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Organic Sources of Sulfur Dioxide, a Reactive Sulfur Species
7 Very Long Chain Acyl-CoA Dehydrogenase Is a DietSensitive Source of Reactive Oxygen Species in Liver Mitochondria 1
Ariel Rodrigues Cardoso and Alicia Juliana Kowaltowski1 1 Institute of Chemistry University of Sao Paulo, Brazil High fat diets and accompanying hepatic steatosis are a staple of modern life. Previous work has shown that this condition is accompanied by enhanced generation of reactive oxygen species (ROS), which may mediate further liver damage. Here we investigated mechanisms leading to enhanced ROS generation following high fat diets (HFD). We found that mitochondria from HFD livers present no differences in maximal respiratory rates and coupling, but generate more ROS specifically when fatty acids are used as substrates. Indeed, many acyl-CoA dehydrogenase isoforms were found to be more highly expressed in HFD livers, although only the very long chain acyl-CoA dehydrogenase (VLCAD) was more functionally active. Studies conducted with permeabilized mitochondria and different chain length acyl-CoA derivatives indicate that VLCAD itself is the source of enhanced ROS production in the mitochondria of HFD animals. Overall, our studies demonstrate that VLCAD is a novel, diet-sensitive, source of mitochondrial ROS.
doi:10.1016/j.freeradbiomed.2012.10.034
8 Disruption of Mitochondrial Superoxide Flux Causes Abnormal Heme Synthesis and Hemoglobin Gene Regulation in Erythroid Precursor Cells 1
Adam J Case , Joshua M Madsen1, David G Motto1, David K Meyerholz1, and Frederick E Domann1 1 The University of Iowa Heme synthesis is a mitochondrial process occurring alongside cellular metabolism. Due to the close proximity of these two processes, the probability of heme intermediates being exposed to metabolic byproducts, particularly reactive oxygen species is II high. In addition, the need for ferrous iron (Fe ) for heme production, Fe-S coordination, and other processes occurring in the mitochondrial matrix suggests that aberrant fluxes of reactive oxygen species in this compartment might perturb normal iron homeostasis. Manganese superoxide dismutase (Sod2) is a mitochondrial anti-oxidant enzyme that governs steady-state levels of the superoxide in the mitochondrial matrix. Here, we report the effect of abnormal mitochondrial superoxide flux on erythrocyte development in vivo. Using hematopoietic stem cellspecific conditional Sod2 knock-out mice we observed increased superoxide concentrations in red cell progeny which caused significant pathologies including impaired heme synthesis, porphyria, and erythrocyte damage. Animals lacking Sod2 expression in erythroid precursors also displayed extramedullary hematopoiesis and systemic iron redistribution. Importantly, we report for the first time to our knowledge a contributing molecular mechanism underlying the phenotype as in vivo inhibition of ferrochelatase by superoxide. The increase in superoxide flux in erythroid precursors also caused abnormal gene regulation of hematopoietic transcription factors, globins, and iron-response genes. Moreover, the erythroid precursors also displayed evidence of global changes of histone post-translational modifications, a likely cause of at least some of the aberrant gene expression noted since globins are especially well-known for the epigenetic basis of their control. From a therapeutic translational perspective, mitochondrially-targeted superoxide-scavenging anti-
oxidants effectively rescued the observed phenotype. Taken together, our findings illuminate the superoxide sensitivity of ferrochelatase and suggest a probable link between mitochondrial redox metabolism and epigenetic control of nuclear gene regulation during mammalian erythropoiesis.
doi:10.1016/j.freeradbiomed.2012.10.035
9 Organic Sources of Sulfur Dioxide, a Reactive Sulfur Species 1
Harinath Chakrapani and Satish Malwal1 1 Indian Institute of Science Education and Research Sulfur dioxide (SO2), a byproduct of combustion of fossil fuels and industrial processes is an environmental pollutant that has severe toxicological effects including on the respiratory tract. Gaseous SO2 is hydrated to produce bisulfite HSO3−, which is converted to 2 sulfite SO3 − in basic pH. S(IV) oxides are also routinely used in the food industry as preservatives and as antibiotics in the wine industry. Sulfur dioxide can participate in both oxidative as well as reductive processes. During its oxidation, sulfur dioxide produces Ɣ Ɣ Ɣ radical intermediates such as including SO3 , SO4 Ǧ and HSO5 Ǧ, which are known to cause DNA damage. Sulfite can break disulfide linkages to produce S-sulfonates with yet incompletely characterized biological consequences. With increased industrialization and greater need for energy, larger populations are exposed to S(IV) oxides for extended durations. Being a gas, obtaining toxicological data using biological model systems and SO2 is cumbersome. Typically, a complex mixture of sulfite and bisulfite is utilized as a surrogate for SO2. Such inorganic exogenous sources lack temporal control over SO2 generation and may not be useful to mimic effects of this toxic gas. Here, we report 1-aryl-1,2-oxathiane-3-oxides, which were designed to undergo thermal extrusion of SO2 as efficient SO2 generators in physiological pH and non-enzymatic conditions with half-lives ranging from 10-56 min with possible applications for controlled generation of S(IV) oxides for biological studies.
doi:10.1016/j.freeradbiomed.2012.10.036
10 Vascular Peroxidase 1 Catalyzes the Formation of Hypohalous Acids: Characterization of Its Substrate Specificity, Enzymatic Properties and Microbicidal Activity
Zehong Cao1, Hong Li1, Guogang Zhang2, Victor J Thannickal1, and 1 Guangjie Cheng 1 2 University of Alabama at Birmingham, Central South University, Changsha, China The heme-containing peroxidase family comprises eight members in humans. Phagocyte-derived myeloperoxidase (MPO) utilizes chloride and bromide, in the presence of hydrogen peroxide (H2O2), to generate HOCl and HOBr, potent oxidizing species that are known to kill invading pathogens. Vascular peroxidase 1 (VPO1) is a new member of the heme-containing peroxidase
SFRBM 2012
S17
Ariel Rodrigues Cardoso and Alicia Juliana Kowaltowski1 1 Institute of Chemistry University of Sao Paulo, Brazil High fat diets and accompanying hepatic steatosis are a staple of modern life. Previous work has shown that this condition is accompanied by enhanced generation of reactive oxygen species (ROS), which may mediate further liver damage. Here we investigated mechanisms leading to enhanced ROS generation following high fat diets (HFD). We found that mitochondria from HFD livers present no differences in maximal respiratory rates and coupling, but generate more ROS specifically when fatty acids are used as substrates. Indeed, many acyl-CoA dehydrogenase isoforms were found to be more highly expressed in HFD livers, although only the very long chain acyl-CoA dehydrogenase (VLCAD) was more functionally active. Studies conducted with permeabilized mitochondria and different chain length acyl-CoA derivatives indicate that VLCAD itself is the source of enhanced ROS production in the mitochondria of HFD animals. Overall, our studies demonstrate that VLCAD is a novel, diet-sensitive, source of mitochondrial ROS.
doi:10.1016/j.freeradbiomed.2012.10.034
8 Disruption of Mitochondrial Superoxide Flux Causes Abnormal Heme Synthesis and Hemoglobin Gene Regulation in Erythroid Precursor Cells 1
Adam J Case , Joshua M Madsen1, David G Motto1, David K Meyerholz1, and Frederick E Domann1 1 The University of Iowa Heme synthesis is a mitochondrial process occurring alongside cellular metabolism. Due to the close proximity of these two processes, the probability of heme intermediates being exposed to metabolic byproducts, particularly reactive oxygen species is II high. In addition, the need for ferrous iron (Fe ) for heme production, Fe-S coordination, and other processes occurring in the mitochondrial matrix suggests that aberrant fluxes of reactive oxygen species in this compartment might perturb normal iron homeostasis. Manganese superoxide dismutase (Sod2) is a mitochondrial anti-oxidant enzyme that governs steady-state levels of the superoxide in the mitochondrial matrix. Here, we report the effect of abnormal mitochondrial superoxide flux on erythrocyte development in vivo. Using hematopoietic stem cellspecific conditional Sod2 knock-out mice we observed increased superoxide concentrations in red cell progeny which caused significant pathologies including impaired heme synthesis, porphyria, and erythrocyte damage. Animals lacking Sod2 expression in erythroid precursors also displayed extramedullary hematopoiesis and systemic iron redistribution. Importantly, we report for the first time to our knowledge a contributing molecular mechanism underlying the phenotype as in vivo inhibition of ferrochelatase by superoxide. The increase in superoxide flux in erythroid precursors also caused abnormal gene regulation of hematopoietic transcription factors, globins, and iron-response genes. Moreover, the erythroid precursors also displayed evidence of global changes of histone post-translational modifications, a likely cause of at least some of the aberrant gene expression noted since globins are especially well-known for the epigenetic basis of their control. From a therapeutic translational perspective, mitochondrially-targeted superoxide-scavenging anti-
oxidants effectively rescued the observed phenotype. Taken together, our findings illuminate the superoxide sensitivity of ferrochelatase and suggest a probable link between mitochondrial redox metabolism and epigenetic control of nuclear gene regulation during mammalian erythropoiesis.
doi:10.1016/j.freeradbiomed.2012.10.035
9 Organic Sources of Sulfur Dioxide, a Reactive Sulfur Species 1
Harinath Chakrapani and Satish Malwal1 1 Indian Institute of Science Education and Research Sulfur dioxide (SO2), a byproduct of combustion of fossil fuels and industrial processes is an environmental pollutant that has severe toxicological effects including on the respiratory tract. Gaseous SO2 is hydrated to produce bisulfite HSO3−, which is converted to 2 sulfite SO3 − in basic pH. S(IV) oxides are also routinely used in the food industry as preservatives and as antibiotics in the wine industry. Sulfur dioxide can participate in both oxidative as well as reductive processes. During its oxidation, sulfur dioxide produces Ɣ Ɣ Ɣ radical intermediates such as including SO3 , SO4 Ǧ and HSO5 Ǧ, which are known to cause DNA damage. Sulfite can break disulfide linkages to produce S-sulfonates with yet incompletely characterized biological consequences. With increased industrialization and greater need for energy, larger populations are exposed to S(IV) oxides for extended durations. Being a gas, obtaining toxicological data using biological model systems and SO2 is cumbersome. Typically, a complex mixture of sulfite and bisulfite is utilized as a surrogate for SO2. Such inorganic exogenous sources lack temporal control over SO2 generation and may not be useful to mimic effects of this toxic gas. Here, we report 1-aryl-1,2-oxathiane-3-oxides, which were designed to undergo thermal extrusion of SO2 as efficient SO2 generators in physiological pH and non-enzymatic conditions with half-lives ranging from 10-56 min with possible applications for controlled generation of S(IV) oxides for biological studies.
doi:10.1016/j.freeradbiomed.2012.10.036
10 Vascular Peroxidase 1 Catalyzes the Formation of Hypohalous Acids: Characterization of Its Substrate Specificity, Enzymatic Properties and Microbicidal Activity
Zehong Cao1, Hong Li1, Guogang Zhang2, Victor J Thannickal1, and 1 Guangjie Cheng 1 2 University of Alabama at Birmingham, Central South University, Changsha, China The heme-containing peroxidase family comprises eight members in humans. Phagocyte-derived myeloperoxidase (MPO) utilizes chloride and bromide, in the presence of hydrogen peroxide (H2O2), to generate HOCl and HOBr, potent oxidizing species that are known to kill invading pathogens. Vascular peroxidase 1 (VPO1) is a new member of the heme-containing peroxidase
SFRBM 2012
S17