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1.
Genes (Basel) ; 12(12)2021 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-34946879

RESUMO

Due to newborn screening for X-linked adrenoleukodystrophy (ALD), and the use of exome sequencing in clinical practice, the detection of variants of unknown significance (VUS) in the ABCD1 gene is increasing. In these cases, functional tests in fibroblasts may help to classify a variant as (likely) benign or pathogenic. We sought to establish reference ranges for these tests in ALD patients and control subjects with the aim of helping to determine the pathogenicity of VUS in ABCD1. Fibroblasts from 36 male patients with confirmed ALD, 26 healthy control subjects and 17 individuals without a family history of ALD, all with an uncertain clinical diagnosis and a VUS identified in ABCD1, were included. We performed a combination of tests: (i) a test for very-long-chain fatty acids (VLCFA) levels, (ii) a D3-C22:0 loading test to study the VLCFA metabolism and (iii) immunoblotting for ALD protein. All ALD patient fibroblasts had elevated VLCFA levels and a reduced peroxisomal ß-oxidation capacity (as measured by the D3-C16:0/D3-C22:0 ratio in the D3-C22:0 loading test) compared to the control subjects. Of the VUS cases, the VLCFA metabolism was not significantly impaired (most test results were within the reference range) in 6/17, the VLCFA metabolism was significantly impaired (most test results were within/near the ALD range) in 9/17 and a definite conclusion could not be drawn in 2/17 of the cases. Biochemical studies in fibroblasts provided clearly defined reference and disease ranges for the VLCFA metabolism. In 15/17 (88%) VUS we were able to classify the variant as being likely benign or pathogenic. This is of great clinical importance as new variants will be detected.

2.
Hum Genomics ; 14(1): 28, 2020 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-32907636

RESUMO

BACKGROUND: Mitochondrial diseases (MDs) are a group of clinically and genetically heterogeneous disorders characterized by defects in oxidative phosphorylation. Since clinical phenotypes of MDs may be non-specific, genetic diagnosis is crucial for guiding disease management. In the current study, whole-exome sequencing (WES) was performed for our paediatric-onset MD cohort of a Southern Chinese origin, with the aim of identifying key disease-causing variants in the Chinese patients with MDs. METHODS: We recruited Chinese patients who had paediatric-onset MDs and a minimum mitochondrial disease criteria (MDC) score of 3. Patients with positive target gene or mitochondrial DNA sequencing results were excluded. WES was performed, variants with population frequency ≤ 1% were analysed for pathogenicity on the basis of the American College of Medical Genetics and Genomics guidelines. RESULTS: Sixty-six patients with pre-biopsy MDC scores of 3-8 were recruited. The overall diagnostic yield was 35% (23/66). Eleven patients (17%) were found to have mutations in MD-related genes, with COQ4 having the highest mutation rate owing to the Chinese-specific founder mutation (4/66, 6%). Twelve patients (12/66, 18%) had mutations in non-MD-related genes: ATP1A3 (n = 3, two were siblings), ALDH5A1, ARX, FA2H, KCNT1, LDHD, NEFL, NKX2-2, TBCK, and WAC. CONCLUSIONS: We confirmed that the COQ4:c.370G>A, p.(Gly124Ser) variant, was a founder mutation among the Southern Chinese population. Screening for this mutation should therefore be considered while diagnosing Chinese patients suspected to have MDs. Furthermore, WES has proven to be useful in detecting variants in patients suspected to have MDs because it helps to obtain an unbiased and precise genetic diagnosis for these diseases, which are genetically heterogeneous.


Assuntos
Predisposição Genética para Doença/genética , Doenças Mitocondriais/genética , Mutação , Sequenciamento Completo do Exoma/métodos , /genética , Criança , China , Estudos de Coortes , Feminino , GTP Fosfo-Hidrolases/genética , Predisposição Genética para Doença/etnologia , Proteínas de Homeodomínio , Humanos , Masculino , Doenças Mitocondriais/diagnóstico , Doenças Mitocondriais/etnologia , Proteínas Mitocondriais/genética , Oxigenases de Função Mista/genética , Proteínas Nucleares , ATPase Trocadora de Sódio-Potássio/genética , Fatores de Transcrição
3.
Pathology ; 52(4): 439-446, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32276787

RESUMO

The objectives were to determine the reference intervals of spot urine copper excretion indexes in pre-school children and to evaluate their utility in screening for Wilson disease (WD). With spot urine collected from a control sample of preschool children (aged 3-7 years, n=153), the reference intervals of spot urine copper excretion indexes and their biological variation were defined. In order to investigate their utility performance in screening for WD in this age group, multiple spot urine samples from six WD patients who were diagnosed at presymptomatic stage were also analysed and compared. Cut-off values useful for detection of WD were defined by receiver operator curve (ROC) analysis. Biological (inter-individual) variation of spot urine copper indexes expressed as coefficient of variation (CVg) were around 60% at this age group, which was moderate and similar to other clinically useful urine tests, such as urine albumin excretion ratio. Spot urine copper excretion strongly correlated with both urine creatinine and osmolality. Linear regression against both creatinine and osmolality showed that ∼94% of data points in healthy preschool children fell within the prediction interval, suggesting that both were useful normalisation factors. ROC showed that copper to osmolality ratio was the best index with an area under curve (AUC) greater than 0.98. Cut-off values of 0.5 µmol/L, 0.1 µmol/mmol and 0.00085 µmol/mOsmol (32 µg/L, 56 µg/g creatinine and 0.054 µg/mOsmol, respectively, in conventional units) for spot urine copper concentration, copper to creatinine ratio and copper to osmolality ratio, respectively, have potential application in the differentiation of WD patients. Based on the data, a new WD screening strategy targeting preschool children is proposed. Application of a bivariate screening strategy using spot urine copper concentration and urine osmolality may be useful in a population-wide screening program for WD among preschool children.


Assuntos
Cobre/urina , Degeneração Hepatolenticular/diagnóstico , Degeneração Hepatolenticular/urina , Urinálise/normas , Variação Biológica Individual , Criança , Pré-Escolar , Creatinina/urina , Feminino , Humanos , Masculino , Programas de Rastreamento/normas , Valores de Referência
4.
NPJ Genom Med ; 4: 18, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31396399

RESUMO

Primary coenzyme Q10 deficiency-7 (COQ10D7) is a rare mitochondrial disease caused by biallelic mutations in COQ4. Here we report the largest cohort of COQ10D7 to date, with 11 southern Chinese patients confirmed with biallelic COQ4 mutations. Five of them have the classical neonatal-onset encephalo-cardiomyopathy, while the others have infantile onset with more heterogeneous clinical presentations. We also identify a founder mutation COQ4 (NM_016035.5): c.370G>A, p.(Gly124Ser) for COQ10D7, suggesting a higher chance of occurrence in the southern Chinese. This study helps improve understanding of the clinical spectrum of this disorder.

5.
Am J Hum Genet ; 105(2): 302-316, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31256877

RESUMO

Members of a paralogous gene family in which variation in one gene is known to cause disease are eight times more likely to also be associated with human disease. Recent studies have elucidated DHX30 and DDX3X as genes for which pathogenic variant alleles are involved in neurodevelopmental disorders. We hypothesized that variants in paralogous genes encoding members of the DExD/H-box RNA helicase superfamily might also underlie developmental delay and/or intellectual disability (DD and/or ID) disease phenotypes. Here we describe 15 unrelated individuals who have DD and/or ID, central nervous system (CNS) dysfunction, vertebral anomalies, and dysmorphic features and were found to have probably damaging variants in DExD/H-box RNA helicase genes. In addition, these individuals exhibit a variety of other tissue and organ system involvement including ocular, outer ear, hearing, cardiac, and kidney tissues. Five individuals with homozygous (one), compound-heterozygous (two), or de novo (two) missense variants in DHX37 were identified by exome sequencing. We identified ten total individuals with missense variants in three other DDX/DHX paralogs: DHX16 (four individuals), DDX54 (three individuals), and DHX34 (three individuals). Most identified variants are rare, predicted to be damaging, and occur at conserved amino acid residues. Taken together, these 15 individuals implicate the DExD/H-box helicases in both dominantly and recessively inherited neurodevelopmental phenotypes and highlight the potential for more than one disease mechanism underlying these disorders.


Assuntos
RNA Helicases DEAD-box/genética , Mutação de Sentido Incorreto , Proteínas de Neoplasias/genética , Transtornos do Neurodesenvolvimento/genética , Transtornos do Neurodesenvolvimento/patologia , RNA Helicases/genética , Feminino , Estudos de Associação Genética , Humanos , Lactente , Recém-Nascido , Masculino , Linhagem , Sequenciamento Completo do Exoma
6.
EMBO Mol Med ; 9(10): 1398-1414, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28729482

RESUMO

Maintenance of genome integrity requires the functional interplay between Fanconi anemia (FA) and homologous recombination (HR) repair pathways. Endogenous acetaldehyde, a product of cellular metabolism, is a potent source of DNA damage, particularly toxic to cells and mice lacking the FA protein FANCD2. Here, we investigate whether HR-compromised cells are sensitive to acetaldehyde, similarly to FANCD2-deficient cells. We demonstrate that inactivation of HR factors BRCA1, BRCA2, or RAD51 hypersensitizes cells to acetaldehyde treatment, in spite of the FA pathway being functional. Aldehyde dehydrogenases (ALDHs) play key roles in endogenous acetaldehyde detoxification, and their chemical inhibition leads to cellular acetaldehyde accumulation. We find that disulfiram (Antabuse), an ALDH2 inhibitor in widespread clinical use for the treatment of alcoholism, selectively eliminates BRCA1/2-deficient cells. Consistently, Aldh2 gene inactivation suppresses proliferation of HR-deficient mouse embryonic fibroblasts (MEFs) and human fibroblasts. Hypersensitivity of cells lacking BRCA2 to acetaldehyde stems from accumulation of toxic replication-associated DNA damage, leading to checkpoint activation, G2/M arrest, and cell death. Acetaldehyde-arrested replication forks require BRCA2 and FANCD2 for protection against MRE11-dependent degradation. Importantly, acetaldehyde specifically inhibits in vivo the growth of BRCA1/2-deficient tumors and ex vivo in patient-derived tumor xenograft cells (PDTCs), including those that are resistant to poly (ADP-ribose) polymerase (PARP) inhibitors. The work presented here therefore identifies acetaldehyde metabolism as a potential therapeutic target for the selective elimination of BRCA1/2-deficient cells and tumors.


Assuntos
Acetaldeído/metabolismo , Proteína BRCA1/metabolismo , Proteína BRCA2/metabolismo , Rad51 Recombinase/metabolismo , Aldeído-Desidrogenase Mitocondrial/genética , Aldeído-Desidrogenase Mitocondrial/metabolismo , Animais , Proteína BRCA1/genética , Proteína BRCA2/genética , Linhagem Celular Tumoral , Dano ao DNA , Anemia de Fanconi/genética , Proteína do Grupo de Complementação D2 da Anemia de Fanconi/genética , Proteína do Grupo de Complementação D2 da Anemia de Fanconi/metabolismo , Fibroblastos , Recombinação Homóloga , Humanos , Camundongos , Camundongos Nus , Rad51 Recombinase/genética , Ensaios Antitumorais Modelo de Xenoenxerto
7.
Pathology ; 46(5): 375-82, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24992243

RESUMO

Inherited metabolic diseases (IMDs) are a large group of rare genetic diseases. The spectrum and incidences of IMDs differ among populations, which has been well characterised in Caucasians but much less so in Chinese. In a setting of a University Hospital Metabolic Clinic in Hong Kong, over 100 patients with IMDs have been seen during a period of 13 years (from 1997 to 2010). The data were used to define the spectrum of diseases in the Southern Chinese population. Comparison with other populations revealed a unique spectrum of common IMDs. Furthermore, the incidence of the common IMDs was estimated by using population carrier frequencies of known recurrent mutations. Locally common diseases (their estimated incidence) include (1) glutaric aciduria type 1 (∼1/60,000), (2) multiple carboxylase deficiency (∼1/60,000), (3) primary carnitine deficiency (∼1/60,000), (4) carnitine-acylcarnitine translocase deficiency (∼1/60,000), (5) glutaric aciduria type 2 (∼1/22,500), (6) citrin deficiency (∼1/17,000), (7) tetrahydrobiopterin-deficient hyperphenylalaninaemia due to 6-pyruvoyl-tetrahydropterin synthase deficiency (∼1/60,000), (8) glycogen storage disease type 1 (∼1/150,000). In addition, ornithine carbamoyltransferase deficiency and X-linked adrenoleukodystrophy are common X-linked diseases. Findings of the disease spectrum and treatment outcome are summarised here which may be useful for clinical practice. In addition, data will also be useful for policy makers in planning of newborn screening programs and resource allocation.


Assuntos
/genética , Erros Inatos do Metabolismo/epidemiologia , Erros Inatos do Metabolismo/genética , China/epidemiologia , Humanos , Incidência , Mutação
8.
World J Pediatr ; 9(4): 361-4, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24146181

RESUMO

BACKGROUND: Recent studies presented a contradictory approach for the investigation of pediatric patients with an isolated increase in alanine transaminase. While classical teaching advised for a thorough investigation, recent studies suggested the yield on further investigation was low and thus not necessary. Yet the approach to the same clinical problem may need to be different due to variable disease prevalence rates among different ethnic populations. For the population with a higher prevalence rate of genetic liver diseases like Wilson's disease, an abnormal liver function may be the first presenting feature for some patients. METHODS: We reviewed 10 Chinese children with Wilson's disease who were diagnosed at a presymptomatic stage because of an isolated persistent elevation of alanine transaminase. RESULTS: All 10 patients did not have overt symptoms of liver impairment or neurological deficit. They were picked up incidentally with an abnormal liver function test. All patients were started on treatment shortly after diagnosis, and they remained well and symptom-free on the latest follow-up. CONCLUSIONS: This case series illustrated that an isolated persistent elevation of alanine transaminase is an important clue to the early diagnosis of pre-symptomatic Wilson's disease. It is particularly relevant in the Asian population where the disease is more prevalent.


Assuntos
Alanina Transaminase/sangue , Degeneração Hepatolenticular/diagnóstico , Adenosina Trifosfatases/genética , Adolescente , Biomarcadores/sangue , Proteínas de Transporte de Cátions/genética , Criança , Pré-Escolar , ATPases Transportadoras de Cobre , Diagnóstico Precoce , Fígado Gorduroso/patologia , Feminino , Hong Kong , Humanos , Masculino , Mutação , Estudos Retrospectivos
9.
Mol Genet Metab ; 107(1-2): 136-44, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22864057

RESUMO

INTRODUCTION: Mucopolysaccharidosis (MPS) type VI (Maroteaux-Lamy syndrome) is a clinically heterogeneous lysosomal storage disorder. It presents significant diagnostic and treatment challenges due to the rarity of the disease and complexity of the phenotype. As information about MPS VI in Asia-Pacific countries is limited, a survey was conducted to assess current practices for diagnosis and management of MPS VI in this region. The participants were selected based on their experience in diagnosing and managing MPS patients. METHODS: The survey comprised 29 structured quantitative or qualitative questions. Follow-up consultations were undertaken to discuss the data further. RESULTS: Thirteen physicians from eight countries or regions (Australia, China, Hong Kong, Japan, Malaysia, Philippines, Taiwan and Thailand) were surveyed. At the time of the survey twenty-two patients with MPS VI were directly treated by the respondents and most (~80%) had rapidly progressing disease. A wide range of medical specialists are involved in managing patients with MPS VI, the most common being orthopedic surgeons, pediatricians and geneticists. The availability/accessibility of diagnostic tools, therapies and national insurance coverage vary greatly across the countries/regions and, in some cases, between different regions within the same country. Currently, there are national MPS management groups in Australia and Japan. Australia, Taiwan and Hong Kong have local guidelines for managing MPS and local MPS registries are available in Australia, Taiwan, and Japan. CONCLUSIONS: This survey highlights differences in the diagnosis and management of MPS VI between Asia-Pacific countries/regions. Important barriers to advancing the identification, understanding and treatment of MPS VI include the paucity of epidemiological information, limited access to laboratory diagnostics and therapies, low disease awareness, and a lack of monitoring and treatment guidelines. There is a clear need to facilitate communications between physicians and establish regional or national disease registries, a multidisciplinary referral network, and a centralized diagnostic and management framework.


Assuntos
Mucopolissacaridose VI/diagnóstico , Mucopolissacaridose VI/terapia , Ásia , Austrália , Pesquisas sobre Serviços de Saúde , Humanos , Médicos , Inquéritos e Questionários
10.
World J Pediatr ; 8(3): 278-80, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21874615

RESUMO

BACKGROUND: Holocarboxylase synthetase deficiency is an inborn error of biotin metabolism leading to multiple carboxylase deficiency which is often biotin responsive. This disease is believed to be relatively common among the Asian population. METHODS: A 6-year-old Vietnamese boy presented with recurrent episodes of severe metabolic acidosis precipitated by intercurrent illnesses. An extensive skin rash was present since the onset of his illness. Multiple carboxylase deficiency was considered a likely diagnosis based on the history and the characteristic skin rash. RESULTS: This diagnosis was later confirmed by urine organic acid and molecular genetic studies. Urine organic acid showed characteristic excretion of glycine conjugates. Serum biotinidase activity was normal. Sequencing of the holocarboxylase synthetase gene revealed the patient being homozygous for a common mutation R508W. The patient showed a dramatic response to biotin within days of its administration. CONCLUSION: This case illustrates a potential highly treatable inborn error of metabolism that can be recognized on clinical grounds and its favorable response to biotin treatment.


Assuntos
Carbono-Nitrogênio Ligases/genética , Deficiência de Holocarboxilase Sintetase/genética , Mutação , /genética , Criança , Diagnóstico Diferencial , Deficiência de Holocarboxilase Sintetase/diagnóstico , Humanos , Masculino , Vietnã
12.
Mol Genet Metab ; 102(2): 222-5, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21131218

RESUMO

The diagnosis of glycogen storage disease (GSD) type IX is often complicated by the complexity of the phosphorylase kinase enzyme (PHK), and molecular analysis is the preferred way to provide definitive diagnosis. Here we reported two novel mutations found in two GSD type IX patients with different residual enzyme activities from Hong Kong, China using genetic analysis and, provided the molecular interpretation of the deficient PHK activity. These two newly described mutations would be useful for the study of future GSD patients.


Assuntos
Doença de Depósito de Glicogênio/genética , Mutação , Fosforilase Quinase/genética , Sequência de Aminoácidos , Sequência de Bases , Criança , Pré-Escolar , Ordem dos Genes , Doença de Depósito de Glicogênio/enzimologia , Hong Kong , Humanos , Fígado/diagnóstico por imagem , Fígado/patologia , Masculino , Ultrassonografia
13.
J Child Neurol ; 26(2): 179-87, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-20823027

RESUMO

This study included 12 Chinese patients with a wide spectrum of phenotypes of tyrosine hydroxylase deficiency. Seven females and 5 males, aged 2.2 to 41 years, had phenotypes ranging from severe type with onset at infancy to mild type with onset after 3 years of age. Patients with the severe type had encephalopathy with poor treatment response or infantile parkinsonism with motor delay. Patients with the less common mild type had dopa-responsive dystonia or a newly recognized predominant symptom of myopathy. Female siblings had more severe phenotypes. The phenotype and treatment outcomes were strongly related to a homovanillic acid level and homovanillic acid/5-hydroxyindolacetic acid ratio of less than 1 in the cerebrospinal fluid. Hyperprolactinemia was found in 50% of the severe cases. Levodopa was the mainstay of treatment, and early addition of selegiline resulted in a remarkable response in some patients. Treatment response for mild-type patients is universally good even with a treatment delay of 10 years after onset of neurological symptoms.


Assuntos
Deficiências Nutricionais/tratamento farmacológico , Dopaminérgicos/uso terapêutico , Levodopa/uso terapêutico , Transtornos Parkinsonianos/tratamento farmacológico , Selegilina/uso terapêutico , Tirosina 3-Mono-Oxigenase/deficiência , Adolescente , Adulto , Criança , Pré-Escolar , Quimioterapia Combinada , Feminino , Humanos , Masculino , Transtornos Parkinsonianos/genética , Fenótipo , Prolactina/sangue , Índice de Gravidade de Doença , Resultado do Tratamento
14.
15.
Pediatr Nephrol ; 24(10): 1989-96, 2009 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-19626343

RESUMO

We retrospectively reviewed the cases of 13 lupus nephritis children with pure membranous glomerulonephritis (MGN; Group A) and ten children with mixed proliferative and membranous nephritis (Group B). The children were identified through a territory-wide survey of patients between 1990 and 2003. All were ethnic Chinese. Age at diagnosis ranged from 3.7 to 18.6 years (Group A) and from 9.6 to 22.1 years (Group B). Female-to-male ratios were 12:1 (Group A) and 9:1 (Group B). Group A patients were more often nephrotic than Group B patients (11/13 vs. 5/10, p = 0.17). The glomerular filtration rate (GFR) at presentation was normal in all but two patients (one from each group). For induction, Group B patients consistently received prednisolone and cyclophosphamide; in contrast, the cytotoxic regimens in Group A patients varied from cyclophosphamide (five patients), mycophenolate mofetil (two patients), azathiorpine plus cyclosporine (one patient), and azathioprine alone (one patient). After a median follow-up of 7.6-7.8 years, one Group A patient had died of fulminant lupus. One survivor in Group B had a GFR < 90 ml/min per 1.73 m(2). Proteinuria persisted in five Group A patients and two Group B patients. In conclusion, Group B patients had good prognosis in terms of survival and proteinuria control. The only death occurred in Group A, and five of the 12 survivors in this group had persistent proteinuria. Further studies are needed to define the best treatment for pure lupus MGN.


Assuntos
Nefrite Lúpica/tratamento farmacológico , Adolescente , Anti-Inflamatórios/uso terapêutico , Azatioprina/uso terapêutico , Criança , Pré-Escolar , Ciclofosfamida/uso terapêutico , Ciclosporinas/uso terapêutico , Quimioterapia Combinada , Feminino , Taxa de Filtração Glomerular , Humanos , Imunossupressores/uso terapêutico , Nefrite Lúpica/fisiopatologia , Masculino , Ácido Micofenólico/análogos & derivados , Ácido Micofenólico/uso terapêutico , Prednisolona/uso terapêutico , Estudos Retrospectivos , Resultado do Tratamento
16.
Clin Chim Acta ; 404(2): 95-9, 2009 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-19265687

RESUMO

BACKGROUND: Multiple acyl-CoA dehydrogenase deficiency (MADD, OMIM 231680) or glutaric aciduria type II (GAII) is an inherited autosomal recessive disease affecting fatty acid, amino acid and choline metabolism, due to mutations in one of three genes namely, electron transfer flavoprotein alpha-subunit, ETFA (OMIM 608053), electron transfer flavoprotein beta-subunit, ETFB (OMIM 130410) and electron transfer flavoprotein dehydrogenase, ETFDH (OMIM 231675). Some MADD patients are responsive to riboflavin treatment with an excellent prognosis. Recently, mutations in ETFDH were found to be responsible for all riboflavin-responsive MADD patients. In this study, we present the clinical features and molecular studies of 2 Chinese families with riboflavin-responsive MADD. METHODS: Genomic DNA was extracted from peripheral blood samples or skin fibroblast cultures from the patients and normal controls. The thirteen exons of ETFDH were amplified by PCR. PCR products were sequenced in both forward and reverse directions. To rule out mutations in other genes, phenotype segregation was studied in the families by microsatellite markers in the proximity of the 3 genes, ETFA, ETFB and ETFDH. RESULTS: Four novel mutations in ETFDH were detected in the 2 families. In family 1, a frame shift mutation, c.1355delG which introduced a premature-termination codon (PTC), I454X in exon 11 of ETFDH was found. Another mutation was a c.250G>A transition in exon 3 of ETFDH, A84T. In family 2, two novel missense mutations were identified, P137S, in exon 4 and G467R in exon 11. No carrier of these four mutations was identified from about 150 alleles of healthy Chinese control subjects. CONCLUSIONS: Four novel mutations (3 missenses and 1 deletion) in ETFDH were found in Chinese families that presented with riboflavin-responsive MADD, which further expands the list of mutations found in patients with riboflavin-responsive MADD. Furthermore, we illustrated the utility of phenotype-genotype segregation in MADD families to prioritize genes for sequencing or to rule out the presence of disease causing mutation in other genes in MADD and other diseases caused by multiple genes.


Assuntos
Flavoproteínas Transferidoras de Elétrons/genética , Proteínas Ferro-Enxofre/genética , Deficiência Múltipla de Acil Coenzima A Desidrogenase/tratamento farmacológico , Deficiência Múltipla de Acil Coenzima A Desidrogenase/genética , Oxirredutases atuantes sobre Doadores de Grupo CH-NH/genética , Pré-Escolar , China , Análise Mutacional de DNA , Flavoproteínas Transferidoras de Elétrons/efeitos dos fármacos , Éxons/genética , Feminino , Mutação da Fase de Leitura/genética , Humanos , Lactente , Proteínas Ferro-Enxofre/efeitos dos fármacos , Repetições de Microssatélites/genética , Mutação de Sentido Incorreto/genética , Oxirredutases atuantes sobre Doadores de Grupo CH-NH/efeitos dos fármacos , Riboflavina/uso terapêutico , Adulto Jovem
18.
J Hum Genet ; 53(1): 55-63, 2008.
Artigo em Inglês | MEDLINE | ID: mdl-18034201

RESUMO

Wilson disease (WD), an autosomal recessive disorder of copper transport, is the most common inherited liver disorder in Hong Kong Chinese. This was the first local study to elucidate the molecular basis and establish an effective DNA-based diagnostic protocol. The ATP7B genes of 65 patients were amplified by polymerase chain reaction (PCR) and sequenced. Haplotype analysis was performed using D13S301, D13S314, and D13S316. The p.L770L/p.R778L status in 660 subjects was determined to estimate WD prevalence. Allele age of p.R778L was determined by the smallest homozygosity region between D13S301 and D13S270. We identified 42 different mutations with 17 being novel. p.R778L (17.3%) was the most prevalent. Exons 2, 8, 12, 13, and 16 harbored 70% mutations. Thirty-two haplotypes were associated with WD chromosomes. The estimated prevalence rate was 1 in 5,400. Three out of 660 normal subjects had p.L770L/p.R778L. In the remaining 657 individuals, neither p.L770L nor p.R778L was found. We characterized a Hong Kong Chinese-specific ATP7B mutation spectrum with great genetic diversity. Exons 2, 8, 12, 13, and 16 should be screened first. The perfect linkage disequilibrium suggested that p.R778L and its private polymorphism p.L770L originated from a single ancestor. This East-Asian-specific mutation p.R778L/p.L770L is aged at least 5,500 years.


Assuntos
Heterogeneidade Genética , Degeneração Hepatolenticular/genética , Mutação , Análise Mutacional de DNA , Frequência do Gene , Haplótipos , Hong Kong , Humanos , Desequilíbrio de Ligação
19.
Clin Chim Acta ; 382(1-2): 25-30, 2007 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-17442290

RESUMO

BACKGROUND: Two separate and complementary assays, total mitochondrial fatty acid beta-oxidation (FAO) flux rate and acylcarnitine profiling, have been used to establish a definitive diagnosis of FAO defects (FAOD) in cultured cells. We developed a novel functional assay for total FAO rate assay by measurement of deuterated water enrichment and to combine it with the conventional acylcarnitine profiling method into a single tracer incubation experiment. METHODS: Skin fibroblasts were incubated in a medium containing universal deuterium-labeled palmitate ((2)H(31)-palmitate) and l-carnitine without glucose supplementation for 96 h. The culture medium was assayed for deuterated water enrichment using isotope ratio mass spectrometry (IRMS) and acylcarnitine profiling by electrospray-ionization tandem mass spectrometry (ESI/MS/MS). RESULTS: The medians of (2)H(2)O enrichment after 96 h of incubation of (2)H(31)-palmitate of the control, other inherited metabolic diseases and FAOD cell lines were 109.9, 102 and 23.1 ppm/mg protein/96 h, respectively. All fibroblasts with FAOD except carnitine uptake defective, multiple acyl-CoA dehydrogenase and short-chain 3-hydroxyacyl-CoA dehydrogenase deficient cells were well separated from the control (<60% control median, p<0.05) and could be identified by IRMS assay. Accumulations of disease-specific acylcarnitines due to blockage in the carnitine cycle and FAO spiral were also demonstrated by acylcarnitine profiling. CONCLUSIONS: This novel functional assay is less time consuming and relatively simple by comparison to other published methods and can be used to investigate patients suspected to have FAO defects.


Assuntos
Carnitina/análogos & derivados , Ácidos Graxos/metabolismo , Erros Inatos do Metabolismo Lipídico/diagnóstico , Mitocôndrias/metabolismo , Espectrometria de Massas por Ionização por Electrospray/métodos , Espectrometria de Massas em Tandem/métodos , Carnitina/análise , Carnitina/metabolismo , Linhagem Celular , Deutério/análise , Deutério/metabolismo , Fibroblastos/metabolismo , Humanos , Erros Inatos do Metabolismo Lipídico/metabolismo , Complexos Multienzimáticos/metabolismo , Oxirredução , Palmitatos/análise , Palmitatos/metabolismo , Pele/citologia , Pele/metabolismo
20.
J Mol Med (Berl) ; 85(4): 351-60, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17160405

RESUMO

Defects in the multimeric enzyme, UDP-N-acetylglucosamine:lysosomal enzyme N-acetylglucosamine-1-phosphotransferase (GNPT), result in the diseases of mucolipidosis (ML). This enzyme generates the mannose 6-phosphate residues on newly synthesized lysosomal enzymes for the efficient receptor-mediated transport to lysosomes. The enzyme contains alpha/beta and gamma subunits. Mutations in the alpha/beta subunit result in the classical ML II and IIIA, while defects in the gamma subunit results in the clinically milder ML IIIC. I-cells, a distinct histological feature characterized by the presence of abnormal cytoplasmic vacuoles, are detected in many cell types, most noticeably, in ML II patients. In this study, we investigated the interactions of the alpha/beta and gamma subunits in the pathogenesis of I-cells. We noted low and deranged alpha/beta subunit expressions in human mucolipidosis cell lines. Unexpectedly, high gamma subunit expressions were also observed. In normal mouse fibroblasts, when alpha/beta subunit was suppressed, abnormal cytoplasmic vacuoles were induced, and up-regulation of the gamma subunit was also observed. On the other hand, suppressing the gamma subunit resulted in biphasic responses of the alpha/beta subunit, while abnormal cytoplasmic vacuoles were not formed, regardless of the expression levels of the alpha/beta subunit. Our data suggest reciprocal feedback mechanisms between alpha/beta and the gamma subunits. A fine balance of the expressions of these subunits may play an important role in the formation of I-cells in this group of lysosomal storage disorders.


Assuntos
Regulação Enzimológica da Expressão Gênica/fisiologia , Lisossomos/enzimologia , Transferases (Outros Grupos de Fosfato Substituídos)/metabolismo , Técnicas de Cultura de Células , Linhagem Celular , Humanos , Mucolipidoses/enzimologia , Transferases (Outros Grupos de Fosfato Substituídos)/genética , Vacúolos
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