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Astrocyte Activation: A Key Step in Rotenone Induced Cytotoxicity and DNA Damage

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Abstract

Astrocytes are the most abundant glial cells, which provide metabolic support for neurons. Rotenone is a botanical pesticide of natural origin, known to exhibit neurotoxic potential via inhibition of mitochondrial complex-I. This study was carried out to explore the effect of rotenone on C6 cells. The cell line C6 derived from rat glioma cells represents astrocyte-like cell. C6 cells were treated with rotenone (0.1, 1 and 10 μM) for 4 h. The effect of rotenone was studied on cell survival (MTT reduction and PI uptake); free radicals (ROS and RNS) and DNA damage (comet assay and Hoechst staining). The glial cell activation and apoptotic cell death was evaluated by expression of Glial fibrillary acidic protein (GFAP) and caspase-3 respectively. The treatment with rotenone resulted in decreased cell survival and increased free radical generation. Altered nuclear morphology and DNA damage were evident following rotenone treatment in Hoechst staining and Comet assay. Rotenone elevated expression of GFAP and caspase-3 that indicates glial cell activation and apoptosis, respectively. We further studied the effect of melatonin, an antioxidant, on the observed toxic effects. Co-incubation of antioxidant, melatonin (300 μM), significantly suppressed rotenone induced above-mentioned effects in C6 cells. Inhibitory effects of melatonin suggest that free radicals play a major role in rotenone induced astrocyte activation and cellular toxicity leading to apoptosis of astroglial cells.

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Abbreviations

DNA:

Deoxyribonucleic acid

EDTA:

Ethylenediamine tetraacetic acid

BSA:

Bovine serum albumin

PBS:

Phosphate buffer saline

PI:

Propidium iodide

RNS:

Reactive nitrogen species

ROS:

Reactive oxygen species

SCGE:

Single cell gel electrophoresis

LMP:

Low melting point

GFAP:

Glial fibrillary acidic protein

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Acknowledgments

Authors are thankful to Dr. A. K. Balapure, Head, Tissue and Cell Culture Unit, CSIR-CDRI and his team for providing cells for experimentation and to Mr. Pradeep Kamat and Miss Aameena Siddiqui for their assistance. Supriya Swarnkar gratefully acknowledges the Council of Scientific and Industrial Research (CSIR), India for research fellowship.

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Authors and Affiliations

  1. Division of Toxicology, CSIR-Central Drug Research Institute, P.O. Box 173, Lucknow, 226001, UP, India

    Supriya Swarnkar, Sarika Singh, Poonam Goswami & Chandishwar Nath

  2. School of Studies in Zoology, Jiwaji University, Gwalior, 474011, MP, India

    Ramesh Mathur & Ishan K. Patro

Authors
  1. Supriya Swarnkar
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  2. Sarika Singh
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  3. Poonam Goswami
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  4. Ramesh Mathur
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  5. Ishan K. Patro
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  6. Chandishwar Nath
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Correspondence to Chandishwar Nath.

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Swarnkar, S., Singh, S., Goswami, P. et al. Astrocyte Activation: A Key Step in Rotenone Induced Cytotoxicity and DNA Damage. Neurochem Res 37, 2178–2189 (2012). https://doi.org/10.1007/s11064-012-0841-y

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  • DOI: https://doi.org/10.1007/s11064-012-0841-y

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