Nobuaki Iwasaki

A nonsense mutation of the MASS1 gene in a family with febrile and afebrile seizures

A naturally occurring mutation of the mass1 (monogenic audiogenic seizure‐susceptible) gene recently has been reported in the Frings mouse strain, which is prone to audiogenic seizures. The human orthologous gene, MASS1, was mapped to chromosome 5q14, for which we previously have reported significant evidence of linkage to febrile seizures (FEB4). We screened for MASS1 mutations in individuals from 48 families with familial febrile seizures and found 25 DNA alterations. None of nine missense polymorphic alleles was significantly associated with febrile seizures; however, a nonsense mutation (S2652X) causing a deletion of the C‐terminal 126 amino acid residues was identified in one family with febrile and afebrile seizures. Our results suggest that a loss‐of‐function mutation in MASS1 might be responsible for the seizure phenotypes, though it is not likely that MASS1 contributed to the cause of febrile seizures in most of our families.

Diphenylarsinic acid poisoning from chemical weapons in Kamisu, Japan

We noted a new clinical syndrome with prominent cerebellar symptoms in apartment building residents in Kamisu, Japan. The well that provided drinking water contained diphenylarsinic acid, a degradation product of diphenylcyanoarsine or diphenylchloroarsine, which were developed for use as chemical weapons, inducing severe vomiting and sneezing. Characteristics of diphenylarsinic acid poisoning include brainstem–cerebellar and cerebral symptoms. Mental retardation associated with brain atrophy in magnetic resonance images was evident in some infants. We must be vigilant to prevent or minimize the effects of further diphenylarsinic acid poisoning in Japan or elsewhere. Ann Neurol 2004;56:741–745

Linkage and association of febrile seizures to the IMPA2 gene on human chromosome 18

Background: Febrile seizures (FSs) are the most common form of childhood seizures, and genetic factors play a role in susceptibility to FS. Objective: To identify novel loci and genes associated with susceptibility to FS. Methods: Study participants were the FS probands and family members of 59 Japanese nuclear families (223 members including 112 affected children). Forty-eight of these families had at least two affected children for which genome-wide linkage screening was carried out. The Genehunter software was used to perform nonparametric multipoint linkage analysis. Mutational and association analyses were conducted in all 59 Japanese FS families. Results: Genotyping data of 407 microsatellite markers suggested linkage of FSs to chromosome 18p11.2 (non-parametric linkage score = 3.68, p = 0.0001). This region includes the IMPA2 gene, which encodes myo-inositol monophosphatase (IMPase) 2. In the phosphatidylinositol-signaling pathway, IMPase is inhibited by lithium, which has a proconvulsant effect, and is stimulated by carbamazepine, an anticonvulsant. A systematic search was performed for mutations in IMPA2 in 24 unrelated randomly selected Japanese FS patients; seven variants were detected. Haplotype analysis revealed an association of a common haplotype in IMPA2 with FSs (p = 0.0009). Conclusion: The authors found a novel locus on chromosome 18p11.2 for febrile seizures (FSs). IMPA2 is likely to be an FS susceptibility gene.

A quantitative analysis of rat central nervous system myelination using the immunohistochemical method for MBP

The temporal order of the myelination of the nervous pathways in 0-42-day-old Wistar rats was quantitatively analyzed using immunohistochemistry with anti-myelin basic protein (MBP) antibody. Immunohistochemistry was performed on paraffin-embedded tissue according to the standard ABC technique. For the objective evaluation of myelination, we converted the level stained with the immunohistochemical method into continuous numbers of 0-256 giving the intensity of myelination, using an image analyzing system. We analyzed nine nervous pathways: corpus callosum, optic tract, internal capsule, spinal tract of trigeminal nerve, inferior cerebellar peduncle, cerebellar white matter, pyramidal tract, medial longitudinal fasciculus, and cuneate fasciculus. The onset of the myelination of the spinal tract of the trigeminal nerve, inferior cerebellar peduncle, medial longitudinal fasciculus and cuneate fasciculus was day 7 (postnatal). That of the corpus callosum, optic tract, internal capsule and cerebellar white matter was day 14, and that of the pyramidal tract was day 21. The time required to reach the level of myelination of day 42 was day 21 for the spinal tract of the trigeminal nerve and the inferior cerebellar peduncle, day 28 for the internal capsule, day 35 for the corpus callosum, optic tract, cerebellar white matter and pyramidal tract, and day 42 for the medial longitudinal fasciculus. Our method using immunohistochemistry with anti-MBP antibody provided a highly sensitive and objective criterion for judging precisely the time and the progress of myelination in each nervous pathway and compare one nervous pathway with another.

A quantitative study of the progress of myelination in the rat central nervous system, using the immunohistochemical method for proteolipid protein

The temporal changes in intensity of myelination of the nervous pathways in 0 to 42-day-old Wistar rats were quantitatively analyzed by immunohistochemistry with anti-proteolipid protein and compared with that obtained by immunohistochemistry with anti-myelin basic protein. Immunohistochemistry was performed on paraffin-embedded tissue according to the standard ABC technique. Intensity of myelination was examined by an image analyzing system. We analyzed nine nervous pathways: corpus callosum, optic tract, internal capsule, spinal tract of the trigeminal nerve, inferior cerebellar peduncle, cerebellar white matter, pyramidal tract, medial longitudinal fasciculus, and cuneate fasciculus. The presence of immunoreactive fibers for proteolipid protein (PLP) in the spinal tract of the trigeminal nerve, medial longitudinal fasciculus and cuneate fasciculus was noted on postnatal day 0. Those of the corpus callosum, inferior cerebellar peduncle, cerebellar white matter, pyramidal tract and internal capsule were noted on day 7, and that of optic tract on day 14. The time required to reach the intensity of myelination of day 42 was day 14 for the cuneate fasciculus, day 21 for the spinal tract of the trigeminal nerve, inferior cerebellar peduncle and medial longitudinal fasciculus, day 28 for the optic and pyramidal tracts, day 35 for the corpus callosum and day 42 for the internal capsule and cerebellar white matter. The appearance of immunoreactive fibers for PLP was usually earlier than that for myelin basic protein (MBP) and the pattern of difference between PLP and MBP can be classified into three groups: (1) their time of appearance and progress are almost the same, as in the optic tract; (2) the appearance and progress of PLP occurs earlier than those of MBP, as in the pyramidal tract; (3) the appearance of PLP occurs earlier than that of MBP, but their progress is the same. Our findings revealed that the time of appearance and progress of myelination as measured by PLP are different among the nervous pathways, and that there is also a difference between PLP and MBP. This difference between PLP and MBP may indicate a functional difference between them.

Volumetric quantification of brain development using MRI

Abstract We devised a three-dimensional method for estimation of cerebral development and myelination which measures cerebral volume using MRI. Accuracy of the system was estimated using cadaver brains. The mean percentage error in the calculated volumes compared with the real volumes was 2.33 %, range 0.00–5.33 %. We applied the method to the volume of both cerebral hemispheres (CH), basal ganglia, thalamus and internal capsule (BT), and myelinated white matter (WM) in 44 neurologically normal individuals (4 months to 28 years of age), 13 patients with spastic motor disturbances (2–25 years of age), and 9 patients with athetotic motor disturbances (2–23 years of age). In the neurologically normal cases, the volumes of CH, BT and WM increased with age; the volume of MW more slowly than that of CH. In cases with spastic motor disturbances, the volumes of CH, BT and WM were between –1.4 and 3.5 SD, –1.0 and –3.5 SD, and 0.0 and –5.2 SD respectively, of those of neurologically-normal cases. On the other hand, 7 of the 9 cases with athetotic motor disturbances were within 2 SD of the volume of CH in neurologically normal cases. Our method for direct measurement of cerebral volume based on serial MRI should be useful for the accurate assessment of brain development and quantitative analysis of delayed myelination.

ICF syndrome in a girl with DNA hypomethylation but without detectable DNMT3B mutation

A 3‐year‐old girl with phenotypic and cytogenetic manifestations of the ICF syndrome and DNA hypomethylation but without DNMT3B gene mutation is described. At age 3 months, she had an apneic spell that left her with spastic paraplegia and severe mental retardation. At age 8 months, she suffered meningococcal meningitis and sepsis. When seen by us at age 3 years with virilization, she had a cleft plate, macroglossia, and an atrial septal defect. An adenoma was surgically removed from the right adrenal cortex. Her serum immunoglobulin levels were normal except IgA at the low normal border. Her lymphocytes showed paracentromeric stretching of chromosomes 1 and 16 in 7% of metaphases, and multiradial figures involving these chromosomes in 1% of cells. Hypomethylation of classical satellite 2 DNA was observed with BstBI digestion, but in a lesser degree than those in the individuals with proven DNMT3B mutations. No mutation was found in the coding and promoter regions of the gene. Several alternative interpretations were considered to explain the low frequencies of chromosomal instabilities and the lower degree of DNA hypomethylation, and undetected DNA3B mutations. A mutation may be present in the gene but undetected, present in other DNA methyltransferases (DNMT) genes or in a DNMT‐associated protein gene.

Molecular Genetics of Febrile Seizures

Summary: Febrile seizures are the most common form of convulsion, occurring in 2–5% of infants in Europe and North America and in 6–9% in Japan. In large families, the febrile seizure (FS) susceptibility trait is inherited by the autosomal dominant pattern with reduced penetrance. Two putative FS loci, FEB1 (chromosome 8q13‐q21) and FEB2 (chromosome 19p13.3) have been mapped. A clinical subset of FS, termed generalized epilepsy with febrile seizures plus (GEFS+), was reported. In GEFS+ families, a mutation in the voltage‐gated sodium channel β1 subunit gene (SCN1B) at chromosome 19q13.1 and two mutations of the same α1 subunit gene (SCN1A) at chromosome 2q24 were identified. These loci are linked to febrile convulsions in large families. We conducted a genome‐wide linkage search for FS in one large family with subsequent linkage confirmation in 39 nuclear families using nonparametric allele‐sharing methods, and found a new FS susceptibility locus, FEB4 (chromosome 5q14‐q15). In contrast to the FEB1, FEB2, and GEFS+ genetic loci, linkage to FEB4 was suggested in nuclear FS families, indicating that FEB4 may be the most common linkage locus in FS families.

Failure to find causal mutations in the GABAA-receptor γ2 subunit (GABRG2) gene in Japanese febrile seizure patients

Recently, mutations in the GABA(A)-receptor gamma2 subunit (GABRG2) gene were identified in two families with generalized epilepsy with febrile seizures plus (GEFS+) and two families with childhood absence epilepsy (CAE) and febrile seizures (FS). We tested the hypothesis that genetic variations in the GABRG2 gene confer susceptibility to FS in the Japanese population. We performed a systematic search for mutations in 94 unrelated Japanese patients with FS and detected six variants (-158C>T, 315C>T, 588T>C, IVS5-55C>T, IVS7+20G>A, and IVS7-141T>A). No non-synonymous mutation was detected. We genotyped three exonic polymorphisms and performed a case control study and a transmission disequilibrium test using 55 independent complete trios with FS and 106 control subjects. None of these polymorphic alleles were significantly associated with FS. Our results indicate that genomic variations of GABRG2 are not likely to be substantially involved in the etiology of FS in the Japanese population.

No evidence for gamma‐interferon mediated haematopoietic inhibition by T cells in aplastic anaemia: an observation in the course of immunosuppressive therapy

Two patients with aplastic anaemia were treated with immunosuppressive agents and peripheral blood T cells were cryopreserved serially. Inhibitory activity of T cells to autologous CFU‐E, γ‐IFN production by T cells and T cell subpopulations were assayed after remission. Inhibitory activity to autologous CFU‐E was not correlated with the numbers or ratios of T cell subpopulations. γ‐IFN production by T cells were within the normal range when inhibitory activity was found. In addition, γ‐IFN production increased after haematopoietic recovery. These findings suggest that γ‐IFN is not a soluble mediator of T cell‐mediated haematopoietic inhibition in aplastic anaemia.