Autism is a pervasive neurodevelopmental disorder diagnosed in early childhood. The genetic factors might play an important role in its pathogenesis. Previous studies revealed that Reelin (RELN) polymorphisms were associated with autism. However, the roles of genes in Reelin signaling pathway for autism are largely unknown. As several knockout mice models in which the Reelin pathway genes (i.e. DAB1, VLDLR/APOER2, FYN/SRC and CRK/CRKL) are deficient have the similar phenotype as the reeler mice (Reelin(-/-)), we hypothesized that the Reelin signaling pathway genes might play roles in the etiology of autism. Therefore, we conducted a family-based association study. Sixty-two tagged single nucleotide polymorphisms (SNPs) covering 15 genes in Reelin pathway were genotyped in 239 trios, and 14 significant SNPs were further investigated in the additional 188 trios. In the total 427 trios, we found significant genetic association between autism and four SNPs in DAB1 (rs12035887 G: p=0.0006; rs3738556 G: p=0.0044; rs1202773 A: p=0.0048; rs12740765 T: p=0.0196). After the Bonferroni correction, SNP rs12035887 remained significant. Furthermore, the haplotype constructed with rs1202773 and rs12023109 in DAB1 showed significant excess transmission in both individual and global haplotype analyses (p=0.0052 and 0.0279, respectively). Our findings suggested that variations in DAB1 involved in the Reelin signaling pathway might contribute to genetic susceptibility to autism with Chinese Han decent, supporting the defect in the Reelin signaling pathway as a predisposition factor for autism.

BACKGROUND

Reelin plays a pivotal role in neurodevelopment and in post-natal synaptic plasticity and has been implicated in the pathogenesis of autism spectrum disorder (ASD). The reelin (RELN) gene expression is significantly decreased in ASD, both in the brain and peripherally. Methylation at the RELN gene promoter is largely triggered at puberty, and hypermethylation has been found in post-mortem brains of schizophrenic and bipolar patients.

METHODS

In this study, we assessed RELN gene methylation status in post-mortem temporocortical tissue samples (BA41/42 or 22) of six pairs of post-puberal individuals with ASD and typically developing subjects, matched for sex (male:female, M:F = 5:1), age, and post-mortem interval.

RESULTS

ASD patients display a significantly higher number of methylated CpG islands and heavier methylation in the 5' region of the RELN gene promoter, spanning from -458 to -223 bp, whereas controls have more methylated CpG positions and greater extent of methylation at the 3' promoter region, spanning from -222 to +1 bp. The most upstream promoter region (-458 to -364 bp) is methylated only in ASD brains, while the most downstream region (-131 to +1 bp) is methylated exclusively in control brains. Within this general framework, three different methylation patterns are discernible, each correlated with different extents of reduction in reelin gene expression among ASD individuals compared to controls.

CONCLUSIONS

The methylation pattern is different in ASD and control post-mortem brains. ASD-specific CpG positions, located in the most upstream gene promoter region, may exert a functional role potentially conferring ASD risk by blunting RELN gene expression.

Both Reelin (RELN) and glutamate decarboxylase 67 (GAD1) have been implicated in the pathophysiology of Autism Spectrum Disorders (ASD). We have previously shown that both mRNAs are reduced in the cerebella (CB) of ASD subjects through a mechanism that involves increases in the amounts of MECP2 binding to the corresponding promoters. In the current study, we examined the expression of RELN, GAD1, GAD2, and several other mRNAs implicated in this disorder in the frontal cortices (FC) of ASD and CON subjects. We also focused on the role that epigenetic processes play in the regulation of these genes in ASD brain. Our goal is to better understand the molecular basis for the down-regulation of genes expressed in GABAergic neurons in ASD brains. We measured mRNA levels corresponding to selected GABAergic genes using qRT-PCR in RNA isolated from both ASD and CON groups. We determined the extent of binding of MECP2 and DNMT1 repressor proteins by chromatin immunoprecipitation (ChIP) assays. The amount of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) present in the promoters of the target genes was quantified by methyl DNA immunoprecipitation (MeDIP) and hydroxyl MeDIP (hMeDIP). We detected significant reductions in the mRNAs associated with RELN and GAD1 and significant increases in mRNAs encoding the Ten-eleven Translocation (TET) enzymes 1, 2, and 3. We also detected increased MECP2 and DNMT1 binding to the corresponding promoter regions of GAD1, RELN, and GAD2. Interestingly, there were decreased amounts of 5mC at both promoters and little change in 5hmC content in these same DNA fragments. Our data demonstrate that RELN, GAD1, and several other genes selectively expressed in GABAergic neurons, are down-regulated in post-mortem ASD FC. In addition, we observed increased DNMT1 and MECP2 binding at the corresponding promoters of these genes. The finding of increased MECP2 binding to the RELN, GAD1 and GAD2 promoters, with reduced amounts of 5mC and unchanged amounts of 5hmC present in these regions, suggests the possibility that DNMT1 interacts with and alters MECP2 binding properties to selected promoters. Comparisons between data obtained from the FC with CB studies showed some common themes between brain regions which are discussed.

Involvement of life stress in Late-Onset Alzheimer's Disease (LOAD) has been evinced in longitudinal cohort epidemiological studies, and endocrinologic evidence suggests involvements of catecholamine and corticosteroid systems in LOAD. Early Life Stress (ELS) rodent models have successfully demonstrated sequelae of maternal separation resulting in LOAD-analogous pathology, thereby supporting a role of insulin receptor signalling pertaining to GSK-3beta facilitated tau hyper-phosphorylation and amyloidogenic processing. Discussed are relevant ELS studies, and findings from three mitogen-activated protein kinase pathways (JNK/SAPK pathway, ERK pathway, p38/MAPK pathway) relevant for mediating environmental stresses. Further considered were the roles of autophagy impairment, neuroinflammation, and brain insulin resistance. For the meta-analytic evaluation, 224 candidate gene loci were extracted from reviews of animal studies of LOAD pathophysiological mechanisms, of which 60 had no positive results in human LOAD association studies. These loci were combined with 89 gene loci confirmed as LOAD risk genes in previous GWAS and WES. Of the 313 risk gene loci evaluated, there were 35 human reports on epigenomic modifications in terms of methylation or histone acetylation. 64 microRNA gene regulation mechanisms were published for the compiled loci. Genomic association studies support close relations of both noradrenergic and glucocorticoid systems with LOAD. For HPA involvement, a CRHR1 haplotype with MAPT was described, but further association of only HSD11B1 with LOAD found; however, association of FKBP1 and NC3R1 polymorphisms was documented in support of stress influence to LOAD. In the brain insulin system, IGF2R, INSR, INSRR, and plasticity regulator ARC, were associated with LOAD. Pertaining to compromised myelin stability in LOAD, relevant associations were found for BIN1, RELN, SORL1, SORCS1, CNP, MAG, and MOG. Regarding epigenetic modifications, both methylation variability and de-acetylation were reported for LOAD. The majority of up-to-date epigenomic findings include reported modifications in the well-known LOAD core pathology loci MAPT, BACE1, APP (with FOS, EGR1), PSEN1, PSEN2, and highlight a central role of BDNF. Pertaining to ELS, relevant loci are FKBP5, EGR1, GSK3B; critical roles of inflammation are indicated by CRP, TNFA, NFKB1 modifications; for cholesterol biosynthesis, DHCR24; for myelin stability BIN1, SORL1, CNP; pertaining to (epi)genetic mechanisms, hTERT, MBD2, DNMT1, MTHFR2. Findings on gene regulation were accumulated for BACE1, MAPK signalling, TLR4, BDNF, insulin signalling, with most reports for miR-132 and miR-27. Unclear in epigenomic studies remains the role of noradrenergic signalling, previously demonstrated by neuropathological findings of childhood nucleus caeruleus degeneration for LOAD tauopathy.

Glioblastoma (GBM) represents the most common and most malignant type of primary brain tumor and significantly contributes to cancer morbidity and mortality. Invasion into the healthy brain parenchyma is a major feature of glioblastoma aggressiveness. Reelin (RELN) is a large secreted extracellular matrix glycoprotein that regulates neuronal migration and positioning in the developing brain and sustains functionality in the adult brain. We here show that both RELN and its main downstream effector DAB1 are silenced in glioblastoma as compared to non-neoplastic tissue and mRNA expression is inversely correlated with malignancy grade. Furthermore, RELN expression is positively correlated with patient survival in two large, independent clinically annotated datasets. RELN silencing occurs via promoter hypermethylation as shown by both database mining and bisulfite sequencing of the RELN promoter. Consequently, treatment with 5'-Azacytidine and trichostatin A induced RELN expression in vitro. On the functional level, we found RELN to regulate glioblastoma cell migration both in a DAB1 (tyrosine phosphorylation)-dependent and -independent fashion, depending on the substrate provided. Moreover, stimulation of RELN signaling strongly reduced proliferation in glioblastoma cells. This phenotype depends on DAB1 stimulation by RELN, as a mutant that lacks all RELN induced tyrosine phosphorylation sites (DAB1-5F) failed to induce a growth arrest. Proteomic analyzes revealed that these effects are mediated by a reduction in E2F targets and dephosphorylation of ERK1/2. Taken together, our data establish a relevance of RELN signaling in glioblastoma pathology and thereby might unearth novel, yet unrecognized treatment options.

Schizophrenia is a common disorder with a high heritability, but its genetic architecture is still elusive. We implemented whole-genome sequencing (WGS) analysis of 8 families with monozygotic (MZ) twin pairs discordant for schizophrenia to assess potential association of de novo mutations (DNMs) or inherited variants with susceptibility to schizophrenia. Eight non-synonymous DNMs (including one splicing site) were identified and shared by twins, which were either located in previously reported schizophrenia risk genes (p.V24689I mutation in TTN, p.S2506T mutation in GCN1L1, IVS3+1G>> T in DOCK1) or had a benign to damaging effect according to in silico prediction analysis. By searching the inherited rare damaging or loss-of-function (LOF) variants and common susceptible alleles from three classes of schizophrenia candidate genes, we were able to distill genetic alterations in several schizophrenia risk genes, including GAD1, PLXNA2, RELN and FEZ1. Four inherited copy number variations (CNVs; including a large deletion at 16p13.11) implicated for schizophrenia were identified in four families, respectively. Most of families carried both missense DNMs and inherited risk variants, which might suggest that DNMs, inherited rare damaging variants and common risk alleles together conferred to schizophrenia susceptibility. Our results support that schizophrenia is caused by a combination of multiple genetic factors, with each DNM/variant showing a relatively small effect size.

A large body of genetic data from schizophrenia-related research has identified an assortment of genes and disturbed pathways supporting involvement of complex genetic components for schizophrenia spectrum and other psychotic disorders. Advances in genetic technology and expanding studies with searchable genomic databases have led to multiple published reports, allowing us to compile a master list of known, clinically relevant, or susceptibility genes contributing to schizophrenia. We searched key words related to schizophrenia and genetics from peer-reviewed medical literature sources, authoritative public access psychiatric websites and genomic databases dedicated to gene discovery and characterization of schizophrenia. Our list of 560 genes were arranged in alphabetical order in tabular form with gene symbols placed on high-resolution human chromosome ideograms. Genome wide pathway analysis using GeneAnalytics was carried out on the resulting list of genes to assess the underlying genetic architecture for schizophrenia. Recognized genes of clinical relevance, susceptibility or causation impact a broad range of biological pathways and mechanisms including ion channels (e.g., CACNA1B, CACNA1C, CACNA1H), metabolism (e.g., CYP1A2, CYP2C19, CYP2D6), multiple targets of neurotransmitter pathways impacting dopamine, GABA, glutamate, and serotonin function, brain development (e.g., NRG1, RELN), signaling peptides (e.g., PIK3CA, PIK4CA) and immune function (e.g., HLA-DRB1, HLA-DQA1) and interleukins (e.g., IL1A, IL10, IL6). This summary will enable clinical and laboratory geneticists, genetic counselors, and other clinicians to access convenient pictorial images of the distribution and location of contributing genes to inform diagnosis and gene-based treatment as well as provide risk estimates for genetic counseling of families with affected relatives.

Otosclerosis is a common form of hearing loss characterized by abnormal bone remodeling in the otic capsule. It is considered a complex disease caused by both genetic and environmental factors. In a previous study, we identified a region on chr7q22.1 located in the RELN gene that is associated with otosclerosis in Belgian-Dutch and French populations. Evidence for allelic heterogeneity was found in this chromosomal region in the form of two independent signals. To confirm this finding, we have completed a replication study that includes four additional populations from Europe (1,141 total samples). Several SNPs in this region replicated in these populations separately. While the power to detect significant association in each population is small, when all four populations are combined, six of seven SNPs replicate and show an effect in the same direction as in the previous populations. We also confirmed the presence of allelic heterogeneity in this region. These data further implicate RELN in the pathogenesis of otosclerosis. Functional research is warranted to determine the pathways through which RELN acts in the pathogenesis of otosclerosis.

Single-cell RNA sequencing (scRNA-seq) has significantly deepened our insights into complex tissues, with the latest techniques capable of processing tens of thousands of cells simultaneously. Analyzing increasing numbers of cells, however, generates extremely large data sets, extending processing time and challenging computing resources. Current scRNA-seq analysis tools are not designed to interrogate large data sets and often lack sensitivity to identify marker genes. With bigSCale, we provide a scalable analytical framework to analyze millions of cells, which addresses the challenges associated with large data sets. To handle the noise and sparsity of scRNA-seq data, bigSCale uses large sample sizes to estimate an accurate numerical model of noise. The framework further includes modules for differential expression analysis, cell clustering, and marker identification. A directed convolution strategy allows processing of extremely large data sets, while preserving transcript information from individual cells. We evaluated the performance of bigSCale using both a biological model of aberrant gene expression in patient-derived neuronal progenitor cells and simulated data sets, which underlines the speed and accuracy in differential expression analysis. To test its applicability for large data sets, we applied bigSCale to assess 1.3 million cells from the mouse developing forebrain. Its directed down-sampling strategy accumulates information from single cells into index cell transcriptomes, thereby defining cellular clusters with improved resolution. Accordingly, index cell clusters identified rare populations, such as reelin (Reln)-positive Cajal-Retzius neurons, for which we report previously unrecognized heterogeneity associated with distinct differentiation stages, spatial organization, and cellular function. Together, bigSCale presents a solution to address future challenges of large single-cell data sets.

The role of epigenetic regulators in the control of adult neurogenesis is largely undefined. We show that the histone demethylase enzyme Kdm5b (Jarid1b) negatively regulates neurogenesis from adult subventricular zone (SVZ) neural stem cells (NSCs) in culture. shRNA-mediated depletion of Kdm5b in proliferating adult NSCs decreased proliferation rates and reduced neurosphere formation in culture. When transferred to differentiation culture conditions, Kdm5b-depleted adult NSCs migrated from neurospheres with increased velocity. Whole-genome expression screening revealed widespread transcriptional changes with Kdm5b depletion, notably the up-regulation of reelin (Reln), the inhibition of steroid biosynthetic pathway component genes and the activation of genes with intracellular transport functions in cultured adult NSCs. Kdm5b depletion increased extracellular reelin concentration in the culture medium and increased phosphorylation of the downstream reelin signaling target Disabled-1 (Dab1). Sequestration of extracellular reelin with CR-50 reelin-blocking antibodies suppressed the increase in migratory velocity of Kdm5b-depleted adult NSCs. Chromatin immunoprecipitation revealed that Kdm5b is present at the proximal promoter of Reln, and H3K4me3 methylation was increased at this locus with Kdm5b depletion in differentiating adult NSCs. Combined the data suggest Kdm5b negatively regulates neurogenesis and represses Reln in neural stem cells from the adult SVZ.

BACKGROUND

Invasive lobular carcinoma (ILC) comprises approximately ~10-20% of breast cancers. In general, multifocal/multicentric (MF/MC) breast cancer has been associated with an increased rate of regional lymph node metastases. Tumor heterogeneity between foci represents a largely unstudied source of genomic variation in those rare patients with MF/MC ILC.

METHODS

We characterized gene expression and copy number in 2 or more foci from 11 patients with MF/MC ILC (all ER+, HER2-) and adjacent normal tissue. RNA and DNA were extracted from 3x1.5 mm cores from all foci. Gene expression (730 genes) and copy number (80 genes) were measured using Nanostring PanCancer and Cancer CNV panels. Linear mixed models were employed to compare expression in tumor versus normal samples from the same patient, and to assess heterogeneity (variability) in expression among multiple ILC within an individual.

RESULTS

35 and 34 genes were upregulated (FC>2) and down-regulated (FC<0.5) respectively in ILC tumor relative to adjacent normal tissue, q<0.05. 9/34 down-regulated genes (FIGF, RELN, PROM1, SFRP1, MMP7, NTRK2, LAMB3, SPRY2, KIT) had changes larger than CDH1, a hallmark of ILC. Copy number changes in these patients were relatively few but consistent across foci within each patient. Amplification of three genes (CCND1, FADD, ORAOV1) at 11q13.3 was present in 2/11 patients in both foci. We observed significant evidence of within-patient between-foci variability (heterogeneity) in gene expression for 466 genes (p<0.05 with FDR 8%), including CDH1, FIGF, RELN, SFRP1, MMP7, NTRK2, LAMB3, SPRY2 and KIT.

CONCLUSIONS

There was substantial variation in gene expression between ILC foci within patients, including known markers of ILC, suggesting an additional level of complexity that should be addressed.

BACKGROUND

Meningioma tumors arise in arachnoid membranes, and are the most reported central nervous system (CNS) tumors worldwide. Up to 20% of grade I meningioma tumors reoccur and currently predictive cancer stem cells (CSCs) markers for aggressive and drug resistant meningiomas are scarce.

METHODS

Meningioma tissues and primary cell lines were investigated using whole transcriptome microarray analysis, immunofluorescence staining of CSCs markers (including CD133, Sox2, Nestin, and Frizzled 9), and drug treatment with cisplatin or etoposide.

RESULTS

Unsupervised hierarchical clustering of six meningioma samples separated tissues into two groups. Analysis identified stem cells related pathways to be differential between the two groups and indicated the de-regulation of the stem cell associated genes Reelin (RELN), Calbindin 1 (CALB1) and Anterior Gradient 2 Homolog (AGR2). Immunofluorescence staining for four tissues confirmed stemness variation in situ. Biological characterization of fifteen meningioma primary cell lines concordantly separated cells into two functionally distinct sub-groups. Pleomorphic cell lines (NG type) grew significantly faster than monomorphic cell lines (G type), had a higher number of cells that express Ki67, and were able to migrate aggressively in vitro. In addition, NG type cell lines had a lower expression of nuclear Caspase-3, and had a significantly higher number of CSCs co-positive for CD133+ Sox2+ or AGR2+ BMI1+. Importantly, these cells were more tolerant to cisplatin and etoposide treatment, showed a lower level of nuclear Caspase-3 in treated cells and harbored drug resistant CSCs.

CONCLUSIONS

Collectively, analyses of tissues and primary cell lines revealed stem cell associated genes as potential targets for aggressive and drug resistant meningiomas.

Over half a century ago, D. S. Falconer first reported a mouse with a reeling gate. Four decades later, the Reln gene was isolated and identified as the cause of the reeler phenotype. Initial studies found that loss of Reelin, a large, secreted glycoprotein encoded by the Reln gene, results in abnormal neuronal layering throughout several regions of the brain. In the years since, the known functions of Reelin signaling in the brain have expanded to include multiple postdevelopmental neuromodulatory roles, revealing an ever increasing body of evidence to suggest that Reelin signaling is a critical player in the modulation of synaptic function. In writing this review, we intend to highlight the most fundamental aspects of Reelin signaling and integrate how these various neuromodulatory effects shape and protect synapses.

Overconsumption of high-fat diets (HFDs) can critically affect synaptic and cognitive functions within telencephalic structures such as the medial prefrontal cortex (mPFC). The underlying mechanisms, however, remain largely unknown. Here we show that adolescence is a sensitive period for the emergence of prefrontal cognitive deficits in response to HFD. We establish that the synaptic modulator reelin (RELN) is a critical mediator of this vulnerability because (1) periadolescent HFD (pHFD) selectively downregulates prefrontal RELN+ cells and (2) augmenting mPFC RELN levels using transgenesis or prefrontal pharmacology prevents the pHFD-induced prefrontal cognitive deficits. We further identify N-methyl-d-aspartate-dependent long-term depression (NMDA-LTD) at prefrontal excitatory synapses as a synaptic signature of this association because pHFD abolishes NMDA-LTD, a function that is restored by RELN overexpression. We believe this study provides the first mechanistic insight into the vulnerability of the adolescent mPFC towards nutritional stress, such as HFDs. Our findings have primary relevance to obese individuals who are at an increased risk of developing neurological cognitive comorbidities, and may extend to multiple neuropsychiatric and neurological disorders in which RELN deficiency is a common feature.

In this study, a genetically diverse panel of 43 mouse strains was exposed to phosgene and genome-wide association mapping performed using a high-density single nucleotide polymorphism (SNP) assembly. Transcriptomic analysis was also used to improve the genetic resolution in the identification of genetic determinants of phosgene-induced acute lung injury (ALI). We prioritized the identified genes based on whether the encoded protein was previously associated with lung injury or contained a nonsynonymous SNP within a functional domain. Candidates were selected that contained a promoter SNP that could alter a putative transcription factor binding site and had variable expression by transcriptomic analyses. The latter two criteria also required that ≥10% of mice carried the minor allele and that this allele could account for ≥10% of the phenotypic difference noted between the strains at the phenotypic extremes. This integrative, functional approach revealed 14 candidate genes that included Atp1a1, Alox5, Galnt11, Hrh1, Mbd4, Phactr2, Plxnd1, Ptprt, Reln, and Zfand4, which had significant SNP associations, and Itga9, Man1a2, Mapk14, and Vwf, which had suggestive SNP associations. Of the genes with significant SNP associations, Atp1a1, Alox5, Plxnd1, Ptprt, and Zfand4 could be associated with ALI in several ways. Using a competitive electrophoretic mobility shift analysis, Atp1a1 promoter (rs215053185) oligonucleotide containing the minor G allele formed a major distinct faster-migrating complex. In addition, a gene with a suggestive SNP association, Itga9, is linked to transforming growth factor β1 signaling, which previously has been associated with the susceptibility to ALI in mice.

Reelin (RELN) is a key molecule for the regulation of neuronal migration in the developing CNS. The reeler mice, which have spontaneous autosomal recessive mutation in the RELN gene, reveal multiple defects in brain development. Morphological, neurochemical and behavioral alterations have been detected in heterozygous reeler (HR) mice, suggesting that not only the presence, but also the level of RELN influences brain development. Several studies implicate an involvement of RELN in the pathophysiology of neuropsychiatric disorders in which an alteration of the cholinergic cortical pathways is implicated as well. Thus, we decided to investigate whether the basal forebrain (BF) cholinergic system is altered in HR mice by examining cholinergic markers at the level of both cell body and nerve terminals. In septal and rostral, but not caudal, basal forebrain region, HR mice exhibited a significant reduction in the number of choline acetyltransferase (ChAT) immunoreactive (ir) cell bodies compared with control mice. Instead, an increase in ChAT ir neurons was detected in lateral striatum. This suggests that an alteration in ChAT ir cell migration which leads to a redistribution of cholinergic neurons in subcortical forebrain regions occurs in HR mice. The reduction of ChAT ir neurons in the BF was paralleled by an alteration of cortical cholinergic nerve terminals. In particular, the HR mice presented a marked reduction of acetylcholinesterase (AChE) staining accompanied by a small reduction of cortical thickness in the rostral dorsomedial cortex, while the density of AChE staining was not altered in the lateral and ventral cortices. Present results show that the cholinergic basalo-cortical system is markedly, though selectively, impaired in HR mice. Rostral sub-regions of the BF and rostro-medial cortical areas show significant decreases of cholinergic neurons and innervation, respectively.

Neuronal network remodeling during critical periods of sensory development might be accompanied by alterations in hypothalamic cell populations. MicroRNAs play a central role in regulating neuronal function, including neural stem cell proliferation, and neuronal migration, maturation and integration into viable circuits by modulating different mRNA targets. Here we investigated the role of miR-138 in cell proliferation and migration in a neuron-enriched hypothalamic cell culture prepared from chicks on embryonic day 16. Ectopic expression of miR-138 enhanced hypothalamic cell migration, but did not affect cell proliferation. As a potential mechanism for miR-138's effect on cell migration, we investigated reelin (Reln) as a direct target of miR-138. Luciferase reporter assay and Ago2-immunoprecipitation experiments confirmed direct binding of miR-138 to the Reln 3'-untranslated region. Ectopic miR-138 abolished Reln levels in hypothalamic cells and enhanced their migration, similar to Reln-antisense DNA. Furthermore, inhibition of Reln expression by miR-138 led to decreased phosphorylation level of the key component of Reln-regulated signaling cascades, Disabled 1. These findings describe miR-138 as a novel regulator of hypothalamic cell migration, acting at least in part via inhibition of Reln expression and leading to the inactivation of Reln signals.

Reelin (RELN) is a large glycoprotein that plays critical roles during and after neuronal development. In a previous study utilizing developing zebrafish, the RELN ortholog was in a subset of genes associated with neuronal function that was altered following exposure to 100ppb Pb at 72h post-fertilization (hpf). Although RELN is associated with neurodevelopment and neurological diseases, it represents a previously uncharacterized target for developmental Pb neurotoxicity. To further investigate Pb-induced expression decrease of RELN, a more detailed analysis was done using in situ hybridization and quantitative PCR (qPCR) on developing zebrafish at multiple developmental time points and at different exposure concentrations. Time points included in the analysis were 24, 36, 48, 60, 72 and 96hpf. The four exposure conditions tested were 10, 50 or 100ppb Pb as well as a control. In situ hybridization demonstrated concentrated expression of the gene in the telencephalon and in midbrain, hindbrain and spinal cord. Expression was less spatially restricted early in development but became isolated to specific cell populations in the central nervous system by 48hpf. The telencephalon demonstrated a high degree of expression throughout development. Pb exposure did not affect the observed location of expression at any of the time points tested. However, qPCR analysis demonstrated that reln expression statistically decreased in embryos exposed to the highest concentration compared to controls at 60hpf. No changes in brain morphology as assessed by measuring brain length and no significant increase in the number of apoptotic cells in the brain were observed at the highest exposure concentration. These data demonstrate that a decrease in reln expression occurs following Pb exposure at a concentration below one that results in changes in brain length or in the number of apoptotic cells in the zebrafish model system during a short window and is likely linked to concurrent developmental processes.

The spreading of neurofibrillary tangles (NFTs), intraneuronal aggregates of highly phosphorylated microtubule-associated protein tau, across the human brain is correlated with the cognitive severity of Alzheimer's disease (AD). To identify genes relevant to NFT expansion defined by the Braak stage, we conducted whole-genome exon array analysis with an exploratory sample set consisting of 213 human post-mortem brain tissue specimens from the entorinal, temporal and frontal cortices of 71 brain-donor subjects: Braak NFT stages 0 (N=13), I-II (N=20), III-IV (N=19) and V-VI (N=19). We identified eight genes, RELN, PTGS2, MYO5C, TRIL, DCHS2, GRB14, NPAS4 and PHYHD1, associated with the Braak stage. The expression levels of three genes, PHYHD1, MYO5C and GRB14, exhibited reproducible association on real-time quantitative PCR analysis. In another sample set, including control subjects (N=30), and in patients with late-onset AD (N=37), dementia with Lewy bodies (N=17) and Parkinson disease (N=36), the expression levels of two genes, PHYHD1 and MYO5C, were obviously associated with late-onset AD. Protein-protein interaction network analysis with a public database revealed that PHYHD1 interacts with MYO5C via POT1, and PHYHD1 directly interacts with amyloid beta-peptide 42. It is thus likely that functional failure of PHYHD1 and MYO5C could lead to AD development.

Sizn1 (Zcchc12) is a transcriptional co-activator that positively modulates bone morphogenic protein (BMP) signaling through its interaction with Smad family members and CBP. We have demonstrated a role for Sizn1 in basal forebrain cholinergic neuron specific gene expression. Furthermore, mutations in SIZN1 have been associated with X-linked mental retardation. Given the defined role of SIZN1 in mental retardation, knowing its complete forebrain expression pattern is essential to further elucidating its role in cognition. To better define the dynamic expression pattern of Sizn1 during forebrain development, we investigated its expression in mouse brain development from embryonic day 8.0 (E8.0) to adult. We found that Sizn1 is primarily restricted to the ventral forebrain including the medial ganglionic eminence, the septum, amygdala, and striatum. In addition, Sizn1 expression is detected in the cortical hem and pallial-subpallial boundary (PSB; anti-hem); both sources of Cajal-Retzius cells. Sizn1 expression in the dorsal forebrain is restricted to a subset of cells in the marginal zone that also express Reln, indicative of Cajal-Retzius cells. These data provide novel information on brain regions and cell types that express Sizn1, facilitating further investigations into the function of Sizn1 in both development and the pathogenesis of mental retardation.

BACKGROUND

The purpose of this study was to address the affects of mood modifying drugs on the transcriptome, in a tissue culture model, using qPCR arrays as a cost effective approach to identifying regulatory networks and pathways that might coordinate the cell response to a specific drug.

METHODS

We addressed the gene expression profile of 90 plus genes associated with human mood disorders using the StellARray™ qPCR gene expression system in the human derived SH-SY5Y neuroblastoma cell line.

RESULTS

Global Pattern Recognition (GPR) analysis identified a total of 9 genes (DRD3(⁎), FOS(†), JUN(⁎), GAD1(⁎†), NRG1(⁎), PAFAH1B3(⁎), PER3(⁎), RELN(⁎) and RGS4(⁎)) to be significantly regulated in response to cellular challenge with the mood stabilisers sodium valproate ((⁎)) and lithium ((†)). Modulation of FOS and JUN highlights the importance of the activator protein 1 (AP-1) transcription factor pathway in the cell response. Enrichment analysis of transcriptional networks relating to this gene set also identified the transcription factor neuron restrictive silencing factor (NRSF) and the oestrogen receptor as an important regulatory mechanism.

CONCLUSIONS

Cell line models offer a window of what might happen in vivo but have the benefit of being human derived and homogenous with regard to cell type.

CONCLUSIONS

This data highlights transcription factor pathways, acting synergistically or separately, in the modulation of specific neuronal gene networks in response to mood stabilising drugs. This model can be utilised in the comparison of the action of multiple drug regimes or for initial screening purposes to inform optimal drug design.

OBJECTIVE

This study aims to identify new genes not described previously that may be relevant in the etiology or pathophysiology of patients with Hirschsprung disease (HD). This was done by identifying differences in gene expression between normal and abnormal segments of bowel in HD patients compared with controls.

METHODS

Full-thickness colonic tissue samples were taken from HD patients, both from the diseased (Ds) and normal segment of the colon (Nr), and from controls (Ct). Samples were further dissected into mucosa (MUC) and muscle (MUS). RNA was extracted and analyzed on Affymetrix Gene Chip Human Gene 1.0 ST arrays. Statistical analyses using ANOVA with a fold change cut off of 2 was applied to detect a number of differentially expressed genes. Selected genes were revalidated by quantitative real-time reverse transcriptase polymerase chain reaction.

RESULTS

Thirty-four samples (18 MUS and 16 MUC) were analyzed. MUC (1.64 ± 0.46 μg/mg) and MUS (0.83 ± 0.48 μg/mg) showed good RNA extraction yield and quality. Of the 24,987 filtered on expression genes, MUS showed 220 genes with expression difference of 2-fold, out of which 120 genes were significant with P ≤ .05. Similarly, MUC demonstrated 206 genes with 2-fold changes and 9 had P ≤ .05. Some genes showing differential expression between groups and therefore subject to further analysis were RELN, GAL, GAP43, NRSN1, and GABRG2.

CONCLUSIONS

Analyzed data showed significant differences in expression of above sets of genes with up- and down-regulation, which has not been described before in HD and could have a role in pathogenesis of this condition.

Studies have suggested that reelin (RELN) polymorphism was associated with the susceptibility of schizophrenia (SZ), but the results remained controversial. Thus, we conducted this meta-analysis to determine whether RELN variants (rs7341475 and rs262355) were associated with SZ risk. Studies were identified through retrieving Web of Science, PubMed and Embase databases from inception to May 2015. The genotype data were extracted to calculate the odds ratios (ORs) and 95% confidence intervals (CIs). For rs7341475, five studies with 4741 SZ patients and 10075 controls are included and the results indicate that carriage of A allele is associated with decreased SZ risk in dominant genetic model (OR=0.90, 95%CI=0.83-0.98) and additive model (OR=0.90, 95% CI=0.84-0.97). Subgroup analysis indicates that the association between rs7341475 and SZ is only significant in Caucasian. For rs262355, four studies with 2017 SZ patients and 3274 controls are included, the results demonstrate that carriage of A allele is associated with increased risk of SZ only in Caucasian (dominant model: OR=1.17, 95%CI=1.01-1.37; additive model OR=1.13, 95%CI=1.02-1.27). This meta-analysis suggests that rs7341475 (A/G) and rs262355 (A/T) polymorphisms in RELN gene are inversely associated with SZ risk.

BACKGROUND

Several reports suggest that the reelin protein could play a role in Alzheimer pathophysiology. This led us to ask whether genetic variability in the reelin pathway may increase the risk of developing Alzheimer disease (AD) or mild cognitive impairment (MCI).

METHODS

This was a case-control study in which neuropsychological tests were administered and peripheral blood samples taken. The study included 121 patients with AD, 94 with MCI, and 198 controls. Forty biallelic variants single nucleotide polimorphisms were genotyped in 8 genes related to reelin signaling pathway using a SNPlex genotyping system, and allele frequencies were compared between patients and controls using χ tests and obtaining odds ratios (OR).

RESULTS

A total of 413 subjects with complete neuropsychological data were analyzed. A significant association between the genotypes RELN (rs528528 and rs2299356), PLK2 (rs15009 and rs702723), and CAMK2A (rs3756577 and rs3822606) and AD or MCI was found. A significant association also was found between the GG genotype at the RELN-rs2299356 and the risk of AD (OR=2.68, P=0.003) and between the AG genotype at the CAMK2A-rs3822606 (OR=2.13, P=0.004). We found a protective effect of the RELN-rs528528 CT genotype and MCI (OR=0.36, P=0.002), and the PLK2-rs15009 CC and GG genotypes and CC genotype at PLK2-rs702723 with OR ranging from 0.40 to 0.57 on AD. These data suggest that TT or CT genotypes at CAMK2A-rs3756577 is associated with risk reduction for AD and MCI ranging from 2 to nearly 8 times.

CONCLUSIONS

Our data suggest a possible relation between certain reelin signaling pathway genotypes and cognitive impairment related to AD.