Oxidative stress, induced by 6-hydroxydopamine, reduces proteasome activities in PC12 cells: implications for the pathogenesis of Parkinson's disease.
ABSTRACT Mutations in familial Parkinson's disease (PD) have been associated with the failure of protein degradation through the ubiquitin-proteasome system (UPS). Impairment of proteasome function has also been suggested to play a role in the pathogenesis of sporadic PD. We examined the proteasome activity in PC12 cells treated with 6-hydroxydopamine (6-OHDA), the dopamine synthetic derivate used in models of PD. We found that 6-OHDA treatment increased protein oxidation, as indicated by carbonyl group accumulation, and increased caspase-3 activity. In addition, there was an increase in trypsin-, chymotrypsin-, and postacidic-like proteasome activities in cells treated with 10-100 microM 6-OHDA, whereas higher doses caused a marked decline. 6-OHDA exposure also increased mRNA expression of the 19S regulatory subunit in a dose-dependent manner, whereas the expression of 20S- and 11S-subunit mRNAs did not change. Administration of the antioxidant N-acetylcysteine to 6-OHDA-treated cells prevented the alteration in proteasome functions. Moreover, reduction in cell viability owing to administration of proteasome inhibitor MG132 or lactacystin was partially prevented by the endogenous antioxidant-reduced glutathione. In conclusion, our data indicate that mild oxidative stress elevates proteasome activity in response to increase in protein damage. Severe oxidative insult might cause UPS failure, which leads to protein aggregation and cell death. Moreover, in the case of UPS inhibition or failure, the blockade of physiological reactive oxygen species production during normal aerobic metabolism is enough to ameliorate cell viability. Control of protein clearance by potent, brain-penetrating antioxidants might act to slow down the progression of PD.
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ABSTRACT: A number of studies suggest that the ubiquitin-proteasome system (UPS) impairment may underlie neuronal death in Parkinson's disease. Celastrol is a neuroprotective agent with anti-inflammatory and antioxidant properties. The aim of this study was to determine whether celastrol may exert neuroprotective effects both in vitro and in vivo under conditions of the lactacystin-induced UPS inhibition. In the in vitro study, mouse primary cortical neurons and neuroblastoma SH-SY5Y cells were incubated with lactacystin for 48 h (2.5 and 10 μg/ml, respectively). The animal study was performed on male Wistar rats injected unilaterally with lactacystin (5 μg/2 μl) into the substantia nigra (SN) pars compacta. In the in vitro study, we did not found any protective effects of celastrol, given either in the pre- or co-treatment mode. Moreover, in the higher concentrations, celastrol itself reduced cell viability, and enhanced the lactacystin-induced cell death in both types of cells. In the in vivo study, none of the celastrol doses (0.3-3 mg/kg) attenuated the lactacystin-induced decrease in the level of dopamine (DA) and its metabolites or protected nigral dopaminergic neurons against the lactacystin-induced degeneration. The highest celastrol dose potentiated the lactacystin-induced decrease in the level of DA and its metabolites in the lesioned striatum, and accelerated the lactacystin-induced increase in the oxidative and total metabolism of DA. Moreover, when given alone, this dose of celastrol bilaterally decreased the number and/or density of dopaminergic neurons in the SN. Our results demonstrate that celastrol does not induce neuroprotective effects under conditions of UPS inhibition.Neurotoxicity Research 05/2014; · 2.87 Impact Factor
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ABSTRACT: The aim of the study was to determine whether the dopamine (DA) precursor L-DOPA attenuates parkinsonian-like symptoms produced by the ubiquitin-proteasome system inhibitor lactacystin. Wistar rats were injected unilaterally with lactacystin (2.5μg/2μl) or 6-OHDA (8μg/2μl) into the substantia nigra (SN) pars compacta. Four weeks after the lesion, the animals were treated chronically with L-DOPA (25 or 50mg/kg) for two weeks. During L-DOPA treatment, the lactacystin-treated rats were tested for catalepsy and forelimb asymmetry. Rotational behavior was evaluated after apomorphine (0.25mg/kg) and L-DOPA in both PD models. After completion of experiments, the animals were killed and the levels of DA and its metabolites in the striatum and SN were assayed. We found that acute L-DOPA administration effectively decreased catalepsy and increased the use of the compromised forelimb in the cylinder test. However, the lactacystin group did not respond to apomorphine or acute L-DOPA administration in the rotational test. Repeated L-DOPA treatment produced contralateral rotations in both PD models, but the number of rotations was much greater in the 6-OHDA-lesioned rats. Both toxins markedly (>90%) reduced the levels of DA and its metabolites in the striatum and SN, while L-DOPA diminished these decreases, especially in the SN. By demonstrating the efficacy of L-DOPA in several behavioral tests, our study confirms the usefulness of the lactacystin lesion as a model of PD. However, marked differences in the rotational response to apomorphine and L-DOPA suggest different mechanisms of neurodegeneration evoked by lactacystin and 6-OHDA.Behavioural brain research 03/2014; 261:79-88. · 3.22 Impact Factor
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ABSTRACT: Environmental toxins, genetic predisposition and old age are major risk factors for Parkinson’s disease (PD). Although the mechanism(s) underlying selective dopaminergic (DA) neurodegeneration remain unclear, molecular studies in both toxin based models and genetic based models of the disease suggest a major etiologic role for mitochondrial dysfunction in the pathogenesis of PD. Further, recent studies have presented clear evidence for a high burden of mtDNA deletions within the substantia nigra neurons in individuals with PD. Ultimately, an understanding of the molecular events which precipitate DA neurodegeneration in idiopathic PD will enable the development of targeted and effective therapeutic strategies. We review recent advances and current understanding of the genetic factors, molecular mechanisms and animal models of PD.Progress in Neurobiology 04/2013; · 9.04 Impact Factor