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Sample records for neural cell growth

  1. Reflectin as a Material for Neural Stem Cell Growth.

    PubMed

    Phan, Long; Kautz, Rylan; Arulmoli, Janahan; Kim, Iris H; Le, Dai Trang T; Shenk, Michael A; Pathak, Medha M; Flanagan, Lisa A; Tombola, Francesco; Gorodetsky, Alon A

    2016-01-13

    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

  2. Short Laminin Peptide for Improved Neural Stem Cell Growth

    PubMed Central

    Li, Xiaowei; Liu, Xiaoyan; Josey, Benjamin; Chou, C. James; Tan, Yu; Zhang, Ning

    2014-01-01

    Human neural stem/progenitor cells (hNSCs) are very difficult to culture and require human or animal source extracellular matrix molecules, such as laminin or collagen type IV, to support attachment and to regulate their survival and proliferation. These extracellular matrix molecules are difficult to purify from human or animal tissues, have high batch-to-batch variability, and may cause an immune response if used in clinical applications. Although several laminin- and collagen IV-derived peptides are commercially available, they do not support long-term hNSC attachment and growth. To solve this problem, we developed a novel peptide sequence with only 12 amino acids based on the Ile-Lys-Val-Ala-Val, or IKVAV, sequence: Ac-Cys-Cys-Arg-Arg-Ile-Lys-Val-Ala-Val-Trp-Leu-Cys. This short peptide sequence, similar to tissue-derived full laminin molecules, supported hNSCs to attach and proliferate to confluence for continuous passage and subculture. This short peptide also directed hNSCs to differentiate into neurons. When conjugated to poly(ethylene glycol) hydrogels, this short peptide benefited hNSC attachment and proliferation on the surface of hydrogels and promoted cell migration inside the hydrogels with maximum enhancement at a peptide density of 10 μM. This novel short peptide shows great promise in artificial niche development for supporting hNSC culture in vitro and in vivo and for promoting hNSC transplantation in future clinical therapy. PMID:24692587

  3. Human neural progenitors express functional lysophospholipid receptors that regulate cell growth and morphology

    PubMed Central

    Hurst, Jillian H; Mumaw, Jennifer; Machacek, David W; Sturkie, Carla; Callihan, Phillip; Stice, Steve L; Hooks, Shelley B

    2008-01-01

    Background Lysophospholipids regulate the morphology and growth of neurons, neural cell lines, and neural progenitors. A stable human neural progenitor cell line is not currently available in which to study the role of lysophospholipids in human neural development. We recently established a stable, adherent human embryonic stem cell-derived neuroepithelial (hES-NEP) cell line which recapitulates morphological and phenotypic features of neural progenitor cells isolated from fetal tissue. The goal of this study was to determine if hES-NEP cells express functional lysophospholipid receptors, and if activation of these receptors mediates cellular responses critical for neural development. Results Our results demonstrate that Lysophosphatidic Acid (LPA) and Sphingosine-1-phosphate (S1P) receptors are functionally expressed in hES-NEP cells and are coupled to multiple cellular signaling pathways. We have shown that transcript levels for S1P1 receptor increased significantly in the transition from embryonic stem cell to hES-NEP. hES-NEP cells express LPA and S1P receptors coupled to Gi/o G-proteins that inhibit adenylyl cyclase and to Gq-like phospholipase C activity. LPA and S1P also induce p44/42 ERK MAP kinase phosphorylation in these cells and stimulate cell proliferation via Gi/o coupled receptors in an Epidermal Growth Factor Receptor (EGFR)- and ERK-dependent pathway. In contrast, LPA and S1P stimulate transient cell rounding and aggregation that is independent of EGFR and ERK, but dependent on the Rho effector p160 ROCK. Conclusion Thus, lysophospholipids regulate neural progenitor growth and morphology through distinct mechanisms. These findings establish human ES cell-derived NEP cells as a model system for studying the role of lysophospholipids in neural progenitors. PMID:19077254

  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. 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. PMID:27212919

  6. 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.

  7. Long-Distance Growth and Connectivity of Neural Stem Cells After Severe Spinal Cord Injury

    PubMed Central

    Lu, Paul; Wang, Yaozhi; Graham, Lori; McHale, Karla; Gao, Mingyong; Wu, Di; Brock, John; Blesch, Armin; Rosenzweig, Ephron S.; Havton, Leif A.; Zheng, Binhai; Conner, James M.; Marsala, Martin; Tuszynski, Mark H.

    2012-01-01

    SUMMARY Neural stem cells (NSCs) expressing GFP were embedded into fibrin matrices containing growth factor cocktails and grafted to sites of severe spinal cord injury. Grafted cells differentiated into multiple cellular phenotypes, including neurons, which extended large numbers of axons over remarkable distances. Extending axons formed abundant synapses with host cells. Axonal growth was partially dependent on mammalian target of rapamycin (mTOR) but not Nogo signaling. Grafted neurons supported formation of electrophysiological relays across sites of complete spinal transection, resulting in functional recovery. Two human stem cell lines (566RSC and HUES7) embedded in growth factor-containing fibrin exhibited similar growth, and 566RSC cells supported functional recovery. Thus, properties intrinsic to early stage neurons can overcome the inhibitory milieu of the injured adult spinal cord to mount remarkable axonal growth resulting in formation of novel relay circuits that significantly improve function. These therapeutic properties extend across stem cell sources and species. PMID:22980985

  8. 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.

  9. Titanium oxide as substrate for neural cell growth.

    PubMed

    Carballo-Vila, Mónica; Moreno-Burriel, Berta; Chinarro, Eva; Jurado, José R; Casañ-Pastor, Nieves; Collazos-Castro, Jorge E

    2009-07-01

    Titanium oxide has antiinflammatory activity and tunable electrochemical behavior that make it an attractive material for the fabrication of implantable devices. The most stable composition is TiO2 and occurs mainly in three polymorphs, namely, anatase, rutile, and brookite, which differ in its crystallochemical properties. Here, we report the preparation of rutile surfaces that permit good adherence and axonal growth of cultured rat cerebral cortex neurons. Rutile disks were obtained by sinterization of TiO2 powders of commercial origin or precipitated from hydrolysis of Ti(IV)-isopropoxide. Commercial powders sintered at 1300-1600 degrees C produced rutile surfaces with abnormal grain growth, probably because of impurities of the powders. Neurons cultured on those surfaces survived in variable numbers and showed fewer neurites than on control materials. On the other hand, rutile sintered from precipitated powders had less contaminants and more homogenous grain growth. By adjusting the thermal treatment it was possible to obtain surfaces performing well as substrate for neuron survival for at least 10 days. Some surfaces permitted normal axonal elongation, whereas dendrite growth was generally impaired. These findings support the potential use of titanium oxide in neuroprostheses and other devices demanding materials with enhanced properties in terms of biocompatibility and axon growth promotion. PMID:18481786

  10. Nitric oxide stimulates the proliferation of neural stem cells bypassing the epidermal growth factor receptor.

    PubMed

    Carreira, Bruno Pereira; Morte, Maria Inês; Inácio, Angela; Costa, Gabriel; Rosmaninho-Salgado, Joana; Agasse, Fabienne; Carmo, Anália; Couceiro, Patrícia; Brundin, Patrik; Ambrósio, António Francisco; Carvalho, Caetana Monteiro; Araújo, Inês Maria

    2010-07-01

    Nitric oxide (NO) was described to inhibit the proliferation of neural stem cells. Some evidence suggests that NO, under certain conditions, can also promote cell proliferation, although the mechanisms responsible for a potential proliferative effect of NO in neural stem cells have remained unaddressed. In this work, we investigated and characterized the proliferative effect of NO in cell cultures obtained from the mouse subventricular zone. We found that the NO donor NOC-18 (10 microM) increased cell proliferation, whereas higher concentrations (100 microM) inhibited cell proliferation. Increased cell proliferation was detected rapidly following exposure to NO and was prevented by blocking the mitogen-activated kinase (MAPK) pathway, independently of the epidermal growth factor (EGF) receptor. Downstream of the EGF receptor, NO activated p21Ras and the MAPK pathway, resulting in a decrease in the nuclear presence of the cyclin-dependent kinase inhibitor 1, p27(KIP1), allowing for cell cycle progression. Furthermore, in a mouse model that shows increased proliferation of neural stem cells in the hippocampus following seizure injury, we observed that the absence of inducible nitric oxide synthase (iNOS(-/-) mice) prevented the increase in cell proliferation observed following seizures in wild-type mice, showing that NO from iNOS origin is important for increased cell proliferation following a brain insult. Overall, we show that NO is able to stimulate the proliferation of neural stem cells bypassing the EGF receptor and promoting cell division. Moreover, under pathophysiological conditions in vivo, NO from iNOS origin also promotes proliferation in the hippocampus. PMID:20506358

  11. Laminin enhances the growth of human neural stem cells in defined culture media

    PubMed Central

    Hall, Peter E; Lathia, Justin D; Caldwell, Maeve A; ffrench-Constant, Charles

    2008-01-01

    Background Human neural stem cells (hNSC) have the potential to provide novel cell-based therapies for neurodegenerative conditions such as multiple sclerosis and Parkinson's disease. In order to realise this goal, protocols need to be developed that allow for large quantities of hNSC to be cultured efficiently. As such, it is important to identify factors which enhance the growth of hNSC. In vivo, stem cells reside in distinct microenvironments or niches that are responsible for the maintenance of stem cell populations. A common feature of niches is the presence of the extracellular matrix molecule, laminin. Therefore, this study investigated the effect of exogenous laminin on hNSC growth. Results To measure hNSC growth, we established culture conditions using B27-supplemented medium that enable neurospheres to grow from human neural cells plated at clonal densities. Limiting dilution assays confirmed that neurospheres were derived from single cells at these densities. Laminin was found to increase hNSC numbers as measured by this neurosphere formation. The effect of laminin was to augment the proliferation/survival of the hNSC, rather than promoting the undifferentiated state. In agreement, apoptosis was reduced in dissociated neurospheres by laminin in an integrin β1-dependent manner. Conclusion The addition of laminin to the culture medium enhances the growth of hNSC, and may therefore aid their large-scale production. PMID:18651950

  12. Vascular endothelial growth factor-expressing neural stem cell for the treatment of neuropathic pain.

    PubMed

    Lee, Hye-Lan; Oh, Jinsoo; Yun, Yeomin; Lee, Hye Yeong; You, Youngsang; Che, Lihua; Lee, Minhyung; Kim, Keung Nyun; Ha, Yoon

    2015-05-01

    Previously, we determined that vascular endothelial growth factor (VEGF) improves the survival of neural stem cells (NSCs) transplanted into an ischemic environment and effectively enhances angiogenesis. Here, we applied NSCs expressing VEGF (SV-VEGF-NSCs) to treat neuropathic pain. In this study, our goal was to verify the therapeutic effect of SV-VEGF-NSCs by transplanting the cells in a sciatic nerve injury model. We compared the amount of VEGF secreted from DsRed-NSCs (control) or SV-VEGF-NSCs and observed that SV-VEGF-NSCs have a much higher expression level of VEGF. We next investigated whether transplantation with SV-VEGF-NSCs aids functional recovery and pain reduction. We confirmed that transplantation with SV-VEGF-NSCs enhances functional recovery, pain reduction, and remyelination as well as the number of blood vessels compared with the control groups. Our results show that VEGF aids functional recovery and pain reduction in a sciatic nerve injury model. PMID:25793634

  13. 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

  14. Temporal changes in the response of SVZ neural stem cells to intraventricular administration of growth factors.

    PubMed

    Ochi, Takashi; Nakatomi, Hirofumi; Ito, Akihiro; Imai, Hideaki; Okabe, Shigeo; Saito, Nobuhito

    2016-04-01

    In vivo growth factor (GF) treatment is a promising approach to enhance the regenerative capacity of neural stem cells (NSCs) for brain repair. However, how exogenous GFs affect endogenous NSCs is not well understood. This study investigated the impact of intraventricular administration of fibroblast growth factor 2 and epidermal growth factor on NSCs in the subventricular zone of intact adult mice. GFs were administered for various periods (3, 7, 10, and 14 days), and the proliferation and neuronal production of NSCs were assessed during and after GF treatment. We found that proliferation of NSCs and their progeny is markedly augmented during the first 7 days after the initiation of GF treatment. GF treatment for longer periods, however, did not lead to further increases in the NSC pool, but rather attenuated such proliferation and inhibited neurogenesis. As a result, the production of new olfactory bulb neurons was increased in animals treated with GFs for 7 days but decreased in animals treated for 14 days. These results show time-dependent changes in the response of NSCs to exogenous GFs and demonstrate that precise control of the duration of GF treatment is important for significant enhancement of neuronal production by NSCs in vivo for brain repair. PMID:26845459

  15. Requirement for neurogenesis to proceed through the division of neuronal progenitors following differentiation of epidermal growth factor and fibroblast growth factor-2-responsive human neural stem cells.

    PubMed

    Ostenfeld, Thor; Svendsen, Clive N

    2004-01-01

    Epidermal growth factor (EGF)- and fibroblast growth factor-2 (FGF-2)-responsive human neural stem cells may provide insight into mechanisms of neural development and have applications in cell-based therapeutics for neurological disease. However, their biology after expansion in vitro is currently poorly understood. Cells grown in either EGF or FGF-2 or a combination of both mitogens displayed characteristically similar levels of transcriptional activation and comparable proliferative profiles with linear cell-cycle kinetics and possessed similar neuronal differentiation capabilities. These data support the view that human neurospheres at later stages of expansion (>10 weeks) are comprised overwhelmingly of a single type of stem cell responsive to both EGF and FGF-2. After mitogen withdrawal and neurosphere plating, bromodeoxyuridine pulse-chase experiments revealed that the stem cells did not undergo differentiation directly into neurons. Instead, most immature neurons arose via the division of emerging progenitor cells in the absence of exogenous EGF or FGF-2. Neurogenesis was abolished by application of high concentrations of either EGF/FGF-2 or the mitotic inhibitor cytosine-b-arabinofuranoside, suggesting that there is an obligatory requirement for at least one round of cell division in the absence of mitogens as a prelude to terminal neuronal differentiation. The differentiation of human neurospheres provides a useful model of human neurogenesis, and the data presented indicate that it proceeds through the division of committed neuronal progenitor cells rather than directly from the neural stem cell. PMID:15342944

  16. Changes in expression and secretion patterns of fibroblast growth factor 8 and Sonic Hedgehog signaling pathway molecules during murine neural stem/progenitor cell differentiation in vitro☆

    PubMed Central

    Lu, Jiang; Lu, Kehuan; Li, Dongsheng

    2012-01-01

    In the present study, we investigated the dynamic expression of fibroblast growth factor 8 and Sonic Hedgehog signaling pathway related factors in the process of in vitro hippocampal neural stem/progenitor cell differentiation from embryonic Sprague-Dawley rats or embryonic Kunming species mice, using fluorescent quantitative reverse transcription-PCR and western blot analyses. Results demonstrated that the dynamic expression of fibroblast growth factor 8 was similar to fibroblast growth factor receptor 1 expression but not to other fibroblast growth factor receptors. Enzyme-linked immunosorbent assay demonstrated that fibroblast growth factor 8 and Sonic Hedgehog signaling pathway protein factors were secreted by neural cells into the intercellular niche. Our experimental findings indicate that fibroblast growth factor 8 and Sonic Hedgehog expression may be related to the differentiation of neural stem/progenitor cells. PMID:25624789

  17. Promotion of Survival and Differentiation of Neural Stem Cells with Fibrin and Growth Factor Cocktails after Severe Spinal Cord Injury

    PubMed Central

    Lu, Paul; Graham, Lori; Wang, Yaozhi; Wu, Di; Tuszynski, Mark

    2014-01-01

    Neural stem cells (NSCs) can self-renew and differentiate into neurons and glia. Transplanted NSCs can replace lost neurons and glia after spinal cord injury (SCI), and can form functional relays to re-connect spinal cord segments above and below a lesion. Previous studies grafting neural stem cells have been limited by incomplete graft survival within the spinal cord lesion cavity. Further, tracking of graft cell survival, differentiation, and process extension had not been optimized. Finally, in previous studies, cultured rat NSCs were typically reported to differentiate into glia when grafted to the injured spinal cord, rather than neurons, unless fate was driven to a specific cell type. To address these issues, we developed new methods to improve the survival, integration and differentiation of NSCs to sites of even severe SCI. NSCs were freshly isolated from embryonic day 14 spinal cord (E14) from a stable transgenic Fischer 344 rat line expressing green fluorescent protein (GFP) and were embedded into a fibrin matrix containing growth factors; this formulation aimed to retain grafted cells in the lesion cavity and support cell survival. NSCs in the fibrin/growth factor cocktail were implanted two weeks after thoracic level-3 (T3) complete spinal cord transections, thereby avoiding peak periods of inflammation. Resulting grafts completely filled the lesion cavity and differentiated into both neurons, which extended axons into the host spinal cord over remarkably long distances, and glia. Grafts of cultured human NSCs expressing GFP resulted in similar findings. Thus, methods are defined for improving neural stem cell grafting, survival and analysis of in vivo findings. PMID:25145787

  18. 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

  19. Direct microfabrication of topographical and chemical cues for the guided growth of neural cell networks on polyamidoamine hydrogels.

    PubMed

    Dos Reis, Gabriel; Fenili, Fabio; Gianfelice, Antonella; Bongiorno, Gero; Marchesi, Davide; Scopelliti, Pasquale Emanuele; Borgonovo, Antonio; Podestà, Alessandro; Indrieri, Marco; Ranucci, Elisabetta; Ferruti, Paolo; Lenardi, Cristina; Milani, Paolo

    2010-08-11

    Cell patterning is an important tool for organizing cells in surfaces and to reproduce in a simple way the tissue hierarchy and complexity of pluri-cellular life. The control of cell growth, proliferation and differentiation on solid surfaces is consequently important for prosthetics, biosensors, cell-based arrays, stem cell therapy and cell-based drug discovery concepts. We present a new electron beam lithography method for the direct and simultaneous fabrication of sub-micron topographical and chemical patterns, on a biocompatible and biodegradable PAA hydrogel. The localized e-beam modification of a hydrogel surface makes the pattern able to adsorb proteins in contrast with the anti-fouling surface. By also exploiting the selective attachment, growth and differentiation of PC12 cells, we fabricated a neural network of single cells connected by neuritis extending along microchannels. E-beam microlithography on PAA hydrogels opens up the opportunity of producing multifunctional microdevices incorporating complex topographies, allowing precise control of the growth and organization of individual cells. PMID:20437406

  20. 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

    2016-04-01

    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. PMID:26913590

  1. Neural induction, neural fate stabilization, and neural stem cells.

    PubMed

    Moody, Sally A; Je, Hyun-Soo

    2002-04-28

    The promise of stem cell therapy is expected to greatly benefit the treatment of neurodegenerative diseases. An underlying biological reason for the progressive functional losses associated with these diseases is the extremely low natural rate of self-repair in the nervous system. Although the mature CNS harbors a limited number of self-renewing stem cells, these make a significant contribution to only a few areas of brain. Therefore, it is particularly important to understand how to manipulate embryonic stem cells and adult neural stem cells so their descendants can repopulate and functionally repair damaged brain regions. A large knowledge base has been gathered about the normal processes of neural development. The time has come for this information to be applied to the problems of obtaining sufficient, neurally committed stem cells for clinical use. In this article we review the process of neural induction, by which the embryonic ectodermal cells are directed to form the neural plate, and the process of neural-fate stabilization, by which neural plate cells expand in number and consolidate their neural fate. We will present the current knowledge of the transcription factors and signaling molecules that are known to be involved in these processes. We will discuss how these factors may be relevant to manipulating embryonic stem cells to express a neural fate and to produce large numbers of neurally committed, yet undifferentiated, stem cells for transplantation therapies. PMID:12805974

  2. 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

  3. Chondroitinase and growth factors enhance activation and oligodendrocyte differentiation of endogenous neural precursor cells after spinal cord injury.

    PubMed

    Karimi-Abdolrezaee, Soheila; Schut, Desiree; Wang, Jian; Fehlings, Michael G

    2012-01-01

    The adult spinal cord harbours a population of multipotent neural precursor cells (NPCs) with the ability to replace oligodendrocytes. However, despite this capacity, proliferation and endogenous remyelination is severely limited after spinal cord injury (SCI). In the post-traumatic microenvironment following SCI, endogenous spinal NPCs mainly differentiate into astrocytes which could contribute to astrogliosis that exacerbate the outcomes of SCI. These findings emphasize a key role for the post-SCI niche in modulating the behaviour of spinal NPCs after SCI. We recently reported that chondroitin sulphate proteoglycans (CSPGs) in the glial scar restrict the outcomes of NPC transplantation in SCI by reducing the survival, migration and integration of engrafted NPCs within the injured spinal cord. These inhibitory effects were attenuated by administration of chondroitinase (ChABC) prior to NPC transplantation. Here, in a rat model of compressive SCI, we show that perturbing CSPGs by ChABC in combination with sustained infusion of growth factors (EGF, bFGF and PDGF-AA) optimize the activation and oligodendroglial differentiation of spinal NPCs after injury. Four days following SCI, we intrathecally delivered ChABC and/or GFs for seven days. We performed BrdU incorporation to label proliferating cells during the treatment period after SCI. This strategy increased the proliferation of spinal NPCs, reduced the generation of new astrocytes and promoted their differentiation along an oligodendroglial lineage, a prerequisite for remyelination. Furthermore, ChABC and GF treatments enhanced the response of non-neural cells by increasing the generation of new vascular endothelial cells and decreasing the number of proliferating macrophages/microglia after SCI. In conclusions, our data strongly suggest that optimization of the behaviour of endogenous spinal NPCs after SCI is critical not only to promote endogenous oligodendrocyte replacement, but also to reverse the otherwise

  4. Epidermal growth factor (EGF) withdrawal masks gene expression differences in the study of pituitary adenylate cyclase-activating polypeptide (PACAP) activation of primary neural stem cell proliferation

    PubMed Central

    Sievertzon, Maria; Wirta, Valtteri; Mercer, Alex; Frisén, Jonas; Lundeberg, Joakim

    2005-01-01

    Background The recently discovered adult neural stem cells, which maintain continuous generation of new neuronal and glial cells throughout adulthood, are a promising and expandable source of cells for use in cell replacement therapies within the central nervous system. These cells could either be induced to proliferate and differentiate endogenously, or expanded and differentiated in culture before being transplanted into the damaged site of the brain. In order to achieve these goals effective strategies to isolate, expand and differentiate neural stem cells into the desired specific phenotypes must be developed. However, little is known as yet about the factors and mechanisms influencing these processes. It has recently been reported that pituitary adenylate cyclase-activating polypeptide (PACAP) promotes neural stem cell proliferation both in vivo and in vitro. Results We used cDNA microarrays with the aim of analysing the transcriptional changes underlying PACAP induced proliferation of neural stem cells. The primary neural stem/progenitor cells used were neurospheres, generated from the lateral ventricle wall of the adult mouse brain. The results were compared to both differentiation and proliferation controls, which revealed an unexpected and significant differential expression relating to withdrawal of epidermal growth factor (EGF) from the neurosphere growth medium. The effect of EGF removal was so pronounced that it masked the changes in gene expression patterns produced by the addition of PACAP. Conclusion Experimental models aiming at transcriptional analysis of induced proliferation in primary neural stem cells need to take into consideration the significant effect on transcription caused by removal of EGF. Alternatively, EGF-free culture conditions need to be developed. PMID:16124881

  5. 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. PMID:26744412

  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. 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. PMID:24933469

  8. Human Neural Stem Cells Overexpressing a Carboxylesterase Inhibit Bladder Tumor Growth.

    PubMed

    Choi, Sung S; Chi, Byung Hoon; Chang, In Ho; Kim, Kyung Do; Lee, Sang-Rae; Kim, Seung U; Lee, Hong J

    2016-06-01

    Bladder cancer is a significant clinical and economic problem. Despite intravesical chemotherapy and immunotherapy, up to 80% of patients with non-muscle-invasive bladder cancer develop recurrent tumors, of which 20% to 30% evolve into more aggressive, potentially lethal tumors. Recently, bladder cancer cells are considered to be mediators of resistance to current therapies and therefore represent strong candidates as biologic targets. No effective chemotherapy has yet been developed for advanced bladder cancer. It is desirable that a drug can be delivered directly and specifically to bladder cancer cells. Stem cells have selective migration ability toward cancer cells, and therapeutic genes can be easily transduced into stem cells. In suicide gene therapy for cancer, stem cells carry a gene encoding a carboxylesterase (CE) enzyme that transforms an inert CPT-11 prodrug into a toxic SN-38 product, a topoisomerase 1 inhibitor. In immunodeficient mice, systemically transplanted HB1.F3.CE stem cells migrated toward the tumor implanted by the TCCSUP bladder cancer cell line, and, in combination with CPT-11, the volume of tumors was significantly reduced. These findings may contribute to the development of a new selective chemotherapeutic strategy against bladder cancer. Mol Cancer Ther; 15(6); 1201-7. ©2016 AACR. PMID:27009215

  9. 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

  10. Transforming growth factor-beta 1 differentially regulates proliferation, morphology, and extracellular matrix expression by three neural crest-derived neuroblastoma cell lines.

    PubMed

    Rogers, S L; Cutts, J L; Gegick, P J; McGuire, P G; Rosenberger, C; Krisinski, S

    1994-04-01

    We reported previously (S. L. Rogers, P. J. Gegick, S. M. Alexander, and P. G. McGuire, Dev. Biol. 151, 191-203, 1992) that transforming growth factor-beta 1 (TGF beta 1) inhibited proliferation, up-regulated fibronectin synthesis, and suppressed melanogenesis in a population of quail neural crest cells in vitro. Here, we report that cell lines derived from the parent SK-N-SH neuroblastoma line (R. A. Ross, B. A. Spengler, and J. L. Biedler, J. Natl. Cancer Inst. 71, 741-747, 1983) respond differentially to TGF beta 1, and their responses provide further insights into the actions of this growth factor on neural crest subpopulations. The SH-EP cell line exhibits primarily nonneuronal traits and responded to TGF beta 1 with increased thymidine uptake after 6 days of culture, increased expression of fibronectin mRNA and protein, and decreased laminin synthesis. Many SH-EP cells also acquired a dramatically elongated morphology, reminiscent of Schwann cells in culture. Thymidine uptake by the neuronal SY5Y cell line was not substantially altered. Neither fibronectin mRNA nor protein was detectable in either TGF beta 1-treated or untreated cultures, although laminin synthesis was upregulated by the growth factor. In TGF beta 1-treated cultures of the intermediate SH-IN cell line, which has been reported to display both neuronal and nonneuronal characteristics, there was marked flattening of many cells, a steady decrease in thymidine uptake, and increased expression of both fibronectin and laminin. The observed responses of SH-IN cells mimic those observed in primary neural crest cultures and appear to represent similar differentiation toward a mesenchymal phenotype. These results substantiate the idea that closely related but diverging neural crest-derived cell types respond selectively to TGF beta 1 and demonstrate that these SK-N-SH-derived cell lines will be useful in experimental approaches that will allow us to infer mechanisms underlying regulation of neural crest

  11. 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

  12. Phenytoin enhances the phosphorylation of epidermal growth factor receptor and fibroblast growth factor receptor in the subventricular zone and promotes the proliferation of neural precursor cells and oligodendrocyte differentiation.

    PubMed

    Galvez-Contreras, Alma Y; Gonzalez-Castaneda, Rocio E; Campos-Ordonez, Tania; Luquin, Sonia; Gonzalez-Perez, Oscar

    2016-01-01

    Phenytoin is a widely used antiepileptic drug that induces cell proliferation in several tissues, such as heart, bone, skin, oral mucosa and neural precursors. Some of these effects are mediated via fibroblast growth factor receptor (FGFR) and epidermal growth factor receptor (EGFR). These receptors are strongly expressed in the adult ventricular-subventricular zone (V-SVZ), the main neurogenic niche in the adult brain. The aim of this study was to determine the cell lineage and cell fate of V-SVZ neural progenitors expanded by phenytoin, as well as the effects of this drug on EGFR/FGFR phosphorylation. Male BALB/C mice received 10 mg/kg phenytoin by oral cannula for 30 days. We analysed the proliferation of V-SVZ neural progenitors by immunohistochemistry and western blot. Our findings indicate that phenytoin enhanced twofold the phosphorylation of EGFR and FGFR in the V-SVZ, increased the number of bromodeoxyuridine (BrdU)+/Sox2+ and BrdU+/doublecortin+ cells in the V-SVZ, and expanded the population of Olig2-expressing cells around the lateral ventricles. After phenytoin removal, a large number of BrdU+/Receptor interacting protein (RIP)+ cells were observed in the olfactory bulb. In conclusion, phenytoin enhanced the phosphorylation of FGFR and EGFR, and promoted the expression of neural precursor markers in the V-SVZ. In parallel, the number of oligodendrocytes increased significantly after phenytoin removal. PMID:26370587

  13. Hepatocyte Growth Factor Activator Inhibitor-1 Is Induced by Bone Morphogenetic Proteins and Regulates Proliferation and Cell Fate of Neural Progenitor Cells

    PubMed Central

    Koivuniemi, Raili; Mäkelä, Johanna; Hokkanen, Marie-Estelle; Bruelle, Céline; Ho, Tho Huu; Ola, Roxana; Korhonen, Laura; Schröder, Jim; Kataoka, Hiroaki; Lindholm, Dan

    2013-01-01

    Background Neural progenitor cells (NPCs) in the developing neuroepithelium are regulated by intrinsic and extrinsic factors. There is evidence that NPCs form a self-supporting niche for cell maintenance and proliferation. However, molecular interactions and cell-cell contacts and the microenvironment within the neuroepithelium are largely unknown. We hypothesized that cellular proteases especially those associated with the cell surface of NPCs play a role in regulation of progenitor cells in the brain. Methodology/Principal Findings In this work, we show that NPCs, isolated from striatal anlage of developing rat brain, express hepatocyte growth factor activator inhibitor-1 and -2 (HAI-1 and HAI-2) that are cell surface-linked serine protease inhibitors. In addition, radial glia cells derived from mouse embryonic stem cells also express HAI-1 and HAI-2. To study the functional significance of HAI-1 and HAI-2 in progenitor cells, we modulated their levels using expression plasmids or silencing RNA (siRNA) transfected into the NPCs. Data showed that overexpression of HAI-1 or HAI-2 decreased cell proliferation of cultured NPCs, whilst their siRNAs had opposite effects. HAI-1 also influenced NPC differentiation by increasing the number of glial fibrillary acidic protein (GFAP) expressing cells in the culture. Expression of HAI-1 in vivo decreased cell proliferation in developing neuroepithelium in E15 old animals and promoted astrocyte cell differentiation in neonatal animals. Studying the regulation of HAI-1, we observed that Bone morphogenetic protein-2 (BMP-2) and BMP-4 increased HAI-1 levels in the NPCs. Experiments using HAI-1-siRNA showed that these BMPs act on the NPCs partly in a HAI-1-dependent manner. Conclusions This study shows that the cell-surface serine protease inhibitors, HAI-1 and HAI-2 influence proliferation and cell fate of NPCs and their expression levels are linked to BMP signaling. Modulation of the levels and actions of HAI-1 in NPCs may be of

  14. Telomerase and estrogen-sensing activities are essential for continued mammary growth in vivo but dispensable for “reprogramming” neural stem cells

    PubMed Central

    George, Andrea L.; Boulanger, Corinne A.; Smith, Gilbert H.

    2016-01-01

    It has been proposed that the erosion of telomere length is a limiting factor in replicative capacity and important in cell senescence. To determine if this activity was essential in the mouse mammary gland in vivo, we serially transplanted mammary fragments from wild type (TER+/+), heterozygous (TER+/−), and homozygous (TER−/−) mammary tissues into the cleared mammary fat pads of immune-compromised nude mice. Individual implants from both homozygous and heterozygous TER null outgrowths showed growth senescence beginning at transplant generation two, earlier than implants from TER+/+ mammary glands which continued to show growth. This result suggests that either mammary epithelial stem cells maintain their telomere length in order to self renew, or that the absence or reduction of telomerase template results in more frequent death/extinction of stem cells during symmetric divisions. A third possibility is the inability of signaling cells in the niche to replicate resulting in reduction of the maintenance signals necessary for stem cell renewal. Consistent with this, examination of senescent outgrowths revealed the absence of estrogen receptor alpha (ERα+) epithelium although progesterone receptor (PR+) cells were abundant. Despite their inability to establish mammary growth in vivo, TER+/− cells were able to direct neural stem cells to mammary cell fates. PMID:27347776

  15. 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. PMID:25957749

  16. 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. PMID:25530343

  17. 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

  18. 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

    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

  19. 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

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

    PubMed

    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

  1. Over-Expression of hNGF in Adult Human Olfactory Bulb Neural Stem Cells Promotes Cell Growth and Oligodendrocytic Differentiation

    PubMed Central

    Marei, Hany E. S.; Althani, Asmaa; Afifi, Nahla; Abd-Elmaksoud, Ahmed; Bernardini, Camilla; Michetti, Fabrizio; Barba, Marta; Pescatori, Mario; Maira, Giulio; Paldino, Emanuela; Manni, Luigi; Casalbore, Patrizia; Cenciarelli, Carlo

    2013-01-01

    The adult human olfactory bulb neural stem/progenitor cells (OBNC/PC) are promising candidate for cell-based therapy for traumatic and neurodegenerative insults. Exogenous application of NGF was suggested as a promising therapeutic strategy for traumatic and neurodegenerative diseases, however effective delivery of NGF into the CNS parenchyma is still challenging due mainly to its limited ability to cross the blood–brain barrier, and intolerable side effects if administered into the brain ventricular system. An effective method to ensure delivery of NGF into the parenchyma of CNS is the genetic modification of NSC to overexpress NGF gene. Overexpression of NGF in adult human OBNSC is expected to alter their proliferation and differentiation nature, and thus might enhance their therapeutic potential. In this study, we genetically modified adult human OBNS/PC to overexpress human NGF (hNGF) and green fluorescent protein (GFP) genes to provide insight about the effects of hNGF and GFP genes overexpression in adult human OBNS/PC on their in vitro multipotentiality using DNA microarray, immunophenotyping, and Western blot (WB) protocols. Our analysis revealed that OBNS/PC-GFP and OBNS/PC-GFP-hNGF differentiation is a multifaceted process involving changes in major biological processes as reflected in alteration of the gene expression levels of crucial markers such as cell cycle and survival markers, stemness markers, and differentiation markers. The differentiation of both cell classes was also associated with modulations of key signaling pathways such MAPK signaling pathway, ErbB signaling pathway, and neuroactive ligand-receptor interaction pathway for OBNS/PC-GFP, and axon guidance, calcium channel, voltage-dependent, gamma subunit 7 for OBNS/PC-GFP-hNGF as revealed by GO and KEGG. Differentiated OBNS/PC-GFP-hNGF displayed extensively branched cytoplasmic processes, a significant faster growth rate and up modulated the expression of oligodendroglia precursor cells

  2. Mechanotransduction of Neural Cells Through Cell-Substrate Interactions.

    PubMed

    Stukel, Jessica M; Willits, Rebecca Kuntz

    2016-06-01

    Neurons and neural stem cells are sensitive to their mechanical and topographical environment, and cell-substrate binding contributes to this sensitivity to activate signaling pathways for basic cell functions. Many transmembrane proteins transmit signals into and out of the cell, including integrins, growth factor receptors, G-protein-coupled receptors, cadherins, cell adhesion molecules, and ion channels. Specifically, integrins are one of the main transmembrane proteins that transmit force across the cell membrane between a cell and its extracellular matrix, making them critical in the study of cell-material interactions. This review focuses on mechanotransduction, defined as the conversion of force a cell generates through cell-substrate bonds to a chemical signal, of neural cells. The chemical signals relay information via pathways through the cellular cytoplasm to the nucleus, where signaling events can affect gene expression. Pathways and the cellular response initiated by substrate binding are explored to better understand their effect on neural cells mechanotransduction. As the results of mechanotransduction affect cell adhesion, cell shape, and differentiation, knowledge regarding neural mechanotransduction is critical for most regenerative strategies in tissue engineering, where novel environments are developed to improve conduit design for central and peripheral nervous system repair in vivo. PMID:26669274

  3. 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. PMID:19369905

  4. Electrokinetic confinement of axonal growth for dynamically configurable neural networks.

    PubMed

    Honegger, Thibault; Scott, Mark A; Yanik, Mehmet F; Voldman, Joel

    2013-02-21

    Axons in the developing nervous system are directed via guidance cues, whose expression varies both spatially and temporally, to create functional neural circuits. Existing methods to create patterns of neural connectivity in vitro use only static geometries, and are unable to dynamically alter the guidance cues imparted on the cells. We introduce the use of AC electrokinetics to dynamically control axonal growth in cultured rat hippocampal neurons. We find that the application of modest voltages at frequencies on the order of 10(5) Hz can cause developing axons to be stopped adjacent to the electrodes while axons away from the electric fields exhibit uninhibited growth. By switching electrodes on or off, we can reversibly inhibit or permit axon passage across the electrodes. Our models suggest that dielectrophoresis is the causative AC electrokinetic effect. We make use of our dynamic control over axon elongation to create an axon-diode via an axon-lock system that consists of a pair of electrode 'gates' that either permit or prevent axons from passing through. Finally, we developed a neural circuit consisting of three populations of neurons, separated by three axon-locks to demonstrate the assembly of a functional, engineered neural network. Action potential recordings demonstrate that the AC electrokinetic effect does not harm axons, and Ca(2+) imaging demonstrated the unidirectional nature of the synaptic connections. AC electrokinetic confinement of axonal growth has potential for creating configurable, directional neural networks. PMID:23314575

  5. Electrokinetic confinement of axonal growth for dynamically configurable neural networks

    PubMed Central

    Honegger, Thibault; Scott, Mark A.; Yanik, Mehmet F.; Voldman, Joel

    2013-01-01

    Axons in the developing nervous system are directed via guidance cues, whose expression varies both spatially and temporally, to create functional neural circuits. Existing methods to create patterns of neural connectivity in vitro use only static geometries, and are unable to dynamically alter the guidance cues imparted on the cells. We introduce the use of AC electrokinetics to dynamically control axonal growth in cultured rat hippocampal neurons. We find that the application of modest voltages at frequencies on the order of 105 Hz can cause developing axons to be stopped adjacent to the electrodes while axons away from the electric fields exhibit uninhibited growth. By switching electrodes on or off, we can reversibly inhibit or permit axon passage across the electrodes. Our models suggest that dielectrophoresis is the causative AC electrokinetic effect. We make use of our dynamic control over axon elongation to create an axon-diode via an axon-lock system that consists of a pair of electrode `gates' that either permit or prevent axons from passing through. Finally, we developed a neural circuit consisting of three populations of neurons, separated by three axon-locks to demonstrate the assembly of a functional, engineered neural network. Action potential recordings demonstrate that the AC electrokinetic effect does not harm axons, and Ca2+ imaging demonstrated the unidirectional nature of the synaptic connections. AC electrokinetic confinement of axonal growth has potential for creating configurable, directional neural networks. PMID:23314575

  6. Fabrication of bioactive conduits containing the fibroblast growth factor 1 and neural stem cells for peripheral nerve regeneration across a 15 mm critical gap.

    PubMed

    Ni, Hsiao-Chiang; Tseng, Ting-Chen; Chen, Jeng-Rung; Hsu, Shan-Hui; Chiu, Ing-Ming

    2013-09-01

    Nerve conduits are often used in combination with bioactive molecules and stem cells to enhance peripheral nerve regeneration. In this study, the acidic fibroblast growth factor 1 (FGF1) was immobilized onto the microporous/micropatterned poly (D, L-lactic acid) (PLA) nerve conduits after open air plasma treatment. PLA substrates grafted with chitosan in the presence of a small amount of gold nanoparticles (nano Au) showed a protective effect on the activity of the immobilized FGF1 in vitro. Different conduits were tested for their ability to bridge a 15 mm critical gap defect in a rat sciatic nerve injury model. Axon regeneration and functional recovery were evaluated by histology, walking track analysis and electrophysiology. Among different conduits, PLA conduits grafted with chitosan-nano Au and the FGF1 after plasma activation had the greatest regeneration capacity and functional recovery in the experimental animals. When the above conduit was seeded with aligned neural stem cells, the efficacy was further enhanced and it approached that of the autograft group. This work suggested that microporous/micropatterned nerve conduits containing bioactive growth factors may be successfully fabricated by micropatterning techniques, open plasma activation, and immobilization, which, combined with aligned stem cells, may synergistically contribute to the regeneration of the severely damaged peripheral nerve. PMID:23880639

  7. 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

  8. Impacts of phosphatase and tensin homology deleted on chromosome ten (PTEN)-inhibiting chitosan scaffold on growth and differentiation of neural stem cells

    PubMed Central

    Guan, Yixiang; Yang, Fu; Yao, Qi; Shi, Jinlong; Wang, Gequan; Gu, Zhikai; Zhou, Fei; Shen, Jianhong

    2015-01-01

    Objective: The aim of this study was to investigate growth and differentiation of neural stem cells (NSCs) on the phosphatase and tensin homology deleted on chromosome ten (PTEN)-inhibitor-adsorbed chitosan scaffold. Methods: NSCs were divide into the chitosan group and the control groups, and performed CCK-8 test on 1st, 3rd and 7th d to compare the proliferation between the 2 groups. The chitosan scaffold adsorbed PTEN inhibitor bpv (pic), and the empty scaffold was used as the control for co-culture of NSCs, immunofluorescence staining was performed on 7th d to detect the differentiation of NSCs on the scaffold. Results: The results of CCK-8 test showed no significant difference in the absorbance between the 2 groups. Immunofluorescence staining showed that the NSCs numbers of the bpv scaffold group were more than the empty scaffold group, among which the anti-glial fibrillary acidic protein (GFAP) positive cells were less than the empty scaffold group, while the anti-β-Tubulin III positive cells were more than the empty scaffold group, the two groups both showed rare anti-receptor-interacting protein (RIP) positive cells. Conclusions: Chitosan scaffold exhibited good compatibility to NSCs, the PTEN-inhibitor-adsorbed chitosan scaffold could promote the migration of NSCs towards the scaffold and their differentiation towards neurons. PMID:26550415

  9. 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

  10. 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

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

    2013-04-01

    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

  11. Primitive neural stem cells from the mammalian epiblast differentiate to definitive neural stem cells under the control of Notch signaling.

    PubMed

    Hitoshi, Seiji; Seaberg, Raewyn M; Koscik, Cheryl; Alexson, Tania; Kusunoki, Susumu; Kanazawa, Ichiro; Tsuji, Shoji; van der Kooy, Derek

    2004-08-01

    Basic fibroblast growth factor (FGF2)-responsive definitive neural stem cells first appear in embryonic day 8.5 (E8.5) mouse embryos, but not in earlier embryos, although neural tissue exists at E7.5. Here, we demonstrate that leukemia inhibitory factor-dependent (but not FGF2-dependent) sphere-forming cells are present in the earlier (E5.5-E7.5) mouse embryo. The resultant clonal sphere cells possess self-renewal capacity and neural multipotentiality, cardinal features of the neural stem cell. However, they also retain some nonneural properties, suggesting that they are the in vivo cells' equivalent of the primitive neural stem cells that form in vitro from embryonic stem cells. The generation of the in vivo primitive neural stem cell was independent of Notch signaling, but the activation of the Notch pathway was important for the transition from the primitive to full definitive neural stem cell properties and for the maintenance of the definitive neural stem cell state. PMID:15289455

  12. Tenuigenin promotes proliferation and differentiation of hippocampal neural stem cells.

    PubMed

    Chen, Yujing; Huang, Xiaobo; Chen, Wenqiang; Wang, Ningqun; Li, Lin

    2012-04-01

    The present study was to investigate the influence of tenuigenin, an active ingredient of Polygala tenuifolia Willd, on the proliferation and differentiation of hippocampal neural stem cells in vitro. Tenuigenin was added to a neurosphere culture and neurosphere growth was measured using MTT assay. The influence of tenuigenin on the proliferation of neural progenitors was examined by Clone forming assay and BrdU detection. In addition, the differentiation of neural stem cells was compared using immunocytochemistry for β III-tubulin and GFAP. The results showed that addition of tenuigenin to the neural stem cell medium increased the number of newly formed neurospheres. More neurons were also obtained when tenuigenin was added in the differentiation medium. These findings suggest that tenuigenin is involved in regulating the proliferation and differentiation of hippocampal neural stem cells. This result may be one of the underlying reasons for tenuigenin's nootropic and anti-aging effects. PMID:22179853

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

    PubMed Central

    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.

    2016-01-01

    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. Significance There is no cure for Alzheimer’s disease (AD) and

  14. Shifts in the Vascular endothelial growth factor (Vegf) isoforms result in transcriptome changes correlated with early neural stem cell proliferation and differentiation in mouse forebrain

    PubMed Central

    Cain, Jacob T.; Berosik, Matthew A.; Snyder, Stephanie D.; Crawford, Natalie F.; Nour, Shirin I.; Schaubhut, Geoffrey J.; Darland, Diane C.

    2014-01-01

    Regulation of neural stem cell (NSC) fate decisions is critical during the transition from a multicellular mammalian forebrain neuroepithelium to the multi-layered neocortex. Forebrain development requires coordinated vascular investment alongside NSC differentiation. Vascular endothelial growth factor A (Vegf) has proven to be a pleiotrophic gene whose multiple protein isoforms regulate a broad range of effects in neurovascular systems. To test the hypothesis that the Vegf isoforms (120, 164, and 188) are required for normal forebrain development, we analyzed the forebrain transcriptome of mice expressing specific Vegf isoforms, Vegf120, VegfF188, or a combination of Vegf120/188. Transcriptome analysis identified differentially expressed genes in embryonic day (E) 9.5 forebrain, a time point preceding dramatic neuroepithelial expansion and vascular investment in the telencephalon. Meta-analysis identified gene pathways linked to chromosome-level modifications, cell fate regulation, and neurogenesis that were altered in Vegf isoform mice. Based on these gene network shifts, we predicted that NSC populations would be affected in later stages of forebrain development. In the E11.5 telencephalon, we quantified mitotic cells [Phospho-Histone H3 (pHH3)-positive] and intermediate progenitor cells (Tbr2/Eomes-positive), observing quantitative and qualitative shifts in these populations. We observed qualitative shifts in cortical layering at P0, particularly with Ctip2-positive cells in layer V. The results identify a suite of genes and functional gene networks that can be used to further dissect the role of Vegf in regulating NSC differentiation and downstream consequences for NSC fate decisions. PMID:24124161

  15. Mesenchymal stem cells expressing neural antigens instruct a neurogenic cell fate on neural stem cells.

    PubMed

    Croft, Adam P; Przyborski, Stefan A

    2009-04-01

    The neurogenic response to injury in the postnatal brain is limited and insufficient for restoration of function. Recent evidence suggests that transplantation of mesenchymal stem cells (MSCs) into the injured brain is associated with improved functional recovery, mediated in part through amplification in the endogenous neurogenic response to injury. In the current study we investigate the interactions between bone marrow-derived MSCs and embryonic neural stem cells (NSCs) plus their differentiated progeny using an in vitro co-culture system. Two populations of MSCs were used, MSCs induced to express neural antigens (nestin+, Tuj-1+, GFAP+) and neural antigen negative MSCs. Following co-culture of induced MSCs with differentiating NSC/progenitor cells a significant increase in Tuj-1+ neurons was detected compared to co-cultures of non-induced MSCs in which an increase in astrocyte (GFAP+) differentiation was observed. The effect was mediated by soluble interactions between the two cell populations and was independent of any effect on cell death and proliferation. Induced and non-induced MSCs also promoted the survival of Tuj-1+ cell progeny in long-term cultures and both promoted axonal growth, an effect also seen in differentiating neuroblastoma cells. Therefore, MSCs provide instructive signals that are able to direct the differentiation of NSCs and promote axonal development in neuronal progeny. The data indicates that the nature of MSC derived signals is dependent not only on their microenvironment but on the developmental status of the MSCs. Pre-manipulation of MSCs prior to transplantation in vivo may be an effective means of enhancing the endogenous neurogenic response to injury. PMID:19159625

  16. Synergistic effects of transplanted adult neural stem/progenitor cells, chondroitinase, and growth factors promote functional repair and plasticity of the chronically injured spinal cord.

    PubMed

    Karimi-Abdolrezaee, Soheila; Eftekharpour, Eftekhar; Wang, Jian; Schut, Desiree; Fehlings, Michael G

    2010-02-01

    The transplantation of neural stem/progenitor cells (NPCs) is a promising therapeutic strategy for spinal cord injury (SCI). However, to date NPC transplantation has exhibited only limited success in the treatment of chronic SCI. Here, we show that chondroitin sulfate proteoglycans (CSPGs) in the glial scar around the site of chronic SCI negatively influence the long-term survival and integration of transplanted NPCs and their therapeutic potential for promoting functional repair and plasticity. We targeted CSPGs in the chronically injured spinal cord by sustained infusion of chondroitinase ABC (ChABC). One week later, the same rats were treated with transplants of NPCs and transient infusion of growth factors, EGF, bFGF, and PDGF-AA. We demonstrate that perturbing CSPGs dramatically optimizes NPC transplantation in chronic SCI. Engrafted NPCs successfully integrate and extensively migrate within the host spinal cord and principally differentiate into oligodendrocytes. Furthermore, this combined strategy promoted the axonal integrity and plasticity of the corticospinal tract and enhanced the plasticity of descending serotonergic pathways. These neuroanatomical changes were also associated with significantly improved neurobehavioral recovery after chronic SCI. Importantly, this strategy did not enhance the aberrant synaptic connectivity of pain afferents, nor did it exacerbate posttraumatic neuropathic pain. For the first time, we demonstrate key biological and functional benefits for the combined use of ChABC, growth factors, and NPCs to repair the chronically injured spinal cord. These findings could potentially bring us closer to the application of NPCs for patients suffering from chronic SCI or other conditions characterized by the formation of a glial scar. PMID:20130176

  17. 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

  18. 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.

  19. 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

    2016-07-01

    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. PMID:27296463

  20. 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.

  1. Factors controlling cardiac neural crest cell migration

    PubMed Central

    Hutson, Mary R

    2010-01-01

    Cardiac neural crest cells originate as part of the postotic caudal rhombencephalic neural crest stream. Ectomesenchymal cells in this stream migrate to the circumpharyngeal ridge and then into the caudal pharyngeal arches where they condense to form first a sheath and then the smooth muscle tunics of the persisting pharyngeal arch arteries. A subset of the cells continues migrating into the cardiac outflow tract where they will condense to form the aorticopulmonary septum. Cell signaling, extracellular matrix and cell-cell contacts are all critical for the initial migration, pauses, continued migration and condensation of these cells. This Review elucidates what is currently known about these factors. PMID:20890117

  2. 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.

  3. Negative chemotaxis does not control quail neural crest cell dispersion.

    PubMed

    Erickson, C A; Olivier, K R

    1983-04-01

    Negative chemotaxis has been proposed to direct dispersion of amphibian neural crest cells away from the neural tube (V. C. Twitty, 1949, Growth 13(Suppl. 9), 133-161). We have reexamined this hypothesis using quail neural crest and do not find evidence for it. When pigmented or freshly isolated neural crest cells are covered by glass shards to prevent diffusion of a "putative" chemotactic agent away from the cells and into the medium, we find a decrease in density of cells beneath the coverslip as did Twitty and Niu (1948, J. Exp. Zool. 108, 405-437). Unlike those investigators, however, we find the covered cells move slower than uncovered cells and that the decrease in density can be attributed to cessation of cell division and increased cell death in older cultures, rather than directed migration away from each other. In cell systems where negative chemotaxis has been demonstrated, a "no man's land" forms between two confronted explants (Oldfield, 1963, Exp. Cell Res. 30, 125-138). No such cell-free space forms between confronted neural crest explants, even if the explants are closely covered to prevent diffusion of the negative chemotactic material. If crest cell aggregates are drawn into capillary tubes to allow accumulation of the putative material, the cells disperse farther, the wider the capillary tube bore. This is contrary to what would be expected if dispersion depended on accumulation of this material. Also, no difference in dispersion is noted between cells in the center of the tubes versus cells near the mouth of the tubes where the tube medium is freely exchanging with external fresh medium. Alternative hypotheses for directionality of crest migration in vivo are discussed. PMID:6832483

  4. Coxsackievirus A16 Infection Induces Neural Cell and Non-Neural Cell Apoptosis In Vitro

    PubMed Central

    Liu, Li; Wei, Zhenhong; Ehrlich, Elana S.; Liu, Guanchen; Li, Jingliang; Liu, Xin; Wang, Hong; Yu, Xiao-fang; Zhang, Wenyan

    2014-01-01

    Coxsackievirus A16 (CA16) is one of the main causative pathogens of hand, foot and mouth disease (HFMD). Viral replication typically results in host cell apoptosis. Although CA16 infection has been reported to induce apoptosis in the human rhabdomyosarcoma (RD) cell line, it remains unclear whether CA16 induces apoptosis in diverse cell types, especially neural cells which have important clinical significance. In the current study, CA16 infection was found to induce similar apoptotic responses in both neural cells and non-neural cells in vitro, including nuclear fragmentation, DNA fragmentation and phosphatidylserine translocation. CA16 generally is not known to lead to serious neurological symptoms in vivo. In order to further clarify the correlation between clinical symptoms and cell apoptosis, two CA16 strains from patients with different clinical features were investigated. The results showed that both CA16 strains with or without neurological symptoms in infected patients led to neural and muscle cell apoptosis. Furthermore, mechanistic studies showed that CA16 infection induced apoptosis through the same mechanism in both neural and non-neural cells, namely via activation of both the mitochondrial (intrinsic) pathway-related caspase 9 protein and the Fas death receptor (extrinsic) pathway-related caspase 8 protein. Understanding the mechanisms by which CA16 infection induces apoptosis in both neural and non-neural cells will facilitate a better understanding of CA16 pathogenesis. PMID:25350381

  5. 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.

  6. 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. PMID:26056571

  7. A phase field model for neural cell chemotropism

    NASA Astrophysics Data System (ADS)

    Najem, Sara; Grant, Martin

    2013-04-01

    Chemotropism is the action of targeting a part of the cell by means of chemical mediators and cues, and subsequently delimiting the pathway that it should undertake. In a neural cell, this initiates axonal elongation. Herein we model this growth, where chemotropic forcing leads the axon, by a phase field method utilizing two dynamical fields assigned respectively to the cell and to its leading edge. Additionally we quantify the condition for the retraction of the axon which takes place when the cell fails to form a synaptic connection.

  8. 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

  9. Monitoring cell growth.

    PubMed

    Strober, W

    2001-05-01

    This appendix provides two protocols for monitoring cell growth. Counting cells using a hemacytometer is tedious but it allows one to effectively distinguish live cells from dead cells (using Trypan Blue exclusion). In addition, this procedure is less subject to errors due to cell clumping or heterogeneity of cell size. The use of an electronic cell counter is quicker and easier than counting cells using a hemacytometer. However, an electronic cell counter as currently constructed does not distinguish live from dead cells in a reliable fashion and is subject to error due to the presence of cell clumps. Overall, the electronic cell counter is best reserved for repetitive and rapid counting of fresh peripheral blood cells and should be used with caution when counting cell populations derived from tissues. PMID:18432653

  10. Signaling mechanisms regulating adult neural stem cells and neurogenesis

    PubMed Central

    Faigle, Roland; Song, Hongjun

    2012-01-01

    Background Adult neurogenesis occurs throughout life in discrete regions of the mammalian brain and is tightly regulated via both extrinsic environmental influences and intrinsic genetic factors. In recent years, several crucial signaling pathways have been identified in regulating self-renewal, proliferation, and differentiation of neural stem cells, as well as migration and functional integration of developing neurons in the adult brain. Scope of review Here we review our current understanding of signaling mechanisms, including Wnt, notch, sonic hedgehog, growth and neurotrophic factors, bone morphogenetic proteins, neurotransmitters, transcription factors, and epigenetic modulators, and crosstalk between these signaling pathways in the regulation of adult neurogenesis. We also highlight emerging principles in the vastly growing field of adult neural stem cell biology and neural plasticity. Major conclusions Recent methodological advances have enabled the field to identify signaling mechanisms that fine-tune and coordinate neurogenesis in the adult brain, leading to a better characterization of both cell-intrinsic and environmental cues defining the neurogenic niche. Significant questions related to niche cell identity and underlying regulatory mechanisms remain to be fully addressed and will be the focus of future studies. General significance A full understanding of the role and function of individual signaling pathways in regulating neural stem cells and generation and integration of newborn neurons in the adult brain may lead to targeted new therapies for neurological diseases in humans. PMID:22982587

  11. Neural commitment of human pluripotent stem cells under defined conditions recapitulates neural development and generates patient-specific neural cells.

    PubMed

    Fernandes, Tiago G; Duarte, Sofia T; Ghazvini, Mehrnaz; Gaspar, Cláudia; Santos, Diana C; Porteira, Ana R; Rodrigues, Gonçalo M C; Haupt, Simone; Rombo, Diogo M; Armstrong, Judith; Sebastião, Ana M; Gribnau, Joost; Garcia-Cazorla, Àngels; Brüstle, Oliver; Henrique, Domingos; Cabral, Joaquim M S; Diogo, Maria Margarida

    2015-10-01

    Standardization of culture methods for human pluripotent stem cell (PSC) neural differentiation can greatly contribute to the development of novel clinical advancements through the comprehension of neurodevelopmental diseases. Here, we report an approach that reproduces neural commitment from human induced pluripotent stem cells using dual-SMAD inhibition under defined conditions in a vitronectin-based monolayer system. By employing this method it was possible to obtain neurons derived from both control and Rett syndrome patients' pluripotent cells. During differentiation mutated cells displayed alterations in the number of neuronal projections, and production of Tuj1 and MAP2-positive neurons. Although investigation of a broader number of patients would be required, these observations are in accordance with previous studies showing impaired differentiation of these cells. Consequently, our experimental methodology was proved useful not only for the generation of neural cells, but also made possible to compare neural differentiation behavior of different cell lines under defined culture conditions. This study thus expects to contribute with an optimized approach to study the neural commitment of human PSCs, and to produce patient-specific neural cells that can be used to gain a better understanding of disease mechanisms. PMID:26123315

  12. 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

  13. Neurogenesis of neural crest-derived periodontal ligament stem cells by EGF and bFGF.

    PubMed

    Fortino, Veronica R; Chen, Ren-Shiang; Pelaez, Daniel; Cheung, Herman S

    2014-04-01

    Neuroregenerative medicine is an ever-growing field in which regeneration of lost cells/tissues due to a neurodegenerative disease is the ultimate goal. With the scarcity of available replacement alternatives, stem cells provide an attractive source for regenerating neural tissue. While many stem cell sources exist, including: mesenchymal stem cells, embryonic stem cells, and induced pluripotent stem cells, the limited cellular potency, technical difficulties, and ethical considerations associated with these make finding alternate sources a desirable goal. Periodontal ligament stem cells (PDLSCs) derived from the neural crest were induced into neural-like cells using a combination of epidermal growth factor, and basic fibroblast growth factor. Morphological changes were evident in our treated group, seen under both light microscopy and scanning electron microscopy. A statistically significant increase in the expression of neuron-specific β-tubulin III and the neural stem/progenitor cell marker nestin, along with positive immunohistochemical staining for glial fibrillary acidic protein, demonstrated the success of our treatment in inducing both neuronal and glial phenotypes. Positive staining for synaptophysin demonstrated neural connections and electrophysiological recordings indicated that when subjected to whole-cell patch clamping, our treated cells displayed inward currents conducted through voltage-gated sodium (Na(+) ) channels. Taken together, our results indicate the success of our treatment in inducing PDLSCs to neural-like cells. The ease of sourcing and expansion, their embryologic neural crest origin, and the lack of ethical implications in their use make PDLSCs an attractive source for use in neuroregenerative medicine. PMID:24105823

  14. Enteric Neurospheres Are Not Specific to Neural Crest Cultures: Implications for Neural Stem Cell Therapies

    PubMed Central

    Cooper, Julie; Kronfli, Rania; Cananzi, Mara; Delalande, Jean-Marie; McCann, Conor; Burns, Alan J.; Thapar, Nikhil

    2015-01-01

    Objectives Enteric neural stem cells provide hope of curative treatment for enteric neuropathies. Current protocols for their harvesting from humans focus on the generation of ‘neurospheres’ from cultures of dissociated gut tissue. The study aims to better understand the derivation, generation and composition of enteric neurospheres. Design Gut tissue was obtained from Wnt1-Cre;Rosa26Yfp/Yfp transgenic mice (constitutively labeled neural crest cells) and paediatric patients. Gut cells were cultured either unsorted (mixed neural crest/non-neural crest), or following FACS selection into neural crest (murine-YFP+ve/human-p75+ve) or non-neural crest (YFP-ve/p75-ve) populations. Cultures and resultant neurospheres were characterized using immunolabelling in vitro and following transplantation in vivo. Results Cultures of (i) unsorted, (ii) neural crest, and (iii) non-neural crest cell populations generated neurospheres similar in numbers, size and morphology. Unsorted neurospheres were highly heterogeneous for neural crest content. Neural crest-derived (YFP+ve/p75+ve) neurospheres contained only neural derivatives (neurons and glia) and were devoid of non-neural cells (i.e. negative for SMA, c-Kit), with the converse true for non-neural crest-derived (YFP-ve/p75-ve) ‘neurospheres’. Under differentiation conditions only YFP+ve cells gave rise to neural derivatives. Both YFP+ve and YFP-ve cells displayed proliferation and spread upon transplantation in vivo, but YFP-ve cells did not locate or integrate within the host ENS. Conclusions Spherical accumulations of cells, so-called ‘neurospheres’ forming in cultures of dissociated gut contain variable proportions of neural crest-derived cells. If they are to be used for ENS cell replacement therapy then improved protocols for their generation, including cell selection, should be sought in order to avoid inadvertent transplantation of non-therapeutic, non-ENS cells. PMID:25799576

  15. 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

  16. Clinical translation of human neural stem cells.

    PubMed

    Tsukamoto, Ann; Uchida, Nobuko; Capela, Alexandra; Gorba, Thorsten; Huhn, Stephen

    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

  17. Neural crest cell evolution: how and when did a neural crest cell become a neural crest cell.

    PubMed

    Muñoz, William A; Trainor, Paul A

    2015-01-01

    As vertebrates evolved from protochordates, they shifted to a more predatory lifestyle, and radiated and adapted to most niches of the planet. This process was largely facilitated by the generation of novel vertebrate head structures, which were derived from neural crest cells (NCC). The neural crest is a unique vertebrate cell population that is frequently termed the "fourth germ layer" because it forms in conjunction with the other germ layers and contributes to a diverse array of cell types and tissues including the craniofacial skeleton, the peripheral nervous system, and pigment cells among many other tissues and cell types. NCC are defined by their origin at the neural plate border, via an epithelial-to-mesenchymal transition (EMT), together with multipotency and polarized patterns of migration. These defining characteristics, which evolved independently in the germ layers of invertebrates, were subsequently co-opted through their gene regulatory networks to form NCC in vertebrates. Moreover, recent data suggest that the ability to undergo an EMT was one of the latter features co-opted by NCC. In this review, we discuss the potential origins of NCC and how they evolved to contribute to nearly all tissues and organs throughout the body, based on paleontological evidence together with an evaluation of the evolution of molecules involved in NCC development and their migratory cell paths. PMID:25662256

  18. 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

  19. 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. PMID:26000486

  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. Isolation, culture and analysis of adult subependymal neural stem cells.

    PubMed

    Belenguer, Germán; Domingo-Muelas, Ana; Ferrón, Sacri R; Morante-Redolat, José Manuel; Fariñas, Isabel

    2016-01-01

    Individual cells dissected from the subependymal neurogenic niche of the adult mouse brain proliferate in medium containing basic fibroblast growth factor (bFGF) and/or epidermal growth factor (EGF) as mitogens, to produce multipotent clonal aggregates called neurospheres. These cultures constitute a powerful tool for the study of neural stem cells (NSCs) provided that they allow the analysis of their features and potential capacity in a controlled environment that can be modulated and monitored more accurately than in vivo. Clonogenic and population analyses under mitogen addition or withdrawal allow the quantification of the self-renewing and multilineage potency of these cells and the identification of the mechanisms involved in these properties. Here, we describe a set of procedures developed and/or modified by our group including several experimental options that can be used either independently or in combination for the ex vivo assessment of cell properties of NSCs obtained from the adult subependymal niche. PMID:27016251

  2. 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

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

    PubMed

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

    2015-12-17

    Neural sequences are a fundamental feature of brain dynamics underlying diverse behaviours, 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

  4. Mechanics of Cell Growth

    PubMed Central

    Ateshian, Gerard A.; Morrison, Barclay; Holmes, Jeffrey W.; Hung, Clark T.

    2012-01-01

    Cell growth describes an essential feature of biological tissues. This growth process may be modeled by using a set of relatively simple governing equations based on the axioms of mass and momentum balance, and using a continuum framework that describes cells and tissues as mixtures of a solid matrix, a solvent and multiple solutes. In this model the mechanics of cell growth is driven by osmotic effects, regulated by the cells’ active uptake of solutes and passive uptake of solvent. By accounting for the anisotropy of the cells’ cytoskeletal structures or extracellular matrix, as well as external constraints, a wide variety of growing shapes may be produced as illustrated in various examples. PMID:22904576

  5. Direct lineage reprogramming to neural cells

    PubMed Central

    Kim, Janghwan; Ambasudhan, Rajesh; Ding, Sheng

    2016-01-01

    Recently we have witnessed an array of studies on direct reprogramming that describe induced inter conversion of mature cell types from higher organisms including human. While these studies reveal an unexpected level of plasticity of differentiated somatic cells, they also provide unprecedented opportunities to develop regenerative therapies for many debilitating disorders and model these ‘diseases-in-a-dish’ for studying their pathophysiology. Here we review the current state of the art in direct lineage reprogramming to neural cells, and discuss the challenges that need to be addressed toward achieving the full potential of this exciting new technology. PMID:22652035

  6. 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

  7. 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 Central

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

    2008-01-01

    Background 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. Results 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-, S100β-, 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. Conclusion 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. PMID:18638414

  8. Regulation of neural stem cells by choroid plexus cells population.

    PubMed

    Roballo, Kelly C S; Gonçalves, Natalia J N; Pieri, Naira C G; Souza, Aline F; Andrade, André F C; Ambrósio, Carlos E

    2016-07-28

    The choroid plexus is a tissue on the central nervous system responsible for producing cerebrospinal fluid, maintaining homeostasis and neural stem cells support; though, all of its functions still unclear. This study aimed to demonstrate the niches of choroid plexus cells for a better understanding of the cell types and functions, using the porcine as the animal model. The collected material was analyzed by histology, immunohistochemistry, and cell culture. The cell culture was characterizated by immunocytochemistry and flow cytometry. Our results showed OCT-4, TUBIII, Nestin, CD45, CD73, CD90 positive expression and GFAP, CD105 negative expression, also methylene blue histological staining confirmed the presence of telocytes cells. We realized that the choroid plexus is a unique and incomparable tissue with different niches of cells as pluripotent, hematopoietic, neuronal progenitors and telocyte cells, which provide its complexity, differentiated functionality and responsibility on brain balance and neural stem cells regulation. PMID:27181512

  9. 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.

  10. FoxOs in neural stem cell fate decision.

    PubMed

    Ro, Seung-Hyun; Liu, Debra; Yeo, Hyeonju; Paik, Ji-hye

    2013-06-01

    Neural stem cells (NSCs) persist over the lifespan of mammals to give rise to committed progenitors and their differentiated cells in order to maintain the brain homeostasis. To this end, NSCs must be able to self-renew and otherwise maintain their quiescence. Suppression of aberrant proliferation or undesired differentiation is crucial to preclude either malignant growth or precocious depletion of NSCs. The PI3K-Akt-FoxO signaling pathway plays a central role in the regulation of multiple stem cells including one in the mammalian brain. In particular, members of FoxO family transcription factors are highly expressed in these stem cells. As an important downstream effector of growth, differentiation, and stress stimuli, mammalian FoxO transcription factor family controls cellular proliferation, oxidative stress response, homeostasis, and eventual maintenance of long-term repopulating potential. The review will focus on the current understanding of FoxO function in NSCs as well as discuss their biological activities that contribute to determining neural stem cell fate. PMID:22902436

  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. VEGF signals induce trailblazer cell identity that drives neural crest migration.

    PubMed

    McLennan, Rebecca; Schumacher, Linus J; Morrison, Jason A; Teddy, Jessica M; Ridenour, Dennis A; Box, Andrew C; Semerad, Craig L; Li, Hua; McDowell, William; Kay, David; Maini, Philip K; Baker, Ruth E; Kulesa, Paul M

    2015-11-01

    Embryonic neural crest cells travel in discrete streams to precise locations throughout the head and body. We previously showed that cranial neural crest cells respond chemotactically to vascular endothelial growth factor (VEGF) and that cells within the migratory front have distinct behaviors and gene expression. We proposed a cell-induced gradient model in which lead neural crest cells read out directional information from a chemoattractant profile and instruct trailers to follow. In this study, we show that migrating chick neural crest cells do not display distinct lead and trailer gene expression profiles in culture. However, exposure to VEGF in vitro results in the upregulation of a small subset of genes associated with an in vivo lead cell signature. Timed addition and removal of VEGF in culture reveals the changes in neural crest cell gene expression are rapid. A computational model incorporating an integrate-and-switch mechanism between cellular phenotypes predicts migration efficiency is influenced by the timescale of cell behavior switching. To test the model hypothesis that neural crest cellular phenotypes respond to changes in the VEGF chemoattractant profile, we presented ectopic sources of VEGF to the trailer neural crest cell subpopulation and show diverted cell trajectories and stream alterations consistent with model predictions. Gene profiling of trailer cells that diverted and encountered VEGF revealed upregulation of a subset of 'lead' genes. Injection of neuropilin1 (Np1)-Fc into the trailer subpopulation or electroporation of VEGF morpholino to reduce VEGF signaling failed to alter trailer neural crest cell trajectories, suggesting trailers do not require VEGF to maintain coordinated migration. These results indicate that VEGF is one of the signals that establishes lead cell identity and its chemoattractant profile is critical to neural crest cell migration. PMID:26278036

  13. Lessons from the embryonic neural stem cell niche for neural lineage differentiation of pluripotent stem cells.

    PubMed

    Solozobova, Valeriya; Wyvekens, Nicolas; Pruszak, Jan

    2012-09-01

    Pluripotent stem cells offer an abundant and malleable source for the generation of differentiated cells for transplantation as well as for in vitro screens. Patterning and differentiation protocols have been developed to generate neural progeny from human embryonic or induced pluripotent stem cells. However, continued refinement is required to enhance efficiency and to prevent the generation of unwanted cell types. We summarize and interpret insights gained from studies of embryonic neuroepithelium. A multitude of factors including soluble molecules, interactions with the extracellular matrix and neighboring cells cooperate to control neural stem cell self-renewal versus differentiation. Applying these findings and concepts to human stem cell systems in vitro may yield more appropriately patterned cell types for biomedical applications. PMID:22628111

  14. 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.

  15. 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. PMID:26478282

  16. Molecular Targeting of Neural Cancer Stem Cells: TTAGGG, You're It!

    PubMed Central

    Hjelmeland, Anita B.; Rich, Jeremy N.

    2011-01-01

    Summary Telomerase is an important mechanism by which cancers escape replicative senescence. In neural tumors, cancer stem cells express telomerase suggesting that this may explain their preferential tumorigenesis. Oligonucleotide telomerase targeting selectively disrupts cancer stem cell growth through the induction of differentiation, adding to the armamentarium of anti-cancer stem cell therapies. PMID:21208901

  17. Coupling actin dynamics to phase-field in modeling neural growth.

    PubMed

    Najem, Sara; Grant, Martin

    2015-06-14

    In this paper we model the growth of a neural cell together with the actin dynamics taking place at its growing region by constructing a phase-field model. This is done by assigning auxiliary fields to different constituents of the cell in order to differentiate them. Specifically, the inner and outer regions of the neural cell are described by ϕ = 1 and ϕ = 0 respectively, whereas the inside and outside of its leading edge are portrayed by ψ = 1 and ψ = 0. This formulation inherently locates the boundary, which is required to determine the evolution of the underlying actin dynamics. Therefore, it provides an alternative to boundary tracking algorithms. Then the equations governing the molecular workings of the cell specifically those of actin are modified in order to satisfy their corresponding boundary conditions. PMID:25943025

  18. 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. PMID:26577681

  19. 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

  20. Skeletal myogenic potential of human and mouse neural stem cells.

    PubMed

    Galli, R; Borello, U; Gritti, A; Minasi, M G; Bjornson, C; Coletta, M; Mora, M; De Angelis, M G; Fiocco, R; Cossu, G; Vescovi, A L

    2000-10-01

    Distinct cell lineages established early in development are usually maintained throughout adulthood. Thus, adult stem cells have been thought to generate differentiated cells specific to the tissue in which they reside. This view has been challenged; for example, neural stem cells can generate cells that normally originate from a different germ layer. Here we show that acutely isolated and clonally derived neural stem cells from mice and humans could produce skeletal myotubes in vitro and in vivo, the latter following transplantation into adult animals. Myogenic conversion in vitro required direct exposure to myoblasts, and was blocked if neural cells were clustered. Thus, a community effect between neural cells may override such myogenic induction. We conclude that neural stem cells, which generate neurons, glia and blood cells, can also produce skeletal muscle cells, and can undergo various patterns of differentiation depending on exposure to appropriate epigenetic signals in mature tissues. PMID:11017170

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

    PubMed Central

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

    2016-01-01

    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. PMID:27073384

  2. 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

  3. 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. PMID:26494823

  4. Adult neural stem cells stake their ground

    PubMed Central

    Lim, Daniel A.; Alvarez-Buylla, Arturo

    2014-01-01

    The birth of new neurons in the walls of the adult brain lateral ventricles has captured the attention of many neuroscientists for over two decades, yielding key insights into the identity and regulation of neural stem cells (NSCs). In the adult ventricular-subventricular zone (V-SVZ), NSCs are a specialized form of astrocyte that generates several types of neurons for the olfactory bulb. Here we discuss recent findings regarding the unique organization of the V-SVZ NSCs niche, the multiple regulatory controls of neuronal production, the distinct regional identities of adult NSCs, and the epigenetic mechanisms that maintain adult neurogenesis. Understanding how V-SVZ NSCs establish and maintain lifelong neurogenesis continues to provide surprising insights into the cellular and molecular regulation of neural development. PMID:25223700

  5. 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

  6. Neural stem cells: Brain building blocks and beyond

    PubMed Central

    Bergström, Tobias

    2012-01-01

    Neural stem cells are the origins of neurons and glia and generate all the differentiated neural cells of the mammalian central nervous system via the formation of intermediate precursors. Although less frequent, neural stem cells persevere in the postnatal brain where they generate neurons and glia. Adult neurogenesis occurs throughout life in a few limited brain regions. Regulation of neural stem cell number during central nervous system development and in adult life is associated with rigorous control. Failure in this regulation may lead to e.g. brain malformation, impaired learning and memory, or tumor development. Signaling pathways that are perturbed in glioma are the same that are important for neural stem cell self-renewal, differentiation, survival, and migration. The heterogeneity of human gliomas has impeded efficient treatment, but detailed molecular characterization together with novel stem cell-like glioma cell models that reflect the original tumor gives opportunities for research into new therapies. The observation that neural stem cells can be isolated and expanded in vitro has opened new avenues for medical research, with the hope that they could be used to compensate the loss of cells that features in several severe neurological diseases. Multipotent neural stem cells can be isolated from the embryonic and adult brain and maintained in culture in a defined medium. In addition, neural stem cells can be derived from embryonic stem cells and induced pluripotent stem cells by in vitro differentiation, thus adding to available models to study stem cells in health and disease. PMID:22512245

  7. Evaluation of neural gene expression in serum treated embryonic stem cells in Alzheimer's patients

    PubMed Central

    Dehghani, Leila; Hashemi-Beni, Batool; Poorazizi, Elahe; Khorvash, Fariborz; Shaygannejad, Vahid; Sedghi, Maryam; Vesal, Sahar; Meamar, Rokhsareh

    2013-01-01

    Background: Previous studies confirmed that neural gene expression in embryonic stem cells (ESC) could influence by chemical compounds through stimulating apoptotic pathway. We aimed to use ESCs-derived neural cells by embryoid body formation as an in vitro model for determination of neural gene expression changes in groups that treated by sera from Alzheimer's patients and compare with healthy individuals. Materials and Methods: ESC line which was derived from the C57BL/6 mouse strain was used throughout this study. ESC-derived neural cells were treated with serum from Alzheimer's patient and healthy individual. Neural gene expression was assessed in both groups by quantitative real-time polymerase chain reaction analysis. The data was analyzed by SPSS Software (version 18). Results: Morphologically, the reducing in neurite out-growth was observed in neural cells in group, which treated by serum from Alzheimer's patient, while neurite growth was natural in appearance in control group. Microtubule-associated protein 2 and glial fibrillary acidic protein expression significantly reduced in the Alzheimer's patient group compared with the control group. Nestin expression did not significantly differ among the groups. Conclusion: Neural gene expression could be reduced in serum treated ESC in Alzheimer's patients. PMID:23961278

  8. [Neural stem cells and Notch signalling].

    PubMed

    Traiffort, Elisabeth; Ferent, Julien

    2015-12-01

    Development and repair of the nervous system are based on the existence of neural stem cells (NSCs) able to generate neurons and glial cells. Among the mechanisms that are involved in the control of embryo or adult NSCs, the Notch signalling plays a major role. In embryo, the pathway participates in the maintenance of NSCs during all steps of development of the central nervous system which starts with the production of neurons also called neurogenesis and continues with gliogenesis giving rise to astrocytes and oligodendrocytes. During the postnatal and adult period, Notch signalling is still present in the major neurogenic areas, the subventricular zone of the lateral ventricles and the subgranular zone of the hippocampus. In these regions, Notch maintains NSC quiescence, contributes to the heterogeneity of these cells and displays pleiotropic effects during the regeneration process occurring after a lesion. PMID:26672665

  9. Neurogenesis of Neural Crest Derived Periodontal Ligament Stem Cells by EGF and bFGF

    PubMed Central

    Fortino, Veronica R.; Chen, Ren-Shiang; Pelaez, Daniel; Cheung, Herman S.

    2013-01-01

    Neuroregenerative medicine is an ever-growing field in which regeneration of lost cells/tissues due to a neurodegenerative disease is the ultimate goal. With the scarcity of available replacement alternatives, stem cells provide an attractive source for regenerating neural tissue. While many stem cell sources exist, including: mesenchymal stem cells (MSCs), embryonic stem cells (ESCs), and induced pluripotent stem cells (iPSCs), the limited cellular potency, technical difficulties, and ethical considerations associated with these make finding alternate sources a desirable goal. Periodontal ligament stem cells (PDLSCs) derived from the neural crest were induced into neural-like cells using a combination of epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF). Morphological changes were evident in our treated group, seen under both light microscopy and scanning electron microscopy (SEM). A statistically significant increase in the expression of neuron-specific β-tubulin III and the neural stem/progenitor cell marker nestin, along with positive immunohistochemical staining for glial fibrillary acidic protein (GFAP), demonstrated the success of our treatment in inducing both neuronal and glial phenotypes. Positive staining for synaptophysin demonstrated neural connections and electrophysiological recordings indicated that when subjected to whole cell patch clamping, our treated cells displayed inward currents conducted through voltage-gated sodium (Na+) channels. Taken together, our results indicate the success of our treatment in inducing PDLSCs to neural-like cells. The ease of sourcing and expansion, their embryologic neural crest origin, and the lack of ethical implications in their use make PDLSCs an attractive source for use in neuroregenerative medicine. PMID:24105823

  10. Stat3 inhibition in neural lineage cells.

    PubMed

    Chiba, Tomohiro; Mack, Laura; Delis, Natalia; Brill, Boris; Groner, Bernd

    2012-06-01

    Abstract Deregulation of signal transducer and activator of transcription 3 (Stat3) is attracting attentions in neurological disorders of elderly populations, e.g., Stat3 is inactivated in hippocampal neurons of Alzheimer's disease (AD) brains, whereas it is often constitutively activated in glioblastoma multiforme (GBM), correlating with poor prognosis. Stat3-inhibiting drugs have been intensively developed for chemotherapy based on the fact that GBM, in many cases, are "addicted" to Stat3 activation. Stat3 inhibitors, however, potentially have unfavorable side effects on postmitotic neurons, normal permanent residents in the central nervous system. It is, therefore, of great importance to address detailed cellular responses of neural lineage cells including normal neurons, astrocytes, and neuronal/glial cancer cell lines to several classes of Stat3 inhibitors focusing on their effective concentrations. Here, we picked up five human and mouse cancer cell lines (Neuro-2a and SH-SY5Y neuroblastoma cell lines and Tu-9648, U-87MG, and U-373MG glioblastoma cell lines) and treated with various Stat3 inhibitors. Among them, Stattic, FLLL31, and resveratrol potently suppressed P-Stat3 and cell viability in all the tested cell lines. Stat3 knockdown or expression of dominant-negative Stat3 further sensitized cells to the inhibitors. Expression of familial AD-related mutant amyloid precursor protein sensitized neuronal cells, not glial cells, to Stat3 inhibitors by reducing P-Stat3 levels. Primary neurons and astrocytes also responded to Stat3 inhibitors with similar sensitivities to those observed in cancer cell lines. Thus, Stat3 inhibitors should be carefully targeted to GBM cells to avoid potential neurotoxicity leading to AD-like neuropsychiatric dysfunctions. PMID:25436682

  11. Zika Virus Infects Human Cortical Neural Progenitors and Attenuates Their Growth.

    PubMed

    Tang, Hengli; Hammack, Christy; Ogden, Sarah C; Wen, Zhexing; Qian, Xuyu; Li, Yujing; Yao, Bing; Shin, Jaehoon; Zhang, Feiran; Lee, Emily M; Christian, Kimberly M; Didier, Ruth A; Jin, Peng; Song, Hongjun; Ming, Guo-Li

    2016-05-01

    The suspected link between infection by Zika virus (ZIKV), a re-emerging flavivirus, and microcephaly is an urgent global health concern. The direct target cells of ZIKV in the developing human fetus are not clear. Here we show that a strain of the ZIKV, MR766, serially passaged in monkey and mosquito cells efficiently infects human neural progenitor cells (hNPCs) derived from induced pluripotent stem cells. Infected hNPCs further release infectious ZIKV particles. Importantly, ZIKV infection increases cell death and dysregulates cell-cycle progression, resulting in attenuated hNPC growth. Global gene expression analysis of infected hNPCs reveals transcriptional dysregulation, notably of cell-cycle-related pathways. Our results identify hNPCs as a direct ZIKV target. In addition, we establish a tractable experimental model system to investigate the impact and mechanism of ZIKV on human brain development and provide a platform to screen therapeutic compounds. PMID:26952870

  12. Imprinted Zac1 in neural stem cells

    PubMed Central

    Daniel, Guillaume; Schmidt-Edelkraut, Udo; Spengler, Dietmar; Hoffmann, Anke

    2015-01-01

    Neural stem cells (NSCs) and imprinted genes play an important role in brain development. On historical grounds, these two determinants have been largely studied independently of each other. Recent evidence suggests, however, that NSCs can reset select genomic imprints to prevent precocious depletion of the stem cell reservoir. Moreover, imprinted genes like the transcriptional regulator Zac1 can fine tune neuronal vs astroglial differentiation of NSCs. Zac1 binds in a sequence-specific manner to pro-neuronal and imprinted genes to confer transcriptional regulation and furthermore coregulates members of the p53-family in NSCs. At the genome scale, Zac1 is a central hub of an imprinted gene network comprising genes with an important role for NSC quiescence, proliferation and differentiation. Overall, transcriptional, epigenomic, and genomic mechanisms seem to coordinate the functional relationships of NSCs and imprinted genes from development to maturation, and possibly aging. PMID:25815116

  13. Neural stem cell therapy for cancer.

    PubMed

    Bagó, Juli Rodriguez; Sheets, Kevin T; Hingtgen, Shawn D

    2016-04-15

    Cancers of the brain remain one of the greatest medical challenges. Traditional surgery and chemo-radiation therapy are unable to eradicate diffuse cancer cells and tumor recurrence is nearly inevitable. In contrast to traditional regenerative medicine applications, engineered neural stem cells (NSCs) are emerging as a promising new therapeutic strategy for cancer therapy. The tumor-homing properties allow NSCs to access both primary and invasive tumor foci, creating a novel delivery platform. NSCs engineered with a wide array of cytotoxic agents have been found to significantly reduce tumor volumes and markedly extend survival in preclinical models. With the recent launch of new clinical trials, the potential to successfully manage cancer in human patients with cytotoxic NSC therapy is moving closer to becoming a reality. PMID:26314280

  14. Human embryonic stem cell differentiation toward regional specific neural precursors.

    PubMed

    Erceg, Slaven; Ronaghi, Mohammad; Stojković, Miodrag

    2009-01-01

    Human embryonic stem cells (hESCs) are self-renewing pluripotent cells that have the capacity to differentiate into a wide variety of cell types. This potentiality represents a promising source to overcome many human diseases by providing an unlimited supply of all cell types, including cells with neural characteristics. Therefore, this review summarizes early neural development and the potential of hESCs to differentiate under in vitro conditions, examining at the same time the potential use of differentiated hESCs for therapeutic applications for neural tissue and cell regeneration. PMID:18845761

  15. Human Embryonic Stem Cell Differentiation Toward Regional Specific Neural Precursors

    PubMed Central

    Erceg, Slaven; Ronaghi, Mohammad; Stojković, Miodrag

    2009-01-01

    Human embryonic stem cells (hESCs) are self-renewing pluripotent cells that have the capacity to differentiate into a wide variety of cell types. This potentiality represents a promising source to overcome many human diseases by providing an unlimited supply of all cell types, including cells with neural characteristics. Therefore, this review summarizes early neural development and the potential of hESCs to differentiate under in vitro conditions, examining at the same time the potential use of differentiated hESCs for therapeutic applications for neural tissue and cell regeneration. PMID:18845761

  16. 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. PMID:12186855

  17. 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

  18. Neural stem and progenitor cells in health and disease

    PubMed Central

    Ladran, Ian; Tran, Ngoc; Topol, Aaron; Brennand, Kristen J.

    2014-01-01

    Neural stem/progenitor cells (NSPCs) have the potential to differentiate into neurons, astrocytes, and/or oligodendrocytes. Because these cells can be expanded in culture, they represent a vast source of neural cells. With the recent discovery that patient fibroblasts can be reprogrammed directly into induced NSPCs, the regulation of NSPC fate and function, in the context of cell-based disease models and patient-specific cell-replacement therapies, warrants review. PMID:24068527

  19. A Supramolecular Gel Approach to Minimize the Neural Cell Damage during Cryopreservation Process.

    PubMed

    Zeng, Jie; Yin, Yixia; Zhang, Li; Hu, Wanghui; Zhang, Chaocan; Chen, Wanyu

    2016-03-01

    The storage method for living cells is one of the major challenges in cell-based applications. Here, a novel supramolecular gel cryopreservation system (BDTC gel system) is introduced, which can observably increase the neural cell viability during cryopreservation process because this system can (1) confine the ice crystal growth in the porous of BDTC gel system, (2) decrease the amount of ice crystallization and cryopreservation system's freezing point, and (3) reduce the change rates of cell volumes and osmotic shock. In addition, thermoreversible BDTC supramolecular gel is easy to be removed after thawing so it does not hinder the adherence, growth, and proliferation of cells. The results of functionality assessments indicate that BDTC gel system can minimize the neural cell damage during cryopreservation process. This method will be potentially applied in cryopreservation of other cell types, tissues, or organs and will benefit cell therapy, tissue engineering, and organs transplantation. PMID:26611502

  20. A probabilistic neural network approach for modeling and classification of bacterial growth/no-growth data.

    PubMed

    Hajmeer, M; Basheer, I

    2002-10-01

    In this paper, we propose to use probabilistic neural networks (PNNs) for classification of bacterial growth/no-growth data and modeling the probability of growth. The PNN approach combines both Bayes theorem of conditional probability and Parzen's method for estimating the probability density functions of the random variables. Unlike other neural network training paradigms, PNNs are characterized by high training speed and their ability to produce confidence levels for their classification decision. As a practical application of the proposed approach, PNNs were investigated for their ability in classification of growth/no-growth state of a pathogenic Escherichia coli R31 in response to temperature and water activity. A comparison with the most frequently used traditional statistical method based on logistic regression and multilayer feedforward artificial neural network (MFANN) trained by error backpropagation was also carried out. The PNN-based models were found to outperform linear and nonlinear logistic regression and MFANN in both the classification accuracy and ease by which PNN-based models are developed. PMID:12133614

  1. Data defining markers of human neural stem cell lineage potential.

    PubMed

    Oikari, Lotta E; Okolicsanyi, Rachel K; Griffiths, Lyn R; Haupt, Larisa M

    2016-06-01

    Neural stem cells (NSCs) and neural progenitor cells (NPCs) are self-renewing and multipotent cells, however, NPCs are considered to be more lineage-restricted with a reduced self-renewing capacity. We present data comparing the expression of 21 markers encompassing pluripotency, self-renewal (NSC) as well as neuronal and glial (astrocyte and oligodendrocyte) lineage specification and 28 extracellular proteoglycan (PG) genes and their regulatory enzymes between embryonic stem cell (ESC)-derived human NSCs (hNSC H9 cells, Thermo Fisher) and human cortex-derived normal human NPCs (nhNPCs, Lonza). The data demonstrates expression differences of multiple lineage and proteoglycan-associated genes between hNSC H9 cells and nhNPCs. Data interpretation of markers and proteoglycans defining NSC and neural cell lineage characterisation can be found in "Cell surface heparan sulfate proteoglycans as novel markers of human neural stem cell fate determination" (Oikari et al. 2015) [1]. PMID:26958640

  2. Data defining markers of human neural stem cell lineage potential

    PubMed Central

    Oikari, Lotta E.; Okolicsanyi, Rachel K.; Griffiths, Lyn R.; Haupt, Larisa M.

    2016-01-01

    Neural stem cells (NSCs) and neural progenitor cells (NPCs) are self-renewing and multipotent cells, however, NPCs are considered to be more lineage-restricted with a reduced self-renewing capacity. We present data comparing the expression of 21 markers encompassing pluripotency, self-renewal (NSC) as well as neuronal and glial (astrocyte and oligodendrocyte) lineage specification and 28 extracellular proteoglycan (PG) genes and their regulatory enzymes between embryonic stem cell (ESC)-derived human NSCs (hNSC H9 cells, Thermo Fisher) and human cortex-derived normal human NPCs (nhNPCs, Lonza). The data demonstrates expression differences of multiple lineage and proteoglycan-associated genes between hNSC H9 cells and nhNPCs. Data interpretation of markers and proteoglycans defining NSC and neural cell lineage characterisation can be found in “Cell surface heparan sulfate proteoglycans as novel markers of human neural stem cell fate determination” (Oikari et al. 2015) [1]. PMID:26958640

  3. Reprogramming of avian neural crest axial identity and cell fate.

    PubMed

    Simoes-Costa, Marcos; Bronner, Marianne E

    2016-06-24

    Neural crest populations along the embryonic body axis of vertebrates differ in developmental potential and fate, so that only the cranial neural crest can contribute to the craniofacial skeleton in vivo. We explored the regulatory program that imbues the cranial crest with its specialized features. Using axial-level specific enhancers to isolate and perform genome-wide profiling of the cranial versus trunk neural crest in chick embryos, we identified and characterized regulatory relationships between a set of cranial-specific transcription factors. Introducing components of this circuit into neural crest cells of the trunk alters their identity and endows these cells with the ability to give rise to chondroblasts in vivo. Our results demonstrate that gene regulatory circuits that support the formation of particular neural crest derivatives may be used to reprogram specific neural crest-derived cell types. PMID:27339986

  4. 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

  5. Neural-Induced Human Mesenchymal Stem Cells Promote Cochlear Cell Regeneration in Deaf Guinea Pigs

    PubMed Central

    Jang, Sujeong; Cho, Hyong-Ho; Kim, Song-Hee; Lee, Kyung-Hwa; Jun, Jae Yeoul; Park, Jong-Seong; Jeong, Han-Seong

    2015-01-01

    Objectives In mammals, cochlear hair cell loss is irreversible and may result in a permanent sensorineural hearing loss. Secondary to this hair cell loss, a progressive loss of spiral ganglion neurons (SGNs) is presented. In this study, we have investigated the effects of neural-induced human mesenchymal stem cells (NI-hMSCs) from human bone marrow on sensory neuronal regeneration from neomycin treated deafened guinea pig cochleae. Methods HMSCs were isolated from the bone marrow which was obtained from the mastoid process during mastoidectomy for ear surgery. Following neural induction with basic fibroblast growth factor and forskolin, we studied the several neural marker and performed electrophysiological analysis. NI-hMSCs were transplanted into the neomycin treated deafened guinea pig cochlea. Engraftment of NI-hMSCs was evaluated immunohistologically at 8 weeks after transplantation. Results Following neural differentiation, hMSCs expressed high levels of neural markers, ionic channel markers, which are important in neural function, and tetrodotoxin-sensitive voltage-dependent sodium currents. After transplantation into the scala tympani of damaged cochlea, NI-hMSCs-injected animals exhibited a significant increase in the number of SGNs compared to Hanks balanced salt solution-injected animals. Transplanted NI-hMSCs were found within the perilymphatic space, the organ of Corti, along the cochlear nerve fibers, and in the spiral ganglion. Furthermore, the grafted NI-hMSCs migrated into the spiral ganglion where they expressed the neuron-specific marker, NeuN. Conclusion The results show the potential of NI-hMSCs to give rise to replace the lost cochlear cells in hearing loss mammals. PMID:26045904

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

    PubMed Central

    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

    2015-01-01

    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. PMID:25799239

  7. 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. PMID:22683799

  8. SIRT1 and Neural Cell Fate Determination.

    PubMed

    Cai, Yulong; Xu, Le; Xu, Haiwei; Fan, Xiaotang

    2016-07-01

    During the development of the central nervous system (CNS), neurons and glia are derived from multipotent neural stem cells (NSCs) undergoing self-renewal. NSC commitment and differentiation are tightly controlled by intrinsic and external regulatory mechanisms in space- and time-related fashions. SIRT1, a silent information regulator 2 (Sir2) ortholog, is expressed in several areas of the brain and has been reported to be involved in the self-renewal, multipotency, and fate determination of NSCs. Recent studies have highlighted the role of the deacetylase activity of SIRT1 in the determination of the final fate of NSCs. This review summarizes the roles of SIRT1 in the expansion and differentiation of NSCs, specification of neuronal subtypes and glial cells, and reprogramming of functional neurons from embryonic stem cells and fibroblasts. This review also discusses potential signaling pathways through which SIRT1 can exhibit versatile functions in NSCs to regulate the cell fate decisions of neurons and glia. PMID:25850787

  9. Ulk4 Regulates Neural Stem Cell Pool.

    PubMed

    Liu, Min; Guan, Zhenlong; Shen, Qin; Flinter, Frances; Domínguez, Laura; Ahn, Joo Wook; Collier, David A; O'Brien, Timothy; Shen, Sanbing

    2016-09-01

    The size of neural stem cell (NSC) pool at birth determines the starting point of adult neurogenesis. Aberrant neurogenesis is associated with major mental illness, in which ULK4 is proposed as a rare risk factor. Little is known about factors regulating the NSC pool, or function of the ULK4. Here, we showed that Ulk4(tm1a/tm1a) mice displayed a dramatically reduced NSC pool at birth. Ulk4 was expressed in a cell cycle-dependent manner and peaked in G2/M phases. Targeted disruption of the Ulk4 perturbed mid-neurogenesis and significantly reduced cerebral cortex in postnatal mice. Pathway analyses of dysregulated genes in Ulk4(tm1a/tm1a) mice revealed Ulk4 as a key regulator of cell cycle and NSC proliferation, partially through regulation of the Wnt signaling. In addition, we identified hemizygous deletion of ULK4 gene in 1.2/1,000 patients with pleiotropic symptoms including severe language delay and learning difficulties. ULK4, therefore, may significantly contribute to neurodevelopmental, neuropsychiatric, and neurodegenerative disorders. Stem Cells 2016;34:2318-2331. PMID:27300315

  10. 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.

  11. Neural tube defects and impaired neural progenitor cell proliferation in Gbeta1-deficient mice.

    PubMed

    Okae, Hiroaki; Iwakura, Yoichiro

    2010-04-01

    Heterotrimeric G proteins are well known for their roles in signal transduction downstream of G protein-coupled receptors (GPCRs), and both Galpha subunits and tightly associated Gbetagamma subunits regulate downstream effector molecules. Compared to Galpha subunits, the physiological roles of individual Gbeta and Ggamma subunits are poorly understood. In this study, we generated mice deficient in the Gbeta1 gene and found that Gbeta1 is required for neural tube closure, neural progenitor cell proliferation, and neonatal development. About 40% Gbeta1(-/-) embryos developed neural tube defects (NTDs) and abnormal actin organization was observed in the basal side of neuroepithelium. In addition, Gbeta1(-/-) embryos without NTDs showed microencephaly and died within 2 days after birth. GPCR agonist-induced ERK phosphorylation, cell proliferation, and cell spreading, which were all found to be regulated by Galphai and Gbetagamma signaling, were abnormal in Gbeta1(-/-) neural progenitor cells. These data indicate that Gbeta1 is required for normal embryonic neurogenesis. PMID:20186915

  12. Transplantation of Neural Stem Cells Cultured in Alginate Scaffold for Spinal Cord Injury in Rats

    PubMed Central

    Sharafkhah, Ali; Koohi-Hosseinabadi, Omid; Semsar-Kazerooni, Maryam

    2016-01-01

    Study Design This study investigated the effects of transplantation of alginate encapsulated neural stem cells (NSCs) on spinal cord injury in Sprague-Dawley male rats. The neurological functions were assessed for 6 weeks after transplantation along with a histological study and measurement of caspase-3 levels. Purpose The aim of this study was to discover whether NSCs cultured in alginate transplantation improve recovery from spinal cord injury. Overview of Literature Spinal cord injury is one of the leading causes of disability and it has no effective treatment. Spinal cord injury can also cause sensory impairment. With an impetus on using stem cells therapy in various central nervous system settings, there is an interest in using stem cells for addressing spinal cord injury. Neural stem cell is one type of stem cells that is able to differentiate to all three neural lineages and it shows promise in spinal injury treatment. Furthermore, a number of studies have shown that culturing NSCs in three-dimensional (3D) scaffolds like alginate could enhance neural differentiation. Methods The NSCs were isolated from 14-day-old rat embryos. The isolated NSCs were cultured in growth media containing basic fibroblast growth factor and endothelial growth factor. The cells were characterized by differentiating to three neural lineages and they were cultured in an alginate scaffold. After 7 days the cells were encapsulated and transplanted in a rat model of spinal cord injury. Results Our data showed that culturing in an alginate 3D scaffold and transplantation of the NSCs could improve neurological outcome in a rat model of spinal cord injury. The inflammation scores and lesion sizes and also the activity of caspase-3 (for apoptosis evaluation) were less in encapsulated neural stem cell transplantation cases. Conclusions Transplantation of NSCs that were cultured in an alginate scaffold led to a better clinical and histological outcome for recovery from spinal cord injury in

  13. Structural and molecular micropatterning of dual hydrogel constructs for neural growth models using photochemical strategies

    PubMed Central

    Horn-Ranney, Elaine L.; Curley, J. Lowry; Catig, Gary C.; Huval, Renee M.; Moore, Michael J.

    2013-01-01

    Chemotactic and haptotactic cues guide neurite growth toward appropriate targets by eliciting attractive or repulsive responses from the neurite growth cones. Here we present an integrated system allowing both structural and molecular micropatterning in dual hydrogel 3D tissue culture constructs for directing in vitro neuronal growth via structural, immobilized, and soluble guidance cues. These tissue culture constructs were fabricated into specifiable geometries using UV light reflected from a digital micromirror device acting as a dynamic photomask, resulting in dual hydrogel constructs consisting of a cell growth-restrictive polyethylene glycol (PEG) boundary with a cell growth-permissive interior of photolabile α-carboxy-2-nitrobenzyl cysteine agarose (CNBC-A). This CNBC-A was irradiated in discrete areas and subsequently tagged with maleimide-conjugated biomolecules. Fluorescent microscopy showed biomolecule binding only at the sites of irradiation in CNBC-A, and confocal microscopy confirmed 3D binding through the depth of the construct. Neurite outgrowth studies showed contained growth throughout CNBC-A. The diffusion rate of soluble fluorescein-bovine serum albumin through the dual hydrogel construct was controlled by PEG concentration and the distance between the protein source and the agarose interior; the timescale for a transient protein gradient changed with these parameters. These findings suggest the dual hydrogel system is a usefulplatform for manipulating a 3D in vitro microenvironment with patterned structural and molecular guidance cues for modeling neural growth and guidance. PMID:22903647

  14. The Role of Direct Current Electric Field-Guided Stem Cell Migration in Neural Regeneration.

    PubMed

    Yao, Li; Li, Yongchao

    2016-06-01

    Effective directional axonal growth and neural cell migration are crucial in the neural regeneration of the central nervous system (CNS). Endogenous currents have been detected in many developing nervous systems. Experiments have demonstrated that applied direct current (DC) electric fields (EFs) can guide axonal growth in vitro, and attempts have been made to enhance the regrowth of damaged spinal cord axons using DC EFs in in vivo experiments. Recent work has revealed that the migration of stem cells and stem cell-derived neural cells can be guided by DC EFs. These studies have raised the possibility that endogenous and applied DC EFs can be used to direct neural tissue regeneration. Although the mechanism of EF-directed axonal growth and cell migration has not been fully understood, studies have shown that the polarization of cell membrane proteins and the activation of intracellular signaling molecules are involved in the process. The application of EFs is a promising biotechnology for regeneration of the CNS. PMID:27108005

  15. ZDHHC3 Tyrosine Phosphorylation Regulates Neural Cell Adhesion Molecule Palmitoylation.

    PubMed

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

    2016-09-01

    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

  16. Stem cell-based therapy in neural repair.

    PubMed

    Hsu, Yi-Chao; Chen, Su-Liang; Wang, Dan-Yen; Chiu, Ing-Ming

    2013-01-01

    Cell-based therapy could aid in alleviating symptoms or even reversing the progression of neurodegenerative diseases and nerve injuries. Fibroblast growth factor 1 (FGF1) has been shown to maintain the survival of neurons and induce neurite outgrowth. Accumulating evidence suggests that combination of FGF1 and cell-based therapy is promising for future therapeutic application. Neural stem cells (NSCs), with the characteristics of self-renewal and multipotency, can be isolated from embryonic stem cells, embryonic ectoderm, and developing or adult brain tissues. For NSC clinical application, several critical problems remain to be resolved: (1) the source of NSCs should be personalized; (2) the isolation methods and protocols of human NSCs should be standardized; (3) the clinical efficacy of NSC transplants must be evaluated in more adequate animal models; and (4) the mechanism of intrinsic brain repair needs to be better characterized. In addition, the ideal imaging technique for tracking NSCs would be safe and yield high temporal and spatial resolution, good sensitivity and specificity. Here, we discuss recent progress and future development of cell-based therapy, such as NSCs, induced pluripotent stem cells, and induced neurons, in neurodegenerative diseases and peripheral nerve injuries. PMID:23806879

  17. 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

  18. Meninges harbor cells expressing neural precursor markers during development and adulthood

    PubMed Central

    Bifari, Francesco; Berton, Valeria; Pino, Annachiara; Kusalo, Marijana; Malpeli, Giorgio; Di Chio, Marzia; Bersan, Emanuela; Amato, Eliana; Scarpa, Aldo; Krampera, Mauro; Fumagalli, Guido; Decimo, Ilaria

    2015-01-01

    Brain and skull developments are tightly synchronized, allowing the cranial bones to dynamically adapt to the brain shape. At the brain-skull interface, meninges produce the trophic signals necessary for normal corticogenesis and bone development. Meninges harbor different cell populations, including cells forming the endosteum of the cranial vault. Recently, we and other groups have described the presence in meninges of a cell population endowed with neural differentiation potential in vitro and, after transplantation, in vivo. However, whether meninges may be a niche for neural progenitor cells during embryonic development and in adulthood remains to be determined. In this work we provide the first description of the distribution of neural precursor markers in rat meninges during development up to adulthood. We conclude that meninges share common properties with the classical neural stem cell niche, as they: (i) are a highly proliferating tissue; (ii) host cells expressing neural precursor markers such as nestin, vimentin, Sox2 and doublecortin; and (iii) are enriched in extracellular matrix components (e.g., fractones) known to bind and concentrate growth factors. This study underlines the importance of meninges as a potential niche for endogenous precursor cells during development and in adulthood. PMID:26483637

  19. 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. PMID:26890524

  20. Disentangling neural cell diversity using single-cell transcriptomics.

    PubMed

    Poulin, Jean-Francois; Tasic, Bosiljka; Hjerling-Leffler, Jens; Trimarchi, Jeffrey M; Awatramani, Rajeshwar

    2016-08-26

    Cellular specialization is particularly prominent in mammalian nervous systems, which are composed of millions to billions of neurons that appear in thousands of different 'flavors' and contribute to a variety of functions. Even in a single brain region, individual neurons differ greatly in their morphology, connectivity and electrophysiological properties. Systematic classification of all mammalian neurons is a key goal towards deconstructing the nervous system into its basic components. With the recent advances in single-cell gene expression profiling technologies, it is now possible to undertake the enormous task of disentangling neuronal heterogeneity. High-throughput single-cell RNA sequencing and multiplexed quantitative RT-PCR have become more accessible, and these technologies enable systematic categorization of individual neurons into groups with similar molecular properties. Here we provide a conceptual and practical guide to classification of neural cell types using single-cell gene expression profiling technologies. PMID:27571192

  1. Using Magnetic Nanoparticles for Gene Transfer to Neural Stem Cells: Stem Cell Propagation Method Influences Outcomes

    PubMed Central

    Pickard, Mark R.; Adams, Christopher F.; Barraud, Perrine; Chari, Divya M.

    2015-01-01

    Genetically engineered neural stem cell (NSC) transplants offer a key strategy to augment neural repair by releasing therapeutic biomolecules into injury sites. Genetic modification of NSCs is heavily reliant on viral vectors but cytotoxic effects have prompted development of non-viral alternatives, such as magnetic nanoparticle (MNPs). NSCs are propagated in laboratories as either 3-D suspension “neurospheres” or 2-D adherent “monolayers”. MNPs deployed with oscillating magnetic fields (“magnetofection technology”) mediate effective gene transfer to neurospheres but the efficacy of this approach for monolayers is unknown. It is important to address this issue as oscillating magnetic fields dramatically enhance MNP-based transfection in transplant cells (e.g., astrocytes and oligodendrocyte precursors) propagated as monolayers. We report for the first time that oscillating magnetic fields enhanced MNP-based transfection with reporter and functional (basic fibroblast growth factor; FGF2) genes in monolayer cultures yielding high transfection versus neurospheres. Transfected NSCs showed high viability and could re-form neurospheres, which is important as neurospheres yield higher post-transplantation viability versus monolayer cells. Our results demonstrate that the combination of oscillating magnetic fields and a monolayer format yields the highest efficacy for MNP-mediated gene transfer to NSCs, offering a viable non-viral alternative for genetic modification of this important neural cell transplant population. PMID:25918990

  2. Neural stem cells and regulation of cell number.

    PubMed

    Sommer, Lukas; Rao, Mahendra

    2002-01-01

    Normal CNS development involves the sequential differentiation of multipotent stem cells. Alteration of the numbers of stem cells, their self-renewal ability, or their proliferative capacity will have major effects on the appropriate development of the nervous system. In this review, we discuss different mechanisms that regulate neural stem cell differentiation. Proliferation signals and cell cycle regulators may regulate cell kinetics or total number of cell divisions. Loss of trophic support and cytokine receptor activation may differentially contribute to the induction of cell death at specific stages of development. Signaling from differentiated progeny or asymmetric distribution of specific molecules may alter the self-renewal characteristics of stem cells. We conclude that the final decision of a cell to self-renew, differentiate or remain quiescent is dependent on an integration of multiple signaling pathways and at each instant will depend on cell density, metabolic state, ligand availability, type and levels of receptor expression, and downstream cross-talk between distinct signaling pathways. PMID:11897403

  3. 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

  4. Arrested neural and advanced mesenchymal differentiation of glioblastoma cells-comparative study with neural progenitors

    PubMed Central

    2009-01-01

    Background Although features of variable differentiation in glioblastoma cell cultures have been reported, a comparative analysis of differentiation properties of normal neural GFAP positive progenitors, and those shown by glioblastoma cells, has not been performed. Methods Following methods were used to compare glioblastoma cells and GFAP+NNP (NHA): exposure to neural differentiation medium, exposure to adipogenic and osteogenic medium, western blot analysis, immunocytochemistry, single cell assay, BrdU incorporation assay. To characterize glioblastoma cells EGFR amplification analysis, LOH/MSI analysis, and P53 nucleotide sequence analysis were performed. Results In vitro differentiation of cancer cells derived from eight glioblastomas was compared with GFAP-positive normal neural progenitors (GFAP+NNP). Prior to exposure to differentiation medium, both types of cells showed similar multilineage phenotype (CD44+/MAP2+/GFAP+/Vimentin+/Beta III-tubulin+/Fibronectin+) and were positive for SOX-2 and Nestin. In contrast to GFAP+NNP, an efficient differentiation arrest was observed in all cell lines isolated from glioblastomas. Nevertheless, a subpopulation of cells isolated from four glioblastomas differentiated after serum-starvation with varying efficiency into derivatives indistinguishable from the neural derivatives of GFAP+NNP. Moreover, the cells derived from a majority of glioblastomas (7 out of 8), as well as GFAP+NNP, showed features of mesenchymal differentiation when exposed to medium with serum. Conclusion Our results showed that stable co-expression of multilineage markers by glioblastoma cells resulted from differentiation arrest. According to our data up to 95% of glioblastoma cells can present in vitro multilineage phenotype. The mesenchymal differentiation of glioblastoma cells is advanced and similar to mesenchymal differentiation of normal neural progenitors GFAP+NNP. PMID:19216795

  5. 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

  6. 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

  7. Nodal Promotes Glioblastoma Cell Growth

    PubMed Central

    De Silva, Tanya; Ye, Gang; Liang, Yao-Yun; Fu, Guodong; Xu, Guoxiong; Peng, Chun

    2012-01-01

    Nodal is a member of the transforming growth factor-β (TGF-β) superfamily that plays critical roles during embryogenesis. Recent studies in ovarian, breast, prostate, and skin cancer cells suggest that Nodal also regulates cell proliferation, apoptosis, and invasion in cancer cells. However, it appears to exert both tumor-suppressing and tumor-promoting effects, depending on the cell type. To further understand the role of Nodal in tumorigenesis, we examined the effect of Nodal in glioblastoma cell growth and spheroid formation using U87 cell line. Treatment of U87 with recombinant Nodal significantly increased U87 cell growth. In U87 cells stably transfected with the plasmid encoding Nodal, Smad2 phosphorylation was strongly induced and cell growth was significantly enhanced. Overexpression of Nodal also resulted in tight spheroid formation. On the other hand, the cells stably transfected with Nodal siRNA formed loose spheroids. Nodal is known to signal through activin receptor-like kinase 4 (ALK4) and ALK7 and the Smad2/3 pathway. To determine which receptor and Smad mediate the growth promoting effect of Nodal, we transfected siRNAs targeting ALK4, ALK7, Smad2, or Smad3 into Nodal-overexpressing cells and observed that cell growth was significantly inhibited by ALK4, ALK7, and Smad3 siRNAs. Taken together, these findings suggest that Nodal may have tumor-promoting effects on glioblastoma cells and these effects are mediated by ALK4, ALK7, and Smad3. PMID:22645523

  8. 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.

  9. 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

  10. The dynamic matching of neural and cognitive growth cycles.

    PubMed

    Peltzer-Karpf, Annemarie

    2012-01-01

    In recent years complex systems biology has developed detailed numerical models mimicking the establishment, modulation, and fine-tuning of neural networks. Current research within the framework of Dynamic Systems Theory (DST) emphasizes the nexus between dynamic cycles in the brain and cognitive development which unfold in a nonlinear way and allow for individual variation. Careful observations over multiple timescales and levels of organization suggest a link to system-specific developmental changes in the central nervous system with more functional specialization opening up more efficient information processing. This can be seen in spurts of EEG energy and altered cortical coherence. Data of age- and experience-related changes in synaptic density and metabolism, shifts in blood flow and improvement of (sub)cortical connections are projected on a dynamic trajectory of cognition moving from diffuse to more refined constructions in the various subsystems, each of which exhibiting its own developmental path. Pending questions are the generation of rules amidst diversity and fluctuation, and the correlation of growth rate and critical mass in developmental dynamics and interaction. PMID:22196112

  11. Neural Crest Stem Cells from Dental Tissues: A New Hope for Dental and Neural Regeneration

    PubMed Central

    Ibarretxe, Gaskon; Crende, Olatz; Aurrekoetxea, Maitane; García-Murga, Victoria; Etxaniz, Javier; Unda, Fernando

    2012-01-01

    Several stem cell sources persist in the adult human body, which opens the doors to both allogeneic and autologous cell therapies. Tooth tissues have proven to be a surprisingly rich and accessible source of neural crest-derived ectomesenchymal stem cells (EMSCs), which may be employed to repair disease-affected oral tissues in advanced regenerative dentistry. Additionally, one area of medicine that demands intensive research on new sources of stem cells is nervous system regeneration, since this constitutes a therapeutic hope for patients affected by highly invalidating conditions such as spinal cord injury, stroke, or neurodegenerative diseases. However, endogenous adult sources of neural stem cells present major drawbacks, such as their scarcity and complicated obtention. In this context, EMSCs from dental tissues emerge as good alternative candidates, since they are preserved in adult human individuals, and retain both high proliferation ability and a neural-like phenotype in vitro. In this paper, we discuss some important aspects of tissue regeneration by cell therapy and point out some advantages that EMSCs provide for dental and neural regeneration. We will finally review some of the latest research featuring experimental approaches and benefits of dental stem cell therapy. PMID:23093977

  12. 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. PMID:26917252

  13. Molecular Diversity Subdivides the Adult Forebrain Neural Stem Cell Population

    PubMed Central

    Giachino, Claudio; Basak, Onur; Lugert, Sebastian; Knuckles, Philip; Obernier, Kirsten; Fiorelli, Roberto; Frank, Stephan; Raineteau, Olivier; Alvarez–Buylla, Arturo; Taylor, Verdon

    2014-01-01

    Neural stem cells (NSCs) in the ventricular domain of the subventricular zone (V-SVZ) of rodents produce neurons throughout life while those in humans become largely inactive or may be lost during infancy. Most adult NSCs are quiescent, express glial markers, and depend on Notch signaling for their self-renewal and the generation of neurons. Using genetic markers and lineage tracing, we identified subpopulations of adult V-SVZ NSCs (type 1, 2, and 3) indicating a striking heterogeneity including activated, brain lipid binding protein (BLBP, FABP7) expressing stem cells. BLBP+ NSCs are mitotically active components of pinwheel structures in the lateral ventricle walls and persistently generate neurons in adulthood. BLBP+ NSCs express epidermal growth factor (EGF) receptor, proliferate in response to EGF, and are a major clonogenic population in the SVZ. We also find BLBP expressed by proliferative ventricular and sub-ventricular progenitors in the fetal and postnatal human brain. Loss of BLBP+ stem/progenitor cells correlates with reduced neurogenesis in aging rodents and postnatal humans. These findings of molecular heterogeneity and proliferative differences subdivide the NSC population and have implications for neurogenesis in the forebrain of mammals during aging. PMID:23964022

  14. Plasticity and neural stem cells in the enteric nervous system.

    PubMed

    Schäfer, Karl-Herbert; Van Ginneken, Chris; Copray, Sjef

    2009-12-01

    The enteric nervous system (ENS) is a highly organized part of the autonomic nervous system, which innervates the whole gastrointestinal tract by several interconnected neuronal networks. The ENS changes during development and keeps throughout its lifespan a significant capacity to adapt to microenvironmental influences, be it in inflammatory bowel diseases or changing dietary habits. The presence of neural stem cells in the pre-, postnatal, and adult gut might be one of the prerequisites to adapt to changing conditions. During the last decade, the ENS has increasingly come into the focus of clinical neural stem cell research, forming a considerable pool of neural crest derived stem cells, which could be used for cell therapy of dysganglionosis, that is, diseases based on the deficient or insufficient colonization of the gut by neural crest derived stem cells; in addition, the ENS could be an easily accessible neural stem cell source for cell replacement therapies for neurodegenerative disorders or traumatic lesions of the central nervous system. PMID:19943347

  15. 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

  16. LIF-dependent primitive neural stem cells derived from mouse ES cells represent a reversible stage of neural commitment.

    PubMed

    Tsang, Wan-Hong; Wang, Bin; Wong, Wing Ki; Shi, Shuo; Chen, Xiao; He, Xiangjun; Gu, Shen; Hu, Jiabiao; Wang, Chengdong; Liu, Pi-Chu; Lu, Gang; Chen, Xiongfong; Zhao, Hui; Poon, Wai-Sang; Chan, Wai-Yee; Feng, Bo

    2013-11-01

    Primitive neural stem cells (NSCs) define an early stage of neural induction, thus provide a model to understand the mechanism that controls initial neural commitment. In this study, we investigated primitive NSCs derived from mouse embryonic stem cells (ESCs). By genome-wide transcriptional profiling, we revealed their unique signature and depicted the molecular changes underlying critical cell fate transitions during early neural induction at a global level. Together with qRT-PCR analysis, our data illustrated that primitive NSCs retained expression of key pluripotency genes Oct4 and Nanog, while exhibiting repression of other pluripotency-related genes Zscan4, Foxp1 and Dusp9 and up-regulation of neural markers Sox1 and Hes1. The early differentiation feature in primitive NSCs was also supported by their intermediate characters on cell cycle profiles. Moreover, re-plating primitive NSCs back to ESC culture condition could reverse them back to ESC stage, as shown by reversible regulation of marker genes, cell cycle profile changes and enhanced embryoid body formation. In addition, our microarray analysis also identified genes differentially expressed in primitive NSCs, and loss-of-function analysis demonstrated that Hes1 and Ccdc141 play important function at this stage, opening up an opportunity to further understand the regulation of early neural commitment. PMID:23973799

  17. 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.

  18. 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

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

    PubMed

    Mundell, Nathan A; Labosky, Patricia A

    2011-02-01

    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

  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. 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

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

    PubMed

    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

  3. 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

  4. 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. PMID:16041760

  5. 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.

  6. 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

  7. 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. PMID:27341268

  8. Understanding How Zika Virus Enters and Infects Neural Target Cells.

    PubMed

    Miner, Jonathan J; Diamond, Michael S

    2016-05-01

    Zika virus is a mosquito-transmitted flavivirus that has become a public health concern because of its ability to cause microcephaly. In this issue of Cell Stem Cell, Tang et al. (2016) and Nowakowski et al. (2016) use human neural stem cell models and single-cell RNA sequencing to investigate Zika virus tropism and potential entry receptors. PMID:27152436

  9. Neural Potential of a Stem Cell Population in the Hair Follicle

    PubMed Central

    Mignone, John L.; Roig-Lopez, Jose L.; Fedtsova, Natalia; Schones, Dustin E.; Manganas, Louis N.; Maletic-Savatic, Mirjana; Keyes, William M.; Mills, Alea A.; Gleiberman, Anatoli; Zhang, Michael Q.; Enikolopov, Grigori

    2013-01-01

    The bulge region of the hair follicle serves as a repository for epithelial stem cells that can regenerate the follicle in each hair growth cycle and contribute to epidermis regeneration upon injury. Here we describe a population of multipotential stem cells in the hair follicle bulge region; these cells can be identified by fluorescence in transgenic nestin-GFP mice. The morphological features of these cells suggest that they maintain close associations with each other and with the surrounding niche. Upon explantation, these cells can give rise to neurosphere-like structures in vitro. When these cells are permitted to differentiate, they produce several cell types, including cells with neuronal, astrocytic, oligodendrocytic, smooth muscle, adipocytic, and other phenotypes. Furthermore, upon implantation into the developing nervous system of chick, these cells generate neuronal cells in vivo. We used transcriptional profiling to assess the relationship between these cells and embryonic and postnatal neural stem cells and to compare them with other stem cell populations of the bulge. Our results show that nestin-expressing cells in the bulge region of the hair follicle have stem cell-like properties, are multipotent, and can effectively generate cells of neural lineage in vitro and in vivo. PMID:17873521

  10. 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.

  11. Characterization of TLX Expression in Neural Stem Cells and Progenitor Cells in Adult Brains

    PubMed Central

    Li, Shengxiu; Sun, Guoqiang; Murai, Kiyohito; Ye, Peng; Shi, Yanhong

    2012-01-01

    TLX has been shown to play an important role in regulating the self-renewal and proliferation of neural stem cells in adult brains. However, the cellular distribution of endogenous TLX protein in adult brains remains to be elucidated. In this study, we used immunostaining with a TLX-specific antibody to show that TLX is expressed in both neural stem cells and transit-amplifying neural progenitor cells in the subventricular zone (SVZ) of adult mouse brains. Then, using a double thymidine analog labeling approach, we showed that almost all of the self-renewing neural stem cells expressed TLX. Interestingly, most of the TLX-positive cells in the SVZ represented the thymidine analog-negative, relatively quiescent neural stem cell population. Using cell type markers and short-term BrdU labeling, we demonstrated that TLX was also expressed in the Mash1+ rapidly dividing type C cells. Furthermore, loss of TLX expression dramatically reduced BrdU label-retaining neural stem cells and the actively dividing neural progenitor cells in the SVZ, but substantially increased GFAP staining and extended GFAP processes. These results suggest that TLX is essential to maintain the self-renewing neural stem cells in the SVZ and that the GFAP+ cells in the SVZ lose neural stem cell property upon loss of TLX expression.Understanding the cellular distribution of TLX and its function in specific cell types may provide insights into the development of therapeutic tools for neurodegenerative diseases by targeting TLX in neural stem/progenitors cells. PMID:22952666

  12. Specification of neural cell fate and regulation of neural stem cell proliferation by microRNAs

    PubMed Central

    Pham, Jacqueline T; Gallicano, G Ian

    2012-01-01

    In the approximately 20 years since microRNAs (miRNAs) were first characterized, they have been shown to play important roles in diverse physiologic functions, particularly those requiring coordinated changes in networks of signaling pathways. The ability of miRNAs to silence expression of multiple gene targets hints at complex connections that research has only begun to elucidate. The nervous system, particularly the brain, and its progenitor cells offer opportunities to examine miRNA function due to the myriad different cell types, numerous functionally distinct regions, and fluidly dynamic connections between them. This review aims to summarize current understanding of miRNA regulation in neurodevelopment, beginning with miRNAs that establish a general neural fate in cells. Particular attention is given to miR-124, the most abundant brain-specific miRNA, along with its key regulators and targets as an example of the potentially far-reaching effects of miRNAs. These modulators and mediators enable miRNAs to subtly calibrate cellular proliferation and differentiation. To better understand their mechanisms of action, miRNA profiles in distinct populations and regions of cells have been examined as well as miRNAs that regulate proliferation of stem cells, a process marked by dramatic morphological shifts in response to temporally subtle and refined shifts in gene expression. To tease out the complex interactions of miRNAs and stem cells more accurately, future studies will require more sensitive methods of assessing miRNA expression and more rigorous models of miRNA pathways. Thorough characterization of similarities and differences in specific miRNAs’ effects in different species is vital to developing better disease models and therapeutics using miRNAs. PMID:23671807

  13. Direct reprogramming of human neural stem cells by OCT4.

    PubMed

    Kim, Jeong Beom; Greber, Boris; Araúzo-Bravo, Marcos J; Meyer, Johann; Park, Kook In; Zaehres, Holm; Schöler, Hans R

    2009-10-01

    Induced pluripotent stem (iPS) cells have been generated from mouse and human somatic cells by ectopic expression of four transcription factors (OCT4 (also called POU5F1), SOX2, c-Myc and KLF4). We previously reported that Oct4 alone is sufficient to reprogram directly adult mouse neural stem cells to iPS cells. Here we report the generation of one-factor human iPS cells from human fetal neural stem cells (one-factor (1F) human NiPS cells) by ectopic expression of OCT4 alone. One-factor human NiPS cells resemble human embryonic stem cells in global gene expression profiles, epigenetic status, as well as pluripotency in vitro and in vivo. These findings demonstrate that the transcription factor OCT4 is sufficient to reprogram human neural stem cells to pluripotency. One-factor iPS cell generation will advance the field further towards understanding reprogramming and generating patient-specific pluripotent stem cells. PMID:19718018

  14. Generation and Applications of Human Pluripotent Stem Cells Induced into Neural Lineages and Neural Tissues

    PubMed Central

    Martinez, Y.; Dubois-Dauphin, M.; Krause, K.-H.

    2012-01-01

    Human pluripotent stem cells (hPSCs) represent a new and exciting field in modern medicine, now the focus of many researchers and media outlets. The hype is well-earned because of the potential of stem cells to contribute to disease modeling, drug screening, and even therapeutic approaches. In this review, we focus first on neural differentiation of these cells. In a second part we compare the various cell types available and their advantages for in vitro modeling. Then we provide a “state-of-the-art” report about two major biomedical applications: (1) the drug and toxicity screening and (2) the neural tissue replacement. Finally, we made an overview about current biomedical research using differentiated hPSCs. PMID:22457650

  15. 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

  16. 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. PMID:27352251

  17. 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.

  18. 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

  19. YAP regulates neural progenitor cell number via the TEA domain transcription factor

    PubMed Central

    Cao, Xinwei; Pfaff, Samuel L.; Gage, Fred H.

    2008-01-01

    Tight control of cell proliferation is essential for proper growth during development and for tissue homeostasis in mature animals. The evolutionarily conserved Hippo pathway restrains proliferation through a kinase cascade that culminates in the inhibition of the transcriptional coactivator YAP. Unphosphorylated YAP activates genes involved in cell proliferation and survival by interacting with a DNA-binding factor. Here we show that during vertebrate neural tube development, the TEA domain transcription factor (TEAD) is the cognate DNA-binding partner of YAP. YAP and TEAD gain of function causes marked expansion of the neural progenitor population, partly owing to their ability to promote cell cycle progression by inducing cyclin D1 and to inhibit differentiation by suppressing NeuroM. Their loss of function results in increased apoptosis, whereas repressing their target genes leads to premature neuronal differentiation. Inhibiting the upstream kinases of the Hippo pathway also causes neural progenitor overproliferation. Thus, the Hippo pathway plays critical roles in regulating neural progenitor cell number by affecting proliferation, fate choice, and cell survival. PMID:19015275

  20. Comparative analysis of neural crest cell death, migration, and function during vertebrate embryogenesis.

    PubMed

    Kulesa, Paul; Ellies, Debra L; Trainor, Paul A

    2004-01-01

    Cranial neural crest cells are a multipotent, migratory population that generates most of the cartilage, bone, connective tissue and peripheral nervous system in the vertebrate head. Proper neural crest cell patterning is essential for normal craniofacial morphogenesis and is highly conserved among vertebrates. Neural crest cell patterning is intimately connected to the early segmentation of the neural tube, such that neural crest cells migrate in discrete segregated streams. Recent advances in live embryo imaging have begun to reveal the complex behaviour of neural crest cells which involve intricate cell-cell and cell-environment interactions. Despite the overall similarity in neural crest cell migration between distinct vertebrates species there are important mechanistic differences. Apoptosis for example, is important for neural crest cell patterning in chick embryos but not in mouse, frog or fish embryos. In this paper we highlight the potential evolutionary significance of such interspecies differences in jaw development and evolution. Developmental Dynamics 229:14-29, 2004. PMID:14699574

  1. An Overview of Protocols for the Neural Induction of Dental and Oral Stem Cells In Vitro.

    PubMed

    Heng, Boon Chin; Lim, Lee Wei; Wu, Wutian; Zhang, Chengfei

    2016-06-01

    To date, various adult stem cells have been identified within the oral cavity, including dental pulp stem cells, dental follicle stem cells, stem cells from apical papilla, stem cells from human exfoliated deciduous teeth, periodontal ligament stem cells, and mesenchymal stem cells from the gingiva. All of these possess neurogenic potential due to their common developmental origin from the embryonic neural crest. Besides the relative ease of isolation of these adult stem cells from readily available biological waste routinely produced during dental treatment, these cells also possess the advantage of immune compatibility in autologous transplantation. In recent years, much interest has been focused on the derivation of neural lineages from these adult stem cells for therapeutic applications in the brain, spinal cord, and peripheral nerve regeneration. In addition, there are also promising nontherapeutic applications of stem cell-derived neurons in pharmacological and toxicological screening of neuroactive drugs, and for in vitro modeling of neurodevelopmental and neurodegenerative diseases. Hence, this review will critically examine the diverse array of in vitro neural induction protocols that have been devised for dental and oral-derived stem cells. These protocols are defined not only by the culture milieu comprising the basal medium plus growth factors, small molecules, and other culture supplements but also by the substrata/surface coatings utilized, the presence of multiple culture stages, the total culture duration, the initial seeding density, and whether the spheroid/neurosphere formation is being utilized to recapitulate the three-dimensional neural differentiation microenvironment that is naturally present physiologically in vivo. PMID:26757369

  2. 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.

  3. 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. PMID:26607204

  4. 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

  5. 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

    2015-10-01

    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

  6. 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

  7. The neural milieu of the developing choroid plexus: neural stem cells, neurons and innervation.

    PubMed

    Prasongchean, Weerapong; Vernay, Bertrand; Asgarian, Zeinab; Jannatul, Nahin; Ferretti, Patrizia

    2015-01-01

    The choroid plexus produces cerebrospinal fluid and plays an important role in brain homeostasis both pre and postnatally. In vitro studies have suggested that cells from adult choroid plexus have stem/progenitor cell-like properties. Our initial aim was to investigate whether such a cell population is present in vivo during development of the choroid plexus, focusing mainly on the chick choroid plexus. Cells expressing neural markers were indeed present in the choroid plexus of chick and also those of rodent and human embryos, both within their epithelium and mesenchyme. ß3-tubulin-positive cells with neuronal morphology could be detected as early as at E8 in chick choroid plexus and their morphological complexity increased with development. Whole mount immunochemistry demonstrated the presence of neurons throughout choroid plexus development and they appeared to be mainly catecholaminergic, as indicated by tyrosine-hydroxylase reactivity. The presence of cells co-labeling for BrdU and the neuroblast marker, doublecortin, in organotypic choroid plexus cultures supported the hypothesis that neurogenesis can occur from neural precursors within the developing choroid plexus. Furthermore, we found that extrinsic innervation is present in the developing choroid plexus, unlike previously suggested. Altogether, our data are consistent with the presence of neural progenitors within the choroid plexus, suggest that at least some of the choroid plexus neurons are born locally, and show for the first time that choroid plexus innervation occurs prenatally. Hence, we propose the existence of a complex neural regulatory network within the developing choroid plexus that may play a crucial role in modulating its function during development as well as throughout life. PMID:25873856

  8. Neural stem cells: plasticity and their transdifferentiation potential.

    PubMed

    Vescovi, Angelo; Gritti, Angela; Cossu, Giulio; Galli, Rossella

    2002-01-01

    The presence of resident stem cells in adult tissues is of fundamental importance for the maintenance of their structural and functional integrity. In fact, throughout life, somatic stem cells attend to the critical function of substituting terminally differentiated cells lost to physiological turnover, injury or disease. Thence, one of the basic dogmata in tissue biology holds that the differentiation potential of an adult stem cell is restricted to the generation of the mature cell lineages found in the tissue to which the stem cell belongs. A plethora of recent evidences from many groups, including ours, is now providing evidence that adult stem cells may possess a broader differentiation repertoire than expected and that their fate potential may not be as tissue specific as once thought. The initial example of an unforeseen, trans-germ layer plasticity - that seems now to emerge as a prototypic functional trait of various somatic stem cells of different origin - has come from the reported awakening of a latent hemopoietic developmental capacity in stem cells isolated from the adult mammalian brain following their transplantation into sub-lethally irradiated mice. More recently, it has been shown that adult neural stem cells can differentiate into a wide array of bodily cells of different origin when injected into the blastocyst and into myogenic cells when transplanted into the adult regenerating skeletal muscle. Moreover, bone marrow stem cells can now give rise to skeletal muscle, hepatic and brain cells, whereas muscle precursors can generate blood cells. In this article, we review some of the basic notions regarding the functional properties of the adult neural stem cells and discuss findings in the expanding area of trans-germ layer conversion, with emphasis on the neural stem cell. PMID:12021492

  9. 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

  10. Isolation and Culture of Neural Crest Cells from Embryonic Murine Neural Tube

    PubMed Central

    Pfaltzgraff, Elise R.; Mundell, Nathan A.; Labosky, Patricia A.

    2012-01-01

    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 1-3. NC also has the unique ability to influence the differentiation and maturation of target organs4-6. 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 tube7-9. 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 embryo10-13. 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 progenitors11,14-20, however isolation of NC progenitors as they emigrate from the neural tube provides optimal preservation of NC cell potential and migratory properties13,21,22. Some protocols employ fluorescence activated cell sorting (FACS) to isolate a NC population enriched for particular progenitors11,13,14,17. 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

  11. Isolation and culture of neural crest cells from embryonic murine neural tube.

    PubMed

    Pfaltzgraff, Elise R; Mundell, Nathan A; Labosky, Patricia A

    2012-01-01

    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. 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. 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

  14. The transcription factor Nerfin-1 prevents reversion of neurons into neural stem cells.

    PubMed

    Froldi, Francesca; Szuperak, Milan; Weng, Chen-Fang; Shi, Wei; Papenfuss, Anthony T; Cheng, Louise Y

    2015-01-15

    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

  15. Endogenous neural precursors influence grafted neural stem cells and contribute to neuroprotection in the Parkinsonian rat

    PubMed Central

    Madhavan, Lalitha; Daley, Brian F; Sortwell, Caryl E; Collier, Timothy J

    2012-01-01

    Neuroprotective and neurorescue effects after neural stem/precursor cell (NPC) transplantation have been reported, but the mechanisms underlying such phenomena are not well understood. Our recent findings in a rat Parkinson’s disease (PD) model indicate that transplantation of NPCs before a 6-hydroxydopamine (6-OHDA) insult can result in nigrostriatal protection which is associated with endogenous NPC proliferation, migration and neurogenesis. Here, we sought to determine whether the observed endogenous NPC response (1) contributes to transplanted NPC - mediated neuroprotection and/or (2) affects graft phenotype and function. Host Fischer 344 rats were administered the antimitotic agent cytosine-β-D-arabinofuranoside (Ara-C) to eliminate actively proliferating endogenous neural precursors before being transplanted with NPCs and treated with 6-OHDA to induce nigrostriatal degeneration. Behavioral and histological analyses demonstrate that the neuroprotective response observed in NPC transplanted animals which had not received Ara-C was significantly attenuated in animals which did receive pre-transplant Ara-C. Also, while grafts in Ara-C treated animals showed no decrease in cell number, they exhibited significantly reduced expression of the neural stem cell regulators nestin and sonic hedgehog. In addition, inhibition of the endogenous NPC response resulted in an exaggerated host glial reaction. Overall, the study establishes for the first time that endogenous NPCs contribute to transplanted NPC-mediated therapeutic effects by affecting both grafted and mature host cells in unique ways. Thus, both endogenous and transplanted NPCs are important in creating an environment suitable for neural protection and rescue, and harnessing their synergistic interaction may lead to the optimization of cell-based therapies for PD. PMID:22417168

  16. Perineural Growth in Head and Neck Squamous Cell Carcinoma: A Review

    PubMed Central

    Roh, Joseph; Muelleman, Thomas; Tawfik, Ossama; Thomas, Sufi M

    2014-01-01

    Perineural growth is a unique route of tumor metastasis that is associated with poor prognosis in several solid malignancies. It is diagnosed by the presence of tumor cells inside the neural space seen on histological or imaging evaluations. Little is known about molecular mechanisms involved in the growth and spread of tumor cells in neural spaces. The poor prognosis associated with perineural growth and lack of targeted approaches necessitates the study of molecular factors involved in communication between tumor and neural cells. Perineural growth rates, shown to be as high as 63% in head and neck squamous cell carcinoma (HNSCC), correlate with increased local recurrence and decreased disease-free survival. Here we describe the literature on perineural growth in HNSCC. In addition, we discuss factors implicated in perineural growth of cancer. These factors include brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), neurotropin-3 and -4, glial cell-line derived neurotrophic factor (GDNF), the neural cell adhesion molecule (NCAM), substance P (SP), and chemokines. We also explore the literature on membrane receptors, including the Trk family and the low-affinity nerve growth factor receptor. This review highlights areas for further study of the mechanisms of perineural invasion which may facilitate the identification of therapeutic targets in HNSCC. PMID:25456006

  17. p73 regulates maintenance of neural stem cell

    SciTech Connect

    Agostini, Massimiliano; Tucci, Paola; Biochemistry Laboratory, IDI-IRCCS, C Chen, Hailan; Knight, Richard A.; Bano, Daniele; Nicotera, Pierluigi; McKeon, Frank; Melino, Gerry; Biochemistry Laboratory, IDI-IRCCS, C/O University of Rome 'Tor Vergata', 00133 Rome

    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.

  18. 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.

  19. Roles of imprinted genes in neural stem cells.

    PubMed

    Hoffmann, Anke; Daniel, Guillaume; Schmidt-Edelkraut, Udo; Spengler, Dietmar

    2014-01-01

    Imprinted genes and neural stem cells (NSC) play an important role in the developing and mature brain. A central theme of imprinted gene function in NSCs is cell survival and G1 arrest to control cell division, cell-cycle exit, migration and differentiation. Moreover, genomic imprinting can be epigenetically switched off at some genes to ensure stem cell quiescence and differentiation. At the genome scale, imprinted genes are organized in dynamic networks formed by interchromosomal interactions and transcriptional coregulation of imprinted and nonimprinted genes. Such multilayered networks may synchronize NSC activity with the demand from the niche resembling their roles in adjusting fetal size. PMID:25431944

  20. 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

  1. 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.

  2. Aebp2 as an epigenetic regulator for neural crest cells.

    PubMed

    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

  3. Quantitative analysis of signaling mechanisms controlling adult neural progenitor cell proliferation.

    PubMed

    Schaffer, David V; O'Neill, Analeah; Hochrein, Lisa; McGranahan, Tresa

    2004-01-01

    Tools of systems engineering and signal dynamics were employed to develop a quantitative model of the intracellular signaling systems involved in adult neural stem cell proliferation, based on pathways elucidated in our experimental systems. Neural progenitors isolated from the adult rat hippocampus are dependent on the basic fibroblast growth factor (FGF-2) and extracellular matrix (ECM) proteins. However, the intracellular effects of these stimuli were previously undetermined. We employed chemical inhibitors of known signal transduction molecules to identify important players in the FGF-2/ECM signal cascade, such as the cyclic AMP responsive element binding protein (CREB), protein kinase B/Akt, and several related molecules. Genetic mutants of these proteins were used to confirm their role in adult neural progenitor proliferation. Proliferation was assayed using the incorporation of a thymidine analog to determine cell doubling rate under various stimuli. Such assays have also uncovered novel synergistic signaling between FGF-2 and ECM components. This research is, to our knowledge, the first to elucidate intracellular signaling pathways for adult neural stem cell proliferation. Upon determination of the pertinent intracellular signaling pathways, quantitative immunoblots were employed to examine the dynamics of these systems. These data, as well as enzyme kinetics information from the literature, are being used to parameterize a dynamic mathematical model of progenitor proliferation events induced by FGF-2. This computational model will be used to predict the biochemical and mechanical signaling inputs necessary to achieve a desired proliferative output from the cells, based on specific extracellular stimuli. It is our hope that this essential quantitative understanding will facilitate the use of adult neural stem cells in medical applications. PMID:17271428

  4. 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

  5. μ- and κ-Opioids Induce the Differentiation of Embryonic Stem Cells to Neural Progenitors*

    PubMed Central

    Kim, Eunhae; Clark, Amy L.; Kiss, Alexi; Hahn, Jason W.; Wesselschmidt, Robin; Coscia, Carmine J.; Belcheva, Mariana M.

    2008-01-01

    Growth factors, hormones, and neurotransmitters have been implicated in the regulation of stem cell fate. Since various neural precursors express functional neurotransmitter receptors, which include G protein-coupled receptors, it is anticipated that they are involved in cell fate decisions. We detected μ-opioid receptor (MOR-1) and κ-opioid receptor (KOR-1) expression and immunoreactivity in embryonic stem (ES) cells and in retinoic acid-induced ES cell-derived, nestin-positive, neural progenitors. Moreover, these G protein-coupled receptors are functional, since [D-Ala2,MePhe4,Gly-ol5]enkephalin, a MOR-selective agonist, and U69,593, a KOR-selective agonist, induce a sustained activation of extracellular signal-regulated kinase (ERK) signaling throughout a 24-h treatment period in undifferentiated, self-renewing ES cells. Both opioids promote limited proliferation of undifferentiated ES cells via the ERK/MAP kinase signaling pathway. Importantly, biochemical and immunofluorescence data suggest that [D-Ala2,MePhe4,Gly-ol5]enkephalin and U69,593 divert ES cells from self-renewal and coax the cells to differentiate. In retinoic acid-differentiated ES cells, opioid-induced signaling features a biphasic ERK activation profile and an opioid-induced, ERK-independent inhibition of proliferation in these neural progenitors. Collectively, the data suggest that opioids may have opposite effects on ES cell self-renewal and ES cell differentiation and that ERK activation is only required by the latter. Finally, opioid modulation of ERK activity may play an important role in ES cell fate decisions by directing the cells to specific lineages. PMID:16954126

  6. Mu- and kappa-opioids induce the differentiation of embryonic stem cells to neural progenitors.

    PubMed

    Kim, Eunhae; Clark, Amy L; Kiss, Alexi; Hahn, Jason W; Wesselschmidt, Robin; Coscia, Carmine J; Belcheva, Mariana M

    2006-11-01

    Growth factors, hormones, and neurotransmitters have been implicated in the regulation of stem cell fate. Since various neural precursors express functional neurotransmitter receptors, which include G protein-coupled receptors, it is anticipated that they are involved in cell fate decisions. We detected mu-opioid receptor (MOR-1) and kappa-opioid receptor (KOR-1) expression and immunoreactivity in embryonic stem (ES) cells and in retinoic acid-induced ES cell-derived, nestin-positive, neural progenitors. Moreover, these G protein-coupled receptors are functional, since [D-Ala(2),MePhe(4),Gly-ol(5)]enkephalin, a MOR-selective agonist, and U69,593, a KOR-selective agonist, induce a sustained activation of extracellular signal-regulated kinase (ERK) signaling throughout a 24-h treatment period in undifferentiated, self-renewing ES cells. Both opioids promote limited proliferation of undifferentiated ES cells via the ERK/MAP kinase signaling pathway. Importantly, biochemical and immunofluorescence data suggest that [D-Ala(2),MePhe(4),Gly-ol(5)]enkephalin and U69,593 divert ES cells from self-renewal and coax the cells to differentiate. In retinoic acid-differentiated ES cells, opioid-induced signaling features a biphasic ERK activation profile and an opioid-induced, ERK-independent inhibition of proliferation in these neural progenitors. Collectively, the data suggest that opioids may have opposite effects on ES cell self-renewal and ES cell differentiation and that ERK activation is only required by the latter. Finally, opioid modulation of ERK activity may play an important role in ES cell fate decisions by directing the cells to specific lineages. PMID:16954126

  7. 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. PMID:26940741

  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. Bone marrow-derived, neural-like cells have the characteristics of neurons to protect the peripheral nerve in microenvironment.

    PubMed

    Guo, Shi-Lei; Zhang, Zhi-Ying; Xu, Yan; Zhi, Yun-Xia; Han, Chang-Jie; Zhou, Yu-Hao; Liu, Fang; Lin, Hai-Yan; Zhang, Chuan-Sen

    2015-01-01

    Effective repair of peripheral nerve defects is difficult because of the slow growth of new axonal growth. We propose that "neural-like cells" may be useful for the protection of peripheral nerve destructions. Such cells should prolong the time for the disintegration of spinal nerves, reduce lesions, and improve recovery. But the mechanism of neural-like cells in the peripheral nerve is still unclear. In this study, bone marrow-derived neural-like cells were used as seed cells. The cells were injected into the distal end of severed rabbit peripheral nerves that were no longer integrated with the central nervous system. Electromyography (EMG), immunohistochemistry, and transmission electron microscopy (TEM) were employed to analyze the development of the cells in the peripheral nerve environment. The CMAP amplitude appeared during the 5th week following surgery, at which time morphological characteristics of myelinated nerve fiber formation were observed. Bone marrow-derived neural-like cells could protect the disintegration and destruction of the injured peripheral nerve. PMID:25861281

  10. 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. PMID:20211178

  11. Intraspinal transplantation of mouse and human neural precursor cells

    PubMed Central

    Weinger, Jason G.; Chen, Lu; Coleman, Ronald; Leang, Ronika; Plaisted, Warren C.; Loring, Jeanne F.; Lane, Thomas E.

    2013-01-01

    This unit describes the preparation and transplantation of human neural precursor cells (hNPCs) and mouse neural precursor cells (mNPCs) into the thoracic region of the mouse spinal cord. The techniques in this unit also describe how to prepare the mouse for surgery by performing a laminectomy to expose the spinal cord for transplantation. Here we show NPCs genetically labeled with eGFP transplanted into the spinal cord of a mouse following viralmediated demyelination can efficiently be detected via eGFP expression. Transplantation of these cells into the spinal cord is an efficacious way to determine their effects in neurological disorders such as multiple sclerosis, Alzheimer's disease, and spinal cord injury. PMID:24510791

  12. 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. PMID:26032608

  13. 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

    2015-01-01

    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. PMID:26568310

  14. The flavonoids hesperidin and rutin promote neural crest cell survival.

    PubMed

    Nones, Jader; Costa, Ana Paula; Leal, Rodrigo Bainy; Gomes, Flávia Carvalho Alcantara; Trentin, Andréa Gonçalves

    2012-11-01

    The neural crest (NC) corresponds to a collection of multipotent and oligopotent progenitors endowed with both neural and mesenchymal potentials. The derivatives of the NC at trunk level include neurons and glial cells of the peripheral nervous system in addition to melanocytes, smooth muscle cells and some endocrine cells. Environmental factors control the fate decisions of NC cells. Despite the well-known influence of flavonoids on the central nervous system, the issue of whether they also influence NC cells has not been yet addressed. Flavonoids are polyphenolic compounds that are integral components of the human diet. The biological activities of these compounds cover a very broad spectrum, from anticancer and antibacterial activities to inhibition of bone reabsorption and modulation of inflammatory response. In the present work, we have investigated the actions of the flavonoids hesperidin, rutin and quercetin on NC cells of quail, in vitro. We show for the first time, that hesperidin and rutin increase the viability of trunk NC cells in culture, without affecting cell differentiation and proliferation. The molecular mechanism of this action is dependent on ERK2 and PI3K pathways. Quercetin had no effect on NC progenitors. Taken together, these results suggest that flavonoids hesperidin and rutin increase NC cell survival, which may be useful against the toxicity of some chemicals during embryonic development. PMID:22855262

  15. Human neural progenitor cells in central nervous system lesions.

    PubMed

    Åkesson, Elisabet; Sundström, Erik

    2016-02-01

    Various immature cells can be isolated from human embryonic and fetal central nervous system (CNS) residual tissue and potentially be used in cell therapy for a number of neurological diseases and CNS insults. Transplantation of neural stem and progenitor cells is essential for replacing lost cells, particularly in the CNS with very limited endogenous regenerative capacity. However, while dopamine released from transplanted cells can substitute the lost dopamine neurons in the experimental models of Parkinson's disease, stem and progenitor cells primarily have a neuroprotective effect, probably through the release of trophic factors. Understanding the therapeutic effects of transplanted cells is crucial to determine the design of clinical trials. During the last few years, a number of clinical trials for CNS diseases and insults such as amyotrophic lateral sclerosis (ALS), stroke, and spinal cord trauma using neural progenitor cells have been initiated. Data from these early studies will provide vital information on the safety of transplanting these cells, which still is a major concern. That the beneficial results observed in experimental models also can be repeated in the clinical setting is highly hoped for. PMID:26803559

  16. DNA topoisomerase IIβ as a molecular switch in neural differentiation of mesenchymal stem cells.

    PubMed

    Isik, Sevim; Zaim, Merve; Yildiz, Mehmet Taha; Negis, Yesim; Kunduraci, Tuba; Karakas, Nihal; Arikan, Gulsum; Cetin, Guven

    2015-02-01

    processes and signal transduction pathways including transcription, translation, cell trafficking, vesicle function, transport, cell morphology, neuron guidance, growth, polarity, and axonal growth. It appears that the deregulation of these pathways may contribute to clarify the further role of topo IIβ in neural differentiation. PMID:25217229

  17. 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

  18. Bone Marrow-Derived, Neural-Like Cells Have the Characteristics of Neurons to Protect the Peripheral Nerve in Microenvironment

    PubMed Central

    Guo, Shi-lei; Zhang, Zhi-ying; Zhi, Yun-xia; Han, Chang-jie; Zhou, Yu-hao; Liu, Fang; Lin, Hai-yan; Zhang, Chuan-sen

    2015-01-01

    Effective repair of peripheral nerve defects is difficult because of the slow growth of new axonal growth. We propose that “neural-like cells” may be useful for the protection of peripheral nerve destructions. Such cells should prolong the time for the disintegration of spinal nerves, reduce lesions, and improve recovery. But the mechanism of neural-like cells in the peripheral nerve is still unclear. In this study, bone marrow-derived neural-like cells were used as seed cells. The cells were injected into the distal end of severed rabbit peripheral nerves that were no longer integrated with the central nervous system. Electromyography (EMG), immunohistochemistry, and transmission electron microscopy (TEM) were employed to analyze the development of the cells in the peripheral nerve environment. The CMAP amplitude appeared during the 5th week following surgery, at which time morphological characteristics of myelinated nerve fiber formation were observed. Bone marrow-derived neural-like cells could protect the disintegration and destruction of the injured peripheral nerve. PMID:25861281

  19. 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. PMID:26177834

  20. 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

  1. Apoptosome inactivation rescues proneural and neural cells from neurodegeneration.

    PubMed

    Cozzolino, M; Ferraro, E; Ferri, A; Rigamonti, D; Quondamatteo, F; Ding, H; Xu, Z S; Ferrari, F; Angelini, D F; Rotilio, G; Cattaneo, E; Carrì, M T; Cecconi, F

    2004-11-01

    Deficiency of the apoptosome component Apaf1 leads to accumulation of supernumerary brain cells in mouse embryos. We observed that neural precursor cells (NPCs) in Apaf1(-/-) embryos escape programmed cell death, proliferate and retain their potential to differentiate. To evaluate the circumstances of Apaf1(-/-) NPC survival and investigate their fate under neurodegenerative conditions, we established cell lines of embryonic origin (ETNA). We found that Apaf1(-/-) NPCs resist common apoptotic stimuli and neurodegenerative inducers such as amyloid-beta peptide (typical of Alzheimer's disease) and mutant G93A superoxide dismutase 1 (typical of familial amyotrophic lateral sclerosis). Similar results were obtained in Apaf1(-/-) primary cells. When death is prevented by Apaf1 deficiency, cytochrome c is released from mitochondria and rapidly degraded by the proteasome, but mitochondria remain intact. Under these conditions, neither activation by cleavage of initiator caspases nor release of alternative apoptotic inducers from mitochondria takes place. In addition, NPCs can still differentiate, as revealed by neurite outgrowth and expression of differentiation markers. Our findings imply that the mitochondrion/apoptosome pathway is the main route of proneural and neural cells to death and that its inhibition prevents them from dismantling in neurodegenerative conditions. Indeed, the ETNA cell model is ideally suited for exploring the potential of novel cell therapies for the treatment of human neurodegenerations. PMID:15257302

  2. MycN Is Critical for the Maintenance of Human Embryonic Stem Cell-Derived Neural Crest Stem Cells.

    PubMed

    Zhang, Jie Ting; Weng, Zhi Hui; Tsang, Kam Sze; Tsang, Lai Ling; Chan, Hsiao Chang; Jiang, Xiao Hua

    2016-01-01

    The biologic studies of human neural crest stem cells (hNCSCs) are extremely challenging due to the limited source of hNCSCs as well as ethical and technical issues surrounding isolation of early human embryonic tissues. On the other hand, vast majority of studies on MycN have been conducted in human tumor cells, thus, the role of MycN in normal human neural crest development is completely unknown. In the present study, we determined the role of MycN in hNCSCs isolated from in vitro-differentiating human embryonic stem cells (hESCs). For the first time, we show that suppression of MycN in hNCSCs inhibits cell growth and cell cycle progression. Knockdown of MycN in hNCSCs increases the expression of Cdkn1a, Cdkn2a and Cdkn2b, which encodes the cyclin-dependent kinases p21CIP1, p16 INK4a and p15INK4b. In addition, MycN is involved in the regulation of human sympathetic neurogenesis, as knockdown of MycN enhances the expression of key transcription factors involved in sympathetic neuron differentiation, including Phox2a, Phox2b, Mash1, Hand2 and Gata3. We propose that unlimited source of hNCSCs provides an invaluable platform for the studies of human neural crest development and diseases. PMID:26815535

  3. Gene expression changes in the course of neural progenitor cell differentiation.

    PubMed

    Gurok, Ulf; Steinhoff, Christine; Lipkowitz, Bettina; Ropers, H-Hilger; Scharff, Constance; Nuber, Ulrike A

    2004-06-30

    The molecular changes underlying neural progenitor differentiation are essentially unknown. We applied cDNA microarrays with 13,627 clones to measure dynamic gene expression changes during the in vitro differentiation of neural progenitor cells that were isolated from the subventricular zone of postnatal day 7 mice and grown in vitro as neurospheres. In two experimental series in which we withdrew epidermal growth factor and added the neurotrophins Neurotrophin-4 or BDNF, four time points were investigated: undifferentiated cells grown as neurospheres, and cells 24, 48, and 96 hr after differentiation. Expression changes of selected genes were confirmed by semiquantitative RT-PCR. Ten different groups of gene expression dynamics obtained by cluster analysis are described. To correlate selected gene expression changes to the localization of respective proteins, we performed immunostainings of cultured neurospheres and of brain sections from adult mice. Our results provide new insights into the genetic program of neural progenitor differentiation and give strong hints to as yet unknown cellular communications within the adult subventricular zone stem cell niche. PMID:15229246

  4. Serum polysialylated neural cell adhesion molecule in childhood neuroblastoma.

    PubMed Central

    Glüer, S.; Schelp, C.; Madry, N.; von Schweinitz, D.; Eckhardt, M.; Gerardy-Schahn, R.

    1998-01-01

    Neuroblastoma cells express the polysialylated form of the neural cell adhesion molecule (NCAM), which normally becomes restricted to a few neural tissues after embryogenesis. In this study, we investigated serum levels of polysialylated NCAM in 14 children with different grades and stages of neuroblastoma using an immunoluminescence assay, and compared the results to 269 healthy control subjects. Simultaneously, the polysialylated NCAM content of the tumours was determined by immunohistochemistry. Serum levels were dramatically elevated (more than sixfold) in children with advanced stages and fatal courses of disease, whereas children with differentiated tumour types and limited disease had low or normal levels. Serum concentrations correlated with the polysialylated NCAM content of the tumours, and they decreased during successful therapy. We therefore suggest polysialylated NCAM to be a useful marker monitoring childhood neuroblastoma. Images Figure 2 Figure 3 PMID:9662259

  5. 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

  6. Neural stem cells attacked by Zika virus.

    PubMed

    Nguyen, Ha Nam; Qian, Xuyu; Song, Hongjun; Ming, Guo-Li

    2016-07-01

    The current outbreak of Zika virus-associated diseases in South America and its threat to spread to other parts of the world has emerged as a global health emergency. Insights from cell and animal models to understand how Zika virus causes severe birth defects may lead to treatments and prevention of these diseases. PMID:27283801

  7. Characterization of neural stem cells and their progeny in the sensory circumventricular organs of adult mouse.

    PubMed

    Furube, Eriko; Morita, Mitsuhiro; Miyata, Seiji

    2015-11-01

    Although evidence has accumulated that neurogenesis and gliogenesis occur in the subventricular zone (SVZ) and subgranular zone (SGZ) of adult mammalian brains, recent studies indicate the presence of neural stem cells (NSCs) in adult brains, particularly the circumventricular regions. In the present study, we aimed to determine characterization of NSCs and their progenitor cells in the sensory circumventricular organs (CVOs), including organum vasculosum of the lamina terminalis, subfornical organ, and area postrema of adult mouse. There were two types of NSCs: tanycyte-like ependymal cells and astrocyte-like cells. Astrocyte-like NSCs proliferated slowly and oligodendrocyte progenitor cells (OPCs) and neural progenitor cells (NPCs) actively divided. Molecular marker protein expression of NSCs and their progenitor cells were similar to those reported in the SVZ and SGZ, except that astrocyte-like NSCs expressed S100β. These circumventricular NSCs possessed the capacity to give rise to oligodendrocytes and sparse numbers of neurons and astrocytes in the sensory CVOs and adjacent brain regions. The inhibition of vascular endothelial growth factor (VEGF) signaling by using a VEGF receptor-associated tyrosine kinase inhibitor AZD2171 largely suppressed basal proliferation of OPCs. A single systemic administration of lipopolysaccharide attenuated proliferation of OPCs and induced remarkable proliferation of microglia. The present study indicates that sensory circumventricular NSCs provide new neurons and glial cells in the sensory CVOs and adjacent brain regions. PMID:25994374

  8. Neural network adapted to wound cell analysis in surgical patients.

    PubMed

    Viljanto, Jouko; Koski, Antti

    2011-01-01

    Assessment of the real state of wound healing of closed surgical wounds is uncertain both clinically and from conventional laboratory tests. Therefore, a novel approach based on early analysis of exactly timed wound cells, computerized further with an artificial neural network, was developed. At the end of routine surgery performed on 481 children under 18 years of age, a specific wound drain Cellstick™ was inserted subcutaneously between the wound edges to harvest wound cells. The Cellsticks™ were removed from 1 to 50 hours, mainly at hour 3 or 24 postsurgery. Immediately, the cellular contents were washed out using a pump constructed for the purpose. After cytocentrifugation, the cells were stained and counted differentially. Based on their relative proportions at selected time intervals, an artificial self-organizing neural map was developed. This was further transformed to a unidirectional linear graph where each node represents one set of relative cell quantities. As early as 3 hours, but more precisely 24 hours after surgery, the location of the nodes on this graph showed individually the patients' initial speed of wound inflammatory cell response. Similarly, timed Cellstick™ specimens from new surgical patients could be analyzed, computerized, and compared with these node values to assess their initial speed in wound inflammatory cell response. Location of the node on the graph does not express the time lapse after surgery but the speed of wound inflammatory cell response in relation to that of other patients. PMID:21362082

  9. Neural stem cells could serve as a therapeutic material for age-related neurodegenerative diseases

    PubMed Central

    Suksuphew, Sarawut; Noisa, Parinya

    2015-01-01

    Progressively loss of neural and glial cells is the key event that leads to nervous system dysfunctions and diseases. Several neurodegenerative diseases, for instance Alzheimer’s disease, Parkinson’s disease, and Huntington’s disease, are associated to aging and suggested to be a consequence of deficiency of neural stem cell pool in the affected brain regions. Endogenous neural stem cells exist throughout life and are found in specific niches of human brain. These neural stem cells are responsible for the regeneration of new neurons to restore, in the normal circumstance, the functions of the brain. Endogenous neural stem cells can be isolated, propagated, and, notably, differentiated to most cell types of the brain. On the other hand, other types of stem cells, such as mesenchymal stem cells, embryonic stem cells, and induced pluripotent stem cells can also serve as a source for neural stem cell production, that hold a great promise for regeneration of the brain. The replacement of neural stem cells, either endogenous or stem cell-derived neural stem cells, into impaired brain is highly expected as a possible therapeutic mean for neurodegenerative diseases. In this review, clinical features and current routinely treatments of age-related neurodegenerative diseases are documented. Noteworthy, we presented the promising evidence of neural stem cells and their derivatives in curing such diseases, together with the remaining challenges to achieve the best outcome for patients. PMID:25815135

  10. 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

  11. Human skin neural crest progenitor cells are susceptible to BRAFV600E-induced transformation

    PubMed Central

    Kumar, SM; Dai, J; Li, S; Yang, R; Yu, H; Nathanson, KL; Liu, S; Zhou, H; Guo, J; Xu, X

    2013-01-01

    Adult stem cells are multipotent and persist in small numbers in adult tissues throughout the lifespan of an organism. Unlike differentiated cells, adult stem cells are intrinsically resistant to senescence. It is unclear how adult stem cells in solid organs respond to oncogenic stimulation and whether these cells have a role in tumor initiation. We report here that expression of BRAFV600E in human neural crest progenitor cells (hNCPCs) did not induce growth arrest as seen in human melanocytes, but instead, increased their cell proliferation capacity. These cells (hNCPCsV600E) acquired anchorage-independent growth ability and were weakly tumorigenic in vivo. Unlike in human melanocytes, BRAFV600E expression in hNCPCs did not induce p16INK4a expression. BRAFV600E induced elevated expression of CDK2, CDK4, MITF and EST1/2 protein in hNCPCs, and also induced melanocytic differentiation of these cells. Furthermore, overexpression of MITF in hNCPCsV600E dramatically increased their tumorigenicity and resulted in fully transformed tumor cells. These findings indicate that hNCPCs are susceptible to BRAFV600E-induced transformation, and MITF potentiates the oncogenic effect of BRAFV600E in these progenitor cells. These results suggest that the hNCPCs are potential targets for BRAFV600E-induced melanocytic tumor formation. PMID:23334329

  12. Prospective identification and culture of rat enteric neural stem cells (ENSCs).

    PubMed

    Gao, Tingting; Chen, Haijiao; Liu, Mei; Ge, Wenliang; Yin, Qiyou

    2016-05-01

    Hirschprung's disease (HD), a very common congenital abnormality in children, occurs mainly due to the congenital developmental defect of the enteric nervous system. The absence of enteric ganglia from the distal gut due to deletion in gut colonization by neural crest progenitor cells may lead to HD. The capacity to identify and isolate the enteric neuronal precursor cells from developing and mature tissues would enable the development of cell replacement therapies for HD. However, a mature method to culture these cells is a challenge. The present study aimed to propose a method to culture enteric neural stem cells (ENSCs) from the DsRed transgenic fetal rat gut. The culture medium used contained 15 % chicken embryo extract, basic fibroblast growth factor, and epidermal growth factor. ENSCs were cultured from embryonic day 18 in DsRed transgenic rat. Under inverted microscope and fluorescence staining, ENSCs proliferated to form small cell clusters on the second day of culture. The neurospheres-like structure were suspended in the medium, and there were some filaments between the adherent cells from day 3 to day 6 of the culture. The neurospheres were formed by ENSCs on day 8 of the culture. Network-like connections were formed between the adherent cells and differentiated cells after adding 10 % FBS. The differentiated cells were positive for neurofilament and glial fibrillary acidic protein antibodies. The present study established a method to isolate and culture ENSCs from E18 DsRed transgenic rats in the terminal stage of embryonic development. This study would offer a way to obtain plenty of cells for the future research on the transplantation of HD. PMID:25407731

  13. Differentiation of isolated human umbilical cord mesenchymal stem cells into neural stem cells

    PubMed Central

    Chen, Song; Zhang, Wei; Wang, Ji-Ming; Duan, Hong-Tao; Kong, Jia-Hui; Wang, Yue-Xin; Dong, Meng; Bi, Xue; Song, Jian

    2016-01-01

    AIM To investigate whether umbilical cord human mesenchymal stem cell (UC-MSC) was able to differentiate into neural stem cell and neuron in vitro. METHODS The umbilical cords were obtained from pregnant women with their written consent and the approval of the Clinic Ethnics Committee. UC-MSC were isolated by adherent culture in the medium contains 20% fetal bovine serum (FBS), then they were maintained in the medium contain 10% FBS and induced to neural cells in neural differentiation medium. We investigated whether UC-MSC was able to differentiate into neural stem cell and neuron in vitro by using flow cytometry, reverse transcriptase-polymerase chain reaction (RT-PCR) and immunofluorescence (IF) analyzes. RESULTS A substantial number of UC-MSC was harvested using the tissue explants adherent method at about 2wk. Flow cytometric study revealed that these cells expressed common markers of MSCs, such as CD105 (SH2), CD73 (SH3) and CD90. After induction of differentiation of neural stem cells, the cells began to form clusters; RT-PCR and IF showed that the neuron specific enolase (NSE) and neurogenic differentiation 1-positive cells reached 87.3%±14.7% and 72.6%±11.8%, respectively. Cells showed neuronal cell differentiation after induced, including neuron-like protrusions, plump cell body, obviously and stronger refraction. RT-PCR and IF analysis showed that microtubule-associated protein 2 (MAP2) and nuclear factor-M-positive cells reached 43.1%±10.3% and 69.4%±19.5%, respectively. CONCLUSION Human umbilical cord derived MSCs can be cultured and proliferated in vitro and differentiate into neural stem cells, which may be a valuable source for cell therapy of neurodegenerative eye diseases. PMID:26949608

  14. Syndecan-4 modulates the proliferation of neural cells and the formation of CaP axons during zebrafish embryonic neurogenesis

    PubMed Central

    Luo, Ning; Li, Hongda; Xiang, Bo; Qiao, Liangjun; He, Jiao; Ji, Yi; Liu, Yuan; Li, Siying; Lu, Ran; Li, Yu; Meng, Wentong; Wu, Yang; Xu, Hong; Mo, Xianming

    2016-01-01

    Syndecan-4 (Syn4), a single-pass transmembrane heparin sulphate proteoglycan (HSPG), plays significant role in the formation of focal adhesions and interacts with many growth factors to regulate cell migration and neural induction. Here, we show the new roles of syndecan-4(syn4) in zebrafish embryonic neurogenesis. Syn4 is broadly and dynamically expressed throughout the early stages of embryonic development. Knockdown of syn4 increases the expression of the marker genes of multiple types of neural cells. The increased expression of the marker genes is resulted from excessive proliferation of the neural cells. In addition, disrupting syn4 expression results in truncated and multiple aberrant branching of caudal primary (CaP) axons. Collectively, these data indicate that Syn4 suppresses the cellular proliferation during neurogenesis and is crucial for the formation of CaP axons during zebrafish embryogenesis. PMID:27143125

  15. 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.

  16. 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

  17. 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

  18. Isolation and characterization of neural stem cells from buffalo.

    PubMed

    Kumar, Kuldeep; Singh, Renu; Kumar, Manish; Agarwal, Pranjali; Mahapatra, P S; Kumar, Ajay; Malakar, Dhruba; Bag, Sadhan

    2014-06-01

    Neural stem cells (NSCs) are primordial, uncommitted cells postulated to give rise to the array of more specialized cells of the central nervous system (CNS). NSCs can self-renew and give rise to neurons, astrocytes and oligodendrocytes. NSCs are found in the CNS of mammalian organisms, and represent a promising resource for both fundamental research and CNS repair. Animal models of CNS damage have highlighted the potential benefit of NSC-based approaches. Present study described that buffalo neural stem cells (Bu-NSCs) were isolated and expanded rapidly from buffalo fetal brain in adherent culture. They were capable of multidifferentiation into neurons, astrocytes, and oligodendrocytes. Bu-NSCs were morphologically homogeneous and possessed high proliferation ability. The population doubled every 128.16 h. Normal buffalo karyotype was unchanged throughout the in vitro culture period. Together, we have isolated and cultured Bu-NSC from fetal brain that showed self-renewal, rapid proliferation and ability to differentiate into cells of nervous system. The availability of such cells may hold great interest for basic and applied neuroscience. PMID:24094244

  19. Low immunogenicity of in vitro-expanded human neural cells despite high MHC expression.

    PubMed

    Odeberg, Jenny; Piao, Jing-Hua; Samuelsson, Eva-Britt; Falci, Scott; Akesson, Elisabet

    2005-04-01

    The ability to expand human neural precursor cells in vitro offers new possibilities for future cell therapies. However, concern over immunologically based rejection of in vitro-expanded human neural cells confounds their use as donor cells. Here, we demonstrate that the expression of human leukocyte antigen (HLA) class I and II molecules, but not the co-stimulatory proteins CD40, CD80 and CD86, substantially increase during expansion of neurospheres. Furthermore, peripheral lymphocytes were unresponsive when co-cultured with in vitro-expanded neural cells. Taken together, these results suggest a low immunogenicity of these cultured human neural cells despite HLA incompatibility and high HLA expression. PMID:15748938

  20. Genetic instability in neural stem cells: an inconvenient truth?

    PubMed

    Harrison, Neil J

    2012-02-01

    The evolutionary struggles from which mutants arise have been documented in almost every living system. In this issue of the JCI, Varela and colleagues extend this list of systems to include neural derivatives of human embryonic stem cells, which they show exhibit a repeated gain of material from chromosome 1q. Although this raises safety issues for therapeutic use of such cells, the frequent observation of a particular change may direct screening strategies for detection and removal of these unwanted cellular variants. PMID:22269327

  1. Co-opting functions of cholinesterases in neural, limb and stem cell development.

    PubMed

    Vogel-Hopker, Astrid; Sperling, Laura E; Layer, Paul G

    2012-02-01

    Acetylcholinesterase (AChE) is a most remarkable protein, not only because it is one of the fastest enzymes in nature, but also since it appears in many molecular forms and is regulated by elaborate genetic networks. As revealed by sensitive histochemical procedures, AChE is expressed specifically in many tissues during development and in many mature organisms, as well as in healthy and diseased states. Therefore it is not surprising that there has been a long-standing search for additional, "non-classical" functions of cholinesterases (ChEs). In principle, AChE could either act nonenzymatically, e.g. exerting cell adhesive roles, or, alternatively, it could work within the frame of classic cholinergic systems, but in non-neural tissues. AChE might be considered a highly co-opting protein, since possibly it combines such various functions within one molecule. By presenting four different developmental cases, we here review i) the expression of ChEs in the neural tube and their close relation to cell proliferation and differentiation, ii) that AChE expression reflects a polycentric brain development, iii) the retina as a model for AChE functioning in neural network formation, and iv) nonneural ChEs in limb development and mature bones. Also, possible roles of AChE in neuritic growth and of cholinergic regulations in stem cells are briefly outlined. PMID:21933123

  2. 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…

  3. 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. PMID:27587351

  4. 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

  5. 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 cells in the zebrafish neural plate as they converge towards the dorsal midline before internalizing to form a neural keel. How these cells are regulated to ensure that they move together as a coherent tissue is unknown. Previous work in other systems has suggested that the underlying mesoderm may play a role in this process but this has not been shown directly in vivo. Results Here we analyze the roles of subjacent mesoderm in the coordination of neural cell movements during convergence of the zebrafish neural plate and neural keel formation. Live imaging demonstrates that the normal highly coordinated movements of neural plate cells are lost in the absence of underlying mesoderm and the movements of internalization and neural tube formation are severely disrupted. Despite this, neuroepithelial polarity develops in the abnormal neural primordium but the resulting tissue architecture is very disorganized. Conclusions We show that the movements of cells in the zebrafish neural plate are highly coordinated during the convergence and internalization movements of neurulation. Our results demonstrate that the underlying mesoderm is required for these coordinated cell movements in the zebrafish neural plate in vivo. PMID:24755297

  6. Comparative Effects of Human Neural Stem Cells and Oligodendrocyte Progenitor Cells on the Neurobehavioral Disorders of Experimental Autoimmune Encephalomyelitis Mice

    PubMed Central

    Bae, Dae-Kwon; Park, Dongsun; Lee, Sun Hee; Yang, Goeun; Kyung, Jangbeen; Kim, Dajeong; Shin, Kyungha; Choi, Ehn-Kyoung; Kim, Gonhyung; Hong, Jin Tae; Kim, Seung U.

    2016-01-01

    Since multiple sclerosis (MS) is featured with widespread demyelination caused by autoimmune response, we investigated the recovery effects of F3.olig2 progenitors, established by transducing human neural stem cells (F3 NSCs) with Olig2 transcription factor, in myelin oligodendrocyte glycoprotein- (MOG-) induced experimental autoimmune encephalomyelitis (EAE) model mice. Six days after EAE induction, F3 or F3.olig2 cells (1 × 106/mouse) were intravenously transplanted. MOG-injected mice displayed severe neurobehavioral deficits which were remarkably attenuated and restored by cell transplantation, in which F3.olig2 cells were superior to its parental F3 cells. Transplanted cells migrated to the injured spinal cord, matured to oligodendrocytes, and produced myelin basic proteins (MBP). The F3.olig2 cells expressed growth and neurotrophic factors including brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), ciliary neurotrophic factor (CNTF), and leukemia inhibitory factor (LIF). In addition, the transplanted cells markedly attenuated inflammatory cell infiltration, reduced cytokine levels in the spinal cord and lymph nodes, and protected host myelins. The results indicate that F3.olig2 cells restore neurobehavioral symptoms of EAE mice by regulating autoimmune inflammatory responses as well as by stimulating remyelination and that F3.olig2 progenitors could be a candidate for the cell therapy of demyelinating diseases including MS. PMID:27429621

  7. Automatic discovery of cell types and microcircuitry from neural connectomics.

    PubMed

    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. PMID:25928186

  8. 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

  9. 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. PMID:25923707

  10. Inducible regulation of GDNF expression in human neural stem cells.

    PubMed

    Wang, ShuYan; Ren, Ping; Guan, YunQian; Zou, ChunLin; Fu, LinLin; Zhang, Yu

    2013-01-01

    Glial cell derived neurotrophic factor (GDNF) holds promises for treating neurodegenerative diseases such as Parkinson's disease. Human neural stem cells (hNSCs) have proved to be a suitable cell delivery vehicle for the safe and efficient introduction of GDNF into the brain. In this study, we used hNSCs-infected with a lentivirus encoding GDNF and the hygromycin resistance gene as such vehicles. A modified tetracycline operator 7 (tetO7) was inserted into a region upstream of the EF1-α promoter to drive GDNF expression. After hygromycin selection, hNSCs were infected with a lentivirus encoding a KRAB-tetracycline repressor fusion protein (TTS). TTS bound to tetO7 and suppressed the expression of GDNF in hNSCs. Upon administration of doxycycline (Dox) the TTS-tetO7 complex separated and the expression of GDNF resumed. The hNSCs infected with GDNF expressed the neural stem cell specific markers, nestin and sox2, and exhibited no significant change in proliferation rate. However, the rate of apoptosis in hNSCs expressing GDNF was lower compared with normal NSCs in response to actinomycin treatment. Furthermore, a higher percentage of Tuj-1 positive cells were obtained from GDNF-producing NSCs under conditions that induced differentiation compared to control NSCs. The inducible expression of GDNF in hNSCs may provide a system for the controllable delivery of GDNF in patients with neurodegenerative diseases. PMID:23269553