Gpx4 protects mitochondrial ATP generation against oxidative damage

Mitochondrial ATP production can be impaired by oxidative stress. Glutathione peroxidase 4 (Gpx4) is an antioxidant defense enzyme found in mitochondria as well as other subcellular organelles that directly detoxifies membrane lipid hydroperoxides. To determine if Gpx4 protects ATP production in vivo, we compared mitochondrial ATP production between wild-type mice and Gpx4 transgenic mice using a diquat model. Diquat (50 mg/kg) significantly decreased mitochondrial ATP synthesis in livers of wild-type mice; however, no decrease in mitochondrial ATP synthesis was detected in Gpx4 transgenic mice after diquat. We observed no differences in activities of mitochondrial respiratory chain complexes between Gpx4 transgenic mice and wild-type mice. However, compared to wild-type mice, diquat-induced loss of mitochondrial membrane potential was attenuated in Gpx4 transgenic mice. Therefore, our results indicate that decreased ATP production under oxidative stress is primarily due to reduced mitochondrial membrane potential and overexpression of Gpx4 maintains mitochondrial membrane potential under oxidative stress.     

The optimized allotopic expression of ND1 or ND4 genes restores respiratory chain complex I activity in fibroblasts harboring mutations in these genes

Leber's Hereditary Optic Neuropathy (LHON) was the first maternally inherited mitochondrial disease identified and is now considered the most prevalent mitochondrial disorder. LHON patients harbor mutations in mitochondrial DNA (mtDNA). In about 90% of cases, the genes involved encode proteins of the respiratory chain complex I. Even though the molecular bases are known since 20 years almost all remains to be done regarding physiopathology and therapy. In this study, we report a severe decrease of complex I activity in cultured skin fibroblasts isolated from two LHON patients harboring mutations in ND4 or ND1 genes. Most importantly, we were able to restore sustainably (a) the ability to grow on galactose, (b) the ATP synthesis rate and (c) the complex I activity, initially impaired in these cells. Our strategy consisted of forcing mRNAs from nuclearly-encoded ND1 and ND4 genes to localize to the mitochondrial surface. The rescue of the respiratory chain defect observed was possible by discreet amounts of hybrid mRNAs and fusion proteins demonstrating the efficiency of their mitochondrial import. Hence, we confirmed here for two mitochondrial genes located in the organelle that the optimized allotopic expression approach represents a powerful tool that could ultimately be applied in human therapy for LHON.     

Mitochondrial ND5 gene variation associated with encephalomyopathy and mitochondrial ATP consumption

Mitochondrial encephalomyopathy and lactic acidosis with strokelike episodes (MELAS) is a severe young onset stroke disorder without effective treatment. We have identified a MELAS patient harboring a 13528A-->G mitochondrial DNA (mtDNA) mutation in the Complex I ND5 gene. This mutation was homoplasmic in mtDNA from patient muscle and nearly homoplasmic (99.9%) in blood. Fibroblasts from the patient exhibited decreased mitochondrial membrane potential (Deltapsim) and increased lactate production, consistent with impaired mitochondrial function. Transfer of patient mtDNA to a new nuclear background using transmitochondrial cybrid fusions confirmed the pathogenicity of the 13528A-->G mutation; Complex I-linked respiration and Deltapsim were both significantly reduced in patient mtDNA cybrids compared with controls. Inhibition of the adenine nucleotide translocase or the F1F0-ATPase with bongkrekic acid or oligomycin caused a loss of potential in patient mtDNA cybrid mitochondria, indicating a requirement for glycolytically generated ATP to maintain Deltapsim. This was confirmed by inhibition of glycolysis with 2-deoxy-D-glucose, which caused depletion of ATP and mitochondrial depolarization in patient mtDNA cybrids. These data suggest that in response to impaired respiration due to the mtDNA mutation, mitochondria consume ATP to maintain Deltapsim, representing a potential pathophysiological mechanism in human mitochondrial disease.     

SIRT3 deregulation is linked to mitochondrial dysfunction in Alzheimer's disease

Alzheimer's disease (AD) is the leading cause of dementia in the elderly. Despite decades of study, effective treatments for AD are lacking. Mitochondrial dysfunction has been closely linked to the pathogenesis of AD, but the relationship between mitochondrial pathology and neuronal damage is poorly understood. Sirtuins (SIRT, silent mating type information regulation 2 homolog in yeast) are NAD‐dependent histone deacetylases involved in aging and longevity. The objective of this study was to investigate the relationship between SIRT3 and mitochondrial function and neuronal activity in AD. SIRT3 mRNA and protein levels were significantly decreased in AD cerebral cortex, and Ac‐p53 K320 was significantly increased in AD mitochondria. SIRT3 prevented p53‐induced mitochondrial dysfunction and neuronal damage in a deacetylase activity‐dependent manner. Notably, mitochondrially targeted p53 (mito‐p53) directly reduced mitochondria DNA‐encoded ND2 and ND4 gene expression resulting in increased reactive oxygen species (ROS) and reduced mitochondrial oxygen consumption. ND2 and ND4 gene expressions were significantly decreased in patients with AD. p53‐ChIP analysis verified the presence of p53‐binding elements in the human mitochondrial genome and increased p53 occupancy of mitochondrial DNA in AD. SIRT3 overexpression restored the expression of ND2 and ND4 and improved mitochondrial oxygen consumption by repressing mito‐p53 activity. Our results indicate that SIRT3 dysfunction leads to p53‐mediated mitochondrial and neuronal damage in AD. Therapeutic modulation of SIRT3 activity may ameliorate mitochondrial pathology and neurodegeneration in AD.     

Loss of parietal cell superoxide dismutase leads to gastric oxidative stress and increased injury susceptibility in mice

Mitochondrial superoxide dismutase (SOD2) prevents accumulation of the superoxide that arises as a consequence of oxidative phosphorylation. However, SOD2 is a target of oxidative/nitrosative inactivation, and reduced SOD2 activity has been demonstrated to contribute to portal hypertensive gastropathy. We investigated the consequences of gastric parietal cell-specific SOD2 deficiency on mitochondrial function and gastric injury susceptibility. Mice expressing Cre recombinase under control of the parietal cell Atpase4b gene promoter were crossed with mice harboring loxP sequences flanking the sod2 gene (SOD2 floxed mice). Cre-positive mice and Cre-negative littermates (controls) were used in studies of SOD2 expression, parietal cell function (ATP synthesis, acid secretion, and mitochondrial enzymatic activity), increased oxidative/nitrosative stress, and gastric susceptibility to acute injury. Parietal cell SOD2 deficiency was accompanied by a 20% (P < 0.05) reduction in total gastric SOD activity and a 93% (P < 0.001) reduction in gastric SOD2 activity. In SOD2-deficient mice, mitochondrial aconitase and ATP synthase activities were impaired by 36% (P < 0.0001) and 44% (P < 0.005), respectively. Gastric tissue ATP content was reduced by 34% (P < 0.002). Basal acid secretion and peak secretagogue (histamine)-induced acid secretion were reduced by 43% (P < 0.0001) and 40% (P < 0.0005), respectively. There was a fourfold (P < 0.02) increase in gastric mucosal apoptosis and 41% (P < 0.001) greater alcohol-induced gastric damage in the parietal cell SOD2-deficient mice. Our findings indicate that loss of parietal cell SOD2 leads to mitochondrial dysfunction, resulting in perturbed energy metabolism, impaired parietal cell function, and increased gastric mucosal oxidative stress. These alterations render the gastric mucosa significantly more susceptible to acute injury.     

中华雏蝗（Chorthippus chinensis Tarb）线粒体基因组分析

采用Lon-PCR扩增线粒体全基因组和保守引物步移法结合克隆方法测定并拼接获得了中华雏蝗（Chorthippus chinensis Tarb）线粒体基因组全序列．序列的注释和分析结果表 明,中华雏蝗线粒体基因组序列全长15 599 bp,共有13个编码蛋白质基因、22个tRNA基因、2个rRNA基因和1个A+T富集区．基因顺序与非洲飞蝗（Locusta migratoria）相同,也发生了2个 tRNA Asp(D)和tRNALys(K)的倒置．13个编码蛋白质基因都使用了ATN作为起始密码子．除ND1以TAG和ND5的终止密码子为不完全的T外,其余11个编码蛋白质基因的终止密码子都为完整的TAA．6种直翅类昆虫13个蛋白质的氨基酸序列的联合数据集构建的系统树与形态分类系统一致,中华雏蝗与非洲飞蝗为姐妹群,并与东方蝼蛄构成一单系群．     

Macrophage Migration Inhibitory Factor Inhibition Is Deleterious for High-Fat Diet-Induced Cardiac Dysfunction

Aims

Development of metabolic syndrome is associated with impaired cardiac performance, mitochondrial dysfunction and pro-inflammatory cytokine increase, such as the macrophage migration inhibitory factor MIF. Depending on conditions, MIF may exert both beneficial and deleterious effects on the myocardium. Therefore, we tested whether pharmacological inhibition of MIF prevented or worsened metabolic syndrome-induced myocardial dysfunction.

Methods and Results

C57BL/6J mice were fed for ten weeks with 60% fat-enriched diet (HFD) or normal diet (ND). MIF inhibition was obtained by injecting mice twice a week with ISO-1, for three consecutive weeks. Then, triglycerides, cholesterol, fat mass, glucose intolerance, insulin resistance, ex vivo cardiac contractility, animal energetic substrate utilization assessed by indirect calorimetry and mitochondrial respiration and biogenesis were evaluated. HFD led to fat mass increase, dyslipidemia, glucose intolerance and insulin resistance. ISO-1 did not alter these parameters. However, MIF inhibition was responsible for HFD-induced cardiac dysfunction worsening. Mouse capacity to increase oxygen consumption in response to exercise was reduced in HFD compared to ND, and further diminished in ISO-1-treated HFD group. Mitochondrial respiration was reduced in HFD mice, treated or not with ISO-1. Compared to ND, mitochondrial biogenesis signaling was upregulated in the HFD as demonstrated by mitochondrial DNA amount and PGC-1α expression. However, this increase in biogenesis was blocked by ISO-1 treatment.

Conclusion

MIF inhibition achieved by ISO-1 was responsible for a reduction in HFD-induced mitochondrial biogenesis signaling that could explain majored cardiac dysfunction observed in HFD mice treated with MIF inhibitor.     

Bacillus anthracis spores influence ATP synthase activity in murine macrophages

Anthrax is an infectious disease caused by toxigenic strains of the Gram-positive bacterium Bacillus anthracis. To identify the mitochondrial proteins that are expressed differently in murine macrophages infected with spores of B. anthracis Sterne, proteomic and MALDI-TOF/MS analyses of uninfected and infected macrophages were conducted. As a result, 13 mitochondrial proteins with different expression patterns were discovered in the infected murine macrophages, and some were identified as ATP5b, NIAP-5, ras-related GTP binding protein B isoform CRAa, along with several unnamed proteins. Among these proteins, ATP5b is related to energy production and cytoskeletal rearrangement, whereas NIAP-5 causes apoptosis of host cells due to binding with caspase-9. Therefore, this paper focused on ATP5b, which was found to be downregulated following infection. The downregulated ATP5b also reduced ATP production in the murine macrophages infected with B. anthracis spores. Consequently, this study represents the first mitochondrial proteome analysis of infected macrophages.     

Oxidative phosphorylation. The relation between the specific binding of trimethlytin and triethyltin to mitochondria and their effects on various mitochondrial functions

1. A binding site (site 1) is present in mitochondria with affinity for trimethyltin and triethyltin adequate for a site to which they could be attached when the processes of energy conservation are inhibited. 2. The quantitative relationships between the binding of trimethyltin and triethyltin to site 1 and their effects on various mitochondrial functions have been examined. 3. ATP synthesis linked to the oxidation of pyruvate, succinate and intramitochondrial substrate, ATP synthesis and oxygen uptake (succinate or pyruvate as substrate) stimulated by uncoupling agents are all inhibited by trimethyltin and triethyltin; when inhibition is less than 50% the ratio (percentage inhibition)/(percentage of binding site 1 complexed) is approx. 10:1. 4. ATP synthesis linked to the oxidation of reduced cytochrome c (ascorbate+NNN'N'-tetramethyl-p-phenylenediamine), ATP hydrolysis and oxygen uptake in the presence of low concentrations of trimethyltin and triethyltin approach zero activity as the proportion of binding site 1 complexed approaches 100%. 5. Possible interpretations of these findings are discussed with reference to published arrangements for coupling of electron transport to ATP synthesis and also to our present knowledge of the chemical and biological specificity of trialkyltin compounds.