Thioredoxin Peroxidase Research Articles

Screening high-efficiency promoter to construct trans-vp28 gene Anabaena sp. PCC7120 against white spot syndrome virus of Litopenaeus vannamei

This study aimed to select high-quality promoters to construct trans-vp28 gene Anabaena sp. PCC7120 and feed Litopenaeus vannamei to assess the effect of L.vannamei against white spot syndrome virus (WSSV). Transgenic algae were created using five plasmids containing PrbcL, Pcpc560, Ptrc, Ptac, and PpsbA. According to the gene expression efficiency and the growth index of transgenic algae, Pcpc560 was determined to be the most efficient promoter. Shrimps were continuously fed trans-vp28 gene Anabaena sp. PCC7120 for one week and then challenged with WSSV. After the challenge, the transgenic algae group (vp28-7120 group) was continuously immunized [continuous immunization for 0 days (vp28-7120-0d); continuous immunization for 2 days (vp28-7120-2d); continuous immunization for 4 days (vp28-7120-4d)]. After seven days, the daily survival rate of each experimental group was continuously tracked. Following the viral challenge, the hepatopancreas samples were assayed for their levels of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), thioredoxin peroxidase (TPX), acid phosphatase (ACP), and alkaline phosphatase (AKP) at varying time intervals. In comparison to the positive control group (challenge and no vaccination) and the wild-type group (challenge, fed wild-type Anabaena sp. PCC7120), the vp28-7120 group (challenge, fed trans-vp28 gene Anabaena sp. PCC7120) exhibited a remarkable increase in survival rates, reaching 50 % (vp28-7120-0d), 76.67 % (vp28-7120-2d), and 80 % (vp28-7120-4d). Furthermore, the vp28-7120 group consistently displayed significantly higher activities of SOD, CAT, GSH-Px, ACP, and AKP, while exhibiting notably lower TPX activity, when compared to the control group. These results indicate that the Pcpc560 promoter effectively elevated the expression level of the exogenous vp28 gene and spurred the growth of the trans-vp28 gene Anabaena sp. PCC7120. Consequently, trans-vp28 gene Anabaena sp. PCC7120 significantly bolstered the immunity of L.vannamei. Therefore, utilizing the Pcpc560 promoter to develop trans-vp28 gene Anabaena sp. PCC7120 based oral vaccine is highly beneficial for industrial-scale cultivation, advancing its commercialization prospects.

Assessing the Role of Antioxidants in Migraine Patients in Erbil City

Migraine is a prevalent chronic condition that is often characterized by recurrent incapacitating headache attacks and related symptoms. The purpose of this investigation was to assess the potential effects of antioxidants on migraine disease. The main goal of the study was to measure the concentrations of many important antioxidants, such as catalase (CAT), superoxide dismutase (SOD), thioredoxin (TRX), and glutathione peroxidase 1 (GPX1). Ninety participants were recruited from the outpatient headache clinic of the neurology department at Al Jumhury Teaching Hospital in Erbil City for this study. In this study, there were 60 patients with migraine and 30 controls. Among the migraine group, there were 45 female and 15 male participants, while the control group consisted of 20 female and 10 male participants, with matching in terms of age and sex. According to study results, both male and female migraine patients had significantly lower levels of enzymes SOD and GPX1 compared to the control group. Additionally, female migraine patients had lower levels of CAT significantly compared to the control group. In male migraine patients, CAT also showed lower levels, but the difference was not statistically significant. On the other hand, TRX showed lower levels in both female and male migraine patients, but the difference was not statistically significant compared to the control group. There was a negative correlation observed between the frequency of headache-related days per month and the serum concentrations of SOD. This implies that as the levels of SOD in the serum decrease, the number of headache-related days per month tends to increase.

Recharacterization of RSL3 reveals that the selenoproteome is a druggable target in colorectal cancer.

Ferroptosis is a non-apoptotic form of cell death resulting from the iron-dependent accumulation of lipid peroxides. Colorectal cancer (CRC) cells accumulate high levels of intracellular iron and reactive oxygen species (ROS) and are thus particularly sensitive to ferroptosis. The compound (S)-RSL3 ([1S,3R]-RSL3) is a commonly used ferroptosis inducing compound that is currently characterized as a selective inhibitor of the selenocysteine containing enzyme (selenoprotein) Gluathione Peroxidase 4 (GPx4), an enzyme that utilizes glutathione to directly detoxify lipid peroxides. However, through chemical controls utilizing the (R) stereoisomer of RSL3 ([1R,3R]-RSL3) that does not bind GPx4, combined with inducible genetic knockdowns of GPx4 in CRC cell lines, we revealed that GPx4 dependency does not always align with (S)-RSL3 sensitivity, questioning the current characterization of GPx4 as the central regulator of ferroptosis. Utilizing affinity pull-down mass spectrometry with chemically modified (S)-RSL3 probes we discovered that the effects of (S)-RSL3 extend far beyond GPx4 inhibition, revealing that (S)-RSL3 is a broad and non-selective inhibitor of selenoproteins. To further investigate the therapeutic potential of broadly disrupting the selenoproteome as a therapeutic strategy in CRC, we employed additional chemical and genetic approaches. We found that the selenoprotein inhibitor auranofin, an FDA approved gold-salt, chemically induced oxidative cell death and ferroptosis in both in-vitro and in-vivo models of CRC. Consistent with these data, we found that AlkBH8, a tRNA-selenocysteine methyltransferase required for the translation of selenoproteins, is essential for the in-vitro growth and xenograft survival of CRC cell lines. In summary, these findings recharacterize the mechanism of action of the most commonly used ferroptosis inducing molecule, (S)-RSL3, and reveal that broad inhibition of selenoproteins is a promising novel therapeutic angle for the treatment of CRC.

Vitamin C as a functional enhancer in the non-specific immune defense, antioxidant capacity and resistance to low-temperature stress of juvenile mud crab, Scylla paramamosain

This experiment was conducted to explore the effects of dietary vitamin C supplementation on non-specific immune defense, antioxidant capacity and resistance to low-temperature stress of juvenile mud crab (Scylla paramamosain). Mud crabs with an initial weight of 14.67 ± 0.13 g were randomly divided into 6 treatments and fed diets with 0.86 (control), 44.79, 98.45, 133.94, 186.36 and 364.28 mg/kg vitamin C, respectively. The experiment consisted of 6 treatments, each treatment was designed with 4 replicates and each replicate was stocked with 8 crabs. After 42 days of feeding experiment, 2 crabs were randomly selected from each replicate, and a total of 8 crabs in each treatment were carried out 72 h low-temperature challenge experiment. The results showed that crabs fed diets with 186.36 and 364.28 mg/kg vitamin C significantly improved the activities of alkaline phosphatase (AKP) and acid phosphatase (ACP) in hemolymph and hepatopancreas (P < 0.05). Crabs fed diet with 133.94 mg/kg vitamin C significantly decreased the concentration of nitric oxide (NO) and the activity of nitric oxide synthase (NOS) in hemolymph (P < 0.05). Diet with 133.94 mg/kg vitamin C was improved the activity of polyphenol oxidase (PPO) and the concentration of albumin (ALB) in hemolymph. Crabs fed diet with 133.94 mg/kg vitamin C showed lower concentration of malondialdehyde (MDA) in hemolymph and hepatopancreas than those fed the other diets. Meanwhile, crabs fed diet with 98.45 mg/kg vitamin C showed higher activity of total superoxide dismutase (T-SOD) in hemolymph, and crabs fed diet with 133.94 mg/kg vitamin C showed higher activity of T-SOD in hepatopancreas. Crabs fed diet with 186.36 mg/kg vitamin C significantly decreased the concentration of reduced glutathione (GSH) and the activity of glutathione peroxidase (GSH-PX) in hepatopancreas (P < 0.05). In normal temperature, crabs fed diets with 133.94 mg/kg vitamin C significantly up-regulated the expression levels of gpx (glutathione peroxidase) and trx (thioredoxin) in hepatopancreas compared with the control treatment (P < 0.05). The highest expression levels of relish, il16 (interleukin 16), caspase 2 (caspase 2), p38 mapk (p38 mitogen-activated protein kinases) and bax (bcl-2 associated x protein) in hepatopancreas were found at crabs fed control diet (P < 0.05). Moreover, crabs fed diet with 133.94 mg/kg vitamin C showed higher expression levels of alf-3 (anti-lipopolysaccharide factor 3) and bcl-2 (B-cell lymphoma 2) in hepatopancreas than those fed the other diets (P < 0.05). Under low-temperature stress, crabs fed diet with 133.94 mg/kg vitamin C significantly improved the expression levels of hsp90 (heat shock protein 90), cat (catalase), gpx, prx (thioredoxin peroxidase) and trx in hepatopancreas (P < 0.05). In addition, dietary with 133.94 vitamin C significantly up-regulated the expression levels of alf-3 and bcl-2 (P < 0.05). Based on two slope broken-line regression analysis of activity of PPO against the dietary vitamin C level, the optimal dietary vitamin C requirement was estimated to be 144.81 mg/kg for juvenile mud crab. In conclusion, dietary 133.94–144.81 mg/kg vitamin C significantly improved the non-specific immune defense, antioxidant capacity and resistance to low-temperature stress of juvenile mud crab.

Development of a subunit vaccine against the cholangiocarcinoma causing Opisthorchis viverrini: a computational approach.

Opisthorchis viverrini is the etiological agent of the disease opisthorchiasis and related cholangiocarcinoma (CCA). It infects fish-eating mammals and more than 10 million people in Southeast Asia suffered from opisthorchiasis with a high fatality rate. The only effective drug against this parasite is Praziquantel, which has significant side effects. Due to the lack of appropriate treatment options and the high death rate, there is a dire need to develop novel therapies against this pathogen. In this study, we designed a multi-epitope chimeric vaccine design against O. viverrini by using immunoinformatics approaches. Non-allergenic and immunogenic MHC-1, MHC-2, and B cell epitopes of three candidate proteins thioredoxin peroxidase (Ov-TPx-1), cathepsin F1 (Ov-CF-1) and calreticulin (Ov-CALR) of O. viverrini, were predicted to construct a potent multiepitope vaccine. The coverage of the HLA-alleles of these selected epitopes was determined globally. Four vaccine constructs made by different adjuvants and linkers were evaluated in the context of their physicochemical properties, antigenicity, and allergenicity. Protein-protein docking and MD simulation found that vaccines 3 was more stable and had a higher binding affinity for TLR2 and TLR4 immune receptors. In-silico restriction cloning of vaccine model led to the formation of plasmid constructs for expression in a suitable host. Finally, the immune simulation showed strong immunological reactions to the engineered vaccine. These findings suggest that the final vaccine construct has the potential to be validated by in vivo and in vitro experiments to confirm its efficacy against the CCA causing O. viverrini.

Comprehensive approach to in silico identification and in vitro validation of anti-filarial hit molecules targeting the dimer interface of thioredoxin peroxidase 1 in Wuchereria bancrofti: a progress in anti-filariasis drug development.

Lymphatic filariasis (LF) remains a significant health challenge for populations in developing countries. LF is a parasitic disease transmitted by mosquitoes, mainly caused by the filarial nematode, Wuchereria bancrofti, prevalent in tropical and subtropical regions. Since the present drugs develop complications, including adverse side effects, lack of specificity, and development of drug resistance, the present study focused on developing the potential anti-filariasis drugs targeting crucial proteins for the nematode life cycle. We have identified the therapeutic compounds by targeting the enzyme thioredoxin peroxidase 1 (WbTPx1), which facilitates the conversion of hydrogen peroxide into water, an essential mechanism by which the nematode survives against oxidative stress in the host. This approach might resolve treatment efficacy and activity difficulties at various stages of filarial parasitic worms. We modeled the structure of WbTPx1 and employed the structure-based virtual screening approach, focusing on the dimer interface region of the protein. ADMET prediction profiles of the non-toxic, top-ranked hits with higher docking scores demonstrate higher affinity to the nematode protein than its human homolog. The molecular dynamic simulation studies show WbTPx1-hit complexes' stability and the intactness of hits in the binding site. Further, in vitro validation of identified hits using Setaria digitata, a cattle nematode, showed better IC50 and higher inhibition than the standard drug ivermectin, indicating the potential to inhibit enzyme activity and the development of drug candidates for controlling LF.

Protein vicinal thiols as intrinsic probes of brain redox states in health, aging, and ischemia

The nature of brain redox metabolism in health, aging, and disease remains to be fully established. Reversible oxidations, to disulfide bonds, of closely spaced (vicinal) protein thiols underlie the catalytic maintenance of redox homeostasis by redoxin enzymes, including thioredoxin peroxidases (peroxiredoxins), and have been implicated in redox buffering and regulation. We propose that non-peroxidase proteins containing vicinal thiols that are responsive to physiological redox perturbations may serve as intrinsic probes of brain redox metabolism. Using redox phenylarsine oxide (PAO)-affinity chromatography, we report that PAO-binding vicinal thiols on creatine kinase B and alpha-enolase from healthy rat brains were preferentially oxidized compared to other selected proteins, including neuron-specific (gamma) enolase, under conditions designed to trap in vivo protein thiol redox states. Moreover, measures of the extents of oxidations of vicinal thiols on total protein, and on creatine kinase B and alpha-enolase, showed that vicinal thiol-linked redox states were stable over the lifespan of rats and revealed a transient reductive shift in these redox couples following decapitation-induced global ischemia. Finally, formation of disulfide-linked complexes between peroxiredoxin-2 and brain proteins was demonstrated on redox blots, supporting a link between protein vicinal thiol redox states and the peroxidase activities of peroxiredoxins. The implications of these findings with respect to underappreciated aspects of brain redox metabolism in health, aging, and ischemia are discussed.

Differential protective impact of peptide vaccine formulae targeting the lung- and liver-stage of challenge Schistosoma mansoni infection in mice

The study aimed to elicit protective immune responses against murine schistosomiasis mansoni at the parasite lung- and liver stage. Two peptides showing amino acid sequence similarity to gut cysteine peptidases, which induce strong memory immune effectors in the liver, were combined with a peptide based on S. mansoni thioredoxin peroxidase (TPX), a prominent lung-stage schistosomula excretory-secretory product, and alum as adjuvant. Only one of the 2 cysteine peptidases-based peptides in a multiple antigenic peptide construct (MAP-3 and MAP-4) appeared to adjuvant protective immune responses induced by the TPX peptide in a MAP form. Production of TPX MAP-specific IgG1 serum antibodies, and increase in lung interleukin-1 (IL-1), uric acid, and reactive oxygen species (ROS) content were associated with significant (P < 0.05) 50 % reduction in recovery of lung-stage larvae. Increase in lung triglycerides and cholesterol levels appeared to provide the surviving worms with nutrients necessary for a stout double lipid bilayer barrier at the parasite-host interface. Surviving worms-released products elicited memory responses to the MAP-3 immunogen, including production of specific IgG1 antibodies and increase in liver IL-33 and ROS. Reduction in challenge worm burden recorded 45 days post infection did not exceed 48 % associated with no differences in parasite egg counts in the host liver and small intestine compared to unimmunized adjuvant control mice. Alum adjuvant assisted the second peptide, MAP-4, in production of IgG1, IgG2a, IgG2b and IgA specific antibodies and increase in liver ROS, but with no protective potential, raising doubt about the necessity of adjuvant addition. Accordingly, different vaccine formulas containing TPX MAP and 1, 2 or 3 cysteine peptidases-derived peptides with or without alum were used to immunize parallel groups of mice. Compared to unimmunized control mice, significant (P < 0.05 to < 0.005) 22 to 54 % reduction in worm burden was recorded in the different groups associated with insignificant changes in parasite egg output. The results together indicated that a schistosomiasis vaccine able to entirely prevent disease and halt its transmission still remains elusive.

A Physiological Approach to Explore How Thioredoxin-Glutathione Reductase (TGR) and Peroxiredoxin (Prx) Eliminate H2O2 in Cysticerci of Taenia.

Peroxiredoxins (Prxs) and glutathione peroxidases (GPxs) are the main enzymes of the thiol-dependent antioxidant systems responsible for reducing the H2O2 produced via aerobic metabolism or parasitic organisms by the host organism. These antioxidant systems maintain a proper redox state in cells. The cysticerci of Taenia crassiceps tolerate millimolar concentrations of this oxidant. To understand the role played by Prxs in this cestode, two genes for Prxs, identified in the genome of Taenia solium (TsPrx1 and TsPrx3), were cloned. The sequence of the proteins suggests that both isoforms belong to the class of typical Prxs 2-Cys. In addition, TsPrx3 harbors a mitochondrial localization signal peptide and two motifs (-GGLG- and -YP-) associated with overoxidation. Our kinetic characterization assigns them as thioredoxin peroxidases (TPxs). While TsPrx1 and TsPrx3 exhibit the same catalytic efficiency, thioredoxin-glutathione reductase from T. crassiceps (TcTGR) was five and eight times higher. Additionally, the latter demonstrated a lower affinity (>30-fold) for H2O2 in comparison with TsPrx1 and TsPrx3. The TcTGR contains a Sec residue in its C-terminal, which confers additional peroxidase activity. The aforementioned aspect implies that TsPrx1 and TsPrx3 are catalytically active at low H2O2 concentrations, and the TcTGR acts at high H2O2 concentrations. These results may explain why the T. crassiceps cysticerci can tolerate high H2O2 concentrations.

Pathway crosstalk between the central metabolic and heme biosynthetic pathways in Phanerochaete chrysosporium

A comprehensive analysis to survey heme-binding proteins produced by the white-rot fungus Phanerochaete chrysosporium was achieved using a biotinylated heme–streptavidin beads system. Mitochondrial citrate synthase (PcCS), glyceraldehyde 3-phosphate dehydrogenase (PcGAPDH), and 2-Cys thioredoxin peroxidase (mammalian HBP23 homolog) were identified as putative heme-binding proteins. Among these, PcCS and PcGAPDH were further characterized using heterologously expressed recombinant proteins. Difference spectra of PcCS titrated with hemin exhibited an increase in the Soret absorbance at 414 nm, suggesting that the axial ligand of the heme is a His residue. The activity of PcCS was strongly inhibited by hemin with Ki oxaloacetate of 8.7 μM and Ki acetyl-CoA of 5.8 μM. Since the final step of heme biosynthesis occurred at the mitochondrial inner membrane, the inhibition of PcCS by heme is thought to be a physiological event. The inhibitory mode of the heme was similar to that of CoA analogues, suggesting that heme binds to PcCS at His347 at the AcCoA–CoA binding site, which was supported by the homology model of PcCS. PcGAPDH was also inhibited by heme, with a lower concentration than that for PcCS. This might be caused by the different location of these enzymes. From the integration of these phenomena, it was concluded that metabolic regulations by heme in the central metabolic and heme synthetic pathways occurred in the mitochondria and cytosol. This novel pathway crosstalk between the central metabolic and heme biosynthetic pathways, via a heme molecule, is important in regulating the metabolic balance (heme synthesis, ATP synthesis, flux balance of the tricarboxylic acid (TCA) cycle and cellular redox balance (NADPH production) during fungal aromatic degradation.Key points• A comprehensive survey of heme-binding proteins in P. chrysosporium was achieved.• Several heme-binding proteins including CS and GAPDH were identified.• A novel metabolic regulation by heme in the central metabolic pathways was found.

The effects of thioredoxin peroxidase from Cysticercus cellulosae excretory-secretory antigens on TGF-β signaling pathway and Th17 cells differentiation in Jurkat cells by transcriptomics.

Thioredoxin peroxidase (TPx) protein from the excretory-secretory antigens (ESAs) of Cysticercus cellulosae (C. cellulosae) has been shown to regulate the differentiation of host Treg and Th17 cells, resulting in an immunosuppressive response dominated by Treg cells. However, the molecular mechanism by which TPx protein from the ESAs of C. cellulosae regulates the imbalance of host Treg/Th17 cell differentiation has not been reported. TPx protein from porcine C. cellulosae ESAs was used to stimulate Jurkat cells activated with PMA and ionomycin at 0, 24, 48, and 72h. Transcriptomic analysis was performed to investigate the signaling pathways associated with Jurkat cells differentiation regulated by TPx protein from C. cellulosae ESAs. Gene Set Enrichment Analysis (GSEA) revealed that TPx protein from porcine C. cellulosae ESAs could induce upregulation of the TGF-β signaling pathway and downregulation of Th17 cell differentiation in Jurkat cells. TPx protein from porcine C. cellulosae ESAs can activate the TGF-β signaling pathway in Jurkat cells, thereby regulating the differentiation of Treg/Th17 cells and leading to an immunosuppressive response dominated by Treg cells, enabling evasion of the host immune attack. This study provides a foundation for further validation of these pathways and further elucidates the molecular mechanisms underlying immune evasion caused by porcine C. cellulosae.

Long live the host! Proteomic analysis reveals possible strategies for parasitic manipulation of its social host.

Parasites with complex life cycles often manipulate the phenotype of their intermediate hosts to increase the probability of transmission to their definitive hosts. Infection with Anomotaenia brevis, a cestode that uses Temnothorax nylanderi ants as intermediate hosts, leads to a multiple-fold extension of host lifespan and to changes in behaviour, morphology and colouration. The mechanisms behind these changes are unknown, as is whether the increased longevity is achieved through parasite manipulation. Here, we demonstrate that the parasite releases proteins into its host with functions that might explain the observed changes. These parasitic proteins make up a substantial portion of the proteome of the hosts' haemolymph, and thioredoxin peroxidase and superoxide dismutase, two antioxidants, exhibited the highest abundances among them. The largest part of the secreted proteins could not be annotated, indicating they are either novel or severely altered during recent coevolution to function in host manipulation. We also detected shifts in the hosts' proteome with infection, in particular an overabundance of vitellogenin-like A in infected ants, a protein that regulates division of labour in Temnothorax ants, which could explain the observed behavioural changes. Our results thus suggest two different strategies that might be employed by this parasite to manipulate its host: secreting proteins with immediate influence on the host's phenotype and altering the host's translational activity. Our findings highlight the intricate molecular interplay required to influence the phenotype of a host and point to potential signalling pathways and genes involved in parasite-host communication.

High Prolactin Concentration Induces Ovarian Granulosa Cell Oxidative Stress, Leading to Apoptosis Mediated by L-PRLR and S-PRLR.

High prolactin (PRL) concentration has been shown to induce the apoptosis of ovine ovarian granulosa cells (GCs), but the underlying mechanisms are unclear. This study aimed to investigate the mechanism of apoptosis induced by high PRL concentration in GCs. Trial 1: The optimal concentration of glutathion was determined according to the detected cell proliferation. The results showed that the optimal glutathione concentration was 5 μmol/mL. Trial 2: 500 ng/mL PRL was chosen as the high PRL concentration. The GCs were treated with 0 ng/mL PRL (C group), 500 ng/mL PRL (P group) or 500 ng/mL PRL, and 5 μmol/mL glutathione (P-GSH group). The results indicated that the mitochondrial respiratory chain complex (MRCC) I-V, ATP production, total antioxidant capacity (T-AOC), superoxide dismutase (SOD), and thioredoxin peroxidase (TPx) in the C group were higher than those in the P group (p < 0.05), while they were lower than those in the P-GSH group (p < 0.05). Compared to the C group, the P group exhibited elevated levels of reactive oxygen species (ROS) and apoptosis (p < 0.05) and increased expression of ATG7 and ATG5 (p < 0.05). However, MRCC I-V, ATP, SOD, A-TOC, TPx, ROS, and apoptosis were decreased after the addition of glutathione (p < 0.05). The knockdown of either L-PRLR or S-PRLR in P group GCs resulted in a significant reduction (p < 0.05) in MRCC I-V, ATP, T-AOC, SOD and TPx, while the overexpression of either receptor showed an opposite trend (p < 0.05). Our findings suggest that high PRL concentrations induce apoptotic cell death in ovine ovarian GCs by downregulating L-PRLR and S-PRLR, activating oxidative stress and autophagic pathways.

Haloxyfop-P-methyl induces immunotoxicity and glucose metabolism disorders and affects the Nrf2/ARE pathway mediated antioxidant system in Chiromantes dehaani

Haloxyfop-P-methyl is used extensively in agricultural production, and its metabolites in soil have potentially toxic effects on aquatic ecosystems. In this study, we explored the toxicity of haloxyfop-P-methyl on Chiromantes dehaani. The results of the 21-day toxicity test showed that haloxyfop-P-methyl decreased the weight gain (WG), specific growth rate (SGR) and hepatosomatic index (HSI). In glucose metabolism, haloxyfop-P-methyl reduced pyruvate, lactate, lactate dehydrogenase and succinate dehydrogenase, but enhanced glucose-6-phosphate dehydrogenase and hexokinase. Furthermore, expression of glucose metabolism-related genes was upregulated. We cloned the full-length CdG6PDH gene, which contains a 1587 bp ORF that encoded a 528 amino acid polypeptide. In antioxidant system, haloxyfop-P-methyl increased glutathione, thioredoxin reductase and thioredoxin peroxidase activities and activated the Nrf2/ARE pathway through upregulation of ERK, JNK, PKC and Nrf2. In immunity, low concentrations haloxyfop-P-methyl, or short-term exposure, upregulated the expression of immune-related genes and enhanced immune-related enzymes activity, while high concentrations or long-term exposure inhibited immune function. In summary, haloxyfop-P-methyl inhibited the growth performance, disrupted glucose metabolism, activated the antioxidant system, and led to immunotoxicity. The results deepen our understanding of the toxicity mechanism of haloxyfop-P-methyl and provide basic biological data for the comprehensive assessment of the risk of haloxyfop-P-methyl to the environment and humans.

Evaluation of the immunoprotective effects of eight recombinant proteins from Baylisascaris schroederi in mice model

BackgroundBaylisascaris schroederi is the most common and harmful intestinal parasitic nematode of giant pandas, causing ascariasis. Although drug deworming is the main measure to control ascariasis in captive giant pandas, prolonged and repeated use of deworming drugs might induce resistance in nematodes and drug residues in giant pandas. Therefore, developing a safe and effective vaccine might provide a novel strategy to prevent ascariasis in captive giant pandas.MethodsFour highly expressed secretome genes encoding excretory and secretory proteins of B. schroederi, including transthyretin-like protein 46 (BsTLP), uncharacterized protein (BsUP), hypothetical protein 1 (BsHP1), and hypothetical protein 2 (BsHP2) and four functional genes [(encoding Galectin (BsGAL), glutathione S-transferase (BsGST), fatty acid-binding protein (BsFABP), and thioredoxin peroxidase (BsTPX)] were identified based on genome and transcriptome databases of B. schroederi and used to construct recombinant proteins via prokaryotic expression. Kunming mice were vaccinated subcutaneously twice with the recombinant proteins (50 μg/mouse) mixed with Quil A adjuvant with a 2-week interval and then orally challenged with 3000 infective eggs. The immunoprotective effects of the eight recombinant proteins on mice were assessed comprehensively using surface lesion histology scores of the mouse liver and lung, larval worm reduction, serum antibody levels (IgG, IgE, IgA, IgG1, and IgG2a), and cytokine production [interferon gamma (IFN-γ), interleukin (IL)-2, IL-4, IL-5, and IL-10].ResultsMice vaccinated with recombinant (r)BsUP (76.5%), rBsGAL (74.7%), and rBsHP2 (71.5%) showed a significant (P < 0.001) reduction in the larval worm rate compared with that in the adjuvant control. Besides, the surface lesions in the liver and lung of the vaccinated mice were alleviated. Serum levels of total IgG, IgE, IgA, IgG1, IgG2a, and cytokines, including IL-10, IL-5, and IFN-γ, were significantly higher (P < 0.001) than those in the control group.ConclusionsThe results showed that candidate three vaccines (rBsUP, rBsGAL, and rBsHP2) could provide effective protection against egg infection in mice associated with a mixed Th1/2-type immune response.Graphical

Proteomic characterization of Opisthorchis felineus exosome-like vesicles and their uptake by human cholangiocytes

The epidemiologically important food-borne trematode Opisthorchis felineus infests the liver biliary tract of fish-eating mammals and causes disorders, including bile duct neoplasia. Many parasitic species release extracellular vesicles (EVs) that mediate host–parasite interaction. Currently, there is no information on O. felineus EVs.Using gel electrophoresis followed by liquid chromatography coupled with tandem mass spectrometry, we aimed to characterize the proteome of EVs released by the adult O. felineus liver fluke. Differential abundance of proteins between whole adult worms and EVs was assessed by semiquantitative iBAQ (intensity-based absolute quantification). Imaging, flow cytometry, inhibitor assays, and colocalization assays were performed to monitor the uptake of the EVs by H69 human cholangiocytes.The proteomic analysis reliably identified 168 proteins (at least two peptides matched a protein). Among major proteins of EVs were ferritin, tetraspanin CD63, helminth defense molecule 1, globin 3, saposin B type domain-containing protein, 60S ribosomal protein, glutathione S-transferase GST28, tubulin, and thioredoxin peroxidase. Moreover, as compared to the whole adult worm, EVs proved to be enriched with tetraspanin CD63, saposin B, helminth defense molecule 1, and Golgi-associated plant pathogenesis-related protein 1 (GAPR1). We showed that EVs are internalized by human H69 cholangiocytes via clathrin-dependent endocytosis, whereas phagocytosis and caveolin-dependent endocytosis do not play a substantial role in this process.Our study describes for the first time proteomes and differential abundance of proteins in whole adult O. felineus worms and EVs released by this food-borne trematode. Studies elucidating the regulatory role of individual components of EVs of liver flukes should be continued to determine which components of EV cargo play the most important part in the pathogenesis of fluke infection and in a closely linked pathology: bile duct neoplasia. SignificanceThe food-borne trematode Opisthorchis felineus is a pathogen that causes hepatobiliary disorders in humans and animals. Our study describes for the first time the release of EVs by the liver fluke O. felineus, their microscopic and proteomic characterization, and internalization pathways by human cholangiocytes. Differential abundance of proteins between whole adult worms and EVs was assessed. EVs are enriched with canonical EV markers as well as parasite specific proteins, i.e. tetraspanin CD63, saposin B, helminth defense molecule 1, and others. Our findings will form the basis of the search for potential immunomodulatory candidates with therapeutic potential in the context of inflammatory diseases, as well as novel vaccine candidates.

Fagonia indica ameliorates chromium-induced nephrotoxicity: Role of antioxidant activity and pro-inflammatory cytokines in in-vivo renoprotection

Chromium (Cr) is an environmental pollutant, has high redox potential, and can exist in various oxidation states, possibly leading to nephrotoxicity. As a potential treatment option, Fagonia indica (F. indica) is an herb remedy traditionally used as a phytomedicine to cure ailments. However, efficient validation of its protective effect and molecular mechanisms has not yet been established. As such, this study aims to investigate the protective effect of F. indica against Cr-induced nephrotoxicity in Swiss mice. Mice were divided into five groups: group I (negative control), group II (F. indica), group III (potassium dichromate [PDC]-treated), group IV (PDC + saline), and group V (PDC + F. indica). Our results demonstrate that group III exhibited decreases in superoxide dismutase (SOD), glutathione s-transferases (GST), glutathione peroxidase (GSH-Px), catalase (CAT), and thioredoxin peroxidase (TPX) levels. Meanwhile, protein carbonyl (PCO) and malondialdehyde (MDA) levels increased in kidney homogenates, increasing the expression of the pro-inflammatory cytokine interleukin-6 (IL-6). This was followed by elevated NF-κB, blood urea nitrogen (BUN), and creatinine serum levels in group III compared with group I. Moreover, histopathological and immunohistochemical examinations demonstrated severe damage to the renal tubular epithelial cells, as well as marked congestion and expressions of caspase-3 and NF-κB. Further, group V showed an improvement in antioxidant activity parameters and reductions in the IL-6, caspase-3, and NF-κB expressions, followed by significant decreases in NF-κB, BUN, and creatinine serum levels. Furthermore, fewer histopathological disturbances were observed compared with untreated group III. Such alterations may be attributed to the antioxidant and anti-inflammatory effects of F. indica. Therefore, our exploration reveals that F. indica is effective in protecting against Cr-induced nephrotoxicity, and it could be applied in the future to human kidney diseases caused by environmental pollutants.

Enhanced stress resistance of Bifidobacterium breve NRBB57 by induction of stress proteins at near-zero growth rates.

Bifidobacterium breve is a common habitant of the human gut and is used as probiotic in functional foods. B. breve has to cope with multiple stress conditions encountered during processing and passage through the human gut, including high temperature, low pH and exposure to oxygen. Additionally, during industrial processing and in the gut, B. breve could encounter nutrient limitation resulting in reduced growth rates that can trigger adaptive stress responses. For this reason, it is important to develop culture methods that elicit resistance to multiple stresses (robustness) encountered by the bacteria. To investigate the impact of caloric restriction on robustness of the probiotic B. breve NRBB57, this strain was grown in lactose-limited chemostat cultures and in retentostat for 21 days, at growth rates ranging from 0.4 h-1 to 0.00081 h-1. Proteomes of cells harvested at different growth rates were correlated to acid, hydrogen peroxide and heat stress survival capacity. Comparative proteome analysis showed that retentostat-grown cells had significantly increased abundance of a variety of stress proteins involved in protein quality maintenance and DNA repair (DnaJ, Hsp90, FtsH, ClpB, ClpP1, ClpC, GroES, RuvB, RecA), as well as proteins involved in oxidative stress defence (peroxiredoxin, ferredoxin, thioredoxin peroxidase, glutaredoxin and thioredoxin reductase). Exposure to three different stress conditions, 45 °C, pH 3, and 10 mM H2O2, showed highest stress resistance of retentostat cells sampled at week 2 and week 3 grown at 0.0018 and 0.00081 h-1. Our findings show that cultivation at near-zero growth rates induces higher abundance of stress defence proteins contributing to the robustness of B. breve NRBB57, thereby offering an approach that may support its production and functionality.

Cloning, Expression and Evaluation of Thioredoxin Peroxidase-1 Antigen for the Serological Diagnosis of Schistosoma mekongi Human Infection.

Schistosoma mekongi, a blood fluke that causes Asian zoonotic schistosomiasis, is distributed in communities along the Mekong River in Cambodia and Lao People's Democratic Republic. Decades of employing numerous control measures including mass drug administration using praziquantel have resulted in a decline in the prevalence of schistosomiasis mekongi. This, however, led to a decrease in sensitivity of Kato-Katz stool microscopy considered as the gold standard in diagnosis. In order to develop a serological assay with high sensitivity and specificity which can replace Kato-Katz, recombinant S. mekongi thioredoxin peroxidase-1 protein (rSmekTPx-1) was expressed and produced. Diagnostic performance of the rSmekTPx-1 antigen through ELISA for detecting human schistosomiasis was compared with that of recombinant protein of S. japonicum TPx-1 (rSjTPx-1) using serum samples collected from endemic foci in Cambodia. The sensitivity and specificity of rSmekTPx-1 in ELISA were 89.3% and 93.3%, respectively, while those of rSjTPx-1 were 71.4% and 66.7%, respectively. In addition, a higher Kappa value of 0.82 calculated between rSmekTPx-1 antigen ELISA and Kato-Katz confirmed better agreement than between rSjTPx-1 antigen ELISA and Kato-Katz (Kappa value 0.38). These results suggest that ELISA with rSmekTPx-1 antigen can be a potential diagnostic method for detecting active human S. mekongi infection.

Trx CDSP32-overexpressing tobacco plants improves cadmium tolerance by modulating antioxidant mechanism

The effects of overexpression of the thioredoxin-like protein CDSP32 (Trx CDSP32) on reactive oxygen species (ROS) metabolism in tobacco leaves exposed to cadmium (Cd) were studied by combining physiological measures and proteomics technology. Thus, the number of differentially expressed proteins (DEPs) in plants overexpressing the Trx CDSP32 gene in tobacco (OE) was observed to be evidently lower than that in wild-type (WT) tobacco under Cd exposure, especially the number of down-regulated DEPs. Cd exposure induced disordered ROS metabolism in tobacco leaves. Although Cd exposure inhibited the activities of superoxide dismutase (SOD), catalase (CAT), and l-ascorbate peroxidase (APX) and the expression of proteins related to the thioredoxin-peroxiredoxin (Trx-Prx) pathway, the increase in the activities of peroxidase (POD), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), glutathione reductase (GR), glutathione peroxidase (GPX), and glutathione S-transferase (GST) and their protein expression levels played an important role in the physiological response to Cd exposure. Notably, Trx CDSP32 was observed to alleviate the decrease in the expression and activities of SOD and CAT caused by Cd exposure and enhance the function of POD. Trx CDSP32 was observed to increase the H2O2 scavenging capacity of the ascorbic acid-glutathione (AsA-GSH) cycle and Trx-Prx pathway under Cd exposure, and it can especially regulate 2-Cys peroxiredoxin (2-Cys Prx) protein expression and thioredoxin peroxidase (TPX) activity. Thus, overexpression of the Trx CDSP32 gene can alleviate the oxidative damage that occurs in tobacco leaves under Cd exposure by modulating antioxidant defense systems.