Transthyretin Thr60Ala Appalachian-type mutation in a Japanese family with familial amyloidotic polyneuropathy.

A Japanese case with familial amyloidotic polyneuropathy (FAP) associated with the transthyretin mutation Thr60Ala (Appalachian-type mutation) is described This is the first reported case of a non-Caucasian harboring this type of TTR mutation. The patient developed severe late-onset restrictive cardiomyopathy as well as sensorimotor and autonomic polyneuropathy, which were essentially similar to the previously reported clinical pictures of Appalachian-type FAP.     

Homozygosity for a Thr575Met missense mutation in the catalytic domain associated with factor XI deficiency

In this study we investigated an asymptomatic 55-year-old Lebanese woman with factor XI deficiency. The F11 gene was analyzed and a cross reacting material positive (CRM+) variant, Thr575Met, was identified in homozygosity in the proband, and in heterozygosity in four of her siblings.     

A Thr359Met mutation in factor VII of a patient with a hereditary deficiency causes defective secretion of the molecule

We elucidated the genetic basis responsible for factor VII deficiency in an Italian woman with a severe bleeding diathesis. In the allele inherited from the patient's father, we identified a G to A mutation at nucleotide 6070 at the 5' splice site of intron 4 and a G to A substitution at nucleotide 10976 resulting in the Arg353Gln polymorphism. The maternal allele demonstrated a C to T substitution at nucleotide 10994 resulting in Thr359Met. The mutation at nucleotide 6070 alters an invariant GT dinucleotide and disrupts normal mRNA processing. To investigate the mechanism by which Thr359Met reduces factor VIl levels, we expressed wild type factor VII cDNA (FVIIwt) and a mutant factor VII cDNA containing the base substitution resulting in Met359 (FVII359M) in Chinese hamster ovary cells (CHO). In cells transfected with the mutant factor VII cDNA, FVII359M accumulated intracellularly, and no factor VII was detected in the media after 3 hours of chase. The carbohydrate side chains associated with FVII359M were sensitive to Endo H digestion, which indicates that the protein is retained in the endoplasmic reticulum. Analysis of cell lysates also showed that FVII359M was associated with the 78 kD protein corresponding to GRP78/BiP. We conclude that a Thr359Met mutation in factor VII results in a severe secretion defect that probably results from abnormal folding of the molecule.     

A Japanese family of typical Loeys-Dietz syndrome with a TGFBR2 mutation

This report describes a Japanese family with vessel and craniofacial abnormalities. Although the clinical findings of the patient's father fulfilled the diagnostic criteria for Marfan syndrome, arterial tortuosity, aneurysms, hypertelorism and a bifid uvula were noted in both the patient and his father. These findings were compatible with the clinical manifestations that were previously reported in Loeys-Dietz syndrome. A molecular genetic analysis demonstrated a heterozygous missense mutation of the transforming growth factor-beta receptor II gene in both the patient and his father, which thus caused Loeys-Dietz syndrome. This is the first Japanese family case report of typical Loeys-Dietz syndrome.     

First Spanish family with familial amyloidotic polyneuropathy associated to TTR Thr49Ile mutation

We present a Spanish patient with familial amyloidotic polyneuropathy associated with the TTR Thr49lle mutation previously described in a Japanese patient. This is the first report in a Caucasian patient and the second in the literature. Age of onset at 66 and the clinical picture were similar to the Japanese patient: sensorimotor polyneuropathy, digestive autonomic disturbances, cardiomyopathy and loss of weight. The mutation was diagnosed by DNA sequencing and induced mutation restriction analysis.     

A novel mutation associated with Jervell and Lange-Nielsen syndrome in a Japanese family.

BACKGROUND: The Jervell and Lange-Nielsen (JLN) syndrome is a variant of long QT syndromes (LQTS) and is associated with congenital deafness. The syndrome is caused by homozygous or compound heterozygous mutations in genes KCNQ1 and KCNE1, which are responsible for encoding the delayed rectifier repolarizing current, I(Ks). METHODS AND RESULTS: A novel and homozygous KCNQ1 mutation in a 23-year-old deaf woman with a prolonged QT interval and recurrent syncope in a Japanese family was identified. Genetic analyses revealed that the proband harbored a KCNQ1 missense mutation (W248F) located in the intracellular S4-S5 linker on both alleles. The same mutation was identified in both maternal and paternal families in a heterozygous manner. However, the family members of both sides had no clinical evidence of LQTS or hearing defects. Functional assays using a heterologous expression system revealed that W248F KCNQ1 plus KCNE1 channels reconstitute hardly measurable I(Ks) currents. In contrast, heterozygous wild-type/W248F KCNQ1 plus KCNE1 channels displayed biophysical properties similar to those of the wild-type KCNQ1 plus KCNE1 channels with a weak dominant-negative effect. CONCLUSION: In this study, we present a family with JLN syndrome. The electrophysiological properties of the mutant I(Ks) channels explain the pathophysiology underlying JLNS.     

First Spanish family with familial amyloidotic polyneuropathy associated to TTR Thr49Ile mutation.

We present a Spanish patient with familial amyloidotic polyneuropathy associated with the TTR Thr49Ile mutation previously described in a Japanese patient. This is the first report in a Caucasian patient and the second in the literature. Age of onset at 66 and the clinical picture were similar to the Japanese patient: sensorimotor polyneuropathy, digestive autonomic disturbances, cardiomyopathy and loss of weight. The mutation was diagnosed by DNA sequencing and induced mutation restriction analysis.     

A Japanese case of familial Mediterranean fever with family history demonstrating a mutation in MEFV

We describe a 17-year-old woman with a family history of FMF who suffered from recurrent fever accompanied by pains in the left chest and abdomen. During a five-year period she experienced attacks about once every six months. The metaraminol provocative test was positive. Genomic DNA extracted from peripheral blood lymphocytes from both her and her parents were analyzed by polymerase chain reaction (PCR), followed by cycle sequencing. We detected a mutation (ATG to ATA) in codon 694 in exon 10 of the FMF gene, MEFV, that resulted in a substitution of isoleucine for methionine (M6941) in both her and her father. This is the first Japanese case of FMF with a mutation in MEFV identified in the family history.     

Angiotensinogen gene polymorphism (Met235Thr) influences visceral obesity and insulin resistance in obese Japanese women

To investigate the relationship between angiotensinogen (AGT) Met235Thr polymorphism (M235T) and human obesity, because AGT is regarded as one of the cytokines produced from adipocytes and serum AGT concentrations are reported to be positively correlated with body mass index. One hundred and twenty obese Japanese women (age, 58.8+/-9.4 years; body mass index, 32.2+/-4.9 kg/m(2)) were enrolled. Angiotensinogen genotypes were determined with a fluorescent allele-specific DNA primer assay system. Subjects were divided into M/M, M/T, and T/T groups. Control subjects comprised 146 healthy age-matched women. Clinical characteristics and the effects of diet and exercise therapy for 6 months were compared among the 3 genotypes. The genotype frequencies of AGT M235T polymorphism were in accordance with the Hardy-Weinberg equation (obese: M/M, 6.7%; M/T, 27.5%; T/T, 65.8%; control: M/M, 6.8%; M/T, 21.2%; T/T, 71.9%). The frequency of the T allele did not differ between obese and control subjects (0.80 vs 0.83). As the number of obese women with M/M genotype was only 8, comparisons of the characteristics and outcomes of weight reduction therapy were performed only between subjects with M/T genotype and T/T genotype. In the T/T group, % body fat and waist circumference at baseline were significantly greater than in the M/T group (36.3%+/-4.8% vs 33.8%+/-4.7%, P=.0105; 107.9+/-10.9 vs 102.6+/-7.9 cm, P=.0428, respectively). Before the weight reduction therapy, significantly higher insulin and higher homeostasis model assessment (HOMA-R) were demonstrated in the T/T group than in the M/T group (9.1+/-5.5 microU/mL vs 5.9+/-4.4 microU/mL, P=.0056; 2.3+/-1.4 vs 1.6+/-1.3, P=.0252, respectively). Both systolic and diastolic blood pressure at baseline in the T/T group tended to be higher than those in the M/T group, but the differences were not significant. No genotype-dependent difference in energy expenditure or outcome of weight reduction therapy was observed with respect to AGT M235T polymorphism. After the diet and exercise therapy, the blood pressure in the T/T group tended to be higher than that in the M/T group, but the difference was not significant. We demonstrated that the T/T genotype of the AGT M235T gene polymorphism was positively related to visceral obesity and hyperinsulinemia in obese Japanese women. Blood pressure did not show genotype-specific differences before or after the treatment. Further studies of the association between obesity and this gene polymorphism should contribute to understanding and treating obesity-related diseases.     

Photobleaching through glass micropipettes: sodium channels without lateral mobility in the sarcolemma of frog skeletal muscle.

Sodium currents were recorded from frog skeletal muscle by using fire-polished micropipettes to electrically isolate and voltage clamp a small patch of sarcolemma. Sodium current amplitude served as an assay for the number of functional sodium channels in the patch. With the pipette as a light guide, these channels were irradiated with ultraviolet (UV) light directed through a quartz fiber into the back end of the pipette. The UV light emerging from the pipette tip caused localized destruction of the sodium channels in the patch, reducing sodium current 3- to 5-fold during a 30-90 s irradiation. If sodium channels could diffuse laterally in the membrane, current from the patch should recover with time as fresh channels enter from neighboring areas. No such recovery was observed during observation for 1 hr after irradiation. Our results set an upper limit of 10(-12) cm2/s for the diffusion coefficient--1/1000th that of rhodopsin, a membrane protein in the cell membrane of retinal rods. It is suggested that sodium channels are anchored in the sarcolemma.     

DKC1 gene mutation in a Taiwanese kindred with X-linked dyskeratosis congenita

Dyskeratosis congenita (DKC) is a rare inherited disease characterized by the triad of abnormal skin pigmentation, nail dystrophy, and mucosal leukoplakia. Recent studies demonstrated mutations in the DKC1 gene encoding a protein named dyskerin, which is a component of human telomerase. In addition to the hypothesized function of pseudouridination in rRNA biosynthesis, ribosomal subunit assembly, and/or centromere/ microtubule binding, lower levels of telomerase activity in cells from patients with X-linked DKC have been observed. We report the mutation analysis of a Taiwanese family with X-linked DKC. The patient was a 19-year-old man who presented with progressive reticulate hyperpigmentation, nail dystrophy, alopecia, leukoplakia of the tongue, and pancytopenia. He died of enterocolitis and Escherichia coli sepsis at the age of 20 years. Only his mother's DNA was available for mutation analysis, which revealed a nucleotide transition of C to T (1058 C --> T), a hotspot mutation in DKC, resulting in an amino acid change from alanine to valine (A353V) in the DKC1 gene. Recent advances in the research of telomerase and its implications in the human aging process and cancer are discussed.     

De novo acute hepatitis B infection in a previously vaccinated liver transplant recipient due to a strain of HBV with a Met 133 Thr mutation in the "a" determinant

De novo HBV infection post-liver transplantation (LT) from an anti-HBc seropositive donor rarely presents as acute failure. We report a 42-year-old Caucasian female, HBsAg and anti-HBc seronegative, with primary biliary cirrhosis who received an allograft from a HBsAg negative, anti-HBc seropositive donor. The patient, previously vaccinated years pre-LT, was re-vaccinated against HBV and 1 year post-LT had an anti-HBs titre of 256 IU/l. Two years post-LT, elevated serum aminotransferases and worsening liver function with an INR of 2.0 developed. The HBsAg became positive, anti-HBs undetectable and serum HBV-DNA >2000 pg/ml by hybridisation assay. Liver biopsy revealed significant ballooning degeneration, piecemeal necrosis and positive immunostaining for HBsAg. Progressive liver failure developed followed by sepsis and terminal multi-organ failure. Subsequent analysis of the predominant HBV strain revealed mutations in the "a" determinant: Met 133 Thr (codon change ATG to ACG) and Asn 131 Thr. CONCLUSION:' Acute de novo HBV infection from an anti-HBc sero-positive donor may occur long after LT despite protective anti-HBs titres post-vaccination secondary to the emergence of "a" determinant mutated strains of HBV.     

肌球蛋白重链Val606Met突变基因型与家族性肥厚型心肌病

目的 研究中国人家族性肥厚型心肌病(HCM)的致病基因突变位点,分析基因型与临床表型的相互关系.方法 对3个家族性HCM的先证者进行β-肌球蛋白重链基因(β-MHC)扫描,聚合酶链反应扩增其功能区外显子片段,双脱氧末端终止法测序.对阳性测序结果者进行家系中其他成员筛查,并分析患者临床表型特点.结果 在其中1个家系发现Val606Met杂合突变,而正常对照组同一位置未见异常,属于在我国HCM家系中首次发现.结论 MYH7基因16号外显子Val606Met错义突变为我国家族性HCM的致病基因之一,其临床表型的异质性提示多因素参与了HCM的发生及外显.     

A novel mutation that leads to a congenital factor XI deficiency in a Japanese family

We have identified a novel mutation leading to a congenital deficiency of the coagulation factor XI (FXI) in a Japanese family. A propositus was a 42-year-old female patient without bleeding tendency. Coagulant activity and the antigen level of FXI in her plasma were below the detectable range. The nucleotide sequences of the FXI gene of this patient were determined by a direct sequence method established in this study. A novel nonsense mutation (CAA; Gly263 --> TAA; stop) was identified in exon 8 of the FXI gene. Her parents are first cousins, and a polymerase chain reaction-restriction-fragment length polymorphism analysis revealed that her parents were heterozygous at this nucleotide position. This patient inherited mutant alleles from her parents and is homozygous at this nucleotide position. The nonsense mutation in the FXI gene is responsible for her deficiency of FXI.     

The N131S mutation in the von Hippel-Lindau gene in a Japanese family with pheochromocytoma and hemangioblastomas

von Hippel-Lindau (VHL) disease (VHLD) is a hereditary autosomal dominant syndrome that causes various benign and malignant tumors. VHLD is caused by mutations in the VHL tumor suppressor gene. Here, we report a mutation in the VHL gene in a Japanese family with VHLD type 2A, characterized by pheochromocytoma (PHE), and hemangioblastomas (HAB) in both the retina and thoracic spinal cord but without renal cell carcinoma (RCC). We identified a heterozygous A to G point mutation at the second base of codon 131 of the VHL protein (pVHL). This mutation was predicted to convert codon 131 from asparagine to serine (N131S). Although most mutations in VHLD type 2A have been detected in the alpha domain of pVHL, the present mutated amino acid was located at the region encoding the beta domain of pVHL. Previous patients with the N131K or N131T mutation in pVHL developed VHLD type 2B with RCC or VHLD type 1 without PHE, respectively. We also identified somatic loss of heterozygosity (LOH) at chromosome 3p25-26 in the adrenal tumor of the patient. The results of our study suggest that not only the location of mutation but also the altered amino acid may be critical for determining the clinical phenotype of VHLD. LOH was associated with the development of PHE in a patient with the N131S mutation in pVHL.     

Type 2 (non-insulin-dependent) diabetes mellitus associated with a mutation of the glucokinase gene in a Japanese family

Summary Mutations were screened for in the glucokinase gene of 25 Japanese patients with Type 2 (non-insulin-dependent) diabetes mellitus. Each exon was scanned by electrophoresis of enzymatically amplified DNA segments under non-denaturing conditions and variants were sequenced. A variant pattern was detected in exon 5 of one patient. Direct sequencing of this exon revealed a single nucleotide substitution in codon 188 (GCTACT) of one of two alleles resulting in the mutation of Ala¹⁸⁸Thr, an invariant residue in the sequence of all mammalian glucokinases and hexokinases. This mutation was not found in 40 normal control subjects. The proband had been diagnosed with Type 2 diabetes at the age of 62 years. Four other members of her family have the same mutation and all have Type 2 diabetes or impaired glucose tolerance. The youngest age at diagnosis of Type 2 diabetes in these other members was 13 years, suggesting that her pedigree was maturity-onset diabetes of the young (MODY). All subjects with the Thr¹⁸⁸ mutation show a decreased insulin secretory response during oral glucose tolerance testing. Mutations in the glucokinase gene associated with Type 2 diabetes have been previously identified in Caucasian (French and British) subjects. This study indicates that mutations in this gene are also implicated in the development of Type 2 diabetes in Asians. Further studies are required to determine the frequency of mutations in glucokinase among Japanese patients with Type 2 diabetes.     

Two antithrombin mutations in a compound heterozygote: Met20Thr and Tyr166Cys

The molecular basis for a family with Type I antithrombin deficiency has been established. Amplification and sequencing of the antithrombin gene identified two mutations: Met20Thr (2523T°C) within exon 2 and Tyr166Cys (5493A→G) within exon 3a. Further analysis indicated that the propositus was a compound heterozygote but in addition provided evidence for phase disruption during the amplification and/or cloning procedure. The Met20Thr mutation appears to be a neutral mutation with no functional consequences. In contrast, the Tyr166Cys mutation is associated with a Type I phenotype.