Disturbance of mitochondrial functions associated with permeability transition pore opening induced by cis-5-tetradecenoic and myristic acids in liver of adolescent rats.

Patients affected by very long-chain acyl-CoA dehydrogenase (VLCAD) deficiency commonly present liver dysfunction whose pathogenesis is poorly known. We demonstrate here that major metabolites accumulating in this disorder, namely cis-5-tetradecenoic acid (Cis-5) and myristic acid (Myr), markedly impair mitochondrial respiration, decreasing ATP production in liver mitochondrial preparations from adolescent rats. Other parameters of mitochondrial homeostasis such as membrane potential (ΔΨm) and Ca2+retention capacity were strongly compromised by these fatty acids, involving induction of mitochondrial permeability transition. The present data indicate that disruption of mitochondrial bioenergetics and Ca2+homeostasis may contribute to the liver dysfunction of VLCAD deficient patients.

Resilient hepatic mitochondrial function and lack of iNOS dependence in diet-induced insulin resistance.

Obesity-derived inflammation and metabolic dysfunction has been related to the activity of the inducible nitric oxide synthase (iNOS). To understand the interrelation between metabolism, obesity and NO., we evaluated the effects of obesity-induced NO. signaling on liver mitochondrial function. We used mouse strains containing mitochondrial nicotinamide transhydrogenase activity, while prior studies involved a spontaneous mutant of this enzyme, and are, therefore, more prone to oxidative imbalance. Wild-type and iNOS knockout mice were fed a high fat diet for 2, 4 or 8 weeks. iNOS knockout did not protect against diet-induced metabolic changes. However, the diet decreased fatty-acid oxidation capacity in liver mitochondria at 4 weeks in both wild-type and knockout groups; this was recovered at 8 weeks. Interestingly, other mitochondrial functional parameters were unchanged, despite significant modifications in insulin resistance in wild type and iNOS knockout animals. Overall, we found two surprising features of obesity-induced metabolic dysfunction: (i) iNOS does not have an essential role in obesity-induced insulin resistance under all experimental conditions and (ii) liver mitochondria are resilient to functional changes in obesity-induced metabolic dysfunction.

Respiratory analysis of coupled mitochondria in cryopreserved liver biopsies.

The aim of this work was to develop a cryopreservation method of small liver biopsies for in situ mitochondrial function assessment. Herein we describe a detailed protocol for tissue collection, cryopreservation, high-resolution respirometry using complex I and II substrates, calculation and interpretation of respiratory parameters. Liver biopsies from cow and rat were sequentially frozen in a medium containing dimethylsulfoxide as cryoprotectant and stored for up to 3 months at -80 °C. Oxygen consumption rate studies of fresh and cryopreserved samples revealed that most respiratory parameters remained unchanged. Additionally, outer mitochondrial membrane integrity was assessed adding cytochrome c, proving that our cryopreservation method does not harm mitochondrial structure. In sum, we present a reliable way to cryopreserve small liver biopsies without affecting mitochondrial function. Our protocol will enable the transport and storage of samples, extending and facilitating mitochondrial function analysis of liver biopsies.

Mechanistic insights into functional characteristics of native crotamine.

The chemical composition of snake venoms is a complex mixture of proteins and peptides that can be pharmacologically active. Crotamine, a cell-penetrating peptide, has been described to have antimicrobial properties and it exerts its effects by interacting selectively with different structures, inducing changes in the ion flow pattern and cellular responses. However, its real therapeutic potential is not yet fully known. Bearing in mind that crotamine is a promising molecule in therapeutics, this study investigated the action of purified molecule in three aspects: I) antibacterial action on different species of clinical interest, II) the effect of two different concentrations of the molecule on platelet aggregation, and III) its effects on isolated mitochondria. Crotamine was purified to homogeneity in a single step procedure using Heparin Sepharose. The molecular mass of the purified enzyme was 4881.4 Da, as determined by mass spectrometry. To assess antibacterial action, changes in the parameters of bacterial oxidative stress were determined. The peptide showed antibacterial activity on Escherichia coli (MIC: 2.0 µg/µL), Staphylococcus aureus (MIC: 8-16 µg/µL) and methicillin-resistant Staphylococcus aureus (MIC: 4.0-8.0 µg/µL), inducing bacterial death by lipid peroxidation and oxidation of target proteins, determined by thiobarbituric acid reactive substances and sulfhydryl groups, respectively. Crotamine induced increased platelet aggregation (IPA) at the two concentrations analyzed (0.1 and 1.4 µg/µL) compared to ADP-induced aggregation of PRP. Mitochondrial respiratory parameters and organelle structure assays were used to elucidate the action of the compound in this organelle. The exposure of mitochondria to crotamine caused a decrease in oxidative phosphorylation and changes in mitochondrial permeability, without causing damage in the mitochondrial redox state. Together, these results support the hypothesis that, besides the antimicrobial potential, crotamine acts on different molecular targets, inducing platelet aggregation and mitochondrial dysfunction.

Mechanisms of the beneficial effect of sevoflurane in liver ischemia/reperfusion injury.

PURPOSE: To evaluate the underlying mechanisms by which sevoflurane protects the liver against ischemia/reperfusion injury evaluate the mechanism by which sevoflurane exerts this protective effect. METHODS: Twenty-six rats were subjected to partial ischemia/reperfusion injury for 1h: one group received no treatment, one group received sevoflurane, and sham group of animals received laparotomy only. Four hours after reperfusion, levels of alanine and aspartate aminotransferases, tumor necrosis factor-a, and interleukins 6 and 10 were measured. Analyses of mitochondrial oxidation and phosphorylation, malondialdehyde content, histology, and pulmonary vascular permeability were performed. RESULTS: Serum levels of alanine and aspartate aminotransferases were significantly lower in the sevoflurane group compared to untreated controls (p<0.05). The sevoflurane group also showed preservation of liver mitochondrial function compared to untreated controls (p<0.05). Sevoflurane administration did not alter increases in serum levels of tumor necrosis factor-a, and interleukins 6 and 10. Sevoflurane treatment significantly reduced the coagulative necrosis induced by ischemia/reperfusion (p<0.05). Pulmonary vascular permeability was preserved in the sevoflurane group compared to untreated controls. CONCLUSION: Sevoflurane administration protects the liver against ischemia/reperfusion injury, via preservation of mitochondrial function, and also preserves lung vascular permeability.

Mechanisms of the beneficial effect of sevoflurane in liver ischemia/reperfusion injury

PURPOSE: To evaluate the underlying mechanisms by which sevoflurane protects the liver against ischemia/reperfusion injury evaluate the mechanism by which sevoflurane exerts this protective effect. METHODS: Twenty-six rats were subjected to partial ischemia/reperfusion injury for 1h: one group received no treatment, one group received sevoflurane, and sham group of animals received laparotomy only. Four hours after reperfusion, levels of alanine and aspartate aminotransferases, tumor necrosis factor-a, and interleukins 6 and 10 were measured. Analyses of mitochondrial oxidation and phosphorylation, malondialdehyde content, histology, and pulmonary vascular permeability were performed. RESULTS: Serum levels of alanine and aspartate aminotransferases were significantly lower in the sevoflurane group compared to untreated controls (p<0.05). The sevoflurane group also showed preservation of liver mitochondrial function compared to untreated controls (p<0.05). Sevoflurane administration did not alter increases in serum levels of tumor necrosis factor-a, and interleukins 6 and 10. Sevoflurane treatment significantly reduced the coagulative necrosis induced by ischemia/reperfusion (p<0.05). Pulmonary vascular permeability was preserved in the sevoflurane group compared to untreated controls. CONCLUSION: Sevoflurane administration protects the liver against ischemia/reperfusion injury, via preservation of mitochondrial function, and also preserves lung vascular permeability.

Effects of hyperbaric oxygen therapy as hepatic preconditioning in rats submitted to hepatic ischemia/reperfusion injury.

PURPOSE: To analyze the role of hyperbaric oxygen therapy as hepatic preconditioning in rats submitted to hepatic ischemia and reperfusion. METHODS: Wistar rats were randomly divided into three groups: SHAM, rats submitted to surgical stress without hepatic ischemia and reperfusion, I/R, rats submitted to total hepatic pedicle ischemia for 30 min, followed by 5 min of reperfusion; HBOI/R, rats submitted to 60 minutes of hyperbaric oxygen therapy at 2 atm and immediately submitted to the experimental protocol of ischemia and reperfusion. Liver function was assessed by measuring serum alanine aminotransferase and aspartate aminotransferase, as well as mitochondrial function by determining states 3 and 4 of mitochondrial respiration, respiratory control rate and mitochondrial permeability transition (mitochondrial swelling). The results were analyzed by the Mann-Whitney test and all P-values <0.05 were considered significant. RESULTS: There were significant differences in serum aspartate aminotransferase values in groups SHAM vs. HBOI/R, I/R vs HBOI/R, alanine aminotransferase in groups SHAM and I/R; State 3 in SHAM groups vs. I/R, SHAM vs. HBOI/R, State 4 in I/R vs HBOI/R groups, respiratory control rate in SHAM vs I/R groups; mitochondrial swelling in SHAM vs. I/R groups, and SHAM vs HBOI/R. CONCLUSION: Hyperbaric preconditioning improved hepatic mitochondrial function and decreased serum markers of liver injury in the ischemia and reperfusion process.

Hyperoxic preconditioning in partial liver ischemia.

PURPOSE: To evaluate the effect of the hyperbaric oxygen (HBO) treatment as a pre-conditioning for I/R effects in the liver ischemia. METHODS: Fifty-seven male Wistar rats (260-300g) were submitted to the following procedures: SHAM; I/R, rats submitted to I/R, consisting of partial ischemia of 70% of the liver for 90 minutes followed by 15 minutes of reperfusion; HBO I/R 1 ATA, 30 minutes of HBO treatment at the pressure of 1 absolute atmosphere (ATA) during the ischemia time. HBO I/R 2 ATA, 30 minutes of HBO (2 ATA) during the ischemia time. Pre HBO I/R 30', rats submitted to 30 minutes of HBO (2 ATA) immediately before the I/R time. Pre HBO I/R 90', rats submitted to 90 minutes of HBO (2 ATA) immediately before the I/R time. RESULTS: There was a significant worsening of all the parameters of mitochondrial energy production (state 3, 4, RCR and Swelling) in the I/R group, when compared to the Sham group (I/R

Effect of hyperbaric hepatic hyperoxia on the liver of rats submitted to intermittent ischemia/reperfusion injury.

PURPOSE: To determine the effect of hyperbaric hyperoxia as hepatic preconditioning on hepatocellular integrity in rats submitted to intermittent hepatic ischemia/reperfusion injury. METHODS: Twenty male Wistar rats were divided into 4 groups (SHAM, I/R, HBO-I/R and CONTROL). The surgical technique consisted of total clamping of the hepatic pedicle for 15 min, followed by reperfusion for 5 min, performed twice. The application of hyperbaric oxygen (HBO) was carried out in a collective chamber (simultaneous exposure of 4 rats) pressurized directly with oxygen at 2 ATA for 60 min. Tissue malondialdehyde (MDA) levels were determined and blood samples were collected for the determination of serum AST and ALT levels. Data were analyzed statistically by the Mann-Whitney test, with the level of significance set at p < 0.05. RESULTS: A statistically significant difference in MDA (p< 0.05) was observed between control and HBO-I/R, but not between control and I/R. Regarding AST, there was a difference between control and I/R and HBO-I/R. Analysis of ALT revealed a significant difference between control and I/R (p<0.05) and between I/R and HBO-I/R, with no difference between control and HBO-IR. CONCLUSION: Hyperoxic preconditioning proved to be favorable regarding alanine transaminase, but not aspartate aminotranserase or malondialdehyde levels.

Effects of hyperbaric oxygen therapy on the liver after injury caused by the hepatic ischemia-reperfusion process.

PURPOSE: To evaluate the effects of hyperbaric oxygen on rats submitted to hepatic ischemia and reperfusion. METHODS: Twenty-three Wistar rats were divided at random into 3 groups: SHAM, rats submitted to surgical and anesthetic stress without induction of hepatic ischemia/reperfurion; I/R, rats submitted to total ischemia of the hepatic pedicle for 25 min followed by 5 min of reperfusion; HBOI/R, rats submitted to 60 min of hyperbaric oxygen therapy at a pressure of 2 absolute atmospheres immediately after the experimental protocol of ischemia/reperfusion. Hepatic function was evaluated by quantitation of serum alanine aminotranferase (ALT) and aspartate aminotransferase (AST), and by mitochondrial function through the determination of states 3 and 4 of mitochondrial respiration, respiratory control ratio (RCR) and mitochondrial swelling. Data were analyzed by the Mann-Whitney test, with the level of significance set at p <0.05. RESULTS: There was a significant difference in state 3 values for the SHAM group vs I/R and I/R vs IRHBO, in state 4 values for the SHAM group vs I/R; and in mitochondrial swelling for the SHAM groups vs I/RHBO, SHAM vs I/R, and IR vs I/RHBO. CONCLUSION: The use of hyperbaric oxygen after I/R improved in a relative manner both the production of energy and the effects on the mitochondrial wall.

Effects of hyperbaric oxygen therapy on the liver after injury caused by the hepatic ischemia-reperfusion process

PURPOSE: To evaluate the effects of hyperbaric oxygen on rats submitted to hepatic ischemia and reperfusion. METHODS: Twenty-three Wistar rats were divided at random into 3 groups: SHAM, rats submitted to surgical and anesthetic stress without induction of hepatic ischemia/reperfurion; I/R, rats submitted to total ischemia of the hepatic pedicle for 25 min followed by 5 min of reperfusion; HBOI/R, rats submitted to 60 min of hyperbaric oxygen therapy at a pressure of 2 absolute atmospheres immediately after the experimental protocol of ischemia/reperfusion. Hepatic function was evaluated by quantitation of serum alanine aminotranferase (ALT) and aspartate aminotransferase (AST), and by mitochondrial function through the determination of states 3 and 4 of mitochondrial respiration, respiratory control ratio (RCR) and mitochondrial swelling. Data were analyzed by the Mann-Whitney test, with the level of significance set at p <0.05. RESULTS: There was a significant difference in state 3 values for the SHAM group vs I/R and I/R vs IRHBO, in state 4 values for the SHAM group vs I/R; and in mitochondrial swelling for the SHAM groups vs I/RHBO, SHAM vs I/R, and IR vs I/RHBO. CONCLUSION: The use of hyperbaric oxygen after I/R improved in a relative manner both the production of energy and the effects on the mitochondrial wall. .

Effect of hyperbaric hepatic hyperoxia on the liver of rats submitted to intermittent ischemia/reperfusion injury

PURPOSE: To determine the effect of hyperbaric hyperoxia as hepatic preconditioning on hepatocellular integrity in rats submitted to intermittent hepatic ischemia/reperfusion injury. METHODS: Twenty male Wistar rats were divided into 4 groups (SHAM, I/R, HBO-I/R and CONTROL). The surgical technique consisted of total clamping of the hepatic pedicle for 15 min, followed by reperfusion for 5 min, performed twice. The application of hyperbaric oxygen (HBO) was carried out in a collective chamber (simultaneous exposure of 4 rats) pressurized directly with oxygen at 2 ATA for 60 min. Tissue malondialdehyde (MDA) levels were determined and blood samples were collected for the determination of serum AST and ALT levels. Data were analyzed statistically by the Mann-Whitney test, with the level of significance set at p < 0.05. RESULTS: A statistically significant difference in MDA (p< 0.05) was observed between control and HBO-I/R, but not between control and I/R. Regarding AST, there was a difference between control and I/R and HBO-I/R. Analysis of ALT revealed a significant difference between control and I/R (p<0.05) and between I/R and HBO-I/R, with no difference between control and HBO-IR. CONCLUSION: Hyperoxic preconditioning proved to be favorable regarding alanine transaminase, but not aspartate aminotranserase or malondialdehyde levels. .

Hyperoxic preconditioning in partial liver ischemia

PURPOSE: To evaluate the effect of the hyperbaric oxygen (HBO) treatment as a pre-conditioning for I/R effects in the liver ischemia. METHODS: Fifty-seven male Wistar rats (260-300g) were submitted to the following procedures: SHAM; I/R, rats submitted to I/R, consisting of partial ischemia of 70% of the liver for 90 minutes followed by 15 minutes of reperfusion; HBO I/R 1 ATA, 30 minutes of HBO treatment at the pressure of 1 absolute atmosphere (ATA) during the ischemia time. HBO I/R 2 ATA, 30 minutes of HBO (2 ATA) during the ischemia time. Pre HBO I/R 30', rats submitted to 30 minutes of HBO (2 ATA) immediately before the I/R time. Pre HBO I/R 90', rats submitted to 90 minutes of HBO (2 ATA) immediately before the I/R time. RESULTS: There was a significant worsening of all the parameters of mitochondrial energy production (state 3, 4, RCR and Swelling) in the I/R group, when compared to the Sham group (I/R <Sham, p<0.05). There was also a significant worsening in state 4, RCR and mitochondrial edema in the Pre HBO I/R 90' group compared to the I/R group. Hepatic enzyme concentrations were significantly higher in the I/R group. CONCLUSION: The use of hyperbaric oxygen before and during I/R did not improve the production of hepatocellular energy reduced by I/R, nor did it prevent the installation of mitochondrial edema induced by Iischemia/reperfusion. .

Effects of hyperbaric oxygen therapy as hepatic preconditioning in rats submitted to hepatic ischemia/reperfusion injury

PURPOSE: To analyze the role of hyperbaric oxygen therapy as hepatic preconditioning in rats submitted to hepatic ischemia and reperfusion. METHODS: Wistar rats were randomly divided into three groups: SHAM, rats submitted to surgical stress without hepatic ischemia and reperfusion, I/R, rats submitted to total hepatic pedicle ischemia for 30 min, followed by 5 min of reperfusion; HBOI/R, rats submitted to 60 minutes of hyperbaric oxygen therapy at 2 atm and immediately submitted to the experimental protocol of ischemia and reperfusion. Liver function was assessed by measuring serum alanine aminotransferase and aspartate aminotransferase, as well as mitochondrial function by determining states 3 and 4 of mitochondrial respiration, respiratory control rate and mitochondrial permeability transition (mitochondrial swelling). The results were analyzed by the Mann-Whitney test and all P-values <0.05 were considered significant. RESULTS: There were significant differences in serum aspartate aminotransferase values in groups SHAM vs. HBOI/R, I/R vs HBOI/R, alanine aminotranferase in groups SHAM and I/R; State 3 in SHAM groups vs. I/R, SHAM vs. HBOI/R, State 4 in I/R vs HBOI/R groups, respiratory control rate in SHAM vs I/R groups; mitochondrial swelling in SHAM vs. I/R groups, and SHAM vs HBOI/R. CONCLUSION: Hyperbaric preconditioning improved hepatic mitochondrial function and decreased serum markers of liver injury in the ischemia and reperfusion process. .

Swimming training induces liver mitochondrial adaptations to oxidative stress in rats submitted to repeated exhaustive swimming bouts.

BACKGROUND AND AIMS: Although acute exhaustive exercise is known to increase liver reactive oxygen species (ROS) production and aerobic training has shown to improve the antioxidant status in the liver, little is known about mitochondria adaptations to aerobic training. The main objective of this study was to investigate the effects of the aerobic training on oxidative stress markers and antioxidant defense in liver mitochondria both after training and in response to three repeated exhaustive swimming bouts. METHODS: Wistar rats were divided into training (n = 14) and control (n = 14) groups. Training group performed a 6-week swimming training protocol. Subsets of training (n = 7) and control (n = 7) rats performed 3 repeated exhaustive swimming bouts with 72 h rest in between. Oxidative stress biomarkers, antioxidant activity, and mitochondria functionality were assessed. RESULTS: Trained group showed increased reduced glutathione (GSH) content and reduced/oxidized (GSH/GSSG) ratio, higher superoxide dismutase (MnSOD) activity, and decreased lipid peroxidation in liver mitochondria. Aerobic training protected against exhaustive swimming ROS production herein characterized by decreased oxidative stress markers, higher antioxidant defenses, and increases in methyl-tetrazolium reduction and membrane potential. Trained group also presented higher time to exhaustion compared to control group. CONCLUSIONS: Swimming training induced positive adaptations in liver mitochondria of rats. Increased antioxidant defense after training coped well with exercise-produced ROS and liver mitochondria were less affected by exhaustive exercise. Therefore, liver mitochondria also adapt to exercise-induced ROS and may play an important role in exercise performance.

Effect of hyperbaric oxygen therapy on liver function during intermittent ischemia.

PURPOSE: To analyze the effects of hyperbaric oxygen therapy on liver function in rats previously subjected to ischemia and reperfusion. METHODS: A randomly distribution of 23 Wistar rats was conducted into three groups: SHAM, animals subjected to surgical stress without restricting blood flow by clamping the hepatic pedicle, IR, rats underwent hepatic vascular occlusion intermittently for two complete cycles of 15 minutes of ischemia followed by 5 min of reperfusion, IR / HBO, rats underwent hepatic pedicle clamping and thereafter exposed to hyperbaric oxygen pressure of 2 absolute atmospheres for 60 minutes. We evaluated liver function through mitochondrial function, determined by the stages 3 and 4 of respiration, respiratory control ratio (RCR) and mitochondrial permeability transition (Swelling). Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were also quantified . We analyzed the results using the Mann-Whitney test and were considered significant all results with p <0.05. RESULTS: There were significant differences between the results of stage 3 in SHAM vs IR group ; of the stage 4 in the groups IR vs SHAM and SHAM vs IR /HBO; of the Respiratory Control Ratio (RCR) in the group IR vs IR / HBO ; of alanine aminotransferase in the groups IR vs SHAM , SHAM vs IR/HBO and IR vs IR / HBO; aspartate aminotransferase in the groups SHAM vs IR and SHAM vs IR / HBO. CONCLUSION: The whole analysis of the mitochondrial function indicators permits us to conclude that the hyperbaric oxygen therapy acted as a protective agent of the mitochondrial function, minimizing the ischemia-reperfusion injury of the hepatic parenchyma.

Effects of hyperbaric oxygen (HBO), as pre-conditioning in liver of rats submitted to periodic liver ischemia/reperfusion.

PURPOSE: to assess the effect of hyperbaric oxygen (HBO) as pre-conditioning on periodic liver ischemia/reperfusion injury. METHODS: Thirty-six male Wistar rats were divided into 4 groups (SHAM, I/R , HBO-I/R and CONTROL). The surgical technique consisted of total clamping of the hepatic pedicle for 15 min followed by twice repeated reperfusion for 5 min (unclamping). HBO was applied in a collective chamber (simultaneous exposure of 4 rats) directly pressurized with oxygen at 2 ATA for 60 min. Hepatic mitochondrial function was determined using samples of the median lobe obtained after exactly 5 min of reperfusion for the analysis of mitochondrial respiration based on the determination of states 3 and 4, the respiratory control ratio and the transition of mitochondrial permeability (mitochondrial swelling).Data were analyzed by the Mann-Whitney test and the level of significance was set at p < 0.05. RESULTS: There was a statistically significant difference (p < 0.05) in state 3 between the CONTROL and I/R and HBO-I/R groups, in state 4 between the CONTROL and I/R and HBO-I/R groups; in respiratory control ratio (RCR) between the CONTROL and I/R and HBO-I/R groups and between the CONTROL and Sham groups, and in mitochondrial swelling between the CONTROL and I/R and HBO-/R groups and between the Sham and I/R and HBO-I/R groups. CONCLUSION: In this process of periodic ischemia and reperfusion, hyperbaric pre-conditioning did not improve significantly hepatic mitochondrial function.

Nitric oxide inhibits succinate dehydrogenase-driven oxygen consumption in potato tuber mitochondria in an oxygen tension-independent manner.

NO (nitric oxide) is described as an inhibitor of plant and mammalian respiratory chains owing to its high affinity for COX (cytochrome c oxidase), which hinders the reduction of oxygen to water. In the present study we show that in plant mitochondria NO may interfere with other respiratory complexes as well. We analysed oxygen consumption supported by complex I and/or complex II and/or external NADH dehydrogenase in Percoll-isolated potato tuber (Solanum tuberosum) mitochondria. When mitochondrial respiration was stimulated by succinate, adding the NO donors SNAP (S-nitroso-N-acetyl-DL-penicillamine) or DETA-NONOate caused a 70% reduction in oxygen consumption rate in state 3 (stimulated with 1 mM of ADP). This inhibition was followed by a significant increase in the Km value of SDH (succinate dehydrogenase) for succinate (Km of 0.77±0.19 to 34.3±5.9 mM, in the presence of NO). When mitochondrial respiration was stimulated by external NADH dehydrogenase or complex I, NO had no effect on respiration. NO itself and DETA-NONOate had similar effects to SNAP. No significant inhibition of respiration was observed in the absence of ADP. More importantly, SNAP inhibited PTM (potato tuber mitochondria) respiration independently of oxygen tensions, indicating a different kinetic mechanism from that observed in mammalian mitochondria. We also observed, in an FAD reduction assay, that SNAP blocked the intrinsic SDH electron flow in much the same way as TTFA (thenoyltrifluoroacetone), a non-competitive SDH inhibitor. We suggest that NO inhibits SDH in its ubiquinone site or its Fe-S centres. These data indicate that SDH has an alternative site of NO action in plant mitochondria.

Effect of hyperbaric oxygen therapy on liver function during intermittent ischemia / Efeito da oxigenoterapia hiperbárica sobre a função hepática na isquemia intermitente

PURPOSE: To analyze the effects of hyperbaric oxygen therapy on liver function in rats previously subjected to ischemia and reperfusion. METHODS: A randomly distribution of 23 Wistar rats was conducted into three groups: SHAM, animals subjected to surgical stress without restricting blood flow by clamping the hepatic pedicle, IR, rats underwent hepatic vascular occlusion intermittently for two complete cycles of 15 minutes of ischemia followed by 5 min of reperfusion, IR / HBO, rats underwent hepatic pedicle clamping and thereafter exposed to hyperbaric oxygen pressure of 2 absolute atmospheres for 60 minutes. We evaluated liver function through mitochondrial function, determined by the stages 3 and 4 of respiration, respiratory control ratio (RCR) and mitochondrial permeability transition (Swelling). Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were also quantified . We analyzed the results using the Mann-Whitney test and were considered significant all results with p <0.05. RESULTS: There were significant differences between the results of stage 3 in SHAM vs IR group ; of the stage 4 in the groups IR vs SHAM and SHAM vs IR /HBO; of the Respiratory Control Ratio (RCR) in the group IR vs IR / HBO ; of alanine aminotransferase in the groups IR vs SHAM , SHAM vs IR/HBO and IR vs IR / HBO; aspartate aminotransferase in the groups SHAM vs IR and SHAM vs IR / HBO. CONCLUSION: The whole analysis of the mitochondiral function indicators permits us to conclude that the hyperbaric oxygen therapy acted as a protective agent of the mitochondrial function, minimizing the ischemia-reperfusion injury of the hepatic parenchyma.

OBJETIVO: Analisar os efeitos da oxigenoterapia hiperbárica sobre a função hepática de ratos previamente submetidos à isquemia e reperfusão. MÉTODOS: Realizou-se a distribuição aleatória de 23 ratos Wistar em três grupos: SHAM, animais submetidos ao estresse cirúrgico sem restrição de fluxo sanguíneo por clampeamento do pedículo hepático; IR ratos submetidos a oclusão vascular hepática de maneira intermitente em dois ciclos completos de 15 minutos de isquemia, seguidos por 5 min de reperfusão; IR/HBO, ratos submetidos ao clampeamento do pedículo hepático e ,ulteriormente , expostos a oxigenoterapia hiperbárica a pressão de 2 atmosferas absolutas durante 60 minutos. Avaliou-se a função hepática através da função mitocondrial, determinada pelos estados 3 e 4 da respiração mitocondrial, razão de controle respiratório(RCR) e transição de permeabilidade mitocondrial (Swelling) .Quantificou-se também alanina aminotransferase e aspartato aminotransferase séricas. Analisou-se os resultados pelo teste de Mann-Whitney e foram considerados significativos os valores de p < 0,05. RESULTADOS:Houve diferença significativa nos valores do estado 3 no grupo SHAM vs IR; estado 4 nos grupos SHAM vs IR, SHAM vsIR/HBO; Razão de Controle Respiratório(RCR) no grupo IR vs IR/HBO; alanina aminotransferase nos grupos SHAM vs IR, SHAM vs IR/HBO, IR vs IR/HBO; aspartato aminotransferase nos grupos SHAM vs IR , SHAM vs IR/HBO. CONCLUSÃO: A análise dos parâmetros mitocondriais como um todo permitiu concluir que a oxigenoterapia hiperbárica atuou como agente protetor da função mitocondrial, minimizando os efeitos deletérios da lesão de isquemia e reperfusão do parênquima hepático.

Effects of hyperbaric oxygen (HBO), as pre-conditioning in liver of rats submitted to periodic liver ischemia/reperfusion / Efeitos da oxigenoterapia hiperbárica como pré-condicionamento em fígados de ratos submetidos à lesão hepática de isquemia/reperfusão intermitente

PURPOSE: to assess the effect of hyperbaric oxygen (HBO) as pre-conditioning on periodic liver ischemia/reperfusion injury. METHODS: Thirty-six male Wistar rats were divided into 4 groups (SHAM, I/R , HBO-I/R and CONTROL). The surgical technique consisted of total clamping of the hepatic pedicle for 15 min followed by twice repeated reperfusion for 5 min (unclamping). HBO was applied in a collective chamber (simultaneous exposure of 4 rats) directly pressurized with oxygen at 2 ATA for 60 min. Hepatic mitochondrial function was determined using samples of the median lobe obtained after exactly 5 min of reperfusion for the analysis of mitochondrial respiration based on the determination of states 3 and 4, the respiratory control ratio and the transition of mitochondrial permeability (mitochondrial swelling).Data were analyzed by the Mann-Whitney test and the level of significance was set at p < 0.05. RESULTS: There was a statistically significant difference (p< 0.05) in state 3 between the CONTROL and I/R and HBO-I/R groups, in state 4 between the CONTROL and I/R and HBO-I/R groups; in respiratory control ratio (RCR) between the CONTROL and I/R and HBO-I/R groups and between the CONTROL and Sham groups, and in mitochondrial swelling between the CONTROL and I/R and HBO-/R groups and between the Sham and I/R and HBO-I/R groups. CONCLUSION: In this process of periodic ischemia and reperfusion, hyperbaric pre-conditioning did not improve significantly hepatic mitochondrial function.

OBJETIVO: Avaliar os efeitos da oxigenoterapia hiperbárica (HBO), como pré-condicionamento, em lesão hepática de isquemia/reperfusão intermitente. MÉTODOS: Foram avaliados 36 ratos Wistar machos, distribuídos em 4 grupos (SHAM, I/R , HBO - I/R e CONTROLE). A técnica operatória consistiu em pinçamento total do pedículo hepático durante 15min, seguido de reperfusão por 5 min (desclampeamento), por duas vezes. A aplicação de HBO foi realizada em câmara coletiva (exposição simultânea de 4 ratos) diretamente pressurizada com oxigênio a 2ATA, durante 60min. Determinou-se a função mitocondrial hepática através de amostras do lobo mediano colhidas com exatos 5min de reperfusão para análise da respiração mitocondrial, através da determinação dos estados 3 e 4, razão de controle respiratório e transição de permeabilidade mitocondrial (intumescimento osmótico - swelling mitocondrial).Os resultados foram analisados pelo teste de Mann-Whitney e foi considerado significativo todo valor de p < 0,05. RESULTADOS: Houve diferença estatistica significativa (p< 0,05) no Estado 3 nos grupos CONTROLE vs I/R e HBO - I/R, no Estado 4 nos grupos CONTROLE vs I/R e HBO - I/R; na Razão de controle respiratório(RCR) nos grupos CONTROLE vs I/R e HBO-IRe CONTROLE vs Sham e no Swelling mitocondrial nos grupos CONTROLE vs I/R e HBO - I/R, I/R vs HBO-IRe Sham vs I/R e HBO-IR. CONCLUSÃO: O pré-condicionamento hiperbárico não melhorou a função mitocondrial hepática significativamente neste processo de isquemia e reperfusão intermitente.