There are some indications that morphine may exert myocardial protective effects under certain conditions. The aim of the present study was to investigate the effect of morphine on viability and oxidative state of H9c2 cells (rat cardiomyoblasts) influenced by oxidative stress that was elicited by exposure to tert-butyl hydroperoxide (t-BHP). Our experiments showed that pretreatment with morphine before the addition of t-BHP markedly improved cell viability. Morphine was able to increase total antioxidant capacity of H9c2 cells and to reduce the production of reactive oxygen species, protein carbonylation, and lipid peroxidation. Cellular damage caused by t-BHP was associated with low levels of p38 MAPK and GSK-3β phosphorylation. Pretreatment with morphine augmented p38 phosphorylation, and the increased phospho-p38/p38 ratio was preserved even in the presence of t-BHP. Morphine did not change the level of GSK-3β phosphorylation, but interestingly, the phospho-GSK-3β/GSK-3β ratio significantly increased after subsequent incubation with t-BHP. Furthermore, morphine exposure resulted in upregulation of the antioxidant enzyme catalase. The protective effect of morphine was abrogated by the addition of the PI3K inhibitor wortmannin and/or p38 MAPK inhibitor SB203580. It can be concluded that morphine may protect H9c2 cells against oxidative stress and that this protection is at least partially mediated through activation of the p38 MAPK and PI3K/GSK-3β pathways.

• Klíčová slova
• Apoptosis, GSK-3β, H9c2 cells, Morphine, Oxidative stress, Reactive oxygen species, Tert-butyl hydroperoxide, p38 MAPK,
• MeSH
• 1-fosfatidylinositol-3-kinasa metabolismus   MeSH
• antioxidancia farmakologie   MeSH
• apoptóza účinky léků   MeSH
• buněčné linie MeSH
• cytoprotekce MeSH
• fosforylace MeSH
• GSK3B metabolismus   MeSH
• karbonylace proteinů účinky léků   MeSH
• kardiomyocyty účinky léků   metabolismus   patologie   MeSH
• krysa rodu Rattus MeSH
• mitogenem aktivované proteinkinasy p38 metabolismus   MeSH
• morfin farmakologie   MeSH
• nekróza MeSH
• oxidační stres účinky léků   MeSH
• oxidancia toxicita   MeSH
• peroxidace lipidů účinky léků   MeSH
• signální transdukce účinky léků   MeSH
• terc-butylhydroperoxid toxicita   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 1-fosfatidylinositol-3-kinasa MeSH
• antioxidancia MeSH
• Gsk3b protein, rat MeSH Prohlížeč
• GSK3B MeSH
• mitogenem aktivované proteinkinasy p38 MeSH
• morfin MeSH
• oxidancia MeSH
• terc-butylhydroperoxid MeSH

Neutrophil gelatinase-associated lipocalin is an extracellular protein produced mostly in kidney. Recently, it has become a promising biomarker of renal damage in vivo. On the other hand, the validation of NGAL as a biomarker for nephrotoxicity estimation in vitro has not been characterized in detail yet. Since the HK-2 cells are frequently used human kidney cell line, we aimed to characterize the production of NGAL in these cells and to evaluate NGAL as a possible marker of cell impairment. We used heavy metals (mercury, cadmium), peroxide, drugs (acetaminophen, gentamicin) and cisplatin to mimic nephrotoxicity. HK-2 cells were incubated with selected compounds for 1-24h and cell viability was measured together with extracellular NGAL production. We proved that HK-2 cells possess a capacity to produce NGAL in amount of 2pg/ml/h. We found a change in cell viability after 24h incubation with all tested toxic compounds. The largest decrease of the viability was detected in mercury, acetaminophen, cisplatin and gentamicin. Unexpectedly, we found also a significant decrease in NGAL production in HK-2 cells treated with these toxins for 24h: to 11±5%, 54±5%, 57±6% and 76±9% respectively, compared with controls (=100%). Our results were followed with qPCR analysis when we found no significant increase in LCN2 gene expression after 24h incubation. We conclude that extracellular NGAL production negatively correlates with HK-2 cell impairment.

• Klíčová slova
• Cell viability, HK-2 cells, In vitro nephrotoxicity, NGAL, Nephrotoxicity markers,
• MeSH
• akutní poškození ledvin chemicky indukované   metabolismus   MeSH
• biologické markery metabolismus   MeSH
• buněčné linie MeSH
• cisplatina toxicita   MeSH
• gentamiciny toxicita   MeSH
• kadmium toxicita   MeSH
• lidé MeSH
• lipokalin-2 genetika   metabolismus   MeSH
• paracetamol toxicita   MeSH
• rtuť toxicita   MeSH
• terc-butylhydroperoxid toxicita   MeSH
• viabilita buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• biologické markery MeSH
• cisplatina MeSH
• gentamiciny MeSH
• kadmium MeSH
• LCN2 protein, human MeSH Prohlížeč
• lipokalin-2 MeSH
• paracetamol MeSH
• rtuť MeSH
• terc-butylhydroperoxid MeSH

Epigallocatechin gallate (EGCG) is a green tea antioxidant with adverse effects on rat liver mitochondria and hepatocytes at high doses. Here, we assessed whether low doses of EGCG would protect these systems from damage induced by tert-butyl hydroperoxide (tBHP). Rat liver mitochondria or permeabilized rat hepatocytes were pretreated with EGCG and then exposed to tBHP. Oxygen consumption, mitochondrial membrane potential (MMP), and mitochondrial retention capacity for calcium were measured. First, 50 μM EGCG or 0.25 mM tBHP alone increased State 4 Complex I-driven respiration, thus demonstrating uncoupling effects; tBHP also inhibited State 3 ADP-stimulated respiration. Then, the coexposure to 0.25 mM tBHP and 50 μM EGCG induced a trend of further decline in the respiratory control ratio beyond that observed upon tBHP exposure alone. EGCG had no effect on MMP and no effect, in concentrations up to 50 μM, on mitochondrial calcium retention capacity. tBHP led to a decline in both MMP and mitochondrial retention capacity for calcium; these effects were not changed by pretreatment with EGCG. In addition, EGCG dose-dependently enhanced hydrogen peroxide formation in a cell- and mitochondria-free medium. Conclusion. Moderate nontoxic doses of EGCG were not able to protect rat liver mitochondria and hepatocytes from tBHP-induced mitochondrial dysfunction.

• MeSH
• hepatocyty cytologie   účinky léků   metabolismus   MeSH
• jaterní mitochondrie účinky léků   metabolismus   MeSH
• katechin analogy a deriváty   farmakologie   MeSH
• krysa rodu Rattus MeSH
• kultivované buňky MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• peroxid vodíku metabolismus   MeSH
• potkani Wistar MeSH
• spotřeba kyslíku účinky léků   MeSH
• terc-butylhydroperoxid toxicita   MeSH
• vápník metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• epigallocatechin gallate MeSH Prohlížeč
• katechin MeSH
• peroxid vodíku MeSH
• terc-butylhydroperoxid MeSH
• vápník MeSH

Oxidative stress and mitochondrial dysfunction play an important role in the pathogenesis of nonalcoholic fatty liver disease and toxic liver injury. The present study was designed to evaluate the effect of exogenous inducer of oxidative stress (tert-butyl hydroperoxide, tBHP) on nonfatty and steatotic hepatocytes isolated from the liver of rats fed by standard and high-fat diet, respectively. In control steatotic hepatocytes, we found higher generation of ROS, increased lipoperoxidation, an altered redox state of glutathione, and decreased ADP-stimulated respiration using NADH-linked substrates, as compared to intact lean hepatocytes. Fatty hepatocytes exposed to tBHP exert more severe damage, lower reduced glutathione to total glutathione ratio, and higher formation of ROS and production of malondialdehyde and are more susceptible to tBHP-induced decrease in mitochondrial membrane potential. Respiratory control ratio of complex I was significantly reduced by tBHP in both lean and steatotic hepatocytes, but reduction in NADH-dependent state 3 respiration was more severe in fatty cells. In summary, our results collectively indicate that steatotic rat hepatocytes occur under conditions of enhanced oxidative stress and are more sensitive to the exogenous source of oxidative injury. This confirms the hypothesis of steatosis being the first hit sensitizing hepatocytes to further damage.

• MeSH
• dieta s vysokým obsahem tuků MeSH
• glutathion metabolismus   MeSH
• hepatocyty účinky léků   metabolismus   MeSH
• krysa rodu Rattus MeSH
• kultivované buňky MeSH
• L-laktátdehydrogenasa metabolismus   MeSH
• malondialdehyd metabolismus   MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• nealkoholová steatóza jater metabolismus   patologie   MeSH
• oxidační stres účinky léků   MeSH
• peroxidace lipidů účinky léků   MeSH
• potkani Wistar MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• terc-butylhydroperoxid toxicita   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• glutathion MeSH
• L-laktátdehydrogenasa MeSH
• malondialdehyd MeSH
• reaktivní formy kyslíku MeSH
• terc-butylhydroperoxid MeSH

The aim of the study was to evaluate time course and dose dependence of peroxidative damage induced by tert-butyl hydroperoxide (tBHP) in rat hepatocytes cultured in suspension and in monolayer. At the lowest (0.1 mM) concentration, decrease of cytosolic glutathione and discharge of mitochondrial membrane potential (MMP) could be detected. Significant increases in leakage of lactate dehydrogenase and in malondialdehyde concentrations together with decrease of pyruvate-dependent respiration were detected at higher tBHP concentrations (above 0.5 mM) and after longer periods of incubation. Changes in plasma membrane integrity were observed at 1 mM concentration of tBHP. Succinate-dependent oxidation was most resistant to peroxidative damages. Opening of the mitochondrial permeability transition pore was responsible for the discharge of mitochondria membrane potential. In the presence of cyclosporine A and succinate, the membrane potential could be restored. Our data showed that the most sensitive indicators of the peroxidative damage are changes of cytosolic glutathione concentration and MMP.

• MeSH
• cytosol účinky léků   metabolismus   MeSH
• glutathion metabolismus   MeSH
• hepatocyty účinky léků   metabolismus   MeSH
• jaterní mitochondrie účinky léků   enzymologie   fyziologie   MeSH
• krysa rodu Rattus MeSH
• kultivované buňky MeSH
• L-laktátdehydrogenasa metabolismus   MeSH
• malondialdehyd metabolismus   MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• oxidační stres účinky léků   fyziologie   MeSH
• oxidancia toxicita   MeSH
• peroxidace lipidů účinky léků   MeSH
• potkani Wistar MeSH
• spotřeba kyslíku účinky léků   MeSH
• terc-butylhydroperoxid toxicita   MeSH
• viabilita buněk účinky léků   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• glutathion MeSH
• L-laktátdehydrogenasa MeSH
• malondialdehyd MeSH
• oxidancia MeSH
• terc-butylhydroperoxid MeSH

The aim of this work was to study the effects of resveratrol (RES) as compared to silymarin (SM) pretreatments on tert-butylhydroperoxide (tBH) induced apoptotic/necrotic markers in hepatocytes. Hepatocyte in cultures (48 h) and in perifused immobilized agarose threads (5h) were used as cellular systems. Hepatocyte apoptosis was estimated morphologically using Annexin-V combined with propidium iodide, or toluidine blue staining. Hepatocyte viability and functionality were evaluated by ALT and urea synthesis. Nitric oxide (NO) and carbon monoxide involvements were also examined. Resveratrol and silymarin reduced tBH-induced hepatocyte toxic effects in short term experiments (5h) as measured by a significant reduction in ALT and NO increase produced by tBH. Both inducible nitric oxide synthase (NOS-2) and hemoxygenase-1 (HO-1) gene expression were increased by tBH and reduced by both RES and SM pretreatments. Morphologically, there were ameliorations in both apoptotic and necrotic markers under RES treatment and were similar to biochemical findings. In addition, RES improved hepatocyte stability in both cellular systems. It may be concluded that resveratrol and sylimarin ameliorative effects on tBH hepatocyte toxicity are comparable; involve NOS-2 and HO-1 expression and should be re-evaluated in various in vitro and in vivo experimental conditions.

• MeSH
• alanintransaminasa metabolismus   MeSH
• analýza rozptylu MeSH
• apoptóza účinky léků   MeSH
• cytoprotekce MeSH
• hemoxygenasa-1 metabolismus   MeSH
• hepatocyty účinky léků   metabolismus   ultrastruktura   MeSH
• imobilizované buňky MeSH
• krysa rodu Rattus MeSH
• kultivované buňky MeSH
• nádorové biomarkery metabolismus   MeSH
• oxid dusnatý metabolismus   MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• potkani Wistar MeSH
• resveratrol MeSH
• silymarin farmakologie   MeSH
• stilbeny farmakologie   MeSH
• synthasa oxidu dusnatého, typ II metabolismus   MeSH
• terc-butylhydroperoxid toxicita   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• alanintransaminasa MeSH
• hemoxygenasa-1 MeSH
• nádorové biomarkery MeSH
• oxid dusnatý MeSH
• resveratrol MeSH
• silymarin MeSH
• stilbeny MeSH
• synthasa oxidu dusnatého, typ II MeSH
• terc-butylhydroperoxid MeSH

Two classes of newly synthesized amphiphilic compounds, phenolic antioxidants ("phenolics") and N-oxides exert in vivo antioxidant effects on live S. cerevisiae cells. Both groups have low toxicity, phenolics being more toxic than N-oxides and compounds with a longer alkyl chain having higher toxicity than those with a shorter alkyl chain. Phenolic antioxidants protect yeast cells exposed to the superoxide producer paraquat and peroxyl generator tert-butylhydroperoxide better than N-oxides at 3-fold higher concentration. Both types of antioxidants enhance the survival of pro-oxidant-exposed cells of S. cerevisiae mutants deficient in cytosolic and/or mitochondrial superoxide dismutase and could be good compounds which mimic the role of superoxide dismutases. The results of measurement of antioxidant activity in an in vitro chemiluminescence test differ from the results obtained in vivo with S. cerevisiae superoxide dismutase mutants. In contrast to their action on live cells, phenolics are less effective than N-oxides in preventing lipid peroxidation of an emulsion of lipids isolated from S. cerevisiae membranes.

• MeSH
• aminy chemie   farmakologie   toxicita   MeSH
• antifungální látky toxicita   MeSH
• antioxidancia chemie   farmakologie   toxicita   MeSH
• delece genu MeSH
• fenoly chemie   farmakologie   toxicita   MeSH
• kvartérní amoniové sloučeniny chemie   farmakologie   toxicita   MeSH
• membránové lipidy metabolismus   MeSH
• mikrobiální viabilita MeSH
• paraquat toxicita   MeSH
• peroxidace lipidů MeSH
• Saccharomyces cerevisiae účinky léků   MeSH
• superoxiddismutasa genetika   MeSH
• terc-butylhydroperoxid toxicita   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• aminy MeSH
• antifungální látky MeSH
• antioxidancia MeSH
• fenoly MeSH
• kvartérní amoniové sloučeniny MeSH
• membránové lipidy MeSH
• paraquat MeSH
• superoxiddismutasa MeSH
• terc-butylhydroperoxid MeSH

The majority of toxic agents act either fully or partially via oxidative stress, the liver, specifically the mitochondria in hepatocytes, being the main target. Maintenance of mitochondrial function is essential for the survival and normal performance of hepatocytes, which have a high energy requirement. Therefore, greater understanding of the role of mitochondria in hepatocytes is of fundamental importance. Mitochondrial function can be analysed in several basic models: hepatocytes cultured in vitro; mitochondria in permeabilised hepatocytes; and isolated mitochondria. The aim of our study was to use all of these approaches to evaluate changes in mitochondria exposed in vitro to a potent non-specific peroxidating agent, tert-butylhydroperoxide (tBHP), which is known to induce oxidative stress. A decrease in the mitochondrial membrane potential (MMP) was observed in cultured hepatocytes treated with tBHP, as illustrated by a significant reduction in Rhodamine 123 accumulation and by a decrease in the fluorescence of the JC-1 molecular probe. Respiratory Complex I in the mitochondria of permeabilised hepatocytes showed high sensitivity to tBHP, as documented by high-resolution respirometry. This could be caused by the oxidation of NADH and NADPH by tBHP, followed by the disruption of mitochondrial calcium homeostasis, leading to the collapse of the MMP. A substantial decrease in the MMP, as determined by tetraphenylphosphonium ion-selective electrode measurements, also confirmed the dramatic impact of tBHP-induced oxidative stress on mitochondria. Swelling was observed in isolated mitochondria exposed to tBHP, which could be prevented by cyclosporin A, which is evidence for the role of mitochondrial permeability transition. Our results demonstrate that all of the above-mentioned models can be used for toxicity assessment, and the data obtained are complementary.

• MeSH
• alternativy testů na zvířatech MeSH
• hepatocyty účinky léků   metabolismus   patologie   MeSH
• jaterní mitochondrie účinky léků   metabolismus   MeSH
• krysa rodu Rattus MeSH
• kultivované buňky MeSH
• kyslík analýza   metabolismus   MeSH
• manometrie MeSH
• membránový potenciál mitochondrií účinky léků   fyziologie   MeSH
• oxidační stres * MeSH
• oxidancia toxicita   MeSH
• potkani Wistar MeSH
• spotřeba kyslíku MeSH
• terc-butylhydroperoxid toxicita   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zduření mitochondrií účinky léků   fyziologie   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kyslík MeSH
• oxidancia MeSH
• terc-butylhydroperoxid MeSH

Amphiphilic 3-(alkanoylamino)propyldimethylamine-N-oxides with different length of the alkyl chain, i.e. different hydrophilic-lipophilic balance, act in micromolar concentrations as SOD mimics by lifting the inhibition of aerobic growth caused by SOD deletions in Saccharomyces cerevisiae. They also enhance the survival of sod mutants of S. cerevisiae exposed to the hydrophilic superoxide-generating prooxidant paraquat and the amphiphilic hydroperoxide-producing tert-butylhydroperoxide (TBHP), and largely prevent TBHP-induced peroxidation of isolated yeast plasma membrane lipids. Unlike the SOD-mimicking effect, the magnitude of these effects depends on the alkyl chain length of the amine-N-oxides, which incorporate into S. cerevisiae membranes, causing fluidity changes in both the hydrophilic surface part of the membrane and the membrane lipid matrix. Unlike wild-type strains, the membranes of sod mutants were found to contain polyunsaturated fatty acids; the sensitivity of the mutants to lipophilic pro-oxidants was found to increase with increasing content of these acids. sod mutants are useful in assessing pro- and antioxidant properties of different compounds.

• MeSH
• antioxidancia farmakologie   MeSH
• dimethylaminy farmakologie   MeSH
• fluidita membrány účinky léků   MeSH
• membránové lipidy chemie   metabolismus   MeSH
• oxidační stres MeSH
• paraquat toxicita   MeSH
• peroxidace lipidů účinky léků   MeSH
• Saccharomyces cerevisiae účinky léků   enzymologie   MeSH
• superoxiddismutasa chemie   genetika   metabolismus   MeSH
• terc-butylhydroperoxid toxicita   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antioxidancia MeSH
• dimethylaminy MeSH
• membránové lipidy MeSH
• paraquat MeSH
• superoxiddismutasa MeSH
• terc-butylhydroperoxid MeSH

The aim of this study was to evaluate the cytoprotective effects upon primary human hepatocytes of silymarin extract and its main flavonolignans following exposure to the cytotoxic actions of model toxins. The conditions for the hepatocyte intoxication were optimised for allyl alcohol, carbon tetrachloride, D-galactosamine and paracetamol. Silymarin extract, silychristin and silydianin did not exert cytotoxicity (10-100 microM), whereas silybin and isosilybin at higher concentrations and after longer incubation periods were cytotoxic. All main flavonolignans of silymarin tested displayed concentration-dependent cytoprotection against the toxic effects of both allyl alcohol and carbon tetrachloride but neither paracetamol nor galactosamine. The best protection was achieved by silydianin and silychristin and to a lesser degree by silymarin, while silybin and isosilybin were less effective. It is concluded that these differing outcomes result from the varying abilities of the Silybum marianum substances tested to stabilize the cell membrane, exert antioxidant properties and exhibit intrinsic toxicity.

• MeSH
• antioxidancia farmakologie   MeSH
• Asteraceae chemie   MeSH
• bifenylové sloučeniny MeSH
• biologické toxiny toxicita   MeSH
• flavonoidy farmakologie   MeSH
• galaktosamin metabolismus   MeSH
• hepatocyty účinky léků   enzymologie   MeSH
• kultivované buňky MeSH
• L-laktátdehydrogenasa metabolismus   MeSH
• lidé MeSH
• lignany farmakologie   MeSH
• neopioidní analgetika farmakologie   MeSH
• paracetamol farmakologie   MeSH
• pikráty metabolismus   MeSH
• propanoly toxicita   MeSH
• proteosyntéza MeSH
• rostlinné extrakty toxicita   MeSH
• scavengery volných radikálů farmakologie   MeSH
• separace buněk MeSH
• terc-butylhydroperoxid toxicita   MeSH
• viabilita buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 1,1-diphenyl-2-picrylhydrazyl MeSH Prohlížeč
• allyl alcohol MeSH Prohlížeč
• antioxidancia MeSH
• bifenylové sloučeniny MeSH
• biologické toxiny MeSH
• flavonoidy MeSH
• galaktosamin MeSH
• L-laktátdehydrogenasa MeSH
• lignany MeSH
• neopioidní analgetika MeSH
• paracetamol MeSH
• pikráty MeSH
• propanoly MeSH
• rostlinné extrakty MeSH
• scavengery volných radikálů MeSH
• terc-butylhydroperoxid MeSH