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Significance: The development and progression of renal diseases, including acute kidney injury (AKI) and chronic kidney disease (CKD), are the result of heterogeneous pathophysiology that reflects a range of environmental factors and, in a lesser extent, genetic mutations. The pathophysiology specific to most kidney diseases is not currently identified; therefore, these diseases are diagnosed based on non-pathological factors. For that reason, pathophysiology-based companion diagnostics for selection of pathophysiology-targeted treatments have not been available, which impedes personalized medicine in kidney disease. Recent Advances: Pathophysiology-targeted therapeutic agents are now being developed for the treatment of redox dysregulation. Redox modulation therapeutics, including bardoxolone methyl, suppresses the onset and progression of AKI and CKD. On the other hand, pathophysiology-targeted diagnostics for renal redox dysregulation are also being developed. Urinary thioredoxin (TXN) is a biomarker that can be used to diagnose tubular redox dysregulation. AKI causes oxidation and urinary excretion of TXN, which depletes TXN from the tubules, resulting in tubular redox dysregulation. Urinary TXN is selectively elevated at the onset of AKI and correlates with the progression of CKD in diabetic nephropathy. Critical Issues: Diagnostic methods should provide information about molecular mechanisms that aid in the selection of appropriate therapies to improve the prognosis of kidney disease. Future Directions: A specific diagnostic method enabling detection of redox dysregulation based on pathological molecular mechanisms is much needed and could provide the first step toward personalized medicine in kidney disease. Urinary TXN is a candidate for a companion diagnostic method to identify responders to redox-modulating therapeutics. Antioxid. Redox Signal. 36, 1051-1065.

Significance: The pathogenesis and progression of allergic inflammation in the respiratory system are closely linked to oxidative stress. Thioredoxin (TRX) is an essential redox balance regulator in organisms and is induced by various oxidative stress factors, including ultraviolet rays, radiation, oxidation, viral infections, ischemia reperfusion, and anticancer agents. Recent Advances: We demonstrated that systemic administration and transgenic overexpression of TRX is useful in a wide variety of in vivo inflammatory respiratory diseases models, such as viral pneumonia, interstitial lung disease, chronic obstructive pulmonary disease, asthma, acute respiratory distress syndrome, and obstructive sleep apnea syndrome, by removing reactive oxygen species, blocking production of inflammatory cytokines, inhibiting migration and activation of neutrophils and eosinophils, and regulating the cellular redox status. In addition, TRX's anti-inflammatory mechanism is different from the mechanisms associated with anti-inflammatory agents, such as glucocorticoids, which regulate the inflammatory reaction in association with suppressing immune responses. Critical Issues: Understanding the molecular mechanism of TRX is very helpful for understanding the role of TRX in respiratory diseases. In this review, we show the protective effect of TRX in various respiratory diseases. In addition, we discuss its anti-allergic and anti-inflammatory molecular mechanism in detail. Future Directions: The application of TRX may be useful for treating respiratory allergic inflammatory disorders. Antioxid. Redox Signal . 32, 785-801.

Aims: Accumulating evidence indicates that oxidative stress is associated with inflammation, and the cellular redox status can determine the sensitivity and the final outcome in response to inflammatory stimuli. To control the redox balance, mammalian cells contain a variety of oxidoreductases belonging to the thioredoxin superfamily. The large number of these enzymes suggests a complex mechanism of redox regulation in mammals, but the precise function of each family member awaits further investigations., Results: We generated mice deficient in transmembrane thioredoxin-related protein (TMX), a transmembrane oxidoreductase in the endoplasmic reticulum (ER). When exposed to lipopolysaccharide (LPS) and d-(+)-galactosamine (GalN) to induce inflammatory liver injury, mutant mice were highly susceptible to the toxicants and developed severe liver damage. LPS-induced production of inflammatory mediators was equivalent in both wild-type and TMX(-/-) mice, whereas neutralization of the proinflammatory cytokine tumor necrosis factor-α suppressed the toxic effects of LPS/GalN in the mutant mice. Liver transcriptional profiles revealed enhanced activation of the p53-signaling pathway in the TMX(-/-) mice after LPS/GalN treatment. Furthermore, TMX deficiency also caused increased sensitivity to thioacetamide, which exerts its hepatotoxicity through the generation of reactive oxygen species., Innovation: The present study is the first to address the role of the oxidoreductase TMX in inflammatory liver injury. The phenotype of mice deficient in TMX suggests a functional link between redox regulation in the ER and susceptibility to oxidative tissue damage., Conclusion: We conclude that TMX plays a major role in host defense under the type of inflammatory conditions associated with oxidative stress.

Thioredoxin (TRX) is a key component of redox regulation and has been indicated to play an essential role in cell survival and growth. Here, we investigated the molecular mechanism of TRX in the regulation of cell survival and growth by using RNA interference (RNAi) in A549 lung cancer and MCF7 breast cancer cells. TRX knockdown did not significantly increase the basal level of cell death without exposure to stress, but CDDP-induced cell death was enhanced. Meanwhile, TRX knockdown resulted in significant cell-cycle arrest at the G(1) phase. Cyclin D1 expression was reduced by TRX knockdown at the protein and mRNA levels. TRX knockdown caused suppression of activation of the cyclin D1 promoter through elements including AP-1. TRX knockdown also reduced the levels of phosphorylated ERK1/2 and the nuclear translocation of ERK 1/2 induced by EGF. These results suggest that TRX is an important regulator of the cell cycle in the G(1) phase via cyclin D1 transcription and the ERK/AP-1 signaling pathways.

Reactive oxygen species (ROS) are the key mediators of pathogenesis in cardiovascular diseases. Members of the thioredoxin superfamily take an active part in scavenging reactive oxygen species, thus playing an essential role in maintaining the intracellular redox status. The alteration in the expression levels of thioredoxin family members and related molecules constitute effective biomarkers in various diseases, including cardiovascular complications that involve oxidative stress. Thioredoxin, glutaredoxin, peroxiredoxin, and glutathione peroxidase, along with their isoforms, are involved in interaction with the members of metabolic and signaling pathways, thus making them attractive targets for clinical intervention. Studies with cells and transgenic animals have supported this notion and raised the hope for possible gene therapy as modern genetic medicine. Of all the molecules, thioredoxins, glutaredoxins, and peroxiredoxins are emphasized, because a growing body of evidence reveals their essential and regulatory role in several steps of redox regulation. In this review, we discuss some pertinent observations regarding their distribution, structure, functions, and interactions with the several survival- and death-signaling pathways, especially in the myocardium.

Thioredoxin binding protein 2 (TBP2) plays a regulatory role in lipid metabolism and immune regulation. We previously reported the effect of TBP2 loss-of-function on lipid metabolism using TBP2 knockout (TBP2KO) mice. In this study, we employed TBP2 transgenic (TBP2TG) mice to analyze the in vivo effect of TBP2 gain-of-function. We revealed a decrease in the percentage of hepatic natural killer T (NKT) cells in TBP2KO mice and an increase in the percentage of hepatic NKT cells in TBP2TG mice. The TBP2KO mice were resistant to concanavalin A (ConA)-induced hepatitis, but they were highly susceptible to other types of hepatitis. TBP2 modulates lipid metabolism as well as NKT cell activity. Moreover, TBP2 expression was increased significantly in klotho-deficient mice, which exhibit a syndrome resembling aging human phenotypes. TBP2 may play multiple roles in lipid metabolism, innate immunity, and aging.

Thioredoxin-1 (TRX) is a small (14 kDa) multifunctional protein with the redox-active site Cys-Gly-Pro-Cys. Macrophage migration inhibitory factor (MIF) is a 12 kDa cytokine belonging to the TRX family. Historically, when we purified TRX from the supernatant of ATL-2 cells, a 12 kDa protein was identified along with TRX, which was later proved to be MIF. Here, we show that TRX and MIF form a complex in the cell and the culture supernatant of ATL-2 cells. Using a BIAcore assay, we confirmed that TRX has a specific affinity with MIF. We also found that extracellular MIF was more effectively internalized into the ATL-2 cells expressing TRX on the cell surface, than the Jurkat T cells which do not express surface TRX. Moreover, anti-TRX antibody blocked the MIF internalization, suggesting that the cell surface TRX is involved in MIF internalization into the cells. Furthermore, anti-TRX antibody inhibited MIF-mediated enhancement of TNF-alpha production from macrophage RAW264.7 cells. These results suggest that the cell surface TRX serves as one of the MIF binding molecules or MIF receptor component and inhibits MIF-mediated inflammatory signals.

In nonalcoholic fatty liver disease, oxidative stress is believed to play a crucial role as a second-hit for the progression of simple steatosis to steatohepatitis. Thioredoxin (TRX) is a potent antioxidant molecule that exerts anti-apoptotic and anti-inflammatory functions. TRX-binding protein-2 (TBP-2) is an endogenous negative regulator of TRX. Deficiency of TBP-2 in mice causes hyperlipidemia, hepatic steatosis, hypoglycemia, and bleeding tendency, resembling Reye syndrome in a fasting/glucose-deficient state. The aim of this study was to investigate the role of TBP-2 in the development of nonalcoholic steatohepatitis (NASH). TBP-2-deficient (TBP-2(-/-)) and wild-type (WT) mice were fed either a normal or methionine-choline-deficient (MCD) diet for up to 10 weeks. Compared with WT mice, TBP-2(-/-) mice showed severe simple steatosis rather than steatohepatitis. However, oxidative stress determined by lipid peroxidation and DNA damage, neutrophil infiltration, and hepatic fibrosis were attenuated in TBP-2(-/-) mice. PCR analysis showed the expressions of fibrosis-inducing and inflammatory cytokine-related genes were less in TBP-2(-/-) mice. Moreover, leptin, SREBP1c, PPARgamma, and adipogenesis-lipogenesis-related genes were upregulated in TBP-2(-/-) mice. These results strongly suggested that TBP-2 might be involved in pathogenesis of NASH in WT mice, and inhibitors of TBP-2 could be useful in the prevention or treatment of NASH.

Thioredoxin, a redox-regulating protein that scavenges reactive oxygen species, appears to show an excellent antiinflammatory effect in treating animal models of various human inflammatory diseases. The aim of this study was to clarify whether thioredoxin is useful for treating inflammatory skin diseases, such as contact dermatitis, caused by epicutaneous exposure to environmental and occupational antigens. The allergic contact hypersensitivity response was suppressed in thioredoxin-transgenic mice. This suppressive effect of thioredoxin appeared to be via the inhibition of the efferent limb of contact hypersensitivity because administration of recombinant thioredoxin suppressed the inflammatory response in the elicitation phase but not in the induction phase. Adoptive-transfer studies revealed that the host environment, but not donor leukocytes, is critical in this suppressive effect. In thioredoxin-transgenic mice, the infiltration of neutrophils in the elicitation site was diminished, whereas the migratory function of cutaneous dendritic cells and hapten-specific cell proliferation were not disturbed. Thioredoxin-transgenic mice had also an attenuated inflammatory response to croton oil. These findings suggest that thioredoxin prevents skin inflammatory responses and could be a suitable candidate for the treatment of contact dermatitis.

Thioredoxin, a key molecule in redox regulation, and many redox enzymes are regulated through the antioxidant responsive element (ARE). To search for antioxidative constituents, we screened extracts from vegetables and found that the extracts of Perilla frutescens and Artemisia princeps have potent thioredoxin-inducing activities. By activity-guided purification of Perilla frutescens extracts, we identified perillaldehyde as a novel thioredoxin inducer. Fragrant unsaturated aldehydes, such as trans-cinnamaldehyde, safranal, 2,4-octadienal, citral, trans-2, cis-6-nonadienal, and trans-2-hexenal showed the ability to activate ARE. Perillaldehyde-induced activation through the ARE was suppressed by the overexpression of wild-type Keap1, whereas sulforaphane-induced activation seemed to be partially suppressed. Mutant Keap1 (R272A/K287A or C273A/C288A) did not suppress this activation. Pretreatment with perillaldehyde reduced the H(2)O(2)-induced cytotoxicity. Thus, we show that fragrant unsaturated aldehydes from edible plants are novel thioredoxin inducers and ARE activators and may be beneficial for protection against oxidative stress-induced cellular damage. These results also suggest that perillaldehyde activates the Nrf2-Keap1 system and that the lysine and arginine residues juxtaposed to the critical cysteine residues of Keap1 are required for signal sensing.

Obstructive sleep apnea (OSA) is associated with increased cardiovascular mortality, and oxidative stress was suggested to play an important role. We hypothesized that the plasma TRX level, a novel oxidative stress marker, is elevated in OSA patients. Plasma TRX and adiponectin levels, which are significantly associated with cardiovascular mortality, were measured in 41 patients with severe OSA before (n = 41) and after (n = 27) nasal continuous positive airway pressure therapy (nCPAP) for 1 month and in 12 subjects without OSA (non-OSA group). The TRX level was significantly higher (p = 0.02) and the adiponectin level was significantly lower (p = 0.02) in the OSA group than in the non-OSA group. After 1 month of nCPAP (n = 27), the TRX level significantly decreased (p = 0.03), and the adiponectin level significantly increased (p = 0.03). Among the 14 patients with untreated OSA, the TRX and adiponectin levels did not significantly change over a 1-month interval. Among the 53 (41 OSA + 12 non-OSA) subjects, the TRX level was positively correlated with the respiratory disturbance index (p = 0.001) and percentage of time with SaO(2) <90% (p = 0.0002). The adiponectin level, but not the TRX level, was correlated with the BMI (n = 53; p = 0.02). Plasma TRX may be a unique marker for evaluating oxidative stress and monitoring the effectiveness of nCPAP in OSA patients.

The authors previously established a transgenic mouse line in the type 1 diabetes model, NOD mouse, in which thioredoxin (TRX), a redox protein, is overexpressed in pancreatic beta cells, and found that TRX overexpression slows the progression of type 1 diabetes. Recent reports on type 2 diabetes suggest that oxidative stress also degrades the function of beta cells. To elucidate whether TRX overexpression can prevent progressive beta cell failure from oxidative stress in type 2 diabetes, the authors transferred the TRX transgene from the NOD mouse onto a mouse model of type 2 diabetes, the db/db mouse. The progression of hyperglycemia and the reduction of body weight gain and insulin content of the db/db mouse were significantly suppressed by the TRX expression. Furthermore, TRX suppressed the reduction of Pdx-1 and MafA expression in the beta cells, which may be one of the cellular mechanisms for protecting beta cells from losing their insulin-secreting capacity. These results showed that TRX can protect beta cells from destruction not only in type 1 but also in type 2 diabetes, and that they provide evidence that oxidative stress plays a crucial role in the deterioration of beta cell function during the progression of type 2 diabetes.

Thioredoxin-1 (TRX) plays important roles in cellular signaling by controlling the redox state of cysteine residues in target proteins. TRX is released in response to oxidative stress and shows various biologic functions from the extracellular environment. However, the mechanism by which extracellular TRX transduces the signal into the cells remains unclear. Here we report that the cysteine modification at the active site of TRX promotes the internalization of TRX into the cells. TRX-C35S, in which the cysteine at residue 35 of the active site was replaced with serine, was internalized more effectively than wild-type TRX in human T-cell leukemia virus-transformed T cells. TRX-C35S bound rapidly to the cell surface and was internalized into the cells dependent on lipid rafts in the plasma membrane. This process was inhibited by wild-type TRX, reducing reagents such as dithiothreitol, and methyl-beta-cyclodextrin, which disrupts lipid rafts. Moreover, the internalized TRX-C35S binds to endogenous TRX, resulting in the generation of intracellular reactive oxygen species (ROS) and enhanced cis-diamine-dichloroplatinum (II) (CDDP)-induced apoptosis via a ROS-mediated pathway involving apoptosis signal-regulating kinase-1 (ASK-1) activation. These findings suggest that the cysteine at the active site of TRX plays a key role in the internalization and signal transduction of extracellular TRX into the cells.

Human thioredoxin-1 (hTrx) exhibits a disulfide reducing activity and was originally identified as a soluble cytokine-like factor secreted from cells of a human T-cell leukemia virus type I (HTLV-I)-transformed cell line. Recent studies have revealed that endogenous Trx plays an important role in cytoprotection against various oxidative stress-associated disorders. However, the function of exogenous Trx is still not fully understood. We report here that a cysteine-modified mutant of recombinant human Trx (rhTrx-C35S) binds to human umbilical vein endothelial cells (HUVECs) as well as stimulated T cells and rapidly enters these cells via lipid rafts. In addition, we found that endogenous Trx is expressed on the surface of HUVECs, including lipid rafts. These events suggest cell-surface Trx as a possible target of rhTrx-C35S. Furthermore, we found that anti-human Trx mouse monoclonal antibody inhibits adherence of LPS-stimulated human peripheral blood polymorphonuclear cells (PMNs) to HUVECs. This adherence was also suppressed by a recombinant human Trx (rhTrx), but not by a mutant rhTrx (rhTrx-C32S/C35S) with no reducing activity. Cell-surface Trx may be involved in the process of interaction between PMNs and HUVECs and a possible target of cysteine-modified exogenous Trx as well as wild-type exogenous Trx through redox regulation.

Oxidative stress has been widely recognized to be involved in the pathogenesis of cardiopulmonary disorders. In ischemic heart diseases, it is involved not only in the development of atherosclerosis but also in ongoing ischemic injury, especially in the reperfusion process. Cardiomyopathy is another cardiac disorder in which oxidative stress is involved. In diabetic cardiomyopathy, homocysteine, a well-known source of oxidative stress, is believed to play major roles in its development. Thioredoxin (TRX) is a redox-acting protein ubiquitously present in the human body. It also is inducible by a wide variety of oxidative stresses. TRX is a multifunctional protein and has anti-inflammatory and antiapoptotic effects, as well as antioxidative effects. It is therefore feasible to think that TRX is a potential therapy for cardiac disease. Moreover, serum TRX is a well-recognized biomarker of various diseases involving oxidative stress, and this is also the case for cardiac disorders. Here we discuss how TRX is useful as a biomarker of and therapeutic agent for cardiopulmonary disorders, especially focusing on ischemic heart disease, myocarditis and oxygen sensing, and acute respiratory distress syndrome.

We show that 1-methyl-4-phenylpyridinium ion (MPP(+)), an active metabolite of 1-methyl-4-phenyl-1,2,3,6- tetrahydropyridine (MPTP), induces cytotoxicity via endoplasmic reticulum (ER)- and mitochondria-mediated pathways, and thioredoxin-1 (TRX-1), a redox-active protein, prevents MPTP-induced neurotoxicity. TRX-1 overexpression suppressed reactive oxygen species and the ATP decline caused by MPP(+) in HepG2 cells. MPP(+) activated caspase-12 in PC12 cells and induced cytotoxicity in HeLa-rho(0) cells lacking mitochondrial DNA, as well as in the parental HeLa-S3 cells. TRX-1-transgenic mice demonstrated significant resistance to caspase-12 activation and the apoptotic decrease of dopaminergic neurons after MPTP administration, compared with wild-type C57BL/6 mice.

Although the essential role of selenium for cellular immune responses is obvious, delineation of the functions is lacking because selenium can either promote or inhibit cell growth, cytokine production, and activation of transcription factor nuclear factor-kappaB (NF-kappaB). Studies with human thioredoxin-1 (Trx-1)-transgenic (Tg) mice were conducted to evaluate the relationship between stimulation of T-cell mitogenic response by sodium selenite and the intracellular Trx-1 levels, and the activities of selenoenzymes and NF-kappaB-DNA binding. Concanavalin A-induced mitogenesis of wild-type mouse splenic cells was stimulated by exposure to low levels of selenite (0.02-0.1 microM), with augmentation of NF-kappaB-DNA binding activity. Treatment with NF-kappaB nuclear translocation inhibitor SN50 or thioredoxin reductase (TR) inhibitor aurothioglucose depressed this stimulatory action. The mitogenic response of Trx-1-Tg mouse splenic cells was enhanced by exposure to relatively high levels of selenite (> or = 0.05 microM), compared with the wild-type mouse. Selenite also augmented TR activity but not cellular glutathione peroxidase activity in the Trx-1-overexpressed cells. These results suggest that the stimulation of T-cell mitogenic response by the physiological levels of selenite is predominantly caused by increased TR activity, which may lead to reduction of Trx-1 dependent on the intracellular expression level and promotion of DNA binding of NF-kappaB.

Thioredoxin-1 (TRX) is a small redox-active protein with antioxidative effects and redox-regulating functions. Cigarette smoking is a major etiological factor in the pathogenesis of a variety of diseases and recruits systemic immune and inflammatory responses. This report demonstrates that TRX attenuates the systemic inflammatory responses induced by cigarette smoking. The mRNA expressions of tumor necrosis factor alpha (TNF-alpha) and macrophage migration inhibitory factor (MIF) were suppressed in the spleen of TRX overexpressing transgenic mice (TRX-tg) exposed to cigarette smoking, compared with control C57BL/6 mice. In addition, protein carbonylation, a marker of cellular protein oxidation, was enhanced by cigarette smoking in the tissues of heart and liver in control mice more than in TRX-tg mice. These findings suggest that TRX may suppress the systemic inflammatory responses against cigarette smoking.

Oxidative stresses are largely mediated by intracellular protein oxidations by reactive oxygen species (ROS). Host cells are equipped with antioxidants that scavenge ROS. The cellular reduction/oxidation (redox) balance is maintained by ROS and antioxidants. Accumulating evidence suggests that the redox balance plays an important role in cellular signaling through the redox modification of cysteine residues in various important components of the signal transduction pathway. Thioredoxin (TRX) is a small protein playing important roles in cellular responses, including cell growth, cell cycle, gene expression, and apoptosis, to maintain the redox circumstance. Moreover, many recent papers have shown that the redox regulation by TRX is deeply involved in the pathogenesis of various oxidative stress-associated disorders. This review focuses on TRX and its related molecules, and discusses the role of TRX-dependent redox regulation in oxidative stress-induced signal transduction.

Chronic pancreatitis (CP) is considered to result from repetitive pancreatic injury, and sustained production of various proinflammatory cytokines and chemokines are closely involved in its pathogenesis. Monocyte chemoattractant protein 1 (MCP-1), a member of the CC chemokine family, is believed to contribute to the progression of CP through monocyte/macrophage recruitment. This study aimed to clarify the protective role of thioredoxin-1 (TRX-1), a redox-regulating protein with antioxidative activity, in MCP-1 production and pancreatic fibrosis using a CP model in transgenic mice overexpressing TRX-1 (TRX-1-TG mice) and wildtype C57BL/6 mice. Experimental CP was induced by repeated administration of cerulein and lipopolysaccharide for 6 weeks. In TRX-1-TG mice, pancreatic atrophy was ameliorated, and histologically detectable inflammatory cell infiltration, glandular atrophy, and pseudotubular complex formation were suppressed. Overexpression of TRX-1 also attenuated pancreatic fibrosis and suppressed the activation of pancreatic stellate cells. Serum levels of MCP-1 and pancreatic expression of transforming growth factor-beta, platelet-derived growth factor, and MCP-1 were reduced in TRX-1-TG mice compared with levels in wild-type mice. Overexpression of TRX-1 also reduced H(2)O(2)-induced MCP-1 production in isolated pancreatic acinar cells. These results indicate that TRX-1 can potentially attenuate pancreatic fibrosis via the suppression of oxidative stress and MCP-1-mediated chronic inflammation.

The lungs are the richest in oxygen among the various organs of the body and are always subject to harmful reactive oxygen species. Regulation of the reduction/oxidation (redox) state is critical for cell viability, activation, proliferation, and organ functions. Although the protective importance of various antioxidants has been reported, few antioxidants have established their clinical usefulness. Thioredoxin (TRX), a key redox molecule, plays crucial roles as an antioxidant and a catalyst in protein disulfide/dithiol exchange. TRX also modulates intracellular signal transduction and exerts antiinflammatory effects in tissues. In addition to its beneficial effects in other organs, the protective effect of TRX in the lungs has been shown against ischemia/ reperfusion injury, influenza infection, bleomycin-induced injury, or lethal inflammation caused by interleukin- 2 and interleukin-18. Monitoring of TRX in the plasma, airway, or lung tissue may be useful for the diagnosis and follow-up of pulmonary inflammation. Promotion/modulation of the TRX system by the administration of recombinant TRX protein, induction of endogenous TRX, or gene therapies can be a therapeutic modality for oxidative stress-associated lung disorders.

Increasing evidence has indicated that the modulation of intracellular redox states has important aspects to cellular events, such as cellular proliferation, activation, growth inhibition, or death via the regulation of intracellular signal transduction and gene expression. Thioredoxin (TRX) is a multifunctional stress-inducible protein, which protects cells from various types of stresses. TRX has not only a scavenging activity of reactive oxygen species, but also a regulating activity of various intracellular molecules including transcription factors. We demonstrated that the serum TRX levels are correlated with the severity of heart failure, and are negatively correlated with left ventricular ejection fractions of patients with heart failure. The expression of TRX is enhanced in endothelial cells and macrophages in human atherosclerotic plaques, in balloon-injured rat arteries, and in damaged cardiomyocytes of rats with acute myocarditis. Overexpression of TRX in transgenic mice attenuates adriamycin-induced cardiotoxicity by reducing oxidative stresses. These findings suggest that TRX and the redox system modulated by TRX have an important role in cellular defense against oxidative stress in cardiovascular diseases.

Reactive oxygen species are produced by various stressors derived from internal and external sources, including endogenous metabolic activities. Glucose metabolism is one of the most primitive sources for energy production for most cells; however, it may at the same time yield hazardous oxidative stress via simultaneous oxidant production. The protective mechanism against oxidative stress is thus an indispensable biological function. Recently, genetic mutation loci affecting life span were isolated from experimental model organisms, and several locus products were found to be closely linked with machinery either producing or defending oxidative stress. Thioredoxin (TRX) is a small protein having strong antioxiradical quenching capabilities and other multiple functions depending on the cellular redox state. In this review, we focus on the role of TRX in the aging process (senescence) as a redox-regulating molecule against oxidative stress. We also discuss the possibility of the TRX system serving as an index marker for cellular proliferation and senescence.

Transgenic (Tg) mice overexpressing human thioredoxin (TRX), a small redox-active protein, were produced to investigate the role of the protein in a variety of stresses. Bone marrow cells from TRX-Tg mice were more resistant to ultraviolet C-induced cytocide compared with those from wild type (WT) C57BL/6 mice. TRX-Tg mice exhibited extended median and maximum life spans compared with WT mice. Telomerase activity in spleen tissues in TRX-Tg mice was higher than that in WT mice. These results suggest that overexpression of TRX results in resistance against oxidative stress and a possible extension of life span without apparent abnormality in mammals.

Human immunodeficiency virus (HIV)-infected individuals are suffering from systemic oxidative stress. Reactive oxygen species act as second messengers for the activation of nuclear factor-kappaB (NF-kappaB), which augments the replication of HIV. Intracellular levels of glutathione (GSH), a major cytosolic antioxidant, in T cells decrease during the disease progression. Another redox-regulating molecule, thioredoxin (TRX), is also transiently down-regulated in the cells by acute HIV infection. In contrast, plasma levels of TRX are elevated in the late stage of HIV infection. Intracellular GSH and plasma TRX can be biomarkers to predict the prognosis of the disease. N-Acetylcysteine (NAC), a prodrug of cysteine that is necessary for GSH synthesis, has been used for HIV infection to prevent the activation of NF-kappaB and the replication of HIV. NAC shows some beneficial effects for HIV-infected individuals, although the intracellular GSH levels in lymphocytes are not significantly restored. The control of imbalanced redox status by antioxidants may be beneficial for the quality of life in HIV infection even in the era after the effective therapy with protease inhibitors has been applied. Redox control will be an important therapeutic strategy for oxidative stress-associated disorders including HIV infection.

Cellular redox is controlled by the thioredoxin (Trx) and glutathione (GSH) systems that scavenge harmful intracellular reactive oxygen species (ROS). Oxidative stress also evokes many intracellular events including apoptosis. There are two major pathways through which apoptosis is induced; one involves death receptors and is exemplified by Fas-mediated caspase-8 activation, and another is the stress- or mitochondria-mediated caspase-9 activation pathway. Both pathways converge on caspase-3 activation, resulting in nuclear degradation and cellular morphological change. Oxidative stress induces cytochrome c release from mitochondria and activation of caspases, p53, and kinases, including apoptosis signal-regulating kinase 1 (ASK1), c-Jun N-terminal kinase, and p38 mitogen-activated protein kinase. Trx inhibits apoptosis signaling not only by scavenging intracellular ROS in cooperation with the GSH system, but also by inhibiting the activity of ASK1 and p38. Mitochondria-specific thioredoxin (Trx-2) and Trx peroxidases (peroxiredoxins) are suggested to regulate cytochrome c release from mitochondria, which is a critical early step in the apoptotis-signaling pathway. dATP/ATP and reducing factors including Trx determine the manifestation of cell death, apoptosis or necrosis, by regulating the activation process and the activity of redox-sensitive caspases. As mitochondria are the most redox-active organelle and indispensable for cells to initiate or inhibit the apoptosis process, the regulation of mitochondrial function is the central focus in the research field of apoptosis and redox.

CD21 serves as a receptor for the Epstein-Barr virus (EBV). In this report, surface expression of CD21 on B and T cells was shown to be suppressed by a thiol-antioxidant, N-acetylcysteine (NAC), in a dose- and time-dependent manner. In contrast, expression of other surface markers, CD25 and CD4 for T cells and CD19 and surface IgM for B cells, was not affected by NAC. When an EBV-negative B-cell line B104 was treated with NAC, the cells were not susceptible to infection with B95-8-derived EBV. The effect of NAC was shown to be irrelevant to the transcriptional levels of CD21 mRNA and the intracellular glutathione levels. Immunoprecipitation study revealed that NAC causes a loss of anti-CD21 monoclonal antibody (HB5) binding to both membrane and soluble CD21, suggesting that NAC modulates the structure of CD21. Other thiol-antioxidants, such as 2-mercaptoethanol, pyrrolidine dithiocarbamate, and glutathione, showed similar effect to NAC on CD21 expression. These results suggest the possible modulation of EBV infection via thiol-dependent redox control of CD21, and thiol-antioxidants may be good candidates for controlling EBV infection.

Thioredoxin reductase (TR) is a flavoenzyme, containing one selenocysteine (Sec) residue at the penultimate carboxyl-terminus, that catalyzes the NADPH-dependent reduction of oxidized thioredoxin. Sec is encoded by the UGA stop codon in the open reading frame of the mRNA, and the conserved stem-loop structure in 3'-untranslated regions functions as the determinant of Sec incorporation instead of termination of translation. The efficiency of Sec incorporation in Sec-containing enzymes in physiological or selenium (Se)-deficient condition remains unclear. To clarify this, we have developed monoclonal antibodies to human TR, and established a sandwich enzyme-linked immunosorbent assay to determine TR protein content. We observed that the specific activity of cytosolic TR in NCI-H441 cells increased with increasing concentrations of Se in a serum-free medium. The specific activity of TR purified from each cytosol was essentially equal to the calculated specific activity of each cytosolic TR. The Se content of TR increased with increasing concentration of Se in the medium, from 0.32 mol/mol of TR subunit (no SE) to 0.98 mol/mol of TR subunit (500 nM Se), and was directly correlated with the specific activity of TR. When calculated from the cytosolic TR specific activity of human peripheral mononuclear cell, the theoretical efficiency of Sec incorporation in physiological conditions is assumed to be 87%.

Thioredoxin (TRX) is a redox-active protein induced by a variety of stress and secreted from cells. Collecting evidence revealed that extracellular TRX shows cytokine- and chemokine-like activities. In the present study, we studied the role of secreted TRX on cellular resistance to cis-diamminedichloroplatinum (II) (CDDP). The CDDP-resistant variants of HeLa cells not only have enhanced expression of intracellular TRX, but also show increased secretion of TRX into the culture medium, compared to the parental cells. The CDDP-resistant cells also exhibit an enhanced L-cystine uptake capability, which results in a significant increase in the intracellular sulfhydryl content, including glutathione (GSH). Exogenous administration of recombinant TRX (rTRX) increases cellular resistance to CDDP and augments the L-cystine uptake in the parental HeLa cells. Moreover, depletion of L-cystine from the culture medium or combined treatment with L-cystine uptake inhibitors increases cellular sensitivity to CDDP in the CDDP-resistant cells. These findings suggest that secreted TRX may play an important role in the acquisition of cellular CDDP resistance through enhancement of the L-cystine uptake activity, and that the L-cystine transport system, as well as the TRX system, may be a novel therapeutic target in CDDP-resistant cancer cells.

We have investigated the thioredoxin (TRX) levels in severely burned patients and the possible origin of TRX, based on the recent understanding that TRX is a potent antioxidant with cytoprotective functions. Serum and plasma samples from burns patients and healthy blood donors were collected during the first 10 post-burn days and analyzed in a sandwich TRX enzyme-linked immunosorbent assay (ELISA). The TRX levels found were correlated to a panel of blood tests. The presence of TRX in platelets was investigated by immunoelectron microscopy and Western blotting. TRX serum levels of the severely burned patients showed a significant increase, with a mean serum TRX concentration on the day of injury of 76.5 +/- 19.5 ng/ml (mean +/- SD) and on post-burn day one 122.6 +/- 66.9 ng/ml, compared to control blood donor levels of 22.7 +/- 12.2 ng/ml (p = 0.0041 and 0.0117, respectively). A second peak of increase was found on post-burn days 7 to 9 with a four- to five-fold rise in concentration compared to controls. TRX elevation correlated well with increased platelet (p = 0.007) and leukocyte counts (p = 0.002). We also demonstrated by immunoelectron microscopy and Western blotting the presence of TRX in platelets. In conclusion, our demonstration of TRX release in burn injuries indicates that the TRX system is involved in a rapid antioxidant defense, coagulation processes, cell growth, and control of the extracellular peroxide tone intimately linked to cytoprotection and wound healing in burns. One of the cell types that delivers TRX promptly and efficiently into the blood may be the platelet.

Although oxygen is essential for promoting energy metabolism and for enhancing cell proliferation, early mouse embryos are very sensitive to high oxygen concentration. Because the tissue-specificity and sequential changes of the expression of antioxidative enzymes in rodent embryos have not been investigated systematically, we examined the ontogenesis of thioredoxin (TRX) and glutaredoxin (GRX) in mouse embryos and fetuses by using immunohistochemical methods. These compounds were found to be localized in most tissues examined, with some tissue specificity and temporal sequence. In many tissues, both TRX and GRX began to be expressed at embryonic day 11 (E11) or E13 and appeared to increase later in development, but the heart and great vessels of E8.5 embryos were already positive for their immunoreactivity. The stage at which the antioxidative enzymes begin to be expressed seems to coincide with the stage at which rodent embryos acquire the capacity of aerobic energy metabolism. Although TRX and GRX were co-localized in many tissues and showed similar sequential changes of expression, their expression patterns were different in the fetal cartilage, suggesting that they may play different roles in endochondral ossification. Their immunoreactivity was not homogeneous in the liver and the epithelium of uriniferous tubules, probably because their expression is associated with the proliferating and metabolic activities of the cell, as suggested by previous investigators. These results suggest that TRX and GRX play some tissue-specific roles in mammalian morphogenesis as well as general roles as antioxidant enzymes.

Thioredoxin (TRX), a thiol-containing protein, is induced by various oxidative stresses. Serum TRX levels were measured with a sandwich enzyme-linked immunosorbent assay kit in 210 hepatitis C virus (HCV)-infected patients, 39 hepatitis B virus (HBV)-infected patients, and 17 healthy volunteers. The effects of hepatoprotective drugs on TRX levels were also examined. The median TRX levels were significantly higher in HCV-infected patients than in controls (34.2 vs. 23.5 ng/ml, respectively; p < 0.005), but were not elevated in HBV-infected patients (26.7 ng/ml). The TRX levels were significantly correlated with serum lipid peroxide levels and indocyanine green exclusion test values, and were markedly decreased following treatment with Stronger Neo-Minophagen C or ursodeoxycholic acid. In conclusion serum TRX levels, a marker of oxidative stress, were higher in patients with HCV infection than those with HBV infection and healthy controls. The therapeutic efficacy of hepatoprotective drugs may be connected with the decrease in oxidative stress in hepatitis patients.

Gene induction by tumor necrosis factor-alpha (TNFalpha) or interleukin-1beta (IL-1beta) is mediated in part by activation of the transcription factor nuclear factor kappaB (NF-kappaB), and requires signal adaptor molecules such as TNF receptor-associated factor (TRAFs). The latter interact with the NF-kappaB-inducing kinase (NIK), which is believed to be part of the IkappaB kinase complex. Although the precise mechanism is to be elucidated, it is well-known that antioxidant treatments inhibit the inflammatory cytokine-induced NF-kappaB activation. Thioredoxin (TRX) is a 12-kDa endogenous protein that regulates various cellular functions by modulating the redox state of proteins, overexpression of this molecule inhibits NF-kappaB activation. To elucidate the roles of TRX in the signal transduction of the cytokines, we investigated the effects of TRX on NF-kappaB activation induced by cytokine treatment or by overexpression of the signaling molecules. Our data show that TRX treatment inhibits NF-kappaB-dependent transcription at the level of downstream of TRAFs and upstream of NIK: TRX inhibited TRAF2-, TRAF5-, and TRAF6-induced NF-kappaB activation but does not inhibit NIK-, IKKalpha-, and MEKK-induced activation. In addition, we show that TRX inhibits NF-kappaB activation in a manner different from that for SAPK (stress activated protein kinase) inhibition.

Human thioredoxin (hTRX)/adult T-cell leukemia (ATL)-derived factor (ADF) was originally reported as an interleukin-2 (IL-2) receptor-alpha-inducing factor produced by human T-cell lymphotropic virus-1-positive (HTLV-1+) cell lines. Growing evidence indicates that hTRX/ADF plays important roles in cellular responses against oxidative stress and is involved in a variety of cellular functions. A high level of hTRX/ADF expression is also observed in other human virus-infected cell lines including those of Epstein-Barr virus (EBV) and human papillomavirus. In this report, we analyzed the effect of hTRX/ADF on lytic amplification and persistent replication of EBV as a model for lytic versus latent phase of viral replication in host cells. Addition of hTRX/ADF clearly suppressed lytic replication of EBV in Raji cells and B95-8 cells induced to the lytic phase of 12-O-tetradecanoylphorbol-13-acetate (TPA), and it prevented the death of these cells evoked by the lytic induction. In contrast, hTRX/ADF did not have any effect on persistent replication in the latent phase. These data indicated that hTRX/ADF prevents EBV-transformed cells from proceeding into the lytic phase and regulates cohabitation of EBV and its host cells.