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Pearson marrow-pancreas syndrome (PS) is a rare mitochondrial disorder. Impaired mitochondrial respiratory chain complexes (MRCC) differ among individuals and organs, which accounts for variable clinical pictures. A subset of PS patients develop 3-methylglutaconic aciduria (3-MGA-uria), but the characteristic symptoms and impaired MRCC remain unknown. Our patient, a girl, developed pancytopenia, hyperlactatemia, steatorrhea, insulin-dependent diabetes mellitus, liver dysfunction, Fanconi syndrome, and 3-MGA-uria. She died from cerebral hemorrhage at 3 years of age. We identified a novel 5.4-kbp deletion of mitochondrial DNA. The enzymatic activities of MRCC I and IV were markedly reduced in the liver and muscle and mildly reduced in skin fibroblasts and the heart. To date, urine organic acid analysis has been performed on 29 PS patients, including our case. Eight patients had 3-MGA-uria, while only one patient did not. The remaining 20 patients were not reported to have 3-MGA-uria. In this paper, we included these 20 patients as PS patients without 3-MGA-uria. PS patients with and without 3-MGA-uria have similar manifestations. Only a few studies have examined the enzymatic activities of MRCC.No clinical characteristics distinguish between PS patients with and without 3-MGA-uria. The correlation between 3-MGA-uria and the enzymatic activities of MRCC remains to be elucidated.• The clinical characteristics of patients with Pearson marrow-pancreas syndrome and 3-methylglutaconic aciduria remain unknown.• No clinical characteristics distinguish between Pearson marrow-pancreas syndrome patients with and without 3-methylglutaconic aciduria.

AIM To examine the contribution of PTPN2 coding variants to the risk of childhood-onset Type 1A diabetes. METHODS PTPN2 mutation analysis was carried out for 169 unrelated Japanese people with childhood-onset Type 1A diabetes. We searched for coding variants that were absent or extremely rare in the general population and were scored as damaging by multiple in silico programs. We performed mRNA analysis and three-dimensional structural prediction of the detected variants, when possible. We also examined possible physical links between these variants and previously reported risk SNPs as well as clinical information from variant-positive children. RESULTS One frameshift variant (p.Q286Yfs*24) and two probably damaging missense substitutions (p.C232W and p.R350Q) were identified in one child each. Of these, p.Q286Yfs*24 and p.C232W were hitherto unreported, while p.R350Q accounted for 2/121,122 alleles of the exome datasets. The p.Q286Yfs*24 variant did not encode stable mRNA, and p.C232W appeared to affect the structure of the tyrosine-protein phosphatase domain. The three variants were physically unrelated to known risk SNPs. The variant-positive children manifested Type 1A diabetes without additional clinical features and invariably carried risk human leukocyte antigen alleles. CONCLUSIONS The results provide the first indication that PTPN2 variants contribute to the risk of Type 1A diabetes, independently of known risk SNPs. PTPN2 coding variants possibly induce non-specific Type 1A diabetes phenotypes in individuals with human leukocyte antigen-mediated disease susceptibility. Our findings warrant further validation.

Introduction: Mitochondrial dysfunction results in a wide range of organ disorders through diverse genetic abnormalities. We herein present the detailed clinical course of an infant admitted for extensive, rapidly progressing white matter lesions and hypertrophic cardiomyopathy due to a BOLA3 gene mutation. Case: A 6-month-old girl with no remarkable family or past medical history until 1 month prior presented with developmental regression and feeding impairment. Ultrasound cardiography and brain magnetic resonance imaging (MRI) respectively disclosed the presence of hypertrophic cardiomyopathy and symmetrical deep white matter lesions. She was transferred to our hospital at age 6 months. High lactate levels in her cerebrospinal fluid suggested mitochondrial dysfunction. Despite vitamin supplementation therapy followed by a ketogenic diet, the patient began exhibiting clusters of myoclonic seizures and respiratory failure. Brain and spinal cord MRI revealed rapid progression of the white matter lesions. She died at 10 months of age. Fibroblasts obtained pre-mortem displayed low mitochondrial respiratory chain complex I and II activity. A homozygous H96R (c. 287 A > G) mutation was identified in the BOLA3 gene. Discussion: No reported case of a homozygous BOLA3 gene mutation has survived past 1 year of life. BOLA3 appears to play a critical role in the electron transport system and production of iron-sulfur clusters that are related to lipid metabolism and enzyme biosynthesis.

Background Carnitine palmitoyltransferase (CPT) II deficiency is one of the most common forms of mitochondrial fatty acid oxidation disorder (FAOD). However, newborn screening (NBS) for this potentially fatal disease has not been established partly because reliable indices are not available. Methods We diagnosed CPT II deficiency in a 7-month-old boy presenting with hypoglycemic encephalopathy, which apparently had been missed in the NBS using C16 and C18:1 concentrations as indices. By referring to his acylcarnitine profile from the NBS, we adopted the (C16 + C18:1)/C2 ratio (cutoff 0.62) and C16 concentration (cutoff 3.0 nmol/mL) as alternative indices for CPT II deficiency such that an analysis of a dried blood specimen collected at postnatal day five retroactively yielded the correct diagnosis. Thereafter, positive cases were assessed by measuring (1) the fatty acid oxidation ability of intact lymphocytes and/or (2) CPT II activity in the lysates of lymphocytes. The diagnoses were then further confirmed by genetic analysis. Results The disease was diagnosed in seven of 21 newborns suspected of having CPT II deficiency based on NBS. We also analyzed the false-negative patient and five symptomatic patients for comparison. Values for the NBS indices of the false-negative, symptomatic patient were lower than those of the seven affected newborns. Although it was difficult to differentiate the false-negative patient from heterozygous carriers and false-positive subjects, the fatty acid oxidation ability of the lymphocytes and CPT II activity clearly confirmed the diagnosis. Among several other indices proposed previously, C14/C3 completely differentiated the seven NBS-positive patients and the false-negative patient from the heterozygous carriers and the false-positive subjects. Genetic analysis revealed 16 kinds of variant alleles. The most prevalent, detected in ten alleles in nine patients from eight families, was c.1148T > A (p.F383Y), a finding in line with those of several previous reports on Japanese patients. Conclusions These findings suggested that CPT II deficiency can be screened by using (C16 + C18:1)/C2 and C16 as indices. An appropriate cutoff level is required to achieve adequate sensitivity albeit at the cost of a considerable increase in the false-positive rate, which might be reduced by using additional indices such as C14/C3.

Medium-chain acyl-Coenzyme A dehydrogenase (MCAD) is involved in the initial step of mitochondrial fatty acid β-oxidation (FAO). Loss of function results in MCAD deficiency, a disorder that usually presents in childhood with hypoketotic hypoglycemia, vomiting and lethargy. While the disruption of mitochondrial fatty acid metabolism is the primary metabolic defect, secondary defects in mitochondrial oxidative phosphorylation (OXPHOS) may also contribute to disease pathogenesis. Therefore, we examined OXPHOS activity and stability in MCAD-deficient patient fibroblasts that have no detectable MCAD protein. We found a deficit in mitochondrial oxygen consumption, with reduced steady-state levels of OXPHOS complexes I, III and IV, as well as the OXPHOS supercomplex. To examine the mechanisms involved, we generated an MCAD knockout (KO) using human 143B osteosarcoma cells. These cells also exhibited defects in OXPHOS complex function and steady-state levels, as well as disrupted biogenesis of newly-translated OXPHOS subunits. Overall, our findings suggest that the loss of MCAD is associated with a reduction in steady-state OXPHOS complex levels, resulting in secondary defects in OXPHOS function which may contribute to the pathology of MCAD deficiency.

2-Methyl-3-hydroxybutyryl-CoA dehydrogenase (2M3HBD) deficiency (HSD10 disease) is a rare inborn error of metabolism, and30 cases have been reported worldwide. This disorder is typically characterized by progressive neurodegenerative disease from 6 to 18 months of age. Here, we report the first patient with this disorder in Asia, with atypical clinical presentation. A 6-year-old boy, who had been well, presented with severe ketoacidosis following a 5-day history of gastroenteritis. Urinary organic acid analysis showed elevated excretion of 2-methyl-3-hydroxybutyrate and tiglylglycine. He was tentatively diagnosed with β-ketothiolase (T2) deficiency. However, repeated enzyme assays using lymphocytes showed normal T2 activity and no T2 mutation was found. Instead, a hemizygous c.460GA (p.A154T) mutation was identified in the HSD17B10 gene. This mutation was not found in 258 alleles from Japanese subjects (controls). A normal level of the HSD17B10 protein was found by immunoblot analysis but no 2M3HBD enzyme activity was detected in enzyme assays using the patient's fibroblasts. These data confirmed that this patient was affected with HSD10 disease. He has had no neurological regression until now. His fibroblasts showed punctate and fragmented mitochondrial organization by MitoTracker staining and had relatively low respiratory chain complex IV activity to those of other complexes.

We report the first case of Leigh syndrome (LS) with Fukuyama congenital muscular dystrophy (FCMD). A neonate suffered from lactic acidosis and subsequently presented with poor feeding, muscle weakness, hypotonia, cardiopulmonary dysfunction, and hydrocephalus. He died at 17 months. The findings of brain magnetic resonance imaging indicated some specific features of both LS and FCMD, and FCMD gene mutation was detected. Decreased mitochondrial respiratory complex I and II activity was noted. Mitochondrial DNA sequencing showed no pathogenic mutation. A case with complex I+II deficiency has rarely been reported, suggesting a nuclear gene mutation.

Influence of hyperphenylalaninemia on lipoproteins in early life remains unclear.We enrolled 24 phenylalanine hydroxylase (PAH)-deficient children who were classified into a phenylketonuria (PKU) group (n=12) lacking PAH activity and a benign hyperphenylalaninemia (HPA) group (n=12) having partial PAH activity, and their 11 non-affected siblings. We measured serum total-cholesterol, low-density lipoprotein (LDL)-cholesterol, and high-density lipoprotein (HDL)-cholesterol levels together with apolipoproteins for the first year of life, and compared them with those of 30 age-matched healthy controls.The affected groups invariably had lower cholesterol levels than non-affected groups. At birth, HDL-cholesterol decrease was greatest and predominated over the LDL-cholesterol decrease: total cholesterol, 28/36% decrease to the control level in HPA/PKU; HDL-cholesterol, 33/51%; LDL-cholesterol, 20/28%. At 3months, the opposite changes were observed: total cholesterol, 16/28%; HDL-cholesterol, 13/23%; LDL-cholesterol, 16/33%. At 12months, LDL were still significantly lower in both groups (8/18%, p.05 and .001), although HDL was significantly decreased only in the PKU group (15%, p.05). Apolipoprotein A-I/A-II and B changed respectively in accordance with HDL-cholesterol and LDL-cholesterol changes. Despite similar phenylalanine levels, the PKU group invariably had lower cholesterol concentrations than the HPA group had.Irrespective of phenylalanine concentrations, lipoprotein synthesis in PAH-deficient children, particularly in PKU children, was suppressed in early life.

Objective Mitochondrial respiratory chain disorder (MRCD) is an intractable disease of infants with variable clinical symptoms. Our goal was to identify the causative mutations in MRCD patients. Methods The subjects were 90 children diagnosed with MRCD by enzyme assay. We analyzed whole mitochondrial DNA (mtDNA) sequences. A cybrid study was performed in two patients. Whole exome sequencing was performed for one of these two patients whose mtDNA variant was confirmed as non-pathogenic. Results Whole mtDNA sequences identified 29 mtDNA variants in 29 patients (13 were previously reported, the other 13 variants and three deletions were novel). The remaining 61 patients had no pathogenic mutations in their mtDNA. Of the 13 patients harboring unreported mtDNA variants, we excluded seven variants by manual curation. Of the remaining six variants, we selected two Leigh syndrome patients whose mitochondrial enzyme activity was decreased in their fibroblasts and performed a cybrid study. We confirmed that m.14439G>A (MT-ND6) was pathogenic, while m.1356A>G (mitochondrial 12S rRNA) was shown to be a non-pathogenic polymorphism. Exome sequencing and a complementation study of the latter patient identified a novel c.55C>T hemizygous missense mutation in the nuclear-encoded gene NDUFA1. Interpretation Our results demonstrate that it is important to perform whole mtDNA sequencing rather than only typing reported mutations. Cybrid assays are also useful to diagnose the pathogenicity of mtDNA variants, and whole exome sequencing is a powerful tool to diagnose nuclear gene mutations as molecular diagnosis can provide a lead to appropriate genetic counseling.

Mitochondrial disorders have the highest incidence among congenital metabolic diseases, and are thought to occur at a rate of 1 in 5000 births. About 25% of the diseases diagnosed as mitochondrial disorders in the field of pediatrics have mitochondrial DNA abnormalities, while the rest occur due to defects in genes encoded in the nucleus. The most important function of the mitochondria is biosynthesis of ATP. Mitochondrial disorders are nearly synonymous with mitochondrial respiratory chain disorder, as respiratory chain complexes serve a central role in ATP biosynthesis. By next-generation sequencing of the exome, we analyzed 104 patients with mitochondrial respiratory chain disorders. The results of analysis to date were 18 patients with novel variants in genes previously reported to be disease-causing, and 27 patients with mutations in genes suggested to be associated in some way with mitochondria, and it is likely that they are new disease-causing genes in mitochondrial disorders. This article is part of a Special Issue entitled Frontiers of Mitochondrial Research.

Large numbers of genes are responsible for Leigh syndrome (LS), making genetic confirmation of LS difficult. We screened our patients with LS using a limited set of 21 primers encompassing the frequently reported gene for the respiratory chain complexes I (ND1–ND6, and ND4L), IV(SURF1), and V(ATP6) and the pyruvate dehydrogenase E1α-subunit. Of 18 LS patients, we identified mutations in 11 patients, including 7 in mDNA (two with ATP6), 4 in nuclear (three with SURF1). Overall, we identified mutations in 61% of LS patients (11/18 individuals) in this cohort. Sanger sequencing with our limited set of primers allowed us a rapid genetic confirmation of more than half of the LS patients and it appears to be efficient as a primary genetic screening in this cohort.

We present characteristic magnetic resonance imaging (MRI) features in a pediatric female patient with chronic inflammatory demyelinating polyradiculoneuropathy (CIDP). Muscle weakness developed at 8 years old and fluctuated during the clinical course over 7 years. Electrophysiological studies showed a demyelination pattern with moderately delayed nerve conduction velocity, as well as dispersion phenomenon. MRI showed marked changes in thickening of the spinal nerve roots and their peripheral nerves in the lumber and brachial plexuses, as well as in the bilateral trigeminal nerves. It is suggested that these MRI features are characteristic and strongly supportive of the diagnosis of CIDP with a prolonged clinical course.

The patient had hepatomegaly with liver dysfunction at the age of 1 month. Magnetic resonance imaging performed at the age of 1 year showed multiple nodules of varying size in his liver. We were able to examine the mitochondrial respiratory chain function in the liver biopsy samples because all other differential diagnoses for hepatic cirrhosis had been ruled out. Complex I and IV activities were below the normal level (

Plasma mannose is suggested to be largely generated from liver glycogen-oriented glucose-6-phosphate. This study examined plasma mannose in glycogen storage disease type Ia (GSD Ia) lacking conversion of glucose-6-phosphate to glucose in the liver. We initially examined fasting-and postprandial 2 h-plasma mannose and other blood carbohydrates and lipids for seven GSD Ia children receiving dietary interventions using cornstarch and six healthy age-matched children. Next, one-day successive intra-individual parameter changes were examined for six affected and two control children. Although there were no significant differences in fasting-and postprandial 2 h-glucose and insulin levels, the mannose level of the affected group was invariably much higher than that of the control group (p

In the search for sequence variants underlying disease, commonly applied filtering steps usually result in a number of candidate variants that cannot further be narrowed down. In autosomal recessive families, disease usually occurs only in one generation so that genetic linkage analysis is unlikely to help. Because homozygous recessive mutations tend to be inherited together with flanking homozygous variants, we developed a statistical method to detect pathogenic variants in autosomal recessive families: We look for differences in patterns of homozygosity around candidate variants between patients and control individuals and expect that such differences are greater for pathogenic variants than random candidate variants. In six autosomal recessive mitochondrial disease families, in which pathogenic homozygous variants have already been identified, our approach succeeded in prioritizing pathogenic mutations. Our method is applicable to single patients from recessive families with at least a few dozen control individuals from the same population; it is easy to use and is highly effective for detecting causative mutations in autosomal recessive families.

Mitochondrial disorders have the highest incidence among congenital metabolic disorders characterized by biochemical respiratory chain complex deficiencies. It occurs at a rate of 1 in 5,000 births, and has phenotypic and genetic heterogeneity. Mutations in about 1,500 nuclear encoded mitochondrial proteins may cause mitochondrial dysfunction of energy production and mitochondrial disorders. More than 250 genes that cause mitochondrial disorders have been reported to date. However exact genetic diagnosis for patients still remained largely unknown. To reveal this heterogeneity, we performed comprehensive genomic analyses for 142 patients with childhood-onset mitochondrial respiratory chain complex deficiencies. The approach includes whole mtDNA and exome analyses using high-throughput sequencing, and chromosomal aberration analyses using high-density oligonucleotide arrays. We identified 37 novel mutations in known mitochondrial disease genes and 3 mitochondria-related genes (MRPS23, QRSL1, and PNPLA4) as novel causative genes. We also identified 2 genes known to cause monogenic diseases (MECP2 and TNNI3) and 3 chromosomal aberrations (6q24.3-q25.1, 17p12, and 22q11.21) as causes in this cohort. Our approaches enhance the ability to identify pathogenic gene mutations in patients with biochemically defined mitochondrial respiratory chain complex deficiencies in clinical settings. They also underscore clinical and genetic heterogeneity and will improve patient care of this complex disorder., Author Summary Mitochondria play a crucial role in ATP biosynthesis and comprise proteins encoded in both the nuclear and mitochondrial genomes. Although more than 250 mitochondrial disease-causing genes have been reported, the exact genetic causes in patients remain largely unknown. Here, we aimed to provide further insights into the pathogenic mechanisms of mitochondrial disorders. We investigated the genes encoded in the nuclear and mitochondrial genomes using comprehensive genomic analysis in 142 patients with mitochondrial respiratory chain complex deficiencies. We identified 3 novel disease-causing mitochondria-related genes (MRPS23, QRSL1, and PNPLA4) as well as other disease-causing genes and novel pathogenic mutations in known mitochondrial disease-causing genes. All pathogenic mutations in this study are validated by genetic and/or functional evidence. Our findings, including the achievement of firm genetic diagnoses for 49 of 142 patients (34.5%), were higher than the general diagnosis rate of approximately 25% and demonstrated the value of comprehensive genomic analysis. Accordingly, we have shed light on the genetic heterogeneity underlying mitochondrial disorders.

In two Japanese infants with neonatal cholestasis, 3-oxo-Δ(4)-steroid 5β-reductase deficiency was diagnosed based on mutations of the SRD5B1 gene. Unusual bile acids such as elevated 3-oxo-Δ(4) bile acids were detected in their serum and urine by gas chromatography-mass spectrometry. We studied effects of oral chenodeoxycholic acid treatment.SRD5B1 gene analysis used peripheral lymphocyte genomic DNA. Diagnosis and treatment of these two patients were investigated retrospectively and prospectively investigated.With respect to SRD5B1, one patient was heterozygous (R266Q, a novel mutation) while the other was a compound heterozygote (G223E/R261C). Chenodeoxycholic acid treatment was effective in improving liver function and decreasing unusual bile acids such as 7α-hydroxy- and 7α,12α-dihydroxy-3-oxo-4-cholen-24-oic acids in serum and urine.Primary bile acid treatment using chenodeoxycholic acid was effective for these patients treated in early infancy before the late stage of chronic cholestatic liver dysfunction.

We report two patients with Leigh syndrome that showed a combination of facial dysmorphism and MRI imaging indicating an SURF1 deficiency, which was confirmed by sequence analysis. Case 1 is a 3-year-old girl with failure to thrive and developmental delay. She presented with tachypnea at rest and displayed facial dysmorphism including frontal bossing, lateral displacement of inner canthi, esotropia, maxillary hypoplasia, slightly upturned nostril, and hypertrichosis dominant on the forehead and extremities. Case 2 is an 8-year-old boy with respiratory failure. He had been diagnosed as selective complex IV deficiency. Case 2 displayed facial dysmorphism and hypertrichosis. Since both patients displayed characteristic facial dysmorphism and MRI findings, we sequenced the SURF1 gene and identified two heterozygous mutations; c.49+1 G>T and c.752_753del in Case 1, and homozygous c.743 C>A in Case 2. For patients with Leigh syndrome showing these facial dysmorphism and hypertrichosis, sequence analysis of the SURF1 gene may be useful.

Mitochondrial respiratory chain disorders are the most common disorders among inherited metabolic disorders. However, there are few published reports regarding the relationship between mitochondrial respiratory chain disorders and sudden unexpected death in infancy. In the present study, we performed metabolic autopsy in 13 Japanese cases of sudden unexpected death in infancy. We performed fat staining of liver and postmortem acylcarnitine analysis. In addition, we analyzed mitochondrial respiratory chain enzyme activity in frozen organs as well as in postmortem cultured fibroblasts. In heart, 11 cases of complex I activity met the major criteria and one case of complex I activity met the minor criteria. In liver, three cases of complex I activity met the major criteria and four cases of complex I activity met the minor criteria. However, these specimens are susceptible to postmortem changes and, therefore, correct enzyme analysis is hard to be performed. In cultured fibroblasts, only one case of complex I activity met the major criteria and one case of complex I activity met the minor criteria. Cultured fibroblasts are not affected by postmortem changes and, therefore, reflect premortem information more accurately. These cases might not have been identified without postmortem cultured fibroblasts. In conclusion, we detected one probable case and one possible case of mitochondrial respiratory chain disorders among 13 Japanese cases of sudden unexpected death in infancy. Mitochondrial respiratory chain disorders are one of the important inherited metabolic disorders causing sudden unexpected death in infancy. We advocate metabolic autopsy with postmortem cultured fibroblasts in sudden unexpected death in infancy cases.

The synthesis of bile salts from cholesterol is a complex biochemical pathway involving at least 16 enzymes. Most inborn errors of bile acid biosynthesis result in excessive formation of intermediates and/or their metabolites that accumulate in blood and are excreted in part in urine. Early detection is important as oral therapy with bile acids results in improvement. In the past, these intermediates in bile acid biosynthesis have been detected in neonatal blood or urine by screening with FAB-MS followed by detailed characterization using GC-MS. Both methods have proved difficult to automate, and currently most laboratories screen candidate samples using LC-MS/MS. Here, we describe a new, simple and sensitive analytical method for the identification and characterization of 39 conjugated and unconjugated bile acids, including Δ(4)-3-oxo- and Δ(4,6)-3-oxo-bile acids (markers for Δ(4)-3-oxo-steroid 5β-reductase deficiency), using liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). In this procedure a concentrated, desalted urinary sample (diluted with ethanol) is injected directly into the LC-ESI-MS/MS, operated with ESI and in the negative ion mode; quantification is obtained by selected reaction monitoring (SRM). To evaluate the performance of our new method, we compared it to a validated method using GC-MS, in the analysis of urine from two patients with genetically confirmed Δ(4)-3-oxo-steroid 5β-reductase deficiency as well as a third patient with an elevated concentration of abnormal conjugated and unconjugated Δ(4)-3-oxo-bile acids. The Δ(4)-3-oxo-bile acids concentration recovered in three patients with 5β-reductase deficiency were 48.8, 58.9, and 49.4 μmol/mmol creatinine, respectively by LC-ESI-MS/MS.

Background: Succinyl-coenzyme A ligase (SUCL) is a mitochondrial enzyme that catalyses the reversible conversion of succinyl-coenzyme A to succinate. SUCL consists of an α subunit, encoded by SUCLG1, and a β subunit, encoded by either SUCLA2 or SUCLG2. Recently, mutations in SUCLG1 or SUCLA2 have been identified in patients with infantile lactic acidosis showing elevated urinary excretion of methylmalonate, mitochondrial respiratory chain (MRC) deficiency, and mitochondrial DNA depletion. Methods: Case description of a Japanese female patient who manifested a neonatal-onset lactic acidosis with urinary excretion of methylmalonic acid. Enzymatic analyses (MRC enzyme assay and Western blotting) and direct sequencing analysis of SUCLA2 and SUCLG1 were performed. Results: MRC enzyme assay and Western blotting showed that MRC complex I was deficient. SUCLG1 mutation analysis showed that the patient was a compound heterozygote for disease-causing mutations (p.M14T and p.S200F). Conclusion: For patients showing neonatal lactic acidosis and prolonged mild methylmalonic aciduria, MRC activities and mutations of SUCLG1 or SUCLA2 should be screened for.

Costello syndrome (CS) is a congenital disease that is characterized by a distinctive facial appearance, failure to thrive, mental retardation and cardiomyopathy. In 2005, we discovered that heterozygous germline mutations in HRAS caused CS. Several studies have shown that CS-associated HRAS mutations are clustered in codons 12 and 13, and mutations in other codons have also been identified. However, a comprehensive comparison of the substitutions identified in patients with CS has not been conducted. In the current study, we identified four mutations (p.G12S, p.G12A, p.G12C and p.G12D) in 21 patients and analyzed the associated clinical manifestations of CS in these individuals. To examine functional differences among the identified mutations, we characterized a total of nine HRAS mutants, including seven distinct substitutions in codons 12 and 13, p.K117R and p.A146T. The p.A146T mutant demonstrated the weakest Raf-binding activity, and the p.K117R and p.A146T mutants had weaker effects on downstream c-Jun N-terminal kinase signaling than did codon 12 or 13 mutants. We demonstrated that these mutant HRAS proteins induced senescence when overexpressed in human fibroblasts. Oncogene-induced senescence is a cellular reaction that controls cell proliferation in response to oncogenic mutation and it has been considered one of the tumor suppression mechanisms in vivo. Our findings suggest that the HRAS mutations identified in CS are sufficient to cause oncogene-induced senescence and that cellular senescence might therefore contribute to the pathogenesis of CS.

The mechanism underlying the development of osteopenia or osteoporosis in longstanding phenylketonuria (PKU) remains to be clarified. We investigated the details of bone metabolism in 21 female and 13 male classical PKU patients aged 20–35 years. Vitamin D (VD), parathyroid hormone (PTH), bone turnover markers, and daily nutrient intake were examined. The patients had lower daily energy and protein intake than did the age-matched controls (22 women, 14 men), but their respective fat, VD, and calcium intake did not differ. Serum 1,25-dihydroxy VD and 25-hydroxy VD levels in female and male patient groups were significantly higher and lower than those in respective control groups (females, P

Fibrodysplasia ossificans progressiva (FOP) is a rare autosomal dominant congenital disorder characterized by progressive heterotopic ossification in muscle tissues. Constitutively activated mutants of a bone morphogenetic protein (BMP) receptor, ALK2, have been identified in patients with FOP. Recently, a novel ALK2 mutation, L196P, was found in the most benign case of FOP reported thus far. In the present study, we examined the biological activities of ALK2(L196P) in vitro. Over-expression of ALK2(L196P) induced BMP-specific activities, including the suppression of myogenesis, the induction of alkaline phosphatase activity, increased BMP-specific luciferase reporter activity, and increased phosphorylation of Smad1/5 but not Erk1/2 or p38. The activities of ALK2(L196P) were higher than those of ALK2(G356D), another mutant ALK2 allele found in patients with FOP and were equivalent to those of ALK2(R206H), a typical mutation found in patients with FOP. ALK2(L196P) was equally or more resistant to inhibitors in comparison to ALK2(R206H). These findings suggest that ALK2(L196P) is an activated BMP receptor equivalent to ALK2(R206H) and that ALK2(L196P) activity may be suppressed in vivo by a novel molecular mechanism in patients with this mutation.

Background/aims To describe the clinical and biological findings of two Japanese siblings with novel MPV17 gene mutations (c.451insC/c.509C > T) manifesting hepatic mitochondrial DNA depletion syndrome. Methods We observed these brothers and sought to determine the efficacy of treatment targeting respiratory chain complex II for the younger brother. Results A 3-month-old boy had presented with profound liver dysfunction, failure to thrive, and watery diarrhea. Although he was then placed on a carbohydrate-rich diet, his liver function thereafter fluctuated greatly in association with viral infections, and rapidly deteriorated to liver failure. He underwent liver transplantation at 17 months of age but died at 22 months of age. The younger brother, aged 47 months at the time of this writing, presented with liver dysfunction from 8 months of age. His transaminase levels also fluctuated considerably fluctuations in association with viral infections. At 31 months of age, treatment with succinate and ubiquinone was initiated together with a lipid-rich diet using ketone milk. Thereafter, his transaminase levels normalized and never fluctuated, and the liver histology improved. Conclusions These cases suggested that the clinical courses of patients with MPV17 mutations are greatly influenced by viral infections and that dietary and pharmaceutical treatments targeting the mitochondrial respiratory chain complex II may be beneficial in the clinical management of MPV17 mutant patients.

The etiology of secretory diarrhea in early life is often unclear. We report a Japanese boy who survived until 3 years of age, despite intractable diarrhea commencing soon after birth. The fecal sodium content was strikingly high (109 mmol/L [normal range, 27-35 mmol/L]) and the osmotic gap was decreased (15 mOsm/kg), consistent with the findings of congenital sodium diarrhea. We examined the mitochondrial respiratory chain function by blue native polyacrylamide gel electrophoresis (BN-PAGE) in-gel enzyme staining, BN-PAGE western blotting, respiratory chain enzyme activity assay, and immunohistochemistry. Liver respiratory chain complex (Co) I activity was undetectable, while other respiratory chain complex activities were increased (Co II, 138%; Co III, 153%; Co IV, 126% versus respective control activities). Liver BN-PAGE in-gel enzyme staining and western blotting showed an extremely weak complex I band, while immunohistochemistry showed extremely weak staining for the 30-kDa subunit of complex I, but normal staining for the 70-kDa subunit of complex II. The patient was, therefore, diagnosed with complex I deficiency. The overall complex I activity of the jejunum was substantially decreased (63% of the control activity). The immunohistochemistry displayed apparently decreased staining of the 30-kDa complex I subunit, together with a slightly enhanced staining of the 70-kDa complex II subunit in intestinal epithelial cells. These data imply that intestinal epithelial cells are also complex I-deficient in this patient. Complex I deficiency is a novel cause of secretory diarrhea and may act via disrupting the supply of adenosine triphosphate (ATP) needed for the maintenance of ion gradients across membranes.

Carnitine palmitoyltransferase II (CPT II) deficiency is an inherited disorder involving beta-oxidation of long-chain fatty acids. CPT II deficiency is a wide-spectrum disorder that includes a lethal neonatal form, an infantile form, and an adult-onset form. However, the ethnic characteristics and the relationship between genotype and clinical manifestation are not well understood. We investigated three non-consanguineous Japanese patients with CPT II deficiency and examined cell lines from 4 unrelated patients and 50 healthy donors. The CPT 2 gene was typed by direct DNA sequencing of polymerase chain reaction-amplified gene products. Case 1 (infantile form) was heterozygous for a phenylalanine to tyrosine substitution at position 383 (p.F383Y) and a novel valine to leucine substitution at 605 (p.V605L). Cases 2, 4, and 5 (infantile form) and case 3 (adult-onset form) were heterozygous for a single mutation at F383Y. Case 6 (adult-onset form) was compound heterozygous at the CPT 2 locus, with deletion of cytosine and thymine at residue 408, resulting in a stop signal at 420 (p.Y408fsX420), and an arginine to cysteine substitution at position 631 (p.R631C). Case 7 (adult-onset form) was homozygous for the p.F383Y mutation. In conclusion, we identified p.F383Y mutations in six of seven patients with CPT II deficiency and two novel variants of the coding gene: p.Y408fsX420 and p.V605L. These mutations differ from those in Caucasian patients, who commonly harbor p.S113L, p.P50H, and p.Q413fsX449 mutations; therefore, our data and those of other Japanese groups suggest that the p.F383Y mutation is significant in Japanese patients with CPT II deficiency.

Background While the association between scoliosis and impairments in cardiac and respiratory function has been well characterized in clinical practice and research, the potential effect of scoliosis on urinary tract structure and renal function has received little attention. Therefore, the purpose of our study was to evaluate the preoperative clinical characteristics of urinary tract structure and renal function in pediatric patients with idiopathic scoliosis, using a combination of blood tests, urinalyses, and imaging procedures. Methods Preoperative measures of urinary tract structure and renal function were obtained for 16 patients, 13 to 17 years old, scheduled for corrective surgery for idiopathic scoliosis. Preoperative assessment included blood test and urinalyses, combined with structural imaging by ultrasound (US), magnetic resonance (MRI), magnetic resonance urography (MRU), and radioisotope tracing (RI), using technetium-99mTC-mercaptoacetyltriglycine (99mTC-MAG3). Differences in blood and urine tests between patients with and without urinary tract obstruction (UTO) were evaluated for significance using Mann-Whitney U test. Results For all 16 patients, blood tests and MRU results were within normal limits. Evidence of dilatation of the renal pelvis was identified by US imaging in 8 (50.0%) patients. UTO was identified by RI in 6 (37.5%) patients. UTO was associated with elevated levels of β2-microglobulin concentration. A urinary β2-microglobulin concentration above 0.7μg/mg Cr differentiated patients with UTO, from those without UTO, with a sensitivity of 100% and specificity of 70%. Conclusions β2-microglobulin concentration may provide a useful marker to screen for asymptomatic UTO in patients with idiopathic scoliosis. This article is protected by copyright. All rights reserved.

Background A nationwide, multicenter and observational study was retrospectively conducted to evaluate the clinical utility of cepharanthin (CEP) therapy for pediatric patients with chronic immune thrombocytopenia (ITP). Methods Clinical and laboratory data of 46 Japanese patients less than 16 years of age were analyzed, who were diagnosed as having chronic ITP in 14 hospitals during the 2001-2011 decade, and were treated with CEP for more than 12 months. Results The median daily dose of CEP was 1 mg/kg (0.12–2). The platelet counts prior to CEP therapy were 20.5 ×109/L (IQR 8.3-53.0), and were then significantly increased to 58.5 ×109/L (IQR 22.8-115.0) and 69.0 ×109/L (IQR 23.0-134.0) after 12 and 24 months of the treatment course, respectively. No life-threatening bleedings or moderate to severe adverse events were reported. Of 38 patients who received both corticosteroids (CS) and CEP, 17 patients (45%) were weaned from CS, and 15 patients (39%) attained the reduced dose of CS. The period from the start of CEP therapy to the stop of CS was median 413 days (range 49–1734) in patients who were weaned from CS. Conclusions CEP alone or combined therapy with CS was found to be useful for the management of pediatric patients with chronic ITP. This article is protected by copyright. All rights reserved.

Aim The purpose of this study was to evaluate the efficacy and safety of LIVALO tablets (pitavastatin) in Japanese male children with heterozygous familial hypercholesterolemia (FH). Methods A multicenter, randomized, double-blind, parallel study was conducted in 14 male children 10-15 years of age with heterozygous FH. Pitavastatin (1 mg/day or 2 mg/day) was administered orally for 52 weeks.The primary endpoint was the percent change in the LDL-cholesterol (LDL-C) concentrations from baseline to endpoint (repeated measures ANCOVA at Weeks 8 and 12). Secondary endpoints included the percentage of patients who achieved the target LDL-C concentration and percent changes in the levels of lipoprotein and lipid parameters at the visit performed at 52 weeks. Results The percent change in LDL-C from baseline (mean 258 mg/dL for all patients) to the endpoint was -27.3% (95%CI; -34.0, -20.5) and -34.3% (95%CI; -41.0, -27.5) in the patients receiving 1 mg and 2 mg of pitavastatin, respectively. Stable reductions in the total cholesterol (TC), non-HDL cholesterol (non-HDL-C), apolipoprotein B (Apo-B) and LDL-C levels and non-HDL-C/HDL-C and Apo-B/Apo-A1 ratios were observed up to 52 weeks in both groups. One patient in each dose group (14%) reached the treatment target level of 130 mg/dL.Adverse events were observed in seven (100%) patients receiving 1 mg and five (71%) patients receiving 2 mg of pitavastatin, although none were considered related to the study treatment. One patient in the 1 mg group reported a musculoskeletal AE; however, it was attributed to recent excessive exercise. Conclusions Pitavastatin significantly reduced the LDL-C levels and was well tolerated when administered at usual adult doses in 14 male children 10-15 years of age with heterozygous FH. Pitavastatin is a promising therapeutic agent for pediatric dyslipidemia with few safety concerns.

Anderson-Fabry (FD) disease is an inborn error of metabolism caused by a deficiency of α-galactosidase A (GLA), a lysosomal enzyme. Many male FD patients display a classic FD phenotype; however, some female patients have neither reduced leukocyte GLA enzyme activity level nor FD symptoms. Thus, GLA gene analysis is especially important for diagnosing suspected FD in female subjects. In this study, we revealed 4 novel GLA gene mutations in 5 independent families using GLA cDNA analysis and multiplex ligation-dependent probe amplification (MLPA) analysis. These distinct mutations included a large deletion mutation from intron 1 to exon 5 (c.195-471_c.691del5.5k, corresponding to g.8508_g.14069del5.5k), an insertion mutation of splicing enhancer sequence in intron 4 (c.639+329_c.639+330ins113, corresponding to g.12627_g.12628ins113), an insertion mutation of retrotransposon L1 in exon 4 (c.634_c.635, corresponding to g.12293_g.12294), and a non-SNP deep intronic point mutation in intron 3 (c.547+395G>C, corresponding to g.11727G>C). It is difficult to detect these mutations with direct sequencing of only the exonic element. When exonic mutations are not found in the GLA gene from suspected FD patients, GLA cDNA and MLPA analyses should be performed to detect large deletion/insertion and intronic mutations including transcription abnormalities.

Defects in the mitochondrial translation apparatus can impair energy production in affected tissues and organs. Most components of this apparatus are encoded by nuclear genes, including GFM2, which encodes a mitochondrial ribosome recycling factor. A few patients with mutations in some of these genes have been reported to date. Here, we present two female siblings with arthrogryposis multiplex congenita, optic atrophy and severe mental retardation. The younger sister had a progressive cerebellar atrophy and bilateral neuropathological findings in the brainstem. Although her cerebrospinal fluid (CSF) levels of lactate and pyruvate were not increased, brain magnetic resonance spectroscopy showed a lactate peak. Additionally, her CSF lactate/pyruvate and serum beta-hydroxybutyrate/acetoacetate ratios were high, and levels of oxidative phosphorylation in skin fibroblasts were reduced. We therefore diagnosed Leigh syndrome. Genomic investigation confirmed the presence of compound heterozygous GFM2 mutations (c.206+4A>G and c.2029-1G>A) in both siblings, causing aberrant splicing with premature stop codons (p.Gly50Glufs*4 and p.Ala677Leufs*2, respectively). These findings suggest that GFM2 mutations could be causative of a phenotype of Leigh syndrome with arthrogryposis multiplex congenita.

OBJECTIVE: Short-chain enoyl-CoA hydratase (ECHS1) is a multifunctional mitochondrial matrix enzyme that is involved in the oxidation of fatty acids and essential amino acids such as valine. Here, we describe the broad phenotypic spectrum and pathobiochemistry of individuals with autosomal-recessive ECHS1 deficiency. METHODS: Using exome sequencing, we identified ten unrelated individuals carrying compound heterozygous or homozygous mutations in ECHS1. Functional investigations in patient-derived fibroblast cell lines included immunoblotting, enzyme activity measurement, and a palmitate loading assay. RESULTS: Patients showed a heterogeneous phenotype with disease onset in the first year of life and course ranging from neonatal death to survival into adulthood. The most prominent clinical features were encephalopathy (10/10), deafness (9/9), epilepsy (6/9), optic atrophy (6/10), and cardiomyopathy (4/10). Serum lactate was elevated and brain magnetic resonance imaging showed white matter changes or a Leigh-like pattern resembling disorders of mitochondrial energy metabolism. Analysis of patients' fibroblast cell lines (6/10) provided further evidence for the pathogenicity of the respective mutations by showing reduced ECHS1 protein levels and reduced 2-enoyl-CoA hydratase activity. While serum acylcarnitine profiles were largely normal, invitro palmitate loading of patient fibroblasts revealed increased butyrylcarnitine, unmasking the functional defect in mitochondrial β-oxidation of short-chain fatty acids. Urinary excretion of 2-methyl-2,3-dihydroxybutyrate - a potential derivative of acryloyl-CoA in the valine catabolic pathway - was significantly increased, indicating impaired valine oxidation. INTERPRETATION: In conclusion, we define the phenotypic spectrum of a new syndrome caused by ECHS1 deficiency. We speculate that both the β-oxidation defect and the block in l-valine metabolism, with accumulation of toxic methacrylyl-CoA and acryloyl-CoA, contribute to the disorder that may be amenable to metabolic treatment approaches.

Contains fulltext : 152388.pdf (Publisher’s version ) (Open Access) S-adenosylmethionine (SAM) is the predominant methyl group donor and has a large spectrum of target substrates. As such, it is essential for nearly all biological methylation reactions. SAM is synthesized by methionine adenosyltransferase from methionine and ATP in the cytoplasm and subsequently distributed throughout the different cellular compartments, including mitochondria, where methylation is mostly required for nucleic-acid modifications and respiratory-chain function. We report a syndrome in three families affected by reduced intra-mitochondrial methylation caused by recessive mutations in the gene encoding the only known mitochondrial SAM transporter, SLC25A26. Clinical findings ranged from neonatal mortality resulting from respiratory insufficiency and hydrops to childhood acute episodes of cardiopulmonary failure and slowly progressive muscle weakness. We show that SLC25A26 mutations cause various mitochondrial defects, including those affecting RNA stability, protein modification, mitochondrial translation, and the biosynthesis of CoQ10 and lipoic acid.

Biochemical diagnosis of mitochondrial respiratory chain disorders requires caution to avoid misdiagnosis of secondary enzyme defects, and can be improved by the use of conservative diagnostic criteria. Pathogenic mutations causing mitochondrial disorders have now been identified in more than 30 mitochondrial DNA (mtDNA) genes encoding respiratory chain subunits, ribosomal- and t-RNAs. mtDNA mutations appear to be responsible for most adult patients with mitochondrial disease and approximately a quarter of paediatric patients. A family history suggesting maternal inheritance is the exception rather than the norm for children with mtDNA mutations, many of whom have de novo mutations. Prenatal diagnosis and pre-implantation genetic diagnosis can be offered to some women at risk of transmitting a mtDNA mutation, particularly those at lower recurrence risk. Mutations in more than 30 nuclear genes, including those encoding for respiratory chain subunits and assembly factors, have now been shown to cause mitochondrial disorders, creating difficulties in prioritising which genes should be studied by mutation analysis in individual patients. A number of approaches offer promise to guide the choice of candidate genes, including Blue Native-PAGE immunoblotting and microarray expression analysis.

Respiratory chain complex I deficiency is a common cause of Leigh's disease (LD) and can be caused by mutations in genes encoded by either nuclear or mitochondrial DNA (mtDNA). Most pathogenic mtDNA mutations act recessively and only cause disease when present at high mutant loads (typically >90%) in tissues such as muscle and brain. Two mitochondrial DNA mutations in complex I subunit genes, G14459A in ND6, and T12706C in ND5, have been associated with complex I deficiency and LD. We report another ND5 mutation, G13513A, in three unrelated patients with complex I deficiency and LD. The G13513A mutation was present at mutant loads of approximately 50% or less in all tissues tested, including multiple brain regions. The threshold mutant load for causing a complex I defect in cultured cells was approximately 30%. Blue Native polyacrylamide gel electrophoresis showed that fibroblasts with 45% G13513A mutant load had approximately 50% of the normal amount of fully assembled complex I. Fibroblasts with greater than 97% of the ND6 G14459A mutation had only 20% fully assembled complex I, suggesting that both mutations disrupt complex I assembly or turnover. We conclude that the G13513A mutation causes a complex I defect when present at unusually low mutant load and may act dominantly.

We report on a 4-year-old boy who died from influenza encephalopathy. The clinical course and microscopic findings of the autopsied liver were compatible with Reye's syndrome. We examined the mitochondrial respiratory chain function by blue native polyacrylamide gel electrophoresis (BN-PAGE), western blotting, and respiratory chain enzyme activity assays. The activity of liver respiratory chain complex (CO) I was markedly decreased (7.2% of the respective control activity); whereas, the other respiratory chain complex activities were substantially normal (CO II, 57.9%; CO III, 122.3%; CO IV, 161.0%). The activities of CO I-IV in fibroblasts were normal (CO I, 82.0%; CO II, 83.1%; CO III, 72.9%; CO IV, 97.3%). The patient was diagnosed with liver-specific complex I deficiency. This inborn disorder may have contributed to the fatal outcome. We propose that relying only on fibroblast respiratory chain complex activities may lead to the misdiagnosis of liver-specific complex I deficiency.

Carnitine palmitoyltransferase I (CPT I) is one of the carnitine cycle enzymes that plays a role in the transportation of long-fatty acids into the mitochondria for beta-oxidation. Hepatic carnitine palmitoyltransferase I (CPT IA) is one of the isozymes of CPT I, and its deficiency results in an autosomal recessive mitochondrial fatty acid oxidation disorder. To date, 19 patients with CPT IA deficiency and 9 CPT IA mutations have been reported. Recently, six novel mutations in the CPT IA gene were reported in Japanese patients with CPT I deficiencies who were clinically diagnosed as having a Reye-like syndrome. One of these mutations was a missense mutation, 1079AG (E360G). The other was a splicing mutation, 2027-2028+2delAAGT, which caused aberrant splicing transcripts, whereas 1876-2028del, 2027-2028insGTCTCTTCC ACTTCTTCC, and 2026-2028del were three aberrant transcripts that kept reading in-frame. In this report, an expression assay using SV40 transformed fibroblasts was performed to investigate the consequences of these two mutations on enzyme activity and protein levels. Molecular analysis in this study revealed that the two mutations 1079AG and 2028+2delAAGT were the disease-causing mutations.

We report on the clinical, neuropathological, and genetic findings of a Japanese case with myocerebrohepatopathy spectrum (MCHS) disorder due to polymerase gamma (POLG) mutations. A girl manifested poor sucking and failure to thrive since 4 months of age and had frequent vomiting and developmental regression at 5 months of age. She showed significant hypotonia and hepatomegaly. Laboratory tests showed hepatocellular dysfunction and elevated protein and lactate levels in the cerebrospinal fluid. Her liver function and neurologic condition exacerbated, and she died at 8 months of age. At autopsy, fatty degeneration and fibrosis were observed in the liver. Neuropathological examination revealed white matter-predominant spongy changes with Alzheimer type II glia and loss of myelin. Enzyme activities of the respiratory chain complex I, III, and IV relative to citrate synthase in the muscle were normal in the biopsied muscle tissue, but they were reduced in the liver to 0%, 10%, and 14% of normal values, respectively. In the liver, the copy number of mitochondrial DNA compared to nuclear DNA was reduced to 3.3% of normal values as evaluated by quantitative polymerase chain reaction. Genetic analysis revealed compound heterozygous mutations for POLG (I1185T/A957V). This case represents the differential involvement of multiple organs and phenotype-specific distribution of brain lesions in mitochondrial DNA depletion disorders.

International audience; Respiratory chain deficiencies exhibit a wide variety of clinical phenotypes resulting from defective mitochondrial energy production through oxidative phosphorylation. These defects can be caused by either mutations in the mtDNA or mutations in nuclear genes coding for mitochondrial proteins. The underlying pathomechanisms can affect numerous pathways involved in mitochondrial physiology. By whole-exome and candidate gene sequencing, we identified 11 individuals from 9 families carrying compound heterozygous or homozygous mutations in GTPBP3, encoding the mitochondrial GTP-binding protein 3. Affected individuals from eight out of nine families presented with combined respiratory chain complex deficiencies in skeletal muscle. Mutations in GTPBP3 are associated with a severe mitochondrial translation defect, consistent with the predicted function of the protein in catalyzing the formation of 5-taurinomethyluridine (τm(5)U) in the anticodon wobble position of five mitochondrial tRNAs. All case subjects presented with lactic acidosis and nine developed hypertrophic cardiomyopathy. In contrast to individuals with mutations in MTO1, the protein product of which is predicted to participate in the generation of the same modification, most individuals with GTPBP3 mutations developed neurological symptoms and MRI involvement of thalamus, putamen, and brainstem resembling Leigh syndrome. Our study of a mitochondrial translation disorder points toward the importance of posttranscriptional modification of mitochondrial tRNAs for proper mitochondrial function.

Background Valproate-induced Fanconi syndrome is a rare adverse effect of valproate. Severely disabled patients who require tube feeding are reported to be susceptible to valproate-induced Fanconi syndrome. Although most patients with valproate-induced Fanconi syndrome are asymptomatic and detected incidentally with findings such as hypophosphatemia, hypouricemia, increased urinary β2-microglobulin, and generalized hyperaminoaciduria, clinical symptoms such as bone fracture, fever, tachypnea, and edema have been reported. Patient description This 15-year-old, severely disabled, tube-fed, male patient with cytochrome oxidase deficiency had taken valproate for 3 years when he developed fever for 3 weeks. Hypophosphatemia, hypouricemia, hypokalemia, increased urinary β2-microglobulin, and generalized hyperaminoaciduria, as well as hypocarnitinemia, were found, indicating that he had Fanconi syndrome. Valproate was the most likely cause of Fanconi syndrome in this patient. After discontinuation of valproate, the fever resolved immediately, and the laboratory findings normalized. Conclusion Valproate-induced Fanconi syndrome should be considered when individuals taking valproate develop fever of unknown origin.

Recently, de novo mutations in TBL1XR1 were found in two patients with autism spectrum disorders. Here, we report on a Japanese girl presenting with West syndrome, Rett syndrome-like and autistic features. Her initial development was normal until she developed a series of spasms at 5 months of age. Electroencephalogram at 7 months showed a pattern of hypsarrhythmia, which led to a diagnosis of West syndrome. Stereotypic hand movements appeared at 8 months of age, and autistic features such as deficits in communication, hyperactivity and excitability were observed later, at 4 years and 9 months. Whole exome sequencing of the patient and her parents revealed a de novo TBL1XR1 mutation [c.209 G>A (p.Gly70Asp)] occurring at an evolutionarily conserved amino acid in an F-box-like domain. Our report expands the clinical spectrum of TBL1XR1 mutations to West syndrome with Rett-like features, together with autistic features.

Galactokinase (GALK) deficiency is an autosomal recessive disorder characterized by hypergalactosemia and cataract formation. Through mass screening of newborn infants, we identified a novel and prevalent GALK variant (designated here as the "Osaka" variant) associated with an A198V mutation in three infants with mild GALK deficiency. GALK activity and the amount of immunoreactive protein in the mutant were both 20% of normal construct in expression analysis. The K(m) values for galactose and ATP-Mg(2+) in erythrocytes with homozygous A198V were similar to those of the healthy adult control subjects. A population study for A198V revealed prevalences of 4.1% in Japanese and 2.8% in Koreans, lower incidence in Taiwanese and Chinese, no incidence in blacks and whites from the United States, and a significantly high frequency (7.8%; P.023) in Japanese individuals with bilateral cataract. This variant probably originated in Japanese and Korean ancestors and is one of the genetic factors that causes cataract in elderly individuals.

Background Recently we proposed the therapeutic potential of pyruvate therapy for mitochondrial diseases. Leigh syndrome is a progressive neurodegenerative disorder ascribed to either mitochondrial or nuclear DNA mutations. Methods In an attempt to circumvent the mitochondrial dysfunction, we orally applied sodium pyruvate and analyzed its effect on an 11-year-old female with Leigh syndrome due to cytochrome c oxidase deficiency accompanied by cardiomyopathy. The patient was administered sodium pyruvate at a maintenance dose of 0.5 g/kg/day and followed up for 1 year. Results The exercise intolerance was remarkably improved so that she became capable of running. Echocardiography indicated improvements both in the left ventricle ejection fraction and in the fractional shortening. Electrocardiography demonstrated amelioration of the inverted T waves. When the pyruvate administration was interrupted because of a gastrointestinal infection, the serum lactate level became elevated and the serum pyruvate level, decreased, suggesting that the pyruvate administration was effective in decreasing the lactate-to-pyruvate ratio. Conclusions These data indicate that pyruvate therapy was effective in improving exercise intolerance at least in a patient with cytochrome c oxidase deficiency. General significance Administration of sodium pyruvate may prove effective for other patients with cytochrome c oxidase deficiency due to mitochondrial or nuclear DNA mutations.

N-acetyl- l -glutamate synthetase (EC 2.3.1.1) is a regulatory enzyme involved in the control of carbamoyl-phosphate synthesis in the mammalian liver. The enzyme is activated specifically by arginine, and the sensitivity of acetylglutamate synthetase to arginine undergoes marked changes after food ingestion. Since the extent of arginine activation of the synthetase — high for the enzyme from fed mice and low for the enzyme from fasted mice — remained much the same during partial purification, these changes appear to be due to a modification of the enzyme molecule itself. When the enzyme preparation with a low sensitivity to arginine activation, partially purified from the liver of starved mice, was incubated with thiol compounds, the sensitivity increased. When the enzyme preparation with a high sensitivity, obtained from the liver of fed mice, was incubated with disulfide compounds, the sensitivity decreased. Diminution and enhancement of arginine sensitivity of a single enzyme preparation were also achieved by the disulfide and thiol treatments, thereby revealing the reversibility of the process. These results suggest that thiol/disulfide interchange may be involved in the regulation of arginine sensitivity of acetylglutamate synthetase in vivo.