RESUMEN
Prevention of atherosclerosis is important because it is a risk factor for cardiovascular diseases globally. One of the causes of atherosclerosis is accumulation of cholesterol and triglycerides in peripheral cells. ATP-binding cassette protein A1 (ABCA1) and G1 (ABCG1) are important in eliminating excess cholesterol from cells including macrophages and forming high-density lipoprotein, which contributes to the prevention and regression of atherosclerosis. Enhanced cholesterol efflux activities of ABCA1 and ABCG1 are expected to prevent the progression of atherosclerosis. ABCA1 and ABCG1 are induced by the LXR/RXR pathway and regulated transcriptionally, post-transcriptionally, and post-translationally. Their mRNAs are destabilized by microRNAs and their cellular localization and degradation are regulated by other proteins and phosphorylation. Furthermore, ABCA1 and ABCG1 suppress the inflammatory responses of macrophages. These proteins are effective targets because their increased activities can suppress cholesterol accumulation and inflammation in macrophages. Moreover, ABCA1 and ABCG1 prevent amyloid ß accumulation; therefore, their increased activity may prevent Alzheimer's disease. Because ABCA1 and ABCG1 are affected by transcriptional, post-transcriptional, and post-translational regulation, the regulatory factors involved could also serve as therapeutic targets. This review highlights that ABCA1 and ABCG1 could be potential therapeutic targets for preventing atherosclerosis by regulating their expression, degradation, and localization.
Asunto(s)
Transportador 1 de Casete de Unión a ATP , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1 , Aterosclerosis/genética , Aterosclerosis/prevención & control , Terapia Molecular Dirigida , Transportador 1 de Casete de Unión a ATP/genética , Transportador 1 de Casete de Unión a ATP/metabolismo , Transportador 1 de Casete de Unión a ATP/fisiología , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/genética , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/metabolismo , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/fisiología , Enfermedad de Alzheimer/prevención & control , Péptidos beta-Amiloides/metabolismo , Aterosclerosis/etiología , Aterosclerosis/metabolismo , Transporte Biológico/genética , Colesterol/metabolismo , Progresión de la Enfermedad , Humanos , Macrófagos/metabolismo , Receptores X Retinoide/metabolismo , Transducción de Señal/genética , Transducción de Señal/fisiología , Transcripción Genética/fisiología , Triglicéridos/metabolismoRESUMEN
BACKGROUND: Aberrant concentration, structure and functionality of High Density Lipoprotein (HDL) are associated with many prevalent diseases, including cardiovascular disease and non-alcoholic fatty liver disease (NAFLD). Mice with liver-specific ablation of Hnf4α (H4LivKO) present steatosis and dyslipidemia by mechanisms that are not completely understood. The aim of this study was to explore the role of liver HNF4A in HDL metabolism and the development of steatosis. METHODS AND RESULTS: Serum and tissue samples were obtained from 6-weeks old H4LivKO mice and their littermate controls. Liver and serum lipids were measured and HDL structure and functionality were assessed. Global gene expression changes in the liver were analyzed by expression arrays, validations were performed by RT-qPCR and DNA-protein interactions were studied by chromatin immunoprecipitation (ChIP). H4LivKO mice presented liver steatosis, increased liver triglyceride content and decreased concentration of serum total cholesterol, HDL cholesterol, triglycerides, phospholipids and cholesteryl esters. Most classes of phospholipids showed significant changes in species ratio and sphingosine-1-phosphate (S1P) levels were reduced. H4LivKO serum was enriched in the smaller, denser HDL particles, devoid of APOA2 and APOM apolipoproteins, exhibiting decreased activity of paraoxonase-1 but retaining macrophage cholesterol efflux capacity and phospho-AKT activation in endothelial cells. Global gene expression analysis revealed the association of liver HNF4A with known and novel regulators of HDL metabolism as well as NAFLD-susceptibility genes. CONCLUSIONS: HNF4A ablation in mouse liver causes hepatic steatosis, perturbations in HDL structure and function and significant global changes in gene expression. This study reveals new targets of HNF4A involved in HDL metabolism and the development of steatosis and enriches our knowledge on HDL functionality in NAFLD.
Asunto(s)
Factor Nuclear 4 del Hepatocito/fisiología , Lipoproteínas HDL/metabolismo , Enfermedad del Hígado Graso no Alcohólico/etiología , Transportador 1 de Casete de Unión a ATP/fisiología , Animales , Arildialquilfosfatasa/metabolismo , Perfilación de la Expresión Génica , Metabolismo de los Lípidos , Lipoproteínas HDL/química , Lisofosfolípidos/sangre , Ratones , Ratones Noqueados , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Esfingosina/análogos & derivados , Esfingosina/sangreRESUMEN
BACKGROUND: The function of miR-20a-5p in pulmonary artery smooth muscle cells (PASMCs) and the underlying mechanism remains largely unknown. METHODS: C57BL/6J mice and PASMCs were used for constructing pulmonary artery hypertension (PAH) animal and cell models, respectively. Reverse transcription polymerase chain reaction (RT-PCR) was employed to detect miR-20a-5p and ATP-binding cassette subfamily A member 1 (ABCA1) messenger RNA expression. CCK-8, Transwell, and TUNEL experiments were used to determine PASMCs proliferation, migration, and apoptosis. The relationship between miR-20a-5p and ABCA1 was detected by luciferase reporter experiment, Western blot analysis, and qRT-PCR. RESULTS: miR-20a-5p was remarkably elevated in PASMCs of PAH mice and human PASMCs treated by hypoxia, while ABCA1 was remarkably decreased. After transfection of miR-20a-5p mimics, PASMCs proliferation and migration were promoted and PASMCs apoptosis was suppressed. ABCA1 was confirmed to be a target of miR-20a-5p and restoration of ABCA1 reversed the function of miR-20a-5p. CONCLUSION: miR-20a-5p enhances the proliferation and migration of PASMCs to promote the development of PAH via targeting ABCA1.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/fisiología , Movimiento Celular/fisiología , Proliferación Celular/fisiología , MicroARNs/fisiología , Músculo Liso Vascular/citología , Arteria Pulmonar/citología , Animales , Apoptosis/fisiología , Células Cultivadas , Humanos , Ratones , Ratones Endogámicos C57BLRESUMEN
Cholesterol-laden macrophage foam cells are a hallmark of atherosclerosis. For that reason, cholesterol metabolism in macrophages has attracted considerable scrutiny, particularly the mechanisms by which macrophages unload surplus cholesterol (a process referred to as "cholesterol efflux"). Many studies of cholesterol efflux in macrophages have focused on the role of ABC transporters in moving cholesterol onto high-density lipoproteins (HDLs), but other mechanisms for cholesterol efflux likely exist. We hypothesized that macrophages have the capacity to unload cholesterol directly onto adjacent cells. To test this hypothesis, we used methyl-ß-cyclodextrin (MßCD) to load mouse peritoneal macrophages with [13C]cholesterol. We then plated the macrophages (in the absence of serum or HDL) onto smooth muscle cells (SMCs) that had been metabolically labeled with [15N]choline. After incubating the cells overnight in the absence of HDL or serum, we visualized 13C and 15N distribution by nanoscale secondary ion mass spectrometry (NanoSIMS). We observed substantial 13C enrichment in SMCs that were adjacent to [13C]cholesterol-loaded macrophages-including in cytosolic lipid droplets of SMCs. In follow-up studies, we depleted "accessible cholesterol" from the plasma membrane of [13C]cholesterol-loaded macrophages with MßCD before plating the macrophages onto the SMCs. After an overnight incubation, we again observed substantial 13C enrichment in the SMCs adjacent to macrophages. Thus, macrophages transfer cholesterol to adjacent cells in the absence of serum or HDL. We suspect that macrophages within tissues transfer cholesterol to adjacent cells, thereby contributing to the ability to unload surplus cholesterol.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/metabolismo , Colesterol/metabolismo , Macrófagos/metabolismo , Transportador 1 de Casete de Unión a ATP/fisiología , Transportadoras de Casetes de Unión a ATP/metabolismo , Animales , Aterosclerosis/metabolismo , Aterosclerosis/fisiopatología , Transporte Biológico , Células Espumosas/metabolismo , Metabolismo de los Lípidos , Lipoproteínas HDL/metabolismo , Macrófagos/fisiología , Macrófagos Peritoneales/metabolismo , Ratones , Ratones Endogámicos C57BL , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Suero/metabolismo , beta-Ciclodextrinas/metabolismoRESUMEN
Purpose: This study aimed to investigate the role and pathophysiological mechanism of ATP binding cassette transporter A1 (ABCA1) in regulating the IOP and aqueous humor outflow. Methods: ABCA1 expression was measured in trabecular meshwork samples obtained from patients with POAG and human donor eyes by Western blot. To further evaluate the functional significance of ABCA1, porcine angular aqueous plexus (AAP) cells, which are equivalent to human Schlemm's canal endothelial cells, were either treated with ABCA1 agonist GW3965 or transduced with lentivirus expressing ABCA1-shRNA. Transendothelial electrical resistance, protein expression, and nitric oxide (NO) concentration were measured. GW3965 was administered by intracameral injection. IOP and aqueous humor outflow facility were also measured. Results: ABCA1 expression was significantly higher in the trabecular meshwork tissue of patients with POAG compared with controls. ABCA1 upregulation in angular aqueous plexus cells decreased the transendothelial electrical resistance in the angular aqueous plexus monolayers accompanied by a 0.56-fold decrease in caveolin-1 expression and a 2.85-fold and 1.17-fold increase in endothelial NO synthase expression and NO concentration, respectively (n = 3, P < 0.05). Conversely, ABCA1 downregulation increased transendothelial electrical resistance and caveolin-1 expression and decreased endothelial NO synthase expression and NO production (n = 3, P < 0.05). GW3965 decreased IOP and significantly increased conventional outflow facility (P < 0.05). Conclusions: Regulation of aqueous humor outflow via the caveolin-1/endothelial NO synthase/NO pathway is a newly defined function of ABCA1 that is different from its traditional role in mediating cholesterol efflux. ABCA1 is a compelling, novel therapeutic candidate for the treatment of glaucoma and ocular hypertension.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/fisiología , Caveolina 1/metabolismo , Glaucoma de Ángulo Abierto/metabolismo , Presión Intraocular/fisiología , Óxido Nítrico Sintasa de Tipo III/metabolismo , Óxido Nítrico/metabolismo , Transducción de Señal/fisiología , Animales , Humor Acuoso/fisiología , Benzoatos/farmacología , Bencilaminas/farmacología , Western Blotting , Impedancia Eléctrica , Células Endoteliales/efectos de los fármacos , Glaucoma de Ángulo Abierto/cirugía , Humanos , Lentivirus/genética , ARN Interferente Pequeño/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Porcinos , Malla Trabecular/metabolismo , Trabeculectomía , TransfecciónRESUMEN
Cholesterol is a critical component of membranes and a precursor for hormones and other signaling molecules. Previously, we showed that unlike astrocytes, glioblastoma cells do not downregulate cholesterol synthesis when plated at high density. In this report, we show that high cell density induces ABCA1 expression in glioblastoma cells, enabling them to get rid of excess cholesterol generated by an activated cholesterol biosynthesis pathway. Because oxysterols are agonists for Liver X Receptors (LXRs), we investigated whether increased cholesterol activates LXRs to maintain cholesterol homeostasis in highly-dense glioblastoma cells. We observed that dense cells had increased oxysterols, which activated LXRß to upregulate ABCA1. Cells with CRISPR-mediated knockdown of LXRß, but not ABCA1, had decreased cell cycle progression and cell survival, and decreased feedback repression of the mevalonate pathway in densely-plated glioma cells. LXRß gene expression poorly correlates with ABCA1 in glioblastoma patients, and expression of each gene correlates with poor patient prognosis in different prognostic subtypes. Finally, gene expression and lipidomics analyses cells revealed that LXRß regulates the expression of immune response gene sets and lipids known to be involved in immune modulation. Thus, therapeutic targeting of LXRß in glioblastoma might be effective through diverse mechanisms.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/fisiología , Neoplasias Encefálicas/patología , Proliferación Celular/fisiología , Glioblastoma/patología , Metabolismo de los Lípidos , Receptores X del Hígado/fisiología , Transportador 1 de Casete de Unión a ATP/genética , Neoplasias Encefálicas/inmunología , Neoplasias Encefálicas/metabolismo , Colesterol/metabolismo , Glioblastoma/inmunología , Glioblastoma/metabolismo , Homeostasis , Humanos , Receptores X del Hígado/metabolismo , Ácido Mevalónico/metabolismo , Transducción de Señal , Transcripción GenéticaRESUMEN
Our previous study has indicated that the parathyroid hormone type 1 receptor (PTHR1) may play important roles in development and progression of osteosarcoma (OS) by regulating Wnt, angiogenesis, and inflammation pathway genes. The goal of this study was to further illuminate the roles of PTHR1 in OS by investigating upstream regulation mechanisms (including microRNA [miRNA] and transcription factors [TFs]) of crucial genes. The microarray dataset GSE46861 was downloaded from the Gene Expression Omnibus database, in which six tumors with short hairpin RNA (shRNA) PTHR1 knockdown (PTHR1.358) and six tumors with shRNA control knockdown (Ren.1309) were collected from mice. Differentially expressed genes (DEGs) between PTHR1.358 and Ren.1309 were identified using the linear models for microarray data (LIMMA) method, and then the miRNA-TF-mRNA regulatory network was constructed using data from corresponding databases, followed by module analysis, to screen crucial regulatory relationships. OS-related human miRNAs were extracted from the curated Osteosarcoma Database. Gene ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were enriched using the Database for Annotation, Visualization, and Integrated Discovery (DAVID) tool. As a result, the miRNA-TF-mRNA regulatory network, including 1049 nodes (516 miRNA, 25 TFs, and 508 DEGs) and 15942 edges (interaction relationships, such as Pparg-Abca1 and miR-590-3p-AXIN2), was constructed, from which three significant modules were extracted and modules 2 and 3 contained interactions between miRNAs/TFs and DEGs such as miR-103-3p-AXIN2, miR-124-3p-AR-Tgfb1i1, and miR-27a-3p-PPARG-Abca1. miR-27a-3p was a known miRNA associated with OS. Abca1, AR, and miR-124-3p were hub genes in the miRNA-TF-mRNA network. Tgfb1i1 was involved in cell proliferation, Abca1 participated in the cholesterol metabolic process, and AXIN2 was associated with the canonical Wnt signaling pathway. Furthermore, we also confirmed upregulation of miR-590-3p and downregulation of AXIN2 in the mouse OS cell line K7M2-WT transfected with PTHR1 shRNA. In conclusion, PTHR1 may play important roles in progression of OS by activating miR-124-3p-AR-Tgfb1i1, miR-27a-3p-PPARG-Abca1, and miR-103/590-3p-AXIN2 axes.
Asunto(s)
Neoplasias Óseas , Osteosarcoma , Receptor de Hormona Paratiroídea Tipo 1/fisiología , Transportador 1 de Casete de Unión a ATP/genética , Transportador 1 de Casete de Unión a ATP/fisiología , Animales , Proteína Axina/genética , Proteína Axina/fisiología , Neoplasias Óseas/genética , Neoplasias Óseas/patología , Proteínas del Citoesqueleto/genética , Proteínas del Citoesqueleto/fisiología , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/fisiología , Progresión de la Enfermedad , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Redes Reguladoras de Genes , Células HEK293 , Humanos , Proteínas con Dominio LIM/genética , Proteínas con Dominio LIM/fisiología , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , MicroARNs/genética , MicroARNs/fisiología , Osteosarcoma/genética , Osteosarcoma/patología , PPAR gamma/genética , PPAR gamma/fisiología , Receptor de Hormona Paratiroídea Tipo 1/genética , Receptores Androgénicos/genética , Receptores Androgénicos/fisiología , Transducción de Señal/genética , Células Tumorales CultivadasRESUMEN
OBJECTIVE: The role of hepatocyte Abca1 (ATP binding cassette transporter A1) in trafficking hepatic free cholesterol (FC) into plasma versus bile for reverse cholesterol transport (RCT) is poorly understood. We hypothesized that hepatocyte Abca1 recycles plasma HDL-C (high-density lipoprotein cholesterol) taken up by the liver back into plasma, maintaining the plasma HDL-C pool, and decreasing HDL-mediated RCT into feces. Approach and Results: Chow-fed hepatocyte-specific Abca1 knockout (HSKO) and control mice were injected with human HDL radiolabeled with 125I-tyramine cellobiose (125I-TC; protein) and 3H-cholesteryl oleate (3H-CO). 125I-TC and 3H-CO plasma decay, plasma HDL 3H-CO selective clearance (ie, 3H-125I fractional catabolic rate), liver radiolabel uptake, and fecal 3H-sterol were significantly greater in HSKO versus control mice, supporting increased plasma HDL RCT. Twenty-four hours after 3H-CO-HDL injection, HSKO mice had reduced total hepatic 3H-FC (ie, 3H-CO hydrolyzed to 3H-FC in liver) resecretion into plasma, demonstrating Abca1 recycled HDL-derived hepatic 3H-FC back into plasma. Despite similar liver LDLr (low-density lipoprotein receptor) expression between genotypes, HSKO mice treated with LDLr-targeting versus control antisense oligonucleotide had slower plasma 3H-CO-HDL decay, reduced selective 3H-CO clearance, and decreased fecal 3H-sterol excretion that was indistinguishable from control mice. Increased RCT in HSKO mice was selective for 3H-CO-HDL, since macrophage RCT was similar between genotypes. CONCLUSIONS: Hepatocyte Abca1 deletion unmasks a novel and selective FC trafficking pathway that requires LDLr expression, accelerating plasma HDL-selective CE uptake by the liver and promoting HDL RCT into feces, consequently reducing HDL-derived hepatic FC recycling into plasma.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/fisiología , Colesterol/metabolismo , Hepatocitos/fisiología , Lipoproteínas HDL/sangre , Receptores de LDL/fisiología , Animales , Transporte Biológico , Femenino , Macrófagos/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
CONTEXT: Elevated serum lipoprotein(a) [Lp(a)] levels are associated with increased cardiovascular disease risk. ABCA1-mediated cholesterol efflux from macrophages may be an antiatherogenic process. Plasminogen (PLG) is a driver of ABCA1-mediated cholesterol efflux, and its action is inhibited by purified human Lp(a). OBJECTIVE: To determine the effects of Lp(a) in human serum on ABCA1 cholesterol efflux. METHODS: Cholesterol efflux capacity (CEC) was measured with two different cell-culture models using serum from 76 patients with either low (<50 mg/dL) or high (>50 mg/dL) Lp(a) levels. RESULTS: Using cAMP-stimulated J774 macrophages or baby hamster kidney fibroblasts overexpressing human ABCA1, we show that CEC was lower in patients with high Lp(a) levels compared with patients with low levels (-30.6%, P = 0.002 vs -24.1%, P < 0.001, respectively). Total-serum CEC negatively correlated with Lp(a) levels (r = -0.433, P = 0.0007 vs r = -0.505, P = 0.0011, respectively). These negative associations persisted after adjusting for serum cholesterol, age, sex, and statin use in a multiple linear regression model (adjusted R2 = 0.413 or 0.405, respectively) and were strengthened when further adjusting for the interaction between Lp(a) and PLG levels (adjusted R2 = 0.465 and 0.409, respectively). Total-serum and isolated Lp(a) from patients with high Lp(a) inhibited PLG-mediated ABCA1 cholesterol efflux. CONCLUSION: Total-serum CEC is reduced in patients with high Lp(a) levels. This is in part due to the inhibition of PLG-mediated ABCA1 cholesterol efflux by Lp(a). Our findings suggest an atherogenic role for Lp(a) through its ability to inhibit CEC.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/fisiología , Colesterol/metabolismo , Lipoproteína(a)/sangre , Lipoproteína(a)/farmacología , Transportador 1 de Casete de Unión a ATP/metabolismo , Adulto , Animales , Transporte Biológico/efectos de los fármacos , Células Cultivadas , Cricetinae , Femenino , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Humanos , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Masculino , Ratones , Persona de Mediana EdadRESUMEN
Proteins differential expression in type 2 diabetes mellitus (T2DM) can be due to etiological factors or pathological changes, such proteins can be utilized as biomarkers. Identification of a marker protein out of thousands became a feasible task during the proteomics era by using liquid chromatography-tandem mass spectrometry (LC-MS/MS). In this study, blood samples were obtained from 80 Bahraini subjects with and without T2DM, a subset was used for proteomic analysis by LC-MS/MS, while all samples were used for ELISA analysis of 3 proteins, TATA-box binding protein-associated factor RNA polymerase-1-C (TAF1C), ceruloplasmin (CERP) and fibronectin (FN). The former 2 proteins were selected from the T2DM-protein-panel identified by LC-MS/MS, and the latter was analyzed for validation of the setting. The main findings of the proteomic analysis are i. Identifications of 62 differentially expressed proteins in T2DM, ii. Upregulation of 71% of the identified proteins. While the ELISA analysis showed that; both TAF1C and FN were significantly increased in T2DM (P0.015 and P0.001, respectively), while CERP was not (P0.088). Logistic regression analysis: i. confirmed the above associations after correction for covariates, ii. Revealed an interaction between age and gender that affect the association of the proteins with T2DM. In conclusion, knowing that TAF1C is a prerequisite in ribosomal biogenesis, our ELISA results are suggestive of increased protein synthesis in T2DM, explaining the observed upregulation of the proteins identified by LC-MSMS. The association between T2DM and TAF1C is a novel finding that might open a new avenue in DM research.
Asunto(s)
Diabetes Mellitus Tipo 2/genética , Proteómica/métodos , Factores Asociados con la Proteína de Unión a TATA/genética , Factor de Transcripción TFIID/genética , Transportador 1 de Casete de Unión a ATP/genética , Transportador 1 de Casete de Unión a ATP/fisiología , Adulto , Biomarcadores , Cromatografía Liquida/métodos , Diabetes Mellitus Tipo 2/fisiopatología , Femenino , Humanos , Masculino , Espectrometría de Masas/métodos , Persona de Mediana Edad , Péptidos , TATA Box/genética , TATA Box/fisiología , Factores Asociados con la Proteína de Unión a TATA/fisiología , Factor de Transcripción TFIID/fisiologíaRESUMEN
Atherosclerosis (AS) is a chronic inflammatory disease of the arterial wall. Macrophages are considered to be closely associated with the development and progression of AS. However, the precise mechanism of miR-17-5p in the macrophages under AS remains incompletely clarified. This study investigated the regulatory effect of miR-17-5p on the inflammation and lipid accumulation in mouse macrophages both in vivo and in vitro. It was found that miR-17-5p was highly expressed with lowered ATP-binding cassette transporterA1 (ABCA1) level in the peripheral blood leucocytes (PBLs) of AS patients. Moreover, the level of miR-17-5p was up-regulated in the macrophages of ApoE-/- mice fed with a high-cholesterol diet. Furthermore, we injected miR-17-5p antagomir into AS mice or transfected miR-17-5p inhibitors into mouse macrophage RAW264.7 cells. Results showed that downregulation of miR-17-5p significantly reduced the production of inflammatory cytokines, inhibited the lipid accumulation and up-regulated ABCA1, and activated peroxisome proliferator-activated receptor (PPAR) γ/Liver X receptor (LXR) α signaling pathway. Additionally, ABCA1 was found to be a target of miR-17-5p by directly binding to 3'-untranslated region (3'-UTR) of its mRNA. Our study indicates a novel regulatory mechanism for miR-17-5p by interacting with ABCA1, which could be a therapy-target for the treatment of AS.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/genética , Transportador 1 de Casete de Unión a ATP/metabolismo , Aterosclerosis/genética , Aterosclerosis/metabolismo , Regulación hacia Abajo , Expresión Génica , Metabolismo de los Lípidos/genética , Macrófagos/metabolismo , MicroARNs/fisiología , Transportador 1 de Casete de Unión a ATP/fisiología , Animales , Aterosclerosis/terapia , Citocinas/metabolismo , Femenino , Humanos , Inflamación , Mediadores de Inflamación/metabolismo , Leucocitos/metabolismo , Receptores X del Hígado/metabolismo , Masculino , Ratones , Ratones Transgénicos , MicroARNs/genética , MicroARNs/metabolismo , Terapia Molecular Dirigida , PPAR gamma/metabolismo , Células RAW 264.7 , Regulación hacia Arriba/genéticaRESUMEN
CONTEXT: Low testosterone levels are associated with an increased incidence of cardiovascular (CV) events, but the underlying biochemical mechanisms are not fully understood. The clinical condition of hypogonadism offers a unique model to unravel the possible role of lipoprotein-associated abnormalities in CV risk. In particular, the assessment of the functional capacities of high-density lipoproteins (HDLs) may provide insights besides traditional risk factors. DESIGN: To determine whether reduced testosterone levels correlate with lipoprotein function, HDL cholesterol (HDL-C) efflux capacity (CEC) and serum cholesterol loading capacity (CLC). PARTICIPANTS: Genetic and idiopathic hypogonadal patients (n = 20) and control subjects (n = 17). RESULTS: Primary and secondary hypogonadal patients presented with lower HDL ATP-binding cassette transporter A1 (ABCA1)-, ATP-binding cassette transporter G1 (ABCG1)-, and aqueous diffusion-mediated CEC (-19.6%, -40.9%, and -12.9%, respectively), with a 16.2% decrement of total CEC. In the whole series, positive correlations between testosterone levels and both total HDL CEC (r2 = 0.359, P = 0.0001) and ABCG1 HDL CEC (r2 = 0.367, P = 0.0001) were observed. Conversely, serum CLC was markedly raised (+43%) in hypogonadals, increased, to a higher extent, in primary vs secondary hypogonadism (18.45 ± 2.78 vs 15.15 ± 2.10 µg cholesterol/mg protein) and inversely correlated with testosterone levels (r2 = 0.270, P = 0.001). HDL-C concentrations did not correlate with either testosterone levels, HDL CEC (total, ABCG1, and ABCA1) or serum CLC. CONCLUSIONS: In hypogonadal patients, proatherogenic lipoprotein-associated changes are associated with lower cholesterol efflux and increased influx, thus offering an explanation for a potentially increased CV risk.
Asunto(s)
Enfermedades Cardiovasculares/etiología , HDL-Colesterol/fisiología , Hipogonadismo/metabolismo , Transportador 1 de Casete de Unión a ATP/fisiología , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/fisiología , Adulto , Colesterol/metabolismo , HDL-Colesterol/sangre , Humanos , Hipogonadismo/complicaciones , Masculino , Persona de Mediana Edad , Testosterona/sangreRESUMEN
RATIONALE: Inhibition of miR-33 reduces atherosclerotic plaque burden, but miR-33 deficient mice are predisposed to the development of obesity and metabolic dysfunction. The proatherogenic effects of miR-33 are thought to be in large part because of its repression of macrophage cholesterol efflux, through targeting of Abca1 (ATP-binding cassette subfamily A member 1). However, targeting of other factors may also be required for the beneficial effects of miR-33, and currently available approaches have not allowed researchers to determine the specific impact of individual miRNA target interactions in vivo. OBJECTIVE: In this work, we sought to determine how specific disruption of Abca1 targeting by miR-33 impacts macrophage cholesterol efflux and atherosclerotic plaque formation in vivo. METHODS AND RESULTS: We have generated a novel mouse model with specific point mutations in the miR-33 binding sites of the Abca1 3'untranslated region, which prevents targeting by miR-33. Abca1 binding site mutant ( Abca1BSM) mice had increased hepatic ABCA1 expression but did not show any differences in body weight or metabolic function after high fat diet feeding. Macrophages from Abca1BSM mice also had increased ABCA1 expression, as well as enhanced cholesterol efflux and reduced foam cell formation. Moreover, LDLR (low-density lipoprotein receptor) deficient animals transplanted with bone marrow from Abca1BSM mice had reduced atherosclerotic plaque formation, similar to mice transplanted with bone marrow from miR-33 knockout mice. CONCLUSION: Although the more pronounced phenotype of miR-33 deficient animals suggests that other targets may also play an important role, our data clearly demonstrate that repression of ABCA1 is primarily responsible for the proatherogenic effects of miR-33. This work shows for the first time that disruption of a single miRNA/target interaction can be sufficient to mimic the effects of miRNA deficiency on complex physiological phenotypes in vivo and provides an approach by which to assess the impact of individual miRNA targets.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/fisiología , Colesterol/metabolismo , Macrófagos/metabolismo , MicroARNs/fisiología , Placa Aterosclerótica/etiología , Transportador 1 de Casete de Unión a ATP/genética , Animales , Sitios de Unión , Ratones , Ratones Noqueados , Receptores de LDL/fisiologíaRESUMEN
BACKGROUND: Cholesterol retention within plasma membranes of macrophages is associated with increased inflammatory signaling. Cholesterol efflux via the transporters ABCA1, ABCG1, and SR-BI to high-density lipoprotein (HDL) particles is a critical mechanism to maintain cellular cholesterol homeostasis. Little is known about the impact of the obese microenvironment on cholesterol efflux capacity (CEC) of macrophages. In this study, the CEC of obese-derived primary adipose-tissue macrophages (ATM) is evaluated and the in vivo microenvironment is modeled in vitro to determine mechanisms underlying modulated CEC. MATERIALS AND METHODS: F4/80+ ATM are labeled with 3 H-cholesterol ex vivo, and CEC and ABCA1/ABCG1 protein levels are determined. Total, ABCA1-dependent, and ABCA1-independent CECs are determined in J774 macrophages polarized to M1 (LPS&IFNγ), M2 (IL-4&IL-13), or metabolic phenotypes (glucose, insulin, and palmitic acid). RESULTS: Obese ATM exhibit enhanced CEC and ABCA1 and ABCG1 expression compared to lean ATM. In contrast, ABCA1-CEC is suppressed from M1 polarized macrophages compared to untreated in vitro, by activation of the JAK/STAT pathway. Incubation of macrophages in vitro in high glucose augments cAMP-induced ABCA1 protein expression and ABCA1-CEC. CONCLUSIONS: These novel findings demonstrate remarkable plasticity of macrophages to respond to their environment with specific modulation of ABCA1 depending on whether classical pro-inflammatory or metabolic cues predominate.
Asunto(s)
Tejido Adiposo/metabolismo , Colesterol/metabolismo , Macrófagos/metabolismo , Obesidad/metabolismo , Transportador 1 de Casete de Unión a ATP/fisiología , Tejido Adiposo/citología , Animales , Células Cultivadas , Señales (Psicología) , Quinasas Janus/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Factores de Transcripción STAT/fisiologíaRESUMEN
Arsenic is one of the most widespread global environmental toxicants associated with endemic poisoning. ATP-binding cassette (ABC) proteins are transmembrane channels that transport and dispose of lipids and metabolic products across the plasma membrane. The majority of ABC family members (including ABCB1 and ABCC1) are reported to play a role in the development of arsenic and drug resistance in mammals. Previously, we established a human arsenic-resistant ECV-304 (AsRE) cell line and identified ABCA1 as a novel arsenic resistance gene. In the current study, we further investigated the potential contribution of ABCA1, ABCB1, and ABCC1 to arsenic resistance through measurement of survival rates and arsenic accumulation in AsRE cells with RNA interference. The arsenic resistance capacity of ABCC1 was the strongest among the three genes, while those of ABCA1 and ABCB1 were similar. Double or triple gene knockdown of ABCA1, ABCB1, and ABCC1 via RNA interference led to a decrease significant in arsenic resistance when ABCA1/ABCB1 or ABCB1/ABCC1 were simultaneously silenced. Interestingly, no differences were evident between cells with ABCA1/ABCC1 and ABCC1 only knockdown. Our findings suggest that ABCA1 and ABCB1 proteins display similar arsenic resistance capabilities and possibly coordinate to promote arsenic resistance in AsRE cells.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/fisiología , Arsénico/farmacología , Resistencia a Medicamentos/genética , Transportador 1 de Casete de Unión a ATP/genética , Subfamilia B de Transportador de Casetes de Unión a ATP/genética , Subfamilia B de Transportador de Casetes de Unión a ATP/fisiología , Arsénico/metabolismo , Transporte Biológico , Línea Celular , Técnicas de Silenciamiento del Gen , Humanos , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Interferencia de ARNRESUMEN
Background: Cholesterol efflux efficiency, reactive oxygen species, and inflammation are closely related to cardiovascular diseases. Our aim was to investigate the effect of propofol on cholesterol-loaded rat aortic endothelial cells after high-density lipoprotein treatment in vitro. Methods and Results: The results showed that propofol promoted cholesterol efflux and ameliorated inflammation and reactive oxygen species overproduction according to the analysis of p65 nuclear translocation and a 2',7'-dichlorofluorescin diacetate assay, respectively. Conclusions: These results provide a possible explanation for the anti-inflammatory, antioxidant, and cholesterol efflux-promoting effects of propofol on rat aortic endothelial cells after incubation with high-density lipoprotein.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/fisiología , Colesterol/metabolismo , Hipnóticos y Sedantes/farmacología , Inflamación , Propofol/farmacología , Especies Reactivas de Oxígeno , Transportador 1 de Casete de Unión a ATP/metabolismo , Transportadoras de Casetes de Unión a ATP , Animales , Aorta , Células Endoteliales , RatasRESUMEN
SCOPE: Chocolate consumption lowers cardiovascular disease risk, which might be attributed to the methylxanthine theobromine. These effects may be mediated through effects on HDL-mediated cholesterol efflux, which may be affected by microRNA (miRNA) levels in the HDL particles. Therefore, the aim of this study is to investigate effects of theobromine consumption on fasting and postprandial cholesterol efflux and miRNAs levels. METHODS AND RESULTS: Thirty overweight and 14 obese healthy men and women participated in this randomized, double-blind crossover study. Participants consumed 500 mg d-1 of theobromine or placebo for 4 weeks. ABCA1-mediated cholesterol efflux was measured using J774 macrophages. MiRNAs levels (miR-92a, miR-223, miR-135a*) were quantified in apolipoprotein B-depleted serum. Theobromine consumption did not affect fasting and postprandial cholesterol efflux. Fasting miR-223 and miR-135a levels were unchanged, while miR-92a levels were decreased (-0.21; p < 0.05). The high-fat meal increased postprandial cholesterol efflux capacity (+4.3 percentage points; p ≤ 0.001), miR-92a (+1.21; p < 0.001), and miR-223 (+1.79; p < 0.001) levels, while a trend was found for miR-135a (+1.08; p = 0.06). CONCLUSION: Theobromine did not improve fasting and postprandial ABCA1-mediated cholesterol efflux capacity, but decreased fasting miR-92a levels. High-fat meal intake increased postprandial cholesterol efflux and the three selected miRNAs levels.
Asunto(s)
HDL-Colesterol/metabolismo , Ayuno/metabolismo , MicroARNs/sangre , Periodo Posprandial , Teobromina/administración & dosificación , Transportador 1 de Casete de Unión a ATP/fisiología , Anciano , Animales , Estudios Cruzados , Método Doble Ciego , Femenino , Humanos , Masculino , Ratones , Persona de Mediana EdadRESUMEN
High levels of circulating TNF and its receptors, TNFR1 and TNFR2, predict the progression of diabetic kidney disease (DKD), but their contribution to organ damage in DKD remains largely unknown. Here, we investigated the function of local and systemic TNF in podocyte injury. We cultured human podocytes with sera collected from DKD patients, who displayed elevated TNF levels, and focal segmental glomerulosclerosis (FSGS) patients, whose TNF levels resembled those of healthy patients. Exogenous TNF administration or local TNF expression was equally sufficient to cause free cholesterol-dependent apoptosis in podocytes by acting through a dual mechanism that required a reduction in ATP-binding cassette transporter A1-mediated (ABCA1-mediated) cholesterol efflux and reduced cholesterol esterification by sterol-O-acyltransferase 1 (SOAT1). TNF-induced albuminuria was aggravated in mice with podocyte-specific ABCA1 deficiency and was partially prevented by cholesterol depletion with cyclodextrin. TNF-stimulated free cholesterol-dependent apoptosis in podocytes was mediated by nuclear factor of activated T cells 1 (NFATc1). ABCA1 overexpression or cholesterol depletion was sufficient to reduce albuminuria in mice with podocyte-specific NFATc1 activation. Our data implicate an NFATc1/ABCA1-dependent mechanism in which local TNF is sufficient to cause free cholesterol-dependent podocyte injury irrespective of TNF, TNFR1, or TNFR2 serum levels.
Asunto(s)
Colesterol/química , Nefropatías Diabéticas/sangre , Glomeruloesclerosis Focal y Segmentaria/sangre , Factores de Transcripción NFATC/fisiología , Síndrome Nefrótico/sangre , Factor de Necrosis Tumoral alfa/fisiología , Transportador 1 de Casete de Unión a ATP/fisiología , Adolescente , Albuminuria/sangre , Animales , Apoptosis , Biopsia , Estudios de Casos y Controles , Niño , Preescolar , Ciclodextrinas/metabolismo , Femenino , Regulación de la Expresión Génica , Tasa de Filtración Glomerular , Humanos , Inflamación , Riñón/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Podocitos/metabolismo , Receptores Tipo I de Factores de Necrosis Tumoral/sangre , Receptores Tipo II del Factor de Necrosis Tumoral/sangre , Esterol O-Aciltransferasa/fisiología , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
Spontaneous preterm delivery (SPTD) with gestational age between 28 and 37 complete weeks was reported to have a genetic predisposition in lipids metabolism. This study aimed to investigate the association between the lipid levels and gene polymorphisms of ABCA1 (rs2422493), APOE (rs7412) and HMGCR (rs12916) in Chinese pregnant women with SPTD. A case-control study was conducted at the baseline randomization in 200 SPTD and 178 healthy full term delivery (FTD) women. Maternal blood lipids were detected close to delivery of fetus in SPTD group and in FTD group with gestational age-matched. Cord blood lipids were detected after delivery in two groups. Three genotypes both in maternal and cord blood were determined by real time PCR. The results showed that the levels of total cholesterol (TCHO), triglyceride (TG), high density lipoprotein (HDL), and low-density lipoprotein cholesterol (LDL) in the maternal blood in the SPTD group were significantly lower than those in the FTD group, while the levels of TCHO, HDL, and LDL in the cord blood in the SPTD group were significantly higher than those in the FTD group. In the SPTD subjects, the levels of TG and LDL in the maternal blood were associated with different genotypes of HMGCR gene rs12916 loci. These results indicate that abnormal lipid metabolism may exist in SPTD women and the premature fetus and the HMGCR gene may be a susceptible gene for SPTD.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/genética , Apolipoproteínas E/genética , Hidroximetilglutaril-CoA Reductasas/genética , Lípidos/sangre , Polimorfismo de Nucleótido Simple/fisiología , Nacimiento Prematuro/genética , Transportador 1 de Casete de Unión a ATP/fisiología , Adulto , Apolipoproteínas E/fisiología , Colesterol/sangre , LDL-Colesterol/sangre , Femenino , Sangre Fetal/química , Predisposición Genética a la Enfermedad , Humanos , Hidroximetilglutaril-CoA Reductasas/fisiología , Lipoproteínas HDL/sangre , Polimorfismo de Nucleótido Simple/genética , Embarazo , Nacimiento Prematuro/sangre , Triglicéridos/sangre , Adulto JovenRESUMEN
RATIONALE: An efficient and reproducible source of genotype-specific human macrophages is essential for study of human macrophage biology and related diseases. OBJECTIVE: To perform integrated functional and transcriptome analyses of human induced pluripotent stem cell-derived macrophages (IPSDMs) and their isogenic human peripheral blood mononuclear cell-derived macrophage (HMDM) counterparts and assess the application of IPSDM in modeling macrophage polarization and Mendelian disease. METHODS AND RESULTS: We developed an efficient protocol for differentiation of IPSDM, which expressed macrophage-specific markers and took up modified lipoproteins in a similar manner to HMDM. Like HMDM, IPSDM revealed reduction in phagocytosis, increase in cholesterol efflux capacity and characteristic secretion of inflammatory cytokines in response to M1 (lipopolysaccharide+interferon-γ) activation. RNA-Seq revealed that nonpolarized (M0) as well as M1 or M2 (interleukin-4) polarized IPSDM shared transcriptomic profiles with their isogenic HMDM counterparts while also revealing novel markers of macrophage polarization. Relative to IPSDM and HMDM of control individuals, patterns of defective cholesterol efflux to apolipoprotein A-I and high-density lipoprotein-3 were qualitatively and quantitatively similar in IPSDM and HMDM of patients with Tangier disease, an autosomal recessive disorder because of mutations in ATP-binding cassette transporter AI. Tangier disease-IPSDM also revealed novel defects of enhanced proinflammatory response to lipopolysaccharide stimulus. CONCLUSIONS: Our protocol-derived IPSDM are comparable with HMDM at phenotypic, functional, and transcriptomic levels. Tangier disease-IPSDM recapitulated hallmark features observed in HMDM and revealed novel inflammatory phenotypes. IPSDMs provide a powerful tool for study of macrophage-specific function in human genetic disorders as well as molecular studies of human macrophage activation and polarization.