Kazushi Miya

Serine racemase is predominantly localized in neurons in mouse brain

D‐Serine is the endogenous ligand for the glycine binding site of the N‐methyl‐D‐aspartate (NMDA)‐type glutamate receptor (GluR) channel and is involved in the regulation of synaptic plasticity, neural network formation, and neurodegenerative disorders. D‐Serine is synthesized from L‐serine by serine racemase (SR), which was first reported to be localized in astrocytes. However, recently, SR mRNA and its protein have been detected in neurons. In this study, we examined the SR distribution in the brain during postnatal development and in cultured cells by using novel SR knockout mice as negative controls. We found that SR is predominantly localized in pyramidal neurons in the cerebral cortex and hippocampal CA1 region. Double immunofluorescence staining revealed that SR signals colocalized with those of the neuron‐specific nuclear protein, but not with the astrocytic markers glial fibrillary acid protein and 3‐phosphoglycerate dehydrogenase. In the striatum, we observed SR expression in γ‐aminobutyric acid (GABA)ergic medium‐spiny neurons. Furthermore, in the adult cerebellum, we detected weak but significant SR signals in GABAergic Purkinje cells. From these findings, we conclude that SR is expressed predominantly in many types of neuron in the brain and plays a key role in the regulation of brain functions under physiological and pathological conditions via the production of the neuromodulator D‐serine. J. Comp. Neurol. 510:641–654, 2008.

PIGA mutations cause early-onset epileptic encephalopathies and distinctive features

Objective: To investigate the clinical spectrum caused by mutations in PIGA at Xp22.2, which is involved in the biosynthesis of the glycosylphosphatidylinositol (GPI) anchor, among patients with early-onset epileptic encephalopathies (EOEEs). Methods: Whole-exome sequencing was performed as a comprehensive genetic analysis for a cohort of 172 patients with EOEEs including early myoclonic encephalopathy, Ohtahara syndrome, and West syndrome, and PIGA mutations were carefully investigated. Results: We identified 4 PIGA mutations in probands showing early myoclonic encephalopathy, West syndrome, or unclassified EOEE. Flow cytometry of blood granulocytes from patients demonstrated reduced expression of GPI-anchored proteins. Expression of GPI-anchored proteins in PIGA-deficient JY5 cells was only partially or hardly restored by transient expression of PIGA mutants with a weak TATA box promoter, indicating a variable loss of PIGA activity. The phenotypic consequences of PIGA mutations can be classified into 2 types, severe and less severe, which correlate with the degree of PIGA activity reduction caused by the mutations. Severe forms involved myoclonus and asymmetrical suppression bursts on EEG, multiple anomalies with a dysmorphic face, and delayed myelination with restricted diffusion patterns in specific areas. The less severe form presented with intellectual disability and treatable seizures without facial dysmorphism. Conclusions: Our study confirmed that PIGA mutations are one genetic cause of EOEE, suggesting that GPI-anchor deficiencies may be an underlying cause of EOEE.

Anti-NMDAR autoimmune encephalitis.

The N-methyl-D-aspartate receptor (NMDAR) is involved in normal physiological and pathological states in the brain. Anti-NMDAR encephalitis is characterized by memory deficits, seizures, confusion, and psychological disturbances in males and females of all ages. This type of encephalitis is often associated with ovarian teratoma in young women, but children are less likely to have tumors. Anti-NMDAR encephalitis is a neuroimmune syndrome in patients with autoantibodies recognizing extracellular epitopes of NMDAR, and the autoantibodies attenuate NMDAR function through the internalization of NMDAR. Following the initial symptoms of inflammation, the patients show the various symptoms such as memory loss, confusion, emotional disturbances, psychosis, dyskinesis, decrease in speech intelligibility, and seizures. About half of these patients improved with immunotherapy including high-dose intravenous corticosteroids and intravenous immunoglobulins is administrated to these patients, but the patients who had no improvement with these therapy require further treatments with rituximab or cyclophosphamide. It is necessary to detect anti-NMDAR antibodies at early stages, because the prognosis of these patients may be improved by early treatment. Recovery is slow, and the patients may have some disturbances in their motor function and cognition. The pathologic mechanism underlying the development of anti-NMDAR encephalitis has been elucidated gradually, but the optimal treatment has not yet been clarified. Further studies are required to clarify in detail the mechanism underlying anti-NMDA encephalitis and to develop effective treatments.

Human bocavirus in patients with encephalitis, Sri Lanka, 2009-2010

We identified human bocavirus (HBoV) DNA by PCR in cerebrospinal fluid from adults and children with encephalitis in Sri Lanka. HBoV types 1, 2, and 3 were identified among these cases. Phylogenetic analysis of HBoV1 strain sequences found no subclustering with strains previously identified among encephalitis cases in Bangladesh.

Clinical and Electrophysiological Effects of D-Serine in a Schizophrenia Patient Positive for Anti-N-Methyl-D-Aspartate Receptor Antibodies

The term anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis refers to an autoimmune disorder in which immunoglobulin G antibodies (ABs) against the NR1 subunit of NMDAR cause receptor internalization and decreased NMDAR-mediated neurotransmission. NMDAR encephalitis predominantly affects women, children, and young adults; occurs with or without tumor association; and is characterized by a predictable set of symptoms including psychosis as a common early feature, disorganized behavior, motor (e.g., catatonia and dyskinesia) manifestations, and seizures. NMDAR encephalitis is responsive to immunotherapy but refractory to antipsychotic medication and is associated with the long-term persistence of behavioral and cognitive deficits. Patients’ ABs decrease the surface density of NMDAR clusters via antibody-mediated capping and internalization resulting in decreased NMDAR-mediated synaptic currents (1,2). This relative NMDAR function loss may underlie the deficits in behavior and cognition that are hallmarks of NMDAR encephalitis. A proportion of clinically diagnosed schizophrenia patients may be seropositive for anti-NMDAR ABs. The seropositivity prevalence rates seem to be 1 in 10 to 20 patients but may differ among patient types (3). Furthermore, similar seropositivity rates were reported among healthy individuals (4), rendering the significance of anti-NMDAR ABs presence less clear. We hypothesized that: 1) seropositive patients can be identified among chronic schizophrenia patients having illness features that are also characteristic manifestations of antiNMDAR encephalitis (3); and 2) NMDAR AB positive patients will respond to treatment with D-serine (DSR), which acts in vivo as NMDAR co-agonist. Anti-NMDAR AB levels were assessed in 17 DSM-IV diagnosed schizophrenia patients who fulfilled the following inclusion criteria: 1) treatment resistance to antipsychotic pharmacotherapy; and 2) at least one of the following: an abrupt start of disease, lack of previous or family psychiatric history, and atypical disease course; or presence of hebephrenic, catatonic, dyskinesia features or seizures unaccounted by a neurological or other disorder. Detection of ABs against extracellular epitopes of NMDAR was performed using a previously described cell-based assay (5). One of the 17 patients was strongly seropositive, at both X200 and X10 dilutions, for both immunoglobulin G and immunoglobulin M anti-NR1 AB isotypes. This was a 67year-old female who, at age 27 after a period of continuous headaches for which no organic basis had been found, had abruptly developed an acute psychosis characterized by grandiose and paranoid delusions, mystical thinking, elated affect, and agitation. Following diagnosis, she never returned to her previous functional level and except for short attempts at living in the community has been hospitalized ever since.

An unmasked mutation of EIF2B2 due to submicroscopic deletion of 14q24.3 in a patient with vanishing white matter disease

Leukodystrophy with vanishing white matter (VWM) is a neurodegenerative disorder with autosomal recessive traits that is caused by alteration of the eukaryotic translation initiation factor‐2B (EIF2B). An 11‐month‐old patient with distinctive features began to exhibit progressive developmental deterioration associated with intractable epilepsy, which was triggered by recurrent acute infectious diseases. Brain magnetic resonance imaging (MRI) revealed abnormal white matter intensity. Chromosomal microarray testing identified a submicroscopic deletion at 14q24.3 that included EIF2B2, the gene encoding one of the subunits of EIF2B. Because the patients clinical findings were distinctive for VWM, compound heterozygous mutations of EIF2B2 were suspected, and subsequent sequencing analysis of the remaining allele unmasked the existence of a novel missense mutation of EIF2B2 (V85W). Some distinctive features including small palpebral fissures, bushy eyebrows, ear abnormalities, small upturned nose, downturned corners of the mouth, and micrognathia may be the common features of the patients with 14q24.3 deletions.

Deep sequencing detects very-low-grade somatic mosaicism in the unaffected mother of siblings with nemaline myopathy

When an expected mutation in a particular disease-causing gene is not identified in a suspected carrier, it is usually assumed to be due to germline mosaicism. We report here very-low-grade somatic mosaicism in ACTA1 in an unaffected mother of two siblings affected with a neonatal form of nemaline myopathy. The mosaicism was detected by deep resequencing using a next-generation sequencer. We identified a novel heterozygous mutation in ACTA1, c.448A>G (p.Thr150Ala), in the affected siblings. Three-dimensional structural modeling suggested that this mutation may affect polymerization and/or actins interactions with other proteins. In this family, we expected autosomal dominant inheritance with either parent demonstrating germline or somatic mosaicism. Sanger sequencing identified no mutation. However, further deep resequencing of this mutation on a next-generation sequencer identified very-low-grade somatic mosaicism in the mother: 0.4%, 1.1%, and 8.3% in the saliva, blood leukocytes, and nails, respectively. Our study demonstrates the possibility of very-low-grade somatic mosaicism in suspected carriers, rather than germline mosaicism.

A de novo interstitial deletion of 8p11.2 including ANK1 identified in a patient with spherocytosis, psychomotor developmental delay, and distinctive facial features

The contiguous gene syndrome involving 8p11.2 is recognized as a combined phenotype of both Kallmann syndrome and hereditary spherocytosis, because the genes responsible for these 2 clinical entities, the fibroblast growth factor receptor 1 (FGFR1) and ankyrin 1 (ANK1) genes, respectively, are located in this region within a distance of 3.2Mb. We identified a 3.7Mb deletion of 8p11.2 in a 19-month-old female patient with hereditary spherocytosis. The identified deletion included ANK1, but not FGFR1, which is consistent with the absence of any phenotype or laboratory findings of Kallmann syndrome. Compared with the previous studies, the deletion identified in this study was located on the proximal end of 8p, indicating a pure interstitial deletion of 8p11.21. This patient exhibited mild developmental delay and distinctive facial findings in addition to hereditary spherocytosis. Thus, some of the genes included in the deleted region would be related to these symptoms.

Epileptic apnea in a patient with inherited glycosylphosphatidylinositol anchor deficiency and PIGT mutations

We report an 11-month-old boy with acetazolamide-responsive epileptic apnea and inherited glycosylphosphatidylinositol (GPI)-anchor deficiency who presented with decreased serum alkaline phosphatase associated with compound PIGT mutations. The patient exhibited congenital anomalies, severe intellectual disability, and seizures, including epileptic apnea with epileptiform discharges from bilateral temporal areas. Brain magnetic resonance imaging revealed delayed myelination and progressive atrophy of the brainstem, cerebellum, and cerebrum. Whole-exome sequencing revealed compound heterozygous mutations in PIGT (c.250G>T, p.Glu84X and c.1096G>T, p.Gly366Trp), which encodes a subunit of the GPI transamidase complex. Flow cytometry revealed decreased expression of CD16 (a GPI anchor protein) on granulocytes, supporting the putative pathogenicity of the mutations. Phenobarbital, clonazepam, and potassium bromide decreased the frequency of tonic seizure and acetazolamide decreased epileptic apnea. To our knowledge, this is the first reported case of intractable seizures accompanied by epileptic apnea associated with GPI anchor deficiency and a compound PIGT mutation.

MODY3, renal cysts, and Dandy-Walker variants with a microdeletion spanning the HNF1A gene

Heterozygous hepatocyte nuclear factor-1-α gene (<italic>HNF1A</italic>) mutations are the most common cause of maturity-onset diabetes of the young (MODY), but they rarely involve extrahepatic manifestations. Renal cysts and diabetes syndrome can be caused by <italic>HNF1B</italic> mutations. No association between MODY3 and Dandy-Walker variants (DWV) has been reported. <italic>HNF1A</italic> mutations might be responsible for renal malformations. In a Japanese girl with glycosuria, developmental delay, mental retardation, renal cysts, and DWV, the <italic>HNF1B</italic> gene had no mutations. Array comparative genomic hybridization analysis identified a de-novo interstitial 12q24.22-q24.31 deletion of 5.6 Mb encompassing the <italic>HNF1A</italic> gene, which is compatible with a diagnosis of MODY3. The variety of phenotypes suggests a novel microdeletion syndrome spanning the <italic>HNF1A</italic> gene. Because <italic>HNF1B</italic> functions as an <italic>HNF1A/HNF1B</italic> heterodimer, haploinsufficient <italic>HNF1A</italic> interacts with a certain <italic>HNF1B</italic> haplotype. The resulting truncated heterodimer might engender renal cysts. More patients with well-defined deletion within 12q.24.31 must be evaluated to produce a detailed genotype-phenotype correlation and to elucidate this emerging microdeletion syndrome.
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