[Show abstract][Hide abstract] ABSTRACT: Limb malformations are rare disorders with high genetic heterogeneity. Although multiple genes/loci have been identified in limb malformations, underlying genetic factors still remain to be determined in most patients.
[Show abstract][Hide abstract] ABSTRACT: The ASCO classification can evaluate the etiology and mechanisms of ischemic stroke more comprehensively and systematically than conventional stroke classification systems such as Trial of Org 10172 in Acute Stroke Treatment (TOAST). Simultaneously, risk factors for cognitive impairment such as arterial sclerosis, leukoaraiosis, and atrial fibrillation can also be gathered and graded using the ASCO classification.
Journal of stroke and cerebrovascular diseases: the official journal of National Stroke Association 08/2014;
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: -Genetic predisposition to life-threatening cardiac arrhythmias such as in congenital long-QT syndrome (LQTS) and catecholaminergic polymorphic ventricular tachycardia (CPVT) represent treatable causes of sudden cardiac death in young adults and children. Recently, mutations in calmodulin (CALM1, CALM2) have been associated with severe forms of LQTS and CPVT, with life-threatening arrhythmias occurring very early in life. Additional mutation-positive cases are needed to discern genotype-phenotype correlations associated with calmodulin mutations. METHODS AND RESULTS: -We employed conventional and next-generation sequencing approaches including exome analysis in genotype-negative LQTS probands. We identified five novel de novo missense mutations in CALM2 in three subjects with LQTS (p.N98S, p.N98I, p.D134H) and two subjects with clinical features of both LQTS and CPVT (p.D132E, p.Q136P). Age of onset of major symptoms (syncope or cardiac arrest) ranged from 1-9 years. Three of five probands had cardiac arrest and one of these subjects did not survive. Although all probands had LQTS, two subjects also exhibited electrocardiographic features consistent with CPVT. The clinical severity among subjects in this series was generally less than that originally reported for CALM1 and CALM2 associated with recurrent cardiac arrest during infancy. Four of five probands responded to beta-blocker therapy whereas one subject with mutation p.Q136P died suddenly during exertion despite this treatment. Mutations affect conserved residues located within calcium binding loops III (p.N98S, p.N98I) or IV (p.D132E, p.D134H, p.Q136P) and caused reduced calcium binding affinity. CONCLUSIONS: -CALM2M mutations can be associated with LQTS and with overlapping features of LQTS and CPVT.
[Show abstract][Hide abstract] ABSTRACT: The heart has a dynamic compensatory mechanism for haemodynamic stress. However, the molecular details of how mechanical forces are transduced in the heart are unclear. Here we show that the transient receptor potential, vanilloid family type 2 (TRPV2) cation channel is critical for the maintenance of cardiac structure and function. Within 4 days of eliminating TRPV2 from hearts of the adult mice, cardiac function declines severely, with disorganization of the intercalated discs that support mechanical coupling with neighbouring myocytes and myocardial conduction defects. After 9 days, cell shortening and Ca(2+) handling by single myocytes are impaired in TRPV2-deficient hearts. TRPV2-deficient neonatal cardiomyocytes form no intercalated discs and show no extracellular Ca(2+)-dependent intracellular Ca(2+) increase and insulin-like growth factor (IGF-1) secretion in response to stretch stimulation. We further demonstrate that IGF-1 receptor/PI3K/Akt pathway signalling is significantly downregulated in TRPV2-deficient hearts, and that IGF-1 administration partially prevents chamber dilation and impairment in cardiac pump function in these hearts. Our results improve our understanding of the molecular processes underlying the maintenance of cardiac structure and function.
[Show abstract][Hide abstract] ABSTRACT: Several types of muscular dystrophy are caused by defective linkage between α-dystroglycan (α-DG) and laminin. Among these, dystroglycanopathy, including Fukuyama-type congenital muscular dystrophy (FCMD), results from abnormal glycosylation of α-DG. Recent studies have shown that like-acetylglucosaminyltransferase (LARGE) strongly enhances the laminin binding activity of α-DG. Therefore, restoration of the α-DG-laminin linkage by LARGE is considered one of the most promising possible therapies for muscular dystrophy. In this study, we generated transgenic mice that overexpress LARGE (LARGE Tg) and crossed them with dy(2 J) mice and fukutin conditional knockout mice, a model for laminin α2-deficient congenital muscular dystrophy (MDC1A) and FCMD, respectively. Remarkably, in both strains, the transgenic overexpression of LARGE resulted in an aggravation of muscular dystrophy. Using morphometric analyses, we found that the deterioration of muscle pathology was caused by suppression of muscle regeneration. Overexpression of LARGE in C2C12 cells further demonstrated defects in myotube formation. Interestingly, a decreased expression of insulin-like growth factor 1 (IGF-1) was identified in both LARGE Tg mice and LARGE-overexpressing C2C12 myotubes. Supplementing the C2C12 cells with IGF-1 restored the defective myotube formation. Taken together, our findings indicate that the overexpression of LARGE aggravates muscular dystrophy by suppressing the muscle regeneration and this adverse effect is mediated via reduced expression of IGF-1.
Human Molecular Genetics 04/2014; · 7.69 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Defects in dystroglycan glycosylation are associated with a group of muscular dystrophies, termed dystroglycanopathies, that include Fukuyama congenital muscular dystrophy (FCMD). It is widely believed that abnormal glycosylation of dystroglycan leads to disease-causing membrane fragility. We previously generated knock-in mice carrying a founder retrotransposal insertion in fukutin, the gene responsible for FCMD, but these mice did not develop muscular dystrophy, which hindered exploring therapeutic strategies. We hypothesized that dysferlin functions may contribute to muscle cell viability in the knock-in mice; however, pathological interactions between glycosylation abnormalities and dysferlin defects remain unexplored. To investigate contributions of dysferlin deficiency to the pathology of dystroglycanopathy, we have crossed dysferlin-deficient dysferlinsjl/sjl mice to the fukutin-knock-in fukutinHp/- and Large-deficient Largemyd/myd mice, which are phenotypically distinct models of dystroglycanopathy. The fukutinHp/- mice do not show a dystrophic phenotype; however, (dysferlinsjl/sjl: fukutinHp/-) mice showed a deteriorated phenotype compared with (dysferlinsjl/sjl: fukutinHp/+) mice. These data indicate that the absence of functional dysferlin in the asymptomatic fukutinHp/- mice triggers disease manifestation and aggravates the dystrophic phenotype. A series of pathological analyses using double mutant mice for Large and dysferlin indicate that the protective effects of dysferlin appear diminished when the dystrophic pathology is severe and also may depend on the amount of dysferlin proteins. Together, our results show that dysferlin exerts protective effects on the fukutinHp/- FCMD mouse model, and the (dysferlinsjl/sjl: fukutinHp/-) mice will be useful as a novel model for a recently proposed antisense oligonucleotide therapy for FCMD.
PLoS ONE 01/2014; 9(9):e106721. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Bardet-Biedl syndrome (BBS) is a rare heterogeneous autosomal recessive disorder characterized by rod-cone dystrophy, postaxial polydactyly, truncal obesity, hypogonadism, learning disability, and renal anomaly that are caused by ciliary dysfunction. 16 genes have been associated with the BBS phenotype. Although recent pathophysiological studies using animal models have shown that ciliary dysfunction may induce hydrocephalus, there have been no reports of BBS with intracranial hypertension. We here describe a 9-year-old Japanese girl who was diagnosed as having BBS and later received renal transplantation due to chronic renal failure. She also exhibited intracranial hypertension, including papilledema and increased intrathecal pressure (260-300mmH2O), but her brain magnetic resonance imaging was normal. No genetic abnormalities were detected by DNA chip analysis or exome sequencing. Her papilledema improved following administration of acetazolamide. This is the first report of a case of BBS complicated with intracranial hypertension and its treatment.
[Show abstract][Hide abstract] ABSTRACT: α-synuclein (SNCA) is an established susceptibility gene for Parkinson's disease (PD), one of the most common human neurodegenerative disorders. Increased SNCA is considered to lead to PD and dementia with Lewy bodies. Four single-nucleotide polymorphisms (SNPs) in SNCA 3' region were prominently associated with PD among different ethnic groups. To examine how these SNPs influence disease susceptibility, we analyzed their potential effects on SNCA gene expression. We found that rs356219 showed allele-specific features. Gel shift assay using nuclear extracts from SH-SY5Y cells showed binding of one or more proteins to the protective allele, rs356219-A. We purified the rs356219-A-protein complex with DNA affinity beads and identified a bound protein using mass spectrometry. This protein, YY1 (Yin Yang 1), is an ubiquitous transcription factor with multiple functions. We next investigated SNCA expression change in SH-SY5Y cells by YY1 transfection. We also analyzed the expression of antisense noncoding RNA (ncRNA) RP11-115D19.1 in SNCA 3'-flanking region, because rs356219 is located in intron of RP11-115D19.1. Little change was observed in SNCA expression levels; however, RP11-115D19.1 expression was prominently stimulated by YY1. In autopsied cortices, positive correlation was observed among RP11-115D19.1, SNCA and YY1 expression levels, suggesting their functional interactions in vivo. Knockdown of RP11-115D19.1 increased SNCA expression significantly in SH-SY5Y cells, suggesting its repressive effect on SNCA expression. Our findings of the protective allele-specific YY1 and antisense ncRNA raised a novel possible mechanism to regulate SNCA expression.Journal of Human Genetics advance online publication, 12 September 2013; doi:10.1038/jhg.2013.90.
Journal of Human Genetics 09/2013; · 2.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Sandhoff disease is a GM2 gangliosidosis caused by mutations in HEXB encoding the β-subunit of β-hexosaminidase A. β-Hexosaminidase A exists as a heterodimer consisting of α- and β-subunits, and requires a GM2 activator protein to hydrolyze GM2. To investigate the molecular pathology in an adult Sandhoff disease patient with an early disease onset, we performed mutation detection, western blot analysis and molecular simulation analysis. The patient had compound heterozygous mutations p.Arg505Gln and p.Ser341ValfsX30. Western blot analysis showed that the amount of mature form of the α- and β-subunits was markedly decreased in the patient. We then performed docking simulation analysis of the α- and β-subunits with p.Arg505Gln, the GM2AP/GM2 complex and β-hexosaminidase A, and GM2 and β-hexosaminidase A. Simulation analysis showed that p.Arg505Gln impaired each step of molecular conformation of the α- and β-subunits heterodimer, the activator protein and GM2. The results indicated that p.Ser341ValfsX30 reduced the amount of β-subunit, and that p.Arg505Gln hampered the maturation of α- and β-subunits, and hindered the catalytic ability of β-hexosaminidase A. In conclusion, various methods including simulation analysis were useful to understand the molecular pathology in Sandhoff disease.Journal of Human Genetics advance online publication, 13 June 2013; doi:10.1038/jhg.2013.68.
Journal of Human Genetics 06/2013; · 2.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Background Multiple-system atrophy is an intractable neurodegenerative disease characterized by autonomic failure in addition to various combinations of parkinsonism, cerebellar ataxia, and pyramidal dysfunction. Although multiple-system atrophy is widely considered to be a nongenetic disorder, we previously identified multiplex families with this disease, which indicates the involvement of genetic components. Methods In combination with linkage analysis, we performed whole-genome sequencing of a sample obtained from a member of a multiplex family in whom multiple-system atrophy had been diagnosed on autopsy. We also performed mutational analysis of samples from members of five other multiplex families and from a Japanese series (363 patients and two sets of controls, one of 520 persons and one of 2383 persons), a European series (223 patients and 315 controls), and a North American series (172 patients and 294 controls). On the basis of these analyses, we used a yeast complementation assay and measured enzyme activity of parahydroxybenzoate-polyprenyl transferase. This enzyme is encoded by the gene COQ2 and is essential for the biosynthesis of coenzyme Q10. Levels of coenzyme Q10 in lymphoblastoid cells and brain tissue were measured on high-performance liquid chromatography. Results We identified a homozygous mutation (M78V-V343A/M78V-V343A) and compound heterozygous mutations (R337X/V343A) in COQ2 in two multiplex families. Furthermore, we found that a common variant (V343A) and multiple rare variants in COQ2, all of which are functionally impaired, are associated with sporadic multiple-system atrophy. The V343A variant was exclusively observed in the Japanese population. Conclusions Functionally impaired variants of COQ2 were associated with an increased risk of multiple-system atrophy in multiplex families and patients with sporadic disease, providing evidence of a role of impaired COQ2 activities in the pathogenesis of this disease. (Funded by the Japan Society for the Promotion of Science and others.).
New England Journal of Medicine 06/2013; 369(3):233-44. · 54.42 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Muscle-eye-brain (MEB) disease is a congenital muscular dystrophy (CMD) phenotype characterized by hypotonia at birth, brain structural abnormalities and ocular malformations. To date, few MEB cases have been reported in China where clinical recognition and genetic confirmatory testing on a research basis are recent developments. Here, we report the clinical and molecular genetics of three MEB disease patients. The patients had different degrees of muscle, eye and brain symptoms, ranging from congenital hypotonia, early-onset severe myopia and mental retardation to mild weakness, independent walking and language problems. This confirmed the expanding phenotypic spectrum of MEB disease with varying degrees of hypotonia, myopia and cognitive impairment. Brain magnetic resonance imaging showed cerebellar cysts, hypoplasia and characteristic brainstem flattening and kinking. Four candidate genes (POMGnT1, FKRP, FKTN and POMT2) were screened, and six POMGnT1 mutations (four novel) were identified, including five missense and one splice site mutation. Pathogenicity of the two novel variants in one patient was confirmed by POMGnT1 enzyme activity assay, protein expression and subcellular localization of mutant POMGnT1 in HeLa cells. Transfected cells harboring this patient's L440R mutant POMGnT1 showed POMGnT1 mislocalization to both the Golgi apparatus and endoplasmic reticulum. We have provided clinical, histological, enzymatic and genetic evidence of POMGnT1 involvement in three unrelated MEB disease patients in China. The identification of novel POMGnT1 mutations and an expanded phenotypic spectrum contributes to an improved understanding of POMGnT1 structure-function relationships, CMD pathophysiology and genotype-phenotype correlations, while underscoring the need to consider POMGnT1 in Chinese MEB disease patients.
MGG Molecular & General Genetics 05/2013; · 2.58 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A group of muscular dystrophies, dystroglycanopathy, is caused by abnormalities in post-translational modifications of dystroglycan (DG). To better understand the pathophysiological roles of DG modification and to establish effective clinical treatment for dystroglycanopathy, we here generated 2 distinct conditional knock-out (cKO) mice for fukutin, the first dystroglycanopathy gene identified for Fukuyama congenital muscular dystrophy. The first dystroglycanopathy model-myofiber-selective fukutin-cKO (MCK-fukutin-cKO) mice-showed mild muscular dystrophy. Forced exercise experiments in presymptomatic MCK-fukutin-cKO mice revealed that myofiber membrane fragility triggered disease manifestation. The second dystroglycanopathy model-muscle precursor cell (MPC)-selective cKO (Myf5-fukutin-cKO) mice-exhibited more severe phenotypes of muscular dystrophy. Using an isolated MPC culture system, we demonstrated, for the first time, that defects in the fukutin-dependent modification of DG lead to impairment of MPC proliferation, differentiation, and muscle regeneration. These results suggest that impaired MPC viability contributes to the pathology of dystroglycanopathy. Since our data suggested that frequent cycles of myofiber degeneration/regeneration accelerate substantial and/or functional loss of MPC, we expected that protection from disease-triggering myofiber degeneration provides therapeutic effects even in mouse models with MPC defects; therefore, we restored fukutin expression in myofibers. Adeno-associated virus (AAV)-mediated rescue of fukutin expression that was limited in myofibers successfully ameliorated the severe pathology even after disease progression. In addition, compared to other gene therapy studies, considerably low AAV titers were associated with therapeutic effects. Together, our findings indicated that fukutin-deficient dystroglycanopathy is a regeneration-defective disorder, and gene therapy is a feasible treatment for the wide range of dystroglycanopathy even after disease progression.
Human Molecular Genetics 04/2013; · 7.69 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Oligomeric forms of amyloid-β peptide (Aβ) are thought to play a pivotal role in the pathogenesis of Alzheimers disease (AD), but the mechanism involved is still unclear. Here, we generated induced pluripotent stem cells (iPSCs) from familial and sporadic AD patients and differentiated them into neural cells. Aβ oligomers accumulated in iPSC-derived neurons and astrocytes in cells from patients with a familial amyloid precursor protein (APP)-E693 mutation and sporadic AD, leading to endoplasmic reticulum (ER) and oxidative stress. The accumulated Aβ oligomers were not proteolytically resistant, and docosahexaenoic acid (DHA) treatment alleviated the stress responses in the AD neural cells. Differential manifestation of ER stress and DHA responsiveness may help explain variable clinical results obtained with the use of DHA treatment and suggests that DHA may in fact be effective for a subset of patients. It also illustrates how patient-specific iPSCs can be useful for analyzing AD pathogenesis and evaluating drugs.
[Show abstract][Hide abstract] ABSTRACT: Protein misfolding and aggregation in the brain have been recognized to be crucial in the pathogenesis of various neurodegenerative diseases, including Alzheimer's, Parkinson's, and the polyglutamine (polyQ) diseases, which are collectively called the "protein misfolding diseases". In the polyQ diseases, an abnormally expanded polyQ stretch in the responsible proteins causes the proteins to misfold and aggregate, eventually resulting in neurodegeneration. Hypothesizing that polyQ protein misfolding and aggregation could be inhibited by molecules specifically binding to the expanded polyQ stretch, we identified polyQ binding peptide 1 (QBP1). We show that QBP1 does, indeed, inhibit misfolding and aggregation of the expanded polyQ protein in vitro. Furthermore overexpression of QBP1 by the crossing of transgenic animals inhibits neurodegeneration in Drosophila models of the polyQ diseases. We also introduce our attempts to deliver QBP1 into the brain by administration using viral vectors and protein transduction domains. Interestingly, recent data suggest that QBP1 can also inhibit the misfolding/aggregation of proteins responsible for other protein misfolding diseases, highlighting the potential of QBP1 as a general therapeutic molecule for a wide range of neurodegenerative diseases. We hope that in the near future, aggregation inhibitor-based drugs will be developed and bring relief to patients suffering from these currently intractable protein misfolding diseases.
Journal of the American Society for Experimental NeuroTherapeutics 03/2013; · 5.38 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Oligomeric forms of amyloid-β peptide (Aβ) are thought to play a pivotal role in the pathogenesis of Alzheimer's disease (AD), but the mechanism involved is still unclear. Here, we generated induced pluripotent stem cells (iPSCs) from familial and sporadic AD patients and differentiated them into neural cells. Aβ oligomers accumulated in iPSC-derived neurons and astrocytes in cells from patients with a familial amyloid precursor protein (APP)-E693Δ mutation and sporadic AD, leading to endoplasmic reticulum (ER) and oxidative stress. The accumulated Aβ oligomers were not proteolytically resistant, and docosahexaenoic acid (DHA) treatment alleviated the stress responses in the AD neural cells. Differential manifestation of ER stress and DHA responsiveness may help explain variable clinical results obtained with the use of DHA treatment and suggests that DHA may in fact be effective for a subset of patients. It also illustrates how patient-specific iPSCs can be useful for analyzing AD pathogenesis and evaluating drugs.
[Show abstract][Hide abstract] ABSTRACT: The Keio Twin Research Center has conducted two longitudinal twin cohort projects and has collected three independent and anonymous twin data sets for studies of phenotypes related to psychological, socio-economic, and mental health factors. The Keio Twin Study has examined adolescent and adult cohorts, with a total of over 2,400 pairs of twins and their parents. DNA samples are available for approximately 600 of these twin pairs. The Tokyo Twin Cohort Project has followed a total of 1,600 twin pairs from infancy to early childhood. The large-scale cross-sectional twin study (CROSS) has collected data from over 4,000 twin pairs, from 3 to 26 years of age, and from two high school twin cohorts containing a total of 1,000 pairs of twins. These data sets of anonymous twin studies have mainly targeted academic performance, attitude, and social environment. The present article introduces the research designs and major findings of our center, such as genetic structures of cognitive abilities, personality traits, and academic performances, developmental effects of genes and environment on attitude, socio-cognitive ability and parenting, genes x environment interaction on attitude and conduct problem, and statistical methodological challenges and so on. We discuss the challenges in conducting twin research in Japan.
Twin Research and Human Genetics 02/2013; 16(1):202-16. · 1.64 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A 36-year-old female with systemic lupus erythematosus and antiphospholipid syndrome was referred to our department because of mild weakness of left arm and an episode of right amaurosis fugax for twenty days. Brain MRI showed right ACA/MCA/PCA border zone infarction on DWI/T2WI/FLAIR and MR angiography (MRA) showed multiple segmental stenosis in right internal carotid artery, right and left middle cerebral artery, and bilateral posterior cerebral arteries. Treatment with oral aspirin (100 mg/day) and continuous infusion of heparin kept her neurological symptoms and signs stable. MRA on 28 days revealed complete recovery of multiple stenotic lesions, thus, diagnosis of reversible cerebral vasoconstriction (RCVS) was made. RCVS should be considered as a cause of neurological deficit in patients with SLE regardless of thunderclap headache.
[Show abstract][Hide abstract] ABSTRACT: A 31-year-old man was referred to our hospital because of progressive tremor and clumsiness in his limbs and trunk. His symptoms were started in the right leg then gradually spread to all extremities as well as his trunk for 2 years. Neurological examinations revealed muscle rigidity with resting tremor predominantly right limbs. Akinesia and retropulsion were positive. Neither pyramidal tract sign nor cerebellar ataxia was detected. Genetic testing showed the expansion of SCA8 CTA/CTG repeats as 28/141 repeats. Though moderate expansion (less than 92) of SCA8 repeats has been reported in healthy subjects and patients with various diseases, the extraordinary long expansion of CTA/CTG repeats in SCA8 gene in our patient could be significantly pathological. 600 mg/day of L-DOPA clearly improved his symptoms. Dedicate follow up of the clinical course of our patient and the accumulation of the further cases is essential.