- Email: [email protected]
Zinc modulates dopamine-induced heme oxygenase 1 expression in human IMR-32 neuroblastoma cells
S164
A. Stepanova et al. / Free Radical Biology and Medicine 120 (2018) S45–S166
4
Cell Biology Program, The Hospital for Sick Children, Toronto, ON, Canada 5 Department of Molecular Biology and Biotechnology, The University of Sheffield, Sheffield, UK 6 Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Oeiras, Portugal 7 CNR Institute of Molecular Biology and Pathology, Rome, Italy
Cytochrome bd is a prokaryotic respiratory terminal oxidase phylogenetically unrelated to haem-copper oxidases. In addition to sustaining energy metabolism, this enzyme is implicated in bacterial protection against oxidative and nitrosative stress. Cytochrome bd is only found in prokaryotes and, as it promotes virulence in several pathogenic bacteria, it is currently recognized as a potential drug target. Escherichia coli and other microbes, including pathogens, colonize the human gut, that is known to be rich in hydrogen sulfide (H2S), while being a major site of reactive nitrogen species production. Depending on its concentration, H2S can have beneficial or detrimental effects in both prokaryotes and eukaryotes. Among the most well known toxic effects of H2S is the inhibition of cellular respiration due to binding to mitochondrial heme copper-type cytochrome c oxidase. Interestingly, working on E. coli mutant strains and purified proteins, we found that bd-type oxidases are insensitive to H2S inhibition, thereby promoting bacterial respiration and growth in the presence of high levels of H2S. The potential impact of this discovery on human (patho)physiology and its possible applications in the field of infectious diseases will be discussed.
E-mail address: [email protected] http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.538
P-392
( ) Epicatechin modulates fatpads changes associated to a high-fatdiet in mice Monica Galleano 1,2,3, Ezequiel Hid 1,2,3, Maria C. Litterio 1,2,3, Jorge E. Toblli 1,2,3, Barbara Piotrkowski 1,2,3, Laura Fischerman 1,2,3, Paula D. Prince 1,2,3, Juana I. Mosse 1,2,3, Cesar G. Fraga 1,2,3 1
Universidad de Buenos Aires, Facultad de Farmacia y Bioquímica, Fisicoquímica, Buenos Aires, Argentina 2 CONICET-Universidad de Buenos Aires, Instituto de Bioquímica y Medicina Molecular (IBIMOL), Buenos Aires, Argentina 3 Laboratorio de Medicina Experimental, Hospital Aleman, Buenos Aires, Argentina
E-mail address: [email protected] Acknowledgements Support: PIP 11220170100585C (MG); UBACyT 20020160100132BA (CF); Packer-Wentz endowment (CF).
http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.539
P-393
Is autophagy involved in the response to oxidative stress of tobacco BY-2 cells overexpressing thioredoxin o1? O. López-Vidal 1, A. Jimenez 1, S. Sevilla 1, L.M. Sandalio 2, A. Ortiz-Espín (*)1 1
Dpt of Stress Biology and Plant Pathology, CEBAS-CSIC, Murcia, Spain 2 Dpt of Biochemistry, Cellular and Molecular Biology of Plants, EEZ-CSIC, Granada, Spain
Autophagy is a catabolic process for cellular remodeling and macromolecular recycling of cytoplasmic components and organelles. Interestingly, autophagy appears as a key mechanism to cope with adverse conditions by removing damaged cellular components, although less information exists about this process in plants. In this sense, little is known about the possible role of redox regulation through thiol oxidoreductases thioredoxins (Trxs), specifically the mitochondrial/nuclear Trxo1. We have previously described a protective role of pea thioredoxin (PsTrxo1) overexpressed in TBY-2 cells treated with H2O2, thus increasing their viability via antioxidants, delaying cell death. In this work, we analyze the possible implication of autophagy as a protective mechanism operating in the PsTrxo1 transformed TBY-2 cells. For this, together with cell viability, different markers such as autophagy-related proteins ATGs are analyzed by Western blot, in parallel to autolysosomes and autophagosomes formation by fluorescence microscopy. The results have shown significant changes in ATG4 and ATG8 contents as well as presence of both lytic structures in H2O2-treated overexpressing PsTrxo1 cells, which suggest that Trxo1 may be also involved in the redox control of some autophagy components.
E-mail address: [email protected] http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.540
P-394 This work focuses on the effects of (-)-epicatechin (EC) on fat pads changes induced by high-fat (HF) diet. C57BL/6 male mice were divided in 4 groups receiving: control diet (10% calories from fat), HF diet (60% of calories), and these diets added with 20 mg EC/kg bw for 15 w. Even when caloric intake was not different among groups, HF increased mice weight. In white fat pads, inflammation (IL-6 and iNOS expression) and oxidative stress (gp91phox expression and MDA content) increases were associated with fat pad expansions. Adipocyte size was not different in HF respect to C group. On the other hand, HF diet induced expansion of aortic perivascular adipose tissue (aPVAT), with an increase of adipocyte size. Adipocytes/area were significantly lower in HF respect to C group. Changes in vacuolization patterns suggest a shift to white adipocyte phenotype, confirmed by a lower UCP-1 expression. All the observed changes in aPVAT were attenuated by EC. Since aPVAT dysfunction is relevant for hypertension and atherosclerosis, the effects of EC support an alternative nutritional strategy to avoid the progression of vascular dysfunction associated to the development of obesity.
Zinc modulates dopamine-induced heme oxygenase 1 expression in human IMR-32 neuroblastoma cells Zachary Kaufman 1, Gabriela Salvador 2, Patricia I. Oteiza 1 1
Departments of Nutrition and Environmental Toxicology, University of California, Davis, CA, USA 2 Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB-UNSCONICET). Bahía Blanca, Argentina
Zinc (Zn) plays a key role in the modulation of neuronal redox status. The present work tested the hypothesis that Zn is necessary for the neuronal defence response against dopamine (DA)-induced oxidative stress, in particular for the upregulation of heme oxigenase 1 (HMOX1).
A. Stepanova et al. / Free Radical Biology and Medicine 120 (2018) S45–S166
Human IMR-32 neuroblastoma cells exposed to a high DA (100 mM) concentration under a condition of Zn deficiency (ZD) showed an impaired capacity to upregulate HMOX1 when measured as mRNA and protein levels. This was reverted in Zn supplemented (ZS) cells. ZD decreased the capacity of IMR-32 cells to upregulate Nrf-2 and ERK1/2 in response to DA. However, chemical inhibition of ERK1/2 in ZS cells did not affect DA-mediated HMOX1 expression. BACH1 is a transcriptional repressor, which is central to the regulation of HMOX1 induction. Bach-1 levels were high in total homogenates and nuclear fraction from ZD cells stimulated with DA compared to ZS cells. BACH1 degradation was impaired in ZD compared to ZS cells, as evaluated by decreased levels of BACH1 ubiquitination upon DA treatment. Results show that Zn is necessary for the neuronal upregulation of HMOX1 upon a DA challenge, which is in part due to a requirement of Zn for the ubiquitin-mediated degradation of BACH1.
E-mail address: [email protected] http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.541
P-395
S165
3
Laboratory of Medicinal Chemistry, University of Antwerp, Belgium Sektion Nephrologie, Medizinische Klinik und Poliklinik III, Universitätsklinikum Carl Gustav Carus, Dresden, Germany 5 Methusalem program; Ghent University, Ghent, Belgium 4
Ferroptosis is an iron dependent form of necrosis, involved in different pathologies such as kidney failure and neurodegenerative diseases. Glutathione peroxidase 4 (GPX4) is the central enzyme protecting the cell from ferroptosis. High-throughput screening led to the discovery of ferrostatin-1 (Fer-1) as a potent in vitro inhibitor of ferroptosis. In vivo however, the molecule suffers from instability. Therefore, we developed ferrostatin-analogues with improved solubility and in vivo stability. To verify their in vivo efficacy, we established an acute mouse model of iron poisoning-induced organ injury. Both an excess of the natural ferroptosis inhibitor vitamin E in the feed and overexpression of GPX4 have a protective effect on this iron poisoning model, indicating it is a valuable experimental model for ferroptosis-induced organ injury. Interestingly, our novel ferrostatin analogues effectively decrease several markers for organ injury after iron poisoning. Conclusively, 1) ferroptosis is a detrimental factor in acute iron poisoning, 2) our novel ferrostatin-analogues reduce the severity of organ injury due to acute iron poisoning, and 3) our third generation of ferrostatin-analogues might represent potential leads for the treatment of ferroptosis-driven pathologies.
Early life stress, stress-resilience/susceptibility and oxidative stress
E-mail address: [email protected]; Tom.Van [email protected]
Samina Salim, Hesong Liu, Fatin Atrooz
http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.543
College of Pharmacy, University of Houston, Houston, TX, USA P-397 Adverse experiences during early life contribute to the development of psychiatric conditions later in life. In fact, young children who experience stressful traumatic event(s) during early life, a sensitive developmental period, are considered highly vulnerable to psychiatric disorders in adult life. Interestingly, not all children who experience stressful events are equally at risk of developing later life psychiatric disorders. Some are resilient in spite of being exposed to the same risk factors, while others are susceptible. The mechanistic basis for resilience/susceptibility is not clear. Using an early life stress model of Sprague Dawley rats, the mechanisms governing this phenotype segregation were revealed. Depression-susceptible phenotype is associated with high oxidative stress, low antioxidant status, defective redox-sensitive Nrf2 transcription factor and hyperactive NF-KB transcription factor, selectively in the pre-frontal cortex (PFC) area of the brain. The resistant phenotype displayed low oxidative stress, high antioxidant status, activated Nrf2 and inactivated NF-KB pathways.
E-mail address: [email protected] http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.542
P-396
Novel third-generation ferrostatins protect against acute iron poisoning-induced organ injury 1,2
1,2
1
June Lee, Young-Sam Keum College of Pharmacy and Integrated Research Institute for Drug Development, Dongguk University, Goyang, Gyeonggi-do, Korea
Heme oxygenase-1 (HO-1) catalyzes the enzymatic degradation of heme to produce three anti-oxidant molecules: carbon monoxide (CO), ferrous ion (Fe2 þ ), and biliverdin. Induction of HO-1 is currently considered as a feasible strategy to treat oxidative stress-related diseases. Here, we report marliolide as a novel inducer of HO-1 in human normal keratinocyte HaCaT cells. Mechanism-based studies demonstrated that the induction of HO-1 by marliolide occurred through activation of NRF2/ARE via direct binding of marliolide to KEAP1. Structure-activity relationship revealed chemical moieties of marliolide critical for induction of HO-1, which renders a support for Michael reaction as a potential mechanism of action. Finally, we observed that marliolide significantly inhibited the papilloma formation in DMBA/TPA–induced mouse skin carcinogenesis model and this event was closely associated with lowering the formation of 8-OH-G and 4-HNE in vivo. Together, we provide the first evidence that marliolide might be effective against oxidative stress-related skin disorders.
E-mail address: [email protected]
3
Samya Van Coillie , Wannes Weyts , Lars Devisscher , Sam Hofmans 3, Sze Men Choi 1,2, Bartosz Wiernicki 1,2, Behrouz Hassannia 1,2, Andreas Linkermann 4, Koen Augustyns 3, Peter Vandenabeele 1,2,5, Tom Vanden Berghe (*)1,2 VIB Center for Inflammation Research, Gent, Belgium Department of Biomedical Molecular Biology; Ghent University, Gent, Belgium
2
Marliolide inhibits skin carcinogenesis by activating NRF2/ARE to induce heme oxygenase-1
http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.544
A. Stepanova et al. / Free Radical Biology and Medicine 120 (2018) S45–S166
4
Cell Biology Program, The Hospital for Sick Children, Toronto, ON, Canada 5 Department of Molecular Biology and Biotechnology, The University of Sheffield, Sheffield, UK 6 Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Oeiras, Portugal 7 CNR Institute of Molecular Biology and Pathology, Rome, Italy
Cytochrome bd is a prokaryotic respiratory terminal oxidase phylogenetically unrelated to haem-copper oxidases. In addition to sustaining energy metabolism, this enzyme is implicated in bacterial protection against oxidative and nitrosative stress. Cytochrome bd is only found in prokaryotes and, as it promotes virulence in several pathogenic bacteria, it is currently recognized as a potential drug target. Escherichia coli and other microbes, including pathogens, colonize the human gut, that is known to be rich in hydrogen sulfide (H2S), while being a major site of reactive nitrogen species production. Depending on its concentration, H2S can have beneficial or detrimental effects in both prokaryotes and eukaryotes. Among the most well known toxic effects of H2S is the inhibition of cellular respiration due to binding to mitochondrial heme copper-type cytochrome c oxidase. Interestingly, working on E. coli mutant strains and purified proteins, we found that bd-type oxidases are insensitive to H2S inhibition, thereby promoting bacterial respiration and growth in the presence of high levels of H2S. The potential impact of this discovery on human (patho)physiology and its possible applications in the field of infectious diseases will be discussed.
E-mail address: [email protected] http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.538
P-392
( ) Epicatechin modulates fatpads changes associated to a high-fatdiet in mice Monica Galleano 1,2,3, Ezequiel Hid 1,2,3, Maria C. Litterio 1,2,3, Jorge E. Toblli 1,2,3, Barbara Piotrkowski 1,2,3, Laura Fischerman 1,2,3, Paula D. Prince 1,2,3, Juana I. Mosse 1,2,3, Cesar G. Fraga 1,2,3 1
Universidad de Buenos Aires, Facultad de Farmacia y Bioquímica, Fisicoquímica, Buenos Aires, Argentina 2 CONICET-Universidad de Buenos Aires, Instituto de Bioquímica y Medicina Molecular (IBIMOL), Buenos Aires, Argentina 3 Laboratorio de Medicina Experimental, Hospital Aleman, Buenos Aires, Argentina
E-mail address: [email protected] Acknowledgements Support: PIP 11220170100585C (MG); UBACyT 20020160100132BA (CF); Packer-Wentz endowment (CF).
http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.539
P-393
Is autophagy involved in the response to oxidative stress of tobacco BY-2 cells overexpressing thioredoxin o1? O. López-Vidal 1, A. Jimenez 1, S. Sevilla 1, L.M. Sandalio 2, A. Ortiz-Espín (*)1 1
Dpt of Stress Biology and Plant Pathology, CEBAS-CSIC, Murcia, Spain 2 Dpt of Biochemistry, Cellular and Molecular Biology of Plants, EEZ-CSIC, Granada, Spain
Autophagy is a catabolic process for cellular remodeling and macromolecular recycling of cytoplasmic components and organelles. Interestingly, autophagy appears as a key mechanism to cope with adverse conditions by removing damaged cellular components, although less information exists about this process in plants. In this sense, little is known about the possible role of redox regulation through thiol oxidoreductases thioredoxins (Trxs), specifically the mitochondrial/nuclear Trxo1. We have previously described a protective role of pea thioredoxin (PsTrxo1) overexpressed in TBY-2 cells treated with H2O2, thus increasing their viability via antioxidants, delaying cell death. In this work, we analyze the possible implication of autophagy as a protective mechanism operating in the PsTrxo1 transformed TBY-2 cells. For this, together with cell viability, different markers such as autophagy-related proteins ATGs are analyzed by Western blot, in parallel to autolysosomes and autophagosomes formation by fluorescence microscopy. The results have shown significant changes in ATG4 and ATG8 contents as well as presence of both lytic structures in H2O2-treated overexpressing PsTrxo1 cells, which suggest that Trxo1 may be also involved in the redox control of some autophagy components.
E-mail address: [email protected] http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.540
P-394 This work focuses on the effects of (-)-epicatechin (EC) on fat pads changes induced by high-fat (HF) diet. C57BL/6 male mice were divided in 4 groups receiving: control diet (10% calories from fat), HF diet (60% of calories), and these diets added with 20 mg EC/kg bw for 15 w. Even when caloric intake was not different among groups, HF increased mice weight. In white fat pads, inflammation (IL-6 and iNOS expression) and oxidative stress (gp91phox expression and MDA content) increases were associated with fat pad expansions. Adipocyte size was not different in HF respect to C group. On the other hand, HF diet induced expansion of aortic perivascular adipose tissue (aPVAT), with an increase of adipocyte size. Adipocytes/area were significantly lower in HF respect to C group. Changes in vacuolization patterns suggest a shift to white adipocyte phenotype, confirmed by a lower UCP-1 expression. All the observed changes in aPVAT were attenuated by EC. Since aPVAT dysfunction is relevant for hypertension and atherosclerosis, the effects of EC support an alternative nutritional strategy to avoid the progression of vascular dysfunction associated to the development of obesity.
Zinc modulates dopamine-induced heme oxygenase 1 expression in human IMR-32 neuroblastoma cells Zachary Kaufman 1, Gabriela Salvador 2, Patricia I. Oteiza 1 1
Departments of Nutrition and Environmental Toxicology, University of California, Davis, CA, USA 2 Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB-UNSCONICET). Bahía Blanca, Argentina
Zinc (Zn) plays a key role in the modulation of neuronal redox status. The present work tested the hypothesis that Zn is necessary for the neuronal defence response against dopamine (DA)-induced oxidative stress, in particular for the upregulation of heme oxigenase 1 (HMOX1).
A. Stepanova et al. / Free Radical Biology and Medicine 120 (2018) S45–S166
Human IMR-32 neuroblastoma cells exposed to a high DA (100 mM) concentration under a condition of Zn deficiency (ZD) showed an impaired capacity to upregulate HMOX1 when measured as mRNA and protein levels. This was reverted in Zn supplemented (ZS) cells. ZD decreased the capacity of IMR-32 cells to upregulate Nrf-2 and ERK1/2 in response to DA. However, chemical inhibition of ERK1/2 in ZS cells did not affect DA-mediated HMOX1 expression. BACH1 is a transcriptional repressor, which is central to the regulation of HMOX1 induction. Bach-1 levels were high in total homogenates and nuclear fraction from ZD cells stimulated with DA compared to ZS cells. BACH1 degradation was impaired in ZD compared to ZS cells, as evaluated by decreased levels of BACH1 ubiquitination upon DA treatment. Results show that Zn is necessary for the neuronal upregulation of HMOX1 upon a DA challenge, which is in part due to a requirement of Zn for the ubiquitin-mediated degradation of BACH1.
E-mail address: [email protected] http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.541
P-395
S165
3
Laboratory of Medicinal Chemistry, University of Antwerp, Belgium Sektion Nephrologie, Medizinische Klinik und Poliklinik III, Universitätsklinikum Carl Gustav Carus, Dresden, Germany 5 Methusalem program; Ghent University, Ghent, Belgium 4
Ferroptosis is an iron dependent form of necrosis, involved in different pathologies such as kidney failure and neurodegenerative diseases. Glutathione peroxidase 4 (GPX4) is the central enzyme protecting the cell from ferroptosis. High-throughput screening led to the discovery of ferrostatin-1 (Fer-1) as a potent in vitro inhibitor of ferroptosis. In vivo however, the molecule suffers from instability. Therefore, we developed ferrostatin-analogues with improved solubility and in vivo stability. To verify their in vivo efficacy, we established an acute mouse model of iron poisoning-induced organ injury. Both an excess of the natural ferroptosis inhibitor vitamin E in the feed and overexpression of GPX4 have a protective effect on this iron poisoning model, indicating it is a valuable experimental model for ferroptosis-induced organ injury. Interestingly, our novel ferrostatin analogues effectively decrease several markers for organ injury after iron poisoning. Conclusively, 1) ferroptosis is a detrimental factor in acute iron poisoning, 2) our novel ferrostatin-analogues reduce the severity of organ injury due to acute iron poisoning, and 3) our third generation of ferrostatin-analogues might represent potential leads for the treatment of ferroptosis-driven pathologies.
Early life stress, stress-resilience/susceptibility and oxidative stress
E-mail address: [email protected]; Tom.Van [email protected]
Samina Salim, Hesong Liu, Fatin Atrooz
http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.543
College of Pharmacy, University of Houston, Houston, TX, USA P-397 Adverse experiences during early life contribute to the development of psychiatric conditions later in life. In fact, young children who experience stressful traumatic event(s) during early life, a sensitive developmental period, are considered highly vulnerable to psychiatric disorders in adult life. Interestingly, not all children who experience stressful events are equally at risk of developing later life psychiatric disorders. Some are resilient in spite of being exposed to the same risk factors, while others are susceptible. The mechanistic basis for resilience/susceptibility is not clear. Using an early life stress model of Sprague Dawley rats, the mechanisms governing this phenotype segregation were revealed. Depression-susceptible phenotype is associated with high oxidative stress, low antioxidant status, defective redox-sensitive Nrf2 transcription factor and hyperactive NF-KB transcription factor, selectively in the pre-frontal cortex (PFC) area of the brain. The resistant phenotype displayed low oxidative stress, high antioxidant status, activated Nrf2 and inactivated NF-KB pathways.
E-mail address: [email protected] http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.542
P-396
Novel third-generation ferrostatins protect against acute iron poisoning-induced organ injury 1,2
1,2
1
June Lee, Young-Sam Keum College of Pharmacy and Integrated Research Institute for Drug Development, Dongguk University, Goyang, Gyeonggi-do, Korea
Heme oxygenase-1 (HO-1) catalyzes the enzymatic degradation of heme to produce three anti-oxidant molecules: carbon monoxide (CO), ferrous ion (Fe2 þ ), and biliverdin. Induction of HO-1 is currently considered as a feasible strategy to treat oxidative stress-related diseases. Here, we report marliolide as a novel inducer of HO-1 in human normal keratinocyte HaCaT cells. Mechanism-based studies demonstrated that the induction of HO-1 by marliolide occurred through activation of NRF2/ARE via direct binding of marliolide to KEAP1. Structure-activity relationship revealed chemical moieties of marliolide critical for induction of HO-1, which renders a support for Michael reaction as a potential mechanism of action. Finally, we observed that marliolide significantly inhibited the papilloma formation in DMBA/TPA–induced mouse skin carcinogenesis model and this event was closely associated with lowering the formation of 8-OH-G and 4-HNE in vivo. Together, we provide the first evidence that marliolide might be effective against oxidative stress-related skin disorders.
E-mail address: [email protected]
3
Samya Van Coillie , Wannes Weyts , Lars Devisscher , Sam Hofmans 3, Sze Men Choi 1,2, Bartosz Wiernicki 1,2, Behrouz Hassannia 1,2, Andreas Linkermann 4, Koen Augustyns 3, Peter Vandenabeele 1,2,5, Tom Vanden Berghe (*)1,2 VIB Center for Inflammation Research, Gent, Belgium Department of Biomedical Molecular Biology; Ghent University, Gent, Belgium
2
Marliolide inhibits skin carcinogenesis by activating NRF2/ARE to induce heme oxygenase-1
http://dx.doi.org/10.1016/j.freeradbiomed.2018.04.544