Mitochondrial Dysfunction: Different Routes to Alzheimer's Disease Therapy

Abstract

Mitochondria are dynamic ATP-generating organelle which contribute to many cellular functions including bioenergetics processes, intracellular calcium regulation, alteration of reduction-oxidation potential of cells, free radical scavenging, and activation of caspase mediated cell death. Mitochondrial functions can be negatively affected by amyloid β peptide (Aβ), an important component in Alzheimer's disease (AD) pathogenesis, and Aβ can interact with mitochondria and cause mitochondrial dysfunction. One of the most accepted hypotheses for AD onset implicates that mitochondrial dysfunction and oxidative stress are one of the primary events in the insurgence of the pathology. Here, we examine structural and functional mitochondrial changes in presence of Aβ. In particular we review data concerning Aβ import into mitochondrion and its involvement in mitochondrial oxidative stress, bioenergetics, biogenesis, trafficking, mitochondrial permeability transition pore (mPTP) formation, and mitochondrial protein interaction. Moreover, the development of AD therapy targeting mitochondria is also discussed.

Full text

Review Article
Mitochondrial Dysfunction: Different Routes to
Alzheimer’s Disease Therapy
Pasquale Picone, Domenico Nuzzo, Luca Caruana, Valeria Scafidi, and Marta Di Carlo
Istituto di Biomedicina ed Immunologia Molecolare (IBIM) “Alberto Monroy,” CNR, via Ugo La Malfa 153,
90146 Palermo, Italy
Correspondence should be addressed to Marta Di Carlo; marta.dicarlo@ibim.cnr.it
Received  March ; Accepted  May ; Published  August 
Academic Editor: Giles E. Hardingham
Copyright ©  Pasquale Picone et a l. is is an open access article distributed under the Creative Commons Attribution License,
which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Mitochondria are dynamic ATP-generating organelle which contribute to many cellular functions including bioenergetics
processes, intracellular calcium regulation, alteration of reduction-oxidation potential of cells, free radical scavenging, and
activation of caspase mediated cell death. Mitochondrial functions can be negatively aected by amyloid 𝛽peptide (A𝛽), an
important component in Alzheimer’s disease (AD) pathogenesis, and A𝛽can interact with mitochondria and cause mitochondrial
dysfunction. One of the most accepted hypotheses for AD onset implicates that mitochondrial dysfunction and oxidative stress
are one of the primary events in the insurgence of the pathology. Here, we examine structural and functional mitochondrial
changesinpresenceofA𝛽. In particular we review data concerning A𝛽import into mitochondrion and its involvement
in mitochondrial oxidative stress, bioenergetics, biogenesis, tracking, mitochondrial permeability transition pore (mPTP)
formation, and mitochondrial protein interaction. Moreover, the development of AD therapy targeting mitochondria is also
discussed.
1. Introduction
Alzheimer’s disease (AD) is an age-related progressive neu-
rodegenerative disorder characterized by impairment of cog-
nitive function. e neuropathology of AD concerns two
neurodegenerative processes: amyloidogenesis, leading to the
presence of extracellular amyloid 𝛽-peptide (A𝛽) deposition,
and neurobrillary degeneration, corresponding to the for-
mation of intracellular tangles composed of phosphorylated
Tau protei n [ ]. e presence of these abnormal structures
leads to neuronal dysfunction and cell death. Many lines
of evidence suggest that the mitochondrion plays a central
role in neurodegenerative diseases, including AD [], and
thesocalledmitochondrialcascadehypothesisproposes
that mitochondrial dysfunction is the primary event in AD
pathology []. Mitochondrion is a cellular organelle required
for bioenergetics processes and is also involved in amino-
acid, lipid, and steroid metabolism, calcium homeostasis,
free radicals production, and apoptosis triggering. In the
brain, where there is the highest energy request and con-
sumption, the number of mitochondria is elevated mainly in
the synapses and impairment could be a serious threat for
neurons survival.
Many aspects of the strict link between mitochondrial
dysfunction and AD remain to be elucidated, but some
evidence indicates that the progressive accumulation of A𝛽
inmitochondriacouldbetherelationshipformitochondria-
mediated toxicity []. e Amyloid Precursor Protein (APP)
was, indeed, found accumulated in the mitochondrial import
channels and A𝛽was found interacting with some mitochon-
drial proteins [].
Here we dissect dierent points in which the mito-
chondrial functionality could be aected by A𝛽presence,
producing evidence from studies on AD human postmortem
brains as well as cellular and AD animal models. Moreover,
we analyze how these vulnerable points for A𝛽-mediated
mitochondrial dysfunction may be distinctive and perhaps
complementary therapeutic intervention targets for AD.
Hindawi Publishing Corporation
Oxidative Medicine and Cellular Longevity
Volume 2014, Article ID 780179, 11 pages
http://dx.doi.org/10.1155/2014/780179

Oxidative Medicine and Cellular Longevity
2. Mitochondria in Neuronal Cells
High metabolic energy is required by human brain for its
function. Since neurons have a limited glycolytic capacity
this kind of cells is extremely dependent on mitochondrial
energy production []. Neurons are cells with a particular
morphology extending their axons and dendrites from mil-
limeters up to a meter. Mitochondria are spread along the
cells probably with a major presence in some regions of
the neurons, as the synapses, that have the highest demand
for ATP production and energy consumption. When a
transmitter is released by a synapsis several ion channels
areopenedinthepostsynapticmembranetopermitinux
of ions. is pumping needs high energy consumption.
Moreover, ATP produced by mitochondria is also consumed,
together with all the normal requirements of a cell, for action
potential to restore ion gradients and axonal transport. An
imaging study established that a quiescent cortical neuron
expends . billion ATP molecules per second []. A cor-
rect distribution of mitochondria into the neuronal regions
in which a local demand for ATP or Ca2+ is necessary
results to be important. us, tracking of mitochondria
is essential for neurons survival. Biochemical and imaging
studies have demonstrated that mitochondrial translocation
involves a motor/adaptor complex formed by microtubule-
based motors proteins, as kinesin and dynein, together with
two mitochondrion-specic proteins, milton and miro [].
Moreover, mitochondrion together with energy metabolism
discussed above plays in neurons a pivotal role in cell survival
and death by regulating apoptotic pathways and contributing
to dierent cellular functions including intracellular calcium
homeostasis, maintaining the cellular redox potential, cell
cycle regulation, and synaptic plasticity []. Mitochondrion
has a role in establishing the polarity by reducing the Ca2+
concentration at the base of the presumptive axon, especially
in neurons, thereby promoting polymerization of micro-
tubules and the rapid neuronal growth and dierentiation,
accompanied by an increased amount of the mitochondria
number per cell. is observation was achieved by studies
in which treatment with chloramphenicol, an inhibitor of
mitochondrial protein synthesis, prevents dierentiation of
the cells whereas oligomycin, an inhibitor of the mitochon-
drial ATP synthase, does not, thus suggesting that increased
mitochondrial mass, but not ATP production, is required
for neuronal dierentiation []. Mitochondrial Ca2+ aects
neurogenesis as suggested by results showing that an increase
of mitochondrial fusion and intramitochondrial Ca2+ levels
is observable when neuroblastoma cells are induced to stop
division and begin dierentiation in neuro-like cells [].
During axons and dendrites dierentiation, their growth
and synaptic junction may be inuenced by mitochondrial
motility and functions. During the formation of axonal
branches mitochondria respond to changes by modifying
their entry into branches; the process does not require an
active growth cone, suggesting the involvement of a dierent
mechanism [].
Furthermore, another intriguing mitochondrial role
occurs during the self-renewal of neural stem cells. Stud-
ies on mouse embryonic stem (ES) cells suggest that the
Nonamyloidgenic
pathway pathway
Amyloidgenic
AICD
CTF𝛼
APP
C-Ter
N-Ter
CTF𝛽
𝛼-Secretase
sAPP𝛼sAPP𝛽
A𝛽aggregates
A𝛽
𝛾-Secretase
𝛽-Secretase
F : Nonamyloidgenic or amyloidgenic pathways are origi-
nated by dierent APP processing: the combined cleavage of 𝛼-and
𝛾-secretase produces the sAPP𝛼and CTF-𝛼fragments preventing
A𝛽generation; in contrast, 𝛽-secretase cleaves in a dierent site of
APP thus originating, together with𝛾-secretase complex, the sAPP𝛽
and A𝛽fragments and the intracellular AICD fragment. A𝛽through
a misfolding step forms brillar aggregates.
proliferative capacity is correlated with low mitochondrial
oxygen consumption and high levels of glycolytic activity
[]. Human ES cells exhibit an anaerobic metabolic prole.
When somatic cells are induced to revert to an ES cell-like
phenotypetheirmitochondriafollowthesamefatechanging
their morphology, subcellular distribution, biogenesis, and
ROS and ATP production [].
On the basis of these observations it is easy to understand
how several inherited diseases are caused by mutations in
mitochondrial DNA, and cells as muscle cells and neurons,
with high energy demands, are the most aected of these dis-
orders. Furthermore, impaired mitochondrial activity causes
the most common neurodegenerative disorders, such as
Alzheimer’s, Parkinson’s and Huntington’s diseases, stroke,
and psychiatric disorders.
3. APP Metabolism and A𝛽Generation
Alzheimer’s disease is a devastating neurodegenerative disor-
der with a progressive cognitive impairment and dementia.
AD pathogenesis is believed to be triggered by the A𝛽
accumulation,whichisduetooverproductionofA𝛽and/or
the failure of the clearance mechanisms. A𝛽is generated by
sequential cleavages of its larger precursor, a protein called
amyloid precursor protein (APP). APP is an integral mem-
brane protein with a single membrane spanning domain,
a large extracellular glycosylated N terminus and a shorter
cytoplasmic C terminus. APP is produced in several dierent
isoforms and the most abundant form in brain (APP) is
produced mainly by neurons and diverges from the longest
one because of the lacking of a kunitz-type protease inhibitor
sequence in its ectodomain [,]. APP processing is divided
into the nonamyloidogenic pathway and the amyloidogenic
pathway (see Figure ). e nonamyloidgenic processing is
initiated by 𝛼-secretases, cleaving within the A𝛽domain

Oxidative Medicine and Cellular Longevity 
and preventing the release of A𝛽[]. Alternatively, 𝛽-
secretase (BACE) cleavage initiates the amyloidogenic path-
way and generates the soluble 𝛽-secreted APP (sAPP𝛽)and
the membrane bound fragment CTF𝛽. Both fragments are
substrates for 𝛾-secretase, a multisubunit protease complex
comprising proteins as presenilin  or  (PS, PS) [–].
e further processing of CTF𝛽releases the amyloidogenic
A𝛽fragment [,]. Interestingly, 𝛾-secretase seems to
cleave the CTF𝛽exactly in the middle of the membrane
domain [], suggesting the hypothesis that the formation
of dierent A𝛽species (A𝛽38,A𝛽40,andA𝛽42) is dependent
on the membrane properties. Each A𝛽species have dierent
lipophilic properties and dierent tendencies to form A𝛽
oligomers and aggregates. Notably, the ratio A𝛽40/A𝛽42 is of
clinical relevance in AD []. However, in AD, the produced
A𝛽misfolds and self-aggregates into oligomers of various
sizes and forms, up to produce diuse amyloid neuritic
plaques. A𝛽oligomers and plaques are potent synaptotoxins,
block proteasome function, inhibit mitochondrial activity,
alter intracellular Ca2+ levels, and stimulate inammatory
processes.
4. Mitochondrial A𝛽
A𝛽, as well as its extracellular canonical localization, has
been found in dierent subcellular compartment including
the endoplasmic reticulum (ER), the Golgi apparatus or the
trans-Golgi network, the early, late, or recycling endosomes,
and the lysosome, where probably is generated []. However,
presence of A𝛽has been also observed in mitochondria
[] and studies from dierent independent groups have
clearly demonstrated that A𝛽progressively accumulates
within mitochondria of both human AD brain and Tg
mouse models for AD []. Moreover, nding indicates
that accumulation of A𝛽in mitochondria begins before the
occurrence of the extracellular deposition as demonstrated
by experiments in which its presence in Tg mA𝛽PP mice
arises as early as - months and increases with aging [].
e presence of A𝛽in mitochondria was also evidenced
by an interesting study in which APP modulates cell death
through interaction with a newly identied mitochondrial
membrane proapoptotic protein, called Appoptosin, involved
inthehemesynthesis[]. ese ndings raise the question
whether A𝛽in mitochondria is in situ generated or imported.
Several observations from dierent groups and experimental
approaches demonstrated that A𝛽is not locally produced.
ishypothesisissupportedbythefactthattheabilityof𝛾-
secretase to cleave APP associated with mitochondria is not
known [,]. us, it is supposed that A𝛽is derived from its
extracellular or intracellular pool and that a cellular track-
ing is involved in the internalization of A𝛽in mitochondria.
Recent ndings, using isolated rat mitochondria, have shown
that a specic uptake mechanism for import of A𝛽in
mitochondria involves the translocase of the outer membrane
(TOM) complex []. is data was supported by the fact
that extracellularly applied A𝛽is internalized in cells and
colocalizes with mitochondrial markers and that is associated
with the mitochondrial inner membrane aer import. Other
evidences come from experiments using confocal microscopy
showing that the A𝛽42 fragments colocalize with complex
II of the respiratory chain, the outer cell membrane, the
mitochondrial membrane, and the mitochondrial chaperon
matrix protein Hsp [,].
5. Mitochondrial Dysfunction and
Oxidative Stress Induced By A𝛽
Mitochondria have a pivotal role in cellular energy
metabolism but are also involved in amino-acid, lipid, and
steroid metabolism, modulation of cellular calcium levels,
production of free radicals, and regulation of apoptosis, key
features of neurodegeneration. Despite extensive research
eorts to understand the pathophysiology of neurological
diseases with respect to mitochondrial dysfunction, the
exact mechanism is still not established. However, it seems
clear that mitochondrial dysfunction plays a well-dened
role in neurodegenerative diseases, making it a topic of
high interest in neuroscience research today. An immediate
consequence of mitochondrial dysfunction is the increase
of reactive oxygen species (ROS) production that promotes
oxidative damage to DNA, RNA, proteins, and lipids.
Mitochondria are the primary cellular consumers of oxygen
and contain numerous redox enzymes capable of transferring
single electrons to oxygen, generating the superoxide
(O2−) molecules. Mitochondria also contain an extensive
antioxidant defense system to detoxify the generated ROS.
When mitochondria are damaged, the antioxidant defense
decreases thus increasing ROS production that can further
damage mitochondria, causing more free radicals generation
and loss or depletion of antioxidant capacity. Under normal
physiological conditions, ROS act as “redox messengers”
regulating intracellular signalling, whereas imbalance of ROS
induces irreversible damage to cellular components leading
to cell death. Mitochondrial ROS can originate from multiple
reactions in the TCA cycle and/or in the respiratory chain.
Several in vitro studies suggested a link between elevated A𝛽
levels, mitochondrial dysfunction, and oxidative stress, all
factors that contribute to AD pathogenesis. Moreover, we
reported that A𝛽is a key factor in free radical generation,
oxidative damage, and mitochondrial dysfunction, activating
a cascade of events leading to neurodegeneration in
neuroblastoma LAN cells [–]andseaurchinmodel
system [].
Mitochondrial A𝛽presence strongly inuences mito-
chondrial respiratory function, ROS production rates, and
alters mitochondrial membrane potential in dierent brain
regions of AD mouse models. Moreover, a dierent mito-
chondrial distribution was evidenced. e elevated A𝛽pres-
ence and the mitochondrial dysfunction were found in
hippocampus and cortex, brain regions devoted to memory,
whereas lower levels were found in striatum and amygdala. A
striking association between mitochondrial impairment and
cognitive dysfunction in the A𝛽PP and A𝛽PP/PS mice was
also found. is is the rst demonstration of an association
between mitochondrial A𝛽levels, mitochondrial dysfunc-
tion, and cognitive impairment in AD transgenic mice [].

Oxidative Medicine and Cellular Longevity
Xie et al. for the rst time showed how the structure and
function of mitochondria changed in the living brain of
transgenic animals developing amyloid deposits, by using
intravital multiphoton imaging, with a range of uorescent
markers. e authors observed that severe impairments were
limitedinthebrainregionsclosetotheA𝛽plaques. In
these regions a decreased number of mitochondria were
found; some of them were dystrophic and fragmented and
thesurvivedonesshowedareducedmembranepotential.
Both neuronal soma and neuritis with oxidative stress show
severe alterations in mitochondrial membrane potential.
ese results provide in vivo evidence that A𝛽plaques can
be focal sources of toxicity leading to severe structural and
functional abnormalities in mitochondria []. Oxidative
stress may activate signalling pathways that alter APP or
Tau processing. For example, oxidative stress increases the
expression of 𝛽-secretase through activation of c-Jun amino-
terminal kinase and p mitogen-activated protein kinase
(MAPK) [] and increases aberrant Tau phosphorylation
by activation of glycogen synthase kinase -𝛽(GSK-𝛽)
[]. Oxidant-induced inactivation of specic molecules may
also be important. Using a proteomic approach, the prolyl
isomerase PIN was found to be particularly sensitive to
oxidative damage, being highly downregulated and oxidized
in hippocampus of AD patients []. e oxidative modi-
cation of PIN was related to the loss of isomerase activity
that is retained critical for neurobrillary tangle forma-
tion. Later studies have demonstrated that PIN catalyses
protein conformational changes that aect both APP and
Tau p roc e s s ing [ ]. Knockout of PIN increases amyloido-
genic APP processing and intracellular A𝛽levels in mice.
PIN-knockout mice also exhibit Tau hyperphosphorylation,
motor and behavioural decits, and neuronal degeneration
[].
Moreover, the cause of mitochondrial dysfunction could
be the alteration of mitochondria-associated ER membrane
(MAM)function.MAMisadynamicsubcompartmentof
ER with a lipid ra-like structure intimately involved in
cholesterol and phospholipid metabolism, calcium home-
ostasis, mitochondrial function and dynamics, bioenergetics,
and cell signaling [–]. MAM is physically, biochemically,
and reversibly associated to mitochondria and this organellar
contact site is a critical intracellular signaling platform that
determines cellular life and death. During cellular stress
situations,likeanalteredcellularredoxstate,theMAMalters
its set of regulatory proteins and consequently alters MAM
functions. Presenilins and 𝛾-secretaseareenrichedinER
subcompartment and it has been hypothesized that genetic
and biochemical alterations in these factors aecting MAM
function should be relevant for increased APP processing
and AD progression. Moreover, in agreement with this
hypothesis both in model systems as PS-knockdown cells and
in broblasts from patientshaving familial (FDA) or sporadic
(SDA) forms of AD, MAM functionality, measured by the
amount of cholesterol and phosphatidylserine production,
is increased [–]. Similarly an increased expression of
MAM-associated proteins is found in postmortem AD brains
[].
6. A𝛽Affects Mitochondrial Bioenergetics
e bioenergetics eect induced by endogenous A𝛽on
mitochondrion was investigated both in WD and PA,
two cell lines used as cellular model for AD []. Functional
impairment of the respiratory chain was found distributed
among the protein complexes especially in complex I and
complex IV. Measurements of ATP concentration showed,
in WD and PA cells, that its synthesis, by oxidative
phosphorylation, was decreased by ∼%, and this loss
was somewhat compensated by glycolysis (Warburg eect).
Compensation proved to be more ecient in WD than in
PA cells in agreement with the highest ROS production of
the latter cell line. Moreover, the mitochondrial membrane
potential was % and % lower in WD and PA cells,
respectively, compared to Chinese hamster ovary (CHO)
controls. Moreover, in dierent AD models, the voltage-
dependent anion channel VDAC, a major component of the
outer mitochondrial membrane that regulates ion uxes and
metabolites,isdamagedasaresultofoxidativestress[]. e
lipid composition of lipid ras, key membrane microdomains
that facilitate the transfer of substrates, and protein-protein
and lipid-protein interactions is altered as a result of the
abnormally low levels of n- long chain polyunsaturated
fatty acids (mainly docosahexaenoic acid) that increase vis-
cosity and augment energy consumption. Abnormal lipid
ra composition may also modify the activity of the key
enzymesthatmodulatethecleavageoftheAPPtoform
toxic A𝛽[]. Cholesterol and sphingolipids, that are rich in
membrane microdomains, change their metabolism during
normalbrainagingandinthebrainsofADpatientsresulting
in accumulation of long chain ceramides and cholesterol
[]. Similarly, exposure of hippocampal neurons to A𝛽
induces membrane oxidative stress that perturbs cholesterol
metabolism and activates sphingomyelinases, resulting in
increased ceramide production. In contrast, treatment of
neurons with 𝛼-tocopherol or an inhibitor of sphingomyelin
synthesis prevents accumulation of ceramides and cholesterol
and protects them against A𝛽induced death [].
All these alterations converge in a severe bioenergetics
mitochondria impairment of the AD cells, with the extent
of mitochondrial dysfunction being correlated with the accu-
mulation of A𝛽and oligomers.
MitochondriaaretheprincipalsitefortheATPpro-
duction oxidative phosphorylation (OXPHOS) system. e
mitochondrial OXPHOS machinery, whose components are
produced by mitochondrial genome, is composed of ve
multisubunit complexes (complexes I–V). Several studies
have shown that direct exposure to A𝛽signicantly impairs
functionality of the mitochondrial electron transport chain
(ETC). e ETC is essential to ATP production and its
constituent enzyme complexes are a major source of ROS
generation, especially when one or more of the enzyme
complexes is inhibited [–]. Moreover, age-related bioen-
ergetics decit was described in female of xTg-AD mice
aged from  to  months []. A decreased activity of
OXPHOS, pyruvate dehydrogenase (PDH), and cytochrome
𝑐oxidase (COX) regulatory enzymes and increased oxidative
stress and lipid peroxidation were found. Most of the eects

Oxidative Medicine and Cellular Longevity 
on mitochondria appeared at the age of  months, whereas
mitochondrial respiration was signicantly decreased at 
months of age. Importantly, mitochondrial bioenergetics
decit preexists at the development of AD pathology in xTg-
AD mice.
APP is known to be alternatively spliced to produce three
major isoforms in the brain: APP, APP, and APP.
Both APP and APP contain the Kunitz protease
inhibitory (KPI) domain, but the former contains also an
extra OX- domain. APP on the other hand lacks both
domains. In AD, upregulation of the KPI-containing APP
isoforms has been reported. Chua et al. found that the KPI-
containing APP signicantly decreased the expression
of three major mitochondrial metabolic enzymes such as
citrate synthase, succinate dehydrogenase, and cytochrome 𝑐
oxidase (COX IV). is reduction lowers the NAD+/NADH
ratio, COX IV activity, and mitochondrial membrane poten-
tial [].
7. Mitochondrial Biogenesis and AD
During the life cycle, the biogenesis of new mitochondria
plays an essential role in maintaining healthy mitochondria
in eukaryotic cells. Mitochondrial biogenesis could also
havethepotentialitytoquicklyrespondtochangesdueto
mitochondrial damage or increased demand in response to
environmental stimuli. Mitochondrial biogenesis is regulated
by the PGC- 𝛼-NRFTFAM pathway. Expression levels of
PGC- 𝛼, NRF , NRF , and TFAM were signicantly
decreased in both AD hippocampal tissue and APPswe M
cells, suggesting that mitochondrial biogenesis was aected
during neurodegeneration. Moreover, it has been demon-
strated that impaired mitochondrial biogenesis contributes
to mitochondrial dysfunction in AD and its enhancing
may represent a potential pharmacologic approach for the
treatment of AD [].
8. Abnormal Mitochondrial Dynamics in AD
Despite traditional knowledge, mitochondria are now con-
sidered highly dynamic organelles that move throughout
a cell and regularly fuse and divide. Increasing evidence
suggests that abnormal mitochondrial dynamics, such as
increased ssion and decreased fusion, are early and key
factors that have been found in neurodegenerative diseases,
such as AD. ese processes observed in axons and dendrites,
as in all cells, allow the exchange of materials between
mitochondria []. Even brief contact can involve fusion and
extensive exchange of proteins in each compartment of the
mitochondrion, as it has been shown in nonneuronal cells
[]. In addition, these abnormal mitochondrial dynamics
have been associated with unusual changes in mitochondria
structure. Abnormal mitochondrial ssion and fusion were
reported in AD []. In this study, the authors aimed to
determine whether APP and A𝛽cause mitochondrial and
neuronal dysfunction through modulation of mitochon-
drial dynamics []. Further, both confocal and electron
microscopy analyses demonstrated that APP overexpression
causes mitochondrial fragmentation in neurons []. Since
the balance of mitochondrial ssion and fusion tightly con-
trols mitochondrial morphology, it was hypothesized that
APP-induced mitochondrial fragmentation was caused by
enhanced ssion and reduced fusion []. Recent studies
show that the fusion dynamin-related protein  (Drp), also
known as dynamin-like protein  (DLP), a protein that
maintains and remodels mammalian mitochondria, interacts
with A𝛽and phosphorylates Tau, leading to excessive mito-
chondrial fragmentation, impaired axonal transport of mito-
chondria, and lastly neuronal damage and cognitive decline
[]. However, in agreement with the idea that mitochondria
dynamics are altered in AD neurons, signicant changes in
the expression of proteins involved in mitochondrial ssion
and fusion were reported. Reduced expression of all the
fusion proteins (i.e., OPA, Mfn, and Mfn) and increased
expression of ssion protein Fis were described [–].
9. A𝛽Interacts and Interferes with
Mitochondrial Proteins
Structurally and functionally intact mitochondria are crucial
for healthy cells. A mitochondrion contains outer and inner
membranes composed of phospholipid bilayers and proteins
having dierent properties and their integrity is essential for
itsgoodfunctionality.Incelldeathmechanisms,asapop-
tosis and necrosis, an increase of mitochondrial membrane
permeability is one of the key events. Mitochondria isolated
from a variety of sources can show an abrupt increase in
the permeability of the inner mitochondrial membrane to
solutes with a molecular mass of less than , Da, which
results in the loss of mitochondrial membrane potential
(ΔΨm), mitochondrial swelling, and rupture of the outer
mitochondrial membrane. is process is better known as
the mitochondrial membrane permeability transition (MPT)
[]. e MPT can be induced under various conditions, such
as exposure of mitochondria to Ca2+ together with inorganic
phosphate. Although the molecular mechanisms of the MPT
are largely unknown, the most widely accepted model is that
it occurs aer the opening of a channel complex that has
been termed the permeability transition pore (PTP), which
is thought to consist of the voltage-dependent anion channel
(VDAC: outer membrane channel), the adenine nucleotide
translocator (ANT: inner membrane channel), cyclophilin D
(CypD), and possibly other molecule(s). CypD, a peptidyl-
prolyl isomerase F, resides in the mitochondrial matrix but
becomes associated with the inner mitochondrial membrane
duringtheMPTanditplaysacentralroleinopening
the mitochondrial membrane permeability transition pore
(mPTP). Recent studies provide evidence that CypD binds to
A𝛽and forms a complex with A𝛽in cortical mitochondria
of AD patients and Tg A𝛽PP mice []. e interaction of
A𝛽with CypD provokes mitochondrial and neuronal pertur-
bation []. is interaction causes the formation of mPTP,
resulting in decreased mitochondrial membrane potential,
compromised mitochondrial respiration function, increased
oxidative stress, release of cytochrome 𝑐, and impaired axonal
mitochondrial transport. It has been suggested that mPTP

Oxidative Medicine and Cellular Longevity
could be a possible therapeutic target. e design of small
molecules able to interfere with A𝛽-CypD complex could,
indeed, decrease A𝛽neurotoxicity eects [].
e involvement of mitochondria in the pathogenic
pathway of A𝛽was also conrmed by specic binding
of A𝛽and A𝛽PP to mitochondrial proteins, which causes
energy impairment and cell physiology defects. Firstly, A𝛽
specically binds to the mitochondrial A𝛽-binding alcohol
dehydrogenase (ABAD), an intracellular enzyme present in
the mitochondrial matrix []. In presence of A𝛽,ABAD
increases cell stress, DNA fragmentation, and ROS genera-
tion induced by A𝛽. Moreover, the expression levels of ABAD
are related to mitochondrial A𝛽levels being ABAD expres-
sion levels signicantly higher in AD-aected brain regions,
as hippocampus, than in healthy brain []. Recent experi-
ments indicate that the – ABAD residues interact with
A𝛽and inhibition of the ABAD-A𝛽interaction signicantly
reduces mitochondrial A𝛽accumulation and protects against
aberrant mitochondrial and neuronal function, improving
spatial learning/memory in Tg AD mice [–]. Proteomics
studies on Tg mAPP/ABAD mice showed that in brain the
ABAD-A𝛽interaction also aects the expression of proteins
such as Ep- (endophilin-), a cytoplasmic SH domain-
containing protein, located in presynaptic nerve termini
[] and Prdx- (peroxiredoxin-), an antioxidant protein
[], both of which were also found to be overexpressed
in human AD brains [,]. Even if the link between
these two proteins and mitochondrial dysfunction remains
unclearsomeevidencesindicatethatincreasedlevelsofEp-
 cause the activation of JNK (c-Jun-N-terminal kinase)
[,]. JNK is a stress kinase that has been linked to A𝛽
production and neurons death []. us, all these data
support the signicance of ABAD to A𝛽induced neuronal
stress. Several enzymes, such as presequence protease (PreP),
a metalloprotease containing an inverted zinc binding motif,
were identied to degrade A𝛽[,]. Intramitochondrial
localization studies demonstrated that PreP is localized
within the mitochondrial matrix. PreP activity is severely
reducedinsamplesofmitochondrialmatrixisolatedfrom
the temporal lobe of AD patients, compared to age-matched
controls []. Moreover, exposure of puried hPreP to an
oxidizing agent, hydrogen peroxide, results in decreased
peptidolytic activity []. During oxidative phosphorylation,
the electron transporters release energy that drives the pro-
duction of ATP. A𝛽may inhibit mitochondrial respiration,
by interacting with the subunit 𝛼of ATP synthase causing
ATP depletion []. Another study reports that A𝛽decreases
cytochrome 𝑐oxidase activity, but the specic mechanism is
still unclear [].Incontrast,anincreaseintheactivityof
the cytochrome 𝑐reductase (complex III) has been reported
[]. Interestingly, complex III appears to be one of the sites
of superoxide radical production. Bobba et al. found that A𝛽
caused a selective defect in complex I activity, associated with
an increase of intracellular ROS (-fold) and an impairment
ofcomplexIV,likelyduetomembranelipidperoxidation.
In addition, a % increase of the GSSG/GSH ratio was
measured in AD brains with respect to age-matched controls
[].
10. Therapeutic Strategies against A𝛽Induced
Mitochondrial Toxicity
e free radical and oxidative stress theory of aging suggests
that oxidative damage is a major player in neuronal degener-
ation and oxidative stress has a well-established pathophysio-
logical feature in AD []. However, up to now use of antiox-
idants in prevention or therapy gives controversial results.
One possible explanation is related to the low permeability
of the blood brain barrier (BBB) to most of the currently
used antioxidants. To overcome these potential diculties,
researchers have formulated new delivery systems, such
as those based on nanoparticles, which might represent a
successful strategy for drug delivery into CNS. Moreover,
on the basis of the consideration that mitochondria are
the major source of ROS and are particularly vulnerable
to oxidative stress, one would predict that a more ecient
therapy could consider use of antioxidants alleviating mito-
chondrial dysfunction. is prompted researchers to develop
antioxidant therapy directed to mitochondrion, through the
development of specically designed mitochondria-targeted
antioxidants [,]. is might be a strategy to overcome
the apparent clinical ineciency of antioxidants that do
not target oxidative stress in this organelle []. e Szeto-
Schiller (SS) peptides, a family of small mitochondria-
targeted antioxidantmolecules, were developed as a potential
treatment for AD []. ese SS peptides display mitochon-
drial accumulation having a sequence motif that allows them
to target mitochondria. ey scavenge H2O2and inhibit lipid
peroxidation. eir antioxidant action can be attributed to
the tyrosine, or dimethyltyrosine (Dmt), that plays a role in
scavenging mitochondrial ROS. SS, in particular, is capable
of entering mitochondria and of concentrating in the inner
mitochondrial membrane, protecting mitochondria against
mPTP formation, swelling, and cytochrome 𝑐release. Using
SS in dierent AD models as Na neuroblastoma cells
treated with A𝛽–, primary neurons from Tg mice and
aged Tg mice, a range of eects on mitochondria were
observed [,]. Increased expression of mitochondrial
ssion genes and decreased expression of fusion genes were
found together with increased number of mitochondria,
indicating that mitochondria fragmentation occurred. On
the basis of these results, the Antipodean Pharmaceuticals
Inc. patented MitoQ, a mitochondria-targeted antioxidant.
is drug is now undergoing phase II clinical trials for the
potential treatment of several diseases in which mitochon-
drial oxidative damage is implicated, including neurodegen-
erative diseases []. MitoQ was designed to accumulate
extensively within mitochondria in vivo in order to increase
the local antioxidant capacity against oxidative damage. e
active drug is the ubiquinone, which is identical to the
antioxidant component of the respiratory chain constituent
coenzyme Q (CoQ). An aliphatic -carbon chain, to
the lipophilic cation triphenylphosphonium, that drives its
selective uptake into mitochondria in a membrane potential-
dependent manner, covalently links this component. Once
internalized by mitochondria, it adsorbs in the phospholipid
bilayers, where it is readily reduced to the active ubiquinol
form MitoQH, which exerts its antioxidant properties [].

Oxidative Medicine and Cellular Longevity 
Prefrontal cortex
Amygdala
Hippocampus
Brainstem
Cerebellum
Mitochondria from the hippocampus and cortex show highest levels of A𝛽
A𝛽
A𝛽
and
degree of mitochondrial dysfunction followed by the striatum, with
only a minimal presence on the amygdala
Impaired mitochondrial
biogenesis
Oxidative
stress
Abnormal mitochondrial
dynamics
Accumulation of A𝛽
(import through TOM)
Alteration of the complexes I, III, and IV
Mitochondrial
Dysfunction
Formation of mPTP
Decit in bioenergetics
F : Mitochondrial dysfunction induced by A𝛽. In AD brain some areas are more sensitive to mitochondrial dysfunction. In the neurons
A𝛽induces mitochondrial dysfunction by using dierent mechanisms. A𝛽is taken up by mitochondria via the TOM complex and imported
in the inner membrane; A𝛽alters the enzyme activity of the respiratory chain complexes I, II, and IV; A𝛽aects mitochondrial dynamics
by impaired balance of ssion and fusion; A𝛽causes formation of mPTP via interaction with CypD; A𝛽induces decreased mitochondrial
respiration; A𝛽aects new mitochondrial biogenesis; A𝛽increases ROS generation.
Since it enters into mitochondria several hundred-fold more
than natural antioxidants, it rapidly neutralizes free radi-
cals at their source, before they reach their targets, thus
showing an improved therapeutic potential []. For all
these proprieties, MitoQ is a promising antioxidant candidate
for AD treatment []. Recently, using cytoplasmic hybrid
(cybrid) neurons from AD and age-matched non-AD human
subjects, it was demonstrated that treatment with the antiox-
idant probucol protects against AD mitochondria-induced
extracellular signal-regulated kinase (ERK) activation and
mitochondrial ssion-fusion imbalances. In fact, inhibition
of ERK activation not only attenuates aberrant mitochondrial
morphology and function but also reestablishes the mito-
chondrial ssion and fusion balance, as conrmed by changes
in expression and distribution of DLP and Mfn [].
Among antioxidant molecules used in prevention and
treatment of AD, a relevant role is played by natural antiox-
idants []. e curry spice curcumin shows antioxidant
[], anti-inammatory [], and amyloid-disaggregating
properties [], eects that have been largely studied both
in vitro and in vivo models. A pilot clinical trial to develop
procedures for testing the eectiveness of curcumin on
slowing AD progression was carried out []. irty-four
human subjects with AD received daily placebo or two
dierent doses of curcumin for  months. At dierent times
of the study, cognitive tests were performed, and blood
samples were analyzed for levels of isoprostane, amyloid
beta protein, metals, and cholesterol. ere was no cognitive
decline in the placebo group, and no improvement was
observed with curcumin. Moreover, curcumin presented low
bioavailability and photodegradation. With such issues in
mind,MarracheandDharformulatedtargetedcurcumin-
loaded NPs (mitochondria-targeted polymeric nanoparticle
system) to provide photostability and enhance mitochondrial
uptake. An in vitro cytotoxicity assay, using A𝛽treated
human neuroblastoma IMR- cells, demonstrated enhanced
neuroprotection with the targeted curcumin NPs compared
with the no-targeted curcumin NPs or free curcumin against
A𝛽, which accounts for the targeted delivery of curcumin
into the mitochondria of cells []. Ferulic acid (FA) is
another natural antioxidant having a neuroprotective eect
against oxidative stress and cell death induced by A𝛽42
oligomers []. FA was also successfully conjugated with solid
lipid nanoparticles to improve its delivery and enhance its
potential antioxidant therapeutic eect []. Moreover, other
data suggest that natural plants such as a standardized Ginkgo
biloba extract or the green tea component epigallocatechin-
-gallate may be promising treatment strategies. Of note, in
addition to their antioxidative properties, these compounds
stabilize mitochondrial functions such as the mitochondrial
membrane potential, ATP levels, and mitochondrial respira-
tory complexes [,]. As discussed above, the interaction
of A𝛽with CypD provokes mitochondrial perturbation and
formation of mPTP events leading to neuronal degeneration.
us, inhibition of mPTP formation by blocking CypD is a
rational target for potential therapeutic AD strategies.

Oxidative Medicine and Cellular Longevity
11. Conclusions
Mitochondrial dysfunction is an early feature of Alzheimer’s
disease. Extracellular or intracellular A𝛽is imported into
the mitochondria through the TOM machinery. e pro-
gressive accumulation of mitochondrial A𝛽is associated
with aberrant mitochondrial functions leading to neuronal
damage and cognitive decline. e mitotoxicity induced by
A𝛽is still not clear but includes numerous mechanisms.
A𝛽induced mitochondrial dysfunction contributes to energy
metabolism impairment, defects in key respiratory enzyme
activity/function, accumulation/generation of mitochondrial
ROS, formation of mPTP, altered mitochondrial biogene-
sis, and dynamics. Binding of A𝛽to mitochondrial pro-
teins (CypD and ABAD) amplies A𝛽induced eects on
mitochondria and neuron functions (Figure ). From these
insights, it is easy to deduce how mitochondrion oers
multiple points to develop strategies against mitochondrial
dysfunction. us, not only antioxidant targeted therapeutic
strategies but also specic mitochondrial targeted therapeutic
strategies should be explored for neuroprotection against A𝛽
toxicity.
Conflict of Interests
e authors declare that there is no conict of interests
regarding the publication of this paper.
Authors’ Contribution
PasqualePiconeandDomenicoNuzzocontributedequallyto
this paper.
Acknowledgments
e authors deeply thank Dr. Daniela Giacomazza for the
critical revision of the paper. is work was supported by
the Italian Ministry of Economy and Finance with the “PNR-
CNR Aging Program –” Project.
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