The active site of thioredoxin-1 (Trx1) is oxidized in cells with increased reactive oxygen species (ROS) and is reduced by thioredoxin reductase-1 (TrxR 1). The purpose of the present study was to determine the extent to which the redox state of Trx 1 is sensitive to changes in these opposing reactions. Trx1 redox state and ROS generation were measured in cells exposed to the TrxR1 inhibitors aurothioglucose (ATG) and monomethylarsonous acid (MMA(III)) and in cells depleted of TrxR1 activity by siRNA knock down. The results showed that all three treatments inhibited TrxR1 activity to similar extents (90% inhibition), but that only MMA(111) exposure resulted in oxidation of Trx 1. Similarly, ROS levels were elevated in response to MMA(III), but not in response to ATG or TrxR I siRNA. Therefore, TrxR 1 inhibition alone was not sufficient to oxidize Trx 1, suggesting that Trx 1-independent pathways should be considered when evaluating pharmacological and toxicological mechanisms involving TrxR1 inhibition. (c) 2008 Elsevier Inc. All rights reserved.