Ephraim Sehayek

Memory deficits and cholinergic impairments in apolipoprotein E-deficient mice ☆

Apolipoprotein E-deficient mice provide a useful system for studying the role of apolipoprotein E (apoE) in the function of distinct neuronal systems. In the present study we focused on the cholinergic system of these mice. This was pursued by measurements of specific biochemical, physiological and cognitive parameters. Morris Water Maze tasks revealed impairments in working memory but not in reference memory of the apoE-deficient mice. Measurements of brain choline acetyltransferase activities revealed them to be markedly lower in the hippocampus and frontal cortex of the apoE-deficient mice than in the corresponding brain areas of the controls, but unaltered in other brain areas. In addition, hypothermia induced by the centrally acting muscarinic agonist, oxotremorine, was reduced in the apoE-deficient mice as compared to controls. These results show that apoE-deficient mice have cholinergic deficits and highlight the importance of this mouse model for studying the interactions between apoE and the cholinergic nervous system.

Significance of High HbA1 Levels in Normal Glucose Tolerance

The significance of high hemoglobin A1 (HbA1) levels (≥8.0%) found in 12.1% of 648 individuals with normal glucose tolerance constituting a part of a representative population sample was examined. Measurement error in HbA1 and/or glucose-tolerance levels was precluded by HbA1 remaining in the same range over 3.5 yr in 89.7% of 29 individuals with initially high and 68.1% of 22 individuals with initially low (<6.5%) HbA1. Rate of deterioration to glucose intolerance (6.9%) in the high group during that period resembled the rate (11.8%) in a control group (n = 279). Fasting plasma glucose significantly accounted for only 2.4% of total HbA1 - population variance. No correlation of HbA1 was found with other correlates of glucose tolerance or with daily caloric intake and physical activity. A small but significant increment in HbA1 was associated with smoking (7.1 vs. 6.8%, P < .01) and with clinically overt atherosclerosis (7.3 vs. 6.9%, P < .01). We conclude that factors unrelated to glucose metabolism are the main determinants of HbA1 level in normal glucose tolerance and play an important role in diabetes as well. These factors have bearing on evaluation of diabetic control by HbA1 and possibly on risk for diabetic complications.

Phosphorylation of tau in apolipoprotein E-deficient mice

It has been suggested that the deleterious effects of the allele E4 of apolipoprotein E (apoE) in Alzheimers disease (AD) are related to its inability to interact with the microtubule associated protein tau and to thereby prevent its hyperphosphorylation. In the present study we investigated the effects of apoE on tau phosphorylation by immunoblot analysis of the levels and extents of phosphorylation of tau of apoE-deficient mice. This revealed that mAb AT8, which is directed against a phosphorylated tau epitope, labels tau of the apoE-deficient mice more intensely than that of control mice and that the opposite occurs with mAb Tau1, which is directed against dephosphorylated tau epitopes. mAb ALZ50 also labeled the tau enriched preparations of the apoE-deficient mice more intensely than those of the controls, whereas the extents of their labeling by the phosphorylation insensitive anti-tau mAb 134 were similar. These results suggest that tau of apoE-deficient mice is hyperphosphorylated.

Biochemical and cognitive studies of apolipoprotein-E-deficient mice

Apolipoprotein-E-deficient mice provide a useful model system for studying the role of apolipoprotein E (apoE) in brain function. In the present study, we characterization the cholinergic function of these mice and the extent of phosphorylation of their cytoskeletal protein τ. Morris water maze tasks revealed deficits in working memory that were accompanied by a specific decrease in hippocampal and cortical choline acetyltransferase activities. Immunoblot experiments utilizing native and dephosphorylated τ and antibodies directed against specific phosphorylated and unphosphorylated τ epitopes revealed that τ of the apoE-deficient mice is hyperphosphorylated. These results show that apoE-deficient mice have cognitive cholinergic and cytoskeletal derangements and point out the importance of this model for studying the role of apoE in neuronal function.

Derangement in stress response of apolipoprotein E-deficient mice☆

Apolipoprotein E (apoE) is associated with familial and sporadic Alzheimers disease (AD). Stress has been identified as a putative risk factor of AD. Thus, in the present study we examined the susceptibility of apoE-deficient mice to stress. The results obtained revealed that the elevation of corticosterone levels in apoE-deficient mice following restraint stress is markedly lower than in controls, and that these mice differ in their behavioral pain response to noxious stimuli in both stress and non-stress conditions. These findings suggest an interplay between apoE and the response to stressful stimuli and provide a model for elucidating the relationship between apoE and susceptibility to stress.

Binding to heparan sulfate is a major event during catabolism of lipoprotein lipase by HepG2 and other cell cultures

Lipoprotein lipase (LPL) is rapidly and efficiently cleared from the circulation by the liver through an as yet unclear mechanism. In the present study, we determined the nature of LPL interactions with the liver parenchimal cell line HepG2 as compared to other cells in culture. Binding, cell association and degradation of 125I-labelled bovine milk LPL by HepG2 cells, normal and low density lipoprotein (LDL) receptor-negative human fibroblasts and Chinese hamster ovary (CHO) cells show similar values irrespective of source and origin. LPL metabolism in HepG2 cells was characterized by a high capacity to degrade the enzyme, an extremely high sensitivity to heparin and was inhibited by 60%-70% after treatment of the cells with sodium chlorate and heparinase (but not chondroitinase). These findings suggested an important role for heparan sulfate in the process of cell interaction and metabolism of LPL. To further clarify the role of heparan sulfate in determining the LPL-cell interactions, we compared the metabolism of LPL in wild type and mutant heparan sulfate-deficient CHO cells. Heparan sulfate-deficient CHO cells show a low capacity to bind and degrade LPL, about 10%-20% that of the wild type cells. In another set of experiments, we sought to determine whether LPL interactions with HepG2 cells are affected by triglyceride-rich lipoproteins. The results clearly show that whereas unlabeled LPL dramatically enhanced the metabolism of radioiodinated very low density lipoprotein (VLDL), unlabeled VLDL had no effect on radioiodinated LPL metabolism in these cells.(ABSTRACT TRUNCATED AT 250 WORDS)

The role of native apolipoprotein B-containing lipoproteins in atherosclerosis: cellular mechanisms

Apolipoprotein B-containing lipoproteins play a central role in the pathogenesis of atherosclerosis. The processes that determine the atherogenicity of these lipoproteins, however, remain unclear. A new development that involves the interactions of lipoproteins, enzymes, apolipoproteins, and cell-surface heparan sulphate may explain, at least in part, the enhanced atherogenicity of these lipoproteins. In addition, the development of specific transgenic apolipoproteins, and apolipoprotein-deficient mice models that develop atherosclerosis, provide new tools for studies on the role of native apolipoprotein-B containing lipoproteins in atheroma formation.

A synthetic heparin-mimicking polyanionic compound binds to the LDL receptor-related protein and inhibits vascular smooth muscle cell proliferation.

A synthetic heparin‐mimicking polyaromatic anionic compound RG‐13577 (polymer of 4‐hydroxyphenoxy acetic acid and formaldehyde ammonium salt, Mr∼5800) exhibits specific binding to vascular smooth muscle cells (SMCs) and inhibits their proliferative response to growth promoting factors. Receptor binding of 14C‐RG‐13577 was efficiently competed by apolipoprotein E3 (apoE), lactoferrin, and the LRP (LDL receptor‐related protein) receptor associated 39 kDa protein (RAP). Unlike cell surface binding of apoE, binding of RG‐13577 to SMCs was not affected by heparin, heparan sulfate degrading enzymes, or low density lipoprotein (LDL). Moreover, wild‐type and heparan sulfate‐deficient Chinese hamster ovary (CHO) cells, as well as normal‐ and LDL receptor negative‐ human skin fibroblasts bind RG‐13577, but not apoE, to a similar extent. On the other hand, homozygous mouse embryonic fibroblasts deficient in the LDL receptor‐related protein (LRP) expressed a markedly reduced binding of RG‐13577 as compared to normal mouse embryonic fibroblasts. These results indicate that RG‐13577 and related compounds bind to the LRP receptor on the surface of vascular SMCs. Addition of lactoferrin to cultured SMCs protected the cells against the antiproliferative effect of compound RG‐13577, suggesting that this inhibition is mediated by RG‐13577 binding to LRP receptors on the SMC surface. Altogether, we have identified a series of synthetic polyaromatic anionic molecules that exhibit specific binding to LRP and therby exert an antiproliferative effect on vascular SMCs. These compounds are applied to suppress SMC proliferation associated with restenosis and accelerated atherosclerosis. J. Cell. Biochem. 81:114–127, 2001.