Familial High-density Lipoprotein Deficiency Research Articles

ABCA1 deficiency causes tissue-specific dysregulation of the SREBP2 pathway in mice

ABCA1 plays an essential role in the formation of high-density lipoprotein (HDL), and its mutations cause Tangier disease (TD), a familial HDL deficiency. In addition to the disappearance of HDL, TD patients exhibit cholesterol deposition in peripheral tissues through a mechanism poorly understood, which may contribute to the development of premature atherosclerosis. We and others previously showed that ABCA1 deficiency causes hyperactivation of the SREBP2 pathway in vitro. Here, we show using Abca1 knockout mice that ABCA1 deficiency leads to tissue-specific dysregulation of SREBP2 activity in a nutritional status-dependent manner, which may underlie the pathophysiology of TD.

The brave new world of genetic testing in the management of the dyslipidaemias.

With the exception of familial hypercholesterolaemia, the value of genetic testing for managing dyslipidaemias is not established. We review the genetics of major dyslipidaemias in context of clinical practice. Genetic testing for familial hypercholesterolaemia is valuable to enhance diagnostic precision, cascade testing, risk prediction and the use of new medications. Hypertriglyceridaemia may be caused by rare recessive monogenic, or by polygenic, gene variants; genetic testing may be useful in the former, for which antisense therapy targeting apoC-III has been approved. Familial high-density lipoprotein deficiency is caused by specific genetic mutations, but there is no effective therapy. Familial combined hyperlipidaemia (FCHL) is caused by polygenic variants for which there is no specific gene testing panel. Familial dysbetalipoproteinaemia is less frequent and commonly caused by APOE ε2ε2 homozygosity; as with FCHL, it is responsive to lifestyle modifications and statins or/and fibrates. Elevated lipoprotein(a) is a quantitative genetic trait whose value in risk prediction over-rides genetic testing; treatment relies on RNA therapeutics. Genetic testing is not at present commonly available for managing dyslipidaemias. Rapidly advancing technology may presage wider use, but its worth will require demonstration of cost-effectiveness and a healthcare workforce trained in genomic medicine.

Structure of the Human Lipid Exporter ABCA1

ABCA1, an ATP-binding cassette (ABC) subfamily A exporter, mediates the cellular efflux of phospholipids and cholesterol to the extracellular acceptor apolipoprotein A-I (apoA-I) for generation of nascent high-density lipoprotein (HDL). Mutations of human ABCA1 are associated with Tangier disease and familial HDL deficiency. Here, we report the cryo-EM structure of human ABCA1 with nominal resolutions of 4.1Å for the overall structure and 3.9Å for the massive extracellular domain. The nucleotide-binding domains (NBDs) display a nucleotide-free state, while the two transmembrane domains (TMDs) contact each other through a narrow interface in the intracellular leaflet of the membrane. In addition to TMDs and NBDs, two extracellular domains of ABCA1 enclose an elongated hydrophobic tunnel. Structural mapping of dozens of disease-related mutations allows potential interpretation of their diverse pathogenic mechanisms. Structural-based analysis suggests a plausible "lateral access" mechanism for ABCA1-mediated lipid export that may be distinct from the conventional alternating-access paradigm.

A complex phenotype in a child with familial HDL deficiency due to a novel frameshift mutation in APOA1 gene (apoA-IGuastalla)

We describe a kindred with high-density lipoprotein (HDL) deficiency due to APOA1 gene mutation in which comorbidities affected the phenotypic expression of the disorder. An overweight boy with hypertriglyceridemia (HTG) and HDL deficiency (HDL cholesterol 0.39mmol/L, apoA-I 40mg/dL) was investigated. We sequenced the candidate genes for HTG (LPL, APOC2, APOA5, GPIHBP1, LMF1) and HDL deficiency (LCAT, ABCA1 and APOA1), analyzed HDL subpopulations, measured cholesterol efflux capacity (CEC) of sera and constructed a model of the mutant apoA-I. No mutations in HTG-related genes, ABCA1 and LCAT were found. APOA1 sequence showed that the proband, his mother and maternal grandfather were heterozygous of a novel frameshift mutation (c.546_547delGC), which generated a truncated protein (p.[L159Afs*20]) containing 177 amino acids with an abnormal C-terminal tail of 19 amino acids. Trace amounts of this protein were detectable in plasma. Mutation carriers had reduced levels of LpA-I, preβ-HDL and large HDL and no detectable HDL-2 in their plasma; their sera had a reduced CEC specifically the ABCA1-mediated CEC. Metabolic syndrome in the proband explains the extremely low HDL cholesterol level (0.31mmol/L), which was half of that found in the other carriers. The proband's mother and grandfather, both presenting low plasma low-density lipoprotein cholesterol, were carriers of the β-thalassemic trait, a condition known to be associated with a reduced low-density lipoprotein cholesterol and a reduced prevalence of cardiovascular disease. This trait might have delayed the development of atherosclerosis related to HDL deficiency. In these heterozygotes for apoA-I truncation, the metabolic syndrome has deleterious effect on HDL system, whereas β-thalassemia trait may delay the onset of cardiovascular disease.

Deficiency in the Lipid Exporter ABCA1 Impairs Retrograde Sterol Movement and Disrupts Sterol Sensing at the Endoplasmic Reticulum

Cellular cholesterol homeostasis involves sterol sensing at the endoplasmic reticulum (ER) and sterol export from the plasma membrane (PM). Sterol sensing at the ER requires efficient sterol delivery from the PM; however, the macromolecules that facilitate retrograde sterol transport at the PM have not been identified. ATP-binding cassette transporter A1 (ABCA1) mediates cholesterol and phospholipid export to apolipoprotein A-I for the assembly of high density lipoprotein (HDL). Mutations in ABCA1 cause Tangier disease, a familial HDL deficiency. Several lines of clinical and experimental evidence suggest a second function of ABCA1 in cellular cholesterol homeostasis in addition to mediating cholesterol efflux. Here, we report the unexpected finding that ABCA1 also plays a key role in facilitating retrograde sterol transport from the PM to the ER for sterol sensing. Deficiency in ABCA1 delays sterol esterification at the ER and activates the SREBP-2 cleavage pathway. The intrinsic ATPase activity in ABCA1 is required to facilitate retrograde sterol transport. ABCA1 deficiency causes alternation of PM composition and hampers a clathrin-independent endocytic activity that is required for ER sterol sensing. Our finding identifies ABCA1 as a key macromolecule facilitating bidirectional sterol movement at the PM and shows that ABCA1 controls retrograde sterol transport by modulating a certain clathrin-independent endocytic process.

A-Subclass ATP-Binding Cassette Proteins in Brain Lipid Homeostasis and Neurodegeneration

The A-subclass of ATP-binding cassette (ABC) transporters comprises 12 structurally related members of the evolutionarily highly conserved superfamily of ABC transporters. ABCA transporters represent a subgroup of “full-size” multispan transporters of which several members have been shown to mediate the transport of a variety of physiologic lipid compounds across membrane barriers. The importance of ABCA transporters in human disease is documented by the observations that so far four members of this protein family (ABCA1, ABCA3, ABCA4, ABCA12) have been causatively linked to monogenetic disorders including familial high-density lipoprotein deficiency, neonatal surfactant deficiency, degenerative retinopathies, and congenital keratinization disorders. Recent research also point to a significant contribution of several A-subfamily ABC transporters to neurodegenerative diseases, in particular Alzheimer’s disease (AD). This review will give a summary of our current knowledge of the A-subclass of ABC transporters with a special focus on brain lipid homeostasis and their involvement in AD.

Familial high-density lipoprotein deficiency states and premature coronary heart disease

High-density lipoprotein cholesterol (HDL-C) values <40 mg/dL have been shown to be an independent risk factor for coronary heart disease (CHD). Rare genetic disorders associated with marked human HDL deficiency include apolipoprotein A-I (apoA-I) deficiency with undetectable plasma apoA-I, which can be due to defects within the APOA1 gene resulting in lack of apoA-I secretion. Such patients have marked HDL deficiency, normal levels of triglycerides (TGs), and low-density lipoprotein cholesterol (LDL-C), and they can have xanthomas and premature CHD. ApoA-I variants with amino acid substitutions, especially in the region of amino acid residues 50–93 and 170–178, have been associated with amyloidosis. Patients with homozygous Tangier disease have defective cellular cholesterol efflux due to mutations in the adenosine-5′-triphosphate (ATP)-binding cassette transporter A1, detectable plasma apoA-I levels, and only pre-β1 HDL in their plasma. They have decreased LDL-C levels and can develop neuropathy and premature CHD. Patients with lecithin:cholesterol acyltransferase deficiency have both pre-β1 and α4 HDL present in their plasma and develop corneal opacities, anemia, proteinuria, and kidney failure. HDL deficiency has been associated with hypertriglyceridemia, obesity, insulin resistance, and diabetes. Common familial disorders associated with premature CHD and low HDL are: (1) dyslipidemia, seen in 15% of families, (2) combined hyperlipidemia, seen in 14% of families, and (3) hypoalphalipoproteinemia, seen in 4% of families. A common feature of all these disorders associated with premature CHD is a marked deficiency of very large α1 HDL. Niacin is currently the most optimal treatment strategy to raise HDL-C and normalize the HDL particle profile in these patients. Hot Topics in Cardiometabolic Disorders Volume 2 • Issue 3 • Year 2011 © FBCommunication Modena (Italy) Cite this article as: Schaefer EJ, Santos RD, Tani M, Schaefer PM, Asztalos BF. Familial high-density lipoprotein deficiency states and premature coronary heart disease. Hot Topics Cardiomet Disord 2011;2(3):7-16. E-mail: ernst.schaefer@tufts.edu Hot Topics in Cardiometabolic Disorders 2011;3:7-16 Copyright © 2011 FBCommunication s.r.l. a socio unico Downloaded from www.hottopicsin.com As shown in Figure 2-upper panel pre-β1 HDL is a very small disk with two molecules of apoA-I forming a belt (in yellow) around approximately 16 molecules of phospholipid (in blue), with a diameter of about 5.6 nm and a molecular weight of about 70 kDa. This particle is converted to small α4 HDL, which is a small disk with 2 molecules of apoA-I, but has about 26 molecules of phospholipid, and about 12 molecules of free cholesterol (in green), with a diameter of about 7.4 nm, and a molecular weight of about 80 kDa. Figure 2-lower panel depicts a model of medium spherical α3 HDL with 2 molecules of apoA-I, 1 molecule of apoA-II, with phospholipid and free cholesterol on its surface, and cholesteryl ester (in green) and TG (in purple) in the HDL core with a diameter of about 8.0 nm. The large spherical α2 HDL contains 4 molecules of apoA-I; 2 molecules of apoA-II, phospholipid, and free cholesterol on its surface; and cholesteryl ester and TG in its core, with a diameter of about 9.2 nm (see Figure 2–lower panel). The α1 HDL is very large and spherical with 8 molecules of apoA-I, phospholipid, and free cholesterol on its surface, and cholesteryl ester and TG in the core of HDL with a diameter of about 11.0 nm (see Figure 2–lower panel) [19]. In collaboration with George Rothblat’s laboratory, the authors studied the roles of HDL particles in ABCA1-, ABCG1-, and scavenger receptor class B type 1 (SRB1) mediated lipid flux [20-23]. It has been documented that only the small precursor HDL (pre-β1) was able to remove lipids (phospholipid and free cholesterol) from cells via the ABCA1 pathway, and that the bidirectional cell-lipid flux via the SRB1 mechanism was mediated by the larger HDL

Tangier Disease in Severely Progressive Coronary and Peripheral Artery Disease

A 37-year-old man was first referred to our lipid clinic in December 2007 for plasma lipid alteration. He presented with large, orange tonsils (Figure 1) and hepatosplenomegaly. There was no evidence of corneal opacities nor of other ocular abnormalities, and there were no nervous system abnormalities as assessed by sensitive and motor electromyography. Figure 1. Orange large tonsils. Hematologic abnormalities included thrombocytopenia and erythrocytes with altered morphology (stomatocytes; Figure 2) and function (decreased osmotic resistance). Serum levels of total cholesterol, triglycerides, and high-density lipoprotein (HDL) cholesterol were 58, 184, and 4 mg/dL, respectively; plasma apolipoprotein A-I concentrations were very low (3.9 mg/dL); and apolipoprotein A-II plasma levels were 1.7 mg/dL. Figure 2. Peripheral blood erythrocytes with numerous stomatocytes. The patient’s father (58 years of age) had serum levels of total cholesterol, HDL cholesterol, and apolipoprotein A-I of 127, 25, and 99.5 mg/dL, respectively. Moreover, he had a …

Effect of ABCA1 mutations on risk for myocardial infarction

The adenosine triphosphate-binding cassette A1 (ABCA1) gene codes for a cellular phospholipid and cholesterol transporter that mediates the initial and essential step in high-density lipoprotein (HDL) biogenesis: the formation of nascent HDL particles. Mutations at the ABCA1 gene locus cause severe familial HDL deficiency and, in the homozygous form, cause Tangier disease. Several studies have investigated the influence of ABCA1 variation on lipid metabolism and coronary heart disease, but they have resulted in controversial and inconsistent results. Genetic variability at the ABCA1 gene has also been associated with increased risk of myocardial infarction. In one study, this association was independent of HDL cholesterol levels, raising the possibility that the measurement of HDL cholesterol levels may not provide adequate information on the functional roles of HDL particles. Nevertheless, genomic screening for complex diseases, such as coronary heart disease, and HDL deficiency in particular, may not add additional information to that gained from conventional global cardiovascular risk stratification.

Multiple Drug Resistance Associated with Function of ABC-Transporters in Diabetes Mellitus: Molecular Mechanism and Clinical Relevance

ATP-binding cassette (ABC) transporters are involved in a variety of physiological processes such as lipid metabolism, ion homeostasis and immune functions. A large number of these proteins have been causatively linked to rare and common human genetic diseases including familial high-density lipoprotein deficiency, retinopathies, cystic fibrosis, diabetes and cardiomyopathies. Furthermore, genetic variations in ABC transporter genes and deregulated expression patterns significantly contribute to drug resistance in human cancer and pancreatic beta cells and alter the pharmacokinetic properties of a variety of drugs. Up-to-date 15 ABC transporters have been identified in human pancreatic beta cells, however only a few of them are identified to date as proteins/genes associated with multidrug resistance (MDR) in diabetes mellitus. Prominent members include the multidrug resistance protein 1 (MRP1/ABCC1), sulfonylurea receptor 1 (SUR1/ABCC8), the multi drug transporter TAP2 and member of the ATP-binding cassette transporter subfamily A (ABCA1). ABCC8 is a subunit of the pancreatic beta-cell K(ATP) channel and plays a key role in the regulation of glucose-induced insulin secretion. Although the physiological role of these transporters to MDR is not yet fully understood, they play an important role in the blood-membrane barrier in pancreatic beta cells. The aim of this article is to provide an overview and to present few examples of drug treatment in MDR in diabetes mellitus associated with function of ABC-transporters.

Physiologically regulated transgenic ABCA1 does not reduce amyloid burden or amyloid-β peptide levels in vivo

ABCA1-deficient mice have low levels of poorly lipidated apolipoprotein E (apoE) and exhibit increased amyloid load. To test whether excess ABCA1 protects from amyloid deposition, we crossed APP/PS1 mice to ABCA1 bacterial artificial chromosome (BAC) transgenic mice. Compared with wild-type animals, the ABCA1 BAC led to a 50% increase in cortical ABCA1 protein and a 15% increase in apoE abundance, demonstrating that this BAC supports modest ABCA1 overexpression in brain. However, this was observed only in animals that do not deposit amyloid. Comparison of ABCA1/APP/PS1 mice with APP/PS1 controls revealed no differences in levels of brain ABCA1 protein, amyloid, Abeta, or apoE, despite clear retention of ABCA1 overexpression in the livers of these animals. To further investigate ABCA1 expression in the amyloid-containing brain, we then compared ABCA1 mRNA and protein levels in young and aged cortex and cerebellum of APP/PS1 and ABCA1/APP/PS1 animals. Compared with APP/PS1 controls, aged ABCA1/APP/PS1 mice exhibited increased ABCA1 mRNA, but not protein, selectively in cortex. Additionally, ABCA1 mRNA levels were not increased before amyloid deposition but were induced only in the presence of extensive Abeta and amyloid levels. These data suggest that an induction of ABCA1 expression may be associated with late-stage Alzheimer's neuropathology.

ABCA7 expression is regulated by cellular cholesterol through the SREBP2 pathway and associated with phagocytosis

ABCA7 is highly homologous to ABCA1 and mediates cellular cholesterol and phospholipid release by apolipoproteins when transfected in vitro. However, expression of ABCA7 was downregulated by increased cellular cholesterol while ABCA1 was upregulated, and the results were consistent by forced expression or downregulation of sterol-responsive/regulatory element (SRE) binding proteins (SREBPs). We analyzed the promoter of the ABCA7 gene and identified the new exon encoding 96 bp (mouse) and 95 bp (human) of the 5' untranslated region and the transcription start site at 1,122 bp (mouse) and 1,260 bp (human) upstream of the initiation methionine codon. At 5' upstream of this exon is the ABCA7 proximal promoter containing multiple binding sites of transcription factors for hematopoiesis and SRE of 9 bp at 212 bp (mouse) and 179 bp (human) upstream of the new exon. The apolipoprotein A-I-mediated lipid release was not influenced by suppression of the endogenous ABCA7 with small interfering RNA in mouse fibroblasts or by its increase in ABCA1-deficient mouse cells. In contrast, phagocytic activity was altered in parallel to the ABCA7 expression in these cells. When phagocytosis was induced, the messages increased for SREBP2, ABCA7, and other SREBP2-regulated proteins. The ABCA1 message decreased in this condition. We conclude that the ABCA7 gene is regulated by sterol in the opposite direction to ABCA1 through SRE/SREBP2 and that expression of ABCA7 by this regulation is associated with phagocytic activity.

Molecular diagnosis of ATP-binding cassette transporter-related diseases

ATP-binding cassette (ABC) transporters are involved in a variety of physiologic processes such as xenobiotic defense, lipid metabolism, ion homeostasis and immune functions. A large number of ABC proteins have been causatively linked to rare and common human genetic diseases including familial high-density lipoprotein deficiency, retinopathies, cystic fibrosis, diabetes and cardiomyopathies. Furthermore, genetic variations in ABC transporter genes and dysregulated expression patterns of these molecules significantly contribute to drug resistance in human cancer cells and alter the pharmacokinetic properties of a variety of drugs. In order to analyze DNA sequence alterations or define disease-associated mRNA expression patterns of the complete ABC transporter superfamily, novel high-throughput molecular methods such as quantitative real-time PCR and DNA microarray analysis are emerging. The aim of this review is to provide an overview and to present some examples of human ABC transporters involved in monogenic diseases, cancer and pharmacogenetics. Methodologic aspects of molecular diagnostics applied to analyze genetic variations, mRNA and protein expression levels and functional characteristics of ABC transporters are discussed.

1115. Apo A-I and ABCA1 Gene Transfer both Increase HDL Cholesterol but Have Opposing Effects on Progression of Early Atherosclerosis in apo E Deficient Mice

Background: High density lipoprotein (HDL) cholesterol levels are negatively correlated with ischemic cardiovascular diseases in humans. Apolipoprotein (apo) A-I is the principal apolipoprotein of HDL and a strong correlation exists between apo A-I and HDL cholesterol levels. The ATP binding cassette transporter I (ABCA1) is defective in patients with Tangier disease and familial HDL deficiency. Indirect evidence suggests that ABCA1 expression in the liver is an important determinant of HDL cholesterol levels. Therefore, the effect of adenoviral human apo A-I (AdA-I), murine ABCA1 (AdABCA1) and combined AdA-I and AdABCA1 transfer on the lipoprotein profile and on progression of atherosclerosis in C57BL/6 apo E deficient mice was evaluated.

Denaturing high-performance liquid chromatography in the detection of ABCA1 gene mutations in familial HDL deficiency

Mutations in the ABCA1 gene are the cause of familial high density lipoprotein deficiency (FHD). Because these mutations are spread over the entire gene, their detection requires the sequencing of all 50 exons. The aim of this study was to validate denaturing high-performance liquid chromatography (DHPLC) in mutation detection as an alternative to systematic sequencing. Exons of the ABCA1 gene were amplified using primers employed for sequencing. Temperatures for DHPLC were deducted from a software and empirically defined for each amplicon. To assess DHPLC reliability, we tested 30 sequence variants found in FHD patients and controls. Combined DHPLC and sequencing was applied to the genotyping of new FHD patients. Most of the amplicons required from two to five temperature conditions to obtain partially denatured DNA over the entire amplicon length. Twenty-nine of the variants found by sequencing were detected by DHPLC (97% sensitivity). The detection of the last variant (in exon 40) required different primers and amplification conditions. DHPLC and sequencing analysis of new FHD patients revealed that all amplicons showing a heteroduplex DHPLC profile contained sequence variants. No variants were detected in amplicons with a homoduplex profile. DHPLC is a sensitive and reliable method for the detection of ABCA1 gene mutations.

LXR-mediated activation of macrophage stearoyl-CoA desaturase generates unsaturated fatty acids that destabilize ABCA1

Abnormal HDL metabolism among patients with diabetes and insulin resistance may contribute to their increased risk of atherosclerosis. ATP binding cassette transporter A1 (ABCA1) mediates the transport of cholesterol and phospholipids from cells to HDL apolipoproteins and thus modulates HDL levels and atherogenesis. Because fatty acids are increased in diabetes, we examined their effects on ABCA1 activity in cultured macrophages. cAMP analogs and ligands for the liver X receptor/retinoid X receptor (LXR/RXR) system can induce Abca1 transcription in murine macrophages. When induced by cAMP, unsaturated but not saturated long-chain fatty acids inhibit apolipoprotein-mediated lipid efflux by destabilizing ABCA1 protein. Here, we show that the saturated fatty acids palmitate and stearate also destabilize ABCA1 when Abca1 is induced by LXR/RXR ligands instead of cAMP. This was associated with increased palmitate and stearate desaturation by stearoyl-CoA desaturase (SCD), another gene product induced by LXR/RXR ligands. The SCD inhibitors conjugated linoleic acid and troglitazone nearly abolished ABCA1 destabilization by palmitate and stearate but not by linoleate. These results suggest that LXR/RXR ligands generate ABCA1-destabilizing monounsaturated fatty acids from their saturated precursors by activating SCD. Thus, with cholesterol-loaded macrophages exposed to saturated fatty acids, activated LXR/RXR may counteract the enhanced ABCA1 transcription by reducing the ABCA1 protein content.

Expression pattern and raft association of NIPSNAP3 and NIPSNAP4, highly homologous proteins encoded by genes in close proximity to the ATP-binding cassette transporter A1

The highly homologous genes NIPSNAP3 and NIPSNAP4, with 87% amino acid identity, are members of the NIPSNAP family with putative roles in vesicular trafficking. NIPSNAP3 mRNA and NIPSNAP4 mRNA and protein were detected in multiple tissues and cells at varying degrees. Interestingly, NIPSNAP3 is most highly expressed in skeletal muscle, where NIPSNAP4 has a low mRNA abundance. NIPSNAP4 was found associated with membranes and partly localized in rafts. The ubiquitous expression of the highly conserved NIPSNAPs and their association with membranes further support an important cellular function of these proteins probably linked to vesicular trafficking. The NIPSNAP3 and NIPSNAP4 genes are located in close proximity to the 3′ end of the ATP-binding cassette transporter A1 (ABCA1), whose mutations cause familial high-density lipoprotein deficiency syndromes. The adjacent genomic location and the finding that ABCA1 is a regulator of vesicular trafficking may indicate a functional relation of these proteins, even though NIPSNAP4 does not interact directly with ABCA1 nor is its expression altered in cells with mutated ABCA1.

Human ATP-binding cassette (ABC) transporters and lipoprotein metabolism.

The ATP-binding cassette (ABC) superfamily of active transporters involves a large number of functionally diverse transmembrane proteins. They transport a variety of substrates including amino acids, lipids, inorganic ions, peptides, saccharides, metals, drugs, and proteins. The ABC transporters not only move a variety of substrates into and out of the cell, but also are also involved in intracellular compartmental transport. Energy derived from the hydrolysis of ATP is used to transport the substrate across the membrane against a concentration gradient. The typical ABC transporter consists of two transmembrane domains and two nucleotide-binding domains. Defects in 14 of these transporters cause 13 genetic diseases (cystic fibrosis, Stargardt disease, adrenoleukodystrophy, Tangier disease, etc.). Mutations in three genes affect lipid levels expressively. Mutations in ABCA1 cause severe HDL deficiency syndromes called Tangier disease and familial high-density lipoprotein deficiency, which are characterized by a severe deficiency or absence of high-density lipoprotein in the plasma. Two other ABCG transporters, ABCG5 and ABCG8, mutations of which cause sitosterolemia, have been identified. The affected individuals absorb and retain plant sterols, as well as shellfish sterols.

Expression, regulation, and activity of ABCA1 in human cell lines

Mutations in the ATP-binding cassette transporter A1 (ABCA1) gene cause familial high-density lipoprotein deficiency and Tangier disease. ABCA1 plays a crucial role in active apolipoprotein A-I (apoA-I) lipidation, a key step in reverse cholesterol transport. We compared ABCA1 transcriptional regulation and cholesterol efflux in human skin fibroblasts, monocyte-derived macrophages and hepatocytes (HepG2). 8-Br-cAMP did not increase ABCA1 transcription in these tissues compared to mouse macrophages. We found that ABCA1 is differentially regulated among tissues. While transcription in HepG2 appears to be constitutive, sterols stimulate ABCA1 transcription in fibroblasts and monocyte-derived macrophages. ApoA-I promoted cholesterol efflux in fibroblasts, macrophages, and HepG2. Cholesterol homeostasis in fibroblasts is tightly regulated, and ABCA1 mRNA closely follows the cellular mass of free cholesterol (dose- and time-dependent manner). To further determine the mechanism used by fibroblasts to maintain sterol balance, we used a competitive inhibition approach with geranylgeranyl pyrophosphate (GGPP) to block the LXR induction pathway. GGPP blocked basal, 22-( R)-hydroxycholesterol- and cholesterol-induced ABCA1 expression. Taken together, these results demonstrate that: (1) ABCA1 expression varies among tissues, and (2) cholesterol conversion to hydroxycholesterol is an important mechanism for the maintenance of cholesterol homeostasis in fibroblasts.

Novel ABCA1 compound variant associated with HDL cholesterol deficiency

The recent discovery of an ATP-binding cassette transporter, ABCA1, as an important regulator of high density lipoprotein (HDL) metabolism and reverse cholesterol transport has facilitated the identification of novel variants associated with HDL cholesterol deficiency states. We identified a subject with HDL cholesterol deficiency (4 mg/dl) who developed and died of complications related to cerebral amyloid angiopathy (CAA). The proband had a compound heterozygous mutation. One mutation was a G3295T substitution with conversion of asparagine to tyrosine (D1099Y) in ABCA1. The single-base substitution at codon 1099 resulted in the abolition of an RsaI cleavage site. The proband and affected individuals having another mutation were heterozygotes for T5966C with phenylalanine converted to serine (F2009S). The presence of the T5966C mutation was detected by restriction digestion with HinfI. These variants were not identified in over 400 chromosomes of healthy subjects. In the kindred, family members heterozygous for the ABCA1 variant exhibited low levels of HDL cholesterol. Direct sequencing of all coding regions and splice site junctions of other HDL candidate genes revealed no additional mutations, indicating that combined defective ABCA1 alleles may result in familial HDL deficiency.