Rat Liver Mitochondrial Function Research Articles

The Novel Arylamidine T-2307 Selectively Disrupts Yeast Mitochondrial Function by Inhibiting Respiratory Chain Complexes.

The novel arylamidine T-2307 exhibits broad-spectrum in vitro and in vivo antifungal activities against clinically significant pathogens. Previous studies have shown that T-2307 accumulates in yeast cells via a specific polyamine transporter and disrupts yeast mitochondrial membrane potential. Further, it has little effect on rat liver mitochondrial function. The mechanism by which T-2307 disrupts yeast mitochondrial function is poorly understood, and its elucidation may provide important information for developing novel antifungal agents. This study aimed to determine how T-2307 promotes yeast mitochondrial dysfunction and to investigate the selectivity of this mechanism between fungi and mammals. T-2307 inhibited the respiration of yeast whole cells and isolated yeast mitochondria in a dose-dependent manner. The similarity of the effects of T-2307 and respiratory chain inhibitors on mitochondrial respiration prompted us to investigate the effect of T-2307 on mitochondrial respiratory chain complexes. T-2307 particularly inhibited respiratory chain complexes III and IV not only in Saccharomyces cerevisiae but also in Candida albicans, indicating that T-2307 acts against pathogenic fungi in a manner similar to that of yeast. Conversely, T-2307 showed little effect on bovine respiratory chain complexes. Additionally, we demonstrated that the inhibition of respiratory chain complexes by T-2307 resulted in a decrease in the intracellular ATP levels in yeast cells. These results indicate that inhibition of respiratory chain complexes III and IV is a key factor for selective disruption of yeast mitochondrial function and antifungal activity.

Effect of bedaquiline on the functions of rat liver mitochondria

The paper considers the effects of bedaquiline (BDQ), an antituberculous preparation of the new generation, on rat liver mitochondria. It was shown that 50 μM BDQ inhibited mitochondrial respiration measured with substrates of complexes I and II (glutamate/malate and succinate/rotenone systems respectively) in the states V3 and VDNP. At the same time, at concentrations below 50 μM, BDQ slightly stimulated respiration with substrates of complex I in the state V2. BDQ was also found to suppress, in a dose-dependent manner, the activity of complex II and the total activity of complexes II + III of the mitochondrial transport chain. It was discovered that at concentrations up to 10 μM, BDQ inhibited H2O2 production in mitochondria. BDQ (10–50 μM) suppressed the opening of Ca2+-dependent CsA-sensitive mitochondrial permeability transition pore. The latter was revealed experimentally as the inhibition of Ca2+/Pi-dependent swelling of mitochondria, suppression of cytochrome c release, and an increase in the Ca2+ capacity of the organelles. BDQ also decreased the rate of mitochondrial energy-dependent K+ transport, which was evaluated by the energy-dependent swelling of mitochondria in a K+ buffer and DNP-induced K+ efflux from the organelles. The possible mechanisms of BDQ effect of rat liver mitochondria are discussed.

Effects of the Nonsteroidal Anti-inflammatory Drug Celecoxib on Mitochondrial Function.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are widely used to treat inflammation and pain. In the present study, we examined the effects of celecoxib, a cyclooxygenase-2 (COX-2)-selective NSAID, on rat liver mitochondrial function. Celecoxib dose-dependently induced mitochondria swelling, which was not suppressed by cyclosporine A (CsA). The oxygen consumption rate in mitochondria-suspended solution was facilitated by the addition of celecoxib, and its uncoupling activity was observed. Celecoxib also suppressed SF6847-induced uncoupling, and appeared to exert inhibitory effects on the electron transport chain. Celecoxib suppressed the state 3 oxygen consumption rate in the presence of ADP. Protein release from the mitochondrial matrix was detected following the addition of celecoxib, and aldehyde dehydrogenase 2 (ALDH2) and hydroxymethylglutaryl-CoA (HMG-CoA) synthase 2 (HMGCS2) bands were confirmed in a Western blot analysis. On the other hand, protein release of cytochrome C (CytC), which is an inducer of apoptosis, from the intermembrane space was not observed. Celecoxib enhanced the membrane permeability of human erythrocytes and synthesized liposomes dose-dependently. It then induced the membrane-involving mitochondrial swelling and suppressed mitochondrial function.

Coenzyme Q10 remarkably improves the bio-energetic function of rat liver mitochondria treated with statins

CoQ10 shares a biosynthetic pathway with cholesterol therefore it can be a potential target of the widely available lipid-lowering agents such as statins. Statins are the most widely prescribed cholesterol-lowering drugs with the ability to inhibit HMG-CoA (3-hydroxy-3-methylglutaryl-coenzyme A) reductase. Preclinical and clinical safety data have shown that statins do not cause serious adverse effects in humans. However, their long-term administration is associated with a variety of myopatic complaints. The aim of this study was to investigate whether CoQ10 supplementation of animals under high fat diet (HFD) treated with statins is able to bypass the mitochondrial metabolic defects or not?Animals were divided into 7 groups and fed with either regular (RD) or HFD during experiments. The first group considered as regular control and fed with a RD. Groups 2–7 including HFD control, CoQ10 (10mg/kg), simvastatin (30mg/kg), atorvastatin (30mg/kg), simvastatin+CoQ10 or atorvastatin+CoQ10 treated orally for 30 days and fed with HFD. At the end of treatments, the animals were killed and blood samples were collected for biochemical examinations. The rat liver mitochondria were isolated and several mitochondrial indices including succinate dehydrogenase activity (SDA), ATP levels, mitochondrial membrane potential (MMP) and mitochondrial permeability transition pore (MPP) were determined.We found that triglyceride (Tg), cholesterol (Chol) and low-density lipoprotein (LDL) were augmented with HFD compared to RD and treatment with statins remarkably lowered the Tg, Chol and LDL levels. Mitochondrial parameters including, SDA, ATP levels, MMP and MPP were reduced with statin treatment and improved by co-administration with CoQ10.

The antiestrogen 4-hydroxytamoxifen protects against isotretinoin-induced permeability transition and bioenergetic dysfunction of liver mitochondria: comparison with tamoxifen

The combination of isotretinoin (13-cis-retinoic acid) with antiestrogens seems to be a promising strategy for cancer chemotherapy. The aim of the study was to evaluate the effects of isotretinoin alone or in combination with 4-hydroxytamoxifen (OHTAM) and with its prodrug tamoxifen (TAM), on the functions of rat liver mitochondria, i.e., mitochondrial permeability transition (MPT), bioenergetic functions and adenine nucleotide translocase (ANT). Isotretinoin (5 nmol/mg protein) induced the Ca²⁺-dependent MPT pore opening in mitochondria energized with succinate, which was prevented by OHTAM, cyclosporine A, TAM and ANT ligands. When mitochondria were energized with glutamate/malate and in the absence of added Ca²⁺ isotretinoin decreased the state 3 respiration, the ATP levels, the active ANT content and increased the lag phase of the phosphorylation cycle, demonstrating that isotretinoin decreased the mitochondrial phosphorylation efficiency. These changes of isotretinoin in bioenergetic parameters were not significant in the presence of succinate. The effects of isotretinoin at 5 nmol/mg protein on the Ca²⁺-dependent MPT and phosphorylative efficacy may be related with interactions with the ANT. Above 10 nmol/mg protein isotretinoin strongly diminished the active ANT content, decreased the Δψ, inhibited the complex I and induced proton leak through the Fo fraction of complex V. The combination of OHTAM with isotretinoin only induced significant changes in the energy production systems at concentrations ≥5 nmol isotretinoin/mg protein. Therefore, our results suggest that isotretinoin-associated liver toxicity is possibly related with mitochondrial dysfunctions and that the combination with OHTAM may contribute to decrease its toxicity.

Effects of all-trans-retinoic acid on the permeability transition and bioenergetic functions of rat liver mitochondria in combination with endoxifen

AimsThe clinical utilization of the combinations of all-trans-retinoic acid (RA) with antiestrogens, which present synergism of action in breast cancer, has been limited by RA adverse effects, including hepatotoxicity, which may be related with mitochondrial damage. This work evaluated the effects of RA alone and in combination with the antiestrogen endoxifen (EDX) on liver mitochondria. Main methodsMitochondrial permeability transition (MPT) was assessed by using Calcium Green-5N fluorescence and a tetraphenylphosphonium selective electrode. Oxidative stress was evaluated by oxygen consumption and thiobarbituric acid method. Mitochondrial bioenergetic was monitored by measuring oxygen consumption and mitochondrial membrane potential (ΔΨ). Osmotic volume changes of mitochondria were followed at 540nm. Key findingsEDX prevents the MPT induced by RA, allowing mitochondria pre-incubated with RA to accumulate Ca2+ and inhibiting the depolarization of ΔΨ. RA above 10nmol/mg protein depresses the phosphorylation capacity of mitochondria, as shown by the increase in the time required for ADP phosphorylation as well as by the decrease in state 3 respiration. At 20nmol/mg protein, RA decreases the ΔΨ and increases the state 4 respiration, suggesting that high concentrations of RA permeabilize the membrane to protons, possibly due to a proton leak through the Fo fraction of complex V. Moreover, the effects of RA on mitochondrial bioenergetics are not changed by EDX. SignificanceRA-induced hepatotoxicity may be related with induction of MPT and alterations in bioenergetic parameters; the combination with EDX, which reduces mitochondrial dysfunction and synergistically potentiates the anticancer activity, may provide a safer therapeutic strategy.

Polyethylenimine-mediated impairment of mitochondrial membrane potential, respiration and membrane integrity: Implications for nucleic acid delivery and gene therapy

The 25kDa branched polyethylenimine (PEI) is a highly efficient synthetic polycation used in transfection protocols, but also triggers mitochondrial-mediated apoptotic cell death processes where the mechanistic issues are poorly understood. We now demonstrate that PEI in a concentration- and time-dependent manner can affect functions (membrane potential, swelling and respiration) and ultrastructural integrity of freshly isolated rat liver mitochondria. The threshold concentration for detection of PEI-mediated impairment of rat liver mitochondrial functions is 3μg/mL, however, lower PEI levels still exert some effects on mitochondrial morphology and respiration, and these may be related to the inherent membrane perturbing properties of this polycation. The PEI-mediated mitochondrial swelling phase is biphasic, with a fast decaying initial period (most prominent from 4μg/mL PEI) followed by a slower, linear swelling response. The slow phase is presumably the result of a time-dependent transition permeability opening in mitochondria initially resistant to swelling/depolarization, but may further be related to PEI-induced nanoscale structural defects and/or formation of pores in the outer membrane. Respiration assessments further suggested that PEI in the presence of exogenous ADP behaves in a similar fashion to a slow-acting inhibitory compound. PEI further shows an uncoupling property that is detectable at low respiration rates. The relevance of these findings to PEI-mediated initiation of intrinsic apoptotic pathway is discussed.

Rat liver mitochondrial membrane characteristics and mitochondrial functions are more profoundly altered by dietary lipid quantity than by dietary lipid quality: effect of different nutritional lipid patterns

Dietary lipids are known to affect the composition of the biological membrane and functions that are involved in cell death and survival. The mitochondrial respiratory chain enzymes are membrane protein complexes whose function depends on the composition and fluidity of the mitochondrial membrane lipid. The present study aimed at investigating the impact of different nutritional patterns of dietary lipids on liver mitochondrial functions. A total of forty-eight Wistar male rats were divided into six groups and fed for 12 weeks with a basal diet, lard diet or fish oil diet, containing either 50 or 300g lipid/kg. The 30% lipid intake increased liver NEFA, TAG and cholesterol levels, increased mitochondrial NEFA and TAG, and decreased phospholipid (PL) levels. SFA, PUFA and unsaturation index (UI) increased, whereas MUFA and trans-fatty acids (FA) decreased in the mitochondrial membrane PL in 30% fat diet-fed rats compared with 5% lipid diet-fed rats. PL UI increased with fish oil diet v. basal and lard-rich diets, and PL trans-FA increased with lard diet v. basal and fish oil diets. The 30% lipid diet intake increased mitochondrial membrane potential, membrane fluidity, mitochondrial respiration and complex V activity, and decreased complex III and IV activities. With regard to lipid quality effects, β-oxidation decreased with the intake of basal or fish oil diets compared with that of the lard diet. The intake of a fish oil diet decreased complex III and IV activities compared with both the basal and lard diets. In conclusion, the characteristics and mitochondrial functions of the rat liver mitochondrial membrane are more profoundly altered by the quantity of dietary lipid than by its quality, which may have profound impacts on the pathogenesis and development of non-alcoholic fatty liver disease.

Comparative study of the effects of 1,3,4-thiadiazolium mesoionic derivatives on energy-linked functions of rat liver mitochondria

The main goal of this work was to investigate the relationship between the effects of three new 1,3,4-thiadiazolium mesoionic derivatives on mitochondrial bioenergetics and their previously described chemical structure and antimelanoma activity. The 4-phenyl-5-(2′-Y, 4′-X or 4′-X-cinnamoyl)-1,3,4-thiadiazolium-2-phenylamine chlorides differed from each other only in the cinnamoyl ring substituent: MI-J, X = OH; MI-F, X = F; MI-2,4diF X = Y = F. The state 3 respiratory rate was strongly decreased by all derivatives, reaching total inhibition of MI-4F and MI-2,4diF (130 nmol mg −1 protein), when glutamate plus malate were used as substrate. State 3 inhibition was less accentuated with succinate as substrate. Analyses of segments of the respiratory chain indicated complexes I and IV as sites inhibited by the derivatives. State 4 respiration was strongly stimulated by the three derivatives, and was characterized as an uncoupling effect, which was more intense for MI-4F. This stimulus was so pronounced that the values of RCC and ADP/O ratio were only calculated for the lowest concentration (6.5 nmol mg −1 protein). In intact mitochondria, the ATPase activity was increased dramatically by ∼120%, ∼207% and ∼261% for MI-J, MI-4F and MI-2,4diF (32.5 nmol mg −1 protein), respectively. In conclusion, the presence of fluorine substituent in the cinnamoyl ring intensifies the effect of mesoionic compounds on mitochondrial functions and, in this context, hydrophobicity is more important than the electronic effect, which was correlated to antimelanoma activity described previously for these compounds.

Comparative effects of 3,4-methylenedioxymethamphetamine and 4-methylthioamphetamine on rat liver mitochondrial function

Ecstasy, which is used as a recreational drug, is a common street name for 3, 4-methylenedioxymethamphetamine (MDMA). Another drug of abuse chemically related, though less common than MDMA, is the amphetamine derivative 4-methylthioamphetamine (MTA). MDMA and MTA induce different systemic and organ-specific effects, including neurotoxicity, hyperthermia, nephrotoxicity, cardiotoxicity and hepatotoxicity. Therefore, it is clear that MDMA and MTA are responsible for inducing organ toxicity. The mechanisms underlying MDMA and MTA-induced hepatotoxicity are multifactorial, and therefore not completely understood. Recent findings indicate interference with cellular bioenergetics as an important toxicological feature of ecstasy. However, less is known about the involvement of mitochondria in MTA-induced hepatotoxicity. Thus, we compared the direct influence of MDMA and MTA on rat liver mitochondrial function [mitochondrial permeability transition (MPT), mitochondrial oxidative stress, and mitochondrial bioenergetics]. It was shown that MTA (from 0.025 up to 0.1 mM) was more potent than MDMA (from 0.2 up to 0.5 mM) in decreasing the sensitivity of rat liver mitochondria to MPT. However, higher concentrations of MTA (from 0.5 up to 2 mM) were highly toxic to mitochondria. MTA simultaneously increased H 2O 2 production in a monoamine oxidase (MAO)-dependent way, and uncoupled and inhibited mitochondrial respiration. In contrast, MDMA had only limited or no effects on these mitochondrial parameters. According to these results, it is possible to postulate that, depending on the concentration, MTA can potentially be more efficient in its effects on liver mitochondria than MDMA and, also, that its harmful effects may contribute to its hepatotoxicity.

Long-term High-fat-diet Feeding Impairs Mitochondrial Biogenesis in Liver of Male and Female Rats

Background/Aims: Mitochondrial biogenesis includes both mitochondrial proliferation and differentiation and its regulation under different physiological conditions is not clear. Given the sexual dimorphism previously found in mitochondrial function, the aim of this study was to investigate the gender-dependent effect of chronic high-fat-diet (HFD) feeding on rat liver mitochondrial function and biogenesis. Methods: Ten-week old male and female rats were fed a HFD (26% fat) or a control diet (2.9% fat) for 26 weeks. Mitochondrial morphology was studied. Mitochondrial DNA and protein content, hydrogen peroxide production, oxidative capacity, antioxidant defenses, as well as markers of oxidative damage and mitochondrial biogenesis were analyzed. Results: Female rats showed higher levels of mitochondrial protein and an enhanced oxidative capacity per mitochondrion than males. In both genders, HFD feeding increased mtDNA content and decreased mitochondrial differentiation markers. Conclusion: In comparison to male rats, females show higher oxidative capacity as a consequence of their greater mitochondrial differentiation under both control and obese status. In response to HFD feeding, the oxidative capacity of the whole mitochondrial population is maintained in both genders. This is obtained by means of an enhancement of mitochondrial proliferation, which counteracts the diet-induced impairment of the function of each mitochondrion.

Effects of ischemia-reperfusion and pretreatment with mildronate on rat liver mitochondrial function

Mildronate (3-(2,2,2-trimethylhydrazinium) propionate), which is mostly used in cardiological practice and is considered an anti-ischemic drug, was designed to inhibit carnitine biosynthesis in order to prevent accumulation of cytotoxic intermediate products of fatty acid beta-oxidation. Recently it was shown that the mitochondrial respiratory chain may also be a target for mildronate action. In this study, we aimed to investigate whether mildronate can protect the liver against a 90-min normothermic ischemia/30-min reperfusion-induced mitochondrial dysfunction. Rats were pre-treated for one or two weeks with mildronate (100mg/kg/day or 200mg/kg/day) or Ringer solution and subjected to ischemia/reperfusion. We found that ischemia/reperfusion caused a decrease in mitochondrial State 3 respiration rate and in the respiratory control index (RCI), and an increase in State 2 respiration rate with succinate, glutamate + malate and palmitoyl-L-carnitine + malate. One or two weeks of pre-treatment of rats with different doses of mildronate did not reduce the ischemia/reperffusion-induced decrease in the State 3 respiration rate or RCI; however, a one week pre-treatment slightly diminished the increase in the State 2 respiration rate with glutamate + malate substrates. The leakage of the liver enzymes, aspartate aminotransferase, alanine aminotransferase and lactate dehydrogenase, was similar in both the untreated and pre-treated with mildronate groups. No steatotic livers were observed in any experimental groups after mildronate pre-treatment. In conclusion, 90min of liver ischemia followed by a 30min reperfusion has a deleterious effect on rat liver mitochondrial function. Mildronate pre-treatment of rats at doses of 100 or 200mg/kg/day for one or two weeks did not prevent ischemia/reperfusion-induced mitochondrial dysfunction and liver injury.

Effects of 5-acetyl(carbamoyl)-6-methylsulfanyl-1,4-dihydropyridine-5-carbonitriles on rat liver mitochondrial function

It is increasingly recognised that mitochondria are potential targets to pharmacological and toxicological actions of membrane-active agents, including some 1,4-dihydropyridines derivatives (DHPs). The 5-acetyl(carbamoyl)-6-methylsulfanyl-1,4-dihydropyridine-5-carbonitriles (OSI-1146, OSI-3701, OSI-3761, and OSI-9642) is a new group of DHPs with minor differences on the molecular structure. It has also been shown that OSI-1146 displays cardiovascular, antioxidant, and antiradical activities, whereas OSI-3701 and OSI-3761 display hepatoprotective activity. Due to their protective properties, this group of DHPs may be potentially useful for the treatment of several pathological processes, including those associated with oxidative stress. However, the cellular targets for their pharmacological actions have not been investigated. The presented study, using isolated rat liver mitochondria was designed to investigate if mitochondria are a cellular target for the pharmacological and/or toxicological actions of these new group of DHPs. We studied the direct influence of these DHPs on rat liver mitochondrial function [bioenergetics, membrane permeability transition (MPT), and oxidative stress]. It was shown that OSI-1146, OSI-3761, and OSI-9642, in the concentration range of up to 200 μM, interfered with mitochondrial bioenergetics by affecting complexes I and II of the mitochondrial respiratory chain, the ATPase activity, and mitochondrial inner membrane permeability to protons. However, the effects of OSI-1146 were higher than those of OSI-3761 and OSI-9642. The remaining compound, OSI-3701, had no effect on the bioenergetic parameters tested. All the compounds increased the susceptibility of mitochondria to MPT, but, OSI-3701, not affecting the bioenergetic parameters, was the most potent. Moreover, all the compounds protected mitochondria against lipid peroxidation induced by the oxidant pair ADP/Fe 2+, but OSI-1146 was also the most potent. In conclusion, our results indicate that mitochondria are the potential intracellular targets for both protective and toxicological actions of the DHP compounds studied, suggesting that the potential use of these compounds as therapeutic agents should carefully consider their toxic effects to mitochondria.

Cisplatin impairs rat liver mitochondrial functions by inducing changes on membrane ion permeability: Prevention by thiol group protecting agents

Cisplatin (CisPt) is the most important platinum anticancer drug widely used in the treatment of head, neck, ovarian and testicular cancers. However, the mechanisms by which CisPt induces cytotoxicity, namely hepatotoxicity, are not completely understood. The goal of this study was to investigate the influence of CisPt on rat liver mitochondrial functions (Ca 2+-induced mitochondrial permeability transition (MPT), mitochondrial bioenergetics, and mitochondrial oxidative stress) to better understand the mechanism underlying its hepatotoxicity. The effect of thiol group protecting agents and some antioxidants against CisPt-induced mitochondrial damage was also investigated. Treatment of rat liver mitochondria with CisPt (20 nmol/mg protein) induced Ca 2+-dependent mitochondrial swelling, depolarization of membrane potential (Δ Ψ), Ca 2+ release, and NAD(P)H fluorescence intensity decay. These effects were prevented by cyclosporine A (CyA), a potent and specific inhibitor of the MPT. In the concentration range of up to 40 nmol/mg protein, CisPt slightly inhibited state 3 and stimulated state 2 and state 4 respiration rates using succinate as respiratory substrate. The respiratory indexes, respiratory control ratio (RCR) and ADP/O ratios, the Δ Ψ, and the ADP phosphorylation rate were also depressed. CisPt induced mitochondrial inner membrane permeabilization to protons (proton leak) but did not induce significant changes on mitochondrial H 2O 2 generation. All the effects induced by CisPt on rat liver mitochondria were prevented by thiol group protecting agents namely, glutathione (GSH), dithiothreitol (DTT), N-acetyl- l-cysteine (NAC) and cysteine (CYS), whereas superoxide-dismutase (SOD), catalase (CAT) and ascorbate (ASC) were without effect. In conclusion, the anticancer drug CisPt: (1) increases the sensitivity of mitochondria to Ca 2+-induced MPT; (2) interferes with mitochondrial bioenergetics by increasing mitochondrial inner membrane permeabilization to H +; (3) does not significantly affect H 2O 2 generation by mitochondria; (4) its mitochondrial damaging effects are protected by thiol group protecting agents. Based on these conclusions, it is possible to hypothesise that small changes on the redox-status of thiol groups, affecting membrane permeability to cations (Ca 2+ and H +) underlie CisPt-induced liver mitochondrial damage, putatively responsible for its hepatotoxicity. Therefore, we propose that CisPt-induced mitochondrial damage and consequent hepatotoxicity could be prevented by using thiol group protecting agents as therapeutic adjuvants.

Assessment of the effect of cyclic chalcone analogues on mitochondrial membrane and DNA

AbstractThe cytotoxic and protective effects of selected synthetic chalcone analogues have been shown in previous studies. We studied their cytotoxic effect on the modification of mitochondrial membrane potential and on DNA. The first spectral information about the methoxy group as well as the dimethylamino substituent in E-2-arylmethylene-1-benzosuberones molecule was obtained by absorption and emission spectra. The cytotoxic effect of both cyclic chalcone analogues on DNA were detected by alkaline single-cell gel electrophoresis. Better fluorescent chalcone analogue E-2-(4′-dimethylamino-benzylidene)-1-benzosuberone was studied further in fresh isolated mitochondria. The decrease of rat liver mitochondria membrane potential (Δψ) was observed by fluorescence emission spectra. For the collapsing of mitochondrial potentials and as the negative control of mitochondrial function the CCCP uncoupler was used. The absorption maximum of the methoxy group was found at a shorter wavelength (λ = 335 nm) than that of the dimethylamino group (λ = 406 nm). The excitation spectra were very similar to the absorption spectra for both molecules but the emission spectra showed a better fluorescence for dimethylamino derivative. After the addition of E-2-(4′-dimethylamino-benzylidene)-1-benzosuberone to the intact mitochondria the decrease of mitochondrial membrane potential Δψ was observed by emisssion fluorescence spectra. Both cyclic chalcone analogues induced DNA damage, which was detected by alkaline comet assay. Mainly the apoptotic cells were detected, but necrotic cells were also present. Similarities in the percentages of DNA migration from the head were observed in both treatment groups. Both benzosuberones, with dimethylamino- and methoxy- substituent, were very active biologically, as shown by DNA results of the comet assay. Due to its better fluorescence properties, only the fluorophore with dimethylamino substituent was selected for further study of the function of rat liver mitochondria. Decline of mitochondrial function as well as mitochondrial DNA damage were evident between experimental and control groups.

Preliminary biological evaluation of poli(amidoamine) (PAMAM) dendrimer G3.5 on selected parameters of rat liver mitochondria

Polyamidoamino (PAMAM) dendrimer of generation 3.5 (G3.5) specific interactions with rat liver mitochondria were studied. Selected parameters of mitochondria (transmembrane potential and uptake of Ca 2+ ions) were investigated after the exposure to G3.5 used at the concentration of 10, 30 and 50 μM and Ca 2+ ions used at 1 mM. The times of preincubation of isolated liver mitochondria with dendrimer were 5, 15 and 30 min at room temperature. The mitochondrial membrane potential was monitored spectrofluorimetrically by fluorescence quenching of Rhodamine 123 (Rh 123). The changes in calcium homeostasis upon dendrimer exposure were detected using flow cytometric analysis with Fluo-3. On the one hand the obtained results revealed an impact of the tested chemical on rat liver mitochondrial function. We found that dendrimer G3.5 in a concentration above 10 μM contributes to the reduction in the transmembrane potential and hinders in the influx of Ca 2+ ions to mitochondria added externally, or accelerates their efflux from mitochondria. The most effective preincubation time was observed to be 15 min for all tested concentrations. On the other hand the combined treatment of dendrimer G3.5 with Ca 2+ demonstrated the protective effect of G3.5 against mitochondrial depolarization caused by calcium ions. These preliminary studies suggest that tested dendrimer can significantly affect the mitochondria and modulate their functionality.

Amaranth seed oil: Effect of oral administration on energetic functions of rat liver mitochondria activated with adrenaline

Amaranth seed oil: Effect of oral administration on energetic functions of rat liver mitochondria activated with adrenaline

Effect of sydnone SYD-1, a mesoionic compound, on energy-linked functions of rat liver mitochondria

An important antitumour effect of SYD-1 (3-[4-chloro-3-nitrophenyl]-1,2,3-oxadiazolium-5-olate) has been shown. We now report the effects of this mesoionic compound on mitochondrial metabolism. SYD-1 (1.5 μmol mg −1 protein) dose-dependently inhibited the respiratory rate by 65% and 40% in state 3 using sodium glutamate and succinate, respectively, as substrates. Phosphorylation efficiency was depressed by SYD-1, as evidenced by stimulation of the state 4 respiratory rate, which was more accentuated with glutamate (∼180%) than with succinate (∼40%), with 1.5 μmol mg −1 protein of SYD-1. As a consequence of the effects on states 3 and 4, the RCC and ADP/O ratios were lowered by SYD-1 using both substrates, although this effect was stronger with glutamate. The formation of membrane electrical potential was inhibited by ∼50% (1.5 μmol SYD-1 mg −1 protein). SYD-1 interfered with the permeability of the inner mitochondrial membrane, as demonstrated by assays of mitochondrial swelling in the presence of sodium acetate and valinomycin +K +. SYD-1 (1.5 μmol mg −1 protein) inhibited glutamate completely and succinate energized-mitochondrial swelling by 80% in preparations containing sodium acetate. The swelling of de-energized mitochondria induced by K + and valinomycin was inhibited by 20% at all concentrations of SYD-1. An analysis of the segments of the respiratory chain suggested that the SYD-1 inhibition site goes beyond the complex I and includes complexes III and IV. Glutamate dehydrogenase was inhibited by 20% with SYD-1 (1.5 μmol mg −1 protein). The hydrolytic activity of complex F 1F o ATPase in intact mitochondria was greatly increased (∼450%) in the presence of SYD-1. Our results show that SYD-1 depresses the efficiency of electron transport and oxidative phosphorylation, suggesting that these effects may be involved in its antitumoural effect.

Treatment with Antimalarials Adversely Affects the Oxidative Energy Metabolism in Rat Liver Mitochondria

Effects of in vivo treatment with the three antimalarials chloroquine, primaquine and quinine on rat liver mitochondrial energy transduction functions were examined. Treatment with all the three antimalarials decreased the state 3 and state 4 respiration rates drastically. The extent of inhibition was higher with pyruvate + malate as the substrate than with glutamate. The antimalarials also acted as uncouplers. The uncoupling effect was seen on site II and site III of phosphorylation; site I was unaffected. As a consequence the ADP phosphorylation rates also decreased significantly. Following antimalarials treatment the basal and Mg2 +stimulated ATPase activities increased while the DNP‐stimulated ATPase activity was reduced by half. Treatment with chloroquine resulted in decreased contents of cytochromes aa3 and b; primaquine and quinine treatments increased the contents of the two cytochromes in 14 day treatment groups.

Effects of deltamethrin on functions of rat liver mitochondria and on native and synthetic model membranes

Deltamethrin (DTM) is a synthetic pyrethroid insecticide used wideworld in agriculture, home pest control, protection of foodstuff, and disease vector control. It has widespread applications in Brazilian agriculture. The effects of DTM on mitochondrial respiratory parameters and on the organization of artificial and native membranes are described. DTM (200 nmol mg −1 protein) on isolated liver mitochondria decreased oxygen consumption of both, state III and state IV, as well as the inner mitochondrial membrane potential (Δ ψ). Analysis of segments of the respiratory chain suggested that the DTM inhibition site is located between complex II and complex III. Mitochondrial swelling, energized or driven by the K + diffusion potential using valinomycin, were partially inhibited by DTM (200 nmol mg −1 protein). Fluorescence polarization of DPH and DPH-PA, probing the core and outer regions, respectively, of dimyristoylphosphatidylcholine (DMPC) and native mitochondrial membranes, indicated that DTM shifts the midpoint phase transition to lower values, besides broadening the phase transition. DTM decreased the lipid order of DMPC bilayers, at temperatures lower than the transition temperature and also caused a disordering effect on native membranes. However at temperatures above the transition temperature, the pesticide increased the rigidity of the membrane. These results suggest that DTM causes perturbations in lipid-lipid and lipid-protein interactions, interferes in transport mechanisms operating at the membrane level, and causes alterations of membrane permeability and mitochondrial enzyme activities. These effects could be associated with the toxicity of deltamethrin.