The glutathione (GSH) redox reaction is critical for defense against cellular reactive oxygen species (ROS). However, direct and real-time monitoring of this reaction in living mammalian cells has been hindered by the lack of a facile method. Herein, we describe a new approach that exploits the GSH biosynthetic pathway and heteronuclear NMR. [U-(13) C]-labeled cysteine was incorporated into GSH in U87 glioblastoma cells, and the oxidation of GSH to GSSG by a ROS-producing agent could be monitored in living cells. Further application of the approach to cells resistant to temozolomide (TMZ), an anti-glioblastoma drug, suggested a possible new resistance mechanism involving neutralization of ROS. This result was corroborated by the observation of up-regulation of glutathione peroxidase 3 (GPx3). This new approach could be easily applied to redox-dependent signaling pathways and drug resistance involving ROS.
Keywords: cancer; drug resistance; glutathione; in-cell NMR; redox chemistry.
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