Science.gov

Sample records for neural cell growth

  1. Conditioned medium from neural stem cells inhibits glioma cell growth.

    PubMed

    Li, Z; Zhong, Q; Liu, H; Liu, P; Wu, J; Ma, D; Chen, X; Yang, X

    2016-10-31

    Malignant glioma is one of the most common brain tumors in the central nervous system. Although the significant progress has been made in recent years, the mortality is still high and 5-year survival rate is still very low. One of the leading causes to the high mortality for glioma patients is metastasis and invasion. An efficient method to control the tumor metastasis is a promising way to treat the glioma. Previous reports indicated that neural stem cells (NSCs) were served as a delivery vector to the anti-glioma therapy. Here, we used the conditioned medium from rat NSCs (NSC-CM) to culture the human glioblastoma cell lines. We found that NSC-CM could inhibit the glioma cell growth, invasion and migration in vitro and attenuate the tumor growth in vivo. Furthermore, this anti-glioma effect was mediated by the inactivation of mitogen activated protein kinase (MAPK) pathway. Above all, this study provided the direct evidence to put forward a simple and efficient method in the inhibition of glioma cells/tumor growth, potentially advancing the anti-glioma therapy.

  2. Reflectin as a Material for Neural Stem Cell Growth

    PubMed Central

    2015-01-01

    Cephalopods possess remarkable camouflage capabilities, which are enabled by their complex skin structure and sophisticated nervous system. Such unique characteristics have in turn inspired the design of novel functional materials and devices. Within this context, recent studies have focused on investigating the self-assembly, optical, and electrical properties of reflectin, a protein that plays a key role in cephalopod structural coloration. Herein, we report the discovery that reflectin constitutes an effective material for the growth of human neural stem/progenitor cells. Our findings may hold relevance both for understanding cephalopod embryogenesis and for developing improved protein-based bioelectronic devices. PMID:26703760

  3. Nerve growth factor promotes in vitro proliferation of neural stem cells from tree shrews.

    PubMed

    Xiong, Liu-Lin; Chen, Zhi-Wei; Wang, Ting-Hua

    2016-04-01

    Neural stem cells promote neuronal regeneration and repair of brain tissue after injury, but have limited resources and proliferative ability in vivo. We hypothesized that nerve growth factor would promote in vitro proliferation of neural stem cells derived from the tree shrews, a primate-like mammal that has been proposed as an alternative to primates in biomedical translational research. We cultured neural stem cells from the hippocampus of tree shrews at embryonic day 38, and added nerve growth factor (100 μg/L) to the culture medium. Neural stem cells from the hippocampus of tree shrews cultured without nerve growth factor were used as controls. After 3 days, fluorescence microscopy after DAPI and nestin staining revealed that the number of neurospheres and DAPI/nestin-positive cells was markedly greater in the nerve growth factor-treated cells than in control cells. These findings demonstrate that nerve growth factor promotes the proliferation of neural stem cells derived from tree shrews.

  4. Nerve growth factor promotes in vitro proliferation of neural stem cells from tree shrews

    PubMed Central

    Xiong, Liu-lin; Chen, Zhi-wei; Wang, Ting-hua

    2016-01-01

    Neural stem cells promote neuronal regeneration and repair of brain tissue after injury, but have limited resources and proliferative ability in vivo. We hypothesized that nerve growth factor would promote in vitro proliferation of neural stem cells derived from the tree shrews, a primate-like mammal that has been proposed as an alternative to primates in biomedical translational research. We cultured neural stem cells from the hippocampus of tree shrews at embryonic day 38, and added nerve growth factor (100 μg/L) to the culture medium. Neural stem cells from the hippocampus of tree shrews cultured without nerve growth factor were used as controls. After 3 days, fluorescence microscopy after DAPI and nestin staining revealed that the number of neurospheres and DAPI/nestin-positive cells was markedly greater in the nerve growth factor-treated cells than in control cells. These findings demonstrate that nerve growth factor promotes the proliferation of neural stem cells derived from tree shrews. PMID:27212919

  5. Growth hormone (GH), brain development and neural stem cells.

    PubMed

    Waters, M J; Blackmore, D G

    2011-12-01

    A range of observations support a role for GH in development and function of the brain. These include altered brain structure in GH receptor null mice, and impaired cognition in GH deficient rodents and in a subgroup of GH receptor defective patients (Laron dwarfs). GH has been shown to alter neurogenesis, myelin synthesis and dendritic branching, and both the GH receptor and GH itself are expressed widely in the brain. We have found a population of neural stem cells which are activated by GH infusion, and which give rise to neurons in mice. These stem cells are activated by voluntary exercise in a GH-dependent manner. Given the findings that local synthesis of GH occurs in the hippocampus in response to a memory task, and that GH replacement improves memory and cognition in rodents and humans, these new observations warrant a reappraisal of the clinical importance of GH replacement in GH deficient states.

  6. Tailored Fringed Platforms Produced by Laser Interference for Aligned Neural Cell Growth.

    PubMed

    Peláez, Ramón J; González-Mayorga, Ankor; Gutiérrez, María C; García-Rama, Concepción; Afonso, Carmen N; Serrano, María C

    2016-02-01

    Ordering neural cells is of interest for the development of neural interfaces. The aim of this work is to demonstrate an easy-to-use, versatile, and cost/time effective laser-based approach for producing platforms that promote oriented neural growth. We use laser interferometry to generate fringed channels with topography on partially reduced graphene oxide layers as a proof-of-concept substrate. We study cell adhesion, morphology, viability, and differentiation in cultures of embryonic neural progenitor cells on platforms with a 9.4 μm period. Results evidence that fringed platforms significantly promote neurite alignment (≈50% at 6 d), while preserving viability and neural differentiation.

  7. Neural Growth Factor Stimulates Proliferation of Spinal Cord Derived-Neural Precursor/Stem Cells

    PubMed Central

    Han, Youngmin

    2016-01-01

    Objective Recently, regenerative therapies have been used in clinical trials (heart, cartilage, skeletal). We don't make use of these treatments to spinal cord injury (SCI) patients yet, but regenerative therapies are rising interest in recent study about SCI. Neural precursor/stem cell (NPSC) proliferation is a significant event in functional recovery of the central nervous system (CNS). However, brain NPSCs and spinal cord NPSCs (SC-NPSCs) have many differences including gene expression and proliferation. The purpose of this study was to investigate the influence of neural growth factor (NGF) on the proliferation of SC-NPSCs. Methods NPSCs (2×104) were suspended in 100 µL of neurobasal medium containing NGF-7S (Sigma-Aldrich) and cultured in a 96-well plate for 12 days. NPSC proliferation was analyzed five times for either concentration of NGF (0.02 and 2 ng/mL). Sixteen rats after SCI were randomly allocated into two groups. In group 1 (SCI-vehicle group, n=8), animals received 1.0 mL of the saline vehicle solution. In group 2 (SCI-NGF group, n=8), the animals received single doses of NGF (Sigma-Aldrich). A dose of 0.02 ng/mL of NGF or normal saline as a vehicle control was intra-thecally injected daily at 24 hour intervals for 7 days. For Immunohistochemistry analysis, rats were sacrificed after one week and the spinal cords were obtained. Results The elevation of cell proliferation with 0.02 ng/mL NGF was significant (p<0.05) but was not significant for 2 ng/mL NGF. The optical density was increased in the NGF 0.02 ng/mL group compared to the control group and NGF 2 ng/mL groups. The density of nestin in the SCI-NGF group was significantly increased over the SCI-vehicle group (p<0.05). High power microscopy revealed that the density of nestin in the SCI-NGF group was significantly increased over the SCI-vehicle group. Conclusion SC-NPSC proliferation is an important pathway in the functional recovery of SCI. NGF enhances SC-NPSC proliferation in vitro and in

  8. Effects of epidermal growth factor on neural crest cells in tissue culture

    SciTech Connect

    Erickson, C.A.; Turley, E.A.

    1987-04-01

    Epidermal growth factor (EGF) stimulates the release of hyaluronic acid (HA) and chondroitin sulfate proteoglycan (CSPG) from quail trunk neural crest cultures in a dose-dependent fashion. It also promotes the expression of cell-associated heparan sulfate proteoglycan (HSPG) as detected by immunofluorescence and immunoprecipitation of the /sup 3/H-labeled proteoglycan. Furthermore, EGF stimulates (/sup 3/H)thymidine incorporation into total cell DNA. These results raise the possibility that EGF or an analogous growth factor is involved in regulation of neural crest cell morphogenesis.

  9. PHBV microspheres as neural tissue engineering scaffold support neuronal cell growth and axon-dendrite polarization.

    PubMed

    Chen, Wenhui; Tong, Yen Wah

    2012-02-01

    Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) microspheres, with properties such as slower degradation and more efficient drug delivery properties, have important benefits for neural tissue engineering. Our previous studies have shown PHBV microspheres to improve cell growth and differentiation. This study aimed to investigate if PHBV microspheres would support neurons to extend these benefits to neural tissue engineering. PHBV microspheres' suitability as neural tissue engineering scaffolds was investigated using PC12 cells, cortical neurons (CNs), and neural progenitor cells (NPCs) to cover a variety of neuronal types for different applications. Microspheres were fabricated using an emulsion-solvent-evaporation technique. DNA quantification, cell viability assays, and immunofluorescent staining were carried out. PC12 cultures on PHBV microspheres showed growth trends comparable to two-dimensional controls. This was further verified by staining for cell spreading. Also, CNs expressed components of the signaling pathway on PHBV microspheres, and had greater axon-dendrite segregation (4.1 times for axon stains and 2.3 times for dendrite stains) than on coverslips. NPCs were also found to differentiate into neurons on the microspheres. Overall, the results indicate that PHBV microspheres, as scaffolds for neural tissue engineering, supported a variety of neuronal cell types and promoted greater axon-dendrite segregation.

  10. Elastic modulus affects the growth and differentiation of neural stem cells

    PubMed Central

    Jiang, Xian-feng; Yang, Kai; Yang, Xiao-qing; Liu, Ying-fu; Cheng, Yuan-chi; Chen, Xu-yi; Tu, Yue

    2015-01-01

    It remains poorly understood if carrier hardness, elastic modulus, and contact area affect neural stem cell growth and differentiation. Tensile tests show that the elastic moduli of Tiansu and SMI silicone membranes are lower than that of an ordinary dish, while the elastic modulus of SMI silicone membrane is lower than that of Tiansu silicone membrane. Neural stem cells from the cerebral cortex of embryonic day 16 Sprague-Dawley rats were seeded onto ordinary dishes as well as Tiansu silicone membrane and SMI silicone membrane. Light microscopy showed that neural stem cells on all three carriers show improved adherence. After 7 days of differentiation, neuron specific enolase, glial fibrillary acidic protein, and myelin basic protein expression was detected by immunofluorescence. Moreover, flow cytometry revealed a higher rate of neural stem cell differentiation into astrocytes on Tiansu and SMI silicone membranes than on the ordinary dish, which was also higher on the SMI than the Tiansu silicone membrane. These findings confirm that all three cell carrier types have good biocompatibility, while SMI and Tiansu silicone membranes exhibit good mechanical homogenization. Thus, elastic modulus affects neural stem cell differentiation into various nerve cells. Within a certain range, a smaller elastic modulus results in a more obvious trend of cell differentiation into astrocytes. PMID:26604916

  11. Cellular Neural Network Models of Growth and Immune of Effector Cells Response to Cancer

    NASA Astrophysics Data System (ADS)

    Su, Yongmei; Min, Lequan

    Four reaction-diffusion cellular neural network (R-D CNN) models are set up based on the differential equation models for the growths of effector cells and cancer cells, and the model of the immune response to cancer proposed by Allison et al. The CNN models have different reaction-diffusion coefficients and coupling parameters. The R-D CNN models may provide possible quantitative interpretations, and are good in agreement with the in vitro experiment data reported by Allison et al.

  12. Effects of Nerve Growth Factor and Basic Fibroblast Growth Factor Promote Human Dental Pulp Stem Cells to Neural Differentiation.

    PubMed

    Zhang, Jinlong; Lian, Min; Cao, Peipei; Bao, Guofeng; Xu, Guanhua; Sun, Yuyu; Wang, Lingling; Chen, Jiajia; Wang, Yi; Feng, Guijuan; Cui, Zhiming

    2017-04-01

    Dental pulp stem cells (DPSCs) were the most widely used seed cells in the field of neural regeneration and bone tissue engineering, due to their easily isolation, lack of ethical controversy, low immunogenicity and low rates of transplantation rejection. The purpose of this study was to investigate the role of basic fibroblast growth factor (bFGF) and nerve growth factor (NGF) on neural differentiation of DPSCs in vitro. DPSCs were cultured in neural differentiation medium containing NGF and bFGF alone or combination for 7 days. Then neural genes and protein markers were analyzed using western blot and RT-PCR. Our study revealed that bFGF and NGF increased neural differentiation of DPSCs synergistically, compared with bFGF and NGF alone. The levels of Nestin, MAP-2, βIII-tubulin and GFAP were the most highest in the DPSCs + bFGF + NGF group. Our results suggested that bFGF and NGF signifiantly up-regulated the levels of Sirt1. After treatment with Sirt1 inhibitor, western blot, RT-PCR and immunofluorescence staining showed that neural genes and protein markers had markedly decreased. Additionally, the ERK and AKT signaling pathway played a key role in the neural differentiation of DPSCs stimulated with bFGF + NGF. These results suggested that manipulation of the ERK and AKT signaling pathway may be associated with the differentiation of bFGF and NGF treated DPSCs. Our date provided theoretical basis for DPSCs to treat neurological diseases and repair neuronal damage.

  13. Growth hormone and cell survival in the neural retina: caspase dependence and independence.

    PubMed

    Harvey, Steve; Baudet, Marie-Laure; Sanders, Esmond J

    2006-11-06

    Growth hormone has recently been shown to be expressed in the retinal ganglion cells of embryonic chicks, in which it induces cell survival during neurogenesis. The mechanism of this action has been examined in neural retina explants from 6-day-old and 8-day-old embryos that were incubated for 48 h in 10 M growth hormone, to reduce the number of spontaneous apoptotic cells. This anti-apoptotic action was accompanied by a reduction in caspase-3 expression and, at embryonic day 8, by reduced expression of apoptosis inducing factor-1, which is caspase independent. These actions were specific, as other genes involved in apoptotic signaling (bcl-2, bcl-x, bid and inhibitor of apoptosis protein-1) were unaffected. These results therefore demonstrate caspase-dependent and caspase-independent pathways in growth hormone-induced retinal cell survival.

  14. Covalent growth factor tethering to direct neural stem cell differentiation and self-organization.

    PubMed

    Ham, Trevor R; Farrag, Mahmoud; Leipzig, Nic D

    2017-04-15

    Tethered growth factors offer exciting new possibilities for guiding stem cell behavior. However, many of the current methods present substantial drawbacks which can limit their application and confound results. In this work, we developed a new method for the site-specific covalent immobilization of azide-tagged growth factors and investigated its utility in a model system for guiding neural stem cell (NSC) behavior. An engineered interferon-γ (IFN-γ) fusion protein was tagged with an N-terminal azide group, and immobilized to two different dibenzocyclooctyne-functionalized biomimetic polysaccharides (chitosan and hyaluronan). We successfully immobilized azide-tagged IFN-γ under a wide variety of reaction conditions, both in solution and to bulk hydrogels. To understand the interplay between surface chemistry and protein immobilization, we cultured primary rat NSCs on both materials and showed pronounced biological effects. Expectedly, immobilized IFN-γ increased neuronal differentiation on both materials. Expression of other lineage markers varied depending on the material, suggesting that the interplay of surface chemistry and protein immobilization plays a large role in nuanced cell behavior. We also investigated the bioactivity of immobilized IFN-γ in a 3D environment in vivo and found that it sparked the robust formation of neural tube-like structures from encapsulated NSCs. These findings support a wide range of potential uses for this approach and provide further evidence that adult NSCs are capable of self-organization when exposed to the proper microenvironment. For stem cells to be used effectively in regenerative medicine applications, they must be provided with the appropriate cues and microenvironment so that they integrate with existing tissue. This study explores a new method for guiding stem cell behavior: covalent growth factor tethering. We found that adding an N-terminal azide-tag to interferon-γ enabled stable and robust Cu-free 'click

  15. Neural Progenitor Cells Promote Axonal Growth and Alter Axonal mRNA Localization in Adult Neurons

    PubMed Central

    Merianda, Tanuja T.; Jin, Ying

    2017-01-01

    Abstract The inhibitory environment of the spinal cord and the intrinsic properties of neurons prevent regeneration of axons following CNS injury. However, both ascending and descending axons of the injured spinal cord have been shown to regenerate into grafts of embryonic neural progenitor cells (NPCs). Previous studies have shown that grafts composed of glial-restricted progenitors (GRPs) and neural-restricted progenitors (NRPs) can provide a permissive microenvironment for axon growth. We have used cocultures of adult rat dorsal root ganglion (DRG) neurons together with NPCs, which have shown significant enhancement of axon growth by embryonic rat GRP and GRPs/NRPs, both in coculture conditions and when DRGs are exposed to conditioned medium from the NPC cultures. This growth-promoting effect of NPC-conditioned medium was also seen in injury-conditioned neurons. DRGs cocultured with GRPs/NRPs showed altered expression of regeneration-associated genes at transcriptional and post-transcriptional levels. We found that levels of GAP-43 mRNA increased in DRG cell bodies and axons. However, hepcidin antimicrobial peptide (HAMP) mRNA decreased in the cell bodies of DRGs cocultured with GRPs/NRPs, which is distinct from the increase in cell body HAMP mRNA levels seen in DRGs after injury conditioning. Endogenous GAP-43 and β-actin mRNAs as well as reporter RNAs carrying axonally localizing 3'UTRs of these transcripts showed significantly increased levels in distal axons in the DRGs cocultured with GRPs/NRPs. These results indicate that axon growth promoted by NPCs is associated not only with enhanced transcription of growth-associated genes but also can increase localization of some mRNAs into growing axons. PMID:28197547

  16. Neural Progenitor Cells Promote Axonal Growth and Alter Axonal mRNA Localization in Adult Neurons.

    PubMed

    Merianda, Tanuja T; Jin, Ying; Kalinski, Ashley L; Sahoo, Pabitra K; Fischer, Itzhak; Twiss, Jeffery L

    2017-01-01

    The inhibitory environment of the spinal cord and the intrinsic properties of neurons prevent regeneration of axons following CNS injury. However, both ascending and descending axons of the injured spinal cord have been shown to regenerate into grafts of embryonic neural progenitor cells (NPCs). Previous studies have shown that grafts composed of glial-restricted progenitors (GRPs) and neural-restricted progenitors (NRPs) can provide a permissive microenvironment for axon growth. We have used cocultures of adult rat dorsal root ganglion (DRG) neurons together with NPCs, which have shown significant enhancement of axon growth by embryonic rat GRP and GRPs/NRPs, both in coculture conditions and when DRGs are exposed to conditioned medium from the NPC cultures. This growth-promoting effect of NPC-conditioned medium was also seen in injury-conditioned neurons. DRGs cocultured with GRPs/NRPs showed altered expression of regeneration-associated genes at transcriptional and post-transcriptional levels. We found that levels of GAP-43 mRNA increased in DRG cell bodies and axons. However, hepcidin antimicrobial peptide (HAMP) mRNA decreased in the cell bodies of DRGs cocultured with GRPs/NRPs, which is distinct from the increase in cell body HAMP mRNA levels seen in DRGs after injury conditioning. Endogenous GAP-43 and β-actin mRNAs as well as reporter RNAs carrying axonally localizing 3'UTRs of these transcripts showed significantly increased levels in distal axons in the DRGs cocultured with GRPs/NRPs. These results indicate that axon growth promoted by NPCs is associated not only with enhanced transcription of growth-associated genes but also can increase localization of some mRNAs into growing axons.

  17. Differentiation of neural stem cells in three-dimensional growth factor-immobilized chitosan hydrogel scaffolds.

    PubMed

    Leipzig, Nic D; Wylie, Ryan G; Kim, Howard; Shoichet, Molly S

    2011-01-01

    The adult central nervous system (CNS) contains adult neural stem/progenitor cells (NSPCs) that possess the ability to differentiate into the primary cell types found in the CNS and to regenerate lost or damaged tissue. The ability to specifically and spatially control differentiation is vital to enable cell-based CNS regenerative strategies. Here we describe the development of a protein-biomaterial system that allows rapid, stable and homogenous linking of a growth factor to a photocrosslinkable material. A bioactive recombinant fusion protein incorporating pro-neural rat interferon-γ (rIFN-γ) and the AviTag for biotinylation was successfully expressed in Escherichia coli and purified. The photocrosslinkable biopolymer, methacrylamide chitosan (MAC), was thiolated, allowing conjugation of maleimide-strepatavidin via Michael-type addition. We demonstrated that biotin-rIFN-γ binds specifically to MAC-streptavidin in stoichiometric yields at 100 and 200 ng/mL in photocrosslinked hydrogels. For cell studies, NSPCs were photo-encapsulated in 100 ng/mL biotin-rIFN-γ immobilized MAC based scaffolds and compared to similar NSPC-seeded scaffolds combining 100 ng/mL soluble biotin-rIFN-γ vs. no growth factor. Cells were cultured for 8 days after which differentiation was assayed using immunohistochemistry for lineage specific markers. Quantification showed that immobilized biotin-rIFN-γ promoted neuronal differentiation (72.8 ± 16.0%) similar to soluble biotin-rIFN-γ (71.8 ± 13.2%). The percentage of nestin-positive (stem/progenitor) cells as well as RIP-positive (oligodendrocyte) cells were significantly higher in scaffolds with soluble vs. immobilized biotin-rIFN-γ suggesting that 3-D immobilization results in a more committed lineage specification.

  18. Cell organization, growth, and neural and cardiac development require αII-spectrin

    PubMed Central

    Stankewich, Michael C.; Cianci, Carol D.; Stabach, Paul R.; Ji, Lan; Nath, Anjali; Morrow, Jon S.

    2011-01-01

    Spectrin α2 (αII-spectrin) is a scaffolding protein encoded by the Spna2 gene and constitutively expressed in most tissues. Exon trapping of Spna2 in C57BL/6 mice allowed targeted disruption of αII-spectrin. Heterozygous animals displayed no phenotype by 2 years of age. Homozygous deletion of Spna2 was embryonic lethal at embryonic day 12.5 to 16.5 with retarded intrauterine growth, and craniofacial, neural tube and cardiac anomalies. The loss of αII-spectrin did not alter the levels of αI- or βI-spectrin, or the transcriptional levels of any β-spectrin or any ankyrin, but secondarily reduced by about 80% the steady state protein levels of βII- and βIII-spectrin. Residual βII- and βIII-spectrin and ankyrins B and G were concentrated at the apical membrane of bronchial and renal epithelial cells, without impacting cell morphology. Neuroepithelial cells in the developing brain were more concentrated and more proliferative in the ventricular zone than normal; axon formation was also impaired. Embryonic fibroblasts cultured on fibronectin from E14.5 (Spna2−/−) animals displayed impaired growth and spreading, a spiky morphology, and sparse lamellipodia without cortical actin. These data indicate that the spectrin–ankyrin scaffold is crucial in vertebrates for cell spreading, tissue patterning and organ development, particularly in the developing brain and heart, but is not required for cell viability. PMID:22159418

  19. High efficacy of clonal growth and expansion of adult neural stem cells.

    PubMed

    Wachs, Frank-Peter; Couillard-Despres, Sebastien; Engelhardt, Maren; Wilhelm, Daniel; Ploetz, Sonja; Vroemen, Maurice; Kaesbauer, Johanna; Uyanik, Goekhan; Klucken, Jochen; Karl, Claudia; Tebbing, Johanna; Svendsen, Clive; Weidner, Norbert; Kuhn, Hans-Georg; Winkler, Juergen; Aigner, Ludwig

    2003-07-01

    Neural stem cells (NSCs) from the adult central nervous system are currently being investigated for their potential use in autologous cell replacement strategies. High expansion rates of NSCs in culture are crucial for the generation of a sufficient amount of cells needed for transplantation. Here, we describe efficient growth of adult NSCs in Neurobasal medium containing B27 supplement under clonal and low-density conditions in the absence of serum or conditioned medium. Expansion of up to 15-fold within 1 week was achieved on low-density NSC cultures derived from the lateral ventricle wall, the hippocampal formation, and the spinal cord of adult rats. A 27% single-cell cloning efficiency in Neurobasal/B27 combination further demonstrates its growth-promoting ability. Multipotency and nontumorgenicity of NSCs were retained despite the high rate of culture expansion. In addition, increased cell survival was obtained when Accutase, instead of trypsin, was used for enzymatic dissociation of NSC cultures. This work provides an important step toward the development of standardized protocols for highly efficient in vitro expansion of NSCs from the adult central nervous system to move more closely to the clinical use of NSCs.

  20. Concentration-dependent effect of nerve growth factor on cell fate determination of neural progenitors.

    PubMed

    Zhang, Lei; Jiang, Hui; Hu, Zhengqing

    2011-10-01

    Stem cell-based spiral ganglion neuron (SGN) replacement therapy has been proposed to be a promising strategy to restore hearing either via replacing degenerated neurons or by improving the efficacy of cochlear implants which rely on functional neurons. However, lack of suitable donor cells and low survival rate of implanted cells are the major obstacles to successful implementation of therapeutic transplantation. The present study investigated the potential of mouse inner ear statoacoustic ganglion (SAG)-derived neural progenitors (NPs) to differentiate toward SGN-like glutamatergic cells and the influence to cell survival and differentiation when nerve growth factor (NGF) was supplied. We found that SAG-NPs could form neurospheres, proliferate, and differentiate into cells expressing neuronal protein neurofilament and β-III tubulin. NGF affected the cell fate of SAG-NP in a concentration-dependent manner in vitro. Low concentration of NGF (2-5 ng/mL) promoted cell proliferation. Medium concentration of NGF (20-40 ng/mL) stimulated cells to differentiate into bi-polar SGN-like cells expressing glutamatergic proteins. High concentration of NGF (100 ng/mL) could rescue cells from induced apoptosis. In the in vivo study, NGF (100 ng/mL) dramatically enhanced SAG-NP survival rate after implantation into adult mammalian inner ear. This finding raises the possibility to further induce these NPs to differentiate into SGN-like neurons in future in vivo study. In conclusion, given the capability of proliferation and differentiation into SGN-like cells with the supplement of NGF in vitro, SAG-NPs can serve as donor cells in stem cell-based SGN replacement therapy. NGF improved the survival of SAG-NPs not only in vitro but also in vivo.

  1. Expression of nerve growth factor in rat stomach. Implications for interactions between endothelial, neural and epithelial cells.

    PubMed

    Tarnawski, A S; Ahluwalia, A; Jones, M K; Brzozowski, T

    2016-12-01

    This study was aimed to determine the expression and localization of nerve growth factor (NGF) in the gastric mucosa. Transmural gastric specimens were obtained from euthanized rats. 1) expression of NGF and TrkA receptor by Western blotting; 2) histological evaluation of gastric wall architecture; 3) expression of NGF using immunostaining. Immunostaining showed strong and differential expression of NGF in neural elements of gastric myenteric and submucosal plexuses; in epithelial cells: mainly in chief and progenitor cells, in enterochromaffin-like (ECL) cells; and, in endothelial cells (ECs) lining blood vessels. We concluded that NGF expression in neural elements, epithelial cells and endothelial cells of blood vessels indicated a complex local interaction between neural, epithelial and endothelial cells that regulated gastric mucosal homeostasis and, likely, the protection against gastric injury and ulcer healing.

  2. The Evaluation of Nerve Growth Factor Over Expression on Neural Lineage Specific Genes in Human Mesenchymal Stem Cells

    PubMed Central

    Mortazavi, Yousef; Sheikhsaran, Fatemeh; Khamisipour, Gholamreza Khamisipour; Soleimani, Masoud; Teimuri, Ali; Shokri, Somayeh

    2016-01-01

    Objective Treatment and repair of neurodegenerative diseases such as brain tumors, spinal cord injuries, and functional disorders, including Alzheimer’s disease, are challenging problems. A common treatment approach for such disorders involves the use of mesenchymal stem cells (MSCs) as an alternative cell source to replace injured cells. However, use of these cells in hosts may potentially cause adverse outcomes such as tumorigenesis and uncontrolled differentiation. In attempt to generate mesenchymal derived neural cells, we have infected MSCs with recombinant lentiviruses that expressed nerve growth factor (NGF) and assessed their neural lineage genes. Materials and Methods In this experimental study, we cloned the NGF gene sequence into a helper dependent lentiviral vector that contained the green fluorescent protein (GFP) gene. The recombinant vector was amplified in DH5 bacterial cells. Recombinant viruses were generated in the human embryonic kidney 293 (HEK-293) packaging cell line with the helper vectors and analyzed under fluorescent microscopy. Bone marrow mesenchymal cells were infected by recombinant viruses for three days followed by assessment of neural differentiation. We evaluated expression of NGF through measurement of the NGF protein in culture medium by ELISA; neural specific genes were quantified by real-time polymerase chain reaction (PCR). Results We observed neural morphological changes after three days. Quantitative PCR showed that expressions of NESTIN, glial derived neurotrophic factor (GDNF), glial fibrillary acidic protein (GFAP) and Microtubule-associated protein 2 (MAP2) genes increased following induction of NGF overexpression, whereas expressions of endogenous NGF and brain derived neural growth factor (BDNF) genes reduced. Conclusion Ectopic expression of NGF can induce neurogenesis in MSCs. Direct injection of MSCs may cause tumorigenesis and an undesirable outcome. Therefore an alternative choice to overcome this obstacle may

  3. Gold- and Silver Nanoparticles Affect the Growth Characteristics of Human Embryonic Neural Precursor Cells

    PubMed Central

    Söderstjerna, Erika; Johansson, Fredrik; Klefbohm, Birgitta; Englund Johansson, Ulrica

    2013-01-01

    Rapid development of nanotechnologies and their applications in clinical research have raised concerns about the adverse effects of nanoparticles (NPs) on human health and environment. NPs can be directly taken up by organs exposed, but also translocated to secondary organs, such as the central nervous system (CNS) after systemic- or subcutaneous administration, or via the olfactory system. The CNS is particularly vulnerable during development and recent reports describe transport of NPs across the placenta and even into brain tissue using in vitro and in vivo experimental systems. Here, we investigated whether well-characterized commercial 20 and 80 nm Au- and AgNPs have an effect on human embryonic neural precursor cell (HNPC) growth. After two weeks of NP exposure, uptake of NPs, morphological features and the amount of viable and dead cells, proliferative cells (Ki67 immunostaining) and apoptotic cells (TUNEL assay), respectively, were studied. We demonstrate uptake of both 20 and 80 nm Au- and AgNPs respectively, by HNPCs during proliferation. A significant effect on the sphere size- and morphology was found for all cultures exposed to Au- and AgNPs. AgNPs of both sizes caused a significant increase in numbers of proliferating and apoptotic HNPCs. In contrast, only the highest dose of 20 nm AuNPs significantly affected proliferation, whereas no effect was seen on apoptotic cell death. Our data demonstrates that both Au- and AgNPs interfere with the growth profile of HNPCs, indicating the need of further detailed studies on the adverse effects of NPs on the developing CNS. PMID:23505470

  4. Transforming growth factor beta1 regulates melanocyte proliferation and differentiation in mouse neural crest cells via stem cell factor/KIT signaling.

    PubMed

    Kawakami, Tamihiro; Soma, Yoshinao; Kawa, Yoko; Ito, Masaru; Yamasaki, Emiko; Watabe, Hidenori; Hosaka, Eri; Yajima, Kenji; Ohsumi, Kayoko; Mizoguchi, Masako

    2002-03-01

    Stem cell factor is essential to the migration and differentiation of melanocytes during embryogenesis based on the observation that mutations in either the stem cell factor gene, or its ligand, KIT, result in defects in coat pigmentation in mice. Stem cell factor is also required for the survival of melanocyte precursors while they are migrating towards the skin. Transforming growth factor beta1 has been implicated in the regulation of both cellular proliferation and differentiation. NCC-melb4, an immortal cloned cell line, was cloned from a mouse neural crest cell. NCC-melb4 cells provide a model to study the specific stage of differentiation and proliferation of melanocytes. They also express KIT as a melanoblast marker. Using the NCC-melb4 cell line, we investigated the effect of transforming growth factor beta1 on the differentiation and proliferation of immature melanocyte precursors. Immunohistochemically, NCC-melb4 cells showed transforming growth factor beta1 expression. The anti-transforming growth factor beta1 antibody inhibited the cell growth, and downregulated the KIT protein and mRNA expression. To investigate further the activation of autocrine transforming growth factor beta1, NCC-melb4 cells were incubated in nonexogenous transforming growth factor beta1 culture medium. KIT protein decreased with anti-transforming growth factor beta1 antibody concentration in a concentration-dependent manner. We concluded that in NCC-melb4 cells, transforming growth factor beta1 promotes melanocyte precursor proliferation in autocrine and/or paracrine regulation. We further investigated the influence of transforming growth factor beta1 in vitro using a neural crest cell primary culture system from wild-type mice. Anti-transforming growth factor beta1 antibody decreased the number of KIT positive neural crest cell. In addition, the anti-transforming growth factor beta1 antibody supplied within the wild-type neural crest explants abolished the growth of the neural

  5. Nerve growth factor-induced changes in neural cell adhesion molecule (N-CAM) in PC12 cells.

    PubMed Central

    Prentice, H M; Moore, S E; Dickson, J G; Doherty, P; Walsh, F S

    1987-01-01

    The effects of nerve growth factor (NGF) on the expression of neural cell adhesion molecule (N-CAM) in PC12 cells were determined. A quantitative immunoassay was used to show that NGF induces a 4- to 5-fold increase in relative N-CAM levels over a 3-day period. This increase could not be mimicked by cholera toxin suggesting that it is not a simple consequence of morphological differentiation. The changes in N-CAM levels induced by NGF were accompanied by changes in N-CAM molecular forms. The 140-kd N-CAM species is the major N-CAM expressed by naive PC12 cells, while NGF-treated cultures express N-CAM species of 180 kd and 140 kd. Northern analysis showed that naive cells express a 6.7-kd N-CAM mRNA species only, while NGF-treated cultures express both a 6.7-kb and a 7.2-kb transcript. As the 6.7-kb and 7.2-kb mRNAs are alternative spliced transcripts of a single gene, this result shows that NGF can activate a neuron-specific splicing mechanism. This is the first description of control of N-CAM expression by a growth factor. Images Fig. 3. Fig. 4. Fig. 5. PMID:3308447

  6. Growth and differentiation of adult hippocampal arctic ground squirrel neural stem cells.

    PubMed

    Drew, Kelly L; McGee, Rebecca C; Wells, Matthew S; Kelleher-Andersson, Judith A

    2011-01-07

    Arctic ground squirrels (Urocitellus parryii, AGS) are unique in their ability to hibernate with a core body temperature near or below freezing. These animals also resist ischemic injury to the brain in vivo and oxygen-glucose deprivation in vitro. These unique qualities provided the impetus to isolate AGS neurons to examine inherent neuronal characteristics that could account for the capacity of AGS neurons to resist injury and cell death caused by ischemia and extremely cold temperatures. Identifying proteins or gene targets that allow for the distinctive properties of these cells could aid in the discovery of effective therapies for a number of ischemic indications and for the study of cold tolerance. Adult AGS hippocampus contains neural stem cells that continue to proliferate, allowing for easy expansion of these stem cells in culture. We describe here methods by which researchers can utilize these stem cells and differentiated neurons for any number of purposes. By closely following these steps the AGS neural stem cells can be expanded through two passages or more and then differentiated to a culture high in TUJ1-positive neurons (~50%) without utilizing toxic chemicals to minimize the number of dividing cells. Ischemia induces neurogenesis and neurogenesis which proceeds via MEK/ERK and PI3K/Akt survival signaling pathways contributes to ischemia resistance in vivo and in vitro (Kelleher-Anderson, Drew et al., in preparation). Further characterization of these unique neural cells can advance on many fronts, using some or all of these methods.

  7. In vivo assessment of guided neural stem cell differentiation in growth factor immobilized chitosan-based hydrogel scaffolds.

    PubMed

    Li, Hang; Koenig, Andrew M; Sloan, Patricia; Leipzig, Nic D

    2014-11-01

    In this study, we demonstrate that a unique growth factor-biomaterial system can offer spatial control of growth factors with sustained signaling to guide the specific lineage commitment of neural stem/progenitor cells (NSPCs) in vivo. First, recombinant fusion proteins incorporating an N-terminal biotin tag and interferon-γ (IFN-γ), platelet derived growth factor-AA (PDGF-AA), or bone morphogenic protein-2 (BMP-2) were immobilized to a methacrylamide chitosan (MAC) based biopolymer via a streptavidin linker to specify NSPC differentiation into neurons, oligodendrocytes, or astrocytes, respectively. MAC was mixed with growth factors (immobilized or adsorbed), acrylated laminin, NSPCs, and crosslinked within chitosan conduits. This system mimics regenerative aspects of the central nervous system ECM, which is largely composed of a crosslinked polysaccharide matrix with cell-adhesive regions, and adds the new functionality of protein sequestration. We demonstrated that these growth factors are maintained at functionally significant levels for 28 d in vitro. In the main study, immobilized treatments were compared to absorbed and control treatments after 28 d in vivo (rat subcutaneous). Masson's Trichrome staining revealed that small collagen capsules formed around the chitosan conduits with an average acceptable thickness of 153.07 ± 6.02 μm for all groups. ED-1 staining showed mild macrophage clustering around the outside of chitosan conduits in all treatments with no macrophage invasion into hydrogel portions. Importantly, NSPC differentiation staining demonstrated that immobilized growth factors induced the majority of cells to differentiate into the desired cell types as compared with adsorbed growth factor treatments and controls by day 28. Interestingly, immobilized IFN-γ resulted in neural rosette-like arrangements and even structures resembling neural tubes, suggesting this treatment can lead to guided dedifferentiation and subsequent neurulation.

  8. A novel cell-permeable RDP-p53 fusion protein for specific inhibition on the growth of cancerous neural cells.

    PubMed

    Wu, Jing; Zhang, Enqi; Fu, Ailing

    2016-09-01

    There is 25-35% mutation rate of p53 in cancerous neural cells and this rate reaches 70-76% in glioma cell line. Complement of wild-type p53 has become a potential strategy for protein therapy of cancerous neural cells. Here we investigated the feasibility of a novel RDP-p53 fusion protein for anti-proliferation of cancerous neural cell and the possible mechanism, which would provide an effective approach for targeted delivery of p53 protein to treat cancerous neural cells. The RDP-p53 fusion proteins are expressed in Escherichia coli, and they are labeled with FITC and rhodamine B by chemical modification. The fluorescence-labeled proteins are added to human hepatocellular carcinoma cells (HepG-2) and human neuroblastoma cells (SH-SY5Y) in order to investigate the possibility of RDP enhancing the cell uptake efficiency into neural cells as a cell-permeable carrier. The inhibitory effect of RDP-p53 on SH-SY5Y and human glioma cells (U251) was evaluated by MTT assay. Moreover, the anti-proliferation mechanism of RDP-p53 was determined by Apoptosis and Necrosis Assay Kit and flow cytometric analysis. The results showed that RDP-p53 could enter SH-SY5Y cells with high efficiency and selectively inhibit the growth of cancerous neural cells, including SH-SY5Y and U251. Also, cell apoptosis pathway and cell-cycle arrest at the G2/M phase were associated with the inhibition mechanism of RDP-p53 according to the data of flow cytometric analysis. RDP-p53 could be a novel antitumor candidate for targeting treatment of cancerous neural cells.

  9. Proliferation and differentiation of neural stem cells irradiated with X-rays in logarithmic growth phase.

    PubMed

    Isono, Mayu; Otsu, Masahiro; Konishi, Teruaki; Matsubara, Kana; Tanabe, Toshiaki; Nakayama, Takashi; Inoue, Nobuo

    2012-07-01

    Exposure of the fetal brain to ionizing radiation causes congenital brain abnormalities. Normal brain formation requires regionally and temporally appropriate proliferation and differentiation of neural stem cells (NSCs) into neurons and glia. Here, we investigated the effects of X-irradiation on proliferating homogenous NSCs prepared from mouse ES cells. Cells irradiated with X-rays at a dose of 1Gy maintained the capabilities for proliferation and differentiation but stopped proliferation temporarily. In contrast, the cells ceased proliferation following irradiation at a dose of >5Gy. These results suggest that irradiation of the fetal brain at relatively low doses may cause congenital brain abnormalities as with relatively high doses.

  10. Response of the sensorimotor cortex of cerebral palsy rats receiving transplantation of vascular endothelial growth factor 165-transfected neural stem cells.

    PubMed

    Tan, Jielu; Zheng, Xiangrong; Zhang, Shanshan; Yang, Yujia; Wang, Xia; Yu, Xiaohe; Zhong, Le

    2014-10-01

    Neural stem cells are characterized by the ability to differentiate and stably express exogenous ge-nes. Vascular endothelial growth factor plays a role in protecting local blood vessels and neurons of newborn rats with hypoxic-ischemic encephalopathy. Transplantation of vascular endothelial growth factor-transfected neural stem cells may be neuroprotective in rats with cerebral palsy. In this study, 7-day-old Sprague-Dawley rats were divided into five groups: (1) sham operation (control), (2) cerebral palsy model alone or with (3) phosphate-buffered saline, (4) vascular endothelial growth factor 165 + neural stem cells, or (5) neural stem cells alone. The cerebral palsy model was established by ligating the left common carotid artery followed by exposure to hypoxia. Phosphate-buffered saline, vascular endothelial growth factor + neural stem cells, and neural stem cells alone were administered into the sensorimotor cortex using the stereotaxic instrument and microsyringe. After transplantation, the radial-arm water maze test and holding test were performed. Immunohistochemistry for vascular endothelial growth factor and histology using hematoxylin-eosin were performed on cerebral cortex. Results revealed that the number of vascular endothelial growth factor-positive cells in cerebral palsy rats transplanted with vascular endothelial growth factor-transfected neural stem cells was increased, the time for finding water and the finding repetitions were reduced, the holding time was prolonged, and the degree of cell degeneration or necrosis was reduced. These findings indicate that the transplantation of vascular endothelial growth factor-transfected neural stem cells alleviates brain damage and cognitive deficits, and is neuroprotective in neonatal rats with hypoxia ischemic-mediated cerebral palsy.

  11. Noncanonical transforming growth factor β (TGFβ) signaling in cranial neural crest cells causes tongue muscle developmental defects.

    PubMed

    Iwata, Jun-ichi; Suzuki, Akiko; Pelikan, Richard C; Ho, Thach-Vu; Chai, Yang

    2013-10-11

    Microglossia is a congenital birth defect in humans and adversely impacts quality of life. In vertebrates, tongue muscle derives from the cranial mesoderm, whereas tendons and connective tissues in the craniofacial region originate from cranial neural crest (CNC) cells. Loss of transforming growth factor β (TGFβ) type II receptor in CNC cells in mice (Tgfbr2(fl/fl);Wnt1-Cre) causes microglossia due to a failure of cell-cell communication between cranial mesoderm and CNC cells during tongue development. However, it is still unclear how TGFβ signaling in CNC cells regulates the fate of mesoderm-derived myoblasts during tongue development. Here we show that activation of the cytoplasmic and nuclear tyrosine kinase 1 (ABL1) cascade in Tgfbr2(fl/fl);Wnt1-Cre mice results in a failure of CNC-derived cell differentiation followed by a disruption of TGFβ-mediated induction of growth factors and reduction of myogenic cell proliferation and differentiation activities. Among the affected growth factors, the addition of fibroblast growth factor 4 (FGF4) and neutralizing antibody for follistatin (FST; an antagonist of bone morphogenetic protein (BMP)) could most efficiently restore cell proliferation, differentiation, and organization of muscle cells in the tongue of Tgfbr2(fl/fl);Wnt1-Cre mice. Thus, our data indicate that CNC-derived fibroblasts regulate the fate of mesoderm-derived myoblasts through TGFβ-mediated regulation of FGF and BMP signaling during tongue development.

  12. Noncanonical Transforming Growth Factor β (TGFβ) Signaling in Cranial Neural Crest Cells Causes Tongue Muscle Developmental Defects*♦

    PubMed Central

    Iwata, Jun-ichi; Suzuki, Akiko; Pelikan, Richard C.; Ho, Thach-Vu; Chai, Yang

    2013-01-01

    Microglossia is a congenital birth defect in humans and adversely impacts quality of life. In vertebrates, tongue muscle derives from the cranial mesoderm, whereas tendons and connective tissues in the craniofacial region originate from cranial neural crest (CNC) cells. Loss of transforming growth factor β (TGFβ) type II receptor in CNC cells in mice (Tgfbr2fl/fl;Wnt1-Cre) causes microglossia due to a failure of cell-cell communication between cranial mesoderm and CNC cells during tongue development. However, it is still unclear how TGFβ signaling in CNC cells regulates the fate of mesoderm-derived myoblasts during tongue development. Here we show that activation of the cytoplasmic and nuclear tyrosine kinase 1 (ABL1) cascade in Tgfbr2fl/fl;Wnt1-Cre mice results in a failure of CNC-derived cell differentiation followed by a disruption of TGFβ-mediated induction of growth factors and reduction of myogenic cell proliferation and differentiation activities. Among the affected growth factors, the addition of fibroblast growth factor 4 (FGF4) and neutralizing antibody for follistatin (FST; an antagonist of bone morphogenetic protein (BMP)) could most efficiently restore cell proliferation, differentiation, and organization of muscle cells in the tongue of Tgfbr2fl/fl;Wnt1-Cre mice. Thus, our data indicate that CNC-derived fibroblasts regulate the fate of mesoderm-derived myoblasts through TGFβ-mediated regulation of FGF and BMP signaling during tongue development. PMID:23950180

  13. Effects of ethanol on transforming growth factor Β1-dependent and -independent mechanisms of neural stem cell apoptosis.

    PubMed

    Hicks, Steven D; Miller, Michael W

    2011-06-01

    Stem cell vitality is critical for the growth of the developing brain. Growth factors can define the survival of neural stem cells (NSCs) and ethanol can affect growth factor-mediated activities. The present study tested two hypotheses: (a) ethanol causes the apoptotic death of NSCs and (b) this effect is influenced by the ambient growth factor. Monolayer cultures of non-immortalized NS-5 cells were exposed to fibroblast growth factor (FGF) 2 or transforming growth factor (TGF) β1 in the absence or presence of ethanol for 48 h. Ethanol killed NSCs as measured by increases in the numbers of ethidium bromide+ and annexin V+ cells and decreases in the number of calcein AM+ (viable) cells. These toxic effects were promoted by TGFβ1. A quantitative polymerase chain reaction array of apoptosis-related mRNAs revealed an ethanol-induced increase (≥2-fold change; p<0.05) in transcripts involved in Fas ligand (FasL) and tumor necrosis factor (TNF) signaling. These effects, particularly the FasL pathway, were potentiated by TGFβ1. Immunocytochemical analyses of NS-5 cells showed that transcriptional alterations translated into consistent up-regulation of protein expression. Experiments with the neocortical proliferative zones harvested from fetal mice exposed to ethanol showed that ethanol activated similar molecular systems in vivo. Thus, ethanol induces NSC death through two distinct molecular mechanisms, one is initiated by TGFβ1 (FasL) and another (through TNF) which is TGFβ1-independent.

  14. Toward Intelligent Synthetic Neural Circuits: Directing and Accelerating Neuron Cell Growth by Self-Rolled-Up Silicon Nitride Microtube Array

    PubMed Central

    2015-01-01

    In neural interface platforms, cultures are often carried out on a flat, open, rigid, and opaque substrate, posing challenges to reflecting the native microenvironment of the brain and precise engagement with neurons. Here we present a neuron cell culturing platform that consists of arrays of ordered microtubes (2.7–4.4 μm in diameter), formed by strain-induced self-rolled-up nanomembrane (s-RUM) technology using ultrathin (<40 nm) silicon nitride (SiNx) film on transparent substrates. These microtubes demonstrated robust physical confinement and unprecedented guidance effect toward outgrowth of primary cortical neurons, with a coaxially confined configuration resembling that of myelin sheaths. The dynamic neural growth inside the microtube, evaluated with continuous live-cell imaging, showed a marked increase (20×) of the growth rate inside the microtube compared to regions outside the microtubes. We attribute the dramatic accelerating effect and precise guiding of the microtube array to three-dimensional (3D) adhesion and electrostatic interaction with the SiNx microtubes, respectively. This work has clear implications toward building intelligent synthetic neural circuits by arranging the size, site, and patterns of the microtube array, for potential treatment of neurological disorders. PMID:25329686

  15. Toward intelligent synthetic neural circuits: directing and accelerating neuron cell growth by self-rolled-up silicon nitride microtube array.

    PubMed

    Froeter, Paul; Huang, Yu; Cangellaris, Olivia V; Huang, Wen; Dent, Erik W; Gillette, Martha U; Williams, Justin C; Li, Xiuling

    2014-11-25

    In neural interface platforms, cultures are often carried out on a flat, open, rigid, and opaque substrate, posing challenges to reflecting the native microenvironment of the brain and precise engagement with neurons. Here we present a neuron cell culturing platform that consists of arrays of ordered microtubes (2.7-4.4 μm in diameter), formed by strain-induced self-rolled-up nanomembrane (s-RUM) technology using ultrathin (<40 nm) silicon nitride (SiNx) film on transparent substrates. These microtubes demonstrated robust physical confinement and unprecedented guidance effect toward outgrowth of primary cortical neurons, with a coaxially confined configuration resembling that of myelin sheaths. The dynamic neural growth inside the microtube, evaluated with continuous live-cell imaging, showed a marked increase (20×) of the growth rate inside the microtube compared to regions outside the microtubes. We attribute the dramatic accelerating effect and precise guiding of the microtube array to three-dimensional (3D) adhesion and electrostatic interaction with the SiNx microtubes, respectively. This work has clear implications toward building intelligent synthetic neural circuits by arranging the size, site, and patterns of the microtube array, for potential treatment of neurological disorders.

  16. Type 1 Fibroblast Growth Factor Receptor in Cranial Neural Crest Cell-derived Mesenchyme Is Required for Palatogenesis*

    PubMed Central

    Wang, Cong; Chang, Julia Yu Fong; Yang, Chaofeng; Huang, Yanqing; Liu, Junchen; You, Pan; McKeehan, Wallace L.; Wang, Fen; Li, Xiaokun

    2013-01-01

    Cleft palate is a common congenital birth defect. The fibroblast growth factor (FGF) family has been shown to be important for palatogenesis, which elicits the regulatory functions by activating the FGF receptor tyrosine kinase. Mutations in Fgf or Fgfr are associated with cleft palate. To date, most mechanistic studies on FGF signaling in palate development have focused on FGFR2 in the epithelium. Although Fgfr1 is expressed in the cranial neural crest (CNC)-derived palate mesenchyme and Fgfr1 mutations are associated with palate defects, how FGFR1 in palate mesenchyme regulates palatogenesis is not well understood. Here, we reported that by using Wnt1Cre to delete Fgfr1 in neural crest cells led to cleft palate, cleft lip, and other severe craniofacial defects. Detailed analyses revealed that loss-of-function mutations in Fgfr1 did not abrogate patterning of CNC cells in palate shelves. However, it upset cell signaling in the frontofacial areas, delayed cell proliferation in both epithelial and mesenchymal compartments, prevented palate shelf elevation, and compromised palate shelf fusion. This is the first report revealing how FGF signaling in CNC cells regulates palatogenesis. PMID:23754280

  17. Co-localization of neural cell adhesion molecule and fibroblast growth factor receptor 2 in early embryo development.

    PubMed

    Vesterlund, Liselotte; Töhönen, Virpi; Hovatta, Outi; Kere, Juha

    2011-01-01

    During development there is a multitude of signaling events governing the assembly of the developing organism. Receptors for signaling molecules such as fibroblast growth factor receptor 2 (FGFR2) enable the embryo to communicate with the surrounding environment and activate downstream pathways. The neural cell adhesion molecule (NCAM) was first characterized as a cell adhesion molecule highly expressed in the nervous system, but recent studies have shown that it is also a signaling receptor. Using a novel single oocyte adaptation of the proximity ligation assay, we here show a close association between NCAM and FGFR2 in mouse oocytes and 2-cell embryos. Real-time PCR analyses revealed the presence of messenger RNA encoding key proteins in downstream signaling pathways in oocytes and early mouse embryos. In summary these findings show a co-localization of NCAM and FGFR2 in early vertebrate development with intracellular signaling pathways present to enable a cellular response.

  18. A Hydrogel Bridge Incorporating Immobilized Growth Factors and Neural Stem/Progenitor Cells to Treat Spinal Cord Injury.

    PubMed

    Li, Hang; Ham, Trevor R; Neill, Nicholas; Farrag, Mahmoud; Mohrman, Ashley E; Koenig, Andrew M; Leipzig, Nic D

    2016-04-06

    Spinal cord injury (SCI) causes permanent, often complete disruption of central nervous system (CNS) function below the damaged region, leaving patients without the ability to regenerate lost tissue. To engineer new CNS tissue, a unique spinal cord bridge is created to deliver stem cells and guide their organization and development with site-specifically immobilized growth factors. In this study, this bridge is tested, consisting of adult neural stem/progenitor cells contained within a methacrylamide chitosan (MAC) hydrogel and protected by a chitosan conduit. Interferon-γ (IFN-γ) and platelet-derived growth factor-AA (PDGF-AA) are recombinantly produced and tagged with an N-terminal biotin. They are immobilized to streptavidin-functionalized MAC to induce either neuronal or oligodendrocytic lineages, respectively. These bridges are tested in a rat hemisection model of SCI between T8 and T9. After eight weeks treatments including chitosan conduits result in a significant reduction in lesion area and macrophage infiltration around the lesion site (p < 0.0001). Importantly, neither immobilized IFN-γ nor PDGF-AA increased macrophage infiltration. Retrograde tracing demonstrates improved neuronal regeneration through the use of immobilized growth factors. Immunohistochemistry staining demonstrates that immobilized growth factors are effective in differentiating encapsulated cells into their anticipated lineages within the hydrogel, while qualitatively reducing glial fibrillary acid protein expression.

  19. Effect of neurosphere size on the growth rate of human neural stem/progenitor cells.

    PubMed

    Mori, Hideki; Ninomiya, Kazuaki; Kino-oka, Masahiro; Shofuda, Tomoko; Islam, Mohammed Omedul; Yamasaki, Mami; Okano, Hideyuki; Taya, Masahito; Kanemura, Yonehiro

    2006-12-01

    Neural stem/progenitor cells (NSPCs) proliferate as aggregates in vitro, but the mechanism of aggregation is not fully understood. Here, we report that aggregation promotes the proliferation of NSPCs. We found that the proliferation rate was linear and depended on the size of the aggregate; that is, the population doubling time of the NSPCs gradually decreased as the diameter approached 250 micro m and flattened to a nearly constant value beyond this diameter. Given this finding, and with the intent of enhancing the efficiency of human NSPC expansion, we induced the NSPCs to form aggregates close to 250 micro m in diameter quickly by culturing them in plates with U-bottomed wells. The NSPCs formed aggregates effectively in the U-bottomed wells, with cell numbers approximately 1.5 times greater than those in the aggregates that formed spontaneously in flat-bottomed wells. In addition, this effect of aggregation involved cell-cell signaling molecules of the Notch1 pathway. In the U-bottomed wells, Hes1 and Hes5, which are target genes of the Notch signal, were expressed at higher levels than in the control, flat-bottomed wells. The amount of cleaved Notch1 was also higher in the cells cultured in the U-bottomed wells. The addition of gamma-secretase inhibitor, which blocks Notch signaling, suppressed cell proliferation in the U-bottomed wells. These results suggest that the three-dimensional architecture of NSPC aggregates would create a microenvironment that promotes the proliferation of human NSPCs.

  20. Growth-associated protein 43 in differentiating peripheral nerve sheath tumors from other non-neural spindle cell neoplasms.

    PubMed

    Chen, Wei-Shen; Chen, Pei-Ling; Lu, Dongsi; Lind, Anne C; Dehner, Louis P

    2014-02-01

    The malignant peripheral nerve sheath tumor is a relatively uncommon type of soft tissue sarcoma arising from a peripheral nerve or extraneural soft tissues and showing nerve sheath differentiation. The diagnosis of malignant peripheral nerve sheath tumor is one of the most challenging tasks in surgical pathology because of its uncommon type (5-10% soft tissue sarcomas), morphologic resemblance to other spindle cell neoplasms and lack of sensitive and specific immunohistochemical markers. The pathologic diagnosis is more straightforward in the clinical setting of neurofibromatosis-1, but problems are mainly centered on the non-neurofibromatosis-1 malignant peripheral nerve sheath tumors. To date, S100 protein is the most widely applied marker in the case of a suspected malignant peripheral nerve sheath tumor, yet its suboptimal sensitivity and its expression in other spindle cell neoplasms, including spindle cell melanoma, clear-cell sarcoma, leiomyosarcoma and monophasic synovial sarcoma, add to the diagnostic conundrum. Growth-associated protein 43 (GAP43), a membrane-associated phosphoprotein expressed in neuronal growth cones and Schwann cell precursors during neural development and axonal regeneration, was applied to a set of nerve sheath and non-nerve sheath spindle cell neoplasms. The findings in this study indicate that GAP43 is expressed in malignant peripheral nerve sheath tumors (n=18/21; 86%) and demonstrates a sensitivity superior to S100 protein (n=13/21; 62%). GAP43 is also positive in neurofibromas (n=17/18; 94%), schwannomas (n=11/12; 92%) and desmoplastic melanomas (n=7/10; 70%). In contrast, it is negative in the non-desmoplastic spindle cell melanomas (n=20/22; 91%). Of the other non-neural soft tissue sarcomas, GAP43 is non-reactive in most leiomyosarcomas (n=14/16; 88%) and clear-cell sarcomas (n=8/8), and only focally positive in monophasic synovial sarcomas (n=3/7; 43%). GAP43 is seemingly a highly sensitive marker for peripheral nerve

  1. Glutamate promotes neural stem cell proliferation by increasing the expression of vascular endothelial growth factor of astrocytes in vitro.

    PubMed

    Liu, C X; Xu, X; Chen, X L; Yang, P B; Zhang, J S; Liu, Y

    2015-09-20

    The high levels of glutamate might involve in neurogenesis after brain injuries. However, the mechanisms are not fully understood. In this study, we investigated the effect of glutamate on the proliferation of rat embryonic neural stem/progenitor cells (NSCs) through regulating the vascular endothelial growth factor (VEGF) expression of astrocytes (ASTs) in vitro, and the cyclin D1 expression of NSCs. The results showed that glutamate promoted the expression and secretion of VEGF of rat astrocytes by activating group I mGluRs. Astrocyte conditioned medium-containing Glu [ACM (30%)] promoted the proliferation of embryonic NSCs compared with normal astrocyte conditioned medium+Glu [N-ACM (30%)+Glu (30 μM)] by increasing cell activity, diameter of neurospheres, bromodeoxyuridine (BrdU) incorporation and cell division; while ACM+VEGF neutralizing antibody [ACM (30%)+VEGF NAb (15 μg/ml)] significantly inhibited the proliferation of embryonic NSCs compared with ACM (30%). ACM (30%) increased the expressions of cyclin D1 and decreased cell death compared with N-ACM (30%)+Glu (30 μM). ACM (30%)+VEGF NAb (15 μg/ml) decreased the expressions of cyclin D1 and increased cell death compared with ACM (30%). These results demonstrated that glutamate could also indirectly promote the proliferation of rat embryonic NSCs through inducing the VEGF expression of ASTs in vitro, and VEGF may increase the expression of cyclin D1. These finding suggest that glutamate may be a major molecule for regulating embryonic NSC proliferation and facilitate neural repair in the process of NSC transplants after brain injuries.

  2. Dynamic change of neural cell adhesion molecule polysialylation on human neuroblastoma (IMR-32) and rat pheochromocytoma (PC-12) cells during growth and differentiation.

    PubMed

    Poongodi, Geetha L; Suresh, Nimmagadda; Gopinath, Subash C B; Chang, Tschining; Inoue, Sadako; Inoue, Yasuo

    2002-08-02

    Polysialic acid (PSA) is a regulatory epitope of neural cell adhesion molecule (NCAM) in homophilic adhesion of neural cells mediated by NCAM, is also known to be re-expressed in several human tumors, thus serves as an oncodevelopmental antigen. In this study, using a recently developed ultrasensitive chemical method in addition to immunochemical methods, growth stage-dependent and retinoic acid (RA)-induced differentiation-dependent changes of PSA expression in human neuroblastoma (IMR-32) and rat pheochromocytoma (PC-12) cells were analyzed both qualitatively and quantitatively. Both IMR-32 and PC-12 cells expressed PSA on NCAM, and the level of PSA expressed per unit weight of cells increased with post-inoculation incubation time. The most prominent feature was seen at the full confluence stage. RA induced neuronal differentiation in both IMR-32 and CP-12 cells that paralleled the change in the PSA level. Chemical analysis revealed the presence of NCAM glycoforms differing in the degree of polymerization (DP) of oligo/polysialyl chains, whose DP was smaller than 40. DP distribution of PSA was different between the cell lines and was changed by the growth stage and the RA treatment. Thus DP analysis of PSA is important in understanding both mechanism and biological significance of its regulated expression.

  3. Liver growth factor promotes the survival of grafted neural stem cells in a rat model of Parkinson's disease.

    PubMed

    Reimers, Diana; Osuna, Cristina; Gonzalo-Gobernado, Rafael; Herranz, Antonio S; Diaz-Gil, Juan Jose; Jimenez-Escrig, Adriano; Asensio, Maria Jose; Miranda, Cristina; Rodriguez-Serrano, Macarena; Bazan, Eulalia

    2012-01-01

    Neural stem cells (NSCs) with self-renewal and multilineage potential are considered good candidates for cell replacement of damaged nerve tissue. Several studies have focused on the ability of the neurotrophic factors coadministration to improve the efficiency of grafted NSCs. Liver growth factor (LGF) is an hepatic mitogen that promotes regeneration of damaged tissues, including brain tissue. It has neurogenic activity and has partially restored the nigrostriatal dopaminergic system in an experimental model of Parkinson's disease. Present results demonstrate that in the dopamine- depleted striatum of 6-hydroxydopamine-lesioned rats, grafted NSCs retained their ability to differentiate into neurons, astrocytes, and oligodendrocytes. NSCs also differentiated into microglia/macrophages and endothelial cells. Thus, 23 ± 5.6% of them were inmunoreactive for isolectin IB4, and a small population integrated into blood vessels, showing an endothelial-like morphology. Intrastriatal infusion of LGF promoted the viability of the implants, and favored their differentiation to an endothelial-like phenotype. Moreover, LGF infusion raised the expression of the anti-apoptotic protein Bcl-2 by 3.9 ± 0.9 fold without affecting the levels of the pro-apoptotic protein Bax. Since LGF-treated rats also showed a significant reduction in apomorphine-induced rotational behavior, our results suggest that administration of this factor might be a convenient treatment for Parkinson's disease cell replacement therapies based on NSCs transplantation.

  4. Human Cortical Neural Stem Cells Expressing Insulin-Like Growth Factor-I: A Novel Cellular Therapy for Alzheimer's Disease.

    PubMed

    McGinley, Lisa M; Sims, Erika; Lunn, J Simon; Kashlan, Osama N; Chen, Kevin S; Bruno, Elizabeth S; Pacut, Crystal M; Hazel, Tom; Johe, Karl; Sakowski, Stacey A; Feldman, Eva L

    2016-03-01

    Alzheimer's disease (AD) is the most prevalent age-related neurodegenerative disorder and a leading cause of dementia. Current treatment fails to modify underlying disease pathologies and very little progress has been made to develop effective drug treatments. Cellular therapies impact disease by multiple mechanisms, providing increased efficacy compared with traditional single-target approaches. In amyotrophic lateral sclerosis, we have shown that transplanted spinal neural stem cells (NSCs) integrate into the spinal cord, form synapses with the host, improve inflammation, and reduce disease-associated pathologies. Our current goal is to develop a similar "best in class" cellular therapy for AD. Here, we characterize a novel human cortex-derived NSC line modified to express insulin-like growth factor-I (IGF-I), HK532-IGF-I. Because IGF-I promotes neurogenesis and synaptogenesis in vivo, this enhanced NSC line offers additional environmental enrichment, enhanced neuroprotection, and a multifaceted approach to treating complex AD pathologies. We show that autocrine IGF-I production does not impact the cell secretome or normal cellular functions, including proliferation, migration, or maintenance of progenitor status. However, HK532-IGF-I cells preferentially differentiate into gamma-aminobutyric acid-ergic neurons, a subtype dysregulated in AD; produce increased vascular endothelial growth factor levels; and display an increased neuroprotective capacity in vitro. We also demonstrate that HK532-IGF-I cells survive peri-hippocampal transplantation in a murine AD model and exhibit long-term persistence in targeted brain areas. In conclusion, we believe that harnessing the benefits of cellular and IGF-I therapies together will provide the optimal therapeutic benefit to patients, and our findings support further preclinical development of HK532-IGF-I cells into a disease-modifying intervention for AD. ©AlphaMed Press.

  5. A re-assessment of long distance growth and connectivity of neural stem cells after severe spinal cord injury.

    PubMed

    Sharp, Kelli G; Yee, Kelly Matsudaira; Steward, Oswald

    2014-07-01

    As part of the NIH "Facilities of Research Excellence-Spinal Cord Injury" project to support independent replication, we repeated key parts of a study reporting robust engraftment of neural stem cells (NSCs) treated with growth factors after complete spinal cord transection in rats. Rats (n=20) received complete transections at thoracic level 3 (T3) and 2weeks later received NSC transplants in a fibrin matrix with a growth factor cocktail using 2 different transplantation methods (with and without removal of scar tissue). Control rats (n=9) received transections only. Hindlimb locomotor function was assessed with the BBB scale. Nine weeks post injury, reticulospinal tract axons were traced in 6 rats by injecting BDA into the reticular formation. Transplants grew to fill the lesion cavity in most rats although grafts made with scar tissue removal had large central cavities. Grafts blended extensively with host tissue obliterating the astroglial boundary at the cut ends, but in most cases there was a well-defined partition within the graft that separated rostral and caudal parts of the graft. In some cases, the partition contained non-neuronal scar tissue. There was extensive outgrowth of GFP labeled axons from the graft, but there was minimal ingrowth of host axons into the graft revealed by tract tracing and immunocytochemistry for 5HT. There were no statistically significant differences between transplant and control groups in the degree of locomotor recovery. Our results confirm the previous report that NSC transplants can fill lesion cavities and robustly extend axons, but reveal that most grafts do not create a continuous bridge of neural tissue between rostral and caudal segments.

  6. A re-assessment of long distance growth and connectivity of neural stem cells after severe spinal cord injury

    PubMed Central

    Sharp, Kelli G.; Yee, Kelly Matsudaira; Steward, Oswald

    2014-01-01

    As part of the NIH “Facilities of Research Excellence—Spinal Cord Injury” project to support independent replication, we repeated key parts of a study reporting robust engraftment of neural stem cells (NSCs) treated with growth factors after complete spinal cord transection in rats. Rats (n = 20) received complete transections at thoracic level 3 (T3) and 2 weeks later received NSC transplants in a fibrin matrix with a growth factor cocktail using 2 different transplantation methods (with and without removal of scar tissue). Control rats (n = 9) received transections only. Hindlimb locomotor function was assessed with the BBB scale. Nine weeks post injury, reticulospinal tract axons were traced in 6 rats by injecting BDA into the reticular formation. Transplants grew to fill the lesion cavity in most rats although grafts made with scar tissue removal had large central cavities. Grafts blended extensively with host tissue obliterating the astroglial boundary at the cut ends, but in most cases there was a well-defined partition within the graft that separated rostral and caudal parts of the graft. In some cases, the partition contained non-neuronal scar tissue. There was extensive outgrowth of GFP labeled axons from the graft, but there was minimal ingrowth of host axons into the graft revealed by tract tracing and immunocy-tochemistry for 5HT. There were no statistically significant differences between transplant and control groups in the degree of locomotor recovery. Our results confirm the previous report that NSC transplants can fill lesion cavities and robustly extend axons, but reveal that most grafts do not create a continuous bridge of neural tissue between rostral and caudal segments. PMID:24747827

  7. Neural stem cells producing an inducible and soluble form of Gas1 target and inhibit intracranial glioma growth.

    PubMed

    López-Ornelas, Adolfo; Vergara, Paula; Segovia, José

    2014-07-01

    Glioblastoma multiforme (GBM) is the most common and lethal primary brain tumor and current treatments have not improved its prognosis. Therefore, new strategies and therapeutic agents should be investigated. Growth arrest specific-1 (Gas1) is a protein that induces cell arrest and apoptosis of gliomas and a soluble form, tGas1, increases these effects acting in both autocrine and paracrine manners. Moreover, neural stem cells (NSCs) can be used as a vehicle to transport therapeutic molecules because they have innate tropism towards tumors. Lentiviral vectors were used to obtain NSCs capable of expressing tGas1 in a regulated manner. The ability of engineered NSCs to track and reach GBM in vivo, produce tGas1, and their efficacy decreasing tumor growth and increasing the overall health and survival time of nude mice implanted with GBM were assessed. The overexpression of tGas1 from NSCs decreased viability and induced cell arrest and apoptosis of GBM cells and also, albeit in a reduced manner, of NSCs themselves. NSCs migrate from one cerebral hemisphere to the contralateral, reach GBM, express the tGas1 transgene when induced by tetracycline and produce the protein. Tumor volume decreased by 77% compared with controls, and tGas1 improved the overall health and increased the survival time of mice implanted with GBM by 75%. We demonstrated that tGas1 has an antineoplastic effect, and the results support the potential of tGas1 as an adjuvant for the treatment of gliomas. Copyright © 2014 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  8. Neural stem cells.

    PubMed

    Kennea, Nigel L; Mehmet, Huseyin

    2002-07-01

    Neural stem cells (NSCs) have the ability to self-renew, and are capable of differentiating into neurones, astrocytes and oligodendrocytes. Such cells have been isolated from the developing brain and more recently from the adult central nervous system. This review aims to provide an overview of the current research in this evolving area. There is now increasing knowledge of the factors controlling the division and differentiation of NSCs during normal brain development. In addition, the cues for differentiation in vitro, and the possibility of transdifferentiation are reviewed. The discovery of these cells in the adult brain has encouraged research into their role during neurogenesis in the normal mature brain and after injury. Lastly other sources of neural precursors are discussed, and the potential for stem cells to be used in cell replacement therapy for brain injury or degenerative brain diseases with a particular emphasis on cerebral ischaemia and Parkinson's disease. Copyright 2002 John Wiley & Sons, Ltd.

  9. Polypyrrole-coated electrospun poly(lactic acid) fibrous scaffold: effects of coating on electrical conductivity and neural cell growth.

    PubMed

    Sudwilai, Thitima; Ng, Jun Jye; Boonkrai, Chatikorn; Israsena, Nipan; Chuangchote, Surawut; Supaphol, Pitt

    2014-01-01

    Neuronal activities play critical roles in both neurogenesis and neural regeneration. In that sense, electrically conductive and biocompatible biomaterial scaffolds can be applied in various applications of neural tissue engineering. In this study, we fabricated a novel biomaterial for neural tissue engineering applications by coating electrospun poly(lactic acid) (PLA) nanofibers with a conducting polymer, polypyrole (PPy), via admicellar polymerization. Optimal conditions for polymerization and preparation of PPy-coated electrospun PLA nanofibers were obtained by comparing results from scanning electron microscopy, X-ray photoelectron spectrometer, and surface conductivity tests. In vitro cell culture experiments showed that PPy-coated electrospun PLA fibrous scaffold is not toxic. The scaffold could support attachment and migration of neural progenitor cells. Neurons derived from progenitor exhibited long neurite outgrowth under electrical stimulation. Our study concluded that PPy-coated electrospun PLA fibers had a good biocompatibility with neural progenitor cells and may serve as a promising material for controlling progenitor cell behaviors and enhancing neural repair.

  10. Carbon-ion beams effectively induce growth inhibition and apoptosis in human neural stem cells compared with glioblastoma A172 cells.

    PubMed

    Isono, Mayu; Yoshida, Yukari; Takahashi, Akihisa; Oike, Takahiro; Shibata, Atsushi; Kubota, Yoshiki; Kanai, Tatsuaki; Ohno, Tatsuya; Nakano, Takashi

    2015-09-01

    Carbon-ion radiotherapy (CIRT) holds promise in the treatment of glioblastoma, an aggressive X-ray-resistant brain tumor. However, since glioblastoma cells show a highly invasive nature, carbon-ion (C-ion) irradiation of normal tissues surrounding the tumor is inevitable. Recent studies have revealed the existence of neural stem cells in the adult brain. Therefore, the damaging effect of C-ion beams on the neural stem cells has to be carefully considered in the treatment planning of CIRT. Here, we investigated the growth and death mode of human neural stem cells (hNSCs) and glioblastoma A172 cells after X-ray or C-ion beam irradiation. The X-ray dose resulting in a 50% growth rate (D(50)) was 0.8 Gy in hNSCs and 3.0 Gy in A172 cells, while the D(50) for C-ion beams was 0.4 Gy in hNSCs and 1.6 Gy in A172 cells; the relative biological effectiveness value of C-ion beams was 2.0 in hNSCs and 1.9 in A172 cells. Importantly, both X-rays and C-ion beams preferentially induced apoptosis, not necrosis, in hNSCs; however, radiation-induced apoptosis was less evident in A172 cells. The apoptosis-susceptible nature of the irradiated hNSCs was associated with prolonged upregulation of phosphorylated p53, whereas the apoptosis-resistant nature of A172 cells was associated with a high basal level of nuclear factor kappa B expression. Taken together, these data indicate that apoptosis is the major cell death pathway in hNSCs after irradiation. The high sensitivity of hNSCs to C-ion beams underscores the importance of careful target volume delineation in the treatment planning of CIRT for glioblastoma. © The Author 2015. Published by Oxford University Press on behalf of The Japan Radiation Research Society and Japanese Society for Radiation Oncology.

  11. Neural progenitor cell implants modulate vascular endothelial growth factor and brain-derived neurotrophic factor expression in rat axotomized neurons.

    PubMed

    Talaverón, Rocío; Matarredona, Esperanza R; de la Cruz, Rosa R; Pastor, Angel M

    2013-01-01

    Axotomy of central neurons leads to functional and structural alterations which largely revert when neural progenitor cells (NPCs) are implanted in the lesion site. The new microenvironment created by NPCs in the host tissue might modulate in the damaged neurons the expression of a high variety of molecules with relevant roles in the repair mechanisms, including neurotrophic factors. In the present work, we aimed to analyze changes in neurotrophic factor expression in axotomized neurons induced by NPC implants. For this purpose, we performed immunofluorescence followed by confocal microscopy analysis for the detection of vascular endothelial growth factor (VEGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and nerve growth factor (NGF) on brainstem sections from rats with axotomy of abducens internuclear neurons that received NPC implants (implanted group) or vehicle injections (axotomized group) in the lesion site. Control abducens internuclear neurons were strongly immunoreactive to VEGF and BDNF but showed a weak staining for NT-3 and NGF. Comparisons between groups revealed that lesioned neurons from animals that received NPC implants showed a significant increase in VEGF content with respect to animals receiving vehicle injections. However, the immunoreactivity for BDNF, which was increased in the axotomized group as compared to control, was not modified in the implanted group. The modifications induced by NPC implants on VEGF and BDNF content were specific for the population of axotomized abducens internuclear neurons since the neighboring abducens motoneurons were not affected. Similar levels of NT-3 and NGF immunolabeling were obtained in injured neurons from axotomized and implanted animals. Among all the analyzed neurotrophic factors, only VEGF was expressed by the implanted cells in the lesion site. Our results point to a role of NPC implants in the modulation of neurotrophic factor expression by lesioned central neurons, which might

  12. Neural Progenitor Cell Implants Modulate Vascular Endothelial Growth Factor and Brain-Derived Neurotrophic Factor Expression in Rat Axotomized Neurons

    PubMed Central

    Talaverón, Rocío; Matarredona, Esperanza R.; de la Cruz, Rosa R.; Pastor, Angel M.

    2013-01-01

    Axotomy of central neurons leads to functional and structural alterations which largely revert when neural progenitor cells (NPCs) are implanted in the lesion site. The new microenvironment created by NPCs in the host tissue might modulate in the damaged neurons the expression of a high variety of molecules with relevant roles in the repair mechanisms, including neurotrophic factors. In the present work, we aimed to analyze changes in neurotrophic factor expression in axotomized neurons induced by NPC implants. For this purpose, we performed immunofluorescence followed by confocal microscopy analysis for the detection of vascular endothelial growth factor (VEGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and nerve growth factor (NGF) on brainstem sections from rats with axotomy of abducens internuclear neurons that received NPC implants (implanted group) or vehicle injections (axotomized group) in the lesion site. Control abducens internuclear neurons were strongly immunoreactive to VEGF and BDNF but showed a weak staining for NT-3 and NGF. Comparisons between groups revealed that lesioned neurons from animals that received NPC implants showed a significant increase in VEGF content with respect to animals receiving vehicle injections. However, the immunoreactivity for BDNF, which was increased in the axotomized group as compared to control, was not modified in the implanted group. The modifications induced by NPC implants on VEGF and BDNF content were specific for the population of axotomized abducens internuclear neurons since the neighboring abducens motoneurons were not affected. Similar levels of NT-3 and NGF immunolabeling were obtained in injured neurons from axotomized and implanted animals. Among all the analyzed neurotrophic factors, only VEGF was expressed by the implanted cells in the lesion site. Our results point to a role of NPC implants in the modulation of neurotrophic factor expression by lesioned central neurons, which might

  13. Growth hormone (GH) is a survival rather than a proliferative factor for embryonic striatal neural precursor cells.

    PubMed

    Regalado-Santiago, Citlalli; López-Meraz, María Leonor; Santiago-García, Juan; Fernández-Pomares, Cynthia; Juárez-Aguilar, Enrique

    2013-10-01

    A possible role of GH during central nervous system (CNS) development has been suggested by the presence of this hormone and its receptor in brain areas before its production by the pituitary gland. Although several effects have been reported for GH, the specific role of this hormone during CNS development remains unclear. Here, we examined the effect of GH on proliferation, survival and neurosphere formation in primary cultures of striatal tissue from 14-day-old (E14) mouse embryos. GH receptor gene expression was confirmed by RT-PCR. Primary cultures of embryonic striatal cells were treated with different doses of GH in serum free media, then the number of neurospheres was determined. To examine the GH effect on proliferation and survival of the striatal primary cultures, bromodeoxyuridine (BrdU) and TUNEL immunoreactivity was conducted. In the presence of the epidermal growth factor (EGF), GH increased the formation of neurospheres, with a maximal response at 10 ng/ml, higher doses were inhibitory. In absence of EGF, GH failed to stimulate neurosphere formation. Proliferation rate in the primary striatal cultures was inhibited by 24 or 48 h incubation with GH. However, in the absence of EGF, GH increased BrdU incorporation. GH treatment decreases the rate of apoptosis of nestin and GFAP positive cells in the primary striatal cultures, enhancing neurosphere formation. Our in vitro data demonstrate that GH plays a survival role on the original population of embryonic striatal cells, improving Neural Precursor Cells (NPCs) expansion. We suggest that this GH action could be predominant during striatal neurodevelopment. © 2013.

  14. Human melanoma-initiating cells express neural crest nerve growth factor receptor CD271.

    PubMed

    Boiko, Alexander D; Razorenova, Olga V; van de Rijn, Matt; Swetter, Susan M; Johnson, Denise L; Ly, Daphne P; Butler, Paris D; Yang, George P; Joshua, Benzion; Kaplan, Michael J; Longaker, Michael T; Weissman, Irving L

    2010-07-01

    The question of whether tumorigenic cancer stem cells exist in human melanomas has arisen in the last few years. Here we show that in melanomas, tumour stem cells (MTSCs, for melanoma tumour stem cells) can be isolated prospectively as a highly enriched CD271(+) MTSC population using a process that maximizes viable cell transplantation. The tumours sampled in this study were taken from a broad spectrum of sites and stages. High-viability cells isolated by fluorescence-activated cell sorting and re-suspended in a matrigel vehicle were implanted into T-, B- and natural-killer-deficient Rag2(-/-)gammac(-/-) mice. The CD271(+) subset of cells was the tumour-initiating population in 90% (nine out of ten) of melanomas tested. Transplantation of isolated CD271(+) melanoma cells into engrafted human skin or bone in Rag2(-/-)gammac(-/-) mice resulted in melanoma; however, melanoma did not develop after transplantation of isolated CD271(-) cells. We also show that in mice, tumours derived from transplanted human CD271(+) melanoma cells were capable of metastatsis in vivo. CD271(+) melanoma cells lacked expression of TYR, MART1 and MAGE in 86%, 69% and 68% of melanoma patients, respectively, which helps to explain why T-cell therapies directed at these antigens usually result in only temporary tumour shrinkage.

  15. Human Melanoma Initiating Cells Express Neural Crest Nerve Growth Factor Receptor CD271

    PubMed Central

    Boiko, Alexander D.; Razorenova, Olga V.; van de Rijn, Matt; Swetter, Susan M.; Johnson, Denise L.; Ly, Daphne P.; Butler, Paris D.; Yang, George P.; Joshua, Benzion; Kaplan, Michael J.; Longaker, Michael T.; Weissman, Irving L.

    2010-01-01

    The question whether tumorigenic cancer stem cells exist in human melanomas has arisen recently1. Here we show that in melanomas, tumor stem cells (MTSC) can be isolated prospectively as a highly enriched CD271+ MTSC population using a process that maximizes viable cell transplantation1,6. In this study the tumors sampled were taken from a broad spectrum of sites and stages. High viability FACS isolated cells resuspended in a matrigel vehicle were implanted into T, B, and NK deficient Rag2−/− γc−/− mice (RG) mice. The CD271+ subset of cells was the tumor initiating population in 9/10 melanomas tested. Transplantation of isolated melanoma cells into engrafted human skin or bone in RG mice resulted in melanoma from CD271+ but not CD271− cells. We also showed that tumors transplanted by CD271+ patient cells were capable of metastasis in-vivo. Importantly, CD271+ melanoma cells lacked expression of TYR, MART and MAGE in 86%, 69% and 68% of melanoma patients respectively suggesting why T cell therapies directed at these antigens usually result in only temporary tumor shrinkage. PMID:20596026

  16. N-Cadherin- and L1-functionalised conducting polymers for synergistic stimulation and guidance of neural cell growth.

    PubMed

    Collazos-Castro, Jorge E; Hernández-Labrado, Gabriel R; Polo, José L; García-Rama, Concepción

    2013-05-01

    Conducting polymers are promising materials for advanced neuroprostheses and neural repair devices. However, these challenging technologies demand stable presentation of multiple biomolecules on the polymer surface and fabrication of scaffolds suitable for implantation. We electrosynthesised poly(3,4-ethylenedioxythiophene) doped with poly[(4-styrenesulfonic acid)-co-(maleic acid)] (PEDOT:PSS-co-MA) on gold-coated surfaces or carbon microfibres, functionalised the polymer by covalent immobilisation of anti-IgG antibodies and subsequent binding of N-Cadherin and L1 recombinant proteins, and used these materials as substrates for culturing cerebral cortex neurons. N-Cadherin and L1 were much more effective than polylysine in promoting axonal elongation and collateralisation on the polymer. However, N-Cadherin also induced cell migration and dendritic extension and branching, whereas L1 inhibited dendrites. Dual functionalisation with N-Cadherin and L1 produced synergistic effects on neuronal growth that could not be achieved with either of the proteins when used alone. PEDOT:PSS-co-MA electrosynthesised on carbon microfibres showed good electrochemical properties and, when biofunctionalised with N-Cadherin or L1, stimulated very long and guided axonal elongation. Finally, electrochemical impedance spectroscopy, cyclic voltammetry and chronoamperometry showed that the good electrical properties of PEDOT:PSS-co-MA were not degraded by covalent peptide attachment, indicating that this polymer is suitable for multiple biofunctionalisation of electroactive surfaces in neuroprosthetic and lesion-bridging applications. Copyright © 2013 Elsevier Ltd. All rights reserved.

  17. Electric field stimulation through a biodegradable polypyrrole-co-polycaprolactone substrate enhances neural cell growth

    PubMed Central

    Nguyen, Hieu T; Wei, Claudia; Chow, Jacqueline K; Nguyen, Alvin; Coursen, Jeff; Sapp, Shawn; Luebben, Silvia; Chang, Emily; Ross, Robert; Schmidt, Christine E

    2014-01-01

    Nerve guidance conduits (NGCs) are FDA-approved devices used to bridge gaps across severed nerve cables and help direct axons sprouting from the proximal end toward the distal stump. In this paper we present the development of a novel electrically conductive, biodegradable NGC made from a polypyrrole-block-polycaprolactone (PPy-PCL) copolymer material laminated with poly(lactic-co-glycolic acid) (PLGA). The PPy-PCL has a bulk conductivity ranging 10–20 S/cm and loses 40 wt% after 7 months under physiologic conditions. Dorsal root ganglia (DRG) grown on flat PPy-PCL/PLGA material exposed to direct current electric fields (EF) of 100 mV/cm for 2 h increased axon growth by 13% (± 2%) towards either electrode of a 2-electrode setup, compared to control grown on identical substrates without EF exposure. Alternating current increased axon growth by 21% (± 3%) without an observable directional preference, compared to the same control group. The results from this study demonstrate PLGA-coated PPy-PCL is a unique biodegradable material that can deliver substrate EF stimulation to improve axon growth for peripheral nerve repair. PMID:23964001

  18. Electric field stimulation through a biodegradable polypyrrole-co-polycaprolactone substrate enhances neural cell growth.

    PubMed

    Nguyen, Hieu T; Sapp, Shawn; Wei, Claudia; Chow, Jacqueline K; Nguyen, Alvin; Coursen, Jeff; Luebben, Silvia; Chang, Emily; Ross, Robert; Schmidt, Christine E

    2014-08-01

    Nerve guidance conduits (NGCs) are FDA-approved devices used to bridge gaps across severed nerve cables and help direct axons sprouting from the proximal end toward the distal stump. In this article, we present the development of a novel electrically conductive, biodegradable NGC made from a polypyrrole-block-polycaprolactone (PPy-PCL) copolymer material laminated with poly(lactic-co-glycolic acid) (PLGA). The PPy-PCL has a bulk conductivity ranging 10-20 S/cm and loses 40 wt % after 7 months under physiologic conditions. Dorsal root ganglia (DRG) grown on flat PPy-PCL/PLGA material exposed to direct current electric fields (EF) of 100 mV/cm for 2 h increased axon growth by 13% (± 2%) toward either electrode of a 2-electrode setup, compared with control grown on identical substrates without EF exposure. Alternating current increased axon growth by 21% (±3%) without an observable directional preference, compared with the same control group. The results from this study demonstrate PLGA-coated PPy-PCL is a unique biodegradable material that can deliver substrate EF stimulation to improve axon growth for peripheral nerve repair.

  19. Smad7 Regulates the Adult Neural Stem/Progenitor Cell Pool in a Transforming Growth Factor β- and Bone Morphogenetic Protein-Independent Manner▿

    PubMed Central

    Krampert, Monika; Chirasani, Sridhar Reddy; Wachs, Frank-Peter; Aigner, Robert; Bogdahn, Ulrich; Yingling, Jonathan M.; Heldin, Carl-Henrik; Aigner, Ludwig; Heuchel, Rainer

    2010-01-01

    Members of the transforming growth factor β (TGF-β) family of proteins modulate the proliferation, differentiation, and survival of many different cell types. Neural stem and progenitor cells (NPCs) in the adult brain are inhibited in their proliferation by TGF-β and by bone morphogenetic proteins (BMPs). Here, we investigated neurogenesis in a hypomorphic mouse model for the TGF-β and BMP inhibitor Smad7, with the hypothesis that NPC proliferation might be reduced due to increased TGF-β and BMP signaling. Unexpectedly, we found enhanced NPC proliferation as well as an increased number of label-retaining cells in vivo. The enhanced proliferation potential of mutant cells was retained in vitro in neurosphere cultures. We observed a higher sphere-forming capacity as well as faster growth and cell cycle progression. Use of specific inhibitors revealed that these effects were independent of TGF-β and BMP signaling. The enhanced proliferation might be at least partially mediated by elevated signaling via epidermal growth factor (EGF) receptor, as mutant cells showed higher expression and activation levels of the EGF receptor. Conversely, an EGF receptor inhibitor reduced the proliferation of these cells. Our data indicate that endogenous Smad7 regulates neural stem/progenitor cell proliferation in a TGF-β- and BMP-independent manner. PMID:20479122

  20. Ensconsin/Map7 promotes microtubule growth and centrosome separation in Drosophila neural stem cells

    PubMed Central

    Gallaud, Emmanuel; Caous, Renaud; Pascal, Aude; Bazile, Franck; Gagné, Jean-Philippe; Huet, Sébastien; Poirier, Guy G.; Chrétien, Denis; Richard-Parpaillon, Laurent

    2014-01-01

    The mitotic spindle is crucial to achieve segregation of sister chromatids. To identify new mitotic spindle assembly regulators, we isolated 855 microtubule-associated proteins (MAPs) from Drosophila melanogaster mitotic or interphasic embryos. Using RNAi, we screened 96 poorly characterized genes in the Drosophila central nervous system to establish their possible role during spindle assembly. We found that Ensconsin/MAP7 mutant neuroblasts display shorter metaphase spindles, a defect caused by a reduced microtubule polymerization rate and enhanced by centrosome ablation. In agreement with a direct effect in regulating spindle length, Ensconsin overexpression triggered an increase in spindle length in S2 cells, whereas purified Ensconsin stimulated microtubule polymerization in vitro. Interestingly, ensc-null mutant flies also display defective centrosome separation and positioning during interphase, a phenotype also detected in kinesin-1 mutants. Collectively, our results suggest that Ensconsin cooperates with its binding partner Kinesin-1 during interphase to trigger centrosome separation. In addition, Ensconsin promotes microtubule polymerization during mitosis to control spindle length independent of Kinesin-1. PMID:24687279

  1. Ensconsin/Map7 promotes microtubule growth and centrosome separation in Drosophila neural stem cells.

    PubMed

    Gallaud, Emmanuel; Caous, Renaud; Pascal, Aude; Bazile, Franck; Gagné, Jean-Philippe; Huet, Sébastien; Poirier, Guy G; Chrétien, Denis; Richard-Parpaillon, Laurent; Giet, Régis

    2014-03-31

    The mitotic spindle is crucial to achieve segregation of sister chromatids. To identify new mitotic spindle assembly regulators, we isolated 855 microtubule-associated proteins (MAPs) from Drosophila melanogaster mitotic or interphasic embryos. Using RNAi, we screened 96 poorly characterized genes in the Drosophila central nervous system to establish their possible role during spindle assembly. We found that Ensconsin/MAP7 mutant neuroblasts display shorter metaphase spindles, a defect caused by a reduced microtubule polymerization rate and enhanced by centrosome ablation. In agreement with a direct effect in regulating spindle length, Ensconsin overexpression triggered an increase in spindle length in S2 cells, whereas purified Ensconsin stimulated microtubule polymerization in vitro. Interestingly, ensc-null mutant flies also display defective centrosome separation and positioning during interphase, a phenotype also detected in kinesin-1 mutants. Collectively, our results suggest that Ensconsin cooperates with its binding partner Kinesin-1 during interphase to trigger centrosome separation. In addition, Ensconsin promotes microtubule polymerization during mitosis to control spindle length independent of Kinesin-1.

  2. Insulin-like Growth Factor-II (IGF-II) and IGF-II Analogs with Enhanced Insulin Receptor-a Binding Affinity Promote Neural Stem Cell Expansion*

    PubMed Central

    Ziegler, Amber N.; Chidambaram, Shravanthi; Forbes, Briony E.; Wood, Teresa L.; Levison, Steven W.

    2014-01-01

    The objective of this study was to employ genetically engineered IGF-II analogs to establish which receptor(s) mediate the stemness promoting actions of IGF-II on mouse subventricular zone neural precursors. Neural precursors from the subventricular zone were propagated in vitro in culture medium supplemented with IGF-II analogs. Cell growth and identity were analyzed using sphere generation and further analyzed by flow cytometry. F19A, an analog of IGF-II that does not bind the IGF-2R, stimulated an increase in the proportion of neural stem cells (NSCs) while decreasing the proportion of the later stage progenitors at a lower concentration than IGF-II. V43M, which binds to the IGF-2R with high affinity but which has low binding affinity to the IGF-1R and to the A isoform of the insulin receptor (IR-A) failed to promote NSC growth. The positive effects of F19A on NSC growth were unaltered by the addition of a functional blocking antibody to the IGF-1R. Altogether, these data lead to the conclusion that IGF-II promotes stemness of NSCs via the IR-A and not through activation of either the IGF-1R or the IGF-2R. PMID:24398690

  3. Designing Neural Networks in Culture: Experiments are described for controlled growth, of nerve cells taken from rats, in predesigned geometrical patterns on laboratory culture dishes.

    PubMed

    Wheeler, Bruce C; Brewer, Gregory J

    2010-03-01

    Technology has advanced to where it is possible to design and grow-with predefined geometry and surprisingly good fidelity-living networks of neurons in culture dishes. Here we overview the elements of design, emphasizing the lithographic techniques that alter the cell culture surface which in turn influences the attachment and growth of the neural networks. Advanced capability in this area makes it possible to design networks of desired complexity. Other issues addressed include the influence of glial cells and media on activity and the potential for extending the designs into three dimensions. Investigators are advancing the art and science of analyzing and controlling through stimulation the function of the neural networks, including the ability to take advantage of their geometric form in order to influence functional properties.

  4. Accelerating proliferation of neural stem/progenitor cells in collagen sponges immobilized with engineered basic fibroblast growth factor for nervous system tissue engineering.

    PubMed

    Ma, Fukai; Xiao, Zhifeng; Chen, Bing; Hou, Xianglin; Han, Jin; Zhao, Yannan; Dai, Jianwu; Xu, Ruxiang

    2014-03-10

    Neural stem/progenitor cells (NS/PCs) play a therapeutic role in nervous system diseases and contribute to functional recovery. However, their efficacy is limited as the majority of cells die post-transplantation. In this study, collagen sponges were utilized as carriers for NS/PCs. Basic fibroblast growth factor (bFGF), a mitogen for NS/PCs, was incorporated into the collagen sponges to stimulate NS/PC proliferation. However, the effect of native bFGF is limited because it diffuses into the culture medium and is lost following medium exchange. To overcome this problem, a collagen-binding polypeptide domain, which has high affinity to collagen, was fused with bFGF to sustain the exposure of NS/PCs within the collagen sponges to bFGF. The results indicated that the number of NS/PCs was significantly higher in collagen sponges incorporating engineered bFGF than in those with native bFGF or the PBS control after 7 days in culture. Here, we designed a natural biological neural scaffold consisting of collagen sponges, engineered bFGF, and NS/PCs. In addition to the effect of proliferated NS/PCs, the engineered bFGF retained in the natural biological neural scaffolds could have a direct effect on nervous system reconstruction. The two aspects of the natural biological neural scaffolds may produce synergistic effects, and so they represent a promising candidate for nervous system repair.

  5. Upregulation of Slc38a1 Gene Along with Promotion of Neurosphere Growth and Subsequent Neuronal Specification in Undifferentiated Neural Progenitor Cells Exposed to Theanine.

    PubMed

    Takarada, Takeshi; Ogura, Masato; Nakamichi, Noritaka; Kakuda, Takami; Nakazato, Ryota; Kokubo, Hiroshi; Ikeno, Shinsuke; Nakamura, Saki; Kutsukake, Takaya; Hinoi, Eiichi; Yoneda, Yukio

    2016-02-01

    We have shown marked promotion of both cluster growth and neuronal specification in pluripotent P19 cells with overexpression of solute carrier 38a1 (Slc38a1), which is responsible for membrane transport of glutamine. In this study, we evaluated pharmacological profiles of the green tea amino acid ingredient theanine, which is a good substrate for glutamine transporters, on proliferation and neuronal specification in neural progenitor cells from embryonic rat neocortex. Sustained exposure to theanine, but not glutamine, accelerated the growth of neurospheres composed of proliferating cells and 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) reducing activity at concentrations of 1-100 μM in undifferentiated progenitor cells. Such prior exposure to theanine promoted spontaneous and induced commitment to a neuronal lineage with concomitant deteriorated astroglial specification. Selective upregulation was seen in the expression of Slc38a1 in progenitor cells cultured with theanine. Similarly significant increases in cluster growth and MTT reducing activity were found in P19 cells cultured with theanine for 4 days. Luciferase activity was doubled in a manner sensitive to the deletion of promoter regions in P19 cells with a luciferase reporter plasmid of the Slc38a1 promoter after sustained exposure to theanine for 4 days. Overexpression of X-box binding protein-1 led to a marked increase in luciferase activity in P19 cells transfected with the Slc38a1 reporter plasmid. These results suggest that theanine accelerates cellular proliferation and subsequent neuronal specification through a mechanism relevant to upregulation of Slc38a1 gene in undifferentiated neural progenitor cells.

  6. Cytosine deaminase-expressing human neural stem cells inhibit tumor growth in prostate cancer-bearing mice.

    PubMed

    Lee, Hong Jun; Doo, Seung Whan; Kim, Dae Hong; Cha, Young Joo; Kim, Jae Heon; Song, Yun Seob; Kim, Seung U

    2013-07-10

    Prostate cancer is the most common malignancy among men. Prostate cancer-related deaths are largely attributable to the development of hormone resistance in the tumor. No effective chemotherapy has yet been developed for advanced prostate cancer. It is desirable if a drug can be delivered directly and specifically to prostate cancer cells. Stem cells have selective migration ability toward cancer cells and therapeutic genes can be easily transduced into stem cells. In one form of gene therapy for cancer, the stem cells carry a gene encoding an enzyme that transforms an inert prodrug into a toxic product. Cytosine deaminase (CD) transforms the pro-drug 5-fluorocytosine into highly cytotoxic 5-fluorouracil (5-FU). The migration of the genetically modified stem cells was monitored by molecular magnetic resonance imaging, after labeling the stem cells with fluorescent magnetic nanoparticles (MNPs). Human neural stem cells encoding CD (HB1.F3.CD) were prepared and labeled with MNP. In tumor-bearing C57B mice, systemically transplanted HB1.F3.CD stem cells migrated toward the tumor and in combination with prodrug 5-FC, the volume of tumor implant was significantly reduced. These findings may contribute to development of a new selective chemotherapeutic strategy against prostate cancer.

  7. Vascular endothelial growth factor influences migration and focal adhesions, but not proliferation or viability, of human neural stem/progenitor cells derived from olfactory epithelium.

    PubMed

    Ramírez-Rodríguez, Gerardo Bernabé; Perera-Murcia, Gerardo Rodrigo; Ortiz-López, Leonardo; Vega-Rivera, Nelly Maritza; Babu, Harish; García-Anaya, Maria; González-Olvera, Jorge Julio

    2017-09-01

    In humans, new neurons are continuously added in the olfactory epithelium even in the adulthood. The resident neural stem/progenitor cells (hNS/PCs-OE) in the olfactory epithelium are influenced by several growth factors and neurotrophins. Among these modulators the vascular endothelial growth factor (VEGF) has attracted attention due its implicated in cell proliferation, survival and migration of other type of neural/stem progenitor cells. Interestingly, VEGFr2 receptor expression in olfactory epithelium has been described in amphibians but not in humans. Here we show that VEGFr is expressed in the hNS/PCs-OE. We also investigated the effect of VEGF on the hNS/PCs-OE proliferation, viability and migration in vitro. Additionally, pharmacological approaches showed that VEGF (0.5 ng/ml)-stimulated migration of hNS/PCs-OE was blocked with the compound DMH4, which prevents the activation of VEGFr2. Similar effects were found with the inhibitors for Rac (EHT1864) and p38MAPK (SB203850) proteins, respectively. These observations occurred with changes in focal adhesion contacts. However, no effects of VEGF on proliferation or viability were found in hNS/PCs-OE. Our results suggest that hNS/PCs-OE respond to VEGF involving VEGFr2, Rac and p38MAPK. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Neural Stem Cells and Glioblastoma

    PubMed Central

    Rispoli, Rossella; Conti, Carlo; Celli, Paolo; Caroli, Emanuela; Carletti, Sandro

    2014-01-01

    Summary Glioblastoma multiforme represents one of the most common brain cancers with a rather heterogeneous cellular composition, as indicated by the term “multiforme". Recent reports have described the isolation and identification of cancer neural stem cells from human adult glioblastoma multiforme, which possess the capacity to establish, sustain, and expand these tumours, even under the challenging settings posed by serial transplantation experiments. Our study focused on the distribution of neural cancer stem cells inside the tumour. The study is divided into three phases: removal of tumoral specimens in different areas of the tumour (centre, periphery, marginal zone) in an operative room equipped with a 1.5 T scanner; isolation and characterization of neural cancer stem cells from human adult glioblastoma multiforme; identification of neural cancer stem cell distribution inside the tumour. PMID:24750704

  9. β-Arrestin1/miR-326 Transcription Unit Is Epigenetically Regulated in Neural Stem Cells Where It Controls Stemness and Growth Arrest

    PubMed Central

    Begalli, Federica; Abballe, Luana; Catanzaro, Giuseppina; Vacca, Alessandra; Napolitano, Maddalena; Tafani, Marco; Giangaspero, Felice; Locatelli, Franco

    2017-01-01

    Cell development is regulated by a complex network of mRNA-encoded proteins and microRNAs, all funnelling onto the modulation of self-renewal or differentiation genes. How intragenic microRNAs and their host genes are transcriptionally coregulated and their functional relationships for the control of neural stem cells (NSCs) are poorly understood. We propose here the intragenic miR-326 and its host gene β-arrestin1 as novel players whose epigenetic silencing maintains stemness in normal cerebellar stem cells. Such a regulation is mediated by CpG islands methylation of the common promoter. Epigenetic derepression of β-arrestin1/miR-326 by differentiation signals or demethylating agents leads to suppression of stemness features and cell growth and promotes cell differentiation. β-Arrestin1 inhibits cell proliferation by enhancing the nuclear expression of the cyclin-dependent kinase inhibitor p27. Therefore, we propose a new mechanism for the control of cerebellar NSCs where a coordinated epigenetic mechanism finely regulates β-arrestin1/miR-326 expression and consequently NSCs stemness and cell growth. PMID:28298929

  10. Human neural stem cells improve cognition and promote synaptic growth in two complementary transgenic models of Alzheimer's disease and neuronal loss.

    PubMed

    Ager, Rahasson R; Davis, Joy L; Agazaryan, Andy; Benavente, Francisca; Poon, Wayne W; LaFerla, Frank M; Blurton-Jones, Mathew

    2015-07-01

    Alzheimer's disease (AD) is the most prevalent age-related neurodegenerative disorder, affecting over 35 million people worldwide. Pathologically, AD is characterized by the progressive accumulation of β-amyloid (Aβ) plaques and neurofibrillary tangles within the brain. Together, these pathologies lead to marked neuronal and synaptic loss and corresponding impairments in cognition. Current treatments, and recent clinical trials, have failed to modify the clinical course of AD; thus, the development of novel and innovative therapies is urgently needed. Over the last decade, the potential use of stem cells to treat cognitive impairment has received growing attention. Specifically, neural stem cell transplantation as a treatment for AD offers a novel approach with tremendous therapeutic potential. We previously reported that intrahippocampal transplantation of murine neural stem cells (mNSCs) can enhance synaptogenesis and improve cognition in 3xTg-AD mice and the CaM/Tet-DT(A) model of hippocampal neuronal loss. These promising findings prompted us to examine a human neural stem cell population, HuCNS-SC, which has already been clinically tested for other neurodegenerative disorders. In this study, we provide the first evidence that transplantation of research grade HuCNS-SCs can improve cognition in two complementary models of neurodegeneration. We also demonstrate that HuCNS-SC cells can migrate and differentiate into immature neurons and glia and significantly increase synaptic and growth-associated markers in both 3xTg-AD and CaM/Tet-DTA mice. Interestingly, improvements in aged 3xTg-AD mice were not associated with altered Aβ or tau pathology. Rather, our findings suggest that human NSC transplantation improves cognition by enhancing endogenous synaptogenesis. Taken together, our data provide the first preclinical evidence that human NSC transplantation could be a safe and effective therapeutic approach for treating AD. © 2014 The Authors. Hippocampus

  11. Dopaminergic Neuronal Differentiation from the Forebrain-Derived Human Neural Stem Cells Induced in Cultures by Using a Combination of BMP-7 and Pramipexole with Growth Factors

    PubMed Central

    Yang, HongNa; Wang, Jing; Wang, Feng; Liu, XiaoDun; Chen, Heng; Duan, WeiMing; Qu, TingYu

    2016-01-01

    Transplantation of dopaminergic (DA) neurons is considered to be the most promising therapeutic strategy for replacing degenerated dopamine cells in the midbrain of Parkinson's disease (PD), thereby restoring normal neural circuit function and slow clinical progression of the disease. Human neural stem cells (hNSCs) derived from fetal forebrain are thought to be the important cell sources for producing DA neurons because of their multipotency for differentiation and long-term expansion property in cultures. However, low DA differentiation of the forebrain-derived hNSCs limited their therapeutic potential in PD. In the current study, we explored a combined application of Pramipexole (PRX), bone morphogenetic proteins 7 (BMP-7), and growth factors, including acidic fibroblast factor (aFGF), forskolin, and phorbol-12-myristae-13-acetate (TPA), to induce differentiation of forebrain-derived hNSCs toward DA neurons in cultures. We found that DA neuron-associated genes, including Nurr1, Neurogenin2 (Ngn2), and tyrosine hydroxylase (TH) were significantly increased after 24 h of differentiation by RT-PCR analysis (p < 0.01). Fluorescent examination showed that about 25% of cells became TH-positive neurons at 24 h, about 5% of cells became VMAT2 (vascular monoamine transporter 2)-positive neurons, and less than 5% of cells became DAT (dopamine transporter)-positive neurons at 72 h following differentiation in cultures. Importantly, these TH-, VMAT2-, and DAT-expressing neurons were able to release dopamine into cultures under both of the basal and evoked conditions. Dopamine levels released by DA neurons produced using our protocol were significantly higher compared to the control groups (P < 0.01), as examined by ELISA. Our results demonstrated that the combination of PRX, BMP-7, and growth factors was able to greatly promote differentiation of the forebrain-derived hNSCs into DA-releasing neurons. PMID:27147976

  12. Nitric oxide from inflammatory origin impairs neural stem cell proliferation by inhibiting epidermal growth factor receptor signaling

    PubMed Central

    Carreira, Bruno P.; Morte, Maria I.; Santos, Ana I.; Lourenço, Ana S.; Ambrósio, António F.; Carvalho, Caetana M.; Araújo, Inês M.

    2014-01-01

    Neuroinflammation is characterized by activation of microglial cells, followed by production of nitric oxide (NO), which may have different outcomes on neurogenesis, favoring or inhibiting this process. In the present study, we investigated how the inflammatory mediator NO can affect proliferation of neural stem cells (NSCs), and explored possible mechanisms underlying this effect. We investigated which mechanisms are involved in the regulation of NSC proliferation following treatment with an inflammatory stimulus (lipopolysaccharide plus IFN-γ), using a culture system of subventricular zone (SVZ)-derived NSCs mixed with microglia cells obtained from wild-type mice (iNOS+/+) or from iNOS knockout mice (iNOS-/-). We found an impairment of NSC cell proliferation in iNOS+/+ mixed cultures, which was not observed in iNOS-/- mixed cultures. Furthermore, the increased release of NO by activated iNOS+/+ microglial cells decreased the activation of the ERK/MAPK signaling pathway, which was concomitant with an enhanced nitration of the EGF receptor. Preventing nitrogen reactive species formation with MnTBAP, a scavenger of peroxynitrite (ONOO-), or using the ONOO- degradation catalyst FeTMPyP, cell proliferation and ERK signaling were restored to basal levels in iNOS+/+ mixed cultures. Moreover, exposure to the NO donor NOC-18 (100 μM), for 48 h, inhibited SVZ-derived NSC proliferation. Regarding the antiproliferative effect of NO, we found that NOC-18 caused the impairment of signaling through the ERK/MAPK pathway, which may be related to increased nitration of the EGF receptor in NSC. Using MnTBAP nitration was prevented, maintaining ERK signaling, rescuing NSC proliferation. We show that NO from inflammatory origin leads to a decreased function of the EGF receptor, which compromised proliferation of NSC. We also demonstrated that NO-mediated nitration of the EGF receptor caused a decrease in its phosphorylation, thus preventing regular proliferation signaling through

  13. Three-dimensional bioprinting of rat embryonic neural cells.

    PubMed

    Lee, Wonhye; Pinckney, Jason; Lee, Vivian; Lee, Jong-Hwan; Fischer, Krisztina; Polio, Samuel; Park, Je-Kyun; Yoo, Seung-Schik

    2009-05-27

    We present a direct cell printing technique to pattern neural cells in a three-dimensional (3D) multilayered collagen gel. A layer of collagen precursor was printed to provide a scaffold for the cells, and the rat embryonic neurons and astrocytes were subsequently printed on the layer. A solution of sodium bicarbonate was applied to the cell containing collagen layer as nebulized aerosols, which allowed the gelation of the collagen. This process was repeated layer-by-layer to construct the 3D cell-hydrogel composites. Upon characterizing the relationship between printing resolutions and the growth of printed neural cells, single/multiple layers of neural cell-hydrogel composites were constructed and cultured. The on-demand capability to print neural cells in a multilayered hydrogel scaffold offers flexibility in generating artificial 3D neural tissue composites.

  14. Mobilization of Neural Stem Cells and Generation of New Neurons in 6-OHDA–lesioned Rats by Intracerebroventricular Infusion of Liver Growth Factor

    PubMed Central

    Gonzalo-Gobernado, Rafael; Reimers, Diana; Herranz, Antonio S.; Díaz-Gil, Juan José; Osuna, Cristina; Asensio, María José; Baena, Silvia; Rodríguez-Serrano, Macarena; Bazán, Eulalia

    2009-01-01

    Neural stem cells with self-renewal and multilineage potential persist in the subventricular zone of the adult mammalian forebrain. These cells remain relatively quiescent but, under certain conditions, can be stimulated, giving rise to new neurons. Liver growth factor (LGF) is a mitogen for liver cells that shows biological activity in extrahepatic sites and is useful for neuroregenerative therapies. The aim of this study was to investigate the potential neurogenic activity of LGF in the 6-hydroxydopamine rat model of Parkinson's disease. Proliferation was significantly increased in the subventricular zone and denervated striatum of rats receiving ICV LGF infusions, and 25% of the proliferating cells were doublecortin-positive neurons. Doublecortin-positive cells with the morphology of migrating neuroblasts were also observed in the dorsal and ventral regions of the striatum of LGF-infused animals. Moreover, some newly generated cells were neuronal nuclei-positive mature neurons. LGF also stimulated microglia and induced astrogliosis, both phenomena associated with generation and migration of new neurons in the adult brain. In summary, our study shows that LGF stimulates neurogenesis when applied intraventricularly in 6-hydroxydopamine–lesioned rats. Considering that this factor also promotes neuronal migration into damaged tissue, we propose LGF as a novel factor useful for neuronal replacement in neurodegenerative diseases. (J Histochem Cytochem 57:491–502, 2009) PMID:19188487

  15. Basic fibroblast growth factor (bFGF) acts intracellularly to cause the transdifferentiation of avian neural crest-derived Schwann cell precursors into melanocytes.

    PubMed

    Sherman, L; Stocker, K M; Morrison, R; Ciment, G

    1993-08-01

    We previously found that cultured neural crest-derived cells from embryonic quail peripheral nerves, which consist mostly of Schwann cell precursors, gave rise to melanocytes following treatment with basic fibroblast growth factor (bFGF) or 12-O-tetradecanoyl phorbol-13-acetate (TPA). Here, we show that antisense deoxyoligonucleotides targeted against two regions of the bFGF mRNA transcript blocked this TPA-induced transdifferentiation of Schwann cell precursors. Neither sense nor scrambled antisense control oligonucleotides had any effect in this regard. TPA increased bFGF protein expression in cell lysates but not in conditioned media from these cultures, and this expression was localized to the nucleus and cytoplasm. Furthermore, bFGF-neutralizing antibodies and inositol-hexakisphosphate (InsP6) both inhibited pigmentation caused by exogenous bFGF, but had no affect on TPA-induced melanogenesis, suggesting that bFGF is not released by these cells. These data indicate that bFGF is necessary for the TPA-induced transdifferentiation of Schwann cell precursors into melanocytes and that bFGF acts via an intracrine mechanism.

  16. Self-renewing and differentiating properties of cortical neural stem cells are selectively regulated by basic fibroblast growth factor (FGF) signaling via specific FGF receptors.

    PubMed

    Maric, Dragan; Fiorio Pla, Alessandra; Chang, Yoong Hee; Barker, Jeffery L

    2007-02-21

    Developmental processes mediating the initiation of lineage commitment from self-renewing neural stem cells (NSCs) remain mostly unclear because of the persisting ambiguity in identifying true NSCs from proliferative lineage-restricted progenitors (LRPs), which are directly or indirectly derived from NSCs. Our multilineage immunohistochemical analyses of early embryonic rat telencephalon at the onset of neurogenesis revealed clear dorsoventral gradients in the emergence of two types of neuronal progenitors (NPs) from multilineage-negative NSCs. Enumeration of NSCs using comprehensive flow cytometric analysis demonstrated that their precipitous decline in vivo involved both active differentiation into NPs and an increased propensity toward apoptosis. Both processes paralleled the dorsoventral changes in fibroblast growth factor receptor (FGFR) expressions. NSCs residing in the dorsal telencephalon coexpressed FGFR1 and FGFR3, whereas those residing in the ventral telencephalon also expressed FGFR2. NSCs exposed to basic fibroblast growth factor (bFGF) in vitro generated four stereotypical clonal expansion states: efficiently self-renewing, inefficiently self-renewing limited by apoptosis, exclusively neurogenic, and multipotential, generating up to five types of LRPs. The plasticity among these expansion states depended on ambient [bFGF], telencephalic developmental stage, and differential activation/inactivation of specific FGFRs. Coactivation of FGFR1 and FGFR3 promoted symmetrical divisions of NSCs (self-renewal), whereas inactivation of either triggered asymmetrical divisions and neurogenesis from these cells. Developmental upregulation of FGFR2 expression correlated with a shift of NSCs into a multipotential state or apoptosis. These results provide new insights regarding the roles of FGFRs in diversification of NSC properties and initiation of neural lineage-restricted differentiation.

  17. [Effect of antepartum taurine supplementation in regulating the activity of Rho family factors and promoting the proliferation of neural stem cells in neonatal rats with fetal growth restriction].

    PubMed

    Li, Xiang-Wen; Li, Fang; Liu, Jing; Wang, Yan; Fu, Wei

    2016-11-01

    To study the possible effect of antepartum taurine supplementation in regulating the activity of Rho family factors and promoting the proliferation of neural stem cells in neonatal rats with fetal growth restriction (FGR), and to provide a basis for antepartum taurine supplementation to promote brain development in children with FGR. A total of 24 pregnant Sprague-Dawley rats were randomly divided into three groups: control, FGR, and taurine (n=8 each ). A rat model of FGR was established by food restriction throughout pregnancy. RT-PCR, immunohistochemistry, and Western blot were used to measure the expression of the specific intracellular markers for neural stem cells fatty acid binding protein 7 (FABP7), Rho-associated coiled-coil containing protein kinase 2 (ROCK2), ras homolog gene family, member A (RhoA), and Ras-related C3 botulinum toxin substrate (Rac). The FGR group had significantly lower OD value of FABP7-positive cells and mRNA and protein expression of FABP7 than the control group, and the taurine group had significantly higher OD value of FABP7-positive cells and mRNA and protein expression of FABP7 than the FGR group (P<0.05). The FGR group had significantly higher mRNA expression of RhoA and ROCK2 than the control group. The taurine group had significantly higher mRNA expression of RhoA and ROCK2 than the control group and significantly lower expression than the FGR group (P<0.05). The FGR group had significantly lower mRNA expression of Rac than the control group. The taurine group had significantly higher mRNA expression of Rac than the FGR and control groups (P<0.05). The FGR group had significantly higher protein expression of RhoA and ROCK2 than the control group. The taurine group had significantly lower protein expression of RhoA and ROCK2 than the FGR group (P<0.05). Antepartum taurine supplementation can promote the proliferation of neural stem cells in rats with FGR, and its mechanism may be related to the regulation of the activity of

  18. Proliferation control in neural stem and progenitor cells

    PubMed Central

    Homem, Catarina CF; Repic, Marko; Knoblich, Juergen A

    2015-01-01

    Neural circuit function can be drastically affected by variations in the number of cells that are produced during development or by a reduction in adult cell number due to disease. Unlike many other organs, the brain is unable to compensate for such changes by increasing cell numbers or altering the size of the cells. For this reason, unique cell cycle and cell growth control mechanisms operate in the developing and adult brain. In Drosophila melanogaster and mammalian neural stem and progenitor cells these mechanisms are intricately coordinated with the developmental age and the nutritional, metabolic and hormonal state of the animal. Defects in neural stem cell proliferation that result in the generation of incorrect cell numbers or defects in neural stem cell differentiation can cause microcephaly or megalencephaly. PMID:26420377

  19. Neural Transcription Factors: from Embryos to Neural Stem Cells

    PubMed Central

    Lee, Hyun-Kyung; Lee, Hyun-Shik; Moody, Sally A.

    2014-01-01

    The early steps of neural development in the vertebrate embryo are regulated by sets of transcription factors that control the induction of proliferative, pluripotent neural precursors, the expansion of neural plate stem cells, and their transition to differentiating neural progenitors. These early events are critical for producing a pool of multipotent cells capable of giving rise to the multitude of neurons and glia that form the central nervous system. In this review we summarize findings from gain- and loss-of-function studies in embryos that detail the gene regulatory network responsible for these early events. We discuss whether this information is likely to be similar in mammalian embryonic and induced pluripotent stem cells that are cultured according to protocols designed to produce neurons. The similarities and differences between the embryo and stem cells may provide important guidance to stem cell protocols designed to create immature neural cells for therapeutic uses. PMID:25234468

  20. Multiple roles of Activin/Nodal, bone morphogenetic protein, fibroblast growth factor and Wnt/β-catenin signalling in the anterior neural patterning of adherent human embryonic stem cell cultures

    PubMed Central

    Lupo, Giuseppe; Novorol, Claire; Smith, Joseph R.; Vallier, Ludovic; Miranda, Elena; Alexander, Morgan; Biagioni, Stefano; Pedersen, Roger A.; Harris, William A.

    2013-01-01

    Several studies have successfully produced a variety of neural cell types from human embryonic stem cells (hESCs), but there has been limited systematic analysis of how different regional identities are established using well-defined differentiation conditions. We have used adherent, chemically defined cultures to analyse the roles of Activin/Nodal, bone morphogenetic protein (BMP), fibroblast growth factor (FGF) and Wnt/β-catenin signalling in neural induction, anteroposterior patterning and eye field specification in hESCs. We show that either BMP inhibition or activation of FGF signalling is required for effective neural induction, but these two pathways have distinct outcomes on rostrocaudal patterning. While BMP inhibition leads to specification of forebrain/midbrain positional identities, FGF-dependent neural induction is associated with strong posteriorization towards hindbrain/spinal cord fates. We also demonstrate that Wnt/β-catenin signalling is activated during neural induction and promotes acquisition of neural fates posterior to forebrain. Therefore, inhibition of this pathway is needed for efficient forebrain specification. Finally, we provide evidence that the levels of Activin/Nodal and BMP signalling have a marked influence on further forebrain patterning and that constitutive inhibition of these pathways represses expression of eye field genes. These results show that the key mechanisms controlling neural patterning in model vertebrate species are preserved in adherent, chemically defined hESC cultures and reveal new insights into the signals regulating eye field specification. PMID:23576785

  1. Identification and characterization of secondary neural tube-derived embryonic neural stem cells in vitro.

    PubMed

    Shaker, Mohammed R; Kim, Joo Yeon; Kim, Hyun; Sun, Woong

    2015-05-15

    Secondary neurulation is an embryonic progress that gives rise to the secondary neural tube, the precursor of the lower spinal cord region. The secondary neural tube is derived from aggregated Sox2-expressing neural cells at the dorsal region of the tail bud, which eventually forms rosette or tube-like structures to give rise to neural tissues in the tail bud. We addressed whether the embryonic tail contains neural stem cells (NSCs), namely secondary NSCs (sNSCs), with the potential for self-renewal in vitro. Using in vitro neurosphere assays, neurospheres readily formed at the rosette and neural-tube levels, but less frequently at the tail bud tip level. Furthermore, we identified that sNSC-generated neurospheres were significantly smaller in size compared with cortical neurospheres. Interestingly, various cell cycle analyses revealed that this difference was not due to a reduction in the proliferation rate of NSCs, but rather the neuronal commitment of sNSCs, as sNSC-derived neurospheres contain more committed neuronal progenitor cells, even in the presence of epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF). These results suggest that the higher tendency for sNSCs to spontaneously differentiate into progenitor cells may explain the limited expansion of the secondary neural tube during embryonic development.

  2. Dysfunction of Platelet-derived Growth Factor Receptor α (PDGFRα) Represses the Production of Oligodendrocytes from Arylsulfatase A-deficient Multipotential Neural Precursor Cells*

    PubMed Central

    Pituch, Katarzyna C.; Moyano, Ana L.; Lopez-Rosas, Aurora; Marottoli, Felecia M.; Li, Guannan; Hu, Chenqi; van Breemen, Richard; Månsson, Jan E.; Givogri, Maria I.

    2015-01-01

    The membrane-bound receptor for platelet-derived growth factor A (PDGFRα) is crucial for controlling the production of oligodendrocytes (OLs) for myelination, but regulation of its activity during OL differentiation is largely unknown. We have examined the effect of increased sulfated content of galactosylceramides (sulfatides) on the regulation of PDGFRα in multipotential neural precursors (NPs) that are deficient in arylsulfatase A (ASA) activity. This enzyme is responsible for the lysosomal hydrolysis of sulfatides. We show that sulfatide accumulation significantly impacts the formation of OLs via deregulation of PDGFRα function. PDGFRα is less associated with detergent-resistant membranes in ASA-deficient cells and showed a significant decrease in AKT phosphorylation. Rescue experiments with ASA showed a normalization of the ratio of long versus short sulfatides, restored PDGFRα levels, corrected its localization to detergent-resistant membranes, increased AKT phosphorylation, and normalized the production of OLs in ASA-deficient NPs. Moreover, our studies identified a novel mechanism that regulates the secretion of PDGFRα in NPs, in glial cells, and in the brain cortex via exosomal shedding. Our study provides a first step in understanding the role of sulfatides in regulating PDGFRα levels in OLs and its impact in myelination. PMID:25605750

  3. Ventrally emigrating neural tube (VENT) cells: a second neural tube-derived cell population.

    PubMed

    Dickinson, Douglas P; Machnicki, Michal; Ali, Mohammed M; Zhang, Zhanying; Sohal, Gurkirpal S

    2004-08-01

    Two embryological fates for cells of the neural tube are well established. Cells from the dorsal part of the developing neural tube emigrate and become neural crest cells, which in turn contribute to the development of the peripheral nervous system and a variety of non-neural structures. Other neural tube cells form the neurons and glial cells of the central nervous system (CNS). This has led to the neural crest being treated as the sole neural tube-derived emigrating cell population, with the remaining neural tube cells assumed to be restricted to forming the CNS. However, this restriction has not been tested fully. Our investigations of chick, quail and duck embryos utilizing a variety of different labelling techniques (DiI, LacZ, GFP and quail chimera) demonstrate the existence of a second neural tube-derived emigrating cell population. These cells originate from the ventral part of the cranial neural tube, emigrate at the exit/entry site of the cranial nerves, migrate in association with the nerves and populate their target tissues. On the basis of its site of origin and route of migration we have named this cell population the ventrally emigrating neural tube (VENT) cells. VENT cells also differ from neural crest cells in that they emigrate considerably after the emigration of neural crest cells, and lack expression of the neural crest cell antigen HNK-1. VENT cells are multipotent, differentiating into cell types belonging to all four basic tissues in the body: the nerve, muscle, connective and epithelium. Thus, the neural tube provides at least two cell populations--neural crest and VENT cells--that contribute to the development of the peripheral nervous system and various non-neural structures. This review describes the origin of the idea of VENT cells, and discusses evidence for their existence and subsequent fates.

  4. EphrinA/EphA-induced ectodomain shedding of neural cell adhesion molecule regulates growth cone repulsion through ADAM10 metalloprotease.

    PubMed

    Brennaman, Leann H; Moss, Marcia L; Maness, Patricia F

    2014-01-01

    EphrinA/EphA-dependent axon repulsion is crucial for synaptic targeting in developing neurons but downstream molecular mechanisms remain obscure. Here, it is shown that ephrinA5/EphA3 triggers proteolysis of the neural cell adhesion molecule (NCAM) by the metalloprotease a disintegrin and metalloprotease (ADAM)10 to promote growth cone collapse in neurons from mouse neocortex. EphrinA5 induced ADAM10 activity to promote ectodomain shedding of polysialic acid-NCAM in cortical neuron cultures, releasing a ~ 250 kDa soluble fragment consisting of most of its extracellular region. NCAM shedding was dependent on ADAM10 and EphA3 kinase activity as shown in HEK293T cells transfected with dominant negative ADAM10 and kinase-inactive EphA3 (K653R) mutants. Purified ADAM10 cleaved NCAM at a sequence within the E-F loop of the second fibronectin type III domain (Leu(671) -Lys(672) /Ser(673) -Leu(674) ) identified by mass spectrometry. Mutations of NCAM within the ADAM10 cleavage sequence prevented EphA3-induced shedding of NCAM in HEK293T cells. EphrinA5-induced growth cone collapse was dependent on ADAM10 activity, was inhibited in cortical cultures from NCAM null mice, and was rescued by WT but not ADAM10 cleavage site mutants of NCAM. Regulated proteolysis of NCAM through the ephrin5/EphA3/ADAM10 mechanism likely impacts synapse development, and may lead to excess NCAM shedding when disrupted, as implicated in neurodevelopmental disorders such as schizophrenia. PSA-NCAM and ephrinA/EphA3 coordinately regulate inhibitory synapse development. Here, we have found that ephrinA5 stimulates EphA3 kinase and ADAM10 activity to promote PSA-NCAM cleavage at a site in its second FNIII repeat, which regulates ephrinA5-induced growth cone collapse in GABAergic and non-GABAergic neurons. These findings identify a new regulatory mechanism which may contribute to inhibitory connectivity.

  5. Enhanced expression of FNDC5 in human embryonic stem cell-derived neural cells along with relevant embryonic neural tissues.

    PubMed

    Ghahrizjani, Fatemeh Ahmadi; Ghaedi, Kamran; Salamian, Ahmad; Tanhaei, Somayeh; Nejati, Alireza Shoaraye; Salehi, Hossein; Nabiuni, Mohammad; Baharvand, Hossein; Nasr-Esfahani, Mohammad Hossein

    2015-02-25

    Availability of human embryonic stem cells (hESCs) has enhanced the capability of basic and clinical research in the context of human neural differentiation. Derivation of neural progenitor (NP) cells from hESCs facilitates the process of human embryonic development through the generation of neuronal subtypes. We have recently indicated that fibronectin type III domain containing 5 protein (FNDC5) expression is required for appropriate neural differentiation of mouse embryonic stem cells (mESCs). Bioinformatics analyses have shown the presence of three isoforms for human FNDC5 mRNA. To differentiate which isoform of FNDC5 is involved in the process of human neural differentiation, we have used hESCs as an in vitro model for neural differentiation by retinoic acid (RA) induction. The hESC line, Royan H5, was differentiated into a neural lineage in defined adherent culture treated by RA and basic fibroblast growth factor (bFGF). We collected all cell types that included hESCs, rosette structures, and neural cells in an attempt to assess the expression of FNDC5 isoforms. There was a contiguous increase in all three FNDC5 isoforms during the neural differentiation process. Furthermore, the highest level of expression of the isoforms was significantly observed in neural cells compared to hESCs and the rosette structures known as neural precursor cells (NPCs). High expression levels of FNDC5 in human fetal brain and spinal cord tissues have suggested the involvement of this gene in neural tube development. Additional research is necessary to determine the major function of FDNC5 in this process.

  6. Human Neural Cell-Based Biosensor

    DTIC Science & Technology

    2011-10-11

    mitochondrial health, reactive oxygen species generation and cell migration in our neural progenitor and differentiated neural cells. These assays...measure reactive oxygen species (ROS) generation in hNP1™ and hN2™ cells under conditions that induce oxidative stress, we are developing an assay

  7. Expression of chondrogenic potential of mouse trunk neural crest cells by FGF2 treatment.

    PubMed

    Ido, Atsushi; Ito, Kazuo

    2006-02-01

    There is a significant difference between the developmental patterns of cranial and trunk neural crest cells in the amniote. Thus, whereas cranial neural crest cells generate bone and cartilage, trunk neural crest cells do not contribute to skeletal derivatives. We examined whether mouse trunk neural crest cells can undergo chondrogenesis to analyze how the difference between the developmental patterns of cranial and trunk neural crest cells arises. Our present data demonstrate that mouse trunk neural crest cells have chondrogenic potential and that fibroblast growth factor (FGF) 2 is an inducing factor for their chondrogenesis in vitro. FGF2 altered the expression patterns of Hox9 genes and Id2, a cranial neural crest cell marker. These results suggest that environmental cues may play essential roles in generating the difference between developmental patterns of cranial and trunk neural crest cells. Copyright 2005 Wiley-Liss, Inc.

  8. Incremental evolution of the neural crest, neural crest cells and neural crest-derived skeletal tissues.

    PubMed

    Hall, Brian K; Gillis, J Andrew

    2013-01-01

    Urochordates (ascidians) have recently supplanted cephalochordates (amphioxus) as the extant sister taxon of vertebrates. Given that urochordates possess migratory cells that have been classified as 'neural crest-like'- and that cephalochordates lack such cells--this phylogenetic hypothesis may have significant implications with respect to the origin of the neural crest and neural crest-derived skeletal tissues in vertebrates. We present an overview of the genes and gene regulatory network associated with specification of the neural crest in vertebrates. We then use these molecular data--alongside cell behaviour, cell fate and embryonic context--to assess putative antecedents (latent homologues) of the neural crest or neural crest cells in ascidians and cephalochordates. Ascidian migratory mesenchymal cells--non-pigment-forming trunk lateral line cells and pigment-forming 'neural crest-like cells' (NCLC)--are unlikely latent neural crest cell homologues. Rather, Snail-expressing cells at the neural plate of border of urochordates and cephalochordates likely represent the extent of neural crest elaboration in non-vertebrate chordates. We also review evidence for the evolutionary origin of two neural crest-derived skeletal tissues--cartilage and dentine. Dentine is a bona fide vertebrate novelty, and dentine-secreting odontoblasts represent a cell type that is exclusively derived from the neural crest. Cartilage, on the other hand, likely has a much deeper origin within the Metazoa. The mesodermally derived cellular cartilages of some protostome invertebrates are much more similar to vertebrate cartilage than is the acellular 'cartilage-like' tissue in cephalochordate pharyngeal arches. Cartilage, therefore, is not a vertebrate novelty, and a well-developed chondrogenic program was most likely co-opted from mesoderm to the neural crest along the vertebrate stem. We conclude that the neural crest is a vertebrate novelty, but that neural crest cells and their

  9. Olfactory ensheathing cells: biology in neural development and regeneration.

    PubMed

    Su, Zhida; He, Cheng

    2010-12-01

    Olfactory ensheathing cells (OECs) constitute a unique population of glia that accompany and ensheath the primary olfactory axons. They are thought to be critical for spontaneous growth of olfactory axons within the developing and adult olfactory nervous system, and have recently emerged as potential candidates for cell-mediated repair of neural injuries. Here, based on the current research, we give an overview of the biology of OECs in neural development and regeneration. This review starts with a detailed description of the cellular and molecular biological properties of OECs. Their functions in olfactory neurogenesis, olfactory axonal growth and olfactory bulb formation are sequently discussed. We also describe therapeutic applications of OECs for the treatment of a variety of neural lesions, including spinal cord injury, stroke, degenerative diseases, and PNS injuries. Finally, we address issues that may foster a better understanding of OECs in neural development and regeneration. Copyright © 2010 Elsevier Ltd. All rights reserved.

  10. Incremental evolution of the neural crest, neural crest cells and neural crest-derived skeletal tissues

    PubMed Central

    Hall, Brian K; Gillis, J Andrew

    2013-01-01

    Urochordates (ascidians) have recently supplanted cephalochordates (amphioxus) as the extant sister taxon of vertebrates. Given that urochordates possess migratory cells that have been classified as ‘neural crest-like’– and that cephalochordates lack such cells – this phylogenetic hypothesis may have significant implications with respect to the origin of the neural crest and neural crest-derived skeletal tissues in vertebrates. We present an overview of the genes and gene regulatory network associated with specification of the neural crest in vertebrates. We then use these molecular data – alongside cell behaviour, cell fate and embryonic context – to assess putative antecedents (latent homologues) of the neural crest or neural crest cells in ascidians and cephalochordates. Ascidian migratory mesenchymal cells – non-pigment-forming trunk lateral line cells and pigment-forming ‘neural crest-like cells’ (NCLC) – are unlikely latent neural crest cell homologues. Rather, Snail-expressing cells at the neural plate of border of urochordates and cephalochordates likely represent the extent of neural crest elaboration in non-vertebrate chordates. We also review evidence for the evolutionary origin of two neural crest-derived skeletal tissues – cartilage and dentine. Dentine is a bona fide vertebrate novelty, and dentine-secreting odontoblasts represent a cell type that is exclusively derived from the neural crest. Cartilage, on the other hand, likely has a much deeper origin within the Metazoa. The mesodermally derived cellular cartilages of some protostome invertebrates are much more similar to vertebrate cartilage than is the acellular ‘cartilage-like’ tissue in cephalochordate pharyngeal arches. Cartilage, therefore, is not a vertebrate novelty, and a well-developed chondrogenic program was most likely co-opted from mesoderm to the neural crest along the vertebrate stem. We conclude that the neural crest is a vertebrate novelty, but that neural

  11. Plant Growth Models Using Artificial Neural Networks

    NASA Technical Reports Server (NTRS)

    Bubenheim, David

    1997-01-01

    In this paper, we descrive our motivation and approach to devloping models and the neural network architecture. Initial use of the artificial neural network for modeling the single plant process of transpiration is presented.

  12. Plant Growth Models Using Artificial Neural Networks

    NASA Technical Reports Server (NTRS)

    Bubenheim, David

    1997-01-01

    In this paper, we descrive our motivation and approach to devloping models and the neural network architecture. Initial use of the artificial neural network for modeling the single plant process of transpiration is presented.

  13. Two outward potassium current types are expressed during the neural differentiation of neural stem cells

    PubMed Central

    Bai, Ruiying; Gao, Guowei; Xing, Ying; Xue, Hong

    2013-01-01

    The electrophysiological properties of potassium ion channels are regarded as a basic index for determining the functional differentiation of neural stem cells. In this study, neural stem cells from the hippocampus of newborn rats were induced to differentiate with neurotrophic growth factor, and the electrophysiological properties of the voltage-gated potassium ion channels were observed. Immunofluorescence staining showed that the rapidly proliferating neural stem cells formed spheres in vitro that expressed high levels of nestin. The differentiated neurons were shown to express neuron-specific enolase. Flow cytometric analysis revealed that the neural stem cells were actively dividing and the percentage of cells in the S + G2/M phase was high. However, the ratio of cells in the S + G2/M phase decreased obviously as differentiation proceeded. Whole-cell patch-clamp recordings revealed apparent changes in potassium ion currents as the neurons differentiated. The potassium ion currents consisted of one transient outward potassium ion current and one delayed rectifier potassium ion current, which were blocked by 4-aminopyridine and tetraethylammonium, respectively. The experimental findings indicate that neural stem cells from newborn rat campus could be cultured and induced to differentiate into functional neurons under defined conditions in vitro. The differentiated neurons expressed two types of outward potassium ion currents similar to those of mature neurons in vivo. PMID:25206577

  14. Perinatal applications of neural stem cells.

    PubMed

    Kennea, Nigel L; Mehmet, Huseyin

    2004-12-01

    The brain, unlike many tissues, has a limited capacity for self-repair and so there has been great interest in the possibility of transplanting neural cells to replace those lost through injury or disease. Encouraging research in humans is already underway examining the possibility of neural cell replacement in adult neurodegenerative conditions such as Parkinson's disease and Huntington disease. In addition, experiments exploring neural stem cell replacement in rodent models of acute stroke, demyelination and spinal cord injury have demonstrated functional improvements in treated animals. When considering perinatal neural stem cell therapy, it should not be overlooked that the immature, developing brain might provide a more favourable environment for stem cell integration. However, considerable advances need to be made both in understanding the basic biology of neural stem cells, including the instructive signals that determine their proliferation and differentiation, and in characterising their responses when transplanted in a damaged or diseased area of the brain.

  15. Microfluidic systems for stem cell-based neural tissue engineering.

    PubMed

    Karimi, Mahdi; Bahrami, Sajad; Mirshekari, Hamed; Basri, Seyed Masoud Moosavi; Nik, Amirala Bakhshian; Aref, Amir R; Akbari, Mohsen; Hamblin, Michael R

    2016-07-05

    Neural tissue engineering aims at developing novel approaches for the treatment of diseases of the nervous system, by providing a permissive environment for the growth and differentiation of neural cells. Three-dimensional (3D) cell culture systems provide a closer biomimetic environment, and promote better cell differentiation and improved cell function, than could be achieved by conventional two-dimensional (2D) culture systems. With the recent advances in the discovery and introduction of different types of stem cells for tissue engineering, microfluidic platforms have provided an improved microenvironment for the 3D-culture of stem cells. Microfluidic systems can provide more precise control over the spatiotemporal distribution of chemical and physical cues at the cellular level compared to traditional systems. Various microsystems have been designed and fabricated for the purpose of neural tissue engineering. Enhanced neural migration and differentiation, and monitoring of these processes, as well as understanding the behavior of stem cells and their microenvironment have been obtained through application of different microfluidic-based stem cell culture and tissue engineering techniques. As the technology advances it may be possible to construct a "brain-on-a-chip". In this review, we describe the basics of stem cells and tissue engineering as well as microfluidics-based tissue engineering approaches. We review recent testing of various microfluidic approaches for stem cell-based neural tissue engineering.

  16. Compensatory adrenal growth - A neurally mediated reflex

    NASA Technical Reports Server (NTRS)

    Dallman, M. F.; Engeland, W. C.; Shinsako, J.

    1976-01-01

    The responses of young rats to left adrenalectomy or left adrenal manipulation were compared to surgical sham adrenalectomy in which adrenals were observed but not touched. At 12 h right adrenal wet weight, dry weight, DNA, RNA, and protein content were increased (P less than 0.05) after the first two operations. Left adrenal manipulation resulted in increased right adrenal weight at 12 h but no change in left adrenal weight. Sequential manipulation of the left adrenal at time 0 and the right adrenal at 12 h resulted in an enlarged right adrenal at 12 h (P less than 0.01), and an enlarged left adrenal at 24 h (P less than 0.05), showing that the manipulated gland was capable of response. Bilateral adrenal manipulation of the adrenal glands resulted in bilateral enlargement of 12 h (P less than 0.01). Taken together with previous results, these findings strongly suggest that compensatory adrenal growth is a neurally mediated reflex.

  17. Human neural stem cells improve cognition and promote synaptic growth in two complementary transgenic models of Alzheimer's disease and neuronal loss

    PubMed Central

    Ager, Rahasson R.; Davis, Joy L.; Agazaryan, Andy; Benavente, Francisca; Poon, Wayne W.; LaFerla, Frank M.

    2015-01-01

    ABSTRACT Alzheimer's disease (AD) is the most prevalent age‐related neurodegenerative disorder, affecting over 35 million people worldwide. Pathologically, AD is characterized by the progressive accumulation of β‐amyloid (Aβ) plaques and neurofibrillary tangles within the brain. Together, these pathologies lead to marked neuronal and synaptic loss and corresponding impairments in cognition. Current treatments, and recent clinical trials, have failed to modify the clinical course of AD; thus, the development of novel and innovative therapies is urgently needed. Over the last decade, the potential use of stem cells to treat cognitive impairment has received growing attention. Specifically, neural stem cell transplantation as a treatment for AD offers a novel approach with tremendous therapeutic potential. We previously reported that intrahippocampal transplantation of murine neural stem cells (mNSCs) can enhance synaptogenesis and improve cognition in 3xTg‐AD mice and the CaM/Tet‐DTA model of hippocampal neuronal loss. These promising findings prompted us to examine a human neural stem cell population, HuCNS‐SC, which has already been clinically tested for other neurodegenerative disorders. In this study, we provide the first evidence that transplantation of research grade HuCNS‐SCs can improve cognition in two complementary models of neurodegeneration. We also demonstrate that HuCNS‐SC cells can migrate and differentiate into immature neurons and glia and significantly increase synaptic and growth‐associated markers in both 3xTg‐AD and CaM/Tet‐DTA mice. Interestingly, improvements in aged 3xTg‐AD mice were not associated with altered Aβ or tau pathology. Rather, our findings suggest that human NSC transplantation improves cognition by enhancing endogenous synaptogenesis. Taken together, our data provide the first preclinical evidence that human NSC transplantation could be a safe and effective therapeutic approach for treating AD. © 2014

  18. Monitoring the Differentiation and Migration Patterns of Neural Cells Derived from Human Embryonic Stem Cells Using a Microfluidic Culture System

    PubMed Central

    Lee, Nayeon; Park, Jae Woo; Kim, Hyung Joon; Yeon, Ju Hun; Kwon, Jihye; Ko, Jung Jae; Oh, Seung-Hun; Kim, Hyun Sook; Kim, Aeri; Han, Baek Soo; Lee, Sang Chul; Jeon, Noo Li; Song, Jihwan

    2014-01-01

    Microfluidics can provide unique experimental tools to visualize the development of neural structures within a microscale device, which is followed by guidance of neurite growth in the axonal isolation compartment. We utilized microfluidics technology to monitor the differentiation and migration of neural cells derived from human embryonic stem cells (hESCs). We co-cultured hESCs with PA6 stromal cells, and isolated neural rosette-like structures, which subsequently formed neurospheres in suspension culture. Tuj1-positive neural cells, but not nestin-positive neural precursor cells (NPCs), were able to enter the microfluidics grooves (microchannels), suggesting that neural cell-migratory capacity was dependent upon neuronal differentiation stage. We also showed that bundles of axons formed and extended into the microchannels. Taken together, these results demonstrated that microfluidics technology can provide useful tools to study neurite outgrowth and axon guidance of neural cells, which are derived from human embryonic stem cells. PMID:24938227

  19. Monitoring the differentiation and migration patterns of neural cells derived from human embryonic stem cells using a microfluidic culture system.

    PubMed

    Lee, Nayeon; Park, Jae Woo; Kim, Hyung Joon; Yeon, Ju Hun; Kwon, Jihye; Ko, Jung Jae; Oh, Seung-Hun; Kim, Hyun Sook; Kim, Aeri; Han, Baek Soo; Lee, Sang Chul; Jeon, Noo Li; Song, Jihwan

    2014-06-01

    Microfluidics can provide unique experimental tools to visualize the development of neural structures within a microscale device, which is followed by guidance of neurite growth in the axonal isolation compartment. We utilized microfluidics technology to monitor the differentiation and migration of neural cells derived from human embryonic stem cells (hESCs). We co-cultured hESCs with PA6 stromal cells, and isolated neural rosette-like structures, which subsequently formed neurospheres in suspension culture. Tuj1-positive neural cells, but not nestin-positive neural precursor cells (NPCs), were able to enter the microfluidics grooves (microchannels), suggesting that neural cell-migratory capacity was dependent upon neuronal differentiation stage. We also showed that bundles of axons formed and extended into the microchannels. Taken together, these results demonstrated that microfluidics technology can provide useful tools to study neurite outgrowth and axon guidance of neural cells, which are derived from human embryonic stem cells.

  20. Neural stem cells and stroke.

    PubMed

    Ding, Dah-Ching; Lin, Chen-Huan; Shyu, Woei-Cherng; Lin, Shinn-Zong

    2013-01-01

    Acute ischemic stroke causes a disturbance of neuronal circuitry and disruption of the blood-brain barrier that can lead to functional disabilities. At present, thrombolytic therapy inducing recanalization of the occluded vessels in the cerebral infarcted area is a commonly used therapeutic strategy. However, only a minority of patients have timely access to this kind of therapy. Recently, neural stem cells (NSCs) as therapy for stroke have been developed in preclinical studies. NSCs are harbored in the subventricular zone (SVZ) as well as the subgranular zone of the brain. The microenvironment in the SVZ, including intercellular interactions, extracellular matrix proteins, and soluble factors, can promote NSC proliferation, self-renewal, and multipotency. Endogenous neurogenesis responds to insults of ischemic stroke supporting the existence of remarkable plasticity in the mammalian brain. Homing and integration of NSCs to the sites of damaged brain tissue are complex morphological and physiological processes. This review provides an update on current preclinical cell therapies for stroke, focusing on neurogenesis in the SVZ and dentate gyrus and on recruitment cues that promote NSC homing and integration to the site of the damaged brain.

  1. Generalized Potential of Adult Neural Stem Cells

    NASA Astrophysics Data System (ADS)

    Clarke, Diana L.; Johansson, Clas B.; Wilbertz, Johannes; Veress, Biborka; Nilsson, Erik; Karlström, Helena; Lendahl, Urban; Frisén, Jonas

    2000-06-01

    The differentiation potential of stem cells in tissues of the adult has been thought to be limited to cell lineages present in the organ from which they were derived, but there is evidence that some stem cells may have a broader differentiation repertoire. We show here that neural stem cells from the adult mouse brain can contribute to the formation of chimeric chick and mouse embryos and give rise to cells of all germ layers. This demonstrates that an adult neural stem cell has a very broad developmental capacity and may potentially be used to generate a variety of cell types for transplantation in different diseases.

  2. Function of Mouse Embryonic Stem Cell-Derived Supporting Cells in Neural Progenitor Cell Maturation and Long Term Cxpansion

    PubMed Central

    Guan, Yunqian; Du, Qing-An; Zhu, Wanwan; Zou, Chunlin; Wu, Di; Chen, Ling; Zhang, Yu Alex

    2013-01-01

    Background In the differentiation of mouse embryonic stem (ES) cells into neurons using the 5-stage method, cells in stage 4 are in general used as neural progenitors (NPs) because of their ability to give rise to neurons. The choice of stage 4 raises several questions about neural progenitors such as the type of cell types that are specifically considered to be neural progenitors, the exact time when these progenitors become capable of neurogenesis and whether neurogenesis is an independent and autonomous process or the result of an interaction between NP cells and the surrounding cells. Methodology/Principal Findings In this study, we found that the confluent monolayer cells and neural sphere like cell clusters both appeared in the culture of the first 14 days and the subsequent 6 weeks. However, only the sphere cells are neural progenitors that give rise to neurons and astrocytes. The NP cells require 14 days to mature into neural lineages fully capable of differentiation. We also found that although the confluent monolayer cells do not undergo neurogenesis, they play a crucial role in the growth, differentiation, and apoptosis of the sphere cells, during the first 14 days and long term culture, by secreted factors and direct cell to cell contact. Conclusions/Significance The sphere cells in stage 4 are more committed to developing into neural progenitors than monolayer cells. Interaction between the monolayer cells and sphere cells is important in the development of stage 4 cell characteristics. PMID:23342136

  3. Capacity of Human Dental Follicle Cells to Differentiate into Neural Cells In Vitro

    PubMed Central

    Ogura, Naomi; Takahashi, Kosuke; Ito, Ko; Suemitsu, Masaaki; Kuyama, Kayo

    2017-01-01

    The dental follicle is an ectomesenchymal tissue surrounding the developing tooth germ. Human dental follicle cells (hDFCs) have the capacity to commit to differentiation into multiple cell types. Here we investigated the capacity of hDFCs to differentiate into neural cells and the efficiency of a two-step strategy involving floating neurosphere-like bodies for neural differentiation. Undifferentiated hDFCs showed a spindle-like morphology and were positive for neural markers such as nestin, β-III-tubulin, and S100β. The cellular morphology of several cells was neuronal-like including branched dendrite-like processes and neurites. Next, hDFCs were used for neurosphere formation in serum-free medium containing basic fibroblast growth factor, epidermal growth factor, and B27 supplement. The number of cells with neuronal-like morphology and that were strongly positive for neural markers increased with sphere formation. Gene expression of neural markers also increased in hDFCs with sphere formation. Next, gene expression of neural markers was examined in hDFCs during neuronal differentiation after sphere formation. Expression of Musashi-1 and Musashi-2, MAP2, GFAP, MBP, and SOX10 was upregulated in hDFCs undergoing neuronal differentiation via neurospheres, whereas expression of nestin and β-III-tubulin was downregulated. In conclusion, hDFCs may be another optimal source of neural/glial cells for cell-based therapies to treat neurological diseases. PMID:28261273

  4. Engraftment and Differentiation of Embryonic Stem Cell–Derived Neural Progenitor Cells in the Cochlear Nerve Trunk: Growth of Processes into the Organ of Corti

    PubMed Central

    Corrales, C. Eduardo; Pan, Luying; Li, Huawei; Liberman, M. Charles; Heller, Stefan; Edge, Albert S.B.

    2007-01-01

    Hearing loss in mammals is irreversible because cochlear neurons and hair cells do not regenerate. To determine whether we could replace neurons lost to primary neuronal degeneration, we injected EYFP-expressing embryonic stem cell–derived mouse neural progenitor cells into the cochlear nerve trunk in immunosuppressed animals 1 week after destroying the cochlear nerve (spiral ganglion) cells while leaving hair cells intact by ouabain application to the round window at the base of the cochlea in gerbils. At 3 days post transplantation, small grafts were seen that expressed endogenous EYFP and could be immunolabeled for neuron-specific markers. Twelve days after transplantation, the grafts had neurons that extended processes from the nerve core toward the denervated organ of Corti. By 64–98 days, the grafts had sent out abundant processes that occupied a significant portion of the space formerly occupied by the cochlear nerve. The neurites grew in fasciculating bundles projecting through Rosenthal’s canal, the former site of spiral ganglion cells, into the osseous spiral lamina and ultimately into the organ of Corti, where they contacted hair cells. Neuronal counts showed a significant increase in neuronal processes near the sensory epithelium, compared to animals that were denervated without subsequent stem cell transplantation. The regeneration of these neurons shows that neurons differentiated from stem cells have the capacity to grow to a specific target in an animal model of neuronal degeneration. PMID:17013931

  5. Human Neural Cell-Based Biosensor

    DTIC Science & Technology

    2013-05-28

    format (96-,384-well) assays, 2) grow as adherent monolayers, and 3) possess a stable karyotype for multiple (>10) passages with a doubling time of ~36...derived neural progenitor cell line working stock has been amplified, characterized for karyotype and evaluated for the expression of neural progenitor...Orlando R, Stice SL. Membrane proteomic signatures of karyotypically normal and abnormal human embryonic stem cell lines and derivatives. Proteomics. 2011

  6. Neural and oligodendrocyte progenitor cells: transferrin effects on cell proliferation

    PubMed Central

    Silvestroff, Lucas; Franco, Paula Gabriela; Pasquini, Juana María

    2013-01-01

    NSC (neural stem cells)/NPC (neural progenitor cells) are multipotent and self-renew throughout adulthood in the SVZ (subventricular zone) of the mammalian CNS (central nervous system). These cells are considered interesting targets for CNS neurodegenerative disorder cell therapies, and understanding their behaviour in vitro is crucial if they are to be cultured prior to transplantation. We cultured the SVZ tissue belonging to newborn rats under the form of NS (neurospheres) to evaluate the effects of Tf (transferrin) on cell proliferation. The NS were heterogeneous in terms of the NSC/NPC markers GFAP (glial fibrillary acidic protein), Nestin and Sox2 and the OL (oligodendrocyte) progenitor markers NG2 (nerve/glia antigen 2) and PDGFRα (platelet-derived growth factor receptor α). The results of this study indicate that aTf (apoTransferrin) is able to increase cell proliferation of SVZ-derived cells in vitro, and that these effects were mediated at least in part by the TfRc1 (Tf receptor 1). Since OPCs (oligodendrocyte progenitor cells) represent a significant proportion of the proliferating cells in the SVZ-derived primary cultures, we used the immature OL cell line N20.1 to show that Tf was able to augment the proliferation rate of OPC, either by adding aTf to the culture medium or by overexpressing rat Tf in situ. The culture medium supplemented with ferric iron, together with aTf, increased the DNA content, while ferrous iron did not. The present work provides data that could have a potential application in human cell replacement therapies for neurodegenerative disease and/or CNS injury that require the use of in vitro amplified NPCs. PMID:23368675

  7. Hypoxic preconditioning enhances neural stem cell transplantation therapy after intracerebral hemorrhage in mice.

    PubMed

    Wakai, Takuma; Narasimhan, Purnima; Sakata, Hiroyuki; Wang, Eric; Yoshioka, Hideyuki; Kinouchi, Hiroyuki; Chan, Pak H

    2016-12-01

    Previous studies have shown that intraparenchymal transplantation of neural stem cells ameliorates neurological deficits in animals with intracerebral hemorrhage. However, hemoglobin in the host brain environment causes massive grafted cell death and reduces the effectiveness of this approach. Several studies have shown that preconditioning induced by sublethal hypoxia can markedly improve the tolerance of treated subjects to more severe insults. Therefore, we investigated whether hypoxic preconditioning enhances neural stem cell resilience to the hemorrhagic stroke environment and improves therapeutic effects in mice. To assess whether hypoxic preconditioning enhances neural stem cell survival when exposed to hemoglobin, neural stem cells were exposed to 5% hypoxia for 24 hours before exposure to hemoglobin. To study the effectiveness of hypoxic preconditioning on grafted-neural stem cell recovery, neural stem cells subjected to hypoxic preconditioning were grafted into the parenchyma 3 days after intracerebral hemorrhage. Hypoxic preconditioning significantly enhanced viability of the neural stem cells exposed to hemoglobin and increased grafted-cell survival in the intracerebral hemorrhage brain. Hypoxic preconditioning also increased neural stem cell secretion of vascular endothelial growth factor. Finally, transplanted neural stem cells with hypoxic preconditioning exhibited enhanced tissue-protective capability that accelerated behavioral recovery. Our results suggest that hypoxic preconditioning in neural stem cells improves efficacy of stem cell therapy for intracerebral hemorrhage.

  8. Applicability of tooth derived stem cells in neural regeneration

    PubMed Central

    Parisi, Ludovica; Manfredi, Edoardo

    2016-01-01

    Within the nervous system, regeneration is limited, and this is due to the small amount of neural stem cells, the inhibitory origin of the stem cell niche and often to the development of a scar which constitutes a mechanical barrier for the regeneration. Regarding these aspects, many efforts have been done in the research of a cell component that combined with scaffolds and growth factors could be suitable for nervous regeneration in regenerative medicine approaches. Autologous mesenchymal stem cells represent nowadays the ideal candidate for this aim, thank to their multipotency and to their amount inside adult tissues. However, issues in their harvesting, through the use of invasive techniques, and problems involved in their ageing, require the research of new autologous sources. To this purpose, the recent discovery of a stem cells component in teeth, and which derive from neural crest cells, has came to the light the possibility of using dental stem cells in nervous system regeneration. In this work, in order to give guidelines on the use of dental stem cells for neural regeneration, we briefly introduce the concepts of regeneration and regenerative medicine, we then focus the attention on odontogenesis, which involves the formation and the presence of a stem component in different parts of teeth, and finally we describe some experimental approaches which are exploiting dental stem cells for neural studies. PMID:28123398

  9. Comparison of Gompertz and neural network models of broiler growth.

    PubMed

    Roush, W B; Dozier, W A; Branton, S L

    2006-04-01

    Neural networks offer an alternative to regression analysis for biological growth modeling. Very little research has been conducted to model animal growth using artificial neural networks. Twenty-five male chicks (Ross x Ross 308) were raised in an environmental chamber. Body weights were determined daily and feed and water were provided ad libitum. The birds were fed a starter diet (23% CP and 3,200 kcal of ME/kg) from 0 to 21 d, and a grower diet (20% CP and 3,200 kcal of ME/ kg) from 22 to 70 d. Dead and female birds were not included in the study. Average BW of 18 birds were used as the data points for the growth curve to be modeled. Training data consisted of alternate-day weights starting with the first day. Validation data consisted of BW at all other age periods. Comparison was made between the modeling by the Gompertz nonlinear regression equation and neural network modeling. Neural network models were developed with the Neuroshell Predictor. Accuracy of the models was determined by mean square error (MSE), mean absolute deviation (MAD), mean absolute percentage error (MAPE), and bias. The Gompertz equation was fit for the data. Forecasting error measurements were based on the difference between the model and the observed values. For the training data, the lowest MSE, MAD, MAPE, and bias were noted for the neural-developed neural network. For the validation data, the lowest MSE and MAD were noted with the genetic algorithm-developed neural network. Lowest bias was for the neural-developed network. As measured by bias, the Gompertz equation underestimated the values whereas the neural- and genetic-developed neural networks produced little or no overestimation of the observed BW responses. Past studies have attempted to interpret the biological significance of the estimates of the parameters of an equation. However, it may be more practical to ignore the relevance of parameter estimates and focus on the ability to predict responses.

  10. Successful elimination of non-neural cells and unachievable elimination of glial cells by means of commonly used cell culture manipulations during differentiation of GFAP and SOX2 positive neural progenitors (NHA) to neuronal cells.

    PubMed

    Witusik, Monika; Piaskowski, Sylwester; Hulas-Bigoszewska, Krystyna; Zakrzewska, Magdalena; Gresner, Sylwia M; Azizi, S Ausim; Krynska, Barbara; Liberski, Pawel P; Rieske, Piotr

    2008-07-19

    Although extensive research has been performed to control differentiation of neural stem cells - still, the response of those cells to diverse cell culture conditions often appears to be random and difficult to predict. To this end, we strived to obtain stabilized protocol of NHA cells differentiation - allowing for an increase in percentage yield of neuronal cells. Uncommitted GFAP and SOX2 positive neural progenitors - so-called, Normal Human Astrocytes (NHA) were differentiated in different environmental conditions to: only neural cells consisted of neuronal [MAP2+, GFAP-] and glial [GFAP+, MAP2-] population, non-neural cells [CD44+, VIMENTIN+, FIBRONECTIN+, MAP2-, GFAP-, S100beta-, SOX2-], or mixture of neural and non-neural cells.In spite of successfully increasing the percentage yield of glial and neuronal vs. non-neural cells by means of environmental changes, we were not able to increase significantly the percentage of neuronal (GABA-ergic and catecholaminergic) over glial cells under several different cell culture testing conditions. Supplementing serum-free medium with several growth factors (SHH, bFGF, GDNF) did not radically change the ratio between neuronal and glial cells--i.e., 1,1:1 in medium without growth factors and 1,4:1 in medium with GDNF, respectively. We suggest that biotechnologists attempting to enrich in vitro neural cell cultures in one type of cells - such as that required for transplantology purposes, should consider the strong limiting influence of intrinsic factors upon extracellular factors commonly tested in cell culture conditions.

  11. [Effects of endothelial cells on renewal and differentiation of neural stem cells].

    PubMed

    Dong, Zhiwu; Su, Le; Mino, Junying

    2007-10-01

    It is well established that neural stem cells (NSCs) are not randomly distributed throughout the brain, but rather are concentrated around blood vessels. Although NSCs lie in a vascular niche, there is no direct evidence for a functional relationship between the NSCs and blood vessel component cells. It is reported that endothelial cells release soluble factors that stimulate the self-renewal of NSCs, inhibit their differentiation, and enhance their neuron production. Endothelial coculture can activate Notch to promote self-renewal. Furthermore, vascular endothelial growth factor (VEGF) plays a significant role in neural cells; it stimulates the growth and differentiation of astrocytes in the central nervous system (CNS). Therefore, beyond their traditional role as structural components of blood vessels, endothelial cells are not only critical component of the neural stem cell niche, but they also are able to enhance neurogenesis, possibly through the secretion of brain-derived neurotrophic factor.

  12. Analysis of Neural Stem Cells from Human Cortical Brain Structures In Vitro.

    PubMed

    Aleksandrova, M A; Poltavtseva, R A; Marei, M V; Sukhikh, G T

    2016-05-01

    Comparative immunohistochemical analysis of the neocortex from human fetuses showed that neural stem and progenitor cells are present in the brain throughout the gestation period, at least from week 8 through 26. At the same time, neural stem cells from the first and second trimester fetuses differed by the distribution, morphology, growth, and quantity. Immunocytochemical analysis of neural stem cells derived from fetuses at different gestation terms and cultured under different conditions showed their differentiation capacity. Detailed analysis of neural stem cell populations derived from fetuses on gestation weeks 8-9, 18-20, and 26 expressing Lex/SSEA1 was performed.

  13. Chondroitinase ABC combined with neural stem/progenitor cell transplantation enhances graft cell migration and outgrowth of growth-associated protein-43-positive fibers after rat spinal cord injury.

    PubMed

    Ikegami, Takeshi; Nakamura, Masaya; Yamane, Junichi; Katoh, Hiroyuki; Okada, Seiji; Iwanami, Akio; Watanabe, Kota; Ishii, Ken; Kato, Fumikazu; Fujita, Hiroshi; Takahashi, Toyomi; Okano, Hirotaka James; Toyama, Yoshiaki; Okano, Hideyuki

    2005-12-01

    We previously reported that the transplantation of neural stem/progenitor cells (NSPCs) can contribute to the repair of injured spinal cord in adult rats and monkeys. In some cases, however, most of the transplanted cells adhered to the cavity wall and failed to migrate and integrate into the host spinal cord. In this study we focused on chondroitin sulfate proteoglycan (CSPG), a known constituent of glial scars that is strongly expressed after spinal cord injury (SCI), as a putative inhibitor of NSPC migration in vivo. We hypothesized that the digestion of CSPG by chondroitinase ABC (C-ABC) might promote the migration of transplanted cells and neurite outgrowth after SCI. An in vitro study revealed that the migration of NSPC-derived cells was inhibited by CSPG and that this inhibitory effect was attenuated by C-ABC pre-treatment. Consistently, an in vivo study of C-ABC treatment combined with NSPC transplantation into injured spinal cord revealed that C-ABC pre-treatment promoted the migration of the transplanted cells, whereas CSPG-immunopositive scar tissue around the lesion cavity prevented their migration into the host spinal cord in the absence of C-ABC pre-treatment. Furthermore, this combined treatment significantly induced the outgrowth of a greater number of growth-associated protein-43-positive fibers at the lesion epicentre, compared with NSPC transplantation alone. These findings suggested that the application of C-ABC enhanced the benefits of NSPC transplantation for SCI by reducing the inhibitory effects of the glial scar, indicating that this combined treatment may be a promising strategy for the regeneration of injured spinal cord.

  14. A brief perspective on neural cell therapy.

    PubMed

    Pruszak, Jan

    2014-01-01

    For a range of nervous system disorders current treatment options remain limited. Focusing on Parkinson's disease as a neurodegenerative entity that affects an increasing quantity of people in our aging societies, we briefly discuss remaining challenges and opportunities that neural stem cell therapy might be able to offer. Providing a snapshot of neural transplantation paradigms, we contemplate possible imminent translational scenarios and discuss critical requirements to be considered before clinical implementation.

  15. VLSI Cells Placement Using the Neural Networks

    SciTech Connect

    Azizi, Hacene; Zouaoui, Lamri; Mokhnache, Salah

    2008-06-12

    The artificial neural networks have been studied for several years. Their effectiveness makes it possible to expect high performances. The privileged fields of these techniques remain the recognition and classification. Various applications of optimization are also studied under the angle of the artificial neural networks. They make it possible to apply distributed heuristic algorithms. In this article, a solution to placement problem of the various cells at the time of the realization of an integrated circuit is proposed by using the KOHONEN network.

  16. Overexpression of MCT8 enhances the differentiation of ES cells into neural progenitors.

    PubMed

    Sugiura, Mika; Nagaoka, Masato; Yabuuchi, Hikaru; Akaike, Toshihiro

    2007-09-07

    Embryonic stem (ES) cell differentiation is regulated by cytokines and growth factors, as well as small-compound chemicals incorporated into cells by transporter proteins. Little is known regarding the effect of transporters on ES cell differentiation. This study focused on the effect of transporters during the neural-lineage differentiation of ES cells. Among the 27 types of SLC family transporters, MCT8 expression was coincident with that of neural stem cell markers, and the overexpression of MCT8 accelerated the differentiation into neural cells. These results suggested that the transporters and their substrates also play a crucial role in the regulation of ES cell differentiation.

  17. Viral susceptibility, transfection and growth of SPB--a fish neural progenitor cell line from the brain of snubnose pompano, Trachinotus blochii (Lacépède).

    PubMed

    Wen, C-M; Ku, C-C; Wang, C-S

    2013-07-01

    This study investigates the susceptibilities of the SPB cell line to fish viruses including giant seaperch iridovirus (GSIV-K1), red sea bream iridovirus (RSIV-Ku), grouper nervous necrosis virus (GNNV-K1), chum salmon reovirus (CSV) and eel herpesvirus (HVA). GSIV-K1, RSIV-Ku and CSV replicated well in SPB cells, with a significant cytopathic effect and virus production. However, the cells were HVA and GNNV refractory. To examine the ability of SPB cells to stably express foreign protein, expression vectors encoding GNNV B1 and B2 fused to enhanced green fluorescent protein (EGFP) and GSIV ORF35L fused to DsRed were constructed and introduced by transfection into SPB cells. Stable transfectants displayed different morphologies compared with SPB and with each other. EGFP-B1 was predominantly localized in the nuclei, EFPF-B2 was distributed throughout the cytoplasm and nucleus, and granular 35L-DsRed was localized with secreted vesicles. The expression of EFPF-B2 in SPB cells produced blebs on the surface, but the cells showing stable expression of EGFP, EGFP-B1 or 35L-DsRed showed normal morphologies. Results show the SPB cells and the transfected cells grow well at temperatures between 20 and 35 °C and with serum-dependent growth. SPB cells are suitable for studies on foreign protein expression and virology.

  18. ALK5-mediated transforming growth factor β signaling in neural crest cells controls craniofacial muscle development via tissue-tissue interactions.

    PubMed

    Han, Arum; Zhao, Hu; Li, Jingyuan; Pelikan, Richard; Chai, Yang

    2014-08-01

    The development of the craniofacial muscles requires reciprocal interactions with surrounding craniofacial tissues that originate from cranial neural crest cells (CNCCs). However, the molecular mechanism involved in the tissue-tissue interactions between CNCCs and muscle progenitors during craniofacial muscle development is largely unknown. In the current study, we address how CNCCs regulate the development of the tongue and other craniofacial muscles using Wnt1-Cre; Alk5(fl/fl) mice, in which loss of Alk5 in CNCCs results in severely disrupted muscle formation. We found that Bmp4 is responsible for reduced proliferation of the myogenic progenitor cells in Wnt1-Cre; Alk5(fl/fl) mice during early myogenesis. In addition, Fgf4 and Fgf6 ligands were reduced in Wnt1-Cre; Alk5(fl/fl) mice and are critical for differentiation of the myogenic cells. Addition of Bmp4 or Fgf ligands rescues the proliferation and differentiation defects in the craniofacial muscles of Alk5 mutant mice in vitro. Taken together, our results indicate that CNCCs play critical roles in controlling craniofacial myogenic proliferation and differentiation through tissue-tissue interactions. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  19. ALK5-Mediated Transforming Growth Factor β Signaling in Neural Crest Cells Controls Craniofacial Muscle Development via Tissue-Tissue Interactions

    PubMed Central

    Han, Arum; Zhao, Hu; Li, Jingyuan; Pelikan, Richard

    2014-01-01

    The development of the craniofacial muscles requires reciprocal interactions with surrounding craniofacial tissues that originate from cranial neural crest cells (CNCCs). However, the molecular mechanism involved in the tissue-tissue interactions between CNCCs and muscle progenitors during craniofacial muscle development is largely unknown. In the current study, we address how CNCCs regulate the development of the tongue and other craniofacial muscles using Wnt1-Cre; Alk5fl/fl mice, in which loss of Alk5 in CNCCs results in severely disrupted muscle formation. We found that Bmp4 is responsible for reduced proliferation of the myogenic progenitor cells in Wnt1-Cre; Alk5fl/fl mice during early myogenesis. In addition, Fgf4 and Fgf6 ligands were reduced in Wnt1-Cre; Alk5fl/fl mice and are critical for differentiation of the myogenic cells. Addition of Bmp4 or Fgf ligands rescues the proliferation and differentiation defects in the craniofacial muscles of Alk5 mutant mice in vitro. Taken together, our results indicate that CNCCs play critical roles in controlling craniofacial myogenic proliferation and differentiation through tissue-tissue interactions. PMID:24912677

  20. Enrichment of skin-derived neural precursor cells from dermal cell populations by altering culture conditions.

    PubMed

    Bayati, Vahid; Gazor, Rohoullah; Nejatbakhsh, Reza; Negad Dehbashi, Fereshteh

    2016-01-01

    As stem cells play a critical role in tissue repair, their manipulation for being applied in regenerative medicine is of great importance. Skin-derived precursors (SKPs) may be good candidates for use in cell-based therapy as the only neural stem cells which can be isolated from an accessible tissue, skin. Herein, we presented a simple protocol to enrich neural SKPs by monolayer adherent cultivation to prove the efficacy of this method. To enrich neural SKPs from dermal cell populations, we have found that a monolayer adherent cultivation helps to increase the numbers of neural precursor cells. Indeed, we have cultured dermal cells as monolayer under serum-supplemented (control) and serum-supplemented culture, followed by serum free cultivation (test) and compared. Finally, protein markers of SKPs were assessed and compared in both experimental groups and differentiation potential was evaluated in enriched culture. The cells of enriched culture concurrently expressed fibronectin, vimentin and nestin, an intermediate filament protein expressed in neural and skeletal muscle precursors as compared to control culture. In addition, they possessed a multipotential capacity to differentiate into neurogenic, glial, adipogenic, osteogenic and skeletal myogenic cell lineages. It was concluded that serum-free adherent culture reinforced by growth factors have been shown to be effective on proliferation of skin-derived neural precursor cells (skin-NPCs) and drive their selective and rapid expansion.

  1. Rethinking cell growth models.

    PubMed

    Kafri, Moshe; Metzl-Raz, Eyal; Jonas, Felix; Barkai, Naama

    2016-11-01

    The minimal description of a growing cell consists of self-replicating ribosomes translating the cellular proteome. While neglecting all other cellular components, this model provides key insights into the control and limitations of growth rate. It shows, for example, that growth rate is maximized when ribosomes work at full capacity, explains the linear relation between growth rate and the ribosome fraction of the proteome and defines the maximal possible growth rate. This ribosome-centered model also highlights the challenge of coordinating cell growth with related processes such as cell division or nutrient production. Coordination is promoted when ribosomes don't translate at maximal capacity, as it allows escaping strict exponential growth. Recent data support the notion that multiple cellular processes limit growth. In particular, increasing transcriptional demand may be as deleterious as increasing translational demand, depending on growth conditions. Consistent with the idea of trade-off, cells may forgo maximal growth to enable more efficient interprocess coordination and faster adaptation to changing conditions. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  2. Antipsychotics promote neural differentiation of human iPS cell-derived neural stem cells.

    PubMed

    Asada, Minoru; Mizutani, Shuki; Takagi, Masatoshi; Suzuki, Hidenori

    2016-11-25

    We investigated the effects of antipsychotics on human induced pluripotent stem cell (hiPSC)-derived neural stem cell (NSC) differentiation. Induction of NSCs from hiPSCs was performed using PSC neural induction medium. Induced NSCs were subsequently cultured in neural differentiation medium containing antipsychotics. Cultured cells were subjected to neural differentiation marker analysis. As previously shown in rodent cells, antipsychotics promoted neural differentiation compared with vehicle treatment. Atypical antipsychotics appear to possess more differentiation induction potential than typical ones. Most NSCs do not express dopamine D2 receptor; however, our in vitro study indicates the clinical potential of antipsychotics could include effects independent of monoamine receptor expression in NSCs. Our study shows NSCs derived from hiPSCs provide opportunity to investigate the underlying direct effect of antipsychotics treatment on NSCs. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Neural stem cell sex dimorphism in aromatase (CYP19) expression: a basis for differential neural fate

    PubMed Central

    Waldron, Jay; McCourty, Althea; Lecanu, Laurent

    2010-01-01

    Purpose Neural stem cell (NSC) transplantation and pharmacologic activation of endogenous neurogenesis are two approaches that trigger a great deal of interest as brain repair strategies. However, the success rate of clinical attempts using stem cells to restore neurologic functions altered either after traumatic brain injury or as a consequence of neurodegenerative disease remains rather disappointing. This suggests that factors affecting the fate of grafted NSCs are largely understudied and remain to be characterized. We recently reported that aging differentially affects the neurogenic properties of male and female NSCs. Although the sex steroids androgens and estrogens participate in the regulation of neurogenesis, to our knowledge, research on how gender-based differences affect the capacity of NSCs to differentiate and condition their neural fate is lacking. In the present study, we explored further the role of cell sex as a determining factor of the neural fate followed by differentiating NSCs and its relationship with a potential differential expression of aromatase (CYP19), the testosterone-metabolizing enzyme. Results Using NSCs isolated from the subventricular zone of three-month-old male and female Long-Evans rats and maintained as neurospheres, we showed that differentiation triggered by retinoic acid resulted in a neural phenotype that depends on cell sex. Differentiated male NSCs mainly expressed markers of neuronal fate, including βIII-tubulin, microtubule associated protein 2, growth-associated protein 43, and doublecortin. In contrast, female NSCs essentially expressed the astrocyte marker glial fibrillary acidic protein. Quantification of the expression of aromatase showed a very low level of expression in undifferentiated female NSCs, whereas aromatase expression in male NSCs was 14-fold greater than the female level. Conclusion Our results confirm our previous data that the neural phenotype acquired by differentiating NSCs largely depends on

  4. Neural syntax: cell assemblies, synapsembles and readers

    PubMed Central

    Buzsáki, György

    2010-01-01

    Summary A widely discussed hypothesis in neuroscience is that transiently active ensembles of neurons, known as ‘cell assemblies’, underlie numerous operations of the brain, from encoding memories to reasoning. However, the mechanisms responsible for the formation and disbanding of cell assemblies and temporal evolution of cell assembly sequences are not well understood. I introduce and review three interconnected topics, which could facilitate progress in defining cell assemblies, identifying their neuronal organization and revealing causal relationships between assembly organization and behavior. First, I hypothesize that cell assemblies are best understood in light of their output product, as detected by ‘reader-actuator’ mechanisms. Second, I suggest that the hierarchical organization of cell assemblies may be regarded as a neural syntax. Third, constituents of the neural syntax are linked together by dynamically changing constellations of synaptic weights (‘synapsembles’). Existing support for this tripartite framework is reviewed and strategies for experimental testing of its predictions are discussed. PMID:21040841

  5. Ocular nerve growth factor administration counteracts the impairment of neural precursor cell viability and differentiation in the brain subventricular area of rats with streptozotocin-induced diabetes.

    PubMed

    Tirassa, Paola; Maccarone, Mattia; Carito, Valentina; De Nicolò, Sara; Fiore, Marco

    2015-05-01

    The ocular administration of nerve growth factor (NGF) as eye drops (oNGF) has been shown to exert protective effects in forebrain-injured animal models, including adult diabetes induced by a single injection of streptozotocin (STZ) (60 mg/kg body weight). This type 1 diabetes model was used in this study to investigate whether oNGF might extend its actions on neuronal precursors localised in the subventricular zone (SVZ). NGF or saline was administrated as eye drops twice daily for 2 weeks in rats with STZ-induced diabetes and healthy control rats. The expression of mature and precursor NGF and the NGF receptors, tropomyosin-related kinase A and neurotrophin receptor p75, and the levels of DNA fragmentation were analysed by ELISA and western blotting. Incorporation of bromodeoxyuridine was used to trace newly formed cells. Nestin, polysialylated neuronal cell adhesion molecule (PSA-NCAM), doublecortin (DCX) and glial fibrillary acidic protein antibodies were used to identify the SVZ cells by confocal microscopy. It was found that oNGF counteracts the STZ-induced cell death and the alteration of mature/pro-NGF expression in the SVZ. It also affects the survival and differentiation of SVZ progenitors. In particular, oNGF counteracts the reduction in the number of cells expressing PSA-NCAM/DCX (neuroblast type A cells) and the related reductions in the number and distribution of nestin/DCX-positive cells (C-type cells), or glia-committed cells (type B cells), observed in the SVZ of diabetic rats. These findings show that oNGF treatment counteracts the effect of type 1 diabetes on neuronal precursors in the SVZ, and further support the neuroprotective and reparative role of oNGF in the brain.

  6. Exfoliated Human Olfactory Neuroepithelium: A Source of Neural Progenitor Cells.

    PubMed

    Jiménez-Vaca, Ana L; Benitez-King, Gloria; Ruiz, Víctor; Ramírez-Rodríguez, Gerardo B; Hernández-de la Cruz, Beatriz; Salamanca-Gómez, Fabio A; González-Márquez, Humberto; Ramírez-Sánchez, Israel; Ortíz-López, Leonardo; Vélez-Del Valle, Cristina; Ordoñez-Razo, Rosa Ma

    2017-04-08

    Neural progenitor cells (NPC) contained in the human adult olfactory neuroepithelium (ONE) possess an undifferentiated state, the capability of self-renewal, the ability to generate neural and glial cells as well as being kept as neurospheres in cell culture conditions. Recently, NPC have been isolated from human or animal models using high-risk surgical methods. Therefore, it was necessary to improve methodologies to obtain and maintain human NPC as well as to achieve better knowledge of brain disorders. In this study, we propose the establishment and characterization of NPC cultures derived from the human olfactory neuroepithelium, using non-invasive procedures. Twenty-two healthy individuals (29.7 ± 4.5 years of age) were subjected to nasal exfoliation. Cells were recovered and kept as neurospheres under serum-free conditions. The neural progenitor origin of these neurospheres was determined by immunocytochemistry and qPCR. Their ability for self-renewal and multipotency was analyzed by clonogenic and differentiation assays, respectively. In the cultures, the ONE cells preserved the phenotype of the neurospheres. The expression levels of Nestin, Musashi, Sox2, and βIII-tubulin demonstrated the neural origin of the neurospheres; 48% of the cells separated could generate neurospheres, determining that they retained their self-renewal capacity. Neurospheres were differentiated in the absence of growth factors (EGF and FGF), and their multipotency ability was maintained as well. We were also able to isolate and grow human neural progenitor cells (neurospheres) through nasal exfoliates (non-invasive method) of the ONE from healthy adults, which is an extremely important contribution for the study of brain disorders and for the development of new therapies.

  7. Neural stem cells: an overview.

    PubMed

    Parati, E A; Pozzi, S; Ottolina, A; Onofrj, M; Bez, A; Pagano, S F

    2004-01-01

    Multipotent stem cells are present in the majority of mammalian tissues where they are a renewable source of specialized cells. According to the several biological portions from which multipotent stem cells can be derived, they are characterized as a) embryonic stem cells (ESCs) isolated from the pluripotent inner-cell mass of the pre-implantation blastocyste-stage embryo; b) multipotent fetal stem cells (FSCs) from aborted fetuses; and c) adult stem cells (ASCs) localized in small zones of several organs known as "niche" where a subset of tissue cells and extracellular substrates can indefinitely house one or more stem cells and control their self-renewal and progeny production in vivo. ECSs have an high self-renewing capacity, plasticity and pluripotency over the years. Pluripotency is a property that makes a stem cell able to give rise to all cell type found in the embryo and adult animals.

  8. In vitro study of the long-term cortisol treatment effects on the growth rate and proliferation of the neural stem/precursor cells.

    PubMed

    Abdanipour, Alireza; Sagha, Mohsen; Noori-Zadeh, Ali; Pakzad, Iraj; Tiraihi, Taki

    2015-02-01

    Adult neural stem/precursor cells (NSPCs) residing in the subventricular zone of the lateral ventricles and the subgranular zone of the dentate gyrus of the hippocampus are involved in the memory formations and psychological problems. It is believed that basal levels of glucocorticoids are essential for neuronal development, plasticity, and survival, while stress-mediated levels of glucocorticoids produce neuronal loss. Degeneration of NSPCs by the apoptotic and necrotic stimuli have great devastating outcomes on the brain and contributes to the pathophysiology of neurological as well as psychological disorders. Using MTT assay, acridine orange, and TUNEL assay, we have demonstrated that cortisol at high and excessive (more than 5 μM) levels had anti-proliferative effects on the NSPCs derived from subventricular and subgranular zones in a dose- and time-dependent manner through apoptosis as well as necrosis. These outcomes can highlight the role of stress-mediated decline of adult neurogenesis in the aging brain and interconnect stress-mediated cortisol secretion with brain aging diseases.

  9. Clinical translation of human neural stem cells

    PubMed Central

    2013-01-01

    Human neural stem cell transplants have potential as therapeutic candidates to treat a vast number of disorders of the central nervous system (CNS). StemCells, Inc. has purified human neural stem cells and developed culture conditions for expansion and banking that preserve their unique biological properties. The biological activity of these human central nervous system stem cells (HuCNS-SC®) has been analyzed extensively in vitro and in vivo. When formulated for transplantation, the expanded and cryopreserved banked cells maintain their stem cell phenotype, self-renew and generate mature oligodendrocytes, neurons and astrocytes, cells normally found in the CNS. In this overview, the rationale and supporting data for pursuing neuroprotective strategies and clinical translation in the three components of the CNS (brain, spinal cord and eye) are described. A phase I trial for a rare myelin disorder and phase I/II trial for spinal cord injury are providing intriguing data relevant to the biological properties of neural stem cells, and the early clinical outcomes compel further development. PMID:23987648

  10. ADULT NEURAL STEM CELLS: RESPONSE TO STROKE INJURY AND POTENTIAL FOR THERAPEUTIC APPLICATIONS

    PubMed Central

    Barkho, Basam Z.; Zhao, Xinyu

    2011-01-01

    The plasticity of neural stem/progenitor cells allows a variety of different responses to many environmental cues. In the past decade, significant research has gone into understanding the regulation of neural stem/progenitor cell properties, because of their promise for cell replacement therapies in adult neurological diseases. Both endogenous and grafted neural stem/progenitor cells are known to have the ability to migrate long distances to lesioned sites after brain injury and differentiate into new neurons. Several chemokines and growth factors, including stromal cell-derived factor-1 and vascular endothelial growth factor, have been shown to stimulate the proliferation, differentiation, and migration of neural stem/progenitor cells, and investigators have now begun to identify the critical downstream effectors and signaling mechanisms that regulate these processes. Both our own lab and others have shown that the extracellular matrix and matrix remodeling factors play a critical role in directing cell differentiation and migration of adult neural stem/progenitor cells within injured sites. Identification of these and other molecular pathways involved in stem cell homing into ischemic areas is vital for the development of new treatments. To ensure the best functional recovery, regenerative therapy may require the application of a combination approach that includes cell replacement, trophic support, and neural protection. Here we review the current state of our knowledge about endogenous adult and exogenous neural stem/progenitor cells as potential therapeutic agents for central nervous system injuries. PMID:21466483

  11. Neural crest cells: from developmental biology to clinical interventions.

    PubMed

    Noisa, Parinya; Raivio, Taneli

    2014-09-01

    Neural crest cells are multipotent cells, which are specified in embryonic ectoderm in the border of neural plate and epiderm during early development by interconnection of extrinsic stimuli and intrinsic factors. Neural crest cells are capable of differentiating into various somatic cell types, including melanocytes, craniofacial cartilage and bone, smooth muscle, and peripheral nervous cells, which supports their promise for cell therapy. In this work, we provide a comprehensive review of wide aspects of neural crest cells from their developmental biology to applicability in medical research. We provide a simplified model of neural crest cell development and highlight the key external stimuli and intrinsic regulators that determine the neural crest cell fate. Defects of neural crest cell development leading to several human disorders are also mentioned, with the emphasis of using human induced pluripotent stem cells to model neurocristopathic syndromes. © 2014 Wiley Periodicals, Inc.

  12. Neural prosthesis in the wake of nanotechnology: controlled growth of neurons using surface nanostructures.

    PubMed

    Lee, J K; Baac, H; Song, S H; Jang, E; Lee, S D; Park, D; Kim, S J

    2006-01-01

    Neural prosthesis has been successfully applied to patients with motional or sensory disabilities for clinical purpose. To enhance the performance of the neural prosthetic device, the electrodes for the biosignal recording or electrical stimulation should be located in closer proximity to target neurons than they are now. Instead of revising the prior implanting surgery to improve the electrical contact of neurons, we propose a technique that can bring the neurons closer to the electrode sites. A new method is investigated that can control the direction of neural cell growth using surface nanostructures. We successfully guide the neurons to the position of the microelectrodes by providing a surface topographical cue presented by the surface nanostructure on a photoresponsive polymer material. Because the surface structure formed by laser holography is reversible and repeatable, the geometrical positioning of the neurons to microelectrodes can be adjusted by applying laser treatment during the surgery for the purpose of improving the performance of neural prosthetic device.

  13. Differentiation of Neural Lineage Cells from Human Pluripotent Stem Cells

    PubMed Central

    Schwartz, Philip H.; Brick, David J.; Stover, Alexander E.; Loring, Jeanne F.; Müller, Franz Josef

    2008-01-01

    Human pluripotent stem cells have the unique properties of being able to proliferate indefinitely in their undifferentiated state and to differentiate into any somatic cell type. These cells are thus posited to be extremely useful for furthering our understanding of both normal and abnormal human development, providing a human cell preparation that can be used to screen for new reagents or therapeutic agents, and generating large numbers of differentiated cells that can be used for transplantation purposes. Critical among the applications for the latter are diseases and injuries of the nervous system, medical approaches to which have been, to date, primarily palliative in nature. Differentiation of human pluripotent stem cells into cells of the neural lineage, therefore, has become a central focus of a number of laboratories. This has resulted in the description in the literature of several dozen methods for neural cell differentiation from human pluripotent stem cells. Among these are methods for the generation of such divergent neural cells as dopaminergic neurons, retinal neurons, ventral motoneurons, and oligodendroglial progenitors. In this review, we attempt to fully describe most of these methods, breaking them down into five basic subdivisions: 1) starting material, 2) induction of loss of pluripotency, 3) neural induction, 4) neural maintenance and expansion, and 5) neuronal/glial differentiation. We also show data supporting the concept that undifferentiated human pluripotent stem cells appear to have an innate neural differentiation potential. In addition, we evaluate data comparing and contrasting neural stem cells differentiated from human pluripotent stem cells with those derived directly from the human brain. PMID:18593611

  14. Neural stem cell transplantation in mouse brain.

    PubMed

    Lee, Jean-Pyo; McKercher, Scott; Muller, Franz-Josef; Snyder, Evan Y

    2008-01-01

    Neural stem cells (NSCs) are the most primordial, least committed cells of the nervous system, and transplantation of these multipotent cells holds the promise of regenerative therapy for many central nervous system (CNS) diseases. This unit describes methods for NSC transplantation into neonatal mouse pups, embryonic mouse brain, and adult mouse brain. A description of options for detection of labeled donor cells in engrafted mouse brain is provided along with an example protocol for detecting lacZ-expressing cells in situ. Also included is a protocol for preparing NSCs for transplantation.

  15. Differentiation state determines neural effects on microvascular endothelial cells

    SciTech Connect

    Muffley, Lara A.; Pan, Shin-Chen; Smith, Andria N.; Ga, Maricar; Hocking, Anne M.; Gibran, Nicole S.

    2012-10-01

    Growing evidence indicates that nerves and capillaries interact paracrinely in uninjured skin and cutaneous wounds. Although mature neurons are the predominant neural cell in the skin, neural progenitor cells have also been detected in uninjured adult skin. The aim of this study was to characterize differential paracrine effects of neural progenitor cells and mature sensory neurons on dermal microvascular endothelial cells. Our results suggest that neural progenitor cells and mature sensory neurons have unique secretory profiles and distinct effects on dermal microvascular endothelial cell proliferation, migration, and nitric oxide production. Neural progenitor cells and dorsal root ganglion neurons secrete different proteins related to angiogenesis. Specific to neural progenitor cells were dipeptidyl peptidase-4, IGFBP-2, pentraxin-3, serpin f1, TIMP-1, TIMP-4 and VEGF. In contrast, endostatin, FGF-1, MCP-1 and thrombospondin-2 were specific to dorsal root ganglion neurons. Microvascular endothelial cell proliferation was inhibited by dorsal root ganglion neurons but unaffected by neural progenitor cells. In contrast, microvascular endothelial cell migration in a scratch wound assay was inhibited by neural progenitor cells and unaffected by dorsal root ganglion neurons. In addition, nitric oxide production by microvascular endothelial cells was increased by dorsal root ganglion neurons but unaffected by neural progenitor cells. -- Highlights: Black-Right-Pointing-Pointer Dorsal root ganglion neurons, not neural progenitor cells, regulate microvascular endothelial cell proliferation. Black-Right-Pointing-Pointer Neural progenitor cells, not dorsal root ganglion neurons, regulate microvascular endothelial cell migration. Black-Right-Pointing-Pointer Neural progenitor cells and dorsal root ganglion neurons do not effect microvascular endothelial tube formation. Black-Right-Pointing-Pointer Dorsal root ganglion neurons, not neural progenitor cells, regulate

  16. Single-cell transcriptome analyses reveal signals to activate dormant neural stem cells.

    PubMed

    Luo, Yuping; Coskun, Volkan; Liang, Aibing; Yu, Juehua; Cheng, Liming; Ge, Weihong; Shi, Zhanping; Zhang, Kunshan; Li, Chun; Cui, Yaru; Lin, Haijun; Luo, Dandan; Wang, Junbang; Lin, Connie; Dai, Zachary; Zhu, Hongwen; Zhang, Jun; Liu, Jie; Liu, Hailiang; deVellis, Jean; Horvath, Steve; Sun, Yi Eve; Li, Siguang

    2015-05-21

    The scarcity of tissue-specific stem cells and the complexity of their surrounding environment have made molecular characterization of these cells particularly challenging. Through single-cell transcriptome and weighted gene co-expression network analysis (WGCNA), we uncovered molecular properties of CD133(+)/GFAP(-) ependymal (E) cells in the adult mouse forebrain neurogenic zone. Surprisingly, prominent hub genes of the gene network unique to ependymal CD133(+)/GFAP(-) quiescent cells were enriched for immune-responsive genes, as well as genes encoding receptors for angiogenic factors. Administration of vascular endothelial growth factor (VEGF) activated CD133(+) ependymal neural stem cells (NSCs), lining not only the lateral but also the fourth ventricles and, together with basic fibroblast growth factor (bFGF), elicited subsequent neural lineage differentiation and migration. This study revealed the existence of dormant ependymal NSCs throughout the ventricular surface of the CNS, as well as signals abundant after injury for their activation.

  17. Growth and splitting of neural sequences in songbird vocal development

    PubMed Central

    Okubo, Tatsuo S.; Mackevicius, Emily L.; Payne, Hannah L.; Lynch, Galen F.; Fee, Michale S.

    2015-01-01

    Neural sequences are a fundamental feature of brain dynamics underlying diverse behaviors, but the mechanisms by which they develop during learning remain unknown. Songbirds learn vocalizations composed of syllables; in adult birds, each syllable is produced by a different sequence of action potential bursts in the premotor cortical area HVC. Here we carried out recordings of large populations of HVC neurons in singing juvenile birds throughout learning to examine the emergence of neural sequences. Early in vocal development, HVC neurons begin producing rhythmic bursts, temporally locked to a ‘prototype’ syllable. Different neurons are active at different latencies relative to syllable onset to form a continuous sequence. Through development, as new syllables emerge from the prototype syllable, initially highly overlapping burst sequences become increasingly distinct. We propose a mechanistic model in which multiple neural sequences can emerge from the growth and splitting of a common precursor sequence. PMID:26618871

  18. Immunological control of adult neural stem cells

    PubMed Central

    Gonzalez-Perez, Oscar; Quiñones-Hinojosa, Alfredo; Garcia-Verdugo, Jose Manuel

    2010-01-01

    Adult neurogenesis occurs only in discrete regions of adult central nervous system: the subventricular zone and the subgranular zone. These areas are populated by adult neural stem cells (aNSC) that are regulated by a number of molecules and signaling pathways, which control their cell fate choices, survival and proliferation rates. For a long time, it was believed that the immune system did not exert any control on neural proliferative niches. However, it has been observed that many pathological and inflammatory conditions significantly affect NSC niches. Even more, increasing evidence indicates that chemokines and cytokines play an important role in regulating proliferation, cell fate choices, migration and survival of NSCs under physiological conditions. Hence, the immune system is emerging is an important regulator of neurogenic niches in the adult brain, which may have clinical relevance in several brain diseases. PMID:20861925

  19. Persistent neural activity in head direction cells

    NASA Technical Reports Server (NTRS)

    Taube, Jeffrey S.; Bassett, Joshua P.; Oman, C. M. (Principal Investigator)

    2003-01-01

    Many neurons throughout the rat limbic system discharge in relation to the animal's directional heading with respect to its environment. These so-called head direction (HD) cells exhibit characteristics of persistent neural activity. This article summarizes where HD cells are found, their major properties, and some of the important experiments that have been conducted to elucidate how this signal is generated. The number of HD and angular head velocity cells was estimated for several brain areas involved in the generation of the HD signal, including the postsubiculum, anterior dorsal thalamus, lateral mammillary nuclei and dorsal tegmental nucleus. The HD cell signal has many features in common with what is known about how neural integration is accomplished in the oculomotor system. The nature of the HD cell signal makes it an attractive candidate for using neural network models to elucidate the signal's underlying mechanisms. The conditions that any network model must satisfy in order to accurately represent how the nervous system generates this signal are highlighted and areas where key information is missing are discussed.

  20. Persistent neural activity in head direction cells

    NASA Technical Reports Server (NTRS)

    Taube, Jeffrey S.; Bassett, Joshua P.; Oman, C. M. (Principal Investigator)

    2003-01-01

    Many neurons throughout the rat limbic system discharge in relation to the animal's directional heading with respect to its environment. These so-called head direction (HD) cells exhibit characteristics of persistent neural activity. This article summarizes where HD cells are found, their major properties, and some of the important experiments that have been conducted to elucidate how this signal is generated. The number of HD and angular head velocity cells was estimated for several brain areas involved in the generation of the HD signal, including the postsubiculum, anterior dorsal thalamus, lateral mammillary nuclei and dorsal tegmental nucleus. The HD cell signal has many features in common with what is known about how neural integration is accomplished in the oculomotor system. The nature of the HD cell signal makes it an attractive candidate for using neural network models to elucidate the signal's underlying mechanisms. The conditions that any network model must satisfy in order to accurately represent how the nervous system generates this signal are highlighted and areas where key information is missing are discussed.

  1. The neural crest and neural crest cells: discovery and significance for theories of embryonic organization.

    PubMed

    Hall, Brian K

    2008-12-01

    The neural crest has long fascinated developmental biologists,and,increasingly over the past decades,evolutionary and evolutionary developmental biologists.The neural crest is the name given to the fold of ectoderm at the junction between neural and epidermal ectoderm in neurula-stage vertebrate embryos.In this sense,the neural crest is a morphological term akin to head fold or limb bud.This region of the dorsal neural tube consists of neural crest cells,a special population(s)of cell,that give rise to an astonishing number of cell types and to an equally astonishing number of tissues and organs.Neural crest cell contributions may be direct - providing cells - or indirect - providing a necessary, often inductive, environment in which other cells develop.The enormous range of cell types produced provides an important source of evidence of the neural crest as a germ layer, bringing the number of germ layers to four - ectoderm,endoderm,mesoderm,and neural crest. In this paper I provide a brief overview of the major phases of investigation into the neural crest and the major players involved,discuss how the origin of the neural crest relates to the origin of the nervous system in vertebrate embryos,discuss the impact on the germ-layer theory of the discovery of the neural crest and of secondary neurulation,and present evidence of the neural crest as the fourth germ layer.A companion paper (Hall, Evol. Biol.2008) deals with the evolutionary origins of the neural crest and neural crest cells.

  2. Phenotype and Stability of Neural Differentiation of Androgenetic Murine ES Cell-Derived Neural Progenitor Cells

    PubMed Central

    Wolber, Wanja; Ahmad, Ruhel; Choi, Soon Won; Eckardt, Sigrid; McLaughlin, K. John; Schmitt, Jessica; Geis, Christian; Heckmann, Manfred; Sirén, Anna-Leena; Müller, Albrecht M.

    2013-01-01

    Uniparental zygotes with two paternal (androgenetic, AG) or two maternal genomes (gynogenetic, GG) cannot develop into viable offsprings but form blastocysts from which pluripotent embryonic stem (ES) cells can be derived. For most organs, it is unclear whether uniparental ES cells can give rise to stably expandable somatic stem cells that can repair injured tissues. Even if previous reports indicated that the capacity of AG ES cells to differentiate in vitro into pan-neural progenitor cells (pNPCs) and into cells expressing neural markers is similar to biparental [normal fertilized (N)] ES cells, their potential for functional neurogenesis is not known. Here we show that murine AG pNPCs give rise to neuron-like cells, which then generate sodium-driven action potentials while maintaining fidelity of imprinted gene expression. Neural engraftment after intracerebral transplantation was achieved only by late (22 days) AG and N pNPCs with in vitro low colony-forming cell (CFC) capacity. However, persisting CFC formation seen, in particular, in early (13 or 16 days) differentiation cultures of N and AG pNPCs correlated with a high incidence of trigerm layer teratomas. As AG ES cells display functional neurogenesis and in vivo stability similar to N ES cells, they represent a unique model system to study the roles of paternal and maternal genomes on neural development and on the development of imprinting-associated brain diseases. PMID:26858862

  3. CXCR4 activation promotes differentiation of human embryonic stem cells to neural stem cells.

    PubMed

    Zhang, Lijun; Hua, Qiuhong; Tang, Kaiyi; Shi, Changjie; Xie, Xin; Zhang, Ru

    2016-11-19

    G protein-coupled receptors (GPCRs) are involved in many fundamental cellular responses such as growth, death, movement, transcription and excitation. Their roles in human stem cell neural specialization are not well understood. In this study, we aimed to identify GPCRs that may play a role in the differentiation of human embryonic stem cells (hESCs) to neural stem cells (NSCs). Using a feeder-free hESC neural differentiation protocol, we found that the expression of several chemokine receptors changed dramatically during the hESC/NSC transition. Especially, the expression of CXCR4 increased approximately 50 folds in NSCs compared to the original hESCs. CXCR4 agonist SDF-1 promoted, whereas the antagonist AMD3100 delayed the neural induction process. In consistence with antagonizing CXCR4, knockdown of CXCR4 in hESCs also blocked the neural induction and cells with reduced CXCR4 were rarely positive for Nestin and Sox1-staining. Taken together, our results suggest that CXCR4 is involved in the neural induction process of hESC and it might be considered as a target to facilitate NSC production from hESCs in regenerative medicine.

  4. Lateral patterns of the neural cell adhesion molecule on the surface of hippocampal cells developing in vitro.

    PubMed

    Krivko, I M; Rusakov, D A; Savina, S V; Skibo, G G; Berezin, V A

    1993-07-01

    In monolayer cultures of hippocampal neurons from newborn rats, an immunocytochemical quantitative study was carried out to investigate age-dependent arrangement of the neural cell adhesion molecules in different parts of cell membranes. On the fifth and 12th day in vitro, neural cell adhesion molecules were labelled with specific antibodies and protein A conjugated to colloidal gold particles. Samples of randomly selected electron micrographs that displayed labelled membrane fragments of cell bodies, growth cones, and axons were numerically analysed for the five- and 12-day in vitro neurons. Neural cell adhesion molecules surface topography was quantitatively described and compared, using a statistical stereological approach. The mean surface density of labelled neural cell adhesion molecules was found to be approximately 2.5 times higher in growth cone membranes relative to somatic and axonal membranes in five-day in vitro neurons. By the 12th day in vitro, this density decreases in somatic membranes (approximately 18%) and increases in axonal membranes (approximately 60%). Representative spectra of lateral intervals between labels as well as images that show typical topography of label on membrane surfaces were simulated. The results revealed regular patterns of neural cell adhesion molecules on the somatic surface and allowed consideration of neural cell adhesion molecules arrangement in a view of membrane adhesion properties. Participation of cytoskeleton in neural cell adhesion molecules rearrangement is discussed.

  5. Human neural progenitor cells promote photoreceptor survival in retinal explants.

    PubMed

    Englund-Johansson, Ulrica; Mohlin, Camilla; Liljekvist-Soltic, Ingela; Ekström, Per; Johansson, Kjell

    2010-02-01

    Different types of progenitor and stem cells have been shown to provide neuroprotection in animal models of photoreceptor degeneration. The present study was conducted to investigate whether human neural progenitor cells (HNPCs) have neuroprotective properties on retinal explants models with calpain- and caspase-3-dependent photoreceptor cell death. In the first experiments, HNPCs in a feeder layer were co-cultured for 6 days either with postnatal rd1 mouse or normal rat retinas. Retinal histological sections were used to determine outer nuclear layer (ONL) thickness, and to detect the number of photoreceptors with labeling for calpain activity, cleaved caspase-3 and TUNEL. The ONL thickness of co-cultured rat and rd1 retinas was found to be almost 10% and 40% thicker, respectively, compared to controls. Cell counts of calpain activity, cleaved caspase-3 and TUNEL labeled photoreceptors in both models revealed a 30-50% decrease when co-cultured with HNPCs. The results represent significant increases of photoreceptor survival in the co-cultured retinas. In the second experiments, for an identification of putative survival factors, or a combination of them, a growth factor profile was performed on conditioned medium. The relative levels of various growth factors were analyzed by densitometric measurements of growth factor array membranes. Following growth factors were identified as most potential survival factors; granulocyte colony stimulating factor (G-CSF), granulocyte-macrophage colony stimulating factor (GMCSF), insulin-like growth factor II (IGF-II), neurotrophic factor 3 (NT-3), placental growth factor (PIGF), transforming growth factors (TGF-beta1 and TGF-beta2) and vascular endothelial growth factor (VEGF-D). HNPCs protect both against calpain- and caspase-3-dependent photoreceptor cell death in the rd1 mouse and against caspase-3-dependent photoreceptor cell death in normal rat retinas in vitro. The protective effect is possibly achieved by a variety of

  6. Cell Growth Enhancement

    NASA Technical Reports Server (NTRS)

    1992-01-01

    Exogene Corporation uses advanced technologies to enhance production of bio-processed substances like proteins, antibiotics and amino acids. Among them are genetic modification and a genetic switch. They originated in research for Jet Propulsion Laboratory. Extensive experiments in cell growth through production of hemoglobin to improve oxygen supply to cells were performed. By improving efficiency of oxygen use by cells, major operational expenses can be reduced. Greater product yields result in decreased raw material costs and more efficient use of equipment. A broad range of applications is cited.

  7. Regulation of endogenous neural stem/progenitor cells for neural repair—factors that promote neurogenesis and gliogenesis in the normal and damaged brain

    PubMed Central

    Christie, Kimberly J.; Turnley, Ann M.

    2012-01-01

    Neural stem/precursor cells in the adult brain reside in the subventricular zone (SVZ) of the lateral ventricles and the subgranular zone (SGZ) of the dentate gyrus in the hippocampus. These cells primarily generate neuroblasts that normally migrate to the olfactory bulb (OB) and the dentate granule cell layer respectively. Following brain damage, such as traumatic brain injury, ischemic stroke or in degenerative disease models, neural precursor cells from the SVZ in particular, can migrate from their normal route along the rostral migratory stream (RMS) to the site of neural damage. This neural precursor cell response to neural damage is mediated by release of endogenous factors, including cytokines and chemokines produced by the inflammatory response at the injury site, and by the production of growth and neurotrophic factors. Endogenous hippocampal neurogenesis is frequently also directly or indirectly affected by neural damage. Administration of a variety of factors that regulate different aspects of neural stem/precursor biology often leads to improved functional motor and/or behavioral outcomes. Such factors can target neural stem/precursor proliferation, survival, migration and differentiation into appropriate neuronal or glial lineages. Newborn cells also need to subsequently survive and functionally integrate into extant neural circuitry, which may be the major bottleneck to the current therapeutic potential of neural stem/precursor cells. This review will cover the effects of a range of intrinsic and extrinsic factors that regulate neural stem/precursor cell functions. In particular it focuses on factors that may be harnessed to enhance the endogenous neural stem/precursor cell response to neural damage, highlighting those that have already shown evidence of preclinical effectiveness and discussing others that warrant further preclinical investigation. PMID:23346046

  8. Macro cell placement with neural net algorithms

    NASA Astrophysics Data System (ADS)

    Storti-Gajani, Giancarlo

    Placement of VLSI (Very Large Scale Integration) macro cells is one of the hard problems encountered in the process of integrated circuits design. Since the problem is essentially NP-complete a solution must be searched for with the aid of heuristics using, maybe, non deterministic strategies. A new algorithm for cell preplacement based on neural nets that may be very well extended to find solution of the final placement problem is presented. Simulations for the part of the algorithm concerning preplacement are carried out on several different examples giving always a sharply decreasing cost function (where cost is evaluated essentially on total length of wires given a rectangular boundary). The direct mapping between neural units and VLSI blocks that is adopted in the algorithm makes the extension to the final placement problem quite simple. Simulation programs are implemented in a interpreted mathematical simulation language and a C language implementation is currently under way.

  9. Reconstitution of a Patterned Neural Tube from Single Mouse Embryonic Stem Cells.

    PubMed

    Ishihara, Keisuke; Ranga, Adrian; Lutolf, Matthias P; Tanaka, Elly M; Meinhardt, Andrea

    2017-01-01

    The recapitulation of tissue development and patterning in three-dimensional (3D) culture is an important dimension of stem cell research. Here, we describe a 3D culture protocol in which single mouse ES cells embedded in Matrigel under neural induction conditions clonally form a lumen containing, oval-shaped epithelial structure within 3 days. By Day 7 an apicobasally polarized neuroepithelium with uniformly dorsal cell identity forms. Treatment with retinoic acid at Day 2 results in posteriorization and self-organization of dorsal-ventral neural tube patterning. Neural tube organoid growth is also supported by pure laminin gels as well as poly(ethylene glycol) (PEG)-based artificial extracellular matrix hydrogels, which can be fine-tuned for key microenvironment characteristics. The rapid generation of a simple, patterned tissue in well-defined culture conditions makes the neural tube organoid a tractable model for studying neural stem cell self-organization.

  10. Adult Palatum as a Novel Source of Neural Crest-Related Stem Cells

    PubMed Central

    Widera, Darius; Zander, Christin; Heidbreder, Meike; Kasperek, Yvonne; Noll, Thomas; Seitz, Oliver; Saldamli, Belma; Sudhoff, Holger; Sader, Robert; Kaltschmidt, Christian; Kaltschmidt, Barbara

    2009-01-01

    Somatic neural and neural crest stem cells are promising sources for cellular therapy of several neurodegenerative diseases. However, because of practical considerations such as inadequate accessibility of the source material, the application of neural crest stem cells is strictly limited. The secondary palate is a highly regenerative and heavily innervated tissue, which develops embryonically under direct contribution of neural crest cells. Here, we describe for the first time the presence of nestin-positive neural crest-related stem cells within Meissner corpuscles and Merkel cell-neurite complexes located in the hard palate of adult Wistar rats. After isolation, palatal neural crest-related stem cells (pNC-SCs) were cultivated in the presence of epidermal growth factor and fibroblast growth factor under serum-free conditions, resulting in large amounts of neurospheres. We used immunocytochemical techniques and reverse transcriptase-polymerase chain reaction to assess the expression profile of pNC-SCs. In addition to the expression of neural crest stem cell markers such as Nestin, Sox2, and p75, we detected the expression of Klf4, Oct4, and c-Myc. pNC-SCs differentiated efficiently into neuronal and glial cells. Finally, we investigated the potential expression of stemness markers within the human palate. We identified expression of stem cell markers nestin and CD133 and the transcription factors needed for reprogramming of somatic cells into pluripotent cells: Sox2, Oct4, Klf4, and c-Myc. These data show that cells isolated from palatal rugae form neurospheres, are highly plastic, and express neural crest stem cell markers. In addition, pNC-SCs may have the ability to differentiate into functional neurons and glial cells, serving as a starting point for therapeutic studies. Stem Cells 2009;27:1899–1910 PMID:19544446

  11. Endothelial cells regulate neural crest and second heart field morphogenesis

    PubMed Central

    Milgrom-Hoffman, Michal; Michailovici, Inbal; Ferrara, Napoleone; Zelzer, Elazar; Tzahor, Eldad

    2014-01-01

    ABSTRACT Cardiac and craniofacial developmental programs are intricately linked during early embryogenesis, which is also reflected by a high frequency of birth defects affecting both regions. The molecular nature of the crosstalk between mesoderm and neural crest progenitors and the involvement of endothelial cells within the cardio–craniofacial field are largely unclear. Here we show in the mouse that genetic ablation of vascular endothelial growth factor receptor 2 (Flk1) in the mesoderm results in early embryonic lethality, severe deformation of the cardio–craniofacial field, lack of endothelial cells and a poorly formed vascular system. We provide evidence that endothelial cells are required for migration and survival of cranial neural crest cells and consequently for the deployment of second heart field progenitors into the cardiac outflow tract. Insights into the molecular mechanisms reveal marked reduction in Transforming growth factor beta 1 (Tgfb1) along with changes in the extracellular matrix (ECM) composition. Our collective findings in both mouse and avian models suggest that endothelial cells coordinate cardio–craniofacial morphogenesis, in part via a conserved signaling circuit regulating ECM remodeling by Tgfb1. PMID:24996922

  12. Endothelial cells regulate neural crest and second heart field morphogenesis.

    PubMed

    Milgrom-Hoffman, Michal; Michailovici, Inbal; Ferrara, Napoleone; Zelzer, Elazar; Tzahor, Eldad

    2014-07-04

    Cardiac and craniofacial developmental programs are intricately linked during early embryogenesis, which is also reflected by a high frequency of birth defects affecting both regions. The molecular nature of the crosstalk between mesoderm and neural crest progenitors and the involvement of endothelial cells within the cardio-craniofacial field are largely unclear. Here we show in the mouse that genetic ablation of vascular endothelial growth factor receptor 2 (Flk1) in the mesoderm results in early embryonic lethality, severe deformation of the cardio-craniofacial field, lack of endothelial cells and a poorly formed vascular system. We provide evidence that endothelial cells are required for migration and survival of cranial neural crest cells and consequently for the deployment of second heart field progenitors into the cardiac outflow tract. Insights into the molecular mechanisms reveal marked reduction in Transforming growth factor beta 1 (Tgfb1) along with changes in the extracellular matrix (ECM) composition. Our collective findings in both mouse and avian models suggest that endothelial cells coordinate cardio-craniofacial morphogenesis, in part via a conserved signaling circuit regulating ECM remodeling by Tgfb1.

  13. Enhanced growth of neural networks on conductive cellulose-derived nanofibrous scaffolds.

    PubMed

    Kuzmenko, Volodymyr; Kalogeropoulos, Theodoros; Thunberg, Johannes; Johannesson, Sara; Hägg, Daniel; Enoksson, Peter; Gatenholm, Paul

    2016-01-01

    The problem of recovery from neurodegeneration needs new effective solutions. Tissue engineering is viewed as a prospective approach for solving this problem since it can help to develop healthy neural tissue using supportive scaffolds. This study presents effective and sustainable tissue engineering methods for creating biomaterials from cellulose that can be used either as scaffolds for the growth of neural tissue in vitro or as drug screening models. To reach this goal, nanofibrous electrospun cellulose mats were made conductive via two different procedures: carbonization and addition of multi-walled carbon nanotubes. The resulting scaffolds were much more conductive than untreated cellulose material and were used to support growth and differentiation of SH-SY5Y neuroblastoma cells. The cells were evaluated by scanning electron microscopy and confocal microscopy methods over a period of 15 days at different time points. The results showed that the cellulose-derived conductive scaffolds can provide support for good cell attachment, growth and differentiation. The formation of a neural network occurred within 10 days of differentiation, which is a promising length of time for SH-SY5Y neuroblastoma cells.

  14. Three-dimensional graphene foam as a biocompatible and conductive scaffold for neural stem cells

    NASA Astrophysics Data System (ADS)

    Li, Ning; Zhang, Qi; Gao, Song; Song, Qin; Huang, Rong; Wang, Long; Liu, Liwei; Dai, Jianwu; Tang, Mingliang; Cheng, Guosheng

    2013-04-01

    Neural stem cell (NSC) based therapy provides a promising approach for neural regeneration. For the success of NSC clinical application, a scaffold is required to provide three-dimensional (3D) cell growth microenvironments and appropriate synergistic cell guidance cues. Here, we report the first utilization of graphene foam, a 3D porous structure, as a novel scaffold for NSCs in vitro. It was found that three-dimensional graphene foams (3D-GFs) can not only support NSC growth, but also keep cell at an active proliferation state with upregulation of Ki67 expression than that of two-dimensional graphene films. Meanwhile, phenotypic analysis indicated that 3D-GFs can enhance the NSC differentiation towards astrocytes and especially neurons. Furthermore, a good electrical coupling of 3D-GFs with differentiated NSCs for efficient electrical stimulation was observed. Our findings implicate 3D-GFs could offer a powerful platform for NSC research, neural tissue engineering and neural prostheses.

  15. Efficient neural differentiation of mouse pluripotent stem cells in a serum-free medium and development of a novel strategy for enrichment of neural cells.

    PubMed

    Verma, Isha; Rashid, Zubin; Sikdar, Sujit K; Seshagiri, Polani B

    2017-10-01

    Pluripotent stem cells (PSCs) offer an excellent model to study neural development and function. Although various protocols have been developed to direct the differentiation of PSCs into desired neural cell types, many of them suffer from limitations including low efficiency, long duration of culture, and the use of expensive, labile, and undefined growth supplements. In this study, we achieved efficient differentiation of mouse PSCs to neural lineage, in the absence of exogenous molecules, by employing a serum-free culture medium containing knockout serum replacement (KSR). Embryoid bodies (EBs) cultured in this medium predominantly produced neural cells which included neural progenitors (15-18%), immature neurons (8-24%), mature neurons (10-26%), astrocytes (27-61%), and oligodendrocytes (∼1%). Different neuronal subtypes including glutamatergic, GABAergic, cholinergic, serotonergic, and dopaminergic neurons were generated. Importantly, neurons generated in the KSR medium were electrically active. Further, the EB scooping strategy, involving the removal of the EB core region from the peripheral EB outgrowth, resulted in the enrichment of PSC-derived neural cells. Taken together, this study provides the evidence that the KSR medium is ideal for the rapid and efficient generation of neural cells, including functional neurons, from PSCs without the requirement of any other additional molecule. Copyright © 2017 ISDN. Published by Elsevier Ltd. All rights reserved.

  16. Adult palatum as a novel source of neural crest-related stem cells.

    PubMed

    Widera, Darius; Zander, Christin; Heidbreder, Meike; Kasperek, Yvonne; Noll, Thomas; Seitz, Oliver; Saldamli, Belma; Sudhoff, Holger; Sader, Robert; Kaltschmidt, Christian; Kaltschmidt, Barbara

    2009-08-01

    Somatic neural and neural crest stem cells are promising sources for cellular therapy of several neurodegenerative diseases. However, because of practical considerations such as inadequate accessibility of the source material, the application of neural crest stem cells is strictly limited. The secondary palate is a highly regenerative and heavily innervated tissue, which develops embryonically under direct contribution of neural crest cells. Here, we describe for the first time the presence of nestin-positive neural crest-related stem cells within Meissner corpuscles and Merkel cell-neurite complexes located in the hard palate of adult Wistar rats. After isolation, palatal neural crest-related stem cells (pNC-SCs) were cultivated in the presence of epidermal growth factor and fibroblast growth factor under serum-free conditions, resulting in large amounts of neurospheres. We used immunocytochemical techniques and reverse transcriptase-polymerase chain reaction to assess the expression profile of pNC-SCs. In addition to the expression of neural crest stem cell markers such as Nestin, Sox2, and p75, we detected the expression of Klf4, Oct4, and c-Myc. pNC-SCs differentiated efficiently into neuronal and glial cells. Finally, we investigated the potential expression of stemness markers within the human palate. We identified expression of stem cell markers nestin and CD133 and the transcription factors needed for reprogramming of somatic cells into pluripotent cells: Sox2, Oct4, Klf4, and c-Myc. These data show that cells isolated from palatal rugae form neurospheres, are highly plastic, and express neural crest stem cell markers. In addition, pNC-SCs may have the ability to differentiate into functional neurons and glial cells, serving as a starting point for therapeutic studies.

  17. Synergistic effects of FGF-2 and Activin A on early neural differentiation of human pluripotent stem cells.

    PubMed

    Mimura, Sumiyo; Suga, Mika; Liu, Yujung; Kinehara, Masaki; Yanagihara, Kana; Ohnuma, Kiyoshi; Nikawa, Hiroki; Furue, Miho K

    2015-09-01

    Neural differentiation is an important target of human embryonic stem cells, which provide a source for cell-based therapy, developmental biology, and pharmaceutical research. Previous studies revealed that inhibition of the bone morphogenetic protein is required for neural induction from human embryonic stem cells. On the contrary, the functions of fibroblast growth factors and Activin/Nodal signaling are controversial. Fibroblast growth factor-2 and Activin/Nodal pathways exert divergent influences on human embryonic stem cell concerning the maintenance of both pluripotency and cellular differentiation. We hypothesized that the combination of fibroblast growth factor-2 and Activin A at various concentrations synergistically exerts diverse effects on cell differentiation. To determine the effects of fibroblast growth factor-2 and Activin A on cellular differentiation into neural lineages, we examined the expression of neural differentiation markers in human embryonic stem cells treated with fibroblast growth factor-2 and/or Activin A at various concentrations in a growth factor-defined serum-free medium in short-term culture. In this study, we provide evidence that fibroblast growth factor-2 and Activin A synergistically regulated the initiation of human embryonic stem cell differentiation into neural cell lineages even though human embryonic stem cells autonomously differentiate into neural cell lineages.

  18. Ketamine-Induced Toxicity in Neurons Differentiated from Neural Stem Cells.

    PubMed

    Slikker, William; Liu, Fang; Rainosek, Shuo W; Patterson, Tucker A; Sadovova, Natalya; Hanig, Joseph P; Paule, Merle G; Wang, Cheng

    2015-10-01

    Ketamine is used as a general anesthetic, and recent data suggest that anesthetics can cause neuronal damage when exposure occurs during development. The precise mechanisms are not completely understood. To evaluate the degree of ketamine-induced neuronal toxicity, neural stem cells were isolated from gestational day 16 rat fetuses. On the eighth day in culture, proliferating neural stem cells were exposed for 24 h to ketamine at 1, 10, 100, and 500 μM. To determine the effect of ketamine on differentiated stem cells, separate cultures of neural stem cells were maintained in transition medium (DIV 6) for 1 day and kept in differentiation medium for another 3 days. Differentiated neural cells were exposed for 24 h to 10 μM ketamine. Markers of cellular proliferation and differentiation, mitochondrial health, cell death/damage, and oxidative damage were monitored to determine: (1) the effects of ketamine on neural stem cell proliferation and neural stem cell differentiation; (2) the nature and degree of ketamine-induced toxicity in proliferating neural stem cells and differentiated neural cells; and (3) to provide information regarding receptor expression and possible mechanisms underlying ketamine toxicity. After ketamine exposure at a clinically relevant concentration (10 μM), neural stem cell proliferation was not significantly affected and oxidative DNA damage was not induced. No significant effect on mitochondrial viability (3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay) in neural stem cell cultures (growth medium) was observed at ketamine concentrations up to 500 μM. However, quantitative analysis shows that the number of differentiated neurons was substantially reduced in 10 μM ketamine-exposed cultures in differentiation medium, compared with the controls. No significant changes in the number of GFAP-positive astrocytes and O4-positive oligodendrocytes (in differentiation medium) were detected from ketamine-exposed cultures

  19. Differentiation state determines neural effects on microvascular endothelial cells

    PubMed Central

    Muffley, Lara A.; Pan, Shin-Chen; Smith, Andria N.; Ga, Maricar; Hocking, Anne M.; Gibran, Nicole S.

    2012-01-01

    Growing evidence indicates that nerves and capillaries interact paracrinely in uninjured skin and cutaneous wounds. Although mature neurons are the predominant neural cell in the skin, neural progenitor cells have also been detected in uninjured adult skin. The aim of this study was to characterize differential paracrine effects of neural progenitor cells and mature sensory neurons on dermal microvascular endothelial cells. Our results suggest that neural progenitor cells and mature sensory neurons have unique secretory profiles and distinct effects on dermal microvascular endothelial cell proliferation, migration, and nitric oxide production. Neural progenitor cells and dorsal root ganglion neurons secrete different proteins related to angiogenesis. Specific to neural progenitor cells were dipeptidyl peptidase-4, IGFBP-2, pentraxin-3, serpin f1, TIMP-1, TIMP-4 and VEGF. In contrast, endostatin, FGF-1, MCP-1 and thrombospondin-2 were specific to dorsal root ganglion neurons. Microvascular endothelial cell proliferation was inhibited by dorsal root ganglion neurons but unaffected by neural progenitor cells. In contrast, microvascular endothelial cell migration in a scratch wound assay was inhibited by neural progenitor cells and unaffected by dorsal root ganglion neurons. In addition, nitric oxide production by microvascular endothelial cells was increased by dorsal root ganglion neurons but unaffected by neural progenitor cells. PMID:22683922

  20. Integrating Biomaterials and Stem Cells for Neural Regeneration.

    PubMed

    Maclean, Francesca L; Rodriguez, Alexandra L; Parish, Clare L; Williams, Richard J; Nisbet, David R

    2016-02-01

    The central nervous system has a limited capacity to regenerate, and thus, traumatic injuries or diseases often have devastating consequences. Therefore, there is a distinct need to develop alternative treatments that can achieve functional recovery without side effects currently observed with some pharmacological treatments. Combining biomaterials with pluripotent stem cells (PSCs), either embryonic or induced, has the potential to revolutionize the treatment of neurodegenerative diseases and traumatic injuries. Biomaterials can mimic the extracellular matrix and present a myriad of relevant biochemical cues through rational design or further functionalization. Biomaterials such as nanofibers and hydrogels, including self-assembling peptide (SAP) hydrogels can provide a superior cell culture environment. When these materials are then combined with PSCs, more accurate drug screening and disease modeling could be developed, and the generation of large number of cells with the appropriate phenotype can be achieved, for subsequent use in vitro. Biomaterials have also been shown to support endogenous cell growth after implantation, and, in particular, hydrogels and SAPs have effectively acted as cell delivery vehicles, increasing cell survival after transplantation. Few studies are yet to fully exploit the combination of PSCs and innovative biomaterials; however, initial studies with neural stem cells, for example, are promising, and, hence, such a combination for use in vitro and in vivo is an exciting new direction for the field of neural regeneration.

  1. Metabolic circuits in neural stem cells

    PubMed Central

    Kim, Do-Yeon; Rhee, Inmoo

    2015-01-01

    Metabolic activity indicative of cellular demand is emerging as a key player in cell fate decision. Numerous studies have demonstrated that diverse metabolic pathways have a critical role in the control of the proliferation, differentiation and quiescence of stem cells. The identification of neural stem/progenitor cells (NSPCs) and the characterization of their development and fate decision process have provided insight into the regenerative potential of the adult brain. As a result, the potential of NSPCs in cell replacement therapies for neurological diseases is rapidly growing. The aim of this review is to discuss the recent findings on the crosstalk among key regulators of NSPC development and the metabolic regulation crucial for the function and cell fate decisions of NSPCs. Fundamental understanding of the metabolic circuits in NSPCs may help to provide novel approaches for reactivating neurogenesis to treat degenerative brain conditions and cognitive decline. PMID:25037158

  2. Neural Stem Cells (NSCs) and Proteomics.

    PubMed

    Shoemaker, Lorelei D; Kornblum, Harley I

    2016-02-01

    Neural stem cells (NSCs) can self-renew and give rise to the major cell types of the CNS. Studies of NSCs include the investigation of primary, CNS-derived cells as well as animal and human embryonic stem cell (ESC)-derived and induced pluripotent stem cell (iPSC)-derived sources. NSCs provide a means with which to study normal neural development, neurodegeneration, and neurological disease and are clinically relevant sources for cellular repair to the damaged and diseased CNS. Proteomics studies of NSCs have the potential to delineate molecules and pathways critical for NSC biology and the means by which NSCs can participate in neural repair. In this review, we provide a background to NSC biology, including the means to obtain them and the caveats to these processes. We then focus on advances in the proteomic interrogation of NSCs. This includes the analysis of posttranslational modifications (PTMs); approaches to analyzing different proteomic compartments, such the secretome; as well as approaches to analyzing temporal differences in the proteome to elucidate mechanisms of differentiation. We also discuss some of the methods that will undoubtedly be useful in the investigation of NSCs but which have not yet been applied to the field. While many proteomics studies of NSCs have largely catalogued the proteome or posttranslational modifications of specific cellular states, without delving into specific functions, some have led to understandings of functional processes or identified markers that could not have been identified via other means. Many challenges remain in the field, including the precise identification and standardization of NSCs used for proteomic analyses, as well as how to translate fundamental proteomics studies to functional biology. The next level of investigation will require interdisciplinary approaches, combining the skills of those interested in the biochemistry of proteomics with those interested in modulating NSC function. © 2016 by The

  3. Neural Stem Cells (NSCs) and Proteomics*

    PubMed Central

    Shoemaker, Lorelei D.; Kornblum, Harley I.

    2016-01-01

    Neural stem cells (NSCs) can self-renew and give rise to the major cell types of the CNS. Studies of NSCs include the investigation of primary, CNS-derived cells as well as animal and human embryonic stem cell (ESC)-derived and induced pluripotent stem cell (iPSC)-derived sources. NSCs provide a means with which to study normal neural development, neurodegeneration, and neurological disease and are clinically relevant sources for cellular repair to the damaged and diseased CNS. Proteomics studies of NSCs have the potential to delineate molecules and pathways critical for NSC biology and the means by which NSCs can participate in neural repair. In this review, we provide a background to NSC biology, including the means to obtain them and the caveats to these processes. We then focus on advances in the proteomic interrogation of NSCs. This includes the analysis of posttranslational modifications (PTMs); approaches to analyzing different proteomic compartments, such the secretome; as well as approaches to analyzing temporal differences in the proteome to elucidate mechanisms of differentiation. We also discuss some of the methods that will undoubtedly be useful in the investigation of NSCs but which have not yet been applied to the field. While many proteomics studies of NSCs have largely catalogued the proteome or posttranslational modifications of specific cellular states, without delving into specific functions, some have led to understandings of functional processes or identified markers that could not have been identified via other means. Many challenges remain in the field, including the precise identification and standardization of NSCs used for proteomic analyses, as well as how to translate fundamental proteomics studies to functional biology. The next level of investigation will require interdisciplinary approaches, combining the skills of those interested in the biochemistry of proteomics with those interested in modulating NSC function. PMID:26494823

  4. Human neural stem cells promote proliferation of endogenous neural stem cells and enhance angiogenesis in ischemic rat brain.

    PubMed

    Ryu, Sun; Lee, Seung-Hoon; Kim, Seung U; Yoon, Byung-Woo

    2016-02-01

    Transplantation of human neural stem cells into the dentate gyrus or ventricle of rodents has been reportedly to enhance neurogenesis. In this study, we examined endogenous stem cell proliferation and angiogenesis in the ischemic rat brain after the transplantation of human neural stem cells. Focal cerebral ischemia in the rat brain was induced by middle cerebral artery occlusion. Human neural stem cells were transplanted into the subventricular zone. The behavioral performance of human neural stem cells-treated ischemic rats was significantly improved and cerebral infarct volumes were reduced compared to those in untreated animals. Numerous transplanted human neural stem cells were alive and preferentially localized to the ipsilateral ischemic hemisphere. Furthermore, 5-bromo-2'-deoxyuridine-labeled endogenous neural stem cells were observed in the subventricular zone and hippocampus, where they differentiated into cells immunoreactive for the neural markers doublecortin, neuronal nuclear antigen NeuN, and astrocyte marker glial fibrillary acidic protein in human neural stem cells-treated rats, but not in the untreated ischemic animals. The number of 5-bromo-2'-deoxyuridine-positive ⁄ anti-von Willebrand factor-positive proliferating endothelial cells was higher in the ischemic boundary zone of human neural stem cells-treated rats than in controls. Finally, transplantation of human neural stem cells in the brains of rats with focal cerebral ischemia promoted the proliferation of endogenous neural stem cells and their differentiation into mature neural-like cells, and enhanced angiogenesis. This study provides valuable insights into the effect of human neural stem cell transplantation on focal cerebral ischemia, which can be applied to the development of an effective therapy for stroke.

  5. Confetti clarifies controversy: neural crest stem cells are multipotent.

    PubMed

    Bronner, Marianne

    2015-03-05

    Neural crest precursors generate diverse cell lineages during development, which have been proposed to arise either from multipotent precursor cells or pools of heterogeneous, restricted progenitors. Now in Cell Stem Cell, Baggiolini et al. (2015) perform rigorous in vivo lineage tracing to show that individual neural crest precursors are multipotent.

  6. A conserved role for non-neural ectoderm cells in early neural development.

    PubMed

    Cajal, Marieke; Creuzet, Sophie E; Papanayotou, Costis; Sabéran-Djoneidi, Délara; Chuva de Sousa Lopes, Susana M; Zwijsen, An; Collignon, Jérôme; Camus, Anne

    2014-11-01

    During the early steps of head development, ectodermal patterning leads to the emergence of distinct non-neural and neural progenitor cells. The induction of the preplacodal ectoderm and the neural crest depends on well-studied signalling interactions between the non-neural ectoderm fated to become epidermis and the prospective neural plate. By contrast, the involvement of the non-neural ectoderm in the morphogenetic events leading to the development and patterning of the central nervous system has been studied less extensively. Here, we show that the removal of the rostral non-neural ectoderm abutting the prospective neural plate at late gastrulation stage leads, in mouse and chick embryos, to morphological defects in forebrain and craniofacial tissues. In particular, this ablation compromises the development of the telencephalon without affecting that of the diencephalon. Further investigations of ablated mouse embryos established that signalling centres crucial for forebrain regionalization, namely the axial mesendoderm and the anterior neural ridge, form normally. Moreover, changes in cell death or cell proliferation could not explain the specific loss of telencephalic tissue. Finally, we provide evidence that the removal of rostral tissues triggers misregulation of the BMP, WNT and FGF signalling pathways that may affect telencephalon development. This study opens new perspectives on the role of the neural/non-neural interface and reveals its functional relevance across higher vertebrates. © 2014. Published by The Company of Biologists Ltd.

  7. Adherent neural stem (NS) cells from fetal and adult forebrain.

    PubMed

    Pollard, Steven M; Conti, Luciano; Sun, Yirui; Goffredo, Donato; Smith, Austin

    2006-07-01

    Stable in vitro propagation of central nervous system (CNS) stem cells would offer expanded opportunities to dissect basic molecular, cellular, and developmental processes and to model neurodegenerative disease. CNS stem cells could also provide a source of material for drug discovery assays and cell replacement therapies. We have recently reported the generation of adherent, symmetrically expandable, neural stem (NS) cell lines derived both from mouse and human embryonic stem cells and from fetal forebrain (Conti L, Pollard SM, Gorba T, Reitano E, Toselli M, Biella G, Sun Y, Sanzone S, Ying QL, Cattaneo E, Smith A. 2005. Niche-independent symmetrical self-renewal of a mammalian tissue stem cell. PLoS Biol 3(9):e283). These NS cells retain neuronal and glial differentiation potential after prolonged passaging and are transplantable. NS cells are likely to comprise the resident stem cell population within heterogeneous neurosphere cultures. Here we demonstrate that similar NS cell cultures can be established from the adult mouse brain. We also characterize the growth factor requirements for NS cell derivation and self-renewal. We discuss our current understanding of the relationship of NS cell lines to physiological progenitor cells of fetal and adult CNS.

  8. Rhesus monkey neural stem cell transplantation promotes neural regeneration in rats with hippocampal lesions.

    PubMed

    Ye, Li-Juan; Bian, Hui; Fan, Yao-Dong; Wang, Zheng-Bo; Yu, Hua-Lin; Ma, Yuan-Ye; Chen, Feng

    2016-09-01

    Rhesus monkey neural stem cells are capable of differentiating into neurons and glial cells. Therefore, neural stem cell transplantation can be used to promote functional recovery of the nervous system. Rhesus monkey neural stem cells (1 × 10(5) cells/μL) were injected into bilateral hippocampi of rats with hippocampal lesions. Confocal laser scanning microscopy demonstrated that green fluorescent protein-labeled transplanted cells survived and grew well. Transplanted cells were detected at the lesion site, but also in the nerve fiber-rich region of the cerebral cortex and corpus callosum. Some transplanted cells differentiated into neurons and glial cells clustering along the ventricular wall, and integrated into the recipient brain. Behavioral tests revealed that spatial learning and memory ability improved, indicating that rhesus monkey neural stem cells noticeably improve spatial learning and memory abilities in rats with hippocampal lesions.

  9. On becoming neural: what the embryo can tell us about differentiating neural stem cells

    PubMed Central

    Moody, Sally A; Klein, Steven L; Karpinski, Beverley A; Maynard, Thomas M; LaMantia, Anthony-Samuel

    2013-01-01

    The earliest steps of embryonic neural development are orchestrated by sets of transcription factors that control at least three processes: the maintenance of proliferative, pluripotent precursors that expand the neural ectoderm; their transition to neurally committed stem cells comprising the neural plate; and the onset of differentiation of neural progenitors. The transition from one step to the next requires the sequential activation of each gene set and then its down-regulation at the correct developmental times. Herein, we review how these gene sets interact in a transcriptional network to regulate these early steps in neural development. A key gene in this regulatory network is FoxD4L1, a member of the forkhead box (Fox) family of transcription factors. Knock-down experiments in Xenopus embryos show that FoxD4L1 is required for the expression of the other neural transcription factors, whereas increased FoxD4L1 levels have three different effects on these genes: up-regulation of neural ectoderm precursor genes; transient down-regulation of neural plate stem cell genes; and down-regulation of neural progenitor differentiation genes. These different effects indicate that FoxD4L1 maintains neural ectodermal precursors in an immature, proliferative state, and counteracts premature neural stem cell and neural progenitor differentiation. Because it both up-regulates and down-regulates genes, we characterized the regions of the FoxD4L1 protein that are specifically involved in these transcriptional functions. We identified a transcriptional activation domain in the N-terminus and at least two domains in the C-terminus that are required for transcriptional repression. These functional domains are highly conserved in the mouse and human homologues. Preliminary studies of the related FoxD4 gene in cultured mouse embryonic stem cells indicate that it has a similar role in promoting immature neural ectodermal precursors and delaying neural progenitor differentiation

  10. Interleukin 6-preconditioned neural stem cells reduce ischaemic injury in stroke mice.

    PubMed

    Sakata, Hiroyuki; Narasimhan, Purnima; Niizuma, Kuniyasu; Maier, Carolina M; Wakai, Takuma; Chan, Pak H

    2012-11-01

    Transplantation of neural stem cells provides a promising therapy for stroke. Its efficacy, however, might be limited because of massive grafted-cell death after transplantation, and its insufficient capability for tissue repair. Interleukin 6 is a pro-inflammatory cytokine involved in the pathogenesis of various neurological disorders. Paradoxically, interleukin 6 promotes a pro-survival signalling pathway through activation of signal transducer and activator of transcription 3. In this study, we investigated whether cellular reprogramming of neural stem cells with interleukin 6 facilitates the effectiveness of cell transplantation therapy in ischaemic stroke. Neural stem cells harvested from the subventricular zone of foetal mice were preconditioned with interleukin 6 in vitro and transplanted into mouse brains 6 h or 7 days after transient middle cerebral artery occlusion. Interleukin 6 preconditioning protected the grafted neural stem cells from ischaemic reperfusion injury through signal transducer and activator of transcription 3-mediated upregulation of manganese superoxide dismutase, a primary mitochondrial antioxidant enzyme. In addition, interleukin 6 preconditioning induced secretion of vascular endothelial growth factor from the neural stem cells through activation of signal transducer and activator of transcription 3, resulting in promotion of angiogenesis in the ischaemic brain. Furthermore, transplantation of interleukin 6-preconditioned neural stem cells significantly attenuated infarct size and improved neurological performance compared with non-preconditioned neural stem cells. This interleukin 6-induced amelioration of ischaemic insults was abolished by transfecting the neural stem cells with signal transducer and activator of transcription 3 small interfering RNA before transplantation. These results indicate that preconditioning with interleukin 6, which reprograms neural stem cells to tolerate oxidative stress after ischaemic reperfusion

  11. Fas Activation Increases Neural progenitor Cell Survival

    PubMed Central

    Knight, Julia C.; Scharf, Eugene L.; Mao-Draayer, Yang

    2015-01-01

    Although there is a sizable amount of research focusing on adult neural progenitor cells (NPCs) as a therapeutic approach for many neurodegenerative diseases, including multiple sclerosis, little is known about the pathways that govern NPC survival and apoptosis. Fas, a member of the death receptor superfamily, plays a well-characterized role in the immune system, but its function in neural stem cells remains uncertain. Our study focuses on the effects of Fas on NPC survival in vitro. Activation of Fas by recombinant Fas ligand (FasL) did not induce apoptosis in murine NPCs in culture. In fact, both an increase in the amount of viable cells and a decrease in apoptotic and dying cells were observed with FasL treatment. Our data indicate that FasL-mediated adult NPC neuroprotection is characterized by a reduction in apoptosis, but not increased proliferation. Further investigation of this effect revealed that the antiapoptotic effects of FasL are mediated by the up-regulation of Birc3, an inhibitor of apoptosis protein (IAP). Conversely, the observed effect is not the result of altered caspase activation or FLIP (Fas-associated death domain-like interleukin-1beta-converting enzyme inhibitory protein) up-regulation, which is known to inhibit caspase-8-mediated cell death in T cells. Our data indicate that murine adult NPCs are resistant to FasL-induced cell death. Activation of Fas increased cell survival by decreasing apoptosis through Birc3 up-regulation. These results describe a novel pathway involved in NPC survival. PMID:19830835

  12. When cells become depressed: focus on neural stem cells in novel treatment strategies against depression.

    PubMed

    Benninghoff, J; Schmitt, A; Mössner, R; Lesch, K-P

    2002-05-01

    Clinical neuroscience enters a new era in understanding the pathophysiology of depressive illness and the mode of action of antidepressant therapy. While elucidation of factors that lead to depression is still in its infancy, biochemical malfunctions appear to have well defined morphological correlations, especially in the hippocampus. Hippocampus is one of the main sites in the brain habouring neural stem cells. Cytokines and neurotrophic factors like brain-derived neurotrophic factor (BDNF) play a pivotal role in neural plasticity and potentially influence growth and migration of these progenitors. Not surprisingly, antidepressant drugs interfering with neurotransmitters such as serotonin (5-HT) influence neurotrophins like BDNF, since 5-HT homeostasis is essential for brain development, neurogenesis, and neuroplasticity as well as complex behavior. In this review, the new area of neural stem cell research and the avenues of ongoing and future research sustaining the development of novel treatments for depression will be explored.

  13. GBM secretome induces transient transformation of human neural precursor cells.

    PubMed

    Venugopal, Chitra; Wang, X Simon; Manoranjan, Branavan; McFarlane, Nicole; Nolte, Sara; Li, Meredith; Murty, Naresh; Siu, K W Michael; Singh, Sheila K

    2012-09-01

    Glioblastoma (GBM) is the most aggressive primary brain tumor in humans, with a uniformly poor prognosis. The tumor microenvironment is composed of both supportive cellular substrates and exogenous factors. We hypothesize that exogenous factors secreted by brain tumor initiating cells (BTICs) could predispose normal neural precursor cells (NPCs) to transformation. When NPCs are grown in GBM-conditioned media, and designated as "tumor-conditioned NPCs" (tcNPCs), they become highly proliferative and exhibit increased stem cell self-renewal, or the unique ability of stem cells to asymmetrically generate another stem cell and a daughter cell. tcNPCs also show an increased transcript level of stem cell markers such as CD133 and ALDH and growth factor receptors such as VEGFR1, VEGFR2, EGFR and PDGFRα. Media analysis by ELISA of GBM-conditioned media reveals an elevated secretion of growth factors such as EGF, VEGF and PDGF-AA when compared to normal neural stem cell-conditioned media. We also demonstrate that tcNPCs require prolonged or continuous exposure to the GBM secretome in vitro to retain GBM BTIC characteristics. Our in vivo studies reveal that tcNPCs are unable to form tumors, confirming that irreversible transformation events may require sustained or prolonged presence of the GBM secretome. Analysis of GBM-conditioned media by mass spectrometry reveals the presence of secreted proteins Chitinase-3-like 1 (CHI3L1) and H2A histone family member H2AX. Collectively, our data suggest that GBM-secreted factors are capable of transiently altering normal NPCs, although for retention of the transformed phenotype, sustained or prolonged secretome exposure or additional transformation events are likely necessary.

  14. Production of chick embryo extract for the cultivation of murine neural crest stem cells.

    PubMed

    Pajtler, Kristian; Bohrer, Anna; Maurer, Jochen; Schorle, Hubert; Schramm, Alexander; Eggert, Angelika; Schulte, Johannes Hubertus

    2010-11-27

    The neural crest arises from the neuro-ectoderm during embryogenesis and persists only temporarily. Early experiments already proofed pluripotent progenitor cells to be an integral part of the neural crest(1). Phenotypically, neural crest stem cells (NCSC) are defined by simultaneously expressing p75 (low-affine nerve growth factor receptor, LNGFR) and SOX10 during their migration from the neural crest(2,3,4,5). These progenitor cells can differentiate into smooth muscle cells, chromaffin cells, neurons and glial cells, as well as melanocytes, cartilage and bone(6,7,8,9). To cultivate NCSC in vitro, a special neural crest stem cell medium (NCSCM) is required(10). The most complex part of the NCSCM is the preparation of chick embryo extract (CEE) representing an essential source of growth factors for the NCSC as well as for other types of neural explants. Other NCSCM ingredients beside CEE are commercially available. Producing CCE using laboratory standard equipment it is of high importance to know about the challenging details as the isolation, maceration, centrifugation, and filtration processes. In this protocol we describe accurate techniques to produce a maximized amount of pure and high quality CEE.

  15. The Neural Cell Adhesion Molecule (NCAM) Promotes Clustering and Activation of EphA3 Receptors in GABAergic Interneurons to Induce Ras Homolog Gene Family, Member A (RhoA)/Rho-associated protein kinase (ROCK)-mediated Growth Cone Collapse.

    PubMed

    Sullivan, Chelsea S; Kümper, Maike; Temple, Brenda S; Maness, Patricia F

    2016-12-16

    Establishment of a proper balance of excitatory and inhibitory connectivity is achieved during development of cortical networks and adjusted through synaptic plasticity. The neural cell adhesion molecule (NCAM) and the receptor tyrosine kinase EphA3 regulate the perisomatic synapse density of inhibitory GABAergic interneurons in the mouse frontal cortex through ephrin-A5-induced growth cone collapse. In this study, it was demonstrated that binding of NCAM and EphA3 occurred between the NCAM Ig2 domain and EphA3 cysteine-rich domain (CRD). The binding interface was further refined through molecular modeling and mutagenesis and shown to be comprised of complementary charged residues in the NCAM Ig2 domain (Arg-156 and Lys-162) and the EphA3 CRD (Glu-248 and Glu-264). Ephrin-A5 induced co-clustering of surface-bound NCAM and EphA3 in GABAergic cortical interneurons in culture. Receptor clustering was impaired by a charge reversal mutation that disrupted NCAM/EphA3 association, emphasizing the importance of the NCAM/EphA3 binding interface for cluster formation. NCAM enhanced ephrin-A5-induced EphA3 autophosphorylation and activation of RhoA GTPase, indicating a role for NCAM in activating EphA3 signaling through clustering. NCAM-mediated clustering of EphA3 was essential for ephrin-A5-induced growth cone collapse in cortical GABAergic interneurons, and RhoA and a principal effector, Rho-associated protein kinase, mediated the collapse response. This study delineates a mechanism in which NCAM promotes ephrin-A5-dependent clustering of EphA3 through interaction of the NCAM Ig2 domain and the EphA3 CRD, stimulating EphA3 autophosphorylation and RhoA signaling necessary for growth cone repulsion in GABAergic interneurons in vitro, which may extend to remodeling of axonal terminals of interneurons in vivo.

  16. Neural stem cells in lead toxicity.

    PubMed

    Chen, W-W; Zhang, X; Huang, W-J

    2016-12-01

    Lead (Pb) exposure in the early stages of neurodevelopment results in long-lasting alterations that ultimately cognitive function and behaviour. The prime targets of lead toxicity are the multipotent neural stem cells (NSCs). The present review will discuss the basic molecular physiology involved in the toxicity mechanisms induced by lead and its resultant counter effects on nervous system and physiology. The article shall help researchers working in the area to design new drugs and therapeutics for the efficient management of neuro-toxic states especially upon prenatal exposure to lead.

  17. Dynamic transcriptional signature and cell fate analysis reveals plasticity of individual neural plate border cells.

    PubMed

    Roellig, Daniela; Tan-Cabugao, Johanna; Esaian, Sevan; Bronner, Marianne E

    2017-03-29

    The 'neural plate border' of vertebrate embryos contains precursors of neural crest and placode cells, both defining vertebrate characteristics. How these lineages segregate from neural and epidermal fates has been a matter of debate. We address this by performing a fine-scale quantitative temporal analysis of transcription factor expression in the neural plate border of chick embryos. The results reveal significant overlap of transcription factors characteristic of multiple lineages in individual border cells from gastrula through neurula stages. Cell fate analysis using a Sox2 (neural) enhancer reveals that cells that are initially Sox2+ cells can contribute not only to neural tube but also to neural crest and epidermis. Moreover, modulating levels of Sox2 or Pax7 alters the apportionment of neural tube versus neural crest fates. Our results resolve a long-standing question and suggest that many individual border cells maintain ability to contribute to multiple ectodermal lineages until or beyond neural tube closure.

  18. Differentiation of nonhuman primate embryonic stem cells along neural lineages

    PubMed Central

    Kuai, Xiao Ling; Gagliardi, Christine; Flaat, Mette; Bunnell, Bruce A.

    2009-01-01

    The differentiation of embryonic stem cells (ESCs) into neurons and glial cells represents a promising cell-based therapy for neurodegenerative diseases. Because the rhesus macaque is physiologically and phylogenetically similar to humans, it is a clinically relevant animal model for ESC research. In this study, the pluripotency and neural differentiation potential of a rhesus monkey ESC line (ORMES6) was investigated. ORMES6 was derived from an in vitro produced blastocyst, which is the same way human ESCs have been derived. ORMES6 stably expressed the embryonic transcription factors POU5F1 (Oct4), Sox2 and NANOG. Stage-specific embryonic antigen 4 (SSEA 4) and the glycoproteins TRA-1-60 and TRA-1-81 were also expressed. The embryoid bodies (EBs) formed from ORMES6 ESCs spontaneously gave rise to cells of three germ layers. After exposure to basic fibroblast growth factor (bFGF) for 14–16 days, columnar rosette cells formed in the EB outgrowths. Sox2, microtubule-associated protein (MAP2), β-tublinIII and glial fibrillary acidic protein (GFAP) genes and Nestin, FoxD3, Pax6 and β-tublinIII antigens were expressed in the rosette cells. Oct4 and NANOG expression were remarkably down-regulated in these cells. After removal of bFGF from the medium, the rosette cells differentiated along neural lineages. The differentiated cells expressed MAP2, β-tublinIII, Neuro D and GFAP genes. Most differentiated cells expressed early neuron-specific antigen β-tublinIII (73±4.7%) and some expressed intermediate neuron antigen MAP2 (18±7.2%). However, some differentiated cells expressed the glial cell antigens A2B5 (7.17%±1.2%), GFAP (4.93±1.9%), S100 (7±3.5%) and O4 (0.2±70.2%). The rosette cells were transplanted into the striatum of immune-deficient NIHIII mice. The cells persisted for approximately 2 weeks and expressed Ki67, NeuN, MAP2 and GFAP. These results demonstrate that the rhesus monkey ESC line ORMES6 retains the pluripotent characteristics of ESCs and can be

  19. Phylogeny of a neural cell adhesion molecule.

    PubMed

    Hall, A K; Rutishauser, U

    1985-07-01

    The phylogeny of adhesion among cells derived from neural tissue has been examined using a combination of functional and immunological analyses. The presence of the neural cell adhesion molecule (NCAM) was evaluated with respect to NCAM-specific antigenic determinants attached to a polypeptide chain with appropriate electrophoretic properties. By these criteria, NCAM-like molecules were detected in all embryonic and adult vertebrates tested, and an adult mollusc, but not in an adult insect, crustacean, or nematode. The functional assays measured adhesiveness by simple aggregation of neural membrane vesicles, as well as by NCAM-specific binding between membranes from different species. The presence of the NCAM antigen in vertebrate membranes correlated with binding activity in both the NCAM-specific and general adhesion assays, implying that the adhesiveness of these membranes largely reflects NCAM-mediated binding. The results also indicate that NCAM function has been conserved during the evolution of vertebrates, and supports the possibility that mechanisms of nerve-nerve, nerve-muscle, and nerve-glial interaction, which have been demonstrated previously to involve NCAM, may be similar for many chordates. Whereas NCAM was not detected in adult fly and worm, these species did express NCAM-like antigens transiently during early development. These results are consistent with the hypothesis that NCAM is required during several periods of development, and that the functions of this molecule in nematodes and insects may be distinct from or a subset of those that occur in vertebrates. The expanded role of the molecule represented by its expression during later stages of vertebrate development may thus have been an important contribution to the evolution of chordates.

  20. Folate receptor alpha is necessary for neural plate cell apical constriction during Xenopus neural tube formation.

    PubMed

    Balashova, Olga A; Visina, Olesya; Borodinsky, Laura N

    2017-03-02

    Folate supplementation prevents up to 70% of neural tube defects (NTDs), which result from a failure of neural tube closure during embryogenesis. The elucidation of the mechanisms underlying folate action has been challenging. This study introduces Xenopus laevis as a model to determine the cellular and molecular mechanisms involved in folate action during neural tube formation. We show that knockdown of folate receptor-α (FRα) impairs neural tube formation and leads to NTDs. FRα knockdown in neural plate cells only is necessary and sufficient to induce NTDs. FRα-deficient neural plate cells fail to constrict, resulting in widening of the neural plate midline and defective neural tube closure. Pharmacological inhibition of folate action by methotrexate during neurulation induces NTDs by inhibiting folate interaction with its uptake systems. Our findings support a model for folate receptor interacting with cell adhesion molecules, thus regulating apical cell membrane remodeling and cytoskeletal dynamics necessary for neural plate folding. Further studies in this organism may unveil novel cellular and molecular events mediated by folate and lead to new means for preventing NTDs.

  1. Neural stem cell differentiation in a cell-collagen-bioreactor culture system.

    PubMed

    Lin, Hsingchi J; O'Shaughnessy, Thomas J; Kelly, Jeremy; Ma, Wu

    2004-11-25

    Neural stem cells and neural progenitors (NSCs/NPs) are capable of self-renewal and can give rise to both neurons and glia. Such cells have been isolated from the embryonic brain and immobilized in three dimensional collagen gels. The collagen-entrapped NSCs/NPs recapitulate CNS stem cell development and form functional synapses and neuronal circuits. However, the cell-collagen constructs from static conditions contain hypoxic, necrotic cores and the cells are short-lived. In the present study, NSCs/NPs isolated from embryonic day 13 rat cortical neuroepithelium are immobilized in type I collagen gels and cultured in NASA-designed rotating wall vessel (RWV) bioreactors for up to 9 weeks. Initially, during the first 2 weeks of culture, a lag phase of cellular growth and differentiation is observed in the RWV bioreactors. Accelerated growth and differentiation, with the cells beginning to form large aggregates (approximately 1 mm in diameter) without death cores, begins during the third week. The collagen-entrapped NSCs/NPs cultured in RWV show active neuronal generation followed by astrocyte production. After 6 weeks in rotary culture, the cell-collagen constructs contain over 10 fold greater nestin+ and GFAP+ cells and two-fold more TuJ1 gene expression than those found in static cultures. In addition, TuJ1+ neurons in RWV culture give rise to extensive neurite outgrowth and considerably more synapsin I+ pre-synaptic puncta surrounding MAP2+ cell bodies and dendrites. These results strongly suggest that the cell-collagen-bioreactor culture system supports long-term NSC/NP growth and differentiation, and RWV bioreactors can be useful in generating neural tissue like constructs, which may have the potential for cell replacement therapy.

  2. Presenilin-1 regulates neural progenitor cell differentiation in the adult brain

    PubMed Central

    Gadadhar, Archana; Marr, Robert; Lazarov, Orly

    2011-01-01

    Presenilin-1 (PS1) is the catalytic core of the aspartyl protease γ-secretase. Previous genetic studies using germ-line deletion of PS1 and conditional knockout mice demonstrated that PS1 plays an essential role in neuronal differentiation during neural development, but it remained unclear whether PS1 plays a similar role in neurogenesis in the adult brain. Here we show that neural progenitor cells infected with lentiviral vectors expressing short interfering RNA (siRNA) for the exclusive knockdown of PS1 in the neurogenic microenvironments, exhibit a dramatic enhancement of cell differentiation. Infected cells differentiated into neurons, astrocytes and oligodendrocytes, suggesting that multipotentiality of neural progenitor cells is not affected by reduced levels of PS1. Neurosphere cultures treated with γ-secretase inhibitors exhibit a similar phenotype of enhanced cell differentiation, suggesting that PS1 function in neural progenitor cells is γ-secretase-dependent. Neurospheres infected with lentiviral vectors expressing siRNA for the targeting of PS1 differentiated even in the presence of the proliferation factors epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF), suggesting that PS1 dominates EFG and bFGF signaling pathways. Reduction of PS1 expression in neural progenitor cells was accompanied by a decrease in epidermal growth factor receptor (EGFR) and β-catenin expression level, suggesting that they are downstream essential transducers of PS1 signaling in adult neural progenitor cells. These findings suggest a physiological role for PS1 in adult neurogenesis, and a potential target for the manipulation of neural progenitor cell differentiation. PMID:21325529

  3. Neural crest cell-derived VEGF promotes embryonic jaw extension

    PubMed Central

    Wiszniak, Sophie; Mackenzie, Francesca E.; Anderson, Peter; Kabbara, Samuela; Ruhrberg, Christiana; Schwarz, Quenten

    2015-01-01

    Jaw morphogenesis depends on the growth of Meckel’s cartilage during embryogenesis. However, the cell types and signals that promote chondrocyte proliferation for Meckel’s cartilage growth are poorly defined. Here we show that neural crest cells (NCCs) and their derivatives provide an essential source of the vascular endothelial growth factor (VEGF) to enhance jaw vascularization and stabilize the major mandibular artery. We further show in two independent mouse models that blood vessels promote Meckel’s cartilage extension. Coculture experiments of arterial tissue with NCCs or chondrocytes demonstrated that NCC-derived VEGF promotes blood vessel growth and that blood vessels secrete factors to instruct chondrocyte proliferation. Computed tomography and X-ray scans of patients with hemifacial microsomia also showed that jaw hypoplasia correlates with mandibular artery dysgenesis. We conclude that cranial NCCs and their derivatives provide an essential source of VEGF to support blood vessel growth in the developing jaw, which in turn is essential for normal chondrocyte proliferation, and therefore jaw extension. PMID:25922531

  4. Role of growth hormone (GH) in the treatment on neural diseases: from neuroprotection to neural repair.

    PubMed

    Arce, Víctor M; Devesa, Pablo; Devesa, Jesús

    2013-08-01

    Growth hormone (GH) is a pleiotropic hormone that exerts important functions in the control of brain development as well as in the regulation neuronal differentiation and function, together with several behavioral and psychological effects that have been linked to its modulatory actions on brain neurotransmitters. In addition, the possibility that GH may play a role on brain repair after injury has been also envisaged, and a number of reports have shown that GH administration following injury confers neuroprotection and accelerates the recovery of some neural functions. In this review we have analyzed the state of the art of GH administration in several neural diseases. Though more studies are still necessary in order to completely understand the importance of GH in these processes, the promising results obtained so far, together with the absence of untoward effects during GH therapy, encourages the development of clinical assays in order to further support the use GH treatment in neural diseases in which neuroprotection and/or neuroregeneration are involved. Copyright © 2013. Published by Elsevier Ireland Ltd.

  5. Cerium oxide nanoparticles inhibit differentiation of neural stem cells.

    PubMed

    Gliga, Anda R; Edoff, Karin; Caputo, Fanny; Källman, Thomas; Blom, Hans; Karlsson, Hanna L; Ghibelli, Lina; Traversa, Enrico; Ceccatelli, Sandra; Fadeel, Bengt

    2017-08-24

    Cerium oxide nanoparticles (nanoceria) display antioxidant properties and have shown cytoprotective effects both in vitro and in vivo. Here, we explored the effects of nanoceria on neural progenitor cells using the C17.2 murine cell line as a model. First, we assessed the effects of nanoceria versus samarium (Sm) doped nanoceria on cell viability in the presence of the prooxidant, DMNQ. Both particles were taken up by cells and nanoceria, but not Sm-doped nanoceria, elicited a temporary cytoprotective effect upon exposure to DMNQ. Next, we employed RNA sequencing to explore the transcriptional responses induced by nanoceria or Sm-doped nanoceria during neuronal differentiation. Detailed computational analyses showed that nanoceria altered pathways and networks relevant for neuronal development, leading us to hypothesize that nanoceria inhibits neuronal differentiation, and that nanoceria and Sm-doped nanoceria both interfere with cytoskeletal organization. We confirmed that nanoceria reduced neuron specific β3-tubulin expression, a marker of neuronal differentiation, and GFAP, a neuroglial marker. Furthermore, using super-resolution microscopy approaches, we could show that both particles interfered with cytoskeletal organization and altered the structure of neural growth cones. Taken together, these results reveal that nanoceria may impact on neuronal differentiation, suggesting that nanoceria could pose a developmental neurotoxicity hazard.

  6. Chondroitin Sulfate Impairs Neural Stem Cell Migration Through ROCK Activation.

    PubMed

    Galindo, Layla T; Mundim, Mayara T V V; Pinto, Agnes S; Chiarantin, Gabrielly M D; Almeida, Maíra E S; Lamers, Marcelo L; Horwitz, Alan R; Santos, Marinilce F; Porcionatto, Marimelia

    2017-05-05

    Brain injuries such as trauma and stroke lead to glial scar formation by reactive astrocytes which produce and secret axonal outgrowth inhibitors. Chondroitin sulfate proteoglycans (CSPG) constitute a well-known class of extracellular matrix molecules produced at the glial scar and cause growth cone collapse. The CSPG glycosaminoglycan side chains composed of chondroitin sulfate (CS) are responsible for its inhibitory activity on neurite outgrowth and are dependent on RhoA activation. Here, we hypothesize that CSPG also impairs neural stem cell migration inhibiting their penetration into an injury site. We show that DCX+ neuroblasts do not penetrate a CSPG-rich injured area probably due to Nogo receptor activation and RhoA/ROCK signaling pathway as we demonstrate in vitro with neural stem cells cultured as neurospheres and pull-down for RhoA. Furthermore, CS-impaired cell migration in vitro induced the formation of large mature adhesions and altered cell protrusion dynamics. ROCK inhibition restored migration in vitro as well as decreased adhesion size.

  7. [Comparisons among different methods of culturing neural stem cells isolated from human fetal cortex].

    PubMed

    Ren, Ping; Guan, Yun Qian; Zhang, Yu

    2007-02-01

    Neural stem cells (NSCs) are proved to be promising cell sources for gene therapy and cell therapy. To pursue optimal conditions for the isolation and culture of neural stem cells residing in human fetal cortex,the cortical tissue was dissociated mechanically and digested with various enzymes. Short-term trypsin digestion combined with pipette dissociation proved to be the suitable method of isolating human NSCs derived from the fetal cortex. Furthermore,DMEM/F12 medium was superior to the neurobasal medium in the aspect of clonal formation. In repeated dissociation experiments,it was found that accutase, instead of trypsin,endowed the NSCs with better growth and efficient neurosphere formation.

  8. REN: a novel, developmentally regulated gene that promotes neural cell differentiation.

    PubMed

    Gallo, Rita; Zazzeroni, Francesca; Alesse, Edoardo; Mincione, Claudia; Borello, Ugo; Buanne, Pasquale; D'Eugenio, Roberta; Mackay, Andrew R; Argenti, Beatrice; Gradini, Roberto; Russo, Matteo A; Maroder, Marella; Cossu, Giulio; Frati, Luigi; Screpanti, Isabella; Gulino, Alberto

    2002-08-19

    Expansion and fate choice of pluripotent stem cells along the neuroectodermal lineage is regulated by a number of signals, including EGF, retinoic acid, and NGF, which also control the proliferation and differentiation of central nervous system (CNS) and peripheral nervous system (PNS) neural progenitor cells. We report here the identification of a novel gene, REN, upregulated by neurogenic signals (retinoic acid, EGF, and NGF) in pluripotent embryonal stem (ES) cells and neural progenitor cell lines in association with neurotypic differentiation. Consistent with a role in neural promotion, REN overexpression induced neuronal differentiation as well as growth arrest and p27Kip1 expression in CNS and PNS neural progenitor cell lines, and its inhibition impaired retinoic acid induction of neurogenin-1 and NeuroD expression. REN expression is developmentally regulated, initially detected in the neural fold epithelium of the mouse embryo during gastrulation, and subsequently throughout the ventral neural tube, the outer layer of the ventricular encephalic neuroepithelium and in neural crest derivatives including dorsal root ganglia. We propose that REN represents a novel component of the neurogenic signaling cascade induced by retinoic acid, EGF, and NGF, and is both a marker and a regulator of neuronal differentiation.

  9. REST regulation of gene networks in adult neural stem cells.

    PubMed

    Mukherjee, Shradha; Brulet, Rebecca; Zhang, Ling; Hsieh, Jenny

    2016-11-07

    Adult hippocampal neural stem cells generate newborn neurons throughout life due to their ability to self-renew and exist as quiescent neural progenitors (QNPs) before differentiating into transit-amplifying progenitors (TAPs) and newborn neurons. The mechanisms that control adult neural stem cell self-renewal are still largely unknown. Conditional knockout of REST (repressor element 1-silencing transcription factor) results in precocious activation of QNPs and reduced neurogenesis over time. To gain insight into the molecular mechanisms by which REST regulates adult neural stem cells, we perform chromatin immunoprecipitation sequencing and RNA-sequencing to identify direct REST target genes. We find REST regulates both QNPs and TAPs, and importantly, ribosome biogenesis, cell cycle and neuronal genes in the process. Furthermore, overexpression of individual REST target ribosome biogenesis or cell cycle genes is sufficient to induce activation of QNPs. Our data define novel REST targets to maintain the quiescent neural stem cell state.

  10. REST regulation of gene networks in adult neural stem cells

    PubMed Central

    Mukherjee, Shradha; Brulet, Rebecca; Zhang, Ling; Hsieh, Jenny

    2016-01-01

    Adult hippocampal neural stem cells generate newborn neurons throughout life due to their ability to self-renew and exist as quiescent neural progenitors (QNPs) before differentiating into transit-amplifying progenitors (TAPs) and newborn neurons. The mechanisms that control adult neural stem cell self-renewal are still largely unknown. Conditional knockout of REST (repressor element 1-silencing transcription factor) results in precocious activation of QNPs and reduced neurogenesis over time. To gain insight into the molecular mechanisms by which REST regulates adult neural stem cells, we perform chromatin immunoprecipitation sequencing and RNA-sequencing to identify direct REST target genes. We find REST regulates both QNPs and TAPs, and importantly, ribosome biogenesis, cell cycle and neuronal genes in the process. Furthermore, overexpression of individual REST target ribosome biogenesis or cell cycle genes is sufficient to induce activation of QNPs. Our data define novel REST targets to maintain the quiescent neural stem cell state. PMID:27819263

  11. Regulation of viability, differentiation and death of human melanoma cells carrying neural stem cell biomarkers: a possibility for neural trans-differentiation.

    PubMed

    Ivanov, Vladimir N; Hei, Tom K

    2015-07-01

    During embryonic development, melanoblasts, the precursors of melanocytes, emerge from a subpopulation of the neural crest stem cells and migrate to colonize skin. Melanomas arise during melanoblast differentiation into melanocytes and from young proliferating melanocytes through somatic mutagenesis and epigenetic regulations. In the present study, we used several human melanoma cell lines from the sequential phases of melanoma development (radial growth phase, vertical growth phase and metastatic phase) to compare: (i) the frequency and efficiency of the induction of cell death via apoptosis and necroptosis; (ii) the presence of neural and cancer stem cell biomarkers as well as death receptors, DR5 and FAS, in both adherent and spheroid cultures of melanoma cells; (iii) anti-apoptotic effects of the endogenous production of cytokines and (iv) the ability of melanoma cells to perform neural trans-differentiation. We demonstrated that programed necrosis or necroptosis, could be induced in two metastatic melanoma lines, FEMX and OM431, while the mitochondrial pathway of apoptosis was prevalent in a vast majority of melanoma lines. All melanoma lines used in the current study expressed substantial levels of pluripotency markers, SOX2 and NANOG. There was a trend for increasing expression of Nestin, an early neuroprogenitor marker, during melanoma progression. Most of the melanoma lines, including WM35, FEMX and A375, can grow as a spheroid culture in serum-free media with supplements. It was possible to induce neural trans-differentiation of 1205Lu and OM431 melanoma cells in serum-free media supplemented with insulin. This was confirmed by the expression of neuronal markers, doublecortin and β3-Tubulin, by significant growth of neurites and by the negative regulation of this process by a dominant-negative Rac1N17. These results suggest a relative plasticity of differentiated melanoma cells and a possibility for their neural trans-differentiation without the

  12. Slit/Robo1 signaling regulates neural tube development by balancing neuroepithelial cell proliferation and differentiation

    SciTech Connect

    Wang, Guang; Li, Yan; Wang, Xiao-yu; Han, Zhe; Chuai, Manli; Wang, Li-jing; Ho Lee, Kenneth Ka; Geng, Jian-guo; Yang, Xuesong

    2013-05-01

    Formation of the neural tube is the morphological hallmark for development of the embryonic central nervous system (CNS). Therefore, neural tube development is a crucial step in the neurulation process. Slit/Robo signaling was initially identified as a chemo-repellent that regulated axon growth cone elongation, but its role in controlling neural tube development is currently unknown. To address this issue, we investigated Slit/Robo1 signaling in the development of chick neCollege of Life Sciences Biocentre, University of Dundee, Dundee DD1 5EH, UKural tube and transgenic mice over-expressing Slit2. We disrupted Slit/Robo1 signaling by injecting R5 monoclonal antibodies into HH10 neural tubes to block the Robo1 receptor. This inhibited the normal development of the ventral body curvature and caused the spinal cord to curl up into a S-shape. Next, Slit/Robo1 signaling on one half-side of the chick embryo neural tube was disturbed by electroporation in ovo. We found that the morphology of the neural tube was dramatically abnormal after we interfered with Slit/Robo1 signaling. Furthermore, we established that silencing Robo1 inhibited cell proliferation while over-expressing Robo1 enhanced cell proliferation. We also investigated the effects of altering Slit/Robo1 expression on Sonic Hedgehog (Shh) and Pax7 expression in the developing neural tube. We demonstrated that over-expressing Robo1 down-regulated Shh expression in the ventral neural tube and resulted in the production of fewer HNK-1{sup +} migrating neural crest cells (NCCs). In addition, Robo1 over-expression enhanced Pax7 expression in the dorsal neural tube and increased the number of Slug{sup +} pre-migratory NCCs. Conversely, silencing Robo1 expression resulted in an enhanced Shh expression and more HNK-1{sup +} migrating NCCs but reduced Pax7 expression and fewer Slug{sup +} pre-migratory NCCs were observed. In conclusion, we propose that Slit/Robo1 signaling is involved in regulating neural tube

  13. Induction of neural differentiation in rat C6 glioma cells with taxol.

    PubMed

    Chao, Chuan-Chuan; Kan, Daphne; Lo, Ta-Hsuan; Lu, Kuo-Shyan; Chien, Chung-Liang

    2015-12-01

    Glioblastoma is a common and aggressive type of primary brain tumor. Several anticancer drugs affect GBM (glioblastoma multiforme) cells on cell growth and morphology. Taxol is one of the widely used antineoplastic drugs against many types of solid tumors, such as breast, ovarian, and prostate cancers. However, the effect of taxol on GBM cells remains unclear and requires further investigation. Survival rate of C6 glioma cells under different taxol concentrations was quantified. To clarify the differentiation patterns of rat C6 glioma cells under taxol challenge, survived glioma cells were characterized by immunocytochemical, molecular biological, and cell biological approaches. After taxol treatment, not only cell death but also morphological changes, including cell elongation, cellular processes thinning, irregular shapes, and fragmented nucleation or micronuclei, occurred in the survived C6 cells. Neural differentiation markers NFL (for neurons), β III-tubulin (for neurons), GFAP (for astrocytes), and CNPase (for oligodendrocytes) were detected in the taxol-treated C6 cells. Quantitative analysis suggested a significant increase in the percentage of neural differentiated cells. The results exhibited that taxol may trigger neural differentiation in C6 glioma cells. Increased expression of neural differentiation markers in C6 cells after taxol treatment suggest that some anticancer drugs could be applied to elimination of the malignant cancer cells as well as changing proliferation and differentiation status of tumor cells.

  14. Bioinformatic analysis of neural stem cell differentiation.

    PubMed

    Goff, Loyal A; Davila, Jonathan; Jörnsten, Rebecka; Keles, Sunduz; Hart, Ronald P

    2007-09-01

    Regulated mRnAs during differentiation of rat neural stem cells were analyzed using the ABi1700 microarray platform. This microarray, while technically advanced, suffers from the difficulty of integrating hybridization results into public databases for systems-level analysis. This is particularly true for the rat array, since many of the probes were designed for transcripts based on predicted human and mouse homologs. using several strategies, we increased the public annotation of the 27,531 probes from 43% to over 65%. To increase the dynamic range of annotation, probes were mapped to numerous public keys from several data sources. consensus annotation from multiple sources was determined for well-scoring alignments, and a confidence-based ranking system established for probes with less agreement across multiple data sources. previous attempts at genomic interpretation using the celera annotation model resulted in poor overlap with expected genomic sequences. since the public keys are more precisely mapped to the genome, we could now analyze the relationships between predicted transcription-factor binding sites and expression clusters. Results collected from a differentiation time course of two neural stem cell clones were clustered using a model-based algorithm. Transcription-factor binding sites were predicted from upstream regions of mapped transcripts using position weight matrices from either JAspAR or TRAnsFAc, and the resulting scores were used to discriminate between observed expression clusters. A classification and regression tree analysis was conducted using cluster numbers as gene identifiers and TFBs scores as predictors, pruning back to obtain a tree with the lowest gene class prediction error rate. Results identify several transcription factors, the presence or absence of which are sufficient to describe clusters of mRnAs changing over time-those that are static, as well as clusters describing cell line differences. public annotation of the AB1700

  15. Generation of diverse neural cell types through direct conversion

    PubMed Central

    Petersen, Gayle F; Strappe, Padraig M

    2016-01-01

    A characteristic of neurological disorders is the loss of critical populations of cells that the body is unable to replace, thus there has been much interest in identifying methods of generating clinically relevant numbers of cells to replace those that have been damaged or lost. The process of neural direct conversion, in which cells of one lineage are converted into cells of a neural lineage without first inducing pluripotency, shows great potential, with evidence of the generation of a range of functional neural cell types both in vitro and in vivo, through viral and non-viral delivery of exogenous factors, as well as chemical induction methods. Induced neural cells have been proposed as an attractive alternative to neural cells derived from embryonic or induced pluripotent stem cells, with prospective roles in the investigation of neurological disorders, including neurodegenerative disease modelling, drug screening, and cellular replacement for regenerative medicine applications, however further investigations into improving the efficacy and safety of these methods need to be performed before neural direct conversion becomes a clinically viable option. In this review, we describe the generation of diverse neural cell types via direct conversion of somatic cells, with comparison against stem cell-based approaches, as well as discussion of their potential research and clinical applications. PMID:26981169

  16. In vivo blockade of neural activity alters dendritic development of neonatal CA1 pyramidal cells.

    PubMed

    Groc, Laurent; Petanjek, Zdravko; Gustafsson, Bengt; Ben-Ari, Yehezkel; Hanse, Eric; Khazipov, Roustem

    2002-11-01

    During development, neural activity has been proposed to promote neuronal growth. During the first postnatal week, the hippocampus is characterized by an oscillating neural network activity and a rapid neuronal growth. In the present study we tested in vivo, by injecting tetanus toxin into the hippocampus of P1 rats, whether this neural activity indeed promotes growth of pyramidal cells. We have previously shown that tetanus toxin injection leads to a strong reduction in the frequency of spontaneous GABA and glutamatergic synaptic currents, and to a complete blockade of the early neural network activity during the first postnatal week. Morphology of neurobiotin-filled CA1 pyramidal cells was analyzed at the end of the first postnatal week (P6-10). In activity-reduced neurons, the total length of basal dendritic tree was three times less than control. The number, but not the length, of basal dendritic branches was affected. The growth impairment was restricted to the basal dendrites. The apical dendrite, the axons, or the soma grew normally during activity deprivation. Thus, the in vivo neural activity in the neonate hippocampus seems to promote neuronal growth by initiating novel branches.

  17. Analysing human neural stem cell ontogeny by consecutive isolation of Notch active neural progenitors.

    PubMed

    Edri, Reuven; Yaffe, Yakey; Ziller, Michael J; Mutukula, Naresh; Volkman, Rotem; David, Eyal; Jacob-Hirsch, Jasmine; Malcov, Hagar; Levy, Carmit; Rechavi, Gideon; Gat-Viks, Irit; Meissner, Alexander; Elkabetz, Yechiel

    2015-03-23

    Decoding heterogeneity of pluripotent stem cell (PSC)-derived neural progeny is fundamental for revealing the origin of diverse progenitors, for defining their lineages, and for identifying fate determinants driving transition through distinct potencies. Here we have prospectively isolated consecutively appearing PSC-derived primary progenitors based on their Notch activation state. We first isolate early neuroepithelial cells and show their broad Notch-dependent developmental and proliferative potential. Neuroepithelial cells further yield successive Notch-dependent functional primary progenitors, from early and midneurogenic radial glia and their derived basal progenitors, to gliogenic radial glia and adult-like neural progenitors, together recapitulating hallmarks of neural stem cell (NSC) ontogeny. Gene expression profiling reveals dynamic stage-specific transcriptional patterns that may link development of distinct progenitor identities through Notch activation. Our observations provide a platform for characterization and manipulation of distinct progenitor cell types amenable for developing streamlined neural lineage specification paradigms for modelling development in health and disease.

  18. Adult Mammalian Neural Stem Cells and Neurogenesis: Five Decades Later

    PubMed Central

    Bond, Allison M.; Ming, Guo-li; Song, Hongjun

    2015-01-01

    Summary Adult somatic stem cells in various organs maintain homeostatic tissue regeneration and enhance plasticity. Since its initial discovery five decades ago, investigations of adult neurogenesis and neural stem cells have led to an established and expanding field that has significantly influenced many facets of neuroscience, developmental biology and regenerative medicine. Here we review recent progress and focus on questions related to adult mammalian neural stem cells that also apply to other somatic stem cells. We further discuss emerging topics that are guiding the field toward better understanding adult neural stem cells and ultimately applying these principles to improve human health. PMID:26431181

  19. Generation of retinal pigment epithelial cells from human embryonic stem cell-derived spherical neural masses.

    PubMed

    Cho, Myung Soo; Kim, Sang Jin; Ku, Seung-Yup; Park, Jung Hyun; Lee, Haksup; Yoo, Dae Hoon; Park, Un Chul; Song, Seul Ae; Choi, Young Min; Yu, Hyeong Gon

    2012-09-01

    Dysfunction and loss of retinal pigment epithelium (RPE) are major pathologic changes observed in various retinal degenerative diseases such as aged-related macular degeneration. RPE generated from human pluripotent stem cells can be a good candidate for RPE replacement therapy. Here, we show the differentiation of human embryonic stem cells (hESCs) toward RPE with the generation of spherical neural masses (SNMs), which are pure masses of hESCs-derived neural precursors. During the early passaging of SNMs, cystic structures arising from opened neural tube-like structures showed pigmented epithelial morphology. These pigmented cells were differentiated into functional RPE by neuroectodermal induction and mechanical purification. Most of the differentiated cells showed typical RPE morphologies, such as a polygonal-shaped epithelial monolayer, and transmission electron microscopy revealed apical microvilli, pigment granules, and tight junctions. These cells also expressed molecular markers of RPE, including Mitf, ZO-1, RPE65, CRALBP, and bestrophin. The generated RPE also showed phagocytosis of isolated bovine photoreceptor outer segment and secreting pigment epithelium-derived factor and vascular endothelial growth factor. Functional RPE could be generated from SNM in our method. Because SNMs have several advantages, including the capability of expansion for long periods without loss of differentiation capability, easy storage and thawing, and no need for feeder cells, our method for RPE differentiation may be used as an efficient strategy for generating functional RPE cells for retinal regeneration therapy.

  20. Prolonged cultivation of hippocampal neural precursor cells shifts their differentiation potential and selects for aneuploid cells.

    PubMed

    Nguyen, The Duy; Widera, Darius; Greiner, Johannes; Müller, Janine; Martin, Ina; Slotta, Carsten; Hauser, Stefan; Kaltschmidt, Christian; Kaltschmidt, Barbara

    2013-12-01

    Neural precursor cells (NPCs) are lineage-restricted neural stem cells with limited self-renewal, giving rise to a broad range of neural cell types such as neurons, astrocytes, and oligodendrocytes. Despite this developmental potential, the differentiation capacity of NPCs has been controversially discussed concerning the trespassing lineage boundaries, for instance resulting in hematopoietic competence. Assessing their in vitro plasticity, we isolated nestin+/Sox2+, NPCs from the adult murine hippocampus. In vitro-expanded adult NPCs were able to form neurospheres, self-renew, and differentiate into neuronal, astrocytic, and oligodendrocytic cells. Although NPCs cultivated in early passage efficiently gave rise to neuronal cells in a directed differentiation assay, extensively cultivated NPCs revealed reduced potential for ectodermal differentiation. We further observed successful differentiation of long-term cultured NPCs into osteogenic and adipogenic cell types, suggesting that NPCs underwent a fate switch during culture. NPCs cultivated for more than 12 passages were aneuploid (abnormal chromosome numbers such as 70 chromosomes). Furthermore, they showed growth factor-independent proliferation, a hallmark of tumorigenic transformation. In conclusion, our findings substantiate the lineage restriction of NPCs from adult mammalian hippocampus. Prolonged cultivation results, however, in enhanced differentiation potential, which may be attributed to transformation events leading to aneuploid cells.

  1. Human placenta-derived mesenchymal stem cells acquire neural phenotype under the appropriate niche conditions.

    PubMed

    Martini, Maristela Maria; Jeremias, Talita da Silva; Kohler, Maria Cecília; Marostica, Lucas Lourenço; Trentin, Andréa Gonçalves; Alvarez-Silva, Marcio

    2013-02-01

    Mesenchymal stem cells (MSCs) are multipotent stem cells with clinical interest. It has been reported that MSCs can be isolated from the human term placenta. We investigated the ability of human placenta-derived MSCs to differentiate into a neural phenotype in coculture assays with astrocytes obtained from neonatal rats. Placenta-derived MSCs were cocultured on a confluent monolayer of astrocytes obtained from the rat cerebellum to evaluate the differences in morphology. The extracellular matrix (ECM) produced by astrocytes as well as the growth factors produced by the astrocyte-conditioned medium were evaluated. The expression of the neural markers glial fibrillate acid protein (GFAP) and Nestin was studied in MSCs by immunocytochemistry. MSCs were able to respond to the astrocyte niche in coculture assays. They expressed the neural markers GFAP, Nestin, or β-Tubulin III, followed by an outgrowth of cell processes. The ECM from astrocytes was not effective in inducing the neural phenotype in MSCs, although the expression of β-Tubulin III was observed. When MSCs were cocultured with cerebellar astrocytes from newborn rats, a neural phenotype was achieved. This was determined by immunocytochemistry to GFAP, Nestin, or β-Tubulin III and by morphological changes. It was achieved without the addition of exogenous differentiation factors. This demonstrates that placenta-derived MSCs may be able to differentiate into neural cell types when in direct contact with a neural environment.

  2. IGF-1 enhances cell proliferation and survival during early differentiation of mesenchymal stem cells to neural progenitor-like cells.

    PubMed

    Huat, Tee Jong; Khan, Amir Ali; Pati, Soumya; Mustafa, Zulkifli; Abdullah, Jafri Malin; Jaafar, Hasnan

    2014-07-22

    There has been increasing interest recently in the plasticity of mesenchymal stem cells (MSCs) and their potential to differentiate into neural lineages. To unravel the roles and effects of different growth factors in the differentiation of MSCs into neural lineages, we have differentiated MSCs into neural lineages using different combinations of growth factors. Based on previous studies of the roles of insulin-like growth factor 1 (IGF-1) in neural stem cell isolation in the laboratory, we hypothesized that IGF-1 can enhance proliferation and reduce apoptosis in neural progenitor-like cells (NPCs) during differentiation of MSCs into NCPs.We induced MSCs differentiation under four different combinations of growth factors: (A) EGF + bFGF, (B) EGF + bFGF + IGF-1, (C) EGF + bFGF + LIF, (D) EGF + bFGF + BDNF, and (E) without growth factors, as a negative control. The neurospheres formed were characterized by immunofluorescence staining against nestin, and the expression was measured by flow cytometry. Cell proliferation and apoptosis were also studied by MTS and Annexin V assay, respectively, at three different time intervals (24 hr, 3 days, and 5 days). The neurospheres formed in the four groups were then terminally differentiated into neuron and glial cells. The four derived NPCs showed a significantly higher expression of nestin than was shown by the negative control. Among the groups treated with growth factors, NPCs treated with IGF-1 showed the highest expression of nestin. Furthermore, NPCs derived using IGF-1 exhibited the highest cell proliferation and cell survival among the treated groups. The NPCs derived from IGF-1 treatment also resulted in a better yield after the terminal differentiation into neurons and glial cells than that of the other treated groups. Our results suggested that IGF-1 has a crucial role in the differentiation of MSCs into neuronal lineage by enhancing the proliferation and reducing the apoptosis in the

  3. Pannexin 1 regulates postnatal neural stem and progenitor cell proliferation

    PubMed Central

    2012-01-01

    Background Pannexin 1 forms ion and metabolite permeable hexameric channels and is abundantly expressed in the brain. After discovering pannexin 1 expression in postnatal neural stem and progenitor cells we sought to elucidate its functional role in neuronal development. Results We detected pannexin 1 in neural stem and progenitor cells in vitro and in vivo. We manipulated pannexin 1 expression and activity in Neuro2a neuroblastoma cells and primary postnatal neurosphere cultures to demonstrate that pannexin 1 regulates neural stem and progenitor cell proliferation likely through the release of adenosine triphosphate (ATP). Conclusions Permeable to ATP, a potent autocrine/paracine signaling metabolite, pannexin 1 channels are ideally suited to influence the behavior of neural stem and progenitor cells. Here we demonstrate they play a robust role in the regulation of neural stem and progenitor cell proliferation. Endogenous postnatal neural stem and progenitor cells are crucial for normal brain health, and their numbers decline with age. Furthermore, these special cells are highly responsive to neurological injury and disease, and are gaining attention as putative targets for brain repair. Therefore, understanding the fundamental role of pannexin 1 channels in neural stem and progenitor cells is of critical importance for brain health and disease. PMID:22458943

  4. Adult vascular smooth muscle cells in culture express neural stem cell markers typical of resident multipotent vascular stem cells.

    PubMed

    Kennedy, Eimear; Mooney, Ciaran J; Hakimjavadi, Roya; Fitzpatrick, Emma; Guha, Shaunta; Collins, Laura E; Loscher, Christine E; Morrow, David; Redmond, Eileen M; Cahill, Paul A

    2014-10-01

    Differentiation of resident multipotent vascular stem cells (MVSCs) or de-differentiation of vascular smooth muscle cells (vSMCs) might be responsible for the SMC phenotype that plays a major role in vascular diseases such as arteriosclerosis and restenosis. We examined vSMCs from three different species (rat, murine and bovine) to establish whether they exhibit neural stem cell characteristics typical of MVSCs. We determined their SMC differentiation, neural stem cell marker expression and multipotency following induction in vitro by using immunocytochemistry, confocal microscopy, fluorescence-activated cell sorting analysis and quantitative real-time polymerase chain reaction. MVSCs isolated from rat aortic explants, enzymatically dispersed rat SMCs and rat bone-marrow-derived mesenchymal stem cells served as controls. Murine carotid artery lysates and primary rat aortic vSMCs were both myosin-heavy-chain-positive but weakly expressed the neural crest stem cell marker, Sox10. Each vSMC line examined expressed SMC differentiation markers (smooth muscle α-actin, myosin heavy chain and calponin), neural crest stem cell markers (Sox10(+), Sox17(+)) and a glia marker (S100β(+)). Serum deprivation significantly increased calponin and myosin heavy chain expression and decreased stem cell marker expression, when compared with serum-rich conditions. vSMCs did not differentiate to adipocytes or osteoblasts following adipogenic or osteogenic inductive stimulation, respectively, or respond to transforming growth factor-β1 or Notch following γ-secretase inhibition. Thus, vascular SMCs in culture express neural stem cell markers typical of MVSCs, concomitant with SMC differentiation markers, but do not retain their multipotency. The ultimate origin of these cells might have important implications for their use in investigations of vascular proliferative disease in vitro.

  5. In Vitro Differentiation of Human iPS Cells into Neural like Cells on a Biomimetic Polyurea.

    PubMed

    Hoveizi, Elham; Ebrahimi-Barough, Somayeh; Tavakol, Shima; Sanamiri, Khadije

    2017-01-01

    Human-induced pluripotent stem cells (hiPSCs) have the pluripotency to differentiate into all three germ layers in vitro and have been considered potent candidates for regenerative medicine as an unlimited source of cells for therapeutic applications. Neural tissue engineering is an important area of research in the field of tissue-engineering especially for neurodegenerative disease. Here, we investigated the use of poly lactic acid/gelatin (PLA/gelatin) scaffold as three-dimensional (3D) system which increase neural cell differentiation. Through neural induction, neural-like cells (NLCs) were derived from hiPSCs on nanofibrous PLA/gelatin scaffold. Enhanced numbers of neural structures and staining of neural markers were observed with hiPS cell-seeded nanofibrous scaffolds when compared with control medium. The results revealed that hiPSCs attach and grow on the nanofibrous PLA/gelatin scaffold, and hiPSCs cultured on scaffold have the potential to differentiate in neuronal cells in the presence of growth factors. The result of this study may have impact in tissue engineering and cells-base therapy of neurodegenerative diseases and have a great potential for wide application.

  6. Nerve growth factor-immobilized electrically conducting fibrous scaffolds for potential use in neural engineering applications.

    PubMed

    Lee, Jae Y; Bashur, Chris A; Milroy, Craig A; Forciniti, Leandro; Goldstein, Aaron S; Schmidt, Christine E

    2012-03-01

    Engineered scaffolds simultaneously exhibiting multiple cues are highly desirable for neural tissue regeneration. To this end, we developed a neural tissue engineering scaffold that displays submicrometer-scale features, electrical conductivity, and neurotrophic activity. Specifically, electrospun poly(lactic acid-co-glycolic acid) (PLGA) nanofibers were layered with a nanometer thick coating of electrically conducting polypyrrole (PPy) presenting carboxylic groups. Then, nerve growth factor (NGF) was chemically immobilized onto the surface of the fibers. These NGF-immobilized PPy-coated PLGA (NGF-PPyPLGA) fibers supported PC12 neurite formation ( 28.0±3.0% of the cells) and neurite outgrowth (14.2 μm median length), which were comparable to that observed with NGF (50 ng/mL) in culture medium ( 29.0±1.3%, 14.4 μm). Electrical stimulation of PC12 cells on NGF-immobilized PPyPLGA fiber scaffolds was found to further improve neurite development and neurite length by 18% and 17%, respectively, compared to unstimulated cells on the NGF-immobilized fibers. Hence, submicrometer-scale fibrous scaffolds that incorporate neurotrophic and electroconducting activities may serve as promising neural tissue engineering scaffolds such as nerve guidance conduits.

  7. Nerve Growth Factor-Immobilized Electrically Conducting Fibrous Scaffolds for Potential Use in Neural Engineering Applications

    PubMed Central

    Lee, Jae Y.; Bashur, Chris A.; Milroy, Craig A.; Forciniti, Leandro; Goldstein, Aaron S.

    2015-01-01

    Engineered scaffolds simultaneously exhibiting multiple cues are highly desirable for neural tissue regeneration. To this end, we developed a neural tissue engineering scaffold that displays submicrometer-scale features, electrical conductivity, and neurotrophic activity. Specifically, electrospun poly(lactic acid-co-glycolic acid) (PLGA) nanofibers were layered with a nanometer thick coating of electrically conducting polypyrrole (PPy) presenting carboxylic groups. Then, nerve growth factor (NGF) was chemically immobilized onto the surface of the fibers. These NGF-immobilized PPy-coated PLGA (NGF-PPyPLGA) fibers supported PC12 neurite formation (28.0±3.0% of the cells) and neurite outgrowth (14.2 µm median length), which were comparable to that observed with NGF (50 ng/mL) in culture medium (29.0±1.3%, 14.4 µm). Electrical stimulation of PC12 cells on NGF-immobilized PPyPLGA fiber scaffolds was found to further improve neurite development and neurite length by 18% and 17%, respectively, compared to unstimulated cells on the NGF-immobilized fibers. Hence, submicrometer-scale fibrous scaffolds that incorporate neurotrophic and electroconducting activities may serve as promising neural tissue engineering scaffolds such as nerve guidance conduits. PMID:21712166

  8. Endothelial Cells Stimulate Self-Renewal and Expand Neurogenesis of Neural Stem Cells

    NASA Astrophysics Data System (ADS)

    Shen, Qin; Goderie, Susan K.; Jin, Li; Karanth, Nithin; Sun, Yu; Abramova, Natalia; Vincent, Peter; Pumiglia, Kevin; Temple, Sally

    2004-05-01

    Neural stem cells are reported to lie in a vascular niche, but there is no direct evidence for a functional relationship between the stem cells and blood vessel component cells. We show that endothelial cells but not vascular smooth muscle cells release soluble factors that stimulate the self-renewal of neural stem cells, inhibit their differentiation, and enhance their neuron production. Both embryonic and adult neural stem cells respond, allowing extensive production of both projection neuron and interneuron types in vitro. Endothelial coculture stimulates neuroepithelial cell contact, activating Notch and Hes1 to promote self-renewal. These findings identify endothelial cells as a critical component of the neural stem cell niche.

  9. Endothelial cells stimulate self-renewal and expand neurogenesis of neural stem cells.

    PubMed

    Shen, Qin; Goderie, Susan K; Jin, Li; Karanth, Nithin; Sun, Yu; Abramova, Natalia; Vincent, Peter; Pumiglia, Kevin; Temple, Sally

    2004-05-28

    Neural stem cells are reported to lie in a vascular niche, but there is no direct evidence for a functional relationship between the stem cells and blood vessel component cells. We show that endothelial cells but not vascular smooth muscle cells release soluble factors that stimulate the self-renewal of neural stem cells, inhibit their differentiation, and enhance their neuron production. Both embryonic and adult neural stem cells respond, allowing extensive production of both projection neuron and interneuron types in vitro. Endothelial coculture stimulates neuroepithelial cell contact, activating Notch and Hes 1 to promote self-renewal. These findings identify endothelial cells as a critical component of the neural stem cell niche.

  10. Bone morphogenetic protein-mediated modulation of lineage diversification during neural differentiation of embryonic stem cells.

    PubMed

    Gossrau, Gudrun; Thiele, Janine; Konang, Rachel; Schmandt, Tanja; Brüstle, Oliver

    2007-04-01

    Embryonic stem cells (ES cells) can give rise to a broad spectrum of neural cell types. The biomedical application of ES cells will require detailed knowledge on the role of individual factors modulating fate specification during in vitro differentiation. Bone morphogenetic proteins (BMPs) are known to exert a multitude of diverse differentiation effects during embryonic development. Here, we show that exposure to BMP2 at distinct stages of neural ES cell differentiation can be used to promote specific cell lineages. During early ES cell differentiation, BMP2-mediated inhibition of neuroectodermal differentiation is associated with an increase in mesoderm and smooth muscle differentiation. In fibroblast growth factor 2-expanded ES cell-derived neural precursors, BMP2 supports the generation of neural crest phenotypes, and, within the neuronal lineage, promotes distinct subtypes of peripheral neurons, including cholinergic and autonomic phenotypes. BMP2 also exerts a density-dependent promotion of astrocyte differentiation at the expense of oligodendrocyte formation. Experiments involving inhibition of the serine threonine kinase FRAP support the notion that these effects are mediated via the JAK/STAT pathway. The preservation of diverse developmental BMP2 effects in differentiating ES cell cultures provides interesting prospects for the enrichment of distinct neural phenotypes in vitro.

  11. Pig Induced Pluripotent Stem Cell-Derived Neural Rosettes Parallel Human Differentiation Into Sensory Neural Subtypes.

    PubMed

    Webb, Robin L; Gallegos-Cárdenas, Amalia; Miller, Colette N; Solomotis, Nicholas J; Liu, Hong-Xiang; West, Franklin D; Stice, Steven L

    2017-04-01

    The pig is the large animal model of choice for study of nerve regeneration and wound repair. Availability of porcine sensory neural cells would conceptually allow for analogous cell-based peripheral nerve regeneration in porcine injuries of similar severity and size to those found in humans. After recently reporting that porcine (or pig) induced pluripotent stem cells (piPSCs) differentiate into neural rosette (NR) structures similar to human NRs, here we demonstrate that pig NR cells could differentiate into neural crest cells and other peripheral nervous system-relevant cell types. Treatment with either bone morphogenetic protein 4 or fetal bovine serum led to differentiation into BRN3A-positive sensory cells and increased expression of sensory neuron TRK receptor gene family: TRKA, TRKB, and TRKC. Porcine sensory neural cells would allow determination of parallels between human and porcine cells in response to noxious stimuli, analgesics, and reparative mechanisms. In vitro differentiation of pig sensory neurons provides a novel model system for neural cell subtype specification and would provide a novel platform for the study of regenerative therapeutics by elucidating the requirements for innervation following injury and axonal survival.

  12. Surface topography during neural stem cell differentiation regulates cell migration and cell morphology.

    PubMed

    Czeisler, Catherine; Short, Aaron; Nelson, Tyler; Gygli, Patrick; Ortiz, Cristina; Catacutan, Fay Patsy; Stocker, Ben; Cronin, James; Lannutti, John; Winter, Jessica; Otero, José Javier

    2016-12-01

    We sought to determine the contribution of scaffold topography to the migration and morphology of neural stem cells by mimicking anatomical features of scaffolds found in vivo. We mimicked two types of central nervous system scaffolds encountered by neural stem cells during development in vitro by constructing different diameter electrospun polycaprolactone (PCL) fiber mats, a substrate that we have shown to be topographically similar to brain scaffolds. We compared the effects of large fibers (made to mimic blood vessel topography) with those of small-diameter fibers (made to mimic radial glial process topography) on the migration and differentiation of neural stem cells. Neural stem cells showed differential migratory and morphological reactions with laminin in different topographical contexts. We demonstrate, for the first time, that neural stem cell biological responses to laminin are dependent on topographical context. Large-fiber topography without laminin prevented cell migration, which was partially reversed by treatment with rock inhibitor. Cell morphology complexity assayed by fractal dimension was inhibited in nocodazole- and cytochalasin-D-treated neural precursor cells in large-fiber topography, but was not changed in small-fiber topography with these inhibitors. These data indicate that cell morphology has different requirements on cytoskeletal proteins dependent on the topographical environment encountered by the cell. We propose that the physical structure of distinct scaffolds induces unique signaling cascades that regulate migration and morphology in embryonic neural precursor cells. J. Comp. Neurol. 524:3485-3502, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  13. ZDHHC3 Tyrosine Phosphorylation Regulates Neural Cell Adhesion Molecule Palmitoylation

    PubMed Central

    Lievens, Patricia Marie-Jeanne; Kuznetsova, Tatiana; Kochlamazashvili, Gaga; Cesca, Fabrizia; Gorinski, Natalya; Galil, Dalia Abdel; Cherkas, Volodimir; Ronkina, Natalia; Lafera, Juri; Gaestel, Matthias

    2016-01-01

    The neural cell adhesion molecule (NCAM) mediates cell-cell and cell-matrix adhesion. It is broadly expressed in the nervous system and regulates neurite outgrowth, synaptogenesis, and synaptic plasticity. Previous in vitro studies revealed that palmitoylation of NCAM is required for fibroblast growth factor 2 (FGF2)-stimulated neurite outgrowth and identified the zinc finger DHHC (Asp-His-His-Cys)-containing proteins ZDHHC3 and ZDHHC7 as specific NCAM-palmitoylating enzymes. Here, we verified that FGF2 controlled NCAM palmitoylation in vivo and investigated molecular mechanisms regulating NCAM palmitoylation by ZDHHC3. Experiments with overexpression and pharmacological inhibition of FGF receptor (FGFR) and Src revealed that these kinases control tyrosine phosphorylation of ZDHHC3 and that ZDHHC3 is phosphorylated by endogenously expressed FGFR and Src proteins. By site-directed mutagenesis, we found that Tyr18 is an FGFR1-specific ZDHHC3 phosphorylation site, while Tyr295 and Tyr297 are specifically phosphorylated by Src kinase in cell-based and cell-free assays. Abrogation of tyrosine phosphorylation increased ZDHHC3 autopalmitoylation, enhanced interaction with NCAM, and upregulated NCAM palmitoylation. Expression of ZDHHC3 with tyrosine mutated in cultured hippocampal neurons promoted neurite outgrowth. Our findings for the first time highlight that FGFR- and Src-mediated tyrosine phosphorylation of ZDHHC3 modulates ZDHHC3 enzymatic activity and plays a role in neuronal morphogenesis. PMID:27247265

  14. Inhibition of FGF signaling accelerates neural crest cell differentiation of human pluripotent stem cells.

    PubMed

    Jaroonwitchawan, Thiranut; Muangchan, Pattamon; Noisa, Parinya

    2016-12-02

    Neural crest (NC) is a transient population, arising during embryonic development and capable of differentiating into various somatic cells. The defects of neural crest development leads to neurocristopathy. Several signaling pathways were revealed their significance in NC cell specification. Fibroblast growth factor (FGF) is recognized as an important signaling during NC development, for instance Xenopus and avian; however, its contributions in human species are remained elusive. Here we used human pluripotent stem cells (hPSCs) to investigate the consequences of FGF inhibition during NC cell differentiation. The specific-FGF receptor inhibitor, SU5402, was used in this investigation. The inhibition of FGF did not found to affect the proliferation or death of hPSC-derived NC cells, but promoted hPSCs to commit NC cell fate. NC-specific genes, including PAX3, SLUG, and TWIST1, were highly upregulated, while hPSC genes, such as OCT4, and E-CAD, rapidly reduced upon FGF signaling blockage. Noteworthy, TFAP-2α, a marker of migratory NC cells, abundantly presented in SU5402-induced cells. This accelerated NC cell differentiation could be due to the activation of Notch signaling upon the blockage of ERK1/2 phosphorylation, since NICD was increased by SU5402. Altogether, this study proposed the contributions of FGF signaling in controlling human NC cell differentiation from hPSCs, the crosstalk between FGF and Notch, and might imply to the influences of FGF signaling in neurocristophatic diseases.

  15. Trunk neural crest cells: formation, migration and beyond.

    PubMed

    Vega-Lopez, Guillermo A; Cerrizuela, Santiago; Aybar, Manuel J

    2017-01-01

    Neural crest cells (NCCs) are a multipotent, migratory cell population that generates an astonishingly diverse array of cell types during vertebrate development. The trunk neural crest has long been considered of particular significance. First, it has been held that the trunk neural crest has a morphogenetic role, acting to coordinate the development of the peripheral nervous system, secretory cells of the endocrine system and pigment cells of the skin. Second, the trunk neural crest additionally has skeletal potential. However, it has been demonstrated that a key role of the trunk neural crest streams is to organize the innervation of the intestine. Although trunk NCCs have a limited capacity for self-renewal, sometimes they become neural-crest-derived tumor cells and reveal the fact that that NCCs and tumor cells share the same molecular machinery. In this review we describe the routes taken by trunk NCCs and consider the signals and cues that pattern these trajectories. We also discuss recent advances in the characterization of the properties of trunk NCCs for various model organisms in order to highlight common themes. Finally, looking to the future, we discuss the need to translate the wealth of data from animal studies to the clinical area in order to develop treatments for neural crest-related human diseases.

  16. Chemo-mechanical control of neural stem cell differentiation

    NASA Astrophysics Data System (ADS)

    Geishecker, Emily R.

    Cellular processes such as adhesion, proliferation, and differentiation are controlled in part by cell interactions with the microenvironment. Cells can sense and respond to a variety of stimuli, including soluble and insoluble factors (such as proteins and small molecules) and externally applied mechanical stresses. Mechanical properties of the environment, such as substrate stiffness, have also been suggested to play an important role in cell processes. The roles of both biochemical and mechanical signaling in fate modification of stem cells have been explored independently. However, very few studies have been performed to study well-controlled chemo-mechanotransduction. The objective of this work is to design, synthesize, and characterize a chemo-mechanical substrate to encourage neuronal differentiation of C17.2 neural stem cells. In Chapter 2, Polyacrylamide (PA) gels of varying stiffnesses are functionalized with differing amounts of whole collagen to investigate the role of protein concentration in combination with substrate stiffness. As expected, neurons on the softest substrate were more in number and neuronal morphology than those on stiffer substrates. Neurons appeared locally aligned with an expansive network of neurites. Additional experiments would allow for statistical analysis to determine if and how collagen density impacts C17.2 differentiation in combination with substrate stiffness. Due to difficulties associated with whole protein approaches, a similar platform was developed using mixed adhesive peptides, derived from fibronectin and laminin, and is presented in Chapter 3. The matrix elasticity and peptide concentration can be individually modulated to systematically probe the effects of chemo-mechanical signaling on differentiation of C17.2 cells. Polyacrylamide gel stiffness was confirmed using rheological techniques and found to support values published by Yeung et al. [1]. Cellular growth and differentiation were assessed by cell counts

  17. Neural Stem Cell Transplantation and CNS Diseases.

    PubMed

    Gonzalez, Rodolfo; Hamblin, Milton H; Lee, Jean-Pyo

    2016-01-01

    In neurological disorders, pathological lesions in the central nervous system (CNS) may be globally dispersed throughout the brain or localized to specific regions. Although native neural stem cells (NSCs) are present in the adult mammalian brain, intrinsic self-repair of injured adult CNS tissue is inadequate or ineffective. The brain's poor regenerative ability may be due to the fact that NSCs are restricted to discrete locations, are few in number, or are surrounded by a microenvironment that does not support neuronal differentiation. Therapeutic potential of NSC transplantation in CNS diseases characterized by global degeneration requires that gene products and/or replaced cells be widely distributed. Global degenerative CNS diseases include inherited pediatric neurodegenerative diseases (inborn errors of metabolism, including lysosomal storage disorders (LSDs), such as Tay-Sachs-related Sandhoff disease), hypoxic or ischemic encephalopathy, and some adult CNS diseases (such as multiple sclerosis). Both mouse and human NSCs express many chemokines and chemokine receptors (including CXCR4 and adhesion molecules, such as integrins, selectins, and immunoglobulins) that mediate homing to sources of inflammatory chemokines, such as SDF-1α. In mammalian brains of all ages, NSCs may be attracted even at a great distance to regions of neurodegeneration. Consequently, NSC transplantation presents a promising strategy for treating many CNS diseases.

  18. Regulated GDNF Delivery in Vivo Using Neural Stem Cells

    DTIC Science & Technology

    2006-04-01

    which do not kill cell bodies within the striatum but induce retrograde death of dopamine bodies in the brain stem showed a level of survival and...Neural Stem Cells PRINCIPAL INVESTIGATOR: Clive Svendsen, Ph.D. CONTRACTING ORGANIZATION: University of Wisconsin...Regulated GDNF Delivery in Vivo Using Neural Stem Cells 5b. GRANT NUMBER DAMD17-03-1-0122 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) 5d. PROJECT

  19. Regulated GDNF Delivery In Vivo using Neural Stem Cells

    DTIC Science & Technology

    2005-04-01

    attached as Appendix 2. Body Task 1. To produce rat and monkey neural stem cells which secrete GDNF under an inducible promoter. a. Assess and optimize GDNF...AD Award Number: DAMD17-03-1-0122 TITLE: Regulated GDNF Delivery In Vivo Using Neural Stem Cells PRINCIPAL INVESTIGATOR: Clive N. Svendsen, Ph.D...analysis of 4 rats for PET. This system is now ready for the new stem cell transplants and carrying out the experiments outlined in year three which

  20. Induction of neural crest cells from mouse embryonic stem cells in a serum-free monolayer culture.

    PubMed

    Aihara, Yuko; Hayashi, Yohei; Hirata, Mitsuhi; Ariki, Nobutaka; Shibata, Shinsuke; Nagoshi, Narihito; Nakanishi, Mio; Ohnuma, Kiyoshi; Warashina, Masaki; Michiue, Tatsuo; Uchiyama, Hideho; Okano, Hideyuki; Asashima, Makoto; Furue, Miho Kusuda

    2010-01-01

    The neural crest (NC) is a group of cells located in the neural folds at the boundary between the neural and epidermal ectoderm. NC cells differentiate into a vast range of cells,including neural cells, smooth muscle cells, bone and cartilage cells of the maxillofacial region, and odontoblasts. The molecular mechanisms underlying NC induction during early development remain poorly understood. We previously established a defined serum-free culture condition for mouse embryonic stem (mES) cells without feeders. Here, using this defined condition, we have developed a protocol to promote mES cell differentiation into NC cells in an adherent monolayer culture. We found that adding bone morphogenetic protein (BMP)-4 together with fibroblast growth factor (FGF)-2 shifts mES cell differentiation into the NC lineage. Furthermore, we have established a cell line designated as P0-6 that is derived from the blastocysts of P0-Cre/Floxed-EGFP mice expressing EGFP in an NC-lineage-specific manner. P0-6 cells cultured using this protocol expressed EGFP. This protocol could be used to help clarify the mechanisms by which cells differentiate into the NC lineage and to assist the development of applications for clinical therapy.

  1. Role of neural precursor cells in promoting repair following stroke

    PubMed Central

    Dibajnia, Pooya; Morshead, Cindi M

    2013-01-01

    Stem cell-based therapies for the treatment of stroke have received considerable attention. Two broad approaches to stem cell-based therapies have been taken: the transplantation of exogenous stem cells, and the activation of endogenous neural stem and progenitor cells (together termed neural precursors). Studies examining the transplantation of exogenous cells have demonstrated that neural stem and progenitor cells lead to the most clinically promising results. Endogenous activation of neural precursors has also been explored based on the fact that resident precursor cells have the inherent capacity to proliferate, migrate and differentiate into mature neurons in the uninjured adult brain. Studies have revealed that these neural precursor cell behaviours can be activated following stroke, whereby neural precursors will expand in number, migrate to the infarct site and differentiate into neurons. However, this innate response is insufficient to lead to functional recovery, making it necessary to enhance the activation of endogenous precursors to promote tissue repair and functional recovery. Herein we will discuss the current state of the stem cell-based approaches with a focus on endogenous repair to treat the stroke injured brain. PMID:23064725

  2. Behavior of neural stem cells in the Alzheimer brain.

    PubMed

    Waldau, B; Shetty, A K

    2008-08-01

    Alzheimer's disease (AD) is characterized by the deposition of beta-amyloid peptides (Abeta) and a progressive loss of neurons leading to dementia. Because hippocampal neurogenesis is linked to functions such as learning, memory and mood, there has been great interest in examining the effects of AD on hippocampal neurogenesis. This article reviews the pertinent studies and tries to unite them in one possible disease model. Early in the disease, oligomeric Abeta may transiently promote the generation of immature neurons from neural stem cells (NSCs). However, reduced concentrations of multiple neurotrophic factors and higher levels of fibroblast growth factor-2 seem to induce a developmental arrest of newly generated neurons. Furthermore, fibrillary Abeta and down-regulation of oligodendrocyte-lineage transcription factor-2 (OLIG2) may cause the death of these nonfunctional neurons. Therefore, altering the brain microenvironment for fostering apt maturation of graft-derived neurons may be critical for improving the efficacy of NSC transplantation therapy for AD.

  3. The influence of electric fields on hippocampal neural progenitor cells.

    PubMed

    Ariza, Carlos Atico; Fleury, Asha T; Tormos, Christian J; Petruk, Vadim; Chawla, Sagar; Oh, Jisun; Sakaguchi, Donald S; Mallapragada, Surya K

    2010-12-01

    The differentiation and proliferation of neural stem/progenitor cells (NPCs) depend on various in vivo environmental factors or cues, which may include an endogenous electrical field (EF), as observed during nervous system development and repair. In this study, we investigate the morphologic, phenotypic, and mitotic alterations of adult hippocampal NPCs that occur when exposed to two EFs of estimated endogenous strengths. NPCs treated with a 437 mV/mm direct current (DC) EF aligned perpendicularly to the EF vector and had a greater tendency to differentiate into neurons, but not into oligodendrocytes or astrocytes, compared to controls. Furthermore, NPC process growth was promoted perpendicularly and inhibited anodally in the 437 mV/mm DC EF. Yet fewer cells were observed in the DC EF, which in part was due to a decrease in cell viability. The other EF applied was a 46 mV/mm alternating current (AC) EF. However, the 46 mV/mm AC EF showed no major differences in alignment or differentiation, compared to control conditions. For both EF treatments, the percent of mitotic cells during the last 14 h of the experiment were statistically similar to controls. Reported here, to our knowledge, is the first evidence of adult NPC differentiation affected in an EF in vitro. Further investigation and application of EFs on stem cells is warranted to elucidate the utility of EFs to control phenotypic behavior. With progress, the use of EFs may be engineered to control differentiation and target the growth of transplanted cells in a stem cell-based therapy to treat nervous system disorders.

  4. Monocarboxylate transporter 8 in neuronal cell growth.

    PubMed

    James, S R; Franklyn, J A; Reaves, B J; Smith, V E; Chan, S Y; Barrett, T G; Kilby, M D; McCabe, C J

    2009-04-01

    Thyroid hormones are essential for the normal growth and development of the fetus, and even small alterations in maternal thyroid hormone status during early pregnancy may be associated with neurodevelopmental abnormalities in childhood. Mutations in the novel and specific thyroid hormone transporter monocarboxylate transporter 8 (MCT8) have been associated with severe neurodevelopmental impairment. However, the mechanism by which MCT8 influences neural development remains poorly defined. We have therefore investigated the effect of wild-type (WT) MCT8, and the previously reported L471P mutant, on the growth and function of human neuronal precursor NT2 cells as well as MCT8-null JEG-3 cells. HA-tagged WT MCT8 correctly localized to the plasma membrane in NT2 cells and increased T(3) uptake in both cell types. In contrast, L471P MCT8 was largely retained in the endoplasmic reticulum and displayed no T(3) transport activity. Transient overexpression of WT and mutant MCT8 proteins failed to induce endoplasmic reticular stress or apoptosis. However, MCT8 overexpression significantly repressed cell proliferation in each cell type in both the presence and absence of the active thyroid hormone T(3) and in a dose-dependent manner. In contrast, L471P MCT8 showed no such influence. Finally, small interfering RNA depletion of endogenous MCT8 resulted in increased cell survival and decreased T(3) uptake. Given that T(3) stimulated proliferation in embryonic neuronal NT2 cells, whereas MCT8 repressed cell growth, these data suggest an entirely novel role for MCT8 in addition to T(3) transport, mediated through the modulation of cell proliferation in the developing brain.

  5. Comparative aspects of adult neural stem cell activity in vertebrates.

    PubMed

    Grandel, Heiner; Brand, Michael

    2013-03-01

    At birth or after hatching from the egg, vertebrate brains still contain neural stem cells which reside in specialized niches. In some cases, these stem cells are deployed for further postnatal development of parts of the brain until the final structure is reached. In other cases, postnatal neurogenesis continues as constitutive neurogenesis into adulthood leading to a net increase of the number of neurons with age. Yet, in other cases, stem cells fuel neuronal turnover. An example is protracted development of the cerebellar granular layer in mammals and birds, where neurogenesis continues for a few weeks postnatally until the granular layer has reached its definitive size and stem cells are used up. Cerebellar growth also provides an example of continued neurogenesis during adulthood in teleosts. Again, it is the granular layer that grows as neurogenesis continues and no definite adult cerebellar size is reached. Neuronal turnover is most clearly seen in the telencephalon of male canaries, where projection neurons are replaced in nucleus high vocal centre each year before the start of a new mating season--circuitry reconstruction to achieve changes of the song repertoire in these birds? In this review, we describe these and other examples of adult neurogenesis in different vertebrate taxa. We also compare the structure of the stem cell niches to find common themes in their organization despite different functions adult neurogenesis serves in different species. Finally, we report on regeneration of the zebrafish telencephalon after injury to highlight similarities and differences of constitutive neurogenesis and neuronal regeneration.

  6. Utilizing stem cells for three-dimensional neural tissue engineering.

    PubMed

    Knowlton, Stephanie; Cho, Yongku; Li, Xue-Jun; Khademhosseini, Ali; Tasoglu, Savas

    2016-05-26

    Three-dimensional neural tissue engineering has made great strides in developing neural disease models and replacement tissues for patients. However, the need for biomimetic tissue models and effective patient therapies remains unmet. The recent push to expand 2D neural tissue engineering into the third dimension shows great potential to advance the field. Another area which has much to offer to neural tissue engineering is stem cell research. Stem cells are well known for their self-renewal and differentiation potential and have been shown to give rise to tissues with structural and functional properties mimicking natural organs. Application of these capabilities to 3D neural tissue engineering may be highly useful for basic research on neural tissue structure and function, engineering disease models, designing tissues for drug development, and generating replacement tissues with a patient's genetic makeup. Here, we discuss the vast potential, as well as the current challenges, unique to integration of 3D fabrication strategies and stem cells into neural tissue engineering. We also present some of the most significant recent achievements, including nerve guidance conduits to facilitate better healing of nerve injuries, functional 3D biomimetic neural tissue models, physiologically relevant disease models for research purposes, and rapid and effective screening of potential drugs.

  7. Neural stem cells: balancing self-renewal with differentiation.

    PubMed

    Doe, Chris Q

    2008-05-01

    Stem cells are captivating because they have the potential to make multiple cell types yet maintain their undifferentiated state. Recent studies of Drosophila and mammalian neural stem cells have shed light on how stem cells regulate self-renewal versus differentiation and have revealed the proteins, processes and pathways that all converge to regulate neural progenitor self-renewal. If we can better understand how stem cells balance self-renewal versus differentiation, we will significantly advance our knowledge of embryogenesis, cancer biology and brain evolution, as well as the use of stem cells for therapeutic purposes.

  8. Development of novel microfluidic platforms for neural stem cell research

    NASA Astrophysics Data System (ADS)

    Chung, Bonggeun

    This dissertation describes the development and characterization of novel microfluidic platforms to study proliferation, differentiation, migration, and apoptosis of neural stem cells (NSCs). NSCs hold tremendous promise for fundamental biological studies and cell-based therapies in human disorders. NSCs are defined as cells that can self-renew yet maintain the ability to generate the three principal cell types of the central nervous system such as neurons, astrocytes, and oligodendrocytes. NSCs therefore have therapeutic possibilities in multiple neurodevelopmental and neurodegenerative diseases. Despite their promise, cell-based therapies are limited by the inability to precisely control their behavior in culture. Compared to traditional culture tools, microfluidic platforms can provide much greater control over cell microenvironments and optimize proliferation and differentiation conditions of cells exposed to combinatorial mixtures of growth factors. Human NSCs were cultured for more than 1 week in the microfluidic device while constantly exposed to a continuous gradient of a growth factor mixture. NSCs proliferated and differentiated in a graded and proportional fashion that varied directly with growth factor concentration. In parallel to the study of growth and differentiation of NSCs, we are interested in proliferation and apoptosis of mouse NSCs exposed to morphogen gradients. Morphogen gradients are fundamental to animal brain development. Nonetheless, much controversy remains about the mechanisms by which morphogen gradients act on the developing brain. To overcome limitations of in-vitro models of gradients, we have developed a hybrid microfluidic platform that can mimic morphogen gradient profiles. Bone morphogenetic protein (BMP) activity in the developing cortex is graded and cortical NSC responses to BMPs are highly dependent on concentration and gradient slope of BMPs. To make novel microfluidic devices integrated with multiple functions, we have

  9. Differentiation of Neural Precursors and Dopaminergic Neurons from Human Embryonic Stem Cells

    PubMed Central

    Zhang, Xiao-Qing; Zhang, Su-Chun

    2010-01-01

    Directed differentiation of human embryonic stem cells (hESCs) to a functional cell type, including neurons, is the foundation for application of hESCs. We describe here a reproducible, chemically-defined protocol that allows directed differentiation of hESCs to nearly pure neuroectodermal cells and neurons. First, hESC colonies are detached from mouse fibroblast feeder layers and form aggregates to initiate the differentiation procedure. Second, after 4 days of suspension culture, the ESC growth medium is replaced with neural induction medium to guide neuroectodermal specification. Third, the differentiating hESC aggregates are attached onto the culture surface at day 6-7, where columnar neural epithelial cells appear and organize into rosettes. Fourth, the neural rosettes are enriched by detaching rosettes and leaving the peripheral flat cells attached, and expanded as neuroepithelial aggregates in the same medium. Finally, the neuroepithelial aggregates are dissociated and differentiated to nearly pure neurons. This stepwise differentiation protocol results in the generation of primitive neuroepithelia at day 8-10, neural progenitors at the 2nd and 3rd week, and postmitotic neurons at the 4th week, which mirrors the early phase of neural development in a human embryo. Identification of the primitive neuroepithelial cells permits efficient patterning of region-specific progenitors and neuronal subtypes such as midbrain dopaminergic neurons. PMID:19907987

  10. Combination cell therapy with mesenchymal stem cells and neural stem cells for brain stroke in rats.

    PubMed

    Hosseini, Seyed Mojtaba; Farahmandnia, Mohammad; Razi, Zahra; Delavari, Somayeh; Shakibajahromi, Benafsheh; Sarvestani, Fatemeh Sabet; Kazemi, Sepehr; Semsar, Maryam

    2015-05-01

    Brain stroke is the second most important events that lead to disability and morbidity these days. Although, stroke is important, there is no treatment for curing this problem. Nowadays, cell therapy has opened a new window for treating central nervous system disease. In some previous studies the Mesenchymal stem cells and neural stem cells. In this study, we have designed an experiment to assess the combination cell therapy (Mesenchymal and Neural stem cells) effects on brain stroke. The Mesenchymal stem cells were isolated from adult rat bone marrow and the neural stem cells were isolated from ganglion eminence of rat embryo 14 days. The Mesenchymal stem cells were injected 1 day after middle cerebral artery occlusion (MCAO) and the neural stem cells transplanted 7 day after MCAO. After 28 days, the neurological outcomes and brain lesion volumes were evaluated. Also, the activity of Caspase 3 was assessed in different groups. The group which received combination cell therapy had better neurological examination and less brain lesion. Also the combination cell therapy group had the least Caspase 3 activity among the groups. The combination cell therapy is more effective than Mesenchymal stem cell therapy and neural stem cell therapy separately in treating the brain stroke in rats.

  11. Functional electrical stimulation-facilitated proliferation and regeneration of neural precursor cells in the brains of rats with cerebral infarction

    PubMed Central

    Xiang, Yun; Liu, Huihua; Yan, Tiebin; Zhuang, Zhiqiang; Jin, Dongmei; Peng, Yuan

    2014-01-01

    Previous studies have shown that proliferation of endogenous neural precursor cells cannot alone compensate for the damage to neurons and axons. From the perspective of neural plasticity, we observed the effects of functional electrical stimulation treatment on endogenous neural precursor cell proliferation and expression of basic fibroblast growth factor and epidermal growth factor in the rat brain on the infarct side. Functional electrical stimulation was performed in rat models of acute middle cerebral artery occlusion. Simultaneously, we set up a placebo stimulation group and a sham-operated group. Immunohistochemical staining showed that, at 7 and 14 days, compared with the placebo group, the numbers of nestin (a neural precursor cell marker)-positive cells in the subgranular zone and subventricular zone were increased in the functional electrical stimulation treatment group. Western blot assays and reverse-transcription PCR showed that total protein levels and gene expression of epidermal growth factor and basic fibroblast growth factor were also upregulated on the infarct side. Prehensile traction test results showed that, at 14 days, prehension function of rats in the functional electrical stimulation group was significantly better than in the placebo group. These results suggest that functional electrical stimulation can promote endogenous neural precursor cell proliferation in the brains of acute cerebral infarction rats, enhance expression of basic fibroblast growth factor and epidermal growth factor, and improve the motor function of rats. PMID:25206808

  12. Multiple phenotypes in Huntington disease mouse neural stem cells.

    PubMed

    Ritch, James J; Valencia, Antonio; Alexander, Jonathan; Sapp, Ellen; Gatune, Leah; Sangrey, Gavin R; Sinha, Saurabh; Scherber, Cally M; Zeitlin, Scott; Sadri-Vakili, Ghazaleh; Irimia, Daniel; Difiglia, Marian; Kegel, Kimberly B

    2012-05-01

    Neural stem (NS) cells are a limitless resource, and thus superior to primary neurons for drug discovery provided they exhibit appropriate disease phenotypes. Here we established NS cells for cellular studies of Huntington's disease (HD). HD is a heritable neurodegenerative disease caused by a mutation resulting in an increased number of glutamines (Q) within a polyglutamine tract in Huntingtin (Htt). NS cells were isolated from embryonic wild-type (Htt(7Q/7Q)) and "knock-in" HD (Htt(140Q/140Q)) mice expressing full-length endogenous normal or mutant Htt. NS cells were also developed from mouse embryonic stem cells that were devoid of Htt (Htt(-/-)), or knock-in cells containing human exon1 with an N-terminal FLAG epitope tag and with 7Q or 140Q inserted into one of the mouse alleles (Htt(F7Q/7Q) and Htt(F140Q/7Q)). Compared to Htt(7Q/7Q) NS cells, HD Htt(140Q/140Q) NS cells showed significantly reduced levels of cholesterol, increased levels of reactive oxygen species (ROS), and impaired motility. The heterozygous Htt(F140Q/7Q) NS cells had increased ROS and decreased motility compared to Htt(F7Q/7Q). These phenotypes of HD NS cells replicate those seen in HD patients or in primary cell or in vivo models of HD. Huntingtin "knock-out" NS cells (Htt(-/-)) also had impaired motility, but in contrast to HD cells had increased cholesterol. In addition, Htt(140Q/140Q) NS cells had higher phospho-AKT/AKT ratios than Htt(7Q/7Q) NS cells in resting conditions and after BDNF stimulation, suggesting mutant htt affects AKT dependent growth factor signaling. Upon differentiation, the Htt(7Q/7Q) and Htt(140Q/140Q) generated numerous Beta(III)-Tubulin- and GABA-positive neurons; however, after 15 days the cellular architecture of the differentiated Htt(140Q/140Q) cultures changed compared to Htt(7Q/7Q) cultures and included a marked increase of GFAP-positive cells. Our findings suggest that NS cells expressing endogenous mutant Htt will be useful for study of mechanisms of HD

  13. Epigenetic regulation of neural stem cell fate during corticogenesis.

    PubMed

    MuhChyi, Chai; Juliandi, Berry; Matsuda, Taito; Nakashima, Kinichi

    2013-10-01

    The cerebral cortex comprises over three quarters of the brain, and serves as structural basis for the sophisticated perceptual and cognitive functions. It develops from common multipotent neural stem cells (NSCs) that line the neural tube. Development of the NSCs encompasses sequential phases of progenitor expansion, neurogenesis, and gliogenesis along with the progression of developmental stages. Interestingly, NSCs steadfastly march through all of these phases and give rise to specific neural cell types in a temporally defined and highly predictable manner. Herein, we delineate the intrinsic and extrinsic factors that dictate the progression and tempo of NSC differentiation during cerebral cortex development, and how epigenetic modifications contribute to the dynamic properties of NSCs.

  14. Mechanical roles of apical constriction, cell elongation, and cell migration during neural tube formation in Xenopus.

    PubMed

    Inoue, Yasuhiro; Suzuki, Makoto; Watanabe, Tadashi; Yasue, Naoko; Tateo, Itsuki; Adachi, Taiji; Ueno, Naoto

    2016-12-01

    Neural tube closure is an important and necessary process during the development of the central nervous system. The formation of the neural tube structure from a flat sheet of neural epithelium requires several cell morphogenetic events and tissue dynamics to account for the mechanics of tissue deformation. Cell elongation changes cuboidal cells into columnar cells, and apical constriction then causes them to adopt apically narrow, wedge-like shapes. In addition, the neural plate in Xenopus is stratified, and the non-neural cells in the deep layer (deep cells) pull the overlying superficial cells, eventually bringing the two layers of cells to the midline. Thus, neural tube closure appears to be a complex event in which these three physical events are considered to play key mechanical roles. To test whether these three physical events are mechanically sufficient to drive neural tube formation, we employed a three-dimensional vertex model and used it to simulate the process of neural tube closure. The results suggest that apical constriction cued the bending of the neural plate by pursing the circumference of the apical surface of the neural cells. Neural cell elongation in concert with apical constriction further narrowed the apical surface of the cells and drove the rapid folding of the neural plate, but was insufficient for complete neural tube closure. Migration of the deep cells provided the additional tissue deformation necessary for closure. To validate the model, apical constriction and cell elongation were inhibited in Xenopus laevis embryos. The resulting cell and tissue shapes resembled the corresponding simulation results.

  15. Neural stem cells and Alzheimer's disease: challenges and hope.

    PubMed

    Zhongling Feng; Gang Zhao; Lei Yu

    2009-01-01

    Alzheimer's disease is characterized by degeneration and dysfunction of synapses and neurons in brain regions critical for learning and memory functions. The endogenous generation of new neurons in certain regions of the mature brain, derived from primitive cells termed neural stem cells, has raised hope that neural stem cells may be recruited for structural brain repair. Stem cell therapy has been suggested as a possible strategy for replacing damaged circuitry and restoring learning and memory abilities in patients with Alzheimer's disease. In this review, we outline the promising investigations that are raising hope, and understanding the challenges behind translating underlying stem cell biology into novel clinical therapeutic potential in Alzheimer's disease.

  16. Facilitating neural stem/progenitor cell niche calibration for neural lineage differentiation by polyelectrolyte multilayer films.

    PubMed

    Lee, I-Chi; Wu, Yu-Chieh

    2014-09-01

    Neural stem/progenitor cells (NSPCs) are a possible candidate for advancing development and lineage control in neural engineering. Differentiated protocols have been developed in this field to generate neural progeny and to establish neural networks. However, continued refinement is required to enhance differentiation specificity and prevent the generation of unwanted cell types. In this study, we fabricated a niche-modulated system to investigate surface effects on NSPC differentiation by the formation of polyelectrolyte multilayer (PEM) films governed by electrostatic interactions of poly-l-glutamine acid as a polyanion and poly-l-lysine as a polycation. The serum- and chemical agent-free system provided a clean and clear platform to observe in isolation the interaction between surface niche and stem cell differentiation. We found that NSPCs were inducible on PEM films of up to eight alternating layers. In addition, neurite outgrowth, neuron percentage, and synaptic function were regulated by layer number and the surface charge of the terminal layer. The average process outgrowth length was over 500μm on PLL/PLGA(n=7.5) only after 3 days of culture. Moreover, the quantity and quality of the differentiated neurons were enhanced as the number of layers increased, especially when the terminal layer was poly-l-lysine. Our results achieve important targets of neural engineering, including long processes, large neural network size, and large amounts of functional neurons. Our methodology for nanoscale control of material deposition can be successfully applied for surface modification, neural niche modulation, and neural engineering applications. Copyright © 2014. Published by Elsevier B.V.

  17. Gap Junction Proteins in the Blood-Brain Barrier Control Nutrient-Dependent Reactivation of Drosophila Neural Stem Cells

    PubMed Central

    Spéder, Pauline; Brand, Andrea H.

    2014-01-01

    Summary Neural stem cells in the adult brain exist primarily in a quiescent state but are reactivated in response to changing physiological conditions. How do stem cells sense and respond to metabolic changes? In the Drosophila CNS, quiescent neural stem cells are reactivated synchronously in response to a nutritional stimulus. Feeding triggers insulin production by blood-brain barrier glial cells, activating the insulin/insulin-like growth factor pathway in underlying neural stem cells and stimulating their growth and proliferation. Here we show that gap junctions in the blood-brain barrier glia mediate the influence of metabolic changes on stem cell behavior, enabling glia to respond to nutritional signals and reactivate quiescent stem cells. We propose that gap junctions in the blood-brain barrier are required to translate metabolic signals into synchronized calcium pulses and insulin secretion. PMID:25065772

  18. Gap junction proteins in the blood-brain barrier control nutrient-dependent reactivation of Drosophila neural stem cells.

    PubMed

    Spéder, Pauline; Brand, Andrea H

    2014-08-11

    Neural stem cells in the adult brain exist primarily in a quiescent state but are reactivated in response to changing physiological conditions. How do stem cells sense and respond to metabolic changes? In the Drosophila CNS, quiescent neural stem cells are reactivated synchronously in response to a nutritional stimulus. Feeding triggers insulin production by blood-brain barrier glial cells, activating the insulin/insulin-like growth factor pathway in underlying neural stem cells and stimulating their growth and proliferation. Here we show that gap junctions in the blood-brain barrier glia mediate the influence of metabolic changes on stem cell behavior, enabling glia to respond to nutritional signals and reactivate quiescent stem cells. We propose that gap junctions in the blood-brain barrier are required to translate metabolic signals into synchronized calcium pulses and insulin secretion. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  19. Human epidermal neural crest stem cells as a source of Schwann cells

    PubMed Central

    Sakaue, Motoharu; Sieber-Blum, Maya

    2015-01-01

    We show that highly pure populations of human Schwann cells can be derived rapidly and in a straightforward way, without the need for genetic manipulation, from human epidermal neural crest stem cells [hEPI-NCSC(s)] present in the bulge of hair follicles. These human Schwann cells promise to be a useful tool for cell-based therapies, disease modelling and drug discovery. Schwann cells are glia that support axons of peripheral nerves and are direct descendants of the embryonic neural crest. Peripheral nerves are damaged in various conditions, including through trauma or tumour-related surgery, and Schwann cells are required for their repair and regeneration. Schwann cells also promise to be useful for treating spinal cord injuries. Ex vivo expansion of hEPI-NCSC isolated from hair bulge explants, manipulating the WNT, sonic hedgehog and TGFβ signalling pathways, and exposure of the cells to pertinent growth factors led to the expression of the Schwann cell markers SOX10, KROX20 (EGR2), p75NTR (NGFR), MBP and S100B by day 4 in virtually all cells, and maturation was completed by 2 weeks of differentiation. Gene expression profiling demonstrated expression of transcripts for neurotrophic and angiogenic factors, as well as JUN, all of which are essential for nerve regeneration. Co-culture of hEPI-NCSC-derived human Schwann cells with rodent dorsal root ganglia showed interaction of the Schwann cells with axons, providing evidence of Schwann cell functionality. We conclude that hEPI-NCSCs are a biologically relevant source for generating large and highly pure populations of human Schwann cells. PMID:26251357

  20. Neural Cell Chip Based Electrochemical Detection of Nanotoxicity

    PubMed Central

    Kafi, Md. Abdul; Cho, Hyeon-Yeol; Choi, Jeong Woo

    2015-01-01

    Development of a rapid, sensitive and cost-effective method for toxicity assessment of commonly used nanoparticles is urgently needed for the sustainable development of nanotechnology. A neural cell with high sensitivity and conductivity has become a potential candidate for a cell chip to investigate toxicity of environmental influences. A neural cell immobilized on a conductive surface has become a potential tool for the assessment of nanotoxicity based on electrochemical methods. The effective electrochemical monitoring largely depends on the adequate attachment of a neural cell on the chip surfaces. Recently, establishment of integrin receptor specific ligand molecules arginine-glycine-aspartic acid (RGD) or its several modifications RGD-Multi Armed Peptide terminated with cysteine (RGD-MAP-C), C(RGD)4 ensure farm attachment of neural cell on the electrode surfaces either in their two dimensional (dot) or three dimensional (rod or pillar) like nano-scale arrangement. A three dimensional RGD modified electrode surface has been proven to be more suitable for cell adhesion, proliferation, differentiation as well as electrochemical measurement. This review discusses fabrication as well as electrochemical measurements of neural cell chip with particular emphasis on their use for nanotoxicity assessments sequentially since inception to date. Successful monitoring of quantum dot (QD), graphene oxide (GO) and cosmetic compound toxicity using the newly developed neural cell chip were discussed here as a case study. This review recommended that a neural cell chip established on a nanostructured ligand modified conductive surface can be a potential tool for the toxicity assessments of newly developed nanomaterials prior to their use on biology or biomedical technologies.

  1. Neural Cell Chip Based Electrochemical Detection of Nanotoxicity.

    PubMed

    Kafi, Md Abdul; Cho, Hyeon-Yeol; Choi, Jeong Woo

    2015-07-02

    Development of a rapid, sensitive and cost-effective method for toxicity assessment of commonly used nanoparticles is urgently needed for the sustainable development of nanotechnology. A neural cell with high sensitivity and conductivity has become a potential candidate for a cell chip to investigate toxicity of environmental influences. A neural cell immobilized on a conductive surface has become a potential tool for the assessment of nanotoxicity based on electrochemical methods. The effective electrochemical monitoring largely depends on the adequate attachment of a neural cell on the chip surfaces. Recently, establishment of integrin receptor specific ligand molecules arginine-glycine-aspartic acid (RGD) or its several modifications RGD-Multi Armed Peptide terminated with cysteine (RGD-MAP-C), C(RGD)₄ ensure farm attachment of neural cell on the electrode surfaces either in their two dimensional (dot) or three dimensional (rod or pillar) like nano-scale arrangement. A three dimensional RGD modified electrode surface has been proven to be more suitable for cell adhesion, proliferation, differentiation as well as electrochemical measurement. This review discusses fabrication as well as electrochemical measurements of neural cell chip with particular emphasis on their use for nanotoxicity assessments sequentially since inception to date. Successful monitoring of quantum dot (QD), graphene oxide (GO) and cosmetic compound toxicity using the newly developed neural cell chip were discussed here as a case study. This review recommended that a neural cell chip established on a nanostructured ligand modified conductive surface can be a potential tool for the toxicity assessments of newly developed nanomaterials prior to their use on biology or biomedical technologies.

  2. Generation and properties of a new human ventral mesencephalic neural stem cell line

    SciTech Connect

    Villa, Ana; Liste, Isabel; Courtois, Elise T.; Seiz, Emma G.; Ramos, Milagros; Meyer, Morten; Juliusson, Bengt; Kusk, Philip

    2009-07-01

    Neural stem cells (NSCs) are powerful research tools for the design and discovery of new approaches to cell therapy in neurodegenerative diseases like Parkinson's disease. Several epigenetic and genetic strategies have been tested for long-term maintenance and expansion of these cells in vitro. Here we report the generation of a new stable cell line of human neural stem cells derived from ventral mesencephalon (hVM1) based on v-myc immortalization. The cells expressed neural stem cell and radial glia markers like nestin, vimentin and 3CB2 under proliferation conditions. After withdrawal of growth factors, proliferation and expression of v-myc were dramatically reduced and the cells differentiated into astrocytes, oligodendrocytes and neurons. hVM1 cells yield a large number of dopaminergic neurons (about 12% of total cells are TH{sup +}) after differentiation, which also produce dopamine. In addition to proneural genes (NGN2, MASH1), differentiated cells show expression of several genuine mesencephalic dopaminergic markers such as: LMX1A, LMX1B, GIRK2, ADH2, NURR1, PITX3, VMAT2 and DAT, indicating that they retain their regional identity. Our data indicate that this cell line and its clonal derivatives may constitute good candidates for the study of development and physiology of human dopaminergic neurons in vitro, and to develop tools for Parkinson's disease cell replacement preclinical research and drug testing.

  3. Regulation of mouse embryonic stem cell neural differentiation by retinoic acid

    PubMed Central

    Kim, Mijeong; Habiba, Ayman; Doherty, Jason M.; Mills, Jason C.; Mercer, Robert W.; Huettner, James E.

    2009-01-01

    Pluripotent mouse embryonic stem cells (ESCs) derived from the early blastocyst can differentiate in vitro into a variety of somatic cell types including lineages from all three embryonic germ layers. Protocols for ES cell neural differentiation typically involve induction by retinoic acid (RA), or by exposure to growth factors or medium conditioned by other cell types. A serum-free differentiation (SFD) medium completely lacking exogenous retinoids was devised that allows for efficient conversion of aggregated mouse ESCs into neural precursors and immature neurons. Neural cells produced in this medium express neuronal ion channels, establish polarity, and form functional excitatory and inhibitory synapses. Brief exposure to RA during the period of cell aggregation speeds neuronal maturation and suppresses cell proliferation. Differentiation without RA yields neurons and neural progenitors with apparent telencephalic identity, whereas cells differentiated with exposure to RA express markers of hindbrain and spinal cord. Transcriptional profiling indicates a substantial representation of transit amplifying neuroblasts in SFD cultures not exposed to RA. PMID:19217899

  4. The stemness of neural crest cells and their derivatives.

    PubMed

    Kunisada, Takahiro; Tezulka, Ken-Ichi; Aoki, Hitomi; Motohashi, Tsutomu

    2014-09-01

    Neural crest cells (NCCs) are unique to vertebrates and emerge from the border of the neural plate and subsequently migrate extensively throughout the embryo after which they differentiate into many types of cells. This multipotency is the main reason why NCCs are regarded as a versatile tool for stem cell biology and have been gathering attention for their potential use in stem cell based therapy. Multiple sets of networks comprised of signaling molecules and transcription factors regulate every developmental phase of NCCs, including maintenance of their multipotency. Pluripotent stem cell lines, such as embryonic stem cells and induced pluripotent stem (iPS) cells, facilitate the induction of NCCs in combination with sophisticated culture systems used for neural stem cells, although at present, clinical experiments for NCC-based cell therapy need to be improved. Unexpectedly, the multipotency of NCCs is maintained after they reach the target tissues as tissue neural crest stem cells (NCSCs) that may contribute to the establishment of NCC-derived multipotential stem cells. In addition, under specific culture conditions, fate-restricted unipotent descendants of NCCs, such as melanoblasts, show multipotency to differentiate into melanocytes, neurons, and glia cells. These properties contribute to the additional versatility of NCCs for therapeutic application and to better understand NCC development. © 2014 Wiley Periodicals, Inc.

  5. Nanomedicine Approaches to Modulate Neural Stem Cells in Brain Repair.

    PubMed

    Santos, Tiago; Boto, Carlos; Saraiva, Cláudia M; Bernardino, Liliana; Ferreira, Lino

    2016-06-01

    We explore the concept of modulating neural stem cells and their niches for brain repair using nanotechnology-based approaches. These approaches include stimulating cell proliferation, recruitment, and differentiation to functionally recover damaged areas. Nanoscale-engineered materials potentially overcome limited crossing of the blood-brain barrier, deficient drug delivery, and cell targeting. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Transient cell–cell interactions in neural circuit formation

    PubMed Central

    Chao, Daniel L.; Ma, Le; Shen, Kang

    2011-01-01

    The wiring of the nervous system requires a complex orchestration of developmental events. Emerging evidence suggests that transient cell–cell interactions often serve as positional cues for axon guidance and synaptogenesis during the assembly of neural circuits. In contrast to the relatively stable cellular interactions between synaptic partners in mature circuits, these transient interactions involve cells that are not destined to be pre- or postsynaptic cells. Here we review the roles of these transient cell–cell interactions in a variety of developmental contexts and describe the mechanisms through which they organize neural connections. PMID:19300445

  7. Mediators in cell growth and differentiation

    SciTech Connect

    Ford, R.J.; Maizel, A.L.

    1985-01-01

    This book contains papers divided among seven sections. The section headings are: Cell Cycle and Control of Cell Growth, Growth Factors for Nonlymphoid Cells, Colony-Stimulating Factors, Stem Cells and Hematopoiesis, Lymphoid Growth Factors, Growth Factors in Neoplasia, Interferon, and Differentiation in Normal and Neoplastic Cells.

  8. Adult subependymal neural precursors, but not differentiated cells, undergo rapid cathodal migration in the presence of direct current electric fields.

    PubMed

    Babona-Pilipos, Robart; Droujinine, Ilia A; Popovic, Milos R; Morshead, Cindi M

    2011-01-01

    The existence of neural stem and progenitor cells (together termed neural precursor cells) in the adult mammalian brain has sparked great interest in utilizing these cells for regenerative medicine strategies. Endogenous neural precursors within the adult forebrain subependyma can be activated following injury, resulting in their proliferation and migration toward lesion sites where they differentiate into neural cells. The administration of growth factors and immunomodulatory agents following injury augments this activation and has been shown to result in behavioural functional recovery following stroke. With the goal of enhancing neural precursor migration to facilitate the repair process we report that externally applied direct current electric fields induce rapid and directed cathodal migration of pure populations of undifferentiated adult subependyma-derived neural precursors. Using time-lapse imaging microscopy in vitro we performed an extensive single-cell kinematic analysis demonstrating that this galvanotactic phenomenon is a feature of undifferentiated precursors, and not differentiated phenotypes. Moreover, we have shown that the migratory response of the neural precursors is a direct effect of the electric field and not due to chemotactic gradients. We also identified that epidermal growth factor receptor (EGFR) signaling plays a role in the galvanotactic response as blocking EGFR significantly attenuates the migratory behaviour. These findings suggest direct current electric fields may be implemented in endogenous repair paradigms to promote migration and tissue repair following neurotrauma.

  9. Electrical Property Characterization of Neural Stem Cells in Differentiation

    PubMed Central

    Sun, He; Chen, Deyong; Li, Zhaohui; Fan, Beiyuan; George, Julian; Xue, Chengcheng; Cui, Zhanfeng; Wang, Junbo

    2016-01-01

    Electrical property characterization of stem cells could be utilized as a potential label-free biophysical approach to evaluate the differentiation process. However, there has been a lack of technology or tools that can quantify the intrinsic cellular electrical markers (e.g., specific membrane capacitance (Cspecific membrane) and cytoplasm conductivity (σcytoplasm)) for a large amount of stem cells or differentiated cells. In this paper, a microfluidic platform enabling the high-throughput quantification of Cspecific membrane and σcytoplasm from hundreds of single neural stem cells undergoing differentiation was developed to explore the feasibility to characterize the neural stem cell differentiation process without biochemical staining. Experimental quantification using biochemical markers (e.g., Nestin, Tubulin and GFAP) of neural stem cells confirmed the initiation of the differentiation process featured with gradual loss in cellular stemness and increased cell markers for neurons and glial cells. The recorded electrical properties of neural stem cells undergoing differentiation showed distinctive and unique patterns: 1) in the suspension culture before inducing differentiation, a large distribution and difference in σcytoplasm among individual neural stem cells was noticed, which indicated heterogeneity that may result from the nature of suspension culture of neurospheres; and 2) during the differentiation in adhering monolayer culture, significant changes and a large difference in Cspecific membrane were located indicating different expressions of membrane proteins during the differentiation process, and a small distribution difference in σcytoplasm was less significant that indicated the relatively consistent properties of cytoplasm during the culture. In summary, significant differences in Cspecific membrane and σcytoplasm were observed during the neural stem cell differentiation process, which may potentially be used as label-free biophysical markers

  10. MIO-M1 cells and similar muller glial cell lines derived from adult human retina exhibit neural stem cell characteristics.

    PubMed

    Lawrence, Jean M; Singhal, Shweta; Bhatia, Bhairavi; Keegan, David J; Reh, Thomas A; Luthert, Philip J; Khaw, Peng T; Limb, Gloria Astrid

    2007-08-01

    Growing evidence suggests that glial cells may have a role as neural precursors in the adult central nervous system. Although it has been shown that Müller cells exhibit progenitor characteristics in the postnatal chick and rat retinae, their progenitor-like role in developed human retina is unknown. We first reported the Müller glial characteristics of the spontaneously immortalized human cell line MIO-M1, but recently we have derived similar cell lines from the neural retina of several adult eye donors. Since immortalization is one of the main properties of stem cells, we investigated whether these cells expressed stem cell markers. Cells were grown as adherent monolayers, responded to epidermal growth factor, and could be expanded indefinitely without growth factors under normal culture conditions. They could be frozen and thawed without losing their characteristics. In the presence of extracellular matrix and fibroblast growth factor-2 or retinoic acid, they acquired neural morphology, formed neurospheres, and expressed neural stem cell markers including betaIII tubulin, Sox2, Pax6, Chx10, and Notch 1. They also expressed markers of postmitotic retinal neurons, including peripherin, recoverin, calretinin, S-opsin, and Brn3. When grafted into the subretinal space of dystrophic Royal College of Surgeons rats or neonatal Lister hooded rats, immortalized cells migrated into the retina, where they expressed various markers of retinal neurons. These observations indicate that adult human neural retina harbors a population of cells that express both Müller glial and stem cell markers and suggest that these cells may have potential use for cell-based therapies to restore retinal function. Disclosure of potential conflicts of interest is found at the end of this article.

  11. Expression of polysialylated neural cell adhesion molecules on adult stem cells after neuronal differentiation of inner ear spiral ganglion neurons.

    PubMed

    Park, Kyoung Ho; Yeo, Sang Won; Troy, Frederic A

    2014-10-17

    During brain development, polysialylated (polySia) neural cell adhesion molecules (polySia-NCAMs) modulate cell-cell adhesive interactions involved in synaptogenesis, neural plasticity, myelination, and neural stem cell (NSC) proliferation and differentiation. Our findings show that polySia-NCAM is expressed on NSC isolated from adult guinea pig spiral ganglion (GPSG), and in neurons and Schwann cells after differentiation of the NSC with epidermal, glia, fibroblast growth factors (GFs) and neurotrophins. These differentiated cells were immunoreactive with mAb's to polySia, NCAM, β-III tubulin, nestin, S-100 and stained with BrdU. NSC could regenerate and be differentiated into neurons and Schwann cells. We conclude: (1) polySia is expressed on NSC isolated from adult GPSG and on neurons and Schwann cells differentiated from these NSC; (2) polySia is expressed on neurons primarily during the early stage of neuronal development and is expressed on Schwann cells at points of cell-cell contact; (3) polySia is a functional biomarker that modulates neuronal differentiation in inner ear stem cells. These new findings suggest that replacement of defective cells in the inner ear of hearing impaired patients using adult spiral ganglion neurons may offer potential hope to improve the quality of life for patients with auditory dysfunction and impaired hearing disorders.

  12. ETOH inhibits embryonic neural stem/precursor cell proliferation via PLD signaling

    SciTech Connect

    Fujita, Yuko; Hiroyama, Masami; Sanbe, Atsushi Yamauchi, Junji; Murase, Shoko; Tanoue, Akito

    2008-05-23

    While a mother's excessive alcohol consumption during pregnancy is known to have adverse effects on fetal neural development, little is known about the underlying mechanism of these effects. In order to investigate these mechanisms, we investigated the toxic effect of ethanol (ETOH) on neural stem/precursor cell (NSC) proliferation. In cultures of NSCs, phospholipase D (PLD) is activated following stimulation with epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF2). Exposure of NSCs to ETOH suppresses cell proliferation, while it has no effect on cell death. Phosphatidic acid (PA), which is a signaling messenger produced by PLD, reverses ETOH inhibition of NSC proliferation. Blocking the PLD signal by 1-butanol suppresses the proliferation. ETOH-induced suppression of NSC proliferation and the protective effect of PA for ETOH-induced suppression are mediated through extracellular signal-regulated kinase signaling. These results indicate that exposure to ETOH impairs NSC proliferation by altering the PLD signaling pathway.

  13. ETOH inhibits embryonic neural stem/precursor cell proliferation via PLD signaling.

    PubMed

    Fujita, Yuko; Hiroyama, Masami; Sanbe, Atsushi; Yamauchi, Junji; Murase, Shoko; Tanoue, Akito

    2008-05-23

    While a mother's excessive alcohol consumption during pregnancy is known to have adverse effects on fetal neural development, little is known about the underlying mechanism of these effects. In order to investigate these mechanisms, we investigated the toxic effect of ethanol (ETOH) on neural stem/precursor cell (NSC) proliferation. In cultures of NSCs, phospholipase D (PLD) is activated following stimulation with epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF2). Exposure of NSCs to ETOH suppresses cell proliferation, while it has no effect on cell death. Phosphatidic acid (PA), which is a signaling messenger produced by PLD, reverses ETOH inhibition of NSC proliferation. Blocking the PLD signal by 1-butanol suppresses the proliferation. ETOH-induced suppression of NSC proliferation and the protective effect of PA for ETOH-induced suppression are mediated through extracellular signal-regulated kinase signaling. These results indicate that exposure to ETOH impairs NSC proliferation by altering the PLD signaling pathway.

  14. Pleiotrophin enhances PDGFB-induced gliomagenesis through increased proliferation of neural progenitor cells.

    PubMed

    Zhang, Lei; Laaniste, Liisi; Jiang, Yiwen; Alafuzoff, Irina; Uhrbom, Lene; Dimberg, Anna

    2016-12-06

    Pleiotrophin (PTN) augments tumor growth by increasing proliferation of tumor cells and promoting vascular abnormalization, but its role in early gliomagenesis has not been evaluated. Through analysis of publically available datasets, we demonstrate that increased PTN mRNA expression is associated with amplification of chromosome 7, identified as one of the earliest steps in glioblastoma development. To elucidate the role of PTN in tumor initiation we employed the RCAS/tv-a model that allows glioma induction by RCAS-virus mediated expression of oncogenes in neural progenitor cells. Intracranial injection of RCAS-PTN did not induce glioma formation when administrated alone, but significantly enhanced RCAS-platelet derived growth factor (PDGF)B-induced gliomagenesis. PTN co-treatment augmented PDGFB-induced Akt activation in neural progenitor cells in vitro, and enhanced neural sphere size associated with increased proliferation. Our data indicates that PTN expression is associated with chromosome 7 gain, and that PTN enhances PDGFB-induced gliomagenesis by stimulating proliferation of neural progenitor cells.

  15. Three-dimensional graphene foam as a biocompatible and conductive scaffold for neural stem cells

    PubMed Central

    Li, Ning; Zhang, Qi; Gao, Song; Song, Qin; Huang, Rong; Wang, Long; Liu, Liwei; Dai, Jianwu; Tang, Mingliang; Cheng, Guosheng

    2013-01-01

    Neural stem cell (NSC) based therapy provides a promising approach for neural regeneration. For the success of NSC clinical application, a scaffold is required to provide three-dimensional (3D) cell growth microenvironments and appropriate synergistic cell guidance cues. Here, we report the first utilization of graphene foam, a 3D porous structure, as a novel scaffold for NSCs in vitro. It was found that three-dimensional graphene foams (3D-GFs) can not only support NSC growth, but also keep cell at an active proliferation state with upregulation of Ki67 expression than that of two-dimensional graphene films. Meanwhile, phenotypic analysis indicated that 3D-GFs can enhance the NSC differentiation towards astrocytes and especially neurons. Furthermore, a good electrical coupling of 3D-GFs with differentiated NSCs for efficient electrical stimulation was observed. Our findings implicate 3D-GFs could offer a powerful platform for NSC research, neural tissue engineering and neural prostheses. PMID:23549373

  16. Pleiotrophin enhances PDGFB-induced gliomagenesis through increased proliferation of neural progenitor cells

    PubMed Central

    Zhang, Lei; Laaniste, Liisi; Jiang, Yiwen; Alafuzoff, Irina; Uhrbom, Lene; Dimberg, Anna

    2016-01-01

    Pleiotrophin (PTN) augments tumor growth by increasing proliferation of tumor cells and promoting vascular abnormalization, but its role in early gliomagenesis has not been evaluated. Through analysis of publically available datasets, we demonstrate that increased PTN mRNA expression is associated with amplification of chromosome 7, identified as one of the earliest steps in glioblastoma development. To elucidate the role of PTN in tumor initiation we employed the RCAS/tv-a model that allows glioma induction by RCAS-virus mediated expression of oncogenes in neural progenitor cells. Intracranial injection of RCAS-PTN did not induce glioma formation when administrated alone, but significantly enhanced RCAS-platelet derived growth factor (PDGF)B-induced gliomagenesis. PTN co-treatment augmented PDGFB-induced Akt activation in neural progenitor cells in vitro, and enhanced neural sphere size associated with increased proliferation. Our data indicates that PTN expression is associated with chromosome 7 gain, and that PTN enhances PDGFB-induced gliomagenesis by stimulating proliferation of neural progenitor cells. PMID:27806344

  17. Cell Biology of Hyphal Growth.

    PubMed

    Steinberg, Gero; Peñalva, Miguel A; Riquelme, Meritxell; Wösten, Han A; Harris, Steven D

    2017-04-01

    Filamentous fungi are a large and ancient clade of microorganisms that occupy a broad range of ecological niches. The success of filamentous fungi is largely due to their elongate hypha, a chain of cells, separated from each other by septa. Hyphae grow by polarized exocytosis at the apex, which allows the fungus to overcome long distances and invade many substrates, including soils and host tissues. Hyphal tip growth is initiated by establishment of a growth site and the subsequent maintenance of the growth axis, with transport of growth supplies, including membranes and proteins, delivered by motors along the cytoskeleton to the hyphal apex. Among the enzymes delivered are cell wall synthases that are exocytosed for local synthesis of the extracellular cell wall. Exocytosis is opposed by endocytic uptake of soluble and membrane-bound material into the cell. The first intracellular compartment in the endocytic pathway is the early endosomes, which emerge to perform essential additional functions as spatial organizers of the hyphal cell. Individual compartments within septated hyphae can communicate with each other via septal pores, which allow passage of cytoplasm or organelles to help differentiation within the mycelium. This article introduces the reader to more detailed aspects of hyphal growth in fungi.

  18. Matrigel supports neural, melanocytic and chondrogenic differentiation of trunk neural crest cells.

    PubMed

    Ramos-Hryb, Ana B; Da-Costa, Meline C; Trentin, Andréa G; Calloni, Giordano W

    2013-01-01

    The neural crest (NC) is composed of highly multipotent precursor cells able to differentiate into both neural and mesenchymal phenotypes. Until now, most studies focusing on NC cell differentiation have been performed with traditional two-dimensional (2D) cell culture systems. However, such culture systems do not reflect the complex three-dimensional (3D) microenvironments of in vivo NC cells. To address this limitation, we have developed a method of Matrigel™ coating to create 2D and 3D microenvironments in the same culture well. When we performed cultures of trunk neural crest cells (TNCCs) on three different lots of basement membrane matrix (Matrigel™), we observed that all analyzed Matrigel™ lots were equally efficient in allowing the appearance of glial cells, neurons, melanocytes, smooth muscle cells and chondrocytes. We further observed that chondrocytes were found predominantly in the 3D microenvironment, whereas smooth muscle cells were almost exclusively located in the 2D microenvironment. Glial cells were present in both environments, but with broader quantities on the 2D surface. Melanocytes and neurons were equally distributed in both 2D and 3D microenvironments, but with distinct morphologies. It is worth noting the higher frequency of chondrocytes detected in this study using the 3D Matrigel™ microenvironment compared to previous reports of chondrogenesis obtained from TNCCs on traditional 2D cultures. In conclusion, Matrigel™ represents an attractive scaffold to study NC multipotentiality and differentiation, since it permits the appearance of the major NC phenotypes.

  19. ERK-dependent and -independent pathways trigger human neural progenitor cell migration

    SciTech Connect

    Moors, Michaela . E-mail: moors@uni-duesseldorf.de; Cline, Jason E. . E-mail: jason.cline@uni-duesseldorf.de; Abel, Josef . E-mail: josef.abel@uni-duesseldorf.de; Fritsche, Ellen . E-mail: ellen.fritsche@uni-duesseldorf.de

    2007-05-15

    Besides differentiation and apoptosis, cell migration is a basic process in brain development in which neural cells migrate several centimeters within the developing brain before reaching their proper positions and forming the right connections. For identifying signaling events that control neural migration and are therefore potential targets of chemicals to disturb normal brain development, we developed a human neurosphere-based migration assay based on normal human neural progenitor (NHNP) cells, in which the distance is measured that cells wander over time. Applying this assay, we investigated the role of the extracellular signal-regulated kinases 1 and 2 (ERK1/2) in the regulation of NHNP cell migration. Exposure to model substances like ethanol or phorbol 12-myristate 13-acetate (PMA) revealed a correlation between ERK1/2 activation and cell migration. The participation of phospho-(P-) ERK1/2 was confirmed by exposure of the cells to the MEK inhibitor PD98059, which directly prohibits ERK1/2 phosphorylation and inhibited cell migration. We identified protein kinase C (PKC) and epidermal growth factor receptor (EGFR) as upstream signaling kinases governing ERK1/2 activation, thereby controlling NHNP cell migration. Additionally, treatments with src kinase inhibitors led to a diminished cell migration without affecting ERK1/2 phosphorylation. Based on these results, we postulate that migration of NHNP cells is controlled via ERK1/2-dependent and -independent pathways.

  20. Neural crest stem cell multipotency requires Foxd3 to maintain neural potential and repress mesenchymal fates

    PubMed Central

    Mundell, Nathan A.; Labosky, Patricia A.

    2011-01-01

    Neural crest (NC) progenitors generate a wide array of cell types, yet molecules controlling NC multipotency and self-renewal and factors mediating cell-intrinsic distinctions between multipotent versus fate-restricted progenitors are poorly understood. Our earlier work demonstrated that Foxd3 is required for maintenance of NC progenitors in the embryo. Here, we show that Foxd3 mediates a fate restriction choice for multipotent NC progenitors with loss of Foxd3 biasing NC toward a mesenchymal fate. Neural derivatives of NC were lost in Foxd3 mutant mouse embryos, whereas abnormally fated NC-derived vascular smooth muscle cells were ectopically located in the aorta. Cranial NC defects were associated with precocious differentiation towards osteoblast and chondrocyte cell fates, and individual mutant NC from different anteroposterior regions underwent fate changes, losing neural and increasing myofibroblast potential. Our results demonstrate that neural potential can be separated from NC multipotency by the action of a single gene, and establish novel parallels between NC and other progenitor populations that depend on this functionally conserved stem cell protein to regulate self-renewal and multipotency. PMID:21228004

  1. Lingo-1 shRNA and Notch signaling inhibitor DAPT promote differentiation of neural stem/progenitor cells into neurons.

    PubMed

    Wang, Jue; Ye, Zhizhong; Zheng, Shuhui; Chen, Luming; Wan, Yong; Deng, Yubin; Yang, Ruirui

    2016-03-01

    Determination of the exogenous factors that regulate differentiation of neural stem/progenitor cells into neurons, oligodendrocytes and astrocytes is an important step in the clinical therapy of spinal cord injury (SCI). The Notch pathway inhibits the differentiation of neural stem/progenitor cells and Lingo-1 is a strong negative regulator for myelination and axon growth. While Lingo-1 shRNA and N-[N-(3, 5-difluorophenacetyl)-1-alanyl]-S-Phenylglycinet-butylester (DAPT), a Notch pathway inhibitor, have been used separately to help repair SCI, the results have been unsatisfactory. Here we investigated and elucidated the preliminary mechanism for the effect of Lingo-1 shRNA and DAPT on neural stem/progenitor cells differentiation. We found that neural stem/progenitor cells from E14 rat embryos expressed Nestin, Sox-2 and Lingo-1, and we optimized the transduction of neural stem/progenitor cells using lentiviral vectors encoding Lingo-1 shRNA. The addition of DAPT decreased the expression of Notch intracellular domain (NICD) as well as the downstream genes Hes1 and Hes5. Expression of NeuN, CNPase and GFAP in DAPT treated cells and expression of NeuN in Lingo-1 shRNA treated cells confirmed differentiation of neural stem/progenitor cells into neurons, oligodendrocytes and astrocytes. These results revealed that while Lingo-1 shRNA and Notch signaling inhibitor DAPT both promoted differentiation of neural stem cells into neurons, only DAPT was capable of driving neural stem/progenitor cells differentiation into oligodendrocytes and astrocytes. Since we were able to show that both Lingo-1 shRNA and DAPT could drive neural stem/progenitor cells differentiation, our data might aid the development of more effective SCI therapies using Lingo-1 shRNA and DAPT.

  2. Regulation of Asymmetric Cell Division in Mammalian Neural Stem and Cancer Precursor Cells.

    PubMed

    Daynac, Mathieu; Petritsch, Claudia K

    Stem and progenitor cells are characterized by their abilities to self-renew and produce differentiated progeny. The balance between self-renewal and differentiation is achieved through control of cell division mode, which can be either asymmetric or symmetric. Failure to properly control cell division mode may result in premature depletion of the stem/progenitor cell pool or abnormal growth and impaired differentiation. In many tissues, including the brain, stem cells and progenitor cells undergo asymmetric cell division through the establishment of cell polarity. Cell polarity proteins are therefore potentially critical regulators of asymmetric cell division. Decrease or loss of asymmetric cell division can be associated with reduced differentiation common during aging or impaired remyelination as seen in demyelinating diseases. Progenitor-like glioma precursor cells show decreased asymmetric cell division rates and increased symmetric divisions, which suggests that asymmetric cell division suppresses brain tumor formation. Cancer stem cells, on the other hand, still undergo low rates of asymmetric cell division, which may provide them with a survival advantage during therapy. These findings led to the hypotheses that asymmetric cell divisions are not always tumor suppressive but can also be utilized to maintain a cancer stem cell population. Proper control of cell division mode is therefore not only deemed necessary to generate cellular diversity during development and to maintain adult tissue homeostasis but may also prevent disease and determine disease progression. Since brain cancer is most common in the adult and aging population, we review here the current knowledge on molecular mechanisms that regulate asymmetric cell divisions in the neural and oligodendroglial lineage during development and in the adult brain.

  3. Cell movements of the deep layer of non-neural ectoderm underlie complete neural tube closure in Xenopus.

    PubMed

    Morita, Hitoshi; Kajiura-Kobayashi, Hiroko; Takagi, Chiyo; Yamamoto, Takamasa S; Nonaka, Shigenori; Ueno, Naoto

    2012-04-01

    In developing vertebrates, the neural tube forms from a sheet of neural ectoderm by complex cell movements and morphogenesis. Convergent extension movements and the apical constriction along with apical-basal elongation of cells in the neural ectoderm are thought to be essential for the neural tube closure (NTC) process. In addition, it is known that non-neural ectoderm also plays a crucial role in this process, as the neural tube fails to close in the absence of this tissue in chick and axolotl. However, the cellular and molecular mechanisms by which it functions in NTC are as yet unclear. We demonstrate here that the non-neural superficial epithelium moves in the direction of tensile forces applied along the dorsal-ventral axis during NTC. We found that this force is partly attributable to the deep layer of non-neural ectoderm cells, which moved collectively towards the dorsal midline along with the superficial layer. Moreover, inhibition of this movement by deleting integrin β1 function resulted in incomplete NTC. Furthermore, we demonstrated that other proposed mechanisms, such as oriented cell division, cell rearrangement and cell-shape changes have no or only minor roles in the non-neural movement. This study is the first to demonstrate dorsally oriented deep-cell migration in non-neural ectoderm, and suggests that a global reorganization of embryo tissues is involved in NTC.

  4. Establishment of Human Neural Progenitor Cells from Human Induced Pluripotent Stem Cells with Diverse Tissue Origins

    PubMed Central

    Fukusumi, Hayato; Shofuda, Tomoko; Bamba, Yohei; Yamamoto, Atsuyo; Kanematsu, Daisuke; Handa, Yukako; Okita, Keisuke; Nakamura, Masaya; Yamanaka, Shinya; Okano, Hideyuki; Kanemura, Yonehiro

    2016-01-01

    Human neural progenitor cells (hNPCs) have previously been generated from limited numbers of human induced pluripotent stem cell (hiPSC) clones. Here, 21 hiPSC clones derived from human dermal fibroblasts, cord blood cells, and peripheral blood mononuclear cells were differentiated using two neural induction methods, an embryoid body (EB) formation-based method and an EB formation method using dual SMAD inhibitors (dSMADi). Our results showed that expandable hNPCs could be generated from hiPSC clones with diverse somatic tissue origins. The established hNPCs exhibited a mid/hindbrain-type neural identity and uniform expression of neural progenitor genes. PMID:27212953

  5. Rapid determination of bacterial abundance, biovolume, morphology, and growth by neural network-based image analysis

    PubMed

    Blackburn; Hagstrom; Wikner; Cuadros-Hansson; Bjornsen

    1998-09-01

    Annual bacterial plankton dynamics at several depths and locations in the Baltic Sea were studied by image analysis. Individual bacteria were classified by using an artificial neural network which also effectively identified nonbacterial objects. Cell counts and frequencies of dividing cells were determined, and the data obtained agreed well with visual observations and previously published values. Cell volumes were measured accurately by comparison with bead standards. The survey included 690 images from a total of 138 samples. Each image contained approximately 200 bacteria. The images were analyzed automatically at a rate of 100 images per h. Bacterial abundance exhibited coherent patterns with time and depth, and there were distinct subsurface peaks in the summer months. Four distinct morphological classes were resolved by the image analyzer, and the dynamics of each could be visualized. The bacterial growth rates estimated from frequencies of dividing cells were different from the bacterial growth rates estimated by the thymidine incorporation method. With minor modifications, the image analysis technique described here can be used to analyze other planktonic classes.

  6. Rapid Determination of Bacterial Abundance, Biovolume, Morphology, and Growth by Neural Network-Based Image Analysis

    PubMed Central

    Blackburn, Nicholas; Hagström, Åke; Wikner, Johan; Cuadros-Hansson, Rocio; Bjørnsen, Peter Koefoed

    1998-01-01

    Annual bacterial plankton dynamics at several depths and locations in the Baltic Sea were studied by image analysis. Individual bacteria were classified by using an artificial neural network which also effectively identified nonbacterial objects. Cell counts and frequencies of dividing cells were determined, and the data obtained agreed well with visual observations and previously published values. Cell volumes were measured accurately by comparison with bead standards. The survey included 690 images from a total of 138 samples. Each image contained approximately 200 bacteria. The images were analyzed automatically at a rate of 100 images per h. Bacterial abundance exhibited coherent patterns with time and depth, and there were distinct subsurface peaks in the summer months. Four distinct morphological classes were resolved by the image analyzer, and the dynamics of each could be visualized. The bacterial growth rates estimated from frequencies of dividing cells were different from the bacterial growth rates estimated by the thymidine incorporation method. With minor modifications, the image analysis technique described here can be used to analyze other planktonic classes. PMID:9726867

  7. Chemically Induced Reprogramming of Somatic Cells to Pluripotent Stem Cells and Neural Cells

    PubMed Central

    Biswas, Dhruba; Jiang, Peng

    2016-01-01

    The ability to generate transplantable neural cells in a large quantity in the laboratory is a critical step in the field of developing stem cell regenerative medicine for neural repair. During the last few years, groundbreaking studies have shown that cell fate of adult somatic cells can be reprogrammed through lineage specific expression of transcription factors (TFs)-and defined culture conditions. This key concept has been used to identify a number of potent small molecules that could enhance the efficiency of reprogramming with TFs. Recently, a growing number of studies have shown that small molecules targeting specific epigenetic and signaling pathways can replace all of the reprogramming TFs. Here, we provide a detailed review of the studies reporting the generation of chemically induced pluripotent stem cells (ciPSCs), neural stem cells (ciNSCs), and neurons (ciN). We also discuss the main mechanisms of actions and the pathways that the small molecules regulate during chemical reprogramming. PMID:26861316

  8. Chemically Induced Reprogramming of Somatic Cells to Pluripotent Stem Cells and Neural Cells.

    PubMed

    Biswas, Dhruba; Jiang, Peng

    2016-02-06

    The ability to generate transplantable neural cells in a large quantity in the laboratory is a critical step in the field of developing stem cell regenerative medicine for neural repair. During the last few years, groundbreaking studies have shown that cell fate of adult somatic cells can be reprogrammed through lineage specific expression of transcription factors (TFs)-and defined culture conditions. This key concept has been used to identify a number of potent small molecules that could enhance the efficiency of reprogramming with TFs. Recently, a growing number of studies have shown that small molecules targeting specific epigenetic and signaling pathways can replace all of the reprogramming TFs. Here, we provide a detailed review of the studies reporting the generation of chemically induced pluripotent stem cells (ciPSCs), neural stem cells (ciNSCs), and neurons (ciN). We also discuss the main mechanisms of actions and the pathways that the small molecules regulate during chemical reprogramming.

  9. Slit/Robo1 signaling regulates neural tube development by balancing neuroepithelial cell proliferation and differentiation.

    PubMed

    Wang, Guang; Li, Yan; Wang, Xiao-yu; Han, Zhe; Chuai, Manli; Wang, Li-jing; Ho Lee, Kenneth Ka; Geng, Jian-guo; Yang, Xuesong

    2013-05-01

    Formation of the neural tube is the morphological hallmark for development of the embryonic central nervous system (CNS). Therefore, neural tube development is a crucial step in the neurulation process. Slit/Robo signaling was initially identified as a chemo-repellent that regulated axon growth cone elongation, but its role in controlling neural tube development is currently unknown. To address this issue, we investigated Slit/Robo1 signaling in the development of chick neCollege of Life Sciences Biocentre, University of Dundee, Dundee DD1 5EH, UKural tube and transgenic mice over-expressing Slit2. We disrupted Slit/Robo1 signaling by injecting R5 monoclonal antibodies into HH10 neural tubes to block the Robo1 receptor. This inhibited the normal development of the ventral body curvature and caused the spinal cord to curl up into a S-shape. Next, Slit/Robo1 signaling on one half-side of the chick embryo neural tube was disturbed by electroporation in ovo. We found that the morphology of the neural tube was dramatically abnormal after we interfered with Slit/Robo1 signaling. Furthermore, we established that silencing Robo1 inhibited cell proliferation while over-expressing Robo1 enhanced cell proliferation. We also investigated the effects of altering Slit/Robo1 expression on Sonic Hedgehog (Shh) and Pax7 expression in the developing neural tube. We demonstrated that over-expressing Robo1 down-regulated Shh expression in the ventral neural tube and resulted in the production of fewer HNK-1(+) migrating neural crest cells (NCCs). In addition, Robo1 over-expression enhanced Pax7 expression in the dorsal neural tube and increased the number of Slug(+) pre-migratory NCCs. Conversely, silencing Robo1 expression resulted in an enhanced Shh expression and more HNK-1(+) migrating NCCs but reduced Pax7 expression and fewer Slug(+) pre-migratory NCCs were observed. In conclusion, we propose that Slit/Robo1 signaling is involved in regulating neural tube development by tightly

  10. Role of morphogens in neural crest cell determination.

    PubMed

    Jones, Natalie C; Trainor, Paul A

    2005-09-15

    The neural crest is a transient, migratory cell population found in all vertebrate embryos that generate a diverse range of cell and tissue derivatives including, but not limited, to the neurons and glia of the peripheral nervous system, smooth muscle, connective tissue, melanocytes, craniofacial cartilage, and bone. Over the past few years, many studies have provided tremendous insights into understanding the mechanisms regulating the induction and migration of neural crest cell development. This review highlights the surprising and perhaps unexpected roles for morphogens in these distinct processes. A comparison of studies performed in several different vertebrates emphasizes the requirement for coordination between multiple signaling pathways in the induction and migration of neural crest cells in the developing embryo. (c) 2005 Wiley Periodicals, Inc.

  11. Neural Network Modeling of Degradation of Solar Cells

    SciTech Connect

    Gupta, Himanshu; Ghosh, Bahniman; Banerjee, Sanjay K.

    2011-05-25

    Neural network modeling has been used to predict the degradation in conversion efficiency of solar cells in this work. The model takes intensity of light, temperature and exposure time as inputs and predicts the conversion efficiency of the solar cell. Backpropagation algorithm has been used to train the network. It is found that the neural network model satisfactorily predicts the degradation in efficiency of the solar cell with exposure time. The error in the computed results, after comparison with experimental results, lies in the range of 0.005-0.01, which is quite low.

  12. Induction of Excess Centrosomes in Neural Progenitor Cells during the Development of Radiation-Induced Microcephaly

    PubMed Central

    Shimada, Mikio; Matsuzaki, Fumio; Kato, Akihiro; Kobayashi, Junya; Matsumoto, Tomohiro; Komatsu, Kenshi

    2016-01-01

    The embryonic brain is one of the tissues most vulnerable to ionizing radiation. In this study, we showed that ionizing radiation induces apoptosis in the neural progenitors of the mouse cerebral cortex, and that the surviving progenitor cells subsequently develop a considerable amount of supernumerary centrosomes. When mouse embryos at Day 13.5 were exposed to γ-rays, brains sizes were reduced markedly in a dose-dependent manner, and these size reductions persisted until birth. Immunostaining with caspase-3 antibodies showed that apoptosis occurred in 35% and 40% of neural progenitor cells at 4 h after exposure to 1 and 2 Gy, respectively, and this was accompanied by a disruption of the apical layer in which mitotic spindles were positioned in unirradiated mice. At 24 h after 1 Gy irradiation, the apoptotic cells were completely eliminated and proliferation was restored to a level similar to that of unirradiated cells, but numerous spindles were localized outside the apical layer. Similarly, abnormal cytokinesis, which included multipolar division and centrosome clustering, was observed in 19% and 24% of the surviving neural progenitor cells at 48 h after irradiation with 1 and 2 Gy, respectively. Because these cytokinesis aberrations derived from excess centrosomes result in growth delay and mitotic catastrophe-mediated cell elimination, our findings suggest that, in addition to apoptosis at an early stage of radiation exposure, radiation-induced centrosome overduplication could contribute to the depletion of neural progenitors and thereby lead to microcephaly. PMID:27367050

  13. Dynamic transcriptional signature and cell fate analysis reveals plasticity of individual neural plate border cells

    PubMed Central

    Roellig, Daniela; Tan-Cabugao, Johanna; Esaian, Sevan; Bronner, Marianne E

    2017-01-01

    The ‘neural plate border’ of vertebrate embryos contains precursors of neural crest and placode cells, both defining vertebrate characteristics. How these lineages segregate from neural and epidermal fates has been a matter of debate. We address this by performing a fine-scale quantitative temporal analysis of transcription factor expression in the neural plate border of chick embryos. The results reveal significant overlap of transcription factors characteristic of multiple lineages in individual border cells from gastrula through neurula stages. Cell fate analysis using a Sox2 (neural) enhancer reveals that cells that are initially Sox2+ cells can contribute not only to neural tube but also to neural crest and epidermis. Moreover, modulating levels of Sox2 or Pax7 alters the apportionment of neural tube versus neural crest fates. Our results resolve a long-standing question and suggest that many individual border cells maintain ability to contribute to multiple ectodermal lineages until or beyond neural tube closure. DOI: http://dx.doi.org/10.7554/eLife.21620.001 PMID:28355135

  14. Stem cell media culture of melanoma results in the induction of a nonrepresentative neural expression profile.

    PubMed

    Anaka, Matthew; Freyer, Claudia; Gedye, Craig; Caballero, Otavia; Davis, Ian D; Behren, Andreas; Cebon, Jonathan

    2012-02-01

    The ability of cell lines to accurately represent cancer is a major concern in preclinical research. Culture of glioma cells as neurospheres in stem cell media (SCM) has been shown to better represent the genotype and phenotype of primary glioblastoma in comparison to serum cell lines. Despite the use of neurosphere-like models of many malignancies, there has been no robust analysis of whether other cancers benefit from a more representative phenotype and genotype when cultured in SCM. We analyzed the growth properties, transcriptional profile, and genotype of melanoma cells grown de novo in SCM, as while melanocytes share a common precursor with neural cells, melanoma frequently demonstrates divergent behavior in cancer stem cell assays. SCM culture of melanoma cells induced a neural lineage gene expression profile that was not representative of matched patient tissue samples and which could be induced in serum cell lines by switching them into SCM. There was no enrichment for expression of putative melanoma stem cell markers, but the SCM expression profile did overlap significantly with that of SCM cultures of glioma, suggesting that the observed phenotype is media-specific rather than melanoma-specific. Xenografts derived from either culture condition provided the best representation of melanoma in situ. Finally, SCM culture of melanoma did not prevent ongoing acquisition of DNA copy number abnormalities. In conclusion, SCM culture of melanoma does not provide a better representation of the phenotype or genotype of metastatic melanoma, and the resulting neural bias could potentially confound therapeutic target identification.

  15. Substrate-mediated reprogramming of human fibroblasts into neural crest stem-like cells and their applications in neural repair.

    PubMed

    Tseng, Ting-Chen; Hsieh, Fu-Yu; Dai, Niann-Tzyy; Hsu, Shan-Hui

    2016-09-01

    Cell- and gene-based therapies have emerged as promising strategies for treating neurological diseases. The sources of neural stem cells are limited while the induced pluripotent stem (iPS) cells have risk of tumor formation. Here, we proposed the generation of self-renewable, multipotent, and neural lineage-related neural crest stem-like cells by chitosan substrate-mediated gene transfer of a single factor forkhead box D3 (FOXD3) for the use in neural repair. A simple, non-toxic, substrate-mediated method was applied to deliver the naked FOXD3 plasmid into human fibroblasts. The transfection of FOXD3 increased cell proliferation and up-regulated the neural crest marker genes (FOXD3, SOX2, and CD271), stemness marker genes (OCT4, NANOG, and SOX2), and neural lineage-related genes (Nestin, β-tubulin and GFAP). The expression levels of stemness marker genes and neural crest maker genes in the FOXD3-transfected fibroblasts were maintained until the fifth passage. The FOXD3 reprogrammed fibroblasts based on the new method significantly rescued the neural function of the impaired zebrafish. The chitosan substrate-mediated delivery of naked plasmid showed feasibility in reprogramming somatic cells. Particularly, the FOXD3 reprogrammed fibroblasts hold promise as an easily accessible cellular source with neural crest stem-like behavior for treating neural diseases in the future.

  16. Regulated GDNF Delivery in Vivo Using Neural Stem Cells

    DTIC Science & Technology

    2007-04-01

    other neurodegenerative models including amyotrophic lateral sclerosis (ALS) and stroke (Kaspar et al., 2003; Cao et al., 2003; Guan et al., 2001...learning more about stem cell drug delivery it may be possible to explore other therapies for war injuries in the future. References Bilak...Neural Stem Cells PRINCIPAL INVESTIGATOR: Clive Svendsen CONTRACTING ORGANIZATION: University of Wisconsin-Madison

  17. Induction of neural stem cell-like cells (NSCLCs) from mouse astrocytes by Bmi1

    SciTech Connect

    Moon, Jai-Hee; Yoon, Byung Sun; Kim, Bona; Park, Gyuman; Jung, Hye-Youn; Maeng, Isaac; Jun, Eun Kyoung; Yoo, Seung Jun; Kim, Aeree; Oh, Sejong; Whang, Kwang Youn; Kim, Hyunggee; Kim, Dong-Wook; Kim, Ki Dong; You, Seungkwon

    2008-06-27

    Recently, Bmi1 was shown to control the proliferation and self-renewal of neural stem cells (NSCs). In this study, we demonstrated the induction of NSC-like cells (NSCLCs) from mouse astrocytes by Bmi1 under NSC culture conditions. These NSCLCs exhibited the morphology and growth properties of NSCs, and expressed NSC marker genes, including nestin, CD133, and Sox2. In vitro differentiation of NSCLCs resulted in differentiated cell populations containing astrocytes, neurons, and oligodendrocytes. Following treatment with histone deacetylase inhibitors (trichostatin A and valproic acid), the potential of NSCLCs for proliferation, dedifferentiation, and self-renewal was significantly inhibited. Our data indicate that multipotent NSCLCs can be generated directly from astrocytes by the addition of Bmi1.

  18. Mechanism of cell fate choice between neural and mesodermal development during early embryogenesis.

    PubMed

    Takemoto, Tatsuya

    2013-06-01

    During early embryogenesis, Sox2 expression distinguishes the neural plate from other embryonic domains, suggesting that the mechanism underlying the activation of the Sox2 gene is highly relevant to the development of this tissue. At the earliest stages of neural plate development, the Sox2 enhancer N1 regulates Sox2 expression in the extending posterior end of the neural plate. The N1 enhancer is initially activated in the axial stem cells, bipotential precursors of both neural and mesodermal lineages, therefore the activation does not immediately lead to Sox2 expression. A population of axial stem cells that remains in the superficial layer starts expressing Sox2, whereas another population that migrates through the primitive streak loses the N1 activity and becomes mesoderm. Multiple signaling cascades and transcription factors, including Wnt, fibroblast growth factor (FGF), bone morphogenetic protein (BMP) and Tbx6, are responsible for the regulation of Sox2 expression in axial stem cells to guide the development of the posterior neural plate and paraxial mesoderm.

  19. Isolation and manipulation of mammalian neural stem cells in vitro.

    PubMed

    Giachino, Claudio; Basak, Onur; Taylor, Verdon

    2009-01-01

    Neural stem cells are potentially a source of cells not only for replacement therapy but also as drug vectors, bringing bioactive molecules into the brain. Stem cell-like cells can be isolated readily from the human brain, thus, it is important to find culture systems that enable expansion in a multipotent state to generate cells that are of potential use for therapy. Currently, two systems have been described for the maintenance and expansion of multipotent progenitors, an adhesive substrate bound and the neurosphere culture. Both systems have pros and cons, but the neurosphere may be able to simulate the three-dimensional environment of the niche in which the cells reside in vivo. Thus, the neurosphere, when used and cultured appropriately, can expand and provide important information about the mechanisms that potentially control neural stem cells in vivo.

  20. Insulin concentration is critical in culturing human neural stem cells and neurons

    PubMed Central

    Rhee, Y-H; Choi, M; Lee, H-S; Park, C-H; Kim, S-M; Yi, S-H; Oh, S-M; Cha, H-J; Chang, M-Y; Lee, S-H

    2013-01-01

    Cell culture of human-derived neural stem cells (NSCs) is a useful tool that contributes to our understanding of human brain development and allows for the development of therapies for intractable human brain disorders. Human NSC (hNSC) cultures, however, are not commonly used, mainly because of difficulty with consistently maintaining the cells in a healthy state. In this study, we show that hNSC cultures, unlike NSCs of rodent origins, are extremely sensitive to insulin, an indispensable culture supplement, and that the previously reported difficulty in culturing hNSCs is likely because of a lack of understanding of this relationship. Like other neural cell cultures, insulin is required for hNSC growth, as withdrawal of insulin supplementation results in massive cell death and delayed cell growth. However, severe apoptotic cell death was also detected in insulin concentrations optimized to rodent NSC cultures. Thus, healthy hNSC cultures were only produced in a narrow range of relatively low insulin concentrations. Insulin-mediated cell death manifested not only in all human NSCs tested, regardless of origin, but also in differentiated human neurons. The underlying cell death mechanism at high insulin concentrations was similar to insulin resistance, where cells became less responsive to insulin, resulting in a reduction in the activation of the PI3K/Akt pathway critical to cell survival signaling. PMID:23928705

  1. Designer Self-Assembling Peptide Nanofiber Scaffolds for Adult Mouse Neural Stem Cell 3-Dimensional Cultures

    PubMed Central

    Gelain, Fabrizio; Bottai, Daniele; Vescovi, Angleo; Zhang, Shuguang

    2006-01-01

    Biomedical researchers have become increasingly aware of the limitations of conventional 2-dimensional tissue cell culture systems, including coated Petri dishes, multi-well plates and slides, to fully address many critical issues in cell biology, cancer biology and neurobiology, such as the 3-D microenvironment, 3-D gradient diffusion, 3-D cell migration and 3-D cell-cell contact interactions. In order to fully understand how cells behave in the 3-D body, it is important to develop a well-controlled 3-D cell culture system where every single ingredient is known. Here we report the development of a 3-D cell culture system using a designer peptide nanofiber scaffold with mouse adult neural stem cells. We attached several functional motifs, including cell adhesion, differentiation and bone marrow homing motifs, to a self-assembling peptide RADA16 (Ac-RADARADARADARADA-COHN2). These functionalized peptides undergo self-assembly into a nanofiber structure similar to Matrigel. During cell culture, the cells were fully embedded in the 3-D environment of the scaffold. Two of the peptide scaffolds containing bone marrow homing motifs significantly enhanced the neural cell survival without extra soluble growth and neurotrophic factors to the routine cell culture media. In these designer scaffolds, the cell populations with β-Tubulin+, GFAP+ and Nestin+ markers are similar to those found in cell populations cultured on Matrigel. The gene expression profiling array experiments showed selective gene expression, possibly involved in neural stem cell adhesion and differentiation. Because the synthetic peptides are intrinsically pure and a number of desired function cellular motifs are easy to incorporate, these designer peptide nanofiber scaffolds provide a promising controlled 3-D culture system for diverse tissue cells, and are useful as well for general molecular and cell biology. PMID:17205123

  2. Isolation and culture of neural crest stem cells from human hair follicles.

    PubMed

    Yang, Ruifeng; Xu, Xiaowei

    2013-04-06

    Hair follicles undergo lifelong growth and hair cycle is a well-controlled process involving stem cell proliferation and quiescence. Hair bulge is a well-characterized niche for adult stem cells. This segment of the outer root sheath contains a number of different types of stem cells, including epithelial stem cells, melanocyte stem cells and neural crest like stem cells. Hair follicles represent an accessible and rich source for different types of human stem cells. We and others have isolated neural crest stem cells (NCSCs) from human fetal and adult hair follicles. These human stem cells are label-retaining cells and are capable of self-renewal through asymmetric cell division in vitro. They express immature neural crest cell markers but not differentiation markers. Our expression profiling study showed that they share a similar gene expression pattern with murine skin immature neural crest cells. They exhibit clonal multipotency that can give rise to myogenic, melanocytic, and neuronal cell lineages after in vitro clonal single cell culture. Differentiated cells not only acquire lineage-specific markers but also demonstrate appropriate functions in ex vivo conditions. In addition, these NCSCs show differentiation potential toward mesenchymal lineages. Differentiated neuronal cells can persist in mouse brain and retain neuronal differentiation markers. It has been shown that hair follicle derived NCSCs can help nerve regrowth, and they improve motor function in mice transplanted with these stem cells following transecting spinal cord injury. Furthermore, peripheral nerves have been repaired with stem cell grafts, and implantation of skin-derived precursor cells adjacent to crushed sciatic nerves has resulted in remyelination. Therefore, the hair follicle/skin derived NCSCs have already shown promising results for regenerative therapy in preclinical models. Somatic cell reprogramming to induced pluripotent stem (iPS) cells has shown enormous potential for

  3. Enumeration of Neural Stem Cells Using Clonal Assays.

    PubMed

    Narayanan, Gunaseelan; Yu, Yuan Hong; Tham, Muly; Gan, Hui Theng; Ramasamy, Srinivas; Sankaran, Shvetha; Hariharan, Srivats; Ahmed, Sohail

    2016-10-04

    Neural stem cells (NSCs) have the ability to self-renew and generate the three major neural lineages - astrocytes, neurons and oligodendrocytes. NSCs and neural progenitors (NPs) are commonly cultured in vitro as neurospheres. This protocol describes in detail how to determine the NSC frequency in a given cell population under clonal conditions. The protocol begins with the seeding of the cells at a density that allows for the generation of clonal neurospheres. The neurospheres are then transferred to chambered coverslips and differentiated under clonal conditions in conditioned medium, which maximizes the differentiation potential of the neurospheres. Finally, the NSC frequency is calculated based on neurosphere formation and multipotency capabilities. Utilities of this protocol include the evaluation of candidate NSC markers, purification of NSCs, and the ability to distinguish NSCs from NPs. This method takes 13 days to perform, which is much shorter than current methods to enumerate NSC frequency.

  4. Enumeration of Neural Stem Cells Using Clonal Assays

    PubMed Central

    Narayanan, Gunaseelan; Yu, Yuan Hong; Tham, Muly; Gan, Hui Theng; Ramasamy, Srinivas; Sankaran, Shvetha; Hariharan, Srivats; Ahmed, Sohail

    2016-01-01

    Neural stem cells (NSCs) have the ability to self-renew and generate the three major neural lineages — astrocytes, neurons and oligodendrocytes. NSCs and neural progenitors (NPs) are commonly cultured in vitro as neurospheres. This protocol describes in detail how to determine the NSC frequency in a given cell population under clonal conditions. The protocol begins with the seeding of the cells at a density that allows for the generation of clonal neurospheres. The neurospheres are then transferred to chambered coverslips and differentiated under clonal conditions in conditioned medium, which maximizes the differentiation potential of the neurospheres. Finally, the NSC frequency is calculated based on neurosphere formation and multipotency capabilities. Utilities of this protocol include the evaluation of candidate NSC markers, purification of NSCs, and the ability to distinguish NSCs from NPs. This method takes 13 days to perform, which is much shorter than current methods to enumerate NSC frequency. PMID:27768074

  5. Human Neural Cell-Based Biosensor

    DTIC Science & Technology

    2012-04-11

    Chilton, Jamie ArunA Biomedical, Inc. 425 River Road Athens, GA 30602 QTR-11102010.4 Director, Naval Research Lab Attn: Code 5596 4555 Overlook...Modification P00001 Submitted by: Dr. Steven L. Stice, Principle Investigator ArunA Biomedical, Inc. 425 River Road Athens, GA 30602 Phone: 706...Progress Report v1.doc ArunA Biomedical, Inc. Page 1 of 1 Summary As a more biologically relevant model of human physiology, human neural progenitor

  6. VEGF-mediated angiogenesis stimulates neural stem cell proliferation and differentiation in the premature brain

    SciTech Connect

    Sun, Jinqiao; Sha, Bin; Zhou, Wenhao; Yang, Yi

    2010-03-26

    This study investigated the effects of angiogenesis on the proliferation and differentiation of neural stem cells in the premature brain. We observed the changes in neurogenesis that followed the stimulation and inhibition of angiogenesis by altering vascular endothelial growth factor (VEGF) expression in a 3-day-old rat model. VEGF expression was overexpressed by adenovirus transfection and down-regulated by siRNA interference. Using immunofluorescence assays, Western blot analysis, and real-time PCR methods, we observed angiogenesis and the proliferation and differentiation of neural stem cells. Immunofluorescence assays showed that the number of vWF-positive areas peaked at day 7, and they were highest in the VEGF up-regulation group and lowest in the VEGF down-regulation group at every time point. The number of neural stem cells, neurons, astrocytes, and oligodendrocytes in the subventricular zone gradually increased over time in the VEGF up-regulation group. Among the three groups, the number of these cells was highest in the VEGF up-regulation group and lowest in the VEGF down-regulation group at the same time point. Western blot analysis and real-time PCR confirmed these results. These data suggest that angiogenesis may stimulate the proliferation of neural stem cells and differentiation into neurons, astrocytes, and oligodendrocytes in the premature brain.

  7. Pulsed DC Electric Field-Induced Differentiation of Cortical Neural Precursor Cells.

    PubMed

    Chang, Hui-Fang; Lee, Ying-Shan; Tang, Tang K; Cheng, Ji-Yen

    2016-01-01

    We report the differentiation of neural stem and progenitor cells solely induced by direct current (DC) pulses stimulation. Neural stem and progenitor cells in the adult mammalian brain are promising candidates for the development of therapeutic neuroregeneration strategies. The differentiation of neural stem and progenitor cells depends on various in vivo environmental factors, such as nerve growth factor and endogenous EF. In this study, we demonstrated that the morphologic and phenotypic changes of mouse neural stem and progenitor cells (mNPCs) could be induced solely by exposure to square-wave DC pulses (magnitude 300 mV/mm at frequency of 100-Hz). The DC pulse stimulation was conducted for 48 h, and the morphologic changes of mNPCs were monitored continuously. The length of primary processes and the amount of branching significantly increased after stimulation by DC pulses for 48 h. After DC pulse treatment, the mNPCs differentiated into neurons, astrocytes, and oligodendrocytes simultaneously in stem cell maintenance medium. Our results suggest that simple DC pulse treatment could control the fate of NPCs. With further studies, DC pulses may be applied to manipulate NPC differentiation and may be used for the development of therapeutic strategies that employ NPCs to treat nervous system disorders.

  8. Pulsed DC Electric Field–Induced Differentiation of Cortical Neural Precursor Cells

    PubMed Central

    Chang, Hui-Fang; Lee, Ying-Shan; Tang, Tang K.; Cheng, Ji-Yen

    2016-01-01

    We report the differentiation of neural stem and progenitor cells solely induced by direct current (DC) pulses stimulation. Neural stem and progenitor cells in the adult mammalian brain are promising candidates for the development of therapeutic neuroregeneration strategies. The differentiation of neural stem and progenitor cells depends on various in vivo environmental factors, such as nerve growth factor and endogenous EF. In this study, we demonstrated that the morphologic and phenotypic changes of mouse neural stem and progenitor cells (mNPCs) could be induced solely by exposure to square-wave DC pulses (magnitude 300 mV/mm at frequency of 100-Hz). The DC pulse stimulation was conducted for 48 h, and the morphologic changes of mNPCs were monitored continuously. The length of primary processes and the amount of branching significantly increased after stimulation by DC pulses for 48 h. After DC pulse treatment, the mNPCs differentiated into neurons, astrocytes, and oligodendrocytes simultaneously in stem cell maintenance medium. Our results suggest that simple DC pulse treatment could control the fate of NPCs. With further studies, DC pulses may be applied to manipulate NPC differentiation and may be used for the development of therapeutic strategies that employ NPCs to treat nervous system disorders. PMID:27352251

  9. Neuroendocrine Cells of the Prostate Derive from the Neural Crest*

    PubMed Central

    Szczyrba, Jaroslaw; Wagner, Mathias; Wandernoth, Petra M.; Aumüller, Gerhard; Wennemuth, Gunther

    2017-01-01

    The histogenesis of prostatic neuroendocrine cells is controversial: a stem cell hypothesis with a urogenital sinus-derived progeny of all prostatic epithelial cells is opposed by a dual origin hypothesis, favoring the derivation of neuroendocrine cells from the neural crest, with the secretory and basal cells being of urogenital sinus origin. A computer-assisted 3D reconstruction was used to analyze the distribution of chromogranin A immunoreactive cells in serial sections of human fetal prostate specimens (gestation weeks 18 and 25). Immunohistochemical double labeling studies with YFP and serotonin antisera combined with electron microscopy were carried out on double-transgenic Wnt1-Cre/ROSA26-YFP mice showing stable YFP expression in all neural crest-derived cell populations despite loss of Wnt1 expression. 3D reconstruction of the distribution pattern of neuroendocrine cells in the human fetal prostate indicates a migration of paraganglionic cells passing the stroma and reaching the prostate ducts. Double-transgenic mice showed 55% double labeling of periurethral neuroendocrine cells expressing both serotonin and YFP, whereas single serotonin labeling was observed in 36% and exclusive YFP labeling in 9%. The results favor the assumption of a major fraction of neural crest-derived neuroendocrine cells in both the human and murine prostates. PMID:28003366

  10. Reinnervation of hair cells by neural stem cell-derived neurons.

    PubMed

    Yuan, Yasheng; Wang, Yang; Chi, Fanglu

    2014-01-01

    Replacement of spiral ganglion neurons would be one prioritized step in an attempt to restore sensory neuronal hearing loss. However, the possibility that transplanted neurons could regenerate new synaptic connections to hair cells has not been explored. The objective of this study was to test whether neural stem cell (NSC)-derived neurons can form synaptic connections with hair cells in vitro. NSCs were mechanically separated from the hippocampus in SD rat embryos (E12-E14) and cultured in a serum-free medium containing basic fibroblast growth factor and epidermal growth factor. Rat NSCs were co-cultured with explants of cochlea sensory epithelia obtained from postnatal Day 3 rats under transway filter membrane. At Day 3, the NSCs began to show chemotactic differentiation and grew toward cochlea sensory epithelia. After 9-day co-culture, neurites of NSC-derived neurons predominantly elongated toward hair cells. Immunohistochemical analyses revealed the fibers overlapped with synapsin and hair cells, indicating the formation of new synaptic connections. After 14-day culture, triple staining revealed the fibers overlapped with PSD95 (postsynaptic density) which is juxtaposed with CtBP2 (presynaptic vesicle), indicating the formation of new ribbon synapse. NSC-derived neurons can make synaptic connections with hair cells and provide a model for studying synaptic plasticity and regeneration. Whether the newly forming synapse is functional merits further electrophysiological study.

  11. Alcohol alters DNA methylation patterns and inhibits neural stem cell differentiation.

    PubMed

    Zhou, Feng C; Balaraman, Yokesh; Teng, MingXiang; Liu, Yunlong; Singh, Rabindra P; Nephew, Kenneth P

    2011-04-01

    Potential epigenetic mechanisms underlying fetal alcohol syndrome (FAS) include alcohol-induced alterations of methyl metabolism, resulting in aberrant patterns of DNA methylation and gene expression during development. Having previously demonstrated an essential role for epigenetics in neural stem cell (NSC) development and that inhibiting DNA methylation prevents NSC differentiation, here we investigated the effect of alcohol exposure on genome-wide DNA methylation patterns and NSC differentiation. Neural stem cells in culture were treated with or without a 6-hour 88 mM ("binge-like") alcohol exposure and examined at 48 hours, for migration, growth, and genome-wide DNA methylation. The DNA methylation was examined using DNA-methylation immunoprecipitation followed by microarray analysis. Further validation was performed using Independent Sequenom analysis.   Neural stem cell differentiated in 24 to 48 hours with migration, neuronal expression, and morphological transformation. Alcohol exposure retarded the migration, neuronal formation, and growth processes of NSC, similar to treatment with the methylation inhibitor 5-aza-cytidine. When NSC departed from the quiescent state, a genome-wide diversification of DNA methylation was observed-that is, many moderately methylated genes altered methylation levels and became hyper- and hypomethylated. Alcohol prevented many genes from such diversification, including genes related to neural development, neuronal receptors, and olfaction, while retarding differentiation. Validation of specific genes by Sequenom analysis demonstrated that alcohol exposure prevented methylation of specific genes associated with neural development [cut-like 2 (cutl2), insulin-like growth factor 1 (Igf1), epidermal growth factor-containing fibulin-like extracellular matrix protein 1 (Efemp1), and SRY-box-containing gene 7 (Sox 7)]; eye development, lens intrinsic membrane protein 2 (Lim 2); the epigenetic mark Smarca2 (SWI/SNF related, matrix

  12. Leptin Enhances Cholangiocarcinoma Cell Growth

    PubMed Central

    Fava, Giammarco; Alpini, Gianfranco; Rychlicki, Chiara; Saccomanno, Stefania; DeMorrow, Sharon; Trozzi, Luciano; Candelaresi, Cinzia; Venter, Julie; Di Sario, Antonio; Marzioni, Marco; Bearzi, Italo; Glaser, Shannon; Alvaro, Domenico; Marucci, Luca; Francis, Heather; Svegliati-Baroni, Gianluca; Benedetti, Antonio

    2008-01-01

    Cholangiocarcinoma is a strongly aggressive malignancy with a very poor prognosis. Effective therapeutic strategies are lacking because molecular mechanisms regulating cholangiocarcinoma cell growth are unknown. Furthermore, experimental in vivo animal models useful to study the pathophysiologic mechanisms of malignant cholangiocytes are lacking. Leptin, the hormone regulating caloric homeostasis, which is increased in obese patients, stimulates the growth of several cancers, such as hepatocellular carcinoma. The aim of this study was to define if leptin stimulates cholangiocarcinoma growth. We determined the expression of leptin receptors in normal and malignant human cholangiocytes. Effects on intrahepatic cholangiocarcinoma (HuH-28) cell proliferation, migration, and apoptosis of the in vitro exposure to leptin, together with the intracellular pathways, were then studied. Moreover, cholangiocarcinoma was experimentally induced in obese fa/fa Zucker rats, a genetically established animal species with faulty leptin receptors, and in their littermates by chronic feeding with thioacetamide, a potent carcinogen. After 24 weeks, the effect of leptin on cholangiocarcinoma development and growth was assessed. Normal and malignant human cholangiocytes express leptin receptors. Leptin increased the proliferation and the metastatic potential of cholangiocarcinoma cells in vitro through a signal transducers and activators of transcription 3–dependent activation of extracellular signal-regulated kinase 1/2. Leptin increased the growth and migration, and was antiapoptotic for cholangiocarcinoma cells. Moreover, the loss of leptin function reduced the development and the growth of cholangiocarcinoma. The experimental carcinogenesis model induced by thioacetamide administration is a valid and reproducible method to study cholangiocarcinoma pathobiology. Modulation of the leptin-mediated signal could be considered a valid tool for the prevention and treatment of

  13. Methylene blue promotes quiescence of rat neural progenitor cells.

    PubMed

    Xie, Luokun; Choudhury, Gourav R; Wang, Jixian; Park, Yong; Liu, Ran; Yuan, Fang; Zhang, Chun-Li; Yorio, Thomas; Jin, Kunlin; Yang, Shao-Hua

    2014-01-01

    Neural stem cell-based treatment holds a new therapeutic opportunity for neurodegenerative disorders. Here, we investigated the effect of methylene blue on proliferation and differentiation of rat neural progenitor cells (NPCs) both in vitro and in vivo. We found that methylene blue inhibited proliferation and promoted quiescence of NPCs in vitro without affecting committed neuronal differentiation. Consistently, intracerebroventricular infusion of methylene blue significantly inhibited NPC proliferation at the subventricular zone (SVZ). Methylene blue inhibited mTOR signaling along with down-regulation of cyclins in NPCs in vitro and in vivo. In summary, our study indicates that methylene blue may delay NPC senescence through enhancing NPCs quiescence.

  14. Axonal alignment and enhanced neuronal differentiation of neural stem cells on graphene-nanoparticle hybrid structures.

    PubMed

    Solanki, Aniruddh; Chueng, Sy-Tsong Dean; Yin, Perry T; Kappera, Rajesh; Chhowalla, Manish; Lee, Ki-Bum

    2013-10-11

    Human neural stem cells (hNSCs) cultured on graphene-nanoparticle hybrid structures show a unique behavior wherein the axons from the differentiating hNSCs show enhanced growth and alignment. We show that the axonal alignment is primarily due to the presence of graphene and the underlying nanoparticle monolayer causes enhanced neuronal differentiation of the hNSCs, thus having great implications of these hybrid-nanostructures for neuro-regenerative medicine.

  15. The Hippo signalling pathway maintains quiescence in Drosophila neural stem cells

    PubMed Central

    Ding, Rouven; Weynans, Kevin; Bossing, Torsten; Barros, Claudia S.; Berger, Christian

    2016-01-01

    Stem cells control their mitotic activity to decide whether to proliferate or to stay in quiescence. Drosophila neural stem cells (NSCs) are quiescent at early larval stages, when they are reactivated in response to metabolic changes. Here we report that cell-contact inhibition of growth through the canonical Hippo signalling pathway maintains NSC quiescence. Loss of the core kinases hippo or warts leads to premature nuclear localization of the transcriptional co-activator Yorkie and initiation of growth and proliferation in NSCs. Yorkie is necessary and sufficient for NSC reactivation, growth and proliferation. The Hippo pathway activity is modulated via inter-cellular transmembrane proteins Crumbs and Echinoid that are both expressed in a nutrient-dependent way in niche glial cells and NSCs. Loss of crumbs or echinoid in the niche only is sufficient to reactivate NSCs. Finally, we provide evidence that the Hippo pathway activity discriminates quiescent from non-quiescent NSCs in the Drosophila nervous system. PMID:26821647

  16. The Hippo signalling pathway maintains quiescence in Drosophila neural stem cells.

    PubMed

    Ding, Rouven; Weynans, Kevin; Bossing, Torsten; Barros, Claudia S; Berger, Christian

    2016-01-29

    Stem cells control their mitotic activity to decide whether to proliferate or to stay in quiescence. Drosophila neural stem cells (NSCs) are quiescent at early larval stages, when they are reactivated in response to metabolic changes. Here we report that cell-contact inhibition of growth through the canonical Hippo signalling pathway maintains NSC quiescence. Loss of the core kinases hippo or warts leads to premature nuclear localization of the transcriptional co-activator Yorkie and initiation of growth and proliferation in NSCs. Yorkie is necessary and sufficient for NSC reactivation, growth and proliferation. The Hippo pathway activity is modulated via inter-cellular transmembrane proteins Crumbs and Echinoid that are both expressed in a nutrient-dependent way in niche glial cells and NSCs. Loss of crumbs or echinoid in the niche only is sufficient to reactivate NSCs. Finally, we provide evidence that the Hippo pathway activity discriminates quiescent from non-quiescent NSCs in the Drosophila nervous system.

  17. Nanoengineering neural stem cells on biomimetic substrates using magnetofection technology.

    PubMed

    Adams, Christopher F; Dickson, Andrew W; Kuiper, Jan-Herman; Chari, Divya M

    2016-10-20

    Tissue engineering studies are witnessing a major paradigm shift to cell culture on biomimetic materials that replicate native tissue features from which the cells are derived. Few studies have been performed in this regard for neural cells, particularly in nanomedicine. For example, platforms such as magnetic nanoparticles (MNPs) have proven efficient as multifunctional tools for cell tracking and genetic engineering of neural transplant populations. However, as far as we are aware, all current studies have been conducted using neural cells propagated on non-neuromimetic substrates that fail to represent the mechano-elastic properties of brain and spinal cord microenvironments. Accordingly, it can be predicted that such data is of less translational and physiological relevance than that derived from cells grown in neuromimetic environments. Therefore, we have performed the first test of magnetofection technology (enhancing MNP delivery using applied magnetic fields with significant potential for therapeutic application) and its utility in genetically engineering neural stem cells (NSCs; a population of high clinical relevance) propagated in biomimetic hydrogels. We demonstrate magnetic field application safely enhances MNP mediated transfection of NSCs grown as 3D spheroid structures in collagen which more closely replicates the intrinsic mechanical and structural properties of neural tissue than routinely used hard substrates. Further, as it is well known that MNP uptake is mediated by endocytosis we also investigated NSC membrane activity grown on both soft and hard substrates. Using high resolution scanning electron microscopy we were able to prove that NSCs display lower levels of membrane activity on soft substrates compared to hard, a finding which could have particular impact on MNP mediated engineering strategies of cells propagated in physiologically relevant systems.

  18. Combined MSC-Secreted Factors and Neural Stem Cell Transplantation Promote Functional Recovery of PD Rats.

    PubMed

    Yao, Yuan; Huang, Chen; Gu, Ping; Wen, Tieqiao

    2016-01-01

    Stem cell transplantation has enormous potential for the treatment of neurodegenerative disorders like Parkinson's disease (PD). Mesenchymal stem cells (MSCs) have attracted much attention because they can secrete a wide variety of cellular factors that promote cell growth. In this study, we prepared a conditioned medium (CM) using lyophilized MSC culture medium that contained the secretome of MSCs and applied this CM to the culture of neural stem cells (CM-NSCs) for the transplantation of PD model rats. Quantitative real-time PCR, Western blot, and immunocytochemistry were used to identify cell differentiation and expression of dopaminergic neuron-specific genes in vitro. Behavioral tests including rotational behavior and MWM training tests were also performed to assess the recovery. Our results indicated that combined treatment of CM and neural stem cell transplantation can significantly reduce apomorphine-induced rotational asymmetry and improve spatial learning ability. The CM-NSCs were able to differentiate into dopaminergic neurons in the ventral tegmental area (VTA) and medial forebrain bundle (MFB), and migrated around the lesion site. They showed a higher activity than untreated NSCs in cell survival, migration, and behavior improvement in the dopa-deficit rat model. These findings suggest that the neural stem cells treated with conditioned medium possess a great potential as a graft candidate for the treatment of Parkinson's disease.

  19. Expression of polysialylated neural cell adhesion molecules on adult stem cells after neuronal differentiation of inner ear spiral ganglion neurons

    SciTech Connect

    Park, Kyoung Ho; Yeo, Sang Won; Troy, Frederic A.

    2014-10-17

    Highlights: • PolySia expressed on neurons primarily during early stages of neuronal development. • PolySia–NCAM is expressed on neural stem cells from adult guinea pig spiral ganglion. • PolySia is a biomarker that modulates neuronal differentiation in inner ear stem cells. - Abstract: During brain development, polysialylated (polySia) neural cell adhesion molecules (polySia–NCAMs) modulate cell–cell adhesive interactions involved in synaptogenesis, neural plasticity, myelination, and neural stem cell (NSC) proliferation and differentiation. Our findings show that polySia–NCAM is expressed on NSC isolated from adult guinea pig spiral ganglion (GPSG), and in neurons and Schwann cells after differentiation of the NSC with epidermal, glia, fibroblast growth factors (GFs) and neurotrophins. These differentiated cells were immunoreactive with mAb’s to polySia, NCAM, β-III tubulin, nestin, S-100 and stained with BrdU. NSC could regenerate and be differentiated into neurons and Schwann cells. We conclude: (1) polySia is expressed on NSC isolated from adult GPSG and on neurons and Schwann cells differentiated from these NSC; (2) polySia is expressed on neurons primarily during the early stage of neuronal development and is expressed on Schwann cells at points of cell–cell contact; (3) polySia is a functional biomarker that modulates neuronal differentiation in inner ear stem cells. These new findings suggest that replacement of defective cells in the inner ear of hearing impaired patients using adult spiral ganglion neurons may offer potential hope to improve the quality of life for patients with auditory dysfunction and impaired hearing disorders.

  20. Human pluripotent stem cell-derived neural constructs for predicting neural toxicity

    PubMed Central

    Schwartz, Michael P.; Hou, Zhonggang; Propson, Nicholas E.; Zhang, Jue; Engstrom, Collin J.; Costa, Vitor Santos; Jiang, Peng; Nguyen, Bao Kim; Bolin, Jennifer M.; Daly, William; Wang, Yu; Stewart, Ron; Page, C. David; Murphy, William L.; Thomson, James A.

    2015-01-01

    Human pluripotent stem cell-based in vitro models that reflect human physiology have the potential to reduce the number of drug failures in clinical trials and offer a cost-effective approach for assessing chemical safety. Here, human embryonic stem (ES) cell-derived neural progenitor cells, endothelial cells, mesenchymal stem cells, and microglia/macrophage precursors were combined on chemically defined polyethylene glycol hydrogels and cultured in serum-free medium to model cellular interactions within the developing brain. The precursors self-assembled into 3D neural constructs with diverse neuronal and glial populations, interconnected vascular networks, and ramified microglia. Replicate constructs were reproducible by RNA sequencing (RNA-Seq) and expressed neurogenesis, vasculature development, and microglia genes. Linear support vector machines were used to construct a predictive model from RNA-Seq data for 240 neural constructs treated with 34 toxic and 26 nontoxic chemicals. The predictive model was evaluated using two standard hold-out testing methods: a nearly unbiased leave-one-out cross-validation for the 60 training compounds and an unbiased blinded trial using a single hold-out set of 10 additional chemicals. The linear support vector produced an estimate for future data of 0.91 in the cross-validation experiment and correctly classified 9 of 10 chemicals in the blinded trial. PMID:26392547

  1. Human pluripotent stem cell-derived neural constructs for predicting neural toxicity.

    PubMed

    Schwartz, Michael P; Hou, Zhonggang; Propson, Nicholas E; Zhang, Jue; Engstrom, Collin J; Santos Costa, Vitor; Jiang, Peng; Nguyen, Bao Kim; Bolin, Jennifer M; Daly, William; Wang, Yu; Stewart, Ron; Page, C David; Murphy, William L; Thomson, James A

    2015-10-06

    Human pluripotent stem cell-based in vitro models that reflect human physiology have the potential to reduce the number of drug failures in clinical trials and offer a cost-effective approach for assessing chemical safety. Here, human embryonic stem (ES) cell-derived neural progenitor cells, endothelial cells, mesenchymal stem cells, and microglia/macrophage precursors were combined on chemically defined polyethylene glycol hydrogels and cultured in serum-free medium to model cellular interactions within the developing brain. The precursors self-assembled into 3D neural constructs with diverse neuronal and glial populations, interconnected vascular networks, and ramified microglia. Replicate constructs were reproducible by RNA sequencing (RNA-Seq) and expressed neurogenesis, vasculature development, and microglia genes. Linear support vector machines were used to construct a predictive model from RNA-Seq data for 240 neural constructs treated with 34 toxic and 26 nontoxic chemicals. The predictive model was evaluated using two standard hold-out testing methods: a nearly unbiased leave-one-out cross-validation for the 60 training compounds and an unbiased blinded trial using a single hold-out set of 10 additional chemicals. The linear support vector produced an estimate for future data of 0.91 in the cross-validation experiment and correctly classified 9 of 10 chemicals in the blinded trial.

  2. Behavior of neural stem cells in the Alzheimer brain

    PubMed Central

    Waldau, B.; Shetty, A. K.

    2013-01-01

    Alzheimer’s disease (AD) is characterized by the deposition of β-amyloid peptides (Aβ) and a progressive loss of neurons leading to dementia. Because hippocampal neurogenesis is linked to functions such as learning, memory and mood, there has been great interest in examining the effects of AD on hippocampal neurogenesis. This article reviews the pertinent studies and tries to unite them in one possible disease model. Early in the disease, oligomeric Aβ may transiently promote the generation of immature neurons from neural stem cells (NSCs). However, reduced concentrations of multiple neurotrophic factors and higher levels of fibroblast growth factor-2 seem to induce a developmental arrest of newly generated neurons. Furthermore, fibrillary Aβ and down-regulation of oligodendrocyte-lineage transcription factor-2 (OLIG2) may cause the death of these nonfunctional neurons. Therefore, altering the brain microenvironment for fostering apt maturation of graft-derived neurons may be critical for improving the efficacy of NSC transplantation therapy for AD. PMID:18500448

  3. Secretome profiling of differentiated neural mes-c-myc A1 cell line endowed with stem cell properties.

    PubMed

    Severino, Valeria; Farina, Annarita; Colucci-D'Amato, Luca; Reccia, Mafalda Giovanna; Volpicelli, Floriana; Parente, Augusto; Chambery, Angela

    2013-11-01

    Neural stem cell proliferation and differentiation play a crucial role in the formation and wiring of neuronal connections forming neuronal circuits. During neural tissues development, a large diversity of neuronal phenotypes is produced from neural precursor cells. In recent years, the cellular and molecular mechanisms by which specific types of neurons are generated have been explored with the aim to elucidate the complex events leading to the generation of different phenotypes via distinctive developmental programs that control self-renewal, differentiation, and plasticity. The extracellular environment is thought to provide instructive influences that actively induce the production of specific neuronal phenotypes. In this work, the secretome profiling of differentiated neural mes-c-myc A1 (A1) cell line endowed with stem cell properties was analyzed by applying a shotgun LC-MS/MS approach. The results provide a list of secreted molecules with potential relevance for the functional and biological features characterizing the A1 neuronal phenotype. Proteins involved in biological processes closely related to nervous system development including neurites growth, differentiation of neurons and axonogenesis were identified. Among them, proteins belonging to extracellular matrix and cell-adhesion complexes as well as soluble factors with well established neurotrophic properties were detected. The presented work provides the basis to clarify the complex extracellular protein networks implicated in neuronal differentiation and in the acquisition of the neuronal phenotype. This article is part of a Special Issue entitled: An Updated Secretome.

  4. Absence of Rybp Compromises Neural Differentiation of Embryonic Stem Cells

    PubMed Central

    Kovacs, Gergo; Szabo, Viktoria; Pirity, Melinda K.

    2016-01-01

    Rybp (Ring1 and Yy1 Binding Protein) is a transcriptional regulator and member of the noncanonical polycomb repressive complex 1 with essential role in early embryonic development. We have previously described that alteration of Rybp dosage in mouse models induced striking neural tube defects (NTDs), exencephaly, and disorganized neurocortex. In this study we further investigated the role of Rybp in neural differentiation by utilising wild type (rybp +/+) and rybp null mutant (rybp −/−) embryonic stem cells (ESCs) and tried to uncover underlying molecular events that are responsible for the observed phenotypic changes. We found that rybp null mutant ESCs formed less matured neurons, astrocytes, and oligodendrocytes from existing progenitors than wild type cells. Furthermore, lack of rybp coincided with altered gene expression of key neural markers including Pax6 and Plagl1 pinpointing a possible transcriptional circuit among these genes. PMID:26788067

  5. Kuwanon V Inhibits Proliferation, Promotes Cell Survival and Increases Neurogenesis of Neural Stem Cells

    PubMed Central

    Kong, Sun-Young; Park, Min-Hye; Lee, Mina; Kim, Jae-Ouk; Lee, Ha-Rim; Han, Byung Woo; Svendsen, Clive N.; Sung, Sang Hyun; Kim, Hyun-Jung

    2015-01-01

    Neural stem cells (NSCs) have the ability to proliferate and differentiate into neurons and glia. Regulation of NSC fate by small molecules is important for the generation of a certain type of cell. The identification of small molecules that can induce new neurons from NSCs could facilitate regenerative medicine and drug development for neurodegenerative diseases. In this study, we screened natural compounds to identify molecules that are effective on NSC cell fate determination. We found that Kuwanon V (KWV), which was isolated from the mulberry tree (Morus bombycis) root, increased neurogenesis in rat NSCs. In addition, during NSC differentiation, KWV increased cell survival and inhibited cell proliferation as shown by 5-bromo-2-deoxyuridine pulse experiments, Ki67 immunostaining and neurosphere forming assays. Interestingly, KWV enhanced neuronal differentiation and decreased NSC proliferation even in the presence of mitogens such as epidermal growth factor and fibroblast growth factor 2. KWV treatment of NSCs reduced the phosphorylation of extracellular signal-regulated kinase 1/2, increased mRNA expression levels of the cyclin-dependent kinase inhibitor p21, down-regulated Notch/Hairy expression levels and up-regulated microRNA miR-9, miR-29a and miR-181a. Taken together, our data suggest that KWV modulates NSC fate to induce neurogenesis, and it may be considered as a new drug candidate that can regenerate or protect neurons in neurodegenerative diseases. PMID:25706719

  6. Induction of osteoblastic differentiation of neural crest-derived stem cells from hair follicles.

    PubMed

    Urano-Morisawa, Eri; Takami, Masamichi; Suzawa, Tetsuo; Matsumoto, Akifumi; Osumi, Noriko; Baba, Kazuyoshi; Kamijo, Ryutaro

    2017-01-01

    The neural crest (NC) arises near the neural tube during embryo development. NC cells migrate throughout the embryo and have potential to differentiate into multiple cell types, such as peripheral nerves, glial, cardiac smooth muscle, endocrine, and pigment cells, and craniofacial bone. In the present study, we induced osteoblast-like cells using whisker follicles obtained from the NC of mice. Hair follicle cells derived from the NC labeled with enhanced green fluorescent protein (EGFP) were collected from protein zero-Cre/floxed-EGFP double transgenic mice and cultured, then treated and cultured in stem cell growth medium. After growth for 14 days, results of flow cytometry analysis showed that 95% of the EGFP-positive (EGFP+) hair follicle cells derived from the NC had proliferated and 76.2% of those expressed mesenchymal stem cells markers, such as platelet-derived growth factor α and stem cell antigen-1, and also showed constitutive expression of Runx2 mRNA. Cells stimulated with bone morphogenetic protein-2 expressed osteocalcin, osterix, and alkaline phosphatase mRNA, resulting in production of mineralized matrices, which were detected by von Kossa and alizarin red staining. Moreover, EGFP+ hair follicle cells consistently expressed macrophage colony-stimulating factor and osteoprotegerin (OPG). Addition of 1α,25-dihydroxyvitamin D3 [1,25(OH)2D3] (10-8 M) to the cultures suppressed OPG expression and induced RANKL production in the cells. Furthermore, multinucleated osteoclasts appeared within 6 days after starting co-cultures of bone marrow cells with EGFP+ cells in the presence of 1,25(OH)2D3 and PGE2. These results suggest that NC-derived hair follicle cells possess a capacity for osteoblastic differentiation and may be useful for developing new bone regenerative medicine therapies.

  7. Single-cell growth analysis in a mixed cell culture

    NASA Astrophysics Data System (ADS)

    Ando, Jun; Bato, Mary Grace P.; Daria, Vincent Ricardo

    2008-06-01

    We perform single cell analysis of cell growth in a mixed cell culture. Two species of yeast cells: Saccharomyces cerevisiae and Candida albicans, are optically trapped using focused continuous-wave near infrared laser. Cell growth for both cells is inhibited only when the two species of cells are in contact with each other. This indicates cell-cell interaction mediated cell growth inhibition mechanism. Single cell level analysis of cell growth studied here contributes to the further understanding of yeast growth arrest in a mixed yeast culture.

  8. Direct reprogramming of somatic cells into neural stem cells or neurons for neurological disorders

    PubMed Central

    Hou, Shaoping; Lu, Paul

    2016-01-01

    Direct reprogramming of somatic cells into neurons or neural stem cells is one of the most important frontier fields in current neuroscience research. Without undergoing the pluripotency stage, induced neurons or induced neural stem cells are a safer and timelier manner resource in comparison to those derived from induced pluripotent stem cells. In this prospective, we review the recent advances in generation of induced neurons and induced neural stem cells in vitro and in vivo and their potential treatments of neurological disorders. PMID:26981072

  9. Glioma gene therapy using induced pluripotent stem cell derived neural stem cells.

    PubMed

    Lee, Esther Xingwei; Lam, Dang Hoang; Wu, Chunxiao; Yang, Jing; Tham, Chee Kian; Ng, Wai Hoe; Wang, Shu

    2011-10-03

    Using neural stem cells (NSCs) with tumor tropic migratory capacity to deliver therapeutic genes is an attractive strategy in eliminating metastatic or disseminated tumors. While different methods have been developed to isolate or generate NSCs, it has not been assessed whether induced pluripotent stem (iPS) cells, a type of pluripotent stem cells that hold great potential for regenerative medicine, can be used as a source for derivation of NSCs with tumor tropism. In this study, we used a conventional lentivirus transduction method to derive iPS cells from primary mouse embryonic fibroblasts and then generated NSCs from the iPS cells. To investigate whether the iPS cell derived NSCs can be used in the treatment of disseminated brain tumors, the cells were transduced with a baculoviral vector containing the herpes simplex virus thymidine kinase suicide gene and injected into the cerebral hemisphere contralateral to a tumor inoculation site in a mouse intracranial human glioma xenograft model. We observed that NSCs expressing the suicide gene were, in the presence of ganciclovir, effective in inhibiting the growth of the glioma xenografts and prolonging survival of tumor-bearing mice. Our findings provide evidence for the feasibility of using iPS cell derived NSCs as cellular vehicles for targeted anticancer gene therapy.

  10. [Differentiation of C17.2 neural stem cells into neural cells induced by serum-free conditioned medium of olfactory ensheathing cells and cell viability detection of differentiated cells].

    PubMed

    Wang, Lei; Duan, Da; Zhao, Zhenyu; Teng, Xiaohua; Liu, Bo; Ge, Lite; Lu, Ming

    2014-05-01

    To study the possibility of the C17.2 neural stem cells (NSCs) differentiating into neural cells induced by serum-free condition medium of olfactory ensheathing cells (OECs) and to detect the cell viability of the differentiated cells. OECs were isloated and cultured from the olfactory bulbs of 3-day-old postnatal mouse to prepare serum-free condition medium of OECs. After C17.2 NSCs were cultured with H-DMEM/F12 medium containing 15% FBS and the cell fusion reached 80%, the 3rd passage cells were induced by serum-free condition medium of OECs in the experimental group, by H-DMEM/F12 in the control group, and non-induced C17.2 NSCs served as the blank control group. The growth condition of cells was observed with inverted microscope. After 5 days, the immunofluorescence staining [microtubule-associated protein 2 (MAP-2) and beta-tubulin-III] and Western blot (Nestin, beta-tubulin-III, and MAP-2) were carried out to identify the neural cells derived from NSCs. The cell viabilities were measured by MTT assay and the quantity of lactate dehydrogenase (LDH) release in the medium. In the experimental group, the C17.2 NSCs bodies began to contract at 24 hours after induction, and the differentiated cells increased obviously with long synapse at 3 days after induction; in the control group, the cell morphology showed no obvious change at 24 hours, cell body shrinkage, condensation of nuclear chromatin, and lysis were observed at 3 days. The immunofluorescence staining showed that beta3-tubulin-III and MAP-2 of C17.2 NSCs were positive at 5 days after induction, and Western blot suggested that the expression of Nestin protein declined significantly and the expressions of beta-tubulin-III and MAP-2 protein were increased in the experimental group, showing significant differences when compared with those in the control group and blank control group (P < 0.05). The LDH release and the cell viability were 130.60% +/- 6.86% and 62.20% +/- 3.82% in the experimental group, and

  11. Isolation and culture of neural crest cells from embryonic murine neural tube.

    PubMed

    Pfaltzgraff, Elise R; Mundell, Nathan A; Labosky, Patricia A

    2012-06-02

    The embryonic neural crest (NC) is a multipotent progenitor population that originates at the dorsal aspect of the neural tube, undergoes an epithelial to mesenchymal transition (EMT) and migrates throughout the embryo, giving rise to diverse cell types. NC also has the unique ability to influence the differentiation and maturation of target organs. When explanted in vitro, NC progenitors undergo self-renewal, migrate and differentiate into a variety of tissue types including neurons, glia, smooth muscle cells, cartilage and bone. NC multipotency was first described from explants of the avian neural tube. In vitro isolation of NC cells facilitates the study of NC dynamics including proliferation, migration, and multipotency. Further work in the avian and rat systems demonstrated that explanted NC cells retain their NC potential when transplanted back into the embryo. Because these inherent cellular properties are preserved in explanted NC progenitors, the neural tube explant assay provides an attractive option for studying the NC in vitro. To attain a better understanding of the mammalian NC, many methods have been employed to isolate NC populations. NC-derived progenitors can be cultured from post-migratory locations in both the embryo and adult to study the dynamics of post-migratory NC progenitors, however isolation of NC progenitors as they emigrate from the neural tube provides optimal preservation of NC cell potential and migratory properties. Some protocols employ fluorescence activated cell sorting (FACS) to isolate a NC population enriched for particular progenitors. However, when starting with early stage embryos, cell numbers adequate for analyses are difficult to obtain with FACS, complicating the isolation of early NC populations from individual embryos. Here, we describe an approach that does not rely on FACS and results in an approximately 96% pure NC population based on a Wnt1-Cre activated lineage reporter. The method presented here is adapted from

  12. Fibroblast growth factor 13 is essential for neural differentiation in Xenopus early embryonic development.

    PubMed

    Nishimoto, Satoko; Nishida, Eisuke

    2007-08-17

    In Xenopus embryonic development, the MEK5-ERK5 pathway, one of the MAPK pathways, lies downstream of SoxD and upstream of Xngnr1 in a signaling pathway regulating neural differentiation. It remains unclear, however, how the MEK5-ERK5 pathway is regulated in Xenopus neural development. As SoxD is a transcription factor, we hypothesized that some growth factor should be induced by SoxD and activate the MEK5-ERK5 pathway. As the expression level of fibroblast growth factor 13 (FGF13) is increased by SoxD, we analyzed the function of FGF13 in neural development. Knockdown of FGF13 with antisense morpholino-oligonucleotides (MOs) results in the reduced head structure and inhibition of neural differentiation. FGF13 MOs inhibit the SoxD-induced expression of Xngnr1 and the Xngnr1-induced expression of NeuroD, suggesting that FGF13 is necessary both upstream and downstream of Xngnr1 in neural differentiation. In addition, FGF13 MOs inhibit the activation of the MEK5-ERK5 pathway by dominant-negative bone morphogenetic protein receptor, a mimicker of neural inducers, indicating that FGF13 is involved in the activation of the MEK5-ERK5 pathway. Together, these results identify a role of FGF13 in Xenopus neural differentiation.

  13. Human Neural Cell-Based Biosensor

    DTIC Science & Technology

    2011-06-11

    astrocytes using defined medium conditions, (3) cell-based methods to detect botulinum toxin, and (4) HTS amenable assays for proliferation...progenitors into dopaminergic neurons, motoneurons and astrocytes using defined medium conditions, (3) cell-based methods to detect botulinum toxin...cell line developed for potential commercial distribution. (3) Development of cell based methods to detect botulinum toxin There has been

  14. Defining a developmental path to neural fate by global expression profiling of mouse embryonic stem cells and adult neural stem/progenitor cells.

    PubMed

    Aiba, Kazuhiro; Sharov, Alexei A; Carter, Mark G; Foroni, Chiara; Vescovi, Angelo L; Ko, Minoru S H

    2006-04-01

    To understand global features of gene expression changes during in vitro neural differentiation, we carried out the microarray analysis of embryonic stem cells (ESCs), embryonal carcinoma cells, and adult neural stem/progenitor (NS) cells. Expression profiling of ESCs during differentiation in monolayer culture revealed three distinct phases: undifferentiated ESCs, primitive ectoderm-like cells, and neural progenitor cells. Principal component (PC) analysis revealed that these cells were aligned on PC1 over the course of 6 days. This PC1 represents approximately 4,000 genes, the expression of which increased with neural commitment/differentiation. Furthermore, NS cells derived from adult brain and their differentiated cells were positioned along this PC axis further away from undifferentiated ESCs than embryonic stem-derived neural progenitors. We suggest that this PC1 defines a path to neural fate, providing a scale for the degree of commitment/differentiation.

  15. Aneuploidy causes premature differentiation of neural and intestinal stem cells

    PubMed Central

    Gogendeau, Delphine; Siudeja, Katarzyna; Gambarotto, Davide; Pennetier, Carole; Bardin, Allison J.; Basto, Renata

    2015-01-01

    Aneuploidy is associated with a variety of diseases such as cancer and microcephaly. Although many studies have addressed the consequences of a non-euploid genome in cells, little is known about their overall consequences in tissue and organism development. Here we use two different mutant conditions to address the consequences of aneuploidy during tissue development and homeostasis in Drosophila. We show that aneuploidy causes brain size reduction due to a decrease in the number of proliferative neural stem cells (NSCs), but not through apoptosis. Instead, aneuploid NSCs present an extended G1 phase, which leads to cell cycle exit and premature differentiation. Moreover, we show that this response to aneuploidy is also present in adult intestinal stem cells but not in the wing disc. Our work highlights a neural and intestine stem cell-specific response to aneuploidy, which prevents their proliferation and expansion. PMID:26573328

  16. Could the endogenous opioid, morphine, prevent neural stem cell proliferation?

    PubMed

    Shoae-Hassani, Alireza; Sharif, Shiva; Tabatabaei, Seyed Abdolreza Mortazavi; Verdi, Javad

    2011-02-01

    In spite of widespread use of morphine to treat pain in patients, little is known about the effects of this opioid on many cells including stem cells. Moreover the studies have been shown controversial results about morphine effects on several kinds of cells. It is well-known that morphine exposure could decrease testosterone levels in brain and spinal cord. Morphine could increase the activity of 5α-redutase, the enzyme that converts testosterone into its respective 5α-redutase derivative dihydrotestosterone (DHT). Also it could increase aromatase activity that converts testosterone to estradiol. Proliferation of neural stem cells was observed in human stem cells after exposure to certain combinations of steroids especially testosterone. On the other hand DHT has negative effect in neural stem cell reproduction. Morphine induces over-expression of p53 gene that could mediate stem cell apoptosis. Therefore we hypothesized that due to reduction in the testosterone levels, elevation in the DHT levels, and over-expression of p53 gene, morphine could prevent neural stem cell proliferation. Copyright © 2010 Elsevier Ltd. All rights reserved.

  17. Receptor regulation of osmolyte homeostasis in neural cells

    PubMed Central

    Fisher, Stephen K; Heacock, Anne M; Keep, Richard F; Foster, Daniel J

    2010-01-01

    The capacity of cells to correct their volume in response to hyposmotic stress via the efflux of inorganic and organic osmolytes is well documented. However, the ability of cell-surface receptors, in particular G-protein-coupled receptors (GPCRs), to regulate this homeostatic mechanism has received much less attention. Mechanisms that underlie the regulation of cell volume are of particular importance to cells in the central nervous system because of the physical restrictions of the skull and the adverse impact that even small increases in cell volume can have on their function. Increases in brain volume are seen in hyponatraemia, which can arise from a variety of aetiologies and is the most frequently diagnosed electrolyte disorder in clinical practice. In this review we summarize recent evidence that the activation of GPCRs facilitates the volume-dependent efflux of osmolytes from neural cells and permits them to more efficiently respond to small, physiologically relevant, reductions in osmolarity. The characteristics of receptor-regulated osmolyte efflux, the signalling pathways involved and the physiological significance of receptor activation are discussed. In addition, we propose that GPCRs may also regulate the re-uptake of osmolytes into neural cells, but that the influx of organic and inorganic osmolytes is differentially regulated. The ability of neural cells to closely regulate osmolyte homeostasis through receptor-mediated alterations in both efflux and influx mechanisms may explain, in part at least, why the brain selectively retains its complement of inorganic osmolytes during chronic hyponatraemia, whereas its organic osmolytes are depleted. PMID:20498228

  18. Induced Pluripotent Stem Cells for Neural Tissue Engineering

    PubMed Central

    Wang, Aijun; Tang, Zhenyu; Park, In-Hyun; Zhu, Yiqian; Patel, Shyam; Daley, George Q.; Song, Li

    2011-01-01

    Induced pluripotent stem cells (iPSCs) hold great promise for cell therapies and tissue engineering. Neural crest stem cells (NCSCs) are multipotent and represent a valuable system to investigate iPSC differentiation and therapeutic potential. Here we derived NCSCs from human iPSCs and embryonic stem cells (ESCs), and investigated the potential of NCSCs for neural tissue engineering. The differentiation of iPSCs and the expansion of derived NCSCs varied in different cell lines, but all NCSC lines were capable of differentiating into mesodermal and ectodermal lineages, including neural cells. Tissue-engineered nerve conduits were fabricated by seeding NCSCs into nanofibrous tubular scaffolds, and used as a bridge for transected sciatic nerves in a rat model. Electrophysiological analysis showed that only NCSC-engrafted nerve conduits resulted in an accelerated regeneration of sciatic nerves at 1 month. Histological analysis demonstrated that NCSC transplantation promoted axonal myelination. Furthermore, NCSCs differentiated into Schwann cells and were integrated into the myelin sheath around axons. No teratoma formation was observed for up to 1 year after NCSC transplantation in vivo. This study demonstrates that iPSC-derived multipotent NCSCs can be directly used for tissue engineering and that the approach that combines stem cells and scaffolds has tremendous potential for regenerative medicine applications. PMID:21514663

  19. The transcription factor Nerfin-1 prevents reversion of neurons into neural stem cells

    PubMed Central

    Froldi, Francesca; Szuperak, Milan; Weng, Chen-Fang; Shi, Wei; Papenfuss, Anthony T.

    2015-01-01

    Cellular dedifferentiation is the regression of a cell from a specialized state to a more multipotent state and is implicated in cancer. However, the transcriptional network that prevents differentiated cells from reacquiring stem cell fate is so far unclear. Neuroblasts (NBs), the Drosophila neural stem cells, are a model for the regulation of stem cell self-renewal and differentiation. Here we show that the Drosophila zinc finger transcription factor Nervous fingers 1 (Nerfin-1) locks neurons into differentiation, preventing their reversion into NBs. Following Prospero-dependent neuronal specification in the ganglion mother cell (GMC), a Nerfin-1-specific transcriptional program maintains differentiation in the post-mitotic neurons. The loss of Nerfin-1 causes reversion to multipotency and results in tumors in several neural lineages. Both the onset and rate of neuronal dedifferentiation in nerfin-1 mutant lineages are dependent on Myc- and target of rapamycin (Tor)-mediated cellular growth. In addition, Nerfin-1 is required for NB differentiation at the end of neurogenesis. RNA sequencing (RNA-seq) and chromatin immunoprecipitation (ChIP) analysis show that Nerfin-1 administers its function by repression of self-renewing-specific and activation of differentiation-specific genes. Our findings support the model of bidirectional interconvertibility between neural stem cells and their post-mitotic progeny and highlight the importance of the Nerfin-1-regulated transcriptional program in neuronal maintenance. PMID:25593306

  20. p73 regulates maintenance of neural stem cell

    SciTech Connect

    Agostini, Massimiliano; Tucci, Paola; Bano, Daniele; Nicotera, Pierluigi; McKeon, Frank; Melino, Gerry

    2010-12-03

    Research highlights: {yields} TAp73 is expressed in neural stem cells and its expression increases following their differentiation. {yields} Neural stem cells from p73 null mice have a reduced proliferative potential. {yields} p73-deficient neural stem cells show reduced expression of members of the Sox-2 and Notch gene families. {yields} Neurogenic areas are reduced in the brains of embryonic and adult p73-/- mice. -- Abstract: p73, a member of the p53 family, is a transcription factor that plays a key role in many biological processes. In the present study, we show that TAp73 is expressed in neural stem cells (NSC) and its expression increases following their differentiation. NSC from p73 null mice have a reduced proliferative potential, together with reduced expression of members of the Sox-2 and Notch gene families known to be important for NSC proliferation. In parallel with this in vitro data, the width of the neurogenic areas was reduced in the brains of embryonic and adult p73-/- mice. These data suggest that p73, and in particular TAp73, is important for maintenance of the NSC pool.

  1. Autocrine regulation of neural crest cell development by steel factor.

    PubMed

    Guo, C S; Wehrle-Haller, B; Rossi, J; Ciment, G

    1997-04-01

    Steel factor (SLF) and its cognate receptor, c-kit, have been implicated in the generation of melanocytes from migrating neural crest (NC) cells during early vertebrate embryogenesis. However, the source of SLF in the early avian embryo and its precise role in melanogenesis are unclear. We report here that NC cells themselves express and release SLF protein, which in turn acts as an autocrine factor to induce melanogenesis in nearby NC cells. These results indicate that NC cell subpopulations play an active role in the determination of their cell fate and suggest a different developmental role for the embryonic microenvironment than what has been previously proposed.

  2. Human Neural Cell-Based Biosensor

    DTIC Science & Technology

    2011-03-11

    neurons, motoneurons and astrocytes using defined medium conditions, (3) cell-based methods to detect botulinum toxin, and (4) fluorescence based assays...medium conditions, (3) cell-based methods to detect botulinum toxin, and (4) fluorescence based assays for proliferation, cell migration, mitochondrial...line will begin shortly. (3) Development of cell based methods to detect botulinum toxin There has been substantial progress in the development

  3. Effect of Monocular Deprivation on Rabbit Neural Retinal Cell Densities

    PubMed Central

    Mwachaka, Philip Maseghe; Saidi, Hassan; Odula, Paul Ochieng; Mandela, Pamela Idenya

    2015-01-01

    Purpose: To describe the effect of monocular deprivation on densities of neural retinal cells in rabbits. Methods: Thirty rabbits, comprised of 18 subject and 12 control animals, were included and monocular deprivation was achieved through unilateral lid suturing in all subject animals. The rabbits were observed for three weeks. At the end of each week, 6 experimental and 3 control animals were euthanized, their retinas was harvested and processed for light microscopy. Photomicrographs of the retina were taken and imported into FIJI software for analysis. Results: Neural retinal cell densities of deprived eyes were reduced along with increasing period of deprivation. The percentage of reductions were 60.9% (P < 0.001), 41.6% (P = 0.003), and 18.9% (P = 0.326) for ganglion, inner nuclear, and outer nuclear cells, respectively. In non-deprived eyes, cell densities in contrast were increased by 116% (P < 0.001), 52% (P < 0.001) and 59.6% (P < 0.001) in ganglion, inner nuclear, and outer nuclear cells, respectively. Conclusion: In this rabbit model, monocular deprivation resulted in activity-dependent changes in cell densities of the neural retina in favour of the non-deprived eye along with reduced cell densities in the deprived eye. PMID:26425316

  4. Effect of Monocular Deprivation on Rabbit Neural Retinal Cell Densities.

    PubMed

    Mwachaka, Philip Maseghe; Saidi, Hassan; Odula, Paul Ochieng; Mandela, Pamela Idenya

    2015-01-01

    To describe the effect of monocular deprivation on densities of neural retinal cells in rabbits. Thirty rabbits, comprised of 18 subject and 12 control animals, were included and monocular deprivation was achieved through unilateral lid suturing in all subject animals. The rabbits were observed for three weeks. At the end of each week, 6 experimental and 3 control animals were euthanized, their retinas was harvested and processed for light microscopy. Photomicrographs of the retina were taken and imported into FIJI software for analysis. Neural retinal cell densities of deprived eyes were reduced along with increasing period of deprivation. The percentage of reductions were 60.9% (P < 0.001), 41.6% (P = 0.003), and 18.9% (P = 0.326) for ganglion, inner nuclear, and outer nuclear cells, respectively. In non-deprived eyes, cell densities in contrast were increased by 116% (P < 0.001), 52% (P < 0.001) and 59.6% (P < 0.001) in ganglion, inner nuclear, and outer nuclear cells, respectively. In this rabbit model, monocular deprivation resulted in activity-dependent changes in cell densities of the neural retina in favour of the non-deprived eye along with reduced cell densities in the deprived eye.

  5. NFIX regulates neural progenitor cell differentiation during hippocampal morphogenesis.

    PubMed

    Heng, Yee Hsieh Evelyn; McLeay, Robert C; Harvey, Tracey J; Smith, Aaron G; Barry, Guy; Cato, Kathleen; Plachez, Céline; Little, Erica; Mason, Sharon; Dixon, Chantelle; Gronostajski, Richard M; Bailey, Timothy L; Richards, Linda J; Piper, Michael

    2014-01-01

    Neural progenitor cells have the ability to give rise to neurons and glia in the embryonic, postnatal and adult brain. During development, the program regulating whether these cells divide and self-renew or exit the cell cycle and differentiate is tightly controlled, and imbalances to the normal trajectory of this process can lead to severe functional consequences. However, our understanding of the molecular regulation of these fundamental events remains limited. Moreover, processes underpinning development of the postnatal neurogenic niches within the cortex remain poorly defined. Here, we demonstrate that Nuclear factor one X (NFIX) is expressed by neural progenitor cells within the embryonic hippocampus, and that progenitor cell differentiation is delayed within Nfix(-/-) mice. Moreover, we reveal that the morphology of the dentate gyrus in postnatal Nfix(-/-) mice is abnormal, with fewer subgranular zone neural progenitor cells being generated in the absence of this transcription factor. Mechanistically, we demonstrate that the progenitor cell maintenance factor Sry-related HMG box 9 (SOX9) is upregulated in the hippocampus of Nfix(-/-) mice and demonstrate that NFIX can repress Sox9 promoter-driven transcription. Collectively, our findings demonstrate that NFIX plays a central role in hippocampal morphogenesis, regulating the formation of neuronal and glial populations within this structure.

  6. Methods for derivation of multipotent neural crest cells derived from human pluripotent stem cells

    PubMed Central

    Avery, John; Dalton, Stephen

    2016-01-01

    Summary Multipotent, neural crest cells (NCCs) produce a wide-range of cell types during embryonic development. This includes melanocytes, peripheral neurons, smooth muscle cells, osteocytes, chondrocytes and adipocytes. The protocol described here allows for highly-efficient differentiation of human pluripotent stem cells to a neural crest fate within 15 days. This is accomplished under feeder-free conditions, using chemically defined medium supplemented with two small molecule inhibitors that block glycogen synthase kinase 3 (GSK3) and bone morphogenic protein (BMP) signaling. This technology is well-suited as a platform to understand in greater detail the pathogenesis of human disease associated with impaired neural crest development/migration. PMID:25986498

  7. Neural cell image segmentation method based on support vector machine

    NASA Astrophysics Data System (ADS)

    Niu, Shiwei; Ren, Kan

    2015-10-01

    In the analysis of neural cell images gained by optical microscope, accurate and rapid segmentation is the foundation of nerve cell detection system. In this paper, a modified image segmentation method based on Support Vector Machine (SVM) is proposed to reduce the adverse impact caused by low contrast ratio between objects and background, adherent and clustered cells' interference etc. Firstly, Morphological Filtering and OTSU Method are applied to preprocess images for extracting the neural cells roughly. Secondly, the Stellate Vector, Circularity and Histogram of Oriented Gradient (HOG) features are computed to train SVM model. Finally, the incremental learning SVM classifier is used to classify the preprocessed images, and the initial recognition areas identified by the SVM classifier are added to the library as the positive samples for training SVM model. Experiment results show that the proposed algorithm can achieve much better segmented results than the classic segmentation algorithms.

  8. Neural stem cells: from neurobiology to clinical applications.

    PubMed

    Andressen, Christian

    2013-01-01

    In spite of increasing numbers of publications about cell replacement therapies in various neurodegenerative diseases, reports on therapeutic benefits are still rare due to the huge array of parameters affecting the clinically relevant outcome. Limiting conditions can be attributed to origin and number of cells used for transplantation, their in vitro storage, propagation and/or predifferentiation. In addition, the ability of these cells for a site directed differentiation and functional integration in sufficient numbers is known to depend on extrinsic factors including intracerebral position of graft(s). Thus, obstacles to the use of cells in replacement therapies of neurological disorders reflect the molecular as well as cellular complexity of affected functional systems. This review will highlight central aspects of cell replacement strategies that are currently regarded as the most limiting issues in respect to survival, cell identity and site directed differentiation as well as functional integration of grafts. Special attention will be paid to neural stem cells, derived from either fetal or adult central nervous tissue. Unravelling the molecular biology of these proliferating cells in combination with instructive environmental cues for their site directed differentiation will pave the way to high reproducibility in collection, propagation, and predifferentiation of transplantable cells in vitro. In addition, this knowledge of intrinsic and extrinsic cues for a site directed neural differentiation during development will broaden the perspective for any pluripotent stem cell, namely embryonic stem and induced pluripotent stem cells, as an alternate source for a cell based therapy of neurodegenerative diseases.

  9. Molecular Analysis of Stromal Cells-Induced Neural Differentiation of Mouse Embryonic Stem Cells.

    PubMed

    Joshi, Ramila; Buchanan, James Carlton; Paruchuri, Sailaja; Morris, Nathan; Tavana, Hossein

    2016-01-01

    Deriving specific neural cells from embryonic stem cells (ESCs) is a promising approach for cell replacement therapies of neurodegenerative diseases. When co-cultured with certain stromal cells, mouse ESCs (mESCs) differentiate efficiently to neural cells. In this study, a comprehensive gene and protein expression analysis of differentiating mESCs is performed over a two-week culture period to track temporal progression of cells from a pluripotent state to specific terminally-differentiated neural cells such as neurons, astrocytes, and oligodendrocytes. Expression levels of 26 genes consisting of marker genes for pluripotency, neural progenitors, and specific neuronal, astroglial, and oligodendrocytic cells are tracked using real time q-PCR. The time-course gene expression analysis of differentiating mESCs is combined with the hierarchal clustering and functional principal component analysis (FPCA) to elucidate the evolution of specific neural cells from mESCs at a molecular level. These statistical analyses identify three major gene clusters representing distinct phases of transition of stem cells from a pluripotent state to a terminally-differentiated neuronal or glial state. Temporal protein expression studies using immunohistochemistry demonstrate the generation of neural stem/progenitor cells and specific neural lineages and show a close agreement with the gene expression profiles of selected markers. Importantly, parallel gene and protein expression analysis elucidates long-term stability of certain proteins compared to those with a quick turnover. Describing the molecular regulation of neural cells commitment of mESCs due to stromal signaling will help identify major promoters of differentiation into specific cell types for use in cell replacement therapy applications.

  10. Molecular Analysis of Stromal Cells-Induced Neural Differentiation of Mouse Embryonic Stem Cells

    PubMed Central

    Joshi, Ramila; Buchanan, James Carlton; Paruchuri, Sailaja; Morris, Nathan; Tavana, Hossein

    2016-01-01

    Deriving specific neural cells from embryonic stem cells (ESCs) is a promising approach for cell replacement therapies of neurodegenerative diseases. When co-cultured with certain stromal cells, mouse ESCs (mESCs) differentiate efficiently to neural cells. In this study, a comprehensive gene and protein expression analysis of differentiating mESCs is performed over a two-week culture period to track temporal progression of cells from a pluripotent state to specific terminally-differentiated neural cells such as neurons, astrocytes, and oligodendrocytes. Expression levels of 26 genes consisting of marker genes for pluripotency, neural progenitors, and specific neuronal, astroglial, and oligodendrocytic cells are tracked using real time q-PCR. The time-course gene expression analysis of differentiating mESCs is combined with the hierarchal clustering and functional principal component analysis (FPCA) to elucidate the evolution of specific neural cells from mESCs at a molecular level. These statistical analyses identify three major gene clusters representing distinct phases of transition of stem cells from a pluripotent state to a terminally-differentiated neuronal or glial state. Temporal protein expression studies using immunohistochemistry demonstrate the generation of neural stem/progenitor cells and specific neural lineages and show a close agreement with the gene expression profiles of selected markers. Importantly, parallel gene and protein expression analysis elucidates long-term stability of certain proteins compared to those with a quick turnover. Describing the molecular regulation of neural cells commitment of mESCs due to stromal signaling will help identify major promoters of differentiation into specific cell types for use in cell replacement therapy applications. PMID:27832161

  11. Alternative generation of CNS neural stem cells and PNS derivatives from neural crest-derived peripheral stem cells.

    PubMed

    Weber, Marlen; Apostolova, Galina; Widera, Darius; Mittelbronn, Michel; Dechant, Georg; Kaltschmidt, Barbara; Rohrer, Hermann

    2015-02-01

    Neural crest-derived stem cells (NCSCs) from the embryonic peripheral nervous system (PNS) can be reprogrammed in neurosphere (NS) culture to rNCSCs that produce central nervous system (CNS) progeny, including myelinating oligodendrocytes. Using global gene expression analysis we now demonstrate that rNCSCs completely lose their previous PNS characteristics and acquire the identity of neural stem cells derived from embryonic spinal cord. Reprogramming proceeds rapidly and results in a homogenous population of Olig2-, Sox3-, and Lex-positive CNS stem cells. Low-level expression of pluripotency inducing genes Oct4, Nanog, and Klf4 argues against a transient pluripotent state during reprogramming. The acquisition of CNS properties is prevented in the presence of BMP4 (BMP NCSCs) as shown by marker gene expression and the potential to produce PNS neurons and glia. In addition, genes characteristic for mesenchymal and perivascular progenitors are expressed, which suggests that BMP NCSCs are directed toward a pericyte progenitor/mesenchymal stem cell (MSC) fate. Adult NCSCs from mouse palate, an easily accessible source of adult NCSCs, display strikingly similar properties. They do not generate cells with CNS characteristics but lose the neural crest markers Sox10 and p75 and produce MSC-like cells. These findings show that embryonic NCSCs acquire a full CNS identity in NS culture. In contrast, MSC-like cells are generated from BMP NCSCs and pNCSCs, which reveals that postmigratory NCSCs are a source for MSC-like cells up to the adult stage.

  12. Neural crest cell signaling pathways critical to cranial bone development and pathology.

    PubMed

    Mishina, Yuji; Snider, Taylor Nicholas

    2014-07-15

    Neural crest cells appear early during embryogenesis and give rise to many structures in the mature adult. In particular, a specific population of neural crest cells migrates to and populates developing cranial tissues. The ensuing differentiation of these cells via individual complex and often intersecting signaling pathways is indispensible to growth and development of the craniofacial complex. Much research has been devoted to this area of development with particular emphasis on cell signaling events required for physiologic development. Understanding such mechanisms will allow researchers to investigate ways in which they can be exploited in order to treat a multitude of diseases affecting the craniofacial complex. Knowing how these multipotent cells are driven towards distinct fates could, in due course, allow patients to receive regenerative therapies for tissues lost to a variety of pathologies. In order to realize this goal, nucleotide sequencing advances allowing snapshots of entire genomes and exomes are being utilized to identify molecular entities associated with disease states. Once identified, these entities can be validated for biological significance with other methods. A crucial next step is the integration of knowledge gleaned from observations in disease states with normal physiology to generate an explanatory model for craniofacial development. This review seeks to provide a current view of the landscape on cell signaling and fate determination of the neural crest and to provide possible avenues of approach for future research.

  13. Isolation and purification of self-renewable human neural stem cells for cell therapy in experimental model of ischemic stroke.

    PubMed

    Azevedo-Pereira, Ricardo L; Daadi, Marcel M

    2013-01-01

    Human embryonic stem cells (hESCs) are pluripotent with a strong self-renewable ability making them a virtually unlimited source of neural cells for structural repair in neurological disorders. Currently, hESCs are one of the most promising cell sources amenable for commercialization of off-shelf cell therapy products. However, along with this strong proliferative capacity of hESCs comes the tumorigenic potential of these cells after transplantation. Thus, the isolation and purification of a homogeneous, population of neural stem cells (hNSCs) are of paramount importance to avoid tumor formation in the host brain. This chapter describes the isolation, neuralization, and long-term perpetuation of hNSCs derived from hESCs through use of specific mitogenic growth factors and the preparation of hNSCs for transplantation in an experimental model of stroke. Additionally, we describe methods to analyze the stroke and size of grafts using magnetic resonance imaging and Osirix software, and neuroanatomical tracing procedures to study axonal remodeling after stroke and cell transplantation.

  14. Aebp2 as an Epigenetic Regulator for Neural Crest Cells

    PubMed Central

    Kim, Hana; Kang, Keunsoo; Ekram, Muhammad B.; Roh, Tae-Young; Kim, Joomyeong

    2011-01-01

    Aebp2 is a potential targeting protein for the mammalian Polycomb Repression Complex 2 (PRC2). We generated a mutant mouse line disrupting the transcription of Aebp2 to investigate its in vivo roles. Aebp2-mutant homozygotes were embryonic lethal while heterozygotes survived to adulthood with fertility. In developing mouse embryos, Aebp2 is expressed mainly within cells of neural crest origin. In addition, many heterozygotes display a set of phenotypes, enlarged colon and hypopigmentation, similar to those observed in human patients with Hirschsprung's disease and Waardenburg syndrome. These phenotypes are usually caused by the absence of the neural crest-derived ganglia in hindguts and melanocytes. ChIP analyses demonstrated that the majority of the genes involved in the migration and development process of neural crest cells are downstream target genes of AEBP2 and PRC2. Furthermore, expression analyses confirmed that some of these genes are indeed affected in the Aebp2 heterozygotes. Taken together, these results suggest that Aebp2 may regulate the migration and development of the neural crest cells through the PRC2-mediated epigenetic mechanism. PMID:21949878

  15. Differentiation of human CD146-positive endometrial stem cells to adipogenic-, osteogenic-, neural progenitor-, and glial-like cells.

    PubMed

    Fayazi, Mehri; Salehnia, Mojdeh; Ziaei, Saeideh

    2015-04-01

    The aim of this study was to investigate the potential differentiation of CD146(+) endometrial stem cells to several lineages. Endometrial stromal cells were cultured using Dulbecco's modified Eagle's medium/Hams F-12 (DMEM/F-12) and were passaged every 7-10 d when cultures reached 80-100% of confluency. The immunophenotypes of single endometrial cells were analyzed using flow cytometry at fourth passage. Then the CD146(+) cells were sorted using magnetic-activated cell sorting, and they were cultured and analyzed for in vitro differentiation to several lineages. Detection of adipocyte- and osteocyte-like cells were assessed by oil red O and alizarin red staining, respectively. For detection of neural progenitor and oligodendrocyte-like cells, the cells were immunostained by neurofilament 68 and oligo2, respectively. The rates of CD90, CD105, CD146, CD31, CD34, and CD9 of cultured endometrial cells were 94.98 ± 3%, 95.77 ± 2.5%, 27.61 ± 2%, 0.79 ± 0.05%, 1.43 ± 0.1%, and 1.01 ± 0.06%, respectively. CD146(+) cells were isolated to high purity. CD146(+)-differentiated cells to adipogenic cell with typical lipid-rich vacuoles and osteogenic cells were observed and confirmed their mesenchymal origin. They also differentiated into neural progenitor and glial differentiation by retinoic acid, basic fibroblast growth factor, and epidermal growth factor signaling molecules, respectively, and confirmed by neurofilament 68 and oligo2 immunocytochemistry. The efficiency of differentiation to neural progenitor and oligodendrocyte-like cells was 90 ± 3.4% and 79 ± 2.8%, respectively. This study showed that CD146(+) cells from human endometrium after in vitro cultivation can differentiate into adipogenic-, osteogenic-, neural progenitor-, and glial-like cells. They may provide available alternative source of stem cells for future cell-based therapies and tissue engineering applications.

  16. Derivation of Neural Precursor Cells from Human Embryonic Stem Cells for DNA Methylomic Analysis.

    PubMed

    Roubal, Ivan; Park, Sun Joo; Kim, Yong

    2016-01-01

    Embryonic stem cells are self-renewing pluripotent cells with competency to differentiate into all three-germ lineages. Many studies have demonstrated the importance of genetic and epigenetic molecular mechanisms in the maintenance of self-renewal and pluripotency. Stem cells are under unique molecular and cellular regulations different from somatic cells. Proper regulation should be ensured to maintain their unique self-renewal and undifferentiated characteristics. Understanding key mechanisms in stem cell biology will be important for the successful application of stem cells for regenerative therapeutic medicine. More importantly practical use of stem cells will require our knowledge on how to properly direct and differentiate stem cells into the necessary type of cells. Embryonic stem cells and adult stem cells have been used as study models to unveil molecular and cellular mechanisms in various signaling pathways. They are especially beneficial to developmental studies where in vivo molecular/cellular study models are not available. We have derived neural stem cells from human embryonic stem cells as a model to study the effect of teratogen in neural development. We have tested commercial neural differentiation system and successfully derived neural precursor cells exhibiting key molecular features of neural stem cells, which will be useful for experimental application.

  17. Efficient cultivation of neural stem cells with controlled delivery of FGF-2.

    PubMed

    Galderisi, U; Peluso, G; Di Bernardo, G; Calarco, A; D'Apolito, M; Petillo, O; Cipollaro, M; Fusco, F R; Melone, M A B

    2013-01-01

    Neural stem cells (NSCs) raised the hope for cell-based therapies in human neurodevelopmental and neurodegenerative diseases. Current research strategies aim to isolate, enrich, and propagate homogeneous populations of neural stem cells. Unfortunately, several concerns with NSC cultures currently may limit their therapeutic promise. Exhaustion of growth factors and/or their uncontrolled release with burst and fall in their concentration may greatly affect the in vitro behavior of NSCs. In this context, we investigate whether a device containing heparan sulfate (HS), which is a co-factor in growth factor-mediated cell proliferation and differentiation, could potentiate and prolong the delivery of fibroblast growth factor-2 (FGF-2) and thus improve in vitro NSC cultivation. We demonstrated that NSCs cultivated in media with a controlled release of FGF-2 from a polyelectrolyte polymer showed a higher proliferation rate, and reduced apoptosis and senescence. In these experimental conditions NSCs preserve their stemness properties for a longer period of time compared with controls. Also of interest is that cell fate properties are conserved as well. The controlled release of FGF-2 reduced the level of oxidative stress and this is associated with a lower level of damaged DNA. This result may explain the reduced level of senescence and apoptosis in NSCs cultivated in the presence of hydrogel-releasing FGF-2.

  18. Three-dimensional hydrogel cell culture systems for modeling neural tissue

    NASA Astrophysics Data System (ADS)

    Frampton, John

    Two-dimensional (2-D) neural cell culture systems have served as physiological models for understanding the cellular and molecular events that underlie responses to physical and chemical stimuli, control sensory and motor function, and lead to the development of neurological diseases. However, the development of three-dimensional (3-D) cell culture systems will be essential for the advancement of experimental research in a variety of fields including tissue engineering, chemical transport and delivery, cell growth, and cell-cell communication. In 3-D cell culture, cells are provided with an environment similar to tissue, in which they are surrounded on all sides by other cells, structural molecules and adhesion ligands. Cells grown in 3-D culture systems display morphologies and functions more similar to those observed in vivo, and can be cultured in such a way as to recapitulate the structural organization and biological properties of tissue. This thesis describes a hydrogel-based culture system, capable of supporting the growth and function of several neural cell types in 3-D. Alginate hydrogels were characterized in terms of their biomechanical and biochemical properties and were functionalized by covalent attachment of whole proteins and peptide epitopes. Methods were developed for rapid cross-linking of alginate hydrogels, thus permitting the incorporation of cells into 3-D scaffolds without adversely affecting cell viability or function. A variety of neural cell types were tested including astrocytes, microglia, and neurons. Cells remained viable and functional for longer than two weeks in culture and displayed process outgrowth in 3-D. Cell constructs were created that varied in cell density, type and organization, providing experimental flexibility for studying cell interactions and behavior. In one set of experiments, 3-D glial-endothelial cell co-cultures were used to model blood-brain barrier (BBB) structure and function. This co-culture system was

  19. Triennial Growth Symposium: neural regulation of feed intake: modification by hormones, fasting, and disease.

    PubMed

    Sartin, J L; Whitlock, B K; Daniel, J A

    2011-07-01

    Appetite is a complex process that results from the integration of multiple signals at the hypothalamus. The hypothalamus receives neural signals; hormonal signals such as leptin, cholecystokinin, and ghrelin; and nutrient signals such as glucose, FFA, AA, and VFA. This effect is processed by a specific sequence of neurotransmitters beginning with the arcuate nucleus and orexigenic cells containing neuropeptide Y or agouti-related protein and anorexigenic cells containing proopiomelanocortin (yielding the neurotransmitter α-melanocyte-stimulating hormone) or cells expressing cocaine amphetamine-related transcript. These so-called first-order neurons act on second-order orexigenic neurons (containing either melanin-concentrating hormone or orexin) or act on anorexigenic neurons (e.g., expressing corticotropin-releasing hormone) to alter feed intake. In addition, satiety signals from the liver and gastrointestinal tract signal through the vagus nerve to the nucleus tractus solitarius to cause meal termination, and in combination with the hypothalamus, integrate the various signals to determine the feeding response. The activities of these neuronal pathways are also influenced by numerous factors such as nutrients, fasting, and disease to modify appetite and hence affect growth and reproduction. This review will begin with the central nervous system pathways and then discuss the ways in which hormones and metabolites may alter the process to affect feed intake with emphasis on farm animals.

  20. Human Neural Cell-Based Biosensor

    DTIC Science & Technology

    2010-12-10

    botulinum toxin, and (4) development of fluorescence based assays for proliferation, mitochondrial function and reactive oxygen species generation as sensor...dopaminergic neurons, motoneurons and astrocytes using defined medium conditions, (3) development of cell-based methods to detect botulinum toxin, and...neurons are the target cell type for botulinum toxin and would be a useful cell type for the detection of this potential bioterrorism agent. Astrocytes

  1. Human Neural Cell-Based Biosensor

    DTIC Science & Technology

    2010-06-11

    mechanism(s)-of- action for known developmental neurotoxicants (Yamauchi, et al., 2007; Lein, P., et al., 2000; Howard, et al., 2005; Audesirk , et...Inc. Page 8 of 9 CONFIDENTIAL REFERENCES Audesirk , T, et al. (1991) Effects of inorganic lead on the differentiation and growth of cultured

  2. Monitoring the growth of the neural representations of new animal concepts.

    PubMed

    Bauer, Andrew James; Just, Marcel Adam

    2015-08-01

    Although enormous progress has recently been made in identifying the neural representations of individual object concepts, relatively little is known about the growth of a neural knowledge representation as a novel object concept is being learned. In this fMRI study, the growth of the neural representations of eight individual extinct animal concepts was monitored as participants learned two features of each animal, namely its habitat (i.e., a natural dwelling or scene) and its diet or eating habits. Dwelling/scene information and diet/eating-related information have each been shown to activate their own characteristic brain regions. Several converging methods were used here to capture the emergence of the neural representation of a new animal feature within these characteristic, a priori-specified brain regions. These methods include statistically reliable identification (classification) of the eight newly acquired multivoxel patterns, analysis of the neural representational similarity among the newly learned animal concepts, and conventional GLM assessments of the activation in the critical regions. Moreover, the representation of a recently learned feature showed some durability, remaining intact after another feature had been learned. This study provides a foundation for brain research to trace how a new concept makes its way from the words and graphics used to teach it, to a neural representation of that concept in a learner's brain.

  3. Neural stem cells: generating and regenerating the brain.

    PubMed

    Gage, Fred H; Temple, Sally

    2013-10-30

    One of the landmark events of the past 25 years in neuroscience research was the establishment of neural stem cells (NSCs) as a life-long source of neurons and glia, a concept that shattered the dogma that the nervous system lacked regenerative power. Stem cells afford the plasticity to generate, repair, and change nervous system function. Combined with reprogramming technology, human somatic cell-derived NSCs and their progeny can model neurological diseases with improved accuracy. As technology advances, we anticipate further important discoveries and novel therapies based on the knowledge and application of these powerful cells.

  4. Tissue growth and tumorigenesis in Drosophila: cell polarity and the Hippo pathway.

    PubMed

    Richardson, Helena E; Portela, Marta

    2017-03-28

    Cell polarity regulation is critical for defining membrane domains required for the establishment and maintenance of the apical-basal axis in epithelial cells (apico-basal polarity), asymmetric cell divisions, planar organization of tissues (planar cell polarity), and the formation of the front-rear axis in cell migration (front-rear polarity). In the vinegar fly, Drosophila melanogaster, cell polarity regulators also interact with the Hippo tissue growth control signaling pathway. In this review we survey the recent Drosophila literature linking cell polarity regulators with the Hippo pathway in epithelial tissue growth, neural stem cell asymmetric divisions and in cell migration in physiological and tumorigenic settings.

  5. High glucose suppresses embryonic stem cell differentiation into neural lineage cells.

    PubMed

    Yang, Penghua; Shen, Wei-bin; Reece, E Albert; Chen, Xi; Yang, Peixin

    2016-04-01

    Abnormal neurogenesis occurs during embryonic development in human diabetic pregnancies and in animal models of diabetic embryopathy. Our previous studies in a mouse model of diabetic embryopathy have implicated that high glucose of maternal diabetes delays neurogenesis in the developing neuroepithelium leading to neural tube defects. However, the underlying process in high glucose-impaired neurogenesis is uncharacterized. Neurogenesis from embryonic stem (ES) cells provides a valuable model for understanding the abnormal neural lineage development under high glucose conditions. ES cells are commonly generated and maintained in high glucose (approximately 25 mM glucose). Here, the mouse ES cell line, E14, was gradually adapted to and maintained in low glucose (5 mM), and became a glucose responsive E14 (GR-E14) line. High glucose induced the endoplasmic reticulum stress marker, CHOP, in GR-E14 cells. Under low glucose conditions, the GR-E14 cells retained their pluripotency and capability to differentiate into neural lineage cells. GR-E14 cell differentiation into neural stem cells (Sox1 and nestin positive cells) was inhibited by high glucose. Neuron (Tuj1 positive cells) and glia (GFAP positive cells) differentiation from GR-E14 cells was also suppressed by high glucose. In addition, high glucose delayed GR-E14 differentiation into neural crest cells by decreasing neural crest markers, paired box 3 (Pax3) and paired box 7 (Pax7). Thus, high glucose impairs ES cell differentiation into neural lineage cells. The low glucose adapted and high glucose responsive GR-E14 cell line is a useful in vitro model for assessing the adverse effect of high glucose on the development of the central nervous system.

  6. Neuroprotection of VEGF-expression neural stem cells in neonatal cerebral palsy rats.

    PubMed

    Zheng, Xiang-Rong; Zhang, Shan-Shan; Yin, Fei; Tang, Jie-Lu; Yang, Yu-Jia; Wang, Xia; Zhong, Le

    2012-04-21

    Cerebral palsy (CP) is a very common neural system development disorder that can cause physical disability in human. Here, we studied the neuroprotective effect of vascular endothelial growth factor (VEGF)-transfected neural stem cells (NSCs) in newborn rats with cerebral palsy (CP). Seven-day-old Sprague-Dawley rats were randomly divided into four groups: sham operation (control group), PBS transplantation (PBS group), VEGF+NSCs transplantation (transgene NSCs group) and NSCs transplantation groups (NSCs group). PBS, Transgene NSCs and NSCs groups respectively received stereotactic injections of PBS, lentiviral vector (pGC-FU-VEGF) infected NSCs or a NSCs suspension in the left sensory-motor cortex 3 days after CP model was established. The NSCs activity, their impacts on neural cell growth and apoptosis, brain development and animal behaviors were examined on the animals up to age 35-days. As expected, unilateral carotid artery occlusion plus hypoxia (cerebral palsy model) resulted in severe neural developmental disorders, including slowed growth, increased in cortical neuron apoptosis, decreased cerebral cortex micro-vessel density and retarded behavior developments. Transplantation of NSCs not only resulted in increases in VEGF protein expression in rat brains, but also largely prevented the behavioral defects and brain tissue pathology that resulted from cerebral palsy procedure, with animals received VEGF transfected NSCs always being marginally better than these received un-transfected cells. In conclusion, NSCs transplantation can partially prevent/slow down the brain damages that are associated with CP in the newborn rats, suggesting a new possible strategy for CP treatment. Copyright © 2012 Elsevier B.V. All rights reserved.

  7. The Neural Cell Adhesion Molecule-Derived Peptide FGL Facilitates Long-Term Plasticity in the Dentate Gyrus in Vivo

    ERIC Educational Resources Information Center

    Dallerac, Glenn; Zerwas, Meike; Novikova, Tatiana; Callu, Delphine; Leblanc-Veyrac, Pascale; Bock, Elisabeth; Berezin, Vladimir; Rampon, Claire; Doyere, Valerie

    2011-01-01

    The neural cell adhesion molecule (NCAM) is known to play a role in developmental and structural processes but also in synaptic plasticity and memory of the adult animal. Recently, FGL, a NCAM mimetic peptide that binds to the Fibroblast Growth Factor Receptor 1 (FGFR-1), has been shown to have a beneficial impact on normal memory functioning, as…

  8. The Neural Cell Adhesion Molecule-Derived Peptide FGL Facilitates Long-Term Plasticity in the Dentate Gyrus in Vivo

    ERIC Educational Resources Information Center

    Dallerac, Glenn; Zerwas, Meike; Novikova, Tatiana; Callu, Delphine; Leblanc-Veyrac, Pascale; Bock, Elisabeth; Berezin, Vladimir; Rampon, Claire; Doyere, Valerie

    2011-01-01

    The neural cell adhesion molecule (NCAM) is known to play a role in developmental and structural processes but also in synaptic plasticity and memory of the adult animal. Recently, FGL, a NCAM mimetic peptide that binds to the Fibroblast Growth Factor Receptor 1 (FGFR-1), has been shown to have a beneficial impact on normal memory functioning, as…

  9. Potential of adult neural stem cells in stroke therapy.

    PubMed

    Andres, Robert H; Choi, Raymond; Steinberg, Gary K; Guzman, Raphael

    2008-11-01

    Despite state-of-the-art therapy, clinical outcome after stroke remains poor, with many patients left permanently disabled and dependent on care. Stem cell therapy has evolved as a promising new therapeutic avenue for the treatment of stroke in experimental studies, and recent clinical trials have proven its feasibility and safety in patients. Replacement of damaged cells and restoration of function can be accomplished by transplantation of different cell types, such as embryonic, fetal or adult stem cells, human fetal tissue and genetically engineered cell lines. Adult neural stem cells offer the advantage of avoiding the ethical problems associated with embryonic or fetal stem cells and can be harvested as autologous grafts from the individual patients. Furthermore, stimulation of endogenous adult stem cell-mediated repair mechanisms in the brain might offer new avenues for stroke therapy without the necessity of transplantation. However, important scientific issues need to be addressed to advance our understanding of the molecular mechanisms underlying the critical steps in cell-based repair to allow the introduction of these experimental techniques into clinical practice. This review describes up-to-date experimental concepts using adult neural stem cells for the treatment of stroke.

  10. Immune Influence on Adult Neural Stem Cell Regulation and Function

    PubMed Central

    Carpentier, Pamela A.; Palmer, Theo D.

    2009-01-01

    Neural stem cells (NSCs) lie at the heart of central nervous system development and repair, and deficiency or dysregulation of NSCs or their progeny can have significant consequences at any stage of life. Immune signaling is emerging as one of the influential variables that define resident NSC behavior. Perturbations in local immune signaling accompany virtually every injury or disease state and signaling cascades that mediate immune activation, resolution, or chronic persistence influence resident stem and progenitor cells. Some aspects of immune signaling are beneficial, promoting intrinsic plasticity and cell replacement, while others appear to inhibit the very type of regenerative response that might restore or replace neural networks lost in injury or disease. Here we review known and speculative roles that immune signaling plays in the postnatal and adult brain, focusing on how environments encountered in disease or injury may influence the activity and fate of endogenous or transplanted NSCs. PMID:19840551

  11. Comparison of the Odontogenic Differentiation Potential of Dental Follicle, Dental Papilla, and Cranial Neural Crest Cells.

    PubMed

    Chen, Gang; Sun, Qince; Xie, Li; Jiang, Zongting; Feng, Lian; Yu, Mei; Guo, Weihua; Tian, Weidong

    2015-07-01

    During tooth development, cells originating from the neural crest serve as precursors to the cells in the dental follicle and dental papilla. Therefore, the current study aimed to understand the associations of cranial neural crest cells (CNCCs), dental follicle cells (DFCs), and dental papilla cells (DPCs) by performing a parallel comparison to evaluate their odontogenic differentiation capacities. In this study, we harvested the 3 cells from C57/green fluorescent protein-positive mice or embryos and compared the cell morphology, surface antigens, microstructures, and gene and protein expression. Under the odontogenic microenvironments provided by treated dentin matrix, the odontogenic differentiations of the 3 cells were further compared in vitro and in vivo. The gene levels of DFCs in neurofilament, tubulin, and nestin were close to the DPCs, and in alkaline phosphatase, osteopontin, dentin matrix protein 1, and dentin sialophosphoprotein were the lowest in the 3 cells. However, Western blot results showed that DFCs possessed more similar protein profiles to CNCCs than DPCs, including collagen 1, transforming growth factor beta 1, osteopontin, neurofilament, and dentin matrix protein 1. Meanwhile, DFCs as 1 source of dental stem cells possessed high potency in odontogenic differentiation in vitro. Moreover, similar dentinlike tissues were observed in all 3 groups in vivo. CNCCs, DFCs, and DPCs possessed different biological characteristics in odontogenic differentiation. Copyright © 2015 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  12. Lgr5 Marks Neural Crest Derived Multipotent Oral Stromal Stem Cells.

    PubMed

    Boddupally, Keerthi; Wang, Guangfang; Chen, Yibu; Kobielak, Agnieszka

    2016-03-01

    It has been suggested that multipotent stem cells with neural crest (NC) origin persist into adulthood in oral mucosa. However their exact localization and role in normal homeostasis is unknown. In this study, we discovered that Lgr5 is expressed in NC cells during embryonic development, which give rise to the dormant stem cells in the adult tongue and oral mucosa. Those Lgr5 positive oral stromal stem cells display properties of NC stem cells including clonal growth and multipotent differentiation. RNA sequencing revealed that adult Lgr5+ oral stromal stem cells express high number of neural crest related markers like Sox9, Twist1, Snai1, Myc, Ets1, Crabp1, Epha2, and Itgb1. Using lineage-tracing experiments, we show that these cells persist more than a year in the ventral tongue and some areas of the oral mucosa and give rise to stromal progeny. In vivo transplantation demonstrated that these cells reconstitute the stroma. Our studies show for the first time that Lgr5 is expressed in the NC cells at embryonic day 9.5 (E9.5) and is maintained during embryonic development and postnataly in the stroma of the ventral tongue, and some areas of the oral mucosa and that Lgr5+ cells participate in the maintenance of the stroma. © 2015 AlphaMed Press.

  13. Bio-printing of collagen and VEGF-releasing fibrin gel scaffolds for neural stem cell culture.

    PubMed

    Lee, Yeong-Bae; Polio, Samuel; Lee, Wonhye; Dai, Guohao; Menon, Lata; Carroll, Rona S; Yoo, Seung-Schik

    2010-06-01

    Time-released delivery of soluble growth factors (GFs) in engineered hydrogel tissue constructs promotes the migration and proliferation of embedded cells, which is an important factor for designing scaffolds that ultimately aim for neural tissue regeneration. We report a tissue engineering technique to print murine neural stem cells (C17.2), collagen hydrogel, and GF (vascular endothelial growth factor: VEGF)-releasing fibrin gel to construct an artificial neural tissue. We examined the morphological changes of the printed C17.2 cells embedded in the collagen and its migration toward the fibrin gel. The cells showed high viability (92.89+/-2.32%) after printing, which was equivalent to that of manually-plated cells. C17.2 cells printed within 1mm from the border of VEGF-releasing fibrin gel showed GF-induced changes in their morphology. The cells printed in this range also migrated toward the fibrin gel, with the total migration distance of 102.4+/-76.1microm over 3days. The cells in the control samples (fibrin without the VEGF or VEGF printed directly in collagen) neither proliferated nor migrated. The results demonstrated that bio-printing of VEGF-containing fibrin gel supported sustained release of the GF in the collagen scaffold. The presented method can be gainfully used in the development of three-dimensional (3D) artificial tissue assays and neural tissue regeneration applications.

  14. Similarity in gene-regulatory networks suggests that cancer cells share characteristics of embryonic neural cells.

    PubMed

    Zhang, Zan; Lei, Anhua; Xu, Liyang; Chen, Lu; Chen, Yonglong; Zhang, Xuena; Gao, Yan; Yang, Xiaoli; Zhang, Min; Cao, Ying

    2017-08-04

    Cancer cells are immature cells resulting from cellular reprogramming by gene misregulation, and redifferentiation is expected to reduce malignancy. It is unclear, however, whether cancer cells can undergo terminal differentiation. Here, we show that inhibition of the epigenetic modification enzyme enhancer of zeste homolog 2 (EZH2), histone deacetylases 1 and 3 (HDAC1 and -3), lysine demethylase 1A (LSD1), or DNA methyltransferase 1 (DNMT1), which all promote cancer development and progression, leads to postmitotic neuron-like differentiation with loss of malignant features in distinct solid cancer cell lines. The regulatory effect of these enzymes in neuronal differentiation resided in their intrinsic activity in embryonic neural precursor/progenitor cells. We further found that a major part of pan-cancer-promoting genes and the signal transducers of the pan-cancer-promoting signaling pathways, including the epithelial-to-mesenchymal transition (EMT) mesenchymal marker genes, display neural specific expression during embryonic neurulation. In contrast, many tumor suppressor genes, including the EMT epithelial marker gene that encodes cadherin 1 (CDH1), exhibited non-neural or no expression. This correlation indicated that cancer cells and embryonic neural cells share a regulatory network, mediating both tumorigenesis and neural development. This observed similarity in regulatory mechanisms suggests that cancer cells might share characteristics of embryonic neural cells. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  15. Neural crest stem cells: discovery, properties and potential for therapy

    PubMed Central

    Achilleos, Annita; Trainor, Paul A

    2012-01-01

    Neural crest (NC) cells are a migratory cell population synonymous with vertebrate evolution. They generate a wide variety of cell and tissue types during embryonic and adult development including cartilage and bone, connective tissue, pigment and endocrine cells as well as neurons and glia amongst many others. Such incredible lineage potential combined with a limited capacity for self-renewal, which persists even into adult life, demonstrates that NC cells bear the key hallmarks of stem and progenitor cells. In this review, we describe the identification, characterization and isolation of NC stem and progenitor cells from different tissues in both embryo and adult organisms. We discuss their specific properties and their potential application in cell-based tissue and disease-specific repair. PMID:22231630

  16. Cell polarity and neurogenesis in embryonic stem cell-derived neural rosettes.

    PubMed

    Banda, Erin; McKinsey, Anna; Germain, Noelle; Carter, James; Anderson, Nickesha Camille; Grabel, Laura

    2015-04-15

    Embryonic stem cells (ESCs) undergoing neural differentiation form radial arrays of neural stem cells, termed neural rosettes. These structures manifest many of the properties associated with embryonic and adult neurogenesis, including cell polarization, interkinetic nuclear migration (INM), and a gradient of neuronal differentiation. We now identify novel rosette structural features that serve to localize key regulators of neurogenesis. Cells within neural rosettes have specialized basal as well as apical surfaces, based on localization of the extracellular matrix receptor β1 integrin. Apical processes of cells in mature rosettes terminate at the lumen, where adherens junctions are apparent. Primary cilia are randomly distributed in immature rosettes and tightly associated with the neural stem cell's apical domain as rosettes mature. Components of two signaling pathways known to regulate neurogenesis in vivo and in rosettes, Hedgehog and Notch, are apically localized, with the Hedgehog effector Smoothened (Smo) associated with primary cilia and the Notch pathway γ-secretase subunit Presenilin 2 associated with the adherens junction. Increased neuron production upon treatment with the Notch inhibitor DAPT suggests a major role for Notch signaling in maintaining the neural stem cell state, as previously described. A less robust outcome was observed with manipulation of Hedgehog levels, though consistent with a role in neural stem cell survival or proliferation. Inhibition of both pathways resulted in an additive effect. These data support a model by which cells extending a process to the rosette lumen maintain neural stem cell identity whereas release from this association, either through asymmetric cell division or apical abscission, promotes neuronal differentiation.

  17. Planar cell polarity-mediated induction of neural stem cell expansion during axolotl spinal cord regeneration.

    PubMed

    Rodrigo Albors, Aida; Tazaki, Akira; Rost, Fabian; Nowoshilow, Sergej; Chara, Osvaldo; Tanaka, Elly M

    2015-11-14

    Axolotls are uniquely able to mobilize neural stem cells to regenerate all missing regions of the spinal cord. How a neural stem cell under homeostasis converts after injury to a highly regenerative cell remains unknown. Here, we show that during regeneration, axolotl neural stem cells repress neurogenic genes and reactivate a transcriptional program similar to embryonic neuroepithelial cells. This dedifferentiation includes the acquisition of rapid cell cycles, the switch from neurogenic to proliferative divisions, and the re-expression of planar cell polarity (PCP) pathway components. We show that PCP induction is essential to reorient mitotic spindles along the anterior-posterior axis of elongation, and orthogonal to the cell apical-basal axis. Disruption of this property results in premature neurogenesis and halts regeneration. Our findings reveal a key role for PCP in coordinating the morphogenesis of spinal cord outgrowth with the switch from a homeostatic to a regenerative stem cell that restores missing tissue.

  18. Smart drugs for smarter stem cells: making SENSe (sphingolipid-enhanced neural stem cells) of ceramide.

    PubMed

    Bieberich, Erhard

    2008-01-01

    Ceramide and its derivative sphingosine-1-phosphate (S1P) are important signaling sphingolipids for neural stem cell apoptosis and differentiation. Most recently, our group has shown that novel ceramide analogs can be used to eliminate teratoma (stem cell tumor)-forming cells from a neural stem cell graft. In new studies, we found that S1P promotes survival of specific neural precursor cells that undergo differentiation to cells expressing oligodendroglial markers. Our studies suggest that a combination of novel ceramide and S1P analogs eliminates tumor-forming stem cells and at the same time, triggers oligodendroglial differentiation. This review discusses recent studies on the function of ceramide and S1P for the regulation of apoptosis, differentiation, and polarity in stem cells. We will also discuss results from ongoing studies in our laboratory on the use of sphingolipids in stem cell therapy. (c) 2008 S. Karger AG, Basel.

  19. Sonic hedgehog elevates N-myc gene expression in neural stem cells.

    PubMed

    Liu, Dongsheng; Wang, Shouyu; Cui, Yan; Shen, Lun; Du, Yanping; Li, Guilin; Zhang, Bo; Wang, Renzhi

    2012-08-05

    Proliferation of neural stem cells is regulated by the secreted signaling molecule sonic hedgehog. In this study, neural stem cells were infected with recombinant adeno-associated virus expressing sonic hedgehog-N-enhanced green fluorescent protein. The results showed that overexpression of sonic hedgehog in neural stem cells induced the increased expression of Gli1 and N-myc, a target gene of sonic hedgehog. These findings suggest that N-myc is a direct downstream target of the sonic hedgehog signal pathway in neural stem cells. Sonic hedgehog and N-myc are important mediators of sonic hedgehog-induced proliferation of neural stem cells.

  20. Purification of kidney epithelial cell growth inhibitors.

    PubMed Central

    Holley, R W; Böhlen, P; Fava, R; Baldwin, J H; Kleeman, G; Armour, R

    1980-01-01

    Two high molecular weight growth inhibitors have been isolated from the culture medium of BSC-1 cells, epithelial cells of African green monkey kidney. The purified kidney epithelial cell growth inhibitors, at ng/ml concentrations, reversibly arrest the growth of BSC-1 cells in the G1 phase of the cell cycle. Their action is selective; they are most active on BSC-1 cells, are less active as inhibitors of the growth of rat lung and human breast epithelial cells, and do not inhibit the growth of 3T3 mouse embryo fibroblasts ad human skin fibroblasts in culture. Their growth inhibitory action on BSC-1 cell cultures is counteracted by epidermal growth factor or calf serum. PMID:6969400

  1. Blood-neural barrier: its diversity and coordinated cell-to-cell communication.

    PubMed

    Choi, Yoon Kyung; Kim, Kyu-Won

    2008-05-31

    The cerebral microvessels possess barrier characteristics which are tightly sealed excluding many toxic substances and protecting neural tissues. The specialized blood-neural barriers as well as the cerebral microvascular barrier are recognized in the retina, inner ear, spinal cord, and cerebrospinal fluid. Microvascular endothelial cells in the brain closely interact with other components such as astrocytes, pericytes, perivascular microglia and neurons to form functional 'neurovascular unit'. Communication between endothelial cells and other surrounding cells enhances the barrier functions, consequently resulting in maintenance and elaboration of proper brain homeostasis. Furthermore, the disruption of the neurovascular unit is closely involved in cerebrovascular disorders. In this review, we focus on the location and function of these various blood-neural barriers, and the importance of the cell-to-cell communication for development and maintenance of the barrier integrity at the neurovascular unit. We also demonstrate the close relation between the alteration of the blood-neural barriers and cerebrovascular disorders.

  2. Neuralized functions cell autonomously to regulate Drosophila sense organ development.

    PubMed

    Yeh, E; Zhou, L; Rudzik, N; Boulianne, G L

    2000-09-01

    Neurogenic genes, including Notch and Delta, are thought to play important roles in regulating cell-cell interactions required for Drosophila sense organ development. To define the requirement of the neurogenic gene neuralized (neu) in this process, two independent neu alleles were used to generate mutant clones. We find that neu is required for determination of cell fates within the proneural cluster and that cells mutant for neu autonomously adopt neural fates when adjacent to wild-type cells. Furthermore, neu is required within the sense organ lineage to determine the fates of daughter cells and accessory cells. To gain insight into the mechanism by which neu functions, we used the GAL4/UAS system to express wild-type and epitope-tagged neu constructs. We show that Neu protein is localized primarily at the plasma membrane. We propose that the function of neu in sense organ development is to affect the ability of cells to receive Notch-Delta signals and thus modulate neurogenic activity that allows for the specification of non-neuronal cell fates in the sense organ.

  3. Angiogenic growth factors in neural embryogenesis and neoplasia.

    PubMed Central

    Zagzag, D.

    1995-01-01

    "Blood vessels have the power to increase within themselves which is according to the necessity whether natural or diseased. As a further proof that this is a general principle, we find that all growing parts are much more vascular than those that are come to their full growth; because growth is an operation beyond the simple support of the part. This is the reason why young animals are more vascular than those that are full grown. This is not peculiar to the natural operation of growth, but applies also to disease and restoration." PMID:7531952

  4. Rapid Induction of Neural Differentiation in Human Umbilical Cord Matrix Mesenchymal Stem Cells by cAMP-elevating Agents

    PubMed Central

    Shahbazi, Atefeh; Safa, Majid; Alikarami, Fatemeh; Kargozar, Saeid; Asadi, Mohammad Hossein; Joghataei, Mohammad Taghi; Soleimani, Mansoureh

    2016-01-01

    Human umbilical cord matrix (hUCM) is considered as a promising source of mesenchymal stem cells (MSCs) due to several advantages over other tissues. The potential of neural differentiation of hUCM-MSCs is of great interest in the context of treating neurodegenerative diseases. In recent years, considerable efforts have been made to establish in vitro conditions for improving the differentiation of hUCM-MSCs toward neuronal cells. In the present study, we evaluated the neural differentiation potential of hUCM-MSCs in the presence of cAMP-elevating agents forskolin and 3-isobutyl-1-methylxanthine (IBMX). hUCM-MSCs were isolated from fetal umbilical cord and characterized by flow cytometry analysis for mesenchymal specific markers. Mesodermal differentiation potential was assessed through selective media with lineage-specific induction factors. For assessment of neural differentiation, cells were cultured in the presence of cAMP-elevating agents for 8 and 24 h. The neuronal differentiated MSCs were characterized for neuronal specific markers by immunocytochemistry and western blotting. Isolated hUCM-MSCs were found positive for mesenchymal markers (CD73, CD90, and CD105) while negative for hematopoietic markers (CD34 and CD45) .Following neural induction, most cells represented neural-like cells morphology. Neural markers including β-tubulin III (Tuj-1), neuron-specific enolase (NSE), microtubule-associated protein-2 (MAP-2) and nestin were expressed in treated cells with respect to control group. The astrocyte specific marker, glial fibrillary acidic protein (GFAP) was also shown by immunofluorescence in treated cells. (These findings demonstrate that hUCM-MSCs have the ability to rapidly differentiate into neural cell types of neuron-like cells and astrocytes by cAMP-elevating agents without the presence of growth factors. PMID:27942503

  5. Rapid Induction of Neural Differentiation in Human Umbilical Cord Matrix Mesenchymal Stem Cells by cAMP-elevating Agents.

    PubMed

    Shahbazi, Atefeh; Safa, Majid; Alikarami, Fatemeh; Kargozar, Saeid; Asadi, Mohammad Hossein; Joghataei, Mohammad Taghi; Soleimani, Mansoureh

    2016-01-01

    Human umbilical cord matrix (hUCM) is considered as a promising source of mesenchymal stem cells (MSCs) due to several advantages over other tissues. The potential of neural differentiation of hUCM-MSCs is of great interest in the context of treating neurodegenerative diseases. In recent years, considerable efforts have been made to establish in vitro conditions for improving the differentiation of hUCM-MSCs toward neuronal cells. In the present study, we evaluated the neural differentiation potential of hUCM-MSCs in the presence of cAMP-elevating agents forskolin and 3-isobutyl-1-methylxanthine (IBMX). hUCM-MSCs were isolated from fetal umbilical cord and characterized by flow cytometry analysis for mesenchymal specific markers. Mesodermal differentiation potential was assessed through selective media with lineage-specific induction factors. For assessment of neural differentiation, cells were cultured in the presence of cAMP-elevating agents for 8 and 24 h. The neuronal differentiated MSCs were characterized for neuronal specific markers by immunocytochemistry and western blotting. Isolated hUCM-MSCs were found positive for mesenchymal markers (CD73, CD90, and CD105) while negative for hematopoietic markers (CD34 and CD45) .Following neural induction, most cells represented neural-like cells morphology. Neural markers including β-tubulin III (Tuj-1), neuron-specific enolase (NSE), microtubule-associated protein-2 (MAP-2) and nestin were expressed in treated cells with respect to control group. The astrocyte specific marker, glial fibrillary acidic protein (GFAP) was also shown by immunofluorescence in treated cells. (These findings demonstrate that hUCM-MSCs have the ability to rapidly differentiate into neural cell types of neuron-like cells and astrocytes by cAMP-elevating agents without the presence of growth factors.

  6. Astaxanthin Improves Stem Cell Potency via an Increase in the Proliferation of Neural Progenitor Cells

    PubMed Central

    Kim, Jeong-Hwan; Nam, Soo-Wan; Kim, Byung-Woo; Choi, Woobong; Lee, Jong-Hwan; Kim, Wun-Jae; Choi, Yung-Hyun

    2010-01-01

    The present study was designed to investigate the question of whether or not astaxanthin improves stem cell potency via an increase in proliferation of neural progenitor cells (NPCs). Treatment with astaxanthin significantly increased proliferation and colony formation of NPCs. For identification of possible activated signaling molecules involved in active cell proliferation occurring after astaxanthin treatment, total protein levels of several proliferation-related proteins, and expression levels of proliferation-related transcription factors, were assessed in NPCs. In Western blot analysis, astaxanthin induced significant activation of phosphatidylinositol 3-kinase (PI3K) and its downstream mediators in a time-dependent manner. Results of RT-PCR analysis showed upregulation of proliferation-related transcription factors and stemness genes. To estimate the relevance of PI3K and mitogen-activated protein, or extracellular signal-regulated kinase kinase (MEK) signaling pathways in cell growth of astaxanthin-treated NPCs, inhibition assays were performed with LY294002, a specific inhibitor of PI3K, and PD98059, a specific inhibitor of MEK, respectively. These results clearly showed that astaxanthin induces proliferation of NPCs via activation of the PI3K and MEK signaling pathways and improves stem cell potency via stemness acting signals. PMID:21614195

  7. Rapamycin promotes Schwann cell migration and nerve growth factor secretion

    PubMed Central

    Liu, Fang; Zhang, Haiwei; Zhang, Kaiming; Wang, Xinyu; Li, Shipu; Yin, Yixia

    2014-01-01

    Rapamycin, similar to FK506, can promote neural regeneration in vitro. We assumed that the mechanisms of action of rapamycin and FK506 in promoting peripheral nerve regeneration were similar. This study compared the effects of different concentrations of rapamycin and FK506 on Schwann cells and investigated effects and mechanisms of rapamycin on improving peripheral nerve regeneration. Results demonstrated that the lowest rapamycin concentration (1.53 nmol/L) more significantly promoted Schwann cell migration than the highest FK506 concentration (100μmol/L). Rapamycin promoted the secretion of nerve growth factors and upregulated growth-associated protein 43 expression in Schwann cells, but did not significantly affect Schwann cell proliferation. Therefore, rapamycin has potential application in peripheral nerve regeneration therapy. PMID:25206862

  8. Inhibition of glycogen synthase kinase-3 (GSK3) promotes the neural differentiation of full-term amniotic fluid-derived stem cells towards neural progenitor cells.

    PubMed

    Gao, Liyang; Zhao, Mingyan; Ye, Wei; Huang, Jinzhi; Chu, Jiaqi; Yan, Shouquan; Wang, Chaojun; Zeng, Rong

    2016-08-01

    The amniotic fluid has a heterogeneous population of cells. Some human amniotic fluid-derived stem (hAFS) cells have been shown to harbor the potential to differentiate into neural cells. However, the neural differentiation efficiency of hAFS cells remains low. In this study, we isolated CD117-positive hAFS cells from amniotic fluid and then examined the pluripotency of these cells through the formation of embryoid bodies (EBs). Additionally, we induced the neural differentiation of these cells using neuroectodermal medium. This study revealed that the GSK3-beta inhibitor SB216763 was able to stimulate the proliferation of CD117-positive hAFS cells without influencing their undifferentiated state. Moreover, SB216763 can efficiently promote the neural differentiation of CD117-positive hAFS cells towards neural progenitor cells in the presence of DMEM/F12 and N2 supplement. These findings provide an easy and low-cost method to maintain the proliferation of hAFS cells, as well as induce an efficacious generation of neural progenitor cells from hAFS cells. Such induction of the neural commitment of hAFS cells may provide an option for the treatment of neurodegenerative diseases by hAFS cells-based therapies.

  9. The early postnatal nonhuman primate neocortex contains self-renewing multipotent neural progenitor cells.

    PubMed

    Homman-Ludiye, Jihane; Merson, Tobias D; Bourne, James A

    2012-01-01

    The postnatal neocortex has traditionally been considered a non-neurogenic region, under non-pathological conditions. A few studies suggest, however, that a small subpopulation of neural cells born during postnatal life can differentiate into neurons that take up residence within the neocortex, implying that postnatal neurogenesis could occur in this region, albeit at a low level. Evidence to support this hypothesis remains controversial while the source of putative neural progenitors responsible for generating new neurons in the postnatal neocortex is unknown. Here we report the identification of self-renewing multipotent neural progenitor cells (NPCs) derived from the postnatal day 14 (PD14) marmoset monkey primary visual cortex (V1, striate cortex). While neuronal maturation within V1 is well advanced by PD14, we observed cells throughout this region that co-expressed Sox2 and Ki67, defining a population of resident proliferating progenitor cells. When cultured at low density in the presence of epidermal growth factor (EGF) and/or fibroblast growth factor 2 (FGF-2), dissociated V1 tissue gave rise to multipotent neurospheres that exhibited the ability to differentiate into neurons, oligodendrocytes and astrocytes. While the capacity to generate neurones and oligodendrocytes was not observed beyond the third passage, astrocyte-restricted neurospheres could be maintained for up to 6 passages. This study provides the first direct evidence for the existence of multipotent NPCs within the postnatal neocortex of the nonhuman primate. The potential contribution of neocortical NPCs to neural repair following injury raises exciting new possibilities for the field of regenerative medicine.

  10. Emergence of the small-world architecture in neural networks by activity dependent growth

    NASA Astrophysics Data System (ADS)

    Gafarov, F. M.

    2016-11-01

    In this paper, we propose a model describing the growth and development of neural networks based on the latest achievements of experimental neuroscience. The model is based on two evolutionary equations. The first equation is for the evolution of the neurons state and the second is for the growth of axon tips. By using the model, we demonstrated the neuronal growth process from disconnected neurons to fully connected three-dimensional networks. For the analysis of the network's connections structure, we used the random graphs theory methods. It is shown that the growth in neural networks results in the formation of a well-known ;small-world; network model. The analysis of the connectivity distribution shows the presence of a strictly non-Gaussian but no scale-free degree distribution for the in-degree node distribution. In terms of the graphs theory, this study developed a new model of dynamic graph.

  11. Direct cell-cell contact with the vascular niche maintains quiescent neural stem cells

    PubMed Central

    Ottone, Cristina; Krusche, Benjamin; Whitby, Ariadne; Clements, Melanie; Quadrato, Giorgia; Pitulescu, Mara E.; Adams, Ralf H.; Parrinello, Simona

    2014-01-01

    The vasculature is a prominent component of the subventricular zone neural stem cell niche. Although quiescent neural stem cells physically contact blood vessels at specialised endfeet, the significance of this interaction is not understood. In contrast, it is well established that vasculature-secreted soluble factors promote lineage progression of committed progenitors. Here we specifically investigated the role of cell-cell contact-dependent signalling in the vascular niche. Unexpectedly, we find that direct cell-cell interactions with endothelial cells enforces quiescence and promotes stem cell identity. Mechanistically, endothelial ephrinB2 and Jagged1 mediate these effects by suppressing cell-cycle entry downstream of mitogens and inducing stemness genes to jointly inhibit differentiation. In vivo, endothelial-specific ablation of either of the genes which encode these proteins, Efnb2 and Jag1 respectively, aberrantly activates quiescent stem cells, resulting in depletion. Thus, we identify the vasculature as a critical niche compartment for stem cell maintenance, furthering our understanding of how anchorage to the niche maintains stem cells within a pro-differentiative microenvironment. PMID:25283993

  12. Collection of neural inducing factors from PA6 cells using heparin solution and their immobilization on plastic culture dishes for the induction of neurons from embryonic stem cells.

    PubMed

    Yamazoe, Hironori; Murakami, Yoshinobu; Mizuseki, Kenji; Sasai, Yoshiki; Iwata, Hiroo

    2005-10-01

    Embryonic stem (ES) cells have the ability to replicate themselves and differentiate into various mature cells. Recently, dopaminergic neurons were efficiently induced from ES cells using mouse stromal cells (PA6 cells) as a feeder cell layer. This simple procedure seems to be very efficient to obtain dopamine-releasing cells for future clinical cell transplantation treatment of Parkinson's disease. In this study, we prepared stock solutions containing neural inducing factors (NIFs) by washing PA6 cells with phosphate-buffered saline containing heparin. ES cells grew successfully in culture media supplemented with 33 v/v% NIFs stock solution, and the rate of neural differentiation of ES cell progeny increased with increasing heparin concentration in the culture media. In addition, NIFs-immobilized surfaces were prepared by exposing polyethyleneimine-modified surfaces to NIFs stock solutions. The NIFs-immobilized culture dish effectively supported cell growth as the culture medium supplemented with NIFs stock did, but its induction effect to dopaminergic neurons from ES cells was much smaller than free NIFs. NIFs stock solutions have two different activities. One can stimulate cell growth and the other induces differentiation of ES cells to the neural fate when heparin existed. The former factors were effectively immobilized on the culture dish, but those that induce differentiation may not be. Further optimization is required.

  13. Hematopoietic Stem Cells in Neural-crest Derived Bone Marrow.

    PubMed

    Jiang, Nan; Chen, Mo; Yang, Guodong; Xiang, Lusai; He, Ling; Hei, Thomas K; Chotkowski, Gregory; Tarnow, Dennis P; Finkel, Myron; Ding, Lei; Zhou, Yanheng; Mao, Jeremy J

    2016-12-21

    Hematopoietic stem cells (HSCs) in the endosteum of mesoderm-derived appendicular bones have been extensively studied. Neural crest-derived bones differ from appendicular bones in developmental origin, mode of bone formation and pathological bone resorption. Whether neural crest-derived bones harbor HSCs is elusive. Here, we discovered HSC-like cells in postnatal murine mandible, and benchmarked them with donor-matched, mesoderm-derived femur/tibia HSCs, including clonogenic assay and long-term culture. Mandibular CD34 negative, LSK cells proliferated similarly to appendicular HSCs, and differentiated into all hematopoietic lineages. Mandibular HSCs showed a consistent deficiency in lymphoid differentiation, including significantly fewer CD229 + fractions, PreProB, ProB, PreB and B220 + slgM cells. Remarkably, mandibular HSCs reconstituted irradiated hematopoietic bone marrow in vivo, just as appendicular HSCs. Genomic profiling of osteoblasts from mandibular and femur/tibia bone marrow revealed deficiencies in several HSC niche regulators among mandibular osteoblasts including Cxcl12. Neural crest derived bone harbors HSCs that function similarly to appendicular HSCs but are deficient in the lymphoid lineage. Thus, lymphoid deficiency of mandibular HSCs may be accounted by putative niche regulating genes. HSCs in craniofacial bones have functional implications in homeostasis, osteoclastogenesis, immune functions, tumor metastasis and infections such as osteonecrosis of the jaw.

  14. Coordinating cell and tissue behavior during zebrafish neural tube morphogenesis.

    PubMed

    Araya, Claudio; Ward, Laura C; Girdler, Gemma C; Miranda, Miguel

    2016-03-01

    The development of a vertebrate neural epithelium with well-organized apico-basal polarity and a central lumen is essential for its proper function. However, how this polarity is established during embryonic development and the potential influence of surrounding signals and tissues on such organization has remained less understood. In recent years the combined superior transparency and genetics of the zebrafish embryo has allowed for in vivo visualization and quantification of the cellular and molecular dynamics that govern neural tube structure. Here, we discuss recent studies revealing how co-ordinated cell-cell interactions coupled with adjacent tissue dynamics are critical to regulate final neural tissue architecture. Furthermore, new findings show how the spatial regulation and timing of orientated cell division is key in defining precise lumen formation at the tissue midline. In addition, we compare zebrafish neurulation with that of amniotes and amphibians in an attempt to understand the conserved cellular mechanisms driving neurulation and resolve the apparent differences among animals. Zebrafish neurulation not only offers fundamental insights into early vertebrate brain development but also the opportunity to explore in vivo cell and tissue dynamics during complex three-dimensional animal morphogenesis. © 2015 Wiley Periodicals, Inc.

  15. Hematopoietic Stem Cells in Neural-crest Derived Bone Marrow

    PubMed Central

    Jiang, Nan; Chen, Mo; Yang, Guodong; Xiang, Lusai; He, Ling; Hei, Thomas K.; Chotkowski, Gregory; Tarnow, Dennis P.; Finkel, Myron; Ding, Lei; Zhou, Yanheng; Mao, Jeremy J.

    2016-01-01

    Hematopoietic stem cells (HSCs) in the endosteum of mesoderm-derived appendicular bones have been extensively studied. Neural crest-derived bones differ from appendicular bones in developmental origin, mode of bone formation and pathological bone resorption. Whether neural crest-derived bones harbor HSCs is elusive. Here, we discovered HSC-like cells in postnatal murine mandible, and benchmarked them with donor-matched, mesoderm-derived femur/tibia HSCs, including clonogenic assay and long-term culture. Mandibular CD34 negative, LSK cells proliferated similarly to appendicular HSCs, and differentiated into all hematopoietic lineages. Mandibular HSCs showed a consistent deficiency in lymphoid differentiation, including significantly fewer CD229 + fractions, PreProB, ProB, PreB and B220 + slgM cells. Remarkably, mandibular HSCs reconstituted irradiated hematopoietic bone marrow in vivo, just as appendicular HSCs. Genomic profiling of osteoblasts from mandibular and femur/tibia bone marrow revealed deficiencies in several HSC niche regulators among mandibular osteoblasts including Cxcl12. Neural crest derived bone harbors HSCs that function similarly to appendicular HSCs but are deficient in the lymphoid lineage. Thus, lymphoid deficiency of mandibular HSCs may be accounted by putative niche regulating genes. HSCs in craniofacial bones have functional implications in homeostasis, osteoclastogenesis, immune functions, tumor metastasis and infections such as osteonecrosis of the jaw. PMID:28000662

  16. Liquid crystal cells and optical fibers in neural network implementation

    NASA Astrophysics Data System (ADS)

    Domanski, Andrzej W.; Buczynski, Ryszard; Sierakowski, Marek W.

    1995-08-01

    Optical binary computer may be as easy to operate as parallel system. For such configuration Boolean logic is not very convenient and therefore neural networks should be introduced. In works leading to the paper we used liquid crystal cells as a standard system of liquid crystalline layer between to conducting electrodes in 'sandwich' geometry. We have used 25 micrometers display cells filled with nematic 6CHBT working on 'twisted nematic' effect. Based on such elements a mode of a simple Hopfield network was set up. More advanced experiments were carried out on a model of neurone with supervised learning. The model consists of four laser diodes pigtailed to the multimode optical fibers with 50 micrometers core diameter. The directional couplers help to control the level of input optical power. Four liquid crystal cells allow to change the transmission level according to superivised learning requirements. All the signals were detected by one photodiode. The presented results of experiments are in excellent agreement with theoretical predictions. An additional study was done to check the possibility to build up a linear neural network with Grossberg layer, a neural network with Kohonen layer, and a counter propagation network with two layers of neurones. We have proved that such models may be set up based on simple liquid crystals cells and optical fiber networks.

  17. L1 Retrotransposition in Neural Progenitor Cells.

    PubMed

    Muotri, Alysson R

    2016-01-01

    Long interspersed nucleotide element 1 (LINE-1 or L1) is a family of non-LTR retrotransposons that can replicate and reintegrate into the host genome. L1s have considerably influenced mammalian genome evolution by retrotransposing during germ cell development or early embryogenesis, leading to massive genome expansion. For many years, L1 retrotransposons were viewed as a selfish DNA parasite that had no contribution in somatic cells. Historically, L1s were thought to only retrotranspose during gametogenesis and in neoplastic processes, but recent studies have shown that L1s are extremely active in the mouse, rat, and human neuronal progenitor cells (NPCs). These de novo L1 insertions can impact neuronal transcriptional expression, creating unique transcriptomes of individual neurons, possibly contributing to the uniqueness of the individual cognition and mental disorders in humans.

  18. Selective Differentiation of Neural Progenitor Cells by High-Epitope Density Nanofibers

    NASA Astrophysics Data System (ADS)

    Silva, Gabriel A.; Czeisler, Catherine; Niece, Krista L.; Beniash, Elia; Harrington, Daniel A.; Kessler, John A.; Stupp, Samuel I.

    2004-02-01

    Neural progenitor cells were encapsulated in vitro within a three-dimensional network of nanofibers formed by self-assembly of peptide amphiphile molecules. The self-assembly is triggered by mixing cell suspensions in media with dilute aqueous solutions of the molecules, and cells survive the growth of the nanofibers around them. These nanofibers were designed to present to cells the neurite-promoting laminin epitope IKVAV at nearly van der Waals density. Relative to laminin or soluble peptide, the artificial nanofiber scaffold induced very rapid differentiation of cells into neurons, while discouraging the development of astrocytes. This rapid selective differentiation is linked to the amplification of bioactive epitope presentation to cells by the nanofibers.

  19. Vagal neural crest cell migratory behavior: A transition between the cranial and trunk crest

    PubMed Central

    Kuo, Bryan R.; Erickson, Carol A.

    2011-01-01

    Migration and differentiation of cranial neural crest cells are largely controlled by environmental cues, whereas pathfinding at the trunk level is dictated by cell-autonomous molecular changes owing to early specification of the premigratory crest. Here, we investigated the migration and patterning of vagal neural crest cells. We show that: 1) vagal neural crest cells exhibit some developmental bias and 2) they take separate pathways to the heart and to the gut. Together these observations suggest that prior specification dictates initial pathway choice. However, when we challenged the vagal neural crest cells with different migratory environments, we observed that the behavior of the anterior vagal neural crest cells (somite-level 1-3) exhibit considerable migratory plasticity whereas the posterior vagal neural crest cells (somite-level 5-7) are more restricted in their behavior. We conclude that the vagal neural crest is a transitional population that has evolved between the head and the trunk. PMID:22016183

  20. Llgl1 Connects Cell Polarity with Cell-Cell Adhesion in Embryonic Neural Stem Cells.

    PubMed

    Jossin, Yves; Lee, Minhui; Klezovitch, Olga; Kon, Elif; Cossard, Alexia; Lien, Wen-Hui; Fernandez, Tania E; Cooper, Jonathan A; Vasioukhin, Valera

    2017-06-05

    Malformations of the cerebral cortex (MCCs) are devastating developmental disorders. We report here that mice with embryonic neural stem-cell-specific deletion of Llgl1 (Nestin-Cre/Llgl1(fl/fl)), a mammalian ortholog of the Drosophila cell polarity gene lgl, exhibit MCCs resembling severe periventricular heterotopia (PH). Immunohistochemical analyses and live cortical imaging of PH formation revealed that disruption of apical junctional complexes (AJCs) was responsible for PH in Nestin-Cre/Llgl1(fl/fl) brains. While it is well known that cell polarity proteins govern the formation of AJCs, the exact mechanisms remain unclear. We show that LLGL1 directly binds to and promotes internalization of N-cadherin, and N-cadherin/LLGL1 interaction is inhibited by atypical protein kinase C-mediated phosphorylation of LLGL1, restricting the accumulation of AJCs to the basolateral-apical boundary. Disruption of the N-cadherin-LLGL1 interaction during cortical development in vivo is sufficient for PH. These findings reveal a mechanism responsible for the physical and functional connection between cell polarity and cell-cell adhesion machineries in mammalian cells. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Control of Neural Stem Cell Survival by Electroactive Polymer Substrates

    PubMed Central

    Lundin, Vanessa; Herland, Anna; Berggren, Magnus

    2011-01-01

    Stem cell function is regulated by intrinsic as well as microenvironmental factors, including chemical and mechanical signals. Conducting polymer-based cell culture substrates provide a powerful tool to control both chemical and physical stimuli sensed by stem cells. Here we show that polypyrrole (PPy), a commonly used conducting polymer, can be tailored to modulate survival and maintenance of rat fetal neural stem cells (NSCs). NSCs cultured on PPy substrates containing different counter ions, dodecylbenzenesulfonate (DBS), tosylate (TsO), perchlorate (ClO4) and chloride (Cl), showed a distinct correlation between PPy counter ion and cell viability. Specifically, NSC viability was high on PPy(DBS) but low on PPy containing TsO, ClO4 and Cl. On PPy(DBS), NSC proliferation and differentiation was comparable to standard NSC culture on tissue culture polystyrene. Electrical reduction of PPy(DBS) created a switch for neural stem cell viability, with widespread cell death upon polymer reduction. Coating the PPy(DBS) films with a gel layer composed of a basement membrane matrix efficiently prevented loss of cell viability upon polymer reduction. Here we have defined conditions for the biocompatibility of PPy substrates with NSC culture, critical for the development of devices based on conducting polymers interfacing with NSCs. PMID:21494605

  2. Coculture with embryonic stem cells improves neural differentiation of adipose tissue-derived stem cells.

    PubMed

    Bahmani, L; Taha, M F; Javeri, A

    2014-07-11

    Embryonic stem (ES) cells secrete some soluble factors which may affect the differentiation potential of adult stem cells toward different lineages. In the present study, we evaluated neural differentiation of mouse adipose tissue-derived stem cells (ADSCs) following coculture with ES cells. For this purpose, ADSCs were induced in a medium supplemented with a synthetic serum replacement and various concentrations of retinoic acid (RA). Then, third-passaged ADSCs were indirectly cocultured with ES cells, and the expression levels of pluripotency markers, OCT4 and Sox2, mesenchymal stem cell markers, CD73 and CD105, and proliferating cell nuclear antigen (PCNA), were assessed in the cocultured ADSCs. Moreover, the control and cocultured ADSCs were differentiated with or without RA treatment. We showed here that 2-week differentiated ADSCs expressed several neuron-specific genes, and RA treatment improved neural differentiation of the ADSCs. The expression levels of OCT4, Sox2 and PCNA were upregulated in the cocultured ADSCs. Moreover, coculture with the ES cells significantly improved neural differentiation of the ADSCs. Treatment of the cocultured ADSCs with RA diminished the expression of neural maturation markers. Coculture with the ES cells efficiently improves neural differentiation of the ADSCs. Non-contact coculture with the ES cells may be used as an efficient strategy to improve differentiation potential of adult stem cells for developmental studies and regenerative medicine.

  3. Artificial Neural Networks, and Evolutionary Algorithms as a systems biology approach to a data-base on fetal growth restriction.

    PubMed

    Street, Maria E; Buscema, Massimo; Smerieri, Arianna; Montanini, Luisa; Grossi, Enzo

    2013-12-01

    One of the specific aims of systems biology is to model and discover properties of cells, tissues and organisms functioning. A systems biology approach was undertaken to investigate possibly the entire system of intra-uterine growth we had available, to assess the variables of interest, discriminate those which were effectively related with appropriate or restricted intrauterine growth, and achieve an understanding of the systems in these two conditions. The Artificial Adaptive Systems, which include Artificial Neural Networks and Evolutionary Algorithms lead us to the first analyses. These analyses identified the importance of the biochemical variables IL-6, IGF-II and IGFBP-2 protein concentrations in placental lysates, and offered a new insight into placental markers of fetal growth within the IGF and cytokine systems, confirmed they had relationships and offered a critical assessment of studies previously performed. Copyright © 2013 Elsevier Ltd. All rights reserved.

  4. The Cellular Prion Protein Controls Notch Signaling in Neural Stem/Progenitor Cells.

    PubMed

    Martin-Lannerée, Séverine; Halliez, Sophie; Hirsch, Théo Z; Hernandez-Rapp, Julia; Passet, Bruno; Tomkiewicz, Céline; Villa-Diaz, Ana; Torres, Juan-Maria; Launay, Jean-Marie; Béringue, Vincent; Vilotte, Jean-Luc; Mouillet-Richard, Sophie

    2017-03-01

    The prion protein is infamous for its involvement in a group of neurodegenerative diseases known as Transmissible Spongiform Encephalopathies. In the longstanding quest to decipher the physiological function of its cellular isoform, PrP(C) , the discovery of its participation to the self-renewal of hematopoietic and neural stem cells has cast a new spotlight on its potential role in stem cell biology. However, still little is known on the cellular and molecular mechanisms at play. Here, by combining in vitro and in vivo murine models of PrP(C) depletion, we establish that PrP(C) deficiency severely affects the Notch pathway, which plays a major role in neural stem cell maintenance. We document that the absence of PrP(C) in a neuroepithelial cell line or in primary neurospheres is associated with drastically reduced expression of Notch ligands and receptors, resulting in decreased levels of Notch target genes. Similar alterations of the Notch pathway are recovered in the neuroepithelium of Prnp(-/-) embryos during a developmental window encompassing neural tube closure. In addition, in line with Notch defects, our data show that the absence of PrP(C) results in altered expression of Nestin and Olig2 as well as N-cadherin distribution. We further provide evidence that PrP(C) controls the expression of the epidermal growth factor receptor (EGFR) downstream from Notch. Finally, we unveil a negative feedback action of EGFR on both Notch and PrP(C) . As a whole, our study delineates a molecular scenario through which PrP(C) takes part to the self-renewal of neural stem and progenitor cells. Stem Cells 2017;35:754-765.

  5. Primitive mesodermal cells with a neural crest stem cell phenotype predominate proliferating infantile haemangioma.

    PubMed

    Itinteang, Tinte; Tan, Swee T; Brasch, Helen; Day, Darren J

    2010-09-01

    Infantile haemangioma is a tumour of the microvasculature characterised by aggressive angiogenesis during infancy and spontaneously gradual involution, often leaving a fibro-fatty residuum. The segmental distribution of a subgroup of infantile haemangioma, especially those associated with midline structural anomalies that constitute posterior fossa malformations-hemangiomas-arterial anomalies-cardiac defects-eye abnormalities-sternal cleft and supraumbilical raphe syndrome (PHACES), led us to investigate whether neural crest cells might be involved in the aetiology of this tumour. Immunohistochemical staining on paraffin embedded infantile haemangioma sections and immunocytochemical staining on cells derived from proliferating haemangioma cultures were performed. The endothelium of proliferating infantile haemangioma contains abundant cells that express the neurotrophin receptor (p75), a cell surface marker that identifies neural crest cells, and also for brachyury, a transcription factor expressed in cells of the primitive mesoderm. The endothelium is also immunoreactive for the haematopoietic stem cell marker, CD133; the endothelial-haematopoietic stem/progenitor marker, CD34; the endothelial cell markers, CD31 and VEGFR-2; and the mesenchymal stem cell markers, CD29 and vimentin. Additionally, immunoreactivity for the transcription factors, Sox 9 and Sox 10, that are expressed by prospective neural crest cells was also observed. Cells from microvessel-like structures were isolated from in vitro cultured haemangioma tissue explants embedded in a fibrin matrix. Immunostaining of these cells showed that they retained expression of the same lineage-specific markers that are detected on the paraffin embedded tissue sections. These data infer that infantile haemangioma is derived from primitive mesoderm and that the cells within the lesion have a neural crest stem cell phenotype, and they express proteins associated with haematopoietic, endothelial, neural crest and

  6. Differential effects of Th1, monocyte/macrophage and Th2 cytokine mixtures on early gene expression for glial and neural-related molecules in central nervous system mixed glial cell cultures: neurotrophins, growth factors and structural proteins

    PubMed Central

    Lisak, Robert P; Benjamins, Joyce A; Bealmear, Beverly; Nedelkoska, Liljana; Yao, Bin; Land, Susan; Studzinski, Diane

    2007-01-01

    Background In multiple sclerosis, inflammatory cells are found in both active and chronic lesions, and it is increasingly clear that cytokines are involved directly and indirectly in both formation and inhibition of lesions. We propose that cytokine mixtures typical of Th1 or Th2 lymphocytes, or monocyte/macrophages each induce unique molecular changes in glial cells. Methods To examine changes in gene expression that might occur in glial cells exposed to the secreted products of immune cells, we have used gene array analysis to assess the early effects of different cytokine mixtures on mixed CNS glia in culture. We compared the effects of cytokines typical of Th1 and Th2 lymphocytes and monocyte/macrophages (M/M) on CNS glia after 6 hours of treatment. Results In this paper we focus on changes with potential relevance for neuroprotection and axon/glial interactions. Each mixture of cytokines induced a unique pattern of changes in genes for neurotrophins, growth and maturation factors and related receptors; most notably an alternatively spliced form of trkC was markedly downregulated by Th1 and M/M cytokines, while Th2 cytokines upregulated BDNF. Genes for molecules of potential importance in axon/glial interactions, including cell adhesion molecules, connexins, and some molecules traditionally associated with neurons showed significant changes, while no genes for myelin-associated genes were regulated at this early time point. Unexpectedly, changes occurred in several genes for proteins initially associated with retina, cancer or bone development, and not previously reported in glial cells. Conclusion Each of the three cytokine mixtures induced specific changes in gene expression that could be altered by pharmacologic strategies to promote protection of the central nervous system. PMID:18088439

  7. Measurement of adherent cell mass and growth

    PubMed Central

    Park, Kidong; Millet, Larry J.; Kim, Namjung; Li, Huan; Jin, Xiaozhong; Popescu, Gabriel; Aluru, N. R.; Hsia, K. Jimmy; Bashir, Rashid

    2010-01-01

    The characterization of physical properties of cells such as their mass and stiffness has been of great interest and can have profound implications in cell biology, tissue engineering, cancer, and disease research. For example, the direct dependence of cell growth rate on cell mass for individual adherent human cells can elucidate the mechanisms underlying cell cycle progression. Here we develop an array of micro-electro-mechanical systems (MEMS) resonant mass sensors that can be used to directly measure the biophysical properties, mass, and growth rate of single adherent cells. Unlike conventional cantilever mass sensors, our sensors retain a uniform mass sensitivity over the cell attachment surface. By measuring the frequency shift of the mass sensors with growing (soft) cells and fixed (stiff) cells, and through analytical modeling, we derive the Young’s modulus of the unfixed cell and unravel the dependence of the cell mass measurement on cell stiffness. Finally, we grew individual cells on the mass sensors and measured their mass for 50+ hours. Our results demonstrate that adherent human colon epithelial cells have increased growth rates with a larger cell mass, and the average growth rate increases linearly with the cell mass, at 3.25%/hr. Our sensitive mass sensors with a position-independent mass sensitivity can be coupled with microscopy for simultaneous monitoring of cell growth and status, and provide an ideal method to study cell growth, cell cycle progression, differentiation, and apoptosis. PMID:21068372

  8. Heparanase confers a growth advantage to differentiating murine embryonic stem cells, and enhances oligodendrocyte formation.

    PubMed

    Xiong, Anqi; Kundu, Soumi; Forsberg, Maud; Xiong, Yuyuan; Bergström, Tobias; Paavilainen, Tanja; Kjellén, Lena; Li, Jin-Ping; Forsberg-Nilsson, Karin

    2017-10-01

    Heparan sulfate proteoglycans (HSPGs), ubiquitous components of mammalian cells, play important roles in development and homeostasis. These molecules are located primarily on the cell surface and in the pericellular matrix, where they interact with a multitude of macromolecules, including many growth factors. Manipulation of the enzymes involved in biosynthesis and modification of HSPG structures alters the properties of stem cells. Here, we focus on the involvement of heparanase (HPSE), the sole endo-glucuronidase capable of cleaving of HS, in differentiation of embryonic stem cells into the cells of the neural lineage. Embryonic stem (ES) cells overexpressing HPSE (Hpse-Tg) proliferated more rapidly than WT ES cells in culture and formed larger teratomas in vivo. In addition, differentiating Hpse-Tg ES cells also had a higher growth rate, and overexpression of HPSE in NSPCs enhanced Erk and Akt phosphorylation. Employing a two-step, monolayer differentiation, we observed an increase in HPSE as wild-type (WT) ES cells differentiated into neural stem and progenitor cells followed by down-regulation of HPSE as these NSPCs differentiated into mature cells of the neural lineage. Furthermore, NSPCs overexpressing HPSE gave rise to more oligodendrocytes than WT cultures, with a concomitant reduction in the number of neurons. Our present findings emphasize the importance of HS, in neural differentiation and suggest that by regulating the availability of growth factors and, or other macromolecules, HPSE promotes differentiation into oligodendrocytes. Copyright © 2016 Elsevier B.V. All rights reserved.

  9. Biphasic influence of Miz1 on neural crest development by regulating cell survival and apical adhesion complex formation in the developing neural tube

    PubMed Central

    Kerosuo, Laura; Bronner, Marianne E.

    2014-01-01

    Myc interacting zinc finger protein-1 (Miz1) is a transcription factor known to regulate cell cycle– and cell adhesion–related genes in cancer. Here we show that Miz1 also plays a critical role in neural crest development. In the chick, Miz1 is expressed throughout the neural plate and closing neural tube. Its morpholino-mediated knockdown affects neural crest precursor survival, leading to reduction of neural plate border and neural crest specifier genes Msx-1, Pax7, FoxD3, and Sox10. Of interest, Miz1 loss also causes marked reduction of adhesion molecules (N-cadherin, cadherin6B, and α1-catenin) with a concomitant increase of E-cadherin in the neural folds, likely leading to delayed and decreased neural crest emigration. Conversely, Miz1 overexpression results in up-regulation of cadherin6B and FoxD3 expression in the neural folds/neural tube, leading to premature neural crest emigration and increased number of migratory crest cells. Although Miz1 loss effects cell survival and proliferation throughout the neural plate, the neural progenitor marker Sox2 was unaffected, suggesting a neural crest–selective effect. The results suggest that Miz1 is important not only for survival of neural crest precursors, but also for maintenance of integrity of the neural folds and tube, via correct formation of the apical adhesion complex therein. PMID:24307680

  10. Amplification of neural stem cell proliferation by intermediate progenitor cells in Drosophila brain development.

    PubMed

    Bello, Bruno C; Izergina, Natalya; Caussinus, Emmanuel; Reichert, Heinrich

    2008-02-19

    In the mammalian brain, neural stem cells divide asymmetrically and often amplify the number of progeny they generate via symmetrically dividing intermediate progenitors. Here we investigate whether specific neural stem cell-like neuroblasts in the brain of Drosophila might also amplify neuronal proliferation by generating symmetrically dividing intermediate progenitors. Cell lineage-tracing and genetic marker analysis show that remarkably large neuroblast lineages exist in the dorsomedial larval brain of Drosophila. These lineages are generated by brain neuroblasts that divide asymmetrically to self renew but, unlike other brain neuroblasts, do not segregate the differentiating cell fate determinant Prospero to their smaller daughter cells. These daughter cells continue to express neuroblast-specific molecular markers and divide repeatedly to produce neural progeny, demonstrating that they are proliferating intermediate progenitors. The proliferative divisions of these intermediate progenitors have novel cellular and molecular features; they are morphologically symmetrical, but molecularly asymmetrical in that key differentiating cell fate determinants are segregated into only one of the two daughter cells. Our findings provide cellular and molecular evidence for a new mode of neurogenesis in the larval brain of Drosophila that involves the amplification of neuroblast proliferation through intermediate progenitors. This type of neurogenesis bears remarkable similarities to neurogenesis in the mammalian brain, where neural stem cells as primary progenitors amplify the number of progeny they generate through generation of secondary progenitors. This suggests that key aspects of neural stem cell biology might be conserved in brain development of insects and mammals.

  11. Neural Stem Cells and Fetal-Onset Hydrocephalus.

    PubMed

    Rodríguez, Esteban M; Guerra, María M

    2017-01-27

    Fetal-onset hydrocephalus is not only a disorder of cerebrospinal fluid (CSF) dynamics, but also a brain disorder. How can we explain the inborn and, so far, irreparable neurological impairment in children born with hydrocephalus? We hypothesize that a cell junction pathology of neural stem cells (NSC) leads to two inseparable phenomena: hydrocephalus and abnormal neurogenesis. All neurons, glial cells, and ependymal cells of the mammalian central nervous system originate from the NSC forming the ventricular zone (VZ) and the neural progenitor cells (NPC) forming the subventricular zone. Several genetic mutations and certain foreign signals all convey into a final common pathway leading to cell junction pathology of NSC and VZ disruption. VZ disruption follows a temporal and spatial pattern; it leads to aqueduct obliteration and hydrocephalus in the cerebral aqueduct, while it results in abnormal neurogenesis in the telencephalon. The disrupted NSC and NPC are released into the CSF and may transform into neurospheres displaying a junctional pathology similar to that of NSC of the disrupted VZ. These cells can then be utilized to investigate molecular alterations underlying the disease and open an avenue into possible NSC therapy.

  12. Control of neural stem cell adhesion and density by an electronic polymer surface switch.

    PubMed

    Saltó, Carmen; Saindon, Emilien; Bolin, Maria; Kanciurzewska, Anna; Fahlman, Mats; Jager, Edwin W H; Tengvall, Pentti; Arenas, Ernest; Berggren, Magnus

    2008-12-16

    Adhesion is an essential parameter for stem cells. It regulates the overall cell density along the carrying surface, which further dictates the differentiation scheme of stem cells toward a more matured and specified population as well as tissue. Electronic control of the seeding density of neural stem cells (c17.2) is here reported. Thin electrode films of poly(3,4-ethylenedioxythiophene) (PEDOT):Tosylate were manufactured along the floor of cell growth dishes. As the oxidation state of the conjugated polymer electrodes was controlled, the seeding density could be varied by a factor of 2. Along the oxidized PEDOT:Tosylate-electrodes, a relatively lower density of, and less tightly bonded, human serum albumin (HSA) was observed as compared to reduced electrodes. We found that this favors adhesion of the specific stem cells studied. Surface analysis experiments, such as photoelectron spectroscopy, and water contact angle measurements, were carried out to investigate the mechanisms responsible for the electronic control of the seeding density of the c17.2 neural stem cells. Further, our findings may provide an opening for electronic control of stem cell differentiation.

  13. Aging differentially affects male and female neural stem cell neurogenic properties

    PubMed Central

    Waldron, Jay; McCourty, Althea; Lecanu, Laurent

    2010-01-01

    Purpose Neural stem cell transplantation as a brain repair strategy is a very promising technology. However, despite many attempts, the clinical success remains very deceiving. Despite clear evidence that sexual dimorphism rules many aspects of human biology, the occurrence of a sex difference in neural stem cell biology is largely understudied. Herein, we propose to determine whether gender is a dimension that drives the fate of neural stem cells through aging. Should it occur, we believe that neural stem cell sexual dimorphism and its variation during aging should be taken into account to refine clinical approaches of brain repair strategies. Methods Neural stem cells were isolated from the subventricular zone of three- and 20-month-old male and female Long-Evans rats. Expression of the estrogen receptors, ERα and ERβ, progesterone receptor, androgen receptor, and glucocorticoid receptor was analyzed and quantified by Western blotting on undifferentiated neural stem cells. A second set of neural stem cells was treated with retinoic acid to trigger differentiation, and the expression of neuronal, astroglial, and oligodendroglial markers was determined using Western blotting. Conclusion We provided in vitro evidence that the fate of neural stem cells is affected by sex and aging. Indeed, young male neural stem cells mainly expressed markers of neuronal and oligodendroglial fate, whereas young female neural stem cells underwent differentiation towards an astroglial phenotype. Aging resulted in a lessened capacity to express neuron and astrocyte markers. Undifferentiated neural stem cells displayed sexual dimorphism in the expression of steroid receptors, in particular ERα and ERβ, and the expression level of several steroid receptors increased during aging. Such sexual dimorphism might explain, at least in part, the sex difference in neural fate we observed in young and old neural stem cells. These results suggest that sex and aging are two factors to be taken

  14. GDNF is a chemoattractant for enteric neural cells.

    PubMed

    Young, H M; Hearn, C J; Farlie, P G; Canty, A J; Thomas, P Q; Newgreen, D F

    2001-01-15

    In situ hybridization revealed that GDNF mRNA in the mid- and hindgut mesenchyme of embryonic mice was minimal at E10.5 but was rapidly elevated at all gut regions after E11, but with a slight delay (0.5 days) in the hindgut. GDNF mRNA expression was minimal in the mesentery and in the pharyngeal and pelvic mesenchyme adjacent to the gut. To examine the effect of GDNF on enteric neural crest-derived cells, segments of E11.5 mouse hindgut containing crest-derived cells only at the rostral ends were attached to filter paper supports and grown in catenary organ culture. With GDNF (100 ng/ml) in the culture medium, threefold fewer neurons developed in the gut explants and fivefold more neurons were present on the filter paper outside the gut explants, compared to controls. Thus, in controls, crest-derived cells colonized the entire explant and differentiated into neurons, whereas in the presence of exogenous GDNF, most crest-derived cells migrated out of the gut explant. This is consistent with GDNF acting as a chemoattractant. To test this idea, explants of esophagus, midgut, superior cervical ganglia, paravertebral sympathetic chain ganglia, or dorsal root ganglia from E11.5-E12.5 mice were grown on collagen gels with a GDNF-impregnated agarose bead on one side and a control bead on the opposite side. Migrating neural cells and neurites from the esophagus and midgut accumulated around the GDNF-impregnated beads, but neural cells in other tissues showed little or no chemotactic response to GDNF, although all showed GDNF-receptor (Ret and GFRalpha1) immunoreactivity. We conclude that GDNF may promote the migration of crest cells throughout the gastrointestinal tract, prevent them from straying out of the gut (into the mesentery and pharyngeal and pelvic tissues), and promote directed axon outgrowth. Copyright 2001 Academic Press.

  15. Cell delamination in the mesencephalic neural fold and its implication for the origin of ectomesenchyme

    PubMed Central

    Lee, Raymond Teck Ho; Nagai, Hiroki; Nakaya, Yukiko; Sheng, Guojun; Trainor, Paul A.; Weston, James A.; Thiery, Jean Paul

    2013-01-01

    The neural crest is a transient structure unique to vertebrate embryos that gives rise to multiple lineages along the rostrocaudal axis. In cranial regions, neural crest cells are thought to differentiate into chondrocytes, osteocytes, pericytes and stromal cells, which are collectively termed ectomesenchyme derivatives, as well as pigment and neuronal derivatives. There is still no consensus as to whether the neural crest can be classified as a homogenous multipotent population of cells. This unresolved controversy has important implications for the formation of ectomesenchyme and for confirmation of whether the neural fold is compartmentalized into distinct domains, each with a different repertoire of derivatives. Here we report in mouse and chicken that cells in the neural fold delaminate over an extended period from different regions of the cranial neural fold to give rise to cells with distinct fates. Importantly, cells that give rise to ectomesenchyme undergo epithelial-mesenchymal transition from a lateral neural fold domain that does not express definitive neural markers, such as Sox1 and N-cadherin. Additionally, the inference that cells originating from the cranial neural ectoderm have a common origin and cell fate with trunk neural crest cells prompted us to revisit the issue of what defines the neural crest and the origin of the ectomesenchyme. PMID:24198279

  16. Neural Correlates of Posttraumatic Growth after Severe Motor Vehicle Accidents

    ERIC Educational Resources Information Center

    Rabe, Sirko; Zollner, Tanja; Maercker, Andreas; Karl, Anke

    2006-01-01

    Frontal brain asymmetry has been associated with emotion- and motivation-related constructs. The authors examined the relationship between frontal brain asymmetry and subjective perception of posttraumatic growth (PTG) after severe motor vehicle accidents (MVAs). Eighty-two survivors of MVAs completed self-report measures of PTG, trait and state…

  17. Prosencephalic neural folds give rise to neural crest cells in the Australian lungfish, Neoceratodus forsteri.

    PubMed

    Kundrát, Martin; Joss, Jean M P; Olsson, Lennart

    2009-03-15

    Here we present a fate map of the prosencephalic neural fold (PNF) for the Australian lungfish. The experimental procedures were carried out on lungfish embryos at Kemp's stage 24 using three different approaches. First, either medial PNF (MPNF) or lateral PNF (LPNF) were ablated and the embryos cultured until they reached Kemp's stage 42 and 44. Ablation of the LPNF provided phenotypes with arrested development of the eye, reduction of periocular pigmentation, frontonasal deformity, and a slightly reduced olfactory organ, whereas the MPNF-ablated phenotypes resulted in arrested development of the cornea and frontonasal deformity. Second, we labeled the mid-axial level of the PNF with vital DiI and traced the migration of labeled cells following culture to Kemp's stage 33. Labeled PNF-derived cells populated a basal layer of the olfactory placode, migrated into the frontonasal region, the antero-dorsal periocular quadrant, and also terminated at positions where the forebrain meninges form at later stages. Third, we examined HNK-1 immunoreactivity in the forebrain-related region. We conclude that in the Australian lungfish: (1) LPNF-derived neuroepithelium gives rise to the basal layer and contributes to the apical layer of the olfactory placode; (2) PNF-derived NC cells appear to give rise to meningeal, periocular, and frontonasal ectomesenchyme and likely infiltrate the olfactory placode as developmental precusors of the terminal nerve; (3) HNK-1 epitope is temporarily expressed in cells of the neural tube, NC cells, and neurogenic placodal cells. Our experiments have provided the first evidence for a premandibular NC stream (sensu Kundrát, 2008) in a fish.

  18. Proliferation and cilia dynamics in neural stem cells prospectively isolated from the SEZ

    NASA Astrophysics Data System (ADS)

    Khatri, Priti; Obernier, Kirsten; Simeonova, Ina K.; Hellwig, Andrea; Hölzl-Wenig, Gabriele; Mandl, Claudia; Scholl, Catharina; Wölfl, Stefan; Winkler, Johannes; Gaspar, John A.; Sachinidis, Agapios; Ciccolini, Francesca

    2014-01-01

    Neural stem cells (NSCs) generate new neurons in vivo and in vitro throughout adulthood and therefore are physiologically and clinically relevant. Unveiling the mechanisms regulating the lineage progression from NSCs to newborn neurons is critical for the transition from basic research to clinical application. However, the direct analysis of NSCs and their progeny is still elusive due to the problematic identification of the cells. We here describe the isolation of highly purified genetically unaltered NSCs and transit-amplifying precursors (TAPs) from the adult subependymal zone (SEZ). Using this approach we show that a primary cilium and high levels of epidermal growth factor receptor (EGFR) at the cell membrane characterize quiescent and cycling NSCs, respectively. However, we also observed non-ciliated quiescent NSCs and NSCs progressing into the cell cycle without up-regulating EGFR expression. Thus, the existence of NSCs displaying distinct molecular and structural conformations provides more flexibility to the regulation of quiescence and cell cycle progression.

  19. Insulin-Like Growth Factor Receptor Signaling is Necessary for Epidermal Growth Factor Mediated Proliferation of SVZ Neural Precursors in vitro Following Neonatal Hypoxia–Ischemia

    PubMed Central

    Alagappan, Dhivyaa; Ziegler, Amber N.; Chidambaram, Shravanthi; Min, Jungsoo; Wood, Teresa L.; Levison, Steven W.

    2014-01-01

    In this study, we assessed the importance of insulin-like growth factor (IGF) and epidermal growth factor (EGF) receptor co-signaling for rat neural precursor (NP) cell proliferation and self-renewal in the context of a developmental brain injury that is associated with cerebral palsy. Consistent with previous studies, we found that there is an increase in the in vitro growth of subventricular zone NPs isolated acutely after cerebral hypoxia–ischemia; however, when cultured in medium that is insufficient to stimulate the IGF type 1 receptor, neurosphere formation and the proliferative capacity of those NPs was severely curtailed. This reduced growth capacity could not be attributed simply to failure to survive. The growth and self-renewal of the NPs could be restored by addition of both IGF-I and IGF-II. Since the size of the neurosphere is predominantly due to cell proliferation we hypothesized that the IGFs were regulating progression through the cell cycle. Analyses of cell cycle progression revealed that IGF-1R activation together with EGFR co-signaling decreased the percentage of cells in G1 and enhanced cell progression into S and G2. This was accompanied by increases in expression of cyclin D1, phosphorylated histone 3, and phosphorylated Rb. Based on these data, we conclude that coordinate signaling between the EGF receptor and the IGF type 1 receptor is necessary for the normal proliferation of NPs as well as for their reactive expansion after injury. These data indicate that manipulations that maintain or amplify IGF signaling in the brain during recovery from developmental brain injuries will enhance the production of new brain cells to improve neurological function in children who are at risk for developing cerebral palsy. PMID:24904523

  20. Generating trunk neural crest from human pluripotent stem cells.

    PubMed

    Huang, Miller; Miller, Matthew L; McHenry, Lauren K; Zheng, Tina; Zhen, Qiqi; Ilkhanizadeh, Shirin; Conklin, Bruce R; Bronner, Marianne E; Weiss, William A

    2016-01-27

    Neural crest cells (NCC) are stem cells that generate different lineages, including neuroendocrine, melanocytic, cartilage, and bone. The differentiation potential of NCC varies according to the level from which cells emerge along the neural tube. For example, only anterior "cranial" NCC form craniofacial bone, whereas solely posterior "trunk" NCC contribute to sympathoadrenal cells. Importantly, the isolation of human fetal NCC carries ethical and scientific challenges, as NCC induction typically occur before pregnancy is detectable. As a result, current knowledge of NCC biology derives primarily from non-human organisms. Important differences between human and non-human NCC, such as expression of HNK1 in human but not mouse NCC, suggest a need to study human NCC directly. Here, we demonstrate that current protocols to differentiate human pluripotent stem cells (PSC) to NCC are biased toward cranial NCC. Addition of retinoic acid drove trunk-related markers and HOX genes characteristic of a posterior identity. Subsequent treatment with bone morphogenetic proteins (BMPs) enhanced differentiation to sympathoadrenal cells. Our approach provides methodology for detailed studies of human NCC, and clarifies roles for retinoids and BMPs in the differentiation of human PSC to trunk NCC and to sympathoadrenal lineages.

  1. Generating trunk neural crest from human pluripotent stem cells

    PubMed Central

    Huang, Miller; Miller, Matthew L.; McHenry, Lauren K.; Zheng, Tina; Zhen, Qiqi; Ilkhanizadeh, Shirin; Conklin, Bruce R.; Bronner, Marianne E.; Weiss, William A.

    2016-01-01

    Neural crest cells (NCC) are stem cells that generate different lineages, including neuroendocrine, melanocytic, cartilage, and bone. The differentiation potential of NCC varies according to the level from which cells emerge along the neural tube. For example, only anterior “cranial” NCC form craniofacial bone, whereas solely posterior “trunk” NCC contribute to sympathoadrenal cells. Importantly, the isolation of human fetal NCC carries ethical and scientific challenges, as NCC induction typically occur before pregnancy is detectable. As a result, current knowledge of NCC biology derives primarily from non-human organisms. Important differences between human and non-human NCC, such as expression of HNK1 in human but not mouse NCC, suggest a need to study human NCC directly. Here, we demonstrate that current protocols to differentiate human pluripotent stem cells (PSC) to NCC are biased toward cranial NCC. Addition of retinoic acid drove trunk-related markers and HOX genes characteristic of a posterior identity. Subsequent treatment with bone morphogenetic proteins (BMPs) enhanced differentiation to sympathoadrenal cells. Our approach provides methodology for detailed studies of human NCC, and clarifies roles for retinoids and BMPs in the differentiation of human PSC to trunk NCC and to sympathoadrenal lineages. PMID:26812940

  2. Live Imaging of Adult Neural Stem Cells in Rodents

    PubMed Central

    Ortega, Felipe; Costa, Marcos R.

    2016-01-01

    The generation of cells of the neural lineage within the brain is not restricted to early development. New neurons, oligodendrocytes, and astrocytes are produced in the adult brain throughout the entire murine life. However, despite the extensive research performed in the field of adult neurogenesis during the past years, fundamental questions regarding the cell biology of adult neural stem cells (aNSCs) remain to be uncovered. For instance, it is crucial to elucidate whether a single aNSC is capable of differentiating into all three different macroglial cell types in vivo or these distinct progenies constitute entirely separate lineages. Similarly, the cell cycle length, the time and mode of division (symmetric vs. asymmetric) that these cells undergo within their lineage progression are interesting questions under current investigation. In this sense, live imaging constitutes a valuable ally in the search of reliable answers to the previous questions. In spite of the current limitations of technology new approaches are being developed and outstanding amount of knowledge is being piled up providing interesting insights in the behavior of aNSCs. Here, we will review the state of the art of live imaging as well as the alternative models that currently offer new answers to critical questions. PMID:27013941

  3. Neural stem cell-based treatment for neurodegenerative diseases.

    PubMed

    Kim, Seung U; Lee, Hong J; Kim, Yun B

    2013-10-01

    Human neurodegenerative diseases such as Parkinson's disease (PD), Huntington's disease (HD), amyotrophic lateral sclerosis (ALS) and Alzheimer's disease (AD) are caused by a loss of neurons and glia in the brain or spinal cord. Neurons and glial cells have successfully been generated from stem cells such as embryonic stem cells (ESCs), mesenchymal stem cells (MSCs) and neural stem cells (NSCs), and stem cell-based cell therapies for neurodegenerative diseases have been developed. A recent advance in generation of a new class of pluripotent stem cells, induced pluripotent stem cells (iPSCs), derived from patients' own skin fibroblasts, opens doors for a totally new field of personalized medicine. Transplantation of NSCs, neurons or glia generated from stem cells in animal models of neurodegenerative diseases, including PD, HD, ALS and AD, demonstrates clinical improvement and also life extension of these animals. Additional therapeutic benefits in these animals can be provided by stem cell-mediated gene transfer of therapeutic genes such as neurotrophic factors and enzymes. Although further research is still needed, cell and gene therapy based on stem cells, particularly using neurons and glia derived from iPSCs, ESCs or NSCs, will become a routine treatment for patients suffering from neurodegenerative diseases and also stroke and spinal cord injury.

  4. Functional identification of neural stem cell-derived oligodendrocytes.

    PubMed

    Grade, Sofia; Agasse, Fabienne; Bernardino, Liliana; Malva, João O

    2012-01-01

    Directing neural stem cells (NSCs) differentiation towards oligodendroglial cell lineage is a crucial step in the endeavor of developing cell replacement-based therapies for demyelinating diseases. Evaluation of NSCs differentiation is mostly performed by methodologies that use fixed cells, like immunocytochemistry, or lysates, like Western blot. On the other hand, electrophysiology allows differentiation studies on living cells, but it is highly time-consuming and endowed with important limitations concerning population studies. Herein, we describe a functional method, based on single cell calcium imaging, which accurately and rapidly distinguishes cell types among NSCs progeny, in living cultures prepared from the major reservoir of NSCs in the postnatal mouse brain, the subventricular zone (SVZ). Indeed, by applying a rational sequence of three stimuli-KCl, histamine, and thrombin-to the heterogeneous SVZ cell population, one can identify each cell phenotype according to its unique calcium signature. Mature oligodendrocytes, the myelin-forming cells of the central nervous system, are the thrombin-responsive cells in SVZ cell culture and display no intracellular calcium increase upon KCl or histamine perfusion. On the other hand, KCl and histamine stimulate neurons and immature cells, respectively. The method described in this chapter is a valuable tool to identify novel pro-oligodendrogenic compounds, which may play an important role in the design of future treatments for demyelinating disorders such as multiple sclerosis.

  5. Clonal amniotic fluid-derived stem cells express characteristics of both mesenchymal and neural stem cells.

    PubMed

    Tsai, Ming-Song; Hwang, Shiaw-Min; Tsai, Yieh-Loong; Cheng, Fu-Chou; Lee, Jia-Ling; Chang, Yu-Jen

    2006-03-01

    Recent evidence has shown that amniotic fluid may be a novel source of fetal stem cells for therapeutic transplantation. We previously developed a two-stage culture protocol to isolate a population of amniotic fluid-derived mesenchymal stem cells (AFMSCs) from second-trimester amniocentesis. AFMSCs maintain the capacity to differentiate into multiple mesenchymal lineages and neuron-like cells. It is unclear whether amniotic fluid contains heterogeneous populations of stem cells or a subpopulation of primitive stem cells that are similar to marrow stromal cells showing the behavior of neural progenitors. In this study, we showed a subpopulation of amniotic fluid-derived stem cells (AF-SCs) at the single-cell level by limiting dilution. We found that NANOG- and POU5F1 (also known as OCT4)-expressing cells still existed in the expanded single cell-derived AF-SCs. Aside from the common mesenchymal characteristics, these clonal AF-SCs also exhibit multiple phenotypes of neural-derived cells such as NES, TUBB3, NEFH, NEUNA60, GALC, and GFAP expressions both before and after neural induction. Most importantly, HPLC analysis showed the evidence of dopamine release in the extract of dopaminergic-induced clonal AF-SCs. The results of this study suggest that besides being an easily accessible and expandable source of fetal stem cells, amniotic fluid will provide a promising source of neural progenitor cells that may be used in future cellular therapies for neurodegenerative diseases and nervous system injuries.

  6. Cell surface beta 1,4-galactosyltransferase functions during neural crest cell migration and neurulation in vivo

    PubMed Central

    1992-01-01

    Mesenchymal cell migration and neurite outgrowth are mediated in part by binding of cell surface beta 1,4-galactosyltransferase (GalTase) to N-linked oligosaccharides within the E8 domain of laminin. In this study, we determined whether cell surface GalTase functions during neural crest cell migration and neural development in vivo using antibodies raised against affinity-purified chicken serum GalTase. The antibodies specifically recognized two embryonic proteins of 77 and 67 kD, both of which express GalTase activity. The antibodies also immunoprecipitated and inhibited chick embryo GalTase activity, and inhibited neural crest cell migration on laminin matrices in vitro. Anti-GalTase antibodies were microinjected into the head mesenchyme of stage 7-9 chick embryos or cranial to Henson's node of stage 6 embryos. Anti-avian GalTase IgG decreased cranial neural crest cell migration on the injected side but did not cross the embryonic midline and did not affect neural crest cell migration on the uninjected side. Anti-avian GalTase Fab crossed the embryonic midline and perturbed cranial neural crest cell migration throughout the head. Neural fold elevation and neural tube closure were also disrupted by Fab fragments. Cell surface GalTase was localized to migrating neural crest cells and to the basal surfaces of neural epithelia by indirect immunofluorescence, whereas GalTase was undetectable on neural crest cells prior to migration. These results suggest that, during early embryogenesis, cell surface GalTase participates during neural crest cell migration, perhaps by interacting with laminin, a major component of the basal lamina. Cell surface GalTase also appears to play a role in neural tube formation, possibly by mediating neural epithelial adhesion to the underlying basal lamina. PMID:1560031

  7. The role of the JAK-STAT pathway in neural stem cells, neural progenitor cells and reactive astrocytes after spinal cord injury.

    PubMed

    Wang, Tianyi; Yuan, Wenqi; Liu, Yong; Zhang, Yanjun; Wang, Zhijie; Zhou, Xianhu; Ning, Guangzhi; Zhang, Liang; Yao, Liwei; Feng, Shiqing; Kong, Xiaohong

    2015-03-01

    Patients with spinal cord injuries can develop severe neurological damage and dysfunction, which is not only induced by primary but also by secondary injuries. As an evolutionarily conserved pathway of eukaryotes, the JAK-STAT pathway is associated with cell growth, survival, development and differentiation; activation of the JAK-STAT pathway has been previously reported in central nervous system injury. The JAK-STAT pathway is directly associated with neurogenesis and glia scar formation in the injury region. Following injury of the axon, the overexpression and activation of STAT3 is exhibited specifically in protecting neurons. To investigate the role of the JAK-STAT pathway in neuroprotection, we summarized the effect of JAK-STAT pathway in the following three sections: Firstly, the modulation of JAK-STAT pathway in proliferation and differentiation of neural stem cells and neural progenitor cells is discussed; secondly, the time-dependent effect of JAK-STAT pathway in reactive astrocytes to reveal their capability of neuroprotection is revealed and lastly, we focus on how the astrocyte-secretory polypeptides (astrocyte-derived cytokines and trophic factors) accomplish neuroprotection via the JAK-STAT pathway.

  8. Interplay between cell growth and cell cycle in plants.

    PubMed

    Sablowski, Robert; Carnier Dornelas, Marcelo

    2014-06-01

    The growth of organs and whole plants depends on both cell growth and cell-cycle progression, but the interaction between both processes is poorly understood. In plants, the balance between growth and cell-cycle progression requires coordinated regulation of four different processes: macromolecular synthesis (cytoplasmic growth), turgor-driven cell-wall extension, mitotic cycle, and endocycle. Potential feedbacks between these processes include a cell-size checkpoint operating before DNA synthesis and a link between DNA contents and maximum cell size. In addition, key intercellular signals and growth regulatory genes appear to target at the same time cell-cycle and cell-growth functions. For example, auxin, gibberellin, and brassinosteroid all have parallel links to cell-cycle progression (through S-phase Cyclin D-CDK and the anaphase-promoting complex) and cell-wall functions (through cell-wall extensibility or microtubule dynamics). Another intercellular signal mediated by microtubule dynamics is the mechanical stress caused by growth of interconnected cells. Superimposed on developmental controls, sugar signalling through the TOR pathway has recently emerged as a central control point linking cytoplasmic growth, cell-cycle and cell-wall functions. Recent progress in quantitative imaging and computational modelling will facilitate analysis of the multiple interconnections between plant cell growth and cell cycle and ultimately will be required for the predictive manipulation of plant growth.

  9. Lack of endothelial cell survivin causes embryonic defects in angiogenesis, cardiogenesis, and neural tube closure.

    PubMed

    Zwerts, Femke; Lupu, Florea; De Vriese, Astrid; Pollefeyt, Saskia; Moons, Lieve; Altura, Rachel A; Jiang, Yuying; Maxwell, Patrick H; Hill, Peter; Oh, Hideyasu; Rieker, Claus; Collen, Désiré; Conway, Simon J; Conway, Edward M

    2007-06-01

    We explored the physiologic role of endothelial cell apoptosis during development by generating mouse embryos lacking the inhibitor of apoptosis protein (IAP) survivin in endothelium. This was accomplished by intercrossing survivin(lox/lox) mice with mice expressing cre recombinase under the control of the endothelial cell specific tie1 promoter (tie1-cre mice). Lack of endothelial cell survivin resulted in embryonic lethality. Mutant embryos had prominent and diffuse hemorrhages from embryonic day 9.5 (E9.5) and died before E13.5. Heart development was strikingly abnormal. Survivin-null endocardial lineage cells could not support normal epithelial-mesenchymal transformation (EMT), resulting in hypoplastic endocardial cushions and in utero heart failure. In addition, 30% of mutant embryos had neural tube closure defects (NTDs) that were not caused by bleeding or growth retardation, but were likely due to alterations in the release of soluble factors from endothelial cells that otherwise support neural stem cell proliferation and neurulation. Thus, regulation of endothelial cell survival, and maintenance of vascular integrity by survivin are crucial for normal embryonic angiogenesis, cardiogenesis, and neurogenesis.

  10. Encapsulated neural stem cell neuronal differentiation in fluorinated methacrylamide chitosan hydrogels.

    PubMed

    Li, Hang; Wijekoon, Asanka; Leipzig, Nic D

    2014-07-01

    Neural stem/progenitor cells (NSPCs) are able to differentiate into the primary cell types (neurons, oligodendrocytes and astrocytes) of the adult nervous system. This attractive property of NSPCs offers a potential solution for neural regeneration. 3D implantable scaffolds should mimic the microstructure and dynamic properties found in vivo, enabling the natural exchange of oxygen, nutrients, and growth factors for cell survival and differentiation. We have previously reported a new class of materials consisting of perfluorocarbons (PFCs) conjugated to methacrylamide chitosan (MAC), which possess the ability to repeatedly take-up and release oxygen at beneficial levels for favorable cell metabolism and proliferation. In this study, the neuronal differentiation responses of NSPCs to fluorinated methacrylamide chitosan (MACF) hydrogels were studied for 8 days. Two treatments, with oxygen reloading or without oxygen reloading, were performed during culture. Oxygen concentration distributions within cell-seeded MACF hydrogels were found to have higher concentrations of oxygen at the edge of the hydrogels and less severe drops in O2 gradient as compared with MAC hydrogel controls. Total cell number was enhanced in MACF hydrogels as the number of conjugated fluorines via PFC substitution increased. Additionally, all MACF hydrogels supported significantly more cells than MAC controls (p < 0.001). At day 8, MACF hydrogels displayed significantly greater neuronal differentiation than MAC controls (p = 0.001), and among MACF groups methacrylamide chitosan with 15 fluorines per addition (MAC(Ali15)F) demonstrated the best ability to promote NSPC differentiation.

  11. Generation of Neural Crest-Like Cells From Human Periodontal Ligament Cell-Derived Induced Pluripotent Stem Cells.

    PubMed

    Tomokiyo, Atsushi; Hynes, Kim; Ng, Jia; Menicanin, Danijela; Camp, Esther; Arthur, Agnes; Gronthos, Stan; Mark Bartold, Peter

    2017-02-01

    Neural crest cells (NCC) hold great promise for tissue engineering, however the inability to easily obtain large numbers of NCC is a major factor limiting their use in studies of regenerative medicine. Induced pluripotent stem cells (iPSC) are emerging as a novel candidate that could provide an unlimited source of NCC. In the present study, we examined the potential of neural crest tissue-derived periodontal ligament (PDL) iPSC to differentiate into neural crest-like cells (NCLC) relative to iPSC generated from a non-neural crest derived tissue, foreskin fibroblasts (FF). We detected high HNK1 expression during the differentiation of PDL and FF iPSC into NCLC as a marker for enriching for a population of cells with NCC characteristics. We isolated PDL iPSC- and FF iPSC-derived NCLC, which highly expressed HNK1. A high proportion of the HNK1-positive cell populations generated, expressed the MSC markers, whilst very few cells expressed the pluripotency markers or the hematopoietic markers. The PDL and FF HNK1-positive populations gave rise to smooth muscle, neural, glial, osteoblastic and adipocytic like cells and exhibited higher expression of smooth muscle, neural, and glial cell-associated markers than the PDL and FF HNK1-negative populations. Interestingly, the HNK1-positive cells derived from the PDL-iPSC exhibited a greater ability to differentiate into smooth muscle, neural, glial cells and adipocytes, than the HNK1-positive cells derived from the FF-iPSC. Our work suggests that HNK1-enriched NCLC from neural crest tissue-derived iPSC more closely resemble the phenotypic and functional hallmarks of NCC compared to the HNK1-low population and non-neural crest iPSC-derived NCLC. J. Cell. Physiol. 232: 402-416, 2017. © 2016 Wiley Periodicals, Inc.

  12. An intrinsic mechanism controls reactivation of neural stem cells by spindle matrix proteins.

    PubMed

    Li, Song; Koe, Chwee Tat; Tay, Su Ting; Tan, Angie Lay Keng; Zhang, Shenli; Zhang, Yingjie; Tan, Patrick; Sung, Wing-Kin; Wang, Hongyan

    2017-07-25

    The switch between quiescence and proliferation is central for neurogenesis and its alteration is linked to neurodevelopmental disorders such as microcephaly. However, intrinsic mechanisms that reactivate Drosophila larval neural stem cells (NSCs) to exit from quiescence are not well established. Here we show that the spindle matrix complex containing Chromator (Chro) functions as a key intrinsic regulator of NSC reactivation downstream of extrinsic insulin/insulin-like growth factor signalling. Chro also prevents NSCs from ire-entering quiescence at later stages. NSC-specific in vivo profiling has dentified many downstream targets of Chro, including a temporal transcription factor Grainy head (Grh) and a neural stem cell quiescence-inducing factor Prospero (Pros). We show that spindle matrix proteins promote the expression of Grh and repress that of Pros in NSCs to govern their reactivation. Our data demonstrate that nuclear Chro critically regulates gene expression in NSCs at the transition from quiescence to proliferation.The spindle matrix proteins, including Chro, are known to regulate mitotic spindle assembly in the cytoplasm. Here the authors show that in Drosophila larval brain, Chro promotes neural stem cell (NSC) reactivation and prevents activated NSCs from entering quiescence, and that Chro carries out such a role by regulating the expression of key transcription factors in the nucleus.

  13. Effect of polyvinylidene fluoride electrospun fiber orientation on neural stem cell differentiation.

    PubMed

    Lins, Luanda C; Wianny, Florence; Livi, Sebastien; Dehay, Colette; Duchet-Rumeau, Jannick; Gérard, Jean-François

    2016-08-29

    Electrospun polymer piezoelectric fibers can be used in neural tissue engineering (NTE) to mimic the physical, biological, and material properties of the native extracellular matrix. In this work, we have developed scaffolds based on polymer fiber architectures for application in NTE. To study the role of such three-dimensional scaffolds, a rotating drum collector was used for electrospinning poly(vinylidene) fluoride (PVDF) polymer at various rotation speeds. The morphology, orientation, polymorphism, as well as the mechanical behavior of the nonaligned and aligned fiber-based architectures were characterized. We have demonstrated that the jet flow and the electrostatic forces generated by electrospinning of PVDF induced local conformation changes which promote the generation of the β-phase. Fiber anisotropy could be a critical feature for the design of suitable scaffolds for NTEs. We thus assessed the impact of PVDF fiber alignment on the behavior of monkey neural stem cells (NSCs). NSCs were seeded on nonaligned and aligned scaffolds and their morphology, adhesion, and differentiation capacities into the neuronal and glial pathways were studied using microscopic techniques. Significant changes in the growth and differentiation capacities of NSCs into neuronal and glial cells as a function of the fiber alignment were evidenced. These results demonstrate that PVDF scaffolds may serve as instructive scaffolds for NSC survival and differentiation, and may be valuable tools for the development of cell- and scaffold-based strategies for neural repair. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2016. © 2016 Wiley Periodicals, Inc.

  14. Enhanced proliferation of PC12 neural cells on untreated, nanotextured glass coverslips

    NASA Astrophysics Data System (ADS)

    Islam, Muhymin; Atmaramani, Rahul; Mukherjee, Siddhartha; Ghosh, Santaneel; Iqbal, Samir M.

    2016-10-01

    Traumatic injury to the central nervous system is a significant health problem. There is no effective treatment available partly because of the complexity of the system. Implementation of multifunctional micro- and nano-device based combinatorial therapeutics can provide biocompatible and tunable approaches to perform on-demand release of specific drugs. This can help the damaged cells to improve neuronal survival, regeneration of axons, and their reconnection to appropriate targets. Nano-topological features induced rapid cell growth is especially important towards the design of effective platforms to facilitate damaged neural circuit reconstruction. In this study, for the first time, feasibility of neuron-like PC12 cell growth on untreated and easy to prepare nanotextured surfaces has been carried out. The PC12 neuron-like cells were cultured on micro reactive ion etched nanotextured glass coverslips. The effect of nanotextured topology as physical cue for the growth of PC12 cells was observed exclusively, eliminating the possible influence(s) of the enhanced concentration of coated materials on the surface. The cell density was observed to increase by almost 200% on nanotextured coverslips compared to plain coverslips. The morphology study indicated that PC12 cell attachment and growth on the nanotextured substrates did not launch any apoptotic machinery of the cell. Less than 5% cells deformed and depicted condensed nuclei with apoptotic bodies on nanotextured surfaces which is typical for the normal cell handling and culture. Enhanced PC12 cell proliferation by such novel and easy to prepare substrates is not only attractive for neurite outgrowth and guidance, but may be used to increase the affinity of similar cancerous cells (ex: B35 neuroblastoma) and rapid proliferation thereafter—towards the development of combinatorial theranostics to diagnose and treat aggressive cancers like neuroblastoma.

  15. Enhanced proliferation of PC12 neural cells on untreated, nanotextured glass coverslips.

    PubMed

    Islam, Muhymin; Atmaramani, Rahul; Mukherjee, Siddhartha; Ghosh, Santaneel; Iqbal, Samir M

    2016-10-14

    Traumatic injury to the central nervous system is a significant health problem. There is no effective treatment available partly because of the complexity of the system. Implementation of multifunctional micro- and nano-device based combinatorial therapeutics can provide biocompatible and tunable approaches to perform on-demand release of specific drugs. This can help the damaged cells to improve neuronal survival, regeneration of axons, and their reconnection to appropriate targets. Nano-topological features induced rapid cell growth is especially important towards the design of effective platforms to facilitate damaged neural circuit reconstruction. In this study, for the first time, feasibility of neuron-like PC12 cell growth on untreated and easy to prepare nanotextured surfaces has been carried out. The PC12 neuron-like cells were cultured on micro reactive ion etched  nanotextured glass coverslips. The effect of nanotextured topology as physical cue for the growth of PC12 cells was observed exclusively, eliminating the possible influence(s) of the enhanced concentration of coated materials on the surface. The cell density was observed to increase by almost 200% on nanotextured coverslips compared to plain coverslips. The morphology study indicated that PC12 cell attachment and growth on the nanotextured substrates did not launch any apoptotic machinery of the cell. Less than 5% cells deformed and depicted condensed nuclei with apoptotic bodies on nanotextured surfaces which is typical for the normal cell handling and culture. Enhanced PC12 cell proliferation by such novel and easy to prepare substrates is not only attractive for neurite outgrowth and guidance, but may be used to increase the affinity of similar cancerous cells (ex: B35 neuroblastoma) and rapid proliferation thereafter-towards the development of combinatorial theranostics to diagnose and treat aggressive cancers like neuroblastoma.

  16. Partial Restoration of Cardiovascular Function by Embryonic Neural Stem Cell Grafts after Complete Spinal Cord Transection

    PubMed Central

    Hou, Shaoping; Tom, Veronica J.; Graham, Lori

    2013-01-01

    High-level spinal cord injury can lead to cardiovascular dysfunction, including disordered hemodynamics at rest and autonomic dysreflexia during noxious stimulation. To restore supraspinal control of sympathetic preganglionic neurons (SPNs), we grafted embryonic brainstem-derived neural stem cells (BS-NSCs) or spinal cord-derived neural stem cells (SC-NSCs) expressing green fluorescent protein into the T4 complete transection site of adult rats. Animals with injury alone served as controls. Implanting of BS-NSCs but not SC-NSCs resulted in recovery of basal cardiovascular parameters, whereas both cell grafts alleviated autonomic dysreflexia. Subsequent spinal cord retransection above the graft abolished the recovery of basal hemodynamics and reflexic response. BS-NSC graft-derived catecholaminergic and serotonergic neurons showed remarkable long-distance axon growth and topographical innervation of caudal SPNs. Anterograde tracing indicated growth of medullar axons into stem cell grafts and formation of synapses. Thus, grafted embryonic brainstem-derived neurons can act as functional relays to restore supraspinal regulation of denervated SPNs, thereby contributing to cardiovascular functional improvement. PMID:24155317

  17. Expression Profiles of the Nuclear Receptors and Their Transcriptional Coregulators During Differentiation of Neural Stem Cells

    PubMed Central

    Androutsellis-Theotokis, A.; Chrousos, G. P.; McKay, R. D.; DeCherney, A. H.; Kino, T.

    2013-01-01

    Neural stem cells (NSCs) are pluripotent precursors with the ability to proliferate and differentiate into 3 neural cell lineages, neurons, astrocytes and oligodendrocytes. Elucidation of the mechanisms underlying these biologic processes is essential for understanding both physiologic and pathologic neural development and regeneration after injury. Nuclear hormone receptors (NRs) and their transcriptional coregulators also play crucial roles in neural development, functions and fate. To identify key NRs and their transcriptional regulators in NSC differentiation, we examined mRNA expression of 49 NRs and many of their coregulators during differentiation (0–5 days) of mouse embryonic NSCs induced by withdrawal of fibroblast growth factor-2 (FGF2). 37 out of 49 NRs were expressed in NSCs before induction of differentiation, while receptors known to play major roles in neural development, such as THRα, RXRs, RORs, TRs, and COUPTFs, were highly expressed. CAR, which plays important roles in xenobiotic metabolism, was also highly expressed. FGF2 withdrawal induced mRNA expression of RORγ, RXRγ, and MR by over 20-fold. Most of the transcriptional coregulators examined were expressed basally and throughout differentiation without major changes, while FGF2 withdrawal strongly induced mRNA expression of several histone deacetylases (HDACs), including HDAC11. Dexamethasone and aldosterone, respectively a synthetic glucocorticoid and natural mineralocorticoid, increased NSC numbers and induced differentiation into neurons and astrocytes. These results indicate that the NRs and their coregulators are present and/or change their expression during NSC differentiation, suggesting that they may influence development of the central nervous system in the absence or presence of their ligands. PMID:22990992

  18. Isolation of Human Neural Stem Cells from the Amniotic Fluid with Diagnosed Neural Tube Defects.

    PubMed

    Chang, Yu-Jen; Su, Hong-Lin; Hsu, Lee-Feng; Huang, Po-Jui; Wang, Tzu-Hao; Cheng, Fu-Chou; Hsu, Li-Wen; Tsai, Ming-Song; Chen, Chih-Ping; Chang, Yao-Lung; Chao, An-Shine; Hwang, Shiaw-Min

    2015-08-01

    Human neural stem cells (NSCs) are particularly valuable for the study of neurogenesis process and have a therapeutic potential in treating neurodegenerative disorders. However, current progress in the use of human NSCs is limited due to the available NSC sources and the complicated isolation and culture techniques. In this study, we describe an efficient method to isolate and propagate human NSCs from the amniotic fluid with diagnosed neural tube defects (NTDs), specifically, anencephaly. These amniotic fluid-derived NSCs (AF-NSCs) formed neurospheres and underwent long-term expansion in vitro. In addition, these cells showed normal karyotypes and telomerase activity and expressed NSC-specific markers, including Nestin, Sox2, Musashi-1, and the ATP-binding cassette G2 (ABCG2). AF-NSCs displayed typical morphological patterns and expressed specific markers that were consistent with neurons, astrocytes, oligodendrocytes, and dopaminergic neurons after proper induction conditions. Furthermore, grafted AF-NSCs improved the physiological functions in a rat stroke model. The ability to isolate and bank human NSCs from this novel source provides a unique opportunity for translational studies of neurological disorders.

  19. Automatic discovery of cell types and microcircuitry from neural connectomics

    PubMed Central

    Jonas, Eric; Kording, Konrad

    2015-01-01

    Neural connectomics has begun producing massive amounts of data, necessitating new analysis methods to discover the biological and computational structure. It has long been assumed that discovering neuron types and their relation to microcircuitry is crucial to understanding neural function. Here we developed a non-parametric Bayesian technique that identifies neuron types and microcircuitry patterns in connectomics data. It combines the information traditionally used by biologists in a principled and probabilistically coherent manner, including connectivity, cell body location, and the spatial distribution of synapses. We show that the approach recovers known neuron types in the retina and enables predictions of connectivity, better than simpler algorithms. It also can reveal interesting structure in the nervous system of Caenorhabditis elegans and an old man-made microprocessor. Our approach extracts structural meaning from connectomics, enabling new approaches of automatically deriving anatomical insights from these emerging datasets. DOI: http://dx.doi.org/10.7554/eLife.04250.001 PMID:25928186

  20. Axonal control of the adult neural stem cell niche.

    PubMed

    Tong, Cheuk Ka; Chen, Jiadong; Cebrián-Silla, Arantxa; Mirzadeh, Zaman; Obernier, Kirsten; Guinto, Cristina D; Tecott, Laurence H; García-Verdugo, Jose Manuel; Kriegstein, Arnold; Alvarez-Buylla, Arturo

    2014-04-03

    The ventricular-subventricular zone (V-SVZ) is an extensive germinal niche containing neural stem cells (NSCs) in the walls of the lateral ventricles of the adult brain. How the adult brain's neural activity influences the behavior of adult NSCs remains largely unknown. We show that serotonergic (5HT) axons originating from a small group of neurons in the raphe form an extensive plexus on most of the ventricular walls. Electron microscopy revealed intimate contacts between 5HT axons and NSCs (B1) or ependymal cells (E1) and these cells were labeled by a transsynaptic viral tracer injected into the raphe. B1 cells express the 5HT receptors 2C and 5A. Electrophysiology showed that activation of these receptors in B1 cells induced small inward currents. Intraventricular infusion of 5HT2C agonist or antagonist increased or decreased V-SVZ proliferation, respectively. These results indicate that supraependymal 5HT axons directly interact with NSCs to regulate neurogenesis via 5HT2C. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. Axonal Control of the Adult Neural Stem Cell Niche

    PubMed Central

    Tong, Cheuk Ka; Chen, Jiadong; Cebrián-Silla, Arantxa; Mirzadeh, Zaman; Obernier, Kirsten; Guinto, Cristina D.; Tecott, Laurence H.; García-Verdugo, Jose Manuel; Kriegstein, Arnold; Alvarez-Buylla, Arturo

    2014-01-01

    SUMMARY The ventricular-subventricular zone (V-SVZ) is an extensive germinal niche containing neural stem cells (NSC) in the walls of the lateral ventricles of the adult brain. How the adult brain’s neural activity influences the behavior of adult NSCs remains largely unknown. We show that serotonergic (5HT) axons originating from a small group of neurons in the raphe form an extensive plexus on most of the ventricular walls. Electron microscopy revealed intimate contacts between 5HT axons and NSCs (B1) or ependymal cells (E1) and these cells were labeled by a transsynaptic viral tracer injected into the raphe. B1 cells express the 5HT receptors 2C and 5A. Electrophysiology showed that activation of these receptors in B1 cells induced small inward currents. Intraventricular infusion of 5HT2C agonist or antagonist increased or decreased V-SVZ proliferation, respectively. These results indicate that supraependymal 5HT axons directly interact with NSCs to regulate neurogenesis via 5HT2C. PMID:24561083

  2. The emerging role of epigenetics in stroke: III. Neural stem cell biology and regenerative medicine.

    PubMed

    Qureshi, Irfan A; Mehler, Mark F

    2011-03-01

    The transplantation of exogenous stem cells and the activation of endogenous neural stem and progenitor cells (NSPCs) are promising treatments for stroke. These cells can modulate intrinsic responses to ischemic injury and may even integrate directly into damaged neural networks. However, the neuroprotective and neural regenerative effects that can be mediated by these cells are limited and may even be deleterious. Epigenetic reprogramming represents a novel strategy for enhancing the intrinsic potential of the brain to protect and repair itself by modulating pathologic neural gene expression and promoting the recapitulation of seminal neural developmental processes. In fact, recent evidence suggests that emerging epigenetic mechanisms are critical for orchestrating nearly every aspect of neural development and homeostasis, including brain patterning, neural stem cell maintenance, neurogenesis and gliogenesis, neural subtype specification, and synaptic and neural network connectivity and plasticity. In this review, we survey the therapeutic potential of exogenous stem cells and endogenous NSPCs and highlight innovative technological approaches for designing, developing, and delivering epigenetic therapies for targeted reprogramming of endogenous pools of NSPCs, neural cells at risk, and dysfunctional neural networks to rescue and restore neurologic function in the ischemic brain.

  3. Mesoderm is required for coordinated cell movements within zebrafish neural plate in vivo

    PubMed Central

    2014-01-01

    Background Morphogenesis of the zebrafish neural tube requires the coordinated movement of many cells in both time and space. A good example of this is the movement of the