Segawa disease, hereditary progressive dystonia with marked diurnal fluctuations or defined dopa-responsive dystonia has age-dependent clinical courses, which are characterized with marked progression in the first one and half decades, its subsiding in the third decade and almost stationary courses after the fourth decade. Also, it has characteristic diurnally fluctuating symptoms, aggravated towards the evening and alleviated after sleep. This autosomally dominantly inherited dystonia is caused by abnormalities of the gene of GTP cyclohydrolase I. The heterozygotic gene's abnormality induces partial decrement of tetrahydrobiopterin and affects synthesis of tyrosine hydroxylase(TH) rather selectively. The reduction of TH induces decrement of dopamine and disfacilitates the D1 receptor-striatal direct pathway. The pathognomonic finding in biochemical examination is the decrease of neopterin in the cerebrospinal fluid(CSF). Levodopa, by replacing dopamine contents at the terminal, alleviates motor symptoms completely and the effects sustain without any side effects. We experienced a girl diagnosed as Segawa disease with typical clinical courses and a decrease of neopterin in the CSF.
Dopamine ; Dystonia* ; Female ; Genetic Diseases, Inborn ; GTP Cyclohydrolase ; Humans ; Levodopa ; Neopterin ; Tyrosine

Dopa-responsive dystonia (DRD) is a clinical syndrome characterized by childhood-onset dystonia and a dramatic response to relatively low doses of levodopa. However, patients with DRD can be misdiagnosed as cerebral palsy or spastic diplegia due to phenotypic variation. Here we report a young woman with DRD who were severely disabled and misdiagnosed as cerebral palsy for over 10 yr. A small dose of levodopa restored wheelchair-bound state to normality. However, thoracolumbar scoliosis has remained as a sequel due to late detection of DRD. Genetic analysis by using PCR-direct sequencing revealed a novel initiation codon mutation (c.1A>T; p.Met1Leu) in GTP cyclohydrolase 1 (GCH1) gene. Although it is known that DRD can be misdiagnosed as cerebral palsy, this case reinforces the importance of differential diagnosis of DRD from cerebral palsy.
Adult ; Cerebral Palsy/diagnosis ; Codon, Initiator ; Diagnosis, Differential ; Dystonic Disorders/*diagnosis/drug therapy/*genetics ; Female ; GTP Cyclohydrolase/*genetics ; Humans ; Levodopa/therapeutic use ; Mutation ; Sequence Analysis, DNA

BACKGROUND: Various vectors have been developed and tried for the delivery of tyrosine hydroxylase (TH) in order to supplement dopamine, which is severely deficient in Parkinson's disease, however, none of the protocols tried have yielded fruitful results that can be applied directly to humans. One of the problems revealed from previous trials was a short duration of expression of the delivered gene, that is, tyrosine hydroxylase. METHODS: To extend the stability and to improve the enzymatic characteristics of the protein, part of the regulatory domain was deleted via PCR technique. The cDNA for regulatory domain-deleted THs (dTH) were sub-cloned into a retroviral vector and the resulting recom-binant retrovirus was used to infect NIH-3T3. After selection, expression levels of TH were determined by Western blot analysis and the enzymatic characteristics were examined. RESULTS: The deletion increased steady state expression level of TH protein by 7-fold for d19TH (TH with amino acids #2-19 are deleted) and 3-fold for d31TH (TH with amino acids #2-31 are deleted. The elevated expression level of d19TH is likely due to the enhanced stability of the protein as determined by a treatment of cycloheximide. The activity of d19TH was also increased approximately by 3-fold but no increase of the L-dopa production was observed. However, the production of L-dopa was dramatically increased when GTP cyclohydrolase I (GTPCH I) was co-transfected suggesting that the activity of d19TH is dependent on the presence of cofactor. d19TH seem to be free of feedback inhibition at low concentration of dopamine (10 nM~1 nM) but more sensitive to the inhibition at high concentration of dopamine (10 mM). CONCLUSIONS: The deletion of 18 amino acids on the regulatory domain increases the stability of the protein, reduces the activity, and frees it from the feedback inhibi-tion by the end product.
Amino Acids ; Blotting, Western ; Cycloheximide ; DNA, Complementary ; Dopamine ; Fruit ; GTP Cyclohydrolase ; Humans ; Levodopa ; Parkinson Disease ; Polymerase Chain Reaction ; Retroviridae ; Staphylococcal Protein A ; Tyrosine 3-Monooxygenase* ; Tyrosine* ; Zidovudine

Tetrahydrobiopterin (BH4) is an essential cofactor for all three nitric oxide synthase (NOS isoforms), which plays an important role in vascular diseases. GTP cyclohydrolase 1 (GCH 1) is the first-step and rate-limiting enzyme for BH4 biosynthesis in its de novo pathway. Common GCH1 gene variant C+243T in the 3'-untranslated region predicts NO excretion. The present study examined the predictive role of GCH 1 gene 3'-UTR C+243T variant in the long-term outcome of ischemic stroke. A total of 142 patients with first-onset ischemic stroke were recruited and detected for genotype of GCH1 3'-UTR C+243T by a TaqMan SNP Genotyping assay. Subsequent vascular events and death were determined over a 5-year follow-up period. The frequency of GCH1 3'-UTR +243 C/T or T/T genotype was significantly increased in patients with endpoint events as compared with those without events (74% vs 57.8%, P=0.06). Cox regression survival analysis indicated that an increased probability of death or new vascular events was found in patients with GCH1 3'-UTR +243 C/T or T/T genotype compared with those with GCH1 3'-UTR C/C genotype (40.6% vs 25.5%), GCH1 3'-UTR +243 C/T or T/T genotype relative to GCH1 3'-UTR C/C genotype was associated with the increased risk of death or vascular events even after adjustment for other risk factors (OR=2.171, 95% CI: 1.066-4.424, P=0.033). It was concluded that GCH1 3'-UTR C+243T variant was an independent predictor of worsening long-term outcomes in patients with first-onset ischemic stroke.
3' Untranslated Regions ; genetics ; Aged ; Brain Ischemia ; genetics ; Female ; GTP Cyclohydrolase ; genetics ; Humans ; Male ; Middle Aged ; Nitric Oxide ; metabolism ; Prognosis ; Risk Factors ; Stroke ; diagnosis ; genetics

OBJECTIVETo identify potential mutation of the GCH1 gene in a Chinese family affected with dopa-responsive dystonia.

METHODSGenomic DNA of patients was extracted from peripheral blood samples. The 6 exons of the GCH1 gene and at least 100 bp of flanking intronic sequences were amplified with PCR. Potential mutations were screened by direct sequencing. Identified mutation was verified with denaturing high performance liquid chromatography (DHPLC) in 100 healthy controls.

RESULTSAll patients were found to be heterozygous for a novel c.597delT (p.Ala200LeufsX5) deletion in the exon 5 of the GCH1 gene. The deletion of T has resulted in formation of a shorter (203 amino acids) truncated non-functional guanosine triphosphate cyclohydrolase I. The same mutation was not found in the 100 controls.

CONCLUSIONA novel GCH1 gene frameshifing mutation probably underlies the dopa-responsive dystonia in this Chinese family.

Adolescent ; Adult ; Base Sequence ; Child ; Dystonic Disorders ; enzymology ; genetics ; Exons ; Female ; Frameshift Mutation ; GTP Cyclohydrolase ; genetics ; Humans ; Male ; Middle Aged ; Molecular Sequence Data ; Pedigree ; Young Adult

BACKGROUND: Parkinson's Disease (PD) is a progressive neurodegenerative disorder characterized by resting tremor, rigidity, and bradykinesia. L-3,4-dihydroxyphenylalanine (L-dopa) has been used for over last 3 decades to treat this disorder, however, its usage is limited due to the reducing effectiveness on time and severe side effects. The best strategy for treating this disorder without serious side effects would be to keep a constant level of dopamine in the brain. This could be achieved by gene or cell therapy using gene(S) involved in dopamine biosynthesis or cells from other individual. For Parkinson's gene therapy, however, there still are controversies on which gene In what combination will yield the best result. In this report, we propose a biochemical background for using GTP cyc]ohydrolase I (GTPCH I) in addition to TH for higher and/or more stable expression of TH. METHODS: TH and GTPCH I cDNA were subcloned into retroviral vectos and resulting recombinant retrovirus packaged in BOSC 23 cells were used to infect NIH-3T3. Confirming successful infections by westers blot analysls, the new cell lines were used to examine steady state TH expression level and TH activity. Furthermore, the effect of ectopic expression of BH4 to the proliferation of these cells were studied. RESULTS: NIH-3T3 cells expressing both TH and GTPCH I showed approximately 10 fold higher expression of TH protein than the cells expressing TH alone. The activity of KNTH2GC6 was approximately 4-6 fold higher than that of striatal tissue and 60 fold higher than KNTH2. Furthermore, growth rate of KNTH2GC6 was strikingly reduced by inhibiting the biosynthesis of BH4. CONCLUSIONS: We showed that the use of GTPCH I in addition to TH not only increased the stability and/or expression of TH protein but also the activity of the enzyme. These improved characteristics of TH protein are very likely due to the expression of BH4 and should be very seriously considered for Parkinson's gene therapy.
Brain ; Cell Line ; Cell- and Tissue-Based Therapy ; DNA, Complementary ; Dopamine ; Genetic Therapy ; GTP Cyclohydrolase* ; Guanosine Triphosphate* ; Hypokinesia ; Levodopa ; Neurodegenerative Diseases ; NIH 3T3 Cells ; Parkinson Disease ; Retroviridae ; Tremor ; Tyrosine 3-Monooxygenase* ; Tyrosine* ; Zidovudine

Recently it was shown that single nucleotide polymorphisms (SNPs) can explain individual variation because of the small changes of the gene expression level and that the 50% decreased expression of an allele might even lead to predisposition to cancer. In this study, we found that a decreased expression of an allele might cause predisposition to genetic disease. Dopa responsive dystonia (DRD) is a dominant disease caused by mutations in GCH1 gene. The sequence analysis of the GCH1 in a patient with typical DRD symptoms revealed two novel missense mutations instead of a single dominant mutation. Family members with either of the mutations did not have any symptoms of DRD. The expression level of a R198W mutant allele decreased to about 50%, suggesting that modestly decreased expression caused by an SNP should lead to predisposition of a genetic disease in susceptible individuals.
Child ; Clubfoot/genetics ; Dopamine/deficiency ; Dystonic Disorders/drug therapy/enzymology/*genetics/physiopathology ; GTP Cyclohydrolase/*genetics/metabolism ; Genes, Recessive ; *Genetic Predisposition to Disease ; Humans ; Levodopa/administration & dosage ; Male ; Mutation, Missense ; Pedigree ; Polymorphism, Genetic

OBJECTIVETo detect mutations of guanosine triphosphate cyclohydrolase I (GCH1) gene in Chinese patients with dopa responsive dystonia (DRD).

METHODSSix sporadic patients with DRD were examined. GCH1 gene mutations were detected using polymerase chain reaction (PCR), DNA sequence analysis and restriction enzyme digestion analysis. One hundred normal people were detected using PCR and restriction enzyme digestion analysis.

RESULTSA new point mutation, 151(G-->A) in exon one was found in a patient. It lead to substitution of a methionine for isoleucine at amino acid 1(M1I). This mutation was not found in normal control people.

CONCLUSIONThe authors report a new heterozygotic point mutation 151(G-->A) in GCH1 gene. There are GCH1 gene mutations in Chinese sporadic patients with DRD.

Asian Continental Ancestry Group ; genetics ; Case-Control Studies ; DNA ; genetics ; DNA Mutational Analysis ; Dihydroxyphenylalanine ; therapeutic use ; Dystonia ; drug therapy ; genetics ; Exons ; genetics ; Female ; GTP Cyclohydrolase ; genetics ; Humans ; Male ; Point Mutation ; genetics ; Polymerase Chain Reaction

OBJECTIVETo study the incidence of various enzyme deficiency in tetrahydrobiopterin (BH4) metabolism and the related gene mutation among the patients with motor disturbance and mental retardation.

METHODSOne hundred patients with unknown motor disturbance and mental retardation were referred to this study. All patients were performed by phenylalanine (Phe) and BH4 loading test, urinary pterin analysis and dihydropteridine reductase (DHPR) activity. Some patients received the dopa treatment for diagnosis of dopa-responsive dystonia (DRD). The analysis of GTP cyclohydrolase 1 gene (GCH1) mutation for DRD patients and the analysis of 6-pyruvoyl tetrahydropterin synthase (PTS) gene mutations for PTS deficient patients were done under the consent from their parents.

RESULTSSeventy of 100 patients had normal basic blood Phe levels, six (6%) patients were diagnosed as DRD. Thirty patients had hyperphenylalaninemia (HPA), eight (8%) were diagnosed as PTS deficiency and 22(22%) were diagnosed as phenylalanine hydroxylase (PAH) deficiency. All patients had normal DHPR activity. The mutation IVS5+3insT of GCH1 was found in 2 patients with DRD. Seven kinds of PTS mutations were found in 8 patients with PTS deficiency, and 75% of the mutations were 259C-->T,286G-->A and 155A-->G.

CONCLUSIONSome patients with unknown motor disturbance and mental retardation may suffer from BH4 metabolism related diseases. Theses patients are necessary to be screened for such kind of diseases in order to confirm the diagnosis.

Adolescent ; Biopterin ; analogs & derivatives ; metabolism ; Child ; Child, Preschool ; Dihydropteridine Reductase ; genetics ; metabolism ; Dystonia ; genetics ; metabolism ; Female ; GTP Cyclohydrolase ; genetics ; metabolism ; Humans ; Infant ; Intellectual Disability ; genetics ; metabolism ; Male ; Mutation ; Phenylalanine Hydroxylase ; genetics ; metabolism ; Phosphorus-Oxygen Lyases ; genetics ; metabolism

OBJECTIVETo explore genetic mutations and clinical features of a pedigree affected with dopa-responsive dystonia.

METHODSPCR and Sanger sequencing were applied to detect mutations of the GCH1 gene among 7 members from the pedigree.

RESULTSThe family was detected to have a known heterozygous mutation of the GCH1 gene (c.550C>T). For the 7 members from the pedigree, the age of onset has ranged from 13 to 60 years. The mother of the proband has carried the same mutation but was still healthy at 80. The symptoms of the other three patients were in slow progression, with diurnal fluctuation which can be improved with sleeping, dystonias of lower limbs, and tremor of both hands. Treatment with small dose of levodopa has resulted in significant improvement of clinical symptoms. By database analysis, the c.550C>T mutation was predicted as probably pathological.

CONCLUSIONThe c.550C>T mutation probably underlies the disease in this pedigree. The clinical phenotypes of family members may be variable for their ages of onset. Some may even be symptom free.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Base Sequence ; DNA Mutational Analysis ; Dystonic Disorders ; enzymology ; genetics ; Female ; GTP Cyclohydrolase ; genetics ; Heterozygote ; Humans ; Male ; Middle Aged ; Molecular Sequence Data ; Mutation ; Pedigree ; Phenotype ; Young Adult