Kenjiro Ono

Curcumin has potent anti‐amyloidogenic effects for Alzheimer's β‐amyloid fibrils in vitro

Inhibition of the accumulation of amyloid β‐peptide (Aβ) and the formation of β‐amyloid fibrils (fAβ) from Aβ, as well as the destabilization of preformed fAβ in the central nervous system, would be attractive therapeutic targets for the treatment of Alzheimers disease (AD). We reported previously that nordihydroguaiaretic acid (NDGA) and wine‐related polyphenols inhibit fAβ formation from Aβ(1–40) and Aβ(1–42) and destabilize preformed fAβ(1–40) and fAβ(1–42) dose‐dependently in vitro. Using fluorescence spectroscopic analysis with thioflavin T and electron microscopic studies, we examined the effects of curcumin (Cur) and rosmarinic acid (RA) on the formation, extension, and destabilization of fAβ(1–40) and fAβ(1–42) at pH 7.5 at 37°C in vitro. We next compared the anti‐amyloidogenic activities of Cur and RA with NDGA. Cur and RA dose‐dependently inhibited fAβ formation from Aβ(1–40) and Aβ(1–42), as well as their extension. In addition, they dose‐dependently destabilized preformed fAβs. The overall activities of Cur, RA, and NDGA were similar. The effective concentrations (EC50) of Cur, RA, and NDGA for the formation, extension, and destabilization of fAβs were in the order of 0.1–1 μM. Although the mechanism by which Cur and RA inhibit fAβ formation from Aβ and destabilize preformed fAβ in vitro remains unclear, they could be a key molecule for the development of therapeutics for AD.

Potent anti-amyloidogenic and fibril-destabilizing effects of polyphenols in vitro: implications for the prevention and therapeutics of Alzheimer's disease.

Cerebral deposition of amyloid β‐peptide (Aβ) in the brain is an invariant feature of Alzheimers disease (AD). A consistent protective effect of wine consumption on AD has been documented by epidemiological studies. In the present study, we used fluorescence spectroscopy with thioflavin T and electron microscopy to examine the effects of wine‐related polyphenols (myricetin, morin, quercetin, kaempferol (+)‐catechin and (–)‐epicatechin) on the formation, extension, and destabilization of β‐amyloid fibrils (fAβ) at pH 7.5 at 37°C in vitro. All examined polyphenols dose‐dependently inhibited formation of fAβ from fresh Aβ(1–40) and Aβ(1–42), as well as their extension. Moreover, these polyphenols dose‐dependently destabilized preformed fAβs. The overall activity of the molecules examined was in the order of: myricetin = morin = quercetin > kaempferol > (+)‐catechin = (–)‐epicatechin. The effective concentrations (EC50) of myricetin, morin and quercetin for the formation, extension and destabilization of fAβs were in the order of 0.1–1 µm. In cell culture experiments, myricetin‐treated fAβ were suggested to be less toxic than intact fAβ, as demonstrated by 3‐[4,5‐dimethylthiazol‐2‐yl]‐2,5‐diphenyltetrazolium bromide assay. Although the mechanisms by which these polyphenols inhibit fAβ formation from Aβ, and destabilize pre‐formed fAβin vitro are still unclear, polyphenols could be a key molecule for the development of preventives and therapeutics for AD.

Structure–neurotoxicity relationships of amyloid β-protein oligomers

Amyloid β-protein (Aβ) oligomers may be the proximate neurotoxins in Alzheimers disease (AD). “Oligomer” is an ill-defined term because many kinds have been reported and they often exist in rapid equilibrium with monomers and higher-order assemblies. We report here results of studies in which specific oligomers have been stabilized structurally, fractionated in pure form, and then studied by using a combination of CD spectroscopy, Thioflavin T fluorescence, EM, atomic force microscopy (AFM), and neurotoxicity assays. Aβ monomers were largely unstructured, but oligomers exhibited order-dependent increases in β-sheet content. EM and AFM data suggest that dimerization and subsequent monomer addition are processes in which significant and asymmetric monomer conformational changes occur. Oligomer secondary structure and order correlated directly with fibril nucleation activity. Neurotoxic activity increased disproportionately (order dependence >1) with oligomer order. The structure–activity correlations reported here significantly extend our understanding of the conformational dynamics, structure, and relative toxicity of pure Aβ oligomers of specific order.

Grape-derived polyphenolics prevent Aβ oligomerization and attenuate cognitive deterioration in a mouse model of Alzheimer's disease

Alzheimers disease (AD) is a neurodegenerative disorder characterized by progressive impairments in memory and cognition. Extracellular accumulation of soluble high-molecular-weight (HMW) Aβ oligomers has been proposed to be largely responsible for AD dementia and memory deficits in the Tg2576 mice, a model of AD. In this study, we found that a naturally derived grape seed polyphenolic extract can significantly inhibit amyloid β-protein aggregation into high-molecular-weight oligomers in vitro. When orally administered to Tg2576 mice, this polyphenolic preparation significantly attenuates AD-type cognitive deterioration coincidentally with reduced HMW soluble oligomeric Aβ in the brain. Our study suggests that grape seed-derived polyphenolics may be useful agents to prevent or treat AD.

REVIEW: Curcumin and Alzheimer's Disease

Curcumin has a long history of use as a traditional remedy and food in Asia. Many studies have reported that curcumin has various beneficial properties, such as antioxidant, antiinflammatory, and antitumor. Because of the reported effects of curcumin on tumors, many clinical trials have been performed to elucidate curcumins effects on cancers. Recent reports have suggested therapeutic potential of curcumin in the pathophysiology of Alzheimers disease (AD). In in vitro studies, curcumin has been reported to inhibit amyloid‐β‐protein (Aβ) aggregation, and Aβ‐induced inflammation, as well as the activities of β‐secretase and acetylcholinesterase. In in vivo studies, oral administration of curcumin has resulted in the inhibition of Aβ deposition, Aβ oligomerization, and tau phosphorylation in the brains of AD animal models, and improvements in behavioral impairment in animal models. These findings suggest that curcumin might be one of the most promising compounds for the development of AD therapies. At present, four clinical trials concerning the effects of curcumin on AD has been conducted. Two of them that were performed in China and USA have been reported no significant differences in changes in cognitive function between placebo and curcumin groups, and no results have been reported from two other clinical studies. Additional trials are necessary to determine the clinical usefulness of curcumin in the prevention and treatment of AD.

Phenolic Compounds Prevent Alzheimer’s Pathology through Different Effects on the Amyloid-β Aggregation Pathway

Inhibition of amyloid-beta (Abeta) aggregation is an attractive therapeutic strategy for Alzheimers disease (AD). Certain phenolic compounds have been reported to have anti-Abeta aggregation effects in vitro. This study systematically investigated the effects of phenolic compounds on AD model transgenic mice (Tg2576). Mice were fed five phenolic compounds (curcumin, ferulic acid, myricetin, nordihydroguaiaretic acid (NDGA), and rosmarinic acid (RA)) for 10 months from the age of 5 months. Immunohistochemically, in both the NDGA- and RA-treated groups, Abeta deposition was significantly decreased in the brain (P < 0.05). In the RA-treated group, the level of Tris-buffered saline (TBS)-soluble Abeta monomers was increased (P < 0.01), whereas that of oligomers, as probed with the A11 antibody (A11-positive oligomers), was decreased (P < 0.001). However, in the NDGA-treated group, the abundance of A11-positive oligomers was increased (P < 0.05) without any change in the levels of TBS-soluble or TBS-insoluble Abeta. In the curcumin- and myricetin-treated groups, changes in the Abeta profile were similar to those in the RA-treated group, but Abeta plaque deposition was not significantly decreased. In the ferulic acid-treated group, there was no significant difference in the Abeta profile. These results showed that oral administration of phenolic compounds prevented the development of AD pathology by affecting different Abeta aggregation pathways in vivo. Clinical trials with these compounds are necessary to confirm the anti-AD effects and safety in humans.

Antioxidant compounds have potent anti-fibrillogenic and fibril-destabilizing effects for α-synuclein fibrils in vitro

The aggregation of α‐synuclein (αS) in the brain has been implicated as a critical step in the development of Lewy body diseases (LBD) and multiple system atrophy (MSA). Various antioxidants not only inhibit the formation of β‐amyloid fibrils (fAβ), but also destabilize preformed fAb in vitro. Using fluorescence spectroscopy with thioflavin S and electron microscopy, here we examined the effects of the antioxidants nordihydroguaiaretic acid, curcumin, rosmarinic acid, ferulic acid, wine‐related polyphenols [tannic acid, myricetin, kaempferol (+)‐catechin and (–)‐epicatechin], docosahexaenoic acid, eicosapentaenoic acid, rifampicin and tetracycline on the formation of αS fibrils (fαS) and on preformed fαS. All molecules, except for docosahexaenoic acid and eicosapentaenoic acid, dose‐dependently inhibited the formation of fαS. Moreover, these molecules dose‐dependently destabilized preformed fαS. The overall activity of the molecules examined was in the order of: tannic acid = nordihydroguaiaretic acid = curcumin = rosmarinic acid = myricetin > kaempferol = ferulic acid > (+)‐catechin = (–)‐epicatechin > rifampicin = tetracycline. These compounds with anti‐fibrillogenic as well as antioxidant activities could be key molecules for the development of preventives and therapeutics for LBD and MSA as well as Alzheimers disease.

Effects of Grape Seed-derived Polyphenols on Amyloid β-Protein Self-assembly and Cytotoxicity

Epidemiological evidence suggests that moderate consumption of red wine reduces the incidence of Alzheimer disease (AD). To study the protective effects of red wine, experiments recently were executed in the Tg2576 mouse model of AD. These studies showed that a commercially available grape seed polyphenolic extract, MegaNatural-AZ (MN), significantly attenuated AD-type cognitive deterioration and reduced cerebral amyloid deposition (Wang, J., Ho, L., Zhao, W., Ono, K., Rosensweig, C., Chen, L., Humala, N., Teplow, D. B., and Pasinetti, G. M. (2008) J. Neurosci. 28, 6388–6392). To elucidate the mechanistic bases for these observations, here we used CD spectroscopy, photo-induced cross-linking of unmodified proteins, thioflavin T fluorescence, size exclusion chromatography, and electron microscopy to examine the effects of MN on the assembly of the two predominant disease-related amyloid β-protein alloforms, Aβ40 and Aβ42. We also examined the effects of MN on Aβ-induced cytotoxicity by assaying 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide metabolism and lactate dehydrogenase activity in Aβ-treated, differentiated pheochromocytoma (PC12) cells. Initial studies revealed that MN blocked Aβ fibril formation. Subsequent evaluation of the assembly stage specificity of the effect showed that MN was able to inhibit protofibril formation, pre-protofibrillar oligomerization, and initial coil → α-helix/β-sheet secondary structure transitions. Importantly, MN had protective effects in assays of cytotoxicity in which MN was mixed with Aβ prior to peptide assembly or following assembly and just prior to peptide addition to cells. These data suggest that MN is worthy of consideration as a therapeutic agent for AD.

Brain-Targeted Proanthocyanidin Metabolites for Alzheimer's Disease Treatment

While polyphenolic compounds have many health benefits, the potential development of polyphenols for the prevention/treatment of neurological disorders is largely hindered by their complexity as well as by limited knowledge regarding their bioavailability, metabolism, and bioactivity, especially in the brain. We recently demonstrated that dietary supplementation with a specific grape-derived polyphenolic preparation (GP) significantly improves cognitive function in a mouse model of Alzheimers disease (AD). GP is comprised of the proanthocyanidin (PAC) catechin and epicatechin in monomeric (Mo), oligomeric, and polymeric forms. In this study, we report that following oral administration of the independent GP forms, only Mo is able to improve cognitive function and only Mo metabolites can selectively reach and accumulate in the brain at a concentration of ∼400 nm. Most importantly, we report for the first time that a biosynthetic epicatechin metabolite, 3′-O-methyl-epicatechin-5-O-β-glucuronide (3′-O-Me-EC-Gluc), one of the PAC metabolites identified in the brain following Mo treatment, promotes basal synaptic transmission and long-term potentiation at physiologically relevant concentrations in hippocampus slices through mechanisms associated with cAMP response element binding protein (CREB) signaling. Our studies suggest that select brain-targeted PAC metabolites benefit cognition by improving synaptic plasticity in the brain, and provide impetus to develop 3′-O-Me-EC-Gluc and other brain-targeted PAC metabolites to promote learning and memory in AD and other forms of dementia.

Nicotine breaks down preformed Alzheimer’s β-amyloid fibrils in vitro

Abstract Background Cerebral deposition of amyloid β-peptide (Aβ) is a major neuropathologic feature in Alzheimer’s disease (AD). A consistent protective effect of smoking on AD has been documented by many case-control studies. It has been suggested that nicotine, a major component of cigarette smoke, protects neurons against Aβ toxicity via the upregulation of nicotinic receptors, as well as via the inhibition of β-amyloid fibril (fAβ) formation from Aβ. Methods We used fluorescence spectroscopy with thioflavin T and electron microscopy to examine the effects of nicotine, pyridine, and N-methylpyrrolidine on the formation, extension, and disruption of fAβ(1-40) and fAβ(1-42) at pH 7.5 at 37°C in vitro. Results Nicotine dose-dependently inhibited fAβ(1-40) and fAβ(1-42) formation from fresh Aβ(1-40) and Aβ(1-42), respectively, as well as the extension reaction of both fAβs. Moreover, nicotine disrupted preformed fAβ(1-40) and fAβ(1-42). These effects of nicotine were observed at concentrations above 10 mmol/L and were similar to those of N-methylpyrrolidine. Conclusions The antiamyloidogenic effect of nicotine may be exerted not only by the inhibition of fAβ formation but also by the disruption of preformed fAβ. Additionally, this effect may be attributed to N-methylpyrrolidine moieties of nicotine.