Melatonin inhibits rotenone-induced SH-SY5Y cell death via the downregulation of Dynamin-Related Protein 1 expression

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Abstract

Previous studies have shown that melatonin can protect cells against rotenone-induced cell death. Yet, the mechanism involved in this protection requires further research. In this study, we aimed to further investigate the effects of melatonin on inhibiting rotenone-induced SH-SY5Y cells and the underlying molecular mechanisms. Human neuroblastoma SH-SY5Y cells were treated with 0.3 or 1 μM rotenone for 6 or 12 h. Cell viability was measured with an MTS assay, the mitochondrial membrane potential was determined with a Rhodamine 123 staining assay, and the protein expression levels of the markers of autophagy, including cytochrome C release (Cyt C), light chain 3B (LC3 B) and Dynamin-Related Protein 1 (Drp1) were analyzed by western blotting. The co-localization of Drp1 and TOM20 proteins in the mitochondria of SH-SY5Y cells was measured by immunofluorescence coupled with confocal microscopy and the overexpression of the Drp1 gene was then conducted. The viability and expression levels of Cyt C and LC3 B in rotenone and melatonin + rotenone-treated Drp1-overexpressed SH-SY5Y cells were analyzed with MTS and western blotting, respectively. We found that rotenone effectively induced SH-SY5Y cell death by causing mitochondrial dysfunction and increasing Cyt C expression. Drp1 expression and its regulation of mitochondrial translocation mediated the rotenone-induced cell death and melatonin inhibited this process. Overexpression of Drp1 protein attenuated melatonin's inhibition of rotenone-induced SH-SY5Y cell death. In conclusion, melatonin effectively inhibits rotenone-induced neuronal cell death via the regulation of Drp1 expression.

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