title: Akt/GSK3β signaling is involved in fipronil-induced apoptotic cell death of human neuroblastoma SH-SY5Y cells.
abstract: Fipronil (FPN) is a phenylpyrazole insecticide acted on insect gamma-aminobutyric acid (GABA) receptors. Although action of FPN is restricted on insect neuronal or muscular transmitter system, a few studies have assessed the effects of this neurotoxicant on neuronal cell death. To determine the mechanisms underlying FPN-induced neuronal cell death, we investigated whether reactive oxygen species (ROS) plays a role in FPN-induced apoptosis, using an in vitro model of human dopaminergic SH-SY5Y cells. FPN was cytotoxic to these cells and its cytotoxicity showed a concentration-dependent manner. Additionally, FPN treatment significantly decreased the tyrosine hydroxylase (TH) expression without change of glutamic acid decarboxylase 65 (GAD65) expression. FPN-induced dopaminergic cell death involved in increase of ROS generation since pretreatment with N-acetyl cysteine (NAC), an anti-oxidant, reduced cell death. After FPN treatment, dopamine (DA) levels decreased significantly in both cell and culture media, and oxidative effects of DA were blocked by NAC pretreatment. We showed that cell death in response to FPN was due to apoptosis since FPN increased cytochrome c release into the cytosol and activated caspase-3. It also led to nuclear accumulation of p53 and reduced the level of Bcl-2 protein in a concentration-dependent manner. Additionally, FPN altered the level of Akt/glycogen synthase kinase-3 (GSK3β) phosphorylation. FPN reduced the Akt phosphorylation on Ser473, and in parallel with the inactivation of Akt, phosphorylation of GSK3β on Ser9 which inactivates GSK3β, decreased after treatment with FPN. Furthermore, inhibition of the GSK3β signal protected the cell against FPN-induced cell death. These results suggest that regulation of GSK3β activity may control the apoptosis induced by FPN-induced oxidative stress associated with neuronal cell death.
authors.affiliation: Department of Pharmacology, College of Medicine, Hanyang University, 133-791 Seoul, Republic of Korea.
authors.forename: Jeong Eun
authors.initials: JE
authors.lastname: Lee
authors.forename: Jin Sun
authors.initials: JS
authors.lastname: Kang
authors.forename: Yeo-Woon
authors.initials: YW
authors.lastname: Ki
authors.forename: Sang-Hun
authors.initials: SH
authors.lastname: Lee
authors.forename: Soo-Jin
authors.initials: SJ
authors.lastname: Lee
authors.forename: Kyung Suk
authors.initials: KS
authors.lastname: Lee
authors.forename: Hyun Chul
authors.initials: HC
authors.lastname: Koh
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D045304:Cytochromes c / Q000378:metabolism; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D007306:Insecticides / Q000494:pharmacology*; D008856:Microscopy, Fluorescence; D009474:Neurons / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011720:Pyrazoles / Q000494:pharmacology*; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
chemical_list: D007306:Insecticides; D019253:Proto-Oncogene Proteins c-bcl-2; D011720:Pyrazoles; D017382:Reactive Oxygen Species; D016159:Tumor Suppressor Protein p53; D045304:Cytochromes c; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D051057:Proto-Oncogene Proteins c-akt; D038362:Glycogen Synthase Kinase 3; D053148:Caspase 3; C082360:fipronil