Author(s): Mirabelli F, Salis A, Marinoni V, Finardi G, Bellomo G,
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Abstract Incubation of isolated rat hepatocytes with menadione (2-methyl-1,4-naphthoquinone) or the thiol oxidant, diamide (azodicarboxylic acid bis(dimethylamide)), resulted in the appearance of numerous plasma membrane protrusions (blebs) preceding cell death. Analysis of the Triton X-100-insoluble fraction (cytoskeleton) extracted from treated cells revealed a dose- and time-dependent increase in the amount of cytoskeletal protein and a concomitant loss of protein thiols. These changes were associated with the disappearance of actin and formation of large-molecular-weight aggregates, when the cytoskeletal proteins were analyzed by polyacrylamide gel electrophoresis under nonreducing conditions. However, if the cytoskeletal proteins were treated with the thiol reductants, dithiothreitol or beta-mercaptoethanol, no changes in the relative abundance of actin or formation of large-molecular-weight aggregates were detected in the cytoskeletal preparations from treated cells. Moreover, addition of dithiothreitol to menadione- or diamide-treated hepatocytes protected the cells from both the appearance of surface blebs and the occurrence of alterations in cytoskeletal protein composition. Our findings show that oxidative stress induced by the metabolism of menadione in isolated hepatocytes causes cytoskeletal abnormalities, of which protein thiol oxidation seems to be intimately related to the appearance of surface blebs.
This article was published in Arch Biochem Biophys
and referenced in Journal of Proteomics & Bioinformatics