Neurotoxic Effect of Fipronil in Neuroblastoma SH-SY5Y Cell Line
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Fipronil is a broad-spectrum insecticide belonging to the phenyl pyrazole chemical family. Fipronil disrupts gamma-aminobutyric acid (GABA) receptors in the central nervous system, thereby blocking GABA-gated chloride channels. Neurotoxic symptoms of fipronil poisoning in humans are typically associated with the antagonism of central GABA receptors. In this study, the cytotoxic effects of fipronil in SH-SY5Y neuroblastoma cell line, the human neural cell line, were investigated. SH-SY5Y cells were exposed to 125, 250, and 500 μM fipronil doses and incubated for 24 and 48 h, and its neurotoxic effect was examined by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl tetrazolium bromide) test, lactate dehydrogenase (LDH) test, trypan blue dye exclusion test, and neutral red uptake assay. Acridine orange-propidium iodide and DAPI (4′,6-diamidino-2-phenylindole dihydrochloride) staining methods were also used to determine the type of cell death. In addition, neurodegenerative changes were also examined by immunofluorescence staining of the cytoskeleton. The amount of neurofilament heavy chain and the degree of phosphorylation of neurofilament subunits were investigated to determine the effect of fipronil on the axonal viability, and we observed a reduction in the amount of neurofilament heavy chain, speculating disruption of the axonal transport leading to cell death. According to our results, we may suggest that fipronil caused cytotoxicity in SH-SY5Y cells.
KeywordsFipronil SH-SY5Y cell line Neurotoxicity Immunofluorescence staining Neurofilament phosphorylation
This work was financially supported by Hacettepe University Scientific Projects Coordination Unit (Project No. FHD-2015-6929). This research is a MSc thesis of Özge Nur Kanat.
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