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Obesity is associated with chronic low-grade inflammation that contributes to defects in energy metabolism and insulin resistance. Suppressor of cytokine signaling (SOCS)-3 expression is increased in skeletal muscle of obese humans. SOCS3 inhibits leptin signaling in the hypothalamus and insulin signal transduction in adipose tissue and the liver. Skeletal muscle is an important tissue for controlling energy expenditure and whole-body insulin sensitivity; however, the physiological importance of SOCS3 in this tissue has not been examined. Therefore, we generated mice that had SOCS3 specifically deleted in skeletal muscle (SOCS MKO). The SOCS3 MKO mice had normal muscle development, body mass, adiposity, appetite, and energy expenditure compared with wild-type (WT) littermates. Despite similar degrees of obesity when fed a high-fat diet, SOCS3 MKO mice were protected against the development of hyperinsulinemia and insulin resistance because of enhanced skeletal muscle insulin receptor substrate 1 (IRS1) and Akt phosphorylation that resulted in increased skeletal muscle glucose uptake. These data indicate that skeletal muscle SOCS3 does not play a critical role in regulating muscle development or energy expenditure, but it is an important contributing factor for inhibiting insulin sensitivity in obesity. Therapies aimed at inhibiting SOCS3 in skeletal muscle may be effective in reversing obesity-related glucose intolerance and insulin resistance. Diabetes 62:56-64, 2013, Obesity is associated with a chronic low-grade inflammatory response that induces defects in energy balance, insulin sensitivity, and lipid metabolism (1). The suppressor of cytokine signaling (SOCS) family of proteins [...]

We assessed structural elements of the retina in individuals with Friedreich ataxia (FRDA) and in mouse models of FRDA, as well as functions of the retinal pigment epithelium (RPE) in FRDA using induced pluripotent stem cells (iPSCs). We analyzed the retina of the FRDA mouse models YG22R and YG8R containing a human FRATAXIN (FXN) transgene by histology. We complemented this work with post-mortem evaluation of eyes from FRDA patients. Finally, we derived RPE cells from patient FRDA-iPSCs to assess oxidative phosphorylation (OXPHOS) and phagocytosis. We showed that whilst the YG22R and YG8R mouse models display elements of retinal degeneration, they do not recapitulate the loss of retinal ganglion cells (RGCs) found in the human disease. Further, RPE cells differentiated from human FRDA-iPSCs showed normal OXPHOS and we did not observe functional impairment of the RPE in Humans.

Mitochondrial DNA haplotypes are associated with various phenotypes, such as altered susceptibility to disease, environmental adaptations, and aging. Accumulating evidence suggests that mitochondrial DNA is essential for cell differentiation and the cell phenotype. However, the effects of different mitochondrial DNA haplotypes on differentiation and development remain to be determined. Using embryonic stem cell lines possessing the same Mus musculuschromosomes but harboring one of Mus musculus, Mus spretus, or Mus terricolormitochondrial DNA haplotypes, we have determined the effects of different mitochondrial DNA haplotypes on chromosomal gene expression, differentiation, and mitochondrial metabolism. In undifferentiated and differentiating embryonic stem cells, we observed mitochondrial DNA haplotype‐specific expression of genes involved in pluripotency, differentiation, mitochondrial energy metabolism, and DNA methylation. These mitochondrial DNA haplotypes also influenced the potential of embryonic stem cells to produce spontaneously beating cardiomyocytes. The differences in gene expression patterns and cardiomyocyte production were independent of ATP content, oxygen consumption, and respiratory capacity, which until now have been considered to be the primary roles of mitochondrial DNA. Differentiation of embryonic stem cells harboring the different mitochondrial DNA haplotypes in a 3D environment significantly increased chromosomal gene expression for all haplotypes during differentiation. However, haplotype‐specific differences in gene expression patterns were maintained in this environment. Taken together, these results provide significant insight into the phenotypic consequences of mitochondrial DNA haplotypes and demonstrate their influence on differentiation and development. We propose that mitochondrial DNA haplotypes play a pivotal role in the process of differentiation and mediate the fate of the cell. STEMCELLS2013;31:703–716

Acquired optic neuropathies are a common cause of blindness in adults, and are associated with characteristic morphological changes at the optic nerve head. Accurate and prompt clinical diagnosis, supplemented with imaging where appropriate, is essential to optimize management of the optic neuropathy and to counsel the patient appropriately on its natural history. History taking, optic disc findings, visual field assessment and imaging of the nerve head and surrounding retinal nerve fiber layer are all paramount to achieving the correct diagnosis. This Review highlights the optic nerve head features that are common to the acquired optic neuropathies, and describes the features that can be used to differentiate these various conditions.

Glaucoma is increasingly recognized as a neurodegenerative disorder, characterized by the accelerated loss of retinal ganglion cells (RGCs) and their axons. Open angle glaucoma prevalence and incidence increase exponentially with increasing age, yet the pathophysiology underlying increasing age as a risk factor for glaucoma is not well understood. Accumulating evidence points to age-related mitochondrial dysfunction playing a key role in the etiology of other neurodegenerative disorders including amyotrophic lateral sclerosis, Alzheimer and Parkinson disease. The 2 major functions of mitochondria are the generation of ATP through oxidative phosphorylation and the regulation of cell death by apoptosis. This review details evidence to support our hypothesis that age-associated mitochondrial dysfunction renders RGCs susceptible to glaucomatous injury by reducing the energy available for repair processes and predisposing RGCs to apoptosis. Eliciting the role of mitochondria in glaucoma pathogenesis may uncover novel therapeutic targets for protecting the optic nerve and preventing vision loss in glaucoma.

Leber's hereditary optic neuropathy (LHON) is a form of blindness caused by mitochondrial DNA (mtDNA) mutations in complex I genes. We report an extensive biochemical analysis of the mitochondrial defects in lymphoblasts and transmitochondrial cybrids harboring the three most common LHON mutations: 3460A, 11778A, and 14484C. Respiration studies revealed that the 3460A mutation reduced the maximal respiration rate 20–28%, the 11778A mutation 30–36%, and the 14484C mutation 10–15%. The respiration defects of the 3460A and 11778A mutations transferred in cybrid experiments linking these defects to the mtDNA. Complex I enzymatic assays revealed that the 3460A mutation resulted in a 79% reduction in specific activity and the 11778A mutation resulted in a 20% reduction, while the 14484C mutation did not affect the complex I activity. The enzyme defect of the 3460A mutation transferred with the mtDNA in cybrids. Overall, these data support the conclusion that the 3460A and 11778A mutants result in complex I defects and that the 14484C mutation causes a much milder biochemical defect. These studies represent the first direct comparison of oxidative phosphorylation defects among all of the primary LHON mtDNA mutations, thus permitting insight into the underlying pathophysiological mechanism of the disease.

The production of in vitroandin vivomodels of mitochondrial DNA (mtDNA) defects is currently limited by a lack of characterized mouse cell mtDNA mutants that may be expected to model human mitochondrial diseases. Here we describe the creation of transmitochondrial mouse (Mus musculus) cells repopulated with mtDNA from different murid species (xenomitochondrial cybrids). The closely related Mus spretusmtDNA is readily maintained when introduced into M. musculusmtDNA-less (ρ0) cells, and the resulting cybrids have normal oxidative phosphorylation (OXPHOS). When the more distantly related Rattus norvegicusmtDNA is transferred to the mouse nuclear background the mtDNA is replicated, transcribed, and translated efficiently. However, function of several OXPHOS complexes that depend on the coordinated assembly of nuclear and mtDNA-encoded proteins is impaired. Complex I activity in the Rattusxenocybrid was 46% of the control mean; complex III was 37%, and complex IV was 78%. These defects combined to restrict maximal respiration to 12–31% of the control and M. spretusxenocybrids, as measured polarographically using isolated cybrid mitochondria. These defects are distinct to those previously reported for human/primate xenocybrids. It should be possible to produce other mouse xenocybrid constructs with less severe OXPHOS phenotypes, to model human mtDNA diseases.

In an attempt to create an animal model of tissue-specif ic mitochondrial disease, we generated ‘knockout’ mice deficient in the heart/muscle isoform of the adenine nucleotide translocator (Ant1). Histological and ultrastructural examination of skeletal muscle from Ant1 null mutants revealed ragged-red muscle fibers and a dramatic proliferation of mitochondria, while examination of the heart revealed cardiac hypertrophy with mitochondrial proliferation. Mitochondria isolated from mutant skeletal muscle exhibited a severe defect in coupled respiration. Ant1mutant adults also had a resting serum lactate level fourfold higher than that of controls, indicative of metabolic acidosis. Significantly, mutant adults manifested severe exercise intolerance. Therefore, Ant1 mutant mice have the biochemical, histological, metabolic and physiological characteristics of mitochondrial myopathy and cardiomyopathy.

Diabetes mellitus (DM) is one of the most common chronic disorders of children and adults. Several reports have suggested an increased incidence of maternal transmission in some forms of DM. Therefore, we tested a pedigree with maternally transmitted DM and deafness for mitochondrial DNA mutations and discovered a 10.4 kilobase (kb) mtDNA deletion. This deletion is unique because it is maternally inherited, removes the light strand origin (OL) of mtDNA replication, inhibits mitochondrial protein synthesis, and is not associated with the hallmarks of mtDNA deletion syndromes. This discovery demonstrates that DM can be caused by mtDNA mutations and suggests that some of the heterogeneity of this disease results from the novel features of mtDNA genetics.

Abstract: Treatment of mouse LMTK− cells with the toxic mitochondrial dye rhodamine 6G (R-6G) at 2.5 μg/ml for 7 days prevented cell growth while maintaining viability, with less than 10−6 cells recovering to form colonies. Pre-treatment of LMTK− cells with R-6G was followed by fusion with enucleated mouse 501–1 cells harboring a homoplasmic point mutation in the mitochondrial DNA (mtDNA) 16S rRNA gene conferring chloramphenicol resistance (CAPR). Cybrids and any surviving unfused LMTK− cells were selected in BrdU with or without CAP and their mtDNAs screened for the presence of the CAPR marker. Approximately 1 colony per 2105 LMTK− cells appeared in the fusion plates selected both with and without CAP. Most clones investigated were confirmed to be cybrids by showing the presence of the generally homoplasmic CAPR mutation, whether or not CAP selection was used. Hence, R-6G pre-treatment permits construction of transmitochondrial cybrid cell lines carrying a variety of mtDNAs, without the need for ρo cell lines.

State III (activated) mitochondrial respiration rates with pyruvate/malate, glutamate/malate, and succinate as substrates were assayed in isolated intact skeletal muscle mitochondria in 29 subjects aged 16-92 years. There was a significant negative correlation between respiration rate and age with all substrates tested. A similar trend was seen for respiratory enzyme activities assayed in muscle homogenate. These findings suggest a substantial fall in mitochondrial oxidative capacity in ageing muscle, which may contribute to reduced exercise capacity in elderly people. Mitochondrial respiratory failure may contribute to the ageing process in other organs.

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A family is presented in which 7 members over 3 generations were affected by cardioskeletal myopathy. A vacuolar myopathy with excessive free and intralysosomal glycogen storage in skeletal and cardiac striated muscle was identified in biopsy studies. Post-mortem studies in several patients revealed changes of a congestive cardiomyopathy with myocardial fibrosis. Acid maltase, phosphorylase, debrancher and lysosomal enzyme screens, and glycolytic enzyme levels in skeletal muscle, were normal in 1 case. This is the third report of non-acid maltase deficient lysosomal glycogen storage disease and adds to previous reports with the presentation of detailed family studies, examination of ante- and post-mortem cardiac histology and reports of detailed glycolytic and lysosomal enzyme analysis. This syndrome is unusual among glycogenoses in having a dominant inheritance pattern.