The mitochondria-targeted antioxidant MitoQ prevents loss of spatial memory retention and early neuropathology in a transgenic mouse model of Alzheimer's disease.
ABSTRACT Considerable evidence suggests that mitochondrial dysfunction and oxidative stress contribute to the progression of Alzheimer's disease (AD). We examined the ability of the novel mitochondria-targeted antioxidant MitoQ (mitoquinone mesylate: [10-(4,5-dimethoxy-2-methyl-3,6-dioxo-1,4-cycloheexadienl-yl) decyl triphenylphosphonium methanesulfonate]) to prevent AD-like pathology in mouse cortical neurons in cell culture and in a triple transgenic mouse model of AD (3xTg-AD). MitoQ attenuated β-amyloid (Aβ)-induced neurotoxicity in cortical neurons and also prevented increased production of reactive species and loss of mitochondrial membrane potential (Δψ(m)) in them. To determine whether the mitochondrial protection conferred by MitoQ was sufficient to prevent the emergence of AD-like neuropathology in vivo, we treated young female 3xTg-AD mice with MitoQ for 5 months and analyzed the effect on the progression of AD-like pathologies. Our results show that MitoQ prevented cognitive decline in these mice as well as oxidative stress, Aβ accumulation, astrogliosis, synaptic loss, and caspase activation in their brains. The work presented herein suggests a central role for mitochondria in neurodegeneration and provides evidence supporting the use of mitochondria-targeted therapeutics in diseases involving oxidative stress and metabolic failure, namely AD.
[show abstract] [hide abstract]
ABSTRACT: The aim of this article was to provide a survey of the clinical development of pharmacotherapy for Alzheimer's disease (AD). A search of PubMed to identify pertinent English-language literature was conducted using the terms Alzheimer's disease AND clinical trials (2003-2008), dementia AND prevention AND clinical trials (2003-2008), and the chemical names of all compounds mentioned in articles on new drugs for AD published since 2005. www.ClinicalTrials.gov was searched for relevant trials. Abstracts of the 2008 International Conference on Alzheimer's Disease (ICAD) were reviewed for relevance, as were pharmaceutical company and AD advocacy Web sites. Articles selected for review were primary reports of data from preclinical studies and clinical trials. A large number of drugs with differing targets and mechanisms of action are under development for the treatment of AD. Phase III trials of Ginkgo biloba, NSAIDs, phenserine, statins, tarenflurbil, tramiprosate, and xaliproden have been completed, none of them demonstrating adequate efficacy. Encouraging results from completed Phase II trials of dimebon, huperzine A, intravenous immunoglobulin, and methylthioninium chloride were reported at ICAD 2008. Nineteen compounds are currently in Phase II trials, and 3 compounds (AN1792, lecozotan SR, and SGS742) failed at this stage of development. Despite disappointing results from recently completed Phase III trials of several novel compounds, the extent and breadth of activity at all phases of clinical development suggest that new pharmacotherapeutic options for the treatment of AD will become available within the next decade.The American Journal of Geriatric Pharmacotherapy 07/2009; 7(3):167-85. · 2.67 Impact Factor
Article: Structure-activity analyses of beta-amyloid peptides: contributions of the beta 25-35 region to aggregation and neurotoxicity.[show abstract] [hide abstract]
ABSTRACT: The neurodegeneration of Alzheimer's disease has been theorized to be mediated, at least in part, by insoluble aggregates of beta-amyloid protein that are widely distributed in the form of plaques throughout brain regions affected by the disease. Previous studies by our laboratory and others have demonstrated that the neurotoxicity of beta-amyloid in vitro is dependent upon its spontaneous adoption of an aggregated structure. In this study, we report extensive structure-activity analyses of a series of peptides derived from both the proposed active fragment of beta-amyloid, beta 25-35, and the full-length protein, beta 1-42. We examine the effects of amino acid residue deletions and substitutions on the ability of beta-amyloid peptides to both form sedimentable aggregates and induce toxicity in cultured hippocampal neurons. We observe that significant levels of peptide aggregation are always associated with significant beta-amyloid-induced neurotoxicity. Further, both N- and C-terminal regions of beta 25-35 appear to contribute to these processes. In particular, significant disruption of peptide aggregation and toxicity result from alterations in the beta 33-35 region. In beta 1-42 peptides, aggregation disruption is evidenced by changes in both electrophoresis profiles and fibril morphology visualized at the light and electron microscope levels. Using circular dichroism analysis in a subset of peptides, we observed classic features of beta-sheet secondary structure in aggregating, toxic beta-amyloid peptides but not in nonaggregating, nontoxic beta-amyloid peptides. Together, these data further define the primary and secondary structures of beta-amyloid that are involved in its in vitro assembly into neurotoxic peptide aggregates and may underlie both its pathological deposition and subsequent degenerative effects in Alzheimer's disease.Journal of Neurochemistry 02/1995; 64(1):253-65. · 4.06 Impact Factor
Article: Mitochondria-targeted antioxidants protect Friedreich Ataxia fibroblasts from endogenous oxidative stress more effectively than untargeted antioxidants.[show abstract] [hide abstract]
ABSTRACT: Friedreich Ataxia (FRDA), the most common inherited ataxia, arises from defective expression of the mitochondrial protein frataxin, which leads to increased mitochondrial oxidative damage. Therefore, antioxidants targeted to mitochondria should be particularly effective at slowing disease progression. To test this hypothesis, we compared the efficacy of mitochondria-targeted and untargeted antioxidants derived from coenzyme Q10 and from vitamin E at preventing cell death due to endogenous oxidative stress in cultured fibroblasts from FRDA patients in which glutathione synthesis was blocked. The mitochondria-targeted antioxidant MitoQ was several hundredfold more potent than the untargeted analog idebenone. The mitochondria-targeted antioxidant MitoVit E was 350-fold more potent than the water soluble analog Trolox. This is the first demonstration that mitochondria-targeted antioxidants prevent cell death that arises in response to endogenous oxidative damage. Targeted antioxidants may have therapeutic potential in FRDA and in other disorders involving mitochondrial oxidative damage.The FASEB Journal 11/2003; 17(13):1972-4. · 5.71 Impact Factor
(AD). We examined the ability of the novel mitochondria-targeted antioxidant MitoQ (mitoquinone mesylate: [10-(4,5-dimethoxy-2-
tical neurons in cell culture and in a triple transgenic mouse model of AD (3xTg-AD). MitoQ attenuated ?-amyloid (A?)-induced
neurotoxicity in cortical neurons and also prevented increased production of reactive species and loss of mitochondrial membrane
of AD-like neuropathology in vivo, we treated young female 3xTg-AD mice with MitoQ for 5 months and analyzed the effect on the
accumulation, astrogliosis, synaptic loss, and caspase activation in their brains. The work presented herein suggests a central role for
Alzheimer’s disease (AD) is the most prevalent of the neurode-
generative diseases and the leading cause of dementia among the
gression of this devastating illness. Extensive evidence indicates
that alterations in glucose metabolism and mitochondrial ener-
getics occur along with the accumulation of oxidative damage
before the cardinal signs of AD pathogenesis in the brains of
transgenic animal models and patients (Hoyer, 1996; Hirai et al.,
2001; Rinaldi et al., 2003; Butterfield et al., 2006; Lin and Beal,
2006; Sultana et al., 2006a,b; Mosconi et al., 2008a; Yao et al.,
2009). Such findings have inspired clinical trials of various anti-
oxidant compounds for treating AD. To date, none of the tested
compounds have proven effective (Sano et al., 1997; Petersen
et al., 2005; Pham and Plakogiannis, 2005; DeKosky et al.,
2008). One possible explanation for these failures is that the
species (RS), perhaps because they did not achieve sufficient con-
centrations at the site of most RS production, mitochondria.
ily traverse the blood–brain barrier and selectively concentrate in
mitochondria (Shigenaga et al., 1994; Halliwell and Gutteridge,
2007). The recently developed, mitochondria-targeted antioxidant
MitoQ (mitoquinone mesylate: [10-(4,5-dimethoxy-2-methyl-3,6-
dioxo-1,4-cycloheexadienl-yl) decyl triphenylphosphonium meth-
anesulfonate]) possesses these qualities (James et al., 2005, 2007;
MitoQ is produced by covalently binding ubiquinone, an
endogenous antioxidant and component of the mitochondrial
electron transport chain to a triphenylphosphonium (TPP?)
cation. MitoQ rapidly crosses the blood–brain barrier and
neuronal membranes and concentrates several hundred-fold
in mitochondria driven by the high membrane potential
across the inner mitochondrial membrane (IMM; Murphy
and Smith, 2007). The ubiquinone moiety is delivered to the
generation (Halliwell and Gutteridge, 2007). The TPP?moi-
ety adsorbs to the matrix side of the IMM, and the ubiquinone
penetrates into the membrane in which it is reduced to the
active antioxidant ubiquinol by respiratory complex II. The
ubiquinol acts as an antioxidant when oxidized to ubiquinone
by RS. Complex II then reduces the ubiquinone to ubiquinol.
MitoQ is a poor substrate for complex I and is negligibly oxi-
dized by complex III. Therefore, it cannot substitute for en-
dogenous ubiquinone in the electron transport chain but
primarily acts as an antioxidant capable of continuous regen-
TheJournalofNeuroscience,November2,2011 • 31(44):15703–15715 • 15703
eration by complex II (James et al., 2005, 2007; Rodriguez-
Cuenca et al., 2010).
To evaluate the therapeutic potential of MitoQ for treating
AD and to examine a role for mitochondria-generated RS in AD
progression, we investigated the effect of MitoQ on ?-amyloid
(A?) toxicity in primary cultures of mouse cortical neurons and
on cognitive deficits and neuropathology in a young triple trans-
genic (3xTg-AD) mouse model of AD. We found that MitoQ
treatment prevented A?-induced oxidative stress and death of
tive stress, A? accumulation, synaptic loss, astrogliosis, and
caspase activation in the brains of 3xTg-AD mice.
Reagents. All MitoQ used in these studies was synthesized as described
previously (Kelso et al., 2001). MitoQ is deliquescent, making pure Mi-
toQ difficult to work with. For this reason, it was adsorbed to cyclodex-
widely used in drug delivery and is not thought to have any significant
effects. For the in vitro experiments, pure MitoQ without cyclodextrin
was used. The control compound, decyl triphenylphosphonium bromide
(dTPP), was purchased from Santa Cruz Biotechnology and was not com-
deliquescent. 5-(and -6)-Chloromethyl-2?, 7?-dicholorodihydrofluorescein
diacetate (CM-H2DCFDA) and tetramethyl rhodamine methyl ester
(TMRM?) were purchased from Invitrogen. A?(22–35)and A?(1–40)pep-
Mice. The 3xTg-AD mouse model used in this study expresses three
mutant human genes: amyloid precursor protein, APPswe; presenilin-1,
PS1M146V; and four-repeat tau, tauP301L(Oddo et al., 2003b). The first
two are associated with early-onset forms of human AD and the latter
with human frontotemporal dementia. These mice and nontransgenic
(nonTg) mice from the same 129/C57BL/6 hybrid background strain
were kindly provided by Dr. Frank LaFerla (University of California,
an age-dependent manner. There is a transient sex divergence in
3xTg-AD pathology, such that young- to middle-aged (6–12 month)
age-matched male 3xTg-AD (Clinton et al., 2007). Furthermore, mito-
chondrial function and oxidative stress have been fully characterized
was to determine whether mitochondria-targeted therapeutics could
prevent the onset of the behavioral and neuropathological hallmarks of
AD in these mice. Therefore, we chose to study only female transgenic
and wild-type mice to minimize variability (attributable to sex differ-
ences) and the number of mice necessary to achieve meaningful results.
The mice were group housed and kept on a 12 h light/dark schedule.
All mice were given ad libitum access to food and water. Starting at 2
(100 ?M) or the negative control for MitoQ (dTPP; 100 ?M) continu-
ously in their drinking water for 5 months. Other age-matched female
3xTg-AD and female nonTg mice received water without MitoQ or
dTPP. When administered to wild-type mice for several months, MitoQ
has no effect on the function of subsequently isolated mitochondria, on
mtDNA, on nuclear or mitochondrial gene expression, on whole-body
metabolism, on whole-body motor function, or on food and liquid con-
sumption (Rodriguez-Cuenca et al., 2010). Each mouse was handled
animal procedures were in accordance with the National Institutes of
Health Guide for the Care and Use of Laboratory Animals.
the cortices were digested in 2 mg/ml trypsin in Ca2?/Mg2?-free HBSS
growth media (Neurobasal-A medium containing 2% B27 supplement,
1% penicillin/streptomycin, 0.1% L-glutamine) and distributed to poly-
plates (Corning). The cells were seeded in 100 ?l of growth media for a
minimum of 2 h in a 5% CO2atmosphere (35°C) in a humidified cell
culture incubator. The media were then aspirated and replaced. Cover-
slips were maintained in growth medium in 35 mM plastic cell culture
replaced at 3–4 d in vitro, and experimental treatments began on day 7.
Cortical neuron survival was determined by blinded counting of phase-
per treatment and was normalized to the average percentage survival of
nontreated, sibling cultures.
Microscopy. Microscopy experiments were conducted with a laser
attached to a Nikon Eclipse TE300 inverted microscope. Cells were ob-
pinhole and gain were maintained at constant levels during each experi-
ment. Laser power was 10% of maximum. Phase-contrast images were
ments) mounted on the Nikon TE-300 microscope and controlled by
MetaMorph software (Molecular Devices).
RS were detected using the redox-sensitive dye CM-H2DCFDA. This
dye is membrane permeant and is trapped in cells by binding of the
chloromethyl group to cellular thiols, primarily in the cytosol. The re-
idation by multiple RS, thus providing an assay of generalized RS
production (Royall and Ischiropoulos, 1993; Halliwell and Gutteridge,
oxidized by RS that lie downstream of dismutation of O2
hydroxyl radicals (OH?), peroxynitrite (ONOO?), and a number of
other radical and nonradical RS. We extensively characterized the use of
this dye in rat and mouse neurons (Kirkland and Franklin, 2001; Kirk-
land et al., 2002). This characterization shows that CM-H2DCFDA is
range, and is not photo-oxidized by the laser power and exposure time
used in these experiments. Cultures were incubated in the appropriate
35°C. They were then washed twice with Leibovitz’s L-15 medium and
with the 488 nm line of the confocal laser. The green photomultiplier
channel of the microscope was used for image acquisition.
The cationic fluorescent probe TMRM?accumulates in mitochondria
because of the high membrane potential across the IMM and, therefore,
low concentrations (20 nM) of TMRM?in experimental medium for 30
At equilibrium, the fluorescence produced by this low concentration of
TMRM?(excitation 543/emission red photomultiplier channel) is a direct
Dye intensities in images were quantified by measuring the raw pixel
intensities in neuronal somas with the region tool of MetaMorph soft-
measured in each neuron was normalized to the average dye intensity of
control nonTg neurons receiving the same concentration of dye for the
(untreated cultures of nonTg cortical neurons plated at the same time).
tal treatments, spatial learning and memory retention were determined
by the Morris water maze (MWM; Morris, 1984) behavioral test. The
??) produced by mitochondria or other sources. However, it is easily
15704 • J.Neurosci.,November2,2011 • 31(44):15703–15715McManusetal.•MitoQPreventsMemoryLossandNeuropathologyin3xTg-ADMice
(4 feet diameter) painted white and filled with water maintained at 24 ?
1°C. Nontoxic, white tempura paint was used to conceal a slightly sub-
merged, circular Plexiglas platform (14 cm diameter). MWM tests were
conducted according to Billings et al. (2005) with minor modifications.
During the acquisition trails, the mice were placed in the tank at one of
four designated start points and allowed 60 s to find and escape onto the
platform using at least three distinct, extra-maze cues on the curtains
surrounding the tank. If a mouse failed to find the platform within the
allotted time, it was manually guided there, and the escape latency was
recorded as 60 s. In either case, the mouse remained on the platform for
30 s to consolidate the spatial cues (Buccafusco, 2001; Arendash et al.,
2006). After this time, the mouse was allowed to rest in a holding cage
fitted with a warm towel for 30 s until the start of the next trial. Each
mouse was given four consecutive trials per day for 7 d (Vorhees and
Williams, 2006). Short- and long-term retention of the spatial training
was assessed in probe trials (platform removed) conducted 1.5 and 24 h,
respectively, after the last acquisition trial. Spatial bias for the platform
location was determined by the number of crosses over the previous
platform location during a 60 s free swim.
Cued acquisition trials were conducted after the last probe trial to
determine whether differences in learning and memory could be attrib-
uted to impaired eyesight, swimming ability, or motivation to escape
platform was made visible by a mounted flag. Multiple parameters in all
MWM tasks were calculated and analyzed by Ethovision XT tracking
software (Noldus Information Technology).
Tissue acquisition. At 7 months of age, mice were killed by cervical
dislocation and intracardially perfused with PBS, pH 7.4. Brains were
rapidly removed and sagittally split. One hemi-brain was fixed in 4%
paraformaldehyde for immunohistochemistry. After removal of the
brainstem and cerebellum, the other hemi-brain was snap frozen in liq-
uid nitrogen and stored at ?80°C for use in biochemical analysis.
Immunohistochemistry. Hemi-brains were fixed for 48 h in 4% para-
formaldehyde, embedded in paraffin, cut into 5 ?m sections, and
mounted on glass slides. Sections were deparaffinized, rehydrated, and
heated in 10 mM sodium citrate, pH 6.0, to ?95°C to assist in antigen
retrieval. The sections were incubated in blocking buffer (150 mM NaCl,
100 mM Tris pH 7.4, 0.1% Triton X-100, 2% BSA) for 1 h at room
temperature, followed by anti-A?(1–42)antibody (1:500; BioSource In-
ternational) overnight at 4°C (Oddo et al., 2003b). A?(1–42)was visual-
ized by using an ABC immunoperoxidase kit from Vector Laboratories
and diaminobenzidine substrate. Images were taken by a CCD camera
mounted on a Nikon TE-300 microscope. Image acquisition was con-
trolled by MetaMorph software.
Immunoblotting. Previously frozen hemi-brains were homogenized in
2% SDS lysis buffer (50 mM Tris, 2 mM EDTA, 150 mM NaCl) supple-
the supernatant was determined by the Bradford assay (Pierce). Equal
amounts of protein from each sample were separated by SDS-PAGE and
transferred to equilibrated PVDF membranes (Millipore). Membranes
were blocked for at least 1 h at room temperature with TBST (10 mM
anti-3-nitrotyrosine (3-NT) (1 ?g/ml; Cayman Chemical), anti-
synaptophysin (1:200 dilution; Sigma), or anti-glial fibrillary acidic pro-
tein (GFAP; 2.5 ?g/ml; Sigma) diluted in 1% nonfat dry milk overnight
at 4°C. The membranes were then washed for 20 min in TBST and then
incubated for 1 h at room temperature in anti-mouse HRP-linked sec-
ondary antibody (1:2000; Bethyl Laboratories), followed by another 20
min wash. The membranes were stripped with Restore Stripping Buffer
(Pierce), washed in TBST, and processed as described above for ?-actin
or ?-tubulin (Sigma) as a loading control. Proteins were detected using
the chemiluminescent SuperSignal substrate (Pierce).
Amyloid(1–42)ELISA. Soluble A?(1–42)was extracted as described by
appropriate amount (150 mg/ml wet weight) of ice-cold 0.6% SDS lysis
buffer (50 mM Tris, 2 mM EDTA, 150 mM NaCl), supplemented with
sonicated and centrifuged at 4°C for 1 h at 100,000 ? g in a Beckman
Coulter Optima TLX-120 ultracentrifuge. The supernatant containing
to the ELISA kit instructions. Briefly, 100 ?l of the samples or A?(1–42)
peptide standards were added to each well in duplicate or triplicate and
incubated overnight at 4°C. The following day, the 96-well plate was
thoroughly washed, incubated in tetramethylbenzidine substrate for 40
a SpectraMax M2 microplate reader. Quantification of soluble A?(1–42)
levels in the samples was achieved by normalizing the A?(1–42)concen-
Bradford protein assay (Pierce).
barbituric acid reactive substances (TBARS). Brain homogenates (0.02
g/ml 50 mM Tris-HCl) were diluted in 2 vol of 15% trichloroacetic acid
and centrifuged at 1000 ? g for 10 min at 4°C. The supernatant was
added to an equal volume of 0.375% TBA in 0.25 M HCl and heated at
assay, 0.015% 2, 6-di-tert-butyl-4-methylphenol was added to the mix-
ture before the acid-heating stage. After cooling and centrifugation at
100 ? g for 5 min, the formation of TBARS was determined by the
absorbance of the colorimetric product at 532 nm by a SpectraMax M2
microplate reader (Molecular Devices). The amount of TBARS in the
samples was calculated from a standard curve produced by hydrolysis of
tetraethoxypropane. Results were normalized to protein concentration
of each sample.
glutathione (GSSG) was determined by a luminescent assay (Promega).
EDTA and briefly centrifuged, and the supernatant was collected. The
ples were then incubated in buffer containing glutathione S-transferase
was quantified by a SpectraMax M2 microplate reader. The linear por-
ratio of GSH/GSSG in the samples.
ufacturer with modifications for tissue homogenates (Liu et al., 2004).
Briefly, brain extracts were prepared by Dounce homogenization in ice-
mM EGTA) and centrifuged for 15 min at 13,000 rpm. Protein concen-
trations of the supernatant were determined using the Bradford assay
(Pierce) to ensure equal loading. Samples were diluted accordingly in
PBS and loaded in duplicate or triplicate into a white-walled, 96-well
plate for 1 h incubation in an equal volume of Caspase-Glo reagent at
room temperature. The luminescence of each sample, a measure of
caspase 3/7 activity, was measured by a SpectraMax M2 microplate
reader and normalized to the respective nonTg control. This assay does
not distinguish between the activities of caspases 3 and 7.
Statistics. Statistical analysis and graph preparation were done with
SigmaPlot 11.1 (Systat Software). Statistical comparisons were made by
tiple comparisons post hoc tests. Error bars are ?SEM.
mitochondria from A?-induced toxicity in N2a cells. They also
demonstrated that MitoQ enhances neurite outgrowth in cul-
McManusetal.•MitoQPreventsMemoryLossandNeuropathologyin3xTg-ADMice J.Neurosci.,November2,2011 • 31(44):15703–15715 • 15705
not present direct evidence of a protective effect of MitoQ on the
mitochondria in these cells. To determine whether MitoQ can
prevent A? toxicity in primary neurons and to explore its effect
on mitochondria, we investigated its effects on mouse cortical
neurons treated with A?(22–35)in cell culture. This A? peptide
fragment contains the biologically active portion of A? (Yanker
tage for in vitro studies of rapidly aggregating in aqueous solu-
tions (Pike et al., 1993). A?(22–35)forms amyloid fibrils in vitro
resembling those of the ?-amyloid protein in senile plaques and
and A?(1–42)in hippocampal and cortical neurons in cell culture
(Pike et al., 1995; Casley et al., 2002b). An even shorter peptide
[A?(25–35)] causes memory impairment when injected into rat
brains (Díaz et al., 2010).
A?(22–35)treatment caused a progressive reduction in cortical
neuron survival that declined by 48 h to ?35% that of untreated
controls (Fig. 1A,B). Low concentrations of MitoQ (1–100 nM)
greatly inhibited this toxicity. dTPP, a compound that is struc-
turally identical to MitoQ but that lacks the ubiquinone moiety,
dTPP did not increase survival of A?(22–35)-treated neurons. Fig-
ure 1C shows that, similar to its effect on A?(22–35), 1 nM MitoQ
also prevented death of cortical neurons treated with full-length
A?(1–40). In other studies, the EC50for MitoQ in cultured cells
logs of concentration (Jauslin et al., 2003). Therefore, our find-
ings in neuronal cells are consistent with other studies.
A?, and its precursor APP, localize, in part, to mitochondria in
increased production of RS, enzyme inhibition, loss of cyto-
evaluate the effect of MitoQ on A?-induced RS and mitochon-
drial damage, cortical neurons in cell culture were treated with
A?(22–35)for 24 h in the presence or absence of MitoQ. A similar
amyloid fragment has been reported to impair mitochondrial
function in isolated brain mitochondria (Canevari et al., 1999).
Confocal microscopy allowed simultaneous assessment in single
cells of changes in RS by the redox-sensitive dye CM-H2DCFDA
and ??mby the potential-dependent fluorescent dye TMRM?
(Fig. 2A). A?(22–35)treatment for 24 h induced a 3.4 ? 0.2-fold
average increase in CM-H2DCFDA intensity, indicating elevated
neuronal RS production. This treatment also significantly depo-
larized ??m(p ? 0.01), suggesting damage to the IMM. MitoQ
(1 and 5 nM) prevented the increased RS caused by A?(22–35)
treatment and maintained ??mat levels indistinguishable from
controls (Fig. 2B,C). A?(22–35)? dTPP-treated neurons had RS
ety of MitoQ was responsible for its protective effects.
A? induces production of O2
tiple RS downstream of these radicals in many cells, including
cortical neurons (Keller et al., 1998; Longo et al., 2000; Keil et al.,
2004; Malinski, 2007; Stepanichev et al., 2008; Díaz et al., 2010).
The A?(22–35)-induced increases in O2
sensitivity of CM-H2DCFDA to oxidation by ONOO?, in com-
??, nitric oxide (NO?), and mul-
??and NO?leads to forma-
bination with the propensity of ONOO?formation attributable
to A?-induced stress, we reasoned that the primary RS detected
in our paradigm was ONOO?(Setsukinai et al., 2003; Malinski,
2007). Treatment of A?(22–35)-exposed cultures with the nitric
oxide synthase (NOS) inhibitor NG-nitro-L-arginine methyl ester
(L-NNA) decreased CM-H2DCFDA fluorescence intensity to that
found in control cultures, confirming that the RS responsible for
CM-H2DCFDA oxidation were primarily nitrogen-associated spe-
A? is reported to increase RS in neurons via multiple mecha-
nisms, including direct A?-induced RS production, elevated
spiratory complex IV), and by causing leakage of electrons from
culture. A, Photomicrographs of untreated cortical neurons and cortical neurons exposed to
C, MitoQ also prevented A?(1–40)-induced death. n ? 9 cultures per condition. After treat-
MitoQ inhibited A?-induced death of C57BL/6 mouse cortical neurons in cell
15706 • J.Neurosci.,November2,2011 • 31(44):15703–15715McManusetal.•MitoQPreventsMemoryLossandNeuropathologyin3xTg-ADMice
the mitochondrial electron transport chain (Hensley et al., 1994;
Canevari et al., 1999; Lustbader et al., 2004; Takuma et al., 2005;
Shelat et al., 2008; Rhein et al., 2009; Saraiva et al., 2010). The
chain is highly dependent on maintenance of the electrochemical
gradient across the IMM (Turrens, 1997). We used the uncou-
pling agent carbonyl cyanide p-trifluoromethoxyphenylhydrazone
(FCCP) to break down this gradient. By decreasing the proton gra-
dient across the IMM, FCCP increases rate of electron transport
through the respiratory chain, thereby inhibiting electron leakage
and consequently mitochondrial O2
the findings of others (Saraiva et al., 2010), brief incubation with
FCCP caused the loss of most CM-H2DCFDA fluorescence in
detected by this dye was primarily of mitochondrial origin
(Duchen, 1999). Together, these results suggest that MitoQ
decreased CM-H2DCFDA oxidation in A?-treated cortical neu-
rons primarily by suppressing RS produced by mitochondria.
??mnot only influences RS generation but other critical mito-
??production. Consistent with
chondrial functions, such as nicotinamide adenine dinucleotide re-
duction, ATP synthesis, and Ca?sequestration, and is therefore
considered the primary dictator of mitochondrial energetics as well
as an indicator of the overall health of mitochondria in neurons
??mat control levels suggests that MitoQ preserved the functional
Treating cultures of cortical neurons obtained from the
caused cell death similar to that shown in Figure 1 for wild-type
for 24 h increased CM-H2DCFDA intensity 3.4 ? 0.2-fold
(10 nM) in the media during A?(22–35)exposure prevented this
increase (0.3 ? 0.19-fold compared with control; p ? 0.1; n ?
49). Therefore, the response of the neurons to A?(22–35)and to
MitoQ was similar to that of wild-type cortical neurons in
There is extensive in vitro evidence that the concentrations of
MitoQ used here have no impact on mitochondrial functions,
such as ATP production, membrane potential, or ROS produc-
tion in cells or mitochondria, but do prevent mitochondrial ox-
idative damage under conditions that lead to oxidative stress
(Kelso et al., 2001; Jauslin et al., 2003; Saretzki et al., 2003; Asin-
Cayuelaa et al., 2004).
age and the pathologies (Adlam et al., 2005; Lowes et al., 2008;
Graham et al., 2009; Supinski et al., 2009; Chacko et al., 2010;
Smith and Murphy 2010). The positive results of the MitoQ
drug-screening experiments (Figs. 1, 2) suggested that MitoQ
tic potential of MitoQ for treating AD and to examine a possible
role for oxidative stress in AD progression, we investigated the
model of AD. This mouse strain develops cognitive dysfunction
as well as both plaque and neurofibrillary tangle pathology in
AD-relevant brain regions in an age-dependent manner, closely
tably, the earliest pathological changes in 3xTg-AD mice involve
mitochondrial impairment and increased oxidative stress (Re-
sende et al., 2008; Yao et al., 2009). Table 1 shows the ages at
which various cognitive pathologies and neuropathologies are
reported to first occur in these mice. Most deficits, other than
simultaneous assessment of RS by the redox-sensitive dye CM-H2DCFDA and ??mby the
CM-H2DCFDA intensity, were much higher in cells treated for 24 h with A?(22–35)(25 ?M)
MitoQ and the NOS inhibitor L-NNA suppressed increased RS caused by A?(22–35)exposure.
MitoQ prevented increased A?-induced RS production and A?-induced ??m
pathologies shown continue to progress as the animals age. A? deposition occurs in cortex, hippocampus, and
McManusetal.•MitoQPreventsMemoryLossandNeuropathologyin3xTg-ADMiceJ.Neurosci.,November2,2011 • 31(44):15703–15715 • 15707
of the lives of these animals. Because of the long period of treat-
ment necessary to test the ability of MitoQ to ameliorate NFT
formation (up to 18 months), we chose to determine its effects
only on the early pathologies.
Spatial retention deficits emerge in 3xTg-AD mice by 4–6
months of age and are reminiscent of spatial learning deficits in
posed to be prodromal for AD (Grundman et al., 2004; Mosconi
et al., 2008a). The mitochondrial dysfunction evident in these
mice also recapitulates multiple parameters found in MCI and
early AD patients, including decreased mitochondrial bioener-
getics, increased oxidative stress, and increased mitochondrial
A? load (Billings et al., 2005; Yao et al., 2009). Consequently,
of prophylactic therapeutics (Billings et al., 2005). Only female
exhibit an earlier and more pronounced AD phenotype than do
males (Clinton et al., 2007). To test the effect of MitoQ on cog-
nitive performance, female 3xTg-AD mice were either left un-
treated or received a continuous supply of 100 ?M MitoQ or 100
?M dTPP in their drinking water from 2–7 months of age. Age-
matched nonTg female mice either received no treatment or 100
4.5 months of treatment, the cognitive performance of these six
groups was determined by performance in the MWM (Morris,
1984). All mice were able to learn the MWM task, because the
average escape latency for each group gradually decreased to
reach a predetermined criterion (?25 s average latency) during
7 d, hidden-platform training trials. Untreated nonTg, nonTg
receiving MitoQ treatment, and 3xTg-AD mice receiving MitoQ
The untreated 3xTg-AD and dTPP-treated 3xTg-AD mice re-
quired additional training to learn the task (Fig. 3A). MitoQ
treatment significantly enhanced 3xTg-AD performance on ac-
quisition trials to a level indistinguishable from nonTg and
nonTg receiving MitoQ treatment (p ? 0.01). Conversely, treat-
ment with the negative control dTPP did not affect the perfor-
mance of 3xTg-AD mice (p ? 0.05 for 3xTg-AD ? dTPP vs
3xTg-AD on all training days). To determine the effect of MitoQ
platform was removed and spatial bias for the previous platform
location in the MWM was analyzed in probe trials conducted 1.5
and 24 h after the last training trial. MitoQ prevented the long-
term retention deficit in 3xTg-AD mice (Fig. 3B). These mice
performed at the same level as nonTg controls. An analogous
trend was evident in the short-term probe.
The effect of MitoQ on rate of learning in the 3xTg-AD mice
could not be explained by a stimulatory effect of MitoQ or by an
effect of MitoQ on vision. Cued acquisition trials in which the
escape platform was made visible to the mice were conducted
average times in which mice in each group reached the platform,
cm/s for each group (p ? 0.84), indicating that motor perfor-
mance was the same for all and that the MitoQ did not have a
stimulatory effect. Rather, MitoQ enhanced learning and spatial
memory retention. Therefore, the 3xTg mice receiving MitoQ
treatment performed as well as nonTg mice on all tasks, indicat-
ing that MitoQ therapy effectively prevented the onset of AD-
associated cognitive decline in them. The negative control dTPP
produced no effect on acquisition or probe trials, indicating that
the cognitive enhancement in MitoQ-treated mice was attribut-
able to the antioxidant moiety.
Resende et al. (2008) reported that several markers for oxidative
stress were higher in the brains of 3- to 5-month-old 3xTg-AD
mice than in nonTg mice of the same age. Among these markers
Methods). Each mouse performed four training trials per day for 7 consecutive days, and the
However, the nonTg, nonTg ? MitoQ, and 3xTg-AD ? MitoQ mice learned the task more
in their drinking water from 2 to 7 months after birth. B, MitoQ treatment prevented loss of
short- and long-term spatial memory retention in the 3xTg-AD mice. Memory retention was
Spatial bias is shown as the number of previous platform location crosses as determined by
video analysis using Ethovision tracking software. MitoQ significantly prevented both short-
same amount of time (p ? 0.7 by ANOVA). n ? 21–38 mice except for the MitoQ-exposed
MitoQ treatment prevented the onset of cognitive deficits in young female
15708 • J.Neurosci.,November2,2011 • 31(44):15703–15715McManusetal.•MitoQPreventsMemoryLossandNeuropathologyin3xTg-ADMice
were decreased levels of GSH, increased levels of GSSG, and in-
creased levels of malondialdehyde (MDA), a marker for lipid
peroxidation. Figure 4A shows that 5 months of MitoQ treat-
ment prevented the decrease in the GSH/GSSG ratios found in
the brains of the 3xTg-AD mice. Figure 4B shows that it had a
similar effect on the increased MDA levels in the brains of these
associated with biological membranes (Bartesaghi et al., 2010).
Nitro-oxidative modification to proteins may dramatically alter
role in AD development. Because of the apparent decrease in
ONOO?-associated RS levels in our in vitro analysis of MitoQ-
treated neurons, we investigated a related effect of MitoQ in vivo
by measuring nitrotyrosine levels in 3xTg-AD mouse brains. Ty-
rosine nitration provides a footprint for free radical-mediated
intermediate (Halliwell and Gutteridge, 2007; Malinski, 2007).
MitoQ prevented an increase in 3-NT in young 3xTg-AD brains
(Fig. 4C). Together, the data indicate that the brains of young
3xTg-AD mice are under oxidative stress and that MitoQ treat-
ment prevented this stress from occurring.
The likely cause of the decline in cognitive function of
3xTg-AD mice with age is an increase in age-related synaptic
dysfunction in their brains (Oddo et al., 2003b). Associated with
this decreased synaptic efficacy is a decline in brain levels of the
ubiquitous presynaptic marker synaptophysin, suggesting an ac-
tual loss of synapses (Blanchard et al., 2010). Figure 5, A and B,
shows that 5 months of MitoQ treatment prevented loss of syn-
aptophysin in young 3xTg-AD brains. This finding suggests that
secondary to preservation of functional synapses.
(MDA levels) in the brains of 3xTg-AD mice (p ? 0.01). MDA levels were determined by the
TBARS assay. n ? 6 brains for each except dTPP in which n ? 3 brains. C, MitoQ treatment
suppressed the development of elevated levels of 3-NT in the brains of 3xTg-AD mice. Top,
of 3xTg-AD mice than they were in the same proteins in nonTg mice. 3-NT levels in these
increase was prevented by MitoQ treatment. Density was determined for entire lanes and is
the synaptic protein synaptophysin (Syn) and increase of the astrocyte marker GFAP in the
brains of 3xTg-AD mice. Blots between lines are all from the same gel. Similar amounts of
McManusetal.•MitoQPreventsMemoryLossandNeuropathologyin3xTg-ADMice J.Neurosci.,November2,2011 • 31(44):15703–15715 • 15709
The brains of human AD patients exhibit extensive astroglio-
sis, a process indicative of neuronal damage. Astrogliosis also
occurs with increasing age in the brains of 3xTg-AD mice (Oddo
et al., 2003a). We used the astrocyte marker GFAP to compare
astrogliosis in the brains of nonTg and 3xTg-AD mice. As re-
ported previously (Oddo et al., 2003a), we found that the brains
nonTg mice (Fig. 5C), indicating that they had more reactive
astrocytes than did the wild-type brains. The brains of 3xTg-AD
animals that had received 5 months of MitoQ treatment had
GFAP levels identical to those found in the wild-type brains.
the pathological events occurring in AD, including NFT forma-
tion, synaptic dysfunction, and neuronal death (Oddo et al.,
2003b; Hardy, 2006). This hypothesis is supported by the early
A? accumulation in the rare, autosomal dominant forms of AD,
by transgenic mice harboring these human mutations (Hardy
and Selkoe, 2002), and by recent studies suggesting that mito-
chondrial A? may be responsible for synaptic degeneration and
the onset of cognitive impairment in AD models of A? enrich-
ment (Zhao et al., 2010). A? appears to enhance production of
Evidence also suggests that not only does A? increase cellular RS
but that these RS may in turn increase production of A? in a
al., 2002, 2005; Lin and Beal, 2006; Quiroz-Baez et al., 2009).
Given the possible role of the mitochondria-generated RS in A?
production, we sought to determine the effect of MitoQ treat-
ment on A? burden (Cardoso et al., 2001; Hansson et al., 2004;
Caspersen et al., 2005; Manczak et al., 2006; Anandatheertha-
varada and Devi, 2007). Intraneuronal A? immunoreactivity is
one of the earliest histopathological events reported in the
3xTg-AD brain (Billings et al., 2005). Figure 6A shows that
7-month-old 3xTg-AD mice that had received MitoQ treatment
for 5 months exhibited reduced intraneuronal staining by an an-
ti-A?(1–42)antibody in their hippocampus and neocortex com-
pared with untreated 3xTg-AD mice. The ability of MitoQ to
decrease soluble A?(1–42)burden in the brains of 3xTg-AD mice
was investigated by ELISA. MitoQ decreased soluble A?(1–42)in
the brains of these animals (Fig. 6B). These findings are consis-
between A? deposition and oxidative stress.
dogenous A? and extracellularly applied A? is mediated, at least
in part, via the intrinsic, mitochondria-dependent apoptotic
pathway (Harada and Sugimoto, 1999; Cardoso et al., 2001; Cas-
ley et al., 2002b; Caspersen et al., 2005; Hansson Petersen et al.,
2008; Cho et al., 2009; Takuma et al., 2009). Recent evidence
suggests that executioner caspases 3 and 7 may contribute to tau,
the protein responsible for forming NFTs, pathology by cleaving
tau after Asp421 to yield a cytotoxic ?tau fragment (Rissman et
al., 2004; Rohn et al., 2008; de Calignon et al., 2010). Because the
intrinsic apoptotic pathway may involve mitochondrial RS gen-
eration (Kirkland et al., 2001), we hypothesized that the decrease
rant caspase activity in the 3xTg-AD model (Rohn et al., 2008).
Using a proluminescent caspase 3/7 substrate, we measured the
effect of MitoQ treatment on caspase 3/7 activity in the brains of
in cerebral caspase 3/7 activity in 7-month-old untreated
completely blocked by treatment with MitoQ during the preced-
ing 5 months (Fig. 7).
The recent development of antioxidants that selectively concen-
trate in mitochondria provides an opportunity to decipher the
impact of mitochondria-generated RS on the pathogenesis and
tive photomicrographs showing immunostaining with an anti-A?(1–42)antibody within the
hippocampus (top) and neocortex (bottom) of nonTg, untreated 3xTg-AD mice and 3xTg-AD
15710 • J.Neurosci.,November2,2011 • 31(44):15703–15715 McManusetal.•MitoQPreventsMemoryLossandNeuropathologyin3xTg-ADMice
al., 2003; Lin and Beal, 2006; Reddy, 2008; Swerdlow and Khan,
2009; Moreira et al., 2010). The most extensively studied com-
pound of this class is the mitochondria-targeted ubiquinone
derivative MitoQ (Smith et al., 2003; James et al., 2007; Rodriguez-
Cuenca et al., 2010). We first investigated the ability of MitoQ to
prevent the A?-induced death of cortical neurons in cell culture.
Low nanomolar concentrations of MitoQ prevented this death.
These concentrations also blocked an A?-induced increase in
NO?-associated RS and A?-induced depolarization of ??m. The
data suggest that the death was associated with mitochondrial
We next tested the ability of MitoQ to inhibit the cognitive defi-
cits that occur in the 3xTg-AD transgenic mouse model of AD
from 2–7 months after birth (Oddo et al., 2003b). MitoQ pre-
vented the decline in spatial memory retention in these animals
during this period. A? deposition and early cognitive impair-
ment are preceded by mitochondrial dysfunction and increased
oxidative stress in 3xTg-AD mouse brains (Resende et al., 2008;
also prevented synaptic loss, astrogliosis, increased A? burden,
and elevated caspase 3/7 activity. These findings are consistent
with a role for A?-induced oxidative stress in mediating the cog-
elevated levels of A?.
reduction of mitochondrial superoxide dismutase (MnSOD)
levels accelerates AD-like pathology in a transgenic mouse
model of AD, whereas overexpression of MnSOD decreases
A?-processing, preserves synaptic integrity, and eliminates
the earliest cognitive deficits in a transgenic mouse AD model
(Li et al., 2004; Esposito et al., 2006; Massaad et al., 2009). The
ubiquinone moiety of MitoQ reacts rapidly with O2
thus, may enhance mitochondrial antioxidant defense by
2009). MitoQ may also prevent damage downstream of O2
Nitro-oxidative modifications to mitochondrial proteins are
associated with pathological changes in mitochondrial redox
state, morphology, and bioenergetics in preclinical and early AD
terfield et al., 2006, 2007; Yao et al., 2009). Evidence supports a
causative role for these modifications in mediating the early syn-
aptic degeneration and cognitive decline in AD (Wang et al.,
Modification of mitochondrial enzymes and components of the
respiratory chain by RS may contribute to impaired glucose me-
tabolism found in preclinical AD subjects (Sultana and Butter-
field, 2009; Yao et al., 2009). Longitudinal neuroimaging studies
reveal the rate of metabolic decline within the medial temporal
lobe predicts the progression of cognitive deterioration from
normal aging to AD and more closely correlates with the level of
oxidative damage than other biomarkers tested in preclinical pa-
tients (Mosconi et al., 2008a,b). This correlation may prove fun-
damental because mitochondrial dysfunction and oxidative
stress may encourage the development of amyloid plaques and
NFTs (Hardy, 2006; Gibson and Shi, 2010).
The predominant pathway of A? production occurs by se-
quential cleavage of APP by ?- and ?-secretases. Metabolic im-
pairment induced by thiamine deficiency or respiratory chain
??degradation by MnSOD (Maroz et al.,
inhibition augments ?-secretase expression and activity (Karup-
pagounder et al., 2009; Zhang et al., 2011). Oxidative stress also
induces ?-secretase upregulation and, consequently, increased
A? generation, both of which are preventable by antioxidant
is present in mitochondria and may also be upregulated by RS
by directly cleaving APP (caspase 3) or via a process involving
?-secretase (caspases 2 and 8; Newcombe et al., 2004; Chae et al.,
2010). Oxidative stress may also contribute to A? accumulation
by inactivating A?-degrading enzymes (Shinall et al., 2005). In-
tracellular A? may then enter the mitochondria in which it in-
duces additional RS and magnifies the metabolic deficit
(Lustbader et al., 2004; Hansson Petersen et al., 2008). MitoQ
treatment may break this putative autocatalytic cycle.
The attenuation of increased caspase 3/7 activity by MitoQ
treatment may modulate tau pathology, because the early cleav-
age of tau by these caspases initiates tau aggregation and phos-
phorylation and may contribute to cognitive decline (Cotman et
al., 2005; de Calignon et al., 2010). As reported by others, ?tau
levels were below the detectable level in our 7-month-old, un-
treated 3xTg-AD mice (data not shown; Rissman et al., 2004). It
will be of interest to treat these mice for longer periods (e.g., 18
months) with MitoQ to determine whether it can ameliorate
development of tau pathology.
The results reported here highlight a possible influence of
mitochondria-induced oxidative stress on cognitive decline and
neuropathology in AD. Failure to address underlying mitochon-
drial deficits may explain the disappointing results of A?-
centered therapy in humans. Recent clinical trials attempting to
decline in AD patients despite significantly decreasing the amy-
a lack of clinical correlation between A? levels and AD progres-
sion and propose that brain atrophy and cerebral metabolic rate
provide the best predictive parameters for clinical changes in AD
(Giannakopoulos et al., 2003; Li et al., 2008; Walhovd et al.,
2010). Thus, A? may not exclusively dictate AD pathogenesis,
and auxiliary approaches may be required for optimal therapeu-
tic benefit and the effective prevention of AD.
Several antioxidants are reported to lessen pathologies found
in transgenic mouse models of AD (Lim et al., 2001; Stackman et
al., 2003; Nicolakakis et al., 2008; Wang et al., 2008a). However,
these antioxidants are nonspecific and should interact with ROS
in all areas of the brain. Our study is a significant step forward
from these in that it shows that an antioxidant that only operates
within mitochondria is protective. In addition, this is the first
demonstration that a mitochondria-targeted therapy is effective
in vivo and opens the way for translation to patients (MitoQ has
been though two phase II trials for other conditions; Snow et al.,
2010). The ability of MitoQ to prevent neuropathologies in
3xTg-AD mice also suggest that in vivo assessments of other
classes of mitochondria-targeted antioxidants for treating AD,
such as SS31 peptide, are warranted (Manczak et al., 2010).
Adlam VJ, Harrison JC, Porteous CM, James AM, Smith RA, Murphy MP,
Sammut IA (2005) Targeting an antioxidant to mitochondria decreases
cardiac ischemia-reperfusion injury. FASEB J 19:1088–1095.
Anandatheerthavarada HK, Devi L (2007) Amyloid precursor protein
McManusetal.•MitoQPreventsMemoryLossandNeuropathologyin3xTg-ADMiceJ.Neurosci.,November2,2011 • 31(44):15703–15715 • 15711
and mitochondrial dysfunction in Alzheimer’s disease. Neuroscientist
AnandatheerthavaradaHK,BiswasG,RobinMA,AvadhaniNG (2003) Mi-
tochondrial targeting and a novel transmembrane arrest of Alzheimer’s
amyloid precursor protein impairs mitochondrial function in neuronal
cells. J Cell Biol 161:41–54.
Anantharaman M, Tangpong J, Keller JN, Murphy MP, Markesbery WR,
Kiningham KK, St Clair DK (2006) ?-Amyloid mediated nitration of
manganese superoxide dismutase: implication for oxidative stress in a
APPNLh/NLhX PS-1P264L/P264Ldouble knock-in mouse model of Alzhei-
mer’s disease. Am J Pathol 168:1608–1618.
Arendash GW, Schleif W, Rezai-Zadeh K, Jackson EK, Zacharia LC, Crac-
chiolo JR, Shippy D, Tan J (2006) Caffeine protects Alzheimer’s mice
against cognitive impairment and reduces brain beta-amyloid produc-
tion. Neuroscience 142:941–952.
Asin-CayuelaJ,ManasAR,JamesAM,SmithRA,MurphyMP (2004) Fine-
Bartesaghi S, Wenzel J, Trujillo M, Lo ´pez M, Joseph J, Kalyanaraman B, Radi
R (2010) Lipid peroxyl radicals mediate tyrosine dimerization and ni-
tration in membranes. Chem Res Toxicol 23:821–835.
BeckmanJS,KoppenolWH (1996) Nitricoxide,superoxide,andperoxyni-
trite: the good, the bad, and the ugly. Am J Physiol 271:C1424–C1437.
BillingsLM,OddoS,GreenKN,McGaughJL,LaFerlaFM (2005) Intraneu-
ronal A? causes the onset of early Alzheimer’s disease-related cognitive
deficits in transgenic mice. Neuron 45:675–688.
Blanchard J, Wanka L, Tung YC, Ca ´rdenas-Aguayo Mdel C, LaFerla FM,
Iqbal K, Grundke-Iqbal I (2010) Pharmacologic reversal of neurogenic
and neuroplastic abnormalities and cognitive impairments without af-
fecting A? and tau pathologies in 3xTg-AD mice. Acta Neuropathol
Buccafusco J (2001) Spatial navigation (water maze) tasks. In: Methods of
ButterfieldDA,PerluigiM,SultanaR (2006) OxidativestressinAlzheimer’s
disease brain: new insights from redox proteomics. Eur J Pharmacol
Butterfield DA, Reed T, Newman SF, Sultana R (2007) Roles of amyloid
?-peptide-associated oxidative stress and brain protein modifications in
the pathogenesis of Alzheimer’s disease and mild cognitive impairment.
Free Radic Biol Med 43:658–677.
Canevari L, Clark JB, Bates TE (1999) ?-Amyloid fragment 25–35 selec-
tively decreases complex IV activity in isolated mitochondria. FEBS Lett
Cardoso SM, Santos S, Swerdlow RH, Oliveira CR (2001) Functional mito-
chondria are required for amyloid ?-mediated neurotoxicity. FASEB J
Casley CS, Canevari L, Land JM, Clark JB, Sharpe MA (2002a) ?-Amyloid
inhibits integrated mitochondrial respiration and key enzyme activities.
J Neurochem 80:91–100.
Casley CS, Land JM, Sharpe MA, Clark JB, Duchen MR, Canevari L (2002b)
?-amyloid fragment 25–35 causes mitochondrial dysfunction in primary
cortical neurons. Neurobiol Dis 10:258–267.
Caspersen C, Wang N, Yao J, Sosunov A, Chen X, Lustbader JW, Xu HW,
Stern D, McKhann G, Yan SD (2005) Mitochondrial A?: a potential
focal point for neuronal metabolic dysfunction in Alzheimer’s disease.
FASEB J 19:2040–2041.
Chacko BK, Reily C, Srivastava A, Johnson MS, Ye Y, Ulasova E, Agarwal A,
Zinn KR, Murphy MP, Kalyanaraman B, Darley-Usmar V (2010) Pre-
vention of diabetic nephropathy in Ins2?/? AkitaJ mice by the
mitochondria-targeted therapy MitoQ. Biochem J 432:9–19.
Chae SS, Yoo CB, Jo C, Yun SM, Jo SA, Koh YH (2010) Caspases-2 and -8
are involved in the presenilin1/gamma-secretase-dependent cleavage of
S-nitrosylation of Drp1 mediates ?-amyloid-related mitochondrial fis-
sion and neuronal injury. Science 324:102–105.
LaFerlaFM (2007) Age-dependentsexualdimorphismincognitionand
stress response in the 3xTg-AD mice. Neurobiol Dis 28:76–82.
Cotman CW, Poon WW, Rissman RA, Blurton-Jones M (2005) The role of
Exp Neurol 64:104–112.
de Calignon A, Fox LM, Pitstick R, Carlson GA, Bacskai BJ, Spires-Jones TL,
HymanBT (2010) Caspaseactivationprecedesandleadstotangles.Na-
DeKosky ST, Williamson JD, Fitzpatrick AL, Kronmal RA, Ives DG, Saxton JA,
ofMemoryStudyInvestigators (2008) Ginkgobilobaforpreventionofde-
Limo ´n ID (2010) The amyloid-?25–35 injection into the CA1 region of
the neonatal rat hippocampus impairs the long-term memory because of
an increase of nitric oxide. Neurosci Lett 468:151–155.
Di Monte D, Ross D, Bellomo G, Eklo ¨w L, Orrenius S (1984) Alterations in
intracellular thiol homeostasis during the metabolism of menadione by
isolated rat hepatocytes. Arch Biochem Biophys 235:334–342.
Duchen MR (1999) Contributions of mitochondria to animal physiology:
from homeostatic sensor to calcium signalling and cell death. J Physiol
Dumont M, Wille E, Stack C, Calingasan NY, Beal MF, Lin MT (2009) Re-
duction of oxidative stress, amyloid deposition, and memory deficit by
manganese superoxide dismutase overexpression in a transgenic mouse
model of Alzheimer’s disease. FASEB J 23:2459–2466.
Esposito L, Raber J, Kekonius L, Yan F, Yu GQ, Bien-Ly N, Puoliva ¨li J,
Scearce-Levie K, Masliah E, Mucke L (2006) Reduction in mitochon-
and accelerates the onset of behavioral changes in human amyloid pre-
cursor protein transgenic mice. J Neurosci 26:5167–5179.
Giannakopoulos P, Herrmann FR, Bussie `re T, Bouras C, Ko ¨vari E, Perl DP,
Morrison JH, Gold G, Hof PR (2003) Tangle and neuron numbers, but
GibsonGE,ShiQ (2010) AmitocentricviewofAlzheimer’sdiseasesuggests
multi-faceted treatments. J Alzheimers Dis 20:S591–S607.
Graham D, Huynh NN, Hamilton CA, Beattie E, Smith RA, Cocheme ´ HM,
Murphy MP, Dominiczak AF (2009) Mitochondria-targeted antioxi-
dant MitoQ10improves endothelial function and attenuates cardiac hy-
pertrophy. Hypertension 54:322–328.
Grundman M, Petersen RC, Ferris SH, Thomas RG, Aisen PS, Bennett DA,
Foster NL, Jack CR Jr, Galasko DR, Doody R, Kaye J, Sano M, Mohs R,
Gauthier S, Kim HT, Jin S, Schultz AN, Schafer K, Mulnard R, van Dyck
Cooperative Study (2004) Mild cognitive impairment can be distin-
guished from Alzheimer disease and normal aging for clinical trials. Arch
Guglielmotto M, Giliberto L, Tamagno E, Tabaton M (2010) Oxidative
stress mediates the pathogenic effect of different Alzheimer’s disease risk
factors. Front Aging Neurosci 2:3.
Halliwell B, Gutteridge JMC (2007) Free radicals in biology and medicine,
Ed 4. Oxford, UK: Oxford UP.
SE, Ito A, Winblad B, Cowburn RF, Thyberg J, Ankarcrona M (2004)
Nicastrin, presenilin, APH-1, and PEN-2 form active ?-secretase com-
plexes in mitochondria. J Biol Chem 279:51654–51660.
Hansson Petersen CA, Alikhani N, Behbahani H, Wiehager B, Pavlov PF,
Alafuzoff I, Leinonen V, Ito A, Winblad B, Glaser E, Ankarcrona M
(2008) The amyloid ?-peptide is imported into mitochondria via the
Acad Sci USA 105:13145–13150.
Harada J, Sugimoto M (1999) Activation of caspase-3 in ?-amyloid-
induced apoptosis of cultured rat cortical neurons. Brain Res
Hardy J (2006) A hundred years of Alzheimer’s disease research. Neuron
Hardy J, Selkoe DJ (2002) The amyloid hypothesis of Alzheimer’s disease:
progress and problems on the road to therapeutics. Science 297:353–356.
Hensley K, Carney JM, Mattson MP, Aksenova M, Harris M, Wu JF, Floyd
RA,ButterfieldDA (1994) Amodelfor?-amyloidaggregationandneu-
rotoxicity based on free radical generation by the peptide: relevance to
Alzheimer’s disease. Proc Natl Acad Sci USA 91:3270–3274.
Hirai K, Aliev G, Nunomura A, Fujioka H, Russell RL, Atwood CS, Johnson
15712 • J.Neurosci.,November2,2011 • 31(44):15703–15715McManusetal.•MitoQPreventsMemoryLossandNeuropathologyin3xTg-ADMice
HarrisPL,JonesPK,PetersenRB,PerryG,SmithMA (2001) Mitochon-
drial abnormalities in Alzheimer’s disease. J Neurosci 21:3017–3023.
Holmes C, Boche D, Wilkinson D, Yadegarfar G, Hopkins V, Bayer A, Jones
RW, Bullock R, Love S, Neal JW, Zotova E, Nicoll JA (2008) Long-term
effects of A? 42 immunisation in Alzheimer’s disease: follow-up of a
randomised, placebo-controlled phase I trial. Lancet 372:216–223.
HoyerS (1996) Oxidativemetabolismdeficienciesinbrainsofpatientswith
Alzheimer’s disease. Acta Neurol Scand Suppl 165:18–24.
James AM, Cocheme ´ HM, Smith RA, Murphy MP (2005) Interactions of
mitochondria-targeted and untargeted ubiquinones with the mito-
chondrial respiratory chain and reactive oxygen species. J Biol Chem
(2007) Interaction of the mitochondria-targeted antioxidant MitoQ with
use of exogenous ubiquinones as therapies and experimental tools. J Biol
Jang JH, Surh YJ (2003) Protective effect of resveratrol on ?-amyloid-
induced oxidative PC12 cell death. Free Radic Biol Med 34:1100–1110.
JauslinML,MeierT,SmithRA,MurphyMP (2003) Mitochondria-targeted
antioxidants protect Friedreich Ataxia fibroblasts from endogenous oxi-
dative stress more effectively than untargeted antioxidants. FASEB J
Beal MF, Gandy SE, Gibson GE (2009) Thiamine deficiency induces
oxidative stress and exacerbates the plaque pathology in Alzheimer’s
mouse model. Neurobiol Aging 30:1587–1600.
Keil U, Bonert A, Marques CA, Scherping I, Weyermann J, Strosznajder JB,
Mu ¨ller-Spahn F, Haass C, Czech C, Pradier L, Mu ¨ller WE, Eckert A
(2004) Amyloid ?-induced changes in nitric oxide production and mi-
tochondrial activity lead to apoptosis. J Biol Chem 279:50310–50320.
Keller JN, Kindy MS, Holtsberg FW, St Clair DK, Yen HC, Germeyer A,
Steiner SM, Bruce-Keller AJ, Hutchins JB, Mattson MP (1998) Mito-
chondrial manganese superoxide dismutase prevents neural apoptosis
and reduces ischemic brain injury: suppression of peroxynitrite produc-
tion, lipid peroxidation, and mitochondrial dysfunction. J Neurosci
Kelso GF, Porteous CM, Coulter CV, Hughes G, Porteous WK, Ledgerwood
EC, Smith RA, Murphy MP (2001) Selective targeting of a redox-active
ubiquinone to mitochondria within cells. Antioxidant and antiapoptotic
properties. J Biol Chem 276:4588–4596.
MC, Seo JH, Choi SH, Suh YH (2002) Amyloid ? peptide induces cyto-
Kirkland RA, Franklin JL (2001) Evidence for redox regulation of cyto-
of protein synthesis and caspase inhibition. J Neurosci 21:1949–1963.
Kirkland RA, Windelborn JA, Kasprzak JM, Franklin JL (2002) A Bax-
induced pro-oxidant state is critical for cytochrome c release during pro-
grammed neuronal death. J Neurosci 22:6480–6490.
Kirkland RA, Saavedra GM, Cummings BS, Franklin JL (2010) Bax regu-
lates production of superoxide in both apoptotic and nonapoptotic neu-
rons: role of caspases. J Neurosci 30:16114–16127.
Li F, Calingasan NY, Yu F, Mauck WM, Toidze M, Almeida CG, Takahashi
RH, Carlson GA, Flint Beal M, Lin MT, Gouras GK (2004) Increased
plaque burden in brains of APP mutant MnSOD heterozygous knockout
mice. J Neurochem 89:1308–1312.
LiangJH,DuJ,XuLD,JiangT,HaoS,BiJ,JiangB (2009) Catalpolprotects
primary cultured cortical neurons induced by A?1–42through a
mitochondrial-dependent caspase pathway. Neurochem Int 55:741–746.
Någren K, Kim BC, Tsui W, de Leon MJ (2008) Regional analysis of
FDG and PIB-PET images in normal aging, mild cognitive impairment,
and Alzheimer’s disease. Eur J Nucl Med Mol Imaging 35:2169–2181.
LimGP,ChuT,YangF,BeechW,FrautschySA,ColeGM (2001) Thecurry
spice curcumin reduces oxidative damage and amyloid pathology in an
Alzheimer transgenic mouse. J Neurosci 21:8370–8377.
Lin MT, Beal MF (2006) Mitochondrial dysfunction and oxidative stress in
neurodegenerative diseases. Nature 443:787–795.
Liu D, Li C, Chen Y, Burnett C, Liu XY, Downs S, Collins RD, Hawiger J
(2004) Nuclear import of proinflammatory transcription factors is re-
quired for massive liver apoptosis induced by bacterial lipopolysaccha-
ride. J Biol Chem 279:48434–48442.
Longo VD, Viola KL, Klein WL, Finch CE (2000) Reversible inactivation of
superoxide-sensitive aconitase in A?1–42-treated neuronal cell lines.
J Neurochem 75:1977–1985.
Lowes DA, Thottakam BM, Webster NR, Murphy MP, Galley HF (2008)
The mitochondria-targeted antioxidant MitoQ protects against organ
damage in a lipopolysaccharide–peptidoglycan model of sepsis. Free
Radic Biol Med 45:1559–1565.
Lustbader JW, Cirilli M, Lin C, Xu HW, Takuma K, Wang N, Caspersen C,
Chen X, Pollak S, Chaney M, Trinchese F, Liu S, Gunn-Moore F, Lue LF,
(2004) ABAD directly links A? to mitochondrial toxicity in Alzheimer’s
disease. Science 304:448–452.
Malinski T (2007) Nitric oxide and nitroxidative stress in Alzheimer’s dis-
ease. J Alzheimers Dis 11:207–218.
Manczak M, Anekonda TS, Henson E, Park BS, Quinn J, Reddy PH (2006)
Mitochondria are a direct site of A? accumulation in Alzheimer’s disease
disease progression. Hum Mol Genet 15:1437–1449.
Manczak M, Mao P, Calkins MJ, Cornea A, Reddy AP, Murphy MP, Szeto
HH,ParkB,ReddyPH (2010) Mitochondria-targetedantioxidantspro-
tect against amyloid-? toxicity in Alzheimer’s disease neurons. J Alzhei-
mers Dis 20:S609–S631.
MarozA,AndersonRF,SmithRA,MurphyMP (2009) Reactivityofubiqui-
cations for in vivo antioxidant activity. Free Radic Biol Med 46:105–109.
Massaad CA, Washington TM, Pautler RG, Klann E (2009) Overexpression
in a mouse model of Alzheimer’s disease. Proc Natl Acad Sci USA
Mazziotti M, Perlmutter DH (1998) Resistance to the apoptotic effect of
aggregated amyloid-? peptide in several different cell types including
neuronal- and hepatoma-derived cell lines. Biochem J 332:517–524.
R,PacelliR,BrunettiA,ZambranoN,RussoT (2007) Essentialrolesfor
Fe65, Alzheimer amyloid precursor-binding protein, in the cellular re-
sponse to DNA damage. J Biol Chem 282:831–835.
Misonou H, Morishima-Kawashima M, Ihara Y (2000) Oxidative stress in-
duces intracellular accumulation of amyloid ?-protein (A?) in human
neuroblastoma cells. Biochemistry 39:6951–6959.
Moreira PI, Zhu X, Wang X, Lee HG, Nunomura A, Petersen RB, Perry G,
SmithMA (2010) Mitochondria:atherapeutictargetinneurodegenera-
tion. Biochim Biophys Acta 1802:212–220.
MorrisR (1984) Developmentsofawater-mazeprocedureforstudyingspa-
tial learning in the rat. J Neurosci Methods 11:47–60.
MosconiL,PupiA,DeLeonMJ (2008a) Brainglucosehypometabolismand
oxidative stress in preclinical Alzheimer’s disease. Ann NY Acad Sci
Mosconi L, De Santi S, Li J, Tsui WH, Li Y, Boppana M, Laska E, Rusinek H,
de Leon MJ (2008b) Hippocampal hypometabolism predicts cognitive
decline from normal aging. Neurobiol Aging 29:676–692.
Murphy MP, Smith RA (2007) Targeting antioxidants to mitochondria by
conjugation to lipophilic cations. Annu Rev Pharmacol Toxicol
Newcombe RE, Blumbergs PC, Sarvestani G, Manavis J, Jones NR (2004)
duction of amyloid A-beta in human acute and chronic compressive my-
leopathy. J Bone Joint Surg Br 86B [Suppl IV]:462.
Nicholls DG, Budd SL (2000) Mitochondria and neuronal survival. Physiol
Nicholls DG, Ferguson SJ (2002) Bioenergetics 3. London, UK: Academic.
Nicholls DG, Ward MW (2000) Mitochondrial membrane potential and
cell death: mortality and millivolts. Trends Neurosci 23:166–174.
Nicolakakis N, Aboulkassim T, Ongali B, Lecrux C, Fernandes P, Rosa-Neto
P, Tong XK, Hamel E (2008) Complete rescue of cerebrovascular func-
tion in aged Alzheimer’s disease transgenic mice by antioxidants and
pioglitazone, a peroxisome proliferator-activated receptor ? agonist.
J Neurosci 28:9287–9296.
OdaA,TamaokaA,ArakiW (2010) Oxidativestressup-regulatespresenilin
1 in lipid rafts in neuronal cells. J Neurosci Res 88:1137–1145.
McManusetal.•MitoQPreventsMemoryLossandNeuropathologyin3xTg-ADMiceJ.Neurosci.,November2,2011 • 31(44):15703–15715 • 15713
Oddo S, Caccamo A, Kitazawa M, Tseng BP, LaFerla FM (2003a) Amyloid
deposition precedes tangle formation in a triple transgenic model of Alz-
heimer’s disease. Neurobiol Aging 24:1063–1070.
Oddo S, Caccamo A, Shepherd JD, Murphy MP, Golde TE, Kayed R,
Metherate R, Mattson MP, Akbari Y, LaFerla FM (2003b) Triple-
transgenic model of Alzheimer’s disease with plaques and tangles.
Petersen RC, Thomas RG, Grundman M, Bennett D, Doody R, Ferris S,
Galasko D, Jin S, Kaye J, Levey A, Pfeiffer E, Sano M, van Dyck CH, Thal
LJ;Alzheimer’sDiseaseCooperativeStudyGroup (2005) VitaminEand
donepezil for the treatment of mild cognitive impairment. N Engl J Med
Pham DQ, Plakogiannis R (2005) Vitamin E supplementation in Alzhei-
Ann Pharmacother 39:2065–2072.
Pike CJ, Burdick D, Walencewicz AJ, Glabe CG, Cotman CW (1993) Neu-
rodegeneration induced by ?-amyloid peptides in vitro: the role of pep-
tide assembly state. J Neurosci 13:1676–1687.
Pike CJ, Walencewicz-Wasserman AJ, Kosmoski J, Cribbs DH, Glabe CG,
Cotman CW (1995) Structure-activity analyses of ?-amyloid peptides:
contributions of the ?25–35 region to aggregation and neurotoxicity.
J Neurochem 64:253–265.
Quiroz-Baez R, Rojas E, Arias C (2009) Oxidative stress promotes JNK-
dependent amyloidogenic processing of normally expressed human APP
chem Int 55:662–670.
ReddyPH (2008) Mitochondrialmedicineforagingandneurodegenerative
diseases. Neuromolecular Med 10:291–315.
Resende R, Moreira PI, Proenc ¸a T, Deshpande A, Busciglio J, Pereira C,
Oliveira CR (2008) Brain oxidative stress in a triple-transgenic mouse
model of Alzheimer disease. Free Radic Biol Med 44:2051–2057.
Rhein V, Baysang G, Rao S, Meier F, Bonert A, Mu ¨ller-Spahn F, Eckert A
(2009) Amyloid-beta leads to impaired cellular respiration, energy pro-
duction and mitochondrial electron chain complex activities in human
neuroblastoma cells. Cell Mol Neurobiol 29:1063–1071.
Rinaldi P, Polidori MC, Metastasio A, Mariani E, Mattioli P, Cherubini A,
Catani M, Cecchetti R, Senin U, Mecocci P (2003) Plasma antioxidants
are similarly depleted in mild cognitive impairment and in Alzheimer’s
disease. Neurobiol Aging 24:915–919.
Rissman RA, Poon WW, Blurton-Jones M, Oddo S, Torp R, Vitek MP,
LaFerlaFM,RohnTT,CotmanCW (2004) Caspase-cleavageoftauisan
early event in Alzheimer disease tangle pathology. J Clin Invest
Rodriguez-CuencaS,Cocheme ´ HM,LoganA,AbakumovaI,PrimeTA,Rose
S, Heales SJ, Lam BY, Neogi SG, McFarlane I, James AM, Smith RA,
Murphy MP (2010) Consequences of long-term oral administration of
the mitochondria-targeted antioxidant MitoQ to wild-type mice. Free
Radic Biol Med 48:161–172.
Rohn TT, Vyas V, Hernandez-Estrada T, Nichol KE, Christie LA, Head E
(2008) Lack of pathology in a triple transgenic mouse model of Alzhei-
mer’s disease after overexpression of the anti-apoptotic protein Bcl-2.
J Neurosci 28:3051–3059.
Royall JA, Ischiropoulos H (1993) Evaluation of 2?,7?-dichlorofluorescin
and dihydrorhodamine 123 as fluorescent probes for intracellular H2O2
in cultured endothelial cells. Arch Biochem Biophys 302:348–355.
Sabbagh MN (2009) Drug development for Alzheimer’s disease: where are
Sano M, Ernesto C, Thomas RG, Klauber MR, Schafer K, Grundman M,
Woodbury P, Growdon J, Cotman CW, Pfeiffer E, Schneider LS, Thal LJ
ment for Alzheimer’s disease. N Engl J Med 336:1216–1222.
De Felice FG (2010) Amyloid-? triggers the release of neuronal hexoki-
nase 1 from mitochondria. PLos One 5:e15230.
Saretzki G, Murphy MP, von Zglinicki T (2003) MitoQ counteracts telo-
mere shortening and elongates lifespan of fibroblasts under mild oxida-
tive stress. Aging Cell 2:141–143.
Setsukinai K, Urano Y, Kakinuma K, Majima HJ, Nagano T (2003) Devel-
opment of novel fluorescence probes that can reliably detect reactive ox-
nyi A, Sun GY (2008) Amyloid beta peptide and NMDA induce ROS
cortical neurons. J Neurochem 106:45–55.
Shigenaga MK, Hagen TM, Ames BN (1994) Oxidative damage and mito-
chondrial decay in aging. Proc Natl Acad Sci USA 91:10771–10778.
Shinall H, Song ES, Hersh LB (2005) Susceptibility of amyloid ? peptide
degrading enzymes to oxidative damage: a potential Alzheimer’s disease
spiral. Biochemistry 44:15345–15350.
Smith RA, Murphy MP (2010) Animal and human studies with the
Smith RA, Porteous CM, Gane AM, Murphy MP (2003) Delivery of bioac-
tive molecules to mitochondria in vivo. Proc Natl Acad Sci USA
Snow BJ, Rolfe FL, Lockhart MM, Frampton CM, O’Sullivan JD, Fung V,
Smith RA, Murphy MP, Taylor KM; Protect Study Group (2010) A
double-blind, placebo-controlled study to assess the mitochondria-
disease. Mov Disord 25:1670–1674.
Prevention of age-related spatial memory deficits in a transgenic mouse
model of Alzheimer’s disease by chronic Ginkgo biloba treatment. Exp
Stepanichev MY, Onufriev MV, Yakovlev AA, Khrenov AI, Peregud DI,
VorontsovaON,LazarevaNA,GulyaevaNV (2008) Amyloid-?(25–35)
increases activity of neuronal NO-synthase in rat brain. Neurochem Int
Sultana R, Butterfield DA (2009) Oxidatively modified, mitochondria-
relevant brain proteins in subjects with Alzheimer disease and mild cog-
nitive impairment. J Bioenerg Biomembr 41:441–446.
Sultana R, Poon HF, Cai J, Pierce WM, Merchant M, Klein JB, Markesbery
WR, Butterfield DA (2006a) Identification of nitrated proteins in Alz-
Sultana R, Boyd-Kimball D, Poon HF, Cai J, Pierce WM, Klein JB, Merchant
M, Markesbery WR, Butterfield DA (2006b) Redox proteomics identi-
fication of oxidized proteins in Alzheimer’s disease hippocampus and
cerebellum: an approach to understand pathological and biochemical
alterations in AD. Neurobiol Aging 27:1564–1576.
Supinski GS, Murphy MP, Callahan LA (2009) MitoQ administration pre-
vents endotoxin-induced cardiac dysfunction. Am J Physiol Regul Integr
Comp Physiol 297:R1095–R1102.
SwerdlowRH,KhanSM (2009) TheAlzheimer’sdiseasemitochondrialcas-
cade hypothesis: an update. Exp Neurol 218:308–315.
Takadera T, Sakura N, Mohri T, Hashimoto T (1993) Toxic effect of
Neurosci Lett 161:41–44.
Takuma K, Yao J, Huang J, Xu H, Chen X, Luddy J, Trillat AC, Stern DM,
Arancio O, Yan SS (2005) ABAD enhances A?-induced cell stress via
mitochondrial dysfunction. FASEB J 19:597–598.
S, Stern DM, Yamada K, Yan SS (2009) RAGE-mediated signaling con-
tributes to intraneuronal transport of amyloid-? and neuronal dysfunc-
tion. Proc Natl Acad Sci USA 106:20021–20026.
Danni O, Smith MA, Perry G, Tabaton M (2002) Oxidative stress in-
creases expression and activity of BACE in NT2 neurons. Neurobiol Dis
Tamagno E, Parola M, Bardini P, Piccini A, Borghi R, Guglielmotto M, San-
toro G, Davit A, Danni O, Smith MA, Perry G, Tabaton M (2005) Beta-
site APP cleaving enzyme up-regulation induced by 4-hydroxynonenal is
mediated by stress-activated protein kinase pathways. J Neurochem
Tong Y, Zhou W, Fung V, Christensen MA, Qing H, Sun X, Song W (2005)
Oxidative stress potentiates BACE1 gene expression and A generation.
J Neural Transm 112:455–469.
Turrens JF (1997) Superoxide production by the mitochondrial respiratory
chain. Biosci Rep 17:3–8.
Velliquette RA, O’Connor T, Vassar R (2005) Energy inhibition elevates
?-secretase levels and activity and is potentially amyloidogenic in APP
15714 • J.Neurosci.,November2,2011 • 31(44):15703–15715 McManusetal.•MitoQPreventsMemoryLossandNeuropathologyin3xTg-ADMice
J Neurosci 25:10874–10883.
Vorhees CV, Williams MT (2006) Morris water maze: procedures for as-
sessing spatial and related forms of learning and memory. Nat Protoc
Walhovd KB, Fjell AM, Brewer J, McEvoy LK, Fennema-Notestine C, Hagler
ing Initiative (2010) Combining MR imaging, positron-emission to-
mography, and CSF biomarkers in the diagnosis and prognosis of
Alzheimer disease. Am J Neuroradiol 31:347–354.
Wang J, Ho L, Zhao W, Ono K, Rosensweig C, Chen L, Humala N, Teplow
DB,PasinettiGM (2008a) Grape-derivedpolyphenolicspreventA?oli-
gomerization and attenuate cognitive deterioration in a mouse model of
Alzheimer’s disease. J Neurosci 28:6388–6392.
Wang X, Su B, Siedlak SL, Moreira PI, Fujioka H, Wang Y, Casadesus G, Zhu
X (2008b) Amyloid-? overproduction causes abnormal mitochondrial
dynamics via differential modulation of mitochondrial fission/fusion
proteins. Proc Natl Acad Sci USA 105:19318–19323.
Yanker BA, Duffy LK, Kirschner DA (1990) Neurotrophic and neurotoxic
effects of amyloid beta protein: reversal by tachykinin neuropeptides.
Yao J, Irwin RW, Zhao L, Nilsen J, Hamilton RT, Brinton RD (2009)
Mitochondrial bioenergetic deficit precedes Alzheimer’s pathology in
female mouse model of Alzheimer’s disease. Proc Natl Acad Sci USA
Zhang Q, Yang G, Li W, Fan Z, Sun A, Luo J, Ke ZJ (2011) Thiamine defi-
tides. Neurobiol Aging 32:42–53.
Zhao XL, Wang WA, Tan JX, Huang JK, Zhang X, Zhang BZ, Wang YH,
YangCheng HY, Zhu HL, Sun XJ, Huang FD (2010) Expression of
?-amyloid induced age-dependent presynaptic and axonal changes in
Drosophila. J Neurosci 30:1512–1522.
McManusetal.•MitoQPreventsMemoryLossandNeuropathologyin3xTg-ADMice J.Neurosci.,November2,2011 • 31(44):15703–15715 • 15715