Oxygen radicals regulate many physiological processes, such as signaling, proliferation, and apoptosis, and thus play a pivotal role in pathophysiology and disease development. There are at least two thioredoxin reductase/ thioredoxin/peroxiredoxin systems participating in the cellular defense against oxygen radicals. At present, relatively little is known about the contribution of individual enzymes to the redox metabolism in different cell types. To begin to address this question, we generated and characterized mice lacking functional mitochondrial thioredoxin reductase (TrxR2). Ubiquitous Cre-mediated inactivation of TrxR2 is associated with embryonic death at embryonic day 13. TrxR2 ؊/؊ embryos are smaller and severely anemic and show increased apoptosis in the liver. The size of hematopoietic colonies cultured ex vivo is dramatically reduced. TrxR2-deficient embryonic fibroblasts are highly sensitive to endogenous oxygen radicals when glutathione synthesis is inhibited. Besides the defect in hematopoiesis, the ventricular heart wall of TrxR2 ؊/؊ embryos is thinned and proliferation of cardiomyocytes is decreased. Cardiac tissue-restricted ablation of TrxR2 results in fatal dilated cardiomyopathy, a condition reminiscent of that in Keshan disease and Friedreich's ataxia. We conclude that TrxR2 plays a pivotal role in both hematopoiesis and heart function.Reactive oxygen species (ROS)-generated mainly as a byproduct of the respiratory chain or by oxidases-are implicated in the pathogenesis and pathophysiology of a variety of human diseases such as cancer, cardiovascular, and degenerative disorders. A variety of cellular antioxidant systems control the balance of free intra-and extracellular oxygen radicals. Previous efforts have addressed the physiological role of superoxide dismutases, catalases, and glutathione (GSH) peroxidases in vivo, but the role of the thioredoxin/thioredoxin reductase/ peroxiredoxin system in ROS removal has only recently attracted attention.Thioredoxins are small redox-active proteins with an essential function in DNA metabolism and repair, transcription, and cell-cell communication (1). Acting through peroxiredoxins, they also efficiently protect cells from oxidative damage (27). Cytosolic (Trx1) and mitochondrial (Trx2) thioredoxins are required for proliferation and protection from apoptosis during early embryogenesis (26). Moreover, in chicken B cells, Trx2 is critically involved in the regulation of mitochondriondependent apoptosis (37). More recently, heart-specific overexpression of dominant-negative Trx1 was shown to be associated with increased oxidative stress and cardiac hypertrophy in mice (39).Trx activities are governed by thioredoxin reductases (TrxRs) that, in turn, use NADPH/H ϩ as the reducing agent (23). TrxRs are members of the pyridine nucleotide-disulfide oxidoreductase family, form homodimers, and possess two interacting redox-active centers. The C-terminal redox center contains a catalytically important selenocysteine (SeCys) (9,17,41). In mammals, three TrxRs...

The glutathione-dependent system is one of the key systems regulating cellular redox balance, and thus cell fate. Cysteine, typically present in its oxidized form cystine in the extracellular space, is regarded as the rate-limiting substrate for glutathione (GSH) synthesis. Cystine is transported into cells by the highly specific amino-acid antiporter system x c À . Since Burkitt's Lymphoma (BL) cells display limited uptake capacity for cystine, and are thus prone to oxidative stress-induced cell death, we stably expressed the substrate-specific subunit of system x c À , xCT, in HH514 BL cells. xCT-overexpressing cells became highly resistant to oxidative stress, particularly upon GSH depletion. Contrary to previous predictions, the increase of intracellular cysteine did not affect the cellular GSH pool, but concomitantly boosted extracellular cysteine concentrations. Even though cells were depleted of bulk GSH, xCT overexpression maintained cellular integrity by protecting against lipid peroxidation, a very early event in cell death progression. Our results show that system x c À protects against oxidative stress not by elevating intracellular GSH levels, but rather creates a reducing extracellular environment by driving a highly efficient cystine/cysteine redox cycle. Our findings show that the cystine/cysteine redox cycle by itself must be viewed as a discrete major regulator of cell survival.

In the present study the developmental profile of ATP-hydrolyzing activity promoted by NTPDase 1, its kinetic properties and the enzyme protein abundance associated with synaptic plasma membrane from rat cerebral cortex were characterized. NTPDase 1 activity increased from birth to day 30; afterwards it decreased and remained unchanged from adulthood (90 days) to senescence (365 days). Kinetic analysis revealed that enzyme exhibited the highest specific activity at day 30 and highest apparent affinity for ATP at day 365; however, V(max)/K(m) values remained unchanged for each age studied. Immunoblot analysis demonstrated that relative abundance of NTPDase 1 is highest at day 15 during ontogeny. The discrepancy between maximum enzyme activity and maximum enzyme protein abundance indicates that NTPDase 1 may have an additional role during development.

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