title: Therapeutic concentrations of mitoxantrone elicit energetic imbalance in H9c2 cells as an earlier event.
abstract: Mitoxantrone (MTX) is a chemotherapeutic agent that emerged as an alternative to anthracycline therapy. However, MTX also causes late cardiotoxicity, being oxidative stress and mitochondrial-impaired function proposed as possible mechanisms. This work aimed to investigate the relevance of these mechanisms to the MTX toxicity in H9c2 cells, using therapeutic concentrations. The observed cytotoxicity of MTX was time and concentration dependent in both lactate dehydrogenase leakage assay and MTT reduction assay. Two therapeutic concentrations (100 nM and 1 μM) and three time points were selected (24, 48, and 96 h) for further studies. Both MTX concentrations caused a significant increase in caspase-3 activity, which was not prevented by inhibiting MTX CYP450-metabolism. Significant decreases were observed in the total and reduced glutathione levels only in MTX 100 nM at 96 h; however, neither alterations in oxidized glutathione nor increases in the malondialdehyde levels were observed at any time or concentrations tested. On the other hand, changes in the intracellular ATP levels, mitochondrial membrane potential, and intracellular calcium levels were observed in both concentrations and all time tested. Noteworthy, decreased levels of ATP-synthase expression and activity and increases in the reactive species generation were observed at 96 h in both working concentrations. However, the radical scavenger N-acetylcysteine or the mitochondrial function enhancer L-carnitine did not prevent MTX cytotoxicity. Thus, this work evidenced the early MTX-induced energetic crisis as a possible key factor in the cell injury.
authors.affiliation: REQUIMTE, Laboratório de Toxicologia, Departamento de Ciências Biológicas, Faculdade de Farmácia, Universidade do Porto, Rua Jorge Viterbo Ferreira, 228, 4050-313, Porto, Portugal, luciana.g.rossato@gmail.com.
authors.forename: Luciana Grazziotin
authors.initials: LG
authors.lastname: Rossato
authors.forename: Vera Marisa
authors.initials: VM
authors.lastname: Costa
authors.forename: Vânia
authors.initials: V
authors.lastname: Vilas-Boas
authors.forename: Maria
authors.initials: M
authors.lastname: de Lourdes Bastos
authors.forename: Anabela
authors.initials: A
authors.lastname: Rolo
authors.forename: Carlos
authors.initials: C
authors.lastname: Palmeira
authors.forename: Fernando
authors.initials: F
authors.lastname: Remião
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000008:administration & dosage*; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004734:Energy Metabolism / Q000187:drug effects* / Q000502:physiology; D008942:Mitoxantrone / Q000008:administration & dosage*; D032383:Myocytes, Cardiac / Q000187:drug effects* / Q000378:metabolism*; D051381:Rats; D013997:Time Factors
chemical_list: D000970:Antineoplastic Agents; D008942:Mitoxantrone