IInvestigation of possible oxidative damage caused by formaldehyde exposure in the rat's heart and aorta tissue / Investigación de posible daño oxidativo causado por la exposición al formaldehído en el corazón de la rata y tejido de la aorta

SUMMARY: Formaldehyde (FA) is a toxic substance used frequently in the field of medicine as well as in many industrial areas. Especially people working in the field of anatomy, histology, and pathology are in high risk group because of the use of the FA. Studies showing the effects of FA on the cardiovascular system are few in number. The purpose of the present study was to investigate the effects of FA exposure, which we believe can cause oxidative stress, on the heart and aorta with various biochemical analyses. A total of 24 Wistar Albino rats were used in our study. We divided the rats into 3 groups as the Control Group (CG), the group exposed to low-dose FA (avg. 1 ppm) (DDG) Group, and the group exposed to high-dose FA (avg. 10 ppm) (YDG). At the end of the subchronic FA exposure, the blood samples, heart and aorta tissues of the rats were taken and subjected to biochemical analyses. As a result of the analyses, statistically significant differences were detected between CG (2.96?0.85 ng/mg), and HDG (2.08?0.77 ng/mg) in aortic tissues in TXNIP analysis (p<0.05). In heart tissues, significant differences were detected between CG (0.73?0.27 ng/mg) and LDG (1.13?0.22 ng/mg) (p<0.05). Statistically significant differences were also detected between CG (1.98?0.31 mM/ml) and YDG (2.43?0.31 mM/ml) in serum MDA analyses (p<0.05). It was shown that subchronic application of FA to LDG rats through inhalation had no effects on apoptosis markers in heart tissues. More studies are required to show FA toxicity and the mechanism of action of pathology on the cardiovascular system. We believe that our study will contribute to clarifying the roles of mild and subchronic exposure of FA in heart and aortic tissues in terms of oxidative stress risk.

RESUMEN: El formaldehído es una sustancia tóxica que se utiliza con frecuencia en el campo de la medicina, así como en muchas áreas industriales. Especialmente las personas que trabajan en el area de la anatomía, y patología se encuentran en el grupo de alto riesgo debido al uso de esta sustancia. Pocos son los estudios que muestran los efectos del formaldehído en el sistema cardiovascular. El propósito del presente estudio fue investigar a través de análisis bioquímicos, los efectos de la exposición a formaldehído, que podría causar estrés oxidativo, en el corazón y la aorta. Se utilizaron un total de 24 ratas Albinas Wistar. Dividimos a las ratas en 3 grupos: grupo control (GC), grupo expuesto a dosis bajas de AG (promedio 1 ppm) (DDG) y grupo expuesto a dosis altas de AG (promedio 10 ppm) (YDG). Al término de la exposición a FA subcrónica, se tomaron muestras de sangre, tejido cardíaco y aorta de las ratas y se sometieron a análisis bioquímicos. Como resultado de los análisis, se detec- taron diferencias estadísticamente significativas entre GC (2,96 ? 0,85 ng / mg) y HDG (2,08 ? 0,77 ng / mg) en los tejidos aórticos en el análisis TXNIP (p <0,05). En los tejidos cardíacos se detectaron diferencias significativas entre GC (0,73 ? 0,27 ng / mg) y LDG (1,13 ? 0,22 ng / mg) (p <0,05). También se detectaron diferencias estadísticamente significativas entre CG (1,98 ? 0,31 mM / ml) y YDG (2,43 ? 0,31 mM / ml) en los análisis de MDA en suero (p <0,05). Se demostró que la aplicación subcrónica de formaldehído a ratas LDG a través de la inhalación no tuvo efectos sobre los marcadores de apoptosis en los tejidos del corazón. Se requieren más estudios para demostrar la toxicidad de los AG y el mecanismo de acción de la patología en el sistema cardiovascular. Creemos que nuestro estudio contribuirá a aclarar las funciones de la exposición leve y subcrónica de formaldehído en los tejidos cardíacos y aórticos en términos de riesgo al estrés oxidativo.

Thioredoxin 1 is upregulated in the bone and bone marrow following experimental myocardial infarction: evidence for a remote organ response.

Ischemia and reperfusion events, such as myocardial infarction (MI), are reported to induce remote organ damage severely compromising patient outcomes. Tissue survival and functional restoration relies on the activation of endogenous redox regulatory systems such as the oxidoreductases of the thioredoxin (Trx) family. Trxs and peroxiredoxins (Prxs) are essential for the redox regulation of protein thiol groups and for the reduction of hydrogen peroxide, respectively. Here, we determined whether experimental MI induces changes in Trxs and Prxs in the heart as well as in secondary organs. Levels and localization of Trx1, TrxR1, Trx2, Prx1, and Prx2 were analyzed in the femur, vertebrae, and kidneys of rats following MI or sham surgery. Trx1 levels were significantly increased in the heart (P = 0.0017) and femur (P < 0.0001) of MI animals. In the femur and lumbar vertebrae, Trx1 upregulation was detected in bone-lining cells, osteoblasts, megakaryocytes, and other hematopoietic cells. Serum levels of Trx1 increased significantly 2 days after MI compared to sham animals (P = 0.0085). Differential regulation of Trx1 in the bone was also detected by immunohistochemistry 1 month after MI. N-Acetyl-cysteine treatment over a period of 1 month induced a significant reduction of Trx1 levels in the bone of MI rats compared to sham and to MI vehicle. This study provides first evidence that MI induces remote organ upregulation of the redox protein Trx1 in the bone, as a response to ischemia-reperfusion injury in the heart.

Relationships between diabetic nephropathy and insulin resistance, inflammation, Trx, Txnip, CysC and serum complement levels.

OBJECTIVE: To investigate the relationships between diabetic nephropathy (DN) and insulin resistance, inflammation, thioredoxin (Trx), thioredoxin-interacting protein (Txnip), Cystatin C (CysC) and serum complement levels. PATIENTS AND METHODS: A total of 119 patients with type 2 diabetes mellitus (T2DM) treated in the Endocrinology Department of our hospital from January 2017 to December 2017 were enrolled as the experiment group, while 30 healthy volunteers were selected as the control group. The expression levels of inflammatory factors, Trx, Txnip, CysC and serum complements in every subject were detected. In addition, the type 2 diabetic nephropathy rat model was established via high-fat diet and injection of low-dose streptozotocin. Blood glucose, insulin resistance indexes and 24h-urinary albumin excretion were measured, and the histomorphological characteristics of the kidney in animals were observed. RESULTS: In clinical subjects, Trx level was notably lower in the simple DM group, early DN group and clinical DN group in comparison with that in the control group. The levels of Txnip and CysC in the simple DM group, early DN group and clinical DN group were remarkably higher than those in the control group. Moreover, the expression levels of TNF-α and IL-6 in the clinical DN group were significantly elevated compared with those in the simple DM group and early DN group. In addition, C1q expression in the clinical DN group was higher than that in the simple DM group and early DN group. In model rats, HOMA-IR was distinctly higher in the DM group and DN group than that in the control group. The ratio of kidney weight to body weight (KW/BW) was evidently higher in the DN group in comparison with that in the control group and DM group. CONCLUSIONS: Insulin resistance, inflammatory factors, and levels of Trx, Txnip, CysC and serum complement C1q are related to the progression of DM.

Redox-dependent and independent effects of thioredoxin interacting protein.

Thioredoxin interacting protein (TXNIP) is an important physiological inhibitor of the thioredoxin (TXN) redox system in cells. Regulation of TXNIP expression and/or activity not only plays an important role in redox regulation but also exerts redox-independent physiological effects that exhibit direct pathophysiological consequences including elevated inflammatory response, aberrant glucose metabolism, cellular senescence and apoptosis, cellular immunity, and tumorigenesis. This review provides a brief overview of the current knowledge concerning the redox-dependent and independent roles of TXNIP and its relevance to various disease states. The implications for the therapeutic targeting of TXNIP will also be discussed.

Reactive oxygen species-regulating proteins peroxiredoxin 2 and thioredoxin, and glyceraldehyde-3-phosphate dehydrogenase are differentially abundant in induced sputum from smokers with lung cancer or asbestos exposure.

Lung cancer is a deadly disease, typically caused by known risk factors, such as tobacco smoke and asbestos exposure. By triggering cellular oxidative stress and altering the antioxidant pathways eliminating reactive oxygen species (ROS), tobacco smoke and asbestos predispose to cancer. Despite easily recognizable high-risk individuals, lung cancer screening and its early detection are hampered by poor diagnostic tools including the absence of proper biomarkers. This study aimed to recognize potential lung cancer biomarkers using induced sputum noninvasively collected from the lungs of individuals in risk of contracting lung cancer. Study groups composed of current and former smokers, who either were significantly asbestos exposed, had lung cancer, or were unexposed and asymptomatic. Screening of potential biomarkers was performed with 52, and five differentially abundant proteins, peroxiredoxin 2 (PRDX2), thioredoxin (TXN), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), extracellular matrix protein 1 (ECM1), and protein S100 A8 (S100A8), were chosen to undergo validation, for their previously known connection with oxidative stress or cancer. Results from the validation in 123 sputa showed that PRDX2, TXN, and GAPDH were differentially abundant in sputa from individuals with lung cancer. TXN had a negative correlation with asbestos exposure, yet a positive correlation with smoking and lung cancer. Thus, tobacco smoking, asbestos exposure, and lung carcinogenesis may disturb the cellular redox state in different ways. A strong correlation was found among PRDX2, TXN, GAPDH, and S100A8, suggesting that these proteins may present a diagnostic biomarker panel to aid recognizing individuals at high risk of contracting lung cancer.

Melatonin inhibits endoplasmic reticulum stress-associated TXNIP/NLRP3 inflammasome activation in lipopolysaccharide-induced endometritis in mice.

Endometritis, an inflammatory response of the uterus tissue, is characterized by the production of inflammatory cytokines and migration of neutrophil (PMN) into the uterus tissue. Melatonin has been demonstrated to have anti-inflammatory and antioxidant effects. The purpose of this study was to investigate the protective effects of melatonin on lipopolysaccharide (LPS)-induced endometritis in mice. An endometritis model was induced by LPS and melatonin was given 1 h before LPS treatment. The results showed that melatonin inhibited LPS-induced pathologic changes, Myeloperoxidase (MPO) activity, and levels of interleukin-1 beta (IL-1ß). Melatonin also inhibited LPS-induced thioredoxin-interacting protein (TXNIP)/NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome and nuclear factor kappa B (NF-κB) activation, reactive oxygen species (ROS) production, and endoplasmic reticulum (ER) stress. Furthermore, melatonin was found to increase AMPK activity. In conclusion, our results demonstrated that melatonin inhibited ER stress-associated TXNIP/NLRP3 inflammasome activation with a regulation of adenosine monophosphate activated protein kinase (AMPK) in LPS-induced endometritis. Melatonin may serve as a promising nutritional supplement for the treatment of endometritis.

Balanced Regulation of Redox Status of Intracellular Thioredoxin Revealed by in-Cell NMR.

To understand how intracellular proteins respond to oxidative stresses, the redox status of the target protein, as well as the intracellular redox potential ( EGSH), which is defined by the concentrations of reduced and oxidized glutathione, should be observed simultaneously within living cells. In this study, we developed a method that can monitor the redox status of thioredoxin (Trx) and EGSH by direct NMR observation of Trx and glutathione within living cells. Unlike the midpoint potential of Trx measured in vitro (∼ -300 mV), the intracellular Trx exhibited the redox transition at EGSH between -250 and -200 mV, the range known to trigger the oxidative stress-mediated signalings. Furthermore, we quantified the contribution of Trx reductase to the redox status of Trx, demonstrating that the redox profile of Trx is determined by the interplay between the elevation of EGSH and the reduction by Trx reductase and other endogenous molecules.

Thioredoxin-1 Negatively Modulates ADAM17 Activity Through Direct Binding and Indirect Reductive Activity.

AIMS: A disintegrin and metalloprotease 17 (ADAM17) modulates signaling events by releasing surface protein ectodomains such as TNFa and the EGFR-ligands. We have previously characterized cytoplasmic thioredoxin-1 (Trx-1) as a partner of ADAM17 cytoplasmic domain. Still, the mechanism of ADAM17 regulation by Trx-1 is unknown, and it has become of paramount importance to assess the degree of influence that Trx-1 has on metalloproteinase ADAM17. RESULTS: Combining discovery and targeted proteomic approaches, we uncovered that Trx-1 negatively regulates ADAM17 by direct and indirect effect. We performed cell-based assays with synthetic peptides and site-directed mutagenesis, and we demonstrated that the interaction interface of Trx-1 and ADAM17 is important for the negative regulation of ADAM17 activity. However, both Trx-1K72A and catalytic site mutant Trx-1C32/35S rescued ADAM17 activity, although the interaction with Trx-1C32/35S was unaffected, suggesting an indirect effect of Trx-1. We confirmed that the Trx-1C32/35S mutant showed diminished reductive capacity, explaining this indirect effect on increasing ADAM17 activity through oxidant levels. Interestingly, Trx-1K72A mutant showed similar oxidant levels to Trx-1C32/35S, even though its catalytic site was preserved. We further demonstrated that the general reactive oxygen species inhibitor, Nacetylcysteine (NAC), maintained the regulation of ADAM17 dependent of Trx-1 reductase activity levels; whereas the electron transport chain modulator, rotenone, abolished Trx-1 effect on ADAM17 activity. INNOVATION: We show for the first time that the mechanism of ADAM17 regulation, Trx-1 dependent, can be by direct interaction and indirect effect, bringing new insights into the cross-talk between isomerases and mammalian metalloproteinases. CONCLUSION: This unexpected Trx-1K72A behavior was due to more dimer formation and, consequently, the reduction of its Trx-1 reductase activity, evaluated through dimer verification, by gel filtration and mass spectrometry analysis. Antioxid. Redox Signal. 29, 717-734.

Mitochondria Targeted and Intracellular Biothiol Triggered Hyperpolarized 129Xe Magnetofluorescent Biosensor.

Biothiols such as gluthathione (GSH), cysteine (Cys), homocysteine (Hcy), and thioredoxin (Trx) play vital roles in cellular metabolism. Various diseases are associated with abnormal cellular biothiol levels. Thus, the intracellular detection of biothiol levels could be a useful diagnostic tool. A number of methods have been developed to detect intracellular thiols, but sensitivity and specificity problems have limited their applications. To address these limitations, we have designed a new biosensor based on hyperpolarized xenon magnetic resonance detection, which can be used to detect biothiol levels noninvasively. The biosensor is a multimodal probe that incorporates a cryptophane-A cage as 129Xe NMR reporter, a naphthalimide moiety as fluorescence reporter, a disulfide bond as thiol-specific cleavable group, and a triphenylphosphonium moiety as mitochondria targeting unit. When the biosensor interacts with biothiols, disulfide bond cleavage leads to enhancements in the fluorescence intensity and changes in the 129Xe chemical shift. Using Hyper-CEST (chemical exchange saturation transfer) NMR, our biosensor shows a low detection limit at picomolar (10-10 M) concentration, which makes a promise to detect thiols in cells. The biosensor can detect biothiol effectively in live cells and shows good targeting ability to the mitochondria. This new approach not only offers a practical technique to detect thiols in live cells, but may also present an excellent in vivo test platform for xenon biosensors.

The paradoxical role of thioredoxin on oxidative stress and aging.

In spite of intensive study, there is still controversy about the free radical or oxidative stress theory of aging, particularly in mammals. Our laboratory has conducted the first detailed studies on the role of thioredoxin (Trx) in the cytosol (Trx1) and in mitochondria (Trx2) on oxidative stress and aging using unique mouse models either overexpressing or down-regulating Trx1 or Trx2. The results generated from our lab and others indicate that: (1) oxidative stress and subsequent changes in signaling pathways could have different pathophysiological impacts at different stages of life; (2) changes in redox-sensitive signaling controlled by levels of oxidative stress and redox state could play more important roles in pathophysiology than accumulation of oxidative damage; (3) changes in oxidative stress and redox state in different cellular compartments (cytosol, mitochondria, or nucleus) could play different roles in pathophysiology during aging, and their combined effects show more impact on aging than changes in either oxidative stress or redox state alone; and (4) the roles of oxidative stress and redox state could have different pathophysiological consequences in different organs/tissues/cells or pathophysiological conditions. To critically test the role of oxidative stress on aging and investigate changes in redox-sensitive signaling pathways, further study is required.

Subcellular localization of six thioredoxins and their antioxidant activity and contributions to biological control potential in Beauveria bassiana.

Thioredoxins (Trx) can detoxify sulfide or act as electron donors in the reduction of disulfide and dithiol to protect yeast cells from ROS damage but remain poorly explored in filamentous fungi. Here we show more Trx homologs in Beauveria bassiana than in many other fungi and examine their functions. This filamentous entomopathogen has six Trx homologs, including four (Txr1-4) evidently localized in cytoplasm, one (Trx5) in nuclear membrane and another (Trx6) in mitochondria. Deletion of each trx had no effect on radial growth on rich or minimal medium but resulted in remarkable transcriptional up-regulation of other partners for compensation. Compared with wild-type, only Δtrx2 was significantly more sensitive to menadione whereas none of six Δtrx mutants was responsive to other oxidants including H2O2. Intriguingly, Δtrx2 showed uniquely a significant increase in total Trx activity in normal cultures but a remarkable decrease in total SOD activity in the cultures grown normally or co-cultivated with menadione. The ratio of reduced/oxidized glutathione accumulated in hyphal cells stressed with menadione decreased to only 0.4 in Δtrx2 from ∼1.0 observed in wild-type and other mutants. The six Δtrx mutants displayed one or more phenotypic changes associated with the fungal biocontrol potential, including conidiation, and germination, thermotolerance, UV-B resistance and virulence of their conidia. All the changes were restored by trx complementation. Taken together, the greater Trx diversity evolutionarily gained by B. bassiana could help it to maintain cellular redox homeostasis and infect insect hosts in diverse habitats.

Reactive oxygen species, glutathione, and thioredoxin influence suberoyl bishydroxamic acid-induced apoptosis in A549 lung cancer cells.

Suberoyl bishydroxamic acid (SBHA) as a histone deacetylase (HDAC) inhibitor can induce apoptosis through the formation of reactive oxygen species (ROS). However, there is no report about the regulation of ROS and antioxidant enzymes in SBHA-treated lung cancer cells. Here, we investigated the toxicological effects of SBHA on the regulations of ROS, glutathione (GSH), and antioxidant enzymes, especially thioredoxin (Trx) in A549 lung cancer cells. SBHA inhibited the growth of A549 cells in time- and dose-dependent manners, and it induced apoptosis which accompanied by the loss of mitochondrial membrane potential (MMP; ΔΨm). SBHA significantly increased ROS levels including O2 (•-) level at 72 h whereas it decreased ROS levels at the early time points (30 min to 3 h). SBHA also induced GSH depletion at 24 and 72 h. N-acetyl cysteine (NAC; a well-known antioxidant) prevented apoptotic cell death and GSH depletion via decreasing ROS in SBHA-treated A549 cells. In addition, SBHA changed the levels of antioxidant-related proteins, especially Trx1. The expression and activity of Trx1 in A549 cells were reduced by SBHA. While the downregulation of Trx1 enhanced cell death, ROS level, and GSH depletion in SBHA-treated A549 cells, the overexpression of Trx1 decreased ROS level in these cells without the prevention of cell death and GSH depletion. In conclusion, SBHA-induced A549 cell death was influenced by changes in ROS and GSH levels. The basal status of Trx1 among other antioxidant proteins was closely correlated with the survival of A549 cells.

Delayed administration of alpha-difluoromethylornithine prevents hippocampus-dependent cognitive impairment after single and combined injury in mice.

Radiation exposure due to radiological terrorism and military circumstances are a continuing threat for the civilian population. In an uncontrolled radiation event, it is likely that there will be other types of injury involved, including trauma. While radiation combined injury is recognized as an area of great significance, overall there is a paucity of information regarding the mechanisms underlying the interactions between irradiation and other forms of injury, or what countermeasures might be effective in ameliorating such changes. The objective of this study was to determine if difluoromethylornithine (DFMO) could reduce the adverse effects of single or combined injury if administered beginning 24 h after exposure. Eight-week-old C57BL/J6 young-adult male mice received whole-body cesium-137 ((137)Cs) irradiation with 4 Gy. Immediately after irradiation, unilateral traumatic brain injury was induced using a controlled cortical impact system. Forty-four days postirradiation, animals were tested for hippocampus-dependent cognitive performance in the Morris water maze. After cognitive testing, animals were euthanized and their brains snap frozen for immunohistochemical assessment of neuroinflammation (activated microglia) and neurogenesis in the hippocampal dentate gyrus. Our data show that single and combined injuries induced variable degrees of hippocampus-dependent cognitive dysfunction, and when given 24 h post trauma, DFMO treatment ameliorated those effects. Cellular changes including neurogenesis and numbers of activated microglia were generally not associated with the cognitive changes. Further analyses also revealed that DFMO increased hippocampal protein levels of the antioxidants thioredoxin 1 and peroxiredoxin 3 compared to vehicle treated animals. While the mechanisms responsible for the improvement in cognition after DFMO treatment are not yet clear, these results constitute a basis for further development of DFMO as a countermeasure for ameliorating the of risks for cognitive dysfunction in individuals subjected to trauma and radiation combined injury.

Expression of thioredoxins and glutaredoxins in human hepatocellular carcinoma: correlation to cell proliferation, tumor size and metabolic syndrome.

Thioredoxins (Trx) and glutaredoxins (Grx) are thiol oxidoreductases that are ubiquitously expressed, and are involved in several biological processes. The expression of thioredoxins and glutaredoxins is induced in many neoplasms, and correlates with prognosis in gallbladder and colorectal carcinoma. The aim of the present study was to examine the expression pattern of these proteins (redoxins) in hepatocellular carcinoma (HCC) and to correlate their levels with clinical features. Paraffin-embedded tissues from 25 patients resected for HCC and 15 patients resected for colorectal carcinoma (CRC) liver metastases were analyzed with immunohistochemistry. Our results showed that Trx1, Trx2 and Grx5 were upregulated in HCCs as compared to the respective surrounding liver. In comparison, almost all redoxins were upregulated in CRC liver metastases, with Trx1 and Grx3 being significantly more increased in the CRC liver metastases than in the primary HCC tumors. In HCC, Trx1 correlated significantly with cell proliferation, and with a trend towards increased levels with micro-vascular invasion, while expression of Trx2 decreased with tumor size. Trx1 levels were lower in tumors of males, smokers, and patients with high alcohol consumption. Grx2 levels were significantly higher in patients with metabolic syndrome. In conclusion, this study illustrates specific correlations of individual redoxins to clinical features of HCC, and implicates the redoxins in the pathogenesis of HCC.

Hyaluronan and N-ERC/mesothelin as key biomarkers in a specific two-step model to predict pleural malignant mesothelioma.

PURPOSE: Diagnosis of malignant mesothelioma is challenging. The first available diagnostic material is often an effusion and biochemical analysis of soluble markers may provide additional diagnostic information. This study aimed to establish a predictive model using biomarkers from pleural effusions, to allow early and accurate diagnosis. PATIENTS AND METHODS: Effusions were collected prospectively from 190 consecutive patients at a regional referral centre. Hyaluronan, N-ERC/mesothelin, C-ERC/mesothelin, osteopontin, syndecan-1, syndecan-2, and thioredoxin were measured using ELISA and HPLC. A predictive model was generated and validated using a second prospective set of 375 effusions collected consecutively at a different referral centre. RESULTS: Biochemical markers significantly associated with mesothelioma were hyaluronan (odds ratio, 95% CI: 8.82, 4.82-20.39), N-ERC/mesothelin (4.81, 3.19-7.93), CERC/mesothelin (3.58, 2.43-5.59) and syndecan-1 (1.34, 1.03-1.77). A two-step model using hyaluronan and N-ERC/mesothelin, and combining a threshold decision rule with logistic regression, yielded good discrimination with an area under the ROC curve of 0.99 (95% CI: 0.97-1.00) in the model generation dataset and 0.83 (0.74-0.91) in the validation dataset, respectively. CONCLUSIONS: A two-step model using hyaluronan and N-ERC/mesothelin predicts mesothelioma with high specificity. This method can be performed on the first available effusion and could be a useful adjunct to the morphological diagnosis of mesothelioma.

Peroxiredoxins, thioredoxin, and Y-box-binding protein-1 are involved in the pathogenesis and progression of dialysis-associated renal cell carcinoma.

Patients with end-stage renal disease are exposed to increased oxidative stress and impairment of antioxidant mechanisms. We focused on dialysis renal cell carcinoma (RCC), including epithelial hyperplasia in acquired cystic disease of the kidney (ACDK). We attempted to obtain insight into the carcinogenesis and tumor progression in terms of cellular defense mechanisms associated with oxidative stress by investigating the expression of antioxidant proteins by immunohistochemistry. We evaluated retrospectively 43 cases of dialysis RCC and, as a control group, 49 cases of sporadic RCC. Peroxiredoxin (Prx) 1, 3, 4, 5, and 6 expression in dialysis RCC was positively correlated with the duration of dialysis. In epithelial hyperplasia, in 17 cases of acquired cystic disease of the kidney, Prxs and thioredoxin were highly expressed. Moreover, in dialysis RCC, Prx 3, 4, and 5 immunoreactivity and nuclear expression of Y-box-binding protein-1 were higher than in sporadic RCC. In dialysis RCC, Prx 3, 4, and 5 immunoreactivity positively correlated with the Fuhrman nuclear grade. These data suggest that oxidative stress during dialysis enhances antioxidant activity, with an inhibiting effect on carcinogenesis. Once cancer has developed, antioxidant activity might have a stimulating effect on the progression of dialysis RCC.

Nuclear expression of thioredoxin-1 in the invasion front is associated with outcome in patients with gallbladder carcinoma.

BACKGROUND: Multifunctional redox protein human thioredoxin (TRX-1) is reduced by thioredoxin reductase (TRX-R). The aim of the present study was to examine the distribution of TRX-1 and TRX-R expressions in gallbladder carcinoma (GBC) to clarify their usefulness as prognostic factors after surgical resection. METHODS: Immunohistochemical staining for TRX-1 and TRX-R was performed in GBC tissue from 38 patients who underwent surgical resection, and TRX-1/TRX-R localization in relation to outcome was examined. RESULTS: TRX-1 protein levels were significantly higher in GBC samples than in cholecystolithiasis samples (P = 0.0174). TRX-1 expression was observed in 100% (38/38) of tumour samples and in the nucleus in 76% (29/38), with nuclear expression in the invasion front observed in 45% (13/29). TRX-R expression was only detected in the cytoplasm of cancer cells and in the invasion front in 28 samples. In all of the samples, the depth of tumour invasion, lymph node metastasis, surgical margin, curability and nuclear expression of TRX-1 in the invasion front were significant prognostic factors by univariate analysis. In 27 selected patients who underwent curative resection, both TRX-1 nuclear expression and TRX-R cytoplasmic expression in the invasion front was a significantly prognostic factor. CONCLUSION: TRX-1 nuclear expression in the GBC invasion front is a significant prognostic marker. Patients with both TRX-1 nuclear expression and TRX-R cytoplasmic expression in the tumour invasion front should be observed carefully even if after curative resection.

Cycling hypoxia up-regulates thioredoxin levels in human MDA-MB-231 breast cancer cells.

The thioredoxin system is a key cellular antioxidant system and is highly expressed in cancer cells, especially in more aggressive and therapeutic resistant tumors. We analysed the expression of the thioredoxin system in the MDA-MB-231 breast cancer cell line under conditions mimicking the tumor oxygen microenvironment. We grew breast cancer cells in either prolonged hypoxia or hypoxia followed by various lengths of reoxygenation and in each case cells were cultured with or without a hypoxic cycling preconditioning (PC) phase preceding the hypoxic growth. Flow cytometry-based assays were used to measure reactive oxygen species (ROS) levels. Cells grown in hypoxia showed a significant decrease in ROS levels compared to normoxic cells, while a significant increase in ROS levels over normoxic cells was observed after 4 h of reoxygenation. The PC pre-treatment did not have a significant effect on ROS levels. Thioredoxin levels were also highest after 4 h of reoxygenation, however cells subjected to PC pre-treatment displayed even higher thioredoxin levels. The high level of intracellular thioredoxin was also reflected on the cell surface. Reporter assays showed that activity of the thioredoxin and thioredoxin reductase gene promoters was also highest in the reoxygenation phase, although PC pre-treatment did not result in a significant increase over non-PC treated cells. The use of a dominant negative Nrf-2 negated the increased thioredoxin promoter activity during reoxygenation. This data suggests that the high levels of thioredoxin observed in tumors may arise due to cycling between hypoxia and reoxygenation.

[Identification of proteins overexpressed in malignant gastric tumors: comparison of the results of 2-De and bioinformatics search].

Comparison of protein expression in intestinal and diffuse stomach tumors by 2D gel electrophoresis led to identification of three proteins (SOD2, S100A6, and TXN), which are overexpressed in tumors as compared to normal controls. It was shown, that overexpression of proteins SOD2 and TXN occurs much more frequently in diffuse tumors than in intestinal ones. A control panel of eleven proteins overexpressed in stomach tumors has been selected based on the data of comparative 2D analysis described in the literature. Bioinformatics search for mRNAs encoding proteins from the control panel in Oncomine database (which contains the results of determination of mRNA transcription level in tumor vs. normal samples) demonstrated the coincidence of proteomic and transcriptomic data for seven out of 11 proteins.