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Sulfur mustard (SM) is a chemical warfare agent that increases oxidative stress in veterans. The literature assessing oxidant/antioxidant parameters in SM-exposed veterans contains conflicting results. A total of 11 relevant studies were identified and screened. Data were extracted, and effect size and heterogeneity were assessed. The analysis revealed significant elevations in levels of malondialdehyde (MDA, an indicator of lipid peroxidation), catalase (CAT), and a reduction in glutathione (GSH) following SM exposure while superoxide dismutase (SOD) and total antioxidant levels did not change. The meta-analysis revealed that the MDA and CAT levels significantly increased in the two post-exposure sampling times (15-20 and 21-33 years) except for theGSH level, which decreased only in the post-exposure sampling time of 21-33 years. The subgroup meta-analysis of the type of analyzed samples indicated that SOD and CAT levels were only increased in the serum/plasma samples, while GSH was decreased. BALF/sputum and erythrocyte samples also revealed significant increases in MDA and SOD levels while GSH level was significantly decreased. This meta-analysis concluded that SM exposure affects the balance between oxidants and antioxidants, promoting oxidative stress that may persist long after exposure. This condition highlights the need for strategies to enhance antioxidant defenses in SM-exposed veterans., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025 Elsevier B.V. All rights reserved.)

Tetrahydrobiopterin (BH4) deficiency is caused by genetic abnormalities that impair its biosynthesis and recycling, which trigger neurochemical, metabolic, and redox imbalances. Low BH4 levels are also associated with hypoxia, reperfusion reoxygenation, endothelial dysfunction, and other conditions that are not genetically determined. The exact cause of changes in BH4 in nongenetic disorders is not entirely understood, but a role for oxidant species has been implicated. The oxidation of BH4 generates several products, including 7,8-dihydrobiopterin (BH2), the accumulation of which is predicted in cells with low dihydrofolate reductase activity. The relative efficiency of oxidant species at causing variations in BH4/BH2 levels in cells furnished with several antioxidant enzymes has not yet been systematically analyzed. This study examined the quantitative changes of BH4/BH2 in cells challenged with several oxidants. We showed that BH2 is not a major product of treatments with hydrogen peroxide or RSL3, as indicated by the moderate effect of dihydrofolate reductase-inhibitor methotrexate on the accumulation of BH2. However, we found a net loss in BH4/BH2, suggesting that products other than BH2 were generated. These reactions were further examined in NOX4-expressing HEK cells producing hydrogen peroxide. These cells showed slightly decreased BH4/BH2 ratios compared with HEK wild-type cells, and, again, methotrexate treatment moderately increased BH2 levels. In contrast, peroxynitrite-producing RAW 264.7 cells showed dramatically decreased BH4 levels without BH2 accumulation. Following the activation of peroxynitrite production with PMA in lipopolysaccharide-treated cells, we also found a significant time-dependent decline in GTPCH-I protein levels. We conclude that hydrogen peroxide is the least effective oxidant species at decreasing intracellular BH4 levels, while peroxynitrite is highly effective by targeting GTPCH-I and BH4 directly. Moreover, we conclude that BH4/BH2 levels are not a determinant of RSL3 cytotoxicity., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

BACKGROUND The aim of this study was to identify the changes in the levels of total oxidant status (TOS) and total antioxidant status (TAS) in gingival crevicular fluid (GCF) samples of patients treated with conventional brackets (CBs) and clear aligners (CAs) over a 30-day period. MATERIAL AND METHODS Thirty-four patients who received non-extraction treatment with either CBs or CAs were included in the study. GCF samples were obtained at baseline (T0) and on days 7 (T1) and 30 (T2) following initial phase of treatment. Clinical periodontal parameters were used to determine periodontal conditions of patients. ELISA method was used to determine TAS and TOS levels in GCF samples. RESULTS In the group treated with CAs, TOS levels in GCF were significantly high at T1 but approached T0 after the first month (T2) of treatment. In contrast, the group treated with CBs showed significantly high TOS levels in GCF at both T1 and T2 compared with T0. No significant changes were observed in GCF TAS data at T1 and T2 in either group (CA and CB), compared with T0. When comparing TOS data between the groups at T1 and T2, the CA group had lower TOS levels than the CB group (Mann-Whitney U with Bonferroni correction). Additionally, the CA group had lower periodontal clinical indexes than did the CB group. CONCLUSIONS Oxidative tissue damage during orthodontic teeth movements may be lower with CA treatment. In cases in which higher oxidative stress is anticipated, CA treatment may be preferred over CB treatment.

Alzheimer's disease is a neurodegenerative disease involving memory impairment, confusion, and behavioural changes. The disease is characterised by the accumulation of amyloid beta plaques and neurofibrillary tangles in the brain, which disrupt normal neuronal function. There is no known cure for Alzheimer's disease and due to increasing life expectancy, occurrence is projected to rise over the coming decades. The causes of Alzheimer's disease are multifactorial with inflammation, oxidative stress, genetic and epigenetic variation, and cerebrovascular abnormalities among the strongest contributors. We review the current literature surrounding inflammation and epigenetics in Alzheimer's disease, with a focus on how oxidants from infiltrating immune cells have the potential to alter DNA methylation profiles in the ageing brain.

The monomeric heme protein myoglobin (Mb) is aberrantly expressed in approximately 40 % of breast tumors. Mb expression is associated with better patient prognosis, yet the molecular mechanisms underlying this effect are unclear. In muscle, Mb's heme moiety confers oxygen storage and delivery. However, prior studies demonstrate that low levels of Mb in cancer cells preclude this function. Several studies propose a fatty acid binding function for Mb via lysine residue K46. Because cancer cells can upregulate fatty acid oxidation (FAO) to fuel cell migration, we tested whether Mb-mediated fatty acid binding modulates FAO and migration. We demonstrate that stable expression of human Mb in MDA-MB-231 breast cancer cells decreases cell migration and FAO. Site-directed mutagenesis of Mb K46 disrupted fatty acid binding but did not improve FAO or migration. Conversely, cells expressing Apo-Mb (with disrupted heme binding) did not show impaired FAO or migration rates, suggesting Mb attenuates FAO and migration via a heme-dependent mechanism rather than through fatty acid binding. Mb's heme-dependent oxidant generation dysregulates migratory gene expression, which is reversed by catalase treatment. Collectively, these data demonstrate that Mb's heme-dependent oxidant production decreases breast cancer cell migration, prompting therapeutic strategies to modulate oxidant production and Mb in tumors., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Thiol/disulphide homeostasis (TDH) has a critical role in many cellular activities such as antioxidant protection. Alterations of oxidative stress in the transition period play an important role in development of some diseases and disorders in dairy cows., Objectives: The purpose of this study is to assess the total antioxidant status (TAS), total oxidant status (TOS), oxidative stress index (OSI), total thiol, native thiol, disulphide and lipid profile in Simmental and Montofon dairy cows (aged 2-3) before and after calving., Methods: Blood samples were collected 233-280 days of pregnancy and the 30 days of post-partum. Serum total thiol, native thiol and disulphide levels were determined as well as TAS, TOS and paraoxonase-1 (PON-1) levels were measured using colorimetric assays. Triglycerides (TG), total cholesterol, HDL cholesterol and LDL cholesterol levels were measured with an automatic analyser., Results: Total thiol (p = 0.038) and disulphide (p = 0.015) levels were higher after calving compare to pregnancy in Montofon. TAS was found lower (p < 0.001), and OSI was higher in both breeds (Montofon p = 0.012, Simmental p = 0.028) after calving than in pregnancy. When compared between pregnancy and after calving levels in the same breed, HDL was found to be higher after calving (p < 0.001) and TG was lower after calving (p = 0.020) in Montofon. PON (p = 0.090), HDL (p < 0.001) and cholesterol levels were found higher (p < 0.001) and TG level was lower (p < 0.001) after calving in Simmental., Conclusions: According to our results, we observed different responses between two breeds before and after calving. There are few studies about TDH in animal research, and this is the first study in the literature that evaluates the TDH along with oxidative stress and lipid profiles in dairy cows in the periparturient and post-partum period., (© 2024 The Author(s). Veterinary Medicine and Science published by John Wiley & Sons Ltd.)

Photoenzymes are light-powered biocatalysts that typically rely on the excitation of cofactors or unnatural amino acids for their catalytic activities 1,2 . A notable natural example is the fatty acid photodecarboxylase, which uses light energy to convert aliphatic carboxylic acids to achiral hydrocarbons 3 . Here we report a method for the design of a non-natural photodecarboxylase based on the excitation of enzyme-bound catalytic intermediates, rather than reliance on cofactor excitation 4 . Iminium ions 5 , transiently generated from enals within the active site of an engineered class I aldolase 6 , can absorb violet light and function as single-electron oxidants. Activation of chiral carboxylic acids, followed by decarboxylation, generates two radicals that undergo stereospecific cross-coupling, yielding products with two stereocentres. Using the appropriate enantiopure chiral substrate, the desired diastereoisomeric product is selectively obtained with complete enantiocontrol. This finding underscores the ability of the active site to transfer stereochemical information from the chiral radical precursor into the product, effectively addressing the long-standing problem of rapid racemization of chiral radicals. The resulting 'memory of chirality' scenario 7 is a rarity in enantioselective radical chemistry., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Fuchs endothelial corneal dystrophy (FECD) stands as the most prevalent primary corneal endothelial dystrophy worldwide, posing a significant risk to corneal homeostasis and clarity. Corneal endothelial cells exhibit susceptibility to oxidative stress, suggesting a nuanced relationship between oxidant-antioxidant imbalance and FECD pathogenesis, irrespective of FECD genotype. Given the constrained availability of corneal transplants, exploration into non-surgical interventions becomes crucial. This encompasses traditional antioxidants, small molecule compounds, biologics, and diverse non-drug therapies, such as gene-related therapy, hydrogen therapy and near infrared light therapy. This review concentrates on elucidating the mechanisms behind oxidant-antioxidant imbalance and the evolution of strategies to restore oxidant-antioxidant balance in FECD. It provides a comprehensive overview of both conventional and emerging therapeutic approaches, offering valuable insights for the advancement of non-surgical treatment modalities. The findings herein might establish a robust foundation for future research and the therapeutic strategy of FECD., Competing Interests: Declaration of competing interest The authors have no financial/non-financial competing interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

An actin binding protein, gelsolin (GSN) has two isoforms, plasma (pGSN) and cytosolic (cGSN). Changes in pGSN and/or cGSN levels have been shown to be associated with the pathogenesis of several diseases. The aim of this study was to evaluate changes in intracellular and extracellular GSNlevels with HIF-1 in animals exposed to chronic sustained hypoxia (CSH), in addition to apoptosis and the cellular redox status. The rats in the Sham group were exposed to 21% O 2 , and the rats in the hypoxia groups were exposed to 13 and 10% O 2 , respectively. Plasma pGSN, HIF-1α, Total Antioxidant Status (TAS) and Total Oxidant Status (TOS), and lung tissue pGSN, HIF-1α, TAS, TOS, GSN levels, and apoptotic cell numbers were measured. HIF-1α levels were found to increase significantly in the tissue, especially in the group with severe hypoxia, both in biochemical and histological examinations. pGSN levels were also significantly decreased in both plasma and tissue. Significant increases in tissue were observed in cGSN. It was observed that while the antioxidant activity was dominant in the tissue, the oxidant activity was dominant in the plasma. In particular, the response to hypoxia regulated by HIF-1 is very important for cellular survival. The results of this study showed that the increase in cGSN and TAS levels in the lung tissue together with HIF-1α can be considered as the activation of mechanisms for cellular protection., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Several recent studies have attempted to understand how fasting has benefits for body health, especially the nervous system. To evaluate the impact of intermittent fasting on body weight, brain neurotransmitters, brain oxidative stress, and brain-derived neurotrophic factor (BDNF) in several areas of the brain, this study was conducted in rats. Thirty male Wistar rats were randomly divided into two groups. Group 1 (15 rats) served as the control and group 2 (15 rats) underwent intermittent fasting (IF; 24 h) for 1, 7, or 15 days. The findings demonstrated that intermittent fasting significantly reduced body weight. In this sense, brain monoamines and amino acids, namely dopamine, glutamate, aspartate, and oxidative stress markers (malondialdehyde and nitric oxide), decreased significantly after 1 day of IF. However, norepinephrine, serotonin, gamma-amino butyric acid, and glycine increased significantly. Additionally, glutathione levels were markedly elevated in IF. Surprisingly, the neuromodulatory effect of intermittent fasting fluctuates depending on the IF period. To support this fluctuation, BDNF levels increased after 1 day in the hippocampus and decreased after 15 days of intermittent fasting in all areas of the brain tested. In conclusion, our results show that intermittent fasting has beneficial influences on the brain; however, prolonged intermittent fasting can also induce some unfavorable physiological outcomes that prevent optimal neurological function., (© 2024. The Author(s).)

Background and Objective: Damage to the hippocampus leads to increased anxiety, memory problems, and learning disabilities. Melatonin (MLT), a hormone secreted by the pineal gland, serves as an antioxidant and provides defense against nerve damage. Omega-3 (ω3) is known for improving brain function. This study aims to examine the impact of melatonin and omega-3, both individually and in combination, on cognitive function, histological changes, and the balance between oxidants and antioxidants in male rats with injuries to the dorsal CA1 hippocampus., Material and Methods: Five rat groups (n = 8) were examined. The sham group was given normal saline via intraperitoneal (ip) and gavage routes. After a local lesion in the hippocampus, the lesion group underwent the same treatment. The MLT group was given melatonin (10 mg/kg, ip), the ω3 group was provided with omega-3 (0.8 g/kg, gavage), and the MLT + ω3 group received both treatments. Injections were administered every other day for 10 days. On the 11th day, behavioral assessments were conducted, and then pyramidal cells were quantified using image analysis software. Serum samples were assessed for levels of oxidants and antioxidants., Results: The results from the open field test indicated a significant increase in distance moved in the Lesion + MLT + ω3 group compared to the lesion group (P < 0.05). Performance in the novel object recognition test showed improvement in the ω3 and MLT + ω3 treated groups compared to the lesion group (P < 0.05). Additionally, social interaction duration notably increased in the ω3, MLT, and MLT + ω3 treated groups compared to the lesion group. The number of degenerated cells in the CA1, CA2, and CA3 areas of the lesion group significantly increased compared to the sham group, but melatonin and omega-3 notably reduced this number (P < 0.05). The serum levels of the antioxidant enzymes,include superoxide dismutase, glutathione peroxidase, and catalase in the lesion group notably changed compared to the sham group, but omega-3 effectively restored them to control levels., Conclusion: According to increase in distance moved, memory function, learning and social interactions of the animal in the behavioral results and the reduction of degenerate cells in the histological results, it can be said that these effects may be part of the neuroprotective effects of melatonin and omega-3. The increase in levels of antioxidant enzymes, particularly omega-3, indicates their promise as therapeutic agents for reducing oxidative stress-induced damage in neurological disorders., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Loss of protein homeostasis (proteostasis) is a common hallmark of aging and age-associated diseases. Considered as the guardian of proteostasis, the proteostasis network (PN) acts to preserve the functionality of proteins during their lifetime. However, its activity declines with age, leading to disease manifestation. While reactive oxygen species (ROS) were traditionally considered culprits in this process, recent research challenges this view. While harmful at high concentrations, moderate ROS levels protect the cell against age-mediated onset of proteotoxicity by activating molecular chaperones, stress response pathways, and autophagy. This review explores the nuanced roles of ROS in proteostasis and discusses the most recent findings regarding the redox regulation of the PN and its potential in extending healthspan and delaying age-related pathologies., Competing Interests: Declaration of interests No interests are declared., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Epilepsy is a disorder characterized by a predisposition to generate seizures. Levetiracetam (LEV) is an antiseizure drug that has demonstrated oxidant-antioxidant effects during the early stages of epilepsy in several animal models. However, the effect of LEV on oxidant-antioxidant activity during long-term epilepsy has not been studied. Therefore, the objective of the present study was to determine the effects of LEV on the concentrations of five antioxidant enzymes and on the levels of four oxidant stress markers in the hippocampus of rats with temporal lobe epilepsy at 5.7 months after status epilepticus (SE). The results revealed that superoxide dismutase (SOD) activity was significantly greater in the epileptic group (EPI) than in the control (CTRL), CTRL + LEV and EPI + LEV groups. No significant differences were found among the groups' oxidant markers. However, the ratios of SOD/hydrogen peroxide (H 2 O 2 ), SOD/glutathione peroxidase (GPx) and SOD/GPx + catalase (CAT) were greater in the EPI group than in the CTRL and EPI + LEV groups. Additionally, there was a positive correlation between SOD activity and GPx activity in the EPI + LEV group. LEV-mediated modulation of the antioxidant system appears to be time dependent; at 5.7 months after SE, the role of LEV may be as a stabilizer of the redox state.

Cyclophosphamide (CTX) is the most commonly used effective alkylating drug in cancer treatment, but its use is restricted because its toxic side effect causes testicular toxicity. CTX disrupts the tissue redox and antioxidant balance and the resulting tissue damage causes oxidative stress. In our study based on this problem, kefir against CTX-induced oxidative stress and testicular toxicity were investigated. Rats were divided into 6 groups: control, 150 mg/kg CTX, 5 and 10 mg/kg kefir, 5 and 10 mg/kg kefir + 150 CTX. While the fermented kefirs were mixed and given to the rats for 12 days, CTX was given as a single dose on the 12th day of the experiment. Testis was scored according to spermatid density, giant cell formation, cells shed into tubules, maturation disorder, and atrophy. According to our biochemical findings, the high levels of total oxidant status (TOS), and the low levels of total antioxidant status (TAS) in the CTX group, which are oxidative stress markers, indicate the toxic effect of CTX, while the decrease in TOS levels and the increase in TAS levels in the kefir groups indicate the protective effect of kefir. In the CTX-administered group, tubules with impaired maturation and no spermatids were observed in the transverse section of the testicle, while in the kefir groups, the presence of near-normal tubule structures and tubule lumens despite CTX showed the protective effect of kefir. In our study, it was observed that kefir had a protective and curative effect on CTX-induced toxicity and oxidative stress and could be a strong protector., (© 2024. The Author(s).)

Objectives: Neutrophils play a key role in ANCA-associated vasculitis, both as targets of autoimmunity and as facilitators of vascular damage. In granulomatosis with polyangiitis (GPA), the data regarding the production of reactive oxygen species (ROS) in neutrophils are unclear. Further, recent data suggests that ROS production could have an anti-inflammatory effect through the regulation of inflammasomes and IL-1-related cytokines. We aimed to analyse ROS production in neutrophils from patients with GPA and investigate its association with IL-1-related cytokines and the autoantigen PR3., Methods: Seventy-two GPA patients with disease flare were included in the NEUTROVASC prospective cohort study. ROS production in whole blood of patients with active GPA was evaluated and compared with that in the same patients in remission or healthy controls. Associations between ROS production, PR3 membrane expression on neutrophils, serum levels of IL-1-related cytokines as well as inflammasome-related proteins were analysed., Results: We observed a robust defect in ROS production by neutrophils from patients with active GPA compared with healthy controls, independent of glucocorticoid treatment. Serum levels of IL-1-related cytokines were significantly increased in GPA patients, particularly in patients with kidney involvement, and levels of these cytokines returned to normal after patients achieved remission. Further, inflammasome-related proteins were significantly dysregulated in the cytosol of neutrophils as well as the serum from GPA patients., Conclusion: Our data suggests that ROS production and regulation of inflammasomes in neutrophils from patients with GPA are disturbed and may be a potential therapeutic target., Trial Registration: ClinicalTrials.gov, https://www.clinicaltrials.gov, NCT01862068., (© The Author(s) 2023. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Oxidative stress has been known about in biological sciences for several decades; however, the understanding of this concept has evolved greatly since its foundation. Over the past years, reactive oxygen species, once viewed as solely deleterious, have become recognized as intrinsic components of life. In contrast, antioxidants, initially believed to be cure-all remedies, have failed to prove their efficacy in clinical trials. Fortunately, research on the health-promoting properties of antioxidants has been ongoing. Subsequent years showed that the former assumption that all antioxidants acted similarly was greatly oversimplified. Redox-active compounds differ in their chemical structures, electrochemical properties, mechanisms of action, and bioavailability; therefore, their efficacy in protecting against oxidative stress also varies. In this review, we discuss the changing perception of oxidative stress and its sources, emphasizing everyday-life exposures, particularly those of dietary origin. Finally, we posit that a better understanding of the physicochemical properties and biological outcomes of antioxidants is crucial to fully utilize their beneficial impact on health.

The extracellular matrix (ECM) is critical to biological architecture and determines cellular properties, function and activity. In many situations it is highly abundant, with collagens and elastin being some of the most abundant proteins in mammals. The ECM comprises of multiple different protein species and sugar polymers, with both different isoforms and post-translational modifications (PTMs) providing a large variety of microenvironments that play a key role in determining tissue structure and health. A number of the PTMs (e.g. cross-links) present in the ECM are critical to integrity and function, whereas others are deleterious to both ECM structure and associated cells. Modifications induced by reactive oxidants and electrophiles have been reported to accumulate in some ECM with increasing age. This accumulation can be exacerbated by disease, and in particular those associated with acute or chronic inflammation, obesity and diabetes. This is likely to be due to higher fluxes of modifying agents in these conditions. In this focused review, the role and effects of oxidants and other electrophiles on ECM are discussed, with a particular focus on the artery wall and atherosclerotic cardiovascular disease. Modifications generated on ECM components are reviewed, together with the effects of these species on cellular properties including adhesion, proliferation, migration, viability, metabolic activity, gene expression and phenotype. Increasing data indicates that ECM modifications are both prevalent in human and mammalian tissues and play an important role in disease development and progression., (© 2024 The Author(s).)

Background: Periodontal diseases are the most frequently diagnosed problem in cats. It has been well-established that periodontal diseases could not only cause various oral health issues but could also contribute to systemic diseases. Oxidative stress is a possible link between systemic diseases and periodontitis. Our study aimed to illustrate the influence of periodontitis on oxidative stress development in cats. Furthermore, the changes in the bacterial flora of the gums were investigated., Methods: Based on the clinical and laboratory examinations, fifty cats were divided into two groups normal (n = 25) and moderate to advanced periodontitis (n = 25). Serum total antioxidant capacity (TAC), total oxidant status (TOS), reduced (GSH) and oxidized glutathione (GSSG) were measured. In addition, samples were taken from the subgingival plaques of all cats for bacterial culture., Results: Serum TOS, GSSG, GSSG to GSH ratio, and oxidative stress index (OSI), calculated as the ratio of TOS to TAC in cats with periodontal disease were significantly higher, and TAC was significantly lower (p < 0.05) compared with controls. The results of bacterial culture indicated that the number of isolated bacterial colonies is higher in patients than in the control group. Additionally, the analysis of these data showed a positive association between periodontal index and oxidative stress., Conclusions: Our results revealed that periodontitis in cats is related to a main oxidative stress. Furthermore, oxidant factors such as TOS and OSI, compared to antioxidant factors, may better indicate the presence of oxidative stress conditions in patients with periodontitis., (© 2024. The Author(s).)

Dramatic metabolic changes during pregnancy and post-partum period resulted in alteration of the biochemical parameters in dromedary she-camels. The current study focused on assessment of stress indicators in post-partum dromedary she-camels on days 14, 28 and 42 post-calving through monitoring the clinical findings, serum steroid hormones, serum or milk oxidant/antioxidant indicators, and milk somatic cell count (SCC) status with reference to serum lipid profile changes. The study also stated several correlations between reproductive cyclicity parameters, stress biomarkers and serum-milk oxidant/antioxidant indicators. The study was conducted on clinically healthy recently calved she-camels (n = 25). They were subjected to clinical and laboratory assays including lipid profiles, serum steroid hormones [Progesterone (P4) and estradiol (E2)], serum or milk oxidant/antioxidant biomarkers [Malondialdehyde (MDA), reduced glutathione (GSH) and cortisol], and milk SCC on days 14, 21 and 28 post-calving. The study concluded the influence of stress as a result of lactation in post-partum period in recently calved she-camels and its relationship with reproductive cyclicity as well as changes in serum steroids, lipid profiles, serum-milk oxidant/antioxidants parameters, and milk SCC that was reflected through significant elevations in serum levels of P4, E2, cortisol, MDA and glucose, and milk values of MDA, cortisol and SCC as well as significant drop in serum levels of GSH, TPs, albumins and globulins on day 14 post-calving comparing with their values particularly on day 42. The study stated variable correlation relationships between reproductive cyclicity parameters, lipid profiles, serum-milk oxidant/antioxidants parameters and milk SCC., (© 2024. The Author(s).)

Diabetes in humans a chronic metabolic disorder characterised by hyperglycaemia, it is associated with an increased risk of cardiovascular disease, disruptions to metabolism and vascular functions. It is also linked to oxidative stress and its complications. Its role in vascular dysfunctions is generally reported without detailed impact on the molecular mechanisms. Potassium ion channel (K + channels) are key regulators of vascular tone, and as membrane proteins, are modifiable by oxidant stress associated with diabetes. This review manuscript examined the impact of oxidant stress on vascular K + channel functions in diabetes, its implication in vascular complications and metabolic and cardiovascular diseases.

Background: Unexplained infertility is defined as the absence of any pathology in the basic evaluation performed in couples who cannot achieve pregnancy after 1 year of unprotected sexual intercourse. The results of tests examining the causes of infertility show no identifiable cause in almost 15% of couples., Aim: The aim of this study was to investigate the effects of reactive oxygen species (ROS) on pregnancy and embryos., Methods: This study included 200 patients, aged between 20-44 years, with unexplained infertility, who had recurrent intrauterine inseminations failures and hence started in vitro fertilization (IVF)/intracytoplasmic sperm injection treatment. Some amounts of waste follicular fluid samples were collected by embryologists from the oocytes of these patients during the ovum pick-up procedure. Next, total antioxidant status (TAS), total oxidant status (TOS), and oxidative stress index (OSI) values were calculated in the biochemistry laboratory., Results: In terms of pregnancy status, both follicular TOS and OSI values were not significantly different in patients with biochemical and clinical pregnancy, whereas TAS values were significantly higher in patients with pregnancy (P < 0.05). In terms of embryo quality, no significant difference was observed in TAS, TOS, and OSI values between grade 1 and 2 embryos, whereas pregnancy rates were significantly higher in patients who received grade 1 embryo transfer (P < 0.05). However, the follicular fluid TAS levels were significantly lower in smoking patients than in those who did not smoke; TOS and OSI levels were significantly higher., Conclusion: This study showed that exposure to oxidative stress might be a causative factor for infertility. In addition, ROS decreased the level of TAS by increasing OSI in the follicular fluid; thus, antioxidant supplementation might be a necessity., (Copyright © 2024 Copyright: © 2024 Nigerian Journal of Clinical Practice.)

A critical feature of the cellular antioxidant response is the induction of gene expression by redox-sensitive transcription factors. In many cells, activating these transcription factors is a dynamic process involving multiple redox steps, but it is unclear how these dynamics should be measured. Here, we show how the dynamic profile of the Schizosaccharomyces pombe Pap1 transcription factor is quantifiable by three parameters: signal amplitude, signal time and signal duration. In response to increasing hydrogen peroxide concentrations, the Pap1 amplitude decreased while the signal time and duration showed saturable increases. In co-response plots, these parameters showed a complex, non-linear relationship to the mRNA levels of four Pap1-regulated genes. We also demonstrate that hydrogen peroxide and tert-butyl hydroperoxide trigger quantifiably distinct Pap1 activation profiles and transcriptional responses. Based on these findings, we propose that different oxidants and oxidant concentrations modulate the Pap1 dynamic profile, leading to specific transcriptional responses. We further show how the effect of combination and pre-exposure stresses on Pap1 activation dynamics can be quantified using this approach. This method is therefore a valuable addition to the redox signalling toolbox that may illuminate the role of dynamics in determining appropriate responses to oxidative stress., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Parabens are being used as preservatives due to their antifungal and antimicrobial effects. They are emerging as aquatic pollutants due to their excessive use in many products. The purpose of this study was to determine the toxic effect of ethyl paraben (C9H10O3) on the hematobiochemical, histological, oxidative, and anti-oxidant enzymatic and non-enzymatic activity; the study also evaluates the potential of ethyl paraben to cause genotoxicity in Rohu Labeo rohita. A number of 15 fish with an average weight of 35.45±1.34g were placed in each group and exposed to ethyl paraben for 21 days. Three different concentrations of ethyl paraben, i.e., T1 (2000μg/L), T2 (4000 μg/L), andT3 (6000 μg/L) on which fish were exposed as compared to the control T0 (0.00 μg/L). Blood was used for hematobiochemical and comet assay. Gills, kidneys, and liver were removed for histological alterations. The results showed a significant rise in all hemato-biochemical parameters such as RBCs, WBCs, PLT count, blood sugar, albumin, globulin, and cholesterol. An increase in aspartate aminotransferase (AST) and alanine transaminase (ALT) levels directed the hepatocytic damage. Histological alterations in the liver, gills and kidneys of fish were found. Ethylparaben induces oxidative stress by suppressing antioxidant enzyme activity such as SOD, GSH, CAT and POD. Based on the comet assay, DNA damage was also observed in blood cells, resulting in genotoxicity. Findings from the present study indicate that ethyl paraben induces hemato-biochemical alterations, tissue damage, oxidative stress, and genotoxicity., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Akmal et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: Although oxidative stress is involved in the pathophysiological process of chronic rhinosinusitis with nasal polyps (CRSwNP), the specific underlying mechanism is still unclear. Whether antioxidant therapy can treat CRSwNP needs further investigation., Methods: Immunohistochemistry, immunofluorescence, western blotting and quantitative polymerase chain reaction (qPCR) analyses were performed to detect the distribution and expression of oxidants and antioxidants in nasal polyp tissues. qPCR revealed correlations between oxidase, antioxidant enzymes and inflammatory cytokine levels in CRSwNP patients. Human nasal epithelial cells (HNEpCs) and primary macrophages were cultured to track the cellular origin of oxidative stress in nasal polyps(NPs) and to determine whether crocin can reduce cellular inflammation by increasing the cellular antioxidant capacity., Results: The expression of NOS2, NOX1, HO-1 and SOD2 was increased in nasal epithelial cells and macrophages derived from nasal polyp tissue. Oxidase levels were positively correlated with those of inflammatory cytokines (IL-5 and IL-6). Conversely, the levels of antioxidant enzymes were negatively correlated with those of IL-13 and IFN-γ. Crocin inhibited M1 and M2 macrophage polarization as well as the expression of NOS2 and NOX1 and improved the antioxidant capacity of M2 macrophages. Moreover, crocin enhanced the ability of antioxidants to reduce inflammation via the KEAP1/NRF2/HO-1 pathway in HNEpCs treated with SEB or LPS. Additionally, we observed the antioxidant and anti-inflammatory effects of crocin in nasal explants., Conclusion: Oxidative stress plays an important role in the development of CRSwNP by promoting various types of inflammation. The oxidative stress of nasal polyps comes from epithelial cells and macrophages. Antioxidant therapy may be a promising strategy for treating CRSwNP., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Zhou, Zhou, Liu, Liu, Meng, Wen, Luo, Liu, Li, Ba and Du.)

Cytochrome P450 enzymes are known to catalyse bimodal oxidation of aliphatic acids via radical intermediates, which partition between pathways of hydroxylation and desaturation 1,2 . Developing analogous catalytic systems for remote C-H functionalization remains a significant challenge 3-5 . Here, we report the development of Cu(I)-catalysed bimodal dehydrogenation/lactonization reactions of synthetically common N-methoxyamides through radical abstractions of the γ-aliphatic C-H bonds. The feasibility of switching from dehydrogenation to lactonization is also demonstrated by altering reaction conditions. The use of a readily available amide as both radical precursor and internal oxidant allows for the development of redox-neutral C-H functionalization reactions with methanol as the sole side product. These C-H functionalization reactions using a Cu(I) catalyst with loading as low as 0.5 mol.% is applied to the diversification of a wide range of aliphatic acids including drug molecules and natural products. The exceptional compatibility of this catalytic system with a wide range of oxidatively sensitive functionality demonstrates the unique advantage of using a simple amide substrate as a mild internal oxidant., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

To establish infections in human hosts, Pseudomonas aeruginosa must overcome innate immune-generated oxidative stress, such as the hypochlorous acid (HOCl) produced by neutrophils. We set out to find specific biomarkers of oxidative stress through the development of a protocol for the metabolic profiling of P. aeruginosa cultures grown in the presence of different oxidants using a novel ionization technique for mass spectrometry, laser desorption rapid evaporative ionization mass spectrometry (LD-REIMS). We demonstrated the ability of LD-REIMS to classify samples as untreated or treated with a specific oxidant with 100% accuracy and identified a panel of 54 metabolites with significantly altered concentrations after exposure to one or more of the oxidants. Key metabolic changes were conserved in P. aeruginosa clinical strains isolated from patients with cystic fibrosis lung infections. These data demonstrated that HOCl stress impacted the Pseudomonas quinolone signal (PQS) quorum sensing system. Ten 2-alkyl-4-quinolones (AHQs) associated with the PQS system were significantly lower in concentration in HOCl-stressed P. aeruginosa cultures, including 2-heptyl-3-hydroxy-4(1H)-quinolone (PQS), the most active signal molecule of the PQS system. The PQS system regulates the production of virulence factors, including pyocyanin and elastase, and their levels were markedly affected by HOCl stress. No pyocyanin was detectable and elastase concentrations were reduced by more than 75% in cultures grown with sub-lethal concentrations of HOCl, suggesting that this neutrophil-derived oxidant may disrupt the ability of P. aeruginosa to establish infections through interference with production of PQS-associated virulence factors., Importance: This work demonstrates that a high-throughput ambient ionization mass spectrometry method can be used successfully to study a bacterial stress response. Its application to the opportunistic pathogen Pseudomonas aeruginosa led to the identification of specific oxidative stress biomarkers, and demonstrated that hypochlorous acid, an oxidant specifically produced by human neutrophils during infection, affects quorum sensing and reduces production of the virulence factors pyocyanin and elastase. No pyocyanin was detectable and elastase levels were reduced by more than 75% in bacteria grown in the presence of hypochlorous acid. This approach has the potential to be widely applicable to the characterization of the stress responses of bacteria., Competing Interests: The authors declare no conflict of interest.

Objectives: To examine the influence of hirudotherapy on parameters of oxidative stress., Methods: The cross-sectional study was conducted from March 29 to September 29, 2021, at the Alanya Research and Training Hospital's Traditional and Complementary Medicine Application Centre, Turkey, and comprised adult volunteers of either gender. The participants were subjected to two sessions of hirudotherapy 4 weeks apart. Total antioxidant status, total oxidant status, oxidative stress index values, ischaemia-modified albumin level, paraoxonase 1, disulfide, native thiol, total thiol, and arylesterase levels were assessed at baseline and after the second hirudotherapy session. Data was analysed using SPSS 15., Results: Of the 50 subjects, 30(60%) were females and 20(40%) were males. The overall mean age was 47.10±15.16 years. Oxidative stress, ischaemia-modified albumin and disulfide levels decreased, but not significantly (p>0.05). The reduction in disulfide levels was significant (p=0.021)., Conclusions: Hirudotherapy, within its limitations, could reduce oxidative stress.

High sodium intake is decisive in the incidence increase and prevalence of hypertension, which has an impact on skeletal muscle functionality. Diazoxide is an antihypertensive agent that inhibits insulin secretion and is an opener of K ATP channels (adosine triphosphate sensitive potasium channels). For this reason, it is hypothesized that moderate-intensity exercise and diazoxide improve skeletal muscle function by reducing the oxidants in hypertensive rats. Male Wistar rats were assigned into eight groups: control (CTRL), diazoxide (DZX), exercise (EX), exercise + diazoxide (EX + DZX), hypertension (HTN), hypertension + diazoxide (HTN + DZX), hypertension + exercise (HTN + EX), and hypertension + exercise + diazoxide (HTN + EX + DZX). To induce hypertension, the rats received 8% NaCl dissolved in water orally for 30 days; in the following 8 weeks, 4% NaCl was supplied to maintain the pathology. The treatment with physical exercise of moderate intensity lasted 8 weeks. The administration dose of diazoxide was 35 mg/kg intraperitoneally for 14 days. Tension recording was performed on the extensor digitorum longus and the soleus muscle. Muscle homogenates were used to measure oxidants using fluorescent probe and the activity of antioxidant systems. Diazoxide and moderate-intensity exercise reduced oxidants and increased antioxidant defenses., (© 2024 The Authors. Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

First in the literature this study aimed to investigate the effects of Tartrazine, a common industrial food dye, on kidney and whether Thymoquinone has a protective effect in tartrazine-induced nephrotoxicity. The study conducted on the rats bred at İnönü University Experimental Animals Production and Research Center. Wistar albino rats were randomly divided into 4 groups, where each group included 8 rats: control, Tartrazine, Thymoquinone, and Tartrazine + Thymoquinone groups. The experiments continued for 3 weeks and then, kidney tissues and blood samples were collected from the rats under anesthesia. Malondialdehyde (MDA), super oxidized dismutase (SOD), total oxidant status (TOS), increase in Oxidative stress index (OSI), glutathione (GSH), Glutathione peroxidase (GSH-Px), catalase (CAT), Total antioxidant status (TAS) levels decreased in the kidney tissues collected from the tartrazine group. Serum Bun and Creatinine levels increased in the tartrazine group. Tartrazine administration damaged and degenerated the glomeruli and cortical distal tubes in the histopathology of kidney tissues, also different degrees of inflammatory cell infiltration were observed in the renal cortex and medulla. Thymoquinone and tartrazine administration improved both biochemical and histopathological parameters. Tartrazine administration induced nephrotoxicity. This could be observed with the increase in oxidant capacity and the deterioration of kidney functions. Thymoquinone was observed to demonstrate strong antioxidant properties. Thymoquinone could be used primarily as a protective agent against Tartrazine-induced toxicity., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Ethnopharmacological Relevance: Ulcerative colitis (UC) which has a global impact on the health care system with its recurrent and incompletely curable characteristics, affects the patients' quality of life. Gilaburu (GB; Viburnum opulus L.) is a fruit with rich polyphenol ingredient which is used ethnobotanically in Türkiye for medicinal purposes (for example, to pass kidney stones, to treat stomach, heart, and liver diseases, hemorrhages, hypertension, ulcers, common cold, tuberculosis, rheumatic and menstrual pain, and diabetes). On the other hand, the effects of GB in the experimental UC model have not been studied., Aim of the Study: This study aimed to explore the potential antioxidant and anti-inflammatory effects of GB fruit extract in improving acetic acid (AA)-induced UC., Materials and Methods: Starting immediately after (AA + GB group) or 1 week before (GB + AA + GB group) the colitis induced by intrarectal AA (5%; v/v) administration, the rats orally received GB (100 mg/kg) once per day for 3 days. The control and AA groups were administered orally saline (1 ml), while the AA + SS group were administered sulfasalazine (SS; 100 mg/kg; orally) as a positive control once per day for 3 days. Distal colonic tissue specimens were obtained for the histological and biochemical [myeloperoxidase (MPO), malondialdehyde (MDA), glutathione (GSH), chemiluminescence (CL), caspase-3, 8-hydroxy-2 ' -deoxyguanosine (8-OHdG), matrix metalloproteinase (MMP)-9, transforming growth factor (TGF)-β 1 , smad-3 and cytokine (tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, IL-8, interferon (IFN)-γ), measurements] evaluations on the 3 rd day., Results: Elevated macroscopic and microscopic damage scores, high tissue wet weight values, increased tissue-associated MPO, MDA, CL, caspase-3, 8-OHdG, cytokines (TNF-α, IL-1β, IL-6, IL-8), MMP-9, TGF-β 1 , smad-3 levels, and decreased GSH values of the AA group were all reversed by GB treatments (AA + GB and GB + AA + GB groups) (p < 0.05-0.001). However, sulfasalazine treatment (AA + SS group) did not change the IL-8, 8-OHdG, MMP-9, and TGF-β 1 measurements significantly., Conclusions: Gilaburu shows both anti-inflammatory and antioxidant effects against AA-induced colonic damage by suppressing neutrophil infiltration, regulating inflammatory mediators, inhibiting reactive species production, lipid peroxidation, and apoptosis, conserving endogenous antioxidant glutathione, and ameliorating oxidative DNA damage. Since the current ulcerative colitis drugs display limited benefits and adverse side effects, potential therapeutic and/or prophylactic role of gilaburu can be evaluated in ulcerative colitis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Nuclear protein trafficking requires the soluble transport factor RanBP1. The subcellular distribution of RanBP1 is dynamic, as the protein shuttles between the nucleus and cytoplasm. To date, the signaling pathways regulating RanBP1 subcellular localization are poorly understood. During interphase, RanBP1 resides mostly in the cytoplasm. We show here that oxidative stress concentrates RanBP1 in the nucleus, and our study defines the underlying mechanisms. Specifically, RanBP1's cysteine residues are not essential for its oxidant-induced relocation. Furthermore, our pharmacological approaches uncover that signaling mediated by epidermal growth factor receptor (EGFR) and protein kinase A (PKA) control RanBP1 localization during stress. In particular, pharmacological inhibitors of EGFR or PKA diminish the oxidant-dependent relocation of RanBP1. Mutant analysis identified serine 60 and tyrosine 103 as regulators of RanBP1 nuclear accumulation during oxidant exposure. Taken together, our results define RanBP1 as a target of oxidative stress and a downstream effector of EGFR and PKA signaling routes. This positions RanBP1 at the intersection of important cellular signaling circuits., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier GmbH.. All rights reserved.)

Objective: Acrylamide (AA) is toxic and forms in food that undergoes high-temperature processing. This study aimed to investigate the effects of AA-induced toxicity on renal tissue in pinealectomized rats and the possible protective effect of exogenous Melatonin (ML) administration., Materials and Methods: Sixty rats were randomized into 6 groups (n = 10): Sham, Sham+AA, Sham+AA+ML, PX, PX+AA, and PX+AA+ML. Sham and pinealectomized rats received AA (25 mg/kg/day orally) and ML (0.5 ml volume at 10 mg/kg/day, intraperitoneal) for 21 days., Results: The results showed that malondialdehyde (MDA), total oxidant status (TOS), oxidative stress index (OSI), tumor necrosis factor-α (TNF-α), and interleukin 1β (IL-1β) levels of the kidney and urea and creatinine levels of serum in the PX (pinealectomy)+AA group were more increased than in the Sham+AA group. In addition, glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), and total antioxidant status (TAS) levels decreased more in the PX+AA group than in the Sham+AA group. Also, we observed more histopathologic damage in the PX+AA group. On the other hand, up-regulation of kidney tissue antioxidants, down-regulation of tissue oxidants, and improvement in kidney function were achieved with ML treatment. Also, histopathological findings such as inflammatory cell infiltration, shrinkage of glomeruli, and dilatation of tubules caused by AA toxicity improved with ML treatment., Conclusion: ML supplementation exhibited adequate nephroprotective effects against the nephrotoxicity of AA on pinealectomized rat kidney tissue function by balancing the oxidant/antioxidant status and suppressing the release of proinflammatory cytokines., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Misfolded proteins are usually refolded to their functional conformations or degraded by quality control mechanisms. When misfolded proteins evade quality control, they can be sequestered to specific sites within cells to prevent the potential dysfunction and toxicity that arises from protein aggregation. Btn2 and Hsp42 are compartment-specific sequestrases that play key roles in the assembly of these deposition sites. Their exact intracellular functions and substrates are not well defined, particularly since heat stress sensitivity is not observed in deletion mutants. We show here that Btn2 and Hsp42 are required for tolerance to oxidative stress conditions induced by exposure to hydrogen peroxide. Btn2 and Hsp42 act to sequester oxidized proteins into defined PQC sites following ROS exposure and their absence leads to an accumulation of protein aggregates. The toxicity of protein aggregate accumulation causes oxidant sensitivity in btn2 hsp42 sequestrase mutants since overexpression of the Hsp104 disaggregase rescues oxidant tolerance. We have identified the Sup35 translation termination factor as an in vivo sequestrase substrate and show that Btn2 and Hsp42 act to suppress oxidant-induced formation of the yeast [PSI+] prion, which is the amyloid form of Sup35. [PSI+] prion formation in sequestrase mutants does not require IPOD (insoluble protein deposit) localization which is the site where amyloids are thought to undergo fragmentation and seeding to propagate their heritable prion form. Instead, both amorphous and amyloid Sup35 aggregates are increased in btn2 hsp42 mutants consistent with the idea that prion formation occurs at multiple intracellular sites during oxidative stress conditions in the absence of sequestrase activity. Taken together, our data identify protein sequestration as a key antioxidant defence mechanism that functions to mitigate the damaging consequences of protein oxidation-induced aggregation., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Carter et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Macrophages count on two O 2 -consuming enzymes to form reactive radical species: NAPDH oxidase 2 (Nox2) and nitric oxide synthase 2 (inducible isoform, iNOS) that produce superoxide radical (O 2 • - ) and nitric oxide ( • NO), respectively. If formed simultaneously, the diffusion-controlled reaction of O 2 • - and • NO yields peroxynitrite, a potent cytotoxic oxidant. In human tissues and cells, the oxygen partial pressure (pO 2 ) normally ranges within 2-14 %, with a typical average pO 2 value for most tissues ca. 5 %. Given that O 2 is a substrate for both Nox2 and iNOS, its tissue and cellular concentration can affect O 2 • - and • NO production. Also, O 2 is a modulator of the macrophage adaptative response and may influence iNOS expression in a hypoxia inducible factor 1-α (HIF1α-)-dependent manner. However, most of the reported experiments in cellula, analyzing the formation and effects of O 2 • - and • NO during macrophage activation and cytotoxicity towards pathogens, have been performed in cells exposed to atmospheric air supplemented with 5 % CO 2 ; under these conditions, most cells are exposed to supraphysiologic oxygen tensions (ca. 20 % O 2 ) which are far from the physiological pO 2 . Here, the role of O 2 as substrate in the oxidative response of J774A.1 macrophages was explored upon exposure to different pO 2 and O 2 • - and • NO formation rates were measured, obtaining a K M of 26 and 42 μM O 2 for Nox2 and iNOS, respectively. Consequently, peroxynitrite formation was influenced by pO 2 , reaching a maximum at ≥ 10 % O 2 , but even at levels as low as 2 % O 2 , a substantial formation rate of this oxidant was detected. Indeed, the cytotoxic capacity of immunostimulated macrophages against the intracellular parasite T. cruzi was significant, even at low pO 2 values, confirming the role of peroxynitrite as a potent oxidizing cytotoxin within a wide range of physiological oxygen tensions., Competing Interests: Declaration of competing interest The authors have no competing interests to declare., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Background: The aim of this study was to quantify serum levels of elafin, a serine protease inhibitor, and to assess its effects on histopathological and biochemical parameters in hepatic ischemia-reperfusion injury., Methods: Forty female Wistar albino rats were divided into five groups: Group 1 served as the control group. Liver ischemia was induced for 30 minutes in the other four groups. An additional 1-hour, 2-hour, and 3-hour reperfusion was induced in Groups 3, 4, and 5, respectively. At the end of the experiment, intracardiac blood samples were obtained for biochemical examination, and tissue samples from the liver were taken for histopathological examination. Levels of elafin, ischemia-modified albumin (IMA), total antioxi-dant status (TAS), and total oxidant status (TOS) were also examined., Results: Serum elafin levels decreased beginning from Group 2, with the lowest level reached in Group 5 (p<0.01). The IMA level was the lowest in the control group and the highest in Group 5 (p<0.01). TOS, aspartate aminotransferase (AST), and alanine amino-transferase (ALT) levels were lowest in the control group and highest in Group 5 (p<0.01). Group 5 had the highest IMA/albumin ratio, although no significant differences were found between these four groups. The lowest TAS level was found in the control group, but a stable and significant increase was not detected in the other groups. No significant differences were found between the groups in terms of alkaline phosphatase (ALP) and albumin levels. A negative correlation was observed between serum elafin levels and AST, ALT, and TOS levels (p<0.01). The number of Grade 1 histopathological results was found to be higher in the groups with reperfusion (Groups 3, 4, 5). In histopathological subgroup analysis, while the elafin level was lower in Grade 1 group, AST, ALT, and TOS levels were higher (p<0.01). Additionally, the IMA/albumin ratio was found to be higher in the Grade 1 group (p=0.02)., Conclusion: In hepatic ischemia-reperfusion injury, elafin levels decreased as the reperfusion time increased. As the reperfusion time increased, both hepatocyte damage and oxidant capacity increased, with a negative correlation observed between these findings and elafin levels. Therefore, elafin may play a protective role in hepatic ischemia-reperfusion injury and could assist clinicians in assessing liver injury.

Acrylamide (ACR) is a toxic chemical frequently encountered in daily life, posing health risks. This study aimed to elucidate the molecular-level mechanism of ACR's toxic effects on testicles and investigate whether Vitamin E can mitigate these effects. A total of 40 adult pregnant rats were utilized, divided into four groups: Control, ACR, Vitamin E, and ACR + Vitamin E. ACR and Vitamin E were administered to the mother rats during pregnancy and lactation, and to the male offspring until the 8th week post-birth. Serum hormone levels, oxidant-antioxidant parameters, histopathological examination of testicular tissue, and mRNA and protein levels of the testicular and liver aromatase gene were analyzed. Spermiogram analysis was conducted on the collected sperm samples from the male offspring. The results revealed that ACR exposure adversely affected hormone levels, oxidant-antioxidant parameters, histological findings, as well as aromatase gene and protein expressions. However, Vitamin E administration effectively prevented the toxic effects of ACR. These findings demonstrate that ACR application significantly impairs the reproductive performance of male offspring rats by increasing liver aromatase activity., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: Cisplatin-containing regimen is an effective treatment for several malignancies. However, cisplatin is an important cause of nephrotoxicity. So, many trials were performed to transplant stem cells systemically or locally to control cisplatin-induced nephrotoxicity. Stem cell therapeutic effect may be dependent on the regulation of inflammation and oxidant stress., Aim: To investigate the effect of human umbilical cord blood-mesenchymal stem cells (hUCB-MSCs) on the histological structure, the oxidant stress, and the inflammatory gene expression in an experimental model of cisplatin-induced nephrotoxicity in rats., Method: The rats were divided into 6 equal groups (each of 10 rats): Group I included normal rats that received no treatment. Group II included healthy rats that received IV hUCB-MSCs. Group III included untreated cisplatin-induced nephrotoxic rats. Group IV included cisplatin-induced nephrotoxic rats that received magnesium (Mg) injections after injury. Group V was injected with hUCB-MSCs after injury. Group VI received both Mg and hUCB-MSCs after injury. In tissue homogenates, reduced glutathione (GSH), superoxide dismutase (SOD), and malondialdehyde (MDA) activities were measured. Quantitative real-time-polymerase chain reaction (qRT-PCR) was performed to assess iNOS, TLR4, and NF-kB gene expression. Hematoxylin and eosin (H&E) staining was performed to study the histological structure of the kidney. Immunohistochemical staining of iNOS and NF-κB was performed, as well., Results: Disturbed kidney functions, oxidative status, and histological structure were seen in the rats that received cisplatin. Treated groups showed improvements in kidney functions, oxidative status, and histological structure, particularly in the combined treatment group., Conclusion: In the cisplatin-induced nephrotoxicity model, hUCB-MSCs could improve the functional and morphological kidney structure by modulation of oxidative and inflammatory status., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Natural products that possess antibiotic and antitumor qualities are often suspected of working through oxidative mechanisms. In this study, two quinone-based small molecules were compared. Menadione, a classic redox-cycling compound, was confirmed to generate high levels of reactive oxygen species inside Escherichia coli . It inactivated iron-cofactored enzymes and blocked growth. However, despite the substantial levels of oxidants that it produced, it was unable to generate significant DNA damage and was not lethal. Streptonigrin, in contrast, was poorer at redox cycling and did not inactivate enzymes or block growth; however, even in low doses, it damaged DNA and killed cells. Its activity required iron and oxygen, and in vitro experiments indicated that its quinone moiety transferred electrons through the adjacent iron atom to oxygen. Additionally, in vitro experiments revealed that streptonigrin was able to damage DNA without inhibition by catalase, indicating that hydrogen peroxide was not involved. We infer that streptonigrin can reduce bound oxygen directly to a ferryl species, which then oxidizes the adjacent DNA, without release of superoxide or hydrogen peroxide intermediates. This scheme allows streptonigrin to kill a bacterial cell without interference by scavenging enzymes. Moreover, its minimal redox-cycling behavior avoids alerting either the OxyR or the SoxRS systems, which otherwise would block killing. This example highlights qualities that may be important in the design of oxidative drugs. These results also cast doubt on proposals that bacteria can be killed by stressors that merely stimulate intracellular O 2 - and H 2 O 2 formation., Competing Interests: Competing interests statement:The authors declare no competing interest.

It has previously been reported that antioxidant vitamins can help reduce the risk of vision loss associated with progression to advanced age-related macular degeneration (AMD), a leading cause of visual impairment among the elderly. Nonetheless, how oxidative stress contributes to the development of choroidal neovascularization (CNV) in some AMD patients and geographic atrophy (GA) in others is poorly understood. Here, we provide evidence demonstrating that oxidative stress cooperates with hypoxia to synergistically stimulate the accumulation of hypoxia-inducible factor (HIF)-1α in the retinal pigment epithelium (RPE), resulting in increased expression of the HIF-1-dependent angiogenic mediators that promote CNV. HIF-1 inhibition blocked the expression of these angiogenic mediators and prevented CNV development in an animal model of ocular oxidative stress, demonstrating the pathological role of HIF-1 in response to oxidative stress stimulation in neovascular AMD. While human-induced pluripotent stem cell (hiPSC)-derived RPE monolayers exposed to chemical oxidants resulted in disorganization and disruption of their normal architecture, RPE cells proved remarkably resistant to oxidative stress. Conversely, equivalent doses of chemical oxidants resulted in apoptosis of hiPSC-derived retinal photoreceptors. Pharmacologic inhibition of HIF-1 in the mouse retina enhanced-while HIF-1 augmentation reduced-photoreceptor apoptosis in two mouse models for oxidative stress, consistent with a protective role for HIF-1 in photoreceptors in patients with advanced dry AMD. Collectively, these results suggest that in patients with AMD, increased expression of HIF-1α in RPE exposed to oxidative stress promotes the development of CNV, but inadequate HIF-1α expression in photoreceptors contributes to the development of GA., Competing Interests: Competing interests statement: A.S. is the cofounder of, holds equity in, is a member of the Board of Directors, and serves as the Chief Executive Officer of HIF Therapeutics, Inc. This arrangement has been reviewed and approved by the Johns Hopkins University in accordance with its conflict of interest policies.

Introduction: The flavonoid-rich fraction of Rosa damascena (FRFRD) contains antioxidant and active compounds. Therefore, this study aimed to investigate the role of FRFRD, rich in quercetin and kaempferol, in liver fibrosis induced by CCl4., Materials and Methods: The FRFRD fraction was separated and standardized by High-Performance Liquid Chromatography (HPLC) based on the levels of quercetin and kaempferol. Liver fibrosis was induced over CCl4 over 12 weeks in 30 male Wistar rats, and three concentrations of FRFRD were administered to them during the last four weeks. Subsequently, after evaluation of liver serum markers and fibrotic parameters, the relative expression of transforming growth factor-beta-1 (TGF-β 1 ), platelet-derived growth factor (PDGF), and lysyl oxidase homolog 2 (Loxl2) genes were assessed, along with the measurement of lysyl oxidase activity and oxidative markers., Results: Fibrotic markers demonstrated progressive recovery of liver damage in the treated group compared to the non-treatment group (p < 0.01). These results were accompanied by a significant decrease in the expression of TGF-β 1 , PDGF, and Loxl2 genes, as well as, a reduction in lysyl oxidase activity (p < 0.001). The antioxidant effects of the treatment were observed through a significant decrease in malondialdehyde (MDA) levels and an increase in catalase enzyme (CAT) and glutathione peroxidase (GPx) activity in the treatment group compared to the fibrotic group (p < 0.01)., Conclusion: The flavonoid-rich fraction of Rosa damascena ameliorates liver damage by affecting collagen cross-linking and lowering oxidative and inflammatory levels., Competing Interests: Declaration of competing interest The authors declare that there are no conflicts of interest, (Copyright © 2023. Published by Elsevier Inc.)

Toxic doses of formaldehyde (FA) can cause oxidative damage and impair energy metabolism. Asprosin (ASP) and subfatin (SUB) are adipokines produced by adipose tissue that help regulate energy metabolism. We investigated the effects of carvacrol (CAR), an antioxidant with hepatoprotective properties, on ASP and SUB in rats exposed to FA using immunohistochemistry and biochemistry. We used 42 male Wistar albino rats divided into six groups of seven: group 1, untreated control; group 2, FA (10 ppm FA by inhalation 8 h/day, 5 days/week); group 3, CAR-20 (20 mg/kg); group 4, CAR-40; group 5, FA (10 ppm FA by inhalation 8 h/day, 5 days/week) + CAR-20 (20 mg/kg); group 6, FA (10 ppm FA by inhalation 8 h/day, 5 days/week) + CAR-40 (40 mg/kg). Levels of ASP and SUB, and total oxidant status (TOS) and total antioxidant status (TAS) in blood and liver tissue were measured using ELISA. ASP and SUB immunoreactivity was assessed using immunohistochemistry. The number of apoptotic cells was determined using the TUNEL method. The number of apoptotic cells in group 2 was increased compared to group 1. TOS in group 2 was increased compared to group 1. The numbers of apoptotic cells and TOS in group 3 were decreased compared to group 1. TOS was decreased in group 6 compared to group 2, but TOS was increased compared to group 1. We found ASP and SUB immunoreactivity in the liver. All alterations were reversed by addition of CAR. It appears that FA disrupts energy metabolism and CAR ameliorates the destructive effects of FA when used at appropriate doses, although CAR might be harmful at high doses.

Metabolic diseases, including obesity, diabetes, and fatty liver disease, are dramatically increasing around the world. Seaweed is low in calories and rich in many active ingredients that are necessary for maintaining good health, and is expected to be effective for preventing metabolic diseases. The purpose of this study was to examine the effects of a traditional Japanese edible seaweed Hypnea asiatica ( H. asiatica ) on obesity, using a mouse model. H. asiatica was dried and powdered, mixed with a high-fat diet, and fed to male C57BL/6J mice for 13 weeks. On the last day of the experiment, blood samples were collected under anesthesia and biochemical parameters such as lipids and adipokines were measured. Liver and adipose tissue were excised, weighed, and oxidant/antioxidant parameters were measured. Some mice were perfused with a fixative solution containing formalin, and tissue specimens were prepared. A glucose tolerance test was used to assess insulin resistance. The inhibition of lipase activity was evaluated in vitro . Thirteen-week supplementation with H. asiatica suppressed body weight gain, body fat accumulation, and blood glucose levels. H. asiatica also improved fatty liver and hypercholesterolemia, and reduced the oxidant and inflammatory parameters of serum and liver. H. asiatica increased fecal triglyceride excretion and polyphenol-rich ethanol extract of H. asiatica inhibited lipase activity in vitro . These results suggest that polysaccharides and polyphenols in H. asiatica may ameliorate obesity and diabetes by inhibiting intestinal fat absorption and reducing oxidative stress and inflammation. H. asiatica may be useful in preventing metabolic diseases such as obesity, diabetes, and fatty liver.

Streptococcus pneumoniae is a commensal bacterium and invasive pathogen that causes millions of deaths worldwide. The pneumococcal vaccine offers limited protection, and the rise of antimicrobial resistance will make treatment increasingly challenging, emphasizing the need for new antipneumococcal strategies. One possibility is to target antioxidant defenses to render S. pneumoniae more susceptible to oxidants produced by the immune system. Human peroxidase enzymes will convert bacterial-derived hydrogen peroxide to hypothiocyanous acid (HOSCN) at sites of colonization and infection. Here, we used saturation transposon mutagenesis and deep sequencing to identify genes that enable S. pneumoniae to tolerate HOSCN. We identified 37 genes associated with S. pneumoniae HOSCN tolerance, including genes involved in metabolism, membrane transport, DNA repair, and oxidant detoxification. Single-gene deletion mutants of the identified antioxidant defense genes sodA , spxB , trxA, and ahpD were generated and their ability to survive HOSCN was assessed. With the exception of Δ ahpD , all deletion mutants showed significantly greater sensitivity to HOSCN, validating the result of the genome-wide screen. The activity of hypothiocyanous acid reductase or glutathione reductase, known to be important for S. pneumoniae tolerance of HOSCN, was increased in three of the mutants, highlighting the compensatory potential of antioxidant systems. Double deletion of the gene encoding glutathione reductase and sodA sensitized the bacteria significantly more than single deletion. The HOSCN defense systems identified in this study may be viable targets for novel therapeutics against this deadly pathogen. IMPORTANCE Streptococcus pneumoniae is a human pathogen that causes pneumonia, bacteremia, and meningitis. Vaccination provides protection only against a quarter of the known S. pneumoniae serotypes, and the bacterium is rapidly becoming resistant to antibiotics. As such, new treatments are required. One strategy is to sensitize the bacteria to killing by the immune system. In this study, we performed a genome-wide screen to identify genes that help this bacterium resist oxidative stress exerted by the host at sites of colonization and infection. By identifying a number of critical pneumococcal defense mechanisms, our work provides novel targets for antimicrobial therapy., Competing Interests: The authors declare no conflict of interest.

Background: Acute respiratory distress syndrome (ARDS) is a life-threatening respiratory failure syndrome with diverse etiologies characterized by increased permeability of alveolar-capillary membranes, pulmonary edema, and acute onset hypoxemia. During the ARDS acute phase, neutrophil infiltration into the alveolar space results in uncontrolled release of reactive oxygen species (ROS) and proteases, overwhelming antioxidant defenses and causing alveolar epithelial and lung endothelial injury. Objectives: To investigate the therapeutic potential of a novel recombinant human Cu-Zn-superoxide dismutase (SOD) fusion protein in protecting against ROS injury and for aerosolized SOD delivery to treat Escherichia coli induced ARDS. Methods: Fusion proteins incorporating human Cu-Zn-SOD (hSOD1), with (pep1-hSOD1-his) and without (hSOD1-his) a fused hyaluronic acid-binding peptide, were expressed in E. coli . Purified proteins were evaluated in in vitro assays with human bronchial epithelial cells and through aerosolized delivery to the lung of an E. coli -induced ARDS rat model. Results: SOD proteins exhibited high SOD activity in vitro and protected bronchial epithelial cells from oxidative damage. hSOD1-his and pep1-hSOD1-his retained SOD activity postnebulization and exhibited no adverse effects in the rat. Pep1-hSOD1-his administered through instillation or nebulization to the lung of an E. coli -induced pneumonia rat improved arterial oxygenation and lactate levels compared to vehicle after 48 hours. Static lung compliance was improved when the pep1-hSOD1-his protein was delivered by instillation. White cell infiltration to the lung was significantly reduced by aerosolized delivery of protein, and reduction of cytokine-induced neutrophil chemoattractant-1, interferon-gamma, and interleukin 6 pro-inflammatory cytokine concentrations in bronchoalveolar lavage was observed. Conclusions: Aerosol delivery of a novel recombinant modified SOD protein reduces oxidant injury and attenuates E. coli induced lung injury in rats. The results provide a strong basis for further investigation of the therapeutic potential of hSOD1 in the treatment of ARDS.

The signals that denote mammalian host environments and dictate the activation of signaling pathways in human-associated microorganisms are often unknown. The transcription regulator Rtg1/3 in the human fungal pathogen Candida albicans is a crucial determinant of host colonization and pathogenicity. Rtg1/3's activity is controlled, in part, by shuttling the regulator between the cytoplasm and nucleus of the fungus. The host signal(s) that Rtg1/3 respond(s) to, however, have remained unclear. Here we report that neutrophil-derived reactive oxygen species (ROS) direct the subcellular localization of this C. albicans transcription regulator. Upon engulfment of Candida cells by human or mouse neutrophils, the regulator shuttles to the fungal nucleus. Using genetic and chemical approaches to disrupt the neutrophils' oxidative burst, we establish that the oxidants produced by the NOX2 complex-but not the oxidants generated by myeloperoxidase-trigger Rtg1/3's migration to the nucleus. Furthermore, screening a collection of C. albicans kinase deletion mutants, we implicate the MKC1 signaling pathway in the ROS-dependent regulation of Rtg1/3 in this fungus. Finally, we show that Rtg1/3 contributes to C. albicans virulence in the nematode Caenorhabditis elegans in an ROS-dependent manner as the rtg1 and rtg3 mutants display virulence defects in wild-type but not in ROS deficient worms. Our findings establish NOX2-derived ROS as a key signal that directs the activity of the pleiotropic fungal regulator Rtg1/3., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Oneissi et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Myeloperoxidase (MPO) is released by neutrophils in inflamed tissues. MPO oxidizes chloride, bromide, and thiocyanate to produce hypochlorous acid (HOCl), hypobromous acid (HOBr), and hypothiocyanous acid (HOSCN), respectively. These oxidants are toxic to pathogens, but may also react with host cells to elicit biological activity and potential toxicity. In cystic fibrosis (CF) and related diseases, increased neutrophil inflammation leads to increased airway MPO and airway epithelial cell (AEC) exposure to its oxidants. In this study, we investigated how equal dose-rate exposures of MPO-derived oxidants differentially impact the metabolome of human AECs (BEAS-2B cells). We utilized enzymatic oxidant production with rate-limiting glucose oxidase (GOX) coupled to MPO, and chloride, bromide (Br - ), or thiocyanate (SCN - ) as substrates. AECs exposed to GOX/MPO/SCN - (favoring HOSCN) were viable after 24 h, while exposure to GOX/MPO (favoring HOCl) or GOX/MPO/Br - (favoring HOBr) developed cytotoxicity after 6 h. Cell glutathione and peroxiredoxin-3 oxidation were insufficient to explain these differences. However, untargeted metabolomics revealed GOX/MPO and GOX/MPO/Br - diverged significantly from GOX/MPO/SCN - for dozens of metabolites. We noted methionine sulfoxide and dehydromethionine were significantly increased in GOX/MPO- or GOX/MPO/Br - -treated cells, and analyzed them as potential biomarkers of lung damage in bronchoalveolar lavage fluid from 5-year-olds with CF (n = 27). Both metabolites were associated with increasing bronchiectasis, neutrophils, and MPO activity. This suggests MPO production of HOCl and/or HOBr may contribute to inflammatory lung damage in early CF. In summary, our in vitro model enabled unbiased identification of exposure-specific metabolite products which may serve as biomarkers of lung damage in vivo. Continued research with this exposure model may yield additional oxidant-specific biomarkers and reveal explicit mechanisms of oxidant byproduct formation and cellular redox signaling., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Diabetes mellitus currently affects ∼10% of the population worldwide, with Type 2 predominating, and this incidence is increasing steadily. Both Type 1 and 2 are complex diseases, involving β-cell death and chronic inflammation, but the pathways involved are unresolved. Chronic inflammation is characterized by increased oxidant formation, with this inducing protein modification, altered function and immunogenicity. Amylin, a peptide hormone co-secreted with insulin by β-cells, has attracted considerable interest for its amyloidogenic properties, however, the effects that oxidants have on amylin aggregation and function are poorly understood. Amylin was exposed in vitro to hypochlorous acid, hydrogen peroxide and peroxynitrous acid/peroxynitrite to investigate the formation of post-translational oxidative modifications (oxPTMs, via mass spectrometry) and fibril formation (via transmission electron microscopy). Amylin free acid (AFA) was also examined to investigate the role of the C-terminal amide in amylin. Oxidant exposure led to changes in aggregate morphology and abundance of oxPTMs in a concentration-dependent manner. The toxicity and immunogenic potential of oxidant-modified amylin or AFA on pancreatic islet cells (INS-1E), human monocyte cell line (THP-1) and monocyte-derived dendritic cells (moDCs) were examined using metabolic activity and cytokine assays, and flow cytometry. No significant changes in vitality or viability were detected, but exposure to oxidant-modified amylin or AFA resulted in altered immunogenicity when compared to the native proteins. THP-1 and moDCs show altered expression of activation markers and changes in cytokine secretion. Furthermore, oxidant-treated amylin and AFA promoted maturation of THP-1 and pre-mature moDCs, as determined by changes in size, and maturation markers., Competing Interests: Declaration of competing interest MJD declares consultancy contracts with Novo Nordisk A/S. This funder had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results. The other authors declare no conflict of interest., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Acute respiratory distress syndrome (ARDS) is a leading cause of respiratory failure and death in patients in the intensive care unit. Experimentally, acute lung injury resolution depends on the repair of mitochondrial oxidant damage by the mitochondrial quality control (MQC) pathways, mitochondrial biogenesis, and mitophagy, but nothing is known about this in the human lung. In a case-control autopsy study, we compared the lungs of subjects dying of ARDS (n = 8; cases) and age-/gender-matched subjects dying of nonpulmonary causes (n = 7; controls). Slides were examined by light microscopy and immunofluorescence confocal microscopy, randomly probing for co-localization of citrate synthase with markers of oxidant stress, mitochondrial DNA damage, mitophagy, and mitochondrial biogenesis. ARDS lungs showed diffuse alveolar damage with edema, hyaline membranes, and neutrophils. Compared with controls, a high degree of mitochondrial oxidant damage was seen in type 2 epithelial (AT2) cells and alveolar macrophages by 8-hydroxydeoxyguanosine and malondialdehyde co-staining with citrate synthase. In ARDS, antioxidant protein heme oxygenase-1 and DNA repair enzyme N-glycosylase/DNA lyase (Ogg1) were found in alveolar macrophages but not in AT2 cells. Moreover, MAP1 light chain-3 (LC3) and serine/threonine-protein kinase (Pink1) staining were absent in AT2 cells, suggesting a mitophagy failure. Nuclear respiratory factor-1 staining was missing in the alveolar region, suggesting impaired mitochondrial biogenesis. Widespread hyperproliferation of AT2 cells in ARDS could suggest defective differentiation into type 1 cells. ARDS lungs show profuse mitochondrial oxidant DNA damage but little evidence of MQC activity in AT2 epithelium. Because these pathways are important for acute lung injury resolution, our findings support MQC as a novel pharmacologic target for ARDS resolution., (Copyright © 2023 United States & Canadian Academy of Pathology. Published by Elsevier Inc. All rights reserved.)