Female rats present higher oxidative damage and inflammation during goitrogenesis.

Analysis of the Metabolic State of Residual Multiple Myeloma Plasma Cells Reveals a Proliferative Subset with Glycolytic Gene Expression

Increased Reactive Oxygen Species and the B-Cell Receptor in Chronic Lymphocytic Leukemia Signaling

In donkeys, the use of frozen-thawed sperm for artificial insemination (AI) leads to low fertility rates. Furthermore, donkey sperm produce a large amount of reactive oxygen species (ROS), and post-AI inflammation induces the formation of neutrophil extracellular traps (NETosis), which further generates many more ROS. These high ROS levels may induce lipid peroxidation in the sperm plasma membrane, thus affecting its integrity. Enzymatic and non-enzymatic antioxidants, mainly found in the seminal plasma (SP), are responsible for maintaining the redox balance. However, this fluid is removed prior to cryopreservation, thereby exposing sperm cells to further oxidative stress. The exogenous addition of antioxidants to the freezing medium can reduce the detrimental effects caused by ROS generation. Therefore, the aim of this study was to evaluate how the addition of different reduced glutathione (GSH) concentrations (control, 2 mM, 4 mM, 6 mM, 8 mM, and 10 mM) to fresh sperm affect their cryotolerance. Total and progressive motility, kinematic parameters and motile sperm subpopulations were significantly (p < 0.05) different from the control in treatments containing 8 mM and 10 mM GSH, but not at lower concentrations. Plasma and acrosome membrane integrity, mitochondrial membrane potential (MMP) and intracellular superoxide levels (O2−) were not affected (p > 0.05) by any GSH concentration. Interestingly, however, the addition of 8 mM or 10 mM GSH reduced (p < 0.05) the percentages of viable sperm with high overall ROS levels compared to the control. In conclusion, frozen-thawed donkey sperm are able to tolerate high GSH concentrations, which differs from what has been observed in other species. This antioxidant capacity suggests that ROS could be important during post-AI and that the impact of using exogenous antioxidants like GSH to improve the sperm resilience to freeze-thawing is limited in this species.

Overexpression of c-Myc results in transformation and multiple other phenotypes, and is accompanied by the deregulation of a large number of target genes. We previously demonstrated that peroxiredoxin 1 (Prdx1), a scavenger of reactive oxygen species (ROS), interacts with a region of the c-Myc transcriptional regulatory domain that is essential for transformation. This results either in the suppression or enhancement of some c-Myc functions and in the altered expression of select target genes. Most notably, c-Myc-mediated transformation is inhibited, implying a tumor suppressor role for Prdx1. Consistent with this, prdx1-/- mice develop age-dependent hemolytic anemias and/or malignancies. We now show that erythrocytes and embryonic fibroblasts from these animals contain higher levels of ROS, and that the latter cells show evidence of c-Myc activation, including the ability to be transformed by a ras oncogene alone. In contrast, other primary cells from prdx1-/- mice do not have elevated ROS, but nonetheless show increased oxidative DNA damage. This apparent paradox can be explained by the fact that ROS localize primarily to the cytoplasm of prdx1+/+ cells, whereas in prdx1-/- cells, much higher levels of nuclear ROS are seen. We suggest that increased DNA damage and tumor susceptibility in prdx1-/- animals results from this shift in intracellular ROS. prdx1-/- mice should be useful in studying the role of oxidative DNA damage in the causation of cancer and its prevention by antioxidants. They should also help in studying the relationship between oncogenes such as c-Myc and DNA damage.

Oxidative Stress Impairs Cell Death by Repressing the Nuclease Activity of Mitochondrial Endonuclease G.

Baicalein is a flavonoid inhibitor of 12-lipoxygenase. Here, we investigated its effect on hydrogen peroxide-induced damage to NG108-15 cells. Hydrogen peroxide activated the mitochondrial apoptotic pathway, decreased Nrf2 expression, increased reactive oxygen species (ROS) levels, reduced viability, and increased cell death after 2-24 h treatment of NG108-15 cells. Co-treatment with hydrogen peroxide and baicalein completely suppressed the activation of mitochondrial apoptotic pathway by upregulating Nrf2 expression and reducing ROS stress and partially inhibited the effects on cell viability and cell death. Silencing of 12-lipoxygenase had a similar protective effect to baicalein on hydrogen peroxide-induced damage by blocking the hydrogen peroxide-induced decrease in Nrf2 expression and increase in ROS levels. Neither protective effect was altered by addition of 12-hydroxyeicosatetraenoic acid, the product of 12-lipoxygenase, suggesting that hydrogen peroxide induced damage via 12-lipoxygenase by another, as yet unknown, mechanism, rather than activating it. Co-treatment of cells with hydrogen peroxide and N-acetylcysteine or the Nrf2 inducer sulforaphane reduced hydrogen peroxide-induced damage in a similar fashion to baicalein, while the Nrf2 inhibitor retinoic acid blocked the protective effect of baicalein. Silencing Nrf2 also inhibited the protective effects of baicalein, sulforaphane, and N-acetylcysteine and resulted in high ROS levels, suggesting ROS elimination was mediated by Nrf2. Taken together our results suggest that baicalein protects cells from hydrogen peroxide-induced activation of the mitochondrial apoptotic pathway by upregulating Nrf2 and inhibiting 12-lipoxygenase to block the increase in ROS levels. Hydrogen peroxide also activates a second mitochondrial dysfunction independent death pathway which is resistant to baicalein.

Using specific stains and confocal microscope imaging, the patterns of mitochondrial distribution, mitochondrial inner membrane potential and reactive oxygen species (ROS) levels during bovine oocyte maturation were determined in the presence or absence of physiological concentrations of linoleic acid (LA; 100µM) or α-linolenic acid (ALA; 50µM). Mitochondrial distribution in control oocytes at 0h was mainly peripheral and changed to a diffused pattern after 1h of culture; this was maintained up to 24h. Mitochondrial clusters were observed during the early hours of maturation (1-4h); the majority of these were arranged in perinuclear fashion. LA supplementation resulted in: (1) delayed redistribution of the mitochondria from a peripheral to a diffuse pattern and a decreased percentages of oocytes showing perinuclear mitochondrial clusters, (2) decreased mitochondrial inner membrane potential at 1 and 24h compared with the control and (3) higher ROS levels, associated with a lower nuclear maturation rate. In contrast, ALA supplementation had no effect on mitochondrial distribution and activity and decreased ROS levels compared with the control; this was associated with an increased nuclear maturation rate. In conclusion, LA induced alterations in mitochondrial distribution and activity as well as increasing ROS levels, which mediate, at least in part, the inhibitory effect on oocyte maturation.

Purpose: Oxidative stress, which refers to the biological damage caused by free radicals produced in excess of innate antioxidant defenses, is indicated in the ocular cancer retinoblastoma (RB). Here we have analysed the differential expression of oxidative stress responsive genes in oxidant-induced RB cells, and in RB tumor tissues.Methods: The study included cultured RB cells, and four RB tumor tissues. The reactive oxygen species (ROS) levels in Y79 cells and the RB tumor induced by hydrogen peroxide were quantified by Dichlorofluorescein (DCF) fluorescence assay. We then analysed the gene expression profile of cultured RB cells induced with hydrogen peroxide (400 µM H2O2 for 8 h) by microarray analysis, and the expression of select genes were validated in Y79 cells and RB tumor tissues by real-time PCR analysis.Results: The oxidant-induced RB tumors showed an average increase in ROS levels of 44-fold compared to induced non-neoplastic donor retina. H2O2-induced RB cell line showed a 3-fold increase in ROS levels. Microarray analysis on RB cell line induced with H2O2 showed differentially regulated genes involved in cellular processes such as: oxidative stress, angiogenesis, lipid metabolism, cell proliferation, and cell signaling pathways. Several up-regulated genes such as SOD, GPX, CAT, CDC25A, CREBBP, JUN, MMP-2, iNOS, CRYAA, RXRA, ACACB and HMGCR were validated by real-time PCR. These results corroborated with the gene expression analysis in RB tumor tissues. Relating the antioxidant gene expression with the clinico-pathologic features of the tumor tissues, we found that the tumor with invasion of choroid, optic nerve and retinal pigment epithelium, had relatively higher ROS levels and minimal antioxidant gene expression, when compared with the tumor with only choroidal invasion.Conclusions: The study suggests active involvement of redox signaling pathways in the pathogenesis of RB. Consideration of oxidative stress components in the clinical management of RB patients is emphasized.

Biochemical, inflammatory, and immunological markers may provide insight into systemic alterations in individuals with or without thyroid dysfunction. This study compared a comprehensive biomarker profile between adults without thyroid disease and those with thyroid dysfunction, and examined gender differences, correlations, and predictive markers. In this case-control study, 120 adults without thyroid disease (cases) and 120 adults with thyroid dysfunction (controls) were enrolled. Demographics, anthropometry, and biochemical parameters-including thyroid hormones, micronutrients, lipid profiles, inflammatory markers, and Epstein-Barr virus PCR-were assessed. Between-group differences were analyzed using the Mann-Whitney U test and the χ² test. Spearman's rho was used to determine correlations, and multivariable linear regression was used to identify predictors of group status. Cases and controls were similar in age (60 ± 12 vs. 60 ± 10.5 years) and body mass index (BMI) (24.85 ± 3.69 vs. 25.58 ± 3.39kg/m²). Compared with controls, cases had significantly lower zinc (97.98 vs. 143.03µg/dL), vitamin D₃ (88.24 vs. 152.76 nmol/L), and HDL (88.75 vs. 152.25mg/dL), but higher serum cholesterol (159.56 vs. 81.44mg/dL), triglycerides (TG) (171.08 vs. 69.92mg/dL), LDL (136.06 vs. 104.94mg/dL), and CRP (140.05 vs. 100.95mg/L) (all p < 0.001). In controls, TSH correlated positively with the Urea Breath Test (UBT) (rho = 0.334) and negatively with LDL (rho=-0.255). In cases, T3 showed a strong correlation with UBT (rho = 0.625) and a moderate correlation with PCR-EBV (rho = 0.417), while TSH correlated with UBT (rho = 0.434) and CRP (rho = 0.264). Regression analysis identified serum TG (β = 76.806, R²=0.462) and cholesterol (β = 64.834, R²=0.337) as the strongest predictors of case-control status. Adults without thyroid disease exhibit distinct biochemical and inflammatory profiles compared with those with thyroid dysfunction, highlighting lipid, micronutrient, and inflammatory markers as potential targets for risk stratification and clinical monitoring. Not applicable.

One tough question induced by the hypoxia in cancer tissue is resistance to anticancer drugs basing on the reactive oxygen species (ROS) mechanism. Furthermore, the hypoxic regions locate in the center of tumor where tumor cells are easily residual and survival due to the poor drug-delivery efficiency even with nanocarriers. In this paper, these problems were well addressed through the rational combination of the enhanced penetration, self-inducing high level of intracellular ROS, and synchronously pH-sensitive drug release, realized by a simple structural and accessible copolymer, poly(poly(ethylene glycol) methyl ether methacrylate-co-(2-methylpropenoic acid-glycerol-cinnamaldehyde)) (PgEMC). For one thing, PgEMC could self-assemble into stable nanoparticles with PEG shell and optimizing diameters of 60 nm to simultaneously facilitate long blood circulation and deep tumor penetration. Second, cinnamylaldehyde moieties could detach from PgEMC NPs in intracellular acidic environment and trigger high level of ROS to allay the doxorubicin (DOX) resistance induced by hypoxia in solid malignancies. Furthermore, the DOX payload in PgEMC NPs could be synchronously released with the intracellular disassembly of PgEMC NPs due to the detaching of cinnamylaldehyde moieties. In 4T1 cells treated with PgEMC/DOX NPs, remarkable elevation of ROS level and enhanced DOX sensitivity in hypoxia environment were observed in in vitro studies. The results of tumor spheroid penetration indicated that 60 nm sized DOX-loaded PgEMC NPs (PgEMC60/DOX) could distribute into deep site of tumor at a high intensity. In vivo studies using a 4T1 breast tumor model, PgEMC60/DOX NPs, showed significant inhibition over 95.4% of the tumor growth. These results reveal that integrating optimizing size, self-inducing ROS, and pH-sensitive drug release into one small-sized nanoparticle can efficiently overcome the poor tumor penetration and hypoxia-induced chemotherapy resistance.

PurposeThe aim of this study is to investigate the relationships among reactive oxygen species (ROS) elevation, histone transition, and seminal cytokine concentrations.MethodsTotal levels of ROS in semen samples from 6560 men were measured. From this sample, 118 cases with high ROS and 106 controls were recruited. Basic semen parameters and histone-to-protamine ratios were analyzed, 400 semen cytokine and receptor alterations were assayed by protein chip, and finally 18 cytokines were validated in each sample using a Bio-Plex Cytokine assay.ResultsThe results showed that the seminal ROS concentration was associated with abnormalities in the sperm histone transition. Compared with controls, 93 cytokines had significant alterations in the high ROS cases, with 14 of them further verified in individual samples. The concentrations of CXCL5, CXCL16, CXCL8, IL-1b, IL-10, CSF3, CCL3, and TNF-α were significantly correlated with the histone transition ratio. In addition, IL-16 showed significantly different concentrations in controls, normal semen with high ROS levels, and abnormal semen with high ROS levels.ConclusionsSemen ROS are associated with abnormalities in sperm histone transition. CXCL5, CXCL8, IL-16, CCL8, CCL22, CCL20, CXCL16, IL-1B, IL-6, IL-7, IL-10, CSF3, CCL3, CCL4, and TNF-α all have elevated concentrations in semen with high ROS levels. These data might help to explain the mechanisms behind the increase in the levels of ROS and seminal cytokines and their relationship with defective spermatogenesis.

OBJECTIVE—Recent studies identified accumulation of reactive oxygen species (ROS) as a common pathway causing insulin resistance. However, whether and how the reduction of ROS levels improves insulin resistance remains to be elucidated. The present study was designed to define this mechanism.RESEARCH DESIGN AND METHODS—We investigated the effect of overexpression of superoxide dismutase (SOD)1 in liver of obese diabetic model (db/db) mice by adenoviral injection.RESULTS—db/db mice had high ROS levels in liver. Overexpression of SOD1 in liver of db/db mice reduced hepatic ROS and blood glucose level. These changes were accompanied by improvement in insulin resistance and reduction of hepatic gene expression of phosphoenol-pyruvate carboxykinase and peroxisome proliferator–activated receptor γ coactivator-1α (PGC-1α), which is the main regulator of gluconeogenic genes. The inhibition of hepatic insulin resistance was accompanied by attenuation of phosphorylation of cAMP-responsive element-binding protein (CREB), which is a main regulator of PGC-1α expression, and attenuation of Jun NH2-terminal kinase (JNK) phosphorylation. Simultaneously, overexpression of SOD1 in db/db mice enhanced the inactivation of forkhead box class O1, another regulator of PGC-1α expression, without the changes of insulin-induced Akt phosphorylation in liver. In hepatocyte cell lines, ROS induced phosphorylation of JNK and CREB, and the latter, together with PGC-1α expression, was inhibited by a JNK inhibitor.CONCLUSIONS—Our results indicate that the reduction of ROS is a potential therapeutic target of liver insulin resistance, at least partly by the reduced expression of PGC-1α.

High levels of reactive oxygen species (ROS) can damage key genetic and cellular processes, but low levels are required for redox cell signaling. Accumulation of ROS is directly linked to genetic damage in aging tissues and implicated in delayed wound healing. In this study, we examined the effect of age and oxygen tension in primary human skin keratinocytes using cell migration and level of ROS as the affected functions. Baseline levels of ROS were higher in young keratinocytes (from 22–35 year old patients) grown at 21% oxygen as compared to 4% oxygen. However, we found the opposite with older keratinocytes (from 78–90 year old patients)– higher levels of ROS at 4% oxygen than at 21% oxygen. Both young and old keratinocytes showed decreased rates of cell migration in response to increased ROS. The in vitro scratch assay demonstrated that cells located at the edge of the scratch greatly increased their ROS levels immediately upon scratching and had declined to near baseline after 24 hours. While young cells showed higher levels of ROS at scratch margins than old cells did, the young cells were less effective at removing the ROS. Furthermore the old keratinocytes were most successful in reducing their ROS levels at the scratch margin when grown at 21% oxygen. Recovery of the cells from increased levels of ROS at their scratch margins correlated with their migration rates. These findings indicate that keratinocytes show an age‐related differential response to both oxygen tension and ROS levels (Supported by a McCord Foundation Research Grant, RO1AR053619, and R21AR053936).

Molecularly Classified High Risk Myeloma Cells with Low ROS Levels and High Nestin Expression Have High Proliferation Index and GEP80 Score,

Compelling evidence shows a strong correlation between accumulation of neurotoxic β-amyloid (Aβ) peptides and oxidative stress in the brains of patients afflicted with Alzheimer disease (AD). One hypothesis for this correlation involves the direct and harmful interaction of aggregated Aβ peptides with enzymes responsible for maintaining normal, cellular levels of reactive oxygen species (ROS). Identification of specific, destructive interactions of Aβ peptides with cellular anti-oxidant enzymes would represent an important step toward understanding the pathogenicity of Aβ peptides in AD. This report demonstrates that exposure of human neuroblastoma cells to cytotoxic preparations of aggregated Aβ peptides results in significant intracellular co-localization of Aβ with catalase, an anti-oxidant enzyme responsible for catalyzing the degradation of the ROS intermediate hydrogen peroxide (H(2)O(2)). These catalase-Aβ interactions deactivate catalase, resulting in increased cellular levels of H(2)O(2). Furthermore, small molecule inhibitors of catalase-amyloid interactions protect the hydrogen peroxide-degrading activity of catalase in Aβ-rich environments, leading to reduction of the co-localization of catalase and Aβ in cells, inhibition of Aβ-induced increases in cellular levels of H(2)O(2), and reduction of the toxicity of Aβ peptides. These studies, thus, provide evidence for the important role of intracellular catalase-amyloid interactions in Aβ-induced oxidative stress and propose a novel molecular strategy to inhibit such harmful interactions in AD.