Jennifer Drake

Evidence of oxidative damage in Alzheimer's disease brain: central role for amyloid β-peptide

Amyloid beta-peptide (Abeta) is heavily deposited in the brains of Alzheimers disease (AD) patients. Free-radical oxidative stress, particularly of neuronal lipids, proteins and DNA, is extensive in those AD brain areas in which Abeta is abundant. Recent research suggests that these observations might be linked, and it is postulated that Abeta-induced oxidative stress leads to neurodegeneration in AD brain. Consonant with this postulate, Abeta leads to neuronal lipid peroxidation, protein oxidation and DNA oxidation by means that are inhibited by free-radical antioxidants. Here, we summarize current research on phospholipid peroxidation, as well as protein and DNA oxidation, in AD brain, and discuss the potential role of Abeta in this oxidative stress.

Evidence that amyloid beta-peptide-induced lipid peroxidation and its sequelae in Alzheimer's disease brain contribute to neuronal death

Amyloid beta-peptide [Abeta(1-42)] is central to the pathogenesis of Alzheimers disease (AD), and the AD brain is under intense oxidative stress, including membrane lipid peroxidation. Abeta(1-42) causes oxidative stress in and neurotoxicity to neurons in mechanisms that are inhibited by Vitamin E and involve the single methionine residue of this peptide. In particular, Abeta induces lipid peroxidation in ways that are inhibited by free radical antioxidants. Two reactive products of lipid peroxidation are the alkenals, 4-hydroxynonenal (HNE) and 2-propenal (acrolein). These alkenals covalently bind to synaptosomal protein cysteine, histidine, and lysine residues by Michael addition to change protein conformation and function. HNE or acrolein binding to proteins introduces a carbonyl to the protein, making the protein oxidatively modified as a consequence of lipid peroxidation. Immunoprecipitation of proteins from AD and control brain, obtained no longer than 4h PMI, showed selective proteins are oxidatively modified in the AD brain. Creatine kinase (CK) and beta-actin have increased carbonyl groups, and Glt-1, a glutamate transporter, has increased binding of HNE in AD. Abeta(1-42) addition to synaptosomes also results in HNE binding to Glt-1, thereby coupling increased Abeta(1-42) in AD brain to increased lipid peroxidation and its sequelae and possibly explaining the mechanism of glutamate transport inhibition known in AD brain. Abeta also inhibits CK. Implications of these findings relate to decreased energy utilization, altered assembly of cytoskeletal proteins, and increased excitotoxicity to neurons by glutamate, all reported for AD. The epsilon-4 allele of the lipid carrier protein apolipoprotein E (APOE) allele is a risk factor for AD. Synaptosomes from APOE knock-out mice are more vulnerable to Abeta-induced oxidative stress (protein oxidation, lipid peroxidation, and ROS generation) than are those from wild-type mice. Further, synaptosomes from allele-specific APOE knock-in mice have tiered vulnerability to Abeta(1-42)-induced oxidative stress, with APOE4 more vulnerable to Abeta(1-42) than are those from APOE2 or APOE3 mice. These results are consistent with the notion of a coupling of the oxidative environment in AD brain and increased risk of developing this disorder. Taken together, the findings from in-vitro studies of lipid peroxidation induced by Abeta(1-42) and postmortem studies of lipid peroxidation (and its sequelae) in AD brain may help explain the APOE allele-related risk for AD, some of the functional and structural alterations in AD brain, and strongly support a causative role of Abeta(1-42)-induced oxidative stress in AD neurodegeneration.

Nutritional approaches to combat oxidative stress in Alzheimer’s disease

Alzheimers disease (AD) brains are characterized by extensive oxidative stress. Additionally, large depositions of amyloid beta-peptide (Abeta) are observed, and many researchers opine that Abeta is central to the pathogenesis of AD. Our laboratory combined these two observations in a comprehensive model for neurodegeneration in AD brains centered around Abeta-induced oxidative stress. Given the oxidative stress in AD and its potentially important role in neurodegeneration, considerable research has been conducted on the use of antioxidants to slow or reverse the pathology and course of AD. One source of antioxidants is the diet. This review examines the literature of the effects of endogenous and exogenous, nutritionally-derived antioxidants in relation to AD. In particular, studies of glutathione and other SH-containing antioxidants, vitamins, and polyphenolic compounds and their use in AD and modulation of Abeta-induced oxidative stress and neurotoxicity are reviewed.

Oxidative stress precedes fibrillar deposition of Alzheimer's disease amyloid β-peptide (1-42) in a transgenic Caenorhabditis elegans model

Alzheimers disease is a progressive, neurodegenerative disorder characterized by senile plaques and neurofibrillary components. Abeta(1-42) is a principal component of senile plaques and is thought to be central to the pathogenesis of the disease. The Alzheimers disease brain is under significant oxidative stress, and the Abeta(1-42) peptide is known to cause oxidative stress in vitro. One controversy in the amyloid hypothesis is whether or not Abeta plaques are required for toxicity. We have employed a temperature-inducible Abeta expression system in Caenorhabditis elegans to create a strain of worms, CL4176, in which Abeta(1-42) is expressed with a non-permissive temperature of 23 degrees C. The CL4176 strain allows examination of the temporal relationship between Abeta expression, oxidative stress, and Abeta fibril formation. CL4176 were under increased oxidative stress, evidenced by increased protein oxidation indexed by increased carbonyl levels, 24 and 32 h after temperature upshift as compared to the control strain, CL1175. The increased oxidative stress in CL4176 occurred in the absence of Abeta fibril formation, consistent with the notion that the toxic species in Abeta toxicity is pre-fibrillar Abeta and not the Abeta fibril. These results are discussed with reference to Alzheimers disease.

Peroxynitrite-induced alterations in synaptosomal membrane proteins : insight into oxidative stress in Alzheimer's disease

Abstract : Peroxynitrite (ONOO‐) is a highly reactive, oxidizing anion with a half‐life of <1 s that is formed by reaction of superoxide radical anion with nitric oxide. Several reports of ONOO‐ ‐induced oxidation of lipids, proteins, DNA, sulfhydryls, and inactivation of key enzymes have appeared. ONOO‐ has also been implicated as playing a role in the pathology of several neurodegenerative disorders, such as Alzheimers disease (AD) and amyotrophic lateral sclerosis, among others. Continuing our laboratorys interest in free radical oxidative stress in brain cells in AD, the present study was designed to investigate the damage to brain neocortical synaptosomal membrane proteins and the oxidation‐sensitive enzyme glutamine synthetase (GS) caused by exposure to ONOO‐. These synaptosomal proteins and GS have previously been shown by us and others to have been oxidatively damaged in AD brain and also following treatment of synaptosomes with amyloid β‐peptide. The results of the current study showed that exposure to physiological levels of ONOO‐ induced significant protein conformational changes, demonstrated using electron paramagnetic resonance in conjunction with a protein‐specific spin label, and caused oxidation of proteins, measured by the increase in protein carbonyls. ONOO‐ also caused inactivation of GS and led to neuronal cell death examined in a hippocampal cell culture system. All these detrimental effects of ONOO‐ were successfully attenuated by the thiol‐containing antioxidant tripeptide glutathione. This research shows that ONOO‐ can oxidatively modify both membranous and cytosolic proteins, affecting both their physical and chemical nature. These findings are discussed with reference to the potential involvement of ONOO‐ in AD neurodegeneration.

Vitamin E and Neurodegenerative Disorders Associated with Oxidative Stress

Abstract Several neurodegenerative disorders are associated with oxidative stress that is manifested by lipid peroxidation, protein oxidation and other markers. Included in these disorders in which oxidative stress is thought to play an important role in their pathogenesis are Alzheimers disease (AD), Parkinsons disease (PD), amyotrophic lateral sclerosis (ALS), tardive dyskinesia, Huntingtons disease (HD), and multiple sclerosis. This review presents some of the chemistry of vitamin E as an antioxidant and summarizes studies in which vitamin E has been employed in these disorders and models thereof.

Elevated oxidative stress in models of normal brain aging and Alzheimer's disease

Age-associated neurodegenerative disorders are becoming more prevalent as the mean age of the population increases in the United States over the next few decades. Both normal brain aging and Alzheimers disease (AD) are associated with oxidative stress. Our laboratory has used a wide variety of physical and biochemical methods to investigate free radical oxidative stress in several models of aging and AD. Beta-amyloid (A beta), the peptide that constitutes the central core of senile plaques in AD brain, is associated with free radical oxidative stress and is toxic to neurons. This review summarizes some of our studies in aging and A beta-associated free radical oxidative stress and on the modulating effects of free radical scavengers on neocortical synaptosomal membrane damage found in aging and A beta-treated systems.

Elevation of brain glutathione by γ-glutamylcysteine ethyl ester protects against peroxynitrite-induced oxidative stress

Elevation of glutathione (GSH) has been recognized as an important method for modulating levels of reactive oxygen species (ROS) in the brain. We investigated the antioxidant properties of γ‐glu‐cys‐ethyl ester (GCEE) in vitro and its ability to increase GSH levels upon in vivo i.p. injection. GCEE displays antioxidant activity similar to GSH as assessed by various in vitro indices such as hydroxyl radical scavenging, dichlorofluorescein fluorescence (DCF), protein specific spin labeling, glutamine synthetase (GS) activity, and protein carbonyls. Intraperitoneal injection of GCEE to gerbils resulted in a 41% increase in brain total GSH levels in vivo as determined by the DTNB‐GSH reductase recycling method. Gerbils injected with buthionine sulfoximine (BSO), an inhibitor of γ‐glutamylcysteine synthetase, had 40% less total brain glutathione. Gerbils injected with BSO followed by a GCEE injection had GSH levels similar to vehicle‐injected controls, suggesting that GCEE upregulates GSH biosynthesis by providing γ‐glutamylcysteine and not cysteine. Cortical synaptosomes from GCEE‐injected animals were less susceptible to peroxynitrite‐induced oxidative damage as assessed by DCF fluorescence, protein‐specific spin labeling, and GS activity. These experiments suggest that GCEE is effective in increasing brain GSH levels and may potentially play an important therapeutic role in attenuating oxidative stress in neurodegenerative diseases associated with oxidative stress such as Alzheimer disease.

The Free Radical Antioxidant Vitamin E Protects Cortical Synaptosomal Membranes from Amyloid β-Peptide(25-35) Toxicity But Not from Hydroxynonenal Toxicity: Relevance to the Free Radical Hypothesis of Alzheimer's Disease

Amyloid β-peptide (Aβ) is a key factor in the neurotoxicity of Alzheimers disease (AD). Recent research has shown that Aβ-mediated neurotoxicity involves free radicals and that Aβ peptides can initiate multiple membrane alterations, including protein oxidation and lipid peroxidation, eventually leading to neuronal cell death. Research also has emphasized the role of 4-hydroxynonenal (HNE), a downstream product of lipid peroxidation, in being able to mimic some of the effects of Aβ peptides. In the current investigation, electron paramagnetic resonance (EPR) studies of spin labeled cortical synaptosomal membrane proteins has been employed to study conformational changes in proteins, spectrophotometric methods have been used to measure protein carbonyl content, and the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay for mitochondrial function has been used to study the effect of vitamin E on samples that were treated with Aβ or HNE. The free radical dependence of β-amyloid-associated toxicity was confirmed by the ability of the free radical scavenger vitamin E to prevent the toxic effects of Aβ. In contrast, HNE was still toxic in the presence of vitamin E. These results support our Aβ-associated free radical model for neurotoxicity in AD brain and are discussed with reference to potential therapeutic strategies for AD.

Role of the proteasome in protein oxidation and neural viability following low‐level oxidative stress

Numerous studies suggest that proteasome inhibition may play a causal role in mediating the increased levels of protein oxidation and neuron death observed in conditions associated with oxidative stress. In the present study we demonstrate that administration of non‐toxic levels of oxidative stress does not result in impairment of 20S/26S proteasome activity, and actually increases the expression of specific proteasome subunits. Non‐toxic levels of oxidative stress were observed to elevate the amount of protein oxidation in the presence of preserved proteasomal function, suggesting that proteasome inhibition may not mediate increases in protein oxidation following low‐level oxidative stress. Preserving basal proteasome function appears to be critical to preventing the neurotoxicity of low‐level oxidative stress, based on the ability of proteasome inhibitor treatment to exacerbate oxidative stress toxicity. Taken together, these data indicate that maintaining neural proteasome function may be critical to preventing neurotoxicity, but not the increase in protein oxidation, following low‐level oxidative stress.