Mutation-specific pathophysiological mechanisms define different neurodevelopmental disorders associated with SATB1 dysfunction.

Whereas large-scale statistical analyses can robustly identify disease-gene relationships, they do not accurately capture genotype-phenotype correlations or disease mechanisms. We use multiple lines of independent evidence to show that different variant types in a single gene, SATB1, cause clinically overlapping but distinct neurodevelopmental disorders. Clinical evaluation of 42 individuals carrying SATB1 variants identified overt genotype-phenotype relationships, associated with different pathophysiological mechanisms, established by functional assays. Missense variants in the CUT1 and CUT2 DNA-binding domains result in stronger chromatin binding, increased transcriptional repression, and a severe phenotype. In contrast, variants predicted to result in haploinsufficiency are associated with a milder clinical presentation. A similarly mild phenotype is observed for individuals with premature protein truncating variants that escape nonsense-mediated decay, which are transcriptionally active but mislocalized in the cell. Our results suggest that in-depth mutation-specific genotype-phenotype studies are essential to capture full disease complexity and to explain phenotypic variability.

Whole genome sequencing of 45 Japanese patients with intellectual disability.

Intellectual disability (ID) is characterized by significant limitations in both intellectual functioning and adaptive behaviors, originating before the age of 18 years. However, the genetic etiologies of ID are still incompletely elucidated due to the wide range of clinical and genetic heterogeneity. Whole genome sequencing (WGS) has been applied as a single-step clinical diagnostic tool for ID because it detects genetic variations with a wide range of resolution from single nucleotide variants (SNVs) to structural variants (SVs). To explore the causative genes for ID, we employed WGS in 45 patients from 44 unrelated Japanese families and performed a stepwise screening approach focusing on the coding variants in the genes. Here, we report 12 pathogenic and likely pathogenic variants: seven heterozygous variants of ADNP, SATB2, ANKRD11, PTEN, TCF4, SPAST, and KCNA2, three hemizygous variants of SMS, SLC6A8, and IQSEC2, and one homozygous variant in AGTPBP1. Of these, four were considered novel. Furthermore, a novel 76 kb deletion containing exons 1 and 2 in DYRK1A was identified. We confirmed the clinical and genetic heterogeneity and high frequency of de novo causative variants (8/12, 66.7%). This is the first report of WGS analysis in Japanese patients with ID. Our results would provide insight into the correlation between novel variants and expanded phenotypes of the disease.

Dominant-negative mutations in alpha-II spectrin cause West syndrome with severe cerebral hypomyelination, spastic quadriplegia, and developmental delay.

A de novo 9q33.3-q34.11 microdeletion involving STXBP1 has been found in one of four individuals (group A) with early-onset West syndrome, severe hypomyelination, poor visual attention, and developmental delay. Although haploinsufficiency of STXBP1 was involved in early infantile epileptic encephalopathy in a previous different cohort study (group B), no mutations of STXBP1 were found in two of the remaining three subjects of group A (one was unavailable). We assumed that another gene within the deletion might contribute to the phenotype of group A. SPTAN1 encoding alpha-II spectrin, which is essential for proper myelination in zebrafish, turned out to be deleted. In two subjects, an in-frame 3 bp deletion and a 6 bp duplication in SPTAN1 were found at the initial nucleation site of the alpha/beta spectrin heterodimer. SPTAN1 was further screened in six unrelated individuals with WS and hypomyelination, but no mutations were found. Recombinant mutant (mut) and wild-type (WT) alpha-II spectrin could assemble heterodimers with beta-II spectrin, but alpha-II (mut)/beta-II spectrin heterodimers were thermolabile compared with the alpha-II (WT)/beta-II heterodimers. Transient expression in mouse cortical neurons revealed aggregation of alpha-II (mut)/beta-II and alpha-II (mut)/beta-III spectrin heterodimers, which was also observed in lymphoblastoid cells from two subjects with in-frame mutations. Clustering of ankyrinG and voltage-gated sodium channels at axon initial segment (AIS) was disturbed in relation to the aggregates, together with an elevated action potential threshold. These findings suggest that pathological aggregation of alpha/beta spectrin heterodimers and abnormal AIS integrity resulting from SPTAN1 mutations were involved in pathogenesis of infantile epilepsy.

A case of osteogenesis imperfecta type II caused by a novel COL1A2 gene mutation: endoscopic third ventriculostomy to prevent hydrocephalus.

Osteogenesis imperfecta (OI) is a hereditary connective tissue disorder caused by defects in type I collagen synthesis. OI is generally classified into four types (I to IV), and the clinical prognosis varies from a lethal outcome for type II and varying deformities for type III to a normal lifespan for the other types. We describe a female patient with biochemically confirmed OI caused by a novel mutation in the COL1A2 gene. Persistence of blue sclerae supported the diagnosis of OI type II. The case was complicated with obstructive hydrocephalus, for which endoscopic third ventriculostomy (ETV) was performed. The ETV was transiently effective for the obstructive hydrocephalus. The patient subsequently developed brain atrophy, partly through ischemic events after the ETV, which appeared to contribute to maintenance of smooth circulation of the cerebrospinal fluid. We conclude that continuous and adequate medical care including ETV can facilitate long-term survival even in lethal OI type II.

[Effects of piracetam therapy in a case of Lance-Adams syndrome].

We report a 17-year-old female patient with Lance-Adams syndrome caused by anoxic encephalopathy during a severe attack of bronchial asthma. She had difficulty in writing because of action myoclonus in her arms. She also exhibited freezing gait and was unable to walk without cane. Although her gait disturbance resembled those seen in patients with parkinsonism secondary to anoxic encephalopathy, surface electromyography revealed that it was caused by action myoclonus in her legs. The presence of giant somatosensory evoked potentials and enhanced cortical reflexes in response to the electrical stimulation to her posterior tibial nerves supported our diagnosis. A combined therapy with valproate sodium, clonazepam and piracetam (15 g/day) was not effective. However, her freezing gait remarkably improved and she was able to walk without help, after the treatment with sufficient dose of piracetam (21 g/day). Cortical hyperexcitability as revealed by electrophysiological examination also improved. We concluded that the combined therapy with antiepileptic drugs and piracetam was effective in the treatment for action myoclonus. However, because the effects seemed dose-related, the dosage of piracetam needed to be increased until the optimum effects were obtained.

Bilirubin encephalopathy: a study of neuronal subpopulations and neurodegenerative mechanisms in 12 autopsy cases.

Bilirubin encephalopathy (BE), which includes acute (kernicterus) and chronic (postkernicteric) forms, results from severe neonatal jaundice. In order to investigate neurodegenerative mechanisms in autopsy cases of BE, we immunohistochemically examined expressions of neurotransmitters, neuropeptides, and calcium-binding proteins in the basal ganglia; and deposition of oxidative products. Expression of tyrosine hydroxylase was reduced in the putamen in cases of acute BE, and in the globus pallidus in cases of acute and chronic postkernicteric BE. Methionine-enkephalin expression was reduced in the external segment of the globus pallidus in cases of acute and chronic postkernicteric BE, and immunoreactivity for substance P was severely altered in both internal and external segments in cases of chronic postkernicteric BE. A decrease in the number of parvalbumin-immunoreactive interneurons in the external segment of the globus pallidus was observed predominantly in cases of acute BE, whereas the number of interneurons immunoreactive for calbindin-D28K was reduced in the putamen in cases of chronic postkernicteric BE. Nuclear immunoreactivity for 8-hydroxy-2'-deoxyguanosine was seen in the putamen in half of the BE cases. These findings indicated that the putamen was impaired in BE and the pallidal external segment was also damaged in the acute form of BE, suggesting that oxidative damage to DNA is implicated in lesions of the basal ganglia.

[Immunohistochemical analysis of hepatic steatosis in autopsy cases of severe motor and intellectual disabilities].

To explore the pathogenesis of hepatic steatosis in severe motor and intellectual disabilities (SMID), we immunohistochemically examined the accumulation of oxidative products in 8 autopsy cases of SMID, 5 of which pathologically showed hepatic steatosis. Three of the 8 cases also demonstrated fibrosis in Glisson's sheath. Serial sections of liver were treated with monoclonal antibodies to oxidative products to proteins, lipids and nucleosides. Immunoreactivity for 4-hydroxynonenal, a marker of oxidative damage to lipids, was increased in the cytoplasm of hepatocytes in 5 cases; 2 with steatosis plus fibrosis, one with only fibrosis, and 2 with only hepatic steatosis. One case showed immunoreactivity for advanced glycation endproduct, a marker of oxidative damage to protein, in hepatocytes. There was no immunoreactivity for 8-hydroxy-2'-deoxyguanosine and hexanoyl lysine, markers of oxidative damage to DNA and lipids, respectively. These findings partly mimic those observed in adult patients with nonalcoholic steatohepatitis. The results suggested the involvement of oxidative damage to lipids in the pathogenesis of hepatic steatosis and/or fibrosis in patients with SMID.

Finger cold-induced vasodilatation, sympathetic skin response, and R-R interval variation in patients with progressive spinal muscular atrophy.

To elucidate autonomic function in spinal muscular atrophy, we evaluated finger cold-induced vasodilatation, sympathetic skin response, and R-R interval variation in 10 patients with spinal muscular atrophy: 7 of type 1, 2 of type 2, and 1 of type 3. Results of finger cold-induced vasodilatation, sympathetic skin response, and R-R interval variation were compared with those of healthy children. Finger cold-induced vasodilatation was abnormal in 6 of 10 patients with spinal muscular atrophy; it was normal in the healthy children. The mean sympathetic skin response latency and amplitude did not differ significantly from those of the healthy children. Amplitudes of sympathetic skin response to sound stimulation were absent or low in all six patients with spinal muscular atrophy. No significant difference was found in the mean R-R interval variation of patients with spinal muscular atrophy and healthy children. Results show that some patients with spinal muscular atrophy have autonomic dysfunction, especially sympathetic nerve hyperactivity, that resembles dysfunction observed in amyotrophic lateral sclerosis.

Autonomic dysfunction in cases of spinal muscular atrophy type 1 with long survival.

In Japan, quite a few patients with spinal muscular atrophy type 1 (SMA type 1) survive with mechanical ventilation. Since a patient with SMA type 1 and continuous artificial ventilation exhibited excessive perspiration and tachycardia, we examined the autonomic functions in three cases of SMA type 1, undergoing mechanical ventilation. Two cases exhibited the common sympathetic-vagal imbalance on R-R interval analysis involving 24-h Holter ECG recordings in addition to an abnormality in finger cold-induced vasodilatation. Furthermore, one case showed blood pressure and heart rate fluctuation with the paroxysmal elevation, and a high plasma concentration of norepinephrine during tachycardia. These findings suggest that autonomic dysfunction should be examined in SMA type 1 patients with long survival, although the pathogenesis remains to be clarified.

Mutations in the genes encoding eukaryotic translation initiation factor 2B in Japanese patients with vanishing white matter disease.

OBJECTIVE: Vanishing white matter disease (VWM) is a chronic, progressive leukoencephalopathy associated with episodes of rapid deterioration following minor stress events such as head traumas or infectious disorders. The white matter of the patients with VWM exhibits characteristic radiological findings. METHOD: The genes encoding all five subunits of eukaryotic translation initiation factor 2B (EIF2B) were analyzed in patients, who were tentatively diagnosed with VWM, by Sanger sequencing. RESULTS: Seven mutations were identified in the genes encoding the subunits 1, 2, 4, and 5 of EIF2B. Among them, one mutation (p.V83E) in the subunit 2 (EIF2B2) was recurrently identified in three alleles, indicating the most common mutation in Japanese patients with VWM. Two patients were homozygous, and the other four patients were compound heterozygous. CONCLUSION: All patients showed white matter abnormalities with various degrees. One patient showed manifestations of end-stage VWM disease. Some patients showed late onset and slow progression associated with brain magnetic resonance imaging displaying T2 high intensity only in the deep white matter. There was clinical heterogeneity among patients with VWM.

Autoimmune neurological disorders associated with group-A beta-hemolytic streptococcal infection.

Although central nervous system (CNS) disorders associated with group-A beta-hemolytic streptococcal (GABHS) infection occur only rarely, Sydenham's chorea is a well-recognized disease that can arise following infection. Children may develop a tic, obsessive compulsive disorder (OCD), and extrapyramidal movement subsequent to GABHS infection. These disorders have been termed pediatric autoimmune neuropsychiatric disorders associated with streptococci (PANDAS). Herein we report one case each of acute disseminated encephalomyelitis (ADEM), PANDAS and subacute encephalitis associated with GABHS infection. To evaluate the pathogenesis of the CNS disorders associated with GABHS infection, we measured levels of neurotransmitters, cytokines, anti-neuronal autoantibodies, and performed immunohistochemistry using patient sera to stain human brain sections. All three cases showed psychiatric behavioral disorders. Immunotherapy was effective, and homovanillic acid levels in the cerebrospinal fluid (CSF) were elevated at the acute stage in all three cases. In each case of ADEM and PANDAS, immunohistochemistry demonstrated neuronal impairment in the basal ganglia during the acute stage. Neuronal immunoreactivity was visualized in the cerebral cortex at the acute stage in the case of subacute encephalitis. There was no direct correlation between immunoreactivity of patient sera on the brain sections and positivity of anti-neuronal autoantibodies or CSF biomarkers. The results suggest that autoimmune responses may modulate neurotransmission, and the use of patient serum for immunohistochemistry is a sensitive screening method for the detection of anti-neuronal autoantibodies in CNS disorders associated with GABHS infection.

Rituximab ameliorates anti-N-methyl-D-aspartate receptor encephalitis by removal of short-lived plasmablasts.

We measured anti-N-methyl-D-aspartate receptor (NMDAR) autoantibody levels and assessed B cell subsets using multicolor flow cytometry of peripheral blood mononuclear cells (PBMCs) from a recurrent anti-NMDAR encephalitis case to evaluate the effectiveness of rituximab treatment. Rituximab depleted CD20(+) fractions of naïve and memory B cell subsets and reduced the number of CD20(-) plasmablasts. This study suggests that short-lived plasmablasts are removed by rituximab-induced depletion of the CD20(+) B cell population. Increased numbers of plasmablasts in PBMCs may be a candidate predictive factor for unfavorable prognosis of anti-NMDAR encephalitis and an indication of when to commence second-line immunotherapy.

An autopsy case presenting repetitive hypoglycemia and unique cortical dysplasia.

In epileptic patients, focal cortical dysplasia (FCD) is pathologically characterized by irregular cortical lamination, blurring of the grey and white matter border and the occurrence of dysplastic cells in the cerebral cortex. Here, we report the case of a 42-year-old male showing developmental delay, transient repetition of hypoglycemic attack and cortical dysplasia, partly mimicking FCD. He had no family history of neurological disorders. He had never been able to stand independently and had always been unable to speak. He developed generalized convulsion in infancy, and then, in the absence of predisposing factors, suffered from repetitive hypoglycemic attacks between the ages of 27 and 38. Various endocrine tests, abdominal CT and brain MRI failed to demonstrate abnormalities. He died of peritonitis. At autopsy, no changes were observed in the pancreas, liver, kidneys, endocrine organs or hypothalamus. In the insular and frontal cortices, many large bizarre cells in the deep layer were observed and perivascular oligodendrocyte satellitosis was present in the adjacent white matter. Unlike FCD, the cortical lamination and the grey-white matter interface were preserved. A well-demarcated pilocytic astrocytoma was present in the brainstem. The cortical dysplasia, consisting of the diffuse occurrence of bizarre cells and the preservation of cortical lamination, is unique and has not been previously reported. Repetition of hypoglycemic attacks within a certain period is also noteworthy, although the relationship of this with the cortical dysplasia is unknown.

Mechanisms of neurodegeneration in neuronal ceroid-lipofuscinoses.

Neuronal ceroid-lipofuscinoses (NCL) are a group of neurodegenerative diseases and autosomal recessive lysosomal storage disorders. We examined the involvement of cell death, oxidative stress, and glutamate excitotoxicity using immunohistochemistry against Bcl-2, Bcl-x, oxidative products to proteins, lipids and DNA, calcium-binding proteins (calbindin-D28K, parvalbumin, calretinin), and glial glutamate transporters (excitatory amino acid transporters 1 and 2), in addition to terminal deoxynucleotidyl transferase-mediated dUTP-nick end labeling (TUNEL) in the brains from three cases of late infantile form of NCL (LINCL) and one case of juvenile form of NCL (JNCL) to investigate the neurodegenerative mechanisms. In the cerebral and cerebellar cortex, all of three LINCL cases demonstrated neurons with TUNEL-immunoreactive nuclei, whereas the JNCL case did not show TUNEL-immunoreactive nuclei. The coexistence of the nuclear TUNEL-immunoreactivity nuclei and cytoplasmic deposition of 4-hydroxy-2-nonenal-modified protein in the frontal cortex and hypoglossal nucleus may suggest a possible interrelationship between DNA fragmentation and lipid oxidation in LINCL. Additionally, glycoxidation of protein and oxidative stress to DNA seemed to be involved in the cerebellar and cerebral degeneration, respectively. Interneurons immunoreactive for calbindin-D28K and parvalbumin were severely reduced in the cerebral cortex, whereas those for calretinin were comparatively well preserved in LINCL, indicating the possibility of altered GABAergic system. The disturbance of expression of glial glutamate transporters seemed to be heterogeneous and mild. These findings suggest the possibility of new treatments for neurodegeneration in LINCL using antioxidative agents and/or GABAergic medications.