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Trent E. Tipple - One of the best experts on this subject based on the ideXlab platform.
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abstract p016 sex differential effects of hyperoxia and thioredoxin reductase 1 inhibition on the kidney endothelin 1 system
Hypertension, 2020Co-Authors: Carmen De Miguel, Katelyn Dunigan, Aleena George, Sara N Biswal, Abigayle Kraus, Trent E. TippleAbstract:The vasoactive peptide endothelin-1 (ET-1) is critical in lung and kidney injury. Notably, renal damageand hyperoxia-induced lung disease are more prevalent in males than females. Aurothioglucose (...
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Thioredoxin Reductase-1 Inhibition Augments Endogenous Glutathione-Dependent Antioxidant Responses in Experimental Bronchopulmonary Dysplasia.
Oxidative Medicine and Cellular Longevity, 2019Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Rachael Wood, Lynette K. Rogers, Trent E. TippleAbstract:Background. Aurothioglucose- (ATG-) mediated inhibition of thioredoxin reductase-1 (TXNRD1) improves alveolarization in experimental murine bronchopulmonary dysplasia (BPD). Glutathione (GSH) mediates susceptibility to neonatal and adult oxidative lung injury. We have previously shown that ATG attenuates hyperoxic lung injury and enhances glutathione- (GSH-) dependent antioxidant defenses in adult mice. Hypothesis. The present studies evaluated the effects of TXNRD1 inhibition on GSH-dependent antioxidant defenses in newborn mice in vivo and lung epithelia in vitro. Methods. Newborn mice received intraperitoneal ATG or saline prior to room air or 85% hyperoxia exposure. Glutamate-cysteine ligase (GCL) catalytic (Gclc) and modifier (Gclm) mRNA levels, total GSH levels, total GSH peroxidase (GPx) activity, and Gpx2 expression were determined in lung homogenates. In vitro, murine transformed club cells (mtCCs) were treated with the TXNRD1 inhibitor auranofin (AFN) or vehicle in the presence or absence of the GCL inhibitor buthionine sulfoximine (BSO). Results. In vivo, ATG enhanced hyperoxia-induced increases in Gclc mRNA levels, total GSH contents, and GPx activity. In vitro, AFN increased Gclm mRNA levels, intracellular and extracellular GSH levels, and GPx activity. BSO prevented AFN-induced increases in GSH levels. Conclusions. Our data are consistent with a model in which TXNRD1 inhibition augments hyperoxia-induced GSH-dependent antioxidant responses in neonatal mice. Discrepancies between in vivo and in vitro results highlight the need for methodologies that permit accurate assessments of the GSH system at the single-cell level.
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Aurothioglucose does not improve alveolarization or elicit sustained nrf2 activation in c57bl 6 models of bronchopulmonary dysplasia
American Journal of Physiology-lung Cellular and Molecular Physiology, 2018Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Lynette K. Rogers, Changchun Ren, Tamas Jilling, Trent E. TippleAbstract:We previously showed that the thioredoxin reductase-1 (TrxR1) inhibitor Aurothioglucose (ATG) improves alveolarization in hyperoxia-exposed newborn C3H/HeN mice. Our data supported a mechanism by w...
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the thioredoxin reductase 1 inhibitor Aurothioglucose enhances glutathione dependent antioxidant responses in a murine model of bronchopulmonary dysplasia
Free Radical Biology and Medicine, 2017Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Rachael Tindell, Trent E. TippleAbstract:Introduction Therapeutic oxygen contributes to the development of bronchopulmonary dysplasia (BPD). The thioredoxin reductase-1 (TrxR1) inhibitor Aurothioglucose (ATG) is protective in our murine BPD model. Improved lung structure in ATG-treated newborn mice was associated with enhanced nuclear factor E2-related factor 2 (Nrf2) activation. TrxR1 inhibitors also enhance glutathione (GSH) levels in models of acute lung injury. Hypothesis The present studies tested the hypothesis that ATG enhances GSH-mediated antioxidant defenses in neonatal lungs. Methods Newborn C3H/HeN mice were dosed with 25mg/kg ATG or saline (SA) i.p. within 12 h of birth and exposed to room air (RA) or hyperoxia (HO, 85% O2). Lung glutamate-cysteine ligase catalytic (Gclc) and modifier (Gclm) subunit levels were assessed by qRT-PCR. Total and oxidized GSH (GSSG) levels were determined by Tietze recycling assay. Data (n=3-6) were analyzed by 2-way ANOVA with Tukey’s post hoc. Results At 24 h, Gclm levels were not different between groups. Gclc levels were 2-fold greater in HO:ATG vs RA:SA (p=0.0456). By 72 h, independent effects of ATG and hyperoxia were detected for Gclm and Gclc. Gclm levels were 1.6-fold higher in HO:ATG vs RA:SA (p=0.0016) and Gclc levels were 2.4 fold greater in HO:ATG vs RA:SA (p=0.0182). At 72 h, total GSH was 1.3 fold greater in HO:SA (p=0.0206) and 1.7 fold greater in HO:ATG (p=0.0002) vs RA:SA. Total GSH levels in HO:ATG lungs were 1.3 fold higher than RA:ATG (p Conclusions In HO-exposed newborn pups, ATG enhanced Gclm, Gclc and lung GSH levels. Given the importance of GSH-mediated antioxidant responses in protection against HO, we speculate that enhanced de novo GSH synthesis may be a mechanism by which ATG improves lung development in our model. TrxR1 inhibitors such as ATG, which increase Nrf2-dependent responses and enhance GSH-mediated antioxidant responses, may represent a novel therapy to decrease BPD development.
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synergistic effects of Aurothioglucose and hyperoxia in neonatal hyperoxic lung injury
Free Radical Biology and Medicine, 2016Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Rachael Tindell, Trent E. TippleAbstract:Oxygen toxicity and antioxidant deficiencies contribute to the development of bronchopulmonary dysplasia (BPD) in preterm infants. We recently demonstrated that a single injection of Aurothioglucose (ATG), a thioredoxin reductase-1 (TrxR1) inhibitor, attenuated lung injury and increased Nrf2 activation in a C3H/HeN mouse model of BPD. Nrf2 activation was greatest in ATG-treated hyperoxia-exposed mice. Additional unpublished data in vivo suggests that ATG treatment also enhances endogenous glutathione (GSH) levels. Hyperoxic sensitivity in newborn mice is strain-dependent, therefore we first tested the hypothesis that the protective effects of ATG are strain independent and dose-dependent. C57Bl/6 mice were pre-treated with a single dose of 0, 25 or 50 mg/kg ATG at 4d of life and were subsequently exposed to room air (RA) or hyperoxia (85% O2) from 4- 14d. Histologic analyses of formalin fixed lungs were performed to evaluate lung development. Our data revealed that hyperoxic exposure resulted in lung developmental abnormalities in saline-treated mice. Preliminary qualitative analyses of H&E-stained lung sections revealed dose-dependent attenuation of ATG treatment on hyperoxia-induced lung development with the most robust protective effect observed in mice pretreated with 50 mg/kg ATG. Studies in vitro tested the hypothesis that ATG inhibits TrxR1, protects against hyperoxia-induced anti-proliferative effects in lung epithelial cells, and enhances cellular GSH levels. Murine transformed club cells (mtCCs) were treated with 0 or 5 μM ATG and were exposed RA or hyperoxia (85% O2) for 24 h. For each condition the following was determined: cell counts, TrxR1 activity via insulin-disulfide reductase assay, and glutathione levels via the glutathione reductase recycling assay. ATG treatment increased viable cell numbers in RA-exposed cells, which directly correlated with a 25% decrease in TrxR1 activity. In addition, ATG-treated hyperoxia-exposed cells TrxR1 activity was 50% less than in control-treated cells and was associated with enhanced total GSH levels. In summary, the effects of ATG on hyperoxia-induced lung developmental deficits are dose-dependent and strain-independent. In vitro, our data reveals synergism between ATG and hyperoxia similar our previous observations in C3H/HeN mice. The inverse correlation between the degree of TrxR1 inhibition and ATG-mediated protection elicited by the combination of ATG and hyperoxia suggests a threshold of TrxR1 inhibition necessary for the protective effects of ATG. Future studies will define dose-response characteristics and mechanisms by which ATG is protective in vivo and in vitro with the overall goal of optimizing TrxR1 inhibition as a viable therapeutic strategy to prevent BPD.
Stephanie B. Wall - One of the best experts on this subject based on the ideXlab platform.
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Aurothioglucose enhances proangiogenic pathway activation in lungs from room air and hyperoxia-exposed newborn mice.
American Journal of Physiology-lung Cellular and Molecular Physiology, 2020Co-Authors: Katelyn Dunigan-russell, Stephanie B. Wall, Vivian Y. Lin, Mary Silverberg, John P. Gotham, Teodora Nicola, Anusha Sridharan, John Snowball, Cassidy DelaneyAbstract:Bronchopulmonary dysplasia (BPD), a long-term respiratory morbidity of prematurity, is characterized by attenuated alveolar and vascular development. Supplemental oxygen and immature antioxidant defenses contribute to BPD development. Our group identified thioredoxin reductase-1 (TXNRD1) as a therapeutic target to prevent BPD. The present studies evaluated the impact of the TXNRD1 inhibitor Aurothioglucose (ATG) on pulmonary responses and gene expression in newborn C57BL/6 pups treated with saline or ATG (25 mg/kg ip) within 12 h of birth and exposed to room air (21% O2) or hyperoxia (>95% O2) for 72 h. Purified RNA from lung tissues was sequenced, and differential expression was evaluated. Hyperoxic exposure altered ~2,000 genes, including pathways involved in glutathione metabolism, intrinsic apoptosis signaling, and cell cycle regulation. The isolated effect of ATG treatment was limited primarily to genes that regulate angiogenesis and vascularization. In separate studies, pups were treated as described above and returned to room air until 14 days. Vascular density analyses were performed, and ANOVA indicated an independent effect of hyperoxia on vascular density and alveolar architecture at 14 days. Consistent with RNA-seq analyses, ATG significantly increased vascular density in room air, but not in hyperoxia-exposed pups. These findings provide insights into the mechanisms by which TXNRD1 inhibitors may enhance lung development.
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Thioredoxin Reductase-1 Inhibition Augments Endogenous Glutathione-Dependent Antioxidant Responses in Experimental Bronchopulmonary Dysplasia.
Oxidative Medicine and Cellular Longevity, 2019Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Rachael Wood, Lynette K. Rogers, Trent E. TippleAbstract:Background. Aurothioglucose- (ATG-) mediated inhibition of thioredoxin reductase-1 (TXNRD1) improves alveolarization in experimental murine bronchopulmonary dysplasia (BPD). Glutathione (GSH) mediates susceptibility to neonatal and adult oxidative lung injury. We have previously shown that ATG attenuates hyperoxic lung injury and enhances glutathione- (GSH-) dependent antioxidant defenses in adult mice. Hypothesis. The present studies evaluated the effects of TXNRD1 inhibition on GSH-dependent antioxidant defenses in newborn mice in vivo and lung epithelia in vitro. Methods. Newborn mice received intraperitoneal ATG or saline prior to room air or 85% hyperoxia exposure. Glutamate-cysteine ligase (GCL) catalytic (Gclc) and modifier (Gclm) mRNA levels, total GSH levels, total GSH peroxidase (GPx) activity, and Gpx2 expression were determined in lung homogenates. In vitro, murine transformed club cells (mtCCs) were treated with the TXNRD1 inhibitor auranofin (AFN) or vehicle in the presence or absence of the GCL inhibitor buthionine sulfoximine (BSO). Results. In vivo, ATG enhanced hyperoxia-induced increases in Gclc mRNA levels, total GSH contents, and GPx activity. In vitro, AFN increased Gclm mRNA levels, intracellular and extracellular GSH levels, and GPx activity. BSO prevented AFN-induced increases in GSH levels. Conclusions. Our data are consistent with a model in which TXNRD1 inhibition augments hyperoxia-induced GSH-dependent antioxidant responses in neonatal mice. Discrepancies between in vivo and in vitro results highlight the need for methodologies that permit accurate assessments of the GSH system at the single-cell level.
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Aurothioglucose does not improve alveolarization or elicit sustained nrf2 activation in c57bl 6 models of bronchopulmonary dysplasia
American Journal of Physiology-lung Cellular and Molecular Physiology, 2018Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Lynette K. Rogers, Changchun Ren, Tamas Jilling, Trent E. TippleAbstract:We previously showed that the thioredoxin reductase-1 (TrxR1) inhibitor Aurothioglucose (ATG) improves alveolarization in hyperoxia-exposed newborn C3H/HeN mice. Our data supported a mechanism by w...
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the thioredoxin reductase 1 inhibitor Aurothioglucose enhances glutathione dependent antioxidant responses in a murine model of bronchopulmonary dysplasia
Free Radical Biology and Medicine, 2017Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Rachael Tindell, Trent E. TippleAbstract:Introduction Therapeutic oxygen contributes to the development of bronchopulmonary dysplasia (BPD). The thioredoxin reductase-1 (TrxR1) inhibitor Aurothioglucose (ATG) is protective in our murine BPD model. Improved lung structure in ATG-treated newborn mice was associated with enhanced nuclear factor E2-related factor 2 (Nrf2) activation. TrxR1 inhibitors also enhance glutathione (GSH) levels in models of acute lung injury. Hypothesis The present studies tested the hypothesis that ATG enhances GSH-mediated antioxidant defenses in neonatal lungs. Methods Newborn C3H/HeN mice were dosed with 25mg/kg ATG or saline (SA) i.p. within 12 h of birth and exposed to room air (RA) or hyperoxia (HO, 85% O2). Lung glutamate-cysteine ligase catalytic (Gclc) and modifier (Gclm) subunit levels were assessed by qRT-PCR. Total and oxidized GSH (GSSG) levels were determined by Tietze recycling assay. Data (n=3-6) were analyzed by 2-way ANOVA with Tukey’s post hoc. Results At 24 h, Gclm levels were not different between groups. Gclc levels were 2-fold greater in HO:ATG vs RA:SA (p=0.0456). By 72 h, independent effects of ATG and hyperoxia were detected for Gclm and Gclc. Gclm levels were 1.6-fold higher in HO:ATG vs RA:SA (p=0.0016) and Gclc levels were 2.4 fold greater in HO:ATG vs RA:SA (p=0.0182). At 72 h, total GSH was 1.3 fold greater in HO:SA (p=0.0206) and 1.7 fold greater in HO:ATG (p=0.0002) vs RA:SA. Total GSH levels in HO:ATG lungs were 1.3 fold higher than RA:ATG (p Conclusions In HO-exposed newborn pups, ATG enhanced Gclm, Gclc and lung GSH levels. Given the importance of GSH-mediated antioxidant responses in protection against HO, we speculate that enhanced de novo GSH synthesis may be a mechanism by which ATG improves lung development in our model. TrxR1 inhibitors such as ATG, which increase Nrf2-dependent responses and enhance GSH-mediated antioxidant responses, may represent a novel therapy to decrease BPD development.
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synergistic effects of Aurothioglucose and hyperoxia in neonatal hyperoxic lung injury
Free Radical Biology and Medicine, 2016Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Rachael Tindell, Trent E. TippleAbstract:Oxygen toxicity and antioxidant deficiencies contribute to the development of bronchopulmonary dysplasia (BPD) in preterm infants. We recently demonstrated that a single injection of Aurothioglucose (ATG), a thioredoxin reductase-1 (TrxR1) inhibitor, attenuated lung injury and increased Nrf2 activation in a C3H/HeN mouse model of BPD. Nrf2 activation was greatest in ATG-treated hyperoxia-exposed mice. Additional unpublished data in vivo suggests that ATG treatment also enhances endogenous glutathione (GSH) levels. Hyperoxic sensitivity in newborn mice is strain-dependent, therefore we first tested the hypothesis that the protective effects of ATG are strain independent and dose-dependent. C57Bl/6 mice were pre-treated with a single dose of 0, 25 or 50 mg/kg ATG at 4d of life and were subsequently exposed to room air (RA) or hyperoxia (85% O2) from 4- 14d. Histologic analyses of formalin fixed lungs were performed to evaluate lung development. Our data revealed that hyperoxic exposure resulted in lung developmental abnormalities in saline-treated mice. Preliminary qualitative analyses of H&E-stained lung sections revealed dose-dependent attenuation of ATG treatment on hyperoxia-induced lung development with the most robust protective effect observed in mice pretreated with 50 mg/kg ATG. Studies in vitro tested the hypothesis that ATG inhibits TrxR1, protects against hyperoxia-induced anti-proliferative effects in lung epithelial cells, and enhances cellular GSH levels. Murine transformed club cells (mtCCs) were treated with 0 or 5 μM ATG and were exposed RA or hyperoxia (85% O2) for 24 h. For each condition the following was determined: cell counts, TrxR1 activity via insulin-disulfide reductase assay, and glutathione levels via the glutathione reductase recycling assay. ATG treatment increased viable cell numbers in RA-exposed cells, which directly correlated with a 25% decrease in TrxR1 activity. In addition, ATG-treated hyperoxia-exposed cells TrxR1 activity was 50% less than in control-treated cells and was associated with enhanced total GSH levels. In summary, the effects of ATG on hyperoxia-induced lung developmental deficits are dose-dependent and strain-independent. In vitro, our data reveals synergism between ATG and hyperoxia similar our previous observations in C3H/HeN mice. The inverse correlation between the degree of TrxR1 inhibition and ATG-mediated protection elicited by the combination of ATG and hyperoxia suggests a threshold of TrxR1 inhibition necessary for the protective effects of ATG. Future studies will define dose-response characteristics and mechanisms by which ATG is protective in vivo and in vitro with the overall goal of optimizing TrxR1 inhibition as a viable therapeutic strategy to prevent BPD.
Katelyn Dunigan - One of the best experts on this subject based on the ideXlab platform.
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abstract p016 sex differential effects of hyperoxia and thioredoxin reductase 1 inhibition on the kidney endothelin 1 system
Hypertension, 2020Co-Authors: Carmen De Miguel, Katelyn Dunigan, Aleena George, Sara N Biswal, Abigayle Kraus, Trent E. TippleAbstract:The vasoactive peptide endothelin-1 (ET-1) is critical in lung and kidney injury. Notably, renal damageand hyperoxia-induced lung disease are more prevalent in males than females. Aurothioglucose (...
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Thioredoxin Reductase-1 Inhibition Augments Endogenous Glutathione-Dependent Antioxidant Responses in Experimental Bronchopulmonary Dysplasia.
Oxidative Medicine and Cellular Longevity, 2019Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Rachael Wood, Lynette K. Rogers, Trent E. TippleAbstract:Background. Aurothioglucose- (ATG-) mediated inhibition of thioredoxin reductase-1 (TXNRD1) improves alveolarization in experimental murine bronchopulmonary dysplasia (BPD). Glutathione (GSH) mediates susceptibility to neonatal and adult oxidative lung injury. We have previously shown that ATG attenuates hyperoxic lung injury and enhances glutathione- (GSH-) dependent antioxidant defenses in adult mice. Hypothesis. The present studies evaluated the effects of TXNRD1 inhibition on GSH-dependent antioxidant defenses in newborn mice in vivo and lung epithelia in vitro. Methods. Newborn mice received intraperitoneal ATG or saline prior to room air or 85% hyperoxia exposure. Glutamate-cysteine ligase (GCL) catalytic (Gclc) and modifier (Gclm) mRNA levels, total GSH levels, total GSH peroxidase (GPx) activity, and Gpx2 expression were determined in lung homogenates. In vitro, murine transformed club cells (mtCCs) were treated with the TXNRD1 inhibitor auranofin (AFN) or vehicle in the presence or absence of the GCL inhibitor buthionine sulfoximine (BSO). Results. In vivo, ATG enhanced hyperoxia-induced increases in Gclc mRNA levels, total GSH contents, and GPx activity. In vitro, AFN increased Gclm mRNA levels, intracellular and extracellular GSH levels, and GPx activity. BSO prevented AFN-induced increases in GSH levels. Conclusions. Our data are consistent with a model in which TXNRD1 inhibition augments hyperoxia-induced GSH-dependent antioxidant responses in neonatal mice. Discrepancies between in vivo and in vitro results highlight the need for methodologies that permit accurate assessments of the GSH system at the single-cell level.
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Aurothioglucose does not improve alveolarization or elicit sustained nrf2 activation in c57bl 6 models of bronchopulmonary dysplasia
American Journal of Physiology-lung Cellular and Molecular Physiology, 2018Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Lynette K. Rogers, Changchun Ren, Tamas Jilling, Trent E. TippleAbstract:We previously showed that the thioredoxin reductase-1 (TrxR1) inhibitor Aurothioglucose (ATG) improves alveolarization in hyperoxia-exposed newborn C3H/HeN mice. Our data supported a mechanism by w...
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the thioredoxin reductase 1 inhibitor Aurothioglucose enhances glutathione dependent antioxidant responses in a murine model of bronchopulmonary dysplasia
Free Radical Biology and Medicine, 2017Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Rachael Tindell, Trent E. TippleAbstract:Introduction Therapeutic oxygen contributes to the development of bronchopulmonary dysplasia (BPD). The thioredoxin reductase-1 (TrxR1) inhibitor Aurothioglucose (ATG) is protective in our murine BPD model. Improved lung structure in ATG-treated newborn mice was associated with enhanced nuclear factor E2-related factor 2 (Nrf2) activation. TrxR1 inhibitors also enhance glutathione (GSH) levels in models of acute lung injury. Hypothesis The present studies tested the hypothesis that ATG enhances GSH-mediated antioxidant defenses in neonatal lungs. Methods Newborn C3H/HeN mice were dosed with 25mg/kg ATG or saline (SA) i.p. within 12 h of birth and exposed to room air (RA) or hyperoxia (HO, 85% O2). Lung glutamate-cysteine ligase catalytic (Gclc) and modifier (Gclm) subunit levels were assessed by qRT-PCR. Total and oxidized GSH (GSSG) levels were determined by Tietze recycling assay. Data (n=3-6) were analyzed by 2-way ANOVA with Tukey’s post hoc. Results At 24 h, Gclm levels were not different between groups. Gclc levels were 2-fold greater in HO:ATG vs RA:SA (p=0.0456). By 72 h, independent effects of ATG and hyperoxia were detected for Gclm and Gclc. Gclm levels were 1.6-fold higher in HO:ATG vs RA:SA (p=0.0016) and Gclc levels were 2.4 fold greater in HO:ATG vs RA:SA (p=0.0182). At 72 h, total GSH was 1.3 fold greater in HO:SA (p=0.0206) and 1.7 fold greater in HO:ATG (p=0.0002) vs RA:SA. Total GSH levels in HO:ATG lungs were 1.3 fold higher than RA:ATG (p Conclusions In HO-exposed newborn pups, ATG enhanced Gclm, Gclc and lung GSH levels. Given the importance of GSH-mediated antioxidant responses in protection against HO, we speculate that enhanced de novo GSH synthesis may be a mechanism by which ATG improves lung development in our model. TrxR1 inhibitors such as ATG, which increase Nrf2-dependent responses and enhance GSH-mediated antioxidant responses, may represent a novel therapy to decrease BPD development.
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synergistic effects of Aurothioglucose and hyperoxia in neonatal hyperoxic lung injury
Free Radical Biology and Medicine, 2016Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Rachael Tindell, Trent E. TippleAbstract:Oxygen toxicity and antioxidant deficiencies contribute to the development of bronchopulmonary dysplasia (BPD) in preterm infants. We recently demonstrated that a single injection of Aurothioglucose (ATG), a thioredoxin reductase-1 (TrxR1) inhibitor, attenuated lung injury and increased Nrf2 activation in a C3H/HeN mouse model of BPD. Nrf2 activation was greatest in ATG-treated hyperoxia-exposed mice. Additional unpublished data in vivo suggests that ATG treatment also enhances endogenous glutathione (GSH) levels. Hyperoxic sensitivity in newborn mice is strain-dependent, therefore we first tested the hypothesis that the protective effects of ATG are strain independent and dose-dependent. C57Bl/6 mice were pre-treated with a single dose of 0, 25 or 50 mg/kg ATG at 4d of life and were subsequently exposed to room air (RA) or hyperoxia (85% O2) from 4- 14d. Histologic analyses of formalin fixed lungs were performed to evaluate lung development. Our data revealed that hyperoxic exposure resulted in lung developmental abnormalities in saline-treated mice. Preliminary qualitative analyses of H&E-stained lung sections revealed dose-dependent attenuation of ATG treatment on hyperoxia-induced lung development with the most robust protective effect observed in mice pretreated with 50 mg/kg ATG. Studies in vitro tested the hypothesis that ATG inhibits TrxR1, protects against hyperoxia-induced anti-proliferative effects in lung epithelial cells, and enhances cellular GSH levels. Murine transformed club cells (mtCCs) were treated with 0 or 5 μM ATG and were exposed RA or hyperoxia (85% O2) for 24 h. For each condition the following was determined: cell counts, TrxR1 activity via insulin-disulfide reductase assay, and glutathione levels via the glutathione reductase recycling assay. ATG treatment increased viable cell numbers in RA-exposed cells, which directly correlated with a 25% decrease in TrxR1 activity. In addition, ATG-treated hyperoxia-exposed cells TrxR1 activity was 50% less than in control-treated cells and was associated with enhanced total GSH levels. In summary, the effects of ATG on hyperoxia-induced lung developmental deficits are dose-dependent and strain-independent. In vitro, our data reveals synergism between ATG and hyperoxia similar our previous observations in C3H/HeN mice. The inverse correlation between the degree of TrxR1 inhibition and ATG-mediated protection elicited by the combination of ATG and hyperoxia suggests a threshold of TrxR1 inhibition necessary for the protective effects of ATG. Future studies will define dose-response characteristics and mechanisms by which ATG is protective in vivo and in vitro with the overall goal of optimizing TrxR1 inhibition as a viable therapeutic strategy to prevent BPD.
Arne Holmgren - One of the best experts on this subject based on the ideXlab platform.
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The production of reactive oxygen species enhanced with the reduction of menadione by active thioredoxin reductase
Metallomics, 2019Co-Authors: Xin Zuo, Ping Cheng, Xiaoyuan Ren, Shibo Sun, Arne HolmgrenAbstract:Cytosolic thioredoxin reductase (TXNRD1) is an important selenoprotein that participates in the reduction of thioredoxin and many other redox-related substrates. The enhancement of ROS production to cause cancer cell death is an effective anticancer strategy. Herein, we found that menadione substantially increased ROS generation via interaction with TXNRD1. To elucidate the mechanism behind this, various TXNRD1 mutant proteins were used to investigate the relationship between ROS production and the reaction between enzymes and menadione. A mutation at the C-terminal active site -GCUG of TXNRD1 to -GSSG or -GC, or the N-terminal active site C59S, C64S, or the deletion of the C-terminal 16 amino acid residues caused the loss of TXNRD1 activity needed for the reduction of menadione and therefore resulted in the loss of the ROS production ability of menadione. In contrast, the mutation of -GCUG to -GCCG resulted in an increase in the TXNRD1 activity towards the reduction of menadione, thus leading to an increase in ROS production. The co-treatment of the TXNRD1 inhibitor Aurothioglucose and menadione could significantly alleviate the efficiency of ROS generation in vitro and increase the viability of A549 cells. Moreover, menadione could be reduced by the glutathione system and caused ROS production with less efficiency. These results demonstrate that TXNRD1 can serve as an effective source to generate ROS, which may provide a novel anticancer method based on the use of menadione.
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glutathione and glutaredoxin act as a backup of human thioredoxin reductase 1 to reduce thioredoxin 1 preventing cell death by Aurothioglucose
Journal of Biological Chemistry, 2012Co-Authors: Huihui Zhang, Arne HolmgrenAbstract:Thioredoxin reductase 1 (TrxR1) in cytosol is the only known reductant of oxidized thioredoxin 1 (Trx1) in vivo so far. We and others found that Aurothioglucose (ATG), a well known active-site inhibitor of TrxR1, inhibited TrxR1 activity in HeLa cell cytosol but had no effect on the viability of the cells. Using a redox Western blot analysis, no change was observed in redox state of Trx1, which was mainly fully reduced with five sulfhydryl groups. In contrast, auranofin killed cells and oxidized Trx1, also targeting mitochondrial TrxR2 and Trx2. Combining ATG with ebselen gave a strong synergistic effect, leading to Trx1 oxidation, reactive oxygen species accumulation, and cell death. We hypothesized that there should exist a backup system to reduce Trx1 when only TrxR1 activity was lost. Our results showed that physiological concentrations of glutathione, NADPH, and glutathione reductase reduced Trx1 in vitro and that the reaction was strongly stimulated by glutaredoxin1. Simultaneous depletion of TrxR activity by ATG and glutathione by buthionine sulfoximine led to overoxidation of Trx1 and loss of HeLa cell viability. In conclusion, the glutaredoxin system and glutathione have a backup role to keep Trx1 reduced in cells with loss of TrxR1 activity. Monitoring the redox state of Trx1 shows that cell death occurs when Trx1 is oxidized, followed by general protein oxidation catalyzed by the disulfide form of thioredoxin.
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Physiological functions of thioredoxin and thioredoxin reductase.
European journal of biochemistry, 2000Co-Authors: Elias S.j. Arnér, Arne HolmgrenAbstract:Thioredoxin, thioredoxin reductase and NADPH, the thioredoxin system, is ubiquitous from Archea to man. Thioredoxins, with a dithiol/disulfide active site (CGPC) are the major cellular protein disulfide reductases; they therefore also serve as electron donors for enzymes such as ribonucleotide reductases, thioredoxin peroxidases (peroxiredoxins) and methionine sulfoxide reductases. Glutaredoxins catalyze glutathione-disulfide oxidoreductions overlapping the functions of thioredoxins and using electrons from NADPH via glutathione reductase. Thioredoxin isoforms are present in most organisms and mitochondria have a separate thioredoxin system. Plants have chloroplast thioredoxins, which via ferredoxin-thioredoxin reductase regulates photosynthetic enzymes by light. Thioredoxins are critical for redox regulation of protein function and signaling via thiol redox control. A growing number of transcription factors including NF-kappaB or the Ref-1-dependent AP1 require thioredoxin reduction for DNA binding. The cytosolic mammalian thioredoxin, lack of which is embryonically lethal, has numerous functions in defense against oxidative stress, control of growth and apoptosis, but is also secreted and has co-cytokine and chemokine activities. Thioredoxin reductase is a specific dimeric 70-kDa flavoprotein in bacteria, fungi and plants with a redox active site disulfide/dithiol. In contrast, thioredoxin reductases of higher eukaryotes are larger (112-130 kDa), selenium-dependent dimeric flavoproteins with a broad substrate specificity that also reduce nondisulfide substrates such as hydroperoxides, vitamin C or selenite. All mammalian thioredoxin reductase isozymes are homologous to glutathione reductase and contain a conserved C-terminal elongation with a cysteine-selenocysteine sequence forming a redox-active selenenylsulfide/selenolthiol active site and are inhibited by goldthioglucose (Aurothioglucose) and other clinically used drugs.
Lynette K. Rogers - One of the best experts on this subject based on the ideXlab platform.
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Thioredoxin Reductase-1 Inhibition Augments Endogenous Glutathione-Dependent Antioxidant Responses in Experimental Bronchopulmonary Dysplasia.
Oxidative Medicine and Cellular Longevity, 2019Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Rachael Wood, Lynette K. Rogers, Trent E. TippleAbstract:Background. Aurothioglucose- (ATG-) mediated inhibition of thioredoxin reductase-1 (TXNRD1) improves alveolarization in experimental murine bronchopulmonary dysplasia (BPD). Glutathione (GSH) mediates susceptibility to neonatal and adult oxidative lung injury. We have previously shown that ATG attenuates hyperoxic lung injury and enhances glutathione- (GSH-) dependent antioxidant defenses in adult mice. Hypothesis. The present studies evaluated the effects of TXNRD1 inhibition on GSH-dependent antioxidant defenses in newborn mice in vivo and lung epithelia in vitro. Methods. Newborn mice received intraperitoneal ATG or saline prior to room air or 85% hyperoxia exposure. Glutamate-cysteine ligase (GCL) catalytic (Gclc) and modifier (Gclm) mRNA levels, total GSH levels, total GSH peroxidase (GPx) activity, and Gpx2 expression were determined in lung homogenates. In vitro, murine transformed club cells (mtCCs) were treated with the TXNRD1 inhibitor auranofin (AFN) or vehicle in the presence or absence of the GCL inhibitor buthionine sulfoximine (BSO). Results. In vivo, ATG enhanced hyperoxia-induced increases in Gclc mRNA levels, total GSH contents, and GPx activity. In vitro, AFN increased Gclm mRNA levels, intracellular and extracellular GSH levels, and GPx activity. BSO prevented AFN-induced increases in GSH levels. Conclusions. Our data are consistent with a model in which TXNRD1 inhibition augments hyperoxia-induced GSH-dependent antioxidant responses in neonatal mice. Discrepancies between in vivo and in vitro results highlight the need for methodologies that permit accurate assessments of the GSH system at the single-cell level.
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Aurothioglucose does not improve alveolarization or elicit sustained nrf2 activation in c57bl 6 models of bronchopulmonary dysplasia
American Journal of Physiology-lung Cellular and Molecular Physiology, 2018Co-Authors: Stephanie B. Wall, Katelyn Dunigan, Lynette K. Rogers, Changchun Ren, Tamas Jilling, Trent E. TippleAbstract:We previously showed that the thioredoxin reductase-1 (TrxR1) inhibitor Aurothioglucose (ATG) improves alveolarization in hyperoxia-exposed newborn C3H/HeN mice. Our data supported a mechanism by w...
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thioredoxin reductase inhibition attenuates neonatal hyperoxic lung injury and enhances nuclear factor e2 related factor 2 activation
American Journal of Respiratory Cell and Molecular Biology, 2016Co-Authors: Stephanie B. Wall, Lynette K. Rogers, Morgan L. Locy, Markus Velten, Changchun Ren, Cynthia L Hill, Trent E. TippleAbstract:Oxygen toxicity and antioxidant deficiencies contribute to the development of bronchopulmonary dysplasia. Aurothioglucose (ATG) and auranofin potently inhibit thioredoxin reductase-1 (TrxR1), and TrxR1 disruption activates nuclear factor E2-related factor 2 (Nrf2), a regulator of endogenous antioxidant responses. We have shown previously that ATG safely and effectively prevents lung injury in adult murine models, likely via Nrf2-dependent mechanisms. The current studies tested the hypothesis that ATG would attenuate hyperoxia-induced lung developmental deficits in newborn mice. Newborn C3H/HeN mice were treated with a single dose of ATG or saline within 12 hours of birth and were exposed to either room air or hyperoxia (85% O2). In hyperoxia, ATG potently inhibited TrxR1 activity in newborn murine lungs, attenuated decreases in body weight, increased the transcription of Nrf2-regulated genes nicotinamide adenine dinucleotide phosphate reduced quinone oxidoreductase-1 (NQO1) and heme oxygenase 1, and attenuated alterations in alveolar development. To determine the impact of TrxR1 inhibition on Nrf2 activation in vitro, murine alveolar epithelial-12 cells were treated with auranofin, which inhibited TrxR1 activity, enhanced Nrf2 nuclear levels, and increased NQO1 and heme oxygenase 1 transcription. Our novel data indicate that a single injection of the TrxR1 inhibitor ATG attenuates hyperoxia-induced alterations in alveolar development in newborn mice. Furthermore, our data support a model in which the effects of ATG treatment likely involve Nrf2 activation, which is consistent with our findings in other lung injury models. We conclude that TrxR1 represents a novel therapeutic target to prevent oxygen-mediated neonatal lung injury.
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the thioredoxin reductase 1 inhibitor Aurothioglucose attenuates lung injury and improves survival in a murine model of acute respiratory distress syndrome
Antioxidants & Redox Signaling, 2014Co-Authors: Rodney D Britt, Trent E. Tipple, Lynette K. Rogers, Morgan L. Locy, Markus VeltenAbstract:Abstract Aims: Inflammation and oxygen toxicity increase free radical production and contribute to the development of acute respiratory distress syndrome (ARDS), which is a significant cause of morbidity and mortality in intensive care patients. We have previously reported increased glutathione (GSH) levels in lung epithelial cells in vitro and attenuated adult murine hyperoxic lung injury in vivo after pharmacological thioredoxin reductase-1 (TrxR1) inhibition. Using a murine ARDS model, we tested the hypothesis that Aurothioglucose (ATG) treatment increases pulmonary GSH levels, attenuates lung injury, and decreases mortality in a GSH-dependent manner. Results: Adult mice received a single intratracheal dose of 0.375 μg/g lipopolysaccharide (LPS) 12 h before a single intraperitoneal injection of 25 mg/kg ATG. Control mice received intratracheal and/or intraperitoneal saline. Mice were then exposed to room air or hyperoxia (>95% O2). Lung injury was assessed by bronchoalveolar lavage protein concentratio...
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Thioredoxin-Related Mechanisms in Hyperoxic Lung Injury in Mice
2013Co-Authors: Stephen E. Welty, Lynette K. Rogers, Thomas N. Hansen, Young-eun Choi, James P. Kehrer, Charles V. SmithAbstract:Reduction of glutathione disulfide (GSSG) to glutathione (GSH) by glutathione reductase (GR) enhances the efficiency of GSH-dependent antioxidant activities. However, GR-deficient (a1Neu) mice are less susceptible to acute lung injury from continuous exposure to. 95 % O 2 (96 h: 6.9 6 0.1 g right lung/kg body versus room air 3.6 6 0.3) than are C3H/HeN control mice (10.6 6 1.3 versus 4.2 6 0.3, P, 0.001). a1Neu mice have greater hepatic thioredoxin (Trx)1 and Trx2 levels than do C3H/HeN mice, suggesting compensation for the absence of GR. a1Neu mice exposed to hyperoxia for 96 hours showed lower levels of inflammatory infiltrates in lungs than did similarly exposed C3H/HeN mice. Pretreatment with Aurothioglucose (ATG), a thioredoxin reductase (TrxR) inhibitor, exacerbated the effects of hyperoxia on lung injury in a1Neu mice (11.6 6 0.8, P, 0.001), but attenuated hyperoxic lung edema and inflammation in C3H/HeN mice (6.3 6 0.4, P, 0.001). No consistent alterations were observed in lung GSH contents or liver GSH or GSSG levels after ATG pretreatment. The data suggest that modulation of Trx/TrxR systems might provide therapeutically useful alterations of cellular resistance to oxidant stresses. The protective effects of ATG against hyperoxic lung injury could prove to be particularly useful therapeutically
