科研之友
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摘要
Tetrazolium salts such as KIT and MTT are widely used to produce formazan for cell proliferation and cytotoxicity assays through bioreductase activity However,. the XTT assay showed significant increase in MDBK cell viability when cells were treated with both 50 and 100 mu M of the pro-oxidant, to t-butylhydroquinone (t-BHQ). although the crystal violet assay showed no cytotoxic effect with these concentrations. and the induction of lipid peioxidation was not observed We investigated the mechanism ot enhancement of XTT substrate reduction after treatment of MDBK cells with t-BHQ. leading to apparent increase in cell viability t-BHQ caused an increase in absorbance at 340 inn in culture medium. suggesting that t-BHQ increases cellular production and release of NADH and/or NADPH Although t-BHQ did not change the NADH concentration in cell culture medium, the addition of NADP(+)-dependent glutathione reductase decreased the XTT reduction to the control level. indicating cellular release of NADPH t-BHQ also increased intracellular glucose-6-phosphate dehydrogenase activity. producing NADPH Taken together. our findings indicate that t-BHQ treatment activates NADPH generating enzymes such as glucose-6-phosphate dehydrogenase followed by release of NADPH in the cell culture medium. resulting in direct XTT reduction by NADPH
- 出版日期2010-3
