Reactive Halogen Species: Role in Living Systems and Current Research Approaches.

Reactive halogen species (RHS) are highly reactive compounds that are normally required for regulation of immune response, inflammatory reactions, enzyme function, etc. At the same time, hyperproduction of highly reactive compounds leads to the development of various socially significant diseases - asthma, pulmonary hypertension, oncological and neurodegenerative diseases, retinopathy, and many others. The main sources of (pseudo)hypohalous acids are enzymes from the family of heme peroxidases - myeloperoxidase, lactoperoxidase, eosinophil peroxidase, and thyroid peroxidase. Main targets of these compounds are proteins and peptides, primarily methionine and cysteine residues. Due to the short lifetime, detection of RHS can be difficult. The most common approach is detection of myeloperoxidase, which is thought to reflect the amount of RHS produced, but these methods are indirect, and the results are often contradictory. The most promising approaches seem to be those that provide direct registration of highly reactive compounds themselves or products of their interaction with components of living cells, such as fluorescent dyes. However, even such methods have a number of limitations and can often be applied mainly for in vitro studies with cell culture. Detection of reactive halogen species in living organisms in real time is a particularly acute issue. The present review is devoted to RHS, their characteristics, chemical properties, peculiarities of interaction with components of living cells, and methods of their detection in living systems. Special attention is paid to the genetically encoded tools, which have been introduced recently and allow avoiding a number of difficulties when working with living systems.

Elucidating the modulatory effect of melatonin on enzyme activity and oxidative stress in wheat: a global meta-analysis.

In our comprehensive meta-analysis, we initially collected 177 publications focusing on the impact of melatonin on wheat. After meticulous screening, 40 published studies were selected, encompassing 558 observations for antioxidant enzymes, 312 for reactive oxygen species (ROS), and 92 for soluble biomolecules (soluble sugar and protein). This analysis revealed significant heterogeneity across studies (I2 > 99% for enzymes, ROS, and soluble biomolecules) and notable publication bias, indicating the complexity and variability in the research field. Melatonin application generally increased antioxidant enzyme activities [superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX)] in wheat, particularly under stress conditions, such as high temperature and heavy-metal exposure. Compared to control, melatonin application increased SOD, POD, CAT, and APX activities by 29.5, 16.96, 35.98, and 171.64%, respectively. Moreover, oxidative stress markers like hydrogen peroxide (H2O2), superoxide anion (O2), and malondialdehyde (MDA) decreased with melatonin by 23.73, 13.64, and 21.91%, respectively, suggesting a reduction in oxidative stress. The analysis also highlighted melatonin's role in improving carbohydrate metabolism and antioxidant defenses. Melatonin showed an overall increase of 12.77% in soluble sugar content, and 22.76% in glutathione peroxidase (GPX) activity compared to the control. However, the effects varied across different wheat varieties, environmental conditions, and application methods. Our study also uncovered complex relationships between antioxidant enzyme activities and H2O2 levels, indicating a nuanced regulatory role of melatonin in oxidative stress responses. Our meta-analysis demonstrates the significant role of melatonin in increasing wheat resilience to abiotic stressors, potentially through its regulatory impact on antioxidant defense systems and stress response.

[The auxiliary diagnostic value of ECP and MPO expression in nasal secretions in different types of rhinitis].

Objective:To evaluate the expression of eosinophil cationic protein and myeloperoxidase in nasal secretions in different types of rhinitis, and to explore their values in the differential diagnosis of different types of rhinitis. Methods:Six hundred and eighty-four subjects were selected, including 62 subjects in the acute rhinitis group, 378 subjects in the allergic rhinitis group, 94 subjects in the vasomotor rhinitis group, 70 subjects in the eosinophilic non-allergic rhinitis group, and 80 subjects in the control group. Nasal secretion samples were collected from the five groups, and the percentages of inflammatory cells were counted by Rachel's staining, and the expression of ECP/MPO was detected by colloidal gold assay. The correlation between the clinical diagnosis, the inflammatory cells in the nasal secretions and the expression of ECP/MPO was analyzed. Results:Nasal cytological smears showed that compared with the control group, the percentage of eosinophils in the AR and NARES groups were significantly higher （P<0.05）, while the percentage of neutrophils was not different （P>0.05）; the percentage of neutrophils was significantly higher in the acute rhinitis group compared with the control group （P<0.05）, while the percentage of eosinophils was not statistically different （P>0.05）; in vasomotor rhinitis group, the eosinophils and neutrophils were not statistically different compared with the control group（P> 0.05）. The colloidal gold results showed that there were differences in the expression of ECP/MPO in different types of rhinitis, among which 49 cases （79.0%） in the acute rhinitis group expressed ECP+/MPO+; 267 cases （70.6%） in the AR group and 56 cases （75.7%） in the NARES group expressed ECP+/MPO-; 80 cases （85.1%） in the vasomotor rhinitis group and 69 cases （86.3%） in the control group expressed ECP-/MPO-. Conclusion:The differences in ECP and MPO expression between different types of rhinitis have certain reference value for the differential diagnosis of different types of rhinitis and the selection of treatment programs.

Hypochlorous acid derived from microglial myeloperoxidase could mediate high-mobility group box 1 release from neurons to amplify brain damage in cerebral ischemia-reperfusion injury.

Myeloperoxidase (MPO) plays critical role in the pathology of cerebral ischemia-reperfusion (I/R) injury via producing hypochlorous acid (HOCl) and inducing oxidative modification of proteins. High-mobility group box 1 (HMGB1) oxidation, particularly disulfide HMGB1 formation, facilitates the secretion and release of HMGB1 and activates neuroinflammation, aggravating cerebral I/R injury. However, the cellular sources of MPO/HOCl in ischemic brain injury are unclear yet. Whether HOCl could promote HMGB1 secretion and release remains unknown. In the present study, we investigated the roles of microglia-derived MPO/HOCl in mediating HMGB1 translocation and secretion, and aggravating the brain damage and blood-brain barrier (BBB) disruption in cerebral I/R injury. In vitro, under the co-culture conditions with microglia BV cells but not the single culture conditions, oxygen-glucose deprivation/reoxygenation (OGD/R) significantly increased MPO/HOCl expression in PC12 cells. After the cells were exposed to OGD/R, MPO-containing exosomes derived from BV2 cells were released and transferred to PC12 cells, increasing MPO/HOCl in the PC12 cells. The HOCl promoted disulfide HMGB1 translocation and secretion and aggravated OGD/R-induced apoptosis. In vivo, SD rats were subjected to 2 h of middle cerebral artery occlusion (MCAO) plus different periods of reperfusion. Increased MPO/HOCl production was observed at the reperfusion stage, accomplished with enlarged infarct volume, aggravated BBB disruption and neurological dysfunctions. Treatment of MPO inhibitor 4-aminobenzoic acid hydrazide (4-ABAH) and HOCl scavenger taurine reversed those changes. HOCl was colocalized with cytoplasm transferred HMGB1, which was blocked by taurine in rat I/R-injured brain. We finally performed a clinical investigation and found that plasma HOCl concentration was positively correlated with infarct volume and neurological deficit scores in ischemic stroke patients. Taken together, we conclude that ischemia/hypoxia could activate microglia to release MPO-containing exosomes that transfer MPO to adjacent cells for HOCl production; Subsequently, the production of HOCl could mediate the translocation and secretion of disulfide HMGB1 that aggravates cerebral I/R injury. Furthermore, plasma HOCl level could be a novel biomarker for indexing brain damage in ischemic stroke patients.

Group A streptococcal collagen-like protein 1 restricts tumor growth in murine pancreatic adenocarcinoma and inhibits cancer-promoting neutrophil extracellular traps.

Introduction: Pancreatic ductal adenocarcinoma (PDAC) is a lethal cancer associated with an immunosuppressive environment. Neutrophil extracellular traps (NETs) were initially described in the context of infection but have more recently been implicated in contributing to the tolerogenic immune response in PDAC. Thus, NETs are an attractive target for new therapeutic strategies. Group A Streptococcus (GAS) has developed defensive strategies to inhibit NETs. Methods: In the present work, we propose utilizing intra-tumoral GAS injection to stimulate anti-tumor activity by inhibiting cancer-promoting NETs. Mice harboring Panc02 or KPC subcutaneous tumors injected with three different M-type GAS strains. Tumors and spleens were harvested at the endpoint of the experiments to assess bacterial colonization and systemic spread, while sera were analyzed for humoral responses toward the streptococcal antigens, especially the M1 and Scl1 proteins. Role of the streptococcal collagen-like protein 1 (Scl1) in anti-PDAC activity was assessed in vivo after intratumoral injection with M1 GAS wild-type, an isogenic mutant strain devoid of Scl1, or a complemented mutant strain with restored scl1 expression. In addition, recombinant Scl1 proteins were tested for NET inhibition using in vitro and ex vivo assays assessing NET production and myeloperoxidase activity. Results: Injection of three different M-type GAS strains reduced subcutaneous pancreatic tumor volume compared to control in two different murine PDAC models. Limitation of tumor growth was dependent on Scl1, as isogenic mutant strain devoid of Scl1 did not reduce tumor size. We further show that Scl1 plays a role in localizing GAS to the tumor site, thereby limiting the systemic spread of bacteria and off-target effects. While mice did elicit a humoral immune response to GAS antigens, tested sera were weakly immunogenic toward Scl1 antigen following intra-tumoral treatment with Scl1-expressing GAS. M1 GAS inhibited NET formation when co-cultured with neutrophils while Scl1-devoid mutant strain did not. Recombinant Scl1 protein inhibited NETs ex vivo in a dose-dependent manner by suppressing myeloperoxidase activity. Discussion: Altogether, we demonstrate that intra-tumoral GAS injections reduce PDAC growth, which is facilitated by Scl1, in part through inhibition of cancer promoting NETs. This work offers a novel strategy by which NETs can be targeted through Scl1 protein and potentiates its use as a cancer therapeutic.

Comparing the oxidative functions of neutrophil myeloperoxidase and cytochrome P450 enzymes in drug metabolism.

Drug metabolism is an essential process that chemically alters xenobiotic substrates to activate or terminate drug activity. Myeloperoxidase (MPO) is a neutrophil-derived haem-containing enzyme that is involved in killing invading pathogens, although consequentially, this same oxidative activity can produce metabolites that damage host tissue and play a role in various human pathologies. Cytochrome P450s (CYPs) are a superfamily of haem-containing enzymes that are significantly involved in the metabolism of drugs by functioning as monooxygenases and can be induced or inhibited, resulting in significant drug-drug interactions that lead to unanticipated adverse drug reactions. In this review, the functions of drug metabolism of MPO and CYPs are explored, along with their involvement and association for common enzymatic pathways by certain xenobiotics. MPO and CYPs metabolize numerous xenobiotics, although few reported studies have made a direct comparison between both enzymes. Additionally, we employed molecular docking to compare the active site and haem prosthetic group of MPO and CYPs, supporting their similar catalytic activities. Furthermore, we performed LCMS analysis and observed a shared hydroxylated mefenamic acid metabolite produced in both enzymatic systems. A proper understanding of the enzymology and mechanisms of action of MPO and CYPs is of significant importance when enhancing the beneficial functions of drugs in health and diminishing their damaging effects on diseases. Therefore, awareness of drugs and xenobiotic substrates involved in MPO and CYPs metabolism pathways will add to the knowledge base to foresee and prevent potential drug interactions and adverse events.

Role of neutrophil myeloperoxidase in the development and progression of high-altitude pulmonary edema.

BACKGROUND: Neutrophil infiltration and hypoxic pulmonary vasoconstriction induced by hypobaric hypoxic stress are vital in high-altitude pulmonary edema (HAPE). Myeloperoxidase (MPO), an important enzyme in neutrophils, is associated with inflammation and oxidative stress and is also involved in the regulation of nitric oxide synthase (NOS), an enzyme that catalyzes the production of the vasodilatory factor nitric oxide (NO). However, the role of neutrophil MPO in HAPE's progression is still uncertain. Therefore, we hypothesize that MPO is involved in the development of HAPE via NOS. METHODS: In Xining, China (altitude: 2260 m), C57BL/6 N wild-type and mpo-/- mice served as normoxic controls, while a hypobaric chamber simulated 7000 m altitude for hypoxia. L-NAME, a nitric oxide synthase (NOS) inhibitor to inhibit NO production, was the experimental drug, and D-NAME, without NOS inhibitory effects, was the control. After measuring pulmonary artery pressure (PAP), samples were collected and analyzed for blood neutrophils, oxidative stress, inflammation, vasoactive substances, pulmonary alveolar-capillary barrier permeability, and lung tissue morphology. RESULTS: Wild-type mice's lung injury scores, permeability, and neutrophil counts rose at 24 and 48 h of hypoxia exposure. Under hypoxia, PAP increased from 12.89 ± 1.51 mmHg under normoxia to 20.62 ± 3.33 mmHg significantly in wild-type mice and from 13.24 ± 0.79 mmHg to 16.50 ± 2.07 mmHg in mpo-/- mice. Consistent with PAP, inducible NOS activity, lung permeability, lung injury scores, oxidative stress response, and inflammation showed more significant increases in wild-type mice than in mpo-/- mice. Additionally, endothelial NOS activity and NO levels decreased more pronouncedly in wild-type mice than in mpo-/- mice. NOS inhibition during hypoxia led to more significant increases in PAP, permeability, and lung injury scores compared to the drug control group, especially in wild-type mice. CONCLUSION: MPO knockout reduces oxidative stress and inflammation to preserve alveolar-capillary barrier permeability and limits the decline in endothelial NOS activity to reduce PAP elevation during hypoxia. MPO inhibition emerges as a prospective therapeutic strategy for HAPE, offering avenues for precise interventions.

An engineered poly(A) tail attenuates gut ischemia/reperfusion-induced acute lung injury.

BACKGROUND: Gut ischemia/reperfusion causes the release of damage-associated molecular patterns, leading to acute lung injury and high mortality. Cold-inducible ribonucleic acid-binding protein is a ribonucleic acid chaperon that binds the polyadenylation tail of messenger ribonucleic acid intracellularly. Upon cell stress, cold-inducible ribonucleic acid-binding protein is released, and extracellular cold-inducible ribonucleic acid-binding protein acts as a damage-associated molecular pattern, worsening inflammation. To inhibit extracellular cold-inducible ribonucleic acid-binding protein, we have recently developed an engineered polyadenylation tail named A12. Here, we sought to investigate the therapeutic potential of A12 in gut ischemia/reperfusion-induced acute lung injury. METHODS: Male C57BL6/J mice underwent superior mesenteric artery occlusion and were treated with intraperitoneal A12 (0.5 nmol/g body weight) or vehicle at the time of reperfusion. Blood and lungs were collected 4 hours after gut ischemia/reperfusion. Systemic levels of extracellular cold-inducible ribonucleic acid-binding protein, interleukin-6, aspartate transaminase, alanine transaminase, and lactate dehydrogenase were determined. The pulmonary gene expression of cytokines (interleukin-6, interleukin-1ß) and chemokines (macrophage-inflammatory protein-2, keratinocyte-derived chemokine) was also assessed. In addition, lung myeloperoxidase, injury score, and cell death were determined. Mice were monitored for 48 hours after gut ischemia/reperfusion for survival assessment. RESULTS: Gut ischemia/reperfusion significantly increased the serum extracellular cold-inducible ribonucleic acid-binding protein levels. A12 treatment markedly reduced the elevated serum interleukin-6, alanine transaminase, aspartate transaminase, and lactate dehydrogenase by 53%, 23%, 23%, and 24%, respectively, in gut ischemia/reperfusion mice. A12 also significantly decreased cytokine and chemokine messenger ribonucleic acids and myeloperoxidase activity in the lungs of gut ischemia/reperfusion mice. Histological analysis revealed that A12 attenuated tissue injury and cell death in the lungs of gut ischemia/reperfusion mice. Finally, administration of A12 markedly improved the survival of gut ischemia/reperfusion mice. CONCLUSION: A12, a novel extracellular cold-inducible ribonucleic acid-binding protein inhibitor, diminishes inflammation and mitigates acute lung injury when employed as a treatment during gut ischemia/reperfusion. Hence, the targeted approach toward extracellular cold-inducible ribonucleic acid-binding protein emerges as a promising therapeutic strategy for alleviating gut ischemia/reperfusion-induced acute lung injury.

Dye-decolorizing peroxidase of Thermobifida halotolerance displays complex kinetics with both substrate inhibition and apparent positive cooperativity.

Dye-decolorizing peroxidases (DyPs) have been intensively investigated for the purpose of industrial dye decolourization and lignin degradation. Unfortunately, the characterization of these peroxidases is hampered by their non-Michaelis-Menten kinetics, exemplified by substrate inhibition and/or positive cooperativity. Although often observed, the underlying mechanisms behind the unusual kinetics of DyPs are poorly understood. Here we studied the kinetics of the oxidation of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), hydroquinones, and anthraquinone dyes by DyP from the bacterium Thermobifida halotolerans (ThDyP) and solved its crystal structure. We also provide rate equations for different kinetic mechanisms explaining the complex kinetics of heme peroxidases. Kinetic studies along with the analysis of the structure of ThDyP suggest that the substrate inhibition is caused by the non-productive binding of ABTS to the enzyme resting state. Strong irreversible inactivation of ThDyP by H2O2 in the absence of ABTS suggests that the substrate inhibition by H2O2 may be caused by the non-productive binding of H2O2 to compound I. Positive cooperativity was observed only with the oxidation of ABTS but not with the two electron-donating substrates. Although the conventional mechanism of cooperativity cannot be excluded, we propose that the oxidation of ABTS assumes the simultaneous binding of two ABTS molecules to reduce compound I to the enzyme resting state, and this causes the apparent positive cooperativity.

Neuropeptide W facilitates chronic gastric ulcer healing by the regulation of cyclooxygenase and NF-κB signaling pathways.

AIMS: Putative beneficial effects of neuropeptide W (NPW) in the early phase of gastric ulcer healing process and the involvement of cyclooxygenase (COX) enzymes were investigated in an acetic acid-induced gastric ulcer model. MAIN METHODS: In anesthetized male Sprague-Dawley rats, acetic acid was applied surgically on the serosa and then a COX-inhibitor (COX-2-selective NS-398, COX-1-selective ketorolac, or non-selective indomethacin; 2 mg/kg/day, 3 mg/kg/day or 5 mg/kg/day; respectively) or saline was injected intraperitoneally. One h after ulcer induction, omeprazole (20 mg/kg/day), NPW (0.1 µg/kg/day) or saline was intraperitoneally administered. Injections of NPW, COX-inhibitors, omeprazole or saline were continued for the following 2 days until rats were decapitated at the end of the third day. KEY FINDINGS: NPW treatment depressed gastric prostaglandin (PG) I2 level, but not PGE2 level. Similar to omeprazole, NPW treatment significantly reduced gastric and serum tumor necrosis factor-alpha and interleukin-1 beta levels and depressed the upregulation of nuclear factor kappa B (NF-κB) and COX-2 expressions due to ulcer. In parallel with the histopathological findings, treatment with NPW suppressed ulcer-induced increases in myeloperoxidase activity and malondialdehyde level and replenished glutathione level. However, the inhibitory effect of NPW on myeloperoxidase activity and NPW-induced increase in glutathione were not observed in the presence of COX-1 inhibitor ketorolac or the non-selective COX-inhibitor indomethacin. SIGNIFICANCE: In conclusion, NPW facilitated the healing of gastric injury in rats via the inhibition of pro-inflammatory cytokine production, oxidative stress and neutrophil infiltration as well as the downregulation of COX-2 protein and NF-κB gene expressions.

Extracellular G-quadruplexes and Z-DNA protect biofilms from DNase I, and G-quadruplexes form a DNAzyme with peroxidase activity.

Many bacteria form biofilms to protect themselves from predators or stressful environmental conditions. In the biofilm, bacteria are embedded in a protective extracellular matrix composed of polysaccharides, proteins and extracellular DNA (eDNA). eDNA most often is released from lysed bacteria or host mammalian cells, and it is the only matrix component most biofilms appear to have in common. However, little is known about the form DNA takes in the extracellular space, and how different non-canonical DNA structures such as Z-DNA or G-quadruplexes might contribute to its function in the biofilm. The aim of this study was to determine if non-canonical DNA structures form in eDNA-rich staphylococcal biofilms, and if these structures protect the biofilm from degradation by nucleases. We grew Staphylococcus epidermidis biofilms in laboratory media supplemented with hemin and NaCl to stabilize secondary DNA structures and visualized their location by immunolabelling and fluorescence microscopy. We furthermore visualized the macroscopic biofilm structure by optical coherence tomography. We developed assays to quantify degradation of Z-DNA and G-quadruplex DNA oligos by different nucleases, and subsequently investigated how these enzymes affected eDNA in the biofilms. Z-DNA and G-quadruplex DNA were abundant in the biofilm matrix, and were often present in a web-like structures. In vitro, the structures did not form in the absence of NaCl or mechanical shaking during biofilm growth, or in bacterial strains deficient in eDNA or exopolysaccharide production. We thus infer that eDNA and polysaccharides interact, leading to non-canonical DNA structures under mechanical stress when stabilized by salt. We also confirmed that G-quadruplex DNA and Z-DNA was present in biofilms from infected implants in a murine implant-associated osteomyelitis model. Mammalian DNase I lacked activity against Z-DNA and G-quadruplex DNA, while Micrococcal nuclease could degrade G-quadruplex DNA and S1 Aspergillus nuclease could degrade Z-DNA. Micrococcal nuclease, which originates from Staphylococcus aureus, may thus be key for dispersal of biofilm in staphylococci. In addition to its structural role, we show for the first time that the eDNA in biofilms forms a DNAzyme with peroxidase-like activity in the presence of hemin. While peroxidases are part of host defenses against pathogens, we now show that biofilms can possess intrinsic peroxidase activity in the extracellular matrix.

Immunohistochemical analysis of myeloperoxidase expression in cutaneous leucocytoclastic vasculitis.

Myeloperoxidase (MPO) is a pro-oxidant enzyme mainly found in the azurophilic granules of neutrophils. It not only displays a key role in the intracellular microbial killing process but also contributes to the extracellular clearance of several pathogens. This study aimed to detect MPO in cutaneous leukocytoclastic vasculitis (LCV) using immunohistochemistry. We retrospectively collected 22 confirmed cases of skin LCV diagnosed in our pathology department over 11 years (2012-2023). Immunohistochemistry was performed using anti-myeloperoxidase antibody (Leica clone 59A5) on the LeicaBond MAX automated system, following manufacturer's instructions. Two pathologists assessed immunohistochemical staining, scoring intensity as weak (+), moderate (++), or strong (+++). Patients' mean age was 56.9 years, with a male-to-female ratio of 1.18. Pathologically, vasculitis involved small blood vessels in all cases. Immunohistochemical analysis showed granular positive MPO staining in 59.1% of cases. Staining intensity was weak in 46.15%, moderate in 46.15%, and strong in 7.69%. Staining was patchy in 84.62% and diffuse in 15.38% of cases. MPO expression, detected in 59.1% of cutaneous LCV tissues, exhibited a patchy and peri-vascular distribution. It holds potential as a diagnostic marker for patients with early or minor histological changes.

In vivo and ex vivo studies support the immunostimulant and immunoprotective effect of Damiana (Turnera diffusa Willd) in Almaco Jack (Seriola rivoliana).

Damiana (Turnera diffusa Willd) was evaluated in vitro for antioxidant and antibacterial activities against Staphylococcus aureus and Streptococcus pyogenes (as a preliminary screening assessment) by high-performance thin-layer chromatography (HPTLC)-Direct bioautography. A study was performed in vivo to evaluate the effects of Damiana enriched diets at 0.5 % on immune parameters in mucus and serum and gene expression in Almaco Jack (Seriola rivoliana) intestine after two and four weeks; an infection with Aeromonas hydrophila at 1x107 colony forming units (CFU) followed and an ex vivo study was carried out using head-kidney leukocytes. Ferric reducing ability of plasma (FRAP) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assays showed high antioxidant activities in Damiana leaves; even in the ABTS assay, Damiana at 300 µg/mL showed similar activity to ascorbic acid - the standard control. Damiana exhibited strong in vitro antimicrobial activity against S. aureus and S. pyogenes. In vivo studies showed a strong enhancement of myeloperoxidase, nitric oxide, superoxide dismutase, and catalase activities in mucus and serum of S. rivoliana supplemented with Damiana; their immunological response enhanced after infection with A. hydrophila. IL-1ß, TNF-α, and IL-10 gene expressions upregulated in the fish intestine challenged with the bacterium. Piscidin and macrophage (MARCO) receptor gene expression up-regulated at week 4 and down-regulated after infection. Intestinal histology results confirm that Damiana not cause inflammation or damage. Finally, the ex vivo study confirmed the immunostimulant and protective effects of Damiana through increased phagocytic, respiratory burst, myeloperoxidase activities and nitric oxide generation before and upon the bacterial encounter. These results support the idea that Damiana has the potential as an immunostimulant additive for diets in aquaculture by enhancing immune parameters and protecting Almaco Jack against A. hydrophila infections upon four weeks of supplementation.

Fe single atoms encapsulated in N, P-codoped carbon nanosheets with enhanced peroxidase-like activity for colorimetric detection of methimazole.

Excessive use of antithyroid drug methimazole (MMI) in pharmaceutical samples can cause hypothyroidism and symptoms of metabolic decline. Hence, it is urgent to develop rapid, low cost and accurate colorimetric method with peroxidase-like nanozymes for determination of MMI in medical, nutrition and pharmaceutical studies. Herein, Fe single atoms were facilely encapsulated into N, P-codoped carbon nanosheets (Fe SAs/NP-CSs) by a simple pyrolysis strategy, as certified by a series of characterizations. UV-vis absorption spectroscopy was employed to illustrate the high peroxidase-mimicking activity of the resultant Fe SAs/NP-CSs nanozyme through the typical catalysis of 3,3',5,5'-tetramethylbenzidine (TMB) oxidation. The catalytic mechanism was scrutionously investigated by the fluorescence spectroscopy and electron paramagnetic resonance (EPR) tests. Additionally, the introduced MMI had the ability to reduce the oxidation of TMB (termed oxTMB) as a peroxidase inhibitor, coupled by fading the blue color. By virtue of the above findings, a visual colorimetric sensor was established for dual detection of methimazole (MMI) with a linear scope of 5-50 mM and a LOD of 1.57 mM, coupled by assay of H2O2 at a linear range of 3-50 mM. According to the irreversible oxidation of the drug, its screening with acceptable results was achieved on the sensing platform even in commercial tablets The Fe SAs/NP-CSs nanozyme holds great potential for clinical diagnosis and drug analysis.

Enhancing Bioethanol Production from Rice Straw through Environmentally Friendly Delignification Using Versatile Peroxidase.

Rice straw (RS), an agricultural residue rich in carbohydrates, has substantial potential for bioethanol production. However, the presence of lignin impedes access to these carbohydrates, hindering efficient carbohydrate-to-bioethanol conversion. Here, we expressed versatile peroxidase (VP), a lignin-degrading enzyme, in Pichia pastoris and used it to delignify RS at 30 °C using a membrane bioreactor that continuously discarded the degraded lignin. Klason lignin analysis revealed that VP-treatment led to 35% delignification of RS. We then investigated the delignified RS by SEC, FTIR, and SEM. The results revealed the changes of RS caused by VP-mediated delignification. Additionally, we compared the saccharification and fermentation yields between RSs treated with and without VP, VP-RS, and Ctrl-RS, respectively. This examination unveiled an improvement in glucose and bioethanol production, VP-RS exhibiting up to 1.5-fold and 1.4-fold production, respectively. These findings underscore the potential of VP for delignifying RS and enhancing bioethanol production through an eco-friendly approach.

Mechanistic investigation and dual-mode colorimetric-chemiluminescent detection of glyphosate based on the specific inhibition of Fe3O4@Cu nanozyme peroxidase-like activity.

In this study, a dual-mode colorimetric/CL nanosensor was developed for glyphosate detection based on the specific inhibition of Fe3O4@Cu peroxidase-like activity. Synthesized Fe3O4@Cu exhibited high levels of peroxidase-like activity that triggered the oxidation of luminol/3,3',5,5'-tetramethyl benzidine dihydrochloride (TMB) to excited-state 3-aminophthalic acid/blue oxTMB, thereby delivering a CL signal/visible colorimetric signal, however, the presence of glyphosate inhibited this activity, resulting in a decrease in signal strength. In-depth investigation revealed that this inhibitory mechanism occurs via two pathways: one in which glyphosate chelates with Fe(III)/Cu(II) and occupy the catalytical active sites of Fe3O4@Cu, thereby decreasing the generation of OH, and another in which glyphosate competes with TMB to consume generated OH, thus reducing the oxidation of TMB. This mechanism formed the basis of our novel dual-mode colorimetric/CL glyphosate nanosensor, which achieved limits of detection (LODs) of 0.086 µg/mL and 0.019 µg/mL in tests, thus demonstrating its significant potential for on-site glyphosate monitoring.

Phenotypic alteration by dengue virus serotype 2 delays neutrophil apoptosis and stimulates the release of prosurvival secretome with immunomodulatory functions.

Neutrophils are the most abundant granuloytes, are phenotypically heterogeneous, and exert detrimental or protective roles during antiviral response. Dengue virus has been reported to activate neutrophils. However, the effect of the dengue virus on the neutrophil phenotypes, survival, and release of inflammatory secretome is yet to be understood. Herein, we investigated the effect of dengue virus serotype 2 (DV-2) on effector functions of naïve neutrophils and studied the impact of its secretome on different immune cells. We found that DV-2 activates purified human neutrophils and causes a significant shift toward the CD16bright/CD62Ldim subtype in a multiplicity of infection and time-dependent manner. These phenotypically altered neutrophils show delayed apoptosis through nuclear factor κB and PI3K pathways and have decreased phagocytic capacity. Treatment of neutrophils with myeloperoxidase and PAD4 inhibitor before DV-2 incubation significantly reduced DV-2-induced double-stranded DNA release, suggesting that myeloperoxidase and PAD4 were involved at early stages for the neutrophil activation and double-stranded DNA release. We also report that DV-2-stimulated neutrophil secretome had a significant effect on viral infection, platelet activation, and naïve neutrophil survival via binding of tumor necrosis factor α to tumor necrosis factor receptor 1/2 receptors. Furthermore, incubation of endothelial cells with the DV-2-stimulated neutrophil secretome potentially inhibits proliferation and wound healing capacity and induces endothelial cell death, which can contribute to endothelial barrier dysfunction. In conclusion, the neutrophil-DV-2 interaction modulates the phenotype of neutrophils and the release of prosurvival and antiviral secretome that may act as a double-edged sword during dengue pathogenesis.

Cellular Senescence Contributes to the Progression of Hyperoxic Bronchopulmonary Dysplasia.

Oxidative stress, inflammation, and endoplasmic reticulum (ER) stress sequentially occur in bronchopulmonary dysplasia (BPD), and all result in DNA damage. When DNA damage becomes irreparable, tumor suppressors increase, followed by apoptosis or senescence. Although cellular senescence contributes to wound healing, its persistence inhibits growth. Therefore, we hypothesized that cellular senescence contributes to BPD progression. Human autopsy lungs were obtained. Sprague-Dawley rat pups exposed to 95% oxygen between Postnatal Day 1 (P1) and P10 were used as the BPD phenotype. N-acetyl-lysyltyrosylcysteine-amide (KYC), tauroursodeoxycholic acid (TUDCA), and Foxo4 dri were administered intraperitoneally to mitigate myeloperoxidase oxidant generation, ER stress, and cellular senescence, respectively. Lungs were examined by histology, transcriptomics, and immunoblotting. Cellular senescence increased in rat and human BPD lungs, as evidenced by increased oxidative DNA damage, tumor suppressors, GL-13 stain, and inflammatory cytokines with decreased cell proliferation and lamin B expression. Cellular senescence-related transcripts in BPD rat lungs were enriched at P10 and P21. Single-cell RNA sequencing showed increased cellular senescence in several cell types, including type 2 alveolar cells. In addition, Foxo4-p53 binding increased in BPD rat lungs. Daily TUDCA or KYC, administered intraperitoneally, effectively decreased cellular senescence, improved alveolar complexity, and partially maintained the numbers of type 2 alveolar cells. Foxo4 dri administered at P4, P6, P8, and P10 led to outcomes similar to TUDCA and KYC. Our data suggest that cellular senescence plays an essential role in BPD after initial inducement by hyperoxia. Reducing myeloperoxidase toxic oxidant production, ER stress, and attenuating cellular senescence are potential therapeutic strategies for halting BPD progression.

Protective effects of topiramate on acetic acid-induced colitis in rats through the inhibition of oxidative stress.

Ulcerative colitis is an intestinal inflammatory condition characterized by a rise in inflammatory mediator production and oxidative stress. Topiramate is an anticonvulsant agent with effectiveness on a wide range of seizures, which is anti-oxidative. This study aims to examine the protective effects of topiramate on acetic acid-induced ulcerative colitis in rats. Rats were randomly divided into four groups as follows: control, acetic acid, acetic acid + topiramate, and acetic acid + dexamethasone groups. Topiramate (100 mg/kg/day) or dexamethasone (2 mg/kg/day) was administered for six consecutive days, and ulcerative colitis was induced on the first day of the study by transrectal administration of 4% acetic acid. Four hours after the last dose of treatments, animals of each group were sacrificed, and colon tissues were removed for further macroscopic, histopathologic, and biochemical analyses. Treatment with topiramate markedly decreased colonic lesions and macroscopic scores as well as the improvement of histopathologic changes. Topiramate also effectively decreased the levels of malondialdehyde and upregulated the activity of anti-oxidative enzymes, including catalase, superoxide dismutase, and glutathione peroxidase. Our results reveal that the administration of topiramate ameliorates acetic acid-induced colitis in rats via anti-oxidative properties, and further studies may introduce it as an effective therapeutic candidate to decrease ulcerative colitis severity.