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Kanazawa-shi, Japan

Shu J.,Tianjin Pediatric Research Institute | Lin S.,Tianjin Pediatric Research Institute | Meng Y.,Tianjin Pediatric Research Institute | Zhang C.,MILS International | And 3 more authors.
Chinese Journal of Medical Genetics | Year: 2015

Objective To detect potential mutation in a Chinese family affected with β-ureidopropinoase deficiency. Methods Genomic DNA was extracted from peripheral blood samples. All exons and flanking intron regions of the UPB1 gene were amplified by PCR and detected by direct sequencing. Results A homozygous mutation c. 977G>A was identified in exon 9 of the UPB1 gene in the proband. Both parents of the proband had heterozygous change of the same site. Conclusion The c. 977G> A mutation of the UPBl gene is responsible for the pathogenesis of the disease in the infant. Source


Meijer J.,Metabolic | Zhang C.,MILS International | Wang X.,Capital Medical University | Dobritzsch D.,Uppsala University | Van Kuilenburg A.B.P.,Metabolic
International Journal of Molecular Sciences | Year: 2016

Dihydropyrimidinase (DHP) deficiency is an autosomal recessive disease caused by mutations in the DPYS gene. Patients present with highly elevated levels of dihydrouracil and dihydrothymine in their urine, blood and cerebrospinal fluid. The analysis of the effect of mutations in DPYS on pre-mRNA splicing is hampered by the fact that DHP is primarily expressed in liver and kidney cells. The minigene approach can detect mRNA splicing aberrations using cells that do not express the endogenous mRNA. We have used a minigene-based approach to analyze the effects of a presumptive pre-mRNA splicing mutation in two newly identified Chinese pediatric patients with DHP deficiency. Mutation analysis of DPYS showed that both patients were compound heterozygous for a novel intronic mutation c.1443+5G>A in intron 8 and a previously described missense mutation c.1001A>G (p.Q334R) in exon 6. Wild-type and the mutated minigene constructs, containing exons 7, 8 and 9 of DPYS, yielded different splicing products after expression in HEK293 cells. The c.1443+5G>A mutation resulted in altered pre-mRNA splicing of the DPYS minigene construct with full skipping of exon 8. Analysis of the DHP crystal structure showed that the deletion of exon 8 severely affects folding, stability and homooligomerization of the enzyme as well as disruption of the catalytic site. Thus, the analysis suggests that the c.1443+5G>A mutation results in aberrant splicing of the pre-mRNA encoding DHP, underlying the DHP deficiency in two unrelated Chinese patients. © 2016 by the authors, licensee MDPI, Basel, Switzerland. Source


Meijer J.,Metabolic | Nakajima Y.,Metabolic | Nakajima Y.,Nagoya City University | Zhang C.,MILS International | And 3 more authors.
Nucleosides, Nucleotides and Nucleic Acids | Year: 2013

β-Ureidopropionase is the third enzyme of the pyrimidine degradation pathway and it catalyzes the conversion of N-carbamyl-β-alanine and N-carbamyl-β-aminoisobutyric acid to β-alanine and β-aminoisobutyric acid, respectively, and ammonia and CO2. To date, only 16 genetically confirmed patients with a complete ß-ureidopropionase deficiency have been reported. Here, we report the clinical, biochemical, and molecular analysis of a newly identified patient with β-ureidopropionase deficiency. Mutation analysis of the UPB1 gene showed that the patient was compound heterozygous for a novel synonymous mutation c.93C >T (p.Gly31Gly) in exon 1 and a previously described missense mutation c.977G >A (p.Arg326Gln) in exon 9. The in silico predicted effect of the synonymous mutation p.Gly31Gly on pre-mRNA splicing was investigated using a minigene approach. Wild-type and the mutated minigene constructs, containing the entire exon 1, intron 1, and exon 2 of UPB1, yielded different splicing products after expression in HEK293 cells. The c.93C >T (p.Gly31Gly) mutation resulted in altered pre-mRNA splicing of the UPB1 minigene construct and a deletion of the last 13 nucleotides of exon 1. This deletion (r.92-104delGCAAGGAACTCAG) results in a frame shift and the generation of a premature stop codon (p.Lys32SerfsX31). Using a minigene approach, we have thus identified the first synonymous mutation in the UPB1 gene, creating a cryptic splice-donor site affecting pre-mRNA splicing. © 2013 Copyright Taylor and Francis Group, LLC. Source


Wen B.,Shandong University | Dai T.,Shandong University | Li W.,Shandong University | Zhao Y.,Shandong University | And 6 more authors.
Journal of Neurology, Neurosurgery and Psychiatry | Year: 2010

Background: Lipid-storage myopathy (LSM), defined by triglyceride accumulation in muscle fibres, is a heterogeneous group of lipid metabolic disorders predominantly affecting skeletal muscle. In the past 15 years, more than 200 cases of LSM have been reported in the Chinese literature, but the accurate pathogenic mechanisms are still unknown. Objective: In order to gain more insight into the metabolic and genetic dysfunctions of LSM, the authors described a group of Chinese patients with LSM who were very responsive to isolated riboflavin treatment (riboflavin responsive LSM, RR-LSM). Methods: Nineteen consecutive LSM patients collected during 1995e2007 in our Neuromuscular Laboratory who were dramatically responsive to riboflavin and presented with proximal muscle weakness, exercise intolerance and elevated serum CK but without episodic encephalopathy were subjected to pathological, biochemical and molecular analysis. Results: On the basis of muscle pathology, all 19 patients were diagnosed as LSM. Seventeen patients were suspected of having multiple acyl-coenzyme A dehydrogenase deficiency (MADD) according to blood acylcarnitine profiles and urine organic acid analysis. Genetic analysis identified 19 novel mutations in ETFDH gene in 18 patients, among which one was homozygote, 16 were compound heterozygotes, and one was a single heterozygote. No pathogenic mutation was detected in ETFA or ETFB genes. Western blot analysis showed there was no significant decrease in ETF:QO expression except for one patient. Conclusions: The research findings suggest that the majority of Chinese patients with RR-LSM are caused by a mild type of MADD with unique myopathy which is due to ETFDH gene mutation. Source


Moriyama M.,Osaka Prefecture University | Fujimoto Y.,Kumamoto University | Fujimoto Y.,Tokushima Bunri University | Rikimaru S.,Tokushima Bunri University | And 23 more authors.
Biochimica et Biophysica Acta - Molecular Basis of Disease | Year: 2015

The mitochondrial aspartate-glutamate carrier isoform 2 (citrin) and mitochondrial glycerol-3-phosphate dehydrogenase (mGPD) double-knockout mouse has been a useful model of human citrin deficiency. One of the most prominent findings has been markedly increased hepatic glycerol 3-phosphate (G3P) following oral administration of a sucrose solution. We aimed to investigate whether this change is detectable outside of the liver, and to explore the mechanism underlying the increased hepatic G3P in these mice. We measured G3P and its metabolite glycerol in plasma and urine of the mice under various conditions. Glycerol synthesis from fructose was also studied using the liver perfusion system. The citrin/mGPD double-knockout mice showed increased urine G3P and glycerol under normal, fed conditions. We also found increased plasma glycerol under fasted conditions, while oral administration of different carbohydrates or ethanol led to substantially increased plasma glycerol. Fructose infusion to the perfused liver of the double-knockout mice augmented hepatic glycerol synthesis, and was accompanied by a concomitant increase in the lactate/pyruvate (L/P) ratio. Co-infusion of either pyruvate or phenazine methosulfate, a cytosolic oxidant, with fructose corrected the high L/P ratio, leading to reduced glycerol synthesis. Overall, these findings suggest that hepatic glycerol synthesis is cytosolic NADH/NAD+ ratio-dependent and reveal a likely regulatory mechanism for hepatic glycerol synthesis following a high carbohydrate load in citrin-deficient patients. Therefore, urine G3P and glycerol may represent potential diagnostic markers for human citrin deficiency. © 2015 Elsevier B.V.. Source

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