Thioredoxin is a multifunctional protein involved in protecting cells against oxidative stress, and it acts as a co-cytokine and chemotactic factor during an immune response. Mechanisms behind thioredoxin release and transport in the blood are unknown, although the presence of thioredoxin in extracellular fluids is indisputable. We investigated the role of circulatory thioredoxin. We have previously found that release of selenoprotein thioredoxin reductase to the blood is induced by oxidative stress, and thioredoxin reductase is present in plasma (Söderberg, Sabaf, and Rosén, Cancer Res. 60, 2281; 2000). Both thioredoxin and protein disulfide isomerase are substrates for thioredoxin reductase, and both are exposed on the surface oflymphocytes and monocytes. Plasma thioredoxin level is elevated by oxidative stress, a condition often seen in burn patients and HIV carriers. Three of our present findings demonstrate that platelets contain thioredoxin and that circulatory thioredoxin is transported primarily in platelets in healthy blood donors. First, thioredoxin, but not thioredoxin reductase was detected in platelets by deconvolution fluorescence microscopy. Second, plasma thioredoxin concentration was sensitive to thrombocytolysis: a significant decrease in thioredoxin was seen in plasma samples (n = 20) pretreated with the platelet degranulation inhibitors theophylline, adenosine, dipyridamol, acetylsalicylic acid, and apyrase (thioredoxin decreased from 28 down to 8 ng/ml; p < 0.0001). Release of thioredoxin from platelets was induced by the thioloxidant diamide but not by platelet degranulation caused by thrombin, a thrombin peptide (SFLLRN) and collagen, ADP, or PMA-ionophore. Third, in thrombocytopenic and thrombocytemic patients, plasma levels of thioredoxin, but not ß- thromboglobulin, were strongly correlated with platelet numbers (correlation coefficient, r = 0.7). Summarizing, we found direct evidence that thioredoxin is present in platelets and is liberated by oxidative stress (diamide). This suggests that platelets are essential for prompt delivery of this cellular reducing agent to sites of injury, where it can activate coagulation factors and subsequently reduce or balance reactive oxygen species released by inflammatory macrophages.