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Colistin-Induced Apoptosis of Neuroblastoma-2a Cells Involves the Generation of Reactive Oxygen Species, Mitochondrial Dysfunction, and Autophagy

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Abstract

Neurotoxicity remains a poorly characterized adverse effect associated with colistin therapy. The aim of the present study was to investigate the mechanism of colistin-induced neurotoxicity using the mouse neuroblastoma2a (N2a) cell line. Colistin treatment (0–200 μM) of N2a neuronal cells induced apoptotic cell death in a dose-dependent manner. Colistin-induced neurotoxicity was associated with a significant increase of reactive oxygen species (ROS) levels, with a concomitant decrease in the activities of superoxide dismutase (SOD), catalase (CAT), and the glutathione (GSH) levels. Mitochondrial dysfunction was evident from the dissipation of membrane potential and the increase of Bax/Bcl-2, followed by the release of cytochrome c (CytC). Caspase-3/7, -8, and -9 activations were also detected. Colistin-induced neurotoxicity significantly increased the gene expression of p53 (1.6-fold), Bax (3.3-fold), and caspase-8 (2.2-fold) (all p < 0.01). The formation of autophagic vacuoles was evident with the significant increases (all p < 0.05 or 0.01) of both of Beclin 1 and LC3B following colistin treatment (50–200 μM). Furthermore, inhibition of autophagy by pretreatment with chloroquine diphosphate (CQ) enhanced colistin-induced apoptosis via caspase activation, which could be attenuated by co-treatment with the pan-caspase inhibitor Z-VAD-FMK. In summary, our study reveals that colistin-induced neuronal cell death involves ROS-mediated oxidative stress and mitochondrial dysfunction, followed by caspase-dependent apoptosis and autophagy. A knowledge base of the neuronal signaling pathways involved in colistin-induced neurotoxicity will greatly facilitate the discovery of neuroprotective agents for use in combination with colistin to prevent this undesirable side effect.

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Acknowledgments

This study was supported by the National Natural Science Foundation of China (award number 31372486). T. V. is supported by a research grant from the National Institute of Allergy and Infectious Diseases of the National Institutes of Health (R01 AI111965). T. V. is also supported by the Australian National Health and Medical Research Council (NHMRC). This study was supported by Chinese Universities Scientific Fund (Award number 2015DY003).

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The authors declare that there are no conflicts of interest.

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Authors and Affiliations

  1. College of Veterinary Medicine, China Agricultural University, 2 Yuanmingyuan West Road, Beijing, 100193, People’s Republic of China

    Chongshan Dai, Shusheng Tang & Xilong Xiao

  2. Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, 381 Royal Parade, Parkville, Victoria, 3052, Australia

    Tony Velkov

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  1. Chongshan Dai
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  2. Shusheng Tang
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  3. Tony Velkov
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  4. Xilong Xiao
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Correspondence to Tony Velkov or Xilong Xiao.

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Dr. Xilong Xiao is the first corresponding author and Dr. Tony Velkov is the co-corresponding author.

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Dai, C., Tang, S., Velkov, T. et al. Colistin-Induced Apoptosis of Neuroblastoma-2a Cells Involves the Generation of Reactive Oxygen Species, Mitochondrial Dysfunction, and Autophagy. Mol Neurobiol 53, 4685–4700 (2016). https://doi.org/10.1007/s12035-015-9396-7

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