title: Keap1 redox-dependent regulation of doxorubicin-induced oxidative stress response in cardiac myoblasts.
abstract: Doxorubicin (DOX) is a widely prescribed treatment for a broad scope of cancers, but clinical utility is limited by the cumulative, dose-dependent cardiomyopathy that occurs with repeated administration. DOX-induced cardiotoxicity is associated with the production of reactive oxygen species (ROS) and oxidation of lipids, DNA and proteins. A major cellular defense mechanism against such oxidative stress is activation of the Keap1/Nrf2-antioxidant response element (ARE) signaling pathway, which transcriptionally regulates expression of antioxidant genes such as Nqo1 and Gstp1. In the present study, we address the hypothesis that an initial event associated with DOX-induced oxidative stress is activation of the Keap1/Nrf2-dependent expression of antioxidant genes and that this is regulated through drug-induced changes in redox status of the Keap1 protein. Incubation of H9c2 rat cardiac myoblasts with DOX resulted in a time- and dose-dependent decrease in non-protein sulfhydryl groups. Associated with this was a near 2-fold increase in Nrf2 protein content and enhanced transcription of several of the Nrf2-regulated down-stream genes, including Gstp1, Ugt1a1, and Nqo1; the expression of Nfe2l2 (Nrf2) itself was unaltered. Furthermore, both the redox status and the total amount of Keap1 protein were significantly decreased by DOX, with the loss of Keap1 being due to both inhibited gene expression and increased autophagic, but not proteasomal, degradation. These findings identify the Keap1/Nrf2 pathway as a potentially important initial response to acute DOX-induced oxidative injury, with the primary regulatory events being the oxidation and autophagic degradation of the redox sensor Keap1 protein.
authors.affiliation: University of Minnesota Medical School Duluth, 1035 University Dr., 252 SMed, Duluth, MN 55812, USA. Electronic address: knordgre@d.umn.edu.
authors.forename: Kendra K S
authors.initials: KK
authors.lastname: Nordgren
authors.affiliation: University of Minnesota Medical School Duluth, 1035 University Dr., 252 SMed, Duluth, MN 55812, USA. Electronic address: kwallace@d.umn.edu.
authors.forename: Kendall B
authors.initials: KB
authors.lastname: Wallace
mesh_terms: D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000633:toxicity; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004317:Doxorubicin / Q000633:toxicity*; D047908:Intracellular Signaling Peptides and Proteins / Q000502:physiology*; D000072019:Kelch-Like ECH-Associated Protein 1; D032386:Myoblasts, Cardiac / Q000187:drug effects* / Q000378:metabolism*; D010084:Oxidation-Reduction / Q000187:drug effects; D018384:Oxidative Stress / Q000187:drug effects* / Q000502:physiology*; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D000903:Antibiotics, Antineoplastic; D047908:Intracellular Signaling Peptides and Proteins; C486610:KEAP1 protein, rat; D000072019:Kelch-Like ECH-Associated Protein 1; D017382:Reactive Oxygen Species; D004317:Doxorubicin
keywords: Autophagy; Cardiotoxicity; Doxorubicin; H9c2; Keap1; Nrf2