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1.
J Lipid Res ; 42(9): 1339-45, 2001 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-11518753

RESUMO

The discovery of the role of the ATP-binding cassette transporter A1 (ABCA1) in mediating apolipoprotein A-I-mediated efflux has led to a dramatic increase in our knowledge of the molecular mechanisms involved in cholesterol efflux and cellular metabolism. In this review, we discuss several aspects of ABCA1 regulation including i) transcriptional regulation, ii) substrate specificity and availability, iii) accessory proteins, iv) acceptor specificity and availability, and v) protein trafficking. The majority of studies of ABCA1 regulation to date have focused on the identification of promoter elements that determine ABCA1 gene transcription. Here we also review the potential functional role of ABCA1 in reverse cholesterol transport. Given the key role that ABCA1 plays in cholesterol homeostasis, it is likely that there are multiple mechanisms for controlling the overall transporter activity of ABCA1.


Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/fisiologia , Regulação da Expressão Gênica , Transportador 1 de Cassete de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/análise , Animais , Apolipoproteína A-I/metabolismo , Colesterol/metabolismo , AMP Cíclico/farmacologia , Citocinas/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Regiões Promotoras Genéticas , Receptores Citoplasmáticos e Nucleares/agonistas , Esteróis/farmacologia , Fatores de Transcrição/agonistas , Transcrição Gênica
2.
J Clin Invest ; 108(2): 303-9, 2001 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-11457883

RESUMO

The discovery of the ABCA1 lipid transporter has generated interest in modulating human plasma HDL levels and atherogenic risk by enhancing ABCA1 gene expression. To determine if increased ABCA1 expression modulates HDL metabolism in vivo, we generated transgenic mice that overexpress human ABCA1 (hABCA1-Tg). Hepatic and macrophage expression of hABCA1 enhanced macrophage cholesterol efflux to apoA-I; increased plasma cholesterol, cholesteryl esters (CEs), free cholesterol, phospholipids, HDL cholesterol, and apoA-I and apoB levels; and led to the accumulation of apoE-rich HDL1. ABCA1 transgene expression delayed 125I-apoA-I catabolism in both liver and kidney, leading to increased plasma apoA-I levels, but had no effect on apoB secretion after infusion of Triton WR1339. Although the plasma clearance of HDL-CE was not significantly altered in hABCA1-Tg mice, the net hepatic delivery of exogenous 3H-CEt-HDL, which is dependent on the HDL pool size, was increased 1.5-fold. In addition, the cholesterol and phospholipid concentrations in hABCA1-Tg bile were increased 1.8-fold. These studies show that steady-state overexpression of ABCA1 in vivo (a) raises plasma apoB levels without altering apoB secretion and (b) raises plasma HDL-C and apoA-I levels, facilitating hepatic reverse cholesterol transport and biliary cholesterol excretion. Similar metabolic changes may modify atherogenic risk in humans.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Bile/metabolismo , Colesterol/metabolismo , Hiperlipoproteinemias/etiologia , Transportador 1 de Cassete de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/biossíntese , Animais , Apolipoproteínas/sangue , Bile/química , Colesterol/análise , Regulação da Expressão Gênica , Humanos , Hiperlipoproteinemias/sangue , Hiperlipoproteinemias/metabolismo , Lipídeos/sangue , Lipoproteínas HDL/sangue , Lipoproteínas HDL/metabolismo , Lipoproteínas VLDL/metabolismo , Fígado/metabolismo , Macrófagos/metabolismo , Camundongos , Camundongos Transgênicos
3.
J Biol Chem ; 276(29): 27584-90, 2001 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-11349133

RESUMO

ABCA1, the ATP-binding cassette protein mutated in Tangier disease, mediates the efflux of excess cellular sterol to apoA-I and thereby the formation of high density lipoprotein. The intracellular localization and trafficking of ABCA1 was examined in stably and transiently transfected HeLa cells expressing a functional human ABCA1-green fluorescent protein (GFP) fusion protein. The fluorescent chimeric ABCA1 transporter was found to reside on the cell surface and on intracellular vesicles that include a novel subset of early endosomes, as well as late endosomes and lysosomes. Studies of the localization and trafficking of ABCA1-GFP in the presence of brefeldin A or monensin, agents known to block intracellular vesicular trafficking, as well as apoA-I-mediated cellular lipid efflux, showed that: (i) ABCA1 functions in lipid efflux at the cell surface, and (ii) delivery of ABCA1 to lysosomes for degradation may serve as a mechanism to modulate its surface expression. Time-lapse fluorescence microscopy revealed that ABCA1-GFP-containing early endosomes undergo fusion, fission, and tubulation and transiently interact with one another, late endocytic vesicles, and the cell surface. These studies establish a complex intracellular trafficking pathway for human ABCA1 that may play important roles in modulating ABCA1 transporter activity and cellular cholesterol homeostasis.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Transportador 1 de Cassete de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/genética , Animais , Apolipoproteína A-I/metabolismo , Transporte Biológico , Células CHO , Compartimento Celular , Cricetinae , Cicloeximida/farmacologia , Endocitose , Proteínas de Fluorescência Verde , Células HeLa , Humanos , Imuno-Histoquímica , Proteínas Luminescentes/genética , Monensin/farmacologia , Proteínas Recombinantes de Fusão/genética , Esteróis/metabolismo , Transfecção
4.
J Biol Chem ; 276(18): 15090-8, 2001 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-11278414

RESUMO

To evaluate the biochemical and molecular mechanisms leading to glomerulosclerosis and the variable development of atherosclerosis in patients with familial lecithin cholesterol acyl transferase (LCAT) deficiency, we generated LCAT knockout (KO) mice and cross-bred them with apolipoprotein (apo) E KO, low density lipoprotein receptor (LDLr) KO, and cholesteryl ester transfer protein transgenic mice. LCAT-KO mice had normochromic normocytic anemia with increased reticulocyte and target cell counts as well as decreased red blood cell osmotic fragility. A subset of LCAT-KO mice accumulated lipoprotein X and developed proteinuria and glomerulosclerosis characterized by mesangial cell proliferation, sclerosis, lipid accumulation, and deposition of electron dense material throughout the glomeruli. LCAT deficiency reduced the plasma high density lipoprotein (HDL) cholesterol (-70 to -94%) and non-HDL cholesterol (-48 to -85%) levels in control, apoE-KO, LDLr-KO, and cholesteryl ester transfer protein-Tg mice. Transcriptome and Western blot analysis demonstrated up-regulation of hepatic LDLr and apoE expression in LCAT-KO mice. Despite decreased HDL, aortic atherosclerosis was significantly reduced (-35% to -99%) in all mouse models with LCAT deficiency. Our studies indicate (i) that the plasma levels of apoB containing lipoproteins rather than HDL may determine the atherogenic risk of patients with hypoalphalipoproteinemia due to LCAT deficiency and (ii) a potential etiological role for lipoproteins X in the development of glomerulosclerosis in LCAT deficiency. The availability of LCAT-KO mice characterized by lipid, hematologic, and renal abnormalities similar to familial LCAT deficiency patients will permit future evaluation of LCAT gene transfer as a possible treatment for glomerulosclerosis in LCAT-deficient states.


Assuntos
Arteriosclerose/enzimologia , Glomerulosclerose Segmentar e Focal/enzimologia , Fosfatidilcolina-Esterol O-Aciltransferase/fisiologia , Animais , Arteriosclerose/fisiopatologia , Sequência de Bases , Primers do DNA , Glomerulosclerose Segmentar e Focal/fisiopatologia , Rim/fisiopatologia , Lipídeos/sangue , Lipoproteínas/sangue , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia Eletrônica , Fosfatidilcolina-Esterol O-Aciltransferase/sangue , Fosfatidilcolina-Esterol O-Aciltransferase/genética , RNA Mensageiro/genética
5.
Am J Med Sci ; 321(1): 66-75, 2001 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11202482

RESUMO

Erdheim-Chester disease (ECD) is a rare multisystem histiocytosis syndrome of unknown cause that usually affects adults. Histiocytic infiltration of multiple end organs produces bone pain, xanthelasma and xanthoma, exophthalmos, diabetes insipidus, and interstitial lung disease. Differential diagnosis includes Langerhans cell histiocytosis, metabolic disorders, malignancy, and sarcoidosis. ECD can be diagnosed using a combination of clinical and histopathologic findings. Sites of involvement include lung, bone, skin, retroorbital tissue, central nervous system, pituitary gland, retroperitoneum, and pericardium. Symmetrical long bone pain with associated osteosclerotic lesions, xanthomas around the eyelids, exophthalmos, and/or diabetes insipidus suggest ECD. Approximately 35% of patients have associated lung involvement, characterized by interstitial accumulations of histiocytic cells and fibrosis in a predominantly perilymphangitic and subpleural pattern. This pattern distinguishes ECD from other histiocytic disorders involving the lung. The diagnosis is confirmed by tissue biopsies that contain histiocytes with non-Langerhans cell features. In general, the clinical course of patients with this disease varies, and the prognosis can be poor despite treatment. Clinical trials for treatment of ECD have not been conducted and treatment is based on anecdotal experience.


Assuntos
Histiocitose de Células não Langerhans/fisiopatologia , Pneumopatias/fisiopatologia , Adulto , Diagnóstico Diferencial , Histiocitose de Células não Langerhans/classificação , Histiocitose de Células não Langerhans/diagnóstico , Histiocitose de Células não Langerhans/terapia , Humanos , Pulmão/patologia , Pneumopatias/classificação , Pneumopatias/diagnóstico , Pneumopatias/terapia , Insuficiência de Múltiplos Órgãos/etiologia , Prognóstico
6.
Biochem Biophys Res Commun ; 280(3): 818-23, 2001 Jan 26.
Artigo em Inglês | MEDLINE | ID: mdl-11162594

RESUMO

ABCAI, a member of the ATP binding cassette family, mediates the efflux of excess cellular lipid to HDL and is defective in Tangier disease. The apolipoprotein acceptor specificity for lipid efflux by ABCAI was examined in stably transfected Hela cells, expressing a human ABCAI-GFP fusion protein. ApoA-I and all of the other exchangeable apolipoproteins tested (apoA-II, apoA-IV, apoC-I, apoC-II, apoC-III, apoE) showed greater than a threefold increase in cholesterol and phospholipid efflux from ABCAI-GFP transfected cells compared to control cells. Expression of ABCAI in Hela cells also resulted in a marked increase in specific binding of both apoA-I (Kd = 0.60 microg/mL) and apoA-II (Kd = 0.58 microg/mL) to a common binding site. In summary, ABCAI-mediated cellular binding of apolipoproteins and lipid efflux is not specific for only apoA-I but can also occur with other apolipoproteins that contain multiple amphipathic helical domains.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Apolipoproteínas/metabolismo , Metabolismo dos Lipídeos , Transportador 1 de Cassete de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/genética , Apolipoproteína A-I/metabolismo , Apolipoproteína A-II/metabolismo , Transporte Biológico Ativo , Expressão Gênica , Proteínas de Fluorescência Verde , Células HeLa , Humanos , Cinética , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Transfecção
7.
Nat Med ; 7(1): 53-8, 2001 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11135616

RESUMO

Peroxisome proliferator-activated receptors (PPARs) are nuclear receptors that regulate lipid and glucose metabolism and cellular differentiation. PPAR-alpha and PPAR-gamma are both expressed in human macrophages where they exert anti-inflammatory effects. The activation of PPAR-alpha may promote foam-cell formation by inducing expression of the macrophage scavenger receptor CD36. This prompted us to investigate the influence of different PPAR-activators on cholesterol metabolism and foam-cell formation of human primary and THP-1 macrophages. Here we show that PPAR-alpha and PPAR-gamma activators do not influence acetylated low density lipoprotein-induced foam-cell formation of human macrophages. In contrast, PPAR-alpha and PPAR-gamma activators induce the expression of the gene encoding ABCA1, a transporter that controls apoAI-mediated cholesterol efflux from macrophages. These effects are likely due to enhanced expression of liver-x-receptor alpha, an oxysterol-activated nuclear receptor which induces ABCA1-promoter transcription. Moreover, PPAR-alpha and PPAR-gamma activators increase apoAI-induced cholesterol efflux from normal macrophages. In contrast, PPAR-alpha or PPAR-gamma activation does not influence cholesterol efflux from macrophages isolated from patients with Tangier disease, which is due to a genetic defect in ABCA1. Here we identify a regulatory role for PPAR-alpha and PPAR-gamma in the first steps of the reverse-cholesterol-transport pathway through the activation of ABCA1-mediated cholesterol efflux in human macrophages.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Colesterol/metabolismo , Células Espumosas/metabolismo , Receptores Citoplasmáticos e Nucleares/agonistas , Fatores de Transcrição/agonistas , Transportador 1 de Cassete de Ligação de ATP , Sequência de Bases , Transporte Biológico , Células Cultivadas , Primers do DNA , Humanos
8.
Proc Natl Acad Sci U S A ; 97(14): 7987-92, 2000 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-10884428

RESUMO

The ABCA1 gene, a member of the ATP-binding cassette A (ABCA1) transporter superfamily, encodes a membrane protein that facilitates the cellular efflux of cholesterol and phospholipids. Mutations in ABCA1 lead to familial high density lipoprotein deficiency and Tangier disease. We report the complete human ABCA1 gene sequence, including 1,453 bp of the promoter, 146,581 bp of introns and exons, and 1 kb of the 3' flanking region. The ABCA1 gene spans 149 kb and comprises 50 exons. Sixty-two repetitive Alu sequences were identified in introns 1-49. The transcription start site is 315 bp upstream of a newly identified initiation methionine codon and encodes an ORF of 6,783 bp. Thus, the ABCA1 protein is comprised of 2,261 aa. Analysis of the 1,453 bp 5' upstream of the transcriptional start site reveals multiple binding sites for transcription factors with roles in lipid metabolism. Comparative analysis of the mouse and human ABCA1 promoter sequences identified specific regulatory elements, which are evolutionarily conserved. The human ABCA1 promoter fragment -200 to -80 bp that contains binding motifs for SP1, SP3, E-box, and AP1 modulates cellular cholesterol and cAMP regulation of ABCA1 gene expression. These combined findings provide insights into ABCA1-mediated regulation of cellular cholesterol metabolism and will facilitate the identification of new pharmacologic agents for the treatment of atherosclerosis in humans.


Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Regiões Promotoras Genéticas , Transportador 1 de Cassete de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/metabolismo , Elementos Alu , Sequência de Aminoácidos , Animais , Sequência de Bases , Sítios de Ligação , Transporte Biológico , Colesterol/metabolismo , Clonagem Molecular , Humanos , Hipolipoproteinemias/genética , Camundongos , Dados de Sequência Molecular , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Especificidade da Espécie , Doença de Tangier/genética , Fatores de Transcrição
9.
Curr Opin Lipidol ; 11(3): 267-75, 2000 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-10882342

RESUMO

In the past several years significant advances have been made in our understanding of lecithin-cholesterol acyltransferase (LCAT) function. LCAT beneficially alters the plasma concentrations of apolipoprotein B-containing lipoproteins, as well as HDL. In addition, its proposed role in facilitating reverse cholesterol transport and modulating atherosclerosis has been demonstrated in vivo. Analysis of LCAT transgenic animals has established the importance of evaluating HDL function, as well as HDL plasma levels, to predict atherogenic risk.


Assuntos
Arteriosclerose/sangue , Colesterol/sangue , Lipoproteínas/sangue , Fosfatidilcolina-Esterol O-Aciltransferase/metabolismo , Animais , Transporte Biológico , Técnicas de Transferência de Genes , Glomerulosclerose Segmentar e Focal/enzimologia , Humanos , Fosfatidilcolina-Esterol O-Aciltransferase/sangue , Fosfatidilcolina-Esterol O-Aciltransferase/genética
10.
J Lipid Res ; 41(5): 667-72, 2000 May.
Artigo em Inglês | MEDLINE | ID: mdl-10787427

RESUMO

Recent in vitro studies have provided evidence that hepatic lipase (HL) facilitates the selective uptake of HDL cholesteryl esters (CE), but the in vivo physiological relevance of this process has not been demonstrated. To evaluate the role that HL plays in facilitating the selective uptake of HDL-CE in vivo, we studied the metabolism of [(3)H]CEt, (125)I-labeled apolipoprotein (apo) A-I, and (131)I-labeled apoA-II-labeled HDL in HL-deficient mice. Kinetic analysis revealed similar catabolism of (125)I-labeled apoA-I (as well as (131)I-labeled apoA-II) in C57BL controls and HL deficient mice, with fractional catabolic rates (FCR) of 2.17 +/- 0.15 and 2.16 +/- 0.11 d(-)(1) (2.59 +/- 0.14 and 2.67 +/- 0.13 d(-)(1), respectively). In contrast, despite similar hepatic scavenger receptor BI expression, HL-deficient mice had delayed clearance of [(3)H]CEt compared to controls (FCR = 3.66 +/- 0.29 and 4.41 +/- 0.18 d(-)(1), P < 0.05). The hepatic accumulation of [(3)H]CEt in HL-deficient mice (62.3 +/- 2.1% of total) was significantly less than in controls (72.7 +/- 3.0%), while the [(3)H]CEt remaining in the plasma compartment increased (20.7 +/- 1.8% and 12.6 +/- 0.5%) (P < 0.05, all). In summary, HL deficiency does not alter the catabolism of apoA-I and apoA-II but decreases the hepatic uptake and the plasma clearance of HDL-CE. These data establish for the first time an important role for HL in facilitating the selective uptake of HDL-CE in vivo.


Assuntos
Ésteres do Colesterol/sangue , HDL-Colesterol/sangue , Lipase/deficiência , Fígado/enzimologia , Animais , Apolipoproteína A-I/sangue , Apolipoproteína A-II/sangue , Transporte Biológico Ativo , Feminino , Cinética , Lipase/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout
12.
Arterioscler Thromb Vasc Biol ; 20(3): 793-800, 2000 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-10712405

RESUMO

To investigate the in vivo role that hepatic lipase (HL) plays in HDL metabolism independently of its lipolytic function, recombinant adenovirus (rAdV) expressing native HL, catalytically inactive HL (HL-145G), and luciferase control was injected in HL-deficient mice. At day 4 after infusion of 2 x 10(8) plaque-forming units of rHL-AdV and rHL-145G-AdV, similar plasma concentrations were detected in postheparin plasma (HL=8.4+/-0.8 microg/mL and HL-145G=8.3+/-0.8 microg/mL). Mice expressing HL had significant reductions of cholesterol (-76%), phospholipids (PL; -68%), HDL cholesterol (-79%), apolipoprotein (apo) A-I (-45%), and apoA-II (-59%; P<0.05 for all), whereas mice expressing HL-145G decreased their cholesterol (-49%), PL (-40%), HDL cholesterol (-42%), and apoA-II (-89%; P<0.005 for all) but had no changes in apoA-I. The plasma kinetics of (125)I-labeled apoA-I HDL, (131)I-labeled apoA-II HDL, and [(3)H]cholesteryl ester (CE) HDL revealed that compared with mice expressing luciferase control (fractional catabolic rate [FCR] in d(-1): apoA-I HDL=1.3+/-0.1; apoA-II HDL=2.1+/-0; CE HDL=4.1+/-0.7), both HL and HL-145G enhanced the plasma clearance of CEs and apoA-II present in HDL (apoA-II HDL=5.6+/-0.5 and 4.4+/-0.2; CE HDL=9.3+/-0. 0 and 8.3+/-1.1, respectively), whereas the clearance of apoA-I HDL was enhanced in mice expressing HL (FCR=4.6+/-0.3) but not HL-145G (FCR=1.4+/-0.4). These combined findings demonstrate that both lipolytic and nonlipolytic functions of HL are important for HDL metabolism in vivo. Our study provides, for the first time, in vivo evidence for a role of HL in HDL metabolism independent of lipolysis and provides new insights into the role of HL in facilitating distinct metabolic pathways involved in the catabolism of apoA-I- versus apoA-II-containing HDL.


Assuntos
HDL-Colesterol/metabolismo , Lipase/genética , Lipase/metabolismo , Lipólise/fisiologia , Fígado/enzimologia , Adenoviridae/genética , Animais , Apolipoproteína A-I/metabolismo , Linhagem Celular , Genes Reporter , Humanos , Radioisótopos do Iodo , Rim/citologia , Luciferases/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Recombinantes de Fusão/genética , Transfecção , Trítio
13.
Arterioscler Thromb Vasc Biol ; 20(2): 450-8, 2000 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-10669643

RESUMO

Elevated low density lipoprotein cholesterol (LDL-C) and reduced high density lipoprotein cholesterol (HDL-C) concentrations are independent risk factors for coronary heart disease. We have previously demonstrated that overexpression of an enzyme with a well established role in HDL metabolism, lecithin:cholesterol acyltransferase (LCAT), in New Zealand White rabbits not only raises HDL-C concentrations but reduces those of LDL-C as well, ultimately preventing diet-induced atherosclerosis. In the present study, the human LCAT gene (hLCAT) was introduced into LDL receptor (LDLr)-deficient (Watanabe heritable hyperlipidemic) rabbits to (1) investigate the role of the LDLr pathway in the hLCAT-mediated reductions of LDL-C and (2) determine the influence of hLCAT overexpression on atherosclerosis susceptibility in an animal model of familial hypercholesterolemia. Heterozygosity or homozygosity for the LDLr defect was determined by polymerase chain reaction, and 3 groups of hLCAT-transgenic (hLCAT+) rabbits that differed in LDLr status were established: (1) LDLr wild-type (LDLr+/+), (2) LDLr heterozygotes (LDLr+/-), and (3) LDLr homozygotes (LDLr-/-). Data for hLCAT+ rabbits were compared with those of nontransgenic (hLCAT-) rabbits of the same LDLr status. Plasma HDL-C concentrations were significantly elevated in the hLCAT+ animals of each LDLr status. However, LDL-C levels were significantly reduced only in hLCAT+/LDLr+/+ and hLCAT+/LDLr+/- rabbits but not in hLCAT+/LDLr-/- rabbits (405+/-14 versus 392+/-31 mg/dL). Metabolic studies revealed that the fractional catabolic rate (FCR, d(-1)) of LDL apolipoprotein (apo) B-100 was increased in hLCAT+/LDLr+/+ (26+/-4 versus 5+/-0) and hLCAT+/LDLr+/- (4+/-1 versus 1+/-0) rabbits, whereas the FCR of LDL apoB-100 in both groups of LDLr-/- rabbits was nearly identical (0.16+/-0.02 versus 0.15+/-0.02). Consistently, neither aortic lipid concentrations nor the extent of aortic atherosclerosis was significantly different between hLCAT+/LDLr-/- and hLCAT-/LDLr-/- rabbits. Significant correlations were observed between the percent of aortic atherosclerosis and both LDL-C (r=0.985) and LDL apoB-100 FCR (-0.745), as well as between LDL-C and LDL apoB-100 FCR (-0.866). These data are the first to establish that LCAT modulates LDL metabolism via the LDLr pathway, ultimately influencing atherosclerosis susceptibility. Moreover, LCAT's antiatherogenic effect requires only a single functional LDLr allele, identifying LCAT as an attractive gene therapy candidate for the majority of dyslipoproteinemic patients.


Assuntos
Arteriosclerose/patologia , Lipoproteínas/sangue , Fosfatidilcolina-Esterol O-Aciltransferase/fisiologia , Receptores de LDL/fisiologia , Animais , Animais Geneticamente Modificados/genética , Aorta/patologia , Apolipoproteína B-100 , Apolipoproteínas B/sangue , Colesterol/sangue , LDL-Colesterol/sangue , Feminino , Humanos , Lipídeos/sangue , Lipoproteínas LDL/sangue , Masculino , Fosfatidilcolina-Esterol O-Aciltransferase/genética , Coelhos
14.
J Biol Chem ; 274(52): 36912-20, 1999 Dec 24.
Artigo em Inglês | MEDLINE | ID: mdl-10601244

RESUMO

Expression of human lecithin cholesterol acyltransferase (LCAT) in mice (LCAT-Tg) leads to increased high density lipoprotein (HDL) cholesterol levels but paradoxically, enhanced atherosclerosis. We have hypothesized that the absence of cholesteryl ester transfer protein (CETP) in LCAT-Tg mice facilitates the accumulation of dysfunctional HDL leading to impaired reverse cholesterol transport and the development of a pro-atherogenic state. To test this hypothesis we cross-bred LCAT-Tg with CETP-Tg mice. On both regular chow and high fat, high cholesterol diets, expression of CETP in LCAT-Tg mice reduced total cholesterol (-39% and -13%, respectively; p < 0.05), reflecting a decrease in HDL cholesterol levels. CETP normalized both the plasma clearance of [(3)H]cholesteryl esters ([(3)H]CE) from HDL (fractional catabolic rate in days(-1): LCAT-Tg = 3.7 +/- 0.34, LCATxCETP-Tg = 6.1 +/- 0.16, and controls = 6.4 +/- 0.16) as well as the liver uptake of [(3)H]CE from HDL (LCAT-Tg = 36%, LCATxCETP-Tg = 65%, and controls = 63%) in LCAT-Tg mice. On the pro-atherogenic diet the mean aortic lesion area was reduced by 41% in LCATxCETP-Tg (21.2 +/- 2.0 micrometer(2) x 10(3)) compared with LCAT-Tg mice (35.7 +/- 2.0 micrometer(2) x 10(3); p < 0.001). Adenovirus-mediated expression of scavenger receptor class B (SR-BI) failed to normalize the plasma clearance and liver uptake of [(3)H]CE from LCAT-Tg HDL. Thus, the ability of SR-BI to facilitate the selective uptake of CE from LCAT-Tg HDL is impaired, indicating a potential mechanism leading to impaired reverse cholesterol transport and atherosclerosis in these animals. We conclude that CETP expression reduces atherosclerosis in LCAT-Tg mice by restoring the functional properties of LCAT-Tg mouse HDL and promoting the hepatic uptake of HDL-CE. These findings provide definitive in vivo evidence supporting the proposed anti-atherogenic role of CETP in facilitating HDL-mediated reverse cholesterol transport and demonstrate that CETP expression is beneficial in pro-atherogenic states that result from impaired reverse cholesterol transport.


Assuntos
Doenças da Aorta/prevenção & controle , Arteriosclerose/prevenção & controle , Proteínas de Transporte/fisiologia , Glicoproteínas , Lipoproteínas HDL/fisiologia , Esterol O-Aciltransferase/fisiologia , Animais , Proteínas de Transferência de Ésteres de Colesterol , Ésteres do Colesterol/metabolismo , Feminino , Humanos , Lipoproteínas HDL/sangue , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Esterol O-Aciltransferase/genética
15.
Proc Natl Acad Sci U S A ; 96(22): 12685-90, 1999 Oct 26.
Artigo em Inglês | MEDLINE | ID: mdl-10535983

RESUMO

Tangier disease is characterized by low serum high density lipoproteins and a biochemical defect in the cellular efflux of lipids to high density lipoproteins. ABC1, a member of the ATP-binding cassette family, recently has been identified as the defective gene in Tangier disease. We report here the organization of the human ABC1 gene and the identification of a mutation in the ABC1 gene from the original Tangier disease kindred. The organization of the human ABC1 gene is similar to that of the mouse ABC1 gene and other related ABC genes. The ABC1 gene contains 49 exons that range in size from 33 to 249 bp and is over 70 kb in length. Sequence analysis of the ABC1 gene revealed that the proband for Tangier disease was homozygous for a deletion of nucleotides 3283 and 3284 (TC) in exon 22. The deletion results in a frameshift mutation and a premature stop codon starting at nucleotide 3375. The product is predicted to encode a nonfunctional protein of 1,084 aa, which is approximately half the size of the full-length ABC1 protein. The loss of a Mnl1 restriction site, which results from the deletion, was used to establish the genotype of the rest of the kindred. In summary, we report on the genomic organization of the human ABC1 gene and identify a frameshift mutation in the ABC1 gene of the index case of Tangier disease. These results will be useful in the future characterization of the structure and function of the ABC1 gene and the analysis of additional ABC1 mutations in patients with Tangier disease.


Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Glicoproteínas/genética , Doença de Tangier/genética , Transportador 1 de Cassete de Ligação de ATP , Animais , Sequência de Bases , DNA , Éxons , Feminino , Humanos , Íntrons , Masculino , Camundongos , Linhagem
16.
Proc Natl Acad Sci U S A ; 96(18): 10158-63, 1999 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-10468579

RESUMO

Receptors that endocytose high-density lipoproteins (HDL) have been elusive. Here yolk-sac endoderm-like cells were used to identify an endocytic receptor for HDL. The receptor was isolated by HDL affinity chromatography and identified as cubilin, the recently described endocytic receptor for intrinsic factor-vitamin B(12). Cubilin antibodies inhibit HDL endocytosis by the endoderm-like cells and in mouse embryo yolk-sac endoderm, a prominent site of cubilin expression. Cubilin-mediated HDL endocytosis is inhibitable by HDL(2), HDL(3), apolipoprotein (apo)A-I, apoA-II, apoE, and RAP, but not by low-density lipoprotein (LDL), oxidized LDL, VLDL, apoC-I, apoC-III, or heparin. These findings, coupled with the fact that cubilin is expressed in kidney proximal tubules, suggest a role for this receptor in embryonic acquisition of maternal HDL and renal catabolism of filterable forms of HDL.


Assuntos
Proteínas de Transporte , Endocitose/fisiologia , Endoderma/fisiologia , Túbulos Renais Proximais/fisiologia , Lipoproteínas HDL/metabolismo , Proteínas de Ligação a RNA , Receptores de Superfície Celular/fisiologia , Receptores de Lipoproteínas/metabolismo , Animais , Apolipoproteína A-I/metabolismo , Apolipoproteína A-II/metabolismo , Apolipoproteína C-III , Apolipoproteínas C/metabolismo , Cromatografia de Afinidade , Humanos , Cinética , Glicoproteínas de Membrana/fisiologia , Camundongos , Microscopia Confocal , Receptores de Lipoproteínas/isolamento & purificação , Saco Vitelino/fisiologia
17.
Nat Genet ; 22(4): 352-5, 1999 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-10431238

RESUMO

Tangier disease (TD) was first discovered nearly 40 years ago in two siblings living on Tangier Island. This autosomal co-dominant condition is characterized in the homozygous state by the absence of HDL-cholesterol (HDL-C) from plasma, hepatosplenomegaly, peripheral neuropathy and frequently premature coronary artery disease (CAD). In heterozygotes, HDL-C levels are about one-half those of normal individuals. Impaired cholesterol efflux from macrophages leads to the presence of foam cells throughout the body, which may explain the increased risk of coronary heart disease in some TD families. We report here refining of our previous linkage of the TD gene to a 1-cM region between markers D9S271 and D9S1866 on chromosome 9q31, in which we found the gene encoding human ATP cassette-binding transporter 1 (ABC1). We also found a change in ABC1 expression level on cholesterol loading of phorbol ester-treated THP1 macrophages, substantiating the role of ABC1 in cholesterol efflux. We cloned the full-length cDNA and sequenced the gene in two unrelated families with four TD homozygotes. In the first pedigree, a 1-bp deletion in exon 13, resulting in truncation of the predicted protein to approximately one-fourth of its normal size, co-segregated with the disease phenotype. An in-frame insertion-deletion in exon 12 was found in the second family. Our findings indicate that defects in ABC1, encoding a member of the ABC transporter superfamily, are the cause of TD.


Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Glicoproteínas/genética , Mutação , Doença de Tangier/genética , Transportador 1 de Cassete de Ligação de ATP , Sequência de Aminoácidos , Apolipoproteínas E/sangue , Sequência de Bases , Cromossomos Humanos Par 9 , Éxons , Feminino , Biblioteca Gênica , Marcadores Genéticos , Humanos , Lipoproteínas HDL , Masculino , Modelos Biológicos , Modelos Genéticos , Dados de Sequência Molecular , Linhagem
18.
J Lipid Res ; 40(7): 1294-303, 1999 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-10393214

RESUMO

Hepatic lipase (HL) plays a major role in high-density lipoprotein (HDL) metabolism both as a lipolytic enzyme and as a ligand. To investigate whether HL enhances the uptake of HDL-cholesteryl ester (CE) via the newly described scavenger receptor BI (SR-BI), we measured the effects of expressing HL and SR-BI on HDL-cell association as well as uptake of 125I-labeled apoA-I and [3H]CE-HDL, by embryonal kidney 293 cells. As expected, HDL cell association and CE selective uptake were increased in SR-BI transfected cells by 2- and 4-fold, respectively, compared to controls (P < 0.001). Cells transfected with HL alone or in combination with SR-BI expressed similar amounts of HL, 20% of which was bound to cell surface proteoglycans. HL alone increased HDL cell association by 2-fold but had no effect on HDL-CE uptake in 293 cells. However, in cells expressing SR-BI, HL further enhanced the selective uptake of CE from HDL by 3-fold (P < 0.001). To determine whether the lipolytic and/or ligand function of HL are required in this process, we generated a catalytically inactive form of HL (HL-145G). Cells co-transfected with HL-145G and SR-BI increased their HDL cell association and HDL-CE selective uptake by 1.4-fold compared to cells expressing SR-BI only (P < 0.03). Heparin abolished the effect of HL-145G on SR-BI-mediated HDL-CE selective uptake.Thus, the enhanced uptake of HDL-CE by HL is mediated by both its ligand role, which requires interaction with proteoglycans, and by lipolysis with subsequent HDL particle remodeling. These results establish HL as a major modulator of SR-BI mediated selective uptake of HDL-CE.


Assuntos
Antígenos CD36/metabolismo , Ésteres do Colesterol/metabolismo , Lipase/metabolismo , Lipoproteínas HDL/metabolismo , Fígado/enzimologia , Proteínas de Membrana , Receptores Imunológicos , Receptores de Lipoproteínas/metabolismo , Antígenos CD36/genética , Catálise , Linhagem Celular , Ensaio de Imunoadsorção Enzimática , Humanos , Lipase/genética , Plasmídeos , Receptores Depuradores , Receptores Depuradores Classe B , Transfecção
19.
Am J Cardiol ; 83(9B): 3F-12F, 1999 May 13.
Artigo em Inglês | MEDLINE | ID: mdl-10357568

RESUMO

There is a growing body of evidence from epidemiologic, clinical, and laboratory data that indicates that elevated triglyceride levels are an independent risk factor for cardiovascular disease. Identification and quantification of atherogenic lipoproteins in patients with hypertriglyceridemia are important steps in the prevention of cardiovascular disease. Increased levels of apoC-III, apoC-I, or apoA-II on the apoB-containing lipoproteins may alter lipoprotein metabolism and result in the accumulation of atherogenic remnants. Hypertriglyceridemic patients at risk for cardiovascular disease often develop a lipoprotein profile characterized by elevated triglyceride, dense LDL, and low HDL cholesterol. Understanding that each of these factors contributes separately to the patient's risk of cardiovascular disease can help physicians provide patients with more effective risk-reduction programs for cardiovascular disease.


Assuntos
Doenças Cardiovasculares/etiologia , Hipertrigliceridemia/complicações , Lipoproteínas/sangue , Apolipoproteínas/metabolismo , LDL-Colesterol/sangue , LDL-Colesterol/metabolismo , Feminino , Humanos , Hipertrigliceridemia/metabolismo , Lipoproteínas/metabolismo , Masculino , Fatores de Risco
20.
J Lipid Res ; 39(12): 2436-42, 1998 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-9831632

RESUMO

We have investigated the role of hepatic lipase (HL) in remnant lipoprotein metabolism independent of lipolysis by using recombinant adenovirus to express native and catalytically inactive HL (HL-145G) in apolipoprotein (apo)E-deficient mice characterized by increased plasma concentrations of apoB-48-containing remnants. In the absence of apoE, the mechanisms by which apoB-48-containing remnants are taken up by either low density lipoprotein (LDL)-receptor or LDL-receptor-related protein (LRP) remain unclear. Overexpression of either native or catalytically inactive HL in apoE-deficient mice led to similar reductions (P > 0.5) in the plasma concentrations of cholesterol (41% and 53%) and non high density lipoprotein (HDL)-cholesterol (41% and 56%) indicating that even in the absence of lipolysis, HL can partially compensate for the absence of apoE in this animal model. Although the clearance of [3H]cholesteryl ether from VLDL was significantly increased (approximately 2-fold; P < 0. 02) in mice expressing native or inactive HL compared to luciferase controls, the fractional catabolic rates (FCR) of [125I-labeled] apoB- very low density lipoprotein (VLDL) in all three groups of mice were similar (P > 0.4, all) indicating selective cholesterol uptake. Hepatic uptake of [3H]cholesteryl ether from VLDL was greater in mice expressing either native HL (87%) or inactive HL-145G (72%) compared to luciferase controls (56%). Our combined findings are consistent with a role for HL in mediating the selective uptake of cholesterol from remnant lipoproteins in apoE-deficient mice, independent of lipolysis. These studies support the concept that hepatic lipase (HL) may serve as a ligand that mediates the interaction between remnant lipoproteins and cell surface receptors and/or proteoglycans. We hypothesize that one of these pathways may involve the interaction of HL with cell surface receptors, such as scavenger receptor (SR)-BI, that mediate the selective uptake of cholesteryl esters.


Assuntos
Ésteres do Colesterol/metabolismo , Lipoproteínas/metabolismo , Fígado/enzimologia , Adenoviridae/genética , Animais , Apolipoproteínas E/deficiência , Catálise , Vetores Genéticos , Masculino , Camundongos
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