse cholesterol transport (RCT). We studied the influence of plasma glucose on expression and function of a key mediator in RCT, the ATP binding cassette transporter-A1 (ABCA1) and expression of its regulators, liver X receptor-α (LXRα) and peroxisome proliferator-activated receptor-γ (PPARγ).Methods and resultsLeukocyte ABCA1, LXRα and PPARγ expression was measured by polymerase chain reaction in 63 men with varying degrees of glucose homeostasis. ABCA1 protein concentrations were measured in leukocytes. In a sub-group of 25 men, ABCA1 function was quantified as apolipoprotein-A1-mediated cholesterol efflux from 2-3 week cultured skin fibroblasts. Leukocyte ABCA1 expression correlated negatively with circulating HbA1c and glucose (rho = -0.41, p<0.001; rho = -0.34, p = 0.006 respectively) and was reduced in Type 2 diabetes (T2DM) (p = 0.03). Leukocyte ABCA1 protein was lower in T2DM (p = 0.03) and positively associated with plasma HDL cholesterol (HDL-C) (rho = 0.34, p = 0.02). Apolipoprotein-A1-mediated cholesterol efflux correlated negatively with fasting glucose (rho = -0.50, p = 0.01) and positively with HDL-C (rho = 0.41, p = 0.02). It was reduced in T2DM compared with controls (p = 0.04). These relationships were independent of LXRα and PPARγ expression.ConclusionsABCA1 expression and protein concentrations in leukocytes, as well as function in cultured skin fibroblasts, are reduced in T2DM. ABCA1 protein concentration and function are associated with HDL-C levels. These findings indicate a glycaemia-related, persistent disruption of a key component of RCT.
| S-EPMC3144880 | biostudies-literature
Project description:ATP-binding cassette (ABC) efflux transporters, including multidrug resistance protein 1 (Mdr1), breast cancer resistance protein (Bcrp), and multidrug resistance-associated proteins (Mrps) extrude chemicals from the brain. Although ABC transporters are critical for blood-brain barrier integrity, less attention has been placed on the regulation of these proteins in brain parenchymal cells such as microglia. Prior studies demonstrate that inflammation after lipopolysaccharide (LPS) treatment alters transporter expression in the livers of mice. Here, we sought to determine the effects of inflammation on the expression and function of transporters in microglia. To test this, the expression and function of ABC efflux transport proteins were quantified in mouse BV-2 microglial cells in response to activation with LPS. Intracellular retention of fluorescent rhodamine 123, Hoechst 33342, and calcein acetoxymethyl ester was increased in LPS-treated microglia, suggesting that the functions of Mdr1, Bcrp, and Mrps were decreased, respectively. LPS reduced Mdr1, Bcrp, and Mrp4 mRNA and protein expression between 40 and 70%. Conversely, LPS increased expression of Mrp1 and Mrp5 mRNA and protein. Immunofluorescent staining confirmed reduced Bcrp and Mrp4 and elevated Mrp1 and Mrp5 protein in activated microglia. Pharmacological inhibition of nuclear factor κB (NF-κB) transcriptional signaling attenuated down-regulation of Mdr1a mRNA and potentiated up-regulation of Mrp5 mRNA in LPS-treated cells. Together, these data suggest that LPS stimulates microglia and impairs efflux of prototypical ABC transporter substrates by altering mRNA and protein expression, in part through NF-κB signaling. Decreased transporter efflux function in microglia may lead to the retention of toxic chemicals and aberrant cell-cell communication during neuroinflammation.
| S-EPMC3500534 | biostudies-literature
Project description:Atherosclerosis is a process of imbalanced lipid metabolism in the vascular walls. The underlying pathology mainly involves the deposition of oxidized lipids in the endothelium and the accumulation of cholesterol in macrophages. Macrophages export excessive cholesterol (cholesterol efflux) through ATP-binding cassette transporter A1 (ABCA1) to counter the progression of atherosclerosis. We synthesized novel chalcone derivatives and assessed their effects and the underlying mechanisms on ABCA1 expression in macrophages. Human THP-1 macrophages were treated with synthetic chalcone derivatives for 24 h. In Western blot and flow cytometry analyses, a chalcone derivative, (E)-1-(3,4-diisopropoxyphenyl)-3-(4-isopropoxy-3-methoxyphenyl)prop- 2-en-1-one (1m), was observed to significantly enhance ABCA1 protein expression in THP-1 cells (10 µM, 24 h). Levels of mRNA of ABCA1 and liver X receptor alpha (LXRα) were quantified using a real-time quantitative polymerase chain reaction technique and were found to be significantly increased after treatment with the novel chalcone derivative 1m. Several microRNAs, including miR155, miR758, miR10b, miR145, miR33, and miR106b, which functionally inhibit ABCA1 expression were suppressed after treatment with 1m. Collectively, 1m increases ABCA1 expression in human THP-1 macrophages. The mechanisms involve the activation of the LXRα-ABCA1 pathway and suppression of certain microRNAs that regulate ABCA1 expression.
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