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Introduction: Chemotherapy, notably docetaxel (Doc), stands as the primary treatment for castration-resistant prostate cancer (CRPC). However, its efficacy is hindered by side effects and chemoresistance. Hypoxia in prostate cancer (PC) correlates with chemoresistance to Doc-induced apoptosis via Heme Oxygenase-1 (HO-1) modulation, a key enzyme in heme metabolism. This study investigated targeting heme degradation pathway via HO-1 inhibition to potentiate the therapeutic efficacy of Doc in PC. Methods: Utilizing diverse PC cell lines, we evaluated HO-1 inhibition alone and with Doc on viability, apoptosis, migration, and epithelial- to- mesenchymal transition (EMT) markers and elucidated the underlying mechanisms. Results: HO-1 inhibition significantly reduced PC cell viability under hypoxic and normoxic conditions, enhancing Doc-induced apoptosis through interconnected mechanisms, including elevated reactive oxygen species (ROS) levels, glutathione cycle disruption, and modulation of Signal Transducer and Activator of Transcription 1 (STAT1) pathway. The interplay between STAT1 and HO-1 suggests its reliance on HO-1 activation. Additionally, a decrease in cell migration and downregulation of EMT markers (vimentin and snail) were observed, indicating attenuation of mesenchymal phenotype. Discussion: In conclusion, the combination of HO-1 inhibition with Doc holds promise for improving therapeutic outcomes and advancing clinical management in PC.
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Purpose: Imaging professionals are occupationally exposed to chronic ionizing radiation (IR) and non-ionizing radiation (NIR). This study aimed to investigate the influence of occupational radiation exposure on oxidative stress and antioxidant levels based on blood biomarkers in different hospital imaging professional groups.Materials and methods: The study groups included 66 imaging professionals occupationally exposed to IR (n = 58, 43 diagnostic radiography (G1), seven nuclear medicine (G2), eight radiation therapy (G3)), and NIR (n = 8, ultrasound imaging (G4)) and 60 non-exposed controls. Blood levels of superoxide (O2â¢-) as an index of oxidative stress, and the antioxidant activities of superoxide dismutase (SOD), glutathione ratio (GSH/GSSG), and catalase (CAT) were measured.Results: The blood values of O2â¢-, SOD, and CAT were significantly higher in imaging professionals occupationally exposed to radiation than in the control group (p < .05), while a significant decrease in the ratio of GSH/GSSG was observed (p < .05). The results from the NIR group were significantly higher compared to IR group.Conclusions: Based on these results, chronic exposure to radiation (IR and NIR) is associated with redox dysregulation that may result in damages to cellular biomolecules including lipids, proteins and DNA. Further studies are needed to determine the impact of redox dysregulation and the need for periodic examination among imaging professionals occupationally exposed to IR and NIR.
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Antioxidantes , Glutationa , Humanos , Antioxidantes/metabolismo , Dissulfeto de Glutationa/metabolismo , Oxirredução , Glutationa/metabolismo , Estresse Oxidativo , Superóxido Dismutase/metabolismo , Radiação Ionizante , Radiação não IonizanteRESUMO
Even in the modern era of combination antiretroviral therapy, aberrations in motor control remain a predominant symptom contributing to age-related functional dependencies (e.g., neurocognitive impairment) in people with HIV (PWH). While recent evidence implicates aberrant mitochondrial redox environments in the modulation of neural oscillatory activity serving motor control in PWH, the contribution of important clinical and demographic factors on this bioenergetic-neural-behavioral pathway is unknown. Herein, we evaluate the predictive capacity of clinical metrics pertinent to HIV (e.g., CD4 nadir, time with viremia) and age on mitochondrial redox-regulated sensorimotor brain-behavior dynamics in 69 virally-suppressed PWH. We used state-of-the-art systems biology and neuroscience approaches, including Seahorse analyzer of mitochondrial energetics, EPR spectroscopy of intracellular oxidant levels, antioxidant activity assays pertinent to superoxide and hydrogen peroxide (H2O2) redox environments, and magnetoencephalographic (MEG) imaging to quantify sensorimotor oscillatory dynamics. Our results demonstrate differential effects of redox systems on the neural dynamics serving motor function in PWH. In addition, measures of immune stability and duration of compromise due to HIV had dissociable effects on this pathway, above and beyond the effects of age alone. Moreover, peripheral measures of antioxidant activity (i.e., superoxide dismutase) fully mediated the relationship between immune stability and current behavioral performance, indicative of persistent oxidative environments serving motor control in the presence of virologic suppression. Taken together, our data suggest that disease-related factors, in particular, are stronger predictors of current redox, neural and behavioral profiles serving motor function, which may serve as effective targets for alleviating HIV-specific alterations in cognitive-motor function in the future.
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Antioxidantes , Infecções por HIV , Humanos , Peróxido de Hidrogênio , Infecções por HIV/tratamento farmacológico , Oxirredução , BiomarcadoresRESUMO
Pancreatic ductal adenocarcinoma (PDAC) is the most prevalent form, accounting for more than 90% of all pancreatic malignancies. In a previous study, we found that hypoxia and chemotherapy induced expression of Heme Oxygenase-1 (HO-1) in PDAC cells and tissues. Arsenic trioxide (ATO) is the first-line chemotherapeutic drug for acute promyelocytic leukemia (APL). ATO increases the generation of reactive oxidative species (ROS) and induces apoptosis in treated cells. The clinical use of ATO for solid tumors is limited due to severe systemic toxicity. In order to reduce cytotoxic side effects and resistance and improve efficacy, it has become increasingly common to use combination therapies to treat cancers. In this study, we used ATO-sensitive and less sensitive PDAC cell lines to test the effect of combining HO-1 inhibitors (SnPP and ZnPP) with ATO on HO-1 expression, cell survival, and other parameters. Our results show that ATO significantly induced the expression of HO-1 in different PDAC cells through the p38 MAPK signaling pathway. ROS production was confirmed using the oxygen-sensitive probes DCFH and DHE, N-acetyl cysteine (NAC), an ROS scavenger, and oxidized glutathione levels (GSSG). Both ATO and HO-1 inhibitors reduced PDAC cell survival. In combined treatment, inhibiting HO-1 significantly increased ATO cytotoxicity, disrupted the GSH cycle, and induced apoptosis as measured using flow cytometry. ATO and HO-1 inhibition modulated autophagy as shown by increased expression of autophagy markers ATG5, p62, and LC3B in PDAC cells. This increase was attenuated by NAC treatment, indicating that autophagy modulation was through an ROS-dependent mechanism. In conclusion, our work explored new strategies that could lead to the development of less toxic and more effective therapies against PDAC by combining increased cellular stress and targeting autophagy.
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Despite virologic suppression, people living with HIV (PLWH) remain at risk for developing cognitive impairment, with aberrations in motor control being a predominant symptom leading to functional dependencies in later life. While the neuroanatomical bases of motor dysfunction have recently been illuminated, the underlying molecular processes remain poorly understood. Herein, we evaluate the predictive capacity of the mitochondrial redox environment on sensorimotor brain-behavior dynamics in 40 virally-suppressed PLWH and 40 demographically-matched controls using structural equation modeling. We used state-of-the-art approaches, including Seahorse Analyzer of mitochondrial function, electron paramagnetic resonance spectroscopy to measure superoxide levels, antioxidant activity assays and dynamic magnetoencephalographic imaging to quantify sensorimotor oscillatory dynamics. We observed differential modulation of sensorimotor brain-behavior relationships by superoxide and hydrogen peroxide-sensitive features of the redox environment in PLWH, while only superoxide-sensitive features were related to optimal oscillatory response profiles and better motor performance in controls. Moreover, these divergent pathways may be attributable to immediate, separable mechanisms of action within the redox environment seen in PLWH, as evidenced by mediation analyses. These findings suggest that mitochondrial redox parameters are important modulators of healthy and pathological oscillations in motor systems and behavior, serving as potential targets for remedying HIV-related cognitive-motor dysfunction in the future.
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Infecções por HIV , Nível de Saúde , Humanos , Encéfalo , MitocôndriasRESUMO
Motor control requires a coordinated ensemble of spatiotemporally precise neural oscillations across a distributed motor network, particularly in the beta range (15 to 30 Hz) to successfully plan and execute volitional actions. While substantial evidence implicates beta activity as critical to motor control, the molecular processes supporting these microcircuits and their inherent oscillatory dynamics remain poorly understood. Among these processes are mitochondrial integrity and the associated redox environments, although their direct impact on human neurophysiological function is unknown. Herein, 40 healthy adults completed a motor sequence paradigm during magnetoencephalography (MEG). MEG data were imaged in the time-frequency domain using a beamformer to evaluate beta oscillatory profiles during distinct phases of motor control (i.e., planning and execution) and subsequent behavior. To comprehensively quantify features of the mitochondrial redox environment, we used state-of-the-art systems biology approaches including Seahorse Analyzer to assess mitochondrial respiration and electron paramagnetic resonance spectroscopy to measure superoxide levels in whole blood as well as antioxidant activity assays. Using structural equation modeling, we tested the relationship between mitochondrial function and sensorimotor brain-behavior dynamics through alterations in the redox environment (e.g., generation of superoxide and alteration in antioxidant defenses). Our results indicated that superoxide-sensitive but not hydrogen peroxide-sensitive features of the redox environment had direct and mediating effects on the bioenergetic-neural pathways serving motor performance in healthy adults. Importantly, our results suggest that alterations in the redox environment may directly impact behavior above and beyond mitochondrial respiratory capacities alone and further may be effective targets for age- and disease-related declines in cognitive-motor function.
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Córtex Sensório-Motor/fisiologia , Adulto , Idoso , Ritmo beta/fisiologia , Feminino , Humanos , Peróxido de Hidrogênio/metabolismo , Magnetoencefalografia , Masculino , Pessoa de Meia-Idade , Mitocôndrias/metabolismo , Modelos Neurológicos , Movimento/fisiologia , Vias Neurais/fisiologia , Oxirredução , Desempenho Psicomotor/fisiologia , Superóxidos/metabolismo , Adulto JovemRESUMO
BACKGROUND: Despite effective combination antiretroviral therapy (cART), people living with HIV (PLWH) remain at risk for developing neurocognitive impairment primarily due to systemic inflammation that persists despite virologic suppression, albeit the mechanisms underlying such inflammation are poorly understood. METHODS: Herein, we evaluate the predictive capacity of the mitochondrial redox environment on circulating neuro- and T-lymphocyte-related inflammation and concomitant cognitive function in 40 virally-suppressed PLWH and 40 demographically-matched controls using structural equation modeling. We used state-of-the-art systems biology approaches including Seahorse Analyzer of mitochondrial function, electron paramagnetic resonance (EPR) spectroscopy to measure superoxide levels, antioxidant activity assays, and Meso Scale multiplex technology to quantify inflammatory proteins in the periphery. FINDINGS: We observed disturbances in mitochondrial function and the redox environment in PLWH compared to controls, which included reduced mitochondrial capacity (t(76) = -1.85, p = 0.034, 95% CI: -∞,-0.13), elevated levels of superoxide (t(75) = 1.70, p = 0.047, 95% CI: 8.01 E 3, ∞) and alterations in antioxidant defense mechanisms (t(74) = 1.76, p = 0.041, 95% CI: -710.92, ∞). Interestingly, alterations in both superoxide- and hydrogen peroxide-sensitive redox environments were differentially predictive of neuro-, but not T-lymphocyte-related inflammatory profiles in PLWH and controls, respectively (ps < 0.026). Finally, when accounting for superoxide-sensitive redox pathways, neuroinflammatory profiles significantly predicted domain-specific cognitive function across our sample (ß = -0.24, p = 0.034, 95% CI: -0.09, -0.004 for attention; ß = -0.26, p = 0.018, 95% CI: -0.10, -0.01 for premorbid function). INTERPRETATION: Our results suggest that precursors to neuroinflammation apparent in PLWH (i.e., mitochondrial function and redox environments) predict overall functionality and cognitive dysfunction and importantly, may serve as a proxy for characterizing inflammation-related functional decline in the future. FUNDING: National Institute of Mental Health, National Institute for Neurological Disorders and Stroke, National Institute on Drug Abuse, National Science Foundation.
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Complexo AIDS Demência/sangue , Estresse Oxidativo , Complexo AIDS Demência/diagnóstico , Adulto , Idoso , Biomarcadores/sangue , Células Cultivadas , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mitocôndrias/metabolismo , Superóxidos/metabolismo , Linfócitos T/metabolismoRESUMO
Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with a poor prognosis. Tumor hypoxia plays an active role in promoting tumor progression, malignancy, and resistance to therapy in PDAC. We present evidence that nab-paclitaxel-gemcitabine (NPG) and/or a hypoxic tumor microenvironment (TME) up-regulate heme oxygenase-1 (HO-1), providing a survival advantage for tumors. Using PDAC cells in vitro and a PDAC mouse model, we found that NPG chemotherapy up-regulated expression of HO-1 in PDAC cells and increased its nuclear translocation. Inhibition of HO-1 with ZnPP and SnPP sensitized PDAC cells to NPG-induced cytotoxicity (p < 0.05) and increased apoptosis (p < 0.05). Additionally, HO-1 expression was increased in gemcitabine-resistant PDAC cells (p < 0.05), and HO-1 inhibition increased GEM-resistant PDAC sensitivity to NPG (p < 0.05). NPG combined with HO-1 inhibitor inhibited tumor size in an orthotopic model. In parallel, HO-1 inhibition abrogated the influx of macrophages and FoxP3+ cells, while increasing the proportion of CD8+ infiltration in the pancreatic tumors. These effects were mediated primarily by reducing expression of the immunosuppressive cytokine IL-10.
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BACKGROUND: A known relationship exists between oxidative stress and preterm birth (PTB). However, few studies have measured oxidative stress prospectively in early or midpregnancy, and no studies have used electron paramagnetic resonance (EPR) spectroscopy prospectively to predict PTB. OBJECTIVE: The purpose of this study was to identify predictive relationships between antioxidants and reactive oxygen species (ROS), specifically, superoxide (O2), peroxynitrite (OONO), and hydroxyl radical (OH), using EPR spectroscopy, measured between 12 and 20 weeks of gestation and compare with the incidence of PTB. METHODS: Blood was obtained from pregnant women (n = 140) recruited from a tertiary perinatal center. Whole blood was analyzed directly for ROS, O2, OONO, and OH using EPR spectroscopy. Red blood cell lysate was used to measure antioxidants. PTB was defined as parturition at <37 weeks of gestation. RESULTS: No differences were found between ROS, O2, OONO, or OH with the incidence of PTB. Catalase activity, glutathione, and reduced/oxidized glutathione ratio were significantly lower with PTB. Logistic regression suggests decreased catalase activity in pregnant women is associated with increased odds of delivering prematurely. DISCUSSION: We prospectively compared antioxidants and specific ROS using EPR spectroscopy in pregnant women between 12 and 20 weeks of gestation with the incidence of PTB. Results are minimal but do suggest that antioxidants-specifically decreased catalase activity-in early pregnancy may be associated with PTB; however, these findings should be cautiously interpreted and may not have clinical significance.
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Idade Gestacional , Estresse Oxidativo , Nascimento Prematuro/epidemiologia , Antioxidantes/análise , Espectroscopia de Ressonância de Spin Eletrônica , Feminino , Humanos , Gravidez , Estudos Prospectivos , Espécies Reativas de Oxigênio/sangueRESUMO
INTRODUCTION: Preeclampsia (PE), one of the most serious complications of pregnancy, is characterized by endothelial dysfunction and hypertension. The pathophysiology of the disease is still unknown; however, evidence suggests that placental and maternal oxidative stress promote the disease process. Several studies have assessed levels of oxidative stress during pregnancy, but after diagnosis of PE. However, few studies have examined oxidative stress before PE diagnosis. Thus, the present work was aimed to gain further insight into the role of oxidative stress prior to diagnosis of PE (i.e. 12-20â¯weeks of gestation) and to further understand and predict PE incidence. METHODS: Blood levels of superoxide (O2-) and erythrocyte antioxidants such as superoxide dismutase (SOD), catalase (CAT), reduced glutathione (GSH) and oxidized glutathione (GSSG) levels were measured in 23 preeclamptic pregnant women and 91 women with normal pregnancies. We further used logistic regression of O2- and each antioxidant level as the main predictor variable for PE risk. RESULTS: CAT activity, GSH, and Total glutathione (TGSH) were significantly lower with All PE pregnant groups, whereas O2- levels were modestly, but significantly, higher in women with mild PE. Logistic regression analysis suggests increased CAT activity in pregnant women is associated with a decreased odds of being preeclamptic. CONCLUSION: CAT is the only antioxidant as shown in our study to be related to the severity of the disease and may be a promising predictor for PE. Further studies are warranted to investigate the use of CAT as a novel therapeutic for PE.
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Estresse Oxidativo , Pré-Eclâmpsia/diagnóstico , Diagnóstico Pré-Natal , Adulto , Estudos de Casos e Controles , Feminino , Glutationa/sangue , Humanos , Estudos Longitudinais , Pré-Eclâmpsia/sangue , Pré-Eclâmpsia/fisiopatologia , Gravidez , Primeiro Trimestre da Gravidez , Segundo Trimestre da Gravidez , Estudos Prospectivos , Superóxido Dismutase/sangueRESUMO
BACKGROUND: Oxidative stress is associated with poor perinatal outcomes. Little is known regarding the longitudinal levels of oxidative stress in the perinatal period or the correlation between maternal and neonatal oxidative stress levels. OBJECTIVE: Describe and compare oxidative stress, specifically superoxide, superoxide dismutase, catalase, and glutathione levels, over the perinatal period. STUDY DESIGN: Longitudinal descriptive design using a convenience sample of medically high- and low-risk pregnant women (n = 140) from a maternal-fetal medicine and general obstetrics practice, respectively. Blood was obtained from women at 12-20 and 24-28 weeks' gestation and during labor, from the umbilical cord at birth, and from neonates at 24-72 hr after birth. Levels of superoxide were measured using electron paramagnetic resonance (EPR) spectroscopy; antioxidants (superoxide dismutase, catalase, and glutathione) were measured using commercial assay kits. Relationships between oxidative stress levels at different time points were examined using nonparametric methods. Pregnancy outcome was collected. RESULTS: Demographic variables, outcome variables, and oxidative stress levels in maternal blood, cord blood, and infants differed between medically high- and low-risk women. Descriptive patterns for oxidative stress measures varied over time and between risk groups. Significant correlations between time points were noted, suggesting intraindividual consistency may exist throughout the perinatal period. However, these correlations were not consistent across each medical risk group. CONCLUSION: EPR spectroscopy is a feasible method for the perinatal population. Results provide new information on perinatal circulating superoxide levels and warrant further investigation into potential relationships between prenatal and neonatal physiologic dysregulation of oxidative stress.
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Antioxidantes/metabolismo , Sangue Fetal/química , Recém-Nascido/sangue , Estresse Oxidativo , Placenta/metabolismo , Adulto , Biomarcadores/sangue , Feminino , Humanos , Estudos Longitudinais , Troca Materno-Fetal , Gravidez , Resultado da Gravidez , Superóxido Dismutase/sangueRESUMO
Bevelacqua and Mortazavi [...].
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Studies have shown an increased risk for a variety of cancers, specifically brain cancer, in healthcare workers occupationally exposed to ionizing radiation. Although the mechanisms mediating these phenomena are not fully understood, ionizing radiation-mediated elevated levels of reactive oxygen species (ROS), oxidative DNA damage, and immune modulation are likely involved. A group of 20 radiation exposed workers and 40 sex- and age-matched non-exposed control subjects were recruited for the study. We measured superoxide (O2â¢-) levels in whole blood of healthcare workers and all other measurements of cytokines, oxidative DNA damage, extracellular superoxide dismutase (EcSOD) activity and reduced/oxidized glutathione ratio (GSH/GSSG) in plasma. Levels of O2â¢- were significantly higher in radiation exposed workers compared to control. Similarly, a significant increase in the levels of interleukin (IL)-6, IL-1α and macrophage inflammatory protein (MIP)-1α in radiation exposed workers compared to control was observed, while there was no significance difference in the other 27 screened cytokines. A significant positive correlation was found between MIP-1α and O2â¢- levels with no correlation in either IL-6 or IL-1α. Further, a dose-dependent relationship with significant O2â¢- production and immune alterations in radiation exposed workers was demonstrated. There was no statistical difference between the groups in terms of oxidative DNA damage, GSH/GSSG levels, or EcSOD activity. Although the biologic significance of cytokines alterations in radiation exposed workers is unclear, further studies are needed for determining the underlying mechanism of their elevation.
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Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive malignancies and has one of the worst prognoses leading to a meager 5-year survival rate of â¼8%. Chemotherapy has had limited success in extending the life span of patients with advanced PDAC due to poor tumor perfusion and hypoxia-induced resistance. Hypoxia reprograms the gene expression profile and upregulates the expression of multiple genes including heme oxygenase-1 (HO-1), which provide survival advantage to PDAC cells. However, the relationships between HO-1, hypoxia, and response to chemotherapy is unclear. Our results showed that hypoxia upregulates the expression of HO-1 in PDAC cells, and HO-1 inhibition using the HO-1 inhibitors zinc protoporphyrin, tin protoporphyrin IX (SnPP), and HO-1 knockout using CRISPR/Cas9 suppresses the proliferation of PDAC cells under hypoxia and sensitize them to gemcitabine under in vitro conditions. Treating orthotopic tumors with SnPP, or SnPP in combination with gemcitabine, significantly reduced the weight of pancreatic tumors (P < 0.05), decreased metastasis and improved the efficacy of gemcitabine treatment (P < 0.05). Mechanistically, inhibition of HO-1 increased the production of reactive oxygen species as demonstrated by increased dihydroethidium, and Mitosox, disrupted glutathione cycle, and enhanced apoptosis. There was significant increase in cleaved caspase-3 staining in tumors after combined treatment with SnPP and gemcitabine comparing to control or gemcitabine alone. In addition, inhibiting HO-1 reduced expression of stemness markers (CD133, and CD44) as compared to control or gemcitabine. Overall, our study may present a novel therapeutic regimen that might be adopted for the treatment of PDAC patients.
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Desoxicitidina/análogos & derivados , Heme Oxigenase-1/antagonistas & inibidores , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/patologia , Animais , Apoptose/efeitos dos fármacos , Biomarcadores Tumorais/metabolismo , Hipóxia Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Desoxicitidina/farmacologia , Desoxicitidina/uso terapêutico , Feminino , Glutationa/metabolismo , Heme Oxigenase-1/metabolismo , Humanos , Masculino , Camundongos Nus , Modelos Biológicos , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Espécies Reativas de Oxigênio/metabolismo , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismo , Células Estromais/patologia , GencitabinaRESUMO
BACKGROUND: A variety of methods and measures have been used to quantify oxidative stress in clinical studies related to preterm birth (PTB), and studies have reported conflicting findings. No integrative reviews have been conducted. OBJECTIVE: To describe specific molecules used as markers of oxidative stress and methods to measure these molecules and to review the literature for associations between oxidative stress and PTB specific to these molecules. METHOD: Systematic literature searches were conducted in June 2015 and updated in 2017 in databases from the Biomedical Reference Collection: Basic Edition, including MEDLINE and clinicaltrials.gov . Articles were included if they described original research published after 2009 and compared PTB or preterm premature rupture of membranes with term birth (TB). RESULTS: Abstracts ( n = 3,107) were reviewed for inclusion/exclusion criteria. Of these, 308 were full-text reviewed, and 30 articles were included in this review. All were identified as nonexperimental. The most common measurements of oxidative stress were quantification of total oxidant or antioxidant status or lipid peroxidation. Studies measuring reactive oxygen species or by-products of oxidative stress reported higher levels of these molecules for preterm specimens compared to TB specimens. Studies measuring antioxidants reported lower levels for these molecules in PTB specimens. Few of the studies had inconclusive findings. DISCUSSION: Findings suggest that an imbalance between oxidants and antioxidants may be associated with PTB. The measurements and findings to date limit interpretation and understanding. Research using multidimensional methods and multidisciplinary teams are necessary to advance research and practice.
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Antioxidantes/análise , Antioxidantes/fisiologia , Biomarcadores/análise , Estresse Oxidativo/fisiologia , Nascimento Prematuro/fisiopatologia , Espécies Reativas de Oxigênio/análise , Adulto , Feminino , Humanos , Recém-Nascido , Gravidez , GestantesRESUMO
The airway epithelium is a broad interface with the environment, mandating well-orchestrated responses to properly modulate inflammation. Classically, autophagy is a homeostatic pathway triggered in response to external cellular stresses, and is elevated in chronic airway diseases. Recent findings highlight the additional role of autophagy in vesicle trafficking and protein secretion, implicating autophagy pathways in complex cellular responses in disease. Th2 cytokines, IL-13 and IL-4, are increased in asthma and other airway diseases contributing to chronic inflammation. Previously, we observed that IL-13 increases reactive oxygen species (ROS) in airway epithelial cells in an autophagy-dependent fashion. Here, we tested our hypothesis that autophagy is required for IL-13-mediated superoxide production via the NADPH oxidase DUOX1. Using a mouse model of Th2-mediated inflammation induced by OVA-allergen, we observed elevated lung amounts of IL-13 and IL-4 accompanied by increased autophagosome levels, determined by LC3BII protein levels and immunostaining. ROS levels were elevated and DUOX1 expression was increased 70-fold in OVA-challenged lungs. To address the role of autophagy and ROS in the airway epithelium, we treated primary human tracheobronchial epithelial cells with IL-13 or IL-4. Prolonged, 7-day treatment increased autophagosome formation and degradation, while brief activation had no effect. Under parallel culture conditions, IL-13 and IL-4 increased intracellular superoxide levels as determined by electron paramagnetic resonance (EPR) spectroscopy. Prolonged IL-13 activation increased DUOX1, localized at the apical membrane. Silencing DUOX1 by siRNA attenuated IL-13-mediated increases in superoxide, but did not reduce autophagy activities. Notably, depletion of autophagy regulatory protein ATG5 significantly reduced superoxide without diminishing total DUOX1 levels. Depletion of ATG5, however, diminished DUOX1 localization at the apical membrane. The findings suggest non-canonical autophagy activity regulates DUOX1-dependent localization required for intracellular superoxide production during Th2 inflammation. Thus, in chronic Th2 inflammatory airway disease, autophagy proteins may be responsible for persistent intracellular superoxide production.
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Autofagia , Oxidases Duais/imunologia , Células Epiteliais/imunologia , Interleucina-13/imunologia , Superóxidos/imunologia , Animais , Linhagem Celular , Oxidases Duais/análise , Humanos , Inflamação/imunologia , Pulmão/imunologia , Camundongos Endogâmicos BALB CRESUMO
Communication between the nucleus and mitochondrion could coordinate many cellular processes. While the mechanisms regulating this communication are not completely understood, we hypothesize that cell cycle checkpoint proteins coordinate the cross-talk between nuclear and mitochondrial functions following oxidative stress. Human normal skin fibroblasts, representative of the G2-phase, were irradiated with 6 Gy of ionizing radiation and assayed for cyclin B1 translocation, mitochondrial function, reactive oxygen species (ROS) levels, and cytotoxicity. In un-irradiated controls, cyclin B1 was found primarily in the nucleus of G2-cells. However, following irradiation, cyclin B1 was excluded from the nucleus and translocated to the cytoplasm and mitochondria. These observations were confirmed further by performing transmission electron microscopy and cell fractionation assays. Cyclin B1 was absent in mitochondria isolated from un-irradiated G2-cells and present in irradiated G2-cells. Radiation-induced translocation of cyclin B1 from the nucleus to the mitochondrion preceded changes in the activities of mitochondrial proteins, that included decreases in the activities of aconitase and the mitochondrial antioxidant enzyme, manganese superoxide dismutase (MnSOD), and increases in complex II activity. Changes in the activities of mito-proteins were followed by an increase in dihydroethidium (DHE) oxidation (indicative of increased superoxide levels) and loss of the mitochondrial membrane potential, events that preceded the restart of the stalled cell cycle and subsequently the loss in cell viability. Comparable results were also observed in un-irradiated control cells overexpressing mitochondria-targeted cyclin B1. These results indicate that MnSOD and cyclin B1 coordinate a cross-talk between nuclear and mitochondrial functions, to regulate a mito-checkpoint during the cell cycle response to oxidative stress.
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It is well established that several forms of cancer associate with significant iron overload. Recent studies have suggested that estrogen (E2) disrupts intracellular iron homeostasis by reducing hepcidin synthesis and maintaining ferroportin integrity. Here, the ability of E2 to alter intracellular iron status and cell growth potential was investigated in MCF-7 cells treated with increasing concentrations of E2. Treated cells were assessed for intracellular iron status, the expression of key proteins involved in iron metabolism, oxidative stress, cell survival, growth, and apoptosis. E2 treatment resulted in a significant reduction in hepcidin expression and a significant increase in hypoxia-inducible factor 1 alpha, ferroportin, transferrin receptor, and ferritin expression; a transient decrease in labile iron pool; and a significant increase in total intracellular iron content mainly at 20 nM/48 h E2 dose. Treated cells also showed increased total glutathione and oxidized glutathione levels, increased superoxide dismutase activity, and increased hemoxygenase 1 expression. Treatment with E2 at 20 nM for 48 h resulted in a significant reduction in cell growth (0.35/1 migration rate) and decreased cell survival (<80%) as compared with controls. Survivin expression significantly increased at 24 h post treatment with 5, 10, or 20 nM; however, that of γ-H2AX increased only after survivin levels dropped and only at the 20 nM E2 dose. Minimal upregulation and splitting of caspase 9 was only evident in cells treated with 20 nM E2; no changes in caspase 3 expression were evident. Although Annexin V staining studies showed that E2 treatment did not induce apoptosis, scanning electron microscopy studies showed marked membrane blebbing at 20 nM/48 h of E2. These findings suggest that estrogen treatment disrupts intracellular iron metabolism and precipitates adverse effects concerning cell viability, membrane integrity, and growth potential.
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Estrogênios/metabolismo , Sobrecarga de Ferro/genética , Ferro/metabolismo , Apoptose/genética , Proteínas de Transporte de Cátions/genética , Pontos de Checagem do Ciclo Celular/genética , Proliferação de Células/genética , Sobrevivência Celular/genética , Histonas/genética , Humanos , Fator 1 Induzível por Hipóxia/genética , Sobrecarga de Ferro/metabolismo , Células MCF-7 , Estresse Oxidativo/genética , Superóxido Dismutase/genéticaRESUMO
OBJECTIVES: Reactive oxygen species (ROS), including superoxide (O2(â¢-)), play an important role in the biological effects of ionizing radiation. The human body has developed different antioxidant systems to defend against excessive levels of ROS. The aim of the present study is to investigate the redox status changes in the blood of radiologic technologists and compare these changes to control individuals. METHODS: We enrolled 60 medical workers: 20 occupationally exposed to ionizing radiation (all radiologic technologists), divided in three subgroups: conventional radiography (CR), computerized tomography (CT), and interventional radiography (IR) and 40 age- and gender-matched unexposed controls. Levels of O2(â¢-) and malondialdehyde (MDA) in blood were measured as an index of redox status, as were the activities of antioxidant enzymes superoxide dismutase (SOD) and catalase. Redox status was also assessed by measuring levels of reduced and oxidized glutathione (GSH, GSSG, respectively). RESULTS: Levels of O2(â¢-) and MDA, and SOD activity in the blood of IR and CT-exposed subjects were significantly higher than both the CR-exposed subjects and control individuals. However, there were no statistically significant differences in the levels of catalase, GSH and ratio of GSH/GSSG between exposed workers and control individuals. DISCUSSION: This study suggests that healthcare workers in CT and IR occupationally exposed to radiation have an elevated circulating redox status as compared to unexposed individuals.
Assuntos
Radiação Ionizante , Antioxidantes/metabolismo , Catalase/metabolismo , Feminino , Glutationa/metabolismo , Dissulfeto de Glutationa/metabolismo , Glutationa Peroxidase/metabolismo , Humanos , Masculino , Malondialdeído/metabolismo , Exposição Ocupacional/efeitos adversos , Oxirredução/efeitos da radiação , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/metabolismo , Superóxidos/metabolismoRESUMO
OBJECTIVE: Endothelial cell (EC) oxidative stress can lead to vascular dysfunction which is an underlying event in the development of cardiovascular disease (CVD). The lack of a potent and bioavailable anti-oxidant enzyme is a major challenge in studies on antioxidant therapy. The objective of this study is to determine whether copper/zinc superoxide dismutase (CuZnSOD or SOD1) after nanoformulation (nanoSOD) can effectively reduce EC oxidative stress and/or vascular inflammation in obesity. METHODS: Human aortic endothelial cells (HAECs) were treated with native- or nanoSOD for 6 h followed by treatment with linoleic acid (LA), a free fatty acid, for 6-24 h. To determine the in vivo relevance, the effectiveness of nanoSOD in reducing vascular cell activation was studied in a mouse model of diet-induced obesity. RESULTS: We noted that nanoSOD was more effectively taken up by ECs than native SOD. Western blot analysis further confirmed that the intracellular accumulation of SOD1 protein was greatly increased upon nanoSOD treatment. Importantly, nanoSOD pretreatment led to a significant decrease in LA-induced oxidative stress in ECs which was associated with a marked increase in SOD enzyme activity in ECs. In vivo studies showed a significant decrease in markers of EC/vascular cell activation and/or inflammation in visceral adipose tissue (VAT), thoracic aorta, and heart collected from nanoSOD-treated mice compared to obese control mice. Interestingly, the expression of metallothionein 2, an antioxidant gene was significantly increased in nanoSOD-treated mice. CONCLUSION: Our data show that nanoSOD is very effective in delivering active SOD to ECs and in reducing EC oxidative stress. Our data also demonstrate that nanoSOD will be a useful tool to reduce vascular cell activation in VAT and aorta in obesity which, in turn, can protect against obesity-associated CVD, in particular, hypertension.