Although amyloid ␤ (A␤) oligomers are presumed to cause synaptic and cognitive dysfunction in Alzheimer's disease (AD), their contribution to other pathological features of AD remains unclear. To address the latter, we generated APP transgenic mice expressing the E693⌬ mutation, which causes AD by enhanced A␤ oligomerization without fibrillization. The mice displayed age-dependent accumulation of intraneuronal A␤ oligomers from 8 months but no extracellular amyloid deposits even at 24 months. Hippocampal synaptic plasticity and memory were impaired at 8 months, at which time the presynaptic marker synaptophysin began to decrease. Furthermore, we detected abnormal tau phosphorylation from 8 months, microglial activation from 12 months, astrocyte activation from 18 months, and neuronal loss at 24 months. These findings suggest that A␤ oligomers cause not only synaptic alteration but also other features of AD pathology and that these mice are a useful model of A␤ oligomer-induced pathology in the absence of amyloid plaques.

Apolipoprotein E (apoE) is a major apolipoprotein in the brain. The ⑀4 allele of apoE is a major risk factor for Alzheimer disease, and apoE deficiency in mice leads to blood-brain barrier (BBB) leakage. However, the effect of apoE isoforms on BBB properties are as yet unknown. Here, using an in vitro BBB model consisting of brain endothelial cells and pericytes prepared from wild-type (WT) mice, and primary astrocytes prepared from human apoE3-and apoE4-knock-in mice, we show that the barrier function of tight junctions (TJs) was impaired when the BBB was reconstituted with primary astrocytes from apoE4-knock-in mice (apoE4-BBB model). The phosphorylation of occludin at Thr residues and the activation of protein kinase C (PKC) in mBECs were attenuated in the apoE4-BBB model compared with those in the apoE3-BBB model. The differential effects of apoE isoforms on the activation of PKC, the phosphorylation of occludin at Thr residues, and TJ integrity were abolished following the treatment with an anti-low density lipoprotein receptor-related protein 1 (LRP1) antibody or a LRP1 antagonist receptor-associated protein. Consistent with the results of in vitro studies, BBB permeability was higher in apoE4-knock-in mice than in apoE3-knock-in mice. Our studies provide evidence that TJ integrity in BBB is regulated by apoE in an isoform-dependent manner. Apolipoprotein E (apoE)2 is a polymorphic glycoprotein with a molecular mass of 34 kDa. Its three isoforms, apoE2, apoE3, and apoE4, are all products of the same gene, which exists as three alleles (⑀2, ⑀3, and ⑀4) at a single locus (1). Among these three isoforms, apoE4 is a major risk factor for Alzheimer disease (AD) (2, 3). ApoE is expressed in several organs, with the liver showing the highest expression level, followed by the brain. In the brain, apoE is a major apolipoprotein and plays a major role in the transportation of lipids as a lipid acceptor (1). ApoE-containing lipoprotein particles are mainly produced by astrocytes and deliver cholesterol and other essential lipids to neurons through low density lipoprotein (LDL) receptor family members (4 -6). A number of studies revealed that astrocytes are involved in the control of endothelium blood-brain barrier (BBB) properties (7,8) and that apoE deficiency leads to BBB leakage (9 -11).BBB is formed by brain endothelial cells and is essential for the protection of the central nervous system from harmful blood molecules and cells (12). Brain endothelial cells form tight junctions (TJs), which are the fundamental characteristics of BBB (13,14). The assembly of TJs requires at least three types of transmembrane protein, namely, occludin, claudin, and junctional adhesion molecule (15). Protein kinases are localized at TJs or interact directly with TJ proteins (16 -18). Among protein kinases, PKC has been shown to regulate the phosphorylation of occludin at its Thr residues and play a crucial role in the assembly and/or maintenance of TJs (19). Cells surrounding brain capillaries, such as astrocytes and pericytes, con...

The E693Delta mutation within the amyloid precursor protein (APP) has been suggested to cause dementia via the enhanced formation of synaptotoxic amyloid beta (Abeta) oligomers. However, this mutation markedly decreases Abeta secretion, implying the existence of an additional mechanism of neuronal dysfunction that is independent of extracellular Abeta. We therefore examined the effects of this mutation on both APP processing to produce Abeta as well as subcellular localization and accumulation of Abeta in transfected HEK293 and COS-7 cells. Both beta- and gamma-cleavage of mutant APP increased, indicating a lack of inhibition in Abeta production. Instead, this mutation promoted Abeta accumulation within cells, including the endoplasmic reticulum (ER), Golgi apparatus, early and late endosomes, lysosomes, and autophagosomes, all of which have been proposed as intracellular sites of Abeta generation and/or degradation, suggesting impairment of APP/Abeta trafficking. Notably, the intracellular mutant Abeta was found to predominantly form oligomers. Concomitant with this accumulation, the ER stress markers Grp78 and phosphorylated eIF2alpha were both strongly induced. Furthermore, the activation of caspase-4 and -3 as well as DNA fragmentation were detected in these cells. These results suggest that mutant Abeta induces alteration of Abeta trafficking and subsequent ER stress-induced apoptosis via enhancement of its intracellular oligomerization. Our findings suggest that Abeta oligomers exhibit toxicity in the extracellular space and within the cells themselves.

Male Tsumura Suzuki obese diabetes (TSOD) mice spontaneously develop obesity and obesity-related metabolic syndrome. Gut dysbiosis, an imbalance of gut microbiota, has been implicated in the pathogenesis of metabolic syndrome, but its mechanisms are unknown. Short-chain fatty acids (SCFAs) are the main fermentation products of gut microbiota and a link between the gut microbiota and the host’s physiology. Here, we investigated a correlation among gut dysbiosis, SCFAs, and metabolic syndrome in TSOD mice. We detected enriched levels of Gram-positive bacteria and corresponding decreases in Gram-negative bacteria in 24-wk-old metabolic syndrome-affected TSOD mice compared with age-matched controls. The abundance of Bacteroidetes species decreased, the abundance of Firmicutes species increased, and nine genera of bacteria were altered in 24-wk-old TSOD mice. The total plasma SCFA level was significantly lower in the TSOD mice than in controls. The major plasma SCFA—acetate—decreased in TSOD mice, whereas propionate and butyrate increased. TSOD mice had no minor SCFAs (valerate and hexanoate) but normal mice did. We thus concluded that gut dysbiosis and consequent disruptions in plasma SCFA profiles occurred in metabolic syndrome-affected TSOD mice. We also propose that the TSOD mouse is a useful model to study gut dysbiosis, SCFAs, and metabolic syndrome.

We previously showed that male Tsumura Suzuki obese diabetes (TSOD) mice, a spontaneous mouse model of metabolic syndrome, manifested gut dysbiosis and subsequent disruption of the type and quantity of plasma short-chain fatty acids (SCFAs), and daily coffee intake prevented nonalcoholic steatohepatitis in this mouse model. Here, we present a preliminary study on whether coffee and its major components, caffeine and chlorogenic acid, would affect the gut dysbiosis and the disrupted plasma SCFA profile of TSOD mice, which could lead to improvement in the liver pathology of these mice. Three mice per group were used. Daily intake of coffee or its components for 16 wk prevented liver lobular inflammation without improving obesity in TSOD mice. Coffee and its components did not repair the altered levels of Gram-positive and Gram-negative bacteria and an increased abundance of Firmicutes in TSOD mice but rather caused additional changes in bacteria in six genera. However, caffeine and chlorogenic acid partially improved the disrupted plasma SCFA profile in TSOD mice, although coffee had no effects. Whether these alterations in the gut microbiome and the plasma SCFA profile might affect the liver pathology of TSOD mice may deserve further investigation.

Lipoprotein lipase (LPL) is a member of a lipase family known to hydrolyze triglyceride molecules in plasma lipoprotein particles. LPL also plays a role in the binding of lipoprotein particles to cell-surface molecules, including sulfated glycosaminoglycans (GAGs). LPL is predominantly expressed in adipose and muscle but is also highly expressed in the brain where its specific roles are unknown. It has been shown that LPL is colocalized with senile plaques in Alzheimer disease (AD) brains, and its mutations are associated with the severity of AD pathophysiological features. In this study, we identified a novel function of LPL; that is, LPL binds to amyloid ␤ protein (A␤) and promotes cell-surface association and uptake of A␤ in mouse primary astrocytes. The internalized A␤ was degraded within 12 h, mainly in a lysosomal pathway. We also found that sulfated GAGs were involved in the LPL-mediated cellular uptake of A␤. Apolipoprotein E was dispensable in the LPL-mediated uptake of A␤. Our findings indicate that LPL is a novel A␤-binding protein promoting cellular uptake and subsequent degradation of A␤. Lipoprotein lipase (LPL)2 catalyzes the hydrolysis of triacylglycerol and mediates the cellular uptake of lipoproteins by functioning as a "bridging molecule" between lipoproteins and sulfated glycosaminoglycans (GAGs) or lipoprotein receptors in blood vessels (1, 2). Sulfated GAGs are side chains of proteoglycans normally found in the extracellular matrix and on the cell surface in the peripheral tissues and brain. Sulfation modifications vary within the GAG chains and are crucial for interaction between GAGs and various protein ligands (3), including LPL (4, 5).It has been shown that LPL is distributed in numerous organs and is highly expressed in the brain (6, 7). Although the catabolic activity of LPL on triacylglycerol is observed in the brain (8), the finding that apolipoprotein CII (apoCII), an essential cofactor for LPL, is not expressed in the brain (9, 10), suggests that LPL has a novel nonenzymatic function in the brain. However, little is known about LPL function in the brain. Interestingly, it has been shown that LPL is accumulated in senile plaques of Alzheimer disease (AD) brains (11). Moreover, SNPs in the coding region of the LPL gene are associated with disease incidence in clinically diagnosed AD subjects, LPL mRNA expression level, brain cholesterol level, and the severity of AD pathologies, including neurofibrillary tangles and senile plaque density (12). These results suggest that LPL may have a physiological role in the brain, whose alternation is associated with the pathogenesis of AD.The occurrence of senile plaques in the brain is one of the pathological hallmarks of AD. They contain extracellular deposits of amyloid ␤ protein (A␤), and the abnormal A␤ deposition or the formation of soluble A␤ oligomers is crucial for AD pathogenesis. A␤ is a physiological peptide whose main species are 40 and 42 amino acids in length, and A␤42 is the predominant specie in senile plaques (13). The A␤ level...

We previously showed that microglial keratan sulfate (KS) was induced in amyotrophic lateral sclerosis. However, the functional roles of the glycan and its synthetic enzyme in neurodegenerative diseases, such as Alzheimer's disease (AD), a progressive disorder, are unclear. In our study, KS modified with sialic acids having a molecular mass of 125-220 kDa and the carbohydrate sulfotransferase GlcNAc6ST1 were up-regulated in the brains of two transgenic mouse models (J20 and Tg2576) and the brains of patients with AD. GlcNAc6ST1-deficient J20 (J20/GlcNAc6ST1) mice demonstrated a complete absence of the microglial sialylated KS. J20/GlcNAc6ST1 primary microglia showed an increased level of amyloid-β phagocytosis and were hyperresponsive to interleukin 4, a potent antiinflammatory cytokine. Moreover, J20/GlcNAc6ST1 mice manifested reduced cerebral amyloid-β deposition. GlcNAc6ST1-synthesizing sialylated KS thus modulates AD pathology. Inhibition of KS synthesis by targeting GlcNAc6ST1 may therefore be beneficial for controlling AD pathogenesis.

Background:The G26R apolipoprotein A-I (apoA-I Iowa ) mutation causes familial amyloid polyneuropathy. Results: ApoA-I Iowa amyloid cellular interaction and cytotoxicity depended on cell surface heparan sulfate (HS). Enzymatic remodeling of HS by extracellular sulfatase mitigated cytotoxicity. Conclusion: Sulfate moieties of cell surface HS are critical for mediating apoA-I amyloid cytotoxicity. Significance: Enzymatic remodeling of HS may be a novel concept for regulating actions of amyloid on cells.