Lipoprotein-like Particles Research Articles

Lipid and fatty acid profile of the retina, retinal pigment epithelium/choroid, and the lacrimal gland, and associations with adipose tissue fatty acids in human subjects

Accumulation of lipids within Bruch's membrane (BrM) and between BrM and retinal pigment epithelium (RPE) accounts for one of the biological changes associated with normal aging and may contribute to the development of age-related maculopathies. The origin of these lipids is still being actively investigated. The relative contribution of plasma lipids and lipids coming from the neural retina remains a matter of controversy. Low-density lipoproteins (LDLs) have been reported to significantly participate in the retina's lipid supply, after active remodeling within RPE. Meanwhile, RPE expresses the enzymatic machinery for synthesizing lipoprotein-like particles. The objective of this study was to establish associations between the fatty acid profile of the ocular structures and adipose tissue as a surrogate for the subjects' past dietary intake. Lipids and fatty acids were analyzed from the neural retina, retinal pigment epithelium (RPE)/choroid, the lacrimal gland, and adipose tissue, collected from 27 human donors (19 women, eight men) aged 59–95 years. DHA concentrations in the neural retina were positively associated with the concentrations in cholesteryl esters (CEs) from RPE/choroid and negatively associated with DHA concentrations in phospholipids (PLs) from RPE/choroid. DHA in orbital fat was positively associated with DHA in the lacrimal gland. No significant association was observed in the other ocular structures. Linoleic acid in orbital fat was positively associated with linoleic acid in the lacrimal gland, followed by the neural retina and CEs from RPE/choroid; it was slightly correlated with PLs from RPE/choroid. Other fatty acids that originate exclusively from the diet such as trans fatty acids were detected in orbital fat, the lacrimal gland, PLs, and CEs from RPE/choroid. DHA in the neural retina was poorly associated with its dietary intake, contrary to other fatty acids such as linoleic acid. Within this context, CEs may be important carriers of fatty acids entering the retina. Although epidemiological studies have reported the benefit of DHA in the prevention of age-related macular degeneration, the leading cause of blindness in Western countries, the relevance of supplementing patients with DHA is questioned.

Morphometric Analysis of Lipoprotein-like Particle Accumulation in Aging Human Macular Bruch’s Membrane

To determine the size and regional distribution of lipoprotein-like particles (LLPs) that accumulate with age in Bruch's membrane (BrM). The quick-freeze/deep-etch method was used to prepare specimens of human BrM (age range, 27-78) for electron microscopic examination. Stereologic methods were used to analyze the resultant micrographs and determine the age-related changes of the LLP volume fraction and diameter distribution in various locations in BrM. The volume fraction occupied by LLPs was found to increase monotonically with age in both the inner collagenous layer (ICL) and elastic layer (EL), but not in the outer collagenous layer (OCL). The mass of total LLP-associated lipids in BrM also increased with age. There was no significant increase in LLP size with age, but there was a modest increase in size with increased volume fraction of LLPs in BrM. The pattern of accumulation of particles was consistent with a retinal pigment epithelium (RPE) source for the LLPs, which explains why once the EL and ICL were filled with particles, LLPs continued to accumulate near the RPE, but no further accumulation was found in the OCL. The quantity of LLP-associated lipids found in BrM accounts for a large portion of the accumulated lipids measured in this tissue.

Age-related changes in human macular Bruch's membrane as seen by quick-freeze/deep-etch

Lipid-containing inclusions have been observed in human Bruch's membrane (BrM) and are postulated to be associated with age-related maculopathy (ARM), a major cause of legal blindness in developed countries. The dehydration associated with specimen preparation for thin-section transmission electron microscopy causes loss of these inclusions. Better preservation of the ultrastructure of the inclusions and tissue is achieved by using a quick-freeze/deep-etch preparation. We use this technique to examine normal human macular BrM in order to characterize the deposition of the lipid-rich inclusions and their age-related accumulation within different layers of the tissue. We find that various inclusions mentioned in other studies can be formed by combinations of three basic structures: lipoprotein-like particles (LLPs), small granules (SGs) and membrane-like structures. These inclusions are associated with collagen and elastic fibrils by fine filaments. In younger eyes, these inclusions are found mostly in the elastic (EL) and outer collageneous layer (OCL) and occupy a small fraction of the interfibrillar spacing. As age increases, LLPs and SGs gradually fill the interfibrillar spacing of the EL and inner collageneous layer (ICL) of the tissue, and later form a new sublayer, the lipid wall, within the boundary region between the basal lamina of retinal pigment epithelium (RPE) and ICL. Because the formation of the lipid wall only occurs after these inclusions fill the ICL, and it seems unlikely that the LLPs can pass through the packed layer, this result suggests a possible RPE origin of the LLPs that make up the lipid wall.

Hepatitis C Lipo-Viro-Particle from Chronically Infected Patients Interferes with TLR4 Signaling in Dendritic Cell

BackgroundHepatitis C virus (HCV) can be purified from serum of chronically-infected patients in the form of Lipo-Viro-Particles (LVP), which are triglycerid-rich lipoprotein-like particles containing viral RNA and proteins. Since LVP is a constant feature of chronically infected patients, we asked whether purified LVP could interfere with the immune response by acting directly on dendritic cell (DC) function.Methods and FindingsWe have analyzed the impact of LVP on the maturation monocyte-derived DC induced by TLR3 or TLR4 ligands. Following incubation with LVP, immature DC supported weak transient HCV-RNA replication and type I IFN synthesis. This, however, did not lead to viral particle production nor to maturation of DC. LVP-treatment prior to TLR3 stimulation by polyI:C only enhanced the secretion of IL-12, IL-6 and TNFα yielding typical mature DC. In contrast, LVP-treated DC activated by the TLR4 ligand LPS yielded phenotypically mature DC with reduced capacity to secrete both pro- and anti-inflammatory cytokines. Their ability to stimulate allogeneic T lymphocytes was strongly affected since activated T cells produced IL-5 and IL-13 instead of IFNγ. Addition of IFNα prevented the effect of LVP on DC function. Restoration of IFNγ secretion by T cells was obtained by blocking ERK activation in DC, while induction of IL-5 and IL-13 secretion was inhibited by blocking the p38-MAPK pathway in DC.ConclusionsLVP can interfere with TLR4-triggered maturation of DC, inducing a shift in DC function that stimulates Th2 cells instead of Th1, by a mechanism that is ERK- and p38-MAPK-dependent. The effect of LVP on DC polarization was reversed by IFNα, providing an additional rationale for the interferon therapy of chronically-infected patients. By acting on TLR4 pathway with LVP, HCV may thus exploit a natural protective mechanism of the liver and the intestine normally used to control inflammation and immunity to commensal microorganisms.

Lipid-Altering Therapies and the Progression of Atherosclerotic Disease

Lipids play a key role in the progression of atherosclerosis, and lipid-lowering therapies have been studied for 30 years in coronary disease. Measurement of the progression of atherosclerosis through carotid intima-media thickness, coronary mean lumen diameter, and, mostly recently, intravascular ultrasound is generally accepted. This article reviews the role of lipid-lowering therapies in changing the rate of atherosclerosis progression in the coronary and carotid circulations. Statins are the primary therapy used to reduce atherosclerosis and cardiovascular events, including strokes and transient ischemic attacks, and have benefits in reducing events in patients undergoing carotid endarterectomy. In contrast, data for other agents, including fibrates and nicotinic acid, in reducing the progression of atherosclerosis are less extensive and not as well known. There is increasing interest in optimizing the whole lipid profile, as this might deliver extra benefits over and above statin therapy alone. Initial proof of this concept has recently come from studies that measured the progression of atherosclerosis and showed that adding nicotinic acid to statin therapy and, more directly, infusion of high-density lipoprotein-like particles reduced progression and indeed might induce regression of the disease. It is likely that the management of significant carotid stenosis will become ever more drug focused and will be customized to the lipid profile of each patient with intervention reserved only for late-stage symptomatic disease.

Comparison of Morphology of Human Macular and Peripheral Bruch's Membrane in Older Eyes

Deposits in macular human Bruch's membrane (BrM) increase with age and have been postulated to be associated with age-related maculopathy. We used two ultrastructural methods to compare these deposits by electron microscopy in macular and peripheral BrM of eight eyes from donors 63–86 years of age. Quick-freeze/deep-etch (QFDE) was used to prepare replicas that showed the ultrastructure of deposits, and osmium–tannic acid–paraphenylenediamine (OTAP) was used to preserve small extracellular lipid particles. We found that an accumulation of lipoprotein-like particles (LLPs) occurred in the peripheral BrM just as it does in the macular region, but with perhaps a somewhat slower time course. The “lipid wall,” reported in macular BrM, was also found occasionally in the peripheral regions. The same processes that lead to age-related accumulation of LLPs in macular BrM appear to also occur in the peripheral regions.

Role of cholesterol in amyloid cascade: cholesterol-dependent modulation of tau phosphorylation and mitochondrial function

Apolipoprotein E (apoE) alleles are important genetic risk factors for Alzheimer's disease (AD), with the epsilon4 allele increasing and the epsilon2 allele decreasing the risk of developing AD. ApoE is the major apolipoprotein that modulates cholesterol transport in the central nervous system, cholesterol being an essential component of membranes for maintaining their structure and functions. Epidemiological studies have suggested a link between serum cholesterol levels and AD development and the potential therapeutic effectiveness of statins for AD; and furthermore, biological studies have shown that amyloid beta-protein (Abeta) secretion is modulated by cellular cholesterol level. However, other lines of evidence show controversial results. In addition to the role of cholesterol in Abeta generation, different interactions of cholesterol with Abeta and its role in AD pathogenesis have been shown, i.e. Abeta affects cholesterol dynamics in neurons, and altered cholesterol metabolism in turn leads to neurodegeneration with abnormally phosphorylated tau (tauopathy). In this review, the reciprocal interactions between cholesterol and Abeta, and the role of cholesterol in tauopathy are discussed. The isoform-specific involvement of apoE in this cascade, in which high-density lipoprotein-like particles are generated and supplied to neurons to maintain cholesterol homeostasis, is also discussed.

Association of a Model Class A (Apolipoprotein) Amphipathic α Helical Peptide with Lipid: HIGH RESOLUTION NMR STUDIES OF PEPTIDE·LIPID DISCOIDAL COMPLEXES

Class A amphipathic helical peptides have been shown to mimic apolipoprotein A-I, the major protein component of high density lipoproteins and have been shown to inhibit atherosclerosis in several dyslipidemic mouse models. Previously we reported the NMR structure of Ac-18A-NH2, the base-line model class A amphipathic helical peptide in a 50% (v/v) trifluoroethanol-d3/water mixture, a membrane-mimic environment (Mishra, V. K., Palgunachari, M. N., Anantharamaiah, G. M., Jones, M. K., Segrest, J. P., and Krishna, N. R. (2001) Peptides 22, 567-573). The peptide Ac-18A-NH2 forms discoidal nascent high density lipoprotein-like particles with 1,2-dimyristoyl-sn-glycero-3-phosphocholine. Because subtle structural changes in the peptide.lipid complexes have been shown to be responsible for their antiatherogenic properties, we undertook high resolution NMR studies to deduce detailed structure of recombinant peptide.1,2-dimyristoyl-sn-glycero-3-phosphocholine complexes. The peptide adopts a well defined amphipathic alpha helical structure in association with the lipid at a 1:1 peptide:lipid weight ratio. Nuclear Overhauser effect spectroscopy revealed a number of intermolecular close contacts between the aromatic residues in the hydrophobic face of the helix and the lipid acyl chain protons. The pattern of observed peptide-lipid nuclear Overhauser effects is consistent with a parallel orientation of the amphipathic alpha helix, with respect to the plane of the lipid bilayer, on the edge of the disc (the belt model). Based on the results of chemical cross-linking and molecular modeling, we propose that peptide helices are arranged in a head to tail fashion to cover the edge of the disc. This arrangement of peptides is also consistent with the pKa values of the Lys residues determined previously. Taken together, these results provide for the first time a high resolution structural view of the peptide.lipid discoidal complexes formed by a class A amphipathic alpha helical peptide.

Expression of ABCG1, but Not ABCA1, Correlates with Cholesterol Release by Cerebellar Astroglia

Central nervous system lipoproteins mediate the exchange of cholesterol between cells and support synaptogenesis and neuronal growth. The primary source of lipoproteins in the brain is astroglia cells that synthesize and secrete apolipoprotein (apo) E in high density lipoprotein-like particles. Small quantities of apoA1, derived from the peripheral circulation, are also present in the brain. In addition to the direct secretion of apoE-containing lipoproteins from astroglia, glia-derived lipoproteins are thought to be formed by cholesterol efflux to extracellular apolipoproteins via ATP-binding cassette (ABC) transporters. We used cultured cerebellar murine astroglia to investigate the relationship among cholesterol availability, apoE secretion, expression of ABCA1 and ABCG1, and cholesterol efflux. In many cell types, cholesterol content, ABCA1 expression, and cholesterol efflux are closely correlated. In contrast, cholesterol enrichment of glia failed to increase ABCA1 expression, although ABCG1 expression and cholesterol efflux to apoA1 were increased. Moreover, the liver X receptor (LXR) agonist TO901317 up-regulated ABCA1 and ABCG1 expression in glia without stimulating cholesterol efflux. Larger lipoproteins were generated when glia were enriched with cholesterol, whereas treatment with the LXR agonist produced smaller particles that were eliminated when the glia were loaded with cholesterol. We also used glia from ApoE(-/-) mice to distinguish between direct lipoprotein secretion and the extracellular generation of lipoproteins. Our observations indicate that partially lipidated apoE, secreted directly by glia, is likely to be the major extracellular acceptor of cholesterol released from glia in a process mediated by ABCG1.

Alzheimer's disease amyloid-beta peptide modulates apolipoprotein E isoform specific receptor binding

The major protein component of the extracellular deposits in Alzheimer's disease (AD) is a 4 kDa peptide termed amyloid-beta (Abeta). This peptide is known to bind apolipoprotein E (apoE), a key mediator of lipoprotein transport, in an isoform specific manner. Whilst these isoform specific effects on apoE are well recognized, the functional significance of this interaction is poorly understood. Here, we investigated the influence of Abeta on apoE-mediated lipoprotein binding to cells using fluorescently tagged lipoprotein-like emulsions. Using this approach, we demonstrate that Abeta enhanced the normally poor binding of apoE2 lipoprotein-like particles to fibroblasts in culture, whilst markedly reducing the binding of apoE3 and apoE4. This suggests that the action of apoE isoforms on cellular lipoprotein or cholesterol metabolism is differentially modulated by Abeta. This also suggests that Abeta may also compromise apoE function in the Alzheimer disease affected brain.

Lipoprotein-like Particles and Cholesteryl Esters in Human Bruch’s Membrane: Initial Characterization

To isolate and characterize cholesteryl ester-containing, lipoprotein-like particles (LLPs) from normal aged human Bruch's membrane (BrM)/choroid (Ch). From BrM/Ch of 20 eyes of 10 donors aged >60 years, LLPs were released by high-salt buffer, fractionated by density gradient ultracentrifugation, and characterized by determining cholesterol, triglyceride, and phospholipid concentration (by enzymatic colorimetry and fluorometry); cholesteryl ester composition (by electrospray ionization mass spectrometry, ESI/MS); and particle morphology (by negative stain electron microscopy). Apolipoprotein (apo) gene expression was determined with RT-PCR, Western blot analysis, and immunofluorescence of retinal-choroidal cryosections. In paraformaldehyde-preserved eyes (20 eyes of 20 donors), cholesteryl ester composition of BrM/Ch, cornea, and sclera was determined by ESI/MS. A pooled fraction of LLP released from BrM/Ch (concentrated total LLP, density [d] < 1.24 g/mL fraction) was fractionated into two peaks. A large Peak 1 (with plasma LDL-HDL density range), containing predominantly phospholipid and unesterified cholesterol, was morphologically heterogeneous. A small Peak 2 (with plasma VLDL density range), enriched with esterified cholesterol, contained approximately 100 nm diameter round electron-lucent particles. Both peaks contained apoB and apoA-I, RPE and retina contained apoA-I mRNA transcripts, and BrM and drusen contained apoA-I immunoreactivity. Peaks 1 and 2, native RPE, and fresh BrM/Ch were cholesteryl linoleate enriched and contained little cholesteryl docosahexaenoate. Preserved BrM/Ch was cholesteryl oleate-enriched, unlike sclera and cornea. BrM/Ch LLP do not resemble plasma lipoproteins in density profile, cholesterol distribution, or morphology. Peak 2 contains EC-rich LLP resembling BrM particles in situ. BrM/Ch cholesteryl esters respond to long-term storage differently than esters of plasma lipoprotein origin accumulated in other ocular tissues. Evidence of intraocular apoB and apoA-I expression supports an emerging hypothesis that the RPE assembles and secretes a large, possibly novel, lipoprotein particle.

Disruption of cholesterol homeostasis in the developing brain as a potential mechanism contributing to the developmental neurotoxicity of ethanol: an hypothesis

While excess cholesterol may have deleterious consequences, as in the case of atherosclerosis, too little cholesterol may endanger the development of the brain. Different degrees of mental retardation are often observed in inborn errors of cholesterol synthesis, such as the Smith-Lemli-Opitz syndrome or in maternal phenylketonuria, where the metabolite of accumulating phenylalanine, phenylacetate, is an inhibitor of cholesterol synthesis. Lack of cholesterol during brain development as a consequence of these genetic defects leads to severe brain damage, microencephaly and mental retardation, which are also hallmarks of the fetal alcohol syndrome (FAS). The brain relies on the in situ synthesis of cholesterol, which occurs mostly in astrocytes. Astrocyte-produced cholesterol is utilized for cell proliferation, or is released, via astrocyte-secreted high density lipoprotein-like particles containing apolipoprotein E, outside the cell, where it is taken up and utilized by neurons for dendrite outgrowth and to form synapses. We propose the hypothesis that ethanol may disrupt cholesterol homeostasis during brain development, and that this effect may be responsible, at least in part, for the central nervous system dysfunctions observed in the FAS, which include altered astrocyte proliferation, neuronal death and diminished synaptic contacts.

Exocrine pancreatic secretion of phospholipid, menaquinone-4, and caveolin-1 in vivo

The exocrine pancreas releases secretory products essential for nutrient assimilation. In addition to digestive enzymes, the release of lipoprotein-like particles containing the membrane trafficking protein caveolin-1 from isolated pancreatic explants has been reported. The present study examined: (1) if gastrointestinal hormones induce the apical secretion of phospholipid in vivo and (2) a potential association of caveolin-1 and the lipid-soluble vitamin K analog menaquinone-4 (MK-4) with these structures. Analysis of isolated acinar cells, purified zymogen granules, and pancreatic juice collected in vivo indicated the presence a caveolin-1 immunoreactive protein that was acutely released in response hormone stimulation. Chloroform-extracted fractions of pancreatic juice also contained high concentrations of MK-4 that was secreted in parallel to protein and phospholipid. The presence of caveolin-1 and MK-4 in the phospholipid fraction of pancreatic juice places these molecules in the secretory pathway of exocrine cells and suggests a physiological role in digestive enzyme synthesis and/or processing.

Lipid formulation as a drug carrier for drug delivery.

In recent years, a Drug Delivery System (DDS), a preparative approach attracts the attention in the development of new drugs. DDS focuses on the regulation of the in vivo dynamics, such as absorption, distribution, metabolism, and elimination, thereby improving the effectiveness and the safety of the drugs by an applicable use of drug preparation technologies. A conventional intravenous dosage form of Amphotericin B (AmB), Fungizone, is the most effective clinically available for treating fungal infections. However, the clinical efficacy of AmB is limited by its adverse effects. Several lipid formulations, such as Liposomal AmB (L-AmB), AmB lipid complex (ABLC), and AmB colloidal dispersion (ABCD), with reduced side effects have been developed. These formulations are reported to have excellent safety and efficacy. However, comparable efficacy can be achieved only when they are administered at high doses than AmB. One of the problems of using these formulations is that they are easily taken up by the reticuloendothelial system (RES). An artificial lipoprotein-like particles, a novel drug carrier Lipid Nano-Sphere (LNS), which is 25 - 50 nm in size and is composed of phospholipids and simple lipid. LNS show a higher plasma concentration of drugs and lower uptake by RES-tissue different forms other lipid base drug carriers. In vitro and in vivo, LNS incorporating AmB, NS-718, shows reduced toxicity, while maintaining activity against fungi. LNS have a unique characteristic as an effective carrier of AmB for treatment of fungal infection.

Apolipoprotein E exhibits isoform-specific promotion of lipid efflux from astrocytes and neurons in culture.

Many studies have shown that apolipoprotein E (apoE) plays important roles in maintaining intracellular lipid homeostasis in nonneuronal cells. However, little is known about the extracellular transport of lipids in the CNS. In this study, we determined whether and to what degree lipid efflux from astrocytes and neurons depended on apoE. Our results showed that exogenously added apoE promoted the efflux of cholesterol and phosphatidylcholine from both astrocytes and neurons in culture, resulting in the generation of high-density lipoprotein-like particles. The order of potency of the apoE isoforms as lipid acceptors was apoE2 > apoE3 = apoE4 in astrocytes and apoE2 > apoE3 > apoE4 in neurons. Treatment with brefeldin A, monensin, and a protein kinase C inhibitor, H7, abolished the ability of apoE to promote cholesterol efflux from cultured astrocytes, without altering apoE-mediated phosphatidylcholine efflux. In contrast, the efflux of both cholesterol and phosphatidylcholine promoted by apoE was abolished following treatment with heparinase or lactoferrin, which block the interaction of apoE with heparan sulfate proteoglycans (HSPGs) or low-density lipoprotein receptor-related protein (LRP), respectively. This study suggests that apoE promotes lipid efflux from astrocytes and neurons in an isoform-specific manner and that cell surface HSPGs and/or HSPG-LRP pathway may mediate this apoE-promoted lipid efflux.

Anderson's disease: exclusion of apolipoprotein and intracellular lipid transport genes.

Anderson's disease is a rare, hereditary hypocholesterolemic syndrome characterized by chronic diarrhea, steatorrhea, and failure to thrive associated with the absence of apo B48-containing lipoproteins. To further define the molecular basis of the disease, we studied 8 affected subjects in 7 unrelated families of North African origin after treatment with a low-fat diet. Lipid loading of intestinal biopsies persisted, but the pattern and extent of loading was variable among the patients. Electron microscopy showed lipoprotein-like particles in membrane-bound compartments, the densities (0.65 to 7.5 particles/mu(2)) and the mean diameters (169 to 580 nm) of which were, in general, significantly larger than in a normal fed subject (0.66 particles/mu(2), 209 nm mean diameter). There were also large lipid particles having diameters up to 7043 nm (average diameters from 368 to 2127 nm) that were not surrounded by a membrane. Rarely, lipoprotein-like particles 50 to 150 nm in diameter were observed in the intercellular spaces. Intestinal organ culture showed that apo B and apo AIV were synthesized with apparently normal molecular weights and that small amounts were secreted in lipid-bound forms (density <1.006 g/mL). Normal microsomal triglyceride transfer protein (MTP) and activity were also detected in intestinal biopsies. Segregation analyses of 4 families excluded, as a cause of the disease, significant regions of the genome surrounding the genes for apo AI, AIV, B, CI, CII, CIII, and E, as were the genes encoding 3 proteins involved in intracellular lipid transport, MTP, and fatty acid binding proteins 1 and 2. The results suggest that a factor other than apoproteins and MTP are important for human intestinal chylomicron assembly and secretion.

Effect of plasminogen activators on human recombinant apolipoprotein(a) having the plasminogen activation cleavage site

The serine-proteinase domain in human apolipoprotein(a) [apo(a)] and plasminogen exhibit 89% sequence identity including the catalytic triad. Cleavage of the Arg 561-Val 562 activation site in plasminogen by either tissue- or urokinase-type plasminogen activator results in formation of the fibrinolytic enzyme plasmin. Apo(a) does not contain measurable amidolytic activity nor can it be activated by plasminogen activators. It has been suggested that the latter finding might be explained by the substitution of the plasminogen Arg-Val activation site by Ser-Ile in apo(a). To investigate if introduction of the Arg-Val activation site in apo(a) might result in sensitivity towards plasminogen activators, we expressed wild-type and Arg-Val mutant recombinant apo(a) [r-apo(a)] in human embryonic kidney and hepatocyte cell lines. Free r-apo(a) and lipoprotein-like particles [r-Lp(a)] were obtained in the culture supernatants of transfected 293 and HepG2 cells, respectively. Incubation of mutant r-apo(a)/r-Lp(a) with plasminogen activators produced neither plasmin-like activity nor cleavage at the Arg-Val activation site, even in the presence of various stimulators of plasminogen activation. Our data suggest that the high selectivity of activators for plasminogen activation requires interactions with regions in plasminogen distant from the activation disulfide loop which are not present in apo(a).

Disruption of the murine lecithin:cholesterol acyltransferase gene causes impairment of adrenal lipid delivery and up-regulation of scavenger receptor class B type I.

Lecithin:cholesterol acyltransferase (LCAT) is the major determinant of the cholesteryl ester (CE) content of high density lipoprotein (HDL) in plasma. The selective uptake of HDL-CE is postulated to participate in delivery of tissue-derived cholesterol both to the liver and steroidogenic tissues. Recent studies comparing mice with similarly low levels of HDL, due to the absence of either of the two major HDL-associated apolipoproteins apoA-I and apoA-II, suggest that apoA-I is crucial in modulating this process, possibly through interaction with scavenger receptor class B type I (SR-BI). Because of the central role of LCAT in determining the size, lipid composition, and plasma concentration of HDL, we have created LCAT-deficient mice by gene targeting to examine the effect of LCAT deficiency on HDL structure and composition and adrenal cholesterol delivery. The HDL in the LCAT-deficient mice was reduced in its plasma concentration (92%) and CE content (96%). The HDL particles were heterogeneous in size and morphology and included numerous discoidal particles, mimicking those observed in LCAT-deficient humans. The adrenals of the male Lcat (-/-) mice were severely depleted of lipid stores, which was associated with a 2-fold up-regulation of the adrenal SR-BI mRNA. These studies demonstrate that LCAT deficiency, similar to apoA-I deficiency, is associated with a marked decrease in adrenal cholesterol delivery and supports the hypothesis that adrenal SR-BI expression is regulated by the adrenal cholesterol.

The role of lipoprotein(a) in atherogenesis and thrombosis.

Lipoprotein(a) [Lp(a)] represents an important independent risk factor for atherosclerotic cardiovascular disease. Lp(a) constitutes a class of low-density lipoprotein-like particles that are structurally heterogeneous due to variability within the distinguishing apoprotein, apolipoprotein(a) [Apo(a)]. Apo(a) bears a high degree of homology to the fibrinolytic zymogen, plasminogen, the parent molecule of the serine protease plasmin. Apo(a) contains a variable number of tandemly repeated triple-loop units called kringles, which appear to mediate Lp(a)'s interactions with fibrin and cell surface receptors. Although the mechanism of its atherogenicity is unknown, Lp(a) has been implicated in the delivery of cholesterol to the injured blood vessel, in blockade of plasmin generation on fibrin and cell surfaces, and as a stimulus for smooth muscle cell proliferation. In addition, new members of the plasminogen/Apo(a) gene family have been defined, creating a potential link between Lp(a) and the control of angiogenesis in both health and disease. Pharmacologic therapy of elevated Lp(a) levels has been only modestly successful; apheresis remains the most effective therapeutic modality.

Concentration of intracellular hepatic apolipoprotein E in Golgi apparatus saccular distensions and endosomes.

The intrahepatic distribution of apolipoprotein E has been assessed by immunogold labeling of cryosections as well as by Western blotting of organelles isolated from liver homogenates. Both techniques supported the prior analytical fractionation studies of Wong (1989) who concluded that intrahepatic apoE was largely endosomal. All endosomal components decorated by gold particles indicative of apoE antigenicity in cryosections appeared filled with lipoprotein-like particles thereby accounting for this prominent morphological feature of isolated liver endosomes. The distribution of gold particles about the hepatic Golgi apparatus revealed a high content of apoE in closely apposed endosomes, ca. 400 nm in diameter, double labeled for apoE and internalized HRP. Remarkably, apoE (but not internalized HRP) was also observed within saccular distensions of all saccules of stacked Golgi cisternae but absent from the flattened saccular components as was also observed for apoB. This contrasted with albumin, the major secretory protein, which was uniformly distributed throughout the hepatic Golgi apparatus. These observations support a growing body of evidence for intra-Golgi sorting of secretory material in hepatic Golgi apparatus. The lack of any immunoreactive apoE or albumin in small 70-90 nm vesicles about the Golgi cisternae suggests limits to current models of vesicle-mediated intra-Golgi transport.