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Proteome analysis reveals roles of L-DOPA in response to oxidative stress in neurons.

Jami MS, Pal R, Hoedt E, Neubert TA, Larsen JP, Møller SG - BMC Neurosci (2014)

Bottom Line: We observed that oxidative stress affects metabolic pathways as well as cytoskeletal integrity and that neuronal cells respond to oxidative conditions by enhancing numerous survival pathways.Oxidative stress changes neuronal metabolic routes and affects cytoskeletal integrity.Further, L-DOPA appears to reverse some H2O2-mediated effects evident at both the proteome and cellular level.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, St John's University, New York, NY, USA. mollers@stjohns.edu.

ABSTRACT

Background: Parkinson's disease (PD) is the second most common neurodegenerative movement disorder, caused by preferential dopaminergic neuronal cell death in the substantia nigra, a process also influenced by oxidative stress. L-3,4-dihydroxyphenylalanine (L-DOPA) represents the main treatment route for motor symptoms associated with PD however, its exact mode of action remains unclear. A spectrum of conflicting data suggests that L-DOPA may damage dopaminergic neurons due to oxidative stress whilst other data suggest that L-DOPA itself may induce low levels of oxidative stress, which in turn stimulates endogenous antioxidant mechanisms and neuroprotection.

Results: In this study we performed a two-dimensional gel electrophoresis (2DE)-based proteomic study to gain further insight into the mechanism by which L-DOPA can influence the toxic effects of H2O2 in neuronal cells. We observed that oxidative stress affects metabolic pathways as well as cytoskeletal integrity and that neuronal cells respond to oxidative conditions by enhancing numerous survival pathways. Our study underlines the complex nature of L-DOPA in PD and sheds light on the interplay between oxidative stress and L-DOPA.

Conclusions: Oxidative stress changes neuronal metabolic routes and affects cytoskeletal integrity. Further, L-DOPA appears to reverse some H2O2-mediated effects evident at both the proteome and cellular level.

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Related in: MedlinePlus

Cell morphology and cell viability of SH-SY5Y cells in response to H2O2, L-DOPA and H2O2/L-DOPA treatments. (A) Cell morphology of SH-SY5Y cells in response to eight hours of 2 mM H2O2, 200 μM L-DOPA and 2 mM H2O2/200 μM L-DOPA treatments. (B) Neutral red cell viability assays of SH-SY5Y cells in response to eight hours of 2 mM H2O2, 200 μM L-DOPA and 2 mM H2O2/200 μM L-DOPA treatments. Each data point is the average of three replicate samples and presented as means ± SD. Standard deviations are indicated by error bars. **p < 0.01.
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Fig1: Cell morphology and cell viability of SH-SY5Y cells in response to H2O2, L-DOPA and H2O2/L-DOPA treatments. (A) Cell morphology of SH-SY5Y cells in response to eight hours of 2 mM H2O2, 200 μM L-DOPA and 2 mM H2O2/200 μM L-DOPA treatments. (B) Neutral red cell viability assays of SH-SY5Y cells in response to eight hours of 2 mM H2O2, 200 μM L-DOPA and 2 mM H2O2/200 μM L-DOPA treatments. Each data point is the average of three replicate samples and presented as means ± SD. Standard deviations are indicated by error bars. **p < 0.01.

Mentions: In order to provide a global overview regarding the mechanism in which L-DOPA may influence toxic effects of H2O2 in neuronal cells we performed cell viability analysis and subsequent proteomic analyses using SH-SY5Y cells. Although primary neurons or dopaminergic neurons would be ideal for this study in relation to PD we selected SH-SY5Y cells based on reproducibility and because they are dopamine beta hydroxylase active. SH-SY5Y cells were grown under control conditions and were exposed to 2 mM H2O2, 200 μM L-DOPA or a combination of the two treatments (2 mM H2O2/200 μM L-DOPA) for eight hours. Cell morphology analysis (Figure 1A) and cell viability assays (Figure 1B) demonstrated that SH-SY5Y cells exposed to L-DOPA showed no effect on morphology or cell viability (Figure 1). By contrast, H2O2 exposure decreased cell viability, but this effect was reversed in response to co-treatment with L-DOPA (Figure 1). This suggested that L-DOPA might have a protective effect towards excess oxidative stress.Figure 1


Proteome analysis reveals roles of L-DOPA in response to oxidative stress in neurons.

Jami MS, Pal R, Hoedt E, Neubert TA, Larsen JP, Møller SG - BMC Neurosci (2014)

Cell morphology and cell viability of SH-SY5Y cells in response to H2O2, L-DOPA and H2O2/L-DOPA treatments. (A) Cell morphology of SH-SY5Y cells in response to eight hours of 2 mM H2O2, 200 μM L-DOPA and 2 mM H2O2/200 μM L-DOPA treatments. (B) Neutral red cell viability assays of SH-SY5Y cells in response to eight hours of 2 mM H2O2, 200 μM L-DOPA and 2 mM H2O2/200 μM L-DOPA treatments. Each data point is the average of three replicate samples and presented as means ± SD. Standard deviations are indicated by error bars. **p < 0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4125692&req=5

Fig1: Cell morphology and cell viability of SH-SY5Y cells in response to H2O2, L-DOPA and H2O2/L-DOPA treatments. (A) Cell morphology of SH-SY5Y cells in response to eight hours of 2 mM H2O2, 200 μM L-DOPA and 2 mM H2O2/200 μM L-DOPA treatments. (B) Neutral red cell viability assays of SH-SY5Y cells in response to eight hours of 2 mM H2O2, 200 μM L-DOPA and 2 mM H2O2/200 μM L-DOPA treatments. Each data point is the average of three replicate samples and presented as means ± SD. Standard deviations are indicated by error bars. **p < 0.01.
Mentions: In order to provide a global overview regarding the mechanism in which L-DOPA may influence toxic effects of H2O2 in neuronal cells we performed cell viability analysis and subsequent proteomic analyses using SH-SY5Y cells. Although primary neurons or dopaminergic neurons would be ideal for this study in relation to PD we selected SH-SY5Y cells based on reproducibility and because they are dopamine beta hydroxylase active. SH-SY5Y cells were grown under control conditions and were exposed to 2 mM H2O2, 200 μM L-DOPA or a combination of the two treatments (2 mM H2O2/200 μM L-DOPA) for eight hours. Cell morphology analysis (Figure 1A) and cell viability assays (Figure 1B) demonstrated that SH-SY5Y cells exposed to L-DOPA showed no effect on morphology or cell viability (Figure 1). By contrast, H2O2 exposure decreased cell viability, but this effect was reversed in response to co-treatment with L-DOPA (Figure 1). This suggested that L-DOPA might have a protective effect towards excess oxidative stress.Figure 1

Bottom Line: We observed that oxidative stress affects metabolic pathways as well as cytoskeletal integrity and that neuronal cells respond to oxidative conditions by enhancing numerous survival pathways.Oxidative stress changes neuronal metabolic routes and affects cytoskeletal integrity.Further, L-DOPA appears to reverse some H2O2-mediated effects evident at both the proteome and cellular level.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, St John's University, New York, NY, USA. mollers@stjohns.edu.

ABSTRACT

Background: Parkinson's disease (PD) is the second most common neurodegenerative movement disorder, caused by preferential dopaminergic neuronal cell death in the substantia nigra, a process also influenced by oxidative stress. L-3,4-dihydroxyphenylalanine (L-DOPA) represents the main treatment route for motor symptoms associated with PD however, its exact mode of action remains unclear. A spectrum of conflicting data suggests that L-DOPA may damage dopaminergic neurons due to oxidative stress whilst other data suggest that L-DOPA itself may induce low levels of oxidative stress, which in turn stimulates endogenous antioxidant mechanisms and neuroprotection.

Results: In this study we performed a two-dimensional gel electrophoresis (2DE)-based proteomic study to gain further insight into the mechanism by which L-DOPA can influence the toxic effects of H2O2 in neuronal cells. We observed that oxidative stress affects metabolic pathways as well as cytoskeletal integrity and that neuronal cells respond to oxidative conditions by enhancing numerous survival pathways. Our study underlines the complex nature of L-DOPA in PD and sheds light on the interplay between oxidative stress and L-DOPA.

Conclusions: Oxidative stress changes neuronal metabolic routes and affects cytoskeletal integrity. Further, L-DOPA appears to reverse some H2O2-mediated effects evident at both the proteome and cellular level.

Show MeSH
Related in: MedlinePlus