Complete nanopore repeat sequencing of SCA27B (GAA-FGF14 ataxia) in Japanese.

BACKGROUND: Although pure GAA expansion is considered pathogenic in SCA27B, non-GAA repeat motif is mostly mixed into longer repeat sequences. This study aimed to unravel the complete sequencing of FGF14 repeat expansion to elucidate its repeat motifs and pathogenicity. METHODS: We screened FGF14 repeat expansion in a Japanese cohort of 460 molecularly undiagnosed adult-onset cerebellar ataxia patients and 1022 controls, together with 92 non-Japanese controls, and performed nanopore sequencing of FGF14 repeat expansion. RESULTS: In the Japanese population, the GCA motif was predominantly observed as the non-GAA motif, whereas the GGA motif was frequently detected in non-Japanese controls. The 5'-common flanking variant was observed in all Japanese GAA repeat alleles within normal length, demonstrating its meiotic stability against repeat expansion. In both patients and controls, pure GAA repeat was up to 400 units in length, whereas non-pathogenic GAA-GCA repeat was larger, up to 900 units, but they evolved from different haplotypes, as rs534066520, located just upstream of the repeat sequence, completely discriminated them. Both (GAA)≥250 and (GAA)≥200 were enriched in patients, whereas (GAA-GCA)≥200 was similarly observed in patients and controls, suggesting the pathogenic threshold of (GAA)≥200 for cerebellar ataxia. We identified 14 patients with SCA27B (3.0%), but their single-nucleotide polymorphism genotype indicated different founder alleles between Japanese and Caucasians. The low prevalence of SCA27B in Japanese may be due to the lower allele frequency of (GAA)≥250 in the Japanese population than in Caucasians (0.15% vs 0.32%-1.26%). CONCLUSIONS: FGF14 repeat expansion has unique features of pathogenicity and allelic origin, as revealed by a single ethnic study.

CSF 14-3-3 zeta(ζ) isoform is associated with tau pathology and cognitive decline in Alzheimer's disease.

14-3-3 is a family of conserved proteins that consist of seven isoforms which are highly expressed in the brain, and 14-3-3 zeta(ζ) is one of the isoforms encoded by the YWHAZ gene. Previous studies demonstrated that 14-3-3ζ is deposited in the neurofibrillary tangles of Alzheimer's disease (AD) brains, and that 14-3-3ζ interacts with tau from the purified neurofibrillary tangles of AD brain extract. The present study examined the cerebrospinal fluid (CSF) 14-3-3ζ levels of 719 participants from the Alzheimer's Disease Neuroimaging Initiative (ADNI), including cognitively normal (CN) participants, patients with mild cognitive impairment (MCI) and patients with AD dementia, and aimed to identify whether CSF 14-3-3ζ is associated with tau pathology. CSF 14-3-3ζ levels were increased in AD, and particularly elevated among tau pathology positive individuals. CSF 14-3-3ζ levels were associated with CSF phosphorylated tau 181 (p-tau) (r = 0.741, P < 0.001) and plasma p-tau (r = 0.293, P < 0.001), which are fluid biomarkers of tau pathology, and could predict tau pathology positive status with high accuracy (area under the receiver operating characteristic curve [AUC], 0.891). CSF 14-3-3ζ levels were also correlated to synaptic biomarker CSF GAP-43 (r = 0.609, P < 0.001) and neuroinflammatory biomarker CSF sTREM-2 (r = 0.507, P < 0.001). High CSF 14-3-3ζ levels at baseline were associated with progressive decline of cognitive function and neuroimaging findings during follow up. In conclusion, this study suggests that CSF 14-3-3ζ is a potential biomarker of AD that may be useful in clinical practice.

CSF 14-3-3ß is associated with progressive cognitive decline in Alzheimer's disease.

Alzheimer's disease is a neurodegenerative disorder characterized pathologically by amyloid-beta plaques, tau tangles and neuronal loss. In clinical practice, the 14-3-3 isoform beta (ß) is a biomarker that aids in the diagnosis of sporadic Creutzfeldt-Jakob disease. Recently, a proteomics study found increased CSF 14-3-3ß levels in Alzheimer's disease patients, suggesting a potential link between CSF 14-3-3ß and Alzheimer's disease. Our present study aimed to further investigate the role of CSF 14-3-3ß in Alzheimer's disease by analysing the data of 719 participants with available CSF 14-3-3ß measurements from the Alzheimer's Disease Neuroimaging Initiative. Higher CSF 14-3-3ß levels were observed in the mild cognitive impairment group compared to the cognitively normal group, with the highest CSF 14-3-3ß levels in the Alzheimer's disease dementia group. This study also found significant associations between CSF 14-3-3ß levels and CSF biomarkers of p-tau, t-tau, pTau/Aß42 ratios and GAP-43, as well as other Alzheimer's disease biomarkers such as Aß-PET. An early increase in CSF 14-3-3ß levels was observed prior to Aß-PET-positive status, and CSF 14-3-3ß levels continued to rise after crossing the Aß-PET positivity threshold before reaching a plateau. The diagnostic accuracy of CSF 14-3-3ß (area under the receiver operating characteristic curve = 0.819) was moderate compared to other established Alzheimer's disease biomarkers in distinguishing cognitively normal Aß pathology-negative individuals from Alzheimer's disease Aß pathology-positive individuals. Higher baseline CSF 14-3-3ß levels were associated with accelerated cognitive decline, reduced hippocampus volumes and declining fluorodeoxyglucose-PET values over a 4-year follow-up period. Patients with mild cognitive impairment and high CSF 14-3-3ß levels at baseline had a significantly increased risk [hazard ratio = 2.894 (1.599-5.238), P < 0.001] of progression to Alzheimer's disease dementia during follow-up. These findings indicate that CSF 14-3-3ß may be a potential biomarker for Alzheimer's disease and could provide a more comprehensive understanding of the underlying pathological changes of Alzheimer's disease, as well as aid in the diagnosis and monitoring of disease progression.

Complete SAMD12 repeat expansion sequencing in a four-generation BAFME1 family with anticipation.

Benign adult familial myoclonic epilepsy type 1 (BAFME1) is an autosomal dominant, adult-onset neurological disease caused by SAMD12 repeat expansion. In BAFME1, anticipation, such as the earlier onset of tremor and/or seizures in the next generation, was reported. This could be explained by intergenerational repeat instability, leading to larger expansions in successive generations. We report a four-generation BAFME1-affected family with anticipation. Using Nanopore long-read sequencing, detailed information regarding the sizes, configurations, and compositions of the expanded SAMD12 repeats across generations was obtained. Unexpectedly, a grandmother-mother-daughter triad showed similar repeat structures but with slight repeat expansions, despite quite variable age of onset of seizures (range: 52-14 years old), implying a complex relationship between the SAMD12 repeat expansion sequence and anticipation. This study suggests that different factor(s) from repeat expansion could modify the anticipation in BAFME1.

CSF GAP-43 as a biomarker of synaptic dysfunction is associated with tau pathology in Alzheimer's disease.

To test whether cerebrospinal fluid (CSF) growth-associated protein 43 (GAP-43) concentration is elevated in Alzheimer's disease (AD) dementia and its associations with other hallmarks of AD, we examined the CSF GAP-43 measurements of 787 participants (245 cognitively normal (CN), 415 individuals with mild cognitive impairment (MCI) and 127 individuals with AD dementia) from the Alzheimer's Disease Neuroimaging Initiative (ADNI) study. Associations were investigated between CSF GAP-43 and clinical diagnosis, Aß/tau/neurodegeneration (AT(N)) status, CSF and blood biomarkers of AD, cognitive measurements and brain neuroimaging findings. CSF GAP-43 levels were increased in patients with AD dementia (mean, 6331.05 pg/ml) compared with the CN (mean, 5001.05 pg/ml) and MCI (mean, 5118.8 pg/ml) (P < 0.001) groups. CSF GAP-43 correlated with CSF phosphorylated tau 181(p-tau) (r = 0.768, P < 0.001), and had high diagnostic accuracy in differentiating tau positive status vs. tau negative status (area under the receiver operating characteristic curve, 0.8606). CSF GAP-43 was particularly elevated among individuals with tau positive status. High CSF GAP-43 was associated with longitudinal deterioration of cognitive scores and brain neuroimaging findings. CSF GAP-43 was associated with a clinical diagnosis of AD dementia and with an individual's tau status, cognitive measurements and findings from neuroimaging. This study implies that CSF GAP-43 as a biomarker of synaptic dysfunction could predict the disease progression of AD patients.

Differential genetic associations and expression of PAPST1/SLC35B2 in bipolar disorder and schizophrenia.

Lithium's inhibitory effect on enzymes involved in sulfation process, such as inhibition of 3'(2')-phosphoadenosine 5'-phosphate (PAP) phosphatase, is a possible mechanism of its therapeutic effect for bipolar disorder (BD). 3'-Phosphoadenosine 5'-phosphosulfate (PAPS) is translocated from cytosol to Golgi lumen by PAPS transporter 1 (PAPST1/SLC35B2), where it acts as a sulfa donor. Since SLC35B2 was previously recognized as a molecule that facilitates the release of D-serine, a co-agonist of N-methyl-D-aspartate type glutamate receptor, altered function of SLC35B2 might be associated with the pathophysiology of BD and schizophrenia (SCZ). We performed genetic association analyses of the SLC35B2 gene using Japanese cohorts with 366 BD cases and 370 controls and 2012 SCZ cases and 2170 controls. We then investigated expression of SLC35B2 mRNA in postmortem brains by QPCR using a Caucasian cohort with 33 BD and 34 SCZ cases and 34 controls and by in situ hybridization using a Caucasian cohort with 37 SCZ and 29 controls. We found significant associations between three SNPs (rs575034, rs1875324, and rs3832441) and BD, and significantly reduced SLC35B2 mRNA expression in postmortem dorsolateral prefrontal cortex (DLPFC) of BD. Moreover, we observed normalized SLC35B2 mRNA expression in BD subgroups who were medicated with lithium. While there was a significant association of SLC35B2 with SCZ (SNP rs2233437), its expression was not changed in SCZ. These findings indicate that SLC35B2 might be differentially involved in the pathophysiology of BD and SCZ by influencing the sulfation process and/or glutamate system in the central nervous system.

A loss-of-function variant in SUV39H2 identified in autism-spectrum disorder causes altered H3K9 trimethylation and dysregulation of protocadherin ß-cluster genes in the developing brain.

Recent evidence has documented the potential roles of histone-modifying enzymes in autism-spectrum disorder (ASD). Aberrant histone H3 lysine 9 (H3K9) dimethylation resulting from genetic variants in histone methyltransferases is known for neurodevelopmental and behavioral anomalies. However, a systematic examination of H3K9 methylation dynamics in ASD is lacking. Here we resequenced nine genes for histone methyltransferases and demethylases involved in H3K9 methylation in individuals with ASD and healthy controls using targeted next-generation sequencing. We identified a novel rare variant (A211S) in the SUV39H2, which was predicted to be deleterious. The variant showed strongly reduced histone methyltransferase activity in vitro. In silico analysis showed that the variant destabilizes the hydrophobic core and allosterically affects the enzyme activity. The Suv39h2-KO mice displayed hyperactivity and reduced behavioral flexibility in learning the tasks that required complex behavioral adaptation, which is relevant for ASD. The Suv39h2 deficit evoked an elevated expression of a subset of protocadherin ß (Pcdhb) cluster genes in the embryonic brain, which is attributable to the loss of H3K9 trimethylation (me3) at the gene promoters. Reduced H3K9me3 persisted in the cerebellum of Suv39h2-deficient mice to an adult stage. Congruently, reduced expression of SUV39H1 and SUV39H2 in the postmortem brain samples of ASD individuals was observed, underscoring the role of H3K9me3 deficiency in ASD etiology. The present study provides direct evidence for the role of SUV39H2 in ASD and suggests a molecular cascade of SUV39H2 dysfunction leading to H3K9me3 deficiency followed by an untimely, elevated expression of Pcdhb cluster genes during early neurodevelopment.

Complete sequencing of expanded SAMD12 repeats by long-read sequencing and Cas9-mediated enrichment.

A pentanucleotide TTTCA repeat insertion into a polymorphic TTTTA repeat element in SAMD12 causes benign adult familial myoclonic epilepsy. Although the precise determination of the entire SAMD12 repeat sequence is important for molecular diagnosis and research, obtaining this sequence remains challenging when using conventional genomic/genetic methods, and even short-read and long-read next-generation sequencing technologies have been insufficient. Incomplete information regarding expanded repeat sequences may hamper our understanding of the pathogenic roles played by varying numbers of repeat units, genotype-phenotype correlations, and mutational mechanisms. Here, we report a new approach for the precise determination of the entire expanded repeat sequence and present a workflow designed to improve the diagnostic rates in various repeat expansion diseases. We examined 34 clinically diagnosed benign adult familial myoclonic epilepsy patients, from 29 families using repeat-primed PCR, Southern blot, and long-read sequencing with Cas9-mediated enrichment. Two cases with questionable results from repeat-primed PCR and/or Southern blot were confirmed as pathogenic using long-read sequencing with Cas9-mediated enrichment, resulting in the identification of pathogenic SAMD12 repeat expansions in 76% of examined families (22/29). Importantly, long-read sequencing with Cas9-mediated enrichment was able to provide detailed information regarding the sizes, configurations, and compositions of the expanded repeats. The inserted TTTCA repeat size and the proportion of TTTCA sequences among the overall repeat sequences were highly variable, and a novel repeat configuration was identified. A genotype-phenotype correlation study suggested that the insertion of even short (TTTCA)14 repeats contributed to the development of benign adult familial myoclonic epilepsy. However, the sizes of the overall TTTTA and TTTCA repeat units are also likely to be involved in the pathology of benign adult familial myoclonic epilepsy. Seven unsolved SAMD12-negative cases were investigated using whole-genome long-read sequencing, and infrequent, disease-associated, repeat expansions were identified in two cases. The strategic workflow resolved two questionable SAMD12-positive cases and two previously SAMD12-negative cases, increasing the diagnostic yield from 69% (20/29 families) to 83% (24/29 families). This study indicates the significant utility of long-read sequencing technologies to explore the pathogenic contributions made by various repeat units in complex repeat expansions and to improve the overall diagnostic rate.

Cooperation of LIM domain-binding 2 (LDB2) with EGR in the pathogenesis of schizophrenia.

Genomic defects with large effect size can help elucidate unknown pathologic architecture of mental disorders. We previously reported on a patient with schizophrenia and a balanced translocation between chromosomes 4 and 13 and found that the breakpoint within chromosome 4 is located near the LDB2 gene. We show here that Ldb2 knockout (KO) mice displayed multiple deficits relevant to mental disorders. In particular, Ldb2 KO mice exhibited deficits in the fear-conditioning paradigm. Analysis of the amygdala suggested that dysregulation of synaptic activities controlled by the immediate early gene Arc is involved in the phenotypes. We show that LDB2 forms protein complexes with known transcription factors. Consistently, ChIP-seq analyses indicated that LDB2 binds to > 10,000 genomic sites in human neurospheres. We found that many of those sites, including the promoter region of ARC, are occupied by EGR transcription factors. Our previous study showed an association of the EGR family genes with schizophrenia. Collectively, the findings suggest that dysregulation in the gene expression controlled by the LDB2-EGR axis underlies a pathogenesis of subset of mental disorders.

Role of an Atypical Cadherin Gene, Cdh23 in Prepulse Inhibition, and Implication of CDH23 in Schizophrenia.

We previously identified quantitative trait loci (QTL) for prepulse inhibition (PPI), an endophenotype of schizophrenia, on mouse chromosome 10 and reported Fabp7 as a candidate gene from an analysis of F2 mice from inbred strains with high (C57BL/6N; B6) and low (C3H/HeN; C3H) PPI levels. Here, we reanalyzed the previously reported QTLs with increased marker density. The highest logarithm of odds score (26.66) peaked at a synonymous coding and splice-site variant, c.753G>A (rs257098870), in the Cdh23 gene on chromosome 10; the c.753G (C3H) allele showed a PPI-lowering effect. Bayesian multiple QTL mapping also supported the same variant with a posterior probability of 1. Thus, we engineered the c.753G (C3H) allele into the B6 genetic background, which led to dampened PPI. We also revealed an e-QTL (expression QTL) effect imparted by the c.753G>A variant for the Cdh23 expression in the brain. In a human study, a homologous variant (c.753G>A; rs769896655) in CDH23 showed a nominally significant enrichment in individuals with schizophrenia. We also identified multiple potentially deleterious CDH23 variants in individuals with schizophrenia. Collectively, the present study reveals a PPI-regulating Cdh23 variant and a possible contribution of CDH23 to schizophrenia susceptibility.

Lipid Pathology of the Corpus Callosum in Schizophrenia and the Potential Role of Abnormal Gene Regulatory Networks with Reduced Microglial Marker Expression.

Structural changes in the corpus callosum have been reported in schizophrenia; however, the underlying molecular mechanism remains unclear. As the corpus callosum is high in lipid content, we analyzed the lipid contents of the corpora callosa from 15 patients with schizophrenia and 15 age- and sex-matched controls using liquid chromatography coupled to tandem mass spectrometry and identified lipid combinations associated with schizophrenia. Real-time quantitative polymerase chain reaction analyses using extended samples (schizophrenia, n = 95; control, n = 91) showed low expression levels of lipid metabolism-related genes and their potential upstream transcription factors in schizophrenia. Subsequent pathway analysis identified a gene regulatory network where nuclear factor of activated T cells 2 (NFATC2) is placed most upstream. We also observed low gene expression levels of microglial markers, inflammatory cytokines, and colony-stimulating factor 1 receptor (CSF1R), which is known to regulate the density of microglia, in the corpus callosum in schizophrenia. The interactions between CSF1R and several genes in the presently identified gene network originating from NFATC2 have been reported. Collectively, this study provides evidence regarding lipid abnormalities in the corpora callosa of patients with schizophrenia and proposes the potential role of impaired "NFATC2-relevant gene network-microglial axis" as its underlying mechanism.

Genetic risks of schizophrenia identified in a matched case-control study.

It has been suggested that dopaminergic neurotransmission plays important roles for the psychotic symptoms and probably etiology of schizophrenia. In our recent preliminary study, we demonstrated that the specific allele combinations of dopamine-related functional single nucleotide polymorphisms (SNPs), rs10770141, rs4680, and rs1800497 could indicate risks for schizophrenia. The present validation study involved a total of 2542 individuals who were age- and sex-matched in a propensity score matching analysis, and the results supported the statistical significances of the proposed genetic risks described in our previous reports. The estimated odds ratios were 1.24 (95% CI 1.06-1.45, p < 0.001) for rs4680, 1.73 (95% CI 1.47-2.02, p < 0.0001) for rs1800497, and 1.79 (95% CI 1.35-2.36, p < 0.0001) for rs10770141. A significant relationship was also revealed among these three polymorphisms and schizophrenia, with corresponding coefficients (p < 0.0001). In this study, we also present a new scoring model for the identification of individuals with the disease risks. Using the cut-off value of 2, our model exhibited sensitivity for almost two-thirds of all of the schizophrenia patients: odds ratio 1.87, 95% CI 1.59-2.19, p < 0.0001. In conclusion, we identified significant associations of dopamine-related genetic combinations with schizophrenia. These findings suggest that some types of dopaminergic neurotransmission play important roles for development of schizophrenia, and this type of approach may also be applicable for other multifactorial diseases, providing a potent new risk predictor.

Peroxisome proliferator-activated receptor α as a novel therapeutic target for schizophrenia.

BACKGROUND: The pathophysiology of schizophrenia, a major psychiatric disorder, remains elusive. In this study, the role of peroxisome proliferator-activated receptor (PPAR)/retinoid X receptor (RXR) families, belonging to the ligand-activated nuclear receptor superfamily, in schizophrenia, was analyzed. METHODS: The PPAR/RXR family genes were screened by exploiting molecular inversion probe (MIP)-based targeted next-generation sequencing (NGS) using the samples of 1,200 Japanese patients with schizophrenia. The results were compared with the whole-genome sequencing databases of the Japanese cohort (ToMMo) and the gnomAD. To reveal the relationship between PPAR/RXR dysfunction and schizophrenia, Ppara KO mice and fenofibrate (a clinically used PPARα agonist)-administered mice were assessed by performing behavioral, histological, and RNA-seq analyses. FINDINGS: Our findings indicate that c.209-2delA, His117Gln, Arg141Cys, and Arg226Trp of the PPARA gene are risk variants for schizophrenia. The c.209-2delA variant generated a premature termination codon. The three missense variants significantly decreased the activity of PPARα as a transcription factor in vitro. The Ppara KO mice exhibited schizophrenia-relevant phenotypes, including behavioral deficits and impaired synaptogenesis in the cerebral cortex. Oral administration of fenofibrate alleviated spine pathology induced by phencyclidine, an N-methyl-d-aspartate (NMDA) receptor antagonist. Furthermore, pre-treatment with fenofibrate suppressed the sensitivity of mice to another NMDA receptor antagonist, MK-801. RNA-seq analysis revealed that PPARα regulates the expression of synaptogenesis signaling pathway-related genes. INTERPRETATION: The findings of this study indicate that the mechanisms underlying schizophrenia pathogenesis involve PPARα-regulated transcriptional machinery and modulation of synapse physiology. Hence, PPARα can serve as a novel therapeutic target for schizophrenia.

A potential role of fatty acid binding protein 4 in the pathophysiology of autism spectrum disorder.

Autism spectrum disorder is a neurodevelopmental disorder characterized by difficulties in social communication and interaction, as well as repetitive and characteristic patterns of behaviour. Although the pathogenesis of autism spectrum disorder is unknown, being overweight or obesity during infancy and low weight at birth are known as risks, suggesting a metabolic aspect. In this study, we investigated adipose tissue development as a pathophysiological factor of autism spectrum disorder by examining the serum levels of adipokines and other metabolic markers in autism spectrum disorder children (n = 123) and typically developing children (n = 92) at 4-12 years of age. Among multiple measures exhibiting age-dependent trajectories, the leptin levels displayed different trajectory patterns between autism spectrum disorder and typically developing children, supporting an adipose tissue-dependent mechanism of autism spectrum disorder. Of particular interest, the levels of fatty acid binding protein 4 (FABP4) were significantly lower in autism spectrum disorder children than in typically developing subjects, at preschool age (4-6 years old: n = 21 for autism spectrum disorder and n = 26 for typically developing). The receiver operating characteristic curve analysis discriminated autism spectrum disorder children from typically developing children with a sensitivity of 94.4% and a specificity of 75.0%. We re-sequenced the exons of the FABP4 gene in a Japanese cohort comprising 659 autism spectrum disorder and 1000 control samples, and identified two rare functional variants in the autism spectrum disorder group. The Trp98Stop, one of the two variants, was transmitted to the proband from his mother with a history of depression. The disruption of the Fabp4 gene in mice evoked autism spectrum disorder-like behavioural phenotypes and increased spine density on apical dendrites of pyramidal neurons, which has been observed in the postmortem brains of autism spectrum disorder subjects. The Fabp4 knockout mice had an altered fatty acid composition in the cortex. Collectively, these results suggest that an 'adipo-brain axis' may underlie the pathophysiology of autism spectrum disorder, with FABP4 as a potential molecule for use as a biomarker.

LDB2 locus disruption on 4p16.1 as a risk factor for schizophrenia and bipolar disorder.

We had previously reported the case of a male patient with schizophrenia, having de-novo balanced translocation. Here, we determined the exact breakpoints in chromosomes 4 and 13. The breakpoint within chromosome 4 was mapped to a region 32.6 kbp upstream of the LDB2 gene encoding Lim domain binding 2. Variant screening in LDB2 revealed a rare novel missense variant in patients with psychiatric disorder.

VLDL-specific increases of fatty acids in autism spectrum disorder correlate with social interaction.

BACKGROUND: Abnormalities of lipid metabolism contributing to the autism spectrum disorder (ASD) pathogenesis have been suggested, but the mechanisms are not fully understood. We aimed to characterize the lipid metabolism in ASD and to explore a biomarker for clinical evaluation. METHODS: An age-matched case-control study was designed. Lipidomics was conducted using the plasma samples from 30 children with ASD compared to 30 typical developmental control (TD) children. Large-scale lipoprotein analyses were also conducted using the serum samples from 152 children with ASD compared to 122 TD children. Data comparing ASD to TD subjects were evaluated using univariate (Mann-Whitney test) and multivariate analyses (conditional logistic regression analysis) for main analyses using cofounders (diagnosis, sex, age, height, weight, and BMI), Spearman rank correlation coefficient, and discriminant analyses. FINDINGS: Forty-eight significant metabolites involved in lipid biosynthesis and metabolism, oxidative stress, and synaptic function were identified in the plasma of ASD children by lipidomics. Among these, increased fatty acids (FAs), such as omega-3 (n-3) and omega-6 (n-6), showed correlations with clinical social interaction score and ASD diagnosis. Specific reductions of very-low-density lipoprotein (VLDL) and apoprotein B (APOB) in serum of ASD children also were found by large-scale lipoprotein analysis. VLDL-specific reduction in ASD was correlated with APOB, indicating VLDL-specific dyslipidaemia associated with APOB in ASD children. INTERPRETATION: Our results demonstrated that the increases in FAs correlated positively with social interaction are due to VLDL-specific degradation, providing novel insights into the lipid metabolism underlying ASD pathophysiology. FUNDING: This study was supported mainly by MEXT, Japan.

Hippocampal sclerosis without visually detectable hippocampal MRI abnormalities: automated subfield volumetric analysis.

PURPOSE: This study aims to investigate hippocampal subfield volumes in patients with hippocampal sclerosis (HS) without visually detectable MRI abnormalities and to determine the diagnostic accuracy using hippocampal subfield volumes. MATERIALS AND METHODS: We examined 46 patients with unilateral HS who had a histopathological diagnosis, and 54 controls. The patients were divided into two groups; visually detectable HS (n = 26) and undetectable HS (n = 20) on MRI. The volumes of hippocampal subfield using FreeSurfer were compared among the three groups. Diagnostic accuracy was calculated as the AUC of ROC using cutoff values for each individual subfield. RESULTS: Compared with the controls, visually detectable HS showed significantly reduced volumes of all the hippocampal subfields and entire hippocampus, whereas visually undetectable HS showed significant atrophy only in the CA3 and hippocampus-amygdala-transition-area. To diagnose visually undetectable HS, the CA3 volumes had AUC of 0.719, which was higher than AUC of 0.614 based on the entire hippocampal volumes. CONCLUSION: Visually undetectable HS demonstrated volume reductions in the CA3. Further, the CA3 volumes was more useful to diagnose visually undetectable HS compared with the entire hippocampal volumes.

A recurrent PJA1 variant in trigonocephaly and neurodevelopmental disorders.

OBJECTIVE: Neurodevelopmental disorders (NDDs) often associate with epilepsy or craniofacial malformations. Recent large-scale DNA analyses identified hundreds of candidate genes for NDDs, but a large portion of the cases still remain unexplained. We aimed to identify novel candidate genes for NDDs. METHODS: We performed exome sequencing of 95 patients with NDDs including 51 with trigonocephaly and subsequent targeted sequencing of additional 463 NDD patients, functional analyses of variant in vitro, and evaluations of autism spectrum disorder (ASD)-like phenotypes and seizure-related phenotypes in vivo. RESULTS: We identified de novo truncation variants in nine novel genes; CYP1A1, C14orf119, FLI1, CYB5R4, SEL1L2, RAB11FIP2, ZMYND8, ZNF143, and MSX2. MSX2 variants have been described in patients with cranial malformations, and our present patient with the MSX2 de novo truncation variant showed cranial meningocele and partial epilepsy. MSX2 protein is known to be ubiquitinated by an E3 ubiquitin ligase PJA1, and interestingly we found a PJA1 hemizygous p.Arg376Cys variant recurrently in seven Japanese NDD patients; five with trigonocephaly and one with partial epilepsy, and the variant was absent in 886 Japanese control individuals. Pja1 knock-in mice carrying p.Arg365Cys, which is equivalent to p.Arg376Cys in human, showed a significant decrease in PJA1 protein amount, suggesting a loss-of-function effect of the variant. Pja1 knockout mice displayed moderate deficits in isolation-induced ultrasonic vocalizations and increased seizure susceptibility to pentylenetetrazole. INTERPRETATION: These findings propose novel candidate genes including PJA1 and MSX2 for NDDs associated with craniofacial abnormalities and/or epilepsy.