Search

A pathological hallmark of Alzheimer's disease (AD) is the presence within neurons and the interneuronal space of aggregates of β-amyloid (Aβ) peptides that originate from an abnormal proteolytic processing of the amyloid precursor protein (APP). The aspartyl proteases that initiate this processing act in the Golgi and endosomal compartments. Here, we show that the neurotransmitter dopamine stimulates the rapid endocytosis and processing of APP and induces apoptosis in neuroblastoma Neuro2A cells over-expressing transgenic human APP (Swedish mutant). Apoptosis could be prevented by impairing Pepstatin-sensitive and acid-dependent proteolysis of APP within endosomal-lysosomal compartments. The γ-secretase inhibitor L685,458 and the α-secretase stimulator phorbol ester elicited protection from dopamine-induced proteolysis of APP and cell toxicity. Our data shed lights on the mechanistic link between dopamine excitotoxicity, processing of APP and neuronal cell death. Since AD often associates with parkinsonian symptoms, which is suggestive of dopaminergic neurodegeneration, the present data provide the rationale for the therapeutic use of lysosomal activity inhibitors such as chloroquine or Pepstatin A to alleviate the progression of AD leading to onset of parkinsonism., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Hydrogen peroxide (H(2)O(2)) is an extremely reactive oxidoradical that is normally produced as a by-product of the mitochondrial activity and also under several metabolic stress conditions. Autophagy, a lysosomal degradation pathway, is triggered by oxidative stress as a defensive response. How autophagy and death pathways are coordinated in cells subjected to oxidative stress is still poorly understood. In human neuroblastoma SH-SY5Y cells, 200microM H(2)O(2) rapidly induced the formation of LC3-positive autophagic vacuoles and of beclin1-Vps34 double-positive macroaggregates. Vacuolar LC3 and beclin1 aggregates did not form when oxidative stress was performed in cells pretreated with 3-methyladenine (3MA), an inhibitor of Vps34, or infected with a recombinant adenovirus expressing a dominant-negative mutant of Vps34. H(2)O(2) provoked the permeabilization of lysosomes (at 30 min) and of mitochondria, the concomitant oligomerization of bax, and eventually (at 2 h), cell death in about 50% of the cell culture. Inactivation of Vps34-dependent autophagy in oxidative-stressed cells abrogated lysosome leakage, bax activation, and caspase-dependent apoptosis and conferred protection for as long as 16 h. Inhibition of caspase activity (by ZVAD-fmk) did not trigger an alternative cell death pathway but rather afforded complete protection from oxidative toxicity, despite the ongoing generation of oxidoradicals and the cellular accumulation of autophagic vacuoles and of leaking lysosomes. On long-term (16 h) exposure to H(2)O(2), signs of necrotic cell death became apparent in LC3-positive cells, which could be prevented by ZVAD-fmk. The present data highlight the pivotal role of autophagy in H(2)O(2)-induced cell death in dopaminergic neuroblastoma cells.

Methamphetamine abuse is toxic to dopaminergic neurons, causing nigrostriatal denervation and striatal dopamine loss. Following methamphetamine exposure, the number of nigral cell bodies is generally preserved, but their cytoplasm features autophagic-like vacuolization and cytoplasmic accumulation of alpha-synuclein-, ubiquitin- and parkin-positive inclusion-like bodies. Whether autophagy is epiphenomenal or it plays a role in the mechanism of methamphetamine toxicity and, in the latter case, whether its role consists of counteracting or promoting the neurotoxic effect remains obscure. We investigated the signaling pathway and the significance (protective vs. toxic) of autophagy activation and the convergence of the autophagic and the ubiquitin-proteasome pathways at the level of the same intracellular bodies in a simple cell model of methamphetamine toxicity. We show that autophagy is rapidly up-regulated in response to methamphetamine. Confocal fluorescence microscopy and immuno-electron microscopy studies demonstrated the presence of alpha-synuclein aggregates in autophagy-lysosomal structures in cells exposed to methamphetamine, a condition compatible with cell survival. Inhibition of autophagy either by pharmacologic or genetic manipulation of the class III Phosphatidylinositol-3 kinase-mediated signaling prevented the removal of alpha-synuclein aggregates and precipitated a bax-mediated mitochondrial apoptosis pathway.

ALS is a devastating neurodegenerative disorder with no effective treatment. In the present study, we found that daily doses of lithium, leading to plasma levels ranging from 0.4 to 0.8 mEq/liter, delay disease progression in human patients affected by ALS. None of the patients treated with lithium died during the 15 months of the follow-up, and disease progression was markedly attenuated when compared with age-, disease duration-, and sex-matched control patients treated with riluzole for the same amount of time. In a parallel study on a genetic ALS animal model, the G93A mouse, we found a marked neuroprotection by lithium, which delayed disease onset and duration and augmented the life span. These effects were concomitant with activation of autophagy and an increase in the number of the mitochondria in motor neurons and suppressed reactive astrogliosis. Again, lithium reduced the slow necrosis characterized by mitochondrial vacuolization and increased the number of neurons counted in lamina VII that were severely affected in saline-treated G93A mice. After lithium administration in G93A mice, the number of these neurons was higher even when compared with saline-treated WT. All these mechanisms may contribute to the effects of lithium, and these results offer a promising perspective for the treatment of human patients affected by ALS.