title: Oxidative stress mediated cytotoxicity of cyanide in LLC-MK2 cells and its attenuation by alpha-ketoglutarate and N-acetyl cysteine.
abstract: Cyanide is a rapidly acting mitochondrial poison that inhibits cellular respiration and energy metabolism leading to histotoxic hypoxia followed by cell death. Cyanide is predominantly a neurotoxin but its toxic manifestations in non-neuronal cells are also documented. This study addresses the oxidative stress mediated cytotoxicity of cyanide in Rhesus monkey kidney epithelial cells (LLC-MK2). Cells were treated with various concentrations of potassium cyanide (KCN) for different time intervals and cytotoxicity was evidenced by increased leakage of intracellular lactate dehydrogenase, mitochondrial dysfunction (MTT assay) and depleted energy status of cells (ATP assay). Cytotoxicity was accompanied by lipid peroxidation indicated by elevated levels of malondialdehyde (MDA), reactive oxygen species (ROS) and reactive nitrogen species (RNS) (DCF-DA staining), diminished cellular antioxidant status (reduced glutathione (GSH), glutathione peroxidase, superoxide dismutase and catalase). These cascading events triggered an apoptotic kind of cell death characterized by oligonucleosomal DNA fragmentation and nuclear fragmentation (Hoechst 33342 staining). Apoptosis was further confirmed by increased caspase-3 activity. Cyanide-induced cytotoxicity, oxidative stress, and DNA fragmentation were prevented by alpha-ketoglutarate (A-KG) and N-acetyl cysteine (NAC). A-KG is a potential cyanide antidote that confers protection by interacting with cyanide to form cyanohydrin complex while NAC is a free radical scavenger and enhances the cellular GSH levels. The study reveals cytotoxicity of cyanide in cells of renal origin and the protective efficacy of A-KG and NAC.
authors.affiliation: Division of Pharmacology and Toxicology, Defence Research and Development Establishment, Jhansi Road, Gwalior 474002, India.
authors.forename: J
authors.initials: J
authors.lastname: Hariharakrishnan
authors.forename: R M
authors.initials: RM
authors.lastname: Satpute
authors.forename: G B K S
authors.initials: GB
authors.lastname: Prasad
authors.forename: R
authors.initials: R
authors.lastname: Bhattacharya
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology*; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D004734:Energy Metabolism / Q000187:drug effects; D004847:Epithelial Cells / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D007656:Ketoglutaric Acids / Q000494:pharmacology*; D015227:Lipid Peroxidation / Q000187:drug effects; D008253:Macaca mulatta; D018384:Oxidative Stress / Q000187:drug effects*; D011190:Potassium Cyanide / Q000633:toxicity*
chemical_list: D000975:Antioxidants; D007656:Ketoglutaric Acids; D011190:Potassium Cyanide; D000111:Acetylcysteine