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Please use this identifier to cite or link to this item:
http://hdl.handle.net/2108/976
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Full metadata record
| DC Field | Value | Language |
|---|
| contributor.advisor | Calissano, Pietro | - |
| contributor.author | Amadoro, Giuseppina | - |
| date.accessioned | 2009-08-03T12:09:56Z | - |
| date.available | 2009-08-03T12:09:56Z | - |
| date.issued | 2009-08-03T12:09:56Z | - |
| identifier.uri | http://hdl.handle.net/2108/976 | - |
| description | 19. ciclo | en |
| description.abstract | E’ stato ipotizzato che l’alterata funzione e/o struttura della proteina tau, una MAP (microtubuleassociated-
protein) particolarmente abbondante nei neuroni, causa la morte di specifiche
popolazioni cellulari in numerosi disordini neurodegenerativi, quali ad esempio le tauopatie e il
morbo di Alzheimer. In particolare, uno squilibrio del rapporto intracellulare delle differenti
isoforme è sufficiente di per sé ad indurre una pronunciata neurodegenerazione, suggerendo che la
precisa stechiometria di tutte le 6 varianti alternative è necessaria per garantire il corretto
adempimento delle sue funzioni fisiologiche, che implicano non esclusivamente l’interazione e la
stabilizzazione dei microtubuli citoscheletrici. Comunque, il meccanismo molecolare attraverso cui
una disfunzione della proteina tau è causa della morte neuronale disordine-associata rimane ancora
da chiarire. A tal fine, noi abbiamo analizzato l’effetto dell’espressione dose-dipendente della tau
umana (htau/h40) e di alcuni dei suoi frammenti N-terminali -privi del dominio di legame ai
microtubuli- in diverse colture primarie neuronali, transdotte ad alta efficienza mediante infezione
adenovirale.
In tale lavoro, noi riportiamo che bassi livelli d’ espressione del dominio di proiezione in membrana
di tau, o tau(1-230), come dell’isoforma più lunga, o tau(1-441), inibiscono moderatamente l’inizio
dell’apoptosi nei granuli del cervelletto, deprivati del siero e del potassio extracellulare. L’effetto
antiapoptotico implica una modulazione positiva della fosforilazione di Akt1/2, una serin-treonin
chinasi che criticamente controlla la cascata delle caspasi effettrici.
Di converso, noi dimostriamo che l’espressione di più alti livelli intracellulari della isoforma più
lunga di htau, e di alcuni dei suoi frammenti N-terminali, in colture neuronali di cervelletto,
ippocampo e corteccia evoca un potente effetto neurotossico NMDAR (N-methyl-D-aspartate
receptor)-mediato e caspasi-indipendente. Al fine di elucidare una possibile via biochimica di
trasduzione del segnale, noi riportiamo che la morte tau-indotta è associata ad un’ eccessiva
stimolazione dei recettori extrasinaptici, in quanto (1) largamente inibita da ifenprodil, un
antagonista altamente selettivo dei recettori NMDA NR2B-arricchiti ed (2) accompagnata dalla
marcata defosforilazione del fattore trascrizionale CREB (cAMP-response-element-bindingprotein),
responsabile dell’espressione di importanti geni di sopravvivenza neuronale. L’attivazione
patologica dell’NMDAR causa una sostenuta, protratta e tardiva fosforilazione di ERK1/2
(extracellular-regulated-kinase 1/2), una MAPK (mitogen-activated-protein kinase) la cui
inibizione significativamente previene la morte tau-mediata. Infine, la stimolazione dell’NMDAR
induce la deleteria attivazione della calpaina-I che, a sua volta, degrada la proteina tau non solo nel
peptide di 17kDa, ma anche in altri peptidi N-terminali più fortemente dannosi. Sulla base dei nostri
esperimenti in vitro, noi ipotizziamo pertanto che il taglio inappropriato della proteina tau all’Nterminale
genera numerosi peptidi tossici i quali, innescando un loop a feedback negativo,
ulteriormente amplificano e propagano il processo di morte. Infatti, l’inibizione della calpaina I
blocca completamente la proteolisi di tau, e quindi la morte neuronale.
I nostri dati dunque rivelano un meccanismo patogenetico che accoppia la neurotossicità taumediata
all’aberrante attivazione dell’NMDAR. Tali osservazioni pertanto non solo risultano
rilevanti in numerosi disordini neurodegenerativi, quali le tauopatie e il morbo di Alzheimer’s
associati ad una marcata eccitotossicità da glutammato, ma provvedono anche un ulteriore supporto
sperimentale all’effetto neuroprotettivo FDA (Food and drug administration)-approvato della
memantina, un antagonista non competitivo del recettore NMDA utilizzato con successo nei trials
clinici sull’uomo, al fine di migliorare le funzioni cognitive di pazienti affetti da tali malattie
dementignene.
2
ABSTRACT
The altered function and/or structure of MAP (microtubule-associated- protein) tau is postulated to
cause cell death in tauopathies and Alzheimer's disease. Any disturbance in intracellular tau ratio
are sufficient to induce neurodegeneration, suggesting that a precise stoichiometry of all 6 isoforms
is necessary to correctly discharge more of their functions, which extend beyond interaction with
microtubules. However, the mechanisms by which tau induces neuronal death remain unclear.
Therefore, we have analyzed the effect of dose-dependent expression of human tau (htau) and of
some of its N-terminal fragments -free of microtubule-binding domain- in primary neuronal cultures
(cerebellum, hippocampus, cortex), by adenovirally-mediated infection.
Here we report that moderate expression levels of the tau (1-230) fragment, as well as of full length
htau, inhibits the onset of apoptosis in serum and potassium-deprived cerebellar granule neurons,
probably acting at the level of Akt1/2-mediated activation of the caspase cascade.
On the contrary, we show that overexpression of more high levels of the longest isoform htau and of
some of its N-terminal fragments in primary neuronal cultures leads to an N-methyl-D-aspartate
receptor (NMDAR)-mediated and caspase-independent cell death. Death signaling likely originates
from stimulation of extrasynaptic NR2B-subunit-containing NMDARs because it is accompanied
by dephosphorylation of cAMP-response-element-binding protein (CREB) and it is inhibited by
ifenprodil. Interestingly, activation of NMDAR leads to a crucial, sustained, and delayed
phosphorylation of extracellular-regulated kinases 1 and 2 (ERK1/2), whose inhibition largely
prevents tau-induced neuronal death. Moreover, NMDAR involvement causes the fatal activation of
calpain I, which, in turn, degrades tau protein into a 17-kDa peptide and possibly other highly toxic
N-terminal peptides. Some of these peptides are hypothesized, on the basis of our in vitro
experiments, to initiate a negative loop, ultimately leading to cell death. Thus, inhibition of calpain I
largely prevents tau degradation and cell death.
Our findings unravel a cellular mechanism linking tau toxicity to NMDAR activation and might be
relevant to Alzheimer's disease and tauopathies, where NMDAR-mediated toxicity is postulated to
play a pivotal role. Finally, these studies provide strong experimental evidences about the
therapeutical efficacy in humans of memantine, an well-tolerated uncompetitive antagonist of
NMDAR, shown to be benefical in the treatment of neurological disorders mediated by
excitotoxicity. | en |
| description.abstract | The altered function and/or structure of MAP (microtubule-associated- protein) tau is postulated to
cause cell death in tauopathies and Alzheimer's disease. Any disturbance in intracellular tau ratio
are sufficient to induce neurodegeneration, suggesting that a precise stoichiometry of all 6 isoforms
is necessary to correctly discharge more of their functions, which extend beyond interaction with
microtubules. However, the mechanisms by which tau induces neuronal death remain unclear.
Therefore, we have analyzed the effect of dose-dependent expression of human tau (htau) and of
some of its N-terminal fragments -free of microtubule-binding domain- in primary neuronal cultures
(cerebellum, hippocampus, cortex), by adenovirally-mediated infection.
Here we report that moderate expression levels of the tau (1-230) fragment, as well as of full length
htau, inhibits the onset of apoptosis in serum and potassium-deprived cerebellar granule neurons,
probably acting at the level of Akt1/2-mediated activation of the caspase cascade.
On the contrary, we show that overexpression of more high levels of the longest isoform htau and of
some of its N-terminal fragments in primary neuronal cultures leads to an N-methyl-D-aspartate
receptor (NMDAR)-mediated and caspase-independent cell death. Death signaling likely originates
from stimulation of extrasynaptic NR2B-subunit-containing NMDARs because it is accompanied
by dephosphorylation of cAMP-response-element-binding protein (CREB) and it is inhibited by
ifenprodil. Interestingly, activation of NMDAR leads to a crucial, sustained, and delayed
phosphorylation of extracellular-regulated kinases 1 and 2 (ERK1/2), whose inhibition largely
prevents tau-induced neuronal death. Moreover, NMDAR involvement causes the fatal activation of
calpain I, which, in turn, degrades tau protein into a 17-kDa peptide and possibly other highly toxic
N-terminal peptides. Some of these peptides are hypothesized, on the basis of our in vitro
experiments, to initiate a negative loop, ultimately leading to cell death. Thus, inhibition of calpain I
largely prevents tau degradation and cell death.
Our findings unravel a cellular mechanism linking tau toxicity to NMDAR activation and might be
relevant to Alzheimer's disease and tauopathies, where NMDAR-mediated toxicity is postulated to
play a pivotal role. Finally, these studies provide strong experimental evidences about the
therapeutical efficacy in humans of memantine, an well-tolerated uncompetitive antagonist of
NMDAR, shown to be benefical in the treatment of neurological disorders mediated by | en |
| format.extent | 10391185 bytes | - |
| format.mimetype | application/pdf | - |
| language.iso | it | en |
| subject.classification | MED/26 Neurologia | en |
| title | Ruolo della proteina tau nella sopravvivenza neuronale | en |
| type | Doctoral thesis | en |
| degree.name | Neuroscienze | en |
| degree.level | Dottorato | en |
| degree.discipline | Facoltà di medicina e chirurgia | en |
| degree.grantor | Università degli studi di Roma Tor Vergata | en |
| date.dateofdefense | A.A. 2005/2006 | en |
| Appears in Collections: | Tesi di dottorato in medicina
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| TESI.pdf | | 10147Kb | Adobe PDF | View/Open |
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