Methylation analysis for postpartum depression: a case control study.

BACKGROUND: Postpartum depression (PPD) is a major depressive disorder that occurs after childbirth. Objective diagnostic and predictive methods for PPD are important for early detection and appropriate intervention. DNA methylation has been recognized as a potential biomarker for major depressive disorder. In this study, we used methylation analysis and peripheral blood to search for biomarkers that could to lead to the development a predictive method for PPD. METHODS: Study participants included 36 pregnant women (18 cases and 18 controls determined after childbirth). Genome-wide DNA methylation profiles were obtained by analysis with an Infinium Human Methylation 450BeadChip. The association of DNA methylation status at each DNA methylation site with PPD was assessed using linear regression analysis. We also conducted functional enrichment analysis of PPD using The Database for Annotation, Visualization and Integrated Discovery 6.8 to explore enriched functional-related gene groups for PPD. RESULTS: In the analysis with postpartum depressed state as an independent variable, the difference in methylation frequency between the postpartum non-depressed group and the postpartum depressed group was small, and sites with genome-wide significant differences were not confirmed. After analysis by The Database for Annotation, Visualization and Integrated Discovery 6.8, we revealed four gene ontology terms, including axon guidance, related to postpartum depression. CONCLUSIONS: These findings may help with the development of an objective predictive method for PPD.

Author Correction: Proteomic analysis of lymphoblastoid cell lines from schizophrenic patients.

The original Article required a few updates; one co-author name, which was given as Hiroki Kiumura, has been updated to Hiroki Kimura. Furthermore, supplementary information has been updated, and grant numbers have been added. These updates have been made to both the PDF and HTML versions of this Article.

Proteomic analysis of lymphoblastoid cell lines from schizophrenic patients.

Although a number of studies have identified several convincing candidate genes or molecules, the pathophysiology of schizophrenia (SCZ) has not been completely elucidated. Therapeutic optimization based on pathophysiology should be performed as early as possible to improve functional outcomes and prognosis; to detect useful biomarkers for SCZ, which reflect pathophysiology and can be utilized for timely diagnosis and effective therapy. To explore biomarkers for SCZ, we employed fluorescence two-dimensional differential gel electrophoresis (2D-DIGE) of lymphoblastoid cell lines (LCLs) (1st sample set: 30 SCZ and 30 CON). Differentially expressed proteins were sequenced by liquid chromatography tandem-mass spectrometry (LC-MS/MS) and identified proteins were confirmed by western blotting (WB) (1st and 2nd sample set: 60 SCZ and 60 CON). Multivariate logistic regression analysis was performed to identify an optimal combination of biomarkers to create a prediction model for SCZ. Twenty protein spots were differentially expressed between SCZ and CON in 2D-DIGE analysis and 22 unique proteins were identified by LC-MS/MS. Differential expression of eight of 22 proteins was confirmed by WB. Among the eight candidate proteins (HSPA4L, MX1, GLRX3, UROD, MAPRE1, TBCB, IGHM, and GART), we successfully constructed logistic regression models comprised of 4- and 6-markers with good discriminative ability between SCZ and CON. In both WB and gene expression analysis of LCL, MX1 showed reproducibly significant associations. Moreover, Mx1 and its related proinflamatory genes (Mx2, Il1b, and Tnf) were also up-regulated in poly I:C-treated mice. Differentially expressed proteins might be associated with molecular pathophysiology of SCZ, including dysregulation of immunological reactions and potentially provide diagnostic and prognostic biomarkers.

Decline of Plasma Concentrations of Interleukin-18 in Severely Malnourished Patients with Anorexia Nervosa: Exploratory Analysis.

Multiple studies on the dynamics of inflammatory cytokines in patients with anorexia nervosa (AN) have been published, although results are not consistent among reports. Thus the pathophysiologic roles of these cytokines are not clear. We performed an exploratory analysis that included (1) comparisons of plasma interleukin-18 (IL-18) concentrations between patients with AN (n = 21) and healthy controls (n = 39), and (2) correlations between body mass index (BMI) and IL-18 concentrations in both groups, exploring the relationship between malnourishment and IL-18. Plasma IL-18 levels were significantly decreased in patients with AN compared with controls. Plasma IL-18 levels correlated to BMI in controls, but not in patients with AN. These results suggest that a decline in plasma IL-18 levels in patients with AN is not only due to malnourishment, but other pathophysiologic changes as well. IL-18 has a role in the brain's reaction to sadness and chronic stress. Therefore, decreased levels of IL-18 may commonly occur in patients with chronic AN.

Juvenile social defeat stress exposure persistently impairs social behaviors and neurogenesis.

Adverse juvenile experiences, including physical abuse, often have negative health consequences later in life. We investigated the influence of social defeat stress exposure as juveniles on neuropsychological behaviors, and the causal role of glucocorticoids in abnormal behaviors and impairment of neurogenesis in mice exposed to the stress. The juvenile (24-day-old) and adult (70-day-old) male C57BL/6J mice were exposed to social defeat stress induced by an aggressive ICR mouse. Social defeat stress exposure as juveniles, even for 1 day, induced persistent social avoidance to the unfamiliar ICR mouse in the social interaction test, but that was not observed in mice exposed to the stress as adults. Social avoidance by the stress exposure as juveniles for 10 consecutive days was observed, when the target mouse was not only unfamiliar ICR but also another C57BL/J mouse, but not an absent or an anesthetized ICR mouse. The stress exposure did not induce anxiety- and depression-like behaviors in spontaneous locomotor activity, elevated plus-maze test, marble-burying test, forced swimming test, or sucrose preference test. Serum corticosterone levels increased immediately after the stress exposure. The hippocampal neurogenesis was suppressed 1 day and 4 weeks after the stress exposure. Administration of mifepristone, a glucocorticoid receptor antagonist, prior to each stress exposure, blocked the persistent social avoidance and suppression of neurogenesis. In conclusion, social avoidance induced by social defeat stress exposure as juveniles are more persistent than that as adults. These social avoidances are associated with suppression of hippocampal neurogenesis via glucocorticoid receptors.

Mutation screening of GRIN2B in schizophrenia and autism spectrum disorder in a Japanese population.

N-methyl-d-aspartate receptors (NMDARs) play a critical role in excitatory synaptic transmission and plasticity in the central nervous systems. Recent genetics studies in schizophrenia (SCZ) show that SCZ is susceptible to NMDARs and the NMDAR signaling complex. In autism spectrum disorder (ASD), several studies report dysregulation of NMDARs as a risk factor for ASD. To further examine the association between NMDARs and SCZ/ASD development, we conducted a mutation screening study of GRIN2B which encodes NR2B subunit of NMDARs, to identify rare mutations that potentially cause diseases, in SCZ and ASD patients (n = 574 and 152, respectively). This was followed by an association study in a large sample set of SCZ, ASD, and normal healthy controls (n = 4145, 381, and 4432, respectively). We identified five rare missense mutations through the mutation screening of GRIN2B. Although no statistically significant association between any single mutation and SCZ or ASD was found, one of its variant, K1292R, is found only in the patient group. To further examine the association between mutations in GRIN2B and SCZ/ASD development, a larger sample size and functional experiments are needed.

Relationship between social support during pregnancy and postpartum depressive state: a prospective cohort study.

Although the association between social support and postpartum depression has been previously investigated, its causal relationship remains unclear. Therefore, we examined prospectively whether social support during pregnancy affected postpartum depression. Social support and depressive symptoms were assessed by Japanese version of Social Support Questionnaire (J-SSQ) and Edinburgh Postnatal Depression Scale (EPDS), among 877 pregnant women in early pregnancy and at one month postpartum. First, J-SSQ was standardized among peripartum women. The J-SSQ was found to have a two-factor structure, with Number and Satisfaction subscales, by exploratory and confirmatory factor analyses. Analysis of covariance was performed to examine how EPDS and J-SSQ scores during pregnancy affected the EPDS score at postpartum. Significant associations were found between postpartum EPDS score and both EPDS and total scores on the Number subscales during pregnancy (ß = 0.488 and -0.054, ps < 0.001). Specifically, this negative correlation was stronger in depressive than non-depressive groups. Meanwhile, total score on Satisfaction subscales was not significantly associated with postpartum EPDS score. These results suggest that having a larger number of supportive persons during pregnancy helps protect against postpartum depression, and that this effect is greater in depressive than non-depressive pregnant women. This finding is expected to be vitally important in preventive interventions.

Identification of Rare, Single-Nucleotide Mutations in NDE1 and Their Contributions to Schizophrenia Susceptibility.

BACKGROUND: Nuclear distribution E homolog 1 (NDE1), located within chromosome 16p13.11, plays an essential role in microtubule organization, mitosis, and neuronal migration and has been suggested by several studies of rare copy number variants to be a promising schizophrenia (SCZ) candidate gene. Recently, increasing attention has been paid to rare single-nucleotide variants (SNVs) discovered by deep sequencing of candidate genes, because such SNVs may have large effect sizes and their functional analysis may clarify etiopathology. METHODS AND RESULTS: We conducted mutation screening of NDE1 coding exons using 433 SCZ and 145 pervasive developmental disorders samples in order to identify rare single nucleotide variants with a minor allele frequency ≤5%. We then performed genetic association analysis using a large number of unrelated individuals (3554 SCZ, 1041 bipolar disorder [BD], and 4746 controls). Among the discovered novel rare variants, we detected significant associations between SCZ and S214F (P = .039), and between BD and R234C (P = .032). Furthermore, functional assays showed that S214F affected axonal outgrowth and the interaction between NDE1 and YWHAE (14-3-3 epsilon; a neurodevelopmental regulator). CONCLUSIONS: This study strengthens the evidence for association between rare variants within NDE1 and SCZ, and may shed light into the molecular mechanisms underlying this severe psychiatric disorder.

Resequencing and association analysis of PTPRA, a possible susceptibility gene for schizophrenia and autism spectrum disorders.

BACKGROUND: The PTPRA gene, which encodes the protein RPTP-α, is critical to neurodevelopment. Previous linkage studies, genome-wide association studies, controlled expression analyses and animal models support an association with both schizophrenia and autism spectrum disorders, both of which share a substantial portion of genetic risks. METHODS: We sequenced the protein-encoding areas of the PTPRA gene for single nucleotide polymorphisms or small insertions/deletions (InDel) in 382 schizophrenia patients. To validate their association with the disorders, rare (minor allele frequency <1%), missense mutations as well as one InDel in the 3'UTR region were then genotyped in another independent sample set comprising 944 schizophrenia patients, 336 autism spectrum disorders patients, and 912 healthy controls. RESULTS: Eight rare mutations, including 3 novel variants, were identified during the mutation-screening phase. In the following association analysis, L59P, one of the two missense mutations, was only observed among patients of schizophrenia. Additionally, a novel duplication in the 3'UTR region, 174620_174623dupTGAT, was predicted to be located within a Musashi Binding Element. MAJOR CONCLUSIONS: No evidence was seen for the association of rare, missense mutations in the PTPRA gene with schizophrenia or autism spectrum disorders; however, we did find some rare variants with possibly damaging effects that may increase the susceptibility of carriers to the disorders.

Factor structure of the Japanese version of the Edinburgh Postnatal Depression Scale in the postpartum period.

BACKGROUND: The Edinburgh Postnatal Depression Scale (EPDS) is a widely used screening tool for postpartum depression (PPD). Although the reliability and validity of EPDS in Japanese has been confirmed and the prevalence of PPD is found to be about the same as Western countries, the factor structure of the Japanese version of EPDS has not been elucidated yet. METHODS: 690 Japanese mothers completed all items of the EPDS at 1 month postpartum. We divided them randomly into two sample sets. The first sample set (n = 345) was used for exploratory factor analysis, and the second sample set was used (n = 345) for confirmatory factor analysis. RESULTS: The result of exploratory factor analysis indicated a three-factor model consisting of anxiety, depression and anhedonia. The results of confirmatory factor analysis suggested that the anxiety and anhedonia factors existed for EPDS in a sample of Japanese women at 1 month postpartum. The depression factor varies by the models of acceptable fit. CONCLUSIONS: We examined EPDS scores. As a result, "anxiety" and "anhedonia" exist for EPDS among postpartum women in Japan as already reported in Western countries. Cross-cultural research is needed for future research.

Novel rare variants in F-box protein 45 (FBXO45) in schizophrenia.

The ubiquitin ligase F-box protein 45 (FBXO45) is critical for synaptogenesis, neuronal migration, and synaptic transmission. FBXO45 is included in the 3q29 microdeletion region that confers a significant risk for schizophrenia, as shown by rare structural variant studies. Thus, FBXO45 is considered a prominent candidate for mediating schizophrenia pathogenesis. Here, we investigated rare, deleterious single nucleotide variants (SNVs) as well as small insertions and deletions (INDELs) in FBXO45 that may contribute to schizophrenia susceptibility. Using Sanger sequencing, we performed mutation screening in FBXO45 exon regions in 337 schizophrenia patients. Novel missense or nonsense variants were followed up with a genetic association study in an independent sample set of 601 schizophrenia patients and 916 controls, a case report for assessing the clinical consequence of the mutations, a pedigree study for measuring mutation inheritance in the proband's family, bioinformatics analyses for evaluating mutation effect on protein structure and function, and mRNA expression analysis for examining mutation transcriptional influence on FBXO45 expression. One heterozygous, novel, and rare missense mutation (R108C) was identified in a single schizophrenia patient and in his healthy mother. At age 20, this patient was diagnosed with paranoid schizophrenia and carried some clinical features of 3q29 deletion phenotypes, including premorbid IQ decline. With follow-up genotyping, this mutation was not found in either the schizophrenia group (0/601) or the healthy control group (0/916). Bioinformatics analyses predicted that R108C probably pathologically impacted the structure and function of the FBXO45 protein. The relative expression of FBXO45 in SCZ case with R108C mutation was relatively low when compared to 50 schizophrenia patients and 52 healthy controls. The R108C mutation in FBXO45 is a rare variant with a modest effect on schizophrenia risk that may disrupt the structure and function of the FBXO45 protein. Our findings also suggest that FBXO45 may be a new attractive candidate gene for schizophrenia.

Definition and refinement of the 7q36.3 duplication region associated with schizophrenia.

Using a very high-resolution oligonucleotide array for copy number variant (CNV) screening of samples comprising schizophrenic patients, we detected a novel CNV within the critical region (NCBI36/hg18, Chr7: 158,630,410-158,719,410) previously shown to be associated with schizophrenia. We investigated the association between the novel CNV identified in the current study and schizophrenia. Three independent samples were used: (1) Screening set, 300 Japanese schizophrenic patients (53.28 ± 14.66 years); (2) Confirmation set, 531 schizophrenic patients (46.03 ± 12.15 years); and (3) 711 healthy controls (47.12 ± 11.03 years). All subjects enrolled in the study were Japanese. Chromosomal position was determined using fluorescence in situ hybridization. We identified a novel duplication within the region associated with schizophrenia identified on 7q36.3 that is adjacent to VIPR2 and is not associated with schizophrenia. In the Japanese population, the 35-kb region that harbors the common, novel CNV should be excluded from the region associated with schizophrenia on 7q36.3.

Differential effects of diazepam, tandospirone, and paroxetine on plasma brain-derived neurotrophic factor level under mental stress.

OBJECTIVES: Serum brain-derived neurotrophic factor (BDNF) levels are reduced in depressed patients, and successful antidepressant treatment leads to increases in BDNF levels. However, little is known about how psychotropic drugs affect the mechanism of the human response to mental stress. We investigated the influence of psychotropic drugs on plasma BDNF levels under mental stress using a driving simulator (DS) task. METHODS: Fourteen healthy male volunteers received one of four drugs, diazepam (5 mg), tandospirone (20 mg), paroxetine (10 mg), and matched placebo, in a double-blind, crossover manner. Subjects were asked to perform the DS task 4 h post-dosing. Plasma BDNF levels were measured before and after the DS task. RESULTS: Plasma BDNF levels under the placebo, diazepam, and tandospirone conditions significantly decreased after the DS task compared with before the task. Conversely, no significant differences in plasma BDNF levels were detected under the paroxetine condition. CONCLUSION: As these three psychotropic drugs have differential effects on plasma BDNF levels under mental stress after 4 h post-dosing, antidepressants, unlike anxiolytics, might have a prompt positive effect on the mental stress response.

Potent inhibitors of amyloid ß fibrillization, 4,5-dianilinophthalimide and staurosporine aglycone, enhance degradation of preformed aggregates of mutant Notch3.

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is caused by mutations in human NOTCH3. We have recently reported that mutant Notch3 shows a greater propensity to form aggregates, and these aggregates resist degradation, leading to accumulation in the endoplasmic reticulum (ER). In this study, we searched for low-molecular compounds that decrease the amount of mutant Notch3 aggregates. Using a cell-based system, we found that degradation of preformed mutant aggregates was enhanced by treatment with either 4,5-dianilinophthalimide (DAPH) or staurosporine aglycone (SA), both of which inhibit amyloid ß (Aß) fibrillization. Regarding other low-molecular compounds interacting with Aß fibrils, thioflavin T (ThT) also enhanced the clearance of mutant Notch3. These findings suggest that DAPH, SA, and ThT are potent reagents to dissociate the preformed aggregates of mutant Notch3 by disruption of intermolecular contacts of misfolded proteins. Our study may provide the basis for the development of a pharmacological therapy for CADASIL.

Mutations in NOTCH3 cause the formation and retention of aggregates in the endoplasmic reticulum, leading to impaired cell proliferation.

Mutations in the human NOTCH3 gene cause cerebral autosomal-dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), but the pathogenic mechanisms of the disorder remain unclear. We investigated the cytotoxic properties of mutant Notch3 using stable cell lines with inducible expression of either wild-type or two mutants p.R133C and p.C185R. We found that both mutants of Notch3 were prone to aggregation and retained in the endoplasmic reticulum (ER). The turnover rates of the mutated Notch3 proteins were strikingly slow, with half-lives greater than 6 days, whereas wild-type Notch3 was rapidly degraded, with a half-life of 0.7 days. The expression of mutant Notch3 also impaired cell proliferation compared with wild-type Notch3. In addition, cell lines expressing mutant Notch3 were more sensitive to proteasome inhibition resulting in cell death. These findings suggest that prolonged retention of mutant Notch3 aggregates in the ER decreases cell growth and increases sensitivity to other stresses. It is also possible that the aggregate-prone property of mutant Notch3 contributes to a pathogenic mechanism underlying CADASIL.

Chronic stress-mutated presenilin 1 gene interaction perturbs neurogenesis and accelerates neurodegeneration.

Recent evidence suggests that supplemental factors coincident with aging and genetic determinants might be involved in the initial progression of Alzheimer's disease (AD). Early studies also indicate that chronic stress decreases hippocampal neurogenesis. Here, we investigate the effect of chronic stress on hippocampal neurogenesis using a transgenic mouse line (Tg) that overexpresses human presenilin 1 (PS1) with a familial AD (FAD)-related mutation in order to elucidate how the combination of chronic stress and mutated genes affects the cytoarchitecture in the hippocampal granule cell layer (GCL), which contributes to spatial learning and memory. Using an original chronic intermittent restraint stress (CIRS) protocol, we examined the effect of stress on hippocampal neurogenesis and neurodegeneration by immunohistochemical analysis. After short-term CIRS, neurodegeneration in Tg mice was significantly increased in the hippocampus with an earlier onset and progression than in the non-stressed Tg mice. Moreover, after long-term CIRS, transitional neurodegeneration appeared to proceed along the neuronal circuit involved in cognitive function in stressed Tg mice. Although the number of Pax6-positive (+) cells (mostly granule neuron precursors) did not significantly decrease during CIRS in both non-Tg and Tg mice, doublecortin (DCX) + neuronal progenitor cells in the GCL were markedly influenced in Tg mice; they were significantly reduced without stress compared with non-stressed non-Tg mice and significantly increased by CIRS compared with non-stressed Tg mice. We conclude from these results that diverse responses against stressful experiences among genetically predisposed individuals could lead to cognitive dysfunction through retardation of neuronal maturation and neurodegeneration.

Distribution of tenascin-c and -X in rat TMJ development.

In the neonatal and postnatal development of rat TMJ, tenascin-C and -X were detected in the muscle, bone matrix, connective tissue around the bone, and blood vessel of rats at E18 (18-days old embryo), 0-, and 5-days postnatal. The reaction of tenascin-X was also found in the connective tissue around the mandibular condyle. The mRNA of tenascin-C (600 bp) and -X (588 bp) was also detected in the developmental muscle with the level of tenascin-C mRNA moderately decreased during development. Therefore, tenascin-C and -X may have different effects on the connective tissue during development of TMJ.

Cholesteryl glucoside-induced protection against gastric ulcer.

The cytoprotective effect of heat shock proteins (HSPs) promises new therapeutic modalities for medical treatment. We examined the anti-ulcer effect of cholesteryl glucoside (1-O-cholesteryl-beta-D-glucopyranoside, CG) on cold-restraint stress-induced gastric ulcer in rats, in terms of its correlative ability to activate heat shock factor (HSF) and to induce HSP70. Rapid induction of CG occurred in animal tissues, especially in stomach, after exposure to stress, indicating that this glycolipid might act as an anti-stress, lipid mediator involved in the very early stages of stress-induced signal transduction. Orally administered CG apparently showed anti-ulcer activity in rats via HSF activation and HSP70 induction. When compared with geranylgeranylacetone (GGA), the well known as an effective, synthetic anti-ulcer agent, CG proved to have the same level of strength on ulcer inhibition. GGA caused CG and HSP70 induction in gastric mucosa, indicating that GGA induced HSP70 via CG production. CG thus might be useful for medical treatment of stress-induced diseases, and as an anti-stress supplement for daily diet.

Steryl glucoside is a lipid mediator in stress-responsive signal transduction.

We previously reported that steryl glucoside (SG) is rapidly induced in cells from molds to humans by exposure to environmental stress (Murakami-Murofushi et al. (1997) J. Biol. Chem., 272, 486-489, Kunimoto et al. (2000) Cell Stress & Chaperones, 5, 3-7), and in mold cells SG production is followed by activation of a certain protein kinase and induction of heat shock proteins (HSP) (Maruya et al. (1997) Cell Struct. Funct., 21, 533-538). To determine the biological significance of SG in stress responsive signal transduction, we added SG to the culture of human fibroblasts and examined its effect on HSP induction. We demonstrated a rapid activation of heat shock transcription factor 1 (HSF1) to bind to heat shock element (HSE) and induction of heat shock protein 70 (HSP70) in fibroblast cells by exposure to exogenously added human major SG, cholesteryl glucoside (CG). In addition, enzyme activity to form CG from cholesterol and UDP-glucose was detected in the homogenate of fibroblast cells. These results strongly suggest that CG acts as a mediator in the early stage of stress responsive signal transduction.