Peroxiredomin-4 ameliorates lipotoxicity-induced oxidative stress and apoptosis in diabetic cardiomyopathy.

Diabetic cardiomyopathy (DCM), one severe complication in the diabetes, leads to high mortality in the diabetic patients. However, the understanding of molecular mechanisms underlying DCM is far from completion. Herein, we investigated the disease-related differences in the proteomes of DCM based on db/db mice and verified the protective roles of peroxiredoxin-4 (Prdx4) in H9c2 cardiomyocytes treated by palmitic acid (PA). Fasting blood glucose (FBG) and cardiac function was detected in the 6-month-old control and diabetic mice. The hearts were then collected and analyzed by a coupled label-free and mass spectrometry approach. In vivo investigation indicated that body weight and FBG of db/db mice markedly increased, and diabetic heart exhibited obvious cardiac hypertrophy and lipid droplet accumulation, and cardiac dysfunction as is indicated by the increases of left ventricle posterior wall thickness in systole (LVPWd) and diastole (LVPWs), and reduction of fractional shortening (FS). We used proteomic analysis and then detected a grand total of 2636 proteins. 175 differentially expressed proteins (DEPs) were markedly detected in the diabetic heart. Thereinto, Prdx4 was markedly down-regulated in the diabetic heart. In vitro experiments revealed that 250 µM PA significantly inhibited viability of H9c2 cell. PA induced much accumulation of lipid droplet in cardiomyocytes and resulted in an increase of mRNA expressions of lipogenic genes (FASN and SCD1) and cardiac hypertrophic genes. Additionally, protein level of Prdx4 evidently reduced in the PA-treated H9c2 cell. It was further found that shRNA-mediated Prdx4 knockdown exacerbated PA-induced oxidative stress and cardiomyocyte apoptosis, whereas overexpressing Prdx4 in the H9c2 cells noteworthily limited PA-induced ROS generation and cardiomyocytes apoptosis. These data collectively reveal the essential role of abnormal Prdx4 in pathological alteration of DCM, and provide potentially therapeutic target for the prevention of DCM.

Peroxiredoxin-1 ameliorates pressure overload-induced cardiac hypertrophy and fibrosis.

BACKGROUND: Previous studies have demonstrated that Peroxiredoxin 1 (Prdx1) is a modulator of physiological and pathophysiological cardiovascular events. However, the roles of Prdx1 in cardiac hypertrophy and heart failure (HF) have barely been explored. Thus, this study aimed to investigate whether Prdx1 participates in cardiac hypertrophy and to elucidate the possible associated mechanisms. METHODS: Mice were subjected to transverse aortic constriction (TAC) for four weeks to induce pathological cardiac hypertrophy. Cardiomyocyte-specific Prdx1 overexpression in mice was achieved using an adeno-associated virus system. Morphological examination; echocardiography; and hemodynamic, biochemical and histological analyses were used to evaluate the roles of Prdx1 in pressure overload-induced cardiac hypertrophy and HF. RESULTS: First, the results showed that Prdx1 expression was noticeably upregulated in hypertrophic mouse hearts and cardiomyocytes with phenylephrine (PE)-induced hypertrophy in vitro. Prdx1 overexpression exerted protective effects against cardiac hypertrophy and fibrosis and ameliorated cardiac dysfunction in mice subjected to pressure overload. In addition, Prdx1 overexpression decreased pressure overload-induced cardiac inflammation and oxidative stress. Further studies demonstrated that Prdx1 overexpression increased the levels of nuclear factor-erythroid 2-related factor 2 (Nrf2) and its downstream antioxidant protein, heme oxygenase-1 (HO-1), in mice. Moreover, Nrf2 knockdown offset the antihypertrophic and anti-oxidative stress effects of Prdx1 overexpression. CONCLUSIONS: Prdx1 protects against pressure overload-induced cardiac hypertrophy and HF by activating Nrf2/HO-1 signaling. These data indicate that targeting Prdx1 may be an attractive pharmacotherapeutic strategy for the treatment of cardiac hypertrophy and HF.

Systematic Characterization of Prognostic Values of Peroxiredoxin Family in Gastric Cancer.

The peroxiredoxin (PRDX) gene family has been reported to participate in regulating occurrence and development of cancerous diseases, but its exact prognostic values in gastric cancer (GC) remain largely elusive. In the current research, we evaluated the prognostic value in predicting overall survival (OS) of each individual PRDX mRNA expression based on patients' cohorts from the Kaplan-Meier (KM) plotter database, which contains clinical information and gene expression data obtained from a total of 876 GC patients. Our results revealed that mRNA expressions of PRDX1, PRDX2, PRDX3, and PRDX4 were significantly associated with worse OS in GC patients, whereas PRDX5 and PRDX6 mRNA expressions were not associated with OS in GC patients. In addition, the prognostic values of PRDXs in the different clinicopathological features according to clinical stages, Lauren classifications, HER2 expression status, differentiation degree, and treatment strategies of GC patients were further evaluated in the KM plotter database. As a result, more potential beneficiaries who may benefit from prognostic assessment using PRDX mRNA expressions were identified. Our results elucidated the exact values of PRDXs in assessing GC prognosis and might provide primary evidence for further study on the mechanism of PRDXs participating in occurrence and development of GC.

Peroxiredoxin 4 attenuates glutamate-induced neuronal cell death through inhibition of endoplasmic reticulum stress.

High concentrations of glutamate induce neurotoxicity by eliciting reactive oxygen species (ROS) generation and intracellular Ca2+ influx. The disruption of Ca2+ homeostasis in the endoplasmic reticulum (ER) evokes ER stress, ultimately resulting in neuronal dysfunction. Additionally, glutamate participates in the development of neurodegenerative diseases, such as Alzheimer's and Parkinson's diseases. Peroxiredoxins (Prxs) are members of a family of antioxidant enzymes that protect cells from neurotoxic factor-induced apoptosis by scavenging hydrogen peroxide (H2O2). Prx4 is located in the ER and controls the redox condition within the ER. The present study investigated the protective effects of Prx4 against glutamate-induced neurotoxicity linked to ER stress. HT22 cells in which Prx4 was either overexpressed or silenced were used to elucidate the protective role of Prx4 against glutamate toxicity. The expression of Prx4 in HT22 cells was significantly increased in response to glutamate treatment, while ROS scavengers and ER chemical chaperones reduced Prx4 levels. Moreover, Prx4 overexpression reduces glutamate-induced apoptosis of HT22 cells by inhibiting ROS formation, Ca2+ influx, and ER stress. Therefore, we conclude that Prx4 has protective effects against glutamate-induced HT22 cell damage. Collectively, these results suggest that Prx4 could contribute to the treatment of neuronal disorders.

Theaflavins alleviate sevoflurane-induced neurocytotoxicity via Nrf2 signaling pathway.

Aim: Sevoflurane could induce apoptosis of rat hippocampal neurons, while theaflavins (TFs) have antioxidant and anti-inflammatory properties. This study aims to explore whether TFs could alleviate sevoflurane-induced neuronal cell injury.Materials and methods: Cells were treated by concentration gradient of sevoflurane and TFs. Cell viability, level of reactive oxygen species (ROS) and apoptosis rate were determined by cell counting kit-8 (CCK-8) and flow cytometry, respectively. Quantitative PCR (qPCR) and western blot were performed to determine mRNA and protein expressions.Results: TFs promoted viability of cells under the treatment of sevoflurane, while it suppressed apoptosis and down-regulated ROS level in a concentration-dependent manner. TFs could also down-regulate expression levels of caspase-3 and caspase-9 and cytosol and intranuclear nuclear factor E2-related factor 2 (Nrf2) in rat hippocampal nerve cells, while it up-regulated those of heme oxygenase 1 (HO-1), NADPH quinine oxidoreductase 1 (NQO1), glutamate cysteine ligase (GCL) and peroxiredoxin 1 (Prx1).Conclusions: Our study suggests that TFs exert protective effects on sevoflurane-induced neurocytotoxicity and therefore could be used as a potential drug for treatment of neuronal injury.

Tumor protein D52 (isoform 3) interacts with and promotes peroxidase activity of Peroxiredoxin 1 in prostate cancer cells implicated in cell growth and migration.

Tumor protein D52 (TPD52) is overexpressed in multiple cancers including prostate cancer due to gene amplification and investigations to understand its role in the pathophysiology of different cancers are continuing. GST pull-down assays and Tandem affinity purification of TPD52 as bait identified novel prey Peroxiredoxin 1 (PRDX1) in prostate cancer (PCa) cells. PRDX1 interaction with TPD52 was confirmed in immunoprecipitation and affinity interaction assays. Mapping of interaction domain indicated that PRDX1 interacts with C-terminal region of TPD52 containing PEST domain between 152 and 179 amino acids, a new binding region of TPD52. Here we show that TPD52 interaction with PRDX1 increased its peroxidase activity and ectopic expression of TPD52 induced dimerization of PRDX1 in PCa cells. Moreover, H2O2 exposure evoked the interaction between TPD52 and PRDX1 while depletion of both proteins led to the accumulation of H2O2 suggesting peroxidase activity is important to maintain oxidative capacity in PCa cells. We also observed that overexpression or downregulation of TPD52 and PRDX1 individually or together affecting PCa cells growth, survival, and migration. Altogether, our results show a novel interaction partner of TPD52 providing new insights of its functions and ascertain the role of TPD52-PRDX1 interaction in PCa progression.

The relapsing fever spirochete Borrelia turicatae persists in the highly oxidative environment of its soft-bodied tick vector.

The relapsing fever spirochete Borrelia turicatae possesses a complex life cycle in its soft-bodied tick vector, Ornithodoros turicata. Spirochetes enter the tick midgut during a blood meal, and, during the following weeks, spirochetes disseminate throughout O. turicata. A population persists in the salivary glands allowing for rapid transmission to the mammalian hosts during tick feeding. Little is known about the physiological environment within the salivary glands acini in which B. turicatae persists. In this study, we examined the salivary gland transcriptome of O. turicata ticks and detected the expression of 57 genes involved in oxidant metabolism or antioxidant defences. We confirmed the expression of five of the most highly expressed genes, including glutathione peroxidase (gpx), thioredoxin peroxidase (tpx), manganese superoxide dismutase (sod-1), copper-zinc superoxide dismutase (sod-2), and catalase (cat) by reverse-transcriptase droplet digital polymerase chain reaction (RT-ddPCR). We also found distinct differences in the expression of these genes when comparing the salivary glands and midguts of unfed O. turicata ticks. Our results indicate that the salivary glands of unfed O. turicata nymphs are highly oxidative environments where reactive oxygen species (ROS) predominate, whereas midgut tissues comprise a primarily nitrosative environment where nitric oxide synthase is highly expressed. Additionally, B. turicatae was found to be hyperresistant to ROS compared with the Lyme disease spirochete Borrelia burgdorferi, suggesting it is uniquely adapted to the highly oxidative environment of O. turicata salivary gland acini.

The overexpression of peroxiredoxin-4 affects the progression of idiopathic pulmonary fibrosis.

BACKGROUND: Acute exacerbation of idiopathic pulmonary fibrosis (AE-IPF) is life-threatening. Several serum biomarkers, such as Krebs von den Lungen-6 (KL-6) and surfactant protein D (SP-D), are clinically used for evaluating AE-IPF, but these biomarkers are not adequate for establishing an early and accurate diagnosis of AE-IPF. Recently, the protective roles of the members of the peroxiredoxin (PRDX) family have been reported in IPF; however, the role of PRDX4 in AE-IPF is unclear. METHODS: Serum levels of PRDX4 protein, KL-6, SP-D and lactate dehydrogenase (LDH) in 51 patients with stable IPF (S-IPF), 38 patients with AE-IPF and 15 healthy volunteers were retrospectively assessed using enzyme-linked immunosorbent assay. Moreover, as an animal model of pulmonary fibrosis, wild-type (WT) and PRDX4-transgenic (Tg) mice were intratracheally administered with bleomycin (BLM, 2 mg/kg), and fibrotic and inflammatory changes in lungs were evaluated 3 weeks after the intratracheal administration. RESULTS: Serum levels of PRDX4 protein, KL-6, SP-D and LDH in patients with S-IPF and AE-IPF were significantly higher than those in healthy volunteers, and those in AE-IPF patients were the highest among the three groups. Using receiver operating characteristic curves, area under the curve values of serum PRDX4 protein, KL-6, SP-D, and LDH for detecting AE-IPF were 0.873, 0.698, 0.675, and 0.906, respectively. BLM-treated Tg mice demonstrated aggravated histopathological findings and poor prognosis compared with BLM-treated WT mice. Moreover, PRDX4 expression was observed in alveolar macrophages and lung epithelial cells of BLM-treated Tg mice. CONCLUSIONS: PRDX4 is associated with the aggravation of inflammatory changes and fibrosis in the pathogenesis of IPF, and serum PRDX4 may be useful in clinical practice of IPF patients.

Rapamycin induces the expression of heme oxygenase-1 and peroxyredoxin-1 in normal hepatocytes but not in tumorigenic liver cells.

Rapamycin (sirolimus) is employed as an immunosuppressant following liver transplant, to inhibit the re-growth of cancer cells following liver resection for hepatocellular carcinoma (HCC), and for the treatment of advanced HCC. Rapamycin also induces the expression of antioxidant enzymes in the liver, suggesting that pretreatment with the drug could provide a potential strategy to reduce ischemia reperfusion injury following liver surgery. The aim of this study was to further investigate the actions of rapamycin in inducing expression of the antioxidant enzymes heme oxygenase-1 (HO-1) and peroxiredoxin-1 (Prx-1) in normal liver and in tumorigenic liver cells. A rat model of segmental hepatic ischemia and reperfusion, cultured freshly-isolated rat hepatocytes, and tumorigenic H4IIE rat liver cells in culture were employed. Expression of HO-1 and Prx-1 was measured using quantitative PCR and western blot. Rapamycin pre-treatment of normal liver in vivo or normal hepatocytes in vitro led to a substantial induction of mRNA encoding HO-1 and Prx-1. The dose-response curve for the action of rapamycin on mRNA expression was biphasic, showing an increase in expression at 0 - 0.1 µM rapamycin but a decrease from maximum at concentrations greater than 0.1 µM. By contrast, in H4IIE cells, rapamycin inhibited the expression of HO-1 and Prx-1 mRNA. Oltipraz, an established activator of transcription factor Nrf2, caused a large induction of HO-1 and Prx-1 mRNA. The dose response curve for the inhibition by rapamycin of HO-1 and Prx-4 mRNA expression, determined in the presence of oltipraz, was monophasic with half maximal inhibition at about 0.01 µM. It is concluded that, at concentrations comparable to those used clinically, pre-treatment of the liver with rapamycin induces the expression of HO-1 and Prx-1. However, the actions of rapamycin on the expression of these two antioxidant enzymes in normal hepatocytes are complex and, in tumorigenic liver cells, differ from those in normal hepatocytes. Further studies are warranted to evaluate preconditioning the livers of patients subject to liver resection or liver transplant with rapamycin as a viable strategy to reduce IR injury following liver surgery.

Epigenetic silencing of the tumor suppressor genes SPI1, PRDX2, KLF4, DLEC1, and DAPK1 in childhood and adolescent lymphomas.

The aim of the study was to investigate the expression and methylation status of seven distinctive genes with tumor suppressing properties in childhood and adolescent lymphomas. A total of 96 patients with Hodgkin Lymphoma (HL, n = 41), Non-Hodgkin Lymphoma (NHL, n = 15), and reactive lymphoid hyperplasia (RLH, n = 40, as controls) are included in the research. The expression status of CDKN2A, SPI1, PRDX2, DLEC1, FOXO1, KLF4 and DAPK1 genes were measured with QPCR method after the RNA isolation from paraffin blocks of tumor tissue and cDNA conversion. DNA isolation was performed from samples with low gene expression followed by methylation PCR study specific to promoter regions of these genes. We found that SPI1, PRDX2, DLEC1, KLF4, and DAPK1 genes are significantly less expressed in patient than the control group (p = 0.0001). However, expression of CDKNA2 and FOXO1 genes in the patient and control groups were not statistically different. The methylation ratios of all genes excluding the CDKN2A and FOXO1 were significantly higher in the HL and NHL groups than the controls (p = 0.0001). We showed that SPI1, PRDX2, DLEC1, KLF4 and DAPK1 genes are epigenetically silenced via hypermethylation in the tumor tissues of children with HL and NHL. As CDKN2A gene was not expressed in both patient and control groups, we conclude that it is not specific to malignancy. As FOXO1 gene was similarly expressed in both groups, its relationship with malignancy could not be established. The epigenetically silenced genes may be candidates for biomarkers or therapeutic targets in childhood and adolescent lymphomas.

Peroxiredoxin 1 (PRDX1) Suppresses Progressions and Metastasis of Osteosarcoma and Fibrosarcoma of Bone.

BACKGROUND Osteosarcoma and fibrosarcoma are malignant tumors with poor prognosis. Peroxiredoxin 1 (PRDX1) is considered to prevent tumors in many malignances. However, few studies have focused on the functions of PRDX1 in osteosarcoma and fibrosarcoma. MATERIAL AND METHODS PRDX1 mRNA in tumors and adjacent tissues of 32 osteosarcoma patients and 16 fibrosarcoma patients was extracted and measured. Proliferation and invasion of MG63 and HT1080 cell lines after silencing or overexpressing PRDX1 were used to detect the role of PRDX1 in metastasis of osteosarcoma and fibrosarcoma. RESULTS PRDX1 mRNA level was lower in tumor tissues than in adjacent tissues of osteosarcoma (F=50.105) and fibrosarcoma (F=28.472) patients, both significantly (P<0.05). Silencing PRDX1 promoted proliferation of MG63 and HT1080 cells, while overexpressing PRDX1 suppressed proliferation after 24 h, 48 h, and 72 h, compared to the control group, both significantly (P<0.05). Silencing PRDX1 increased invasive cells of MG63 (F=246.218) and HT1080 (F=245.602), while overexpressing PRDX1 decreased invasive cells of both, compared to the control, and the difference was significant (P<0.05). CONCLUSIONS PRDX1 expression is low in osteosarcoma and fibrosarcoma tumors. PRDX1 suppressed the progression and metastasis of osteosarcoma and fibrosarcoma cells.

The regulatory role of Nrf2 in antioxidants phase2 enzymes and IL-17A expression in patients with ulcerative colitis.

BACKGROUND: Reactive oxygen species (ROS) is one of the pathogenic factors responsible for intestinal injury in Ulcerative colitis (UC). Nuclear factor erythroid-2 related factor 2 (Nrf2) plays a critical role against ROS factors to conserve epithelial integrity. This study aimed to localize Nrf2 and IL-17A protein in the inflamed mucosa of patients with ulcerative colitis. The gene expression of Nrf2 was also correlated with GST-A4 and PRDX1. MATERIALS AND METHODS: A total of 20 patients and 20 healthy controls with definite UC based on the clinical criteria were enrolled for this study. The expression pattern of Nrf2 and IL-17A protein was compared in inflamed and non-inflamed colonic biopsies by immunohistochemical staining. Nrf2, GST-A4 and PRDX1 gene expression were determined by real-time polymerase chain reaction (RT-PCR). RESULTS: In inflamed colonic biopsies, an increased level of Nrf2 protein factor was detected in epithelial cells. Conversely, IL-17A protein was presented more in mononuclear cells in mucosa and lamina propria regions. A significant increase of Nrf2, GST-A4 gene expression was observed in both mild and severe patients with ulcerative colitis. GST-A4 gene expression indicated a high exponential rate in logistic regression. CONCLUSION: Oxidative stress in inflamed colonic tissue can induce Nrf2 gene expression. The performance of Nrf2 transcription factor may lead to the induction of GST-A4 and PRDX1. IL-17A is less detected in intestinal inflammation, presenting Nrf2 factor. The present findings suggest that Nrf2 function in the gut plays a role in arresting both inflammatory response and oxidative damages of UC.

High Expression of Peroxiredoxin 1 Is Associated with Epithelial-Mesenchymal Transition Marker and Poor Prognosis in Gastric Cancer.

BACKGROUND Recent studies show that peroxiredoxin 1 (Prdx1) contributes to the progression and poor prognosis of carcinoma through multiple mechanisms. However, there is little information on its expression and prognostic value in gastric cancer. This study investigated the expression of Prdx1 in gastric cancer, along with evaluating its clinical-pathological and prognostic importance. MATERIAL AND METHODS A total of 189 pairs of gastric cancer and paracarcinomatous tissues were assessed for Prdx1 expression and its association with clinical characteristics. The molecular mechanism was further investigated through in vitro experimentation. RESULTS The mRNA and protein levels of Prdx1 in the GC tissues were higher than in the peri-tumor tissues. We also found that high Prdx1 expression was positively correlated with the lymph node invasion and poor prognosis. It also served as an autonomous prognostic factor for patients with gastric cancer. Moreover, Prdx1 regulates the invasion and metastasis of GC cell lines through inhibiting E-Ca expression. CONCLUSIONS Prdx1 can promote epithelial-mesenchymal transition and gastric cancer progression. Therefore, it might be a therapeutic target and prognostic indicator for gastric cancer patients.

Galectin-3 down-regulates antioxidant peroxiredoxin-4 in human cardiac fibroblasts: a new pathway to induce cardiac damage.

Galectin-3 (Gal-3) is increased in heart failure (HF) and promotes cardiac fibrosis and inflammation. We investigated whether Gal-3 modulates oxidative stress in human cardiac fibroblasts, in experimental animal models and in human aortic stenosis (AS). Using proteomics and immunodetection approaches, we have identified that Gal-3 down-regulated the antioxidant peroxiredoxin-4 (Prx-4) in cardiac fibroblasts. In parallel, Gal-3 increased peroxide, nitrotyrosine, malondialdehyde, and N-carboxymethyl-lysine levels and decreased total antioxidant capacity. Gal-3 decreased prohibitin-2 expression without modifying other mitochondrial proteins. Prx-4 silencing increased oxidative stress markers. In Gal-3-silenced cells and in heart from Gal-3 knockout mice, Prx-4 was increased and oxidative stress markers were decreased. Pharmacological inhibition of Gal-3 with modified citrus pectin restored cardiac Prx-4 as well as prohibitin-2 levels and improved oxidative status in spontaneously hypertensive rats. In serum from 87 patients with AS, Gal-3 negatively correlated with total antioxidant capacity and positively correlated with peroxide. In myocardial biopsies from 26 AS patients, Gal-3 up-regulation paralleled a decrease in Prx-4 and in prohibitin-2. Cardiac Gal-3 inversely correlated with Prx-4 levels in myocardial biopsies. These data suggest that Gal-3 decreased Prx-4 antioxidant system in cardiac fibroblasts, increasing oxidative stress. In pathological models presenting enhanced cardiac Gal-3, the decrease in Prx-4 expression paralleled increased oxidative stress. Gal-3 blockade restored Prx-4 expression and improved oxidative stress status. In AS, circulating levels of Gal-3 could reflect oxidative stress. The alteration of the balance between antioxidant systems and reactive oxygen species production could be a new pathogenic mechanism by which Gal-3 induces cardiac damage in HF.

Mutation in GNE Downregulates Peroxiredoxin IV Altering ER Redox Homeostasis.

GNE myopathy is a rare neuromuscular genetic disorder characterized by early adult onset and muscle weakness due to mutation in sialic acid biosynthetic enzyme, UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE). More than 180 different GNE mutations are known all over the world with unclear pathomechanism. Although hyposialylation of glycoproteins is speculated to be the major cause, but cellular mechanism leading to loss of muscle mass has not yet been deciphered. Besides sialic acid biosynthesis, GNE affects other cellular functions such as cell adhesion and apoptosis. In order to understand the effect of mutant GNE protein on cellular functions, differential proteome profile of HEK293 cells overexpressing pathologically relevant recombinant mutant GNE protein (D207V and V603L) was analyzed. These cells, along with vector control and wild-type GNE-overexpressing cells, were subjected to two-dimensional gel electrophoresis coupled with mass spectrometry (MALDI-TOF/TOF MS/MS). In the study, 10 differentially expressed proteins were identified. Progenesis same spots software revealed downregulation of peroxiredoxin IV (PrdxIV), an ER-resident H2O2 sensor that regulates neurogenesis. Significant reduction in mRNA and protein levels of PrdxIV was observed in GNE mutant cell lines compared with vector control. However, neither total reactive oxygen species was altered nor H2O2 accumulation was observed in GNE mutant cell lines. Interestingly, ER redox state was significantly affected due to reduced normal GNE enzyme activity. Our study indicates that downregulation of PrdxIV affects ER redox state that may contribute to misfolding and aggregation of proteins in GNE myopathy.

Nrf2-peroxiredoxin I axis in polymorphous adenocarcinoma is associated with low matrix metalloproteinase 2 level.

Polymorphous adenocarcinoma (PAC) is a malignant epithelial neoplasm that affects almost exclusively the minor salivary glands, generally described as having a relatively good prognosis. Aberrant nuclear factor erythroid 2 (NF-E2)-related factor (Nrf2) activation in tumor cells has been associated with induction of antioxidant enzymes, such as peroxiredoxin I (Prx I) and increased matrix metalloproteinase (MMP) expression. In this context, the aim of the present study was to evaluate the expression of Nrf2 and correlate it with Prx I and MMP-2 secretion in PAC. Thirty-one cases of PAC from oral biopsies were selected and immunohistochemically analyzed for Nrf2 and Prx I. MMP-2 quantification was performed on primary cell cultures derived from PAC. Oral squamous cell carcinoma (OSCC) cell cultures were used as control. A high immunoexpression of Nrf2 was observed in both the cytoplasm and the nucleus of neoplastic cells from PAC. Nuclear staining for Nrf2 suggested its activation in the majority of the PAC cells, which was confirmed by the high expression of its target gene, Prx I. Quantification of MMP-2 secretion showed lower levels in PAC cell cultures when compared to OSCC cell cultures (p < 0.05). In conclusion, although Nrf2 overexpression has been frequently associated with high-grade malignancies, such relationship is not infallible and, in fact, the opposite may occur in low-grade tumors, such as PAC of minor salivary glands.

Proteome analysis reveals differential expression of proteins involved in triacylglycerol accumulation by Rhodococcus jostii RHA1 after addition of methyl viologen.

Rhodococcus jostii RHA1 is able to degrade toxic compounds and accumulate high amounts of triacylglycerols (TAG) upon nitrogen starvation. These NADPH-dependent processes are essential for the adaptation of rhodococci to fluctuating environmental conditions. In this study, we used an MS-based, label-free and quantitative proteomic approach to better understand the integral response of R. jostii RHA1 to the presence of methyl viologen (MV) in relation to the synthesis and accumulation of TAG. The addition of MV promoted a decrease of TAG accumulation in comparison to cells cultivated under nitrogen-limiting conditions in the absence of this pro-oxidant. Proteomic analyses revealed that the abundance of key proteins of fatty acid biosynthesis, the Kennedy pathway, glyceroneogenesis and methylmalonyl-CoA pathway, among others, decreased in the presence of MV. In contrast, some proteins involved in lipolysis and ß-oxidation of fatty acids were upregulated. Some metabolic pathways linked to the synthesis of NADPH remained activated during oxidative stress as well as under nitrogen starvation conditions. Additionally, exposure to MV resulted in the activation of complete antioxidant machinery comprising superoxide dismutases, catalases, mycothiol biosynthesis, mycothione reductase and alkyl hydroperoxide reductases, among others. Our study suggests that oxidative stress response affects TAG accumulation under nitrogen-limiting conditions through programmed molecular mechanisms when both stresses occur simultaneously.

OxyR2 Functions as a Three-state Redox Switch to Tightly Regulate Production of Prx2, a Peroxiredoxin of Vibrio vulnificus.

The bacterial transcriptional regulator OxyR is known to function as a two-state redox switch. OxyR senses cellular levels of H2O2 via a "sensing cysteine" that switches from the reduced to a disulfide state upon H2O2 exposure, inducing the expression of antioxidant genes. The reduced and disulfide states of OxyR, respectively, bind to extended and compact regions of DNA, where the reduced state blocks and the oxidized state allows transcription and further induces target gene expression by interacting with RNA polymerase. Vibrio vulnificus OxyR2 senses H2O2 with high sensitivity and induces the gene encoding the antioxidant Prx2. In this study, we used mass spectrometry to identify a third redox state of OxyR2, in which the sensing cysteine was overoxidized to S-sulfonated cysteine (Cys-SO3H) by high H2O2 in vitro and in vivo, where the modification deterred the transcription of prx2 The DNA binding preferences of OxyR25CA-C206D, which mimics overoxidized OxyR2, suggested that overoxidized OxyR2 binds to the extended DNA site, masking the -35 region of the prx2 promoter. These combined results demonstrate that OxyR2 functions as a three-state redox switch to tightly regulate the expression of prx2, preventing futile production of Prx2 in cells exposed to high levels of H2O2 sufficient to inactivate Prx2. We further provide evidence that another OxyR homolog, OxyR1, displays similar three-state behavior, inviting further exploration of this phenomenon as a potentially general regulatory mechanism.

[Xenopus laevis peroxiredoxins: Gene expression during development and characterization of the enzymes].

Reactive oxygen species (ROS) are produced via catabolic and anabolic processes during normal embryonic development, and ROS content in the cell is maintained at a certain level. Peroxiredoxins are a family of selenium-independent peroxidases and play a key role in maintaining redox homeostasis of the cell. In addition to regulating the ROS level, peroxiredoxins are involved in intracellular and intercellular signaling, cell differentiation, and tissue development. The time course of peroxiredoxin gene (prx1-6) expression was studied in Xenopus laevis during early ontogeny (Nieuwkoop and Faber stages 10-63). The highest expression level was observed for prx1 at these developmental stages. The prx1, prx3, and prx4 expression level changed most dramatically in response to oxidative stress artificially induced in X. laevis embryos. In X. laevis adults, prx1-6 were all intensely expressed in all organs examined, the prx1 expression level being the highest. The X. laevis prx1-6 genes were cloned and expressed in Escherichia coli, and physico-chemical characteristics were compared for the recombinant enzymes. The highest peroxidase activity and thermal stability were observed for Prx1 and Prx2. It was assumed that Prx1 plays a leading role in X. laevis early development.

Proteomic analysis of ginsenoside Re attenuates hydrogen peroxide-induced oxidative stress in human umbilical vein endothelial cells.

Ginsenoside Re is an active component in ginseng that has attracted much attention because of its evident therapeutic effects on the cardiovascular system. However, little basic information is available on the mechanisms and pharmacological effects of ginsenoside Re. The potential mechanisms and protective effects of Re on H2O2-induced oxidative injury in human umbilical vein endothelial cells (HUVECs) were investigated in this study. An oxidative injury model was established using H2O2. The anti-oxidative effects of Re were determined using a series of experiments, such as MTT and anti-oxidative indicator assays. The potential protective mechanisms of Re were explored at the proteomic level, and differentially expressed proteins were validated by quantitative real-time polymerase chain reaction and western blotting. Results indicated that Re could be a potential anti-oxidant to protect HUVECs against oxidative stress damage. Proteomic analysis showed that the expression of 23 protein spots was upregulated in Re and H2O2 groups to resist oxidative stress, 15 of which were identified by their mass spectrum. These upregulated proteins were involved in stress response, anti-oxidative systems, protein synthesis, regulation of transcription and post-translational modifications, and repair of mitochondrial functions. This study may provide new insights into the mechanisms of ginsenoside Re in protecting the cardiovascular system.