Heat shock may increase oxidative stress due to increased production of reactive oxygen species (ROS) and/or the promotion of cellular oxidation events. Cytosolic NADP+-dependent isocitrate dehydrogenase (ICDH) in U937 cells produces NADPH, an essential reducing equivalent for the antioxidant system. The protective role of ICDH against heat shock in U937 cells was investigated in control and cells treated with oxlalomalate, a competitive inhibitor of ICDH. Upon exposure to heat shock, the viability was lower and the protein oxidation, lipid peroxidation and oxidative DNA damage were higher in oxalomalate-treated cells as compared to control cells. We also observed the significant increase in the endogenous production of ROS, as measured by the oxidation of 2'7'-dichlorodihydrofluorescin in U937 cells treated with oxalomalate. These results suggest that ICDH plays an important role as an antioxidant defense enzyme in cellular defense against heat shock through the removal of ROS.