PRDX1 and MTH1 cooperate to prevent ROS-mediated inhibition of telomerase.

Telomerase counteracts telomere shortening and cellular senescence in germ, stem, and cancer cells by adding repetitive DNA sequences to the ends of chromosomes. Telomeres are susceptible to damage by reactive oxygen species (ROS), but the consequences of oxidation of telomeres on telomere length and the mechanisms that protect from ROS-mediated telomere damage are not well understood. In particular, 8-oxoguanine nucleotides at 3' ends of telomeric substrates inhibit telomerase in vitro, whereas, at internal positions, they suppress G-quadruplex formation and were therefore proposed to promote telomerase activity. Here, we disrupt the peroxiredoxin 1 (PRDX1) and 7,8-dihydro-8-oxoguanine triphosphatase (MTH1) genes in cancer cells and demonstrate that PRDX1 and MTH1 cooperate to prevent accumulation of oxidized guanine in the genome. Concomitant disruption of PRDX1 and MTH1 leads to ROS concentration-dependent continuous shortening of telomeres, which is due to efficient inhibition of telomere extension by telomerase. Our results identify antioxidant systems that are required to protect telomeres from oxidation and are necessary to allow telomere maintenance by telomerase conferring immortality to cancer cells.

Telomerase can't handle the stress.

Telomerase counteracts the telomere shortening that occurs with each round of cell division. In normal human cells, telomerase is repressed, leading to telomere shortening that triggers replicative senescence. However, in most tumors, telomerase is up-regulated and is essential for telomere maintenance and tumor cell growth. Although long considered a viable target for tumor therapy, successful inhibition of telomerase in cancer therapy remains to be described. In this issue of Genes & Development, Ahmed and Lingner (pp. 658-669) uncover a vulnerability in telomerase upon exposure of cancer cells to oxidative stress. It has long been known that telomeres are sensitive to damage by reactive oxygen species (ROS), but the impact of oxidation on telomerase function in living cells was not known. Using gene knockouts in colon cancer cells, the investigators demonstrate that the antioxidant enzyme peroxiredoxin 1 (PRDX1) and the nudix phosphohydrolase superfamily enzyme (MTH1) cooperate to retain, upon oxidative stress, telomeres in a telomerase-extendible state. Considering that cancer cells are more vulnerable to ROS than noncancer cells, this work may open new avenues targeting telomeres and telomerase in tumor cells.

Chronic exposure to low-dose deltamethrin can lead to colon tissue injury through PRDX1 inactivation-induced mitochondrial oxidative stress injury and gut microbial dysbiosis.

OBJECTIVE: To date, it is unclear whether deltamethrin (DLM) intake causes damage to colon tissue. Hence, in this study, we aimed to clarify the effect of long-term exposure to low-dose DLM on colon tissues, and its potential mechanisms. METHODS: Mice were treated with DLM (0.2 mg/kg/day) or DLM combined with N-acetyl-l-cysteine (NAC) (50 mg/kg/day) for 8 weeks. Human colon cancer cells (HCT-116) were treated with DLM (0, 25, 50, or 100 µM), NAC (2 mM), or overexpression plasmids targeting peroxiredoxin 1 (PRDX1) for 48 h. DLM was detected using a DLM rapid detection card. Colon injury was evaluated using haematoxylin and eosin staining and transmission electron microscopy. Apoptosis was determined using immunofluorescence staining (IF), western blotting (WB) and flow cytometry (FC) assays. MitoTracker, JC-1, and glutathione (GSH) detection were used to detect mitochondrial oxidative stress. Intestinal flora were identified by 16 S rDNA sequencing. RESULTS: DLM accumulation was detected in the colon tissue and faeces of mice following long-term intragastric administration. Interestingly, our results showed that, even at a low dose, long-term intake of DLM resulted in severe weight loss and decreased the disease activity index scores and colon length. The results of IF, WB, and FC showed that DLM induced apoptosis in the colon tissue and cells. MitoTracker, JC-1, and GSH assays showed that DLM increased mitochondrial stress in colonic epithelial cells. Mechanistic studies have shown that increased mitochondrial stress and apoptosis are mediated by PRDX1 inhibition. Further experiments showed that PRDX1 overexpression significantly reduced DLM-induced oxidative stress injury and apoptosis. In addition, we observed that chronic exposure to DLM altered the composition of the intestinal flora in mice, including an increase in Odoribacter and Bacteroides and a decrease in Lactobacillus. The gut microbial richness decreased after DLM exposure in mice. Supplementation with NAC both in vivo and in vitro alleviated DLM-induced oxidative stress injury, colonic epithelial cell apoptosis, and gut microbial dysbiosis. CONCLUSION: Chronic exposure to DLM, even at small doses, can cause damage to the colon tissue, which cannot be ignored. The production and use of pesticides such as DLM should be strictly regulated during agricultural production.

Effect of peroxiredoxin 1 on the regulation of trophoblast function by affecting autophagy and oxidative stress in preeclampsia.

PURPOSE: PE is a pregnancy-specific syndrome and one of the main causes of maternal, fetal, and neonatal mortality. PRDX1 is an antioxidant that regulates cell proliferation, differentiation, and apoptosis. The aim of this study is to investigate the effect of PRDX1 on the regulation of trophoblast function by affecting autophagy and oxidative stress in preeclampsia. METHODS: Western blotting, RT-qPCR, and immunofluorescence were used to examine the expression of PRDX1 in placentas. PRDX1-siRNA was transfected to knockdown PRDX1 in HTR-8/SVneo cells. The biological function of HTR-8/SVneo cells was detected by wound healing, invasion, tube formation, CCK-8, EdU, flow cytometry, and TUNEL assays. Western blotting was used to detect the protein expression of cleaved-Caspase3, Bax, LC3II, Beclin1, PTEN, and p-AKT. DCFH-DA staining was used to detect ROS levels by flow cytometry. RESULTS: PRDX1 was significantly decreased in placental trophoblasts in PE patients. Following the exposure of HTR-8/SVneo cells to H2O2, PRDX1 expression was significantly decreased, LC3II and Beclin1 expression was notably increased, and ROS level was also markedly increased. PRDX1 knockdown impaired migration, invasion, and tube-formation abilities and promoted apoptosis, which was accompanied by an increased expression of cleaved-Caspase3 and Bax. PRDX1 knockdown induced a significant decrease in LC3II and Beclin1 expression, along with an elevated p-AKT expression and a decreased PTEN expression. PRDX1 knockdown increased intracellular ROS levels, and NAC attenuated PRDX1 knockdown-induced apoptosis. CONCLUSION: PRDX1 regulated trophoblast function through the PTEN/AKT signaling pathway to affect cell autophagy and ROS level, which provided a potential target for the treatment of PE.

Peroxiredoxin 1 Interacts with TBK1/IKKε and Negatively Regulates Pseudorabies Virus Propagation by Promoting Innate Immunity.

Peroxiredoxin 1 (PRDX1) is a cellular antioxidant enzyme that is crucial for diverse fundamental biological processes, such as autophagy, inflammation, and carcinogenesis. However, molecular mechanisms underpinning its diverse roles are not well understood. Here, we report that PRDX1 positively regulates interferon (IFN) induction and that pseudorabies virus (PRV) targets PRDX1 to evade IFN induction. PRV UL13 encodes a serine/threonine kinase important for PRV infection, although its biological function remains obscure. We identified PRDX1 as a UL13-interacting protein. Virological and biochemical assays demonstrate that PRDX1 promotes IFN induction by interacting with TANK-binding kinase 1 (TBK1) and IκB kinase ε (IKKε). Conversely, UL13 accelerates PRDX1 degradation via the ubiquitin-proteosome pathway in a kinase-dependent manner. In doing so, PRV inhibits IFN induction during productive infection, which requires PRDX1 expression. This study uncovers an essential role of PRDX1 in the innate immune response and reveals a new viral immune evasion strategy to counteract cellular defenses. IMPORTANCE PRV interacts with numerous cellular proteins during productive infection. Here, we demonstrated the interaction of viral protein UL13 with the antioxidant enzyme PRDX1, which functions in multiple signal transduction pathways. We found that PRDX1 participates in the type I IFN pathway by interacting with TBK1 and IKKε, thereby negatively regulating PRV propagation. However, UL13 ubiquitinates PRDX1, which routes PRDX1 into proteasomes for degradation and effectively reduces its expression. These results illuminate the fundamental role that PRDX1 plays in the IFN pathway, and they identify a potential target for the control of PRV infection.

Peroxiredoxin 1 Controls Ovulation and Ovulated Cumulus-Oocyte Complex Activity through TLR4-Derived ERK1/2 Signaling in Mice.

Peroxiredoxins (PRDXs) are expressed in the ovary and during ovulation. PRDX1 activity related to the immuno-like response during ovulation is unknown. We investigated the roles of Prdx1 on TLR4 and ERK1/2 signaling from the ovulated cumulus-oocyte complex (COC) using Prdx1-knockout (K/O) and wild-type (WT) mice. Ovulated COCs were collected 12 and 16 h after pregnant mare serum gonadotropin/hCG injection. PRDX1 protein expression and COC secretion factors (Il-6, Tnfaip6, and Ptgs2) increased 16 h after ovulated COCs of the WT mice were obtained. We treated the ovulated COCs in mice with LPS (0.5 µg/mL) or hyaluronidase (Hya) (10 units/mL) to induce TLR4 activity. Intracellular reactive oxygen species (ROS), cumulus cell apoptosis, PRDX1, TLR4/P38/ERK1/2 protein expression, and COC secretion factors' mRNA levels increased in LPS- and Hya-treated COCs. The ERK inhibitor (U0126) and Prdx1 siRNA affected TLR4/ERK1/2 expression. The number and cumulus expansion of ovulated COCs by ROS were impaired in Prdx1 K/O mice but not in WT ones. Prdx1 gene deletion induced TLR4/P38/ERK1/2 expression and cumulus expansion genes. These results show the controlling roles of PRDX1 for TLR4/P38/ERK1/2 signaling activity in ovulated mice and the interlink of COCs with ovulation.

Decoy receptor 2 mediation of the senescent phenotype of tubular cells by interacting with peroxiredoxin 1 presents a novel mechanism of renal fibrosis in diabetic nephropathy.

Premature senescence of renal tubular epithelial cell (RTEC), which is involved in kidney fibrosis, is a key event in the progression of diabetic nephropathy. However, the underlying mechanism remains unclear. Here we investigated the role and mechanism of decoy receptor 2 (DcR2) in kidney fibrosis and the senescent phenotype of RTEC. DcR2 was specifically expressed in senescent RTEC and associated with kidney fibrosis in patients with diabetic nephropathy and mice with streptozotocin-induced with diabetic nephropathy. Knockdown of DcR2 decreased the expression of α-smooth muscle actin, collagen I, fibronectin and serum creatinine levels in streptozotocin-induced mice. DcR2 knockdown also inhibited the expression of senescent markers p16, p21, senescence-associated beta-galactosidase and senescence-associated heterochromatic foci and promoted the secretion of a senescence-associated secretory phenotype including IL-6, TGF-ß1, and matrix metalloproteinase 2 in vitro and in vivo. However, DcR2 overexpression showed the opposite effects. Quantitative proteomics and validation studies revealed that DcR2 interacted with peroxiredoxin 1 (PRDX1), which regulated the cell cycle and senescence. Knockdown of PRDX1 upregulated p16 and cyclin D1 while downregulating cyclin-dependent kinase 6 expression in vitro, resulting in RTEC senescence. Furthermore, PRDX1 knockdown promoted DcR2-induced p16, cyclin D1, IL-6, and TGF-ß1 expression, whereas PRDX1 overexpression led to the opposite results. Subsequently, DcR2 regulated PRDX1 phosphorylation, which could be inhibited by the specific tyrosine kinase inhibitor genistein. Thus, DcR2 mediated the senescent phenotype of RTEC and kidney fibrosis by interacting with PRDX1. Hence, DcR2 may act as a potential therapeutic target for the amelioration of diabetic nephropathy progression.

Peroxiredoxin 1, a Novel HBx-Interacting Protein, Interacts with Exosome Component 5 and Negatively Regulates Hepatitis B Virus (HBV) Propagation through Degradation of HBV RNA.

Hepatitis B virus (HBV) infection is a major risk factor for the development of chronic liver diseases, including cirrhosis and hepatocellular carcinoma (HCC). A growing body of evidence suggests that HBV X protein (HBx) plays a crucial role in viral replication and HCC development. Here, we identified peroxiredoxin 1 (Prdx1), a cellular hydrogen peroxide scavenger, as a novel HBx-interacting protein. Coimmunoprecipitation analysis coupled with site-directed mutagenesis revealed that the region from amino acids 17 to 20 of the HBx, particularly HBx Cys17, is responsible for the interaction with Prdx1. Knockdown of Prdx1 by siRNA significantly increased the levels of intracellular HBV RNA, HBV antigens, and extracellular HBV DNA, whereas knockdown of Prdx1 did not increase the activities of HBV core, enhancer I (Enh1)/X, preS1, and preS2/S promoters. Kinetic analysis of HBV RNA showed that knockdown of Prdx1 inhibited HBV RNA decay, suggesting that Prdx1 reduces HBV RNA levels posttranscriptionally. The RNA coimmunoprecipitation assay revealed that Prdx1 interacted with HBV RNA. The exosome component 5 (Exosc5), a member of the RNA exosome complexes, was coimmunoprecipitated with Prdx1, suggesting its role in regulation of HBV RNA stability. Taken together, these results suggest that Prdx1 and Exosc5 play crucial roles in host defense mechanisms against HBV infection.IMPORTANCE Hepatitis B virus (HBV) infection is a major global health problem. HBx plays important roles in HBV replication and viral carcinogenesis through its interaction with host factors. In this study, we identified Prdx1 as a novel HBx-binding protein. We provide evidence suggesting that Prdx1 promotes HBV RNA decay through interaction with HBV RNA and Exosc5, leading to downregulation of HBV RNA. These results suggest that Prdx1 negatively regulates HBV propagation. Our findings may shed new light on the roles of Prdx1 and Exosc5 in host defense mechanisms in HBV infection.

Prdx1 promotes the loss of primary cilia in esophageal squamous cell carcinoma.

BACKGROUND: Loss of primary cilia is frequently observed in tumor cells, suggesting that the absence of this organelle may promote tumorigenesis through aberrant signal transduction, the inability to exit the cell cycle, and promotion of tumor cell invasion. Primary cilia loss also occurs in esophageal squamous cell carcinoma (ESCC) cells, but the molecular mechanisms that explain how ESCC cells lose primary cilia remain poorly understood. METHODS: Inhibiting the expression of Prdx1 in the ESCC cells to detect the up-regulated genes related to cilium regeneration and down-regulated genes related to cilium disassembly by Gene chip. And, mice and cell experiments were carried to confirm the role of the HEF1-Aurora A-HDAC6 signaling axis in ESCC. RESULTS: In this study, we found that silencing Peroxiredoxin 1 (Prdx1) restores primary cilia formation, and over-expressing Prdx1 induces primary cilia loss in ESCC cells. We also showed that the expression of Prdx1 regulates the action of the HEF1-Aurora A-HDAC6 signaling axis to promote the disassembly of primary cilia, and suppression of Prdx1 results in decreased tumor formation and tumor mass volume in vivo. CONCLUSIONS: These results suggest that Prdx1 is a novel regulator of primary cilia formation in ESCC cells.

PRDX1 enhances cerebral ischemia-reperfusion injury through activation of TLR4-regulated inflammation and apoptosis.

Stroke is still a leading cause of death across the world. Despite various signals or molecules that contribute to the pathophysiological process have been investigated, the exact molecular mechanisms revealing stroke damage still remain to be explored. Peroxiredoxin 1 (PRDX1) has been identified as a stress-induced macrophage redox protein with multiple functions. Although PRDX1 is a critical factor related to the regulation of immunity, inflammation, apoptosis and oxidative stress, its effects on cerebral ischemia-reperfusion (I-R) injury were presently unclear. In the study, by using a mouse model of I-R injury, we found that PRDX1 expression was up-regulated during I-R injury in a time-dependent manner. Additionally, PRDX1-knockout mice showed reduced infarction area and alleviated neuropathological scores with decreased brain water contents. Furthermore, cell death and inflammatory response in mice with cerebral I-R injury were markedly attenuated by PRDX1 knockout, which were associated with the blockage of Caspase-3 and nuclear factor-κB (NF-κB) signaling pathways. Mechanistically, PRDX1-regulated cerebral I-R injury was through the promotion of toll-like receptor-4 (TLR4), as proved by the evidence that TLR4 suppression abrogated the exacerbated effect of TLR4 on inflammatory response and apoptosis in oxygen and glucose deprivation (OGD)-treated primary microglial cells. These data demonstrated that PRDX1 contributed to cerebral stroke by interacting with TLR4, providing an effective therapeutic approach for cerebral I-R injury.

Innate differences and colostrum-induced alterations of jejunal mucosal proteins in piglets with intra-uterine growth restriction.

Mammalian neonates undergo rapid transitions from a sterile uterine environment with a continuous intravenous supply of nutrients to a microbe-rich environment with intermittent ingesting of colostrum/milk via the gut. Currently, little is known about the colostrum-induced alterations of intestinal mucosal proteins in piglets with intra-uterine growth restriction (IUGR). In this study, we sought to investigate the innate differences and effects of colostrum on alterations in small-intestinal proteomes of IUGR piglets. Two IUGR (approximately 0·9 kg) and two normal-birth weight (NBW; approximately 1·3 kg) piglets were obtained from each of six sows at birth. One half (n 12; 6 IUGR v. 6 NBW) of the selected newborn piglets were killed to obtain jejunum samples, and the other half (n 12; 6 IUGR v. 6 NBW) of the newborn piglets were allowed to suckle colostrum from their own mothers for 24 h before jejunum sample collection. On the basis of proteomic analysis, we identified thirty-one differentially expressed proteins in the jejunal mucosa between IUGR and normal neonates before or after colostrum consumption. The intestinal proteins altered by colostrum feeding play important roles in the following: (1) increasing intestinal integrity, transport of nutrients, energy metabolism, protein synthesis, immune response and, therefore, cell proliferation; and (2) decreasing oxidative stress, and therefore cell apoptosis, in IUGR neonates. However, colostrum only partially ameliorated the inferior status of the jejunal mucosa in IUGR neonates. These findings provide the first evidence in intestinal protein alterations of IUGR neonates in response to colostrum ingestion, and thus render new insights into the mechanisms responsible for impaired growth in IUGR neonates and into new nutritional intervention strategies.

Peroxiredoxin-1 of macrophage is critical for mycobacterial infection and is controlled by early secretory antigenic target protein through the activation of p38 MAPK.

Early secretory antigenic target protein (ESAT-6) is an important virulent factor which plays a crucial role in Mycobacterium tuberculosis (MTB) pathogenesis. Here, we demonstrate the role of ESAT-6 in phagocytosis and intracellular survival of mycobacteria through a mechanism mediated by regulation of a host protein; Peroxiredoxin-1 (Prdx-1). Prdx-1 is an anti-apoptotic and stress response protein which protects cells from damage by ROS and H2O2. The J774 A.1 cells infected with MTB or over-expressing ESAT-6 through eukaryotic promoter vector showed elevated expression of Prdx-1. Further investigation revealed that the up-regulation of Prdx-1 is mediated through the activation of one of the MAP kinases, p38. The NRF-2, a transcriptional activator of Prdx-1 is translocated to the nucleus upon phosphorylation by p38 and subsequently, regulates expression of Prdx-1. Inhibition of the p38 MAPK by a specific inhibitor, SB203580, abrogates the ESAT-6 mediated induction of Prdx-1 expression as well as the phosphorylation of NRF-2 in a time-dependent manner. The inhibition of Prdx-1 expression by specific siRNA in J774 A.1 cells resulted in the reduced bacterial uptake and intracellular survival of the mycobacteria. This is the first report proclaiming that the ESAT-6 regulates Prdx-1 which is involved in the increase of mycobacterial uptake and survival. The intermediate mechanisms involve the increased Prdx-1 production in macrophages through the activation of p38 and NRF-2 dependent signaling.

Characterization of the Translationally Controlled Tumor Protein (TCTP) Interactome Reveals Novel Binding Partners in Human Cancer Cells.

Translationally controlled tumor protein (TCTP) is a highly conserved housekeeping protein present in eukaryotic organisms. It is involved in regulating many fundamental processes and plays a critical role in tumor reversion and tumorigenesis. Increasing evidence suggests that TCTP plays a role in the regulation of cell fate determination and is a promising therapeutic target for cancer. To decipher the exact mechanisms by which TCTP functions and how all these functions are integrated, we analyzed the interactome of TCTP in HeLa cells by coimmunoprecipitation (IP) and mass spectrometry (MS). A total of 98 proteins were identified. We confirmed the in vitro and in vivo association of TCTP with six of the identified binding proteins using reciprocal IP and bimolecular fluorescence complementation (BiFC) analysis, respectively. Moreover, TCTP interacted with Y-box-binding protein 1 (YBX1), and their interaction was localized to the N-terminal region of TCTP and the 1-129 amino acid (aa) residues of YBX1. The YBX1 protein plays an important role in cell proliferation, RNA splicing, DNA repair, drug resistance, and stress response to extracellular signals. These data suggest that the interaction of TCTP with YBX1 might cooperate or coordinate their functions in the control of diverse regulatory pathways in cancer cells. Taken together, our results not only reveal a large number of TCTP-associated proteins that possess pleiotropic functions, but also provide novel insights into the molecular mechanisms of TCTP in tumorigenesis.

Redox proteomics reveal stress responsive proteins linking peroxiredoxin-1 status in glioma to chemosensitivity and oxidative stress.

The combined deletion of chromosomal arms 1p and 19q has been described as a prognostic marker for oligodendroglial tumors. These tumors show a better response to chemotherapy and radiotherapy. Recently, we found a lower abundance of peroxiredoxin 1 (PRDX1) in oligodendroglial tumors with 1p/19q deletion, suggesting a potential role of this enzyme in the clearance of therapy induced reactive oxygen species (ROS). Here, we confirmed the importance of PRDX1 in tumor cell survival by PRDX1 knockdown and overexpression in A-172 cells treated with the alkylating agent bis-chloroethyl nitrosourea (BCNU). Overexpression of PRDX1 resulted in a higher resistance of cells to BCNU treatment. In addition, BCNU challenged cells showed higher levels of ROS in PRDX1 knockdown cells. We applied a modified version of the redox two dimensional difference gel electrophoresis approach to analyze ROS mediated effects on protein thiols after BCNU treatment by labeling protein thiols with fluorescent dyes. Altogether eleven proteins were identified showing PRDX1 dependent altered labeling, many of them have been previously linked to stress response processes. Furthermore, 30 additional potentially redox active proteins were identified. The majority of them is involved in therapy associated processes like cellular stress response, DNA damage and regulation of cell death and therewith suggests that tumor cells maintain a network of redox sensitive proteins to escape chemotherapy. This article is part of a Special Issue entitled: Medical Proteomics.

PRDX1 is involved in palmitate induced insulin resistance via regulating the activity of p38MAPK in HepG2 cells.

Studies have identified that type 2 diabetes mellitus (T2DM) patients displayed higher levels of plasma peroxiredoxin1(PRDX1) than non-diabetics. However, the impact of PRDX1 on insulin resistance and the underlying mechanism remains totally unknown. Here, we investigated the influence of PRDX1 on hepatic insulin resistance. We showed that the protein and mRNA levels of PRDX1 were significantly elevated under insulin-resistant conditions. In addition, we showed that interference of PRDX1 ameliorated palmitate-induced insulin resistance in HepG2 cells, which was indicated by elevated phosphorylation of protein kinase B (AKT) and of glycogen synthase kinase-3 (GSK3ß). Furthermore, the expression of phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase), two key gluconeogenic enzymes, were down-regulated following PRDX1 depletion. Accordingly, glucose uptake was suppressed in PRDX1-interferred HepG2 cells. In addition, Over-expression of PRDX1 enhanced PA-induced insulin resistance in HepG2 cells. Moreover, we found that knocking down PRDX1 improves insulin sensitivity and decreased the activation of p38 mitogen-activated protein kinase (p38MAPK). Our results demonstrate that PRDX1 can induce hepatic insulin resistance by activating p38MAPK signaling and identifies potential targets for new treatments.

Identification of RAB2A and PRDX1 as the potential biomarkers for oral squamous cell carcinoma using mass spectrometry-based comparative proteomic approach.

Despite the recent advances in diagnostic and therapeutic strategies, oral squamous cell carcinoma (OSCC) remains a major health burden. Protein biomarker discovery for early detection will help to improve patient survival rate in OSCC. Mass spectrometry-based proteomics has emerged as an excellent approach for detection of protein biomarkers in various types of cancers. In the current study, we have used 4-Plex isobaric tags for relative and absolute quantitation (iTRAQ)-based shotgun quantitative proteomic approach to identify proteins that are differentially expressed in cancerous tissues compared to normal tissues. The high-resolution mass spectrometric analysis resulted in identifying 2,074 proteins, among which 288 proteins were differentially expressed. Further, it was noticed that 162 proteins were upregulated, while 125 proteins were downregulated in OSCC-derived cancer tissue samples as compared to the adjacent normal tissues. We identified some of the known molecules which were reported earlier in OSCC such as MMP-9 (8.4-fold), ZNF142 (5.6-fold), and S100A7 (3.5-fold). Apart from this, we have also identified some novel signature proteins which have not been reported earlier in OSCC including ras-related protein Rab-2A isoform, RAB2A (4.6-fold), and peroxiredoxin-1, PRDX1 (2.2-fold). The immunohistochemistry-based validation using tissue microarray slides in OSCC revealed overexpression of the RAB2A and PRDX1 gene in 80 and 68 % of the tested clinical cases, respectively. This study will not only serve as a resource of candidate biomarkers but will contribute towards the existing knowledge on the role of the candidate molecules towards disease progression and therapeutic potential.

Expression of Peroxiredoxin 1 After Traumatic Spinal Cord Injury in Rats.

Reactive astrogliosis and microgliosis after spinal cord injury (SCI) contribute to glial scar formation that impedes axonal regeneration. The mechanisms underlying reactive astrocyte and microglia proliferation upon injury remain partially understood. Peroxiredoxin 1 (PRDX1) is an antioxidant participating in cell proliferation, differentiation, and apoptosis. However, PRDX1 functions in SCI-induced astrocyte and microglia proliferation are unknown. In this study, we established an acute spinal cord contusion injury model in adult rats to investigate the potential role of PRDX1 during the pathological process of SCI. We found the palpable expression increase of PRDX1 after SCI by western blot and immunohistochemistry staining. Double immunofluorescence staining showed that PRDX1 expression mainly increased in astrocytes and microglia. In addition, PRDX1/proliferating cell nuclear antigen (PCNA) colocalized in astrocytes and microglia. Furthermore, PCNA expression also elevated after SCI, as well as was positively correlated with PRDX1 expression. In vitro, PRDX1 expression in primary rat spinal cord astrocytes and microglia changed in a concentration- and time-dependent manner according to LPS treatment. In addition, PRDX1 knockdown in astrocytes and microglia resulted in the decrease of PCNA expression after LPS stimulation, showing that PRDX1 promoted astrocyte and microglia proliferation after inflammation. Our results suggested that PRDX1 might play a crucial role in astrocyte and microglia proliferation after SCI.

[Prdx1 regulates macrophage polarization by maintaining mitochondrial homeostasis].

In order to investigate the role of Prdx1 in macrophage polarization, mouse leukemia cells of monocyte macrophage (RAW264.7) were treated with lipopolysaccharides (LPS)+ interferon gamma (IFNγ) or IL-4 to induce type 1 macrophage (M1) and type 1 macrophage (M2) macrophages, respectively. The Prdx1 gene knockout cells (Prdx1-/-) were used for the study. Flow cytometry was conducted to detect M1/M2 macrophage markers, and ELISA kits were used to measure M1/M2 cytokine levels. Inducible nitric-oxide synthase (iNOS) activity, arginase-1 (Arg-1) activity, and oxidative damage were also assessed. The Seahorse XFe24 Extracellular Flux Analyzer was employed to measure extracellular acidification rate and oxygen consumption rate. The mitochondrial membrane potential was analyzed using the mitochondrial membrane potential dye (JC-1) fluorescent probe, and mitochondrial superoxide was detected through fluorescence staining. Additionally, the impact of adding a mitochondrial reactive oxygen species (ROS) scavenger on RAW264.7 macrophage polarization was examined. The results demonstrated an increase in ROS, hydrogen peroxide, and 8-hydroxy-2 deoxyguanosine (8-OHDG). Cytotoxicity and mitochondrial toxic effects, including mitochondrial superoxide accumulation, decreased adenosine-triphosphate (ATP) production, reduced mitochondrial membrane potential, and decreased mitochondrial DNA copy number, were observed. Furthermore, down-regulation of translocase of inner mitochondrial membrane 23 (TIM23) mitochondrial protein and mitochondrial stress protein heat shock protein 60 (HSP60) was noted. The extra cellular acidification rate (ECAR) in M1 macrophage polarization in RAW264.7 cells was increased, while oxygen consumption rate (OCR) in M2 macrophages was reduced. These findings indicate that Prdx1 knockout in RAW264.7 cells can inhibit M2 macrophage polarization but promote M1 macrophage polarization by impairing mitochondrial function and reducing oxidative phosphorylation.

Furanodienone induces apoptosis via regulating the PRDX1/MAPKs/p53/caspases signaling axis through NOX4-derived mitochondrial ROS in colorectal cancer cells.

Furanodienone, a biologically active constituent of sesquiterpenes isolated from Rhizome Curcumae, has been reported to induce apoptosis in human colorectal cancer (CRC) cells by promoting the generation of reactive oxygen species (ROS). However, the source of ROS and how it manipulates apoptosis in CRC cells remains to be elucidated. Herein, we assessed the potential role of the well-known sources of intracellular ROS-mitochondrial electron transport chain and the nicotinamide adenine dinucleotide phosphate oxidases (NOXs), on furanodienone-induced cell death. The results indicated that furanodienone substantially increased the levels of mitochondrial ROS, which were subsequently eliminated by the general NOX inhibitor. Specifically, the nuclear factor kappa-B (NF-κB) translocation triggered a significant rise in the expression of NOX4, an isoform of the NOXs family, upon furanodienone treatment. Nevertheless, the specific NOX4 inhibitor GLX351322 attenuated cell apoptosis and mitochondrial ROS production. As a result, ROS burst induced by furanodienone suppressed the expression of peroxiredoxin1 (PRDX1), a redox signaling protein overexpressed in CRC cells, through a nuclear factor-erythroid-2-related factor 2 (Nrf2)-dependent pathway, thus amplifying the mitogen-activated protein kinases (MAPKs)/p53-mediated apoptotic signaling by increasing the p-p38, p-JNK levels, as well as the cleaved caspases -3, -8 and -9. In vivo experiments further confirmed the anti-proliferative impact of PRDX1 following furanodienone treatment. In summary, the study demonstrated that furanodienone-induced apoptosis in CRC cells is initiated by mitochondrial ROS derived from NOX4, which targeted the PRDX1 and activated the downstream MAPKs/p53-mediated caspase-dependent signaling pathway. Our findings may provide novel insights into the development of adjuvant drugs for CRC treatment and therapeutic applications.

DDAH1 recruits peroxiredoxin 1 and sulfiredoxin 1 to preserve its activity and regulate intracellular redox homeostasis.

Growing evidence suggests that dimethylarginine dimethylaminohydrolase 1 (DDAH1), a crucial enzyme for the degradation of asymmetric dimethylarginine (ADMA), is closely related to oxidative stress during the development of multiple diseases. However, the underlying mechanism by which DDAH1 regulates the intracellular redox state remains unclear. In the present study, DDAH1 was shown to interact with peroxiredoxin 1 (PRDX1) and sulfiredoxin 1 (SRXN1), and these interactions could be enhanced by oxidative stress. In HepG2 cells, H2O2-induced downregulation of DDAH1 and accumulation of ADMA were attenuated by overexpression of PRDX1 or SRXN1 but exacerbated by knockdown of PRDX1 or SRXN1. On the other hand, DDAH1 also maintained the expression of PRDX1 and SRXN1 in H2O2-treated cells. Furthermore, global knockout of Ddah1 (Ddah1-/-) or liver-specific knockout of Ddah1 (Ddah1HKO) exacerbated, while overexpression of DDAH1 alleviated liver dysfunction, hepatic oxidative stress and downregulation of PRDX1 and SRXN1 in CCl4-treated mice. Overexpression of liver PRDX1 improved liver function, attenuated hepatic oxidative stress and DDAH1 downregulation, and diminished the differences between wild type and Ddah1-/- mice after CCl4 treatment. Collectively, our results suggest that the regulatory effect of DDAH1 on cellular redox homeostasis under stress conditions is due, at least in part, to the interaction with PRDX1 and SRXN1.