Protective Effect of Isorhynchophylline Against β-Amyloid-Induced Neurotoxicity in PC12 Cells

Beta-amyloid peptide (Aβ), a major protein component of senile plaques, has been considered as a critical cause in the pathogenesis of Alzheimer’s disease (AD). Modulation of the Aβ-induced neurotoxicity has emerged as a possible therapeutic approach to ameliorate the onset and progression of AD. The present study aimed to evaluate the protective effect of isorhynchophylline, an oxindole alkaloid isolated from a Chinese herb Uncaria rhynchophylla, on Aβ-induced neurotoxicity in cultured rat pheochromocytoma (PC12) cells. The results showed that pretreatment with isorhynchophylline significantly elevated cell viability, decreased the levels of intracellular reactive oxygen species and malondialdehyde, increased the level of glutathione, and stabilized mitochondrial membrane potential in Aβ_25-35-treated PC12 cells. In addition, isorhynchophylline significantly suppressed the formation of DNA fragmentation and the activity of caspase-3 and moderated the ratio of Bcl-2/Bax. These results indicate that isorhynchophylline exerts a neuroprotective effect against Aβ_25-35-induced neurotoxicity in PC12 cells, at least in part, via inhibiting oxidative stress and suppressing the mitochondrial pathway of cellular apoptosis.     

Copper in the brain and Alzheimer’s disease

Alzheimer’s disease (AD) is the most common form of neurodegenerative disease. The brain is particularly vulnerable to oxidative damage induced by unregulated redox-active metals such as copper and iron, and the brains of AD patients display evidence of metal dyshomeostasis and increased oxidative stress. The colocalisation of copper and amyloid β (Aβ) in the glutamatergic synapse during NMDA-receptor-mediated neurotransmission provides a microenvironment favouring the abnormal interaction of redox-potent Aβ with copper under conditions of copper dysregulation thought to prevail in the AD brain, resulting in the formation of neurotoxic soluble Aβ oligomers. Interactions between Aβ oligomers and copper can further promote the aggregation of Aβ, which is the core component of extracellular amyloid plaques, a central pathological hallmark of AD. Copper dysregulation is also implicated in the hyperphosphorylation and aggregation of tau, the main component of neurofibrillary tangles, which is also a defining pathological hallmark of AD. Therefore, tight regulation of neuronal copper homeostasis is essential to the integrity of normal brain functions. Therapeutic strategies targeting interactions between Aβ, tau and metals to restore copper and metal homeostasis are discussed.     

Mitochondria and Alzheimer’s disease: amyloid-β peptide uptake and degradation by the presequence protease,hPreP

Several lines of evidence suggest mitochondrial dysfunction as a possible underlying mechanism of Alzheimer’s disease (AD). Accumulation of the amyloid-β peptide (Aβ), a neurotoxic peptide implicated in the pathogenesis of AD, has been detected in brain mitochondria of AD patients and AD transgenic mouse models. In vitro evidence suggests that the Aβ causes mitochondrial dysfunction e.g. oxidative stress, mitochondrial fragmentation and decreased activity of cytochrome c oxidase and TCA cycle enzymes. Here we review the link between mitochondrial dysfunctions and AD. In particular we focus on the mechanism for Aβ uptake by mitochondria and on the recently identified Aβ degrading protease in human brain mitochondria.     

Copper catalysed oxidation of amino acids and Alzheimer's disease

Summary Metal-catalyzed oxidation (MCO) can lead to damage of bio-molecules and is implicated in neurodegenerative diseases, such as Alzheimer's disease (AD). The amino acid residues, tyrosine, histidine and methionine, have been proposed to play important roles in metal mediated oxidative stress and subsequent reactions of amyloid β peptide (Aβ) a major contributor in the pathogenesis of AD. The MCO of Aβ residues, particularly histidine, methionine and tyrosine, and reviewed. MCO of Aβ histidine and tyrosine residues can facilitate oligomerization and may play a role in both amyloid formation and Aβ neurotoxicity. Further work is needed to determine the importance of Aβ oxidation in AD and the role of Aβ oxidation products and oxidative stress in disease progression. The mechanisms of Aβ MCO are complex and multiple reaction products can form. Further study is needed to determine the mechanisms by which Aβ MCO occursin vivo. In addition, new analytical methods are required to monitor the formation of Aβ MCO products formed during AD. The copper-H₂O₂ redox system provides a chemical model by which Aβ MCO can be studiedin vitro and can be used to produce oxidatively modified amino acid residues for use as standards in developing new analytical methods to monitor Aβ MCO.     

Beta amyloid peptide: from different aggregation forms to the activation of different biochemical pathways

Amyloid beta peptide (Aβ) is the major component of amyloid plaques in the brain of individuals affected by Alzheimer’s disease (AD). The formation of the plaques is due to an overproduction of Aβ by APP processing, its precursor, and to its ability to convert under specific conditions from its soluble form into highly ordered fibrillar aggregates. Although neuronal degeneration occurs near the amyloid plaques, some studies have suggested that intermediates such as protofibrils or simple oligomers are also involved in AD pathogenesis and even appear to be the more dangerous species in the onset of the pathology. Further, toxic properties of aggregates of different size have been investigated and the obtained results support the hypothesis that different aggregate sizes can induce different degeneration pathways. In the present review some of the knowledge about the biochemical routes of Aβ processing and production and the relationship among Aβ and oxidative stress, metal homeostasis, inflammatory process, and cell death are summarized. Moreover, current strategies addressing both fibrillogenesis process and different Aβ altered biochemical pathways utilized for therapies are described.     

Multifunctional roles of enolase in Alzheimer's disease brain: beyond altered glucose metabolism

Enolase enzymes are abundantly expressed, cytosolic carbon-oxygen lyases known for their role in glucose metabolism. Recently, enolase has been shown to possess a variety of different regulatory functions, beyond glycolysis and gluconeogenesis, associated with hypoxia, ischemia, and Alzheimer's disease (AD). AD is an age-associated neurodegenerative disorder characterized pathologically by elevated oxidative stress and subsequent damage to proteins, lipids, and nucleic acids, appearance of neurofibrillary tangles and senile plaques, and loss of synapse and neuronal cells. It is unclear if development of a hypometabolic environment is a consequence of or contributes to AD pathology, as there is not only a significant decline in brain glucose levels in AD, but also there is an increase in proteomics identified oxidatively modified glycolytic enzymes that are rendered inactive, including enolase. Previously, our laboratory identified α-enolase as one the most frequently up-regulated and oxidatively modified proteins in amnestic mild cognitive impairment (MCI), early-onset AD, and AD. However, the glycolytic conversion of 2-phosphoglycerate to phosphoenolpyruvate catalyzed by enolase does not directly produce ATP or NADH; therefore it is surprising that, among all glycolytic enzymes, α-enolase was one of only two glycolytic enzymes consistently up-regulated from MCI to AD. These findings suggest enolase is involved with more than glucose metabolism in AD brain, but may possess other functions, normally necessary to preserve brain function. This review examines potential altered function(s) of brain enolase in MCI, early-onset AD, and AD, alterations that may contribute to the biochemical, pathological, clinical characteristics, and progression of this dementing disorder.     

Lymphocyte mitochondria: toward identification of peripheral biomarkers in the progression of Alzheimer disease

Alzheimer disease (AD) is an age-related neurodegenerative condition. AD is histopathologically characterized by the presence of three main hallmarks: senile plaques (rich in amyloid-β peptide), neuronal fibrillary tangles (rich in phosphorylated tau protein), and synapse loss. However, definitive biomarkers for this devastating disease in living people are still lacking. In this study, we show that levels of oxidative stress markers are significantly increased in the mitochondria isolated from lymphocytes of subjects with mild cognitive impairment (MCI) compared to cognitively normal individuals. Further, an increase in mitochondrial oxidative stress in MCI is associated with MMSE score, vitamin E components, and β-carotene. Further, a proteomics approach showed that alterations in the levels of thioredoxin-dependent peroxide reductase, myosin light polypeptide 6, and ATP synthase subunit β might be important in the progression and pathogenesis of AD. Increased understanding of oxidative stress and protein alterations in easily obtainable peripheral tissues will be helpful in developing biomarkers to combat this devastating disorder.     

Altered mitochondria,energy metabolism,voltage-dependent anion channel,and lipid rafts converge to exhaust neurons in Alzheimer’s disease

β-amyloid (Aβ) deposition, in the form of plaques and amyloid angiopathy, and hyper-phosphorylated tau deposition forming neurofibrillary tangles, dystrophic neurites around β-amyloid plaques and neuropil threads, are neuropathological hallmarks of Alzheimer’s disease (AD) that accumulate in the brain with disease progression. These changes are accompanied by progressive loss of synapses and nerve cell death. Progressive cognitive impairment and dementia are the main neurological deficits. In addition, there is cumulative evidence demonstrating other metabolic disturbances that impair cell function and hamper neuron viability. The main components of the mitochondria are altered: complex IV of the respiratory chain is reduced; complex V which metabolizes ADP to form ATP is oxidatively damaged and functionally altered; and voltage-dependent anion channel VDAC, a major component of the outer mitochondrial membrane that regulates ion fluxes, is damaged as a result of oxidative stress. Mitochondria are a major source of reactive oxygen species that promote oxidative damage to DNA, RNA, proteins and lipids. Protein targets of oxidative damage are, among others, several enzymatic components of the glycolysis, lipid metabolism and cycle of the citric acid that fuel oxidative phosphorylation, mitochondrial respiration and energy production. The lipid composition of lipid rafts, key membrane specializations that facilitate the transfer of substrates, and protein-protein and lipid-protein interactions, is altered as a result of the abnormally low levels of n-3 long chain polyunsaturated fatty acids (mainly docosahexaenoic acid) that increase viscosity and augment energy consumption. Abnormal lipid raft composition may also modify the activity of key enzymes that modulate the cleavage of the amyloid precursor protein to form toxic Aβ. This is further complicated by the disruption of the complex VDAC with estrogen receptor α at the caveolae which participates, under physiological conditions, in the protection against β-amyloid. Together, all these alterations converge in reduced energy production and increased energy demands in altered cells. Cell exhaustion is suggested as being a determining element to interpret impaired neuron function, reduced molecular turnover, and enhanced cell death.     

Cryptotanshinione Inhibits β-Amyloid Aggregation and Protects Damage from β-Amyloid in SH-SY5Y Cells

The deposition of amyloid β-protein (Aβ) fibrils into plaques within the brain parenchyma and along cerebral blood vessels is a hallmark of Alzheimer’s disease (AD). Aβ42 oligomers and fibrils cause the breakdown of neural circuits, neuronal death and eventually dementia. Drugs that inhibit Aβ42 aggregation may be a novel direction in AD drug discovery. Cryptotanshinone (CTS), an active component of the medicinal herb Salvia miltiorrhiza, has been shown to improve learning and memory in several pharmacological models of AD. However, the effects of CTS on the Aβ aggregation and toxicity are unclear. The current work shows the effectiveness of CTS on the inhibition of Aβ42 aggregation and toxicity to human neuroblastoma cells. In this study, we demonstrated that CTS can inhibit Aβ42 spontaneous aggregation using thioflavin T fluorescence assay and transmission electron microscopy. Furthermore, we investigated the effects of CTS on Aβ-induced oxidative cell death in cultured SH-SY5Y cells. MTT and lactate dehydrogenase assays showed that CTS reduced the cytotoxicity induced by Aβ42. CTS also dramatically reduced Aβ42-induced cellular apoptosis and increased level of reactive oxygen species in these cells. Our study suggests that CTS may be useful in the inhibition or prevention of AD development and progression.     

Cytoplasmic tail adaptors of alzheimer’s amyloid-β protein precursor

Alzheimer’s disease is characterized pathologically by senile plaques in the brain. The major component of senile plaques is amyloid-β (Aβ), which is cleaved from Alzheimer’s Aβ protein precursor (AβPP). Recently, information regarding the cytoplasmic tail of AβPP has started to emerge, opening up various insights into the physiological roles of AβPP and its pathological role in Alzheimer’s disease. The cytoplasmic domain of AβPP shares the evolutionarily conserved GYENPTY motif, which binds to a number of adaptor proteins containing the phosphotyrosine interaction domain (PID). Among the PID-containing proteins, this article focuses on four groups of adaptor proteins of AβPP: Fe65, X11, mDab1, and c-Jun N-terminal kinase-interacting protein 1b/islet-brain 1. These two authors contributed equally to this study.     

Mitochondria,cholesterol and amyloid β peptide: a dangerous trio in Alzheimer disease

The molecular mechanisms of Alzheimer’s disease (AD) are not fully understood. Extensive evidence from experimental models has involved the overgeneration and accumulation of toxic amyloid β peptides (Aβ) in the onset and progression of the disease. The amyloidogenic processing of amyloid precursor protein into pathogenic Aβ fragments is thought to occur in specific domains of the plasma membrane and favored by cholesterol enrichment. Intracellular Aβ accumulation is known to induce oxidative stress, predominantly via mitochondria targeting of toxic Aβ. Recent evidence using mouse models of cholesterol loading has demonstrated that the specific mitochondrial cholesterol pool sensitizes neurons to Aβ-induced oxidant cell death and caspase-independent apoptosis due to selective mitochondrial GSH (mGSH) depletion induced by cholesterol-mediated perturbation of mitochondrial membrane dynamics. mGSH replenishment by permeable precursors such as glutathione ethyl ester protected against Aβ-mediated neurotoxicity and inflammation. Thus, these novel data expand the pathogenic role of cholesterol in AD indicating that in addition to fostering Aβ generation, mitochondrial cholesterol determines Aβ neurotoxicity via mGSH regulation.     

Amino acids variations in Amyloid-β peptides,mitochondrial dysfunction,and new therapies for Alzheimer’s disease

Soluble oligomers and/or aggregates of Amyloid-β (Aβ) are viewed by many as the principal cause for neurodegeneration in Alzheimer’s disease (AD). However, the mechanism by which Aβ and its aggregates cause neurodegeneration is not clear. The toxicity of Aβ has been attributed to its hydrophobicity. However, many specific mitochondrial cytopathologies e.g., loss of complex IV, loss of iron homeostasis, or oxidative damage cannot be explained by Aβ’s hydrophobicity. In order to understand the role of Aβ in these cytopathologies we hypothesized that Aβ impairs specific metabolic pathways. We focused on heme metabolism because it links iron, mitochondria, and Aβ. We generated experimental evidence showing that Aβ alters heme metabolism in neuronal cells. Furthermore, we demonstrated that Aβ binds to and depletes intracellular regulatory heme (forming an Aβ-heme complex), which provides a strong molecular connection between Aβ and heme metabolism. We showed that heme depletion leads to key cytopathologies identical to those seen in AD including loss of iron homeostasis and loss of mitochondrial complex IV. Aβ-heme exhibits a peroxidase-like catalytic activity, which catalytically accelerates oxidative damage. Interestingly, the amino acids sequence of rodent Aβ (roAβ) and human Aβ (huAβ) is identical except for three amino acids within the hydrophilic region, which is also the heme-binding motif that we identified. We found that huAβ, unlike roAβ, binds heme tightly and forms a peroxidase. Although, roAβ and huAβ equally form fibrils and aggregates, rodents do not develop AD-like neuropathology. These findings led us to propose a new mechanism for mitochondrial dysfunction and huAβ’s neurotoxicity. This mechanism prompted the development of methylene blue (MB), which increased heme synthesis, complex IV, and mitochondrial function. Thus, MB may delay the onset and progression of AD and serve as a lead to develop novel drugs to treat AD.     

Heat Shock Proteins and Amateur Chaperones in Amyloid-Beta Accumulation and Clearance in Alzheimer’s Disease

The pathologic lesions of Alzheimer’s disease (AD) are characterized by accumulation of protein aggregates consisting of intracellular or extracellular misfolded proteins. The amyloid-β (Aβ) protein accumulates extracellularly in senile plaques and cerebral amyloid angiopathy, whereas the hyperphosphorylated tau protein accumulates intracellularly as neurofibrillary tangles. “Professional chaperones”, such as the heat shock protein family, have a function in the prevention of protein misfolding and subsequent aggregation. “Amateur” chaperones, such as apolipoproteins and heparan sulfate proteoglycans, bind amyloidogenic proteins and may affect their aggregation process. Professional and amateur chaperones not only colocalize with the pathological lesions of AD, but may also be involved in conformational changes of Aβ, and in the clearance of Aβ from the brain via phagocytosis or active transport across the blood–brain barrier. Thus, both professional and amateur chaperones may be involved in the aggregation, accumulation, persistence, and clearance of Aβ and tau and in other Aβ-associated reactions such as inflammation associated with AD lesions, and may, therefore, serve as potential targets for therapeutic intervention.     

Dietary Supplementation with tBHQ,an Nrf2 Stabilizer Molecule,Confers Neuroprotection Against Apoptosis in Amyloid β-Injected Rat

Nuclear factor erythroid 2-related factor 2 (Nrf2) coordinates the up-regulation of cytoprotective genes via the antioxidant response element (ARE). There is significant evidence that oxidative stress is a critical event in the pathogenesis of AD. Considering the protective role of Nrf2 against oxidative injury, we studied to determine whether in vivo toxicity of amyloid β (Aβ) can be attenuated by tBHQ, an Nrf2 stabilizer, Using an Aβ injection model. We demonstrated that pre-activation of endogenous Nrf2 by tBHQ attenuated Aβ-induced caspase-3 expression. tBHQ enhanced GSH, decreased MDA level, and inhibited NF-κB. This investigation provides the first documentation of tBHQ’s neuroprotective effect through decrease of Aβ accumulation in rat brain. Our results show the involvement of Hsp-70 in this protective effect. In summary tBHQ treatment for 1 week prior to Aβ injection protected against the oxidative damage, apoptosis and Aβ accumulation in rats.     

Role of A β and the α 7 nicotinic acetylcholine receptor in regulating synaptic plasticity in Alzheimer's disease

Alzheimer's disease (AD) is caused by the accumulation of β-amyloid protein (Aβ) in the brain. The aggregation of β-amyloid protein to higher molecular weight fibrillar forms is also considered to be an important step in the pathogenesis of the disease. The memory problems associated with AD are likely to be caused by changes in synaptic plasticity. Recent studies suggest that Aβ binds to the α 7 nicotinic acetylcholine receptor (α 7 nAChR), which plays an important role in synaptic plasticity and memory. A loop domain localized towards the C-terminus of the extracellular region of the receptor has been identified as forming part of a putative Aβ-binding site. In cell culture experiments, the binding of Aβ to the α 7 nAChR has been found to cause an increase in the level of acetylcholinesterase, which is also increased around amyloid plaques in the AD brain. These studies indicate that the Aβ-binding site on the α 7 nAChR receptor is an important new target for therapeutic development in AD.     

Cholesterol Potentiates β-Amyloid-Induced Toxicity in Human Neuroblastoma Cells: Involvement of Oxidative Stress

Alterations in brain cholesterol concentration and metabolism seem to be involved in Alzheimer’s disease (AD). In fact, several experimental studies have reported that modification of cholesterol content can influence the expression of the amyloid precursor protein (APP) and amyloid β peptide (Aβ) production. However, it remains to be determined if changes in neuronal cholesterol content may influence the toxicity of Aβ peptides and the mechanism involved. Aged mice, AD patients and neurons exposed to Aβ, show a significant increase in membrane-associated oxidative stress. Since Aβ is able to promote oxidative stress directly by catalytically producing H₂O₂ from cholesterol, the present work analyzed the effect of high cholesterol incorporated into human neuroblastoma cells in Aβ-mediated neurotoxicity and the role of reactive oxygen species (ROS) generation. Neuronal viability was studied also in the presence of 24S-hydroxycholesterol, the main cholesterol metabolite in brain, as well as the potential protective role of the lipophilic statin, lovastatin. Special issue article in honor of Dr. Ricardo Tapia.     

Roles of amyloid beta-peptide-associated oxidative stress and brain protein modifications in the pathogenesis of Alzheimer's disease and mild cognitive impairment

Oxidative stress has been implicated to play a crucial role in the pathogenesis of a number of diseases, including neurodegenerative disorders, cancer, and ischemia, just to name a few. Alzheimer disease (AD) is an age-related neurodegenerative disorder that is recognized as the most common form of dementia. AD is histopathologically characterized by the presence of extracellular amyloid plaques, intracellular neurofibrillary tangles, the presence of oligomers of amyloid beta-peptide (Abeta), and synapse loss. In this review we discuss the role of Abeta in the pathogenesis of AD and also the use of redox proteomics to identify oxidatively modified brain proteins in AD and mild cognitive impairment. In addition, redox proteomics studies in in vivo models of AD centered around human Abeta(1-42) are discussed.     

The role of cytochrome <Emphasis Type="Italic">c</Emphasis> oxidase deficiency in ROS and amyloid plaque formation

The multiple dysfunctional changes associated with a brain affected with Alzheimer’s disease (AD) makes the understanding of primary pathogenic mechanisms challenging. Mitochondrial dysfunction has been associated with almost every neurodegenerative disease and neurodegenerative-related event. Alzheimer’s disease is no exception with data suggesting mitochondrial malfunctions ranging from improper organelle dynamics, defective oxidative phosphorylation (OXPHOS), oxidative stress, and harmful beta amyloid (Aβ) associations with the mitochondria. A major change often associated with AD is impairment of the electron transport chain at complex IV: cytochrome c oxidase (COX). This mini-review concentrates on recent work by our group that sheds light on the role COX deficiency plays in the pathophysiology of AD using a transgenic mouse model. Results suggest that neuronal COX deficiency does not increase oxidative stress and nor increases amyloidal formations in vivo. Conclusions from this work also suggest that Aβ formation is a cause of COX deficiency as opposed to the consequence.     

Proteomic identification of nitrated brain proteins in amnestic mild cognitive impairment: a regional study

Oxidative stress is an imbalance between the level of antioxidants and oxidants in a cell. Oxidative stress has been shown in brain of subjects with mild cognitive impairment (MCI) as well Alzheimer's disease (AD). MCI is considered as a transition phase between control and AD. The focus of the current study was to identify nitrated proteins in the hippocampus and inferior parietal lobule (IPL) brain regions of subjects with amnestic MCI using proteomics. The identified nitrated proteins in MCI brain were compared to those previously reported to be nitrated and oxidatively modified in AD brain, a comparison that might provide an invaluable insight into the progression of the disease.     

Amyloid β-protein toxicity and oxidative stress in Alzheimer’s disease

Alzheimer’s disease (AD) is a neurodegenerative disorder characterized by loss of memory and progressive decline of cognitive abilities. Although the pathogenesis of this disease is not known and is still under intensive investigation, there are several hypotheses which address certain aspects of the disease. This review focuses on the oxidative-stress hypothesis of AD and on novel antioxidative approaches to an effective neuroprotection for the prevention and therapy of this neurodegenerative disorder. The toxicity of the AD-associated amyloid β-protein (Aβ), the induction of oxidative stress by Aβ in neurons, and potential sources of oxidative events in brain tissue are discussed. Received: 20 February 1997 / Accepted: 9 May 1997