The Role of Peroxiredoxin Family in Cancer Signaling

Peroxiredoxins (Prxs) are antioxidant enzymes that protect cells from oxidative stress by reducing intracellular accumulation of reactive oxygen species (ROS). In mammalian cells, the six Prx isoforms are ubiquitously expressed in diverse intracellular locations. They are involved in the regulation of various physiological processes including cell growth, differentiation, apoptosis, immune response and metabolism as well as intracellular ROS homeostasis. Although there are increasing evidences that Prxs are involved in carcinogenesis of many cancers, their role in cancer is controversial. The ROS levels in cancer cells are increased compared to normal cells, thus promoting cancer development. Nevertheless, for various cancer types, an overexpression of Prxs has been found to be associated with poor patient prognosis, and an increasing number of studies have reported that tumorigenesis is either facilitated or inhibited by regulation of cancer-associated signaling pathways. This review summarizes Prx isoforms and their basic functions, the relationship between the expression level and the physiological role of Prxs in cancer cells, and their roles in regulating cancer-associated signaling pathways.

Interaction between transcriptional factor E26 transformation specific 1 and peroxiredoxin 1 in nicotine-induced oral precancerous lesion cells / 中华口腔医学杂志

Objective@#To investigate the interaction between nuclear transcriptional factor E26 transformation specific 1 (Ets1) and peroxiredoxin 1 (Prx1) in nicotine-induced oral precancerous lesion cells.@*Methods@#Human oral precancerous lesion dysplastic oral keratinocyte (DOK) cells were cultured and divided into nicotine group, control group, knockdown group and knockdown control group. The nicotine group, knockdown group and knockdown control group were treated with 1 μmol/L nicotine for 7 days while the control group was untreated. Western blotting, co-immunoprecipitation (Co-IP) and chromatin immunoprecipitation (ChIP) were performed to detect Prx1 and Ets1 protein expression, Prx1 and Ets1 protein interaction, combined activity of Ets1 with PRDX1 gene promoter region in nicotine group and control group DOK cells. In nicotine group, DOK cells were transfected with siRNA or lentivirus to knockdown Ets1 and Prx1 expression. Prx1 and Ets1 protein expression was examined by Western blotting.@*Results@#Nicotine increased the expression of Prx1 and Ets1 protein in DOK cells. The relative expression of Prx1 and Ets1 was 0.71±0.02, 0.12±0.01 in nicotine group and 0.53±0.06, 0.01±0.01 in control group (P=0.009, P=0.000). Co-IP showed that Prx1 could form protein complex with Ets1. The expression of Prx1 and Ets1 complex protein was increased in nicotine group. ChIP revealed that nicotine upregulated the combination of transcriptional factor Ets1 with PRDX1 gene promoter region, and the enrichment fold was 80.9±19.7 in nicotine group and 13.8±1.2 in control group (P=0.004). Ets1 and Prx1 protein expression was knocked down. The relative expression of Ets1 and Prx1 was 0.60±0.06, 0.48±0.03 in knockdown group and 0.83±0.08, 0.80±0.06 in knockdown control group (P=0.016, P=0.002). Ets1 knockdown suppressed the expression of Prx1 (P=0.002). Conversely, Prx1 knockdown also inhibited the expression of Ets1 significantly (P=0.000).@*Conclusions@#In oral precancerous lesion cells, Ets1 directly regulates Prx1 expression and nicotine might promote the development of oral precancerous lesion by magnifying the positive feedback signal pathway between Ets1 and Prx1.

Efeito pró-oxidante do hidroperóxido de urato sobre proteínas sensíveis às alterações redox: implicações na resposta inflamatória / Pro-oxidant effect of urate hydroperoxide on redoxsensitive proteins: implications on inflammatory reponse

O hidroperóxido de urato (HOOU) é o produto da oxidação do ácido úrico por peroxidases. Sua produção é favorecida durante a inflamação e hiperuricemia, uma vez que há grande quantidade de ácido úrico, peroxidases inflamatórias e superóxido. Neste sentido, o objetivo deste estudo foi avaliar o efeito do hidroperóxido de urato sobre proteínas sensíveis à modulação redox em um ambiente inflamatório asséptico e outro que imita infecção. Assim, nesta tese comparou-se a estrutura química do HOOU obtido fotoquimicamente daquele obtido através da catálise enzimática pela mieloperoxidase. A obtenção do HOOU por foto-oxidação permitiu o melhor isolamento do composto. Este oxidante foi capaz de reagir especificamente com os aminoácidos contendo enxofre (metionina e cisteína). Neste sentido, foi investigada sua reatividade com tiol-peroxidases detoxificadoras de peróxido, a peroxiredoxina 1 e 2 (Prx1 e Prx2). O HOOU apresentou cinética rápida de reação com a Prx1, k = 4,9 × 105 M-1s-1 e Prx2, k = 2,3 × 106 M-1s-1, o que as torna um provável alvo celular, além disso, foi capaz de oxidar a Prx2 de eritrócitos humanos, mostrando ser capaz de atravessar a membrana plasmática. Além das Prxs, a albumina do soro também desempenha papel importante na homeostase redox. O HOOU foi capaz de oxidar a albumina com constante de velocidade de 0,2 × 102 M-1s- 1. Outra tiol-proteína com importante função na homeostase e sinalização redox é a tioredoxina (Trx). A Trx foi oxidada pelo HOOU com constante de reação de 2,8 × 102 M-1s-1 e foi liberada juntamente com a Prx1 e Prx2 das células de macrófagos humanos (linhagem THP-1) quando estas células foram incubadas com HOOU. A liberação dessas proteínas é reconhecidamente um sinal de estresse celular. Assim o HOOU pode estar envolvido na exacerbação do estresse oxidativo em ambiente inflamatório. Quando neutrófilos (linhagem HL- 60) e macrófagos humanos (linhagem THP-1) foram incubados na presença de ácido úrico e Pseudomonas aeruginosa houve uma diminuição na produção de ácido hipocloroso (HOCl). Isto se deveu à competição entre ácido úrico e cloreto pela mieloperoxidase e resultou em menor atividade microbicida pelas células, demonstrando que a formação do HOOU não contribui e, ao contrário, prejudica a atividade microbicida das células inflamatórias. Dessa forma, a oxidação do ácido úrico e formação do hidroperóxido de urato tanto altera a atividade microbicida das células inflamtárias, quanto leva à oxidação de tiósproteínas importantes para manutenção da homeostase redox. Assim, o HOOU pode ser o responsável pelos efeitos pró-oxidantes e pró-inflamatórios do ácido úrico solúvel, e isso indica que o papel antioxidante do ácido úrico deve ser revisto em situações de inflamação.

Urate hydroperoxide (HOOU) is the product of the oxidation of uric acid by peroxidases. The formation of HOOU is favored during inflammation and in hyperuricemia, where there is plenty amount of uric acid, inflammatory peroxidases and superoxide. Therefore, the aim of the present study was to evaluate the effect of urate hydroperoxide on redox sensitive proteins in an inflammatory environment and another that mimics infection. In this thesis the chemical structure of the HOOU produced by photo-oxidation was compared to that obtained by myeloperoxidase catalysis. The chemical production of HOOU allowed a better purification of the compound. This oxidant was able to specifically react with sulfur containing amino acids (methionine and cysteine). In this sense, its reactivity with peroxiredoxins (Prx1 and Prx2) was investigated. HOOU reacted fast with Prx1 k = 4.9 × 105 M-1s-1 and Prx2 k = 2.3 × 106 M-1s-1. In addition, HOOU was able to oxidize Prx2 from intact erythrocytes at the same extend as does hydrogen peroxide. Albumin is an important thiol-containing protein to redox homeostasis in plasma. HOOU was able to oxidize albumin with a rate constant of 0.2 × 102 M-1s-1. Another protein with important function in redox homeostasis is thioredoxin (Trx). Trx was oxidized by HOOU with a rate constant of 2.8 × 102 M-1s-1 and was released together with Prx1 and Prx2 from human macrophages cells (THP-1 cell line) that were incubated with HOOU. The release of these proteins is a signal of cellular stress. Thus, HOOU may be involved in the exacerbation of oxidative stress in inflammatory environments. When neutrophil (HL-60 cell line) and macrophages (THP-1 cell line) were incubated with uric acid and Pseudomonas aeruginosa there was a decrease in hypochlorous acid (HOCl) production because of the competition between chloride and uric acid by myeloperoxidase. It decreased HOCl and impaired the microbicidal activity of the cells, showing that HOOU does not contribute in bacteria clearance. Therefore, the oxidation of uric acid to urate hydroperoxide impairs microbicidal activity and oxidizes thiol-proteins in inflammatory cells contributing to a pro-oxidant status. In this context, the antioxidant role of uric acid in inflammatory response should be reviwed.

Estudos estruturais e funcionais de Tsa1 de Saccharomyces cerevisiae: um modelo biológico para o estudo de inibidores de crescimento celular para leucemia linfoide aguda / Functional and structural evaluation of Tsa1 from Saccharomyces cerevisiae: a biological model for study of cellular growth inhibitors for acute lymphocytic leukemia

As peroxirredoxinas (Prx) são enzimas antioxidantes que se destacam pela capacidade de decompor uma grande variedade de hidroperóxidos com elevada eficiência (106-108M-1s-1), mantendo essas moléculas em níveis adequados à homeostase celular. Entretanto, já foi demonstrado que em diversos tipos tumorais os níveis de Prx são extremamente aumentados e experimentos envolvendo sua inativação resultam na diferenciação ou apoptose de células tumorais. Recentemente, foi descoberto um diterpenóide denominado adenantina que seria o primeiro inibidor para as Prx1 e Prx2 de humanos e foi demonstrada que sua aplicação em células de leucemia mieloide aguda promoveu diferenciação ou apoptose dessas células. Nesse contexto, o presente trabalho apresenta duas vertentes: 1) A caracterização das alterações estruturais e funcionais promovidas pela ligação da adenantina ao sítio ativo das Prx utilizando Tsa1 de Saccharomyces cerevisiae como modelo biológico, em função da sua alta similaridade com Prx2 de humanos; 2) Avaliação da atividade antitumoral dose dependente de adenantina sobre as linhagens celulares REH e MOLT-4 de leucemia linfoide aguda. No que concerne à primeira linha de investigação, nossos resultados revelam que Tsa1 é suscetível à inibição por adenantina, uma vez que o tratamento reduziu em ~66 % a velocidade de decomposição de peróxido de hidrogênio. Adicionalmente, a mutação da Thr44 de Tsa1, pertencente à chamada tríade catalítica, por uma Ser resultou em uma proteína mais suscetível a alterações na estrutura secundária e à inibição da atividade peroxidásica em função da ligação com adenantina, apresentando uma diminuição de ~85% na velocidade de reação. Características semelhantes foram observadas para a proteoforma Tsa2 de S. cerevisiae, que carreia naturalmente a substituição da Thr44 pela Ser. Análises de sequências de Prx em bancos de dados revelaram que majoritariamente proteínas contendo Ser são encontradas em organismos procariotos, muitos deles patogênicos. Finalmente, demonstramos por meio de ensaios citotoxicidade que as bactérias Staphylococcus aureus e Staphylococcus epidermidis, que possuem uma Ser na tríade catalítica, têm seu crescimento inibido pelo tratamento com adenantina (IC50 de 460µM e 77µM, respectivamente), enquanto que para Escherichia coli, que possui Thr nessa posição, a toxicidade da adenantina foi bastante baixa (não foi possível determinar o IC50 nas condições utilizadas). Dessa forma, os dados apresentados neste trabalho demonstram o potencial da utilização da adenantina tanto como antibiótico quanto como antileucêmico

Peroxiredoxins (Prx) are antioxidant enzymes which stand out due the ability to decompose a wide variety of hydroperoxides with high efficiency (106-108M-1s-1) maintaining these molecules at suitable levels to cellular homeostasis and participating in several signaling events. However, it has been shown that, in many tumor types, Prx levels are extremely increased and experiments involving its inactivation have resulted in differentiation or apoptosis of tumor cells. It was recently found a diterpenoid, called adenanthin, that would be the first human Prx1 and Prx2 inhibitor and it was demonstrated that its application in acute myeloid leukemia cells was able to promote differentiation or apoptosis. In this context, this work presents two lines of research: 1) Characterization of structural and functional changes promoted by adenanthin binding to Prx active site using Tsa1 from Saccharomyces cerevisiae as biological model, due to its high similarity to human Prx2. 2) Evaluation of adenanthin dose-dependent antitumor activity over the acute lymphoid leukemia cell lines REH and MOLT-4. As regards the first line of research, our result reveal that Tsa1 is susceptible to inhibition by adenanthin, since the treatment with this binder reduced the hydrogen peroxide decomposition velocity in ~ 66%. In addition, the replacement of Thr44 from Tsa1, aminoacid belonging to the so-called catalytic triad, by a Ser resulted in a protein more susceptible to alterations in secondary structure and to peroxidase activity inhibition in function of adenanthin binding, presenting ~85% of decrease in reaction velocity. Similar characteristics were observed for Tsa2 proteoform from S. cerevisiae, which naturally carries the substitution of Thr44 by Ser. Prx sequences analyzes in databases revealed that mostly Ser-containing proteins are found in prokaryotic organisms, many of them pathogenic ones. Finally, we demonstrate through cytotoxicity assays that the bacteria Staphylococcus aureus and Staphylococcus epidermidis, which have a Ser in catalytic triad, have their growth inhibited by adenanthin treatment (IC50 of 460µM and 77µM, respectively), whereas for Escherichia Coli, which has Thr at that position, the tocyxicity of adenanthin was quite low (it was not possible to determine the IC50 under the used conditions). Regarding the second line of investigation, we found that adenanthin is able to induce the death of leukemic cell lines REH and MOLT-4, and for the last one, there was an unexpected proliferation of cells treated by the longest incubation period (72 hours), characterizing a possible indication of differentiation process. In this sense, the data presented here demonstrate the potential of adenanthin use in both antibiotic and antileukemic treatment

Effects of Prdx4 protein expression on migration and invasion of HeLa cells / 中国病理生理杂志

AIM:To investigate the effects of peroxiredoxin 4 ( Prdx4) protein expression levels on the migra-tion and invasion of human cervical cancer HeLa cells.METHODS:The plasmid pcDNA3.0-HA-Prdx4 was transfected into HeLa cells.The HeLa cells were infected with LV-Prdx4 RNAi vector to establish stable Prdx4 shRNA HeLa cells. The change in the expression of Prdx4 protein was validated by Western blotting.The wound-healing assay, and Transwell migration and invasion assays were performed to detect the migration and invasion of HeLa cells, respectively.RESULTS:The expression of Prdx4 protein was up-regulated in the HeLa cells after transfection with pcDNA3.0-HA-Prdx4 plasmid ( P<0.05), whereas it was down-regulated in the Prdx4 shRNA HeLa cells (P<0.05).The abilities of migration and inva-sion were significantly increased in Prdx4-overexpressing HeLa cells compared with non-transfected and mock plasmid trans-fected control groups ( P<0.01) .When Prdx4 was knocked down by shRNA, the migration and invasion of the HeLa cells were remarkably repressed compared with blank control group and negative control group ( P<0.01 ) .CONCLUSION:The up-regulation of Prdx4 expression facilitates the migration and invasion of HeLa cells, and the down-regulation of Prdx4 expression inhibits the migration and invasion of HeLa cells, indicating that Prdx4 may be a potential molecular target for cervical cancer therapy.

Peroxiredoxin isoforms are associated with cardiovascular risk factors in type 2 diabetes mellitus

The production of oxygen free radicals in type 2 diabetes mellitus contributes to the development of complications, especially the cardiovascular-related ones. Peroxiredoxins (PRDXs) are antioxidant enzymes that combat oxidative stress. The aim of this study was to investigate the associations between the levels of PRDX isoforms (1, 2, 4, and 6) and cardiovascular risk factors in type 2 diabetes mellitus. Fifty-three patients with type 2 diabetes mellitus (28F/25M) and 25 healthy control subjects (7F/18M) were enrolled. We measured the plasma levels of each PRDX isoform and analyzed their correlations with cardiovascular risk factors. The plasma PRDX1, -2, -4, and -6 levels were higher in the diabetic patients than in the healthy control subjects. PRDX2 and -6 levels were negatively correlated with diastolic blood pressure, fasting blood sugar, and hemoglobin A1c. In contrast, PRDX1 levels were positively correlated with low-density lipoprotein and C-reactive protein levels. PRDX4 levels were negatively correlated with triglycerides. In conclusion, PRDX1, -2, -4, and -6 showed differential correlations with a variety of traditional cardiovascular risk factors. These results should encourage further research into the crosstalk between PRDX isoforms and cardiovascular risk factors.

The Peroxiredoxins protein family and related disease / 医学研究生学报

Peroxiredoxins(Prxs) are a family of antioxidant protein that have been identified in prokaryotes and eukaryotes. As antioxidants, Prxs protein contains an active site cysteine that is sensitive to oxidation by H 2 O2, eliminate active oxygen that exist in normal tissues and cells, protect cells from oxidative damage induced by reactive oxygen species ( ROS). Prxs protein is a known free radical scavenger, and has been shown to play a role in several diseases. In this review, recent advances on the study of Prxs protein family and tumor related diseases are reviewed, which is expected to provide new ideas for the diagnosis and treatment of the related clinical diseases.

Effects of change in expression level of Prdx4 protein on proliferation and apoptosis of cervical cancer HeLa cells / 肿瘤

Objective: To investigate the effects of change in expression level of peroxiredoxin 4 (Prdx4) protein on proliferation and apoptosis of cervical cancer HeLa cells. Methods: The combination plasmid pcDNA3.0-HA-Prdx4 (the empty vector pcDNA3.0-HA was used as the negative control) and three small interference RNAs (siRNAs) against human Prdx4 gene (the negative control siRNA was used as the negative control) were transfected into HeLa cells, respectively. The change in the expression of Prdx4 protein was validated by Western blotting, then MTT assay and flow cytometry (FCM) were performed to detect the proliferation activity and apoptosis of HeLa cells, respectively. Results: The expression level of Prdx4 protein was up-regulated in HeLa cells after transfection with pcDNA3.0-HA-Prdx4 plasmid (P 0.05). But three siRNAs-mediated interference of Prdx4 protein expression significantly decreased the proliferation activity and increased the apoptotic rate of HeLa cells as compared with the negative control group (transfected with the negative control siRNA) (all P < 0.05). Conclusion: Although the up-regulation of Prdx4 expression has no effect on the cell proliferation and apoptosis of HeLa cells, the down-regulation of Prdx4 expression can inhibit the cell proliferation and promote the apoptosis. These findings indicate that Prdx4 gene may become a candidate molecular target for cervical cancer therapy.

Thyroid and Hydrogen Peroxide / 한양의대학술지

The thyroid gland produces high concentrations of hydrogen peroxide (H2O2) which is used for the biosynthesis of the thyroid hormones, triiodothyronine and thyroxine. The most important step in thyroid hormone synthesis is oxidation and organification of iodide, which itself is a very potent oxidant. To oxidize iodide, thyrocytes generate high intracellular concentrations of H2O2 using the NADPH-dependent dual oxidase enzymes (DUOX) and remove used H2O2 by various well-characterized antioxidant systems, such as those involving glutathione peroxidases and peroxiredoxins. Although thyrocytes have very intricate and efficient systems to regulate H2O2 generation and removal, leakage of H2O2 may be inevitable because of the high intracellular concentrations of H2O2 in thyrocytes. Leakage of the highly reactive oxidant H2O2 is likely to cause the damage to various intracellular proteins and DNA that has been demonstrated to be a causative factor in the induction of various diseases, such as thyroid cancer, chronic thyroiditis and hyperthyroidism. Therefore, ongoing investigation of the systems regulating H2O2 production and elimination in thyrocytesis essential for understanding the regulation and process of thyroid hormone synthesis and also for elucidating pathogenetic mechanisms underlying various autoimmune thyroid diseases and the initiation and promotion of thyroid cancers.

Protective Effects of Peroxiredoxin on Hydrogen Peroxide Induced Oxidative Stress and Apoptosis in Cardiomyocytes

BACKGROUND AND OBJECTIVES: The redox system is an important anti-oxidative system composed of thioredoxin, thioredoxin reductase, and peroxiredoxin (PRx). The fine details of PRx expression and its protective effects in various cells in cardiovascular tissue under oxidative stress created by hydrogen peroxide have not been fully elucidated. SUBJECTS AND METHODS: Oxidative stress was induced by adding hydrogen peroxide at 0.25 mM for 2 hours to rat neonatal cardiomyocytes (rCMCs), rat vascular smooth muscle cells (rVSMCs), and human umbilical vein endothelial cells (HUVECs). Apoptosis was quantified by flow cytometry and the expression patterns of the six PRx isoforms were evaluated by western blotting in the three cell lines after hydrogen peroxide stimulation. Apoptosis and the cell survival signal pathway were evaluated by PRx1 gene delivery using lentiviral vector in hydrogen peroxide stimulated rCMCs versus green fluorescence protein gene delivery. RESULTS: Hydrogen peroxide induced 25% apoptosis in rCMCs. Furthermore, the PRx1 and 5 isoforms were found to be overexpressed in hydrogen peroxide treated rCMCs, and PRx1 overexpression by gene delivery was found to reduce hydrogen peroxide induced rCMCs apoptosis significantly. In addition, this effect was found to originate from cell survival pathway modification. CONCLUSION: Hydrogen peroxide induced significant oxidative stress in rCMCs, rVSMCs, and HUVECs, and PRx1 overexpression using a lentiviral vector system significantly reduced hydrogen peroxide induced rCMCs apoptosis by upregulation of cell survival signals and downregulation of apoptotic signals. These findings suggest that PRx1 could be used as a treatment strategy for myocardial salvage in conditions of oxidative stress.

Expression of peroxiredoxin I regulated by gonadotropins in the rat ovary / 대한생식의학회지

OBJECTIVE: Peroxiredoxins (Prxs) play an important role in regulating cellular differentiation and proliferation in several types of mammalian cells. This report examined the expression of Prx isotype I in the rat ovary after hormone treatment. METHODS: Immature rats were injected with 10 IU of pregnant mare's serum gonadotropin (PMSG) to induce the growth of multiple preovulatory follicles and 10 IU of human chorionic gonadotropin (hCG) to induce ovulation. Immature rats were also treated with diethylstilbestrol (DES), an estrogen analogue, to induce the growth of multiple immature follicles. Northern blot analysis was performed to detect gene expression. Cell-type specific localization of Prx I mRNA were detected by in situ hybridization analysis. RESULTS: During follicle development, ovarian Prx I gene expression was detected in 3-day-old rats and had increased in 21-day-old rats. The levels of Prx I mRNA slightly declined one to two days following treatment with DES. A gradual increase in Prx I gene expression was observed in ovaries obtained from PMSG-treated immature rats. Furthermore, hCG treatment of PMSG-primed rats resulted in a gradual stimulation of Prx I mRNA levels by 24 hours (2.1-fold increase) following treatment, which remained high until 72 hours following treatment. In situ hybridization analysis revealed the expression of the Prx I gene in the granulosa cells of PMSG-primed ovaries and in the corpora lutea of ovaries stimulated with hCG for 72 hours. CONCLUSION: These results demonstrate the gonadotropin and granulosa cell-specific stimulation of Prx I gene expression, suggesting its role as a local regulator of follicle development.

Nuclear factor E2-related factor 2 Dependent Overexpression of Sulfiredoxin and Peroxiredoxin III in Human Lung Cancer

BACKGROUND/AIMS: Oxidative stress results in protein oxidation and is implicated in carcinogenesis. Sulfiredoxin (Srx) is responsible for the enzymatic reversal of inactivated peroxiredoxin (Prx). Nuclear factor E2-related factor 2 (Nrf2) binds to antioxidant responsive elements and upregulates the expression of Srx and Prx during oxidative stress. We aimed to elucidate the biological functions and potential roles of Srx in lung cancer. METHODS: To study the roles of Srx and Prx III in lung cancer, we compared the protein levels of Nrf2, Prxs, thioredoxin, and Srx in 40 surgically resected human lung cancer tissues using immunoblot and immunohistochemical analyses. Transforming growth factor-beta1, tumor necrosis factor-alpha, and camptothecin treatment were used to examine Prx III inactivation in Mv1Lu mink lung epithelial cells and A549 lung cancer cells. RESULTS: Prx I and Prx III proteins were markedly overexpressed in lung cancer tissues. A significant increase in the oxidized form of a cysteine sulfhydryl at the catalytic site of Prxs was found in carcinogenic lung tissue compared to normal lung tissue. Densitometric analyses of immunoblot data revealed significant Srx expression, which was higher in squamous cell carcinoma tissue (60%, 12/20) than in adenocarcinoma (20%, 4/20). Also, Nrf2 was present in the nuclear compartment of cancer cells. CONCLUSIONS: Srx and Prx III proteins were markedly overexpressed in human squamous cell carcinoma, suggesting that these proteins may play a protective role against oxidative injury and compensate for the high rate of mitochondrial metabolism in lung cancer.

Selectively Decreased Expression of Peroxiredoxins Induced by Silica in Pulmonary Epithelial Cells

BACKGROUND/AIMS: Peroxiredoxin (Prx) belongs to a ubiquitous family of antioxidant enzymes that regulates many cellular processes through intracellular oxidative signal transduction pathways. Silica-induced lung damage involves reactive oxygen species (ROS) that trigger subsequent toxic effects and inflammatory responses in alveolar epithelial cells resulting in fibrosis. Therefore, we investigated the role of Prx in the development of lung oxidant injury caused by silicosis, and determined the implication of ROS in that process. METHODS: Lung epithelial cell lines A549 and WI26 were treated with 1% silica for 0, 24, or 48 hours, following pretreatment of the A549 cells with N-acetyl-L-cysteine and diphenylene iodonium and no pretreatment of the WI26 cells. We transfected an HA-ubiquitin construct into the A549 cell line and then analyzed the cells via Western blotting and co-immunoprecipitation. RESULTS: Silica treatment induced cell death in the A549 lung epithelial cell line and selectively degraded Prx I without impairing protein synthesis in the A549 cells, even when the ROS effect was blocked chemically by N-acetyl-L-cysteine. A co-immunoprecipitation study revealed that Prx I did not undergo ubiquitination. CONCLUSIONS: Silica treatment induces a decrease of Prx I expression in lung epithelial cell lines regardless of the presence of ROS. The silica-induced degradation of Prx does not involve the ubiquitin-proteasomal pathway.

Characterization of Peroxiredoxins in the Gray matter in the spinal cord after Acute Immobilization Stress

PURPOSE: Many stresses produce reactive oxygen species and bring about mechanism of antioxidant reaction. Cytokine and a neurotransmitter through the cell membrane, as well as signal transduction through the cell membrane, are used for various pathological condition of the brain, such as neurodegenerative disease. There are several antioxidant enzymes in cells (superoxcide dismutase, glutathion peroxidasae, peroxiredoxin catalase, etc.) METHODS: This study used single- or double-label immunohistochemical techniques to analyze mouse spinal neuron cells expressing Prx I and Prx III after acute mobilization stress. RESULTS: Prx I was observed in dendritic cell of the gray matter of the spinal cord, and Prx III was observed in the cytoplasm of the GM of the spinal cord. CONCLUSION: The results of this study will help to explain differences of expression in the distributions of the peroxiredoxin enzymes of the spinal cord.

Characterization of Peroxiredoxins in the Gray matter in the spinal cord after Acute Immobilization Stress

PURPOSE: Many stresses produce reactive oxygen species and bring about mechanism of antioxidant reaction. Cytokine and a neurotransmitter through the cell membrane, as well as signal transduction through the cell membrane, are used for various pathological condition of the brain, such as neurodegenerative disease. There are several antioxidant enzymes in cells (superoxcide dismutase, glutathion peroxidasae, peroxiredoxin catalase, etc.) METHODS: This study used single- or double-label immunohistochemical techniques to analyze mouse spinal neuron cells expressing Prx I and Prx III after acute mobilization stress. RESULTS: Prx I was observed in dendritic cell of the gray matter of the spinal cord, and Prx III was observed in the cytoplasm of the GM of the spinal cord. CONCLUSION: The results of this study will help to explain differences of expression in the distributions of the peroxiredoxin enzymes of the spinal cord.

Oxidative Inactivation of Peroxiredoxin Isoforms by H2O2 in Pulmonary Epithelial, Macrophage, and other Cell Lines with their Subsequent Regeneration / 결핵및호흡기질환

BACKGROUND: Peroxiredoxins (Prxs) are a relatively newly recognized, novel family of peroxidases that reduce H2O2 and alkylhydroperoxide into water and alcohol, respectively. There are 6 known isoforms of Prxs present in human cells. Normally, Prxs exist in a head-to-tail homodimeric state in a reduced form. However, in the presence of excess H2O2, it can be oxidized on its catalytically active cysteine site into inactive oxidized forms. This study surveyed the types of the Prx isoforms present in the pulmonary epithelial, macrophage, endothelial, and other cell lines and observed their response to oxidative stress. METHODS: This study examined the effect of exogenous, excess H2O2 on the Prxs of established cell lines originating from the pulmonary epithelium, macrophages, and other cell lines, which are known to be exposed to high oxygen partial pressures or are believed to be subject to frequent oxidative stress, using non-reducing SDS polyacrylamide electrophoresis (PAGE) and 2 dimensional electrophoresis. RESULT: The addition of excess H2O2 to the culture media of the various cell-lines caused the immediate inactivation of Prxs, as evidenced by their inability to form dimers by a disulfide cross linkage. This was detected as a subsequent shift to its monomeric forms on the non-reducing SDS PAGE. These findings were further confirmed by 2 dimensional electrophoresis and immunoblot analysis by a shift toward a more acidic isoelectric point (pI). However, the subsequent reappearance of the dimeric Prxs with a comparable, corresponding decrease in the monomeric bands was noted on the non-reducing SDS PAGE as early as 30 minutes after the H2O2 treatment suggesting regeneration after oxidation. The regenerated dimers can again be converted to the inactivated form by a repeated H2O2 treatment, indicating that the protein is still catalytically active. The recovery of Prxs to the original dimeric state was not inhibited by a pre-treatment with cycloheximide, nor by a pretreatment with inhibitors of protein synthesis, which suggests that the reappearance of dimers occurs via a regeneration process rather than via the de novo synthesis of the active protein. CONCLUSION: The cells, in general, appeared to be equipped with an established system for regenerating inactivated Prxs, and this system may function as a molecular "on-off switch" in various oxidative signal transduction processes. The same mechanisms might applicable other proteins associated with signal transduction where the active catalytic site cysteines exist.