Autophagy protects the rotenone-induced cell death in alpha-synuclein overexpressing SH-SY5Y cells.
ABSTRACT Loss of dopaminergic cells induced by alpha-synuclein accumulation in substantia nigra causes the development of Parkinson's disease (PD). To date, although autophagy has been implicated in the pathology of PD, the molecular mechanism is still unclear. To study the role of autophagy in PD pathogenesis, we established stable SH-SY5Y cell lines overexpressing wild-type or mutant alpha-synuclein proteins (A30P or A53T). Overexpression of mutant alpha-synuclein induced some protein aggregates and cell death in the absence of drug. LC3-II protein, a critical marker for autophagy, was produced in an autophagy-dependent manner. The rotenone-induced cell death was interrupted by autophagy stimulation. Autophagy activation also restored the mitochondrial membrane potential (MMP) impaired by rotenone in mutant alpha-synuclein expressing cells. Additionally, autophagy activation significantly relieved rotenone-induced ROS accumulation and HIF-1alpha expression in neuronal cells expressing mutant alpha-synuclein proteins. These findings indicate that autophagy plays an important scavenger role against harmful influence of toxic protein aggregates produced in rotenone-treated cells.
- SourceAvailable from: Alexander V Panov[show abstract] [hide abstract]
ABSTRACT: Chronic systemic complex I inhibition caused by rotenone exposure induces features of Parkinson's disease (PD) in rats, including selective nigrostriatal dopaminergic degeneration and formation of ubiquitin- and alpha-synuclein-positive inclusions (Betarbet et al., 2000). To determine underlying mechanisms of rotenone-induced cell death, we developed a chronic in vitro model based on treating human neuroblastoma cells with 5 nm rotenone for 1-4 weeks. For up to 4 weeks, cells grown in the presence of rotenone had normal morphology and growth kinetics, but at this time point, approximately 5% of cells began to undergo apoptosis. Short-term rotenone treatment (1 week) elevated soluble alpha-synuclein protein levels without changing message levels, suggesting that alpha-synuclein degradation was retarded. Chronic rotenone exposure (4 weeks) increased levels of SDS-insoluble alpha-synuclein and ubiquitin. After a latency of >2 weeks, rotenone-treated cells showed evidence of oxidative stress, including loss of glutathione and increased oxidative DNA and protein damage. Chronic rotenone treatment (4 weeks) caused a slight elevation in basal apoptosis and markedly sensitized cells to further oxidative challenge. In response to H2O2, there was cytochrome c release from mitochondria, caspase-3 activation, and apoptosis, all of which occurred earlier and to a much greater extent in rotenone-treated cells; caspase inhibition provided substantial protection. These studies indicate that chronic low-grade complex I inhibition caused by rotenone exposure induces accumulation and aggregation of alpha-synuclein and ubiquitin, progressive oxidative damage, and caspase-dependent death, mechanisms that may be central to PD pathogenesis.Journal of Neuroscience 08/2002; 22(16):7006-15. · 6.91 Impact Factor
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ABSTRACT: Alpha-synuclein (alpha-syn) is a major component of inclusion bodies in Parkinson's disease (PD) and other synucleinopathies. To clarify the possible roles of alpha-syn in the molecular pathogenesis of neurodegenerative diseases, we have established a novel cellular model based on the differentiation of SH-SY5Y cells that overexpress alpha-syn. In the presence of ferrous iron, differentiation of the cells led to the formation of large perinuclear inclusion bodies, which developed from scattered small aggregates seen in undifferentiated cells. The iron-induced alpha-syn-positive inclusions co-localized largely with ubiquitin, and some of them were positive for nitrotyrosine, lipid, gamma-tubulin and dynein. Notably, treatment with nocodazole, a microtubule depolymerizing agent, interrupted the aggregate formation but led to a concomitant increase of apoptotic cells. Therefore, it appears that an intracellular retrograde transport system via microtubules plays a crucial role in the aggregate formation and also that the aggregates may represent a cytoprotective response against noxious stimuli. This cellular model will enable better understanding of the molecular pathomechanisms of synucleinopathy.Brain Research 08/2004; 1013(1):51-9. · 2.88 Impact Factor
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ABSTRACT: Studies in yeast are providing critical insights into the mechanisms of neurodegeneration in Parkinson's disease (PD). A recent study shows that disruption of vesicular trafficking between the endoplasmic reticulum (ER) and the Golgi, caused by the overexpression and/or aggregation of alpha-synuclein, is linked to degeneration of dopamine neurons. Overexpression of proteins that are known to enhance ER-to-Golgi transport rescue defective trafficking in yeast, worm, fly, and cellular models of PD.ACS Chemical Biology 09/2006; 1(7):420-4. · 5.44 Impact Factor