Báo cáo khoa học: Post-ischemic brain damage: targeting PARP-1 within the ischemic neurovascular units as a realistic avenue to stroke treatment pptx

MINIREVIEW
Post-ischemic brain damage: targeting PARP-1 within the
ischemic neurovascular units as a realistic avenue to
stroke treatment
Flavio Moroni and Alberto Chiarugi
Department of Preclinical and Clinical Pharmacology, University of Florence, Italy
Therapeutic strategies aimed at reducing brain dam-
age after ischemic stroke have been a major focus of
academic and industrial research for the past
30 years. Two primary therapeutic approaches have
been intensively studied: the first can be defined as
the ‘vascular approach’ and its main goal is the rapid
re-opening of occluded blood vessels so that oxygen
and nutrients may return to the ischemic region. The
second may be defined as the ‘cellular approach’ and
is based on the possibility of interfering with the sig-
naling pathways, leading to loss of neurons and dam-
age of other cellular elements present in the affected
brain region [1,2]. Efforts directed at developing effec-
tive vascular therapy led to clinically useful
procedures and have clearly demonstrated that it is
possible to reduce, selectively, brain damage and
neurologic disability by administering recombinant tis-
sue plasminogen activator within 3 h from when the
stroke symptoms first start. Conversely, the cellular
approach has been so far clinically unsuccessful, and
none of the numerous neuroprotective strategies that
have been tested in clinical trials have reached the
clinical arena [3,4].
Exciting, radical, suicidal and
inflamed – the many pathways of

ischemic brain injury
The enormous body of information on ischemic neuro-
degeneration in different experimental stroke models
has shed light on the complex signaling pathways and
molecular events responsible for neuronal damage
Keywords
blood brain barrier; endothelium;
inflammation; ischemia; microglia;
neuroprotection; neurovascular unit;
PARP-1; pericytes; stroke
Correspondence
F. Moroni, Dipartimento di Farmacologia,
Viale Pieraccini 6, 50139 Firenze, Italy
Fax: +39 055 4271226
Tel: +39 055 4271280
E-mail: flavio.moroni@unifi.it
(Received 3 July 2008, revised 11
September 2008, accepted 14 October
2008)
doi:10.1111/j.1742-4658.2008.06768.x
Stroke is the third leading cause of death in industrialized countries but
efficacious stroke treatment is still an unmet need. Preclinical research indi-
cates that different molecules afford protection from ischemic neurodegen-
eration, but all clinical trials conducted so far have inexorably failed.
Critical re-evaluation of experimental data shows that all the components
of the neurovascular unit, such as neurons, glia, endothelia and basal mem-
branes, must be protected during the ischemic insult to obtain substantial
and long-lasting neuroprotection. Here, we propose the nuclear enzyme
poly(ADP-ribose) polymerase (PARP-1) as a key effector of cell death in
the various elements of the neurovascular units, and assert that drugs

inhibiting PARP-1 may therefore represent valuable tools for pharmacolog-
ical treatment of stroke patients.
Abbreviations
AIF, apoptosis-inducing factor; BBB, blood–brain barrier; HMGB1, high-mobility-group protein box 1; IL, interleukin; MMP, matrix
metalloproteinase; NMDA, N-methyl-
D-aspartate; PARG, poly(ADP-ribose) glycohydrolase; PARP, poly(ADP-ribose) polymerase; PARP-1,
poly(ADP-ribose) polymerase 1; ROS, reactive oxygen species; TNF-a, tumor necrosis factor-a.
36 FEBS Journal 276 (2009) 36–45 ª 2008 The Authors Journal compilation ª 2008 FEBS
when blood flow to a brain region drops below a criti-
cal threshold and when it returns because of vessel
re-opening and tissue reperfusion. In the past, particu-
lar attention was directed to derangement of excitatory
amino acid-mediated neurotransmission that became,
for years, the main target for neuroprotection.
Hypoxia ⁄ ischemia increases the concentrations of
extracellular glutamate [5,6] with excessive stimulation
of ionotropic and metabotropic glutamate receptors,
which initiates a chain of events leading to excitotoxic
neuronal death [7,8]. This concept is strongly sup-
ported by the observation that, in a number of in vitro
and in vivo experimental models of ischemia, glutamate
receptor antagonists, acting either on ionotropic
[N-methyl-d-aspartate (NMDA) or Gk alpha-amino-3-
hydroxy-5-methyl-4-isoxazolone propinate] or on group
I metabotropic receptors, are effective neuroprotective
agents [9–13]. Unfortunately, however, none of the
glutamate receptor antagonists tested in clinical trials
showed positive results or had an acceptable benefit ⁄
side effects ratio.
Triggered by the excitotoxic events as well as by

impairment of mitochondrial respiration, a burst of
reactive oxygen species (ROS) and reactive nitrogen
species typically occurs within the ischemic brain tis-
sue. Again, inhibition of radical formation as well as
of radical scavengers provides significant neuroprotec-
tion in animal stroke models. Agents acting as free-
radical scavengers therefore have been repeatedly
proposed as useful drugs for stroke therapy, but most
were rapidly discarded because of cardiovascular toxic-
ity. Recently, however, the spin-trap nitrone NXY-059
from AstraZeneca reached the clinical arena with some
success [14]. The putative neuroprotectant is probably
n-t-butyl hydroxylamine and ⁄ or its parent spin-trap
2-methyl-2-nitrosopropane, produced by hydrolysis of
NXY-059. Unfortunately, the positive outcome of the
first clinical trial was not confirmed in a second clinical
trial, and NXY-059 development was dropped, leaving
widespread scepticism in the field regarding the possi-
bility of obtaining ischemic neuroprotection in humans
[15].
Apoptotic mechanisms also contribute to ischemic
neuronal demise. This suicidal form of neurodegenera-
tion seems to occur mainly in specific types of brain
ischemia, including the global type of brain ischemia.
Also, activation of the apoptotic program typically
occurs in a delayed manner in brain regions present in
the surroundings of the ischemic core (the so-called
‘penumbra’, see below) and is thought to be a key com-
ponent of time-dependent brain infarct evolution [1].
Yet, strategies aimed at inhibiting the several apoptotic

effectors have not been exploited at the clinical level.
Another event widely recognized to be of key patho-
genetic relevance to post-ischemic brain damage is
immune activation of resident glial cells and leukocytes
infiltrating from blood vessels [16,17]. In this regard,
several therapeutic approaches aimed at counteracting
post-ischemic immune activation and infiltration have
been tested in clinical trials. Some, such as the anti-leu-
kocyte adhesion molecules enlimonab and HU23F2G,
proved inefficacious and harmful, respectively. Others,
such as the interleukin (IL)-1 receptor antagonist, pro-
vided inconclusive results. Failure might be a result of
the fact that both protective as well as detrimental
effects of the inflammatory response during ischemic
neurodegeneration have been reported [18].
Critical re-evaluation of drug
development in stroke
Preclinical studies clearly show that it is feasible to
protect the brain from ischemic injury by means of
pharmacological or genetic approaches aimed at tar-
geting the molecular mechanisms involved in ischemic
neurodegeneration. Hence, because there are no appar-
ent reasons why these strategies should not be effective
in humans, it is reasonable to predict that effective
neuroprotective strategies identified at the preclinical
level also reach clinical practice. Then, the question is
why has this not yet happened? An increasing body of
literature is accumulating on this subject, and several
critical points that have been identified are the past
and, unfortunately, present criteria and methodologies

used for drug development in the stroke field [3,4,19].
To summarize, it is now clear that animal models
should closely reproduce the complex cardiovascular
and cerebral pathophysiology of stroke patients, and
neuroprotection should be evaluated on a long-lasting
and functional basis, rather than on an acute and his-
tological basis. Also, careful and rigorous selection of
patients with salvageable tissue [evidenced using mag-
netic resonance imaging as the presence of an area of
hypoperfusion larger than that of altered water diffu-
sion (the latter is an index of necrosis), the so-called
‘Perfusion ⁄ Diffusion (PWI ⁄ DWI) mismatch’] should
be conducted before treating them with an anti-stroke
drug candidate [4]. Finally, the concepts of ‘pleiotypic
drugs’ (i.e. drugs with several mechanisms of action)
and ‘synergistic combinatorial drug therapy’ emerge as
key requisites for efficacious stroke treatment [4].
Indeed, one of the possible reasons for the lack of clin-
ical efficacy of drugs tested in clinical trials for brain
ischemia is their selective mechanism of action. For
instance, glutamate antagonists act exclusively (or pre-
dominantly) on neurons. So, even if neurons are the
F. Moroni and A. Chiarugi PARP-1 and the ischemic neurovascular unit
FEBS Journal 276 (2009) 36–45 ª 2008 The Authors Journal compilation ª 2008 FEBS 37
first cell type to lose their function when blood supply
is insufficient, the other cell types present in the ner-
vous tissue are of the utmost importance to support
neuronal functioning. When capillaries and glia are
damaged, neurons cannot survive in spite of protection
from excitotoxic insults. Similarly, selective blocking of

apoptosis or inflammation within the ischemic tissue
cannot provide protection when the other detrimental
events are unrestricted. As a whole, efficacious stroke
treatment needs concomitant targeting of the various
pathogenetic events actively contributing to neurode-
generation in cells localized within the ischemic
penumbra.
Penumbra and the neurovascular unit
The ischemic brain region may be divided into a zone
in which blood flow is completely absent (‘ischemic
core’) and a peripheral zone in which collateral ves-
sels supply only a fraction of the oxygen and glucose
required for the normal activity of neural cells (‘ische-
mic penumbra’) [2,20]. While all cell types in the core
region undergo typical necrotic features and die form-
ing an infarct zone, the ischemic penumbra may
initially retain its morphological integrity, even if its
functions (i.e. electrical activity, synthetic processes,
bioenergetic functions, etc.) are temporally lost. How-
ever, if sufficient blood flow eventually returns to the
ischemic region within a reasonable time (hours) it is
possible to rescue this area, thus limiting the neuro-
logical damage. It is now clear that in order to
obtain full functional recovery, not only neurons, but
all cell types (i.e. astrocytes, microglia, oligodendro-
cytes, endothelial cells, muscle cells, pericytes) and
structures (mainly basal membranes) present in the
‘penumbra area’ should be rescued [21,22]. Thus,
ischemic neuroprotection can be achieved only if the
classic, oversimplified strategy ‘save the neurons’ is

changed into ‘save neural and stromal cells’. Overall,
neural and stromal cells are grouped into a functional
entity: the so-called ‘neurovascular unit’. Operatively,
the latter is a very complex network of functions
brought about by different cells and aimed at main-
taining the homeostatic milieu necessary for normal
brain activities. Protection of the components of the
neurovascular unit seems therefore essential to reduce
brain damage and neurological deficits after a stroke.
To achieve this, different strategies have been pro-
posed and evaluated in preclinical settings. Yet, con-
comitant targeting of all the components of the
neurovascular units adds substantial complexity to
the feasibility of obtaining ischemic neuroprotection
by pharmacological approaches and, as mentioned
above, general scepticism permeated the field. As
outlined below, we claim that poly(ADP-ribose) poly-
merase 1 (PARP-1) inhibitors are among the most
efficacious protectants of the neurovascular unit
currently available.
PARP-1 activation and cell death in the
neurovascular unit
Poly(ADP-ribose) polymerases (PARPs) are NAD-
dependent enzymes that are able to catalyse the trans-
fer of ADP-ribose units from NAD to substrate
proteins, thereby contributing to the control of geno-
mic integrity, cell cycle and gene expression [23].
Among PARPs, nuclear PARP-1 is a DNA damage-
activated enzyme of 113 kDa molecular mass and is
the most abundant and commonly studied member of

the family. Its enzymatic activity leads to poly(ADP
ribose) formation, and it was first described over
40 years ago in liver cell nuclei incubated with NAD
and ATP in Paul Mandel’s laboratory in Strasburg
[24]. Although this seminal observation was made in a
neuroscience laboratory, for the following 30 years,
research on PARP-1 was exclusively carried out by
researchers mainly involved in genome stability, DNA
repair and cancer. The neuroscience community
ignored PARP-1 until the early 1990s when it was
shown that it mediates glutamate-induced and nitric
oxide-induced neuronal death [25,26]. Excellent work
carried out in the following years uncovered several
molecular events causally linking PARP-1 activation to
ischemic cell death [27]. As for the triggers of PARP-1
hyperactivity during ischemia, ROS-dependent DNA
damage is thought to play a major role. However,
Ca
2+
-dependent and kinase-dependent PARP-1 activa-
tion might also contribute [28–30]. Ambiguity also
exists regarding the molecular mechanisms underlying
the detrimental role of the enzyme in ischemic brain
injury [31,32]. Indeed, although we know in part the
mechanisms activated by PARP-1 and triggering
neurotoxicity, which of these is causally involved in
PARP-1-dependent ischemic neurodegeneration still
needs to be elucidated.
Experimental data demonstrate that, upon different
stresses, activation of PARP-1 can exert detrimental

effects in every cell type of the neurovascular unit
(Fig. 1). Given that the ischemic challenge mimics
these stresses, we reason that during brain ischemia
PARP-1-dependent cytotoxicity occurs in all the com-
ponents of the neurovascular unit. It is obvious that
triggers, time courses and final effects of PARP-1
activation in endothelial, muscle and glial cells, as well
as in infiltrating leukocytes, are different from those
PARP-1 and the ischemic neurovascular unit F. Moroni and A. Chiarugi
38 FEBS Journal 276 (2009) 36–45 ª 2008 The Authors Journal compilation ª 2008 FEBS
occurring in neurons. Regardless, the hyperactivation
of PARP-1 in each single component of the neurovas-
cular unit triggers dysfunction ⁄ cytotoxicity and, indi-
rectly, severely affects the functioning of neighbouring
neurons. As a whole, PARP-1-dependent derangement
of the integrity of the neurovascular unit is caused by
the enzyme’s ability to prompt an increase of blood–
brain barrier (BBB) permeability, the release of pro-
inflammatory mediators, mitochondrial dysfunction
and bioenergetic failure, as well as the activation of
specific apoptotic pathways.
PARP-1, endothelia and post-ischemic
BBB breakdown
Ischemia causes rapid structural changes and break-
down of the BBB, allowing plasma exudation and
immune cell infiltration, which contribute to ischemic
brain damage [22]. Very early after the onset of brain
ischemia, and especially after a reperfusion period,
abundant free radicals are generated in macrophages,
endothelial cells, perycites, astrocytes, microglia and

neurons, causing significant damage to brain capillaries
and disruption of the BBB [33]. Free radicals formed
both inside and outside the vessels prompt genotoxic
stress and activate PARP-1 in endothelial cells. Under
conditions of chronic hypoxia, PARP-1 activation
within endothelia triggers cell proliferation and slowly
developing brain damage. The molecular mechanisms
of cell proliferation include the generation and release
of ROS from NADPH oxidase and mitochondria, sus-
tained increase of the cytosolic Ca
2+
concentration
and finally nuclear translocation of mitogen-activated
protein kinase kinase ⁄ extracellular regulated protein
kinase with cell cycle activation [34]. Conversely,
during ischemia, PARP-1 hyperactivation causes endo-
thelial cell death. The latter occurs because of cellular
accumulation of the PARP-1 product poly(ADP-
ribose), which causes translocation of apoptosis-induc-
ing factor (AIF) from mitochondria to the nucleus and
activation of a caspase-independent programmed cell-
death pathway [35–37]. Accordingly, the potent
PARP-1 inhibitor, PJ34, administered to rats with
transient focal brain ischemia, preserves the integrity
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Fig. 1. The role of PARP-1 within the ischemic neurovascular units. PARP-1 exerts its detrimental role within the ischemic neurovascular
unit by promoting necrosis and AIF-dependent apoptosis in neurons, astrocytes and endothelial cells. PARP-1 also plays a key role in
immune activation and migration of microglial cells upon different noxious stimuli to the central nervous system. The expression of adhesion
molecules by endothelial cells is also promoted by PARP-1-dependent transcriptional activation, thereby promoting leukocyte recruitment
within the ischemic brain tissue and their detrimental effects on ischemic injury. Hence, the pharmacological inhibition of the enzyme exerts

ischemic neuroprotection by targeting several pleiotypic events of pathogenetic relevance to post-ischemic brain damage. X, adhesion mole-
cules. ADP-ribose monomers are depicted as black circles binding to the transient receptor potential melastatin-2 receptor.
F. Moroni and A. Chiarugi PARP-1 and the ischemic neurovascular unit
FEBS Journal 276 (2009) 36–45 ª 2008 The Authors Journal compilation ª 2008 FEBS 39
of endothelial tight junctions and decreases the expres-
sion of the adhesion molecule intercellular adhesion
molecule-1, thus limiting leukocyte infiltration and the
subsequent inflammatory damage to the ischemic brain
[35,38]. It has also been proposed that post-ischemic
PARP-1 activation contributes to increased expression
of matrix metalloproteinases (MMPs), a group of zinc-
containing proteases with key roles in matrix degrada-
tion and disruption of capillary permeability during
stoke [39]. Indeed, pharmacological PARP-1 inhibition
reduces MMP-9 expression levels in plasma and brain
[40], prevents brain matrix degradation, reduces
delayed increase of BBB permeability and edema for-
mation, preserves endothelial tight junction proteins
and decreases delayed infiltration of leukocytes into
the brain of rats with middle cerebral artery occlusion
[41]. The key role of PARP-1 hyperactivation in endo-
thelial dysfunction in experimental models of diabetes
underscores the pathogenetic relevance of the enzyme
to disorders of this key component of the neurovascu-
lar unit [42]. Accordingly, gene array studies have
demonstrated that upregulation of inflammatory genes
is hampered in PARP-1
) ⁄ )
endothelial cells exposed to
tumor necrosis factor-alfa (TNF-a) [43]. Taken

together, these findings point to basal PARP-1 activity
as central to homeostatic regulation of endothelial
function, whereas its hyperactivation appears causal
to BBB damage and immune cell infiltration during
ischemia.
PARP-1, glia and post-ischemic
inflammatory events
Activation of resident immune cells as well as infiltra-
tion of leukocytes within the ischemic area lead to
excessive release of inflammatory mediators and ensu-
ing worsening of brain damage. In keeping with this,
astrocytes, microglia and blood-derived leukocytes
contribute to ischemic neurodegeneration, whereas
immunosuppressant strategies able to reduce the
inflammatory response decrease infarct volumes in dif-
ferent stroke models [16,17]. Microglial cells are resi-
dent brain macrophages displaying a ‘resting’ highly
ramified phenotype. Upon ischemic challenge, before
neuronal damage can be morphologically detected [44],
microglia assume amoeboid morphology and acquire
phagocytic activity, producing ROS and other inflam-
matory ⁄ cytotoxic factors such as nitric oxide, prosta-
noids, TNF-a, IL-1b and MMPs. Astrocytes and
infiltrating leukocytes within the ischemic brain tissue
also contribute to the synthesis and release of pro-
inflammatory mediators [17]. It is now widely accepted
that the latter are responsible for disruption of the
capillary basal lamina, opening of the BBB and infil-
tration of blood-borne leukocytes. This prompts a
vicious circle comprising waves of release of cytotoxic

inflammatory products, cell death and recruit-
ment ⁄ activation of blood or bystander immune cells.
Eventually, the neuroimmune response causes collapse
of the structures and functions of the neurovascular
unit [16,17,45].
Again, PARP-1 plays a key role in this scenario.
Indeed, numerous reports demonstrate that PARP-1
activity promotes the neuroimmune response thanks
to its ability to assist transcriptional activation and
epigenetic remodeling in immune cells. In this light, it
has been speculated that ischemic neuroprotection
afforded by PARP inhibitors is at least partially med-
iated by their anti-inflammatory properties [46].
Indeed, PARP inhibitors decrease expression of
inflammatory markers ⁄ mediators such as CD11b,
cyclooxygenase-2, inducible nitric oxide synthase,
TNF-a, IL-1b, IL-6, intracellular adhesion molecule-1,
interferon-gamma and E-selectin in different models
of neurodegeneration [40,47–55]. Remarkably, these
molecules actively contribute to ischemic neurodegen-
eration. A key role for PARP-1 in microglia activa-
tion and migration towards injured neurons has also
been reported [56]. Reduced expression of pro-inflam-
matory mediators is probably a result of the fact that
inflammatory transcription factors such as nuclear
factor-kappaB, activator protein-1 and nuclear factor
of activated T-cells are positively regulated by
PARP-1. PARP-1 protein per se , as well as its enzy-
matic activity, promote transcription factor binding
to DNA as well as supramolecular complex formation

containing several transcription-regulating proteins
and RNA polymerase II [23,53,57]. These findings
taken together may explain why post-treatment with
PARP-1 inhibitors reduces the neuroimmune response
in different stroke models [58–60].
Recently, the tetracycline, minocycline, has been
proposed as a clinically relevant tool to limit post-
ischemic brain damage because of its ability to inhibit
microglia activation. Minocycline is indeed able to
reduce brain infarct volumes in preclinical models
[61], as well as neurological impairment in stroke
patients [62]. Interestingly, it has recently been
reported that minocycline is a powerful inhibitor of
PARP-1 [63]. Whether PARP-1 inhibition underpins
the drug’s neuroprotective effects in stroke patients is
currently unknown. Yet, given that minocycline has
been largely used without significant side effects, these
observations indicate that acute inhibition of PARP-1
in vivo might be a rather safe procedure and could be
proposed to preserve the integrity of the ischemic
PARP-1 and the ischemic neurovascular unit F. Moroni and A. Chiarugi
40 FEBS Journal 276 (2009) 36–45 ª 2008 The Authors Journal compilation ª 2008 FEBS
neurovascular unit and limit post-ischemic brain
damage in humans.
PARP-1 and post-ischemic death in
neurons
Excitotoxicity and PARP-1 activation have been caus-
ally linked since 1994 when it was reported that gluta-
mate increases poly(ADP-ribose) synthesis and causes
a type of cell death that is prevented by both NMDA

antagonists and PARP-1 inhibitors [25,26]. The pro-
posed molecular events underlying these observations
include: overactivation of NMDA glutamate receptors
with consequent intracellular Ca
2+
influx; and subse-
quent ROS production mainly caused by neuronal
nitric oxide synthase activity, which, in turn, triggers
DNA damage-dependent hyperactivation of PARP-1,
depletion of intracellular NAD and ATP stores, and
neuronal death [26]. PARP-1 activation may also occur
in neurons without NMDA receptor activation, as
increases of intracellular [Ca
2+
] triggered by K
+
-
induced depolarization or inositol 3-phosphate-recep-
tor activation are sufficient to trigger poly(ADP-ribose)
formation [28,64]. In keeping with this toxic cascade of
events, neurons obtained from PARP-1-deficient mice
are resistant to NMDA toxicity and to oxygen and
glucose deprivation [65]. It was also shown that
NMDA-induced overload of cytosolic Ca
2+
not only
activates neuronal nitric oxide synthase in the cytosol,
but is also responsible for mitochondrial ROS produc-
tion [66], which contributes to DNA damage and fur-
ther activation of PARP-1 [67,68]. Substantial DNA

damage, evaluated by means of the comet assay, is
present in cells isolated from the rat ischemic cortex or
caudate. NMDA receptor antagonists reduce the
extent of the damage and provide ischemic neuropro-
tection, while PARP inhibitors decrease infarct vol-
umes without affecting the severity of DNA damage
[69]. These observations suggest that NMDA receptor
channel openings, ROS formation, DNA damage and
PARP activation are sequential crucial steps in the
process leading to neuronal death. They also indicate
that stroke protection can be achieved without reduc-
ing DNA damage. Energy failure following PARP-1
activation is not only caused by NAD resynthesis but
also by glycolysis block because of NAD depletion,
which results in reduced synthesis of both glycolysis-
derived ATP and mitochondrial energetic substrates
[70]. Accordingly, tricarboxylic acid cycle substrates or
extracellular NAD supplementation protect neurons
from excessive PARP-1 activation [71], whereas
PARP-1 inhibitors prevent ischemia-induced NAD
+
depletion and reduce ischemic brain injury [72]. In
apparent contrast to the hypothesis that PARP-1
worsens ischemic neurodegeneration by reducing ATP
levels within the injured tissue, however, ischemia-
induced energy derangement is similar in the affected
brain areas of PARP-1
+ ⁄ +
and PARP-1
) ⁄ )

mice,
despite the latter showing significant reduction of
ischemic volumes [73].
Controversy still exists on the molecular mecha-
nisms involved in PARP-1-dependent neuronal death
during ischemia. In this regard it has been very
recently reported that exposure of cultured neurons
to poly (ADP-ribose) is sufficient to trigger nuclear
translocation of mitochondrial AIF and cell demise
[74]. The poly(ADP-ribose)-degrading enzyme, poly
(ADP-ribose) glycohydrolase (PARG), should be, in
principle, a neuroprotective agent [75]. Consistently,
PARG-110 kDa
) ⁄ )
or PARG
+ ⁄ )
mice show increased
sensitivity to brain ischemia [36,76]. Also, PARP-1
activity seems to be essential for AIF release within
neurons of the infarct area, and AIF-deficient (Harle-
quin) mice are less sensitive to post-ischemic brain
damage [77]. Data therefore point to PARP-1 activ-
ity-dependent AIF release from mitochondria as a
key molecular event underlying ischemic neuronal
death. Interestingly, the ADP-ribose monomers origi-
nating from the polymer degradation through PARG
might also contribute to neuronal demise by activat-
ing transient receptor potential melastatin-2 receptors
and massive Ca
2+

influx [78,79]. Finally, the finding
that, when released in the extracellular space, high-
mobility-group protein box 1 (HMGB1) promotes the
neuroinflammatory response and worsens brain ische-
mia [80–82], along with evidence that PARP-1 pro-
motes HMGB1 release [83] (but also see [82]),
indicate that HMGB1 may mediate, in part, the toxic
effect of PARP-1 hyperactivation within the ischemic
brain tissue. Overall, a wealth of evidence points to
the synthesis of poly (ADP-ribose) within ischemic
neurons as a crucial event contributing to derange-
ment of the neurovascular unit.
Conclusion
To reduce brain damage after stroke it is not sufficient
to protect neurons from excitotoxic insults, but it is
mandatory to rescue all cellular and structural compo-
nents of the neurovascular unit. As outlined above,
PARP-1 activation during brain ischemia plays a detri-
mental role in all cell types of the neurovascular unit.
Inhibitors of PARP-1 might therefore represent a class
of ‘pleiotypic drugs’, which are considered the most
promising tools for pharmacological treatment of
stroke. Also, the different temporal kinetics of PARP-1
F. Moroni and A. Chiarugi PARP-1 and the ischemic neurovascular unit
FEBS Journal 276 (2009) 36–45 ª 2008 The Authors Journal compilation ª 2008 FEBS 41
activation within the components of the neurovascular
unit would warrant a significant ‘window of opportu-
nity’ to be harnessed for the treatment of stroke
patients. Remarkably, the clinical relevance of PARP-1
inhibitors in stroke treatment is emphasized by the fact

that these drugs are well tolerated by patients enrolled
in clinical trials for treatment of tumor malignancies
or coronary bypass, and that, theoretically, anti-stroke
treatment with PARP-1 inhibitors would require an
acute, 4–6-day treatment. This, of course, would
reduce the risk of side effects. The latter might be fur-
ther reduced by the forthcoming development of
PARP isoform-specific inhibitors [84]. In conclusion,
preclinical and clinical data indicate that PARP-1 is a
very promising target for ischemic neuroprotection,
and PARP-1 inhibitors represent a realistic new avenue
to stroke treatment.
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