Development Of Neural Stem Cells Research Articles

SOX19b regulates the premature neuronal differentiation of neural stem cells through EZH2-mediated histone methylation in neural tube development of zebrafish

ObjectiveNeural tube defects (NTDs) are the most serious and common birth defects in the clinic. The SRY-related HMG box B1 (SoxB1) gene family has been implicated in different processes of early embryogenesis. Sox19b is a maternally expressed gene in the SoxB1 family that is found in the region of the presumptive central nervous system (CNS), but its role and mechanism in embryonic neural stem cells (NSCs) during neural tube development have not yet been explored. Considering that Sox19b is specific to bony fish, we intended to investigate the role and mechanism of Sox19b in neural tube development in zebrafish embryos.Material and methodsMorpholino (MO) antisense oligonucleotides were used to construct a Sox19b loss-of-function zebrafish model. The phenotype and the expression of related genes were analysed by in situ hybridization and immunolabelling. Epigenetic modifications were detected by western blot and chromatin immunoprecipitation.ResultsIn this study, we found that zebrafish embryos exhibited a reduced or even deleted forebrain phenotype after the expression of the Sox19b gene was inhibited. Moreover, we found for the first time that knockdown of Sox19b reduced the proliferation of NSCs; increased the transcription levels of Ngn1, Ascl1, HuC, Islet1, and cyclin-dependent kinase (CDK) inhibitors; and led to premature differentiation of NSCs. Finally, we found that knockdown of Sox19b decreased the levels of EZH2/H3K27me3 and decreased the level of H3K27me3 at the promoters of Ngn1 and ascl1a.ConclusionTogether, our data demonstrate that Sox19b plays an essential role in early NSC proliferation and differentiation through EZH2-mediated histone methylation in neural tube development. This study established the role of transcription factor Sox19b and epigenetic factor EZH2 regulatory network on NSC development, which provides new clues and theoretical guidance for the clinical treatment of neural tube defects.

Trim71 Links an Ancient MicroRNA Pathway to Neural Stem Cell Development and Human Congenital Hydrocephalus

Abstract INTRODUCTION Congenital hydrocephalus (CH), the most common brain malformation that affects 1:1000 live births, remains poorly treated owing to incomplete understanding of disease pathogenesis. Identification of novel CH disease genes is a powerful approach to understanding the molecular genetics of CH. Whole exome sequencing in humans has identified TRIM71 as a bona fide CH disease gene. Trim71 is the major target of the ancient let-7 pathway that famously regulates cell differentiation and organismal development via microRNA-mediated gene silencing in Caenorhabditis elegans. Previous works have implicated Trim71 in the regulation of embryonic neural stem cell (NSC) fate, yet its cellular and molecular function in NSCs remain poorly understood. METHODS To gain a better understanding of Trim71 as a paradigm of NSC involvement in CH, we have generated a mouse model that harbors the point mutation in Trim71 that results in human CH (R595H) using CRISPR-Cas9. RESULTS We found that a subset of mice heterozygous for the Trim71 R595H mutant allele developed severe communicating hydrocephalus with accompanying macrocephaly by 4 wk of age. CONCLUSION Investigation into the function of Trim71 and its CH-causing mutations using our novel mouse models presents outstanding opportunities to increase understanding of human NSC regulation, brain development, and CH pathogenesis.

Chronic Ethanol Exposure Alters DNA Methylation in Neural Stem Cells: Role of Mouse Strain and Sex.

Prenatal alcohol exposure (PAE) is considered as a risk factor for the development of fetal alcohol spectrum disorders (FASD). Evidence indicates that PAE affects epigenetic mechanisms (such as DNA methylation) and alters the normal differentiation and development of neural stem cells (NSC) in the fetal brain. However, PAE effects depend on several factors such as sex and strain of the studied subjects. Here, we investigated whether murine sex and strain contribute to the effects of chronic ethanol exposure on DNA methylation machinery of differentiating NSC. Further, the effects of PAE on glial lineage (including both astrocytes and oligodendrocytes) in a sex- and strain-dependent manner have not been studied yet. To examine the effects of chronic ethanol exposure on gliogenesis, we exposed differentiating NSC to glio-inductive culture conditions. Applying a standard in vitro model system, we treated male and female differentiating NSC (obtained from the forebrain of CD1 and C57BL/6 embryos at embryonic day 14.5) with chronic ethanol exposure (70mM) for 8days. We show that ethanol induces global DNA hypomethylation, while altering the expression of DNA methylation-related genes in a sex- and strain-specific manner. The observed change in cellular DNA methylation levels was associated with altered expression of glial markers CNPASE, GFAP, and OLIG2 in CD1 (but not C57BL/6) cells. We conclude that the impact of ethanol effect on DNA methylation is dependent on cellular sex and strain. Also, ethanol impact on neural stem cell fate commitment was only detected in cells isolated from CD1 mouse strain, but not in C57BL/6 cells. The results of the current study provide evidence that sex and strain of rodents (C57BL/6 and CD1) during gestation are important factors, which affect alcohol effects on NSC differentiation and DNA methylation. Results of this study may also help in interpreting data on the developmental toxicity of many compounds during the gestational period.

Development of glial restricted human neural stem cells for oligodendrocyte differentiation in vitro and in vivo

In this study, we have developed highly expandable neural stem cells (NSCs) from HESCs and iPSCs that artificially express the oligodendrocyte (OL) specific transcription factor gene Zfp488. This is enough to restrict them to an exclusive oligodendrocyte progenitor cell (OPC) fate during differentiation in vitro and in vivo. During CNS development, Zfp488 is induced during the early stages of OL generation, and then again during terminal differentiation of OLs. Interestingly, the human ortholog Znf488, crucial for OL development in human, has been recently identified to function as a dorsoventral pattering regulator in the ventral spinal cord for the generation of P1, P2/pMN, and P2 neural progenitor domains. Forced expression of Zfp488 gene in human NSCs led to the robust generation of OLs and suppression of neuronal and astrocyte fate in vitro and in vivo. Zfp488 expressing NSC derived oligodendrocytes are functional and can myelinate rat dorsal root ganglion neurons in vitro, and form myelin in Shiverer mice brain in vivo. After transplantation near a site of demyelination, Zfp488 expressing hNSCs migrated to the lesion and differentiated into premyelinating OLs. A certain fraction also homed in the subventricular zone (SVZ). Zfp488-ZsGreen1-hNSC derived OLs formed compact myelin in Shiverer mice brain seen under the electron microscope. Transplanted human neural stem cells (NSC) that have the potential to differentiate into functional oligodendrocytes in response to remyelinating signals can be a powerful therapeutic intervention for disorders where oligodendrocyte (OL) replacement is beneficial.

Deleting Adult Neural Stem Cells Ameliorates Synaptic and Cognitive Deficits in Animal Models of Alzheimer's Disease

The development of adult neural stem cells (aNSCs) was impaired in the hippocampus of animal models of Alzheimer’s disease (AD). However, whether and how aNSCs and new neurons derived from aNSCs affect AD neuropathology is unclear. Here, we found that genetic or drug-induced ablation of aNSCs significantly ameliorated deficits of synaptic plasticity and cognitive functions in two mouse models of AD. On the other hand, whole cell recording revealed that deleting aNSCs prevented the increased inhibition in the dentate granule cells of AD mice. Furthermore, deleting aNSCs did not affect Aβ levels but prevented the calbindin depletion in the hippocampus of AD mice. Knocking down calbindin in the hippocampus abolished the effects of deleting aNSCs on synaptic and cognitive functions in AD mice. Taken together, our data suggest that deleting aNSCs improved synaptic plasticity and cognitive functions in AD mice by regulating the activity of dentate granule cells via calbindin.

Development of Ependymal and Postnatal Neural Stem Cells and Their Origin from a Common Embryonic Progenitor

The adult mouse brain contains an extensive neurogenic niche in the lateral walls of the lateral ventricles. This epithelium, which has a unique pinwheel organization, contains multiciliated ependymal (E1) cells and neural stem cells (B1). This postnatal germinal epithelium develops from the embryonic ventricular zone, but the lineage relationship between E1 and B1 cells remains unknown. Distinct subpopulations of radial glia (RG) cells in late embryonic and early postnatal development either expand their apical domain >11-fold to form E1 cells or retain small apical domains that coalesce into the centers of pinwheels to form B1 cells. Using independent methods of lineage tracing, we show that individual RG cells can give rise to clones containing E1 and B1 cells. This study reveals key developmental steps in the formation of the postnatal germinal niche and the shared cellular origin of E1 and B1 cells.

Hippocampal neural stem cells undergo fate switch following intermittent hypoxia

Adult neurogenesis is a phenomenon widely recognized as a process supporting neurophysiology of the hippocampus. We recently reported that 30 days of intermittent hypoxia (IH), a principal consequence for sleep apnea, stimulates the expansion of hippocampal neural stem cells (NSCs), yet suppresses the generation of adult‐born neurons. This study investigates how IH impacts NSCs by using a shorter duration of IH. We hypothesize that 10 days of IH (IH10) redirects the course of development of NSCs. We examined how NSCs in the neurogenic niche of the dentate gyrus are affected by IH10. Immunohistochemical analysis revealed that the number of NSCs (Sox2+ cells) was not increased following IH10, yet appears to increase in quiescent NSCs (i.e., Sox2+ cells that co‐express the astrocytic marker, S100b, and do not develop into adult‐born neurons). Fewer NSCs were observed transitioning into intermediate neuronal progenitors (INPs, SOX2+/TBR2+ cells) and a reduction in the total number of INPs (TBR2+ cells) was also noted. These experiments suggest that the impact of IH on cells involved with adult neurogenesis is cell‐type specific. Our findings provide further support that untreated sleep apnea may impact the fate of pluripotent cellular constituents in the central nervous system.Support or Funding InformationThis work was supported by NIH R01 NS10742101 (AJG)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Chromodomain Helicase DNA-Binding Protein 7 Is Suppressed in the Perinecrotic/Ischemic Microenvironment and Is a Novel Regulator of Glioblastoma Angiogenesis.

Tumorigenic and non-neoplastic tissue injury occurs via the ischemic microenvironment defined by low oxygen, pH, and nutrients due to blood supply malfunction. Ischemic conditions exist within regions of pseudopalisading necrosis, a pathological hallmark of glioblastoma (GBM), the most common primary malignant brain tumor in adults. To recapitulate the physiologic microenvironment found in GBM tumors and tissue injury, we developed an in vitro ischemic model and identified chromodomain helicase DNA-binding protein 7 (CHD7) as a novel ischemia-regulated gene. Point mutations in the CHD7 gene are causal in CHARGE syndrome (a developmental disorder causing coloboma, heart defects, atresia choanae, retardation of growth, and genital and ear anomalies) and interrupt the epigenetic functions of CHD7 in regulating neural stem cell maintenance and development. Using our ischemic system, we observed microenvironment-mediated decreases in CHD7 expression in brain tumor-initiating cells and neural stem cells. Validating our approach, CHD7 was suppressed in the perinecrotic niche of GBM patient and xenograft sections, and an interrogation of patient gene expression datasets determined correlations of low CHD7 with increasing glioma grade and worse patient outcomes. Segregation of GBM by molecular subtype revealed a novel observation that CHD7 expression is elevated in proneural versus mesenchymal GBM. Genetic targeting of CHD7 and subsequent gene ontology analysis of RNA sequencing data indicated angiogenesis as a primary biological function affected by CHD7 expression changes. We validated this finding in tube-formation assays and vessel formation in orthotopic GBM models. Together, our data provide further understanding of molecular responses to ischemia and a novel function of CHD7 in regulating angiogenesis in both neoplastic and non-neoplastic systems. Stem Cells 2019;37:453-462.

Possible Role of PHD Inhibitors as Hypoxia-Mimicking Agents in the Maintenance of Neural Stem Cells’ Self-Renewal Properties

Hypoxia is the most critical factor for maintaining stemness. During embryonic development, neural stem cells (NSCs) reside in hypoxic niches, and different levels of oxygen pressure and time of hypoxia exposure play important roles in the development of NSCs. Such hypoxic niches exist in adult brain tissue, where the neural precursors originate. Hypoxia-inducible factors (HIFs) are key transcription heterodimers consisting of regulatory α-subunits (HIF-1α, HIF-2α, HIF-3α) and a constitutive β-subunit (HIF-β). Regulation of downstream targets determines the fate of NSCs. In turn, the stability of HIFs-α is regulated by prolyl hydroxylases (PHDs), whose activity is principally modulated by PHD substrates like oxygen (O2), α-ketoglutarate (α-KG), and the co-factors ascorbate (ASC) and ferrous iron (Fe2+). It follows that the transcriptional activity of HIFs is actually determined by the contents of O2, α-KG, ASC, and Fe2+. In normoxia, HIFs-α are rapidly degraded via the ubiquitin-proteasome pathway, in which PHDs, activated by O2, lead to hydroxylation of HIFs-α at residues 402 and 564, followed by recognition by the tumor suppressor protein von Hippel–Lindau (pVHL) as an E3 ligase and ubiquitin labeling. Conversely, in hypoxia, the activity of PHDs is inhibited by low O2 levels and HIFs-α can thus be stabilized. Hence, suppression of PHD activity in normoxic conditions, mimicking the effect of hypoxia, might be beneficial for preserving the stemness of NSCs, and it is clinically relevant as a therapeutic approach for enhancing the number of NSCs in vitro and for cerebral ischemia injury in vivo. This study will review the putative role of PHD inhibitors on the self-renewal of NSCs.

DRG-Derived Neural Progenitors Differentiate into Functional Enteric Neurons Following Transplantation in the Postnatal Colon

Cell therapy has great promise for treating gastrointestinal motility disorders caused by intestinal nervous system (ENS) diseases. However, appropriate sources, other than enteric neural stem cells and human embryonic stem cells, are seldom reported. Here, we show that neural progenitors derived from the dorsal root ganglion (DRG) of EGFP mice survived, differentiated into enteric neurons and glia cells, migrated widely from the site of injection, and established neuron-muscle connections following transplantation into the distal colon of postnatal mice. The exogenous EGFP+ neurons were physiologically functional as shown by the activity of calcium imaging. This study shows that that other tissues besides the postnatal bowel harbor neural crest stem cells or neural progenitors that have the potential to differentiate into functional enteric neurons in vivo and can potentially be used for intestinal nerve regeneration. These DRG-derived neural progenitor cells may be a choice for cell therapy of ENS disease as an allograft. The new knowledge provided by our study is important for the development of neural crest stem cell and cell therapy for the treatment of intestinal neuropathy.

Human neural stem cell transplantation improves cognition in a murine model of Alzheimer\u2019s disease

Stem cell transplantation offers a potentially transformative approach to treating neurodegenerative disorders. The safety of cellular therapies is established in multiple clinical trials, including our own in amyotrophic lateral sclerosis. To initiate similar trials in Alzheimer’s disease, efficacious cell lines must be identified. Here, we completed a preclinical proof-of-concept study in the APP/PS1 murine model of Alzheimer’s disease. Human neural stem cell transplantation targeted to the fimbria fornix significantly improved cognition in two hippocampal-dependent memory tasks at 4 and 16 weeks post-transplantation. While levels of synapse-related proteins and cholinergic neurons were unaffected, amyloid plaque load was significantly reduced in stem cell transplanted mice and associated with increased recruitment of activated microglia. In vitro, these same neural stem cells induced microglial activation and amyloid phagocytosis, suggesting an immunomodulatory capacity. Although long-term transplantation resulted in significant functional and pathological improvements in APP/PS1 mice, stem cells were not identified by immunohistochemistry or PCR at the study endpoint. These data suggest integration into native tissue or the idea that transient engraftment may be adequate for therapeutic efficacy, reducing the need for continued immunosuppression. Overall, our results support further preclinical development of human neural stem cells as a safe and effective therapy for Alzheimer’s disease.

Advance of ependymal cells repairing spinal cord injury

Spinal cord injury (SCI) has remained a challenging area for scientists. In recent years, with the development and research of endogenous neural stem cells in the central nervous system, the use of endogenous neural stem cells (ENSCs) in the treatment of brain diseases has made some progress, but almost no activation of spinal cord ENSCs to repair spinal cord function recovery report. To identify the identity and distribution of ENSCs in the adult spinal cord, to analyze their fate mechanism, to find its active intervention target, has become a new idea for the treatment of spinal cord injury. Recent studies have shown that the ependymal cells of the spinal cord have the potential of neural stem cells, the current research progress is summarized. Key words: Ependymal cell; Spinal cord injury; Endogenous neural stem cell; Niche; Neurogenesis; Wound repair

Comparative Analysis of microRNA Expression Profiles of Exosomes Derived from Normal and Hypoxic Preconditioning Human Neural Stem Cells by Next Generation Sequencing.

Stroke recovery is associated with neural stem cell (NSC) development and neurovascular unit reconstruction. The exosome, as an important intercellular player in neurovascular communication, mediates neuro-restorative events by transferring exosomal protein and RNA cargoes. In this study, we explored the role of exosomal microRNAs (miRNAs) in human NSCs (hNSCs), and analyzed the expression profiles of miRNAs in hNSC-derived and hypoxic preconditioning hNSC-derived exosomes with the help of next generation sequencing (NGS). The results demonstrated that a certain proportion of miRNAs were differentially expressed in both exosomes. In addition, target gene prediction and Gene Ontology (GO) enrichment analysis showed that these genes were associated with differential miRNAs primarily participating in biological processes (regulation of cellular process), cellular component (intracellular membrane-bounded organelle), and molecular function (binding). Kyoto Encyclopedia of Genes and Genomes pathway (KEGG) pathway enrichment data suggested that most of targeted genes involved in PI3K-Akt, Hippo, MAPK, mTOR, and Endocytosis etc. signaling pathways. We identified the interesting and important expressed miRNA and considered that miR-98-3p might be a special hNSC-derived exosomal-miRNA which was significantly downregulated under hypoxic preconditioning. The hNSCs-derived exosomes were capable of modulating gene expression or promoting stroke therapy. We observed that after hypoxic preconditioning, the functions of these exosomes were changed, and exosomal-miRNAs expression profile was different. In summary, our study suggested that hNSC-derived exosomal miRNAs including hypoxic preconditioning exosomal miRNAs provided a new strategy for the diagnosis and treatment of stroke patients.

MiR-124 promotes proliferation and neural differentiation of neural stem cells through targeting DACT1 and activating Wnt/β-catenin pathways.

Neural stem cells (NSCs) are multipotent and undifferentiated cells with the potential to differentiate into neuronal lineages and gliocytes. NSCs have the ability to generate and regenerate the brain, indicating the possibility of cell-based therapies for neurological disorders. miR-124 has been demonstrated as a modulator in the survival, expansion, and differentiation of NSCs. However, the underlying molecular mechanisms of miR-124 in NSC development are still far from being understood. The expressions of miR-124, dishevelled binding antagonist of beta-catenin 1 (DACT1), ki-67, Nestin, β-tubulin III, glial fibrillary acidic protein (GFAP), β-catenin, cyclinD1, and glycogen synthase kinase-3β (GSK-3β) were examined by qRT-PCR or western blot. Bioinformatics and Dual-Luciferase reporter assay were used to identify the interaction between miR-124 and DACT1. MTS analysis was performed to measure the viability of NSCs. Enhanced expression of miR-124 and lowered expression of DACT1 were observed during a 14-day NSC differentiation period. DACT1 was verified as a direct target of miR-124. Moreover, overexpression of miR-124 promoted NSC proliferation and induced neuron-specific differentiation, presented as increased cell viability, higher neurosphere number, elevated ki-67, Nestin, β-tubulin III expressions, and decreased GFAP expression. Similarly, DACT1 downregulation facilitated proliferation and neuronal differentiation of NSCs. Furthermore, DACT1 overexpression impaired miR-124-induced proliferation and differentiation of NSCs. Additionally, miR-124 stimulated Wnt/β-catenin signaling via suppressing DACT1 expression. miR-124 promoted proliferation and induced NSC differentiation to neurons by activation of Wnt/β-catenin pathway via targeting DACT1, providing a potential target and aiding the development of cell-based therapies for neurological disorders.

Defective Microglia-Neuronal Communication during Demyelinating Disease Correlates with Altered Neurogenesis

Abstract Inflammatory demyelination is a hallmark of the pathology of multiple sclerosis (MS) and microglia are considered responsible for exacerbating myelin loss. The link between inflammation and neural stem cell (NSC) development, key events during the remyelination process, is being increasingly recognized. However, the mechanisms that connect microglia, myelination and inflammatory-mediated oxidative damage are unclear. Fractalkine (FKN), a neuronal derived chemokine, and its unique receptor, CX3CR1, expressed by microglia, are highly abundant in the healthy brain, and our laboratory has reported severe CNS demyelination and neuronal damage in EAE-induced CX3CR1-KO mice. Our hypothesis is that CX3CR1-defective signaling promotes inflammatory-mediated damage to neuron and myelin-forming cells. We found that expression of the adhesive defective human variant hCX3CR1-M280 caused enhanced disease severity, increased demyelination and exacerbated neuronal loss, mirroring the findings we observed in CX3CR1KO mice. Also, enhanced demyelination was observed after 4 wk of cuprizone treatment. Densities of NG2+ glia were comparable between WT and CX3CR1KO mice, but a significant reduction in DCX+ cells in the subgranular zone of the hippocampus was observed. In addition, lipocalin-2 an acute phase protein, linked to resolution of inflammation by reducing iron-induced toxicity, was found upregulated in cuprizone-induced demyelinated lesions of WT but not CX3CR1KO or hCX3CR1M280 mice. Thus, the FKN/CX3CR1 signaling pathway in microglia plays an under-appreciated role in MS, perhaps via anti-inflammatory processes that enhance neurogenesis through regulation of oxidative damage.

Update on the Role of Stem Cells in the Treatment of Hirschsprung Disease.

Stem cells possess the ability of self-renewal and the potency to differentiate into multiple cell lineages. Somatic stem cells are present in adult tissues, but they usually exhibit limited differentiation capacity and life span. On the other hand, somatic cells from adult tissues can be reprogrammed into induced pluripotent stem cells (iPSCs) that retain a full differentiation capacity with unlimited self-renewal ability. Autologous origin of iPSCs makes them an ideal source of cells for regenerative medicine to replenish the missing or damaged cells in the patients. iPSCs nowadays have also been widely used to build human disease models to study pathological mechanisms of the diseases. Hirschsprung disease (HSCR) is a congenital disorder caused by defects in the development of enteric neural crest stem cells. The failures of the ENCCs to proliferate, differentiate, and/or migrate lead to the absence of enteric neurons in the distal colon, resulting in colonic motility dysfunction. The lack of effective treatment for HSCR urges continuous efforts to develop new therapies for this congenital disorder. In this review, we will discuss the potential applications of somatic stem cells and iPSCs for the cell-based therapy of HSCR. We will also highlight the recent advances in stem cell research for the establishment of human HSCR models for the development of novel therapies.

Validation and bioinformatic analysis of propofol-induced differentially expressed microRNAs in primary cultured neural stem cells

Propofol, a widely used intravenous anesthetic, was previously considered as a neuroprotective agent. Recently, however, accumulating evidence suggests that it may cause neurotoxicity, especially in the development of neural stem cells (NSCs). The potential mechanisms contributing to propofol-induced neurotoxicity during neurogenesis, such as those involving microRNAs (miRNAs), are still unknown. In this study, a total of 27 differentially expressed miRNAs were identified in our initial screen and 6 miRNAs were validated by qRT-PCR. Three miRNAs were up-regulated (miR-377-5p, miR-194-3p and miR-143-5p), and three were down-regulated (miR-3583-3p, miR-466b-5p and miR-410-5p). Following gene ontology and KEGG pathway enrichment analysis, Gabbr1, Canca1b and Gabbr2, which are enriched in the GABAergic synapse pathway, were selected as genes potentially playing a role in propofol-induced neurotoxicity. Gabbr1 and Cacna1b, which are targeted by miRNAs that are up-regulated following propofol exposure, showed decreased expression at the mRNA and protein levels. Gabbr2, targeted by miRNAs that were down-regulated following treatment with propofol, was up-regulated at both the levels of mRNA and protein expression. The two clusters of miRNAs that show differential expression following propofol exposure may act in a synergistic manner to regulate several genes simultaneously during the development of NSCs. Our results may contribute to clarify the molecular mechanism and provide potential therapeutic targets for propofol induced neurotoxicity.

Deciphering the Dynamical Origin of Mixed Population during Neural Stem Cell Development

Neural stem cells (NSCs) often give rise to a mixed population of cells during differentiation. However, the dynamical origin of these mixed states is poorly understood. In this article, our mathematical modeling study demonstrates that the bone morphogenetic protein 2 (BMP2) mediated disparate differentiation dynamics of NSCs in central and peripheral nervous systems essentially function through two distinct bistable switches that are mutually interconnected via a mushroom-like bifurcation. Stochastic simulations of the model reveal that the mixed population originates due to the existence of these bistable switching regulations and that the maintenance of such mixed states depends on the level of stochastic fluctuations of the system. It further demonstrates that due to extrinsic variability, cells in an NSC population can dynamically transit from mushroom to a unique isola kind of bifurcation state, which essentially extends the range of the BMP2-driven mixed population state during differentiation. Importantly, the model predicts that by individually altering the expression level of key regulatory proteins, the NSCs can be converted entirely to a preferred phenotype for BMP2 doses that previously resulted in a mixed population. Our findings show that efficient neuronal regeneration can be achieved by systematically maneuvering the differentiation dynamics.

Developmental neurotoxic effects of a low dose of TCE on a 3-D neurosphere system.

Trichloroethylene (TCE) is one of the industrial toxic byproducts that now persist in the air, soil, and water. Several studies have already illustrated the toxic effect of high doses of TCE on the biological functions of several organs. This study aims to highlight the toxic impact of a low dose of TCE (1 μmol/L) on the development of rat neural stem cells (NSCs). The subventricular zones (SVZ) of rat pup's brains were collected and minced, and the harvested cells were cultured in the presence of neural growth factors B27/N2 to develop neurospheres. The cells were then exposed to a dose of 1 μmol/L TCE for 1 or 2 weeks. The outcomes indicated a remarkable inhibitory effect of TCE on the differentiation capacity of NSCs, which was confirmed by down-regulation of the astrocyte marker GFAP The inhibitory effect of TCE on the proliferation of NSCs was identified by the reductions in neurosphere diameter, Ki67 expression, and cell cycle arrest at the G1/S phase. Immunolabelling with annexin V indicated the proapoptotic effect of TCE exposure. PCR results revealed a TCE-mediated suppression of the expression of the antioxidant enzyme SOD1. This paper illustrates, for the first time, a detailed examination of the toxic effects of an environmentally low dose of TCE on NCSs at the transcriptional, translational, and functional levels.

Expression of a SOX1 overlapping transcript in neural differentiation and cancer models

ABSTRACTSOX1 is a member of the SOXB1 subgroup of transcription factors involved in early embryogenesis, CNS development and maintenance of neural stem cells. The structure and regulation of the human SOX1 locus has been less studied than that of SOX2, another member of the SOXB1 subgroup for which an overlapping transcript has been reported. Here we report that the SOX1 locus harbours a SOX1 overlapping transcript (SOX1-OT), and describe expression, splicing variants and detection of SOX1-OT in different stem and cancer cells. RT-PCR and RACE experiments were performed to detect and characterise the structure of SOX1-OT in neuroprogenitor cultures and across different cancer cell lines. SOX1-OT was found to present a complex structure including several unannotated exons, different transcript variants and at least two potential transcription start sites. SOX1-OT was found to be highly expressed in differentiated neural stem cells across different time points of differentiation, and its expression correlated with SOX1 gene expression. Concomitant expression of SOX1 and SOX1-OT was further observed in several cancer cell models. While the function of this transcript is unknown, the regulatory role reported for other lncRNAs strongly suggests a possible role for SOX1-OT in regulating SOX1 expression, as previously observed for SOX2. The elucidation of the genetic and regulatory context governing SOX1 expression will contribute to clarifying its role in stem cell differentiation and tumorigenesis.