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During systemic inflammation, indoleamine 2,3-dioxygenase 1 (IDO1) becomes expressed in endothelial cells where it uses hydrogen peroxide (H2O2) to oxidize L-tryptophan to the tricyclic hydroperoxide, cis-WOOH, that then relaxes arteries via oxidation of protein kinase G 1α. Here we show that arterial glutathione peroxidases and peroxiredoxins that rapidly eliminate H2O2, have little impact on relaxation of IDO1-expressing arteries, and that purified IDO1 forms cis-WOOH in the presence of peroxiredoxin 2. cis-WOOH oxidizes protein thiols in a selective and stereospecific manner. Compared with its epimer trans-WOOH and H2O2, cis-WOOH reacts slower with the major arterial forms of glutathione peroxidases and peroxiredoxins while it reacts more readily with its target, protein kinase G 1α. Our results indicate a paradigm of redox signaling by H2O2 via its enzymatic conversion to an amino acid-derived hydroperoxide that ‘escapes’ effective reductive inactivation to engage in selective oxidative activation of key target proteins., A major question in redox signaling is how H2O2 oxidizes target protein thiols in the presence of glutathione peroxidases and peroxiredoxins. We reveal signaling by H2O2 via its enzymatic conversion to an alkyl hydroperoxide that stereo-specifically escapes peroxidases and oxidizes target proteins.

The peroxiredoxins (PRXs) constitute a ubiquitous antioxidant. Growing evidence in neurodegenerative disorders such as Parkinson’s disease (PD) or Alzheimer’s disease (AD) has highlighted a crucial role for PRXs against neuro-oxidation. Chorea-acanthocytosis/Vps13A disease (ChAc) is a devastating, life-shortening disorder characterized by acanthocytosis, neurodegeneration and abnormal proteostasis. We recently developed a Vps13a−/− ChAc-mouse model, showing acanthocytosis, neurodegeneration and neuroinflammation which could be restored by LYN inactivation. Here, we show in our Vps13a−/− mice protein oxidation, NRF2 activation and upregulation of downstream cytoprotective systems NQO1, SRXN1 and TRXR in basal ganglia. This was associated with upregulation of PRX2/5 expression compared to wild-type mice. PRX2 expression was age-dependent in both mouse strains, whereas only Vps13a−/− PRX5 expression was increased independent of age. LYN deficiency or nilotinib-mediated LYN inhibition improved autophagy in Vps13a−/− mice. In Vps13a−/−; Lyn−/− basal ganglia, absence of LYN resulted in reduced NRF2 activation and down-regulated expression of PRX2/5, SRXN1 and TRXR. Nilotinib treatment of Vps13a−/− mice reduced basal ganglia oxidation, and plasma PRX5 levels, suggesting plasma PRX5 as a possible ChAc biomarker. Our data support initiation of therapeutic Lyn inhibition as promptly as possible after ChAc diagnosis to minimize development of irreversible neuronal damage during otherwise inevitable ChAc progression.

Albeit endometriosis is one of the most common gynecological diseases, its diagnosis and treatment remain controversial. The reasons behind this include: 1) multifactorial pathogenesis and insufficiently studied mechanisms of endometriosis; 2) relatively low diagnostic value of minimally invasive examination in relation to this disease; 3) inefficiency of current therapeutic approaches in many patient settings. In our opinion, uncovering the causes of endometriosis and factors promoting its progression is the cornerstone of its successful management. Here we review the lessons from genome-wide and candidate gene association studies, discuss the expression of regulatory miRNAs and describe the role of heat shock protein 90, annexin A2, and peroxiredoxin 2 in controlling DNA integrity in the eutopic endometrium. Further, we highlight the role of cytokeratin-19 in urine as a feasible diagnostic marker of endometriosis. Clinicians and basic researchers concur that the molecular basis of endometriosis is still in its infancy and current understanding of its pathophysiology remains poor. Recent progress in -omics approaches and bioinformatics paved the way for complex investigations of regulated cell death, proliferation, cell invasion and angiogenesis, opening the avenue for the novel approaches to treat endometriosis. Yet, the diversity of symptoms and an absence of sensitive and specific biomarkers frequently delay and complicate the diagnosis. In addition, surgery represents the only appropriate option to reliably confirm the diagnosis and to establish the disease extent, reducing patient adherence and postponing the start of the treatment. In this review, we discuss challenges in the diagnosis of endometriosis as well as relevant and potentially informative biomarkers.

In Alzheimer’s disease, reactive oxygen species (ROS) are generated by the deposition of amyloid-beta oligomers (AβOs), which represent one of the important causes of neuronal cell death. Additionally, AβOs are known to induce autophagy via ROS induction. Previous studies have shown that autophagy upregulation aggravates neuronal cell death. In this study, the effects of peroxiredoxin 2 (Prx2), a member of the peroxidase family of antioxidant enzymes, on regulating AβO-mediated autophagy were investigated. Prx2 decreased AβO-mediated oxidative stress and autophagy in N2a-APPswe cells. Further, we examined the relationship between the neuronal protective effect of Prx2 and a decrease in autophagy. Similar to the effects of N-acetyl cysteine, Prx2 decreased AβO-induced ROS and inhibited p62 protein expression levels by downregulating the activation of NRF2 and its translocation to the nucleus. In addition, treatment with 3-methyladenine, an autophagy inhibitor, ameliorates neuronal cell death. Overall, these results demonstrate that the Prx2-induced decrease in autophagy was associated with the inhibition of ROS via the ROS–NRF2–p62 pathway in N2a-APPswe cells. Therefore, our results revealed that Prx2 is a potential therapeutic target in anti-Alzheimer therapy.

To explore gene expression and serum profile of selected antioxidant markers during periparturient period in dromedary camels, blood samples were collected from sixty apparently healthy pregnant female at − 14, 0, and + 14 days of expected date of delivery. Superoxide dismutase1 (SOD1), superoxide dismutase 3 (SOD3), catalase (CAT), peroxiredoxin 2 (PRDX2), peroxiredoxin 3 (PRDX3), peroxiredoxin 4 (PRDX4), peroxiredoxin 6 (PRDX6), glutathione peroxidase (GPX), and alkyl hydroperoxide reductase/thiol-specific antioxidant (AhpC/TSA) markers showed upregulations at (− 14) and (+ 14) compared to their values at calving. A significant upregulation of oxidative stress-responsive kinase 1 (OXSR1), stress-associated endoplasmic reticulum proteins (SERP2), and stress-induced phosphoprotein (STIP1) was also reported at (− 14) compared to their corresponding values at calving and (+ 14). At (− 14), serum GSH levels were low, and increased gradually until early lactation. Total antioxidant capacity (TAC) concentrations were low at (− 14) and at parturition, and the peak values were observed at (+ 14), whereas an opposite trend was noted for Malondialdehyde (MDA) levels. SOD activity was lower at (0) and (+ 14) compared to at (− 14). Glutathione peroxidase (GSH-Px) was also low at (0) and (+ 14) days postpartum compared to that at (− 14). Linear discriminant analysis (LDA) showed that gene expression and serum profile of antioxidant markers could discriminate peri-parturient period. The overall correctly classified percent was 62.0% and 48.1% for results of gene expression and serum parameters respectively. Profound alteration in antioxidant parameters could be a biomarker to that helps follow-up health during the peri-parturient period in order to minimize the incidence and severity of diseases and, consequently, to build up a good management protocol.

Hibernation is a strategy used by some mammals to survive harsh winter conditions. Many small mammals, such as the little brown bat, Myotis lucifugus, enter a long-term state of hibernation characterized by a period of deep torpor that can range from days to weeks. Torpid bats undergo metabolic rate depression that not only results in physiological changes but also promotes biochemical changes that favor survival. The present study utilizes multiplex technology to assess key early apoptosis markers and a select group of antioxidant enzymes in muscle, heart, and liver in euthermic controls and torpid bats. Muscle showed a significant decrease in the proapoptotic c-Jun N-terminal kinase and p53 and the antioxidant enzyme catalase but a significant increase in peroxiredoxin 2 levels. The heart responded similarly, with most proapoptotic proteins (caspase 8/9 and p53) remaining at low levels, while the antiapoptotic Bcl-2 protein significantly increased during torpor. There was no significant change in the antioxidant enzymes measured during torpor in the heart compared with the controls. The liver showed increases in catalase and Mn superoxide dismutase 2 enzymes during torpor, which correlated with activation of select antiapoptotic proteins and suppression of levels of proapoptotic ones. Overall, our data demonstrate that antiapoptotic and antioxidant defense responses have organ-specific regulation during torpor in bats. The induction of key antioxidant enzymes and antiapoptotic proteins may function as protective mechanisms that are necessary for surviving torpor.

Background and purpose Oxidative stress and neuronal apoptosis play key roles in traumatic brain injury. We investigated the protective effects of astaxanthin against traumatic brain injury and its underlying mechanisms of action. Experimental approach A weight-drop model of traumatic brain injury in vivo and hydrogen peroxide exposure in vitro model were established. Brain oedema, behaviour tests, western blot, biochemical analysis, lesion volume, histopathological study and cell viability were performed. Key results Astaxanthin significantly reduced oxidative insults on Days 1, 3 and 7 after traumatic brain injury. Neuronal apoptosis was also ameliorated on Day 3. Additionally, astaxanthin improved neurological functions up to 3 weeks after traumatic brain injury. Astaxanthin treatment dramatically enhanced the expression of peroxiredoxin 2 (Prx2), nuclear factor-erythroid 2-related factor 2 (NRF2/Nrf2) and sirtuin 1 (SIRT1), while it down-regulated the phosphorylation of apoptosis signal-regulating kinase 1 (ASK1) and p38. Inhibition of Prx2 by siRNA injection reversed the beneficial effects of astaxanthin against traumatic brain injury. Additionally, Nrf2 knockout prevented the neuroprotective effects of astaxanthin in traumatic brain injury. In contrast, overexpression of Prx2 in Nrf2 knockout mice attenuated the secondary brain injury after traumatic brain injury. Moreover, inhibiting SIRT1 by EX527 dramatically inhibited the neuroprotective effects of astaxanthin and suppressed SIRT1/Nrf2/Prx2/ASK1/p38 pathway both in vivo and in vitro. Conclusion and implications Astaxanthin improved the neurological functions and protected the brain from injury after traumatic brain injury, primarily by reducing oxidative stress and neuronal death via SIRT1/Nrf2/Prx2/ASK1/p38 signalling pathway and might be a new candidate to ameliorate traumatic brain injury.

Responding appropriately to exercise is essential to maintenance of skeletal muscle mass and function at all ages and particularly during aging. Here, a hypothesis is presented that a key component of the inability of skeletal muscle to respond effectively to exercise in aging is a denervation-induced failure of muscle redox signalling. This novel hypothesis proposes that an initial increase in oxidation in muscle mitochondria leads to a paradoxical increase in the reductive state of specific cysteines of signalling proteins in the muscle cytosol that suppresses their ability to respond to normal oxidising redox signals during exercise. The following are presented for consideration:Transient loss of integrity of peripheral motor neurons occurs repeatedly throughout life and is normally rapidly repaired by reinnervation, but this repair process becomes less efficient with aging. Each transient loss of neuromuscular integrity leads to a rapid, large increase in mitochondrial peroxide production in the denervated muscle fibers and in neighbouring muscle fibers. This peroxide may initially act to stimulate axonal sprouting and regeneration, but also stimulates retrograde mitonuclear communication to increase expression of a range of cytoprotective proteins in an attempt to protect the fiber and neighbouring tissues against oxidative damage. The increased peroxide within mitochondria does not lead to an increased cytosolic peroxide, but the increases in adaptive cytoprotective proteins include some located to the muscle cytosol which modify the local cytosol redox environment to induce a more reductive state in key cysteines of specific signalling proteins. Key adaptations of skeletal muscle to exercise involve transient peroxiredoxin oxidation as effectors of redox signalling in the cytosol. This requires sensitive oxidation of key cysteine residues. In aging, the chronic change to a more reductive cytosolic environment prevents the transient oxidation of peroxiredoxin 2 and hence prevents essential adaptations to exercise, thus contributing to loss of muscle mass and function. Experimental approaches suitable for testing the hypothesis are also outlined., Graphical abstract Image 1, Highlights • It is hypothesised that denervation leads to failed redox signalling and attenuated muscle exercise responses in aging. • Loss of peripheral motor neurons leads to increased mitochondrial peroxide in denervated and neighbouring muscle fibers. • This peroxide stimulates increased expression of cytoprotective proteins to protect the fiber against oxidative damage. • Increased mitochondrial peroxide does not increase cytosolic peroxide, but adaptations reduce the cytosol redox state. • Key adaptations of skeletal muscle to exercise involve transient peroxiredoxin oxidation in the cytosol. • Peroxiredoxin oxidation is suppressed during aging by a chronic change to a more reductive local cytosolic environment.

Purpose Hodgkin's disease (HD) is a type of cancer affecting men in the reproductive age with potential consequences on their fertility status. This study aims to analyze sperm parameters, alterations in proteomic profiles and validate selected protein biomarkers of spermatozoa in men with HD undergoing sperm banking before cancer therapy. Materials and methods Semen analysis was carried out in healthy fertile donors (control, n=42), and patients diagnosed with HD (patients, n=38) before cancer therapy. We compared proteomic profiles of spermatozoa from donors (n=3) and patients (n=3) using LTQ-Orbitrap Elite hybrid MS system. Results A total of 1,169 proteins were identified by global proteomic in both groups. The ingenuity pathway analysis revealed that differentially expressed proteins involved in capacitation, acrosome reaction, binding of sperm to the zona pellucida, sperm motility, regulation of sperm DNA damage, and apoptosis were significantly downregulated in HD patients. Validation of proteins implicated in sperm fertility potential by Western Blot demonstrated that peroxiredoxin 2 (PRDX 2) was underexpressed (p=0.015), and transferrin (p=0.045) and SERPIN A5 (p=0.010) protein levels were overexpressed in spermatozoa of men with HD. Conclusions Findings of this study indicates that the key proteins involved in sperm fertility potential are significantly altered in men with HD, which provides substantial explanation for the observed low sperm quality in HD subjects prior to cancer therapy. Furthermore, our results suggest PRDX 2, transferrin and SERPIN A5 as possible candidate proteins for assessing sperm quality in HD patients prior to cancer therapy.

Leucine-rich repeat kinase 2 (LRRK2) is a complex protein kinase involved in a diverse set of functions. Mutations in LRRK2 are a common cause of autosomal dominant familial Parkinson’s disease. Peroxiredoxin 2 (PRDX2) belongs to a family of anti-oxidants that protect cells from oxidative stress. Importantly, PRDX2 is a cytoplasmic protein, similar to Leucine-rich repeat kinase 2, which localizes predominantly in the cytosol. Here, we demonstrated that Leurice-rich repeat kinase 2 phosphorylates PRDX2 in Drosophila, leading to a loss of dopaminergic neurons, climbing ability and shortened lifespan. These pathogenic phenotypes in the LRRK2 Drosophila were rescued with transgenic expression of PRDX2. Chetomin, a PRDX2 mimic, belongs to a class of epidithio-diketopiperazine fungal secondary metabolites (containing a dithiol group that has hydrogen peroxide-reducing activity). As proof of principle, we demonstrated that Chetomin recapitulated the rescue in these mutant Drosophila. Our findings suggest that Chetomin can be a potential therapeutic compound in LRRK2 linked Parkinson’s disease.

Abdominal aortic aneurysm (AAA) is an inflammatory vascular disease characterized by structural deterioration of the aorta caused by inflammation and oxidative stress, leading to aortic dilatation and rupture. Peroxiredoxin 2 (PRDX2), an antioxidant enzyme, has been reported as a potential negative regulator of inflammatory vascular diseases, and it has been identified as a protein that is increased in patients with ruptured AAA compared to patients with nonruptured AAA. In this study, we demonstrated that PRDX2 was a pivotal factor involved in the inhibition of AAA progression. PRDX2 levels were increased in AAA compared with those in normal aortas in both humans and mice. Ultrasound imaging revealed that the loss of PRDX2 accelerated the development of AAA in the early stages and increased AAA incidence in mice infused with angiotensin II (Ang II). Prdx2−/− mice infused with Ang II exhibited increased aortic dilatation and maximal aortic diameter without a change in blood pressure. Structural deterioration of the aortas from Prdx2−/− mice infused with Ang II was associated with increases in the degradation of elastin, oxidative stress, and intramural thrombi caused by microhemorrhages, immature neovessels, and the activation of matrix metalloproteinases compared to that observed in controls. Moreover, an increase in inflammatory responses, including the production of cell adhesion molecules and the accumulation of inflammatory cells and proinflammatory cytokines due to PRDX2 deficiency, accelerated Ang II-induced AAA progression. Our data confirm that PRDX2 plays a role as a negative regulator of the pathological process of AAA and suggest that increasing PRDX2 activity may be a novel strategy for the prevention and treatment of AAA., Abdominal aortic aneurysm: Potential enzyme biomarker identified An enzyme with antioxidant properties may provide a biomarker and therapeutic agent to help treat abdominal aortic aneurysm (AAA). AAA involves the structural deterioration of the aorta through chronic inflammation and oxidative stress, and can trigger life-threatening artery rupture. An antioxidant enzyme called peroxiredoxin 2 (PRDX2) is increased in patients with ruptures, but whether its role in AAA is beneficial or detrimental is unclear. Goo Taeg Oh at the Ewha Womans University in Seoul, Jong-Gil Park at the Korea Research Institute of Bioscience and Biotechnology, Daejeon, South Korea, and co-workers examined the effect of PRDX2 on AAA progression. PRDX2 suppressed structural damage in mice, limiting artery dilation and protein degradation. Loss of PRDX2 accelerated AAA development. Measuring levels of PRDX2 may indicate AAA severity in patients, while boosting the enzyme could repair aortic damage.

Background: Oxidative stress from elevated reactive oxygen species (ROS) has been reported to induce cell apoptosis and may provide a means to target cancer cells. Celastrol is a natural bioactive compound that was recently shown to increase ROS levels and cause apoptosis in cancer cells. However, the underlying mechanism for this cytotoxic action remains unclear and direct molecular targets of Celastrol have not been identified. Methods: Proteome microarray, surface plasmon resonance, isothermal titration calorimetry and molecular simulation were used to identify the molecular target of Celastrol. Binding and activity assays were used to validate the interaction of Celastrol with target protein in cell-free and gastric cancer cell lysates. We then assessed target transcript levels in in biopsy specimens obtained from patients with gastric cancer. Gastric cancer growth-limiting and cytotoxic activity of Celastrol was evaluated in BALB/c nu/nu mice. Results: Our data show that Celastrol directly binds to an antioxidant enzyme, peroxiredoxin-2 (Prdx2), which then inhibits its enzyme activity at both molecular and cellular level. Inhibition of Prdx2 by Celastrol increased cellular ROS levels and led to ROS-dependent endoplasmic reticulum stress, mitochondrial dysfunction, and apoptosis in gastric cancer cells. Functional tests demonstrated that Celastrol limits gastric cancer cells, at least in part, through targeting Prdx2. Celastrol treatment of mice implanted with gastric cancer cells also inhibited tumor growth, associated with Prdx2 inhibition and increased ROS. Analysis of human gastric cancer also showed increased Prdx2 levels and correlation with survival. Conclusion: Our studies have uncovered a potential Celastrol-interacting protein Prdx2 and a ROS-dependent mechanism of its action. The findings also highlight Prdx2 as a potential target for the treatment of gastric cancer.

Colorectal cancer (CRC) is a prevalent worldwide disease in which the antioxidant enzyme peroxiredoxin 2 (PRDX2) plays an important role. To investigate the molecular mechanism of PRDX2 in CRC, we performed bioinformatics analysis of The Cancer Genome Atlas (TCGA) datasets and Gene Expression Omnibus (GEO) DataSets (accession no. GSE81429). Our results suggest that PRDX2 is associated with cell-cycle progression and autophagy activated by the P38 MAPK/FOXO signaling pathway. Using a short-hairpin RNA vector, we verified that PRDX2 is essential for CRC cell proliferation and S-phase progression. Immunostaining, electron microscopy and western blotting assays verified the effect of PRDX2 knockdown on autophagy flux and p38 activation. The P38 activator dehydrocorydaline chloride partially rescued the effects of sh-PRDX2 on the expression of proteins related to cell-cycle progression and autophagy. We verified the correlation between PRDX2 expression and the expression of an array of cell-cycle and autophagy-related genes using data from an independent set of 222 CRC patient samples. A mouse xenoplast model was consistent with in vitro results. Our results suggest that PRDX2 promotes CRC cell-cycle progression via activation of the p38 MAPK pathway.

Background Hepatocellular carcinoma (HCC) has a high incidence and mortality that epitomizes one of the prominent causes of cancer-related death globally. Novel therapeutic approaches are therefore required. Reactive oxygen species (ROS) are necessary for maintaining cell cycle. Although ROS is involved in HCC progression, hydrogen peroxide (H2O2) has anti-proliferative effect on HCC. Method HCC Huh-7 cells were cultured and incubated with various concentrations of H2O2. Paraoxonase activity, levels of malondialdehyde, glutathione and protein oxidation were measured in treated and untreated Huh-7 cells. Furthermore, untreated and treated Huh-7 cells were subjected to two dimensional gel electrophoresis and identified protein spots which were differentially expressed by LC-MS/MS analysis. qRT-PCR was performed to validate the identified proteins. Results H2O2 depleted glutathione (GSH) with the concomitant up-regulation of GSTP1 and Prx2. H2O2 also increased malondialdehyde and protein oxidation, decreased the activity of paraoxonase in Huh-7 cells. Conclusion H2O2 could be used as a novel therapeutic agent that might be beneficial in inducing cell cytotoxicity and hence suppress HCC proliferation.

Background and Purpose— Our recent study demonstrated that release of Prx2 (peroxiredoxin 2) from red blood cells (RBCs) is involved in the inflammatory response and brain injury after intracerebral hemorrhage. The current study investigated the role of extracellular Prx2 in hydrocephalus development after experimental intraventricular hemorrhage. Methods— There were 4 parts in this study. First, Sprague-Dawley rats received an intraventricular injection of lysed RBC or saline and were euthanized at 1 hour for Prx2 measurements. Second, rats received an intraventricular injection of Prx2, deactivated Prx2, or saline. Third, lysed RBC was coinjected with conoidin A, a Prx2 inhibitor, or vehicle. Fourth, rats received Prx2 injection and were treated with minocycline or saline (i.p.). The effects of Prx2 and the inhibitors were examined using magnetic resonance imaging assessing ventriculomegaly, histology assessing ventricular wall damage, and immunohistochemistry to assess inflammation, particularly at the choroid plexus. Results— Intraventricular injection of lysed RBC resulted in increased brain Prx2 and hydrocephalus. Intraventricular injection of Prx2 alone caused hydrocephalus, ventricular wall damage, activation of choroid plexus epiplexus cells (macrophages), and an accumulation of neutrophils. Conoidin A attenuated lysed RBC-induced injury. Systemic minocycline treatment reduced the epiplexus cell activation and hydrocephalus induced by Prx2. Conclusions— Prx2 contributed to the intraventricular hemorrhage-induced hydrocephalus, probably by inducing inflammatory responses in choroid plexus and ventricular wall damage.

Both reactive nitrogen and oxygen species (RNS and ROS), such as nitric oxide, peroxynitrite, and hydrogen peroxide, have been implicated as mediators of pancreatic β-cell damage during the pathogenesis of autoimmune diabetes. While β-cells are thought to be vulnerable to oxidative damage due to reportedly low levels of antioxidant enzymes, such as catalase and glutathione peroxidase, we have shown that they use thioredoxin reductase to detoxify hydrogen peroxide. Thioredoxin reductase is an enzyme that participates in the peroxiredoxin antioxidant cycle. Peroxiredoxins are expressed in β-cells and, when overexpressed, protect against oxidative stress, but the endogenous roles of peroxiredoxins in the protection of β-cells from oxidative damage are unclear. Here, using either glucose oxidase or menadione to continuously deliver hydrogen peroxide, or the combination of dipropylenetriamine NONOate and menadione to continuously deliver peroxynitrite, we tested the hypothesis that β-cells use peroxiredoxins to detoxify both of these reactive species. Either pharmacological peroxiredoxin inhibition with conoidin A or specific depletion of cytoplasmic peroxiredoxin 1 ( Prdx1) using siRNAs sensitizes INS 832/13 cells and rat islets to DNA damage and death induced by hydrogen peroxide or peroxynitrite. Interestingly, depletion of peroxiredoxin 2 ( Prdx2) had no effect. Together, these results suggest that β-cells use cytoplasmic Prdx1 as a primary defense mechanism against both ROS and RNS.

Objective: To examine if peroxiredoxin 2 (Prx2) deficiency aggravates high-fat diet-induced insulin resistance. Material and methods: Insulin sensitivity was measured in Prx2 knockout (KO) and wild-type (WT) littermates using the hyperinsulinemic-euglycemic clamp. Results: Whole body glucose turnover, glucose uptake, and levels of glucose transporter 4 (Glut4) protein in the skeletal muscle were found to be lower. This was followed by increased expression of oxidative stress markers in Prx2 KO mice than that in WT mice in the control diet group. Although, a 12-week high-fat diet induced insulin resistance and enhanced oxidative stress in both genotypes, there was no difference between WT and Prx2 KO mice with respect to insulin sensitivity and the level of oxidative stress markers. Accordingly, the levels of phosphorylated Akt and Glut4 were similar between the two genotypes. Conclusion: These results suggest that Prx2 does not affect high-fat diet-induced oxidative stress and insulin resistance in mice.

To investigate the role of PRDX2 in esophageal carcinoma (ESCA). The expression of PRDX2 was detected in ESCA tissues. And PRDX2 expression in two ESCA cell lines was knocked down. Cell proliferation, metastasis and invasion were detected in these cells. Here, we found that PRDX2 expression was significantly increased in ESCA tissues and was associated with a poor prognosis in ESCA patients. In addition, PRDX2 expression was significantly associated with pathological grading, infiltration degree and 5-year survival time in ESCA patients. Next, we knocked down PRDX2 expression by PRDX2-shRNA transfection in two ESCA cell lines, Eca-109 and TE-1. Proliferation analysis indicated that in vitro PRDX2 knockdown decreased growth and clone formation of ESCA cells. Scratch and transwell assays indicated that cell migration and invasion were significantly inhibited by PRDX2 knockdown. In addition, PRDX2 knockdown inhibited cell cycle of ESCA cells and down-regulated Cyclin D1-CDK4/6. Moreover, PRDX2 knockdown regulated proteins involved in mitochondrial-dependent apoptosis, including increased Bax and Caspase9/3 and decreased Bcl2. Mechanism investigation indicated that PRDX2 knockdown led to inactivation of Wnt/β-catenin and AKT pathways. Our data suggest that PRDX2 may function as an oncogene in the development of ESCA via regulating Wnt/β-catenin and AKT pathways. Our study fills a gap in the understanding of the role of PRDX2 in ESCA and provides a potential target for ESCA treatment.

Background: Currently, the identification of Clara cell and type II alveolar epithelial cell-type cancer cells requires electron microscopy, which is a time-consuming and expensive process involving a complicated tissue sampling procedure. The aim of this study was to identify unique biomarkers for Clara cell and type II alveolar epithelial cell-type lung cancer cells, respectively, with proteomic profiling. Methods: Six human lung adenocarcinoma cell lines (A549, NCI-H358, NCI-H1650, HCC827, NCI-H1395, and NCI-H1975) were investigated for their ultrastructural characteristics. The differentially expressed proteins (DEPs) were screened between NCI-H358 cells (Clara cell type) and A549 cells (type II alveolar epithelial cell type) using two-dimensional difference gel electrophoresis (2D-DIGE) and matrix- assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS/MS), and then they were validated by western blot. The protein expression levels of endoplasmic reticulum oxidoreductin 1-α (ERO1L), Clara cell 10-kD protein (CC10), and surfactant protein C (SP-C) were also determined in the six cell lines assayed. Results: NCI-H358 cells featured Clara cell differentiation; A549, NCI-H1975, and HCC827 cells had characteristics of type II alveolar epithelial cells; and NCI-H1395 and NCI-H1650 cells had no differentiation characteristics of any lung adenocarcinoma cell type. Five DEPs including ubiquitin carboxyl- terminal hydrolase isozyme L1 (UCHL1), cytokeratin 19 (CK19), cytokeratin 8 (CK8), ERO1L, and peroxiredoxin 2 (PRDX2) between NCI-H358 and A549 cells were identified for further validation; however, none of them showed suitability as an effective biomarker. Similarly, CC10 and SP-C were not appropriate biomarkers. Conclusions: Cytological subtypes of NCI-H1975 and HCC827 cells were identified, but no promising biomarker was discovered in the present study.

Oxidative stress and neuroinflammation are closely related to the pathological processes of neurological disorders. Peroxiredoxin 2 (Prdx2) is an abundant antioxidant enzyme in the central nervous system. Prdx2 reduces the production of reactive oxygen species and participates in regulating various signaling pathways in neurons by catalyzing hydrogen peroxide (H2O2), thereby protecting neurons against oxidative stress and an inflammatory injury. However, the spillage of Prdx2, as damage-associated molecular patterns, accelerates brain damage after stroke by activating an inflammatory response. The post-translational modifications of Prdx2 also affect its enzyme activity. This review focuses on the effects of Prdx2 and its molecular mechanisms in various neurological disorders.

Hydrogen peroxide (H2O2) acts as a signalling molecule by oxidising cysteine thiols in proteins. Recent evidence has established a role for cytosolic peroxiredoxins in transmitting H2O2-based oxidation to a multitude of target proteins. Moreover, it is becoming clear that peroxiredoxins fulfil their function in organised microdomains, where not all interactors are covalently bound. However, most studies aimed at identifying peroxiredoxin interactors were based on methods that only detect covalently linked partners. Here, we explore the applicability of two thiol-disulphide independent in-cell trapping methodological approaches in combination with mass spectrometry for the identification of interaction partners of peroxiredoxin 2 (Prdx2). The first is biotin-dependent proximity-labelling (BioID) with a biotin ligase A (BirA*)-fused Prdx2, which has never been applied on redox-active proteins. The second is crosslinker co-immunoprecipitation with an N-terminally His-tagged Prdx2. During the initial characterisation of the tagged Prdx2 constructs, we found that the His-tag, but not BirA*, compromises the peroxidase and signalling activities of Prdx2. Further, the Prdx2 interactors identified with each approach showed little overlap. We therefore concluded that BioID is a more reliable method than crosslinker co-immunoprecipitation. After a stringent mass spec data filtering, BioID identified 13 interactors under elevated H2O2 conditions, including subunit five of the COP9 signalosome complex (CSN5). The Prdx2:CSN5 interaction was further confirmed in a proximity ligation assay. Taken together, our results demonstrate that BioID can be used as a method for the identification of interactors of Prdxs, and that caution should be exercised when interpreting protein-protein interaction results using tagged Prdxs., Graphical abstract Image 1, Highlights • BioID is a thiol-disulphide independent method that can be used to trap Prdx2 interactors. • Prdx2 interacts with subunit five of the COP9 signalosome complex (CSN5). • N-terminally His-tagged Prdx2 displays compromised peroxidase and signalling activity. • The tag attached to Prdx2 influences its interactome.

Glutathione peroxidase 1 (Gpx1) and peroxiredoxin 2 (Prdx2) belong to the thiol peroxidase family of antioxidants, and have been studied for their antioxidant functions and roles in cancers. However, the physiological significance of Gpx1 and Prdx2 during vertebrate embryogenesis are lacking. Currently, we investigated the functional roles of Gpx1 and Prdx2 during vertebrate embryogenesis using Xenopus laevis as a vertebrate model. Our investigations revealed the zygotic nature of gpx1 having its localization in the eye region of developing embryos, whereas prdx2 exhibited a maternal nature and were localized in embryonic ventral blood islands. Furthermore, the gpx1-morphants exhibited malformed eyes with incompletely detached lenses. However, the depletion of prdx2 has not established its involvement with embryogenesis. A molecular analysis of gpx1-depleted embryos revealed the perturbed expression of a cryba1-lens-specific marker and also exhibited reactive oxygen species (ROS) accumulation in the eye regions of gpx1-morphants. Additionally, transcriptomics analysis of gpx1-knockout embryos demonstrated the involvement of Wnt, cadherin, and integrin signaling pathways in the development of malformed eyes. Conclusively, our findings indicate the association of gpx1 with a complex network of embryonic developmental pathways and ROS responses, but detailed investigation is a prerequisite in order to pinpoint the mechanistic details of these interactions.

Label-free quantitative mass spectrometry was used to analyze the differences in the granulation tissue protein expression profiles of patients with diabetic foot ulcers (DFUs) before and after negative-pressure wound therapy (NPWT) to understand how NPWT promotes the healing of diabetic foot wounds. A total of three patients with DFUs hospitalized for Wagner grade 3 were enrolled. The patients received NPWT for one week. The granulation tissue samples of the patients prior to and following NPWT for one week were collected. The protein expression profiles were analyzed with label-free quantitative mass spectrometry and the differentially expressed proteins (DEPs) in the DFU patients prior to and following NPWT for one week were identified. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses were conducted to annotate the DEPs and DEP-associated signaling pathways. Western blotting and ELISA were performed to validate the results. By comparing the differences in the protein profiles of granulation tissue samples prior to and following NPWT for one week, 36 proteins with significant differences were identified (P

Increasing evidence has shown a correlation between chronic periodontitis (CP) and Alzheimer’s disease (AD). Nevertheless, there is still a lack of direct evidence, and especially key molecules to connect the two diseases. This study aims to investigate potential protein links between CP and AD within the inflammatory aspect. The hippocampus of CP model mice and controls were collected, and changes in protein expression were evaluated using two-dimensional differential in-gel electrophoresis (2D-DIGE) analysis combined with liquid chromatography tandem mass spectrometry. A total of 15 differentially expressed proteins were identified in CP model mice, as compared with the controls. Among them, S100-A9, transthyretin, Cofilin 2, peroxiredoxin 2, and lipocalin-2 were validated by Western blot according to their dual function both in inflammation and AD. Based on 2D-DIGE analysis, CP animal model had higher levels of S100-A9, Cofilin 2, peroxiredoxin 2, and lipocalin-2 compared to controls. The level of Cofilin 2, one of the well-established proteins in the pathology of AD, was strongly correlated with the time course of CP pathology, indicating a specific molecular correlation between CP and AD. Moreover, the in vivo results showed the level of Cofilin 2 increased significantly along with a prominent increase of the phosphorylation of protein phosphatase 2 (PP2A) and tau protein in the cell lysates of Porphyromonas gingivalis (P.g-LPS)-treated SK-N-SH APPwt cells. Cofilin 2 inhibition resulted in a sharp decrease in PP2A dependent of tau phosphorylation. Furthermore, tumor growth factor (TGF)-β1 was one of the most important inflammatory cytokines for the Pg-LPS-induced Cofilin 2 upregulation in SK-N-SH APPwt cells. These results showed inflammation served as the bond between CP and AD, whereas inflammatory related proteins could be the key linkers between the two diseases. Determining the association between CP and AD at the molecular mechanism will not only hold the direct evidence of the association between the two diseases but also provide a new way of preventing and treating AD: the effective prevention and treatment of CP could serve as a useful method to alleviate the development of AD.

Background Circular RNAs (circRNAs) play important roles in cancer development and progression. The purpose of this study is to identify aberrantly expressed circRNAs in gastric cancer (GC), unravel their roles in GC progression, and provide new targets for GC diagnosis and therapy. Methods Bioinformatic analyses were performed to identify the aberrantly expression of hsa_circ_0061137 (termed as circDIDO1) in GC. Gain- and loss-of-function studies were performed to examine the biological roles of circDIDO1 in GC progression. Tagged RNA affinity purification, mass spectrometry, immunofluorescence, co-immunoprecipitation, and Western blot were used to identify circRNA-interacting and circRNA-encoded proteins. RNA sequencing, qRT-PCR, and Western blot were performed to analyze circRNA-regulated downstream target genes and signaling pathways. Mouse tumor models were used to analyze the effects of circDIDO1 on GC growth and metastasis. Results CircDIDO1 was transcribed from human DIDO1 (death-inducer obliterator 1) gene and formed by back-splicing of exons 2–6 of the linear transcript. circDIDO1 was down-regulated in GC tissues and its low levels were associated with larger tumor size, distal metastasis, and poor prognosis. CircDIDO1 overexpression inhibited while knockdown promoted GC cell proliferation, migration and invasion. CircDIDO1 overexpression suppressed GC growth and metastasis in mouse tumor models. Mechanistically, circDIDO1 encoded a novel 529aa protein that directly interacted with poly ADP-ribose polymerase 1 (PARP1) and inhibited its activity. CircDIDO1 also specifically bound to peroxiredoxin 2 (PRDX2) and promoted RBX1-mediated ubiquitination and degradation of PRDX2, which led to the inactivation of its downstream signaling pathways. Conclusions CircDIDO1 is a new circRNA that has tumor suppressor function in GC and it may serve as a potential prognostic biomarker and therapeutic target for GC.

Background: The increasing incidence and mortality rate of HCC is a major concern, especially for developing countries of the world. Hence, extensive research is being carried out in order to explore new approaches for developing successful therapeutic strategies for HCC. The controversial role of oxidative stress in the prognosis and treatment of various diseases such as cancer has become an area of great interest and intrigue for many scientists throughout the world. Objective: We aim to investigate the role of induced oxidative stress on the suppression of HCC Huh-7 cancerous cells as a therapeutic approach. Methods: Induction of oxidative stress via H2O2 treatment produced cell cytotoxicity in a dose dependent manner and also led to the overexpression of GSTP-1 and PRX-2. The expression of GSTP- 1 and PRX-2 was compared in HCC Huh-7 treated, untreated cells and normal hepatocytes using immunocytochemistry. Furthermore, the effects of oxidative stress on cell cycle arrest were also studied through flow cytometry. Results: Our study demonstrated the inhibition of cancer cell proliferation as a result of H2O2 induction by arresting the cell cycle at the G2 phase. Conclusion: The induction of oxidative stress could be a potential therapeutic approach for treating HCC in the future. GSTP-1 and PRX-2 can serve as substantial therapeutic targets for the treatment of HCC.

Fresh meat quality is adversely affected by protein oxidation. However, a fundamental understanding of the diverse factors that influence protein oxidation in postmortem muscle remains elusive. Peroxiredoxin-2 (Prdx2), an antioxidant protein, is more abundant in tough meat based on instrumental tenderness; however, the role of Prdx2 in postmortem skeletal muscle is unknown. Therefore, the objective was to develop a method to purify Prdx2 from the diaphragm, psoas major, and longissimus lumborum. Proteins soluble at low ionic strength were extracted, dialyzed, clarified, and loaded onto a Q-Sepharose anion exchange column equilibrated with TEM (pH 7.4). In all preparations, Prdx2 eluted between about 75 and 115 mM NaCl. Immunoreactive fractions were dialyzed against TEM (pH 8.0), clarified, and loaded onto a DEAE-650S anion exchange column. In all preparations, Prdx2 eluted between approximately 55 and 75 mM NaCl. Immunoreactive fractions were concentrated and loaded onto a Superose-12 size exclusion column. Prdx2 was detected between 14 and 16 mL, and these fractions were concentrated and reduced with 0.5% 2-mercaptoethanol. A final pass over the Superose-12 column was conducted, and Prdx2 was detected in 2 peaks from 11–12 mL and 15–16 mL. Fractions 15–16 were pooled and retained for further experiments. The elution profile of Prdx2 in all 3 muscles was similar. The iden- tification of the primary protein was confirmed with liquid chromatography with tandem mass spectrometry. The purity of Prdx2 off the final Superose-12 column was approximately 33%, 52%, and 47% pure in the diaphragm, psoas major, and longissimus lumborum, respectively. This is the first report of a method to partially purify Prdx2 from skeletal muscle.

Peroxiredoxins (Prxs) are ubiquitously expressed peroxidases that reduce hydrogen peroxide or alkyl peroxide production in cells. Prxs are released from cells in response to various stress conditions, and they function as damage-associated molecular pattern molecules. However, the secretory mechanism of Prxs and their roles have not been elucidated. Thus, we aimed to determine whether inflammasome activation is a secretory mechanism of Prxs and subsequently identify the effect of the secreted Prxs on activation of the classical complement pathway. Using J774A.1, a murine macrophage cell line, we demonstrated that NLRP3 inflammasome activation induces Prx1, Prx2, Prx5, and Prx6 secretion in a caspase-1 dependent manner. Using HEK293T cells with a transfection system, we revealed that the release of Prx1 and Prx2 relies on gasdermin-D (GSDMD)-mediated secretion. Next, we confirmed the binding of both Prx1 and Prx2 to C1q; however, only Prx2 could induce the C1q-mediated classical complement pathway activation. Collectively, our results suggest that inflammasome activation is a secretory mechanism of Prxs and that GSDMD is a mediator of their secretion. Moreover, secreted Prx1 and Prx2 bind with C1q, but only Prx2 mediates the classical complement pathway activation.

Intravenous infusion of high dose (>10 g) vitamin C (IVC) is a common alternative cancer therapy. IVC results in millimolar levels of circulating ascorbate, which is proposed to generate cytotoxic quantities of H2O2 in the presence of transition metal ions. In this study we report on the in vitro and in vivo effects of millimolar ascorbate on erythrocytes. Addition of ascorbate to whole blood increased erythrocyte intracellular ascorbate approximately 35-fold. Within 10 min of ascorbate addition, we detected increased oxidation of erythrocyte peroxiredoxin 2 (Prx2), a major thiol antioxidant protein and a sensitive marker of H2O2 production. Up to 50% of Prx2 was present in the oxidised form after 60 min. The presence of extracellular catalase, removal of plasma or the addition of a metal chelator did not prevent ascorbate-induced Prx2 oxidation, suggesting that the H2O2 responsible for Prx2 oxidation was generated within the erythrocyte. Ascorbate is known to increase the rate of haemoglobin autoxidation and H2O2 production. Through spectral monitoring of oxidised haemoglobin we estimated a generation rate of 15 μM H2O2/min inside erythrocytes. We also investigated changes in erythrocyte ascorbate concentration and Prx2 oxidation following IVC infusion in a cohort of patients with cancer. Plasma ascorbate levels ranged from 7.8 to 35 mM immediately post infusion, while erythrocyte ascorbate levels reached 1.5–3.4 mM 4 h after beginning infusion. Transient oxidation of erythrocyte Prx2 was observed. We conclude that erythrocytes accumulate ascorbate during IVC infusion, providing a significant reservoir of ascorbate, and this ascorbate increases H2O2 generation within the cells. The consequence of increased erythrocyte Prx2 oxidation warrants further investigation., Graphical abstract Image 1

Hydrogen peroxide undergoes an equilibrium reaction with bicarbonate/CO2 to produce peroxymonocarbonate (HCO4-). Peroxymonocarbonate is more reactive with thiols than H2O2 but it makes up only a small fraction of the H2O2 in physiological bicarbonate buffers so the increase in rate of oxidation of low molecular weight thiols is modest. However, for some thiol proteins such as protein tyrosine phosphatases, the rate enhancement is very much greater. We have investigated the effect of bicarbonate/CO2 on the oxidation of peroxiredoxins (Prdxs) 2 and 3. Using an assay in which reduced Prdx2 inhibits oxidation of horseradish peroxidase by H2O2, we saw no difference between phosphate and bicarbonate buffers (pH 7.4). However, hyperoxidation of both Prdxs in bicarbonate was considerably enhanced. Hyperoxidation involves the reaction of the sulfenic acid formed at the active site with a second H2O2, and prevents its condensation to a disulfide. Using LC/MS analysis, we determined that the presence of 25 mM bicarbonate/CO2 increased the ratio of hyperoxidation compared with condensation 6-fold for Prdx2 and 11-fold for Prdx3. These results imply that Prdx hyperoxidation will occur more readily under physiological conditions than appreciated from in vitro experiments, which seldom use bicarbonate buffers. They also raise the possibility that variations in bicarbonate concentration could provide a mechanism for regulating the cellular level of active Prdxs.

Peroxiredoxins (Prx) are enzymes that efficiently reduce hydroperoxides through active participation of cysteine residues (C(P), C(R)). The first step in catalysis, the reduction of peroxide substrate, is fast, 10(7) − 10(8) M(−1)s(−1) for human Prx2. In addition, the high intracellular concentration of Prx positions them not only as good antioxidants but also as central players in redox signaling pathways. These biological functions can be affected by post-translational modifications that could alter the peroxidase activity and/or interaction with other proteins. In particular, inactivation by hyperoxidation of C(P), which occurs when a second molecule of peroxide reacts with the C(P) in the sulfenic acid form, modulates their participation in redox signaling pathways. The higher sensitivity to hyperoxidation of some Prx has been related to the presence of structural motifs that disfavor disulfide formation at the active site, making the C(P) sulfenic acid more available for hyperoxidation or interaction with a redox protein target. We previously reported that treatment of human Prx2 with peroxynitrite results in tyrosine nitration, a post-translational modification on non-catalytic residues, yielding a more active peroxidase with higher resistance to hyperoxidation. In this work, studies on different mutants of hPrx2 confirm that the presence of the tyrosyl side-chain of Y193, belonging to the C-terminal YF motif of eukaryotic Prx, is necessary to observe the increase in Prx2 resistance to hyperoxidation. Moreover, our results underline the critical role of this structural motif on the rate of disulfide formation that determines the differential participation of Prx in redox signaling pathways.

Peroxiredoxin (Prx), also named thioredoxin peroxidase (TPx), is a selenium independent antioxidant enzyme that can protect organisms from oxidative damage caused by reactive oxygen species (ROS) and is important for immune responses. In this study, the molecular cloning and characterization of a Prx2 homologue (CiPrx2) were described from grass carp (Ctenoplicuyngodon idella). The full-length cDNA of CiPrx2 was 1163 by containing 5'-untranslated region (UTR) of 52 bp, a 3'-UTR of 517 by with the putative polyadenylation consensus signal (AATAAA), an open reading frame (ORF) of 594 by encoding polypeptides of 197 amino acids with a predicted molecular mass of 21.84 kDa and theoretical isoelectric point of 5.93. The analysis results of multiple sequence alignment and phylogenetic tree confirmed that CiPrx2 belong to the typical 2-Cys Prx subfamily. The CiPrx2 mRNA was ubiquitously expressed in all tested tissues. The temporal expression of CiPrx2 were differentially induced infected with grass carp reovirus (GCRV), polyinosinic:polycytidylic acid (poly I:C) and lipopolysaccharide (LPS) in liver and spleen. Subcellular localization of CiPrx2-GFP fusion proteins were only distributed in the cytoplasm. The purified recombinant CiPrx2 possessed an apparent antioxidant activity and could protect DNA against oxidative damage. Finally, CiPrx2 proteins could obviously inhibit H2O2 and heavy metal toxicity. However, further researches are needed to better understand the regulation of CiPrx2 under oxidative stresses.

Protein-tyrosine phosphatases (PTPs) counteract protein tyrosine phosphorylation and cooperate with receptor-tyrosine kinases in the regulation of cell signaling. PTPs need to undergo oxidative inhibition for activation of cellular cascades of protein-tyrosine kinase phosphorylation following growth factor stimulation. It has remained enigmatic how such oxidation can occur in the presence of potent cellular reducing systems. Here, using in vitro biochemical assays with purified, recombinant protein, along with experiments in the adenocarcinoma cell line A431, we discovered that bicarbonate, which reacts with H(2)O(2) to form the more reactive peroxymonocarbonate, potently facilitates H(2)O(2)-mediated PTP1B inactivation in the presence of thioredoxin reductase 1 (TrxR1), thioredoxin 1 (Trx1), and peroxiredoxin 2 (Prx2) together with NADPH. The cellular experiments revealed that intracellular bicarbonate proportionally dictates total protein phosphotyrosine levels obtained after stimulation with epidermal growth factor (EGF) and that bicarbonate levels directly correlate with the extent of PTP1B oxidation. In fact, EGF-induced cellular oxidation of PTP1B was completely dependent on the presence of bicarbonate. These results provide a plausible mechanism for PTP inactivation during cell signaling and explain long-standing observations that growth factor responses and protein phosphorylation cascades are intimately linked to the cellular acid–base balance.

Background This study aims to investigate the expression of thioredoxin 1, peroxiredoxin 1 and peroxiredoxin 2 in bulky cervical squamous carcinoma and its predictive role in cisplatin-based neoadjuvant chemotherapy. Methods Initially, the expression of thioredoxin 1, peroxiredoxin 1 and peroxiredoxin 2 protein was analyzed in 13 human cervical squamous cancer tissues and their paired adjacent non-cancerous tissues by western-blotting and immunohistochemistry. Then, correlation between the expression of thioredoxin 1, peroxiredoxin 1, peroxiredoxin 2 and responses to cisplatin-based neoadjuvant chemotherapy was analyzed in 35 paired tumor samples (pre- and post-chemotherapy) from bulky cervical squamous cancer patients by immunohistochemistry. Results A clinical response occurred in 48.6% (17/35) of patients, including 14.3% (5/35) with a complete response and 34.3% (12/35) with a partial response. The expression of thioredoxin 1, peroxiredoxin 1 and peroxiredoxin 2 was much higher in cervical squamous cancer tissues compared with paired adjacent non-cancerous tissues by western-blotting and immunohistochemistry. Additionally, the expression of thioredoxin 1, peroxiredoxin 1 and peroxiredoxin 2 was significantly up-regulated in post-chemotherapy tissues compared to pre-chemotherapy cervical cancer tissues. High levels of thioredoxin 1, peroxiredoxin 1 and peroxiredoxin 2 were associated with a poor chemotherapy response in cervical squamous cancer patients. Conclusions Thioredoxin 1, peroxiredoxin 1 and peroxiredoxin 2 are frequently over-expressed in cervical squamous cancer. High expression levels of these proteins were related to a poor response to cisplatin-based neoadjuvant chemotherapy. The present study is the first report that thioredoxin peroxidase system may serve as a prediction of the responses to neoadjuvant chemotherapy in cervical squamous cancer.

Oxidative damages have adverse effects on mammals. Growing studies have focused on exploring new antioxidants. Here, we report that Lactobacillus frumenti increases the total antioxidation capacity activities and decreases the total reactive oxygen species levels in porcine intestinal epithelial cells. Comparative proteomics revealed that expressions of peroxiredoxin 2, isocitrate dehydrogenase 1, NAD(P)H dehydrogenase quinone 1, antioxidant protein 1, and metallothionein-2A, which are associated with antioxidant defense system, were significantly increased with L. frumenti treatment. In germ-free mice, L. frumenti treatment also remarkably improves the intestinal antioxidant capacity. We further illustrated that nitric oxide production-mediated by nitric oxide synthase 1 activation is essential for L. frumenti-induced improvements in intestinal epithelial antioxidant capacity and barrier function. This study suggested that L. frumenti may be a potential probiotic used to prevent oxidative stress-induced aging and diseases in mammals.-Nie, Y., Hu, J., Hou, Q., Zheng, W., Zhang, X., Yang, T., Ma, L., Yan, X. Lactobacillus frumenti improves antioxidant capacity via nitric oxide synthase 1 in intestinal epithelial cells.

Fetal growth restriction (FGR) is caused by placental insufficiency and can lead to short and long‑term neurodevelopmental delays. Taurine, one of the most abundant amino acids in the brain, is critical for the normal growth and development of the nervous system; however, the mechanistic role of taurine in neural growth and development remains unknown. The present study investigated the role of taurine in FGR. Specifically, we explored the proteomic profiles of fetal rats at 6 h postpartum by two‑dimensional difference gel electrophoresis combined with matrix assisted laser desorption ionization‑time‑of‑flight (TOF)/TOF tandem mass spectrometry; the findings were verified via reverse transcription‑quantitative polymerase chain reaction. A total of 31 differentially expressed protein spots were selected. Among these, 31 were matched, including dihydropyrimidinase‑related protein 2 and , CRK and peroxiredoxin 2. Functional analysis using the Gene Ontology database and Ingenuity Pathway Analysis demonstrated that the differentially expressed proteins were mainly associated with neuronal differentiation, 'metabolic process', 'biological regulation' and developmental processes. The present study identified several proteins that were differentially expressed in rats with FGR in the presence or absence of taurine administration. The results of the present study suggest a potential role for taurine in the treatment and prevention of FGR.

Cancer stem cells (CSC) with low levels of reactive oxygen species (ROS) are resistant to conventional chemotherapy or radiation therapy. Peroxiredoxin 2 (Prx2) is a redox regulatory protein that plays a key role in maintaining ROS homeostasis in the tumor microenvironment. However, despite the role of Prx2 in ROS-mediated signal transduction, the association of Prx2 with stemness via ROS in CSC has not been thoroughly investigated. In this study, we investigated the link between Prx2 and CSC stemness through regulation of ROS levels in hepatocellular carcinoma (HCC) cells. ROS induced CSC stemness reduction and downregulated stem cell markers in Huh7 and SK-HEP1 cells. Prx2 knockdown decreased CSC sphere formation and expression of stem cell makers with increasing intracellular ROS levels. This effect was reversed by the ROS scavengers NAC and GSH in Prx2 knockdown cells. Conversely, we found that Prx2 overexpression promotes CSC stemness and the peroxidase activity of Prx2 is essential for CSC stemness using peroxidase inactive mutant, Prx2C51/172S. More importantly, the hyperoxidation-resistant mutant (Prx2ΔYF), which has a constant ROS scavenging activity even at high concentrations of ROS, increased the CSC stemness and expression of stem cell markers more than Prx2WT under oxidative stress. Taken together, our findings demonstrate that Prx2 links ROS homeostasis to CSC stemness; Prx2 is a mediator between ROS homeostasis and CSC stemness.

Zika virus (ZIKV) has been endemic in Southeast Asian countries for several years, but the presence of the virus has not been associated with significant outbreaks of infection unlike other countries around the world where the Asian lineage ZIKV was introduced recently. However, few studies have been undertaken using the endemic virus. The Thai isolate was shown to have a similar tissue tropism to an African isolate of ZIKV, albeit that the Thai isolate infected cells at a lower level as compared to the African isolate. To further understand the pathogenesis of the Thai isolate, a 2D-gel proteomic analysis was undertaken of ZIKV infected LLC-MK2 cells. Seven proteins (superoxide dismutase [Mn], peroxiredoxin 2, ATP synthase subunit alpha, annexin A5 and annexin A1, carnitine o-palmitoyltransferase 2 and cytoskeleton-associated protein 2) were identified as differentially regulated. Of four proteins selected for validation, three (superoxide dismutase [Mn], peroxiredoxin 2, ATP synthase subunit alpha, and annexin A1) were shown to be differentially regulated at both the transcriptional and translational levels. The proteins identified were primarily involved in energy production both directly, and indirectly through mediation of autophagy, as well as in the response to oxidative stress, possibly occurring as a consequence of increased energy production. This study provides further new information on the pathogenesis of ZIKV.

Red blood cells (RBCs) are exposed to exogenous reactive oxygen species in the circulatory system. To this end, the interactions between the different hemoglobin (Hb) subunits and peroxiredoxin 2, which is a ubiquitous member of the antioxidant enzymes that also controls the cytokine-induced peroxide levels, were assessed. We predicted by the increment of diversity with quadratic discriminant analysis (IDQD) that peroxiredoxin2 (Prx2) could interact with the hemoglobin alpha, beta and gamma subunits but not with the delta subunit. Coimmunoprecipitation (co-IP), electrospray ionization quadrupole time of flight (ESI-Q-TOF) mass spectrometry, Western blotting and X-ray absorption fine structure (XAFS) spectroscopy were performed to verify these predictions. The results showed that Prx2 was a member of the beta-globin immunoprecipitating complex that existed in hemoglobin A, hemolysate-hemoglobin A, hemoglobin A-hemoglobin A2, hemolysate-hemoglobin A-hemoglobin A2 and hemoglobin A2 but not in hemolysate-hemoglobin A2. Adding Prx2 to hemoglobin A altered the second shell of iron embedded in hemoglobin A. Therefore, Prx2 interacts with hemoglobin A (Alpha2Beta2) and hemoglobin F (Alpha2Gamma2) but not with hemoglobin A2 (Alpha2Delta2).

Background and Purpose:Early erythrolysis occurs within the hematoma following intracerebral hemorrhage (ICH), and the release of erythrocyte cytoplasmic proteins such as hemoglobin and Prx2 (peroxiredoxin 2) can cause brain injury. Complement activation can induce erythrolysis. This study determined the function of complement component 3 (C3) in erythrolysis in hematoma and brain injury after ICH in mice.Methods:This study has 3 parts. First, ICH was induced in adult male C3-sufficient and deficient mice and animals were euthanized on days 1, 3, 7, and 28 for immunohistochemistry after magnetic resonance imaging and behavioral testing. Second, C3-sufficient and deficient mice with ICH were euthanized on day 1 for Western blot analysis. Third, C3-sufficient mice received injections of PBS and Prx2. Mice underwent both magnetic resonance imaging and behavioral tests on day 1 and were then euthanized. Brains were harvested for immunohistochemistry and Fluoro-Jade C staining.Results:Erythrolysis occurred in the hematoma in C3-sufficient and deficient mice on day 3 following ICH. C3-deficient mice had less erythrolysis, brain swelling, and neuronal degeneration in the acute phase and less brain atrophy in the chronic phase. There were fewer neurological deficits on days 3, 7, and 28 in C3-deficient mice. C3-deficient mice also had less extracellular Prx2 release. Moreover, Prx2 induced brain edema and brain injury and recruited macrophage scavenger receptor-1- and CD4-positive cells following ICH in mice.Conclusions:C3-deficient mice had less severe erythrolysis and brain injury following ICH compared with C3-sufficient mice. Prx2 released after erythrolysis can cause brain damage and neuroinflammation in mice.

ObjectivesEfficiency of pork production is critical as demand for animal protein rises with increases in global populations and income. Feed efficiency and response to disease challenge are affected by stressors that increase reactive oxygen species in muscle. Reactive oxygen species are detrimental to growth, as they require the expenditure of energy for tissue repair that could be used for weight gain. Peroxiredoxin-2 (Prdx-2) is an antioxidant enzyme that converts hydrogen peroxide to water using reactive cysteines, thus mitigating oxidation. Prdx-2 can exist in multiple oxidation states (reduced, oxidized, and hyperoxidized) and quantenary structures (dimer and decamer). Prdx-2 concentration is greater in more color stable muscles and may be an indicator of tenderness. The objective of this study was to determine differences in Prdx-2 profile and protein oxidation between pigs that differed in feed efficiency and health status. Materials and MethodsPigs selected for differing feed efficiency based on residual feed intake (RFI) were used in a 2 × 2 factorial design for this study. At 50 ± 7 kg in body weight, high RFI (less efficient) and low RFI (more efficient) barrows were distributed between 2 rooms in the same building. One room was inoculated with a dual respiratory/enteric bacterial health challenge. At 21 d post-infection (projected peak of illness), pigs were necropsied (total = 24, n = 6 per group) and longissimus muscle samples were collected. Reduced and nonreduced sarcoplasmic protein samples (β-mercaptoethanol) were used for western blot analyses. Reduced samples were used to quantify total Prdx-2, while nonreduced samples were used to evaluate hyperoxidized peroxiredoxin, Prdx-2 decamer, and Prdx-2 profile in nonreducing gels (2 distinct bands analyzed). Carbonyl content and diagonal gel electrophoresis were performed on sarcoplasmic protein samples to determine protein oxidation. Statistical analysis was performed using SAS v. 9.4 (SAS Inst. Inc., Cary, NC) with fixed effects of RFI line, infection status, and RFI line × infection status interaction. Gel repetition and necropsy day were included as random effects. ResultsInfection status was significant for the Prdx-2 band comparison in nonreducing gels (P = 0.014) indicating potential differences in posttranslational modifications. Compared to low RFI pigs, high RFI pigs had greater total Prdx-2 (P = 0.035), greater Prdx-2 decamer (P = 0.0007), greater Prdx-2 in the second band of nonreducing gels (P = 0.0006), and less hyperoxidized peroxiredoxin relative to the total immunoreactive protein (P = 0.028). An interaction existed between RFI line and infection status for the Prdx-2 band comparison (P = 0.02), with high RFI challenged pigs having greater ratios, and high RFI control pigs having lesser ratios relative to low RFI pigs. Visual analysis of diagonal gels showed more proteins with intermolecular disulfide formation in low RFI muscle samples. No significant difference was seen for carbonyl content. ConclusionDifferences exist in Prdx-2 profile based on RFI line and infection status. High and low RFI pigs may respond differently to health challenges at the molecular level based on differences in antioxidant protein response. By defining Prdx-2 in livestock muscle, we may be better able to manage animals to meet demand for meat without increasing inputs.

The choroid plexus (CP) is the primary source of cerebrospinal fluid in the central nervous system. Recent evidence indicates that inflammatory pathways at the CP may be involved in hydrocephalus development. Peroxiredoxin 2 (Prx2) is a major component of red blood cells. Extracellular Prx2 is proinflammatory, and its release after red blood cell lysis may contribute to hydrocephalus after intraventricular hemorrhage. This study aimed to identify alterations in CP macrophages and dendritic cells following intracerebroventricular Prx2 injection and investigate the relationship between macrophages/dendritic cells and hydrocephalus. There were two parts to this study. In the first part, adult male Sprague–Dawley rats received an intracerebroventricular injection of Prx2 or saline. In the second part, Prx2 was co-injected with clodronate liposomes or control liposomes. All animals were euthanized at 24 h after magnetic resonance imaging. Immunohistochemistry was used to evaluate macrophages in CP, magnetic resonance imaging to quantify hydrocephalus, and histology to assess ventricular wall damage. The intracerebroventricular injection of Prx2 not only increased the OX-6 positive cells, but it also altered their location in the CP and immunophenotype. Co-injecting clodronate liposomes with Prx2 decreased the number of macrophages and simultaneously attenuated Prx2-induced hydrocephalus and ventricular wall damage. These results suggest that CP macrophages play an essential role in CP inflammation-induced hydrocephalus. These macrophages may be a potential therapeutic target in post-hemorrhagic hydrocephalus.

Peroxiredoxin 2 (PRDX2), an inhibitor of reactive oxygen species (ROS), is potentially involved in the progression of atherosclerosis (AS). The aim of this study was to explore the role and mechanism of PRDX2 in AS. The expression of PRDX2 was evaluated in 14 human carotid artery tissues with or without AS. The results showed that the positive reaction of PRDX2 was observed in the carotid artery vascular smooth muscle cells (CAVSMCs). To assess the mechanism by which PRDX2 may function in AS, the CAVSMCs were transfected with pEX4-PRDX2 and si-PRDX2. The catalase, hydrogen peroxide (H2O2) scavenger, was used to further confirm that PRDX2-induced inhibitory effects might be mediated through reducing ROS levels. Phenotype alteration and functional testing included transcription testing, immunostaining, and expression studies. The drug of MAPK signaling pathway inhibitors SB203580, SP600125, and PD98059 was used to evaluate the underlying mechanism. In this study, we found that the protein level of PRDX2 and the level of H2O2 were higher in the human AS carotid artery tissues than in the normal carotid artery tissues, accompanied with the activation of MAPK signaling pathway. The up-regulation of PRDX2 in the CAVSMCs significantly decreased the expression of ROS, collagen type I (COL I), collagen type III (COL III), vascular cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1) and inhibited the proliferation, migration, and transformation of the CAVSMCs. The up-regulation of PRDX2 reversed the effect of the CAVSMCs treated with tumor necrosis factor-α (TNF-α). In addition, PRDX2 down-regulation promoted the protein levels of p-p38, p-JNK, and p-ERK, which was confirmed in relevant MAPK inhibitor treatment experiments. Our results suggest a protective role of PRDX2, as a scavenger of ROS, in AS progression through inhibiting the VSMC phenotype alteration and function via MAPK signaling pathway.

Reactive oxygen species (ROS) released in cells are signaling molecules but can also modify signaling proteins. Red blood cells perform a major role in maintaining the balance of the redox in the blood. The main cytosolic protein of RBC is hemoglobin (Hb), which accounts for 95-97%. Most other proteins are involved in protecting the blood cell from oxidative stress. Hemoglobin is a major factor in initiating oxidative stress within the erythrocyte. RBCs can also be damaged by exogenous oxidants. Hb autoxidation leads to the generation of a superoxide radical, of which the catalyzed or spontaneous dismutation produces hydrogen peroxide. Both oxidants induce hemichrome formation, heme degradation, and release of free iron which is a catalyst for free radical reactions. To maintain the redox balance, appropriate antioxidants are present in the cytosol, such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and peroxiredoxin 2 (PRDX2), as well as low molecular weight antioxidants: glutathione, ascorbic acid, lipoic acid, α-tocopherol, β-carotene, and others. Redox imbalance leads to oxidative stress and may be associated with overproduction of ROS and/or insufficient capacity of the antioxidant system. Oxidative stress performs a key role in CKD as evidenced by the high level of markers associated with oxidative damage to proteins, lipids, and DNA in vivo. In addition to the overproduction of ROS, a reduced antioxidant capacity is observed, associated with a decrease in the activity of SOD, GPx, PRDX2, and low molecular weight antioxidants. In addition, hemodialysis is accompanied by oxidative stress in which low-biocompatibility dialysis membranes activate phagocytic cells, especially neutrophils and monocytes, leading to a respiratory burst. This review shows the production of ROS under normal conditions and CKD and its impact on disease progression. Oxidative damage to red blood cells (RBCs) in CKD and their contribution to cardiovascular disease are also discussed.

Peroxiredoxin-2 (Prx2) is the third most abundant cytoplasmic protein in red blood cells. Prx2 belongs to a well-known family of antioxidants, the peroxiredoxins (Prxs), that are widely expressed in mammalian cells. Prx2 is a typical, homodimeric, 2-Cys Prx that uses two cysteine residues to accomplish the task of detoxifying a vast range of organic peroxides, H2O2, and peroxynitrite. Although progress has been made on functional characterization of Prx2, much still remains to be investigated on Prx2 post-translational changes. Here, we first show that Prx2 is Tyrosine (Tyr) phosphorylated by Syk in red cells exposed to oxidation induced by diamide. We identified Tyr-193 in both recombinant Prx2 and native Prx2 from red cells as a specific target of Syk. Bioinformatic analysis suggests that phosphorylation of Tyr-193 allows Prx2 conformational change that is more favorable for its peroxidase activity. Indeed, Syk-induced Tyr phosphorylation of Prx2 enhances in vitro Prx2 activity, but also contributes to Prx2 translocation to the membrane of red cells exposed to diamide. The biologic importance of Tyr-193 phospho-Prx2 is further supported by data on red cells from a mouse model of humanized sickle cell disease (SCD). SCD is globally distributed, hereditary red cell disorder, characterized by severe red cell oxidation due to the pathologic sickle hemoglobin. SCD red cells show Tyr-phosphorylated Prx2 bound to the membrane and increased Prx2 activity when compared to healthy erythrocytes. Collectively, our data highlight the novel link between redox related signaling and Prx2 function in normal and diseased red cells.

The diagnostic peripheral biomarkers are still lacking for the bipolar II disorder (BD-II). We used isobaric tags for relative and absolute quantification technology to identify five upregulated candidate proteins [matrix metallopeptidase 9 (MMP9), phenylalanyl-tRNA synthetase subunit beta (FARSB), peroxiredoxin 2 (PRDX2), carbonic anhydrase 1 (CA-1), and proprotein convertase subtilisin/kexin type 9 (PCSK9)] for the diagnosis of BD-II. We analysed the differences in the plasma levels of these candidate proteins between BD-II patients and controls (BD-II, n = 185; Controls, n = 186) using ELISA. To establish a diagnostic model for the prediction of BD-II, the participants were divided randomly into a training group (BD-II, n = 149; Controls, n = 150) and a testing group (BD-II, n = 36; Controls, n = 36). Significant increases were found in all five protein levels between BD-II and controls in the training group. Logistic regression was analysed to form the composite probability score of the five proteins in the training group. Receiver-operating characteristic curve analysis revealed the diagnostic validity of the probability score [area under curve (AUC) = 0.89, P P

Pancreatic neuroendocrine neoplasms (pNENs) account for 2&ndash, 3% of pancreatic malignancies. Peroxiredoxins (Prdxs), which are major cellular antioxidants, are involved in multiple oncogenic signaling pathways. We investigated the role of peroxiredoxin-2 in QGP-1 human pNEN cell line and patient-derived pNEN tissue. To validate the cancer stem cell-like cell characteristics of QGP-1 cells in spheroid culture, in vitro analyses and xenografting were performed. Furthermore, immunohistochemical staining was conducted to verify the overexpression of Prdx2 in pNEN tissue. Prdx2 expression was high at the mRNA and protein levels in QGP-1 cells. Prdx2 was also overexpressed in patient-derived pNEN tissue. Silencing of Prdx2 using siRNA induced overexpression and phosphorylation of ERK and AKT in QGP-1. Cell proliferation was increased by treating QGP-1 cells with siPrdx2, and the IC50 of everolimus increased suggesting resistance to everolimus. Interestingly, QGP-1 spheroid cells, which exhibited cancer stem cell-like features, exhibited lower expression of Prdx2 and mTOR. The results suggest that Prdx2 expression level and its activity may be a potential predictive biomarker for therapeutic response or resistance to everolimus in pNEN.

Mitochondria are strategically trafficked throughout the cell by the action of microtubule motors, the actin cytoskeleton and adapter proteins. The intracellular positioning of mitochondria supports subcellular levels of ATP, Ca2+ and reactive oxygen species (ROS, i.e. hydrogen peroxide, H2O2). Previous work from our group showed that deletion of the mitochondrial adapter protein Miro1 leads to perinuclear clustering of mitochondria, leaving the cell periphery devoid of mitochondria which compromises peripheral energy status. Herein, we report that deletion of Miro1 significantly restricts subcellular H2O2 levels to the perinuclear space which directly affects intracellular responses to elevated mitochondrial ROS. Using the genetically encoded H2O2-responsive fluorescent biosensor HyPer7, we show that the highest levels of subcellular H2O2 map to sites of increased mitochondrial density. Deletion of Miro1 or disruption of microtubule dynamics with Taxol significantly reduces peripheral H2O2 levels. Following inhibition of mitochondrial complex 1 with rotenone we observe elevated spikes of H2O2 in the cell periphery and complementary oxidation of mitochondrial peroxiredoxin 3 (PRX3) and cytosolic peroxiredoxin 2 (PRX2). Conversely, in cells lacking Miro1, rotenone did not increase peripheral H2O2 or PRX2 oxidation but rather lead to increased nuclear H2O2 and an elevated DNA-damage response. Lastly, local levels of HyPer7 oxidation correlate with the size and abundance of focal adhesions (FAs) in MEFs and cells lacking Miro1 have significantly smaller focal adhesions and reduced phosphorylation levels of vinculin and p130Cas compared to Miro1+/+ MEFs. Together, we present evidence that the intracellular distribution of mitochondria influences subcellular H2O2 levels and local cellular responses dependent on mitochondrial ROS., Graphical abstract Image 1

Peroxiredoxin-2 (PRX-2) is known to be released from erythrocytes and induce brain damage after intracerebral hemorrhage (ICH); lipocalin-2 (LCN-2) is involved in neuroinflammation following ICH. This study examined the role of LCN-2 in PRX-2 induced brain injury and involved three parts. In the first part, adult male C57BL/6 wild-type (WT), LCN-2 heterozygous (LCN-2 HET), and LCN-2 knockout (LCN-2 KO) mice received either an intracaudate injection of recombinant PRX-2 or saline. In the second part, adult male C57BL/6 WT and male LCN-2 KO mice received recombinant PRX-2 with either recombinant mouse LCN-2 protein or control. In the third part, adult female C57BL/6 WT, LCN-2 HET, and LCN-2 KO mice received recombinant PRX-2. Behavioral tests, and T2- and T2*- weighted magnetic resonance imaging was obtained for all mice. Mice were then euthanized, and their brains used for Western blotting, histology and immunohistochemistry. Intracerebral PRX-2 injections resulted in increased expression of LCN-2 protein. PRX-2-induced brain swelling, neutrophil infiltration, microglia/macrophage activation, neuronal cell death, and neurological deficits were reduced in male LCN-2 HET and LCN-2 KO mice (P 0.01) compared to WT and were exacerbated by exogenous LCN-2 co-injection. Additionally, intracerebral PRX-2 injections caused brain injury and neurological deficits in female WT mice; effects reduced in female LCN-2 KO mice. In conclusion, intracerebral injection of PRX-2 upregulates LCN-2, and LCN-2 is crucial in the effects of PRX-2 on neutrophil infiltration and microglia/macrophage activation, and ultimately brain damage.

Red blood cell death or erythrocyte apoptosis (eryptosis) is generally mediated by oxidative stress, energy depletion, heavy metals exposure, or xenobiotics. As erythrocytes are a major target for oxidative stress due to their primary function as O2-carrying cells, they possess an efficient antioxidant defense system consisting of glutathione peroxidase (GPx), superoxide dismutase (SOD), catalase (CAT), and peroxiredoxin 2 (Prx2). The oxidative stress-mediated activation of the Ca2+-permeable cation channel results in Ca2+ entry into the cells and subsequent cell death. Herein, we describe for the first time that selenium compounds having intramolecular diselenide or selenenyl sulfide moieties can prevent the oxidative stress-induced eryptosis by exhibiting an unusual Prx2-like redox activity under conditions when the cellular Prx2 and CAT enzymes are inhibited.