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The antileishmanial activity of xanthohumol but not of resveratrol is mediated by mitochondrial inhibition
C. Nyakas / Free Radical Biology and Medicine 96 (2016) S21–S31
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The antileishmanial activity of xanthohumol but not of resveratrol is mediated by mitochondrial inhibition Lars Gille 1, Alexandra Lackova 1, Katrin Staniek 1, Silvia Steinbauer 1, Gerald Pichler 1, Walter Jäger 2, Lianet Monzote 3 1
Institute of Pharmacology and Toxicology, Department of Biomedical Sciences, University of Veterinary Medicine, Vienna, Austria 2 Division of Clinical Pharmacy and Diagnostics, University of Vienna, Vienna, Austria 3 Parasitology Department, Institute of Tropical Medicine “Pedro Kouri”, Havana, Cuba
Resveratrol (Res) and xanthohumol (Xan) are naturally occurring antioxidants and constituents of human nutrition. Besides effects on mammalian cells also their effect on mammalian pathogens, such as protozoa, is of interest. Leishmania are eukaryotic protozoal organisms which infect humans and other mammals, threatening millions of people in (sub-) tropical regions. The treatment of leishmaniasis is difficult due to toxicity and resistance development for current drugs. For Res and Xan little is known about their actions on leishmanial parasites and their mitochondria as putative drug target. Therefore, we compared the antileishmanial activity of Res and Xan and their modes of action with respect to mitochondria. Antileishmanial activities of xenobiotics were assessed in Leishmania amazonensis amastigotes (LaA) and compared to peritoneal macrophages from mouse (PMM) using viability assays. Furthermore, mechanistic studies regarding mitochondrial functions were conducted in L. tarentolae promastigotes (LtP) and mitochondrial fractions isolated from LtP. Assays of individual mitochondrial complex activities, membrane potential and superoxide radical formation were performed by photometry, fluorimetry and electron spin resonance spectroscopy, respectively. Xan and Res demonstrated antileishmanial activity in LaA (IC50: Xan 6.9 7 0.3 mM, Res 13.9 7 1.7 mM) while they had less influence on the viability of PMM (IC50: Xan 69.5 7 9.8 mM, Res 4 438 mM). In contrast to Res, Xan strongly inhibited oxygen consumption in LtP. Further studies demonstrated that this was based on the inhibition of the mitochondrial electron transfer complex II/III by Xan. This effect was not observed with Res. However, both Xan and Res resulted in mitochondrial uncoupling. Neither Xan nor Res increased mitochondrial superoxide release in LtP. These data show that the antioxidants Res and Xan may have antileishmanial activity, however, only in case of Xan mitochondrial inhibition is involved in the mechanism. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.171 O-05
Generation of oxidative stress as an anticancer therapeutic strategy Tiantian He 1,2, Elie Hatem 1,2, Laurence Vernis 1,2, Meng-Er Huang 1,2 1
Centre National de la Recherche Scientifique, UMR3348 "Genotoxic Stress and Cancer", Orsay, France 2 Institut Curie, Centre de Recherche, Orsay, France
Aim: Intracellular reactive oxygen species (ROS) and reductive/oxidative (redox) homeostasis play a key role in regulating cell survival/
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death. Among the key players of redox system, peroxiredoxin 1 (PRX1) is a major antioxidant and signaling regulator. PRX1 is frequently over-expressed in various cancer cells, which is thought to be associated with carcinogenesis, metastasis and drug resistance. We have investigated whether and how modulations of intracellular redox system through PRX1 knockdown affect cancer cell sensitivity to a set of chemotherapy drugs or ROS-generating molecules. Materials and Methods: Cancer cell lines with PRX1 gene transiently or stably knocked-down and corresponding controls were exposed to a set of anticancer molecules, including vinblastine, taxol, doxorubicin, daunorubicin, actinomycin D, 5-fluorouracil, βlapachone (β-lap) and vitamin K3 (vitK3). Cytotoxic effects were evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)-based assay, cell clonogenic assay, annexin V/propidium iodide double staining. Global ROS accumulation and compartment-specific H2O2 generation were determined respectively by redox-sensitive chemical probes and H2O2-sensitive sensor HyPer. Oxidation of endogenous antioxidant proteins including TRX1, TRX2 and PRX3 was monitored by redox western blot. Results: PRX1 knockdown significantly and specifically enhances several cancer cell lines’ sensitivity to β-lap and vitK3, two structurally related quinone molecules. Interestingly, β-lap triggers NADPH:quinone oxidoreductase (NQO1)-dependent ROS generation and activation of mitogen-activated protein kinase pathway leading to cell death. On the other hand, PRX1 knockdown up-regulates NRH:quinone oxidoreductase 2 (NQO2) activity which is largely responsible for vitK3-induced ROS accumulation and cell death. Conclusion: Our data provide evidence that PRX1 could be an interesting anticancer target for modulating intracellular redox environment. More generally, these data support the view that redox modulation and enhanced generation of oxidative stress could be an alternative anticancer strategy to enhance cancer cell sensitivity to ROS-generating drugs or to overcome some types of drug resistance. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.172 O-06
Novel role of sirtuin related mechanisms in overloadinduced hypertrophy of skeletal muscle in rat Erika Koltai 1, Zoltán Bori 1, Clovis Chaber 2, Hervé Dubouchaud 2, Tomoko Funakoshi 3, Shuichi Machida 3, Zsolt Radák 1 1
University of Physical Education, Research Institute of Sport Sciences, Budapest, Hungary 2 Université Joseph Fourier, Laboratoire de Bioénergétique Fondamentale et Appliquée, Grenoble, France 3 Department of Exercise Physiology, Graduate School of Health and Sports Science & Medicine, Juntendo University, Tokyo, Japan
Introduction: Skeletal muscle is a dynamic organ, which can change its metabolic rate by manyfold during intensive contractions. The multi-nucleated myocytes as a result of resistance training can readily increase their size, indeed, the role of satellite cells in this process is still contradictory. The major member of sirtuin protein family, SIRT1 has been implicated as one of the key regulator in cell metabolism, but its involvement in muscle hypertrophy has not been investigated. SIRT1 has been shown to deacetylate eNOS, FOXO1 and activate Akt (Lee and Goldberg, 2013). Therefore, we suggested that SIRT1 can be a potential regulator of prosynthetic
O-04
The antileishmanial activity of xanthohumol but not of resveratrol is mediated by mitochondrial inhibition Lars Gille 1, Alexandra Lackova 1, Katrin Staniek 1, Silvia Steinbauer 1, Gerald Pichler 1, Walter Jäger 2, Lianet Monzote 3 1
Institute of Pharmacology and Toxicology, Department of Biomedical Sciences, University of Veterinary Medicine, Vienna, Austria 2 Division of Clinical Pharmacy and Diagnostics, University of Vienna, Vienna, Austria 3 Parasitology Department, Institute of Tropical Medicine “Pedro Kouri”, Havana, Cuba
Resveratrol (Res) and xanthohumol (Xan) are naturally occurring antioxidants and constituents of human nutrition. Besides effects on mammalian cells also their effect on mammalian pathogens, such as protozoa, is of interest. Leishmania are eukaryotic protozoal organisms which infect humans and other mammals, threatening millions of people in (sub-) tropical regions. The treatment of leishmaniasis is difficult due to toxicity and resistance development for current drugs. For Res and Xan little is known about their actions on leishmanial parasites and their mitochondria as putative drug target. Therefore, we compared the antileishmanial activity of Res and Xan and their modes of action with respect to mitochondria. Antileishmanial activities of xenobiotics were assessed in Leishmania amazonensis amastigotes (LaA) and compared to peritoneal macrophages from mouse (PMM) using viability assays. Furthermore, mechanistic studies regarding mitochondrial functions were conducted in L. tarentolae promastigotes (LtP) and mitochondrial fractions isolated from LtP. Assays of individual mitochondrial complex activities, membrane potential and superoxide radical formation were performed by photometry, fluorimetry and electron spin resonance spectroscopy, respectively. Xan and Res demonstrated antileishmanial activity in LaA (IC50: Xan 6.9 7 0.3 mM, Res 13.9 7 1.7 mM) while they had less influence on the viability of PMM (IC50: Xan 69.5 7 9.8 mM, Res 4 438 mM). In contrast to Res, Xan strongly inhibited oxygen consumption in LtP. Further studies demonstrated that this was based on the inhibition of the mitochondrial electron transfer complex II/III by Xan. This effect was not observed with Res. However, both Xan and Res resulted in mitochondrial uncoupling. Neither Xan nor Res increased mitochondrial superoxide release in LtP. These data show that the antioxidants Res and Xan may have antileishmanial activity, however, only in case of Xan mitochondrial inhibition is involved in the mechanism. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.171 O-05
Generation of oxidative stress as an anticancer therapeutic strategy Tiantian He 1,2, Elie Hatem 1,2, Laurence Vernis 1,2, Meng-Er Huang 1,2 1
Centre National de la Recherche Scientifique, UMR3348 "Genotoxic Stress and Cancer", Orsay, France 2 Institut Curie, Centre de Recherche, Orsay, France
Aim: Intracellular reactive oxygen species (ROS) and reductive/oxidative (redox) homeostasis play a key role in regulating cell survival/
S23
death. Among the key players of redox system, peroxiredoxin 1 (PRX1) is a major antioxidant and signaling regulator. PRX1 is frequently over-expressed in various cancer cells, which is thought to be associated with carcinogenesis, metastasis and drug resistance. We have investigated whether and how modulations of intracellular redox system through PRX1 knockdown affect cancer cell sensitivity to a set of chemotherapy drugs or ROS-generating molecules. Materials and Methods: Cancer cell lines with PRX1 gene transiently or stably knocked-down and corresponding controls were exposed to a set of anticancer molecules, including vinblastine, taxol, doxorubicin, daunorubicin, actinomycin D, 5-fluorouracil, βlapachone (β-lap) and vitamin K3 (vitK3). Cytotoxic effects were evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)-based assay, cell clonogenic assay, annexin V/propidium iodide double staining. Global ROS accumulation and compartment-specific H2O2 generation were determined respectively by redox-sensitive chemical probes and H2O2-sensitive sensor HyPer. Oxidation of endogenous antioxidant proteins including TRX1, TRX2 and PRX3 was monitored by redox western blot. Results: PRX1 knockdown significantly and specifically enhances several cancer cell lines’ sensitivity to β-lap and vitK3, two structurally related quinone molecules. Interestingly, β-lap triggers NADPH:quinone oxidoreductase (NQO1)-dependent ROS generation and activation of mitogen-activated protein kinase pathway leading to cell death. On the other hand, PRX1 knockdown up-regulates NRH:quinone oxidoreductase 2 (NQO2) activity which is largely responsible for vitK3-induced ROS accumulation and cell death. Conclusion: Our data provide evidence that PRX1 could be an interesting anticancer target for modulating intracellular redox environment. More generally, these data support the view that redox modulation and enhanced generation of oxidative stress could be an alternative anticancer strategy to enhance cancer cell sensitivity to ROS-generating drugs or to overcome some types of drug resistance. http://dx.doi.org/10.1016/j.freeradbiomed.2016.04.172 O-06
Novel role of sirtuin related mechanisms in overloadinduced hypertrophy of skeletal muscle in rat Erika Koltai 1, Zoltán Bori 1, Clovis Chaber 2, Hervé Dubouchaud 2, Tomoko Funakoshi 3, Shuichi Machida 3, Zsolt Radák 1 1
University of Physical Education, Research Institute of Sport Sciences, Budapest, Hungary 2 Université Joseph Fourier, Laboratoire de Bioénergétique Fondamentale et Appliquée, Grenoble, France 3 Department of Exercise Physiology, Graduate School of Health and Sports Science & Medicine, Juntendo University, Tokyo, Japan
Introduction: Skeletal muscle is a dynamic organ, which can change its metabolic rate by manyfold during intensive contractions. The multi-nucleated myocytes as a result of resistance training can readily increase their size, indeed, the role of satellite cells in this process is still contradictory. The major member of sirtuin protein family, SIRT1 has been implicated as one of the key regulator in cell metabolism, but its involvement in muscle hypertrophy has not been investigated. SIRT1 has been shown to deacetylate eNOS, FOXO1 and activate Akt (Lee and Goldberg, 2013). Therefore, we suggested that SIRT1 can be a potential regulator of prosynthetic