Serial Review: Causes and Consequences of Oxidative Stress inAlzheimers DiseaseGuest Editors: Mark A. Smith and George Perry

AMYLOID- AND SERVE ANTIOXIDANT FUNCTIONS IN THE AGINGAND ALZHEIMER BRAIN

MARK A. SMITH,* GEMMA CASADESUS, JAMES A. JOSEPH, and GEORGE PERRY**Institute of Pathology, Case Western Reserve University, Cleveland, OH, USA; and USDA-Human Nutrition Research Center on

Aging at Tufts University, Boston, MA, USA

(Received 25 February 2002; Revised 11 July 2002; Accepted 11 July 2002)

AbstractHistorically, amyloid- and (tau), the major components of senile plaques and neurofibrillary tangles,respectively, have been considered central mediators of the pathogenesis of Alzheimer disease. Therefore, efforts tounderstand disease mechanisms have concentrated on understanding either the processes involved in amyloid-deposition as senile plaques or on the phosphorylation and aggregation of as neurofibrillary tangles. However, in lightof recent evidence, such lesion-centric approaches look to be inappropriate. In fact, rather than initiators of diseasepathogenesis, the lesions occur consequent to oxidative stress and function as a primary line of antioxidant defense.Given this, it is perhaps not surprising that the increased sensitivity to oxidative stress in the aged brain, even in controlindividuals, is invariably marked by the appearance of both amyloid- and tau. Additionally, in Alzheimer disease,where chronic oxidative stress persists and is superimposed upon an age-related vulnerable environment, one wouldpredict, and there is, an increased lesion load. The notion that amyloid- and function as protective components bringsinto serious question the rationale of current therapeutic efforts targeted toward lesion removal. 2002 ElsevierScience Inc.

KeywordsAlzheimer disease, Amyloid-, Antioxidant, Free radical, Phosphorylation, Redox-active metals, Tau

INTRODUCTION

Amyloid- and the microtubule-associate protein tau ()are, without a shadow of a doubt, the best-studied pro-teins relating to the pathogenesis of Alzheimer disease(AD). While perhaps not surprising since the pathologi-cal diagnosis of Alzheimer disease is dependent uponboth amyloid- and depositions [1,2], the amalgam-ation of diagnostic and mechanistic views relating to thedisease has unfortunately led researchers astray. Amy-loid- and are crucial diagnostic indicators; however,as we discuss below, their mechanistic importance hasfar less to do with their consequences than with the

factors that led to their formation. Thus, the goal of thisreview is to present an alternative hypothesis for the roleof amyloid- and neurofibrillary tangle (NFT) depositionin this disease.

Amyloid-

The overwhelming view currently held concerningdisease pathogenesis is the premise that amyloid-causes the disease [3]. Champions of the amyloid-hypothesis argue that diagnostic, clinical, and patholog-ical studies all support a central role for amyloid-. First,amyloid- is an obligate feature of the disease and cor-relates, albeit weakly, with disease severity [4]. Second,amyloid- is often, though not always, found in regionsof the brain that degenerate during the disease and amy-loid- is a potent neurotoxic agent in vitro. Third, andmost touted, both in vivo in the disease and in vitro intransfected cells, amyloid- is increased by all of themutated genes (including the source of amyloid-,

1This article is part of a series of reviews on Causes and Conse-quences of Oxidative Stress in Alzheimers Disease. The full list ofpapers may be found on the homepage of the journal.

Address correspondence to: Mark A. Smith, Ph.D., Institute ofPathology, Case Western Reserve University, 2085 Adelbert Road,Cleveland, OH 44106, USA; Tel; (216) 368-3670; Fax: (216) 368-8964; E-Mail: mas21@po.cwru.edu.

Free Radical Biology & Medicine, Vol. 33, No. 9, pp. 11941199, 2002Copyright 2002 Elsevier Science Inc.Printed in the USA. All rights reserved

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APP) that are associated with the autosomal dominantinheritance of AD [5]. However, the aforementionedpoints are, at best, circumstantial and may also be used asevidence that amyloid- is playing a protective role.First, the correlation of amyloid- with dementia (i.e.,neuronal dysfunction and loss) appears to be a protectivecompensation mounted in response to the underlyingdisease process [6]. Indeed, there is insurmountable ev-idence that oxidative stress is one of, if not the, earliestpathological alteration in the disease [7,8]. Importantly,neurons respond to oxidative stress, both in vitro and invivo, by increasing amyloid- production [9], and thisincreased amyloid- is associated with a consequentreduction in oxidative stress [7,8]. Proteins, such asamyloid-, that are induced under oxidative conditionsand act to lessen oxidative damage, are typically thoughtof as antioxidants and, in this regard, we recently dem-onstrated that amyloid- is a bona fide antioxidant thatcan act as a potent superoxide dismutase [10].

Viewing amyloid- as a protective response elementprovides a valid mechanism for why the brains of almosteveryone over the age of 40, an age, coincidently, whereredox alterations first manifest [8], contain amyloid-deposits. The alternate view, that everyone at mid-life ison the verge of developing AD is not only unsound froma biological perspective but also does a great disserviceto the large percentage of cognitively normal aged indi-viduals whose brains contain amyloid- loads equivalentto patients with AD [11]. Indeed, although fibrillar oraggregated forms of amyloid-, like those present in thesenile plaques, cause cytotoxicity in vitro [12], the pres-ence and density of amyloid- in vivo correlates weaklywith the onset and severity of AD [13]. Instead, thepresence of the soluble form of amyloid- in the brainmay be a better predictor of the disease [14]. Specifi-cally, SDS-stable oligomers and not monomers of thisform of amyloid- seem to play an important role, asshown by augmented presence of these oligomers duringthe expression of mutations in APP or presenilin [15],as well as by their capacity to inhibit neuronal plasticityparameters (LTP) in vivo when micro-injected into thebrains of rodents [16]. Conversely, amyloid- is notalways present in the brains of cognitively normal el-derly. Nevertheless, these differences can be explained interms of genetic and environmental variability acrossindividuals. One possibility is that some people are ge-netically endowed with more efficient endogenous anti-oxidant defense systems and thus age better/slower. Al-ternatively, these people may have supplemented theirdiets with foods rich in antioxidants throughout theirlifespan, compensating for age-related declines in theseendogenous systems and thus slowing down oxidativestress-related declines seen during aging [1719]. Ifamyloid- and NFT deposition provides an antioxidant

function, it is likely that these processes will be recruitedduring times when oxidative stress is high and the en-dogenous antioxidant-defenses are compromised. Never-theless, if these systems remain relatively efficient or aresupported by exogenous antioxidant supplementation,the presence of amyloid- and NFT may not be neces-sary, and thus lead to little amyloid- and NFT deposi-tion. Preliminary data from in vitro studies support thishypothesis. Incubation of primary cortical neurons withblueberry extract, a fruit rich in antioxidants [1719],prevents phosphorylation when neurons are presentedwith an oxidative stress insult (Casadesus, Smith, andJoseph, in preparation), analogous to the effects of en-dogenous antioxidants [20].

Moreover, unbiased stereological counting indicatesthat during normal aging there may be little or no cellloss despite, as pointed out above, the presence of anincreasing number of plaques [21]. This testifies to theprotective function of amyloid-. Importantly, even thehyper-physiologic levels of amyloid- found in engi-neered AD transgenic mice [22] only lead to senileplaque formation in middle-aged mice and are, like theirhuman counterparts, preceded by oxidative stress [2325]. Taken together, these findings indicate that amy-loid- is not driving the pathogenic process, rather it is aconsequence of the pathogenic oxidative process thatserves an antioxidant function. In normal aging, theproduction and deposition of amyloid- successfullystaves off age-related redox imbalances, but in AD wherethere is a profound and chronic redox imbalance, thepresence of amyloid-, even at high levels, proves in-sufficient.

The idea that amyloid- is protective represents amajor paradigm shift but is really not surprising. Neuro-nal degeneration is associated with a number of re-sponses including the induction of heat shock proteinssuch as heme oxygenase-1 [26] and ubiquitin [27,28]that, like amyloid-, show a relationship with cognitivedecline. Why is it that only amyloid- is consideredpathogenic? The answer, according to the proponents ofthe amyloid- hypothesis is that amyloid- is neurotoxicin vitro and is associated with neuronal loss in vivo.However, that amyloid- is age- and region-specific isfurther consistent with a key role in redox homeostasis.Cell culture studies showing neurotoxicity may be arti-facts of in vitro conditions [29], an aspect further but-tressed by the fact that neither isolated senile plaques norimmobilized amyloid- elicit neurotoxicity in vivo or invitro [3032]. Nonetheless, as a bioactive substance, weshould remain cognizant that antioxidants can also serveas pro-oxidants, dependent upon the conditions and that,in certain nonphysiological circumstances, amyloid-can be made damaging [33,34]. In this regard, and al-though the capacity of amyloid- to induce oxidative

1195A and as antioxidants

stress remains controversial [35], recent data illustratethat the oxidant properties of amyloid- may stem fromits capacity to interact with transition metals and mediatetoxicity via redox-active ions that precipitates lipid per-oxidation and cellular oxidative stress [29].

Based on the evidence presented above, the few re-ports demonstrating, under some circumstances, neuro-nal loss in some transgenic mice with amyloid- deposits[36] argue only that amyloid- is a bioactive substance.Indeed, there is little evidence in the literature showingbehavioral deficits in mice transgenic for only APPmutations. The most consistent findings of behavioraldeficits have been shown in mice transgenic for morethan one mutation, e.g., APP/PS1 [37,38], and eventhen it is superimposed upon an aged environment.

Finally, the notion that genetic linkage mutationscauses increased amyloid- causes disease is simplistic,pays scant regard for biological and cellular homeostasis,and is clearly not as direct as often assumed, even in thecase of mutations in APP. While it is true that APPand presenilin-1 and -2 mutations are deterministic forthe onset of AD and that these mutations produce in-creases in amyloid- in the brain, it is against logic toconclude with certainty that there is a casual relationshipbetween the two. For example, and perhaps not surpris-ing, in light of the antioxidant function of amyloid-,oxidative stress is among the best inducers of APPprotein expression and consequent amyloid- production[9]. Therefore, if a mutation causes oxidant stress thenone would also find an increase in amyloid-. Indeed,together with the in vivo findings showing that increasedamyloid- deposition is associated with reduced oxida-tive stress [7,8], the finding of increased amyloid- withthe mutants simply highlights that perturbations in thecellular system lead to a protective response, i.e., in-creased amyloid- production.

Our arguments supporting amyloid- as a crucialantioxidant defense mechanism are extremely relevant tocurrent pharmacological efforts targeted at either remov-ing amyloid- or lessening amyloid- production, andwill likely leave neurons without one of their fundamen-tal compensatory responses to aging and disease.

Tau

As we show for amyloid-, even those aspects of ADthought most deleterious may actually play an importantrole in antioxidant defenses and this aspect appliesequally well to [8]. The accumulation of phosphory-lated as NFT in neurons appears to be an analogousprotective antioxidant response, since quantitative anal-ysis of the extent of oxidative damage in AD shows theoxidative damage is actually reduced in those neuronswith the most cytopathology [8]. For example, recent

data suggests that most neuronal loss in AD occurs priorto NFT deposition [39,40]. Interestingly, most neuronalloss during this period occurs when the levels of oxida-tive stress are highest and subsequent deposition of NFTdecreases these levels [41].

The importance of this protective aspect of isbrought to the fore if it is considered that protection ofcritical cellular components from oxidants can bethrough the incorporation of damage to others. Whilethe exhaustion of cellular reductants is usually used as ameasure of antioxidant potential, cellular macromole-cules may share a similar function. Consistent with thisview is the physiological modification of and neuro-filament proteins by lipid peroxidation products and car-bonyls [42,43]. Indeed, oxidative stress and attendantmodification of by products of oxidative stress includ-ing HNE [44] as well as other cytotoxic carbonyls [45],though leading to protein aggregation as NFT, enablessuch neurons to survive decades [46]. Intriguingly, al-though cytoskeletal proteins such as and neurofila-ments have a long half-life, the same extent of carbonylmodification is found throughout the normal aging pro-cess as well as along the length of the axon [47]. Thissuggests that the oxidative modification of cytoskeletalproteins is under tight regulation. Both and neurofila-ment protein appear uniquely adapted to oxidative attackdue to their high content of lysine-serine-proline (KSP)domains. Exposure of these domains on the protein sur-face is affected by extensive phosphorylation of serineresidues, resulting in an oxidative sponge of surface-modifiable lysine residues [47]. Since phosphorylationplays this pivotal role in redox balance, it is perhaps notsurprising that oxidative stress, through activation ofMAP kinase pathways, leads to phosphorylation [4851], nor that conditions associated with chronic oxidantstress, such as AD, are invariably associated with exten-sive phosphorylation of cytoskeletal elements. Indeed,other neurological conditions where phosphorylated and neurofilament protein accumulations occur, alsoshow evidence of oxidative adducts, e.g., progressivesupranuclear palsy [52] and frontal temporal dementia[53]. Given this protective role of phosphorylation, it isnot surprising that embryonic neurons that survive treat-ment with oxidants have more phospho- immunoreac-tivity relative to those that die [54]. Further, since hemeoxygenase induction and expression are opposing[20,44], the reduced oxidative damage in neurons with accumulation may be a part of the antioxidant function ofphosphorylated .

SUMMARY

While amyloid- and are often viewed as harbingersof disease, the observed decrease in oxidative damage

1196 M. A. SMITH et al.

with amyloid- and accumulation points towards analternate, contrary interpretation that they represent im-portant survival responses (Fig. 1) [6,37,5557]. If thisproves to be the case, current efforts targeted at theirelimination will actually exacerbate the disease.

Acknowledgements Work in the authors laboratories was funded bythe National Institutes of Health and the Alzheimers Association.

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