Hepatic mitochondria in diet-induced obese mice (stereology and molecular analysis) / Mitocondrias hepáticas en ratones obesos inducidos por la dieta (estereología y análisis molecular)

SUMMARY: The world population is going through an obesity epidemic that has severe consequences for the health system. This study focused on studying hepatic mitochondria in obese animals induced by a high-fat (HF) diet and used the model-based stereology in electron micrographs for the quantitative study. Besides, the gene expressions of molecular markers of mitochondrial biogenesis carnitine palmitoyltransferase 1a (Cpt 1α), mitochondrial transcription factor a (Tfam), uncoupling protein 3 (Ucp 3), and nuclear respiratory factor 1 (Nrf 1) were analyzed. The HF diet caused a weight gain of +1820 % comparing the control group (C) with the HF group (from 0.32±0.31 g to 5.5±0.39 g, P<0.001). The HF group showed fat droplets in the hepatocyte cytoplasm (steatosis) and less dense and large mitochondria in transmission electron microscopy. The mitochondria size (cross-section) did not show a significant difference between the groups C and HF. However, the mitochondria numerical density per area was 30 % less, the mitochondrial surface density (outer membrane) was 20 % less, and the mitochondrial volume density was 22 % less in the HF group than the C group. The gene expressions of molecular markers of mitochondrial biogenesis Cpt 1α, Tfam, Ucp 3, and Nrf 1 decreased in the HF group compared to the C group. The quantitative results match perfectly with the molecular ones of mitochondrial biogenesis markers. In the future, it will be crucial to verify if and how these data recover with the reduction of obesity, which would be of significant interest given the current obesity epidemic that affects the world population.

RESUMEN: La población mundial atraviesa una epidemia de obesidad que tiene graves consecuencias para el sistema de salud. Este estudio se centró en el análisis de las mitocondrias hepáticas en animales obesos inducidos por una dieta alta en grasas (HF) y utilizó la estereología basada en modelos en micrografías electrónicas para el estudio cuantitativo. Además, se analizaron las expresiones génicas de los marcadores moleculares de la biogénesis mitocondrial carnitina palmitoiltransferasa 1a (Cpt 1α), factor de transcripción mitocondrial a (Tfam), proteína desacoplante 3 (Ucp 3) y factor respiratorio nuclear 1 (Nrf 1). La dieta HF provocó un aumento de peso de +1820 % comparando el grupo de control (C) con el grupo HF (de 0,32 ± 0,31 g a 5,5 ± 0,39 g, P <0,001). El grupo HF mostró gotas de grasa en el citoplasma de los hepatocitos (esteatosis) y mitocondrias menos densas y grandes en la microscopía electrónica de transmisión. El tamaño de las mitocondrias (sección transversal) no mostró una diferencia significativa entre los grupos C y HF. Sin embargo, la densidad numérica de mitocondrias por área fue 30% menor, la densidad de superficie mitocondrial (membrana externa) fue 20 % menor y la densidad de volumen mitocondrial fue 22 % menor en el grupo HF que en el grupo C. Las expresiones génicas de los marcadores moleculares de la biogénesis mitocondrial Cpt 1α, Tfam, Ucp 3 y Nrf 1 disminuyeron en el grupo HF en comparación con el grupo C. Los resultados cuantitativos coinciden perfectamente con los moleculares de los marcadores de biogénesis mitocondrial. En el futuro, será crucial verificar si estos datos se recuperan y cómo se recuperan con la reducción de la obesidad, lo que sería de gran interés dada la actual epidemia de obesidad que afecta a la población mundial.

Evaluation of the therapeutic effect of methylene blue on the liver of rats submitted to ischemia and reperfusion

Abstract Purpose: To analyze the effect of methylene blue (MB) therapy during the liver ischemia-reperfusion injury (I/R) process. Methods: Thirty-five male Wistar rats were used, (70%) submitted to partial ischemia (IR) or not (NIR) (30%) were obtained from the same animal. These animals were divided into six groups: 1) Sham (SH), 2) Sham with MB (SH-MB); 3) I/R, submitted to 60 minutes of partial ischemia and 15 minutes of reperfusion; 4) NI/R, without I/R obtained from the same animal of group I/R; 5) I/R-MB submitted to I/R and MB and 6) NI/R-MB, without I/R. Mitochondrial function was evaluated. Osmotic swelling of mitochondria as well as the determination of malondialdehyde (MDA) was evaluated. Serum (ALT/AST) dosages were also performed. MB was used at the concentration of 15mg/kg, 15 minutes before hepatic reperfusion. Statistical analysis was done by the Mann Whitney test at 5%. Results: State 3 shows inhibition in all ischemic groups. State 4 was increased in all groups, except the I/R-MB and NI/R-MB groups. RCR showed a decrease in all I/R and NI/R groups. Mitochondrial osmotic swelling showed an increase in all I/R NI/R groups in the presence or absence of MB. About MDA, there was a decrease in SH values in the presence of MB and this decrease was maintained in the I/R group. AST levels were increased in all ischemic with or without MB. Conclusions: The methylene blue was not able to restore the mitochondrial parameters studied. Also, it was able to decrease lipid peroxidation, preventing the formation of reactive oxygen species.

Rivastigmine prevents injury induced by ischemia and reperfusion in rat liver

Abstract Purpose: To evaluate whether pre-treatment with rivastigmine is able to attenuate the I/R induced lesions in rat liver. Methods: SHAM animals or those submitted to I/R, non-treated or pre-treated with rivastigminine (2mg/kg) either 50 or 15 minutes before ischemia, were used. After I/R protocol, these animals were killed and their livers were harvested to measurement of the mitochondrial swelling as well as the malondialdehyde (MDA), nitrite and nitrate tissue concentration. Blood was also harvested for serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) determinations. Results: I/R promoted a significant increase of mitochondrial swelling in the studied animals. This increase of mitochondrial swelling was partially prevented by rivastigmine, but only if administered 50 minutes before ischemia. No significant modification of MDA, nitrite or nitrate tissue concentrations was observed in consequence of I/R, followed or not by rivastigmine treatments. In addition, I/R elevated both AST and ALT. These elevations of serum enzymes were not reversed by the different rivastigmine treatments. Conclusions: Rivastigmine administered 50 minutes before ischemia attenuates I/R-induced mitochondrial swelling, that indicates liver injury. This protective effect may be related to a greater stimulation of α7nAChR present in the Kupffer cells by the non-methabolized ACh, leading to an attenuation of I/R-induced inflammation.

Liver ischemia and reperfusion injury. Pathophysiology and new horizons in preconditioning and therapy

Abstract It is well known that during hepatic operative procedures, it is often critical that the irrigation is interrupted to avoid possible bleeding, blood transfusions, variable intensities, and their short and long-term consequences. It was believed in the past that the flow interruption should not exceed 20 minutes, which limited the use of this maneuver. However, it has been postulated that ischemia could be maintained for more than 60 minutes in healthy livers. The present paper review includes: 1) A brief introduction to justify the rationale of the review design; 2) Aspects of the pathophysiology of the three stages of the liver ischemia-reperfusion injury; 3) The innate and acquired immunity; 4) Oxidative stress; 5) Apoptosis and autophagy, Some essential biomarkers (Tumor Necrosis Factor-α, nitric oxide, metalloproteinases); and, finally; 6) Preventive ("cheating") strategies, non-pharmacological and pharmacological options to treat the liver IR injury.

Physical training reverses changes in hepatic mitochondrial diameter of Alloxan-induced diabetic rats / Treinamento físico reverte alterações no diâmetro de mitocôndrias hepáticas de ratos diabéticos induzidos pela Aloxana

ABSTRACT Objective To investigate the effects of physical training on metabolic and morphological parameters of diabetic rats. Methods Wistar rats were randomized into four groups: sedentary control, trained control, sedentary diabetic and trained diabetic. Diabetes mellitus was induced by Alloxan (35mg/kg) administration for sedentary diabetic and Trained Diabetic Groups. The exercise protocol consisted of swimming with a load of 2.5% of body weight for 60 minutes per day (5 days per week) for the trained control and Trained Diabetic Groups, during 6 weeks. At the end of the experiment, the rats were sacrificed and blood was collected for determinations of serum glucose, insulin, albumin and total protein. Liver samples were extracted for measurements of glycogen, protein, DNA and mitochondrial diameter determination. Results The sedentary diabetic animals presented decreased body weight, blood insulin, and hepatic glycogen, as well as increased glycemia and mitochondrial diameter. The physical training protocol in diabetic animals was efficient to recovery body weight and liver glycogen, and to decrease the hepatic mitochondrial diameter. Conclusion Physical training ameliorated hepatic metabolism and promoted important morphologic adaptations as mitochondrial diameter in liver of the diabetic rats.

RESUMO Objetivo Investigar os efeitos do treinamento físico nos parâmetros morfológicos e metabólicos de ratos diabéticos. Métodos Ratos Wistar foram randomizados para quatro grupos: controle sedentário, controle treinado, diabético sedentário e diabético treinado. Diabetes mellitus foi induzido por administração de Aloxana (35mg/kg) nos Grupos Diabético Sedentário e diabético treinado. O protocolo de treinamento físico incluiu natação com carga de 2,5% do peso corporal, por 60 minutos por dia (5 dias por semana) para os Grupos Controle Treinado e diabético treinado, durante 6 semanas. Ao final do experimento, os ratos foram sacrificados, e o sangue foi coletado para determinação das concentrações séricas de glicose, insulina, albumina e proteínas totais. Amostras do fígado foram coletadas para determinação do glicogênio, proteínas, DNA e diâmetro mitocondrial. Resultados O Grupo Sedentário Diabético apresentou redução no peso corporal, insulinemia e glicogênio hepático, além de maior glicemia e diâmetro mitocondrial hepático. O protocolo de treinamento físico em animais diabéticos foi eficiente para restaurar o peso corporal e o glicogênio hepático, além de reduzir o diâmetro mitocondrial hepático. Conclusão O treinamento físico melhorou o metabolismo hepático e promoveu importantes adaptações morfológicas, como no diâmetro mitocondrial no fígado de animais diabéticos.

Antioxidant and Anti-aging Activities of Silybum Marianum Protein Hydrolysate in Mice Treated with D-galactose / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>In the present study, we investigated the antioxidant and anti-aging effects of Silybum marianum protein hydrolysate (SMPH) in D-galactose-treated mice.</p><p><b>METHODS</b>D-galactose (500 mg/kg body weight) was intraperitoneally injected daily for 7 weeks to accelerate aging, and SMPH (400, 800, 1,200 mg/kg body weight, respectively) was simultaneously administered orally. The antioxidant and anti-aging effects of SMPH in the liver and brain were measured by biochemical assays. Transmission electron microscopy (TEM) was performed to study the ultrastructure of liver mitochondri.</p><p><b>RESULTS</b>SMPH decreased triglyceride and cholesterol levels in the D-galactose-treated mice. It significantly elevated the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), and total antioxidant capacity (T-AOC), which were suppressed by D-galactose. Monoamine oxidase (MAO) and malondialdehyde (MDA) levels as well as the concentrations of caspase-3 and 8-OHdG in the liver and brain were significantly reduced by SMPH. Moreover, it increased Bcl-2 levels in the liver and brain. Furthermore, SMPH significantly attenuated D-galactose-induced liver mitochondrial dysfunction by improving the activities of Na+-K+-ATPase and Ca2+-Mg2+-ATPase as well as mitochondrial membrane potential (ΔΨm) and fluidity. TEM showed that the degree of liver mitochondrial damage was significantly decreased by SMPH.</p><p><b>CONCLUSION</b>The results indicated that SMPH protects against D-galactose-induced accelerated aging in mice through its antioxidant and anti-aging activities.</p>

Effect of irradiation with different laser wavelengths on oxidative stress of non-hepatectomized rats

PURPOSE: To assess the effect of two laser wavelengths, either separate or combined, on intact rat livers. METHOD: Nineteen male Wistar rats (200-300 g) were submitted to laser irradiation at 5 different sites on the liver surface.Wavelengths 660 and 780 nm were used, with a dose of irradiation of 60 J/cm2/site.The animals were divided into the groups:control (C) and animals irradiated with 660 nm laser (L1), with 780 nm laser (L2) or withboth wavelengths (L3).Mitochondrial function, mitochondrial swelling, and hepatocellular malondialdehyde (MDA) levels were determined.Data were analyzed by the Mann-Whitney test, with the level of significance set at 5%. RESULTS: There was a reduction of ADP-activated respiration (state 3) in group L1 compared to group C (p=0.0016), whereas the values of group L2 were similar to control.Group L3 also showed a reduction of state 3 (p=0.0159).There was a reduction of RCR in group L1 compared to control (p=0.0001) and to group L2 (p=0.0040).Mitochondrial swelling only differed between group L3 and control (p=0.0286).There was a increase in MDA levels in group L3 compared to control (p=0.0476) and to group L2 (p=0.0286) and in group L1 compared to group L2 (p=0.0132). CONCLUSION: Although laser irradiation reduced mitochondrial function,it did not interfere with the hepatocellular energy status.

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.

Mitochondrial ATPase activity and membrane fluidity changes in rat liver in response to intoxication with Buckthorn (Karwinskia humboldtiana)

BACKGROUND: Karwinskia humboldtiana (Kh) is a poisonous plant of the rhamnacea family. To elucidate some of the subcellular effects of Kh toxicity, membrane fluidity and ATPase activities as hydrolytic and as proton-pumping activity were assessed in rat liver submitochondrial particles. Rats were randomly assigned into control non-treated group and groups that received 1,1.5 and 2 g/Kg body weight of dry powder of Kh fruit, respectively. Rats were euthanized at day 1 and 7 after treatment. RESULTS: Rats under Kh treatment at all dose levels tested, does not developed any neurologic symptoms. However, we detected alterations in membrane fluidity and ATPase activity. Lower dose of Kh on day 1 after treatment induced higher mitochondrial membrane fluidity than control group. This change was strongly correlated with increased ATPase activity and pH gradient driven by ATP hydrolysis. On the other hand, membrane fluidity was hardly affected on day 7 after treatment with Kh. Surprisingly, the pH gradient driven by ATPase activity was significantly higher than controls despite an diminution of the hydrolytic activity of ATPase. CONCLUSIONS: The changes in ATPase activity and pH gradient driven by ATPase activity suggest an adaptive condition whereby the fluidity of the membrane is altered.

Hemoplasma prevalence and hematological abnormalities associated with infection in three different cat populations from Southern Brazil / Prevalência da infecção por hemoplasmas e alterações hematológicas associadas à infecção em três diferentes populações de gatos do sul do Brasil

Three hemoplasma species are recognized in domestic cats: Mycoplasma haemofelis, ‘Candidatus Mycoplasma haemominutum’ and ‘Candidatus Mycoplasma turicensis’. We report the prevalence and hematological abnormalities of hemoplasma infection in 369 domestic cats from three different populations (blood donors, hospitalized cats and shelter cats) from Southern Brazil. Complete blood counts were performed at the time of blood collection, and DNA was extracted and tested by conventional PCR for each hemoplasma species. A total of 79 samples (21.40%) were positive for at least one species. The most prevalent hemoplasma was ‘Candidatus Mycoplasma haemominutum’, with 50/369 (13.55%) positive cats, followed by ‘Candidatus Mycoplasma turicensis’, 10/369 (2.71%), and Mycoplasma haemofelis, 8/369 (2.16%). Mycoplasma haemofelis and ‘Candidatus Mycoplasma haemominutum’ coinfection was observed in 4/369 (1.08%), whereas ‘Candidatus Mycoplasma haemominutum’ and ‘Candidatus Mycoplasma turicensis’ in 5/369 (1.35%). Three cats (0.81%) were infected with all three hemoplasmas. There was no association between infection and the different populations. Anemia was associated with Mycoplasma haemofelis and ‘Candidatus Mycoplasma haemominutum’, but not with ‘Candidatus Mycoplasma turicensis’. Male cats and cats with outdoor access were more likely to be infected. Although ‘Candidatus Mycoplasma haemominutum’ is believed to cause minimal or no hematological alterations, the infected cats studied herein were more likely to be anemic.

Três espécies de hemoplasmas são reconhecidas em gatos domésticos: Mycoplasma haemofelis, ‘Candidatus Mycoplasma haemominutum’ e ‘Candidatus Mycoplasma turicensis’. A prevalência e alterações hematológicas associadas à infecção por hemoplasmas foi estudada, em 369 gatos domésticos de três populações distintas (doadores de sangue, hospitais e gatos de abrigo) do Sul do Brasil. Foram realizados hemogramas completos no momento da coleta de sangue e as amostras tiveram seu DNA extraído e testado por PCR convencional para cada espécie de hemoplasmas. Setenta e nove amostras (21,40%) foram positivas para pelo menos uma espécie. O mais prevalente foi ‘Candidatus Mycoplasma haemominutum’ com 50/369 (13,55%) gatos positivos, seguidos por ‘Candidatus Mycoplasma turicensis’ com 10/369 (2,71%) e Mycoplasma haemofelis com 8/369 (2,16%). Coinfecção por Mycoplasma haemofelis e ‘Candidatus Mycoplasma haemominutum’ foi observada em 4/369 (1,08%), enquanto ‘Candidatus Mycoplasma haemominutum’ e ‘Candidatus Mycoplasma turicensis’ coinfectaram 5/369 (1,35%) gatos. Três (0,81%) gatos apresentaram infecção pelos três hemoplasmas. Não houve associação entre a infecção e as diferentes populações. Anemia foi associada com a infecção por Mycoplasma haemofelis e ‘Candidatus Mycoplasma haemominutum’, mas não com ‘Candidatus Mycoplasma turicensis’. Gatos machos e com acesso à rua apresentaram maior probabilidade de serem infectados. Embora se acredite que ‘Candidatus Mycoplasma haemominutum’ possa causar alterações hematológicas mínimas ou ausentes, gatos infectados encontrados neste estudo foram mais propensos à anemia.

Root Canal Penetration of a Sodium Hypochlorite Mixture Using Sonic or Ultrasonic Activation

The purpose of this ex vivo study was to determine, in "open" and "closed" systems, whether the design has an influence on the penetration length of sodium hypochlorite mixed with a radiopaque contrast medium, measured in millimeters, when delivered using positive pressure (PP) and using sonic (SI) or passive ultrasonic (PUI) activation. Sixty single-rooted teeth were divided into two groups: open and closed systems (n=30). Root canal shaping was performed to a working length of 17 mm. The samples were divided into three sub-groups (n=10) according to irrigant delivery and activation: PP, and SI or PUI activation. By using radiographs, penetration length was measured, and vapor lock was assessed. For the closed group, the penetration distance means were: PP 15.715 (±0.898) mm, SI 16.299 (±0.738) mm and PUI 16.813 (±0.465) mm, with vapor lock occurring in 53.3% of the specimens. In the open group, penetration to 17 mm occurred in 97.6% of the samples, and no vapor lock occurred. Irrigant penetration and distribution evaluation using open and closed systems provide significantly different results. For closed systems, PUI is the most effective in delivering the irrigant to working length, followed by SI.

O objetivo deste estudo in vivo foi determinar, para os sistemas "abertos" e "fechados", se o design tem influência na penetração, em milímetros, do hipoclorito de sódio misturado com um meio radiopaco quando empregado na ativação com pressão positiva (PP) e ativação sônica (SI) ou ultrassônica passiva (PUI). Sessenta dentes unirradiculares foram divididos em dois grupos: sistema aberto e sistema fechado (n=30). Os canais radiculares foram trabalhados até um comprimento de trabalho de 17 mm. Os grupos foram subdivididos em três subgrupos (n=10) de acordo com a solução irrigadora e a ativação: PP, e ativação SI ou PUI. Usando radiografias, a distância de penetração foi medida e avaliado o vapor contido. Para o grupo fechado, as distâncias médias de penetração foram PP 15,715 (±0,898) mm, SI 16,299 (±0,738) mm e PUI 16,813 (±0,465) mm e houve vapor contido em 53,3% das amostras. No grupo aberto, houve penetração de 17 mm em 97.6% das amostras, sem contenção de vapor. A penetração do irrigante e avaliação da distribuição usando sistemas aberto e fechado produziram resultados significativamente diferentes. Para os sistemas fechados, PUI é mais eficaz para levar o irrigante até preencher o comprimento de trabalho, seguido por SI.

Fiebre chikungunya en México: caso confirmado y apuntes para la respuesta epidemiológica / Chikungunya fever in Mexico: confirmed case and notes on the epidemiologic response

La fiebre chikungunya (CHIK) es una enfermedad viral transmitida al ser humano por el mismo vector del dengue, el mosquito Aedes. Además de fiebre y fuertes dolores articulares, produce otros síntomas como mialgias, cefalea, náuseas, cansancio y exantema. No tiene tratamiento específico; el manejo terapéutico de los pacientes se enfoca en el alivio de los síntomas. Históricamente se han reportado brotes de grandes proporciones; incluso desde 2010 se llegó a considerar como una potencial epidemia emergente. En 2013 se introdujo a las islas del Caribe y recientemente se ha reportado en el continente americano. En este trabajo se describe el primer caso confirmado de chikungunya en México, en el municipio de Tlajomulco de Zúñiga, Jalisco, en mayo de 2014, importado de la isla Antigua y Barbuda, en el Caribe, por una mujer de 39 años de edad.

Chikungunya fever (CHIK) is a viral disease transmitted to human beings by the same vector as dengue -the Aedes mosquito. Besides fever and severe pain in the joints, it produces other symptoms such as myalgias, headache, nausea, fatigue and exanthema. There is no specific treatment for it; the therapeutic management of patients focuses on symptom relief. Historically, outbreaks of large proportions have been reported; even since 2010 it was considered to be a potential emerging epidemic. In 2013 it was introduced into the islands of the Caribbean, and it has recently been reported in the American continent. This paper describes the first confirmed case of chikungunya in Mexico -in the municipality of Tlajomulco de Zúñiga, Jalisco, in May, 2014-, which was imported from the Caribbean island of Antigua and Barbuda by a 39 year-old woman.

Curcumin restores mitochondrial functions and decreases lipid peroxidation in liver and kidneys of diabetic db/db mice

BACKGROUND: Nitrosative and oxidative stress play a key role in obesity and diabetes-related mitochondrial dysfunction. The objective was to investigate the effect of curcumin treatment on state 3 and 4 oxygen consumption, nitric oxide (NO) synthesis, ATPase activity and lipid oxidation in mitochondria isolated from liver and kidneys of diabetic db/db mice. RESULTS: Hyperglycaemia increased oxygen consumption and decreased NO synthesis in liver mitochondria isolated from diabetic mice relative to the control mice. In kidney mitochondria, hyperglycaemia increased state 3 oxygen consumption and thiobarbituric acid-reactive substances (TBARS) levels in diabetic mice relative to control mice. Interestingly, treating db/db mice with curcumin improved or restored these parameters to normal levels; also curcumin increased liver mitochondrial ATPase activity in db/db mice relative to untreated db/db mice. CONCLUSIONS: These findings suggest that hyperglycaemia modifies oxygen consumption rate, NO synthesis and increases TBARS levels in mitochondria from the liver and kidneys of diabetic mice, whereas curcumin may have a protective role against these alterations.

Efeito do diazóxido nas lesões da isquemia/reperfusão hepática: estudo experimental em ratos / Effect of diazoxide in the lesions of ischemia/ reperfusion liver: experimental study in rats

INTRODUÇÃO: A lesão de isquemia/reperfusão hepática ocorre durante cirurgias hepáticas de grande porte, transplante de fígado e no trauma abdominal. A lesão de isquemia/reperfusão hepática ocasiona lesões no fígado e pode desencadear uma síndrome inflamatória sistêmica com lesões de órgãos a distância. Estudos anteriores demonstraram que o diazóxido protege outros órgãos (coração, rins, cérebro) da lesão de isquemia/reperfusão destes órgãos. OBJETIVO: Investigar o efeito da administração do diazóxido na lesão de isquemia/reperfusão hepática. MÉTODOS: Ratos Wistar machos foram divididos em 3 grupos. Em 2 grupos, os animais foram submetidos à isquemia hepática parcial realizada por clampeamento do pedículo dos lobos mediano e lateral anterior esquerdo durante uma hora sob ventilação mecânica. Grupo Salina (n=26): ratos receberam solução salina e Grupo Diazóxido (n=26): ratos receberam diazóxido EV ( 3.5mg/kg ) 15 minutos antes da reperfusão hepática. No terceiro grupo, Grupo Controle (n = 22 ), os ratos foram submetidos apenas à anestesia e manipulação cirúrgica. Quatro e 24 horas após os procedimentos, amostras de sangue foram recolhidas para determinações de AST, ALT, TNF-alfa, IL-6, IL-10, de nitrito/nitrato, creatinina. Amostras teciduais do fígado foram analisadas para dosagem do malondialdeído (MDA), para o estudo das funções oxidativas e fosforilativas mitocondriais, e para a análise histológica. Pela coleta de tecido pulomonar, a permeabilidade vascular pulmonar e a atividade da mieloperoxidade (MPO) também foram determinados. RESULTADOS: Quatro horas após, a reperfusão o Grupo Diazóxido apresentou elevações de AST, ALT, TNF-alfa, IL-6, IL-10 e níveis séricos de nitrito/nitrato significativamente menores que o Grupo Controle (p < 0,05). Observou-se uma redução significativa da disfunção mitocondrial hepática no Grupo Diazóxido em comparação com o Grupo Controle (p < 0,05). Não foram observadas diferenças no conteúdo de MDA fígado, na creatinina sérica e...

INTRODUCTION: Significant liver ischemia/reperfusion injury can occur during hepatic surgeries, liver transplantation and abdominal trauma. Hepatic ischemia/reperfusion can trigger a local and systemic inflammatory syndrome. Previous studies have shown that diazoxide protects other organs (heart, kidneys, brain) from ischemia/reperfusion injury. AIM: To investigate the effect of diazoxide administration on liver ischemic/reperfusion injury. METHODS: Wistar male rats were divided into 3 groups. In two groups the rats underwent partial liver ischemia performed by clamping the pedicle from medium and left anterior lateral segments during an hour under mechanical ventilation. Saline Group (n=26): rats received saline and Diazoxide Group (n=26): rats received IV diazoxide (3.5mg/kg) 15 minutes before liver reperfusion. The third group, the Control Group (n=22) the rats underwent only anesthesia and surgical manipulation. Four and 24 hours after the procedure blood were collected for determinations of AST, ALT, TNF-alfa, IL-6, IL-10, nitrite/nitrate, creatinine. Liver tissues were assembled for mitochondrial oxidation and phosphorylation, malondialdehyde (MDA) content, and histologic analysis. Pulmonary vascular permeability and myeloperoxidade (MPO) were also determined. RESULTS: Four hours after reperfusion Diazoxide Group presented elevation of AST, ALT, TNF-alfa, IL-6, IL-10 and nitrite/nitrate serum levels significantly lower than Control Group (p < 0.05). A significant reduction on liver mitochondrial dysfunction were observed in Diazoxide Group compared to Control Group (p < 0.05). No differences in liver MDA content,serum creatinine and in pulmonary vascular permeability and MPO activity were observed between groups. Twenty four hours after reperfusion Diazoxide Group showed a reduction of AST, ALT and TGF?1 serum levels when compared to Control group (p < 0.05). CONCLUSION: Diazoxide maintains liver mitochondrial function, increases liver...

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. .

Mollusc C-reactive protein crosses species barrier and reverses hepatotoxicity of lead in rodent models.

Achatina fulica C-reactive protein (ACRP) reversed the toxic effects of lead nitrate both in vivo in mice and in vitro in rat hepatocytes restoring the basal level of cell viability, lipid peroxidation, reduced glutathione and superoxides. Cytotoxicity was also significantly ameliorated in rat hepatocytes by in vitro pre-treatments with individual subunits (60, 62, 90 and 110 kDa) of ACRP. Annexin V-Cy3/CFDA dual staining showed significant reduction in the number of apoptotic hepatocytes pre-treated with ACRP. ACRP induced restoration of mitochondrial membrane potential was remarkable. ACRP pre-treatment prevented Pb-induced apoptosis mediated by caspase activation. The antagonistic effect of ACRP may be due to scavenging of reactive oxygen species which maintained the homeostasis of cellular redox potential as well as reduced glutathione status. The results suggest that ACRP crosses the species barrier and it may be utilized as a viable exogenous agent of cytoprotection against heavy metal related toxicity.