Selective destabilization of soluble amyloid beta oligomers by divalent metal ions

Aggregation of the amyloid beta (Abeta) peptide yields both fibrillar precipitates and soluble oligomers, and is associated with Alzheimer's disease (AD). In vitro, Cu(2+) and Zn(2+) strongly bind Abeta and promote its precipitation. However, less is known about their interactions with the soluble oligomers, which are thought to be the major toxic species responsible for AD. Using fluorescence correlation spectroscopy to resolve the various soluble species of Abeta, we show that low concentrations of Cu(2+) (1 microM) and Zn(2+) (4 microM) selectively eliminate the oligomeric population (within approximately 2h), while Mg(2+) displays a similar effect at a higher concentration (60 microM). This uncovers a new aspect of Abeta-metal ion interactions, as precipitation is not substantially altered at these low metal ion concentrations. Our results suggest that physiological concentrations of Cu(2+) and Zn(2+) can critically alter the stability of the toxic Abeta oligomers and can potentially control the course of neurodegeneration.     

Redox cycling of iron by Abeta42

The amyloid cascade hypothesis and oxidative damage have been inextricably linked in the neurodegeneration that is characteristic of Alzheimer's disease. We have investigated this link and sought to suggest a mechanism whereby the precipitation of Abeta42 might contribute to the redox cycling of iron and hence the generation of reactive oxygen species via Fenton-like chemistry. We have shown that the critical step in the auto-oxidation of Fe(II) under the near-physiological conditions of our study involved the generation of H2O2 via O2.- and that Abeta42 influenced Fenton chemistry through aggregation state-specific binding of both Fe(II) and Fe(III). The net result of these interactions was the delayed precipitation of kinetically redox-inactive Fe(OH)3(s) such that Fe(II)/Fe(III) were cycled in redox-active forms over a substantially longer time period than if peptide had been absent from preparations. The addition of physiologically significant concentrations of either Cu(II) or Zn(II) reduced the role played by Abeta42 in the Fe(II)/Fe(III) redox cycle whereas a pathophysiologically significant concentration of Al(III) potentiated the redox cycle in favour of Fe(II) whether or not Cu(II) or Zn(II) was additionally present. The results support the notion that oxidative damage in the immediate vicinity of, for example, senile plaques, may be the result of Fenton chemistry catalysed by the codeposition of Abeta42 with metals such as Fe(II)/Fe(III) and Al(III).     

Evidence that the beta-amyloid plaques of Alzheimer's disease represent the redox-silencing and entombment of abeta by zinc

Abeta binds Zn(2+), Cu(2+), and Fe(3+) in vitro, and these metals are markedly elevated in the neocortex and especially enriched in amyloid plaque deposits of individuals with Alzheimer's disease (AD). Zn(2+) precipitates Abeta in vitro, and Cu(2+) interaction with Abeta promotes its neurotoxicity, correlating with metal reduction and the cell-free generation of H(2)O(2) (Abeta1-42 > Abeta1-40 > ratAbeta1-40). Because Zn(2+) is redox-inert, we studied the possibility that it may play an inhibitory role in H(2)O(2)-mediated Abeta toxicity. In competition to the cytotoxic potentiation caused by coincubation with Cu(2+), Zn(2+) rescued primary cortical and human embryonic kidney 293 cells that were exposed to Abeta1-42, correlating with the effect of Zn(2+) in suppressing Cu(2+)-dependent H(2)O(2) formation from Abeta1-42. Since plaques contain exceptionally high concentrations of Zn(2+), we examined the relationship between oxidation (8-OH guanosine) levels in AD-affected tissue and histological amyloid burden and found a significant negative correlation. These data suggest a protective role for Zn(2+) in AD, where plaques form as the result of a more robust Zn(2+) antioxidant response to the underlying oxidative attack.     

Cu（Ⅱ） Potentiation of Alzheimer Aβ1-40 Cytotoxicity and Transition on its Secondary Structure

Mounting evidence has shown that dyshomeostasis of the redox-active biometals such as Cuand Fe can lead to oxidative stress,which plays a key role in the neuropathology of Alzheimer's disease(AD).Here we demonstrate that with the formation of Cu(Ⅱ)·Aβ1-40 complexes,copper markedly potentiatesthe neurotoxicity exhibited by β-amyloid peptide (Aβ).A greater amount of hydrogen peroxide was releasedwhen Cu(Ⅱ)·Aβ1-40 complexes was added to the xanthine oxidase/xanthine system detected by potassiumiodide spectrophotometry.Copper bound to Aβ1-40 was observed by electron paramagnetic resonance(EPR) spectroscopy.Circular dichroism (CD) studies indicated that copper chelation could cause a structuraltransition of Aβ.The addition of copper to Aβ introduced an increase on β-sheet as well as α-helix,whichmay be responsible for the aggregation of Aβ.We hypothesized that Aβ aggregation induced by copper maybe responsible for local injury in AD.The interaction between Cu~(2 ) and Aβ also provides a possible mechanismfor the enrichment of metal ions in amyloid plaques in the AD brain.     

Protective Effects of Vitamin E against Oxidative Damage Induced by Aβ1-40Cu（Ⅱ） Complexes

β-amyloid peptide (Aβ) is considered to be responsible for the formation of senile plaques,which is the hallmark of Alzheimer's disease (AD).Oxidative stress,manifested by protein oxidation andlipid peroxidation,among other alterations,is a characteristic of AD brain.A growing body of evidence hasbeen presented in support of Aβ_(1-40) forming an oligomeric complex that binds copper at a CuZn superoxidedismutase-like binding site. Aβ_(1-40)Cu(Ⅱ) complexes generate neurotoxic hydrogen peroxide (H_2O_2) from O_2via Cue reduction,though the precise reaction mechanism is unclear.The toxicity of Aβ_(1-40) or the Aβ_(1-40)Cu(Ⅱ)complexes to cultured primary cortical neurons was partially attenuated when ( )-α-tocopherol (vitamin E)as free radical antioxidant was added at a concentration of 100 μM.The data derived from lactate dehydro-genase (LDH) release and the formation of H_2O_2 confirmed the results from the MTT assay.These findingsindicate that copper binding to Aβ_(1-40) can give rise to greater production of H_2O_2, which leads to a break-down in the integrity of the plasma membrane and subsequent neuronal death.Groups treated with vitaminE exhibited much slighter damage,suggesting that vitamin E plays a key role in protecting neuronal cellsfrom dysfunction or death.     

Copper mediates dityrosine cross-linking of Alzheimer's amyloid-beta

We have previously reported that amyloid Abeta, the major component of senile plaques in Alzheimer's disease (AD), binds Cu with high affinity via histidine and tyrosine residues [Atwood, C. S., et al. (1998) J. Biol. Chem. 273, 12817-12826; Atwood, C. S., et al. (2000) J. Neurochem. 75, 1219-1233] and produces H(2)O(2) by catalyzing the reduction of Cu(II) or Fe(III) [Huang, X., et al. (1999) Biochemistry 38, 7609-7616; Huang, X., et al. (1999) J. Biol. Chem. 274, 37111-37116]. Incubation with Cu induces the SDS-resistant oligomerization of Abeta [Atwood, C. S., et al. (2000) J. Neurochem. 75, 1219-1233], a feature characteristic of neurotoxic soluble Abeta extracted from the AD brain. Since residues coordinating Cu are most vulnerable to oxidation, we investigated whether modifications of these residues were responsible for Abeta cross-linking. SDS-resistant oligomerization of Abeta caused by incubation with Cu was found to induce a fluorescence signal characteristic of tyrosine cross-linking. Using ESI-MS and a dityrosine specific antibody, we confirmed that Cu(II) (at concentrations lower than that associated with amyloid plaques) induces the generation of dityrosine-cross-linked, SDS-resistant oligomers of human, but not rat, Abeta peptides. The addition of H2O2 strongly promoted Cu-induced dityrosine cross-linking of Abeta1-28, Abeta1-40, and Abeta1-42, suggesting that the oxidative coupling is initiated by interaction of H2O2 with a Cu(II) tyrosinate. The dityrosine modification is significant since it is highly resistant to proteolysis and is known to play a role in increasing structural strength. Given the elevated concentration of Cu in senile plaques, our results suggest that Cu interactions with Abeta could be responsible for causing the covalent cross-linking of Abeta in these structures.     

Preventing metal-mediated oxidative DNA damage with selenium compounds

Copper and iron are two widely studied transition metals associated with hydroxyl radical (˙OH) generation, oxidative damage, and disease development. Because antioxidants ameliorate metal-mediated DNA damage, DNA gel electrophoresis assays were used to quantify the ability of ten selenium-containing compounds to inhibit metal-mediated DNA damage by hydroxyl radical. In the Cu(I)/H(2)O(2) system, selenocystine, selenomethionine, and methyl-selenocysteine inhibit DNA damage with IC(50) values ranging from 3.34 to 25.1 μM. Four selenium compounds also prevent DNA damage from Fe(II) and H(2)O(2). Additional gel electrophoresis experiments indicate that Cu(I) or Fe(II) coordination is responsible for the selenium antioxidant activity. Mass spectrometry studies show that a 1?:?1 stoichiometry is the most common for iron and copper complexes of the tested compounds, even if no antioxidant activity is observed, suggesting that metal coordination is necessary but not sufficient for selenium antioxidant activity. A majority of the selenium compounds are electroactive, regardless of antioxidant activity, and the glutathione peroxidase activities of the selenium compounds show no correlation to DNA damage inhibition. Thus, metal binding is a primary mechanism of selenium antioxidant activity, and both the chemical functionality of the selenium compound and the metal ion generating damaging hydroxyl radical significantly affect selenium antioxidant behavior.     

Oxidative stress induces intracellular accumulation of amyloid beta-protein (Abeta) in human neuroblastoma cells

Several lines of evidence suggest that enhanced oxidative stress is involved in the pathogenesis and/or progression of Alzheimer's disease (AD). Amyloid beta-protein (Abeta) that composes senile plaques, a major neuropathological hallmark of AD, is considered to have a causal role in AD. Thus, we have studied the effect of oxidative stress on Abeta metabolism within the cell. Here, we report that oxidative stress induced by H(2)O(2) (100-250 microM) caused an increase in the levels of intracellular Abeta in human neuroblastoma SH-SY5Y cells. Treatment with 200 microM H(2)O(2) caused significant decreases in the protein levels of full-length beta-amyloid precursor protein (APP) and its COOH-terminal fragment that is generated by beta-cleavage, while the gene expression of APP was not altered under these conditions. A pulse-chase experiment further showed a decrease in the half-life of this amyloidogenic COOH-terminal fragment but not in that of nonamyloidogenic counterpart in the H(2)O(2)-treated cells. These results suggest that oxidative stress promotes intracellular accumulation of Abeta through enhancing the amyloidogenic pathway.     

Polyphenol tannic acid inhibits hydroxyl radical formation from Fenton reaction by complexing ferrous ions

Tannic acid (TA), a plant polyphenol, has been described as having antimutagenic, anticarcinogenic and antioxidant activities. Since it is a potent chelator of iron ions, we decided to examine if the antioxidant activity of TA is related to its ability to chelate iron ions. The degradation of 2-deoxyribose induced by 6 microM Fe(II) plus 100 microM H2O2 was inhibited by TA, with an I50 value of 13 microM. Tannic acid was over three orders of magnitude more efficient in protecting against 2-deoxyribose degradation than classical *OH scavengers. The antioxidant potency of TA was inversely proportional to Fe(II) concentration, demonstrating a competition between H2O2 and AT for reaction with Fe(II). On the other hand, the efficiency of TA was nearly unchanged with increasing concentrations of the *OH detector molecule, 2-deoxyribose. These results indicate that the antioxidant activity of TA is mainly due to iron chelation rather than *OH scavenging. TA also inhibited 2-deoxyribose degradation mediated by Fe(III)-EDTA (iron = 50 microM) plus ascorbate. The protective action of TA was significantly higher with 50 microM EDTA than with 500 microM EDTA, suggesting that TA removes Fe(III) from EDTA and forms a complex with iron that cannot induce *OH formation. We also provided evidence that TA forms a stable complex with Fe(II), since excess ferrozine (14 mM) recovered 95-96% of the Fe(II) from 10 microM TA even after a 30-min exposure to 100-500 microM H2O2. Addition of Fe(III) to samples containing TA caused the formation of Fe(II)n-TA, complexes, as determined by ferrozine assays, indicating that TA is also capable of reducing Fe(III) ions. We propose that when Fe(II) is complexed to TA, it is unable to participate in Fenton reactions and mediate *OH formation. The antimutagenic and anticarcinogenic activity of TA, described elsewhere, may be explained (at least in part) by its capacity to prevent Fenton reactions.     

Effects of 1,4-dihydropyridine derivatives (cerebrocrast, gammapyrone, glutapyrone, and diethone) on mitochondrial bioenergetics and oxidative stress: a comparative study

The potential protective action of 1,4-dihydropyridine derivatives (cerebrocrast, gammapyrone, glutapyrone, and diethone) against oxidative stress was assessed on mitochondrial bioenergetics, inner membrane anion channel (IMAC), Ca2+-induced opening of the permeability transition pore (PTP), and oxidative damage induced by the oxidant pair adenosine diphosphate (ADP)/Fe2+ (lipid peroxidation) of mitochondria isolated from rat liver. By using succinate as the respiratory substrate, respiratory control ratio (RCR), ADP to oxygen ratio (ADP/O), state 3, state 4, and uncoupled respiration rates were not significantly affected by gammapyrone, glutapyrone, and diethone concentrations up to 100 microM. Cerebrocrast at concentrations higher than 25 microM depressed RCR, ADP/O, state 3, and uncoupled respiration rates, but increased three times state 4 respiration rate. The transmembrane potential (deltapsi) and the phosphate carrier rate were also decreased. At concentrations lower than 25 microM, cerebrocrast inhibited the mitochondrial IMAC and partially prevented Ca2+-induced opening of the mitochondrial PTP, whereas gammapyrone, glutapyrone, and diethone were without effect. Cerebrocrast, gammapyrone, and glutapyrone concentrations up to 100 microM did not affect ADP/Fe2+-induced lipid peroxidation of rat liver mitochondria, while very low diethone concentrations (up to 5 microM) inhibited it in a dose-dependent manner, as measured by oxygen consumption and thiobarbituric acid reactive substances formation. Diethone also prevented deltapsi dissipation due to lipid peroxidation initiated by ADP/Fe2+. It can be concluded that: none of the compounds interfere with mitochondrial bioenergetics at concentrations lower than 25 microM; cerebrocrast was the only compound that affected mitochondrial bioenergetics, but only for concentrations higher than 25 microM; at concentrations that did not affect mitochondrial bioenergetics (< or = 25 microM), only cerebrocrast inhibited the IMAC and partially prevented Ca2+-induced opening of the PTP; diethone was the only compound that expressed antioxidant activity at very low concentrations (< or = 5 microM). Cerebrocrast acting as an inhibitor of the IMAC and diethone acting as an antioxidant could provide effective protective roles in preventing mitochondria from oxidative damage, favoring their therapeutic interest in the treatment of several pathological situations known to be associated with cellular oxidative stress.     

Toxic levels of amyloid beta peptide do not induce VEGF synthesis

Alzheimer's disease is a neurodegenerative disorder associated with progressive loss of cognitive function and memory. Amyloid beta peptide (Abeta) is the major component of senile plaques and is known to exert its cytotoxic effect mainly by producing H2O2. Vascular endothelial growth factor (VEGF) is elevated in the cerebrospinal fluid (CSF) and brain of AD patients, and H2O2 is one of the factors that induce VEGF. Therefore, we tested whether Abeta might be responsible for the increased VEGF synthesis. We found that Abeta induced the production of H2O2 in vitro. Comparison of the amount of H2O2 required to induce VEGF synthesis in HN33 cells and the amount of H2O2 produced by 10 muM Abeta1-42 in vitro suggested that a toxic concentration of Abeta might induce VEGF synthesis in these cells. However, toxic concentrations of Abeta failed to induce VEGF synthesis in several cell systems. They also had no effect on antioxidant enzymes such as glutathione peroxidase, catalase, and peroxiredoxin in HN33 cells. Cu2+, Zn2+ and Fe3+ are known to accumulate in the brains of AD patients and promote aggregation of Abeta, and Cu2+ by itself induces synthesis of VEGF. However, there was no synergistic effect between Cu2+ and Abeta1-42 in the induction of VEGF synthesis and Zn2+ and Fe3+ also had no effect on the synthesis of VEGF, alone or in combination with Abeta.     

Curcumin inhibits formation of amyloid beta oligomers and fibrils, binds plaques, and reduces amyloid in vivo

Alzheimer's disease (AD) involves amyloid beta (Abeta) accumulation, oxidative damage, and inflammation, and risk is reduced with increased antioxidant and anti-inflammatory consumption. The phenolic yellow curry pigment curcumin has potent anti-inflammatory and antioxidant activities and can suppress oxidative damage, inflammation, cognitive deficits, and amyloid accumulation. Since the molecular structure of curcumin suggested potential Abeta binding, we investigated whether its efficacy in AD models could be explained by effects on Abeta aggregation. Under aggregating conditions in vitro, curcumin inhibited aggregation (IC(50) = 0.8 microM) as well as disaggregated fibrillar Abeta40 (IC(50) = 1 microM), indicating favorable stoichiometry for inhibition. Curcumin was a better Abeta40 aggregation inhibitor than ibuprofen and naproxen, and prevented Abeta42 oligomer formation and toxicity between 0.1 and 1.0 microM. Under EM, curcumin decreased dose dependently Abeta fibril formation beginning with 0.125 microM. The effects of curcumin did not depend on Abeta sequence but on fibril-related conformation. AD and Tg2576 mice brain sections incubated with curcumin revealed preferential labeling of amyloid plaques. In vivo studies showed that curcumin injected peripherally into aged Tg mice crossed the blood-brain barrier and bound plaques. When fed to aged Tg2576 mice with advanced amyloid accumulation, curcumin labeled plaques and reduced amyloid levels and plaque burden. Hence, curcumin directly binds small beta-amyloid species to block aggregation and fibril formation in vitro and in vivo. These data suggest that low dose curcumin effectively disaggregates Abeta as well as prevents fibril and oligomer formation, supporting the rationale for curcumin use in clinical trials preventing or treating AD.     

Binding of Zn(II), Cu(II), and Fe(II) ions to Alzheimer's A beta peptide studied by fluorescence.

Binding of Zn(II), Cu(II) and Fe(II) ions to A beta1-40, A beta1-42 and a single tryptophan mutant of Abeta 1-40 in solution at pH 7.4 was studied by fluorescent titration. Job plots and fitting of titration curves revealed formation of 1:1 and 1:2 peptide-metal complexes. For dimeric peptides A beta1-40 and A betaF4W the order of metal to peptide affinities is Fe < Cu > Zn, which is in agreement with the Irving-Williams series of complex stability. The affinity of A beta1-42 for Fe increases dramatically upon aggregation: K(D) changes from ca. 100 to ca. 0.2 microM.     

Iron(III)- and copper(II) complexes of an asymmetric, pentadentate salen-like ligand bearing a pendant carboxylate group

The equilibrium and solution structural properties of the iron(III) and copper(II) complexes of an asymmetric salen-like ligand (N,N'-bis(2-hydroxybenzyl)-2,3-diamino-propionic acid, H(3)bhbdpa) bearing a pendant carboxylate group were characterized in aqueous solution by potentiometric, pH-dependent electron paramagnetic resonance (EPR) and UV-Vis (UV-Visible) measurements. In the equimolar systems the pentadentate ligand forms very stable, differently protonated mononuclear complexes with both metal ions. In the presence of iron(III) {NH, PhO(-), COO(-)}, {2NH, 2PhO(-), COO(-)} and {2NH, 2PhO(-), COO(-), OH(-)} coordinated complexes are dominant. The EPR titrations reflected the presence of microscopic complex formation pathways, leading to the formation of binding isomers in case of Cu(H(2)bhbdpa)(+), Cu(Hbhbdpa) and Cu(bhbdpa)(-). The {2NH, 2PhO(-)+COO(-)/H(2)O} coordinated Cu(bhbdpa) is the only species between pH 6-11. At twofold excess of metal ion dinuclear complexes were detected with both iron(III) and copper(II). In presence of iron(III) a mu-carboxylato-mu-hydroxo-bridged dinuclear complex (Fe(2)(bhbdpa)(OH)(3)) is formed from Fe(H(2)bhbdpa)(2+) through overlapping proton release processes, providing one of the rare examples for the stabilization of an endogenous carboxylate bridged diiron core in aqueous solution. The complex Cu(2)(bhbdpa)(+) detected in the presence of copper(II) is a paramagnetic (S=1) species with relatively weakly coupled metal ions.     

The redox chemistry of the Alzheimer's disease amyloid beta peptide

There is a growing body of evidence to support a role for oxidative stress in Alzheimer's disease (AD), with increased levels of lipid peroxidation, DNA and protein oxidation products (HNE, 8-HO-guanidine and protein carbonyls respectively) in AD brains. The brain is a highly oxidative organ consuming 20% of the body's oxygen despite accounting for only 2% of the total body weight. With normal ageing the brain accumulates metals ions such iron (Fe), zinc (Zn) and copper (Cu). Consequently the brain is abundant in antioxidants to control and prevent the detrimental formation of reactive oxygen species (ROS) generated via Fenton chemistry involving redox active metal ion reduction and activation of molecular oxygen. In AD there is an over accumulation of the Amyloid beta peptide (Abeta), this is the result of either an elevated generation from amyloid precursor protein (APP) or inefficient clearance of Abeta from the brain. Abeta can efficiently generate reactive oxygen species in the presence of the transition metals copper and iron in vitro. Under oxidative conditions Abeta will form stable dityrosine cross-linked dimers which are generated from free radical attack on the tyrosine residue at position 10. There are elevated levels of urea and SDS resistant stable linked Abeta oligomers as well as dityrosine cross-linked peptides and proteins in AD brain. Since soluble Abeta levels correlate best with the degree of degeneration [C.A. McLean, R.A. Cherny, F.W. Fraser, S.J. Fuller, M.J. Smith, K. Beyreuther, A.I. Bush, C.L. Masters, Soluble pool of Abeta amyloid as a determinant of severity of neurodegeneration in Alzheimer's disease, Ann. Neurol. 46 (1999) 860-866] we suggest that the toxic Abeta species corresponds to a soluble dityrosine cross-linked oligomer. Current therapeutic strategies using metal chelators such as clioquinol and desferrioxamine have had some success in altering the progression of AD symptoms. Similarly, natural antioxidants curcumin and ginkgo extract have modest but positive effects in slowing AD development. Therefore, drugs that target the oxidative pathways in AD could have genuine therapeutic efficacy.     

Methionine regulates copper/hydrogen peroxide oxidation products of Abeta.

Metal-catalysed oxidation (MCO) may play a causative role in the pathogenesis of Alzheimer's disease (AD). Amyloid beta peptide (Abeta), the major biomarker of AD, in the presence of copper ions reduces Cu(2+) to Cu(+) and catalyses the formation of H(2)O(2) that subsequently induces radicals through Fenton chemistry. Abeta is also subject to attack by free radicals, where the presence of Cu(2+) in conjunction with H(2)O(2) catalyses oxygenation, primarily at the methionine sulfur atom. This work investigates MCO of Abeta, to gain further insight into the role of oxidative stress in AD. By combining a fluorescence assay with gel electrophoresis to monitor MCO reactions of Abeta (1-28) in the presence and absence of methionine it was determined that methionine can both protect some residues against MCO and promote the oxidation of Tyr(10) specifically. Electrospray ionization mass spectrometric analysis of methionine MCO products indicated the formation of methionine sulfoxide, methionine sulfone and related hydroxylated products. Similar products could be formed from the oxidation of Met(35) of Abeta and may relate to changes in properties of the peptide following MCO.     

Iron(II) and hydrogen peroxide detoxification by human H-chain ferritin. An EPR spin-trapping study

Overexpression of human H-chain ferritin (HuHF) is known to impart a degree of protection to cells against oxidative stress and the associated damage to DNA and other cellular components. However, whether this protective activity resides in the protein's ability to inhibit Fenton chemistry as found for Dps proteins has never been established. Such inhibition does not occur with the related mitochondrial ferritin which displays much of the same iron chemistry as HuHF, including an Fe(II)/H(2)O(2) oxidation stoichiometry of approximately 2:1. In the present study, the ability of HuHF to attenuate hydroxyl radical production by the Fenton reaction (Fe(2+) + H(2)O(2) --> Fe(3+) + OH(-) + *OH) was examined by electron paramagnetic resonance (EPR) spin-trapping methods. The data demonstrate that the presence of wild-type HuHF during Fe(2+) oxidation by H(2)O(2) greatly decreases the amount of .OH radical produced from Fenton chemistry whereas the ferroxidase site mutant 222 (H62K + H65G) and human L-chain ferritin (HuLF) lack this activity. HuHF catalyzes the pairwise oxidation of Fe(2+) by the detoxification reaction [2Fe(2+) + H(2)O(2) + 2H(2)O --> 2Fe(O)OH(core) + 4H(+)] that occurs at the ferroxidase site of the protein, thereby preventing the production of hydroxyl radical. The small amount of *OH radical that is produced in the presence of ferritin (相似文献   

Protective effect of quercetin in primary neurons against Abeta(1-42): relevance to Alzheimer's disease

Quercetin, a flavonoid found in various foodstuffs, has antioxidant properties and increases glutathione (GSH) levels and antioxidant enzyme function. Considerable attention has been focused on increasing the intracellular GSH levels in many diseases, including Alzheimer's disease (AD). Amyloid beta-peptide [Abeta(1-42)], elevated in AD brain, is associated with oxidative stress and neurotoxicity. We aimed to investigate the protective effects of quercetin on Abeta(1-42)-induced oxidative cell toxicity in cultured neurons in the present study. Decreased cell survival in neuronal cultures treated with Abeta(1-42) correlated with increased free radical production measured by dichlorofluorescein fluorescence and an increase in protein oxidation (protein carbonyl, 3-nitrotyrosine) and lipid peroxidation (protein-bound 4-hydroxy-2-nonenal). Pretreatment of primary hippocampal cultures with quercetin significantly attenuated Abeta(1-42)-induced cytotoxicity, protein oxidation, lipid peroxidation and apoptosis. A dose-response study suggested that quercetin showed protective effects against Abeta(1-42) toxicity by modulating oxidative stress at lower doses, but higher doses were not only non-neuroprotective but also toxic. These findings provide motivation to test the hypothesis that quercetin may provide a promising approach for the treatment of AD and other oxidative-stress-related neurodegenerative diseases.     

Metal ions differentially influence the aggregation and deposition of Alzheimer's beta-amyloid on a solid template

The abnormal deposition and aggregation of beta-amyloid (Abeta) on brain tissues are considered to be one of the characteristic neuropathological features of Alzheimer's disease (AD). Environmental conditions such as metal ions, pH, and cell membranes are associated with Abeta deposition and plaque formation. According to the amyloid cascade hypothesis of AD, the deposition of Abeta42 oligomers as diffuse plaques in vivo is an important earliest event, leading to the formation of fibrillar amyloid plaques by the further accumulation of soluble Abeta under certain environmental conditions. In order to characterize the effect of metal ions on amyloid deposition and plaque growth on a solid surface, we prepared a synthetic template by immobilizing Abeta oligomers onto a N-hydroxysuccinimide ester-activated solid surface. According to our study using ex situ atomic force microscopy (AFM), Fourier transform infrared spectroscopy (FT-IR), and thioflavin T (ThT) fluorescence spectroscopy, Cu2+ and Zn2+ ions accelerated both Abeta40 and Abeta42 deposition but resulted only in the formation of "amorphous" aggregates. In contrast, Fe3+ induced the deposition of "fibrillar" amyloid plaques at neutral pH. Under mildly acidic environments, the formation of fibrillar amyloid plaques was not induced by any metal ion tested in this work. Using secondary ion mass spectroscopy (SIMS) analysis, we found that binding Cu ions to Abeta deposits on a solid template occurred by the possible reduction of Cu ions during the interaction of Abeta with Cu2+. Our results may provide insights into the role of metal ions on the formation of fibrillar or amorphous amyloid plaques in AD.     

Generation of *OH initiated by interaction of Fe2+ and Cu+ with dioxygen; comparison with the Fenton chemistry

Iron and copper toxicity has been presumed to involve the formation of hydroxyl radical (*OH) from H2O2 in the Fenton reaction. The aim of this study was to verify that Fe2+-O2 and Cu+-O2 chemistry is capable of generating *OH in the quasi physiological environment of Krebs-Henseleit buffer (KH), and to compare the ability of the Fe2+-O2 system and of the Fenton system (Fe2+ + H2O2) to produce *OH. The addition of Fe2+ and Cu+ (0-20 microM) to KH resulted in a concentration-dependent increase in *OH formation, as measured by the salicylate method. While Fe3+ and Cu2+ (0-20 microM) did not result in *OH formation, these ions mediated significant *OH production in the presence of a number of reducing agents. The *OH yield from the reaction mediated by Fe2+ was increased by exogenous Fe3+ and Cu2+ and was prevented by the deoxygenation of the buffer and reduced by superoxide dismutase, catalase, and desferrioxamine. Addition of 1 microM, 5 microM or 10 microM Fe2+ to a range of H2O2 concentrations (the Fenton system) resulted in a H2O2-concentration-dependent rise in *OH formation. For each Fe2+ concentration tested, the *OH yield doubled when the ratio [H2O2]:[Fe2+] was raised from zero to one. In conclusion: (i) Fe2+-O2 and Cu+-O2 chemistry is capable of promoting *OH generation in the environment of oxygenated KH, in the absence of pre-existing superoxide and/or H2O2, and possibly through a mechanism initiated by the metal autoxidation; (ii) The process is enhanced by contaminating Fe3+ and Cu2+; (iii) In the presence of reducing agents also Fe3+ and Cu2+ promote the *OH formation; (iv) Depending on the actual [H2O2]:[Fe2+] ratio, the efficiency of the Fe2+-O2 chemistry to generate *OH is greater than or, at best, equal to that of the Fe2+-driven Fenton reaction.