title: Cytotoxicity and ultrastructural changes in H9c2(2-1) cells treated with pavetamine, a novel polyamine.
abstract: Intake of pavetamine, a novel polyamine, synthesized by certain rubiaceous plants, is the cause of gousiekte ("Quick disease") in ruminants. The disease is characterized by a latent period of 4-8 weeks, followed by heart failure. The aim of this study was to firstly investigate the cytotoxicity in H9c2(2-1) cells using the MTT (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide) and LDH (lactate dehydrogenase) release assays. Maximum cell death occurred after pavetamine exposure of cells for 72h at a concentration of 200muM (55%+/-9.84), as measured by the MTT assay. LDH release was only observed after 72h exposure to pavetamine. Secondly, the ultrastructural changes induced by pavetamine in H9c2(2-1) cells were investigated. Changes in the mitochondria and sarcoplasmic reticula were observed. The nucleus was not affected during the first 48h exposure of cells to pavetamine and no chromatin condensation occurred. However, after 72h exposure to pavetamine, the nucleus became fragmented and membrane blebbing occurred. It was concluded that the ultimate cell death of H9c2(2-1) cells treated with pavetamine, was through necrosis and not apoptosis. Thirdly, the effect of pavetamine on the mitochondrial membrane potential (DeltaPsi) was evaluated by using the JC-1 (5,5',6,6'-Tetrachloro-1,1',3,3'-tetraethyl-imidacarbocyanine iodide) and TMRM (tetramethylrhodamine methyl ester perchlorate) probes. Pavetamine treatment led to significant hyperpolarization of the mitochondrial membrane potential. Cyclosporin A (CsA), an inhibitor of the mitochondrial permeability transition pore, did not reduce the cytotoxicity of pavetamine significantly, indicating that the MPTP (mitochondrial permeability transition pore) plays no role in the cytotoxicity of pavetamine.
authors.affiliation: Food, Feed and Veterinary Public Health Programme, Agriculture Research Council-Onderstepoort Veterinary Institute, Private Bag X5, Onderstepoort, Pretoria, Gauteng 0110, South Africa. ellisc@arc.agric.za
authors.forename: C E
authors.initials: CE
authors.lastname: Ellis
authors.forename: D
authors.initials: D
authors.lastname: Naicker
authors.forename: K M
authors.initials: KM
authors.lastname: Basson
authors.forename: C J
authors.initials: CJ
authors.lastname: Botha
authors.forename: R A
authors.initials: RA
authors.lastname: Meintjes
authors.forename: R A
authors.initials: RA
authors.lastname: Schultz
mesh_terms: D000818:Animals; D016923:Cell Death / Q000187:drug effects; D002460:Cell Line; D003603:Cytotoxins / Q000633:toxicity*; D004305:Dose-Response Relationship, Drug; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D046529:Microscopy, Electron, Transmission; D008931:Mitochondria, Muscle / Q000187:drug effects* / Q000648:ultrastructure; D033681:Mitochondrial Membrane Transport Proteins / Q000037:antagonists & inhibitors; D000083162:Mitochondrial Permeability Transition Pore; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000648:ultrastructure; D009336:Necrosis / Q000139:chemically induced; D018515:Plant Leaves / Q000737:chemistry; D011073:Polyamines / Q000302:isolation & purification / Q000633:toxicity*; D051381:Rats; D019666:Rubiaceae / Q000737:chemistry; D012519:Sarcoplasmic Reticulum / Q000187:drug effects* / Q000648:ultrastructure; D013997:Time Factors
chemical_list: D003603:Cytotoxins; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D011073:Polyamines; C462301:pavetamine