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Oral Administration of Rotenone using a Gavage and Image Analysis of Alpha-synuclein Inclusions in the Enteric Nervous System
Published on: October 26, 2010
Roflupram protects against rotenone-induced neurotoxicity and facilitates α-synuclein degradation in Parkinson's
Wen-Li Dong1, Jia-Hong Zhong1, Yun-Qing Chen1
1Guangdong Provincial Key Laboratory of New Drug Screening, School of Pharmaceutical Sciences, Southern Medical University, Guangzhou, 510515, China.
Abstract:
We have previously shown that roflupram (ROF) protects against MPP+-induced neuronal damage in models of Parkinson's disease (PD). Since impaired degradation of α-synuclein (α-syn) is one of the key factors that lead to PD, here we investigated whether and how ROF affects the degradation of α-syn in rotenone (ROT)-induced PD models in vivo and in vitro. We showed that pretreatment with ROF (10 μM) significantly attenuated cell apoptosis and reduced the level of α-syn in ROT-treated SH-SY5Y cells. Furthermore, ROF significantly enhanced the lysosomal function, as evidenced by the increased levels of mature cathepsin D (CTSD) and lysosomal-associated membrane protein 1 (LAMP1) through increasing NAD+/NADH and the expression of sirtuin 1 (SIRT1). Pretreatment with an SIRT1 inhibitor selisistat (SELI, 10 μM) attenuated the neuroprotection of ROF, ROF-reduced expression of α-syn, and ROF-increased expression levels of LAMP1 and mature CTSD. Moreover, inhibition of CTSD by pepstatin A (20 μM) attenuated ROF-reduced expression of α-syn. In vivo study was conducted in mice exposed to ROT (10 mg·kg-1·d-1, i.g.) for 6 weeks; then, ROT-treated mice received ROF (0.5, 1, or 2 mg·kg-1·d-1; i.g.) for four weeks. ROF significantly ameliorated motor deficits, which was accompanied by increased expression levels of tyrosine hydroxylase, SIRT1, mature CTSD, and LAMP1, and a reduced level of α-syn in the substantia nigra pars compacta. Taken together, these results demonstrate that ROF exerts a neuroprotective action and reduces the α-syn level in PD models. The mechanisms underlying ROF neuroprotective effects appear to be associated with NAD+/SIRT1-dependent activation of lysosomal function.
Insights
Roflunpram (ROF) reduces alpha-synuclein levels and protects neurons in Parkinson
Area of Science:
- Neuroscience
- Pharmacology
- Cell Biology
Background:
- Parkinson's disease (PD) is characterized by impaired alpha-synuclein (α-syn) degradation.
- Roflunpram (ROF) has previously shown neuroprotective effects against MPP+-induced neuronal damage.
- Investigating ROF's impact on α-syn degradation is crucial for understanding its therapeutic potential in PD.
Purpose of the Study:
- To determine if and how ROF affects α-syn degradation in rotenone (ROT)-induced Parkinson's disease models.
- To elucidate the underlying molecular mechanisms of ROF's neuroprotective action, focusing on lysosomal function and SIRT1.
- To evaluate ROF's efficacy in both in vitro and in vivo models of PD.
Main Methods:
- In vitro: SH-SY5Y cells treated with ROT and ROF, assessing cell apoptosis, α-syn levels, and lysosomal markers (CTSD, LAMP1).
- Molecular analysis involved measuring NAD+/NADH ratios and SIRT1 expression, with interventions using SIRT1 inhibitor (selisistat) and CTSD inhibitor (pepstatin A).
- In vivo: Mice exposed to ROT, followed by ROF treatment, evaluating motor deficits and protein expression in the substantia nigra pars compacta (tyrosine hydroxylase, SIRT1, CTSD, LAMP1, α-syn).
Main Results:
- ROF pretreatment attenuated ROT-induced cell apoptosis and reduced α-syn levels in SH-SY5Y cells.
- ROF enhanced lysosomal function by increasing mature CTSD and LAMP1, mediated by NAD+/NADH and SIRT1 activation.
- Inhibition of SIRT1 or CTSD counteracted ROF's protective effects and reduction in α-syn.
- In vivo, ROF ameliorated motor deficits in ROT-treated mice, increasing tyrosine hydroxylase, SIRT1, CTSD, and LAMP1, while decreasing α-syn in the substantia nigra.
Conclusions:
- Roflunpram (ROF) demonstrates significant neuroprotective effects and reduces α-synuclein levels in both in vitro and in vivo models of Parkinson's disease.
- The neuroprotective mechanism of ROF involves the activation of lysosomal function via the NAD+/SIRT1 pathway.
- These findings highlight ROF as a potential therapeutic agent for Parkinson's disease by targeting α-syn degradation and enhancing cellular clearance pathways.
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