The Experts below are selected from a list of 1974 Experts worldwide ranked by ideXlab platform
David G. Nicholls - One of the best experts on this subject based on the ideXlab platform.
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mild uncoupling does not decrease mitochondrial superoxide levels in cultured cerebellar granule neurons but decreases spare respiratory capacity and increases toxicity to glutamate and oxidative stress
Journal of Neurochemistry, 2007Co-Authors: Linda Johnsoncadwell, Mika B. Jekabsons, A Wang, B M Polster, David G. NichollsAbstract:Cultured rat cerebellar granule neurons were incubated with low nanomolar concentrations of the Protonophore carbonylcyanide-p-trifluoromethoxyphenyl hydrazone (FCCP) to test the hypothesis that 'mild uncoupling' could be neuroprotective by decreasing oxidative stress. To quantify the uncoupling, respiration and mitochondrial membrane potential (Deltapsi(m)) were determined in parallel as a function of FCCP concentration. Deltapsi(m) dropped by less than 10 mV before respiratory control was lost. Conditions for the valid estimation of matrix superoxide levels were determined from the rate of oxidation of the matrix-targeted fluorescent probe MitoSOX. No significant change in the level of matrix superoxide could be detected on addition of FCCP while respiratory control was retained, although cytoplasmic superoxide levels measured by dihydroethidium oxidation increased. 'Mild uncoupling' by 30 nmol/L FCCP did not alleviate neuronal dysregulation induced by glutathione depletion and significantly enhanced that due to menadione-induced oxidative stress. Low Protonophore concentrations enhanced N-methyl-d-aspartate receptor-induced delayed calcium deregulation consistent with a decrease in the spare respiratory capacity available to match the bioenergetic demand of chronic receptor activation. It is concluded that the 'mild uncoupling' hypothesis is not supported by this model.
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mitochondrial membrane potential and hydroethidine monitored superoxide generation in cultured cerebellar granule cells
FEBS Letters, 1997Co-Authors: Samantha L. Budd, Roger F. Castilho, David G. NichollsAbstract:Mitochondrial depolarisation has been reported to enhance the generation of superoxide anion (O2.-) in a number of cell preparations while an inhibition has been observed with isolated mitochondria. Cerebellar granule cells equilibrated with > 1 microM hydroethidine (dihydroethidium) which is oxidised to the fluorescent ethidium cation by O2.- showed a large increase in fluorescence on Protonophore addition. However, controls showed the fluorescent enhancement to be a consequence of release of unbound preformed ethidium from the mitochondrial matrix within the cell with resultant fluorescent enhancement. This ambiguity was removed by the use of low (1 microM) concentrations of dye in which case generated ethidium remained bound within the mitochondria. Under these conditions antimycin A, but not Protonophore addition, produced an increase in fluorescence. It is concluded that excess ethidium acts as an indicator of mitochondrial membrane potential obscuring the monitoring of O2.- and that certain experiments employing this indicator in cells may require re-evaluation.
Roger F. Castilho - One of the best experts on this subject based on the ideXlab platform.
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underestimation of the maximal capacity of the mitochondrial electron transport system in oligomycin treated cells
PLOS ONE, 2016Co-Authors: Juliana S Ruas, Edilene S Siqueirasantos, Ignacio Amigo, Erika Rodriguessilva, Alicia J Kowaltowski, Roger F. CastilhoAbstract:The maximal capacity of the mitochondrial electron transport system (ETS) in intact cells is frequently estimated by promoting Protonophore-induced maximal oxygen consumption preceded by inhibition of oxidative phosphorylation by oligomycin. In the present study, human glioma (T98G and U-87MG) and prostate cancer (PC-3) cells were titrated with different concentrations of the Protonophore CCCP to induce maximal oxygen consumption rate (OCR) within respirometers in a conventional growth medium. The results demonstrate that the presence of oligomycin or its A-isomer leads to underestimation of maximal ETS capacity. In the presence of oligomycin, the spare respiratory capacity (SRC), i.e., the difference between the maximal and basal cellular OCR, was underestimated by 25 to 45%. The inhibitory effect of oligomycin on SRC was more pronounced in T98G cells and was observed in both suspended and attached cells. Underestimation of SRC also occurred when oxidative phosphorylation was fully inhibited by the ATP synthase inhibitor citreoviridin. Further experiments indicated that oligomycin cannot be replaced by the adenine nucleotide translocase inhibitors bongkrekic acid or carboxyatractyloside because, although these compounds have effects in permeabilized cells, they do not inhibit oxidative phosphorylation in intact cells. We replaced CCCP by FCCP, another potent Protonophore and similar results were observed. Lower maximal OCR and SRC values were obtained with the weaker Protonophore 2,4-dinitrophenol, and these parameters were not affected by the presence of oligomycin. In permeabilized cells or isolated brain mitochondria incubated with respiratory substrates, only a minor inhibitory effect of oligomycin on CCCP-induced maximal OCR was observed. We conclude that unless a previously validated protocol is employed, maximal ETS capacity in intact cells should be estimated without oligomycin. The inhibitory effect of an ATP synthase blocker on potent Protonophore-induced maximal OCR may be associated with impaired metabolism of mitochondrial respiratory substrates.
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mitochondrial membrane potential and hydroethidine monitored superoxide generation in cultured cerebellar granule cells
FEBS Letters, 1997Co-Authors: Samantha L. Budd, Roger F. Castilho, David G. NichollsAbstract:Mitochondrial depolarisation has been reported to enhance the generation of superoxide anion (O2.-) in a number of cell preparations while an inhibition has been observed with isolated mitochondria. Cerebellar granule cells equilibrated with > 1 microM hydroethidine (dihydroethidium) which is oxidised to the fluorescent ethidium cation by O2.- showed a large increase in fluorescence on Protonophore addition. However, controls showed the fluorescent enhancement to be a consequence of release of unbound preformed ethidium from the mitochondrial matrix within the cell with resultant fluorescent enhancement. This ambiguity was removed by the use of low (1 microM) concentrations of dye in which case generated ethidium remained bound within the mitochondria. Under these conditions antimycin A, but not Protonophore addition, produced an increase in fluorescence. It is concluded that excess ethidium acts as an indicator of mitochondrial membrane potential obscuring the monitoring of O2.- and that certain experiments employing this indicator in cells may require re-evaluation.
Kim D. Janda - One of the best experts on this subject based on the ideXlab platform.
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Dual Protonophore–Chitinase Inhibitors Dramatically Affect O. volvulus Molting
2015Co-Authors: Major Gooyit, Nancy Tricoche, Sara Lustigman, Kim D. JandaAbstract:The L3-stage-specific chitinase OvCHT1 has been implicated in the development of Onchocerca volvulus, the causative agent of onchocerciasis. Closantel, a known anthelmintic drug, was previously discovered as a potent and specific OvCHT1 inhibitor. As closantel is also a known Protonophore, we performed a simple scaffold modulation to map out the structural features that are relevant for its individual or dual biochemical roles. Furthermore, we present that either OvCHT1 inhibition or protonophoric activity was capable of affecting O. volvulus L3 molting and that the presence of both activities in a single molecule yielded more potent inhibition of the nematode’s developmental process
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dual Protonophore chitinase inhibitors dramatically affect o volvulus molting
Journal of Medicinal Chemistry, 2014Co-Authors: Major Gooyit, Nancy Tricoche, Sara Lustigman, Kim D. JandaAbstract:The L3-stage-specific chitinase OvCHT1 has been implicated in the development of Onchocerca volvulus, the causative agent of onchocerciasis. Closantel, a known anthelmintic drug, was previously discovered as a potent and specific OvCHT1 inhibitor. As closantel is also a known Protonophore, we performed a simple scaffold modulation to map out the structural features that are relevant for its individual or dual biochemical roles. Furthermore, we present that either OvCHT1 inhibition or protonophoric activity was capable of affecting O. volvulus L3 molting and that the presence of both activities in a single molecule yielded more potent inhibition of the nematode’s developmental process.
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design synthesis and biological activities of closantel analogues structural promiscuity and its impact on onchocerca volvulus
Journal of Medicinal Chemistry, 2011Co-Authors: Amanda L. Garner, Christian Gloeckner, Nancy Tricoche, Sara Lustigman, Joseph S Zakhari, Moses Samje, Fidelis Chongwa, Kim D. JandaAbstract:Onchocerciasis, or river blindness, is a neglected tropical disease that affects more than 37 million people worldwide, primarily in Africa and Central and South America. We have disclosed evidence that the larval-stage-specific chitinase, OvCHT1, may be a potential biological target for affecting nematode development. On the basis of screening efforts, closantel, a known anthelmintic drug, was discovered as a potent and highly specific OvCHT1 inhibitor. Originally, closantel’s anthelmintic mode of action was believed to rely solely on its role as a proton ionophore; thus, the impact of each of its biological activities on O. volvulus L3 molting was investigated. Structure–activity relationship studies on an active closantel fragment are detailed, and remarkably, by use of a simple salicylanilide scaffold, compounds acting only as Protonophores or chitinase inhibitors were identified. From these data, unexpected synergistic Protonophore and chitinase inhibition activities have also been found to be critic...
Rachel J Perry - One of the best experts on this subject based on the ideXlab platform.
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uncoupling hepatic oxidative phosphorylation reduces tumor growth in two murine models of colon cancer
Cell Reports, 2018Co-Authors: Yongliang Wang, Xianman Zhang, Gerald I Shulman, William E. Damsky, Curtis J. Perry, Ali Nasiri, Aviva Rabincourt, Michael Pollak, Rachel J PerryAbstract:Obesity is associated with colon cancer pathogenesis, but the underlying mechanism is actively debated. Here, we confirm that diet-induced obesity promotes tumor growth in two murine colon cancer models and show that this effect is reversed by an orally administered controlled-release mitochondrial Protonophore (CRMP) that acts as a liver-specific uncoupler of oxidative phosphorylation. This agent lowered circulating insulin, and the reduction of tumor growth was abrogated by an insulin infusion raising plasma insulin to the level of high-fat-fed mice. We also demonstrate that hyperinsulinemia increases glucose uptake and oxidation in vivo in tumors and that CRMP reverses these effects. This study provides evidence that perturbations of whole-organism energy balance or hepatic energy metabolism can influence neoplastic growth. Furthermore, the data show that glucose uptake and utilization by cancers in vivo are not necessarily constitutively high but rather may vary according to the hormonal milieu.
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Uncoupling Hepatic Oxidative Phosphorylation Reduces Tumor Growth in Two Murine Models of Colon Cancer
'Elsevier BV', 2018Co-Authors: Yongliang Wang, Xianman Zhang, Gerald I Shulman, Ali R. Nasiri, William E. Damsky, Curtis J. Perry, Aviva Rabin-court, Michael N. Pollak, Rachel J PerryAbstract:Summary: Obesity is associated with colon cancer pathogenesis, but the underlying mechanism is actively debated. Here, we confirm that diet-induced obesity promotes tumor growth in two murine colon cancer models and show that this effect is reversed by an orally administered controlled-release mitochondrial Protonophore (CRMP) that acts as a liver-specific uncoupler of oxidative phosphorylation. This agent lowered circulating insulin, and the reduction of tumor growth was abrogated by an insulin infusion raising plasma insulin to the level of high-fat-fed mice. We also demonstrate that hyperinsulinemia increases glucose uptake and oxidation in vivo in tumors and that CRMP reverses these effects. This study provides evidence that perturbations of whole-organism energy balance or hepatic energy metabolism can influence neoplastic growth. Furthermore, the data show that glucose uptake and utilization by cancers in vivo are not necessarily constitutively high but rather may vary according to the hormonal milieu. : Wang et al. demonstrate that diet-induced hyperinsulinemia increases colon adenocarcinoma tumor glucose uptake and oxidation in mice. They further demonstrate that reversal of hyperinsulinemia by a liver-specific mitochondrial Protonophore is sufficient to reverse the obesity-induced acceleration of tumor growth. Keywords: colon adenocarcinoma, insulin, insulin resistance, glucose metabolism, uncouplin
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controlled release mitochondrial Protonophore reverses diabetes and steatohepatitis in rats
Science, 2015Co-Authors: Rachel J Perry, Dongyan Zhang, Xianman Zhang, James L Boyer, Gerald I ShulmanAbstract:Nonalcoholic fatty liver disease (NAFLD) is a major factor in the pathogenesis of type 2 diabetes (T2D) and nonalcoholic steatohepatitis (NASH). The mitochondrial Protonophore 2,4 dinitrophenol (DNP) has beneficial effects on NAFLD, insulin resistance, and obesity in preclinical models but is too toxic for clinical use. We developed a controlled-release oral formulation of DNP, called CRMP (controlled-release mitochondrial Protonophore), that produces mild hepatic mitochondrial uncoupling. In rat models, CRMP reduced hypertriglyceridemia, insulin resistance, hepatic steatosis, and diabetes. It also normalized plasma transaminase concentrations, ameliorated liver fibrosis, and improved hepatic protein synthetic function in a methionine/choline–deficient rat model of NASH. Chronic treatment with CRMP was not associated with any systemic toxicity. These data offer proof of concept that mild hepatic mitochondrial uncoupling may be a safe and effective therapy for the related epidemics of metabolic syndrome, T2D, and NASH.
Lech Wojtczak - One of the best experts on this subject based on the ideXlab platform.
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fatty acid induced uncoupling of oxidative phosphorylation is partly due to opening of the mitochondrial permeability transition pore
FEBS Letters, 1998Co-Authors: Mariusz R Wieckowski, Lech WojtczakAbstract:Addition of myristate at low concentration (30–60 nmol/mg protein) to energized rat liver mitochondria resulted in dissipation of the electric membrane potential which, in Ca2+-free media, could be partly reversed by carboxyatractyloside but not by cyclosporin A. In contrast, in mitochondria preloaded with Ca2+ this energy-dissipating effect of fatty acid was partly prevented or reversed by cyclosporin A or ADP. In sucrose media, myristate, but not the Protonophore carbonyl cyanide m-chlorophenylhydrazone, induced swelling of Ca2+-loaded mitochondria which was inhibited by cyclosporin A and ADP. We conclude that long-chain fatty acids may induce opening of the mitochondrial permeability transition pore not only because of their protonophoric effect mediated by mitochondrial anion carriers [Skulachev, V.P., FEBS Lett. 294 (1991) 158–162; Wieckowski, M.R. and Wojtczak, L., Biochem. Biophys. Res. Commun. (1997) 232, 414–417] but also by a direct interacton with the pore assembly.
