Nicotera, Pierluigi
E-Mail-Adresse
ORCID
Geburtsdatum
Berufsbeschreibung
Nachname
Vorname
Name
Suchergebnisse Publikationen
The dawning of a new age of toxicology
Die Toxikologie steht in einer Welt, in der wir Tausenden von Chemikalien und Millionen von deren Mischungen ausgesetzt sind, vor enormen Herausforderungen. Als Antwort darauf müssen radikal neue Ansätze entwickelt werden, um die Sicherheit der Bevölkerung zu gewährleisten. Ein Meilenstein in dieser Richtung ist die Vision des Nationalen Forschungsrates der USA Toxizitätstestung im 21. Jahrhundert: Eine Vision und eine Strategie . Gegenwärtig testet eine Allianz, die zwischen dem NTP* und dem NCGC des NIH sowie dem NCCT der EPA gebildet wurde, ob diese neue Strategie realistischerweise eine Basis für künftige Entscheidungen zum Schutz der öffentlichen Gesundheit sein kann. Die Vision setzt einen Paradigmenwechsel im Ansatz von Sicherheitsevaluationen voraus und stellt die traditionell angewandten Verfahren auf den Kopf. Wo bisher Tierexperimente die wichtigste Technologie waren, setzt die Zukunftsvision auf in vitro und in silico Ansätze, die auf menschlichem Material beruhen. Toxizitätstests beginnen heutzutage relativ blind mit einem Tierexperiment, dem dann nur manchmal mechanistische Studien folgen, während die neu vorgeschlagene Strategie die Sicherheitsevaluation von unten her beginnt: Zunächst würden mit in vitro Tests die durch Giftstoffe gestörten Stoffwechsel- und Regulationswege identifiziert; erst in einem möglichen zweiten Schritt kämen dann begrenzte und streng fokussierte Tierversuche dazu, um eventuell noch offene Wissenslücken zu schließen. Dies bedeutet nicht weniger als eine Umkrempelung der Toxikologie von einer hauptsächlich beschreibenden Tätigkeit und Hilfsdisziplin für Behörden wieder hin zu einer Naturwissenschaft mit all ihren Dimensionen. Der Hintergrund und die Konsequenzen werden hier insbesondere für Leser mit einem Interesse auch an parallelen Europäischen Trends beschrieben.
Disialoganglioside GD3 induced oligodendrocyte death and GD3 release by microglia
Differential Effects of Bcl-2 on Cell Death Triggered under ATP-Depleting Conditions
The intracellular ATP concentration decides on the onset of either apoptosis or necrosis in Jurkat cells exposed to death stimuli. Bcl-2 can block apoptotic demise, which occurs preferably under conditions of high cellular ATP levels. Here, we investigated the effects of Bcl-2 on the necrotic type of cell demise that prevails under conditions of energy loss. ATP levels were modulated by using mitochondrial inhibitors, such as rotenone or S-nitrosoglutathione, in medium either lacking glucose or supplemented with glucose to stimulate glycolytic ATP generation. Under conditions of ATP depletion, staurosporine (STS) induced >90% necrosis in vector control-transfected cells, whereas bcl-2-transfected cells were protected. Thus, the antiapoptotic protein Bcl-2 can reduce the overall amount of cell death in ATP-depleted cells regardless whether it occurs by apoptosis or necrosis. Cytochrome c release, normally preceding STS-induced necrosis, was also inhibited by Bcl-2. However, Bcl-2 did not prevent an initial STS-induced drop of the mitochondrial membrane potential (DeltaPsi(m)). Therefore, the mechanisms whereby Bcl-2 prevents cell death and favors retention of cytochrome c in the mitochondria require neither the maintenance of mitochondrial DeltaPsi nor the maintenance of normal ATP levels.
Calpain inhibitors prevent nitric oxidetriggered excitotoxic apoptosis
The pathogenesis of some neurodegenerative disorders has been linked to excitotoxicity, excess generation of nitric oxide (NO) and apoptosis. Here, we used a model of NO-triggered neuronal apoptosis that was strictly dependent on autocrine NMDA receptor (NMDA-R) activation and intracellular Ca2 increase. We investigated the efficiency and potentially beneficial effects of calpain inhibition. Three calpain inhibitors that prevented intracellular fodrin proteolysis also blocked apoptotic features such as decrease in mitochondrial membrane potential, chromatin breakdown, and subsequent death of cerebellar granule neurons exposed to NO donors (S-nitroso- L-glutathione, S-nitroso-N-acetyl D,L-penicillamine, and diethylamino- diazenolate-2-oxide). Since inhibitors did not interfere with NMDA-R activation, we suggest that block of calpains blunts NO-triggered neuronal apoptosis by stopping the cascade downstream of primary autocrine excitotoxic events.
Botulinum neurotoxin C initiates two different programs for neurite degeneration and neuronal apoptosis
Clostridial neurotoxins are bacterial endopeptidases that cleave the major SNARE proteins in peripheral motorneurons. Here, we show that disruption of synaptic architecture by botulinum neurotoxin C1 (BoNT/C) in central nervous system neurons activates distinct neurodegenerative programs in the axo-dendritic network and in the cell bodies. Neurites degenerate at an early stage by an active caspase-independent fragmentation characterized by segregation of energy competent mitochondria. Later, the cell body mitochondria release cytochrome c, which is followed by caspase activation, apoptotic nuclear condensation, loss of membrane potential, and, finally, cell swelling and lysis. Recognition and scavenging of dying processes by glia also precede the removal of apoptotic cell bodies, in line with a temporal and spatial segregation of different degenerative processes. Our results suggest that, in response to widespread synaptic damage, neurons first dismantle their connections and finally undergo apoptosis, when their spatial relationships are lost.
Cascade of Caspase Activation in Potassium-Deprived Cerebellar Granule Neurons : Targets for Treatment with Peptide and Protein Inhibitors of Apoptosis
Cerebellar granule neurons (CGN) cultured in the presence of serum and depolarizing potassium concentrations undergo apoptosis when switched to serum-free medium containing physiological potassium concentrations. Here we show that processing of the key protease, caspase-3, depends on the activation of caspase-9, but not of caspase-8. Selective peptide inhibitors of caspase-9 block processing of caspase-3 and caspase-8 and inhibit apoptosis, whereas a selective inhibitor of caspase-8 blocks neither processing of caspase-3 nor cell death. The data obtained with peptide inhibitors were confirmed by adenovirally mediated ectopic expression of the cytokine response modifier A (crmA), the baculovirus protein p35, and the X chromosome-linked inhibitor of apoptosis (XIAP). Further, caspase-8-activating death receptors do not mediate apoptosis in CGN and potassium withdrawal-induced apoptosis evolves unaltered in gld or lpr mice, which harbor mutations in the CD95/CD95 ligand system. Thus, neuronal apoptosis triggered by potassium deprivation is death receptor-independent but involves the mitochondrial pathway of caspase activation.
Disialoganglioside GD3 is released by microglia and induces oligodendrocyte apoptosis
Increased brain ganglioside levels are a hallmark of various neuroinflammatory pathologies. Here, we provide evidence that murine microglia can secrete disialoganglioside GD3 upon exposure to inflammatory stimuli. Comparison of different neural cell types revealed a particular and specific sensitivity of oligodendrocytes towards exogenous GD3. Oligodendrocyte death triggered by GD3 was preceded by degeneration of cellular processes, and associated with typical features of apoptosis, such as chromatin condensation, exposure of phosphatidylserine, release of cytochrome c from mitochondria, and loss of mitochondrial membrane potential, followed by the loss of plasma membrane integrity and detachment of disintegrated oligodendrocytes. Overexpression of bcl-2 partially protected oligodendrocytes from death. In contrast, treatment with the pan-caspase inhibitor zVAD-fmk did not prevent phosphatidylserine exposure, chromatin margination at the nuclear periphery, and death, although caspase-3 was blocked. Thus, GD3 produced by microglia under neuroinflammatory conditions may function as a novel mediator triggering mitochondria-mediated, but caspase-independent, apoptosis-like death of oligodendrocytes.
Monocular deprivation (MD)-induced apoptosis in the lateral geniculate nucleus (LGN) of new-born mice involves poly(ADP)ribose polymerase (PARP)
Apoptosis in Caspase-inhibited Neurons
Background: There is growing evidence of apoptosis in neurodegenerative disease. However, it is still unclear whether the pathological manifestations observed in slow neurodegenerative diseases are due to neuronal loss or whether they are related to independent degenerative events in the axodendritic network. It also remains elusive whether a single, caspase-based executing system involving caspases is responsible for neuronal loss by apoptosis. Materials and Methods: Long-term exposure to the microtubule-disassembling agent, colchicine, was used to disrupt the axodendritic network and eventually trigger caspase-3-mediated apoptosis in cultures of cerebellar granule cells. For this model, we investigated the role of Bcl-2 and caspases in neurite degeneration and death of neuronal somata. Results: Early degeneration of the axodendritic network occurred by a Bcl-2 and caspase-independent mechanism. Conversely, apoptosis of the cell body was delayed by Bcl-2 and initially blocked by caspase inhibition. However, when caspase activity was entirely blocked by zVAD-fmk, colchicine-exposed neurons still underwent delayed cell death characterized by cytochrome c release, chromatin condensation to irregularly shaped clumps, DNA-fragmentation, and exposure of phosphatidylserine. Inhibitors of the proteasome reduced these caspase-independent apoptotic-like features of the neuronal soma. Conclusion: Our data suggest that Bcl-2 dependent and caspase-mediated death programs account only partially for neurodegenerative changes in injured neurons. Blockage of the caspase execution machinery may only temporarily rescue damaged neurons and classical apoptotic features can still appear in caspase-inhibited neurons.