Antioxidant treatment improves in vivo cardiac and skeletal muscle bioenergetics in patients with Friedreich's ataxia

Friedreich's ataxia (FA) is the most common form of autosomal recessive spinocerebellar ataxia and is often associated with a cardiomyopathy. The disease is caused by an expanded intronic GAA repeat, which results in deficiency of a mitochondrial protein called frataxin. In the yeast YFH1 knockout model of the disease there is evidence that frataxin deficiency leads to a severe defect of mitochondrial respiration, intramitochondrial iron accumulation, and associated production of oxygen free radicals. Recently, the analysis of FA cardiac and skeletal muscle samples and in vivo phosphorus magnetic resonance spectroscopy (31P-MRS) has confirmed the deficits of respiratory chain complexes in these tissues. The role of oxidative stress in FA is further supported by the accumulation of iron and decreased aconitase activities in cardiac muscle. We used 31P-MRS to evaluate the effect of 6 months of antioxidant treatment (Coenzyme Q10 400 mg/day, vitamin E 2,100 IU/day) on cardiac and calf muscle energy metabolism in 10 FA patients. After only 3 months of treatment, the cardiac phosphocreatine to ATP ratio showed a mean relative increase to 178% (p = 0.03) and the maximum rate of skeletal muscle mitochondrial ATP production increased to 139% (p = 0.01) of their respective baseline values in the FA patients. These improvements, greater in prehypertrophic hearts and in the muscle of patients with longer GAA repeats, were sustained after 6 months of therapy. The neurological and echocardiographic evaluations did not show any consistent benefits of the therapy after 6 months. This study demonstrates partial reversal of a surrogate biochemical marker in FA with antioxidant therapy and supports the evaluation of such therapy as a disease-modifying strategy in this neurodegenerative disorder.     

Creatine has no beneficial effect on skeletal muscle energy metabolism in patients with single mitochondrial DNA deletions: a placebo-controlled, double-blind 31P-MRS crossover study

The purpose of our randomized, double-blind, placebo-controlled crossover study in 15 patients with chronic progressive external ophthalmoplegia (CPEO) or Kearns-Sayre syndrome (KSS) because of single large-scale mitochondrial (mt) DNA deletions was to determine whether oral creatine (Cr) monohydrate can improve skeletal muscle energy metabolism in vivo. Each treatment phase with Cr in a dosage of 150 mg/kg body weight/day or placebo lasted 6 weeks. The effect of Cr was estimated by phosphorus-31 magnetic resonance spectroscopy ((31)P-MRS), clinical and laboratory tests. (31)P-MRS analysis prior to treatment showed clear evidence of severe mitochondrial dysfunction. However, there were no relevant changes in (31)P-MRS parameters under Cr. In particular, phosphocreatine (PCr)/ATP at rest did not increase, and there was no facilitation of post-exercise PCr recovery. Clinical scores and laboratory tests did not alter significantly under Cr, which was tolerated without major side-effects in all patients. Cr supplementation did not improve skeletal muscle oxidative phosphorylation in our series of patients. However, one explanation for our negative findings may be the short study duration or the limited number of patients included.     

Effects of sleep deprivation: The phosphorus metabolism in the human brain measured by 31P-magnetic resonance spectroscopy

Sleep deprivation (SD) has an antidepressant effect in some, but not all, patients with depression, although its biological mechanisms have not yet been characterized. We previously reported altered brain phosphorus metabolism measured by phosphorus-31 magnetic resonance spectroscopy (31P-MRS) in patients with bipolar depression. We preliminarily examined effects of SD on phosphorus metabolism in the frontal lobes of 15 normal subjects using 31P-MRS. No significant differences of membrane phospholipid metabolism, high-energy phosphate metabolism and intracellular pH were found between before and after SD in these subjects. Further studies will be necessary to elucidate the physiological mechanism of SD for depressive patients.     

Aerobic conditioning in patients with mitochondrial myopathies: physiological, biochemical, and genetic effects

Aerobic training has been shown to increase work and oxidative capacity in patients with mitochondrial myopathies, but the mechanisms underlying improvement are not known. We evaluated physiological (cycle exercise, 31P-MRS), biochemical (enzyme levels), and genetic (proportion of mutant/wild-type genomes) responses to 14 weeks of bicycle exercise training in 10 patients with heteroplasmic mitochondrial DNA (mtDNA) mutations. Training increased peak work and oxidative capacities (20-30%), systemic arteriovenous O2 difference (20%), and 31P-MRS indices of metabolic recovery (35%), consistent with enhanced muscle oxidative phosphorylation. Mitochondrial volume in vastus lateralis biopsies increased significantly (50%) and increases in deficient respiratory chain enzymes were found in patients with Complex I (36%) and Complex IV (25%) defects, whereas decreases occurred in 2 patients with Complex III defects (approximately 20%). These results suggest that the cellular basis of improved oxygen utilization is related to training-induced mitochondrial proliferation likely resulting in increased levels of functional, wild-type mtDNA. However, genetic analysis indicated the proportion of wild-type mtDNA was unchanged (3/9) or fell (6/9), suggesting a trend toward preferential proliferation of mutant genomes. The long-term implications of training-induced increases in mutant relative to wild-type mtDNA, despite positive physiological and biochemical findings, need to be assessed before aerobic training can be proposed as a general treatment option.     

Neurochemical Investigation of the Schizophrenic Brain by in Vivo Phosphorus Magnetic Resonance Spectroscopy

Abnormal phospholipid metabolisms may play important roles in the pathophysiology of schizophrenia. Phosphorus magnetic resonance spectroscopy (³¹P-MRS) offers a new method for studying phosphorus-related metabolism in vivo. A decrease in the level of phosphomonoesters (PME) and an increase in the level of phosphodiesters (PDE) has been demonstrated in the prefrontal lobe of neuroleptic-naive schizophrenic patients. Most of the studies in medicated schizophrenic patients have shown decreased PME and/or increased PDE. The decreased PME in the frontal lobe appears to be associated with negative symptoms and poor working memory performance. ¹H-decoupled ³¹P-MRS revealed a reduction in the phosphocholine element of PME and an elevation in the mobile phospholipids of PDE in the prefrontal region of medicated schizophrenic patients. PDE were elevated in the temporal lobes of neuroleptic-naive schizophrenic patients, and this increase was partially normalized by haloperidol administration. Data about the temporal lobes of medicated schizophrenic patients have not been consistent. Except for the reduction in the adenosine triphosphate (ATP) in the basal ganglia and the correlation between the increase in the frontal lobe phosphocreatine (PCr) and negative symptomatology, data related to changes in high-energy phosphates are contradictory. No consensus on the effect of neuroleptics on phosphorus metabolites has been achieved. Methodological problems inherent in ³¹P-MRS may have contributed to the confusion in understanding available data. Future directions of MRS studies are suggested in the last section of the paper.     

Abnormal brain energy metabolism shown by in vivo phosphorus magnetic resonance spectroscopy in patients with chronic liver disease

We used phosphorus magnetic resonance spectroscopy (31P-MRS) to assess in vivo the brain bioenergetics of 28 patients with liver cirrhosis. Seven had clinical hepatic encephalopathy (HE), nine hepatocellular carcinoma. 31P-MRS was performed by the DRESS localisation technique on occipital lobes. Brain phosphocreatine was significantly reduced in patients with or without overt HE, and inorganic phosphate was increased in both groups of patients. The cytosolic phosphorylation potential (PP), the relative rate of oxidative metabolism and the regulatory [ADP] were all abnormal. Brain PP was inversely correlated with serum ammonia concentration only in patients without liver cancer. The degree of bioenergetic failure was significantly higher in the presence of overt encephalopathy. We conclude that patients with liver cirrhosis had a derangement of brain energy metabolism, and that 31P-MRS offers a non-invasive method for investigating the underlying mechanisms of HE, with relevant implications in the identification and management of this condition.     

Effects of Erythropoietin on Frataxin Levels and Mitochondrial Function in Friedreich Ataxia �C a Dose�CResponse Trial

Friedreich ataxia (FRDA) is an autosomal recessive inherited neurodegenerative disorder leading to reduced expression of the mitochondrial protein frataxin. Previous studies showed frataxin upregulation in FRDA following treatment with recombinant human erythropoietin (rhuEPO). Dose-response interactions between frataxin and rhuEPO have not been studied until to date. We administered escalating rhuEPO single doses (5,000, 10,000 and 30,000?IU) in monthly intervals to five adult FRDA patients. Measurements of frataxin, serum erythropoietin levels, iron metabolism and mitochondrial function were carried out. Clinical outcome was assessed using the "Scale for the assessment and rating of ataxia". We found maximal erythropoietin serum concentrations 24?h after rhuEPO application which is comparable to healthy subjects. Frataxin levels increased significantly over 3?months, while ataxia rating did not reveal clinical improvement. All FRDA patients had considerable ferritin decrease. NADH/NAD ratio, an indicator of mitochondrial function, increased following rhuEPO treatment. In addition to frataxin upregulation in response to continuous low-dose rhuEPO application shown in previous studies, our results indicate for a long-lasting frataxin increase after single high-dose rhuEPO administration. To detect frataxin-derived neuroprotective effects resulting in clinically relevant improvement, well-designed studies with extended time frame are required.     

Neurochemical investigation of the schizophrenic brain by in vivo phosphorus magnetic resonance spectroscopy.

Abnormal phospholipid metabolisms may play important roles in the pathophysiology of schizophrenia. Phosphorus magnetic resonance spectroscopy (31P-MRS) offers a new method for studying phosphorus-related metabolism in vivo. A decrease in the level of phosphomonoesters (PME) and an increase in the level of phosphodiesters (PDE) has been demonstrated in the prefrontal lobe of neuroleptic-naive schizophrenic patients. Most of the studies in medicated schizophrenic patients have shown decreased PME and/or increased PDE. The decreased PME in the frontal lobe appears to be associated with negative symptoms and poor working memory performance. 1H-decoupled 31P-MRS revealed a reduction in the phosphocholine element of PME and an elevation in the mobile phospholipids of PDE in the prefrontal region of medicated schizophrenic patients. PDE were elevated in the temporal lobes of neuroleptic-naive schizophrenic patients, and this increase was partially normalized by haloperidol administration. Data about the temporal lobes of medicated schizophrenic patients have not been consistent. Except for the reduction in the adenosine triphosphate (ATP) in the basal ganglia and the correlation between the increase in the frontal lobe phosphocreatine (PCr) and negative symptomatology, data related to changes in high-energy phosphates are contradictory. No consensus on the effect of neuroleptics on phosphorus metabolites has been achieved. Methodological problems inherent in 31P-MRS may have contributed to the confusion in understanding available data. Future directions of MRS studies are suggested in the last section of the paper.     

Implications of lipid biology for the pathogenesis of schizophrenia

OBJECTIVE: Preclinical and clinical data suggest that lipid biology is integral to brain development and neurodegeneration. Both aspects are proposed as being important in the pathogenesis of schizophrenia. The purpose of this paper is to examine the implications of lipid biology, in particular the role of essential fatty acids (EFA), for schizophrenia. METHODS: Medline databases were searched from 1966 to 2001 followed by the cross-checking of references. RESULTS: Most studies investigating lipids in schizophrenia described reduced EFA, altered glycerophospholipids and an increased activity of a calcium-independent phospholipase A2 in blood cells and in post-mortem brain tissue. Additionally, in vivo brain phosphorus-31 Magnetic Resonance Spectroscopy (31P-MRS) demonstrated lower phosphomonoesters (implying reduced membrane precursors) in first- and multi-episode patients. In contrast, phosphodiesters were elevated mainly in first-episode patients (implying increased membrane breakdown products), whereas inconclusive results were found in chronic patients. EFA supplementation trials in chronic patient populations with residual symptoms have demonstrated conflicting results. More consistent results were observed in the early and symptomatic stages of illness, especially if EFA with a high proportion of eicosapentaenoic acid was used. CONCLUSION: Peripheral blood cell, brain necropsy and 31P-MRS analysis reveal a disturbed lipid biology, suggesting generalized membrane alterations in schizophrenia. 31P-MRS data suggest increased membrane turnover at illness onset and persisting membrane abnormalities in established schizophrenia. Cellular processes regulating membrane lipid metabolism are potential new targets for antipsychotic drugs and might explain the mechanism of action of treatments such as eicosapentaenoic acid.     

The pathogenesis of Friedreich ataxia and the structure and function of frataxin

Understanding the role of frataxin in mitochondria is key to an understanding of the pathogenesis of Friedreich ataxia. Frataxins are small essential proteins whose deficiency causes a range of metabolic disturbances, which include oxidative stress, deficit of iron-sulphur clusters, and defects in heme synthesis, sulfur amino acid and energy metabolism, stress response, and mitochondrial function. Structural studies carried out on different orthologues have shown that the frataxin fold consists of a flexible N-terminal region present only in eukaryotes and in a highly conserved C-terminal globular domain. Frataxins bind iron directly but with very unusual properties: iron coordination is achieved solely by glutamates and aspartates exposed on the protein surface. It has been suggested that frataxin function is that of a ferritin-like protein, an iron chaperone of the ironsulphur cluster machinery and heme metabolism and/or a controller of cellular oxidative stress. To understand FRDA pathogenesis and to design novel therapeutic strategies, we must first precisely identify the cellular role of frataxin.     

Magnetic resonance spectroscopy in schizophrenia

Numerous studies have shown alterations of some structures and/or cerebral functions in patients with schizophrenia. However, the nature of the neurobiological process which could be at the origin of schizophrenic symptoms is still unknown. Magnetic resonance spectroscopy (MRS) is a unique technique which allows us to estimate the concentrations of endogenous substances which contain natural paramagnetic nuclei such as phosphorus (31P) and hydrogen (proton or 1H). The non invasive character of this technique, the absence of side effects, and the possibility of repetitive evaluations allowing for longitudinal studies, make possible MRS studies on the in vivo cerebral metabolism in schizophrenia. The prefrontal cortex, the hippocampus and the basal ganglia have all been implicated in the pathophysiology of schizophrenia. Therefore these brain regions have been frequently studied using MRS. Both proton and phosphorus spectroscopy have been used to study schizophrenia. Compounds that are detectable by 1H-MRS include N-acetyl aspartate (NAA), choline (Cho), creatine (Cr) and myo-inositol (ml). A deficit in NAA has been consistently shown in both the frontal and temporal lobes suggesting neuronal loss in these areas. Compounds detectable by 31P-MRS include phosphomonoesters (PMEs) and phosphodiesters (PDEs), which largely represent metabolites generated by lipid turnover. 31P-MRS can also detect certain energy-containing phosphorus metabolites such as phosphocreatine (PCr) and nucleotide triphosphates. Decreased levels of PMEs and increased levels of PDEs have been consistently described in the prefrontal lobes suggesting an alteration of phospholipid metabolism. The purpose of this review is to summarize the research on schizophrenia using MRS, to show the utility of this technique in understanding schizophrenia.     

Mesenchymal Stem Cell-Derived Factors Restore Function to Human Frataxin-Deficient Cells

Friedreich’s ataxia is an inherited neurological disorder characterised by mitochondrial dysfunction and increased susceptibility to oxidative stress. At present, no therapy has been shown to reduce disease progression. Strategies being trialled to treat Friedreich’s ataxia include drugs that improve mitochondrial function and reduce oxidative injury. In addition, stem cells have been investigated as a potential therapeutic approach. We have used siRNA-induced knockdown of frataxin in SH-SY5Y cells as an in vitro cellular model for Friedreich’s ataxia. Knockdown of frataxin protein expression to levels detected in patients with the disorder was achieved, leading to decreased cellular viability, increased susceptibility to hydrogen peroxide-induced oxidative stress, dysregulation of key anti-oxidant molecules and deficiencies in both cell proliferation and differentiation. Bone marrow stem cells are being investigated extensively as potential treatments for a wide range of neurological disorders, including Friedreich’s ataxia. The potential neuroprotective effects of bone marrow-derived mesenchymal stem cells were therefore studied using our frataxin-deficient cell model. Soluble factors secreted by mesenchymal stem cells protected against cellular changes induced by frataxin deficiency, leading to restoration in frataxin levels and anti-oxidant defences, improved survival against oxidative stress and stimulated both cell proliferation and differentiation down the Schwann cell lineage. The demonstration that mesenchymal stem cell-derived factors can restore cellular homeostasis and function to frataxin-deficient cells further suggests that they may have potential therapeutic benefits for patients with Friedreich’s ataxia.     

Energy metabolism in muscle paresis and recovery studied by31P-MR spectroscopy: A case report

We report a³¹P Magnetic Resonance Spectroscopy (³¹P-MRS) study on the changes of energy metabolism in human leg anterior and lateral compartment muscles with paresis due to compression from a herniated lumbar disc at the L₄-L₅ level and recovery to normality. A low phosphocreatine to inorganic phosphate ratio due to both decreased phosphocreatine and increased inorganic phosphate contents and a normal intracellular pH were the features of muscles with paresis. Changes of³¹P-MRS parameters were followed during 18 weeks of treatment with physical therapy until complete recovery. Results show that³¹P-MRS is a useful clinical tool for detecting even small biochemical changes that may occur in muscles and for checking the effects of therapy.     

Friedreich's ataxia: past, present and future

Friedreich's ataxia (FRDA) is an autosomal recessive inherited disorder characterized by progressive gait and limb ataxia, dysarthria, areflexia, loss of vibratory and position sense, and a progressive motor weakness of central origin. Additional features include hypertrophic cardiomyopathy and diabetes. Large GAA repeat expansions in the first intron of the FXN gene are the most common mutation underlying FRDA. Patients show severely reduced levels of a FXN-encoded mitochondrial protein called frataxin. Frataxin deficiency is associated with abnormalities of iron metabolism: decreased iron-sulfur cluster (ISC) biogenesis, accumulation of iron in mitochondria and depletion in the cytosol, enhanced cellular iron uptake. Some models have also shown reduced heme synthesis. Evidence for oxidative stress has been reported. Respiratory chain dysfunction aggravates oxidative stress by increasing leakage of electrons and the formation of superoxide. In vitro studies have demonstrated that Frataxin deficient cells not only generate more free radicals, but also show a reduced capacity to mobilize antioxidant defenses. The search for experimental drugs increasing the amount of frataxin is a very active and timely area of investigation. In cellular and in animal model systems, the replacement of frataxin function seems to alleviate the symptoms or even completely reverse the phenotype. Therefore, drugs increasing the amount of frataxin are attractive candidates for novel therapies. This review will discuss recent findings on FRDA pathogenesis, frataxin function, new treatments, as well as recent animal and cellular models. Controversial aspects are also discussed.     

Iron dysregulation in Friedreich ataxia

Friedreich ataxia is the most common hereditary ataxia. The signs and symptoms of the disorder derive from decreased expression of the protein frataxin, which is involved in iron metabolism. Frataxin chaperones iron for iron-sulfur cluster biogenesis and detoxifies iron in the mitochondrial matrix. Decreased expression of frataxin is associated with impairments of iron-sulfur cluster biogenesis and heme synthesis, as well as with mitochondrial dysfunction and oxidative stress. Compounds currently in clinical trials are directed toward improving mitochondrial function and lessening oxidative stress. Iron chelators and compounds that increase frataxin expression are under evaluation. Further elucidation of frataxin's function should lead to additional therapeutic approaches.     

A 31P-magnetic resonance spectroscopy and biochemical study of the movbr mouse: Potential model for the mitochondrial encephalomyopathies

³¹P-magnetic resonance spectroscopy (³¹P-MRS) provides new biochemical information on mitochondrial disorders affecting brain and muscle. To elucidate the mechanisms of mitochondrial abnormalities, however, animal models are needed. We assessed the mo^vbr (mottled viable brindled) mouse for its value in studying (1) energetics of a mitochondrial disorder and (2) ³¹P-MRS changes associated with mitochondrial abnormalities in vivo. The maximal activity of succinate-cytochrome c reductase was significantly reduced in mo^vbr muscle compared to controls, whereas cytochrome oxidase activity was only reduced in mo^vbr brain. ³¹P-MRS of mo^vbr brain showed an increased pH, but no changes in any metabolite ratios. The phosphocreatine (PCr) recovery rate after exercise was reduced in muscles from mo^vbr mice, indicating impairment of oxidative metabolism. We conclude that mo^vbr brain and muscle tissue have biochemical abnormalities consistent with mitochondrial impairment. The PCr recovery rate, measured by ³¹P-MRS, was sensitive to the muscle abnormality. This strain is best described as having chronic mitochondrial dysfunction. © 1997 John Wiley & Sons, Inc. Muscle Nerve 20: 1352–1359, 1997     

Increase of phosphodiesters during neuroleptic treatment of schizophrenics: a longitudinal 31P-magnetic resonance spectroscopic study.

BACKGROUND: Increased levels of phosphodiesters (PDE%) and reduced relative concentrations of phosphomonoesters (PME%) have been reported in unmedicated schizophrenics, whereas findings in brain of medicated patients were not consistent. METHODS: We determined in vivo the metabolism of phospholipids and high-energy phosphates in the left and right frontal lobes of 8 patients with schizophrenia using 31P-magnetic resonance spectroscopy (31P-MRS). Serial investigations were performed first after a neuroleptic-free period (mean 7.5 +/- 1.9 days) and second, after neuroleptic treatment (mean 20.6 +/- 11.1 days). RESULTS: PDE% increased significantly in the left frontal lobe (32.0 +/- 5.9% versus 36.9 +/- 5.6%, p = .009) after medication. All other parameters showed no significant differences. CONCLUSIONS: Our study suggests that neuroleptics do not decrease phospholipase A2 activity in schizophrenia. Individual neuroleptics may have different effects on phospholipase A2 activity as indicated by animal studies. An influence of neuroleptics on high-energy phosphates cannot be confirmed by our data.     

Simultaneous surface electromyography (SEMG) and 31P-MR spectroscopy measurements of the lumbar back muscle during isometric exercise

The aim of this study was to demonstrate the feasibility of simultaneous surface electromyography (SEMG) and 31P-MR spectroscopy (31P-MRS) measurements on the back muscle of volunteers during the performance of an isometric exercise. Six volunteers (three male, three female) performed a modified Biering-S?rensen test inside a 1.5 T MR scanner while simultaneously recording SEMG signals. A surface coil was used for 31P-MRS with a CSI sequence. Spectra were collected with a voxel resolution of 40 mm x 40 mm x 100 mm and a temporal resolution of 30 s during periods of rest, sustained muscle contraction and recovery. The duration of muscle contraction was 150 s. SEMG analysis yielded a decrease of the mean SEMG frequency of approximately 20%. The SEMG amplitudes were constant or increased up to approximately 150% during exercise. 31P-MRS showed a maximum decrease of the phosphocreatine (PCr) amplitude down to approximately 32% of its initial value. Simultaneously, a doubling of the inorganic phosphate (Pi) signal was observed. The present study demonstrates that simultaneous SEMG and 31P-MRS measurements of the back muscle are feasible during isometric exercises.     

Abnormal in vivo skeletal muscle energy metabolism in Huntington's disease and dentatorubropallidoluysian atrophy

We studied in vivo muscle energy metabolism in patients with Huntington's disease (HD) and dentatorubropallidoluysian atrophy (DRPLA) using 31P magnetic resonance spectroscopy (MRS). Twelve gene-positive HP patients (4 presymptomatic patients) and 2 gene-positive DRPLA patients (1 presymptomatic patient) were studied. 31P-MRS at rest showed a reduced phosphocreatine-to-inorganic phosphate ratio in the symptomatic HD patients and DRPLA patient. Muscle adenosine triphosphate/(phosphocreatine + inorganic phosphate) at rest was significantly reduced in both groups of symptomatic and presymptomatic HD subjects and was below the normal range in the 2 DRPLA subjects. During recovery from exercise, the maximum rate of mitochondrial adenosine triphosphate production was reduced by 44% in symptomatic HD patients and by 35% in presymptomatic HD carriers. The maximum rate of mitochondrial adenosine triphosphate production in muscle was also reduced by around 46% in the 2 DRPLA subjects. Our findings show that HD and DRPLA share a deficit of in vivo mitochondrial oxidative metabolism, supporting a role for mitochondrial dysfunction as a factor involved in the pathogenesis of these polyglutamine repeat-mediated neurodegenerative disorders. The identification of 31P-MRS abnormalities may offer a surrogate biochemical marker by which to study disease progression and the effects of treatment in HD and DRPLA.