Protein Restriction Without Strong Caloric Restriction Decreases Mitochondrial Oxygen Radical Production and Oxidative DNA Damage in Rat Liver

Previous studies have shown that caloric restriction decreases mitochondrial oxygen radical production and oxidative DNA damage in rat organs, which can be linked to the slowing of aging rate induced by this regime. These two characteristics are also typical of long-lived animals. However, it has never been investigated if those decreases are linked to the decrease in the intake of calories themselves or to decreases in specific dietary components. In this study the possible role of the dietary protein was investigated. Using semipurified diets, the ingestion of proteins of Wistar rats was decreased by 40% below that of controls while the other dietary components were ingested at the same level as in animals fed ad libitum. After seven weeks in this regime the liver of the protein restricted animals showed 30–40% decreases in mitochondrial production of reactive oxygen species (ROS) and in oxidative damage to nuclear and mitochondrial DNA. The decreases in ROS generation occurred specifically at complex~I. They also occurred without changes in mitochondrial oxygen consumption. Instead, there was a decrease in the percent free radical leak (the percentage of total electron flow leading to ROS generation in the respiratory chain). These results are strikingly similar to those previously obtained after 40% caloric restriction in the liver of Wistar rats. Thus, the results suggest that part of the decrease in aging rate induced by caloric restriction can be due to the decreased intake of proteins acting through decreases in mitochondrial ROS production and oxidative DNA damage. Interestingly, these tissue oxidative stress-linked parameters can be lowered by restricting only the intake of dietary protein, probably a more feasible option than caloric restriction for adult humans.     

氧化损伤与内质网应激在四氯化碳致大鼠肝脂肪变性中的作用机制

肝脂肪变性是长期饮酒、肥胖、药物中毒等致脂肪肝形成过程中重要的中间阶段,严 重的脂肪堆积会导致肝细胞坏死或肝硬化,但是有关肝脂肪变性的分子机理目前仍不十分清 楚.本实验利用四氯化碳建立大鼠肝脂肪变性模型,四氯化碳处理组较对照组肝脏丙二醛含 量增加68%,内质网应激标志蛋白GRP78 mRNA水平和蛋白质水平表达均明显增加;人肝癌细胞株HepG2体外培养中,加入四氯化碳处理后内质网发生应激,并导致SREBP-1表达增加且活化.结果表明,四氯化碳导致的肝脂肪变性与肝细胞的氧化损伤和内质网应激有关,其分子机理可能为内质网应激发生后促进SREBP-1转录因子的表达与活化,SREBP-1在细胞核内参与生脂相关酶如HMG CoA 还原酶等基因的诱导表达,生脂相关酶含量的增加进一步使肝细胞甘油三酯、胆固醇合成增加,脂质的异常堆积导致了肝脂肪变性的发生.     

Dietary Restriction at Old Age Lowers Mitochondrial Oxygen Radical Production and Leak at Complex I and Oxidative DNA Damage in Rat Brain

Previous studies in mammalian models indicate that the rate of mitochondrial reactive oxygen species ROS production and the ensuing modification of mitochondrial DNA (mtDNA) link oxidative stress to aging rate. However, there is scarce information concerning this in relation to caloric restriction (CR) in the brain, an organ of maximum relevance for ageing. Furthermore, it has never been studied if CR started late in life can improve those oxidative stress-related parameters. In this investigation, rats were subjected during 1 year to 40% CR starting at 24 months of age. This protocol of CR significantly decreased the rate of mitochondrial H₂O₂ production (by 24%) and oxidative damage to mtDNA (by 23%) in the brain below the level of both old and young ad libitum-fed animals. In agreement with the progressive character of aging, the rate of H₂O₂ production of brain mitochondria stayed constant with age. Oxidative damage to nuclear DNA increased with age and this increase was fully reversed by CR to the level of the young controls. The decrease in ROS production induced by CR was localized at Complex I and occurred without changes in oxygen consumption. Instead, the efficiency of brain mitochondria to avoid electron leak to oxygen at Complex I was increased by CR. The mechanism involved in that increase in efficiency was related to the degree of electronic reduction of the Complex I generator. The results agree with the idea that CR decreases aging rate in part by lowering the rate of free radical generation of mitochondria in the brain.     

Effects of Selenium with Vitamin E and Melatonin on Cadmium-Induced Oxidative Damage in Rat Liver and Kidneys

The present study was performed to determine the protective effects of melatonin alone and vitamin E with selenium combination against cadmium-induced oxidative damage in rat liver. A total of 60 male rats were equally divided into five groups, one of which acted as control receiving subcutaneous injections of physiological saline. The remaining four groups were treated with subcutaneous injections of cadmium chloride at a dose of 1 mg/kg weight. The first study group received no treatment. The second group was treated with a combination of 60 mg/kg vitamin E and 1 mg/kg sodium selenite. Group 3 was treated with 10 mg/kg melatonin, and the four group received a combination of vitamin E, sodium selenite, and melatonin at the doses mentioned above. After 1 month, the animals were killed, and liver and kidneys were excised for histopathological inspection and determination of tissue malondialdehyde and the activity of superoxide dismutase. The animals receiving no treatment showed significantly higher malondialdehyde levels and reduced activity of superoxide dismutase (p < 0.05). Treatment with antioxidants resulted in a significant reduction in malondialdehyde when compared to nontreated animals (p < 0.05) and increase in the enzyme activity that was almost the same as the controls. The pathological findings were also in parallel with the results of the biochemical analysis. In conclusion, all the agents tested had protective effects against cadmium-induced oxidative damage.     

Copper-1,10-Phenanthroline-Induced Apoptosis in Liver Carcinoma Bel-7402 Cells Associates with Copper Overload,Reactive Oxygen Species Production,Glutathione Depletion and Oxidative DNA Damage

The mechanism of cytotoxicity on liver carcinoma Bel-7402 cells induced by copper-1,10-phenanthroline, Cu(OP)₂, has been studied. Cell viability and apoptotic rate were examined in cells treated with Cu(OP)₂ or Cu²⁺ alone. It was found that the apoptosis induced by Cu(OP)₂ could not be induced by Cu²⁺ or OP alone in our experimental conditions. Total copper content in cells was measured by atomic absorption spectrophotometry, and the abnormal elevation of intracellular copper transported by lipophilic OP ligand may play the role of initial factor in the apoptosis, which caused subsequent redox state changes in cells. Intracellular levels of reactive oxygen species (ROS) were detected by fluorescent probe 2′,7′-dichlorofluorescein diacetate (DCFH-DA). Reduced (GSH) and total glutathione (GSSG + GSH) were determined by High-performance liquid chromatography (HPLC) after derivatization, and the ratios of GSH/GSSG were subsequently calculated. The overproduction of ROS and the decreased GSH/GSSG ratio were observed in cells which represented the occurrence of oxidative stress in the apoptosis. Oxidative DNA damage was also found in cells treated with Cu(OP)₂ in the early stage of the apoptosis, and it suggests that the activation of DNA repair system may be involved in the pathway of the apoptosis induced by Cu(OP)₂.     

Oxidative Damage and Mutagenesis in Saccharomyces cerevisiae: Genetic Studies of Pathways Affecting Replication Fidelity of 8-Oxoguanine

Oxidative damage to DNA constitutes a major threat to the faithful replication of DNA in all organisms and it is therefore important to understand the various mechanisms that are responsible for repair of such damage and the consequences of unrepaired damage. In these experiments, we make use of a reporter system in Saccharomyces cerevisiae that can measure the specific increase of each type of base pair mutation by measuring reversion to a Trp+ phenotype. We demonstrate that increased oxidative damage due to the absence of the superoxide dismutase gene, SOD1, increases all types of base pair mutations and that mismatch repair (MMR) reduces some, but not all, types of mutations. By analyzing various strains that can revert only via a specific CG → AT transversion in backgrounds deficient in Ogg1 (encoding an 8-oxoG glycosylase), we can study mutagenesis due to a known 8-oxoG base. We show as expected that MMR helps prevent mutagenesis due to this damaged base and that Pol η is important for its accurate replication. In addition we find that its accurate replication is facilitated by template switching, as loss of either RAD5 or MMS2 leads to a significant decrease in accurate replication. We observe that these ogg1 strains accumulate revertants during prolonged incubation on plates, in a process most likely due to retromutagenesis.     

High central venous oxygen saturation is associated with mitochondrial dysfunction in septic shock: A prospective observational study

To test the hypothesis that an impaired mitochondrial function is associated with altered central venous oxygen saturation (ScvO₂), venous-to-arterial carbon dioxide tension difference (delta PCO₂) or serum lactate in sepsis patients. This prospective cohort study was conducted in a single tertiary emergency department between April 2017 and March 2019. Patients with suspected sepsis were included in the study. Serum lactate was obtained in sepsis, ScvO₂ and delta PCO₂ were evaluated in septic shock patients. Mitochondrial function was determined from the peripheral blood mononuclear cells. Forty-six patients with suspected sepsis were included. Of these, twenty patients were septic shock. Mitochondrial oxidative stress levels were increased in the high ScvO₂ group (ScvO₂ > 80%, n = 6), compared with the normal (70%-80%, n = 9) and low ScvO₂ (<70%, n = 5) groups. A strong linear relationship was observed between the mitochondrial oxidative stress and ScvO₂ (r = .75; P = .01). However, mitochondrial respiration was increased in the low ScvO₂ group. In addition, mitochondrial complex II protein levels were significantly decreased in the high ScvO₂ group (P < .05). Additionally, there was no correlation between serum lactate, delta PCO₂, and mitochondria oxidative stress or mitochondria function. ScvO₂ can be potentially useful for developing new therapeutics to reduce mitochondrial dysfunction in septic shock patient.