title: Methotrexate induced mitochondrial injury and cytochrome c release in rat liver hepatocytes.
abstract: Methotrexate (MTX) is a folic acid antagonist that is widely used to treat a variety of diseases. One of the most serious side effects of MTX therapy is hepatotoxicity. The potential molecular cytotoxic mechanisms of MTX toward isolated rat hepatocytes were investigated using Accelerated Cytotoxicity Mechanism Screening (ACMS) techniques. A concentration and time dependent increase in cytotoxicity and reactive oxygen species (ROS) formation and a decrease in mitochondrial membrane potential (MMP) were observed with MTX. Furthermore, a significant increase in MTX (300 μM)-induced cytotoxicity and ROS formation were observed when glutathione (GSH)-depleted hepatocytes were used whereas addition of N-acetylcysteine (a GSH precursor) decreased cytotoxicity. Catalase inactivation also increased MTX-induced cytotoxicity, while the direct addition of catalase to the hepatocytes decreased cytotoxicity. MTX treatment in isolated rat mitochondria caused swelling and significantly decreased adenosine triphosphate (ATP) and GSH content, and cytochrome c release. Potent antioxidants such as mesna, resveratrol and Trolox decreased MTX-induced cytotoxicity and ROS formation and increased MMP. This study suggests that MTX-induced cytotoxicity caused by ROS formation and GSH oxidation leads to oxidative stress and mitochondrial injury in rat hepatocytes.
authors.affiliation: a Department of Pharmacology & Toxicology, University of Toronto, Toronto, Canada.
authors.forename: Abdullah
authors.initials: A
authors.lastname: Al Maruf
authors.affiliation: a Department of Pharmacology & Toxicology, University of Toronto, Toronto, Canada.|b Faculty of Pharmacy , University of Toronto , Toronto , Canada.
authors.forename: Peter J
authors.initials: PJ
authors.lastname: O'Brien
authors.affiliation: c Faculty of Pharmacy , Shahid Beheshti University of Medical Sciences , Tehran , Iran.
authors.forename: Parvaneh
authors.initials: P
authors.lastname: Naserzadeh
authors.affiliation: c Faculty of Pharmacy , Shahid Beheshti University of Medical Sciences , Tehran , Iran.
authors.forename: Rozhina
authors.initials: R
authors.lastname: Fathian
authors.affiliation: d Department of Pharmacology and Toxicology , School of Pharmacy, Ardabil University of Medical Science , Ardabil , Iran.
authors.forename: Ahmad
authors.initials: A
authors.lastname: Salimi
authors.affiliation: c Faculty of Pharmacy , Shahid Beheshti University of Medical Sciences , Tehran , Iran.
authors.forename: Jalal
authors.initials: J
authors.lastname: Pourahmad
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D002374:Catalase / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D005493:Folic Acid Antagonists / Q000633:toxicity*; D005978:Glutathione / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D015227:Lipid Peroxidation / Q000187:drug effects; D008297:Male; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008727:Methotrexate / Q000633:toxicity*; D008930:Mitochondria, Liver / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D008933:Mitochondrial Swelling / Q000187:drug effects; D018384:Oxidative Stress / Q000187:drug effects*; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism; D013997:Time Factors
chemical_list: D000975:Antioxidants; D005493:Folic Acid Antagonists; D017382:Reactive Oxygen Species; D000255:Adenosine Triphosphate; D045304:Cytochromes c; D002374:Catalase; D005978:Glutathione; D008727:Methotrexate
keywords: Hepatotoxicity; glutathione; mitochondrial injury; oxidative stress; reactive oxygen species