Alterations in mitochondrial dynamics, including their intracellular trafficking, are common early manifestations of neuronal degeneration. However, current methodologies used to study mitochondrial trafficking events rely on parameters that are primarily altered in later stages of neurodegeneration. Our objective was to establish a reliable applied statistical analysis to detect early alterations in neuronal mitochondrial trafficking. We propose a novel quantitative analysis of mitochondria trajectories based on innovative movement descriptors, including straightness, efficiency, anisotropy, and kurtosis. We evaluated time- and dose-dependent alterations in trajectory descriptors using biological data from differentiated SH-SY5Y cells treated with the mitochondrial toxicants 6-hydroxydopamine and rotenone. MitoTracker Red CMXRos-labelled mitochondria movement was analyzed by total internal reflection fluorescence microscopy followed by computational modelling to describe the process. Based on the aforementioned trajectory descriptors, this innovative analysis of mitochondria trajectories provides insights into mitochondrial movement characteristics and can be a consistent and sensitive method to detect alterations in mitochondrial trafficking occurring in the earliest time points of neurodegeneration.

• MeSH
• adrenergní látky škodlivé účinky   MeSH
• buněčná diferenciace MeSH
• lidé MeSH
• mitochondriální dynamika * MeSH
• mitochondrie účinky léků   patologie   MeSH
• neuroblastom chemicky indukované   patologie   MeSH
• neurony účinky léků   patologie   MeSH
• oxidopamin škodlivé účinky   MeSH
• rotenon škodlivé účinky   MeSH
• rozpřahující látky škodlivé účinky   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

According to the common view, weak acid uncouplers increase proton conductance of biological (and phospholipid bilayer) membranes, thus effecting H+ fluxes driven by their electrochemical gradients. Under certain conditions, however, uncouplers can induce unexpected effects opposite to the dissipation of H+ gradients. Results are presented here demonstrating CCCP-induced proton influx into Saccharomyces cerevisiae cytosol driven by the electrochemical potentials of CCCP and its CCCP- anions, independent of electrochemical H+-gradient. Another view of week acid uncouplers' action is proposed that is logically consistent with these observations.

• MeSH
• biologický transport účinky léků   MeSH
• karbonylkyanid-m-chlorfenylhydrazon farmakologie   MeSH
• koncentrace vodíkových iontů účinky léků   MeSH
• membránové potenciály * účinky léků   MeSH
• protony * MeSH
• rozpřahující látky farmakologie   MeSH
• Saccharomyces cerevisiae metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH

Ellipticine is a DNA-damaging agent acting as a prodrug whose pharmacological efficiencies and genotoxic side effects are dictated by activation with cytochrome P450 (CYP). Over the last decade we have gained extensive experience in using pure enzymes and various animal models that helped to identify CYPs metabolizing ellipticine. In this review we focus on comparison between the in vitro and in vivo studies and show a necessity of both approaches to obtain valid information on CYP enzymes contributing to ellipticine metabolism. Discrepancies were found between the CYP enzymes activating ellipticine to 13-hydroxy- and 12-hydroxyellipticine generating covalent DNA adducts and those detoxifying this drug to 9-hydroxy- and 7-hydroellipticine in vitro and in vivo. In vivo, formation of ellipticine-DNA adducts is dependent not only on expression levels of CYP3A, catalyzing ellipticine activation in vitro, but also on those of CYP1A that oxidize ellipticine in vitro mainly to the detoxification products. The finding showing that cytochrome b5 alters the ratio of ellipticine metabolites generated by CYP1A1/2 and 3A4 explained this paradox. Whereas the detoxification of ellipticine by CYP1A and 3A is either decreased or not changed by cytochrome b5, activation leading to ellipticine-DNA adducts increased considerably. We show that (I) the pharmacological effects of ellipticine mediated by covalent ellipticine-derived DNA adducts are dictated by expression levels of CYP1A, 3A and cytochrome b5, and its own potency to induce these enzymes in tumor tissues, (II) animal models, where levels of CYPs are either knocked out or induced are appropriate to identify CYPs metabolizing ellipticine in vivo, and (III) extrapolation from in vitro data to the situation in vivo is not always possible, confirming the need for these animal models.

• MeSH
• antitumorózní látky farmakologie   MeSH
• cytochrom P-450 CYP1A1 nedostatek   genetika   metabolismus   MeSH
• elipticiny farmakologie   MeSH
• hepatocyty účinky léků   metabolismus   MeSH
• krysa rodu rattus MeSH
• myši MeSH
• poškození DNA * MeSH
• rozpřahující látky farmakologie   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

We compared the effect of alpha-tocopheryl succinate (TOS) on succinate-dependent respiration in rat liver mitochondria, homogenate and permeabilized hepatocytes in both a coupled and uncoupled state. In isolated mitochondria, a significant inhibitory effect was observed at a concentration of 5 microM, in liver homogenate at 25 microM and in permeabilized hepatocytes at 50 microM. The inhibitory effect of TOS on succinate respiration in an uncoupled state was less pronounced than in a coupled state in all the experimental models tested. When the concentration dependence of the TOS inhibitory effect was tested, the most sensitive in both states were isolated mitochondria; the most resistant were permeabilized hepatocytes.

• MeSH
• alfa-tokoferol metabolismus   farmakologie   MeSH
• buněčná membrána metabolismus   MeSH
• buněčné dýchání účinky léků   MeSH
• časové faktory MeSH
• energetický metabolismus účinky léků   MeSH
• hepatocyty účinky léků   metabolismus   MeSH
• jaterní mitochondrie účinky léků   metabolismus   MeSH
• karbonylkyanid-p-trifluormethoxyfenylhydrazon farmakologie   MeSH
• oxidativní fosforylace účinky léků   MeSH
• permeabilita buněčné membrány MeSH
• potkani Wistar MeSH
• respirační komplex II metabolismus   MeSH
• rozpřahující látky farmakologie   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH

Whether active UCP1 can reduce ROS production in brown-fat mitochondria is presently not settled. The issue is of principal significance, as it can be seen as a proof- or disproof-of-principle concerning the ability of any protein to diminish ROS production through membrane depolarization. We therefore undertook a comprehensive investigation of the significance of UCP1 for ROS production, by comparing the ROS production in brown-fat mitochondria isolated from wildtype mice (that display membrane depolarization) or from UCP1(-/-) mice (with a high membrane potential). We tested the significance of UCP1 for glycerol-3-phosphate-supported ROS production by three methods (fluorescent dihydroethidium and the ESR probe PHH for superoxide, and fluorescent Amplex Red for hydrogen peroxide), and followed ROS production also with succinate, acyl-CoA or pyruvate as substrate. We studied the effects of the reverse electron flow inhibitor rotenone, the UCP1 activity inhibitor GDP, and the uncoupler FCCP. We also examined the effect of a physiologically induced increase in UCP1 amount. We noted GDP effects that were not UCP1-related. We conclude that only ROS production supported by exogenously added succinate was affected by the presence of active UCP1; ROS production supported by any other tested substrate (including endogenously generated succinate) was unaffected. This conclusion indicates that UCP1 is not involved in control of ROS production in brown-fat mitochondria. Extrapolation of these data to other tissues would imply that membrane depolarization may not necessarily decrease physiologically relevant ROS production. This article is a part of a Special Issue entitled: 18th European Bioenergetics Conference (Biochim. Biophys. Acta, Volume 1837, Issue 7, July 2014).

• MeSH
• elektronová paramagnetická rezonance MeSH
• glycerolfosfáty farmakologie   MeSH
• guanosindifosfát farmakologie   MeSH
• hnědá tuková tkáň metabolismus   MeSH
• imunoblotting MeSH
• iontové kanály genetika   metabolismus   MeSH
• karbonylkyanid-p-trifluormethoxyfenylhydrazon farmakologie   MeSH
• kyselina jantarová farmakologie   MeSH
• kyselina pyrohroznová farmakologie   MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• mitochondriální proteiny genetika   metabolismus   MeSH
• mitochondrie účinky léků   metabolismus   fyziologie   MeSH
• myši inbrední C57BL MeSH
• myši knockoutované MeSH
• nízká teplota MeSH
• peroxid vodíku metabolismus   MeSH
• protonové ionofory farmakologie   MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• spotřeba kyslíku účinky léků   MeSH
• superoxidy metabolismus   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

A series of twenty substituted 2-hydroxy-3-[(2-aryloxyethyl)amino]propyl 4-[(alkoxycarbonyl)amino]benzoates were prepared and characterized. As similar compounds have been described as potential antimycobacterials, primary in vitro screening of the synthesized carbamates was also performed against two mycobacterial species. 2-Hydroxy-3-[2-(2,6-dimethoxyphenoxy)ethylamino]-propyl 4-(butoxycarbonylamino)benzoate hydrochloride, 2-hydroxy-3-[2-(4-methoxyphenoxy)ethylamino]-propyl 4-(butoxycarbonylamino)benzoate hydrochloride, and 2-hydroxy-3-[2-(2-methoxyphenoxy)ethylamino]-propyl 4-(butoxycarbonylamino)benzoate hydrochloride showed higher activity against M. avium subsp. paratuberculosis and M. intracellulare than the standards ciprofloxacin, isoniazid, or pyrazinamide. Cytotoxicity assay of effective compounds was performed using the human monocytic leukaemia THP-1 cell line. Compounds with predicted amphiphilic properties were also tested for their effects on the rate of photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. All butyl derivatives significantly stimulated the rate of PET, indicating that the compounds can induce conformational changes in thylakoid membranes resulting in an increase of their permeability and so causing uncoupling of phosphorylation from electron transport.

• MeSH
• antibakteriální látky chemická syntéza   farmakologie   MeSH
• benzoáty chemická syntéza   farmakologie   MeSH
• karbamáty chemická syntéza   farmakologie   MeSH
• Mycobacterium avium subsp. paratuberculosis účinky léků   MeSH
• rozpřahující látky chemická syntéza   farmakologie   MeSH
• Spinacia oleracea účinky léků   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The overproduction of β-amyloid (Aβ) fragments in transgenic APPswe/PS1dE9 mice results in formation of amyloid deposits in the cerebral cortex and hippocampus starting around four months of age and leading to cognitive impairment much later. We have previously found an age and transgene-dependent weakening of muscarinic receptor-mediated transmission that was not present in young (6-10-week-old) animals but preceded both amyloid deposits and cognitive deficits. Now we investigated immediate and prolonged in vitro effects of non-aggregated Aβ(1-42) on coupling of individual muscarinic receptor subtypes expressed in CHO (Chinese hamster ovary) cells and their underlying mechanisms. Immediate application of 1 μM Aβ(1-42) had no effect on the binding of the muscarinic antagonist N-methylscopolamine or the agonist carbachol. In contrast, 4-day treatment of CHO cells expressing the M1 muscarinic receptor with 100 nM Aβ(1-42) significantly changed the binding characteristics of the muscarinic agonist carbachol and reduced the extent of the M1 receptor-stimulated breakdown of phosphatidylinositol while it did not demonstrate overt toxic effects. The treatment had no influence on the expression of either G-proteins or muscarinic receptors. In concert, we found no change in the gene expression of muscarinic receptor subtypes and gene or protein expression of the G(s), G(q/11), and G(i/o) G-proteins in the cerebral cortex of young adult APPswe/PS1dE9 mice that demonstrate high concentrations of soluble Aβ(1-42) and impaired muscarinic receptor-mediated G-protein activation. Our results provide strong evidence that the initial injurious effects of Aβ(1-42) on M1 muscarinic receptor-mediated transmissionis is due to compromised coupling of the receptor with G(q/11) G-protein.

• MeSH
• amyloidní beta-protein antagonisté a inhibitory   metabolismus   MeSH
• CHO buňky MeSH
• Cricetulus MeSH
• křečci praví MeSH
• kultivované buňky MeSH
• lidé MeSH
• myši transgenní MeSH
• myši MeSH
• peptidové fragmenty antagonisté a inhibitory   metabolismus   MeSH
• receptor muskarinový M1 metabolismus   MeSH
• receptory spřažené s G-proteiny antagonisté a inhibitory   metabolismus   MeSH
• rozpřahující látky farmakologie   MeSH
• vazba proteinů účinky léků   fyziologie   MeSH
• zvířata MeSH
• Check Tag
• křečci praví MeSH
• lidé MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Myocardial ischemia/reperfusion (IR) injury leads to structural changes in the heart muscle later followed by functional decline due to progressive fibrous replacement. Hence approaches to minimize IR injury are devised, including ischemic pre-and postconditioning. Mild uncoupling of oxidative phosphorylation is one of the mechanisms suggested to be cardioprotective as chemical uncoupling mimics ischemic preconditioning. Uncoupling protein 2 is proposed to be the physiological counterpart of chemical uncouplers and is thought to be a part of the protective machinery of cardiomyocytes. Morphological changes in the mitochondrial network likely accompany the uncoupling with mitochondrial fission dampening the signals leading to cardiomyocyte death. Here we review recent data on the role of uncoupling in cardioprotection and propose that low concentrations of dietary polyphenols may elicit the same cardioprotective effect as dinitrophenol and FCCP, perhaps accounting for the famed "French paradox".

• MeSH
• buněčná smrt účinky léků   MeSH
• dinitrofenoly terapeutické užití   MeSH
• fenoly terapeutické užití   MeSH
• flavonoidy terapeutické užití   MeSH
• iontové kanály metabolismus   MeSH
• ischemické přivykání MeSH
• karbonylkyanid-p-trifluormethoxyfenylhydrazon terapeutické užití   MeSH
• kardiomyocyty metabolismus   patologie   MeSH
• kardiotonika terapeutické užití   MeSH
• lidé MeSH
• mitochondriální proteiny metabolismus   MeSH
• myokard metabolismus   patologie   MeSH
• oxidativní fosforylace MeSH
• reperfuzní poškození myokardu farmakoterapie   metabolismus   patologie   MeSH
• rozpřahující látky terapeutické užití   MeSH
• signální transdukce účinky léků   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• práce podpořená grantem MeSH
• přehledy MeSH

Background: Discovery of uncoupling protein 2 (UCP2) in 1997 and demonstration of its wide tissue expressionhas triggered an important question about controlled oxidative phosphorylation uncoupling and the physiologicalfunction of this process. Uncoupling protein 2 (UcP2) is a mitochondrial protein that can infl uence the mitochondrialmembrane potential and hence the production of reactive oxygen species by mitochondria. It is also thought to beinvolved in apoptotic signaling pathways and it has been suggested to be important in cardio- and neuroprotection.Methods and results: We examined the recent literature (2003–2007) in the MedLine database for evidence linkingp38, one of the stress-related protein kinases, with modulation of UCP2 expression in the heart. While two reportsclearly demonstrate p38 as down-regulating UcP2 expression, only circumstantial evidence exists for cardiomyocytes.Confl icting results on p38-regulated cardiomyocyte survival after ischemia leave an open venue for hypotheses on thediff erential regulation of protein expression, including UCP2. Conclusions: Reviewing the evidence connecting UCP2 and its cytoprotective activities, we propose a tissue specifi clink that may explain the variable infl uence of p38 via modulation of UCP2 expression.

• MeSH
• financování organizované MeSH
• iontové kanály MeSH
• ischemická choroba srdeční enzymologie   MeSH
• kardiomyocyty metabolismus   účinky léků   MeSH
• kardiotonika farmakologie   terapeutické užití   MeSH
• lidé MeSH
• medicína založená na důkazech MeSH
• MEDLINE využití   MeSH
• mitochondriální proteiny MeSH
• mitogenem aktivované proteinkinasy p38 MeSH
• oxidativní fosforylace MeSH
• reaktivní formy kyslíku chemie   metabolismus   MeSH
• rozpřahující látky chemie   MeSH
• viabilita buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• přehledy MeSH

Complex I, i.e. proton-pumping NADH:quinone oxidoreductase, is an essential component of the mitochondrial respiratory chain but produces superoxide as a side-reaction. However, conditions for maximum superoxide production or its attenuation are not well understood. Unlike for Complex III, it has not been clear whether a Complex I-derived superoxide generation at forward electron transport is sensitive to membrane potential or protonmotive force. In order to investigate this, we used Amplex Red for H(2)O(2) monitoring, assessing the total mitochondrial superoxide production in isolated rat liver mitochondria respiring at state 4 as well as at state 3, namely with exclusive Complex I substrates or with Complex I substrates plus succinate. We have shown for the first time, that uncoupling diminishes rotenone-induced H(2)O(2) production also in state 3, while similar attenuation was observed in state 4. Moreover, we have found that 5-(N-ethyl-N-isopropyl) amiloride is a real inhibitor of Complex I H(+) pumping (IC(50) of 27 microM) without affecting respiration. It also partially prevented suppression by FCCP of rotenone-induced H(2)O(2) production with Complex I substrates alone (glutamate and malate), but nearly completely with Complexes I and II substrates. Sole 5-(N-ethyl-N-isopropyl) amiloride alone suppressed 20% and 30% of total H(2)O(2) production, respectively, under these conditions. Our data suggest that Complex I mitochondrial superoxide production can be attenuated by uncoupling, which means by acceleration of Complex I H(+) pumping due to the respiratory control. However, when this acceleration is prevented by 5-(N-ethyl-N-isopropyl) amiloride inhibition, no attenuation of superoxide production takes place.

• MeSH
• amilorid analogy a deriváty   farmakologie   MeSH
• biologické modely MeSH
• buněčné dýchání účinky léků   MeSH
• financování organizované MeSH
• jaterní mitochondrie enzymologie   účinky léků   MeSH
• krysa rodu rattus MeSH
• kyselina glutamová farmakologie   MeSH
• kyselina jantarová farmakologie   MeSH
• maláty farmakologie   MeSH
• peroxid vodíku metabolismus   MeSH
• potkani Wistar MeSH
• protonové pumpy metabolismus   MeSH
• respirační komplex I metabolismus   MeSH
• rozpřahující látky farmakologie   MeSH
• superoxidy metabolismus   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• zvířata MeSH