Fetal Neural Stem Cells Research Articles

Photoactive Poly(3-hexylthiophene) Nanoweb for Optoelectrical Stimulation to Enhance Neurogenesis of Human Stem Cells.

Optoelectrical manipulation has recently gained attention for cellular engineering; however, few material platforms can be used to efficiently regulate stem cell behaviors via optoelectrical stimulation. In this study, we developed nanoweb substrates composed of photoactive polymer poly(3-hexylthiophene) (P3HT) to enhance the neurogenesis of human fetal neural stem cells (hfNSCs) through photo-induced electrical stimulation.Methods: The photoactive nanoweb substrates were fabricated by self-assembled one-dimensional (1D) P3HT nanostructures (nanofibrils and nanorods). The hfNSCs cultured on the P3HT nanoweb substrates were optically stimulated with a green light (539 nm) and then differentiation of hfNSCs on the substrates with light stimulation was examined. The utility of the nanoweb substrates for optogenetic application was tested with photo-responsive hfNSCs engineered by polymer nanoparticle-mediated transfection of an engineered chimeric opsin variant (C1V1)-encoding gene.Results: The nanoweb substrates provided not only topographical stimulation for activating focal adhesion signaling of hfNSCs, but also generated optoelectrical stimulation via photochemical and charge-transfer reactions upon exposure to 539 nm wavelength light, leading to significantly enhanced neuronal differentiation of hfNSCs. The optoelectrically stimulated hfNSCs exhibited mature neuronal phenotypes with highly extended neurite formation and functional neuron-like electrophysiological features of sodium currents and action potentials. Optoelectrical stimulation with 539 nm light simultaneously activated both C1V1-modified hfNSCs and nanoweb substrates, which upregulated the expression and activation of voltage-gated ion channels in hfNSCs and further increased the effect of photoactive substrates on neuronal differentiation of hfNSCs.Conclusion: The photoactive nanoweb substrates developed in this study may serve as platforms for producing stem cell therapeutics with enhanced neurogenesis and neuromodulation via optoelectrical control of stem cells.

The effect of spontaneous locomotor activity restriction on the course of spasticity syndrome after spinal cord injury and NeuroGeltm associated with neural stem cells implantation

Objective: To examine the effect of NeuroGelTM and xenogenic fetal neural stem cells (NSC) coimplantation coupled with the liberalization of spontaneous locomotor activity on the rat's hind limb motor function recovery and spastisity syndrome dynamic after experimental spinal cord injury. Materials and methods. Animals: outbred albino rats (males, 5.5 months, 250 g); experimental groups: 1 – spinal cord injury only (n=16); 2 – spinal cord injury + immediate homotopical transplantation of NeuroGelTM (n=20); 3 – spinal cord injury + analogous transplantation of NeuroGelTM in association with fetal (E17) mouse NSC (n=12). Group 3 included a subgroup 3sc, the animals of which (n=6) were kept in small cages (30×40×10 cm), and subgroup 3lc, whose animals (n=6) were kept in large cages (40×30×15 cm). Model of injury – left-side spinal cord hemisection at Т11; the duration of observation – 28 weeks; monitoring of the ipsilateral hindlimb (IHL) function indicator (FI) and spasticity indicator (SI) – the Вasso-Вeattie-Вresnahan scale (BBB) and Ashworth scale, respectively. Rersults. Restriction of spontaneous locomotor activity slows paretic limb motor function recovery during 1st month, reduces the duration of significant recovery in the late period of injury. At the 28th week of observation IHL FI in group 1 was 1,6±0,5 points of ВВВ, in group 2 – 8,4±0,9 points , in group 3 – 13,1±0,9 points , in subgroup 3sc – 12,6±1,4 points, in subgroup 3lc - 13,7±0,8 points of ВВВ. At this term of observation IHL SI in group 1 was 2,5±0,4 points of the Ashworth scale, in groupe 2 – 1,7±0,2 points, in group 3 – 1,3±0,1 points, in subgroup 3sc – 1,4±0,2, in subgroup 3lc – 1,2±0,1 points of Ashworth scale. Significant difference of IHL SI values between group 2 and group 1 was found at the 7th day, 5th–7th and 12th-24th week, between group 3 and group 1 – at the 1st-2nd, 6th-7th and 16th-28th week, between group 3 and group 2 – at the 1st-2nd and 5th week, between subgroup 3lc and group 2 – at the 2nd and 5th week, between subgroup 3sc and group 2 – at the 2nd week, between subgroup 3lc and subgroup 3sc - was not found. Conclusion. Restriction of spontaneous locomotor activity in case of the tissue engineering–restorative intervention after spinal cord injury complicates the regeneration process, accelerates the formation of stable spasticity syndrome.

Subarachnoid Hemorrhage Promotes Proliferation, Differentiation, and Migration of Neural Stem Cells via BDNF Upregulation.

Patients who suffer from subarachnoid hemorrhage (SAH) usually have long-term neurological impairments. Endogenous neurogenesis might play a potential role in functional recovery after SAH; however, the underlying neurogenesis mechanism is still unclear. We assessed the extent of neurogenesis in the subventricular zone (SVZ) to better understand the neurogenesis mechanism after SAH. We performed a rat model of SAH to examine the extent of neurogenesis in the SVZ and assessed functional effects of the neurotrophic factors in the cerebrospinal fluid (CSF) on neural stem cells (NSCs) after SAH. In this study, the proliferation, differentiation, and migratory capacities of NSCs in the SVZ were significantly increased on days 5 and 7 post SAH. Furthermore, treatment of cultured rat fetal NSCs with the CSF collected from rats on days 5 and 7 post SAH enhanced their proliferation, differentiation, and migration. Enzyme-linked immunosorbent assay (ELISA) of the CSF detected a marked increase in the concentration of brain-derived neurotrophic factor (BDNF). Treating the cultured NSCs with recombinant BDNF (at the same concentration as that in the CSF) or with CSF from SAH rats, directly, stimulated proliferation, differentiation, and migration to a similar extent. BDNF expression was upregulated in the SVZ of rats on days 5 and 7 post SAH, and BDNF release occurred from NSCs, astrocytes, and microglia in the SVZ. These results indicate that SAH triggers the expression of BDNF, which promotes the proliferation, differentiation, and migration of NSCs in the SVZ after SAH.

Zika Virus Infects Neural Progenitors in the Adult Mouse Brain and Alters Proliferation

Zika virus (ZIKV)-related neuropathology is an important global health concern. Several studies have shown that ZIKV can infect neural stem cells in the developing brain, but infection in the adult brain has not been examined. Two areas in the adult mouse brain contain neural stem cells: the subventricular zone of the anterior forebrain and the subgranular zone of the hippocampus. Here, using 6-week-old mice triply deficient in interferon regulatory factor (IRF) as a model, we show that blood-borne ZIKV administration can lead to pronounced evidence ofZIKV infection in these adult neural stem cells, leading to cell death and reduced proliferation. Our data therefore suggest that adult as well as fetal neural stem cells are vulnerable to ZIKV neuropathology. Thus, although ZIKV is considered a transient infection in adult humans without marked long-term effects, there may in fact be consequences of exposure in the adult brain.

L-MYC Expression Maintains Self-Renewal and Prolongs Multipotency ofPrimary Human Neural Stem Cells.

SummaryPre-clinical studies indicate that neural stem cells (NSCs) can limit or reverse CNS damage through direct cell replacement, promotion of regeneration, or delivery of therapeutic agents. Immortalized NSC lines are in growing demand due to the inherent limitations of adult patient-derived NSCs, including availability, expandability, potential for genetic modifications, and costs. Here, we describe the generation and characterization of a new human fetal NSC line, immortalized by transduction with L-MYC (LM-NSC008) that in vitro displays both self-renewal and multipotent differentiation into neurons, oligodendrocytes, and astrocytes. These LM-NSC008 cells were non-tumorigenic in vivo, and migrated to orthotopic glioma xenografts in immunodeficient mice. When administered intranasally, LM-NSC008 distributed specifically to sites of traumatic brain injury (TBI). These data support the therapeutic development of immortalized LM-NSC008 cells for allogeneic use in TBI and other CNS diseases.

Zika Virus NS4A and NS4B Proteins Deregulate Akt-mTOR Signaling in Human Fetal Neural Stem Cells to Inhibit Neurogenesis and Induce Autophagy

The current widespread outbreak of Zika virus (ZIKV) infection has been linked to severe clinical birth defects, particularly microcephaly, warranting urgent study of the molecular mechanisms underlying ZIKV pathogenesis. Akt-mTOR signaling is one of the key cellular pathways essential for brain development and autophagy regulation. Here, we show that ZIKV infection of human fetal neural stem cells (fNSCs) causes inhibition of the Akt-mTOR pathway, leading to defective neurogenesis and aberrant activation of autophagy. By screening the three structural proteins and seven nonstructural proteins present in ZIKV, we found that two, NS4A and NS4B, cooperatively suppress the Akt-mTOR pathway and lead to cellular dysregulation. Corresponding proteins from the closely related dengue virus do not have the same effect on neurogenesis. Thus, our study highlights ZIKV NS4A and NS4B as candidate determinants of viral pathogenesis and identifies a mechanism of action for their effects, suggesting potential targets for anti-ZIKV therapeutic intervention.

RGDS- and SIKVAVS-Modified Superporous Poly(2-hydroxyethyl methacrylate) Scaffolds for Tissue Engineering Applications.

Three-dimensional hydrogel supports for mesenchymal and neural stem cells (NSCs) are promising materials for tissue engineering applications such as spinal cord repair. This study involves the preparation and characterization of superporous scaffolds based on a copolymer of 2-hydroxyethyl and 2-aminoethyl methacrylate (HEMA and AEMA) crosslinked with ethylene dimethacrylate. Ammonium oxalate is chosen as a suitable porogen because it consists of needle-like crystals, allowing their parallel arrangement in the polymerization mold. The amino group of AEMA is used to immobilize RGDS and SIKVAVS peptide sequences with an N-γ-maleimidobutyryloxy succinimide ester linker. The amount of the peptide on the scaffold is determined using 125 I radiolabeled SIKVAVS. Both RGDS- and SIKVAVS-modified poly(2-hydroxyethyl methacrylate) scaffolds serve as supports for culturing human mesenchymal stem cells (MSCs) and human fetal NSCs. The RGDS sequence is found to be better for MSC and NSC proliferation and growth than SIKVAVS.

Human neural stem cell-induced endothelial morphogenesis requires autocrine/paracrine and juxtacrine signaling.

Transplanted neural stem cells (NSC) interact with the host brain microenvironment. A neovascularization is commonly observed in the vicinity of the cell deposit, which is correlated with behavioral improvements. To elucidate the signaling mechanisms between human NSCs and endothelial cells (ECs), these were cocultured in an in vitro model in which NSC-induced endothelial morphogenesis produced a neurovascular environment. Soluble (autocrine/paracrine) and contact–mediated (juxtacrine) signaling molecules were evaluated for two conditionally immortalized fetal NSC lines derived from the cortical anlage (CTXOE03) and ganglionic eminence (STROC05), as well as an adult EC line (D3) derived from the cerebral microvasculature of a hippocampal biopsy. STROC05 were 4 times as efficient to induce endothelial morphogenesis compared to CTXOE03. The cascade of reciprocal interactions between NSCs and ECs in this process was determined by quantifying soluble factors, receptor mapping, and immunocytochemistry for extracellular matrix molecules. The mechanistic significance of these was further evaluated by pharmacological blockade. The sequential cell-specific regulation of autocrine/paracrine and juxtacrine signaling accounted for the differential efficiency of NSCs to induce endothelial morphogenesis. These in vitro studies shed new light on the reciprocal interactions between NSCs and ECs, which are pivotal for our mechanistic understanding of the efficacy of NSC transplantation.

Quantitative Phosphoproteomic Study Reveals that Protein Kinase A Regulates Neural Stem Cell Differentiation Through Phosphorylation of Catenin Beta-1 and Glycogen Synthase Kinase 3β.

Protein phosphorylation is central to the understanding of multiple cellular signaling pathways responsible for regulating the self-renewal and differentiation of neural stem cells (NSCs). Here we performed a large-scale phosphoproteomic analysis of rat fetal NSCs using strong cation exchange chromatography prefractionation and citric acid-assisted two-step enrichment with TiO2 strategy followed by nanoLC-MS/MS analysis. Totally we identified 32,546 phosphosites on 5,091 phosphoproteins, among which 23,945 were class I phosphosites, and quantified 16,000 sites during NSC differentiation. More than 65% of class I phosphosites were novel when compared with PhosphoSitePlus database. Quantification results showed that the early and late stage of NSC differentiation differ greatly. We mapped 69 changed phosphosites on 20 proteins involved in Wnt signaling pathway, including S552 on catenin beta-1 (Ctnnb1) and S9 on glycogen synthase kinase 3β (Gsk3β). Western blotting and real-time PCR results proved that Wnt signaling pathway plays critical roles in NSC fate determination. Furthermore, inhibition and activation of PKA dramatically affected the phosphorylation state of Ctnnb1 and Gsk3β, which regulates the differentiation of NSCs. Our data provides a valuable resource for studying the self-renewal and differentiation of NSCs. Stem Cells 2016;34:2090-2101.

Neural Stem Cells Transplantation as a Neuroprotective Strategy for Amyotrophic Lateral Sclerosis (ALS)

Introduction Amiotrophic lateral sclerosis (ALS) is a neurodegenerative disease with a prevalence rate of up to 7.4/100,000 and the overall risk of developing ALS over a lifetime is 1:400. Most patients die from respiratory failure following a course of progressive weakness. There is no effective therapy. Novel therapeutic strategies might be directed at replacing or repairing the damaged motor neurons; Cell therapy is emerging as a potential therapeutic option in ALS. The primary objective of our study was to verify the safety and tolerability of transplantation of human fetal neural stem cells (hNSCs) directly into the spinal cord of humans. The study was approved by the SuperiorNational Health Institute. Material and Methods 18 patients with definite ALS were recruited and treated. Patients were included if they had ALS of spinal onset with severe functional impairment of the lower limbs and mild functional impairment of the upper limbs without signs of respiratory failure. The patients were monitored by clinical evaluation which included the ALS-FRS scale, Norris score, bulbar score, and MRC strength scale. Respiratory assessment included clinical evaluation, pulmonary function tests, arterial blood gases analysis, and nocturnal cardio-respiratory monitoring. Neurophysiological assessments were made including EMG, and somatosensory evoked potentials. The neuroradiological assessment consisted of MRI of spinal cord and brain before and after Gadolinium DTA infusion. A clinical psychologist performed psychological evaluation including an interview and psychological tests. Human fetal brain tissue specimens, all derived from the forebrain, were routinely collected from fetuses at gestational ages greater than the 8th post-conceptional week. They were immediately dissected and used to generate hNSC lines under sterile conditions. Dissociation of brain tissue, primary culturing and cell propagation were performed according to the procedure described previously by Vescovi and colleagues. Six patients received hNSCs microinjections into the dorsal (T7-T9) cord tract (3 of them received unilateral hNSCs microinjection while the remaining ones received bilateral microinjections); the other twelve patients received hNSCs microinjections into the cervical (C5-C6) cord tract (3 of them received unilateral hNSCs microinjection while the remaining ones received bilateral microinjections). Results No patients manifested severe adverse events such as death, respiratory failure or permanent post-surgical neurological deficits. Minor adverse events were: intercostal pain (2 patients) which was reversible after a mean period of 3 days after surgery, arm and leg sensory dysesthesia (6 patients) which was reversible after a mean period of 6 weeks after surgery. No patient manifested bladder and bowel dysfunction, or arm and leg motor deficit. All patients showed a good acceptance of the procedure and no significant modifications of the psychological status or quality of life were observed. Conclusions Our results appear to demonstrate that the procedures of fetal stem cells transplantation into the spinal cord of humans are safe and well tolerated by ALS patients. The minimal side effects and the absence of detrimental effects on neurological function support further research in stem cell transplantation in carefully monitored patients with ALS.

LSD1 Mediates Neuronal Differentiation of Human Fetal Neural Stem Cells by Controlling the Expression of a Novel Target Gene, HEYL.

Histone-modifying enzymes dynamically regulate the chromatin status and have been implicated in the fate specification of stem cells, including neural stem cells (NSCs), which differentiate into three major cell types: neurons, astrocytes, and oligodendrocytes. Lysine-specific demethylase 1 (LSD1, also known as KDM1A) catalyzes the demethylation of H3K4me1/2 and H3K9me1/2, and it was recently suggested that functional disruption of LSD1 links to various human diseases. However, the mechanism by which LSD1 regulates human neural development remains unclear. Here, we present evidence that specific inhibition of LSD1 suppresses the neurogenesis of cultured human fetal NSCs (hfNSCs) isolated from the human fetal neocortex. Notably, we found that LSD1 directly associates with the promoter of the HEYL gene, and controls the demethylation of H3K4me2, which is accompanied by repression of HEYL expression during hfNSC neuronal differentiation. Furthermore, we also showed that HEYL expression is sufficient to inhibit the neuronal differentiation of hfNSCs. This mechanism seems to be primate-specific because mouse NSCs do not exhibit the LSD1 inhibitor-induced upregulation of Heyl. Our findings suggest that LSD1 plays an important role in primate neurogenesis and may contribute to the characterization of an evolved primate brain. Stem Cells 2016;34:1872-1882.

Neuregulin-1 attenuates cognitive function impairments in a transgenic mouse model of Alzheimer's disease.

The neuregulin (NRG) family of epidermal growth factor-related proteins is composed of a wide variety of soluble and membrane-bound proteins that exert their effects via the tyrosine kinase receptors ErbB2-ErbB4. In the nervous system, the functions of NRG1 are essential for peripheral myelination, the establishment and maintenance of neuromuscular and sensorimotor systems and the plasticity of cortical neuronal circuits. In the present study, we report that an intracerebroventricular infusion of NRG1 attenuated cognitive impairments in 13-month-old Tg2576 mice, an animal model of Alzheimer's disease (AD). In addition, according to Golgi-Cox staining, NRG1 rescued the reduction in the number of dendritic spines detected in the brains of Tg2576 mice compared with vehicle (PBS)-infused mice. This result was also corroborated in vitro as NRG1 attenuated the oligomeric amyloid beta peptide1-42 (Aβ1-42)-induced decrease in dendritic spine density in rat primary hippocampal neuron cultures. NRG1 also alleviated the decrease in neural differentiation induced by oligomeric Aβ1-42 in mouse fetal neural stem cells. Collectively, these results suggest that NRG1 has a therapeutic potential for AD by alleviating the reductions in dendritic spine density and neurogenesis found in AD brains.

Abstracts from The 12th Symposium of the International Neurotrauma Society February 1-4, 2016 Cape Town, South Africa.

Abstracts from The 12th Symposium of the International Neurotrauma Society February 1-4, 2016 Cape Town, South Africa.

Nanoparticle-mediated transcriptional modification enhances neuronal differentiation of human neural stem cells following transplantation in rat brain

Strategies to enhance survival and direct the differentiation of stem cells in vivo following transplantation in tissue repair site are critical to realizing the potential of stem cell-based therapies. Here we demonstrated an effective approach to promote neuronal differentiation and maturation of human fetal tissue-derived neural stem cells (hNSCs) in a brain lesion site of a rat traumatic brain injury model using biodegradable nanoparticle-mediated transfection method to deliver key transcriptional factor neurogenin-2 to hNSCs when transplanted with a tailored hyaluronic acid (HA) hydrogel, generating larger number of more mature neurons engrafted to the host brain tissue than non-transfected cells. The nanoparticle-mediated transcription activation method together with an HA hydrogel delivery matrix provides a translatable approach for stem cell-based regenerative therapy.

Multiparameter Characterization Confirms Apoptosis as the Primary Cause of Reduced Self-renewal Capacity in Cultured Human Fetal Neural Stem Cells

Background: Human fetal striatum-derived neural stem cells (hfsNSCs) are important in regenerative medicine; however, their ability to self-renew diminishes quickly following passages in culture. Typically when hfsNSC-derived neurospheres are dissociated by accutase, more than 90% of the cells survive, but only 6-8% of the cells are able to form secondary neurospheres. Our hypothesis is that the hfsNSCs that are unable to form new neurospheres become apoptotic. Methods/Results: Because the NSC apoptosis process has never been characterized in detail, we characterized hfsNSC apoptosis using multiparameter analysis and determined that the majority of hfsNSCs undergo apoptosis after passaging, which leads to a reduction in self-renewal. The replacement of trituration with vortexing decreases apoptosis, increases self-renewal, and does not affect NSC differentiation. When we used live cell staining with Annexin V, Hoechst 33342, and PI together, the apoptotic index was in agreement with what could be obtained using fixed-cell staining methods, including TUNEL and activated caspase-3 immunocytochemistry. NSC apoptosis could be divided into 9 stage types based on our live cell assay. Several types during early and late stages had similar staining profiles that could be further discriminated based on cell size. Conclusion: Apoptosis largely contributes to the low self-renewal of neurospheres, and replacing trituration with vortexing aided in alleviating NSC apoptosis. Multiparameter analysis is required for the identification of NSC apoptosis, particularly when live cell staining is used.

Characterization of Neurogenic Potential of Dental Pulp Stem Cells Cultured in Xeno/Serum-Free Condition: In Vitro and In Vivo Assessment.

Neural stem cells (NSCs) have a high potency for differentiation to neurons and glial cells for replacement of damaged cells and paracrine effects for the regeneration and remyelination of host axons. Dental pulp is known to have a potential to differentiate into neural-like cells; therefore, dental pulp may be used as an autologous cell source for neural repair. In this study, we selectively expanded stem cells from human dental pulp in an initial culture using NSC media under xeno- and serum-free conditions. At the initial step of primary culture, human dental pulp was divided into two groups according to the culture media: 10% fetal bovine serum medium group (FBS group) and NSC culture medium group (NSC group). In the NSC group relative to the FBS group, the expression of NSC markers and the concentrations of leukemia inhibitory factor, nerve growth factor, and stem cell factor were higher, although their expression levels were lower than those of human fetal NSCs. The transplanted cells of the NSC group survived well within the normal brain and injured spinal cord of rats and expressed nestin and Sox2. Under the xeno- and serum-free conditions, autologous human dental pulp-derived stem cells might prove useful for clinical cell-based therapies to repair damaged neural tissues.

Factor-Reduced Human Induced Pluripotent Stem Cells Efficiently Differentiate into Neurons Independent of the Number of Reprogramming Factors.

Reprogramming of somatic cells into induced pluripotent stem cells (iPSCs) by overexpression of the transcription factors OCT4, SOX2, KLF4, and c-Myc holds great promise for the development of personalized cell replacement therapies. In an attempt to minimize the risk of chromosomal disruption and to simplify reprogramming, several studies demonstrated that a reduced set of reprogramming factors is sufficient to generate iPSC. We recently showed that a reduction of reprogramming factors in murine cells not only reduces reprogramming efficiency but also may worsen subsequent differentiation. To prove whether this is also true for human cells, we compared the efficiency of neuronal differentiation of iPSC generated from fetal human neural stem cells with either one (OCT4; hiPSC1F-NSC) or two (OCT4, KLF4; hiPSC2F-NSC) reprogramming factors with iPSC produced from human fibroblasts using three (hiPSC3F-FIB) or four reprogramming factors (hiPSC4F-FIB). After four weeks of coculture with PA6 stromal cells, neuronal differentiation of hiPSC1F-NSC and hiPSC2F-NSC was as efficient as iPSC3F-FIB or iPSC4F-FIB. We conclude that a reduction of reprogramming factors in human cells does reduce reprogramming efficiency but does not alter subsequent differentiation into neural lineages. This is of importance for the development of future application of iPSC in cell replacement therapies.

多価不飽和脂肪酸による神経幹細胞の分化制御機構の解明

多価不飽和脂肪酸による神経幹細胞の分化制御機構の解明

BacMam-mediated gene transfer into neural cell types

Modified baculoviruses containing a mammalian promoter, known as BacMam, have emerged as a key method for labeling diverse cell types, owing to its high transduction efficiencies, minimal cytopathic effects and the ability to carry DNA fragments larger than 40 KB. The transient expression without viral replication or integration into the mammalian genome makes BacMam an ideal method to label primary and adult stem cells. In the current study, we describe how BacMam enables efficient transduction and robust gene expression in neural stem cells (NSC) and progenitor cells. Utilizing the H9 human embryonic stem cell (hESC), we generated neurospheres, neural rosettes and Neural Stem Cells (NSC) and transduced them with BacMam 2.0 GFP. Commercially available human NSCs (h-NSC) and rat fetal NSCs (rNSCs) were also transduced with BacMam 2.0 GFP. Primary rat neurons and post-differentiated human NSCs were transduced with either BacMam 2.0 GFP or Plasma Membrane-GFP BacMam 2.0 and Synaptophysin-RFP BacMam 2.0 to assess the capability of gene expression in mature neural cell types. Using a GFP marker, we demonstrated that the BacMam virus can consistently transduce neural stem cells with high efficiency and low toxicity. Transduction was also enabled in differentiated NSC, and the expression in primary neurons was dependent on the age of the culture, with preferential labeling of astrocytes in older cultures. BacMam mediated gene delivery provides a superior platform for introducing diverse gene elements, creating assay-ready cells among young neuronal cells and mature astrocytes.

New insight into LSD1 function in human cortical neurogenesis.

The cerebral cortex of primates has evolved massively and intricately in comparison to that of other species. Accumulating evidence indicates that this is caused by changes in cell biological features of neural stem cells (NSCs), which differentiate into neurons and glial cells during development. The fate of NSCs during rodent cortical development is stringently regulated by epigenetic factors, such as histone modification enzymes, but the role of these factors in human corticogenesis is largely unknown. We have recently discovered that a lysine-specific demethylase 1 (LSD1), which catalyzes the demethylation of methyl groups in the histone tail, plays a unique role in human fetal NSCs (hfNSCs). We show that, unlike the role previously reported in mice, LSD1 in hfNSCs is necessary for neuronal differentiation and controls the expression of HEYL, one of the NOTCH target genes, by modulating the methylation level of histones on its promoter region. Interestingly, LSD1-regulation of Heyl expression is not observed in mouse NSCs. Furthermore, we first demonstrated that HEYL is able to maintain the undifferentiated state of hfNSCs. Our findings provide a new insight indicating that LSD1 may be a key player in the development and characterization of the evolved cerebral cortex.