State Of Chronic Oxidative Stress Research Articles

Ozone Environmental Pollution: Relationship between the Intestine and Neurodegenerative Diseases.

Repeated exposure to environmental ozone causes a chronic state of oxidative stress. This state is present in chronic degenerative diseases and induces a loss of control of the inflammatory response. Redox system dysfunction and failures in control of inflammatory responses are involved in a vicious circle that maintains and increases the degenerative process. The intestine also responds to secondary reactive species formed by exposure to ozone doses, generating noxious stimuli that increase degenerative damage. This review aims to elucidate how environmental pollution, mainly by ozone, induces a state of chronic oxidative stress with the loss of regulation of the inflammatory response, both in the intestine and in the brain, where the functionality of both structures is altered and plays a determining role in some neurodegenerative and chronic degenerative diseases. For this purpose, we searched for information on sites such as the Cochrane Library Database, PubMed, Scopus, and Medscape. Reviewing the data published, we can conclude that environmental pollutants are a severe health problem. Ozone pollution has different pathways of action, both molecular and systemic, and participates in neurodegenerative diseases such as Parkinson's and Alzheimer's disease as well in bowel diseases as Inflammatory Bowel Disease, Crohn's Disease, and Irritable Bowel Syndrome.

Reduction of plasma homocysteine and 8-iso-prostaglandin F2α concentrations in hypercholesterolemic rats treated with clove essential oil

Cardiovascular diseases are associated with a chronic state of oxidative stress that is induced by fat accumulation. The current study aims to investigate the potential effects of oral consumption of clove essential oil on the lipid profile, oxidative stress markers and antioxidant enzyme activity, in hypercholesterolemic rats. Two groups of Wistar rats were fed a cholesterol-enriched diet supplemented (CEO) or not (HC) with 4 mg/kg b.w of clove essential oil, for six weeks. A control group (C) was fed a standard diet during the experiment. After six weeks, in clove essential oil treated as compared to untreated hypercholesterolemic rats, daily supplementation with essential oil attenuated serum levels of total cholesterol (-57%), LDL-cholesterol (-28%), triacyglycerols (-26%), total homocysteine (-17%) and 8-Iso-prostaglandin F2α (-39%). Furthermore, TBARS values were decreased in LDL (-37%), liver (-45%) and adipose tissue (-34%). Interestingly, clove essential oil increased respectively thioredoxin reductase activity in liver and adipose tissue by +67% and +66%, respectively. These results demonstrate the therapeutic potential of clove essential oil to reduce dyslipidemia, oxidative stress by increasing the thioredoxin reductase activity.

Ozone Pollution, Oxidative Stress, Regulatory T Cells and Antioxidants.

Ozone pollution, is a serious health problem worldwide. Repeated exposure to low ozone doses causes a loss of regulation of the oxidation–reduction systems, and also induces a chronic state of oxidative stress. This fact is of special importance for the regulation of different systems including the immune system and the inflammatory response. In addition, the oxidation–reduction balance modulates the homeostasis of these and other complex systems such as metabolism, survival capacity, cell renewal, and brain repair, etc. Likewise, it has been widely demonstrated that in chronic degenerative diseases, an alteration in the oxide-reduction balance is present, and this alteration causes a chronic loss in the regulation of the immune response and the inflammatory process. This is because reactive oxygen species disrupt different signaling pathways. Such pathways are related to the role of regulatory T cells (Treg) in inflammation. This causes an increase in chronic deterioration in the degenerative disease over time. The objective of this review was to study the relationship between environmental ozone pollution, the chronic state of oxidative stress and its effect on Treg cells, which causes the loss of regulation in the inflammatory response as well as the role played by antioxidant systems in various pathologies.

P982: MYELOID-DERIVED SUPPRESSOR CELLS: CHARACTERIZATION IN PERIPHERAL BLOOD AND SPLEEN TISSUE OF PATIENTS WITH MYELOFIBROSIS

Background: Myeloid-derived suppressor cells (MDSCs) are immature myeloid cells that accumulate in patients with malignancies, sepsis or chronic inflammation. Features of MDSCs are: myeloid origin, immature state, strong ability to reduce cytotoxic functions of T/NK cells, potential to differentiate into endothelial cells favoring neoangiogenesis. MDSCs are conventionally divided into polymorphonuclear (PMN)- and monocytic (M)-MDSCs. PMN-MDSCs are characterized by the production of reactive oxygen species and arginase-1 (Arg-1), that directly inhibits T-cell proliferation and activation, while M-MDSCs predominantly express inducible nitric oxide synthase and produce nitric oxide. Myelofibrosis (MF) is characterized by clonal expansion of a hematopoietic stem cell, extramedullary hematopoiesis, bone marrow (BM) fibrosis, splenomegaly, BM and splenic neoangiogenesis and an acquired mutation of JAK2, CALR or MPL gene. Inflammation plays a relevant role in MF pathogenesis, as proven by high levels of inflammatory cytokines with prognostic significance and by a state of chronic oxidative stress. Aims: To characterize circulating MDSCs and assess their localization and quantification in spleen samples from MF patients. Methods: Peripheral blood mononuclear cells (PBMCs) from MF patients (n=15) and healthy subjects (HDs; n=10) were stained with surface antibodies (CD11b, CD14, HLA-DR, CD15, CD33) and intracellular Arg-1 and analysed by flow cytometry. Formalin-fixed paraffin-embedded samples of spleen specimens (MF n=41; HDs n=20) were immunostained with anti-Arg-1 antibody using the automated platform Dako Omnis Envision Flex with the EnVision FLEX, High pH, HRP Rabbit/Mouse High pH [GV800] revelation kit. Results: In PBMCs, the percentage of PMN-MDSCs (identified as CD11b+HLA-DRlow/-CD33dimCD14-CD15+) was higher (p=0) in patients (median 9.2, range 1-59) than HDs (median 1.7, range 0.1-4.5) and directly correlated with the allelic burden (R=0.86, p=0.006) in JAK2-mutated patients. Patients in pre-fibrotic phase (n=7) had levels of PMN-MDSCs (median 9.2, range 1-31.4) comparable to HDs; on the contrary, the percentage of PMN-MDSCs in patients with BM fibrosis (n=8; median 13.3, range 3.5-59) was higher (p=0) than in HDs. All PMN-MDSCs were Arg-1+, both in MF and in HDs. Gene expression studies are ongoing. The expression of the CXCR4 receptor on PMN-MDSCs was lower, although not statistically significant, in MF (median 1.1, range 0-9.3) than in HDs (median 2.5, range 0-11.9). PMN-MDSCs in MF inversely correlated (R=-0.89 p=0.007) with CXCR4 expressed on CD34+ cells. The percentage of M-MDSCs (identified as CD11b+HLA-DRlow/-CD33hiCD14+CD15-) was higher (p=0.015) in HDs (median 0.49, range 0.25-1.5) than in MF (median 0.29, range 0.02-0.76). Immunohistochemical staining of spleen sections showed a higher (p=0) percentage of Arg-1+ cells (calculated on total spleen cells) in MF (median 20, range 5-60) than in HDs (median 7, range 2-20); Arg-1+ cells were present in all patients and predominantly located in the red pulp in MF patients, while in HDs were detected in the white pulp. Summary/Conclusion: The identification of MDSCs and their relation with the mutational status suggest a new mechanism of MF pathogenesis, either related to the chronic inflammatory status of patients or as players of the neoangiogenesis that characterizes the disease. Moreover, the low expression of CXCR4 on PMN-MDSCs could be, on one hand, related to the low expression of this receptor on CD34+ cells or suggest a MF-specific recruitment mechanism, other than that observed in solid tumors.

The Role of the Axis TGF-β1/Mir-382-5p/SOD2 in the Induction of Oxidative Stress in CD34+ Hematopoietic Stem/Progenitor Cells from Primary Myelofibrosis

Philadelphia negative chronic myeloproliferative neoplasms (MPNs) are acquired clonal disorders of hematopoietic stem cell. Among MPNs, Primary Myelofibrosis (PMF) carries the worst prognosis with severe clinical hallmarks, including bone marrow failure, extramedullary hematopoiesis and splenomegaly. MNPs are intimately associated with high level of pro-inflammatory cytokines (i.e., transforming growth factor-β1, TGF-β1) and excess generation of Reactive Oxygen Species (ROS) that are, at least in part, responsible for disease progression. Recently, extensive studies have provided unequivocal evidences that differentially expressed microRNAs (miRNAs) may play a role in MNP's pathogenesis. The miRNA expression profile performed on CD34+ hematopoietic stem/progenitor cells (HSPCs) obtained from PMF patients (R. Norfo et al., Blood, 2014), allowed us to identify miR-382-5p as up-regulated miRNA. Overexpression experiments using miRNA mimic in normal CD34+ cells have demonstrated its central role in granulocyte fate decision (R. Zini et al., Stem Cell and Dev, 2016).In this study, to establish the role of miR-382-5p in PMF pathogenesis, we performed a gene expression profile (GEP) in cord blood CD34+ cells overexpressing miR-382-5p by means of Affymetrix HG-U219 microarray platform. We identified 75 downregulated genes in miR-382-5p overexpressing cells compared to controls. Then, we pointed out 6 genes downregulated both upon miRNA overexpression and in PMF CD34+ cells compared to healthy donor. Among them, we selected the anti-oxidant superoxide dismutase 2 (SOD2), depicted as the most favorable predicted target according to different prediction algorithms such as Target Scan, miRNA.org and miRDB. Luciferase reporter assay, performed in K562 cells, confirmed SOD2 as a real miR-382-5p target. Then, we showed that miR-382-5p overexpression leads to SOD2 down-regulation both at mRNA and protein level and reduced SOD activity in CB CD34+ cells. Furthermore, we demonstrated that miR-382-5p induced ROS accumulation as compared to control (42.1±1.5% vs 31.8±1.7%, p<0.05), increased DNA damages (measured by 8-OHdG level) and enhanced CD34+ cells proliferation. Afterwards, to confirm that miR-382-5p play a role in PMF pathogenesis, we performed inhibition experiments in PMF CD34+ cells using miR-382-5p antagomiR. Our data revealed that miR-382-5p silencing restored SOD2 expression and activity, induced ROS disposal (50.1±3.8% vs 62.1±3.2%, p<0,05), decreased DNA oxidation and impaired cell proliferation.Then, to verify whether miR-382-5p could be related to chronic inflammatory state typical of PMF, we treated CD34+ cells with the pro-inflammatory cytokine TGF-β1, already described up-regulated cytokine in PMF serum, and involved in PMF pathogenesis (A. Agarwal et al., Stem Cell Investig, 2016). We showed that TGF-β1 treatment induced miR-382-5p expression, which in turn suppresses SOD2 both at mRNA and protein level and impaired its activity leading to ROS over-production (54.7±1.1% vs 36.7±1.3% 24h after TGF-β1 treatment, p<0.05, 72.4±0.7% vs 51.9±2.0% 48h after TGF-β1 treatment, p<0.01). Finally, to further confirm the involvement of the axis TGF-β1/miR-382-5p/SOD2 in the induction of oxidative stress, we inhibited TGF-β1 pathway in PMF CD34+ cells using 500nM of Galunisertib, a specific TGF-β-receptor I kinase inhibitor, already demonstrated to be capable of attenuating the fibrotic phenotype in MPN mouse model (L.Yue et al., Blood, 2015). As a result, TGF-β1 inhibition reduced miR-382-5p expression, increasing SOD activity and its expression both at mRNA and protein level. Of note, the TGF-β-receptor I kinase inhibitor significantly reduced ROS accumulation 24h after Galunisertib treatment, in PMF CD34+ cells as compared to untreated sample (33.1±7.2% vs 45.3±6.3%, p< 0.05).Taken together, our data showed that miR-382-5p enhanced oxidative stress and oxidative-DNA-damages in normal and PMF CD34+ cells through the modulation of its target SOD2. We showed for the first time that the axis TGF-β1/miR-382-5p/SOD2 is responsible, at least in part, for ROS over-production that contribute to a state of chronic oxidative stress and inflammation, which are typical features of PMF. Furthermore, our results suggested that Galunisertib might be an effective therapy to reduce abnormal oxidative stress sustained by increased level of TGF-β1 in PMF CD34+ cells. [Display omitted] DisclosuresVannucchi:Novartis: Honoraria, Speakers Bureau; Shire: Speakers Bureau.

N-Acetylcysteine Reverses Antiretroviral-Mediated Microglial Activation by Attenuating Autophagy-Lysosomal Dysfunction.

Successful suppression of viral replication by combined antiretroviral therapy (cART) in HIV-1 infected individuals is paradoxically also accompanied by an increased prevalence of HIV-associated neurocognitive disorders (HAND) in these individuals. HAND is characterized by a state of chronic oxidative stress and inflammation. Microglia are extremely sensitive to a plethora of stimuli, including viral proteins and cART. The current study aimed to assess the effects of cART-mediated oxidative stress on the induction of inflammatory responses in microglia. In the present study, we chose a combination of three commonly used antiretroviral drugs—tenofovir disoproxil fumarate, emtricitabine, and dolutegravir. We demonstrated that exposure of microglia to the chosen cART cocktail induced generation of reactive oxygen species, subsequently leading to lysosomal dysfunction and dysregulated autophagy, ultimately resulting in the activation of microglia. Intriguingly, the potent antioxidant, N-acetylcysteine, reversed the damaging effects of cART. These in vitro findings were further corroborated in vivo wherein cART-treated HIV transgenic (Tg) rats demonstrated increased microglial activation, exaggerated lysosome impairment, and dysregulated autophagy in the prefrontal cortices compared with HIV Tg rats not exposed to cART. Similar to in vitro findings, the treatment of HIV Tg rats with N-acetylcysteine also mitigated the deleterious effects of cART. Taken together, our findings suggest that oxidative stress-mediated lysosomal dysfunction plays a critical role in the pathogenesis of HAND in drug-treated HIV-infected individuals and that antioxidant-mediated mitigation of oxidative stress could thus be considered as an adjunctive therapeutic strategy for ameliorating/dampening some of the neurological complications of HAND.

Oxidative Stress, Antioxidant Defenses, COVID-19 and Pollution

Patients with degenerative diseases present a chronic oxidative stress state, which puts them at a disadvantage when facing viral infections such as COVID-19. This is because there is a close relationship between redox signaling and this inflammatory response. Therefore, chronic changes in the redox balance cause alterations in the regulation of the immune system. An inflammatory response that must be reparative and self-limited loses its function and remains over time. In a chronic state of oxidative stress, there is a deficiency of antioxidants. This results in low levels of hormones, vitamins and trace elements, which are essential for the regulation of these systems. Furthermore, low levels of antioxidants imply a diminished capacity for a regulated inflammatory responses are much more vulnerable to a cytokine storm that mainly attacks the lungs, since they present a vicious circle between the null or diminished response of the antioxidant systems and the loss of regulation of the inflammatory process. Therefore, these patients are at a disadvantage in counteracting the response of defense systems to infection from SAR-COV19. A plausible option may be to restore the levels of Vitamins A, B, C, D, E and of essential trace elements such as manganese, selenium, zinc, in the body, which are key to either preventing or reducing the severity of the response of the immune system to the disease caused by SAR-CoV2. For the present review, we searched the specific sites of the Cochrane library database, PubMed and Medscape. The inclusion criteria were documents written in English or Spanish, published during the last 10 years.

Exercise and oxidative stress in type 2 diabetes mellitus

Sedentarism, overweight and malnutrition generate an increase in the production of reactive oxygen species leading to a state of chronic oxidative stress. In patients with type 2 diabetes mellitus, oxidative stress alters pancreatic insulin secretion and the actions of the hormone on target cells, contributing to the development of micro and macrovascular complications. During physical exertion a state of transient oxidative stress occurs. As a consequence, the organism generates multiple physiological adaptations to these repetitive stimuli. Physical exercise is beneficial for type 2 diabetes mellitus but there is a paucity of information about the effects of physical exercise on biomarkers of oxidative stress in patients with the disease. We herein try to elucidate if the effects of exercise on oxidative stress can help in the prevention and treatment of type 2 diabetes mellitus and which is the most effective modality of physical exercise to reduce oxidative stress markers.

Action of Phytochemicals on Insulin Signaling Pathways Accelerating Glucose Transporter (GLUT4) Protein Translocation.

Diabetes is associated with obesity, generally accompanied by a chronic state of oxidative stress and redox imbalances which are implicated in the progression of micro- and macro-complications like heart disease, stroke, dementia, cancer, kidney failure and blindness. All these complications rise primarily due to consistent high blood glucose levels. Insulin and glucagon help to maintain the homeostasis of glucose and lipids through signaling cascades. Pancreatic hormones stimulate translocation of the glucose transporter isoform 4 (GLUT4) from an intracellular location to the cell surface and facilitate the rapid insulin-dependent storage of glucose in muscle and fat cells. Malfunction in glucose uptake mechanisms, primarily contribute to insulin resistance in type 2 diabetes. Plant secondary metabolites, commonly known as phytochemicals, are reported to have great benefits in the management of type 2 diabetes. The role of phytochemicals and their action on insulin signaling pathways through stimulation of GLUT4 translocation is crucial to understand the pathogenesis of this disease in the management process. This review will summarize the effects of phytochemicals and their action on insulin signaling pathways accelerating GLUT4 translocation based on the current literature.

Glycyrrhizin ameliorates high fat diet-induced obesity in rats by activating NrF2 pathway

AimObesity based on insulin resistance is a state of chronic oxidative stress and inflammation that are highly regulated through nuclear factor Erythroid 2-related factor 2 (NrF2) pathway. Materials and methods70 male Wistar rats were randomized into two models. The prophylactic model was 10weeks and rats were grouped into: normal group, GL group (received glycyrrhizin 50mg/kg/day orally along with normal pellet diet), HFD group and HFD+ GL group (received glycyrrhizin along with HFD). The treatment model was 14weeks and rats were grouped into: normal group, HFD group and HFD+GL group (received glycyrrhizin from the week 10). Key findingsGlycyrrhizin decreased significantly rat weights and insulin resistance, normalized lipid profile and reduced significantly the adipocytes size in adipose tissue and lipid deposition in the liver tissue through histopathologic examination. Furthermore, glycyrrhizin ameliorated obesity-induced oxidative stress which indicated by significant decrease in liver malondialdehyde level (P<0.001) and increase in the total antioxidant capacity (P<0.001). Interestingly, molecular mechanism of glycyrrhizin was explored, that included significant reduction of liver gluconeogenic enzymes mRNA expression (P<0.001), a significant increase of liver insulin receptor, NrF2 and homooxygenase-1 mRNA expressions (P<0.001) and significant increase and nuclear translocation of NrF2 in liver tissue. SignificanceGlycyrrhizin ameliorates HFD-induced obesity in rats that may be attributed to its ability to increase insulin receptor expression and to activate NrF2 and subsequent homooxygenase-1 pathway. Thus, this work represents a safe natural compound (glycyrrhizin) that has a great role either as prophylaxis or treatment for insulin resistance related to obesity.

An Investigation of the Antioxidant Capacity in Extracts from Moringa oleifera Plants Grown in Jamaica.

Moringa oleifera trees grow well in Jamaica and their parts are popularly used locally for various purposes and ailments. Antioxidant activities in Moringa oleifera samples from different parts of the world have different ranges. This study was initiated to determine the antioxidant activity of Moringa oleifera grown in Jamaica. Dried and milled Moringa oleifera leaves were extracted with ethanol/water (4:1) followed by a series of liquid–liquid extractions. The antioxidant capacities of all fractions were tested using a 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay. IC50 values (the amount of antioxidant needed to reduce 50% of DPPH) were then determined and values for the extracts ranged from 177 to 4458 μg/mL. Extracts prepared using polar solvents had significantly higher antioxidant capacities than others and may have clinical applications in any disease characterized by a chronic state of oxidative stress, such as sickle cell anemia. Further work will involve the assessment of these extracts in a sickle cell model of oxidative stress.

Abstract 304: CLIC1 membrane insertion is a pivotal regulator of glioblastoma stem cell G1-S transition by promoting an increase of chloride permeability

Abstract Glioblastoma (GBM) is the most aggressive and lethal brain tumor and, despite aggressive surgery and adjuvant radiotherapy and/or chemotherapy, the prognosis remains invariantly poor. As for most of solid and hematological malignancies, it was demonstrated that the bulk of tumor cells in GBM is generated by a rare fraction of self-renewing, multipotent cancer stem cells (CSCs) and the persistence of CSCs within the tumor mass is considered the main determinant of GBM development, progression, recurrence and radio- or chemoresistance. Thus, one of the main goals of current research is to identify specific biological mechanisms or intracellular pathways of CSCs whose pharmacological targeting might affect their survival and proliferation. In particular, little is known about the possibility that the molecular mechanisms underlying cell-cycle control in GBM CSCs are endowed with specific and unique features as compared with normal cells. Our study is based on the observation that GBM cells express higher levels of chloride intracellular channel 1 (CLIC1) as compared to nonmalignant brain cells and that in CSCs CLIC1 is mainly localized in the membrane forming an active channel. Conversely, in physiological conditions CLIC1 is mainly a cytoplasmic protein only transiently translocating to the membrane. We recently showed that the different level of activity of CLIC1 in CSCs and normal mesenchymal stem cells confers CLIC1-targeting drugs (for example the biguanide metformin) selective cytotoxicity toward tumor cells. Here we report, that in response to stress conditions, CLIC1 increases the probability to modify its structure going from a cytoplasmic hydrophilic form to a transmembrane conformation. Once in the membrane, CLIC1 acts as a chloride permeability, participating, together with NADPH oxidase, to the generation of a chronic state of oxidative stress that favor the transition between G1 and S phase. The peculiarity of CLIC1 exposure on the external face of the GMB CSC plasma membrane support the idea that this protein could represent a main determinant of the cell cycle progression in this tumor cell subpopulation and thus an accessible and relevant pharmacological target to eradicate CSCs in GBM. Note: This abstract was not presented at the meeting. Citation Format: Ivan Verduci, Valentina Carlini, Federica M. Raciti, Matteo Conti, Federica Barbieri, Tullio Florio, Michele Mazzanti. CLIC1 membrane insertion is a pivotal regulator of glioblastoma stem cell G1-S transition by promoting an increase of chloride permeability [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 304. doi:10.1158/1538-7445.AM2017-304

Brain neurochemical and hemodynamic findings in the NY1DD mouse model of mild sickle cell disease.

To characterize the cerebral profile associated with sickle cell disease (SCD), we used in vivo proton MRI and MRS to quantify hemodynamics and neurochemicals in the thalamus of NY1DD mice, a mild model of SCD, and compared them with wild-type (WT) control mice. Compared with WT mice, NY1DD mice at steady state had elevated cerebral blood flow (CBF) and concentrations of N-acetylaspartate (NAA), glutamate (Glu), alanine, total creatine and N-acetylaspartylglutamate. Concentrations of glutathione (GSH) at steady state showed a negative correlation with BOLD signal change in response to 100% oxygen, a marker for oxidative stress, and mean diffusivity assessed using diffusion-tensor imaging, a marker for edematous inflammation. In NY1DD mice, elevated basal CBF was correlated negatively with [NAA], but positively with concentration of glutamine ([Gln]). Immediately after experimental hypoxia (at reoxygenation after 18hours of 8% O2 ), concentrations of NAA, Glu, GSH, Gln and taurine (Tau) increased only in NY1DD mice. [NAA], [Glu], [GSH] and [Tau] all returned to baseline levels two weeks after the hypoxic episode. The altered neurochemical profile in the NY1DD mouse model of SCD at steady state and following experimental hypoxia/reoxygenation suggests a state of chronic oxidative stress leading to compensatory cerebral metabolic adjustments.

The Effect of Chronic Ozone Exposure on the Activation of Endoplasmic Reticulum Stress and Apoptosis in Rat Hippocampus.

The chronic exposure to low doses of ozone, like in environmental pollution, leads to a state of oxidative stress, which has been proposed to contribute to neurodegenerative disorders, including Alzheimer’s disease (AD). It induces an increase of calcium in the endoplasmic reticulum (ER), which produces ER stress. On the other hand, different studies show that, in diseases such as Alzheimer’s, there exist disturbances in protein folding where ER plays an important role. The objective of this study was to evaluate the state of chronic oxidative stress on ER stress and its relationship with apoptotic death in the hippocampus of rats exposed to low doses of ozone. We used 108 male Wistar rats randomly divided into five groups. The groups received one of the following treatments: (1) Control (air); (2) Ozone (O3) 7 days; (3) O3 15 days; (4) O3 30 days; (5) O3 60 days; and (6) O3 90 days. Two hours after each treatment, the animals were sacrificed and the hippocampus was extracted. Afterwards, the tissue was processed for western blot and immunohistochemistry using the following antibodies: ATF6, 78 kDa glucose-regulated protein (GRP78) and caspase 12. It was also subjected to terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and electronic microscopy. Our results show an increase in ATF6, GRP78 and caspase 12 as well as ER ultrastructural alterations and an increase of TUNEL positive cells after 60 and 90 days of exposure to ozone. With the obtained results, we can conclude that oxidative stress induced by chronic exposure to low doses of ozone leads to ER stress. ER stress activates ATF6 inducing the increase of GRP78 in the cytoplasm, which leads to the increase in the nuclear translocation of ATF6. Finally, the translocation creates a vicious cycle that, together with the activation of the cascade for apoptotic cell death, contributes to the maintenance of ER stress. These events potentially contribute in the neurodegeneration processes in diseases like AD.

Obesity, Oxidative Stress, and Their Effect on Serum Heme Oxygenase-1 Concentrations and Insulin in Children Aged 3 to 5 Years in a Pediatric Hospital of the Ministry of Health CDMX.

Obesity during early stages of life may condition states of oxidative stress. Heme oxygenase-1 (HO-1) is an enzyme involved in oxidative metabolism; it has antioxidant and anti-inflammatory functions and is related in sensitivity to insulin. However, a high concentration of this enzyme has been described to cause alterations such as insulin resistance. The objective of this work was to study the relationship between obesity, oxidative stress, HO-1, and insulin in children aged 3 to 5 years. To achieve our objective, we studied a control group of children (n = 50) and a group of obese children (n = 50) who underwent an anthropometric evaluation. Additionally, we quantified peroxidized lipids, oxidized low-density lipoproteins (Ox-LDLs), oxidized and reduced glutathione, HO-1, and insulin. We also calculated the homeostasis model assessment of insulin resistance (HOMA-IR), HOMA-%B, and HOMA-%S indices. According to the data distribution, nonparametric and Spearman's rank correlation coefficient tests were conducted. The results demonstrate that obese children show a statistically relevant increase in BMI/age, serum concentrations of peroxidized lipids, Ox-LDLs, oxidized glutathione, HO-1, and insulin (p < 0.005). In addition, there was an increase in the HOMA-IR and HOMA-%B (p < 0.0001) indices and a decrease of reduced glutathione, as well as a reduction in the HOMA-%S, compared with the children of the control group (p < 0.003). With the results obtained, we can conclude that obese preschool children show a chronic state of oxidative stress, an increase of HO-1, and an incipient state of insulin resistance. Finally, the increased reactive oxygen species could be one of the leading factors involved in insulin resistance and Ox-LDL increase from the preschool stage.

Abstract 3293: The TLR-2 signal pathway is enhanced in myeloproliferative neoplasm - related to the increased inflammatory cytokine and pathogenesis

Abstract Introduction Although MPNs have been regarded as stem cell disease. Recent evidence suggest that chronic inflammation induces a state of chronic oxidative stress with elevated reactive oxygen species (ROS) and then induce alternation in epigenetic changes or DNA mutations.. Also, MPNs are associated with constitutional symptoms and high levels of circulating inflammatory cytokines. The etiology of this increase in inflammatory cytokines remains elusive, we investigated the role of TLRs (Toll-like receptors) and related cytokines in MPN as they are known to play a role in innate and adaptive immunity in inflammation. Materials and Methods: TLR assay: TLR-2, 4, 7, 9 quantification was performed by TLR staining of 5×105 mononuclear cells (MNC) which were incubated with 5 μg/ml of conjugated TLR-2, 4, 7, 9 antibody and assayed by flow cytometry. Monocyte-derived dendritic cells (mdDC) and Inflammatory Cytokines. mdDC were generated from isolated monocytes (anti-CD14-coated magnetic beads) then by incubation with IL-4 and GM-CSF for 4 days. Immature mdDC (1×104/well) were either left untreated or stimulated with Pam3CSK4, in 96 well plates. After 48h incubation, cells were stained with fluorescence-labeled mAb specific to CD80, CD83 (BD Biosciences). Cytokine levels were determined in supernatants harvested after 48h using the MSD (Meso Scale Discovery) System. Results. 1) Levels of TLR-2 were significantly elevated in patients with MPNs especially in those with polycythemia vera and essential thrombocythemia, but only marginally elevated in myelofibrosis patients. In all patient subsets, TLR-4, 7, 9 levels did not differ significantly from controls. 2) Maturation of mdDC were done by stimulation of TLR-2 ligand, Pam3CSK4. Higher numbers of CD83+ cells were generated more from patients with elevated TLR levels, confirming these MPN patients have elevated TLR-2 levels which portend the ability to generate more DC by TLR-2 ligand. 3) In patients with elevated TLR-2 level, Pam3CSK4 stimulated more cytokines including IFNγ, IL12p70, TNFα, IL6, IL8, IL10, and IL13 than those patients with low levels of TLR-2. 4). Moreover, patients with elevated TLR-2 levels have more of IL8, TNFα, IL13, IL6, in the plasma, consistent with inflammatory cytokines which is a likely consequence of elevated TLR-2. Conclusion. TLR-2 and not TLR-4, 7, 9 were found to be significantly elevated in MPNs. Stimulation of the TLR-2 ligand promotes the generation of more mature DCs, greater inflammatory cytokines production and Higher plasma inflammatory cytokines are associated with patients with higher TLR-2 levels. We conclude that TLR-2 is important in the production of inflammatory cytokines in MPNs and this evidence suggests that MPNs are in a TLR induced chronic inflammatory state and may play a significant role in the pathogenesis of MPNs. Citation Format: Guanfang Shi, Maryna Yarotska, Hui chen, Cherif Abdelmalek, Ching Wong, Madhumati Kalaovar, VLadimir Gotlieb, Jen Chin Wang. The TLR-2 signal pathway is enhanced in myeloproliferative neoplasm - related to the increased inflammatory cytokine and pathogenesis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3293.

The Protective Effect of Antioxidants Consumption on Diabetes and Vascular Complications.

Obesity and diabetes is generally accompanied by a chronic state of oxidative stress, disequilibrium in the redox balance, implicated in the development and progression of complications such as micro- and macro-angiopathies. Disorders in the inner layer of blood vessels, the endothelium, play an early and critical role in the development of these complications. Blunted endothelium-dependent relaxation and/or contractions are quietly associated to oxidative stress. Thus, preserving endothelial function and oxidative stress seems to be an optimization strategy in the prevention of vascular complications associated with diabetes. Diet is a major lifestyle factor that can greatly influence the incidence and the progression of type 2 diabetes and cardiovascular complications. The notion that foods not only provide basic nutrition but can also prevent diseases and ensure good health and longevity is now attained greater prominence. Some dietary and lifestyle modifications associated to antioxidative supply could be an effective prophylactic means to fight against oxidative stress in diabesity and complications. A significant benefit of phytochemicals (polyphenols in wine, grape, teas), vitamins (ascorbate, tocopherol), minerals (selenium, magnesium), and fruits and vegetables in foods is thought to be capable of scavenging free radicals, lowering the incidence of chronic diseases. In this review, we discuss the role of oxidative stress in diabetes and complications, highlight the endothelial dysfunction, and examine the impact of antioxidant foods, plants, fruits, and vegetables, currently used medication with antioxidant properties, in relation to the development and progression of diabetes and cardiovascular complications.

Exercise Modulates Oxidative Stress and Inflammation in Aging and Cardiovascular Diseases.

Despite the wealth of epidemiological and experimental studies indicating the protective role of regular physical activity/exercise training against the sequels of aging and cardiovascular diseases, the molecular transducers of exercise/physical activity benefits are not fully identified but should be further investigated in more integrative and innovative approaches, as they bear the potential for transformative discoveries of novel therapeutic targets. As aging and cardiovascular diseases are associated with a chronic state of oxidative stress and inflammation mediated via complex and interconnected pathways, we will focus in this review on the antioxidant and anti-inflammatory actions of exercise, mainly exerted on adipose tissue, skeletal muscles, immune system, and cardiovascular system by modulating anti-inflammatory/proinflammatory cytokines profile, redox-sensitive transcription factors such as nuclear factor kappa B, activator protein-1, and peroxisome proliferator-activated receptor gamma coactivator 1-alpha, antioxidant and prooxidant enzymes, and repair proteins such as heat shock proteins, proteasome complex, oxoguanine DNA glycosylase, uracil DNA glycosylase, and telomerase. It is important to note that the effects of exercise vary depending on the type, intensity, frequency, and duration of exercise as well as on the individual's characteristics; therefore, the development of personalized exercise programs is essential.

HPV16 E6 and E7 proteins induce a chronic oxidative stress response via NOX2 that causes genomic instability and increased susceptibility to DNA damage in head and neck cancer cells.

Human papillomavirus (HPV) is the causative agent of a subgroup of head and neck cancer characterized by an intrinsic radiosensitivity. HPV initiates cellular transformation through the activity of E6 and E7 proteins. E6 and E7 expression is necessary but not sufficient to transform the host cell, as genomic instability is required to acquire the malignant phenotype in HPV-initiated cells. This study reveals a key role played by oxidative stress in promoting genomic instability and radiosensitivity in HPV-positive head and neck cancer. By employing an isogenic human cell model, we observed that expression of E6 and E7 is sufficient to induce reactive oxygen species (ROS) generation in head and neck cancer cells. E6/E7-induced oxidative stress is mediated by nicotinamide adenine dinucleotide phosphate oxidases (NOXs) and causes DNA damage and chromosomal aberrations. This mechanism for genomic instability distinguishes HPV-positive from HPV-negative tumors, as we observed NOX-induced oxidative stress in HPV-positive but not HPV-negative head and neck cancer cells. We identified NOX2 as the source of HPV-induced oxidative stress as NOX2 silencing significantly reduced ROS generation, DNA damage and chromosomal aberrations in HPV-positive cells. Due to their state of chronic oxidative stress, HPV-positive cells are more susceptible to DNA damage induced by ROS and ionizing radiation (IR). Furthermore, exposure to IR results in the formation of complex lesions in HPV-positive cells as indicated by the higher amount of chromosomal breakage observed in this group of cells. These results reveal a novel mechanism for sustaining genomic instability in HPV-positive head and neck tumors and elucidate its contribution to their intrinsic radiosensitivity.

Abstract 4558: Using reduction mammoplasty tissues from healthy young women living in USA's high breast cancer (BC) risk posing environment to open up to study the “black box” of latent stages of breast carcinogenesis; a key to develop strategies for BC

Abstract Knowledge of latent stages of breast carcinogenesis is limited to when histological changes are already present due to a lack of; a) cytochemical marker(s) needed to identify loci in histologically normal breast tissues where procarcinogenic conditions exist and; b) microscopic systems needed to geno- and phenotype cells in such loci. In 2 recent publications we presented evidence that protein adducts of 4-hydroxy-2-nonenal (4HNE), markers of a procarcinogenic state of chronic oxidative stress (OxS), might provide the needed fiducial marker. Specifically, a) we documented the presence of 4HNE immunopositive (4HNE+) mammary epithelial (ME) cells in terminal ductal units (TDLUs) in breast tissues of healthy teenagers and young women and their association with altered expression of OxS-related mRNAs and; b) presented Fourier-transform infrared microspectroscopic evidence of differences in biomolecular profile of ME cells and associated intra- and interlobular stromal cells in 4HNE+ vs. 4HNE- TDLUs, i.e., of cells of origin of BCs and cells in their supportive stromal microenvironment. Here we present further evidence of molecular changes in breast tissues of young women (ages 17-30 y) associated with 4HNE adducts. A comparison of the transcriptome of breast tissues with many 4HNE+ vs. few 4HNE+ TDLUs (controls) by Illumina 24,000 gene platform and SABioscience 84 gene Cell Death PCR Array revealed reprogramming of many homeostatic mechanism in 4HNE+ TDLUs. Notably, a) DASL platform identified transcripts for 395 genes that differed (p = &amp;lt;.05) between 4HNE+ vs. 4HNE- tissues, transcripts that were assigned by GO bioinformatics tool to 21 molecular mechanism and; b) Cell Death array identified 30 genes that differed &amp;gt;2 fold between 4HNE+ vs. 4HNE- tissues. The findings support using 4HNE adducts to identify in histologically normal breast tissues loci harboring cells that might initiate a cancer. Our findings fit an evolutionary model of carcinogenesis and reflect adaptive responses to stressors that serve to preserve tissue integrity, e.g., by developing resistance to OxS and to apoptosis, responses however that over time can lead to the emergence of a cancer. Our recently acquired Toponome Imaging System (TIS) enables identifying such responses to chronic OxS implied by the presence of 4HNE adducts. TIS is the first high throughput fluorescence robotic microscopic system that can provide a map, a toponome, of up to 100 proteins at 40nm functional resolution in individual cells in a tissue section. In ongoing experiments we are using TIS and antibodies against up to 21 proteins to test the postulate that 4HNE+ TDLUs are enriched with cells with an apoptosis resistant phenotype, a postulate supported by our transcriptional data and predicted by the evolutionary model of carcinogenesis. Citation Format: Judith Weisz, Debra Shearer, Anna C. Salzberg. Using reduction mammoplasty tissues from healthy young women living in USA's high breast cancer (BC) risk posing environment to open up to study the “black box” of latent stages of breast carcinogenesis; a key to develop strategies for BC [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4558. doi:10.1158/1538-7445.AM2015-4558