Catherine A. Reardon

Unique lipoproteins secreted by primary astrocytes from wild type, apoE (-/-), and human apoE transgenic mice.

Composition of central nervous system lipoproteins affects the metabolism of lipoprotein constituents within the brain. The ε4 allele of apolipoprotein E (apoE) is a risk factor for Alzheimer’s disease via an unknown mechanism(s). As glia are the primary central nervous system cell type that synthesize apoE, we characterized lipoproteins secreted by astrocytes from wild type (WT), apoE (−/−), and apoE transgenic mice expressing human apoE3 or apoE4 in a mouseapoE (−/−) background. Nondenaturing size exclusion chromatography demonstrates that WT, apoE3, and apoE4 astrocytes secrete particles the size of plasma high density lipoprotein (HDL) composed of phospholipid, free cholesterol, and protein, primarily apoE and apoJ. However, the lipid:apoE ratio of particles containing human apoE is significantly lower than WT. ApoE localizes across HDL-like particle sizes. ApoJ localizes to the smallest HDL-like particles.ApoE (−/−) astrocytes secrete little phospholipid or free cholesterol despite comparable apoJ expression, suggesting that apoE is required for normal secretion of astrocyte lipoproteins. Further, particles were not detected in apoE (−/−) samples by electron microscopy. Nondenaturing immunoprecipitation experiments indicate that apoE and apoJ reside predominantly on distinct particles. These studies suggest that apoE expression influences the unique structure of astrocyte lipoproteins, a process further modified by apoE species.

Apolipoprotein E Receptors Mediate the Effects of β-Amyloid on Astrocyte Cultures

We have previously shown that β-amyloid (Aβ) induces astrocyte activation in vitro and that this reaction is attenuated by the addition of exogenous apolipoprotein E (apoE)-containing particles. However, the effects of Aβ on endogenous apoE and apoJ levels and the potential role of apoE receptors in astrocyte activation have not been addressed. Three activating stimuli (lipopolysaccharide, dibutyryl cAMP, and aged Aβ 1–42) were used to induce activation of rat astrocyte cultures, as assessed by changes in morphology and an increase in interleukin-1β. However, only Aβ also induced ∼50% reduction in the amount of released apoE and apoJ and an 8-fold increase in the levels of cell-associated apoE and apoJ. Experiments using two concentrations of receptor-associated protein, an inhibitor of apoE receptors with a differential affinity for the low density lipoprotein receptor (LDLR) and the LDLR-related protein (LRP), suggest that LRP mediates Aβ-induced astrocyte activation, whereas LDLR mediates the Aβ-induced changes in apoE levels. Receptor-associated protein had no effect on apoJ levels or on activation by either dibutyryl cAMP or lipopolysaccharide. These data suggest that apoE receptors translate the presence of extracellular Aβ into cellular responses, both initiating and modulating the inflammatory response induced by Aβ.

Association of human, rat, and rabbit apolipoprotein E with β-amyloid

In humans, apolipoprotein E (apoE) has three major isoforms, E2 (Cys112, Cys158), E3 (Cys112, Arg158), and E4 (Arg112, Arg158). While ϵ4 is a genetic risk factor for Alzheimers disease (AD), ϵ2 may protect against late‐onset AD. Using native preparations of apoE from conditioned tissue culture media or plasma lipoproteins, we have previously shown that when equivalent amounts of apoE3 or E4 were incubated with β‐amyloid (Aβ), apoE3 formed 20 times as much SDS‐stable complex with the peptide as apoE4. This preferential binding of Aβ to apoE3 was abolished when apoE was purified by a process which includes delipidation and denaturation. Here we expand these observations to include Aβ binding to lipoprotein‐associated and purified apoE2. Lipoproteins isolated from the plasma of individuals homozygous for either ϵ2 or ϵ3 were incubated with Aβ(1‐40). SDS‐stable complex formation was analyzed by a non‐reducing gel shift assay, followed by immunoblotting with either Aβ or apoE antibodies. ApoE2:Aβ complex formation was comparable to apoE3:Aβ in both native and purified preparations of apoE. In addition, lipoprotein‐associated rat apoE (Arg112, Arg158), like human apoE4, did not form complex with Aβ, while lipoprotein‐associated rabbit apoE (Cys112, Arg158) did bind the peptide. These binding studies provide one possible explanation for protective effects of both apoE2 and E3 against the development of Alzheimers disease. J. Neurosci. Res. 49:9–18, 1997.

HDL apolipoprotein-related peptides in the treatment of atherosclerosis and other inflammatory disorders.

Elevations of HDL levels or modifying the inflammatory properties of HDL are being evaluated as possible treatment of atherosclerosis, the underlying mechanism responsible for most cardiovascular diseases. A promising approach is the use of small HDL apoprotein-related mimetic peptides. A number of peptides mimicking the repeating amphipathic α-helical structure in apoA-I, the major apoprotein in HDL, have been examined in vitro and in animal models. Several peptides have been shown to reduce early atherosclerotic lesions, but not more mature lesions unless coadministered with statins. These peptides also influence the vascular biology of the vessel wall and protect against other acute and chronic inflammatory diseases. The biologically active peptides are capable of reducing the pro-inflammatory properties of LDL and HDL, likely due to their high affinity for oxidized lipids. They are also capable of influencing other processes, including ABCA1 mediated activation of JAK-2 in macrophages, which may contribute to their anti-atherogenic function. The initial studies involved monomeric 18 amino acid peptides, but tandem peptides are being investigated for their anti-atherogenic and anti-inflammatory properties as they more closely resemble the repeating structure of apoA-I. Peptides based on other HDL associated proteins such as apoE, apoJ and SAA have also been studied. Their mechanism of action appears to be distinct from the apoA-I based mimetics.

Relationships of plasma and hepatic variables with rates of plasma low-density lipoprotein apolipoprotein B metabolism in baboons fed low- and high-fat diets

These studies were conducted to determine relationships of plasma low-density lipoprotein (LDL) cholesterol concentrations and hepatic mRNA levels for apolipoprotein (apo) B, LDL receptor, and hepatic hydroxymethyl glutaryl coenzyme A (HMG CoA) synthase with plasma LDL apo B production and catabolic rates in baboons maintained on a low-cholesterol, low-fat chow diet and on a high-cholesterol, high-fat (HCHF) diet. Twelve baboons with LDL cholesterol levels ranging from low to high on the HCHF diet but with similar high-density lipoprotein (HDL) cholesterol levels were selected from a colony of selectively bred pedigreed baboons. LDL apo B turnover and hepatic mRNA concentrations for apo B, LDL receptor, and HMG CoA synthase were measured on a chow diet and again on a HCHF diet fed for 14 weeks. LDL apo B fractional catabolic rates decreased and production rates increased on the HCHF diet. Hepatic mRNA concentrations for apo B were not affected by the HCHF diet. Hepatic LDL receptor and HMG CoA synthase mRNA concentrations decreased on the HCHF diet as compared with the chow diet. LDL apo B fractional catabolic rate was negatively correlated with plasma cholesterol, LDL cholesterol, LDL apo B, and LDL apo B production and positively correlated with hepatic LDL receptor and HMG CoA synthase mRNA concentrations and with plasma LDL triglyceride to cholesterol ratio on the chow diet but not on the HCHF diet. LDL apo B production was positively correlated with plasma cholesterol, LDL cholesterol, and LDL apo B on the HCHF diet and negatively correlated with LDL triglyceride to cholesterol ratio on both chow and HCHF diets.(ABSTRACT TRUNCATED AT 250 WORDS)

Biosynthesis of Rat Apolipoprotein E

Apolipoprotein E biosynthesis is of particular interest for several reasons. It is a major apolipoprotein of both primary lipoproteins secreted by the liver, namely VLDL and nascent HDL (Marsh 1976; Felker et al. 1977). Unlike the other major apolipoproteins which are made in both the liver and the intestine, significant amounts of apolipoprotein E do not appear to be made in the intestine (Wu and Windmueller 1979). Apolipoprotein E exhibits a genetically influenced polymorphism, which may have a relationship to certain forms of dyslipoproteinemia (Utermann et al. 1977). The plasma concentration of apolipoprotein E and its distribution amongst lipoprotein fractions is significantly modified by the feeding of cholesterol to several species, including rat (Mahley and Holcombe 1977; Weisgraber et al. 1977) and man (Mahley et al. 1978). The most pronounced changes are in the formation of B-VLDL, enriched in apolipoprotein E, and of relatively large quantities of HDLC, in which apolipoprotein E is the major apoprotein. The latter lipoprotein shows particular affinity for the low density lipoprotein receptor of fibroblasts in culture (Innerarity and Mahley 1978) and readily transports cholesterol into the cell, effecting the same regulation of intracellular cholesterol metabolism first described in association with the incorporation of LDL cholesterol into the cell (Innerarity and Mahley 1978; Goldstein and Brown 1977).

Natural killer cells in atherosclerosis

Publisher Summary Atherosclerosis is a chronic inflammatory disorder mediated by hyperlipidaemia. Cells of both the innate and adaptive immune system play important roles in atherogenesis. Natural killer (NK) cells have been detected in atherosclerotic lesions in humans and mice, although at low levels, but their role, if any, in atherogenesis is only beginning to be examined. Murine models of atherogenesis have been used to examine if NK cells contribute to atherogenesis via their cytotoxic properties and/or capacity to produce pro-atherogenic cytokines. Atherosclerosis is now well recognized to be a specific example of chronic inflammation induced, at least in mice and to some extent in humans, by hypercholesterolemia and probably oxidative modification of the levated cholesterol containing lipoproteins in the plasma. Cells of both the innate immune system and the adaptive immune system also play important roles in atherogenesis. Natural killer (NK) cells are among the immune cells that are of potential interest in relation to atherosclerosis. They have been found in small numbers in atherosclerotic plaques, produce cytokines that are known to influence atherogenesis and are present in the liver, which have important functions in the production and clearance of lipoprotein particles. It is evident that the role of NK cells, important elements of the innate immune response system, has received only modest attention in relation to atherosclerosis. These cells have the potential to affect both lipoprotein homeostasis, taking note of the relatively high content of NK cells in the liver, the major organ affecting lipoprotein metabolism, and influence the inflammatory response at the level of the vessel wall.