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The Neuroprotective Effect of Erythropoietin on Rotenone-Induced Neurotoxicity in SH-SY5Y Cells Through the Induction of Autophagy

Molecular Neurobiology volume 53, pages 3812–3821 (2016)Cite this article

Abstract

Currently, the autophagy pathway is thought to be important for the pathogenesis of Parkinson’s disease (PD), and the modulation of autophagy may be a novel strategy for the treatment of this disease. Erythropoietin (EPO) has been reported to have neuroprotective effects through anti-oxidative, anti-apoptotic, and anti-inflammatory mechanisms, and it has also been shown to modulate autophagy signaling in an oxygen toxicity model. Therefore, we investigated the effects of EPO on autophagy markers and evaluated its neuroprotective effect on rotenone-induced neurotoxicity. We adapted the rotenone-induced neurotoxicity model to SH-SY5Y cells as an in vitro model of PD. We measured cell viability using MTT and annexin V/propidium iodide assays and measured intracellular levels of reactive oxygen species. Immunofluorescence analysis was performed to measure the expression of LC3 and α-synuclein. Intracellular signaling proteins associated with autophagy were examined by immunoblot analysis. EPO mono-treatment increased the levels of mammalian target of rapamycin (mTOR)-independent/upstream autophagy markers, including Beclin-1, AMPK, and ULK-1. Rotenone treatment of SH-SY5Y cells reduced their viability, increased reactive oxygen species levels, and induced apoptosis and α-synuclein expression, and simultaneous exposure to EPO significantly reduced these effects. Rotenone enhanced mTOR expression and suppressed Beclin-1 expression, indicating suppression of the autophagy system. However, combined treatment with EPO restored Beclin-1 expression and decreased mTOR expression. EPO protects against rotenone-induced neurotoxicity in SH-SY5Y cells by enhancing autophagy-related signaling pathways. The experimental evidence for the EPO-induced neuroprotection against rotenone-induced dopaminergic neurotoxicity may significantly impact the development of future PD treatment strategies.

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Acknowledgments

This research was financially sponsored by Hanmi Pharmaceutical Co, Ltd through Foundation for Industry Cooperation, University of Ulsan (2015-0598). However, the study was investigator initiated, and the sponsor had no involvement in the study design, in the collection, analysis, and interpretation of data, and in the writing the manuscript. Hanmi Pharmaceutical also had no input into the decision to submit this article for publication. The authors had full access to all the data in the study, and the corresponding author had the final responsibility to submit the manuscript for publication.

This work was also funded by Korea Institute of Science and Technology Natural Products Research Center, and the Bio-Synergy Research Project (NRF-2012M3A9C4048793) of the Ministry of Science, ICT, and Future Planning of the government of the Republic of Korea.

Conflict of Interest

Neither any of the authors have anything to disclose regarding this manuscript nor do they have any potential conflicts of interest to report concerning this article.

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    1. Department of Neurology, Gangneung Asan Hospital, University of Ulsan College of Medicine, Bangdong-ri, Sacheon-myeon, Gangneung, Gangwon-do, 210-711, Republic of Korea

      Wooyoung Jang, Kwang Deog Jo & Moon Kyu Lee

    2. Natural Products Research Center, Korea Institute of Science and Technology, 679 Saimdang-ro, Gangneung, 210-340, Republic of Korea

      Hee Ju Kim, Huan Li & Hyun Ok Yang

    3. Biomedical Research Center, Gangneung Asan Hospital, Bangdong-ri, Sacheon-myeon, Gangneung, 210-711, Gangwon-do, Republic of Korea

      Wooyoung Jang & Hee Ju Kim

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    1. Wooyoung Jang
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    Correspondence to Wooyoung Jang or Hyun Ok Yang.

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    Wooyoung Jang and Hee Ju Kim contributed equally to this work.

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    Jang, W., Kim, H.J., Li, H. et al. The Neuroprotective Effect of Erythropoietin on Rotenone-Induced Neurotoxicity in SH-SY5Y Cells Through the Induction of Autophagy. Mol Neurobiol 53, 3812–3821 (2016). https://doi.org/10.1007/s12035-015-9316-x

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    • DOI: https://doi.org/10.1007/s12035-015-9316-x

    Keywords

    • Erythropoietin
    • Parkinson’s disease
    • Autophagy
    • Neuroprotection