Abstract
The effects of doxorubicin on the cellular biochemistry of the HeLa cell using 1H spin echo nuclear magnetic resonance spectroscopy (NMR) of the intact and viable cell in conjunction with dual wavelength HPLC of cell lysates is reported. Directly dose-related changes were observed in lactate and reduced glutathione concentration. Doxorubicin induces a time-dependent depletion of the cytosolic pool of glutathione and a change in the glycolytic pattern of the cell. The glutathione depletion could be partially reversed by controlled pre-treatment of the cells with N-acetylcysteine and cysteine, the protection being linked to the intracellular concentration of the thiol.
Highlights
The semiquinone can react with molecular oxygen (Hodgson et al, 1983) to produce the superoxide radical anion which is responsible for the extranuclear toxicity
Membrane-binding of the free radical products of doxorubicin metabolism and the subsequent alteration in both conformation and function is a mechanism with some significant implications for cardiac toxicity and tumour response (Hodgson et al, 1983)
Doxorubicin can act as an alkylating agent since the semiquinone derivative can undergo an intramolecular electron transfer with the release of the daunosamine anion (Hodgson et al, 1983)
Summary
Tetrabutylammonium hydroxide 40% w/w, reduced glutathione, cysteine and N-acetylcysteine were obtained from Sigma Chemical Company Ltd (UK), HPLC grade methanol from Rathburn Chemicals Ltd, Walkerburn (UK), Deuterium oxide (gold label) from Aldrich Chemical Company, Resorcinol and other chemical AnalaR grade were obtained from BDH Chemicals Ltd Poole, Dorset, UK. AL-KABBAN et al. Ltd, Maidenhead, Berks, UK (these are a preassembled version of the Amicon MPS-1 ultrafiltration apparatus). IXRPMI 1640 medium (Dutch modification) (with 20mM HEPES buffer, I g 1 1 sodium chloride) and trypsin 2.5% in Hanks balanced salt solution were obtained from Flow Laboratories (UK); foetal bovine serum (FBS) and Lglutamine from Gibco (UK) Ltd. Doxorubicin was a generous gift from Farmitalia (Italy)
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