The Pseudomonas secretory product pyocyanin inhibits catalase activity in human lung epithelial cells

1 Research Service and 4 Department of Internal Medicine, Veterans Affairs Medical Center-Iowa City; 2 Department of Internal Medicine and 3 The Free Radical and Radiation Biology Program of the Department of Radiation Oncology, The Roy G. and Lucille A. Carver College of Medicine, The University of Iowa, Iowa City, Iowa 52242 Submitted 23 June 2003 ; accepted in final form 14 July 2003 Pyocyanin, produced by Pseudomonas aeruginosa , has many deleterious effects on human cells that relate to its ability to generate reactive oxygen species (ROS), such as superoxide and hydrogen peroxide. Human cells possess several mechanisms to protect themselves from ROS, including manganese superoxide dismutase (MnSOD), copper zinc superoxide dismutase (CuZnSOD), and catalase. Given the link between pyocyanin-mediated epithelial cell injury and oxidative stress, we assessed pyocyanin's effect on MnSOD, CuZnSOD, and catalase levels in the A549 human alveolar epithelial cell line and in normal human bronchial epithelial cells. In both cell types, CuZnSOD and MnSOD were unaltered, but over 24 h pyocyanin significantly decreased cellular catalase activity and protein content. Pyocyanin also decreased catalase mRNA. Overexpression of MnSOD in A549 cells prevented pyocyanin-mediated loss of catalase protein, but catalase activity still declined. Furthermore, pyocyanin decreased catalase activity, but not protein, in A549 cells overexpressing human catalase. These data suggest a direct effect of pyocyanin on catalase activity. Addition of pyocyanin to catalase in a cell-free system also decreased catalase activity. Mammalian catalase binds four NADPH molecules, helping maintain enzyme activity. Spin-trapping data suggest that pyocyanin directly oxidizes this NADPH, producing superoxide. We conclude that pyocyanin may decrease cellular catalase activity via both transcriptional regulation and direct inactivation of the enzyme. Decreased cellular catalase activity and failure to augment MnSOD could contribute to pyocyanin-dependent cytotoxicity. hydrogen peroxide; superoxide dismutase; NADPH; cystic fibrosis; oxidant Address for reprint requests and other correspondence: B. E. Britigan, Univ. of Iowa Hospitals and Clinics, Dept. of Internal Medicine, Div. of Infectious Diseases, 200 Hawkins Dr., SW54, GH, Iowa City, IA 52242 (E-mail: bradley-britigan{at}uiowa.edu ).