- Email: [email protected]
20 Methylene blue enhances specific mitochondrial functions in vitro and in vivo
Abstracts / Mitochondrion 10 (2010) 200–242
18 Methylene chloride effects on mitochondria G. Garrabou a,*, C. Morén a, M. Nicolàs a, J.C. Trullàs b, S. Mondón c, V. Navarro c, J. Jou c, S. Nogué d, Ò. Miró a,b, J. Casademont a, F. Cardellach a,e a Mitochondrial Research Laboratory, IDIBAPS, University of Barcelona and CIBER de Enfermedades Raras, CIBERER, Barcelona, Spain; b Emergency, Hospital Clínic of Barcelona, Barcelona, Catalonia, Spain; c Psychiatry, Hospital Clínic of Barcelona, Barcelona, Catalonia, Spain; d Toxicology, Hospital Clínic of Barcelona, Barcelona, Catalonia, Spain; e Internal Medicine Department, Hospital Clínic of Barcelona, Barcelona, Catalonia, Spain
Hypothesis: Overdose of volatile inhaled drugs, although not common in developed countries, can have fatal consequences. Methylene chloride (MC) is a common dissolvent that once metabolised in liver generates carbon monoxide (CO). CO is known to bind heme groups like those present in myoglobin, cytochrome p450, hemoglobin (with consequent carboxihemoglobin – COHb – formation and hypoxia development) and mitochondrial cytochrome c oxidase (COX). COX is an essential enzyme for oxygen consumption and energy production which chronic impairment could underlie clinical symptoms of MC abuse. Methods: We studied clinical and mitochondrial consequences of chronic MC abuse in a non-smoker hair lacquer sniffer woman. Mitochondrial function test consisted in measuring oxygen consumption (polarography) and COX activity (spectrophotometry) in isolated mononuclear cells (ficoll density gradient centrifugation). Results: Clinical analysis revealed hepatic and renal abnormalities, deficits in respiration function tests and neuropsicological alterations compatible with cognitive frontal deficits. COHb levels were increased (5.4%; normality [0–3]). Mitochondrial function tests showed inhibition of COX activity (55 nmol/min mg protein; normality [61–187]) without repercussion on mitochondrial oxygen consumption. Conclusions: Multiorganic involvement and mitochondrial damage characterise hair lacquer abuse. All their components are inert except for MC, which is metabolised into CO causing hypoxia and mitochondrial lesion. Further studies are required to assess contribution of both CO-mediated consequences into multiorganic damage and whether these effects are reversible or not. Supported by: Fundació la Marató de TV3 020210 and 020631, FIPSE 36612/06, FIS 40381/04 and 41239/04, Suports a Grups de Recerca de la Generalitat de Catalunya 2005/SGR/0300 and CIBER de Enfermedades Raras (initiative of the ISCIII). doi:10.1016/j.mito.2009.12.017
19 Role of three different variants of mitochondrial ribosomal S18s in mitochondrial translation Gurler Akpinar *, Ruilin Zhang, Min-Joon Han, Ching-Chung Ko, Hasan Koc, Emine Cavdar Koc Department of Biochemistry & Molecular Biology, Pennsylvania State, University, University Park, PA, United States
Mitochondria produce over 90% of the energy used by mammalian cells through the process of electron transfer. The formation of the membrane complexes essential for oxidative phosphorylation requires the expression of numerous genes, most of which are in the nucleus. However, 13 of the protein components for the electron transfer chain and the ATP synthase are the products of genes present in the mitochondrial DNA. The synthesis of these proteins are carried out by an essential and specific protein synthesizing system, mitochondrial ribosomes, within this organelle. All of the mammalian mitochondrial ribosomal proteins are encoded in nuclear genes and imported into mitochondria where they assemble with the mitochondrially transcribed rRNAs into ribosomes. We have identified all the proteins of mitochondrial ribosomes of which half have homologs in bacterial ribosomes and the remainder constitute a new class of ribosomal proteins. The functions of the many bacterial homologs are also conserved in protein synthesis in the mitochon-
205
dria. One of our most interesting findings was the presence of three different variants of S18s (MRP-S18s) which are homologous to bacterial ribosomal S18 protein in the small subunit of the mitochondrial ribosome. This intriguing observation implies that mitochondria contain three different sub-populations of the small ribosomal subunit (28S) to synthesize only 13 proteins and may imply a novel regulatory mechanism of protein synthesis in mammalian mitochondria. In this study, we confirm the presence of all three different variants of MRP-S18s in mitochondria obtained from various bovine tissues and mammalian cell lines. Moreover, differential expression of these S18s was observed in different tissues and this observation may relate effectiveness of mitochondrial translation to energy production in different tissues and cells. doi:10.1016/j.mito.2009.12.018
20 Methylene blue enhances specific mitochondrial functions in vitro and in vivo Hani Atamna a,*, Afshin Gharib b Department of Basic Sciences, Neuroscience, The Commonwealth Medical College, Scranton, PA 18510, United States; b Dominican University of California, San Rafael, CA 94901, United States a
We found that Methylene blue (MB) is very effective delaying cellular senescence and enhancing mitochondrial activity of primary human embryonic fibroblasts. MB also enhanced specific functions of the mitochondria. At nanomolar concentrations, MB increased: the activity of mitochondrial cytochrome c oxidase (complex IV), heme synthesis, cell resistance to oxidants and in addition delayed cellular senescence. We extended our in vitro findings to the investigation of the effect of long-term intake of MB in old mice. We administered MB, in the drinking water ð250 lMÞ, to old mice for 90 days. Remarkably, MB prevents the age-related decline in cognitive function and spatial memory. MB also prevents the age-related decline in grip strength. Interestingly, MB results in 100% and 50% increases in complex IV activities in the brains and hearts of old mice, respectively. We also found a 39% decrease in brain monoamine oxidase activity in old mice treated with MB. The age-related decline in protein content of the brain was prevented by MB. The activity of brain NQO1, on the other hand, was not affected by MB. These in vivo results are consistent with the in vitro findings and suggest that MB might delay neurodegenerative and physical impairments associated with mitochondrial dysfunctions. MB may serve as a leading compound to develop additional anti-aging agents. doi:10.1016/j.mito.2009.12.019
21 Mitochondrial outer membrane associated cholesterol transport is essential for steroidogenesis Dilip Debnath, Himangshu S. Bose * Department of Biomedical Sciences, Mercer University School of Medicine and Memorial Health University Medical Center, Savannah, GA 31404, United States
Mitochondrial resident steroidogenic acute regulatory protein, StAR, is required for the rapid import of cholesterol into the mitochondria of adrenal and gonadal cells, permitting rapid synthesis and release of steroid hormones in response to tropic hormones. Mutant StAR cannot move cholesterol into mitochondria, resulting an inborn child disorder called congenital lipoid adrenal hyper plasia (lipoid CAH), in which fetuses born shortly after birth due to saltloosing crisis. StAR acts exclusively on the outer mitochondrial membrane (OMM) and binds cholesterol with 1:1 stoichiometry, but each molecule of StAR appears to be responsible for the influx of many molecules of cholesterol. Despite physiologic, biochemical
18 Methylene chloride effects on mitochondria G. Garrabou a,*, C. Morén a, M. Nicolàs a, J.C. Trullàs b, S. Mondón c, V. Navarro c, J. Jou c, S. Nogué d, Ò. Miró a,b, J. Casademont a, F. Cardellach a,e a Mitochondrial Research Laboratory, IDIBAPS, University of Barcelona and CIBER de Enfermedades Raras, CIBERER, Barcelona, Spain; b Emergency, Hospital Clínic of Barcelona, Barcelona, Catalonia, Spain; c Psychiatry, Hospital Clínic of Barcelona, Barcelona, Catalonia, Spain; d Toxicology, Hospital Clínic of Barcelona, Barcelona, Catalonia, Spain; e Internal Medicine Department, Hospital Clínic of Barcelona, Barcelona, Catalonia, Spain
Hypothesis: Overdose of volatile inhaled drugs, although not common in developed countries, can have fatal consequences. Methylene chloride (MC) is a common dissolvent that once metabolised in liver generates carbon monoxide (CO). CO is known to bind heme groups like those present in myoglobin, cytochrome p450, hemoglobin (with consequent carboxihemoglobin – COHb – formation and hypoxia development) and mitochondrial cytochrome c oxidase (COX). COX is an essential enzyme for oxygen consumption and energy production which chronic impairment could underlie clinical symptoms of MC abuse. Methods: We studied clinical and mitochondrial consequences of chronic MC abuse in a non-smoker hair lacquer sniffer woman. Mitochondrial function test consisted in measuring oxygen consumption (polarography) and COX activity (spectrophotometry) in isolated mononuclear cells (ficoll density gradient centrifugation). Results: Clinical analysis revealed hepatic and renal abnormalities, deficits in respiration function tests and neuropsicological alterations compatible with cognitive frontal deficits. COHb levels were increased (5.4%; normality [0–3]). Mitochondrial function tests showed inhibition of COX activity (55 nmol/min mg protein; normality [61–187]) without repercussion on mitochondrial oxygen consumption. Conclusions: Multiorganic involvement and mitochondrial damage characterise hair lacquer abuse. All their components are inert except for MC, which is metabolised into CO causing hypoxia and mitochondrial lesion. Further studies are required to assess contribution of both CO-mediated consequences into multiorganic damage and whether these effects are reversible or not. Supported by: Fundació la Marató de TV3 020210 and 020631, FIPSE 36612/06, FIS 40381/04 and 41239/04, Suports a Grups de Recerca de la Generalitat de Catalunya 2005/SGR/0300 and CIBER de Enfermedades Raras (initiative of the ISCIII). doi:10.1016/j.mito.2009.12.017
19 Role of three different variants of mitochondrial ribosomal S18s in mitochondrial translation Gurler Akpinar *, Ruilin Zhang, Min-Joon Han, Ching-Chung Ko, Hasan Koc, Emine Cavdar Koc Department of Biochemistry & Molecular Biology, Pennsylvania State, University, University Park, PA, United States
Mitochondria produce over 90% of the energy used by mammalian cells through the process of electron transfer. The formation of the membrane complexes essential for oxidative phosphorylation requires the expression of numerous genes, most of which are in the nucleus. However, 13 of the protein components for the electron transfer chain and the ATP synthase are the products of genes present in the mitochondrial DNA. The synthesis of these proteins are carried out by an essential and specific protein synthesizing system, mitochondrial ribosomes, within this organelle. All of the mammalian mitochondrial ribosomal proteins are encoded in nuclear genes and imported into mitochondria where they assemble with the mitochondrially transcribed rRNAs into ribosomes. We have identified all the proteins of mitochondrial ribosomes of which half have homologs in bacterial ribosomes and the remainder constitute a new class of ribosomal proteins. The functions of the many bacterial homologs are also conserved in protein synthesis in the mitochon-
205
dria. One of our most interesting findings was the presence of three different variants of S18s (MRP-S18s) which are homologous to bacterial ribosomal S18 protein in the small subunit of the mitochondrial ribosome. This intriguing observation implies that mitochondria contain three different sub-populations of the small ribosomal subunit (28S) to synthesize only 13 proteins and may imply a novel regulatory mechanism of protein synthesis in mammalian mitochondria. In this study, we confirm the presence of all three different variants of MRP-S18s in mitochondria obtained from various bovine tissues and mammalian cell lines. Moreover, differential expression of these S18s was observed in different tissues and this observation may relate effectiveness of mitochondrial translation to energy production in different tissues and cells. doi:10.1016/j.mito.2009.12.018
20 Methylene blue enhances specific mitochondrial functions in vitro and in vivo Hani Atamna a,*, Afshin Gharib b Department of Basic Sciences, Neuroscience, The Commonwealth Medical College, Scranton, PA 18510, United States; b Dominican University of California, San Rafael, CA 94901, United States a
We found that Methylene blue (MB) is very effective delaying cellular senescence and enhancing mitochondrial activity of primary human embryonic fibroblasts. MB also enhanced specific functions of the mitochondria. At nanomolar concentrations, MB increased: the activity of mitochondrial cytochrome c oxidase (complex IV), heme synthesis, cell resistance to oxidants and in addition delayed cellular senescence. We extended our in vitro findings to the investigation of the effect of long-term intake of MB in old mice. We administered MB, in the drinking water ð250 lMÞ, to old mice for 90 days. Remarkably, MB prevents the age-related decline in cognitive function and spatial memory. MB also prevents the age-related decline in grip strength. Interestingly, MB results in 100% and 50% increases in complex IV activities in the brains and hearts of old mice, respectively. We also found a 39% decrease in brain monoamine oxidase activity in old mice treated with MB. The age-related decline in protein content of the brain was prevented by MB. The activity of brain NQO1, on the other hand, was not affected by MB. These in vivo results are consistent with the in vitro findings and suggest that MB might delay neurodegenerative and physical impairments associated with mitochondrial dysfunctions. MB may serve as a leading compound to develop additional anti-aging agents. doi:10.1016/j.mito.2009.12.019
21 Mitochondrial outer membrane associated cholesterol transport is essential for steroidogenesis Dilip Debnath, Himangshu S. Bose * Department of Biomedical Sciences, Mercer University School of Medicine and Memorial Health University Medical Center, Savannah, GA 31404, United States
Mitochondrial resident steroidogenic acute regulatory protein, StAR, is required for the rapid import of cholesterol into the mitochondria of adrenal and gonadal cells, permitting rapid synthesis and release of steroid hormones in response to tropic hormones. Mutant StAR cannot move cholesterol into mitochondria, resulting an inborn child disorder called congenital lipoid adrenal hyper plasia (lipoid CAH), in which fetuses born shortly after birth due to saltloosing crisis. StAR acts exclusively on the outer mitochondrial membrane (OMM) and binds cholesterol with 1:1 stoichiometry, but each molecule of StAR appears to be responsible for the influx of many molecules of cholesterol. Despite physiologic, biochemical