Pike See Cheah

Neurodevelopmental and neuropsychiatric behaviour defects arise from 14-3-3ζ deficiency.

Complex neuropsychiatric disorders are believed to arise from multiple synergistic deficiencies within connected biological networks controlling neuronal migration, axonal pathfinding and synapse formation. Here, we show that deletion of 14-3-3ζ causes neurodevelopmental anomalies similar to those seen in neuropsychiatric disorders such as schizophrenia, autism spectrum disorder and bipolar disorder. 14-3-3ζ-Deficient mice displayed striking behavioural and cognitive deficiencies including a reduced capacity to learn and remember, hyperactivity and disrupted sensorimotor gating. These deficits are accompanied by subtle developmental abnormalities of the hippocampus that are underpinned by aberrant neuronal migration. Significantly, 14-3-3ζ-deficient mice exhibited abnormal mossy fibre navigation and glutamatergic synapse formation. The molecular basis of these defects involves the schizophrenia risk factor, DISC1, which interacts isoform specifically with 14-3-3ζ. Our data provide the first evidence of a direct role for 14-3-3ζ deficiency in the aetiology of neurodevelopmental disorders and identifies 14-3-3ζ as a central risk factor in the schizophrenia protein interaction network.

Expression of Notch-1 receptor and its ligands Jagged-1 and Delta-1 in amoeboid microglia in postnatal rat brain and murine BV-2 cells

Notch‐1 receptor signaling pathway is involved in neuronal and glial differentiation. Its involvement in microglial functions, however, has remained elusive. This study reports the localization of Notch‐1 receptor immunoreactivity in the amoeboid microglial cells (AMC) in the postnatal rat brain. By immunofluorescence, Notch‐1 receptor was colocalized with its ligands, Jagged‐1 and Delta‐1, in the AMC in the corpus callosum and subventricular zone. Notch‐1 immunopositive cells were confirmed to be microglia labeled by OX42 and lectin. Immunoexpression of Notch‐1 receptor was progressively reduced with age. Western blot analysis showed that Notch‐1 protein level in the corpus callosum in which the AMC were heavily populated was concomitantly decreased. In postnatal rats challenged with lipopolysaccharide (LPS), Notch‐1 receptor immunofluorescence in AMC was noticeably enhanced. Furthermore, Notch‐1 protein level in the corpus callosum was increased as revealed by Western blotting analysis. In primary microglial culture treated with LPS, mRNA expression of Notch‐1 and its ligand Jagged‐1 was upregulated but that of Delta‐1 was reduced. The expression pattern of Notch‐1 and its ligands was confirmed in murine BV‐2 cells. Furthermore, Notch‐1 neutralization with its antibody reduced its protein expression. More importantly, neutralization of Notch‐1 concomitantly suppressed the mRNA expression of IL‐6, IL‐1, M‐CSF, and iNOS; TNF‐α, mRNA expression, however, was enhanced. Western blot confirmed the changes of protein level of the above except for IL‐6, which remained relatively unaltered. It is concluded that Notch‐1 signaling in the AMC and LPS‐activated microglia/BV‐2 cells modulates the expression of proinflammatory cytokines and nitric oxide.

Molecular genetics of the developing neuroendocrine hypothalamus

Formation of the mammalian endocrine system and neuroendocrine organs involves complex regulatory networks resulting in a highly specialized cell system able to secrete a diverse array of peptide hormones. The hypothalamus is located in the mediobasal region of the brain and acts as a gateway between the endocrine and nervous systems. From an endocrinology perspective, the parvicellular neurons of the hypothalamus are of particular interest as they function as a control centre for several critical physiological processes including growth, metabolism and reproduction by regulating hormonal signaling from target cognate cell types in the anterior pituitary. Delineating the genetic program that controls hypothalamic development is essential for complete understanding of parvicellular neuronal function and the etiology of congenital disorders that result from hypothalamic-pituitary axis dysfunction. In recent years, studies have shed light on the interactions between signaling molecules and activation of transcription factors that regulate hypothalamic cell fate commitment and terminal differentiation. The aim of this review is to summarize the recent molecular and genetic findings that have advanced our understanding of the emergence of the known important hypophysiotropic signaling molecules in the hypothalamus. We have focused on reviewing the literature that provides evidence of the dependence on expression of specific genes for the normal development and function of the cells that secrete these neuroendocrine factors, as well as studies of the elaboration of the spatial or temporal patterns of changes in gene expression that drive this development.

Deep sequencing analysis of the developing mouse brain reveals a novel microRNA

BackgroundMicroRNAs (miRNAs) are small non-coding RNAs that can exert multilevel inhibition/repression at a post-transcriptional or protein synthesis level during disease or development. Characterisation of miRNAs in adult mammalian brains by deep sequencing has been reported previously. However, to date, no small RNA profiling of the developing brain has been undertaken using this method. We have performed deep sequencing and small RNA analysis of a developing (E15.5) mouse brain.ResultsWe identified the expression of 294 known miRNAs in the E15.5 developing mouse brain, which were mostly represented by let-7 family and other brain-specific miRNAs such as miR-9 and miR-124. We also discovered 4 putative 22-23 nt miRNAs: mm_br_e15_1181, mm_br_e15_279920, mm_br_e15_96719 and mm_br_e15_294354 each with a 70-76 nt predicted pre-miRNA. We validated the 4 putative miRNAs and further characterised one of them, mm_br_e15_1181, throughout embryogenesis. Mm_br_e15_1181 biogenesis was Dicer1-dependent and was expressed in E3.5 blastocysts and E7 whole embryos. Embryo-wide expression patterns were observed at E9.5 and E11.5 followed by a near complete loss of expression by E13.5, with expression restricted to a specialised layer of cells within the developing and early postnatal brain. Mm_br_e15_1181 was upregulated during neurodifferentiation of P19 teratocarcinoma cells. This novel miRNA has been identified as miR-3099.ConclusionsWe have generated and analysed the first deep sequencing dataset of small RNA sequences of the developing mouse brain. The analysis revealed a novel miRNA, miR-3099, with potential regulatory effects on early embryogenesis, and involvement in neuronal cell differentiation/function in the brain during late embryonic and early neonatal development.

Molecular networks involved in mouse cerebral corticogenesis and spatio-temporal regulation of Sox4 and Sox11 novel antisense transcripts revealed by transcriptome profiling

BackgroundDevelopment of the cerebral cortex requires highly specific spatio-temporal regulation of gene expression. It is proposed that transcriptome profiling of the cerebral cortex at various developmental time points or regions will reveal candidate genes and associated molecular pathways involved in cerebral corticogenesis.ResultsSerial analysis of gene expression (SAGE) libraries were constructed from C57BL/6 mouse cerebral cortices of age embryonic day (E) 15.5, E17.5, postnatal day (P) 1.5 and 4 to 6 months. Hierarchical clustering analysis of 561 differentially expressed transcripts showed regionalized, stage-specific and co-regulated expression profiles. SAGE expression profiles of 70 differentially expressed transcripts were validated using quantitative RT-PCR assays. Ingenuity pathway analyses of validated differentially expressed transcripts demonstrated that these transcripts possess distinctive functional properties related to various stages of cerebral corticogenesis and human neurological disorders. Genomic clustering analysis of the differentially expressed transcripts identified two highly transcribed genomic loci, Sox4 and Sox11, during embryonic cerebral corticogenesis. These loci feature unusual overlapping sense and antisense transcripts with alternative polyadenylation sites and differential expression. The Sox4 and Sox11 antisense transcripts were highly expressed in the brain compared to other mouse organs and are differentially expressed in both the proliferating and differentiating neural stem/progenitor cells and P19 (embryonal carcinoma) cells.ConclusionsWe report validated gene expression profiles that have implications for understanding the associations between differentially expressed transcripts, novel targets and related disorders pertaining to cerebral corticogenesis. The study reports, for the first time, spatio-temporally regulated Sox4 and Sox11 antisense transcripts in the brain, neural stem/progenitor cells and P19 cells, suggesting they have an important role in cerebral corticogenesis and neuronal/glial cell differentiation.

Functional transcriptome analysis of the postnatal brain of the Ts1Cje mouse model for Down syndrome reveals global disruption of interferon-related molecular networks

BackgroundThe Ts1Cje mouse model of Down syndrome (DS) has partial triplication of mouse chromosome 16 (MMU16), which is partially homologous to human chromosome 21. These mice develop various neuropathological features identified in DS individuals. We analysed the effect of partial triplication of the MMU16 segment on global gene expression in the cerebral cortex, cerebellum and hippocampus of Ts1Cje mice at 4 time-points: postnatal day (P)1, P15, P30 and P84.ResultsGene expression profiling identified a total of 317 differentially expressed genes (DEGs), selected from various spatiotemporal comparisons, between Ts1Cje and disomic mice. A total of 201 DEGs were identified from the cerebellum, 129 from the hippocampus and 40 from the cerebral cortex. Of these, only 18 DEGs were identified as common to all three brain regions and 15 were located in the triplicated segment. We validated 8 selected DEGs from the cerebral cortex (Brwd1, Donson, Erdr1, Ifnar1, Itgb8, Itsn1, Mrps6 and Tmem50b), 18 DEGs from the cerebellum (Atp5o, Brwd1, Donson, Dopey2, Erdr1, Hmgn1, Ifnar1, Ifnar2, Ifngr2, Itgb8, Itsn1, Mrps6, Paxbp1, Son, Stat1, Tbata, Tmem50b and Wrb) and 11 DEGs from the hippocampus (Atp5o, Brwd1, Cbr1, Donson, Erdr1, Itgb8, Itsn1, Morc3, Son, Tmem50b and Wrb). Functional clustering analysis of the 317 DEGs identified interferon-related signal transduction as the most significantly dysregulated pathway in Ts1Cje postnatal brain development. RT-qPCR and western blotting analysis showed both Ifnar1 and Stat1 were over-expressed in P84 Ts1Cje cerebral cortex and cerebellum as compared to wild type littermates.ConclusionsThese findings suggest over-expression of interferon receptor may lead to over-stimulation of Jak-Stat signaling pathway which may contribute to the neuropathology in Ts1Cje or DS brain. The role of interferon mediated activation or inhibition of signal transduction including Jak-Stat signaling pathway has been well characterized in various biological processes and disease models including DS but information pertaining to the role of this pathway in the development and function of the Ts1Cje or DS brain remains scarce and warrants further investigation.

DRD and GRIN2B polymorphisms and their association with the development of impulse control behaviour among Malaysian Parkinson’s disease patients

BackgroundImpulse control disorder (ICD) and behaviours (ICB) represent a group of behavioural disorders that have become increasingly recognised in Parkinson’s disease (PD) patients who previously used dopaminergic medications, particularly dopamine agonists and levodopa. It has been suggested that these medications can lead to the development of ICB through the abnormal modulation of dopaminergic transmission and signalling in the mesocorticolimbic dopaminergic system. Several studies have reported an association between polymorphisms in the dopamine receptor (DRD) and N-methyl-D-aspartate 2B (GRIN2B) genes with the development of ICB in PD (PD-ICB) patients. Thus, this study aimed to investigate the association of selected polymorphisms within the DRD and GRIN2B genes with the development of ICB among PD patients using high resolution melt (HRM) analysis.MethodWe used high resolution melt (HRM) analysis to genotype 11 polymorphisms in 5 DRD genes [DRD1 (rs4532, rs4867798 and rs265981), DRD2 (ANKK1 rs1800497, rs104894220 and rs144999500), DRD3 (rs3732783 and rs6280), DRD4 (rs1800443), and DRD5 (rs144132215)] and 1 polymorphism in GRIN2B (rs7301328) in PD patients with (cases, n = 52) and without (controls, n = 39) ICB. Cases were obtained from two tertiary movement disorder centres [UKMMC (n = 9) and UMMC (n = 43)]. At both centres, the diagnosis of ICB was made using the QUIP questionnaire. Controls were recruited from PD patients who attended UKMMC and were found to be negative for ICB using the QUIP questionnaire.ResultsThe HRM analysis showed that 7 of 11 polymorphisms [DRD1 (rs4532, rs4867798, and rs265981), DRD2 (ANKK1 rs1800497), DRD3 (rs3732783 and rs6280), and GRIN2B (rs7301328)] exhibited a clear distinction between wild-type and variant alleles. Variants of DRD2/ANKK1 rs1800497 (OR = 3.77; 95% CI, 1.38-10.30; p = 0.0044), DRD1 rs4867798 (OR = 24.53; 95% CI, 1.68-357.28; p = 0.0054), DRD1 rs4532 (OR = 21.33; 95% CI, 1.97-230.64; p = 0.0024), and GRIN2B rs7301328 (OR = 25.07; 95% CI, 1.30-483.41; p = 0.0097) were found to be associated with an increased risk of developing ICB among PD patients.ConclusionOur findings suggest that polymorphisms in dopamine [DRD1 (rs4532 and rs4867798) and DRD2/ANKK1 rs1800497] and glutamate (GRIN2B rs7301328) receptor genes confer increased risk of ICB development among PD patients.

Expression of the murine transcription factor SOX3 during embryonic and adult neurogenesis.

Previous studies have shown that Sox3 is expressed in nascent neuroprogenitor cells and is functionally required in mammals for development of the dorsal telencephalon and hypothalamus. However, Sox3 expression during embryonic and adult neurogenesis has not been examined in detail. Using a SOX3-specific antibody, we show that murine SOX3 expression is maintained throughout telencephalic neurogenesis and is restricted to progenitor cells with neuroepithelial and radial glial morphologies. We also demonstrate that SOX3 is expressed within the adult neurogenic regions and is coexpressed extensively with the neural stem cell marker SOX2 indicating that it is a lifelong marker of neuroprogenitor cells. In contrast to the telencephalon, Sox3 expression within the developing hypothalamus is upregulated in developing neurons and is maintained in a subset of differentiated hypothalamic cells through to adulthood. Together, these data show that Sox3 regulation is region-specific, consistent with it playing distinct biological roles in the dorsal telencephalon and hypothalamus.

Congenital Hydrocephalus and Abnormal Subcommissural Organ Development in Sox3 Transgenic Mice

Congenital hydrocephalus (CH) is a life-threatening medical condition in which excessive accumulation of CSF leads to ventricular expansion and increased intracranial pressure. Stenosis (blockage) of the Sylvian aqueduct (Aq; the narrow passageway that connects the third and fourth ventricles) is a common form of CH in humans, although the genetic basis of this condition is unknown. Mouse models of CH indicate that Aq stenosis is associated with abnormal development of the subcommmissural organ (SCO) a small secretory organ located at the dorsal midline of the caudal diencephalon. Glycoproteins secreted by the SCO generate Reissners fibre (RF), a thread-like structure that descends into the Aq and is thought to maintain its patency. However, despite the importance of SCO function in CSF homeostasis, the genetic program that controls SCO development is poorly understood. Here, we show that the X-linked transcription factor SOX3 is expressed in the murine SCO throughout its development and in the mature organ. Importantly, overexpression of Sox3 in the dorsal diencephalic midline of transgenic mice induces CH via a dose-dependent mechanism. Histological, gene expression and cellular proliferation studies indicate that Sox3 overexpression disrupts the development of the SCO primordium through inhibition of diencephalic roof plate identity without inducing programmed cell death. This study provides further evidence that SCO function is essential for the prevention of hydrocephalus and indicates that overexpression of Sox3 in the dorsal midline alters progenitor cell differentiation in a dose-dependent manner.

Histomorphometric profile of the corneal response to short-term reverse-geometry orthokeratology lens wear in primate corneas: a pilot study.

Purpose: To investigate the histological changes in primate cornea induced by short-term overnight orthokeratology (OK). Methods: Nine young adult primates were used. One animal served as negative control. The remaining 8 animals wore reverse-geometry OK lenses for periods of 4, 8, 16, and 24 hours on 1 eye with the other eye as control. Central and midperipheral corneal thickness, as well as ultrastructural changes in corneal epithelium, stroma and endothelium in response to OK lenses, were evaluated. Results: OK significantly reduced the thickness of the central cornea in all treatment groups. The central corneal thinning was both stromal and epithelial in origin. Substantial midperipheral corneal thickening was seen in 16-hour and 24-hour lens-wear groups and this effect was both stromal and epithelial in origin as well. Histology evidence indicated the primary epithelial response in the central cornea was compression of cells that resulted in wing cells becoming shorter and basal cells being squatted rather than lost or migration of cell layers. These pronounced cell shape changes occurred without compromising the structural integrity of the desmosomes. The thickened corneal epithelium has normal cell layers. The squamous cells have larger surface sizes and are composed of oval instead of flattened nuclei. This implied delayed surface cell exfoliation at the thickened midperipheral epithelium. Physical presence of OK lens over the cornea did not influence the microstructures of microvilli and microplicae, endothelium, and collagen distribution. Conclusions: The primate cornea, particularly the corneal epithelium, responds rapidly to the application of reverse-geometry OK lenses with significant epithelial cell shape alterations with short-term OK lens wear. This finding suggests that the corneal epithelium is moldable in response to the physical forces generated by the OK lenses.