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Sample records for neural cells exposed

  1. From the Cover: Exposing Imidacloprid Interferes With Neurogenesis Through Impacting on Chick Neural Tube Cell Survival.

    PubMed

    Liu, Meng; Wang, Guang; Zhang, Shi-Yao; Zhong, Shan; Qi, Guo-Long; Wang, Chao-Jie; Chuai, Manli; Lee, Kenneth Ka Ho; Lu, Da-Xiang; Yang, Xuesong

    2016-09-01

    As a neonicotinoid pesticide, imidacloprid is widely used to control insects in agriculture and fleas on domestic animals. However, it is not known whether imidacloprid exposure negatively affects neurogenesis during embryonic development. In this study, using a chick embryo model, we investigated the effects of imidacloprid exposure on neurogenesis at the earliest stage and during late-stage embryo development. Exposing HH0 chick embryos to imidacloprid in EC culture caused neural tube defects (NTDs) and neuronal differentiation dysplasia as determined by NF/Tuj1 labeling. Furthermore, we found that F-actin accumulation on the apical side of the neural tube was suppressed by exposure to imidacloprid, and the expression of BMP4 and Shh on the dorsal and ventral sides of the neural tubes, respectively, were also reduced, which in turn affects the dorsolateral hinge points during bending of the neural plate. In addition, exposure to imidacloprid reduced cell proliferation and increased cell apoptosis, as determined by pHIS3 labeling and TUNEL staining, respectively, also contributing to the malformation. We obtained similar results in late-stage embryos exposed to imidacloprid. Finally, a bioinformatics analysis was employed to determine which genes identified in this study were involved in NTDs. The experimental evidence and bioinformatics analysis suggested that imidacloprid exposure during chick embryo development could increase the risk of NTDs and neural dysplasia.

  2. From the Cover: Exposing Imidacloprid Interferes With Neurogenesis Through Impacting on Chick Neural Tube Cell Survival.

    PubMed

    Liu, Meng; Wang, Guang; Zhang, Shi-Yao; Zhong, Shan; Qi, Guo-Long; Wang, Chao-Jie; Chuai, Manli; Lee, Kenneth Ka Ho; Lu, Da-Xiang; Yang, Xuesong

    2016-09-01

    As a neonicotinoid pesticide, imidacloprid is widely used to control insects in agriculture and fleas on domestic animals. However, it is not known whether imidacloprid exposure negatively affects neurogenesis during embryonic development. In this study, using a chick embryo model, we investigated the effects of imidacloprid exposure on neurogenesis at the earliest stage and during late-stage embryo development. Exposing HH0 chick embryos to imidacloprid in EC culture caused neural tube defects (NTDs) and neuronal differentiation dysplasia as determined by NF/Tuj1 labeling. Furthermore, we found that F-actin accumulation on the apical side of the neural tube was suppressed by exposure to imidacloprid, and the expression of BMP4 and Shh on the dorsal and ventral sides of the neural tubes, respectively, were also reduced, which in turn affects the dorsolateral hinge points during bending of the neural plate. In addition, exposure to imidacloprid reduced cell proliferation and increased cell apoptosis, as determined by pHIS3 labeling and TUNEL staining, respectively, also contributing to the malformation. We obtained similar results in late-stage embryos exposed to imidacloprid. Finally, a bioinformatics analysis was employed to determine which genes identified in this study were involved in NTDs. The experimental evidence and bioinformatics analysis suggested that imidacloprid exposure during chick embryo development could increase the risk of NTDs and neural dysplasia. PMID:27444676

  3. Oxidative stress response in neural stem cells exposed to different superparamagnetic iron oxide nanoparticles.

    PubMed

    Pongrac, Igor M; Pavičić, Ivan; Milić, Mirta; Brkić Ahmed, Lada; Babič, Michal; Horák, Daniel; Vinković Vrček, Ivana; Gajović, Srećko

    2016-01-01

    Biocompatibility, safety, and risk assessments of superparamagnetic iron oxide nanoparticles (SPIONs) are of the highest priority in researching their application in biomedicine. One improvement in the biological properties of SPIONs may be achieved by different functionalization and surface modifications. This study aims to investigate how a different surface functionalization of SPIONs - uncoated, coated with d-mannose, or coated with poly-l-lysine - affects biocompatibility. We sought to investigate murine neural stem cells (NSCs) as important model system for regenerative medicine. To reveal the possible mechanism of toxicity of SPIONs on NSCs, levels of reactive oxygen species, intracellular glutathione, mitochondrial membrane potential, cell-membrane potential, DNA damage, and activities of SOD and GPx were examined. Even in cases where reactive oxygen species levels were significantly lowered in NSCs exposed to SPIONs, we found depleted intracellular glutathione levels, altered activities of SOD and GPx, hyperpolarization of the mitochondrial membrane, dissipated cell-membrane potential, and increased DNA damage, irrespective of the surface coating applied for SPION stabilization. Although surface coating should prevent the toxic effects of SPIONs, our results showed that all of the tested SPION types affected the NSCs similarly, indicating that mitochondrial homeostasis is their major cellular target. Despite the claimed biomedical benefits of SPIONs, the refined determination of their effects on various cellular functions presented in this work highlights the need for further safety evaluations. This investigation helps to fill the knowledge gaps on the criteria that should be considered in evaluating the biocompatibility and safety of novel nanoparticles. PMID:27217748

  4. Persistent oxidative stress in human neural stem cells exposed to low fluences of charged particles.

    PubMed

    Baulch, Janet E; Craver, Brianna M; Tran, Katherine K; Yu, Liping; Chmielewski, Nicole; Allen, Barrett D; Limoli, Charles L

    2015-08-01

    Exposure to the space radiation environment poses risks for a range of deleterious health effects due to the unique types of radiation encountered. Galactic cosmic rays are comprised of a spectrum of highly energetic nuclei that deposit densely ionizing tracks of damage along the particle trajectory. These tracks are distinct from those generated by the more sparsely ionizing terrestrial radiations, and define the geometric distribution of the complex cellular damage that results when charged particles traverse the tissues of the body. The exquisite radiosensitivity of multipotent neural stem and progenitor cells found within the neurogenic regions of the brain predispose the central nervous system to elevated risks for radiation induced sequelae. Here we show that human neural stem cells (hNSC) exposed to different charged particles at space relevant fluences exhibit significant and persistent oxidative stress. Radiation induced oxidative stress was found to be most dependent on total dose rather than on the linear energy transfer of the incident particle. The use of redox sensitive fluorogenic dyes possessing relative specificity for hydroxyl radicals, peroxynitrite, nitric oxide (NO) and mitochondrial superoxide confirmed that most irradiation paradigms elevated reactive oxygen and nitrogen species (ROS and RNS, respectively) in hNSC over a 1 week interval following exposure. Nitric oxide synthase (NOS) was not the major source of elevated nitric oxides, as the use of NOS inhibitors had little effect on NO dependent fluorescence. Our data provide extensive evidence for the capability of low doses of charged particles to elicit marked changes in the metabolic profile of irradiated hNSC. Radiation induced changes in redox state may render the brain more susceptible to the development of neurocognitive deficits that could affect an astronaut's ability to perform complex tasks during extended missions in deep space. PMID:25800120

  5. Persistent oxidative stress in human neural stem cells exposed to low fluences of charged particles

    PubMed Central

    Baulch, Janet E.; Craver, Brianna M.; Tran, Katherine K.; Yu, Liping; Chmielewski, Nicole; Allen, Barrett D.; Limoli, Charles L.

    2015-01-01

    Exposure to the space radiation environment poses risks for a range of deleterious health effects due to the unique types of radiation encountered. Galactic cosmic rays are comprised of a spectrum of highly energetic nuclei that deposit densely ionizing tracks of damage along the particle trajectory. These tracks are distinct from those generated by the more sparsely ionizing terrestrial radiations, and define the geometric distribution of the complex cellular damage that results when charged particles traverse the tissues of the body. The exquisite radiosensitivity of multipotent neural stem and progenitor cells found within the neurogenic regions of the brain predispose the central nervous system to elevated risks for radiation induced sequelae. Here we show that human neural stem cells (hNSC) exposed to different charged particles at space relevant fluences exhibit significant and persistent oxidative stress. Radiation induced oxidative stress was found to be most dependent on total dose rather than on the linear energy transfer of the incident particle. The use of redox sensitive fluorogenic dyes possessing relative specificity for hydroxyl radicals, peroxynitrite, nitric oxide (NO) and mitochondrial superoxide confirmed that most irradiation paradigms elevated reactive oxygen and nitrogen species (ROS and RNS, respectively) in hNSC over a 1 week interval following exposure. Nitric oxide synthase (NOS) was not the major source of elevated nitric oxides, as the use of NOS inhibitors had little effect on NO dependent fluorescence. Our data provide extensive evidence for the capability of low doses of charged particles to elicit marked changes in the metabolic profile of irradiated hNSC. Radiation induced changes in redox state may render the brain more susceptible to the development of neurocognitive deficits that could affect an astronaut’s ability to perform complex tasks during extended missions in deep space. PMID:25800120

  6. Oxidative stress response in neural stem cells exposed to different superparamagnetic iron oxide nanoparticles

    PubMed Central

    Pongrac, Igor M; Pavičić, Ivan; Milić, Mirta; Brkić Ahmed, Lada; Babič, Michal; Horák, Daniel; Vinković Vrček, Ivana; Gajović, Srećko

    2016-01-01

    Biocompatibility, safety, and risk assessments of superparamagnetic iron oxide nanoparticles (SPIONs) are of the highest priority in researching their application in biomedicine. One improvement in the biological properties of SPIONs may be achieved by different functionalization and surface modifications. This study aims to investigate how a different surface functionalization of SPIONs – uncoated, coated with d-mannose, or coated with poly-l-lysine – affects biocompatibility. We sought to investigate murine neural stem cells (NSCs) as important model system for regenerative medicine. To reveal the possible mechanism of toxicity of SPIONs on NSCs, levels of reactive oxygen species, intracellular glutathione, mitochondrial membrane potential, cell-membrane potential, DNA damage, and activities of SOD and GPx were examined. Even in cases where reactive oxygen species levels were significantly lowered in NSCs exposed to SPIONs, we found depleted intracellular glutathione levels, altered activities of SOD and GPx, hyperpolarization of the mitochondrial membrane, dissipated cell-membrane potential, and increased DNA damage, irrespective of the surface coating applied for SPION stabilization. Although surface coating should prevent the toxic effects of SPIONs, our results showed that all of the tested SPION types affected the NSCs similarly, indicating that mitochondrial homeostasis is their major cellular target. Despite the claimed biomedical benefits of SPIONs, the refined determination of their effects on various cellular functions presented in this work highlights the need for further safety evaluations. This investigation helps to fill the knowledge gaps on the criteria that should be considered in evaluating the biocompatibility and safety of novel nanoparticles. PMID:27217748

  7. Gypenosides Protected the Neural Stem Cells in the Subventricular Zone of Neonatal Rats that Were Prenatally Exposed to Ethanol

    PubMed Central

    Dong, Lun; Yang, Kun-Qi; Fu, Wen-Yan; Shang, Zhen-Hua; Zhang, Qing-Yu; Jing, Fang-Miao; Li, Lin-Lin; Xin, Hua; Wang, Xiao-Jing

    2014-01-01

    Fetal alcohol spectrum disorder (FASD) can cause severe mental retardation in children who are prenatally exposed to ethanol. The effects of prenatal and early postnatal ethanol exposure on adult hippocampal neurogenesis have been investigated; however, the effects of prenatal ethanol exposure on the subventricular zone (SVZ) have not. Gypenosides (GPs) have been reported to have neuroprotective effects in addition to other bioactivities. The effects of GPs on neural stem cells (NSCs) in the FASD model are unknown. Here, we test the effect of prenatal ethanol exposure on the neonatal SVZ, and the protection potential of GPs on NSCs in FASD rats. Our results show that prenatal ethanol exposure can suppress the cell proliferation and differentiation of neural stem cells in the neonatal SVZ and that GPs (400 mg/kg/day) can significantly increase the cell proliferation and differentiation of neural stem cells inhibited by ethanol. Our data indicate that GPs have neuroprotective effects on the NSCs and can enhance the neurogenesis inhibited by ethanol within the SVZ of neonatal rats. These findings provide new evidence for a potential therapy involving GPs for the treatment of FASD. PMID:25464383

  8. Protective effects of 1α,25-Dihydroxyvitamin D3 on cultured neural cells exposed to catalytic iron.

    PubMed

    Uberti, Francesca; Morsanuto, Vera; Bardelli, Claudio; Molinari, Claudio

    2016-06-01

    Recent studies have postulated a role for vitamin D and its receptor on cerebral function, and anti-inflammatory, immunomodulatory and neuroprotective effects have been described; vitamin D can inhibit proinflammatory cytokines and nitric oxide synthesis during various neurodegenerative insults, and may be considered as a potential drug for the treatment of these disorders. In addition, iron is crucial for neuronal development and neurotransmitter production in the brain, but its accumulation as catalytic form (Fe(3+)) impairs brain function and causes the dysregulation of iron metabolism leading to tissue damage due to the formation of toxic free radicals (ROS). This research was planned to study the role of vitamin D to prevent iron damage in neuroblastoma BE(2)M17 cells. Mechanisms involved in neurodegeneration, including cell viability, ROS production, and the most common intracellular pathways were studied. Pretreatment with calcitriol (the active form of vitamin D) reduced cellular injury induced by exposure to catalytic iron.

  9. Neuroprotective effects of human mesenchymal stem cells on neural cultures exposed to 6-hydroxydopamine: implications for reparative therapy in Parkinson's disease.

    PubMed

    Cova, Lidia; Bossolasco, Patrizia; Armentero, Marie-Therese; Diana, Valentina; Zennaro, Eleonora; Mellone, Manuela; Calzarossa, Cinzia; Cerri, Silvia; Deliliers, Giorgio Lambertenghi; Polli, Elio; Blandini, Fabio; Silani, Vincenzo

    2012-03-01

    Stem cell (SC) transplantation represents a promising tool to treat neurodegenerative disorders, such as Parkinson's disease (PD), but positive therapeutic outcomes require elucidation of the biological mechanisms involved. Therefore, we investigated human Mesenchymal SCs (hMSCs) ability to protect murine differentiated Neural SCs (mdNSCs) against the cytotoxic effects of 6-hydroxydopamine (6-OHDA) in a co-culture model mimicking the in vivo neurovascular niche. The internalization of 6-OHDA mainly relies on its uptake by the dopamine active transporter (DAT), but its toxicity could also involve other pathways. We demonstrated that mdNSCs consistently expressed DAT along the differentiative process. Exposure to 6-OHDA did not affect hMSCs, but induced DAT-independent apoptosis in mdNSCs with generation of reactive oxygen species and caspases 3/7 activation. The potential neuroprotective action of hMSCs on mdNSCs exposed to 6-OHDA was tested in different co-culture conditions, in which hMSCs were added to mdNSCs prior to, simultaneously, or after 6-OHDA treatment. In the presence of the neurotoxin, the majority of mdNSCs acquired an apoptotic phenotype, while co-cultures with hMSCs significantly increased their survival (up to 70%) in all conditions. Multiplex human angiogenic array analysis on the conditioned media demonstrated that cytokine release by hMSCs was finely modulated. Moreover, sole growth factor addition yielded a similar neuroprotective effect on mdNSCs. In conclusion, our findings demonstrate that hMSCs protect mdNSCs against 6-OHDA neurotoxicity, and rescue cells from ongoing neurodegeneration likely through the release of multiple cytokines. Our findings provide novel insights for the development of therapeutic strategies designed to counteract the neurodegenerative processes of PD.

  10. Functional Consequences of Radiation-Induced Oxidative Stress in Cultured Neural Stem Cells and the Brain Exposed to Charged Particle Irradiation

    PubMed Central

    Tseng, Bertrand P.; Giedzinski, Erich; Izadi, Atefeh; Suarez, Tatiana; Lan, Mary L.; Tran, Katherine K.; Acharya, Munjal M.; Nelson, Gregory A.; Raber, Jacob; Parihar, Vipan K.

    2014-01-01

    Abstract Aims: Redox homeostasis is critical in regulating the fate and function of multipotent cells in the central nervous system (CNS). Here, we investigated whether low dose charged particle irradiation could elicit oxidative stress in neural stem and precursor cells and whether radiation-induced changes in redox metabolism would coincide with cognitive impairment. Results: Low doses (<1 Gy) of charged particles caused an acute and persistent oxidative stress. Early after (<1 week) irradiation, increased levels of reactive oxygen and nitrogen species were generally dose responsive, but were less dependent on dose weeks to months thereafter. Exposure to ion fluences resulting in less than one ion traversal per cell was sufficient to elicit radiation-induced oxidative stress. Whole body irradiation triggered a compensatory response in the rodent brain that led to a significant increase in antioxidant capacity 2 weeks following exposure, before returning to background levels at week 4. Low dose irradiation was also found to significantly impair novel object recognition in mice 2 and 12 weeks following irradiation. Innovation: Data provide evidence that acute exposure of neural stem cells and the CNS to very low doses and fluences of charged particles can elicit a persisting oxidative stress lasting weeks to months that is associated with impaired cognition. Conclusions: Exposure to low doses of charged particles causes a persistent oxidative stress and cognitive impairment over protracted times. Data suggest that astronauts subjected to space radiation may develop a heightened risk for mission critical performance decrements in space, along with a risk of developing long-term neurocognitive sequelae. Antioxid. Redox Signal. 20, 1410–1422. PMID:23802883

  11. Neural Tube Defects In Mice Exposed To Tap Water

    PubMed Central

    Mallela, Murali K; Werre, Stephen R; Hrubec, Terry C

    2010-01-01

    In May of 2006 we suddenly began to observe neural tube defects (NTDs) in embryos of untreated control mice. We hypothesized the mice were being exposed unknowingly to a teratogenic agent and investigated the cause. Our results suggested that NTDs were not resulting from bedding material, feed, strain or source of the mice. Additionally, mice were negative for routine and comprehensive screens of pathogens. To further test whether the NTDs resulted from infectious or genetic cause localized to our facility, we obtained three strains of timed pregnant mice from commercial suppliers located in 4 different states. All strains and sources of mice arrived in our laboratory with NTDs, implying that commercially available mice were possibly exposed to a teratogen prior to purchase. Our investigation eventually concluded that exposure to tap water was causing the NTDs. The incidence of NTDs was greatest in purchased mice provided tap water and lowest in purchased mice provided distilled deionized water (DDI). Providing mice DDI water for two generations (F2-DDI) eliminated the NTDs. When F2-DDI mice were provided tap water from three different urban areas prior to breeding, their offspring again developed NTDs. Increased length of exposure to tap water significantly increased the incidence of NTDs. These results indicate that a contaminant in municipal tap water is likely causing NTDs in mice. The unknown teratogen appears to have a wide geographic distribution but has not yet been identified. Water analysis is currently underway to identify candidate contaminants that might be responsible for the malformations. PMID:20549630

  12. Ozone exposed epithelial cells modify cocultured natural killer cells

    PubMed Central

    Müller, Loretta; Brighton, Luisa E.

    2013-01-01

    Ozone (O3) causes significant adverse health effects worldwide. Nasal epithelial cells (NECs) are among the first sites within the respiratory system to be exposed to inhaled air pollutants. They recruit, activate, and interact with immune cells via soluble mediators and direct cell-cell contacts. Based on our recent observation demonstrating the presence of natural killer (NK) cells in nasal lavages, the goal of this study was to establish a coculture model of NECs and NK cells and examine how exposure to O3 modifies this interaction. Flow cytometry analysis was used to assess immunophenotypes of NK cells cocultured with either air- or O3-exposed NECs. Our data show that coculturing NK cells with O3-exposed NECs decreased intracellular interferon-γ (IFN-γ), enhanced, albeit not statistically significant, IL-4, and increased CD16 expression on NK cells compared with air controls. Additionally, the cytotoxicity potential of NK cells was reduced after coculturing with O3-exposed NECs. To determine whether soluble mediators released by O3-exposed NECs caused this shift, apical and basolateral supernatants of air- and O3-exposed NECs were used to stimulate NK cells. While the conditioned media of O3-exposed NECs alone did not reduce intracellular IFN-γ, O3 enhanced the expression of NK cell ligands ULBP3 and MICA/B on NECs. Blocking ULBP3 and MICA/B reversed the effects of O3-exposed NECs on IFN-γ production in NK cells. Taken together, these data showed that interactions between NECs and NK cells in the context of O3 exposure changes NK cell activity via direct cell-cell interactions and is dependent on ULBP3/MICA/B expressed on NECs. PMID:23241529

  13. Diabetes and apoptosis: neural crest cells and neural tube

    PubMed Central

    Chappell, James H.; Dan Wang, Xiao

    2016-01-01

    Birth defects resulting from diabetic pregnancy are associated with apoptosis of a critical mass of progenitor cells early during the formation of the affected organ(s). Insufficient expression of genes that regulate viability of the progenitor cells is responsible for the apoptosis. In particular, maternal diabetes inhibits expression of a gene, Pax3, that encodes a transcription factor which is expressed in neural crest and neuroepithelial cells. As a result of insufficient Pax3, cardiac neural crest and neuroepithelial cells undergo apoptosis by a process dependent on the p53 tumor suppressor protein. This, then provides a cellular explanation for the cardiac outflow tract and neural tube and defects induced by diabetic pregnancy. PMID:19333760

  14. Embryonic stem cell neurogenesis and neural specification.

    PubMed

    Germain, Noélle; Banda, Erin; Grabel, Laura

    2010-10-15

    The prospect of using embryonic stem cell (ESC)-derived neural progenitors and neurons to treat neurological disorders has led to great interest in defining the conditions that guide the differentiation of ESCs, and more recently induced pluripotent stem cells (iPSCs), into neural stem cells (NSCs) and a variety of neuronal and glial subtypes. Over the past decade, researchers have looked to the embryo to guide these studies, applying what we know about the signaling events that direct neural specification during development. This has led to the design of a number of protocols that successfully promote ESC neurogenesis, terminating with the production of neurons and glia with diverse regional addresses and functional properties. These protocols demonstrate that ESCs undergo neural specification in two, three, and four dimensions, mimicking the cell-cell interactions, patterning, and timing that characterizes the in vivo process. We therefore propose that these in vitro systems can be used to examine the molecular regulation of neural specification.

  15. 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.

  16. 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

  17. Neural stem cells-trends and advances.

    PubMed

    English, Denis; Sharma, Neel K; Sharma, Kaushal; Anand, Akshay

    2013-04-01

    For many years, accepted dogma held that brain is a static organ with no possibility of regeneration of cells in injured or diseased human brain. However, recent preclinical reports have shown regenerative potential of neural stem cells using various injury models. This has resulted in renewed hope for those suffering from spinal cord injury and neural damage. As the potential of stem cell therapy gained impact, these claims, in particular, led to widespread enthusiasm that acute and chronic injury of the nervous system would soon be a problem of the past. The devastation caused by injury or diseases of the brain and spinal cord led to wide premature acceptance that "neural stem cells (NSCs)" derived from embryonic, fetal or adult sources would soon be effective in reversing neural and spinal trauma. However, neural therapy with stem cells has not been realized to its fullest extent. Although, discrete population of regenerative stem cells seems to be present in specific areas of human brain, the function of these cells is unclear. However, similar cells in animals seem to play important role in postnatal growth as well as recovery of neural tissue from injury, anoxia, or disease.

  18. Neural Stem Cells and Ischemic Brain

    PubMed Central

    Zhang, Zhenggang; Chopp, Michael

    2016-01-01

    Stroke activates neural stem cells in the ventricular-subventricular zone (V/SVZ) of the lateral ventricle, which increases neuroblasts and oligodendrocyte progenitor cells (OPCs). Within the ischemic brain, neural stem cells, neuroblasts and OPCs appear to actively communicate with cerebral endothelial cells and other brain parenchymal cells to mediate ischemic brain repair; however, stroke-induced neurogenesis unlikely plays any significant roles in neuronal replacement. In this mini-review, we will discuss recent findings how intercellular communications between stroke-induced neurogenesis and oligodendrogenesis and brain parenchymal cells could potentially facilitate brain repair processes. PMID:27488979

  19. 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

  20. 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.

  1. Embryonic stem cell neurogenesis and neural specification.

    PubMed

    Germain, Noélle; Banda, Erin; Grabel, Laura

    2010-10-15

    The prospect of using embryonic stem cell (ESC)-derived neural progenitors and neurons to treat neurological disorders has led to great interest in defining the conditions that guide the differentiation of ESCs, and more recently induced pluripotent stem cells (iPSCs), into neural stem cells (NSCs) and a variety of neuronal and glial subtypes. Over the past decade, researchers have looked to the embryo to guide these studies, applying what we know about the signaling events that direct neural specification during development. This has led to the design of a number of protocols that successfully promote ESC neurogenesis, terminating with the production of neurons and glia with diverse regional addresses and functional properties. These protocols demonstrate that ESCs undergo neural specification in two, three, and four dimensions, mimicking the cell-cell interactions, patterning, and timing that characterizes the in vivo process. We therefore propose that these in vitro systems can be used to examine the molecular regulation of neural specification. PMID:20589755

  2. 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

  3. Enhanced emotion regulation capacity and its neural substrates in those exposed to moderate childhood adversity

    PubMed Central

    Schweizer, Susanne; Walsh, Nicholas D.; Stretton, Jason; Dunn, Valerie J.; Goodyer, Ian M.; Dalgleish, Tim

    2016-01-01

    Individuals exposed to childhood adversities (CA) present with emotion regulation (ER) difficulties in later life, which have been identified as risk and maintenance factors for psychopathologies. However, it is unclear if CA negatively impacts on ER capacity per se or whether observed regulation difficulties are a function of the challenging circumstances in which ER is being deployed. In this longitudinal study, we aimed to clarify this association by investigating the behavioral and neural effects of exposure to common moderate CA (mCA) on a laboratory measure of ER capacity in late adolescence/young adulthood. Our population-derived samples of adolescents/young adults (N = 53) were administered a film-based ER-task during functional magnetic resonance imaging that allowed evaluation of ER across mCA-exposure. mCA-exposure was associated with enhanced ER capacity over both positive and negative affect. At the neural level, the better ER of negative material in those exposed to mCA was associated with reduced recruitment of ER-related brain regions, including the prefrontal cortex and temporal gyrus. In addition mCA-exposure was associated with a greater down-regulation of the amygdala during ER of negative material. The implications of these findings for our understanding of the effects of mCA on the emergence of resilience in adolescence are discussed. PMID:26341903

  4. Neural autoantibodies and neurophysiologic abnormalities in patients exposed to molds in water-damaged buildings.

    PubMed

    Campbell, Andrew W; Thrasher, Jack D; Madison, Roberta A; Vojdani, Aristo; Gray, Michael R; Johnson, Al

    2003-08-01

    Adverse health effects of fungal bioaerosols on occupants of water-damaged homes and other buildings have been reported. Recently, it has been suggested that mold exposure causes neurological injury. The authors investigated neurological antibodies and neurophysiological abnormalities in patients exposed to molds at home who developed symptoms of peripheral neuropathy (i.e., numbness, tingling, tremors, and muscle weakness in the extremities). Serum samples were collected and analyzed with the enzyme-linked immunosorbent assay (ELISA) technique for antibodies to myelin basic protein, myelin-associated glycoprotein, ganglioside GM1, sulfatide, myelin oligodendrocyte glycoprotein, alpha-B-crystallin, chondroitin sulfate, tubulin, and neurofilament. Antibodies to molds and mycotoxins were also determined with ELISA, as reported previously. Neurophysiologic evaluations for latency, amplitude, and velocity were performed on 4 motor nerves (median, ulnar, peroneal, and tibial), and for latency and amplitude on 3 sensory nerves (median, ulnar, and sural). Patients with documented, measured exposure to molds had elevated titers of antibodies (immunoglobulin [Ig]A, IgM, and IgG) to neural-specific antigens. Nerve conduction studies revealed 4 patient groupings: (1) mixed sensory-motor polyneuropathy (n = 55, abnormal), (2) motor neuropathy (n = 17, abnormal), (3) sensory neuropathy (n = 27, abnormal), and (4) those with symptoms but no neurophysiological abnormalities (n = 20, normal controls). All groups showed significantly increased autoantibody titers for all isotypes (IgA, IgM, and IgG) of antibodies to neural antigens when compared with 500 healthy controls. Groups 1 through 3 also exhibited abnormal neurophysiologic findings. The authors concluded that exposure to molds in water-damaged buildings increased the risk for development of neural autoantibodies, peripheral neuropathy, and neurophysiologic abnormalities in exposed individuals. PMID:15259425

  5. 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

  6. Disrupted insula-based neural circuit organization and conflict interference in trauma-exposed youth.

    PubMed

    Marusak, Hilary A; Etkin, Amit; Thomason, Moriah E

    2015-01-01

    Childhood trauma exposure is a potent risk factor for psychopathology. Emerging research suggests that aberrant saliency processing underlies the link between early trauma exposure and later cognitive and socioemotional deficits that are hallmark of several psychiatric disorders. Here, we examine brain and behavioral responses during a face categorization conflict task, and relate these to intrinsic connectivity of the salience network (SN). The results demonstrate a unique pattern of SN dysfunction in youth exposed to trauma (n = 14) relative to comparison youth (n = 19) matched on age, sex, IQ, and sociodemographic risk. We find that trauma-exposed youth are more susceptible to conflict interference and this correlates with higher fronto-insular responses during conflict. Resting-state functional connectivity data collected in the same participants reveal increased connectivity of the insula to SN seed regions that is associated with diminished reward sensitivity, a critical risk/resilience trait following stress. In addition to altered intrinsic connectivity of the SN, we observed altered connectivity between the SN and default mode network (DMN) in trauma-exposed youth. These data uncover network-level disruptions in brain organization following one of the strongest predictors of illness, early life trauma, and demonstrate the relevance of observed neural effects for behavior and specific symptom dimensions. SN dysfunction may serve as a diathesis that contributes to illness and negative outcomes following childhood trauma.

  7. Immunological control of adult neural stem cells

    PubMed Central

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

    2010-01-01

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

  8. 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.

  9. Neural crest development: the interplay between morphogenesis and cell differentiation.

    PubMed

    Erickson, C A; Reedy, M V

    1998-01-01

    The final pattern of tissues established during embryogenesis reflects the outcome of two developmental processes: differentiation and morphogenesis. Avian neural crest cells are an excellent system in which to study this interaction. In the first phase of neural crest cell migration, neural crest cells separate from the neural epithelium via an epithelial-mesenchymal transformation. We present three models to account for this process: (1) separation by asymmetric mitosis, (2) separation by generating tractional force in order to rupture cell adhesions and (3) loss of expression or function of cell-cell adhesion molecules that keep the presumptive neural crest cells tethered to the neural epithelium. Evidence is presented that the segregation of the neural crest lineage apart from the neural epithelium is caused by the epithelial-mesenchymal transformation. Once they have detached from the neural tube, neural crest cells take two pathways in the trunk of the chick embryo: (1) the ventral path between the neural tube and somite, where neural crest cells give rise to neurons and glial cells of the peripheral nervous systems, and (2) the dorsolateral path between the ectoderm and dermamyotome of the somite, where they differentiate into pigment cells of the skin. We present data to suggest that the migration and differentiation along the ventral path is controlled primarily by environmental cues, which we refer to as the environment-directed model of neural crest morphogenesis. Conversely, only melanoblasts can migrate into the dorsolateral space, and the ability to invade that path is dependent upon their early specification as melanoblasts. We call this the phenotype-directed model for neural crest cell migration and suggest that this latter model for the positioning of neural crest derivatives in the embryo may be more common than previously suspected. These observations invite a re-examination of patterning of other crest derivates, which previously were believed

  10. Characterization of the phosphatidylserine-exposing subpopulation of sickle cells.

    PubMed

    de Jong, K; Larkin, S K; Styles, L A; Bookchin, R M; Kuypers, F A

    2001-08-01

    Phosphatidylserine (PS), exclusively present in the inner monolayer of the normal red blood cell (RBC) membrane, is exposed in subpopulations of sickle cells. PS-exposing RBCs were found predominantly among the densest and the very light sickle cells. Within the light RBC fraction, PS exposure was found on reticulocytes, transferrin receptor-expressing reticulocytes, and mature RBCs. The last subset contained low-density valinomycin-resistant RBCs, previously shown to have high Na(+) and low K(+) content. This subpopulation contained the highest percentage of PS-exposing cells. The PS-exposing sickle cells did not show the sustained high cytosolic Ca(++) levels that have been shown to activate scramblase activity. Data from this study indicate that PS exposure can occur at different stages in the life of the sickle RBC and that it correlates with the loss of aminophospholipid translocase activity, the only common denominator of the PS-exposing cells. The additional requirement of scramblase activation may occur during transient increases in cytosolic Ca(++). (Blood. 2001;98:860-867)

  11. 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

  12. Matrix regulators in neural stem cells functions

    PubMed Central

    Wade, Anna; McKinney, Andrew; Phillips, Joanna J.

    2014-01-01

    Background Neural stem/progenitor cells (NSPCs) reside within a complex and dynamic extracellular microenvironment, or niche. This niche regulates fundamental aspects of their behavior during normal neural development and repair. Precise yet dynamic regulation of NSPC self-renewal, migration, and differentiation is critical and must persist over the life of an organism. Scope of Review In this review, we summarize some of the major components of the NSPC niche and provide examples of how cues from the extracellular matrix regulate NSPC behaviors. We use proteoglycans to illustrate the many diverse roles of the niche in providing temporal and spatial regulation of cellular behavior. Major Conclusions The NSPC niche is comprised of multiple components that include; soluble ligands, such as growth factors, morphogens, chemokines, and neurotransmitters, the extracellular matrix, and cellular components. As illustrated by proteoglycans, a major component of the extracellular matrix, the NSPC niche provides temporal and spatial regulation of NSPC behaviors. General Significance The factors that control NSPC behavior are vital to understand as we attempt to modulate normal neural development and repair. Furthermore, an improved understanding of how these factors regulate cell proliferation, migration, and differentiation, crucial for malignancy, may reveal novel anti-tumor strategies. PMID:24447567

  13. 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

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

    PubMed

    Bronner, Marianne

    2015-03-01

    Neural crest precursors generate diverse cell lineages during development, which have been proposed to arise either from multipotent precursor cells or pools of heterogeneous, restricted progenitors. Now in Cell Stem Cell, Baggiolini et al. (2015) perform rigorous in vivo lineage tracing to show that individual neural crest precursors are multipotent. PMID:25748927

  15. 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

  16. 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

  17. 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

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

    PubMed

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

    2014-11-01

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

  19. Embryonic stem cell-derived neural stem cells fuse with microglia and mature neurons.

    PubMed

    Cusulin, Carlo; Monni, Emanuela; Ahlenius, Henrik; Wood, James; Brune, Jan Claas; Lindvall, Olle; Kokaia, Zaal

    2012-12-01

    Transplantation of neural stem cells (NSCs) is a novel strategy to restore function in the diseased brain, acting through multiple mechanisms, for example, neuronal replacement, neuroprotection, and modulation of inflammation. Whether transplanted NSCs can operate by fusing with microglial cells or mature neurons is largely unknown. Here, we have studied the interaction of a mouse embryonic stem cell-derived neural stem (NS) cell line with rat and mouse microglia and neurons in vitro and in vivo. We show that NS cells spontaneously fuse with cocultured cortical neurons, and that this process requires the presence of microglia. Our in vitro data indicate that the NS cells can first fuse with microglia and then with neurons. The fused NS/microglial cells express markers and retain genetic and functional characteristics of both parental cell types, being able to respond to microglia-specific stimuli (LPS and IL-4/IL-13) and to differentiate to neurons and astrocytes. The NS cells fuse with microglia, at least partly, through interaction between phosphatidylserine exposed on the surface of NS cells and CD36 receptor on microglia. Transplantation of NS cells into rodent cortex results in fusion with mature pyramidal neurons, which often carry two nuclei, a process probably mediated by microglia. The fusogenic role of microglia could be even more important after NSC transplantation into brains affected by neurodegenerative diseases associated with microglia activation. It remains to be elucidated how the occurrence of the fused cells will influence the functional outcome after NSC transplantation in the diseased brain.

  20. Unleashing Cancer Cells on Surfaces Exposing Motogenic IGDQ Peptides.

    PubMed

    Corvaglia, Valentina; Marega, Riccardo; De Leo, Federica; Michiels, Carine; Bonifazi, Davide

    2016-01-20

    Thiolated peptides bearing the Ile-Gly-Asp (IGD) motif, a highly conserved sequence of fibronectin, are used for the preparation of anisotropic self-assembled monolayers (SAM gradients) to study the whole-population migratory behavior of metastatic breast cancer cells (MDA-MB-231 cells). Ile-Gly-Asp-Gln-(IGDQ)-exposing SAMs sustain the adhesion of MDA-MB-231 cells by triggering focal adhesion kinase phosphorylation, similarly to the analogous Gly-Arg-Gly-Asp-(GRGD)-terminating surfaces. However, the biological responses of different cell lines interfaced with the SAM gradients show that only those exposing the IGDQ sequence induce significant migration of MDA-MB-231 cells. In particular, the observed migratory behavior suggests the presence of cell subpopulations associated with a "stationary" or a "migratory" phenotype, the latter determining a considerable cell migration at the sub-cm length scale. These findings are of great importance as they suggest for the first time an active role of biological surfaces exposing the IGD motif in the multicomponent orchestration of cellular signaling involved in the metastatic progression.

  1. A role for adherons in neural retina cell adhesion

    PubMed Central

    1983-01-01

    Embryonic chick neural retina cells release glycoprotein complexes, termed adherons, into their culture medium. When absorbed onto the surface of petri dishes, neural retina adherons increase the initial rate of neural retina cell adhesion; they also stimulate the rate of cell-cell aggregation. Adheron-stimulated adhesion is tissue specific, and the spontaneous aggregation of neural retina cells is inhibited by monovalent Fab' fragments prepared from an antiserum against neural retina adherons. Therefore cell surface antigenic determinants shared with adherons are involved in normal cell-cell adhesions. The particles from the heterogeneous neural retina population contain many proteins and several glycosaminoglycans. The adherons migrate as a symmetrical 12S peak on sucrose gradients and are predominantly 15-nm spheres when examined by electron microscopy. Finally, the specific activity of neural retina adherons increases from embryonic days 7 through 12 and then declines. These results suggest that glycoprotein particles may be involved in some of the adhesive interactions between neural retina cells and between the cells and their environment. PMID:6187755

  2. 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

  3. Functional repair with neural stem cells.

    PubMed

    Sinden, J D; Stroemer, P; Grigoryan, G; Patel, S; French, S J; Hodges, H

    2000-01-01

    Approval to commence phase I/II clinical trials with neural stem cells requires proof of concept in well-accepted animal models of human neurological disease or injury. We initially showed that the conditionally immortal MHP36 line of hippocampal origin (derived from the H-2Kb-tsA58 transgenic mouse) was effective in repopulating CA1 neurons in models of global ischaemia and repairing cognitive function, and have now shown that this line is multifunctional. MHP36 cells are effective in restoring spatial memory deficits in rats after excitotoxic lesions of the cholinergic projections to cortex and hippocampus and in rats showing cognitive impairments due to normal ageing. Moreover, grafts of MHP36 cells are effective in reversing sensory and motor deficits and reducing lesion volume as a consequence of occlusion of the middle cerebral artery, the major cause of stroke. In contrast, MHP36 cell grafts were unable to repair motor asymmetries in rats with unilateral 6-hydroxydopamine lesions of the nigrostriatal dopamine system, the prototype rodent model of Parkinson's disease. These data show that conditionally immortal neuroepithelial stem cells are multifunctional, being able to repair diverse types of brain damage. However, there are limitations to this multifunctionality, suggesting that lines from different regions of the developing brain will be required to treat different brain diseases. ReNeuron is currently developing human neuroepithelial stem cell lines from different brain regions and with similar reparative properties to our murine lines. PMID:11131543

  4. 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

  5. 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

  6. 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

  7. 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

  8. 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.

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

    PubMed

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

    2015-05-15

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

  11. 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.

  12. 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

  13. Directed Differentiation of Human Embryonic Stem Cells into Neural Progenitors.

    PubMed

    Banda, Erin; Grabel, Laura

    2016-01-01

    A variety of protocols have been used to produce neural progenitors from human embryonic stem cells. We have focused on a monolayer culture approach that generates neural rosettes. To initiate differentiation, cells are plated in a serum-free nutrient-poor medium in the presence of a BMP inhibitor. Depending on the cell line used, additional growth factor inhibitors may be required to promote neural differentiation. Long-term culture and addition of the Notch inhibitor DAPT can promote terminal neuronal differentiation. Extent of differentiation is monitored using immunocytochemistry for cell type-specific markers.

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

    PubMed

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

    2015-02-25

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

  15. Comparative genomic hybridization study of arsenic-exposed and non-arsenic-exposed urinary transitional cell carcinoma

    SciTech Connect

    Hsu, L.-I; Chiu, Allen W.; Pu, Y.-S.; Wang, Y.-H.; Huan, Steven K.; Hsiao, C.-H.; Hsieh, F.-I; Chen, C.-J.

    2008-03-01

    To compare the differences in DNA aberrations between arsenic-exposed and non-arsenic-exposed transitional cell carcinoma (TCC), we analyzed 19 arsenic-exposed and 29 non-arsenic-exposed urinary TCCs from Chi-Mei Hospital using comparative genomic hybridization. DNA aberrations were detected in 42 TCCs including 19 arsenic-exposed and 23 non-arsenic-exposed TCCs. Arsenic-exposed TCCs had more changes than unexposed TCCs (mean {+-} SD, 6.6 {+-} 2.9 vs. 2.9 {+-} 2.2). Arsenic exposure was significantly associated with the number of DNA aberrations after adjustment for tumor stage, tumor grade and cigarette smoking in multiple regression analysis. The most frequent DNA gains, which were strikingly different between arsenic-exposed and non-arsenic-exposed TCCs, included those at 1p, 4p, 4q and 8q. A much higher frequency of DNA losses in arsenic-exposed TCCs compared with non-arsenic-exposed TCCs was observed in 10q, 11p and 17p. Chromosomal loss in 17p13 was associated not only with arsenic exposure, but also with tumor stage and grade. The p53 immunohistochemistry staining showed that chromosome 17p13 loss was associated with either p53 no expression (25%) or p53 overexpression (75%). The findings suggest that long-term arsenic exposure may increase the chromosome abnormality in TCC, and 17p loss plays an important role in arsenic-induced urinary carcinogenesis.

  16. Synaptopathy in the noise-exposed and aging cochlea: Primary neural degeneration in acquired sensorineural hearing loss.

    PubMed

    Kujawa, Sharon G; Liberman, M Charles

    2015-12-01

    The classic view of sensorineural hearing loss (SNHL) is that the "primary" targets are hair cells, and that cochlear-nerve loss is "secondary" to hair cell degeneration. Our recent work in mouse and guinea pig has challenged that view. In noise-induced hearing loss, exposures causing only reversible threshold shifts (and no hair cell loss) nevertheless cause permanent loss of >50% of cochlear-nerve/hair-cell synapses. Similarly, in age-related hearing loss, degeneration of cochlear synapses precedes both hair cell loss and threshold elevation. This primary neural degeneration has remained hidden for three reasons: 1) the spiral ganglion cells, the cochlear neural elements commonly assessed in studies of SNHL, survive for years despite loss of synaptic connection with hair cells, 2) the synaptic terminals of cochlear nerve fibers are unmyelinated and difficult to see in the light microscope, and 3) the degeneration is selective for cochlear-nerve fibers with high thresholds. Although not required for threshold detection in quiet (e.g. threshold audiometry or auditory brainstem response threshold), these high-threshold fibers are critical for hearing in noisy environments. Our research suggests that 1) primary neural degeneration is an important contributor to the perceptual handicap in SNHL, and 2) in cases where the hair cells survive, neurotrophin therapies can elicit neurite outgrowth from spiral ganglion neurons and re-establishment of their peripheral synapses. This article is part of a Special Issue entitled .

  17. Neural progenitor cells regulate microglia functions and activity.

    PubMed

    Mosher, Kira I; Andres, Robert H; Fukuhara, Takeshi; Bieri, Gregor; Hasegawa-Moriyama, Maiko; He, Yingbo; Guzman, Raphael; Wyss-Coray, Tony

    2012-11-01

    We found mouse neural progenitor cells (NPCs) to have a secretory protein profile distinct from other brain cells and to modulate microglial activation, proliferation and phagocytosis. NPC-derived vascular endothelial growth factor was necessary and sufficient to exert at least some of these effects in mice. Thus, neural precursor cells may not only be shaped by microglia, but also regulate microglia functions and activity.

  18. Neural crest cells: a model for invasive behavior.

    PubMed

    Tucker, Richard P

    2004-02-01

    Neural crest cells are the embryonic precursors of the neurons and glia of the peripheral nervous system, pigment cells, and connective tissue in the face, neck and heart. They are induced near the junction of the neural plate and embryonic ectoderm and undergo an epithelial to mesenchymal transition (EMT). Neural crest cells then display invasive behavior and migrate into the surrounding tissues along specific pathways. Neural crest cells are amenable to study in tissue culture, and the molecules that regulate their development can be studied in vivo with antisense techniques as well as with the expression of gain and loss-of-function constructs. Mutations in factors that regulate neural crest cell survival or differentiation can lead to cell death or the premature cessation of their migration, resulting in craniofacial abnormalities, pigmentation defects and the absence of enteric neurons. This paper reviews recent advances in our understanding of neural crest cell induction and migration, emphasizing both avian and amphibian models. Cell facts: The embryonic progenitors of pigment cells, the neurons and glia of the peripheral nervous system, as well as connective tissue in the face, neck and heart. Induced to form at the boundary of the neuroepithelium and embryonic ectoderm. Expression of the transcription factors Snail, Slug and FoxD3 leads to delamination from the neural tube. Invasive motility not unlike that of tumor cells can be studied in vitro. Express proteases, distinctive cell surface receptors and glycoproteins to acquire an invasive phenotype. Mutations of transcription factors expressed by the neural crest or in other factors that inhibit their premature differentiation can lead to survival and migration-associated birth defects.

  19. Chromosome aberrations in ataxia telangiectasia cells exposed to heavy ions

    NASA Astrophysics Data System (ADS)

    Kawata, T.; Cucinotta, F.; George, K.; Wu, H.; Shigematsu, N.; Furusawa, Y.; Uno, T.; Isobe, K.; Ito, H.

    Understanding of biological effects of heavy ions is important to assess healt h risk in space. One of the most important issues may be to take into account individual susceptibility. Ataxia telangiectasia (A-T) cells are known to exhibit abnormal responses to radiations but the mechanism of hyper radiosensitivity of A-T still remains unknown. We report chromosome aberrations in normal human fibroblasts and AT fibroblasts exposed to low- and high-LET radiations. A chemical-induced premature chromosome condensation (PCC) technique combined with chromosome- painting technique was applied to score chromosome aberrations in G2/M-phase cells. Following gamma irradiation, GM02052 cells were approximately 5 times more sensitive to g-rays than AG1522 cells. GM02052 cells had a much higher frequency of deletions and misrejoining than AG1522 cells. When the frequency of complex type aberrations was compared, GM02052 cells showed more than 10 times higher frequency than AG1522 cells. The results will be compared with those obtained from high-LET irradiations.

  20. The efficiency of photovoltaic cells exposed to pulsed laser light

    NASA Technical Reports Server (NTRS)

    Lowe, R. A.; Landis, G. A.; Jenkins, P.

    1993-01-01

    Future space missions may use laser power beaming systems with a free electron laser (FEL) to transmit light to a photovoltaic array receiver. To investigate the efficiency of solar cells with pulsed laser light, several types of GaAs, Si, CuInSe2, and GaSb cells were tested with the simulated pulse format of the induction and radio frequency (RF) FEL. The induction pulse format was simulated with an 800-watt average power copper vapor laser and the RF format with a frequency-doubled mode-locked Nd:YAG laser. Averaged current vs bias voltage measurements for each cell were taken at various optical power levels and the efficiency measured at the maximum power point. Experimental results show that the conversion efficiency for the cells tested is highly dependent on cell minority carrier lifetime, the width and frequency of the pulses, load impedance, and the average incident power. Three main effects were found to decrease the efficiency of solar cells exposed to simulated FEL illumination: cell series resistance, LC 'ringing', and output inductance. Improvements in efficiency were achieved by modifying the frequency response of the cell to match the spectral energy content of the laser pulse with external passive components.

  1. 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

  2. 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.

  3. 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.

  4. 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.

  5. 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

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

    PubMed

    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. Combination Cell Therapy with Mesenchymal Stem Cells and Neural Stem Cells for Brain Stroke in Rats

    PubMed Central

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

    2015-01-01

    Objectives Brain stroke is the second most important events that lead to disability and morbidity these days. Although, stroke is important, there is no treatment for curing this problem. Nowadays, cell therapy has opened a new window for treating central nervous system disease. In some previous studies the Mesenchymal stem cells and neural stem cells. In this study, we have designed an experiment to assess the combination cell therapy (Mesenchymal and Neural stem cells) effects on brain stroke. Method and Materials The Mesenchymal stem cells were isolated from adult rat bone marrow and the neural stem cells were isolated from ganglion eminence of rat embryo 14 days. The Mesenchymal stem cells were injected 1 day after middle cerebral artery occlusion (MCAO) and the neural stem cells transplanted 7 day after MCAO. After 28 days, the neurological outcomes and brain lesion volumes were evaluated. Also, the activity of Caspase 3 was assessed in different groups. Result The group which received combination cell therapy had better neurological examination and less brain lesion. Also the combination cell therapy group had the least Caspase 3 activity among the groups. Conclusions The combination cell therapy is more effective than Mesenchymal stem cell therapy and neural stem cell therapy separately in treating the brain stroke in rats. PMID:26019759

  8. Cloning assay thresholds on cells exposed to ultrafast laser pulses

    NASA Astrophysics Data System (ADS)

    Koenig, Karsten; Riemann, Iris; Fischer, Peter; Becker, Thomas P.; Oehring, Hartmut; Halbhuber, Karl-Juergen

    1999-06-01

    The influence of the peak power, laser wavelength and the pulse duration of near infrared (NIR) ultrashort laser pulses on the reproduction behavior of Chinese hamster ovary (CHO) cells has been studied. In particular we determined the cloning efficiency of single cell pairs after exposure to ultrashort laser pulses with an intensity in the range of GW/cm2 and TW/cm2. A total of more than 3500 non- labeled cells were exposed to a highly focused scanning beam of a multiphoton laser microscope with 60 microsecond pixel dwell time per scan. The beam was provided by a tunable argon ion laser pumped mode-locked 76 MHz Titanium:Sapphire laser as well as by a compact solid-state laser based system (Vitesse) at a fixed wavelength of 800 nm. Pulse duration (tau) was varied in the range of 100 fs to 4 ps by out-of-cavity pulse- stretching units consisting of SF14 prisms and blazed gratings. Within an optical (laser power) window CHO cells could be scanned for hours without severe impact on reproduction behavior, morphology and vitality. Ultrastructural studies reveal that mitochondria are the major targets of intense destructive laser pulses. Above certain laser power P thresholds, CHO cells started to delay or failed to undergo cell division and, in part, to develop uncontrolled cell growth (giant cell formation). The damage followed a P2/(tau) relation which is typical for a two-photon excitation process. Therefore, cell damage was found to be more pronounced at shorter pulses. Due to the same P2/(tau) relation for the efficiency of fluorescence excitation, two- photon microscopy of living cells does not require extremely short femtosecond laser pulses nor pulse compression units. Picosecond as well as femtosecond layers can be used as efficient light sources in safe two photon fluorescence microscopy. Only in three photon fluorescence microscopy, femtosecond laser pulses are advantageous over picosecond pulses.

  9. Cloning assay thresholds on cells exposed to ultrafast laser pulses

    NASA Astrophysics Data System (ADS)

    Koenig, Karsten; Riemann, Iris; Fischer, Peter; Becker, Thomas P.; Oehring, Hartmut; Halbhuber, Karl-Juergen

    1999-06-01

    The influence of the peak power, laser wavelength and the pulse duration of near infrared ultrashort laser pulses on the reproduction behavior of Chinese hamster ovary (CHO) cells has been studied. In particular, we determined the cloning efficiency of single cell pairs after exposure to ultrashort laser pulses with an intensity in the range of GW/cm2 and TW/cm2. A total of more than 3500 non- labeled cells were exposed to a highly focused scanning beam of a multiphoton laser microscope with 60 microsecond(s) pixel dwell time per scan. The beam was provided by a tunable argon ion laser pumped mode-locked 76 MHz Titanium:Sapphire laser as well as by a compact solid-state laser based system (Vitesse) at a fixed wavelength of 800 nm. Pulse duration (tau) was varied in the range of 100 fs to 4 ps by out-of- cavity pulse-stretching units consisting of SF14 prisms and blazed gratings. Within an optical (laser power) window CHO cells could be scanned for hours without severe impact on reproduction behavior, morphology and vitality. Ultrastructural studies reveal that mitochondria are the major targets of intense destructive laser pulses. Above certain laser power P thresholds, CHO cells started to delay or failed to undergo cell division and, in part, to develop uncontrolled cell growth (giant cell formation). The damage followed a P2/(tau) relation which is typical for a two- photon excitation process. Therefore, cell damage was found to be more pronounced at shorter pulses. Due to the same P2/(tau) relation for the efficiency of fluorescence excitation, two-photon microscopy of living cells does not require extremely short femtosecond laser pulses nor pulse compression units. Picosecond as well as femtosecond lasers can be used as efficient light sources in safe two photon fluorescence microscopy. Only in three photon fluorescence microscopy, femtosecond laser pulses are advantageous over picosecond pulses.

  10. 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.

  11. Drosophila neural stem cells in brain development and tumor formation.

    PubMed

    Jiang, Yanrui; Reichert, Heinrich

    2014-01-01

    Neuroblasts, the neural stem cells in Drosophila, generate the complex neural structure of the central nervous system. Significant progress has been made in understanding the mechanisms regulating the self-renewal, proliferation, and differentiation in Drosophila neuroblast lineages. Deregulation of these mechanisms can lead to severe developmental defects and the formation of malignant brain tumors. Here, the authors review the molecular genetics of Drosophila neuroblasts and discuss some recent advances in stem cell and cancer biology using this model system.

  12. Induced neural stem cells (iNSCs) in neurodegenerative diseases.

    PubMed

    Hermann, Andreas; Storch, Alexander

    2013-09-01

    Recent advances in somatic cell reprogramming is one of the most important developments in neuroscience in the last decades since it offers for the first time the opportunity to work with disease/patient-specific neurons or other neural cell types. Induced pluripotent stem cells (iPSCs) can be differentiated into all cell types of the body enabling investigations not only on neurons but also on muscle or endothelial cells which are cell types often also of great interest in neurodegenerative diseases. The novel technology of direct lineage conversion of somatic cells into neurons (induced neurons; iNs) or into expandable multipotent neural stem cells (induced neural stem cells; iNSCs) provides interesting alternatives to the iPSC technology. These techniques have the advantage of easier cell culture, but only neurons (iNs) or neuroectodermal cells (iNSCs) can be generated. Although there are several open questions coming along with these new neural cell types, they hold great promises for both cell replacement and cell modelling of neurodegenerative diseases.

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

    PubMed

    Zhongling Feng; Gang Zhao; Lei Yu

    2009-01-01

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

  14. Neural Crest As the Source of Adult Stem Cells

    PubMed Central

    Pierret, Chris; Spears, Kathleen; Maruniak, Joel A.; Kirk, Mark D.

    2012-01-01

    Recent studies suggest that adult stem cells can cross germ layer boundaries. For example, bone marrow-derived stem cells appear to differentiate into neurons and glial cells, as well as other types of cells. How can stem cells from bone marrow, pancreas, skin, or fat become neurons and glia; in other words, what molecular and cellular events direct mesodermal cells to a neural fate? Transdifferentiation, dediffereniation, and fusion of donor adult stem cells with fully differentiated host cells have been proposed to explain the plasticity of adult stem cells. Here we review the origin of select adult stem cell populations and propose a unifying hypothesis to explain adult stem cell plasticity. In addition, we outline specific experiments to test our hypothesis. We propose that peripheral, tissue-derived, or adult stem cells are all progeny of the neural crest. PMID:16646675

  15. Effects of acute hypoxia/acidosis on intracellular pH in differentiating neural progenitor cells.

    PubMed

    Nordström, Tommy; Jansson, Linda C; Louhivuori, Lauri M; Akerman, Karl E O

    2012-06-21

    The response of differentiating mouse neural progenitor cells, migrating out from neurospheres, to conditions simulating ischemia (hypoxia and extracellular or intracellular acidosis) was studied. We show here, by using BCECF and single cell imaging to monitor intracellular pH (pH(i)), that two main populations can be distinguished by exposing migrating neural progenitor cells to low extracellular pH or by performing an acidifying ammonium prepulse. The cells dominating at the periphery of the neurosphere culture, which were positive for neuron specific markers MAP-2, calbindin and NeuN had lower initial resting pH(i) and could also easily be further acidified by lowering the extracellular pH. Moreover, in this population, a more profound acidification was seen when the cells were acidified using the ammonium prepulse technique. However, when the cell population was exposed to depolarizing potassium concentrations no alterations in pH(i) took place in this population. In contrast, depolarization caused an increase in pH(i) (by 0.5 pH units) in the cell population closer to the neurosphere body, which region was positive for the radial cell marker (GLAST). This cell population, having higher resting pH(i) (pH 6.9-7.1) also responded to acute hypoxia. During hypoxic treatment the resting pH(i) decreased by 0.1 pH units and recovered rapidly after reoxygenation. Our results show that migrating neural progenitor cells are highly sensitive to extracellular acidosis and that irreversible damage becomes evident at pH 6.2. Moreover, our results show that a response to acidosis clearly distinguishes two individual cell populations probably representing neuronal and radial cells.

  16. Synthesis of protein in intestinal cells exposed to cholera toxin

    SciTech Connect

    Peterson, J.W.; Berg, W.D. Jr.; Coppenhaver, D.H.

    1987-11-01

    The mechanism by which cyclic adenosine monophosphate (AMP), formed by intestinal epithelial cells in response to cholera toxin, ultimately results in alterations in water and electrolyte transport is poorly understood. Several studies have indicated that inhibitors of transcription or translation block much of the transport of ions and water in the intestine and edema formation in tissue elicited by cholera toxin. Data presented in this study confirmed the inhibitory effects of cycloheximide on cholera toxin-induced fluid accumulation in the rabbit intestinal loop model. Neither cycloheximide nor actinomycin D altered the amount of cyclic AMP that accumulated in intestinal cells and Chinese hamster ovary cells exposed to cholera toxin. An increase in (/sup 3/H) leucine incorporation was readily demonstrable in intestinal epithelial cells from rabbits challenged with Vibrio cholerae. Similarly, intestinal epithelial cells incubated with cholera toxin for 4 hr synthesized substantially more protein than controls as determined by relative incorporation of (/sup 35/S) methionine. Most of the new protein synthesized in response to cholera toxin was membrane associated and of high molecular weight. The possible significance of the toxin-induced protein relative to cholera pathogenesis was discussed.

  17. Role of polarized cell divisions in zebrafish neural tube formation.

    PubMed

    Clarke, Jon

    2009-04-01

    Development of epithelial cell polarity and morphogenesis of a central lumen are essential prerequisites for the formation of the vertebrate neural tube. In teleost fish embryos this first involves the formation of a solid neural rod structure that then undergoes a process of cavitation to form a lumen. This process is initiated from a neural plate that has a distinct organization compared to other vertebrates, and involves complex cell intercalations and rearrangements. A key element is a mode of polarized cell division that generates daughters with mirror-image apico-basal polarity. These mirror-symmetric divisions have powerful morphogenetic influence because when they occur in ectopic locations they orchestrate the development of ectopic apical and basal specializations and the development of ectopic neural tubes.

  18. 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.

  19. 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

  20. Polyamines and polyamine biosynthesis in cells exposed to hyperthermia

    SciTech Connect

    Gerner, E.W.; Stickney, D.G.; Herman, T.S.; Fuller, D.J.

    1983-02-01

    The issue of how polyamines act to sensitize cultured cells to the lethal effects of hyperthermia was investigated using Chinese hamster cells which were induced to express thermotolerance. Intracellular levels of these naturally occurring polycations were manipulated in certain situations by treating whole cells with methylglyoxal bis-(guanylhydrazone), an inhibitor of the S-adenosyl-L-methionine decarboxylases. Exogenous spermine as low as 100 ..mu..M in the culture media dramatically sensitized cells expressing thermotolerance to the lethal effects of subsequent 42/sup 0/C exposures. When thermotolerance was differentially induced in cultures exposed to 42.4/sup 0/C by varying the rate of heating from 37 to 42.4/sup 0/C, the most resistant cells and the highest levels of intracellular spermidine and spermine. This finding was explainable in part by the observation that the putrescine-dependent S-adenosyl-L-methionine decarboxylase activity was minimally affected in cells expressng the greatest degree of thermotolerance. When this enzyme activity was inhibited by drug, lowered intracellular polyamine levels did not correspond with subsequent survival responses to heat. Interestingly, cultures treated with methylglyoxal bis-(guanylhydrazone) 24 hr previous to heat exposure showed a reduced capacity to express rate of heating-induced thermotolerance. Together, these results demonstrate that the polyamines, especially spermidine and spermine, enhance hyperthermia-induced cell killing by some mechanism involving the plasma membrane. Further, our data suggest that methylglyoxal bis-(guanylhydrazone) can act to affect thermal responses by a mechanism(s) other than modification of intracellular polyamine levels.

  1. 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

  2. Development of neural crest cells expressing nerve growth factor receptors

    SciTech Connect

    Greiner, C.A.

    1987-01-01

    The present study examines the ontogeny of the nerve growth factor receptor of neural crest cells in vitro and the phenotypic nature of the neural crest cells expressing this receptor. /sup 125/I-NGF binding assays and autoradiographic and immunofluorescence techniques have demonstrated the presence of a subpopulation of quail neural crest cells that express specific NGF receptors after 3-4 days in vitro. This subpopulations represents approximately 28% of the cells in 5-day primary cultures and 30-35% of the cells in secondary cultures; these cells generally exhibited a flattened, phase-dark morphology. Approximately one-third of these cells also labeled with a 2 hr pulse of /sup 3/H thymidine. Catecholamine-containing neural crest cells generally lacked NGF receptors. NGF receptor-positive cells also failed to demonstrate somatostatin-, neuron-specific enolase-, or S-100-like immunoreactivity. Melanocytes do not appear to express NGF receptors. Exogenous nerve growth factor did not influence the morphology or mitotic status of the cells in culture.

  3. 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.

  4. 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

  5. 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

  6. 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

  7. 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.

  8. Functional activity of mitochondria in cultured neural precursor cells.

    PubMed

    Plotnikov, E Yu; Marei, M V; Podgornyi, O V; Aleksandrova, M A; Zorov, D B; Sukhikh, G T

    2006-01-01

    We studied mitochondrial transmembrane potential of neural precursor cells forming neurospheres in culture. Uneven energization of mitochondria in neurosphere cells was detected. Heterogeneity of cells by the mitochondrial potential increased with neurosphere enlargement during culturing. Decrease in the mitochondrial potential in the central cells in large spheres, presumably caused by insufficient diffusion of oxygen and nutrients, can provoke their damage and death. Population of cells with high mitochondrial potential responded to addition of the nuclear dye by a decrease in mitochondrial potential, which can indicate functioning of ABCG2 complex in these cells, characteristic of undifferentiated stem cells. These data will help to create optimum conditions for culturing of neural stem cells for the maintenance of their maximum functional and proliferative activity. PMID:16929986

  9. 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

  10. 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

  11. Polymerization of the conducting polymer poly(3,4-ethylenedioxythiophene) (PEDOT) around living neural cells

    PubMed Central

    Richardson-Burns, Sarah M.; Hendricks, Jeffrey L.; Foster, Brian; Povlich, Laura K.; Kim, Dong-Hwan; Martin, David C.

    2007-01-01

    In this paper we describe interactions between neural cells and the conducting polymer poly(3,4-ethylenedioxythiophene (PEDOT) toward development of electrically conductive biomaterials intended for direct, functional contact with electrically-active tissues such as the nervous system, heart, and skeletal muscle. We introduce a process for polymerizing PEDOT around living cells and describe a neural cell-templated conducting polymer coating for microelectrodes and a hybrid conducting polymer-live neural cell electrode. We found that neural cells could be exposed to working concentrations (0.01 M) of the EDOT monomer for as long as 72 hours while maintaining 80% cell viability. PEDOT could be electrochemically deposited around neurons cultured on electrodes using 0.5-1 μA/mm2 galvanostatic current. PEDOT polymerized on the electrode and surrounded the cells, covering cell processes. The polymerization was impeded in regions where cells were well-adhered to the substrate. The cells could be removed from the PEDOT matrix to generate a neural cell-templated biomimetic conductive substrate with cell-shaped features that were cell-attracting. Live cells embedded within the conductive polymer matrix remained viable for at least 120 hours following polymerization. Dying cells primarily underwent apoptotic cell death. PEDOT, PEDOT+live neurons, and neuron-templated PEDOT coatings on electrodes significantly enhanced the electrical properties as compared to the bare electrode as indicated by decreased electrical impedance of 1-1.5 orders of magnitude at 0.01-1 kHz and significantly increased charge transfer capacity. PEDOT coatings showed a decrease of the phase angle of the impedance from roughly 80 degrees for the bare electrode to 5-35 degrees at frequencies >0.1 kHz. Equivalent circuit modeling indicated that PEDOT-coated electrodes were best described by R(C(RT)) circuit. We found that an RC parallel circuit must be added to the model for PEDOT+live neuron and neuron

  12. 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

  13. Cellular and molecular effects of high-LET radiation on human neural stem cells and neurons

    NASA Astrophysics Data System (ADS)

    Vazquez, M.; Guida, P.; Green, L.; Chang, P.; Otto, S.

    Because successful operations in space depend in part on the performance capabilities of astronauts, radiation-induced neurological damage could jeopardize the successful completion of mission requirements, as well as have long-term consequences on the health of astronauts. As such, understanding the nature of this risk may be vital to the effective performance of astronauts during future missions in space. This paper describes the neural cell responses to conventional and charged particles radiation in cell culture systems. One of the goals is to characterize radiation-induced neural cell damage pathways; especially those related to apoptosis induction and its modification by pharmacological manipulation. Our laboratory utilizes the method of flow cytometry to measure the induction of apoptosis and necrosis in cells. Neural stem cells (NT2) were exposed to the different ions; we measured a dose-dependent induction of apoptosis. NT2 cells were exposed to graded doses of 1 and 5 GeV/n Fe, 0.29 GeV/n C, 1 GeV/n Ti, and 0.6 GeV/n Si ions and samples were taken at 48 hours after exposure. The percentage of apoptotic cells in culture was measured by FITC-Annexin V by flow cytometry. Similar data obtained from NT2 cells exposed to 255 MeV/n protons and 137Cs are included for comparison. Preliminary RBE calculations demonstrated that iron ions are more effective in inducing apoptosis. Exposure of cells to ionizing radiation produces changes in the expression of many genes as cells react to this insult. At present, the identities of the molecular changes that occur in response to HZE radiation remain largely unknown. In an effort to reveal this information, we screened an array (Superarray) of p53-related genes with RNA purified from NT2 cells mock irradiated or exposed to 50 cGy of 1 GeV/n iron ions. Preliminary results indicated that the expression of numerous critical genes was altered 3 hours after HZE radiation exposure. By performing Western blot analysis on NT2

  14. Abeta40 promotes neuronal cell fate in neural progenitor cells.

    PubMed

    Chen, Y; Dong, C

    2009-03-01

    Sequential cleavage of the amyloid precursor protein (APP) by beta- and then gamma- secretase gives rise to Abeta(1-40) (Abeta40), a major species of Abeta (beta-amyloid) produced by neurons under physiological conditions. Abeta(1-42) (Abeta42), a minor species of Abeta, is also produced by a similar but less understood mechanism of the gamma-secretase. The physiological functions of these Abeta species remain to be defined. In this report, we demonstrate that freshly prepared soluble Abeta40 significantly promotes neurogenesis in primary neural progenitor cells (NPCs). First, Abeta40 increases neuronal markers as determined by NeuN expression and Tuj1 promoter activity, differing from Abeta42, which induces astrocyte markers in NPCs. Second, Abeta40 induces neuronal differentiation at the end of S-phase in the cell cycle. Third, Abeta40 promotes NPC entry into S-phase, playing a role in NPC self-renewal. Interestingly, Abeta40 does not significantly increase apoptotic indexes such as DNA condensation and DNA fragmentation. In addition, Abeta40 does not augment caspase-3 activation in NeuN(+) or nestin(+) cells. Collectively, this report provides strong evidence that Abeta40 is a neurogenic factor and suggests that the debilitated function of Abeta40 in neurogenesis may account for the shortage of neurons in Alzheimer's disease.

  15. Morphine Inhibited the Rat Neural Stem Cell Proliferation Rate by Increasing Neuro Steroid Genesis.

    PubMed

    Feizy, Navid; Nourazarian, Alireza; Rahbarghazi, Reza; Nozad Charoudeh, Hojjatollah; Abdyazdani, Nima; Montazersaheb, Soheila; Narimani, Mohamadreza

    2016-06-01

    Up to present, a large number of reports unveiled exacerbating effects of both long- and short-term administration of morphine, as a potent analgesic agent, on opium-addicted individuals and a plethora of cell kinetics, although contradictory effect of morphine on different cells have been introduced until yet. To address the potent modulatory effect of morphine on neural multipotent precursors with emphasis on endogenous sex-related neurosteroids biosynthesis, we primed the rat neural stem cells isolated from embryonic rat telencephalon to various concentrations of morphine including 10, 20, 50 and 100 µM alone or in combination with naloxone (100 µM) over period of 72 h. Flow cytometric Ki-67 expression and Annexin-V/PI based necrosis and apoptosis of exposed cells were evaluated. The total content of dihydrotestosterone and estradiol in cell supernatant was measured by ELISA. According on obtained data, both concentration- and time-dependent decrement of cell viability were orchestrated thorough down-regulation of ki-67 and simultaneous up-regulation of Annexin-V. On the other hand, the addition of naloxone (100 µM), as Mu opiate receptor antagonist, could blunt the morphine-induced adverse effects. It also well established that time-course exposure of rat neural stem cells with morphine potently could accelerate the endogenous dihydrotestosterone and estradiol biosynthesis. Interestingly, naloxone could consequently attenuate the enhanced neurosteroidogenesis time-dependently. It seems that our results discover a biochemical linkage between an accelerated synthesis of sex-related steroids and rat neural stem cells viability. PMID:26830291

  16. Neural cell apoptosis induced by microwave exposure through mitochondria-dependent caspase-3 pathway.

    PubMed

    Zuo, Hongyan; Lin, Tao; Wang, Dewen; Peng, Ruiyun; Wang, Shuiming; Gao, Yabing; Xu, Xinping; Li, Yang; Wang, Shaoxia; Zhao, Li; Wang, Lifeng; Zhou, Hongmei

    2014-01-01

    To determine whether microwave (MW) radiation induces neural cell apoptosis, differentiated PC12 cells and Wistar rats were exposed to 2.856 GHz for 5 min and 15 min, respectively, at an average power density of 30  mW/cm². JC-1 and TUNEL staining detected significant apoptotic events, such as the loss of mitochondria membrane potential and DNA fragmentation, respectively. Transmission electron microscopy and Hoechst staining were used to observe chromatin ultrastructure and apoptotic body formation. Annexin V-FITC/PI double staining was used to quantify the level of apoptosis. The expressions of Bax, Bcl-2, cytochrome c, cleaved caspase-3 and PARP were examined by immunoblotting or immunocytochemistry. Caspase-3 activity was measured using an enzyme-linked immunosorbent assay. The results showed chromatin condensation and apoptotic body formation in neural cells 6h after microwave exposure. Moreover, the mitochondria membrane potential decreased, DNA fragmentation increased, leading to an increase in the apoptotic cell percentage. Furthermore, the ratio of Bax/Bcl-2, expression of cytochrome c, cleaved caspase-3 and PARP all increased. In conclusion, microwave radiation induced neural cell apoptosis via the classical mitochondria-dependent caspase-3 pathway. This study may provide the experimental basis for further investigation of the mechanism of the neurological effects induced by microwave radiation.

  17. Sonic hedgehog rescues cranial neural crest from cell death induced by ethanol exposure.

    PubMed

    Ahlgren, Sara C; Thakur, Vijaya; Bronner-Fraser, Marianne

    2002-08-01

    Alcohol is a teratogen that induces a variety of abnormalities including brain and facial defects [Jones, K. & Smith, D. (1973) Lancet 2, 999-1001], with the exact nature of the deficit depending on the time and magnitude of the dose of ethanol to which developing fetuses are exposed. In addition to abnormal facial structures, ethanol-treated embryos exhibit a highly characteristic pattern of cell death. Dying cells are observed in the premigratory and migratory neural crest cells that normally populate most facial structures. The observation that blocking Sonic hedgehog (Shh) signaling results in similar craniofacial abnormalities prompted us to examine whether there was a link between this aspect of fetal alcohol syndrome and loss of Shh. We demonstrate that administration of ethanol to chick embryos results in a dramatic loss of Shh, as well as a loss of transcripts involved in Shh signaling pathways. In contrast, other signaling molecules examined do not demonstrate such dramatic changes. Furthermore, we demonstrate that both the ethanol-induced cranial neural crest cell death and the associated craniofacial growth defect can be rescued by application of Shh. These data suggest that craniofacial anomalies resulting from fetal alcohol exposure are caused at least partially by loss of Shh and subsequent neural crest cell death.

  18. Neural Cell Apoptosis Induced by Microwave Exposure Through Mitochondria-dependent Caspase-3 Pathway

    PubMed Central

    Zuo, Hongyan; Lin, Tao; Wang, Dewen; Peng, Ruiyun; Wang, Shuiming; Gao, Yabing; Xu, Xinping; Li, Yang; Wang, Shaoxia; Zhao, Li; Wang, Lifeng; Zhou, Hongmei

    2014-01-01

    To determine whether microwave (MW) radiation induces neural cell apoptosis, differentiated PC12 cells and Wistar rats were exposed to 2.856GHz for 5min and 15min, respectively, at an average power density of 30 mW/cm2. JC-1 and TUNEL staining detected significant apoptotic events, such as the loss of mitochondria membrane potential and DNA fragmentation, respectively. Transmission electron microscopy and Hoechst staining were used to observe chromatin ultrastructure and apoptotic body formation. Annexin V-FITC/PI double staining was used to quantify the level of apoptosis. The expressions of Bax, Bcl-2, cytochrome c, cleaved caspase-3 and PARP were examined by immunoblotting or immunocytochemistry. Caspase-3 activity was measured using an enzyme-linked immunosorbent assay. The results showed chromatin condensation and apoptotic body formation in neural cells 6h after microwave exposure. Moreover, the mitochondria membrane potential decreased, DNA fragmentation increased, leading to an increase in the apoptotic cell percentage. Furthermore, the ratio of Bax/Bcl-2, expression of cytochrome c, cleaved caspase-3 and PARP all increased. In conclusion, microwave radiation induced neural cell apoptosis via the classical mitochondria-dependent caspase-3 pathway. This study may provide the experimental basis for further investigation of the mechanism of the neurological effects induced by microwave radiation. PMID:24688304

  19. Ghrelin regulates cell cycle-related gene expression in cultured hippocampal neural stem cells.

    PubMed

    Chung, Hyunju; Park, Seungjoon

    2016-08-01

    We have previously demonstrated that ghrelin stimulates the cellular proliferation of cultured adult rat hippocampal neural stem cells (NSCs). However, little is known about the molecular mechanisms by which ghrelin regulates cell cycle progression. The purpose of this study was to investigate the potential effects of ghrelin on cell cycle regulatory molecules in cultured hippocampal NSCs. Ghrelin treatment increased proliferation assessed by CCK-8 proliferation assay. The expression levels of proliferating cell nuclear antigen and cell division control 2, well-known cell-proliferating markers, were also increased by ghrelin. Fluorescence-activated cell sorting analysis revealed that ghrelin promoted progression of cell cycle from G0/G1 to S phase, whereas this progression was attenuated by the pretreatment with specific inhibitors of MEK/extracellular signal-regulated kinase 1/2, phosphoinositide 3-kinase/Akt, mammalian target of rapamycin, and janus kinase 2/signal transducer and activator of transcription 3. Ghrelin-induced proliferative effect was associated with increased expression of E2F1 transcription factor in the nucleus, as determined by Western blotting and immunofluorescence. We also found that ghrelin caused an increase in protein levels of positive regulators of cell cycle, such as cyclin A and cyclin-dependent kinase (CDK) 2. Moreover, p27(KIP1) and p57(KIP2) protein levels were reduced when cell were exposed to ghrelin, suggesting downregulation of CDK inhibitors may contribute to proliferative effect of ghrelin. Our data suggest that ghrelin targets both cell cycle positive and negative regulators to stimulate proliferation of cultured hippocampal NSCs. PMID:27325242

  20. The noise exposed factory workers: the prevalence of sensori-neural hearing loss and their use of personal hearing protection devices.

    PubMed

    Maisarah, S Z; Said, H

    1993-09-01

    A total of 524 industrial workers were studied. They consisted of 442 noise exposed and 82 non-noise exposed workers. The purpose was to compare the prevalence of sensori-neural hearing loss among the noise exposed and the non-noise exposed workers, to study their knowledge on the hazard of noise to hearing and the workers' attitude towards the hearing protection devices. The prevalence of sensori-neural hearing loss was significantly higher among the noise exposed workers, i.e., 83% versus 31.7% (p < 0.01). However, the prevalence of hearing impairment was much lower for both groups, being 30.1% for the noise exposed and 3.7% for the non-noise exposed group. Although hearing protection devices were provided to 80.5% of the workers, only 5.1% were wearing them regularly. The possibility of developing hearing loss due to exposure to excessive noise was only known by 35.5% of the noise exposed workers. This awareness was found to have a positive correlation with the workers' compliance to the hearing protection devices. Our findings highlight the need for workers to be educated on the hazards of excessive noise exposure to hearing.

  1. 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

  2. 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.

  3. 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

  4. Derivation of Neural Stem Cells from Mouse Induced Pluripotent Stem Cells.

    PubMed

    Karanfil, Işıl; Bagci-Onder, Tugba

    2016-01-01

    Neural stem cells (NSCs) derived from induced pluripotent stem cells offer therapeutic tools for neurodegenerative diseases. This review focuses on embryoid body (EB)-mediated stem cell culture techniques used to derive NSCs from mouse induced pluripotent stem cells (iPSCs). Generation of healthy and stable NSCs from iPSCs heavily depends on standardized in vitro cell culture systems that mimic the embryonic environments utilized during neural development. Specifically, the neural induction and expansion methods after EB formation are described in this review.

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

    PubMed Central

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

    2016-01-01

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

  6. Human dental mesenchymal stem cells and neural regeneration.

    PubMed

    Xiao, Li; Tsutsui, Takeki

    2013-09-01

    Nerve tissue presents inherent difficulties for its effective regeneration. Stem cell transplantation is considered an auspicious treatment for neuronal injuries. Recently, human dental mesenchymal stem cells (DMSCs) have received extensive attention in the field of regenerative medicine due to their accessibility and multipotency. Since their origin is within the neural crest, they can be differentiated into neural crest-derived cells including neuron and glia cells both in vitro and in vivo. DMSCs are also able to secrete a wide variety of neurotrophins and chemokines, which promote neuronal cells to survival and differentiation. Experimental evidence has shown that human DMSCs engraftment recovered neuronal tissue damage in animal models of central nervous system injuries. Human DMSCs can be a new hope for treatment of nervous system diseases and deficits such as spinal cord injury, stroke and Parkinson's disease.

  7. Receptor regulation of osmolyte homeostasis in neural cells.

    PubMed

    Fisher, Stephen K; Heacock, Anne M; Keep, Richard F; Foster, Daniel J

    2010-09-15

    The capacity of cells to correct their volume in response to hyposmotic stress via the efflux of inorganic and organic osmolytes is well documented. However, the ability of cell-surface receptors, in particular G-protein-coupled receptors (GPCRs), to regulate this homeostatic mechanism has received much less attention. Mechanisms that underlie the regulation of cell volume are of particular importance to cells in the central nervous system because of the physical restrictions of the skull and the adverse impact that even small increases in cell volume can have on their function. Increases in brain volume are seen in hyponatraemia, which can arise from a variety of aetiologies and is the most frequently diagnosed electrolyte disorder in clinical practice. In this review we summarize recent evidence that the activation of GPCRs facilitates the volume-dependent efflux of osmolytes from neural cells and permits them to more efficiently respond to small, physiologically relevant, reductions in osmolarity. The characteristics of receptor-regulated osmolyte efflux, the signalling pathways involved and the physiological significance of receptor activation are discussed. In addition, we propose that GPCRs may also regulate the re-uptake of osmolytes into neural cells, but that the influx of organic and inorganic osmolytes is differentially regulated. The ability of neural cells to closely regulate osmolyte homeostasis through receptor-mediated alterations in both efflux and influx mechanisms may explain, in part at least, why the brain selectively retains its complement of inorganic osmolytes during chronic hyponatraemia, whereas its organic osmolytes are depleted. PMID:20498228

  8. p73 regulates maintenance of neural stem cell

    SciTech Connect

    Agostini, Massimiliano; Tucci, Paola; Bano, Daniele; Nicotera, Pierluigi; McKeon, Frank; Melino, Gerry

    2010-12-03

    Research highlights: {yields} TAp73 is expressed in neural stem cells and its expression increases following their differentiation. {yields} Neural stem cells from p73 null mice have a reduced proliferative potential. {yields} p73-deficient neural stem cells show reduced expression of members of the Sox-2 and Notch gene families. {yields} Neurogenic areas are reduced in the brains of embryonic and adult p73-/- mice. -- Abstract: p73, a member of the p53 family, is a transcription factor that plays a key role in many biological processes. In the present study, we show that TAp73 is expressed in neural stem cells (NSC) and its expression increases following their differentiation. NSC from p73 null mice have a reduced proliferative potential, together with reduced expression of members of the Sox-2 and Notch gene families known to be important for NSC proliferation. In parallel with this in vitro data, the width of the neurogenic areas was reduced in the brains of embryonic and adult p73-/- mice. These data suggest that p73, and in particular TAp73, is important for maintenance of the NSC pool.

  9. 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

  10. Effect of Monocular Deprivation on Rabbit Neural Retinal Cell Densities

    PubMed Central

    Mwachaka, Philip Maseghe; Saidi, Hassan; Odula, Paul Ochieng; Mandela, Pamela Idenya

    2015-01-01

    Purpose: To describe the effect of monocular deprivation on densities of neural retinal cells in rabbits. Methods: Thirty rabbits, comprised of 18 subject and 12 control animals, were included and monocular deprivation was achieved through unilateral lid suturing in all subject animals. The rabbits were observed for three weeks. At the end of each week, 6 experimental and 3 control animals were euthanized, their retinas was harvested and processed for light microscopy. Photomicrographs of the retina were taken and imported into FIJI software for analysis. Results: Neural retinal cell densities of deprived eyes were reduced along with increasing period of deprivation. The percentage of reductions were 60.9% (P < 0.001), 41.6% (P = 0.003), and 18.9% (P = 0.326) for ganglion, inner nuclear, and outer nuclear cells, respectively. In non-deprived eyes, cell densities in contrast were increased by 116% (P < 0.001), 52% (P < 0.001) and 59.6% (P < 0.001) in ganglion, inner nuclear, and outer nuclear cells, respectively. Conclusion: In this rabbit model, monocular deprivation resulted in activity-dependent changes in cell densities of the neural retina in favour of the non-deprived eye along with reduced cell densities in the deprived eye. PMID:26425316

  11. 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.

  12. Developing neural stem cell-based treatments for neurodegenerative diseases.

    PubMed

    Byrne, James A

    2014-05-30

    Owing to the aging of the population, our society now faces an impending wave of age-related neurodegenerative pathologies, the most significant of which is Alzheimer's disease. Currently, no effective therapies for Alzheimer's disease have been developed. However, recent advances in the fields of neural stem cells and human induced pluripotent stem cells now provide us with the first real hope for a cure. The recent discovery by Blurton-Jones and colleagues that neural stem cells can effectively deliver disease-modifying therapeutic proteins throughout the brains of our best rodent models of Alzheimer's disease, combined with recent advances in human nuclear reprogramming, stem cell research, and highly customized genetic engineering, may represent a potentially revolutionary personalized cellular therapeutic approach capable of effectively curing, ameliorating, and/or slowing the progression of Alzheimer's disease.

  13. Microparticle Shedding from Neural Progenitor Cells and Vascular Compartment Cells Is Increased in Ischemic Stroke

    PubMed Central

    Chiva-Blanch, Gemma; Suades, Rosa; Crespo, Javier; Peña, Esther; Padró, Teresa; Jiménez-Xarrié, Elena; Martí-Fàbregas, Joan; Badimon, Lina

    2016-01-01

    Purpose Ischemic stroke has shown to induce platelet and endothelial microparticle shedding, but whether stroke induces microparticle shedding from additional blood and vascular compartment cells is unclear. Neural precursor cells have been shown to replace dying neurons at sites of brain injury; however, if neural precursor cell activation is associated to microparticle shedding, and whether this activation is maintained at long term and associates to stroke type and severity remains unknown. We analyzed neural precursor cells and blood and vascular compartment cells microparticle shedding after an acute ischemic stroke. Methods Forty-four patients were included in the study within the first 48h after the onset of stroke. The cerebral lesion size was evaluated at 3–7 days of the stroke. Circulating microparticles from neural precursor cells and blood and vascular compartment cells (platelets, endothelial cells, erythrocytes, leukocytes, lymphocytes, monocytes and smooth muscle cells) were analyzed by flow cytometry at the onset of stroke and at 7 and 90 days. Forty-four age-matched high cardiovascular risk subjects without documented vascular disease were used as controls. Results Compared to high cardiovascular risk controls, patients showed higher number of neural precursor cell- and all blood and vascular compartment cell-derived microparticles at the onset of stroke, and after 7 and 90 days. At 90 days, neural precursor cell-derived microparticles decreased and smooth muscle cell-derived microparticles increased compared to levels at the onset of stroke, but only in those patients with the highest stroke-induced cerebral lesions. Conclusions Stroke increases blood and vascular compartment cell and neural precursor cell microparticle shedding, an effect that is chronically maintained up to 90 days after the ischemic event. These results show that stroke induces a generalized blood and vascular cell activation and the initiation of neuronal cell repair process

  14. Alternative generation of CNS neural stem cells and PNS derivatives from neural crest-derived peripheral stem cells.

    PubMed

    Weber, Marlen; Apostolova, Galina; Widera, Darius; Mittelbronn, Michel; Dechant, Georg; Kaltschmidt, Barbara; Rohrer, Hermann

    2015-02-01

    Neural crest-derived stem cells (NCSCs) from the embryonic peripheral nervous system (PNS) can be reprogrammed in neurosphere (NS) culture to rNCSCs that produce central nervous system (CNS) progeny, including myelinating oligodendrocytes. Using global gene expression analysis we now demonstrate that rNCSCs completely lose their previous PNS characteristics and acquire the identity of neural stem cells derived from embryonic spinal cord. Reprogramming proceeds rapidly and results in a homogenous population of Olig2-, Sox3-, and Lex-positive CNS stem cells. Low-level expression of pluripotency inducing genes Oct4, Nanog, and Klf4 argues against a transient pluripotent state during reprogramming. The acquisition of CNS properties is prevented in the presence of BMP4 (BMP NCSCs) as shown by marker gene expression and the potential to produce PNS neurons and glia. In addition, genes characteristic for mesenchymal and perivascular progenitors are expressed, which suggests that BMP NCSCs are directed toward a pericyte progenitor/mesenchymal stem cell (MSC) fate. Adult NCSCs from mouse palate, an easily accessible source of adult NCSCs, display strikingly similar properties. They do not generate cells with CNS characteristics but lose the neural crest markers Sox10 and p75 and produce MSC-like cells. These findings show that embryonic NCSCs acquire a full CNS identity in NS culture. In contrast, MSC-like cells are generated from BMP NCSCs and pNCSCs, which reveals that postmigratory NCSCs are a source for MSC-like cells up to the adult stage.

  15. Derivation of Neural Precursor Cells from Human Embryonic Stem Cells for DNA Methylomic Analysis.

    PubMed

    Roubal, Ivan; Park, Sun Joo; Kim, Yong

    2016-01-01

    Embryonic stem cells are self-renewing pluripotent cells with competency to differentiate into all three-germ lineages. Many studies have demonstrated the importance of genetic and epigenetic molecular mechanisms in the maintenance of self-renewal and pluripotency. Stem cells are under unique molecular and cellular regulations different from somatic cells. Proper regulation should be ensured to maintain their unique self-renewal and undifferentiated characteristics. Understanding key mechanisms in stem cell biology will be important for the successful application of stem cells for regenerative therapeutic medicine. More importantly practical use of stem cells will require our knowledge on how to properly direct and differentiate stem cells into the necessary type of cells. Embryonic stem cells and adult stem cells have been used as study models to unveil molecular and cellular mechanisms in various signaling pathways. They are especially beneficial to developmental studies where in vivo molecular/cellular study models are not available. We have derived neural stem cells from human embryonic stem cells as a model to study the effect of teratogen in neural development. We have tested commercial neural differentiation system and successfully derived neural precursor cells exhibiting key molecular features of neural stem cells, which will be useful for experimental application.

  16. [Human pluripotent stem cell and neural differentiation].

    PubMed

    Wataya, Takafumi; Muguruma, Keiko; Sasai, Yoshiki

    2008-10-01

    Recovery of lost brain function is an important issue in medical studies because neurons of the central nervous system (CNS) have poor potential for regeneration. Since few CNS diseases can be treated completely by medicines, regenerative therapy by using stem cells should be studied as a new type of therapeutic intervention. The efficacy of cell replacement therapy in Parkinson's disease has been well investigated. Several studies on fetal tissue transplantation have revealed that quantity and purity of transplanted cells are necessary for recovery of symptoms. SFEB (Serum-free floating culture of embryoid body-like aggregates) method is capable of inducing multi-potential CNS progenitors that can be steered to differentiate into region-specific tissues. On the basis of the existing knowledge of embryology, we have succeeded in the generating of various types of neurons such as telencephalic, cerebeller (Purkinje and granule cells), retinal (photoreceptor cells) and hypothalamic neurons. Application of this culture method to human ES (hES) cells is necessary for clinical purpose: however, poor survival of hES cells in SFEB culture might limit the possibility of using these cells for future medical applications. We found that a selective Rho-associated kinase (ROCK) inhibitor, Y-27632, markedly diminished the dissociation-induced apoptosis of hES cells and enabled the cells to form aggregates in SFEB culture. For both mouse and human ES cells, SFEB culture is a favorable method that can generate large amounts of region-specific neurons. However, stem cell-based therapy continues to face several obstacles. It is important that researchers in the basic sciences and clinical medicine should discuss these problems together to overcome both scientific and ethical issues related to stem cells.

  17. Role of ubiquitin ligases in neural stem and progenitor cells.

    PubMed

    Naujokat, Cord

    2009-01-01

    Ubiquitin ligases are central components of the ubiquitin-proteasome system (UPS), the major machinery for regulated proteolysis in eukaryotic cells. Proteins essential for regulating development, differentiation, proliferation, cell cycling, apoptosis, gene transcription, and signal transduction undergo posttranslational processing via selection by ubiquitin ligases and subsequent controlled proteolysis by the 26S proteasome, the proteolytic unit of the UPS. Neural stem cells (NSCs) are self-renewing multipotent cells of the embryonic and adult mammalian central nervous system. In the last few years, NSCs have generated considerable interest because of their potential to repair neurological damage in preclinical models of stroke, spinal cord injury, and neurodegenerative disease. Recent evidence reveals a central role of ubiquitin ligases in controlling the development, survival, differentiation, and programming of neural stem and progenitor cells. Here the current knowledge of the role and function of ubiquitin ligases in neural stem and progenitor cells is reviewed and insight into an important mechanism of NSC homeostasis by regulated proteolysis is provided. PMID:19479207

  18. Impaired NK Cell Activation and Chemotaxis toward Dendritic Cells Exposed to Complement-Opsonized HIV-1

    PubMed Central

    Ellegård, Rada; Crisci, Elisa; Andersson, Jonas; Shankar, Esaki M.; Nyström, Sofia; Hinkula, Jorma

    2015-01-01

    Mucosa resident dendritic cells (DCs) may represent one of the first immune cells that HIV-1 encounters during sexual transmission. The virions in body fluids can be opsonized with complement factors because of HIV-mediated triggering of the complement cascade, and this appears to influence numerous aspects of the immune defense targeting the virus. One key attribute of host defense is the ability to attract immune cells to the site of infection. In this study, we investigated whether the opsonization of HIV with complement (C-HIV) or a mixture of complement and Abs (CI-HIV) affected the cytokine and chemokine responses generated by DCs, as well as their ability to attract other immune cells. We found that the expression levels of CXCL8, CXCL10, CCL3, and CCL17 were lowered after exposure to either C-HIV or CI-HIV relative to free HIV (F-HIV). DCs exposed to F-HIV induced higher cell migration, consisting mainly of NK cells, compared with opsonized virus, and the chemotaxis of NK cells was dependent on CCL3 and CXCL10. NK cell exposure to supernatants derived from HIV-exposed DCs showed that F-HIV induced phenotypic activation (e.g., increased levels of TIM3, CD69, and CD25) and effector function (e.g., production of IFNγ and killing of target cells) in NK cells, whereas C-HIV and CI-HIV did not. The impairment of NK cell recruitment by DCs exposed to complement-opsonized HIV and the lack of NK activation may contribute to the failure of innate immune responses to control HIV at the site of initial mucosa infection. PMID:26157174

  19. Impaired NK Cell Activation and Chemotaxis toward Dendritic Cells Exposed to Complement-Opsonized HIV-1.

    PubMed

    Ellegård, Rada; Crisci, Elisa; Andersson, Jonas; Shankar, Esaki M; Nyström, Sofia; Hinkula, Jorma; Larsson, Marie

    2015-08-15

    Mucosa resident dendritic cells (DCs) may represent one of the first immune cells that HIV-1 encounters during sexual transmission. The virions in body fluids can be opsonized with complement factors because of HIV-mediated triggering of the complement cascade, and this appears to influence numerous aspects of the immune defense targeting the virus. One key attribute of host defense is the ability to attract immune cells to the site of infection. In this study, we investigated whether the opsonization of HIV with complement (C-HIV) or a mixture of complement and Abs (CI-HIV) affected the cytokine and chemokine responses generated by DCs, as well as their ability to attract other immune cells. We found that the expression levels of CXCL8, CXCL10, CCL3, and CCL17 were lowered after exposure to either C-HIV or CI-HIV relative to free HIV (F-HIV). DCs exposed to F-HIV induced higher cell migration, consisting mainly of NK cells, compared with opsonized virus, and the chemotaxis of NK cells was dependent on CCL3 and CXCL10. NK cell exposure to supernatants derived from HIV-exposed DCs showed that F-HIV induced phenotypic activation (e.g., increased levels of TIM3, CD69, and CD25) and effector function (e.g., production of IFNγ and killing of target cells) in NK cells, whereas C-HIV and CI-HIV did not. The impairment of NK cell recruitment by DCs exposed to complement-opsonized HIV and the lack of NK activation may contribute to the failure of innate immune responses to control HIV at the site of initial mucosa infection. PMID:26157174

  20. In vitro neural differentiation of CD34 (+) stem cell populations in hair follicles by three different neural induction protocols.

    PubMed

    Najafzadeh, Nowruz; Sagha, Mohsen; Heydari Tajaddod, Shirin; Golmohammadi, Mohammad Ghasem; Massahi Oskoui, Nasim; Deldadeh Moghaddam, Maryam

    2015-02-01

    Differentiation of hair follicle stem cells (HFSCs) into neurons and glial cells represents a promising cell-based therapy for neurodegenerative diseases. The hair follicle bulge area is reported as a putative source of new stem cell population for many years. In vitro studies have implicated neural differentiation of HFSCs. Here, we report the identification and purification of CD34 (+) cells from hair follicle by magnetic activated cell sorting (MACS). We next determined the cytotoxic effects of all-trans retinoic acid (RA) by using cell viability assays. Moreover, the neural differentiation potential of CD34 (+) cells was evaluated in the presence of RA, serum-free condition, and neural differentiation medium (NDM) treatments by using immunocytochemistry and reverse transcription polymerase chain reaction (RT-PCR). Our results showed that the isolated CD34 (+) stem cells were 12% of the total cells in the bulge area, and the neural cells derived from the stem cells expressed nestin, microtubule-associated protein 2 (MAP2), and glial fibrillary acidic protein (GFAP). Interestingly, all the neural induction media supported neuronal differentiation most effectively, but treatment with serum-free medium significantly increased the number of GFAP-positive glial cells. Moreover, increasing RA concentration (≥10 μM) leads to increased cell death in the cells, but a lower concentration of RA (1 μM) treatment results in a decrease in CD34-expressing stem cells. These findings show an instructive neuronal effect of three neural induction media in HFSCs, indicating the important role of this induction media in the specification of the stem cells toward a neural phenotype.

  1. 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

  2. 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.

  3. [Neural differentiation of pluripotent stem cells and application for metal-induced neural toxicity study].

    PubMed

    Taniguchi, Yuki; Tobe, Takao; Hayami, Hideaki; Okamoto, Yoshinori; Ueda, Koji; Takada, Tatsuyuki; Kojima, Nakao

    2014-01-01

    Metals are effectively used in biological systems under the strict regulation for exploiting their specific and broad reactivities. For example, manganese (Mn) can induce catecholamines-mediated oxidative biological damage in cooperation with iron (Fe) and/or copper (Cu). In children, the damage could induce developmental disorders such as attention deficit hyperactivity disorder (ADHD). We hypothesize that infant neurons are more labile to metals than adult ones due to the prematured protection systems and sensitive differentiating cells. An experimental system reconstituting neural differentiation is expected to assess the influences of endogenous/exogenous factors including metals. In this study, we investigated an impact of Mn together with Fe and dopamine (DA) on neural differentiation of mouse embryonic stem cells (mESCs). The differentiation of mESCs was initiated by embryoid bodies (EBs) formation in the presence of all-trans retinoic acid, and then EBs were treated with Mn, Fe and/or DA. Then, the mRNA levels of neural differentiation marker genes (Nestin, Emx2, Mtap2, Th, Olig2 and Gfap) were examined using realtime RT-PCR analysis. Mn or DA alone reduced Mtap2, Th and Olig2 expression levels and increased Nestin. Moreover, combined treatment of Mn and DA also increased Nestin expression level. On the other hand, Fe alone reduced Mtap2, Th and Olig2 expression levels, and increased Emx2. Combined treatments of Fe with Mn or DA also tended to increase Emx2 expression level. These effects emerged at about 100 times less concentration than that inducing cytotoxicity in human neuroblastoma. The present study showed that Mn inhibits neural development, and that our mESCs system can be a useful tool to elucidate the toxicity mechanism as well as to evaluate the effects of metals and chemicals on differentiating cells. PMID:24989467

  4. Mammalian neural stem-cell renewal: nature versus nurture.

    PubMed

    Arsenijevic, Yvan

    2003-02-01

    Recent data show that the final events of mammalian brain organogenesis may depend in part on the direct control of neural stem cell (NSC) proliferation and survival. Environmental and intrinsic factors play a role throughout development and during adulthood to regulate NSC proliferation. The NSCs acquire new competences throughout development, including adulthood, and this change in competence is region-specific. The factors controlling NSC survival, undifferentiated state, proliferation, and cell-cycle number are beginning to be identified, but the links between them remain unclear. However, current knowledge should help to formulate an understanding of how a stem cell can generate a new stem cell. PMID:12668902

  5. Proteome analysis of proliferative response of bystander cells adjacent to cells exposed to ionizing radiation

    PubMed Central

    Gerashchenko, Bogdan I.; Yamagata, Akira; Oofusa, Ken; Yoshizato, Katsutoshi; de Toledo, Sonia M.; Howell, Roger W.

    2010-01-01

    Recently (Cytometry 2003, 56A, 71–80), we reported that direct cell-to-cell contact is required for stimulating proliferation of bystander rat liver cells (WB-F344) cocultured with irradiated cells, and neither functional gap junction intercellular communication nor long-range extracellular factors appear to be involved in this proliferative bystander response (PBR). The molecular basis for this response is unknown. Confluent monolayers of WB-F344 cells were exposed to 5-Gray (Gy) of γ-rays. Irradiated cells were mixed with unirradiated cells and co-cultured for 24 h. Cells were harvested and protein expression was examined using 2-DE. Protein expression was also determined in cultures of unirradiated and 5-Gy irradiated cells. Proteins were identified by MS. Nucleophosmin (NPM)-1, a multifunctional nucleolar protein, was more highly expressed in bystander cells than in either unirradiated or 5-Gy irradiated cells. Enolase-α, a glycolytic enzyme, was present in acidic and basic variants in unirradiated cells. In bystander and 5-Gy irradiated cells, the basic variant was weakly expressed, whereas the acidic variant was overwhelmingly present. These data indicate that the presence of irradiated cells can affect NPM-1 and enolase-α in adjacent bystander cells. These proteins appear to participate in molecular events related to the PBR and suggest that this response may involve cellular defense, proliferation, and metabolism. PMID:17514680

  6. Proteome analysis of proliferative response of bystander cells adjacent to cells exposed to ionizing radiation.

    PubMed

    Gerashchenko, Bogdan I; Yamagata, Akira; Oofusa, Ken; Yoshizato, Katsutoshi; de Toledo, Sonia M; Howell, Roger W

    2007-06-01

    Recently (Cytometry 2003, 56A, 71-80), we reported that direct cell-to-cell contact is required for stimulating proliferation of bystander rat liver cells (WB-F344) cocultured with irradiated cells, and neither functional gap junction intercellular communication nor long-range extracellular factors appear to be involved in this proliferative bystander response (PBR). The molecular basis for this response is unknown. Confluent monolayers of WB-F344 cells were exposed to 5-Gray (Gy) of gamma-rays. Irradiated cells were mixed with unirradiated cells and co-cultured for 24 h. Cells were harvested and protein expression was examined using 2-DE. Protein expression was also determined in cultures of unirradiated and 5-Gy irradiated cells. Proteins were identified by MS. Nucleophosmin (NPM)-1, a multifunctional nucleolar protein, was more highly expressed in bystander cells than in either unirradiated or 5-Gy irradiated cells. Enolase-alpha, a glycolytic enzyme, was present in acidic and basic variants in unirradiated cells. In bystander and 5-Gy irradiated cells, the basic variant was weakly expressed, whereas the acidic variant was overwhelmingly present. These data indicate that the presence of irradiated cells can affect NPM-1 and enolase-alpha in adjacent bystander cells. These proteins appear to participate in molecular events related to the PBR and suggest that this response may involve cellular defense, proliferation, and metabolism.

  7. Arsenic and Its Methylated Metabolites Inhibit the Differentiation of Neural Plate Border Specifier Cells.

    PubMed

    McCoy, Christopher R; Stadelman, Bradley S; Brumaghim, Julia L; Liu, Jui-Tung; Bain, Lisa J

    2015-07-20

    Exposure to arsenic in food and drinking water has been correlated with adverse developmental outcomes, such as reductions in birth weight and neurological deficits. Additionally, studies have shown that arsenic suppresses sensory neuron formation and skeletal muscle myogenesis, although the reason why arsenic targets both of these cell types in unclear. Thus, P19 mouse embryonic stem cells were used to investigate the mechanisms by which arsenic could inhibit cellular differentiation. P19 cells were exposed to 0, 0.1, or 0.5 μM sodium arsenite and induced to form embryoid bodies over a period of 5 days. The expression of transcription factors necessary to form neural plate border specifier (NPBS) cells, neural crest cells and their progenitors, and myocytes and their progenitors were examined. Early during differentiation, arsenic significantly reduced the transcript and protein expression of Msx1 and Pax3, both needed for NPBS cell formation. Arsenic also significantly reduced the protein expression of Sox 10, needed for neural crest progenitor cell production, by 31-50%, and downregulated the protein and mRNA levels of NeuroD1, needed for neural crest cell differentiation, in a time- and dose-dependent manner. While the overall protein expression of transcription factors in the skeletal muscle lineage was not changed, arsenic did alter their nuclear localization. MyoD nuclear translocation was significantly reduced on days 2-5 between 15 and 70%. At a 10-fold lower concentration, monomethylarsonous acid (MMA III) appeared to be just as potent as inorganic arsenic at reducing the mRNA levels Pax3 (79% vs84%), Sox10 (49% vs 65%), and Msx1 (56% vs 56%). Dimethylarsinous acid (DMA III) also reduced protein and transcript expression, but the changes were less dramatic than those with MMA or arsenite. All three arsenic species reduced the nuclear localization of MyoD and NeuroD1 in a similar manner. The early changes in the differentiation of neural plate border

  8. Immortalized neural progenitor cells for CNS gene transfer and repair.

    PubMed

    Martínez-Serrano, A; Björklund, A

    1997-11-01

    Immortalized multipotent neural stem and progenitor cells have emerged as a highly convenient source of tissue for genetic manipulation and ex vivo gene transfer to the CNS. Recent studies show that these cells, which can be maintained and genetically transduced as cell lines in culture, can survive, integrate and differentiate into both neurons and glia after transplantation to the intact or damaged brain. Progenitors engineered to secrete trophic factors, or to produce neurotransmitter-related or metabolic enzymes can be made to repopulate diseased or injured brain areas, thus providing a new potential therapeutic tool for the blockade of neurodegenerative processes and reversal of behavioural deficits in animal models of neurodegenerative diseases. With further technical improvements, the use of immortalized neural progenitors may bring us closer to the challenging goal of targeted and effective CNS repair.

  9. Cell polarity and neurogenesis in embryonic stem cell-derived neural rosettes.

    PubMed

    Banda, Erin; McKinsey, Anna; Germain, Noelle; Carter, James; Anderson, Nickesha Camille; Grabel, Laura

    2015-04-15

    Embryonic stem cells (ESCs) undergoing neural differentiation form radial arrays of neural stem cells, termed neural rosettes. These structures manifest many of the properties associated with embryonic and adult neurogenesis, including cell polarization, interkinetic nuclear migration (INM), and a gradient of neuronal differentiation. We now identify novel rosette structural features that serve to localize key regulators of neurogenesis. Cells within neural rosettes have specialized basal as well as apical surfaces, based on localization of the extracellular matrix receptor β1 integrin. Apical processes of cells in mature rosettes terminate at the lumen, where adherens junctions are apparent. Primary cilia are randomly distributed in immature rosettes and tightly associated with the neural stem cell's apical domain as rosettes mature. Components of two signaling pathways known to regulate neurogenesis in vivo and in rosettes, Hedgehog and Notch, are apically localized, with the Hedgehog effector Smoothened (Smo) associated with primary cilia and the Notch pathway γ-secretase subunit Presenilin 2 associated with the adherens junction. Increased neuron production upon treatment with the Notch inhibitor DAPT suggests a major role for Notch signaling in maintaining the neural stem cell state, as previously described. A less robust outcome was observed with manipulation of Hedgehog levels, though consistent with a role in neural stem cell survival or proliferation. Inhibition of both pathways resulted in an additive effect. These data support a model by which cells extending a process to the rosette lumen maintain neural stem cell identity whereas release from this association, either through asymmetric cell division or apical abscission, promotes neuronal differentiation.

  10. Planar cell polarity-mediated induction of neural stem cell expansion during axolotl spinal cord regeneration.

    PubMed

    Rodrigo Albors, Aida; Tazaki, Akira; Rost, Fabian; Nowoshilow, Sergej; Chara, Osvaldo; Tanaka, Elly M

    2015-11-14

    Axolotls are uniquely able to mobilize neural stem cells to regenerate all missing regions of the spinal cord. How a neural stem cell under homeostasis converts after injury to a highly regenerative cell remains unknown. Here, we show that during regeneration, axolotl neural stem cells repress neurogenic genes and reactivate a transcriptional program similar to embryonic neuroepithelial cells. This dedifferentiation includes the acquisition of rapid cell cycles, the switch from neurogenic to proliferative divisions, and the re-expression of planar cell polarity (PCP) pathway components. We show that PCP induction is essential to reorient mitotic spindles along the anterior-posterior axis of elongation, and orthogonal to the cell apical-basal axis. Disruption of this property results in premature neurogenesis and halts regeneration. Our findings reveal a key role for PCP in coordinating the morphogenesis of spinal cord outgrowth with the switch from a homeostatic to a regenerative stem cell that restores missing tissue.

  11. 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

  12. Biliary epithelium and liver B cells exposed to bacteria activate intrahepatic MAIT cells through MR1

    PubMed Central

    Jeffery, Hannah C.; van Wilgenburg, Bonnie; Kurioka, Ayako; Parekh, Krishan; Stirling, Kathryn; Roberts, Sheree; Dutton, Emma E.; Hunter, Stuart; Geh, Daniel; Braitch, Manjit K.; Rajanayagam, Jeremy; Iqbal, Tariq; Pinkney, Thomas; Brown, Rachel; Withers, David R.; Adams, David H.; Klenerman, Paul; Oo, Ye H.

    2016-01-01

    Background & Aims Mucosal-Associated Invariant T (MAIT) cells are innate-like T cells characterised by the invariant TCR-chain, Vα7.2-Jα33, and are restricted by MR1, which presents bacterial vitamin B metabolites. They are important for antibacterial immunity at mucosal sites; however, detailed characteristics of liver-infiltrating MAIT (LI-MAIT) and their role in biliary immune surveillance remain unexplored. Methods The phenotype and intrahepatic localisation of human LI-MAIT cells was examined in diseased and normal livers. MAIT cell activation in response to E. coli-exposed macrophages, biliary epithelial cells (BEC) and liver B cells was assessed with/without anti-MR1. Results Intrahepatic MAIT cells predominantly localised to bile ducts in the portal tracts. Consistent with this distribution, they expressed biliary tropic chemokine receptors CCR6, CXCR6, and integrin αEβ7. LI-MAIT cells were also present in the hepatic sinusoids and possessed tissue-homing chemokine receptor CXCR3 and integrins LFA-1 and VLA-4, suggesting their recruitment via hepatic sinusoids. LI-MAIT cells were enriched in the parenchyma of acute liver failure livers compared to chronic diseased livers. LI-MAIT cells had an activated, effector memory phenotype, expressed α4β7 and receptors for IL-12, IL-18, and IL-23. Importantly, in response to E. coli-exposed macrophages, liver B cells and BEC, MAIT cells upregulated IFN-γ and CD40 Ligand and degranulated in an MR1-dependent, cytokine-independent manner. In addition, diseased liver MAIT cells expressed T-bet and RORγt and the cytokines IFN-γ, TNF-α, and IL-17. Conclusions Our findings provide the first evidence of an immune surveillance effector response for MAIT cells towards BEC in human liver; thus they could be manipulated for treatment of biliary disease in the future. PMID:26743076

  13. Effects of ganglioside GM1 and neural growth factor on neural stem cell proliferation and differentiation.

    PubMed

    Wang, Q; Song, Y H; Tang, Z; Wang, Z P; Xu, Q; Bao, N

    2016-01-01

    Neurogenesis, recovery from nerve injury, neurodegeneration, and Parkinson's disease affect people's health, yet the underlying molecular mechanisms remain elusive. Here, we investigated the effect of ganglioside GM1 and neural growth factor (NGF) on neural stem cell (NSC) proliferation and differentiation in vitro to provide a scientific basis for comprehensive treatment of nervous system diseases via NSC application. As widely applied methods of relatively high accuracy, cell counts and 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assays were used to assess NSC proliferation. In addition, western blotting was employed to determine NSC differentiation. Cell counts and MTT assays demonstrated that in epidermal growth factor (EGF)- and basic fibroblast growth factor (bFGF)-containing medium, a high concentration of GM1, but not NGF, significantly elevated NSC proliferation. In NSC cultures lacking EGF and bFGF, cell counts and MTT values were significantly increased compared to those in the negative control group on days 4, 7, and 10 after GM1 (25, 100, and 200 ng/mL) but not NGF (25, 50, 100, and 200 ng/mL) treatment. Western blotting revealed significantly increased expression of nestin (an NSC marker) in NSCs treated with GM1, and upregulation of glial fibrillary acidic protein (a glial cell marker) and neuron-specific enolase (a neuron marker) in those administered NGF. Our results suggest that GM1 and NGF induce NSC proliferation and differentiation, respectively, in a dose-dependent manner. PMID:27525911

  14. Comparing neural networks: a benchmark on growing neural gas, growing cell structures, and fuzzy ARTMAP.

    PubMed

    Heinke, D; Hamker, F H

    1998-01-01

    This article compares the performance of some recently developed incremental neural networks with the wellknown multilayer perceptron (MLP) on real-world data. The incremental networks are fuzzy ARTMAP (FAM), growing neural gas (GNG) and growing cell structures (GCS). The real-world datasets consist of four different datasets posing different challenges to the networks in terms of complexity of decision boundaries, overlapping between classes, and size of the datasets. The performance of the networks on the datasets is reported with respect to measure classification error, number of training epochs, and sensitivity toward variation of parameters. Statistical evaluations are applied to examine the significance of the results. The overall performance ranks in the following descending order: GNG, GCS, MLP, FAM. PMID:18255809

  15. Inhibition of glycogen synthase kinase-3 (GSK3) promotes the neural differentiation of full-term amniotic fluid-derived stem cells towards neural progenitor cells.

    PubMed

    Gao, Liyang; Zhao, Mingyan; Ye, Wei; Huang, Jinzhi; Chu, Jiaqi; Yan, Shouquan; Wang, Chaojun; Zeng, Rong

    2016-08-01

    The amniotic fluid has a heterogeneous population of cells. Some human amniotic fluid-derived stem (hAFS) cells have been shown to harbor the potential to differentiate into neural cells. However, the neural differentiation efficiency of hAFS cells remains low. In this study, we isolated CD117-positive hAFS cells from amniotic fluid and then examined the pluripotency of these cells through the formation of embryoid bodies (EBs). Additionally, we induced the neural differentiation of these cells using neuroectodermal medium. This study revealed that the GSK3-beta inhibitor SB216763 was able to stimulate the proliferation of CD117-positive hAFS cells without influencing their undifferentiated state. Moreover, SB216763 can efficiently promote the neural differentiation of CD117-positive hAFS cells towards neural progenitor cells in the presence of DMEM/F12 and N2 supplement. These findings provide an easy and low-cost method to maintain the proliferation of hAFS cells, as well as induce an efficacious generation of neural progenitor cells from hAFS cells. Such induction of the neural commitment of hAFS cells may provide an option for the treatment of neurodegenerative diseases by hAFS cells-based therapies.

  16. 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

  17. 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.

  18. L1 Retrotransposition in Neural Progenitor Cells.

    PubMed

    Muotri, Alysson R

    2016-01-01

    Long interspersed nucleotide element 1 (LINE-1 or L1) is a family of non-LTR retrotransposons that can replicate and reintegrate into the host genome. L1s have considerably influenced mammalian genome evolution by retrotransposing during germ cell development or early embryogenesis, leading to massive genome expansion. For many years, L1 retrotransposons were viewed as a selfish DNA parasite that had no contribution in somatic cells. Historically, L1s were thought to only retrotranspose during gametogenesis and in neoplastic processes, but recent studies have shown that L1s are extremely active in the mouse, rat, and human neuronal progenitor cells (NPCs). These de novo L1 insertions can impact neuronal transcriptional expression, creating unique transcriptomes of individual neurons, possibly contributing to the uniqueness of the individual cognition and mental disorders in humans. PMID:26895053

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

    PubMed

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

    2014-06-01

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

  20. 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. Proteome-wide analysis of neural stem cell differentiation to facilitate transition to cell replacement therapies.

    PubMed

    Zizkova, Martina; Sucha, Rita; Tyleckova, Jirina; Jarkovska, Karla; Mairychova, Katerina; Kotrcova, Eva; Marsala, Martin; Gadher, Suresh Jivan; Kovarova, Hana

    2015-02-01

    Neurodegenerative diseases are devastating disorders and the demands on their treatment are set to rise in connection with higher disease incidence. Knowledge of the spatiotemporal profile of cellular protein expression during neural differentiation and definition of a set of markers highly specific for targeted neural populations is a key challenge. Intracellular proteins may be utilized as a readout for follow-up transplantation and cell surface proteins may facilitate isolation of the cell subpopulations, while secreted proteins could help unravel intercellular communication and immunomodulation. This review summarizes the potential of proteomics in revealing molecular mechanisms underlying neural differentiation of stem cells and presents novel candidate proteins of neural subpopulations, where understanding of their functionality may accelerate transition to cell replacement therapies.

  2. 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

  3. 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.

  4. Cytoplasmic myosin-exposed apoptotic cells appear with caspase-3 activation and enhance CLL cell viability.

    PubMed

    Cui, X; Zhang, L; Magli, A R; Catera, R; Yan, X-J; Griffin, D O; Rothstein, T L; Barrientos, J; Kolitz, J E; Allen, S L; Rai, K R; Chiorazzi, N; Chu, C C

    2016-01-01

    The degree of chronic lymphocytic leukemia (CLL) B-cell antigen receptor (BCR) binding to myosin-exposed apoptotic cells (MEACs) correlates with worse patient outcomes, suggesting a link to disease activity. Therefore, we studied MEAC formation and the effects of MEAC binding on CLL cells. In cell line studies, both intrinsic (spontaneous or camptothecin-induced) and extrinsic (FasL- or anti-Fas-induced) apoptosis created a high percent of MEACs over time in a process associated with caspase-3 activation, leading to cytoplasmic myosin cleavage and trafficking to cell membranes. The involvement of common apoptosis pathways suggests that most cells can produce MEACs and indeed CLL cells themselves form MEACs. Consistent with the idea that MEAC formation may be a signal to remove dying cells, we found that natural IgM antibodies bind to MEACs. Functionally, co-culture of MEACs with CLL cells, regardless of immunoglobulin heavy-chain variable region gene mutation status, improved leukemic cell viability. Based on inhibitor studies, this improved viability involved BCR signaling molecules. These results support the hypothesis that stimulation of CLL cells with antigen, such as those on MEACs, promotes CLL cell viability, which in turn could lead to progression to worse disease. PMID:26220042

  5. Marmoset induced pluripotent stem cells: Robust neural differentiation following pretreatment with dimethyl sulfoxide.

    PubMed

    Qiu, Zhifang; Mishra, Anuja; Li, Miao; Farnsworth, Steven L; Guerra, Bernadette; Lanford, Robert E; Hornsby, Peter J

    2015-07-01

    The marmoset is an important nonhuman primate model for regenerative medicine. For experimental autologous cell therapy based on induced pluripotent (iPS) cells in the marmoset, cells must be able to undergo robust and reliable directed differentiation that will not require customization for each specific iPS cell clone. When marmoset iPS cells were aggregated in a hanging drop format for 3 days, followed by exposure to dual SMAD inhibitors and retinoic acid in monolayer culture for 3 days, we found substantial variability in the response of different iPS cell clones. However, when clones were pretreated with 0.05-2% dimethyl sulfoxide (DMSO) for 24 hours, all clones showed a very similar maximal response to the directed differentiation scheme. Peak responses were observed at 0.5% DMSO in two clones and at 1% DMSO in a third clone. When patterns of gene expression were examined by microarray analysis, hierarchical clustering showed very similar responses in all 3 clones when they were pretreated with optimal DMSO concentrations. The change in phenotype following exposure to DMSO and the 6 day hanging drop/monolayer treatment was confirmed by immunocytochemistry. Analysis of DNA content in DMSO-exposed cells indicated that it is unlikely that DMSO acts by causing cells to exit from the cell cycle. This approach should be generally valuable in the directed neural differentiation of pluripotent cells for experimental cell therapy. PMID:26070112

  6. Elimination of the geomagnetic field stimulates the proliferation of mouse neural progenitor and stem cells.

    PubMed

    Fu, Jing-Peng; Mo, Wei-Chuan; Liu, Ying; Bartlett, Perry F; He, Rong-Qiao

    2016-09-01

    Living organisms are exposed to the geomagnetic field (GMF) throughout their lifespan. Elimination of the GMF, resulting in a hypogeomagnetic field (HMF), leads to central nervous system dysfunction and abnormal development in animals. However, the cellular mechanisms underlying these effects have not been identified so far. Here, we show that exposure to an HMF (<200 nT), produced by a magnetic field shielding chamber, promotes the proliferation of neural progenitor/stem cells (NPCs/NSCs) from C57BL/6 mice. Following seven-day HMF-exposure, the primary neurospheres (NSs) were significantly larger in size, and twice more NPCs/NSCs were harvested from neonatal NSs, when compared to the GMF controls. The self-renewal capacity and multipotency of the NSs were maintained, as HMF-exposed NSs were positive for NSC markers (Nestin and Sox2), and could differentiate into neurons and astrocyte/glial cells and be passaged continuously. In addition, adult mice exposed to the HMF for one month were observed to have a greater number of proliferative cells in the subventricular zone. These findings indicate that continuous HMF-exposure increases the proliferation of NPCs/NSCs, in vitro and in vivo. HMF-disturbed NPCs/NSCs production probably affects brain development and function, which provides a novel clue for elucidating the cellular mechanisms of the bio-HMF response. PMID:27484904

  7. Recombinant spider silk matrices for neural stem cell cultures.

    PubMed

    Lewicka, Michalina; Hermanson, Ola; Rising, Anna U

    2012-11-01

    Neural stem cells (NSCs) have the capacity to differentiate into neurons, astrocytes, and oligodendrocytes. Accordingly, NSCs hold great promise in drug screening and treatment of several common diseases. However, a major obstacle in applied stem cell research is the limitation of synthetic matrices for culturing stem cells. The objective of this study was to evaluate the suitability of recombinant spider silk (4RepCT) matrices for growth of NSCs. NSCs isolated from the cerebral cortices of mid-gestation rat embryos were cultured on either 4RepCT matrices or conventional poly-L-ornithine and fibronectin (P + F) coated polystyrene plates. From 48 h of culture, no significant differences in cell proliferation or viability were detected in NSC cultures on 4RepCT compared to control matrices (polystyrene plates coated with P + F). The NSCs retained an undifferentiated state, displaying low or no staining for markers of differentiated cells. Upon stimulation NSCs grown on 4RepCT differentiated efficiently into neuronal and astrocytic cells to virtually the same degree as control cultures, but a slightly less efficient oligodendrocyte differentiation was noted. We suggest that recombinant spider silk matrices provide a functional microenvironment and represent a useful tool for the development of new strategies in neural stem cell research. PMID:22863380

  8. 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.

  9. Inherited effects of low-dose exposure to methylmercury in neural stem cells.

    PubMed

    Bose, Raj; Onishchenko, Natalia; Edoff, Karin; Janson Lang, Ann Marie; Ceccatelli, Sandra

    2012-12-01

    Methylmercury (MeHg) is an environmental contaminant with recognized neurotoxic effects, particularly to the developing nervous system. In the present study, we show that nanomolar concentrations of MeHg can induce long-lasting effects in neural stem cells (NSCs). We investigated short-term direct and long-term inherited effects of exposure to MeHg (2.5 or 5.0 nM) using primary cultures of rat embryonic cortical NSCs. We found that MeHg had no adverse effect on cell viability but reduced NSC proliferation and altered the expression of cell cycle regulators (p16 and p21) and senescence-associated markers. In addition, we demonstrated a decrease in global DNA methylation in the exposed cells, indicating that epigenetic changes may be involved in the mechanisms underlying the MeHg-induced effects. These changes were observed in cells directly exposed to MeHg (parent cells) and in their daughter cells cultured under MeHg-free conditions. In agreement with our in vitro data, a trend was found for decreased cell proliferation in the subgranular zone in the hippocampi of adult mice exposed to low doses of MeHg during the perinatal period. Interestingly, this impaired proliferation had a measurable impact on the total number of neurons in the hippocampal dentate gyrus. Importantly, this effect could be reversed by chronic antidepressant treatment. Our study provides novel evidence for programming effects induced by MeHg in NSCs and supports the idea that developmental exposure to low levels of MeHg may result in long-term consequences predisposing to neurodevelopmental disorders and/or neurodegeneration.

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

    PubMed

    Suksuphew, Sarawut; Noisa, Parinya

    2015-03-26

    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.

  11. Neural stem cell-based treatment for neurodegenerative diseases.

    PubMed

    Kim, Seung U; Lee, Hong J; Kim, Yun B

    2013-10-01

    Human neurodegenerative diseases such as Parkinson's disease (PD), Huntington's disease (HD), amyotrophic lateral sclerosis (ALS) and Alzheimer's disease (AD) are caused by a loss of neurons and glia in the brain or spinal cord. Neurons and glial cells have successfully been generated from stem cells such as embryonic stem cells (ESCs), mesenchymal stem cells (MSCs) and neural stem cells (NSCs), and stem cell-based cell therapies for neurodegenerative diseases have been developed. A recent advance in generation of a new class of pluripotent stem cells, induced pluripotent stem cells (iPSCs), derived from patients' own skin fibroblasts, opens doors for a totally new field of personalized medicine. Transplantation of NSCs, neurons or glia generated from stem cells in animal models of neurodegenerative diseases, including PD, HD, ALS and AD, demonstrates clinical improvement and also life extension of these animals. Additional therapeutic benefits in these animals can be provided by stem cell-mediated gene transfer of therapeutic genes such as neurotrophic factors and enzymes. Although further research is still needed, cell and gene therapy based on stem cells, particularly using neurons and glia derived from iPSCs, ESCs or NSCs, will become a routine treatment for patients suffering from neurodegenerative diseases and also stroke and spinal cord injury.

  12. Platelet microparticles promote neural stem cell proliferation, survival and differentiation.

    PubMed

    Hayon, Yael; Dashevsky, Olga; Shai, Ela; Varon, David; Leker, Ronen R

    2012-07-01

    Platelet microparticles (PMP) are small subcellular fragments, shed upon platelet activation. PMP host a variety of cytokines and growth factor that were previously shown to affect angiogenesis and postischemic tissue regeneration. This study attempted to explore the effect of PMP on neural stem cell (NSC) proliferation, survival and differentiation. Cells were grown as neurospheres and treated with PMP, or relevant growth factors, sphere size and cell fates were evaluated. PMP treatment led to larger neurospheres with increased cell survival. PMP treatment was comparable with the effect of acceptable single growth factors such as fibroblastic growth factor (FGF), vascular endothelial growth factor (VEGF) and platelet derived growth factor (PDGF). PMP treatment also increased the differentiation potential of NSC to glia and neurons. Specific growth factor inhibitors only partly blocked these effects, which were associated with increments in ERK and Akt phosphorylation. In this study, we show that various growth factors contained within the PMP promote neuronal cell proliferation, survival and differentiation. The results suggest a role for platelet microparticles in augmenting endogenous neural progenitor and stem cells angiogenesis and neurogenesis that might be utilized for treatment following brain injury.

  13. 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

  14. 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

  15. Spatial and temporal control of cell aggregation efficiently directs human pluripotent stem cells towards neural commitment.

    PubMed

    Miranda, Cláudia C; Fernandes, Tiago G; Pascoal, Jorge F; Haupt, Simone; Brüstle, Oliver; Cabral, Joaquim M S; Diogo, Maria Margarida

    2015-10-01

    3D suspension culture is generally considered a promising method to achieve efficient expansion and controlled differentiation of human pluripotent stem cells (hPSCs). In this work, we focused on developing an integrated culture platform for expansion and neural commitment of hPSCs into neural precursors using 3D suspension conditions and chemically-defined culture media. We evaluated different inoculation methodologies for hPSC expansion as 3D aggregates and characterized the resulting cultures in terms of aggregate size distribution. It was demonstrated that upon single-cell inoculation, after four days of culture, 3D aggregates were composed of homogenous populations of hPSC and were characterized by an average diameter of 139 ± 26 μm, which was determined to be the optimal size to initiate neural commitment. Temporal analysis revealed that upon neural specification it is possible to maximize the percentage of neural precursor cells expressing the neural markers Sox1 and Pax6 after nine days of culture. These results highlight our ability to define a robust method for production of hPSC-derived neural precursors that minimizes processing steps and that constitutes a promising alternative to the traditional planar adherent culture system due to a high potential for scaling-up. PMID:25866360

  16. Webs, cell assemblies, and chunking in neural nets: introduction.

    PubMed

    Wickelgren, W A

    1999-03-01

    This introduction to Wickelgren (1992), describes a theory of idea representation and learning in the cerebral cortex and seven properties of Hebb's (1949) formulation of cell assemblies that have played a major role in all such neural net models. Ideas are represented in the cerebral cortex by webs (innate cell assemblies), using sparse coding with sparse, all-or-none, innate linking. Recruiting a web to represent a new idea is called chunking. The innate links that bind the neurons of a web are basal dendritic synapses. Learning modifies the apical dendritic synapses that associate neurons in one web to neurons in another web.

  17. Selective migration of neuralized embryonic stem cells to stem cell factor and media conditioned by glioma cell lines

    PubMed Central

    Serfozo, Peter; Schlarman, Maggie S; Pierret, Chris; Maria, Bernard L; Kirk, Mark D

    2006-01-01

    Background Pluripotent mouse embryonic stem (ES) cells can be induced in vitro to become neural progenitors. Upon transplantation, neural progenitors migrate toward areas of damage and inflammation in the CNS. We tested whether undifferentiated and neuralized mouse ES cells migrate toward media conditioned by glioma cell lines (C6, U87 & N1321) or Stem Cell Factor (SCF). Results Cell migration assays revealed selective migration by neuralized ES cells to conditioned media as well as to synthetic SCF. Migration of undifferentiated ES cells was extensive, but not significantly different from that of controls (Unconditioned Medium). RT-PCR analysis revealed that all the three tumor cell lines tested synthesized SCF and that both undifferentiated and neuralized ES cells expressed c-kit, the receptor for SCF. Conclusion Our results demonstrate that undifferentiated ES cells are highly mobile and that neural progenitors derived from ES cells are selectively attracted toward factors produced by gliomas. Given that the glioma cell lines synthesize SCF, SCF may be one of several factors that contribute to the selective migration observed. PMID:16436212

  18. In Vivo Generation of Neural Stem Cells Through Teratoma Formation.

    PubMed

    Hong, Yean Ju; Kim, Jong Soo; Choi, Hyun Woo; Song, Hyuk; Park, Chankyu; Do, Jeong Tae

    2016-09-01

    Pluripotent stem cells have the potential to differentiate into all cell types of the body in vitro through embryoid body formation or in vivo through teratoma formation. In this study, we attempted to generate in vivo neural stem cells (NSCs) differentiated through teratoma formation using Olig2-GFP transgenic embryonic stem cells (ESCs). After 4 to 6 weeks of injection with Olig2-GFP transgenic ESCs, Olig2-GFP(+) NSCs were identified in teratomas formed in immunodeficient mice. Interestingly, 4-week-old teratomas contained higher percentage of Olig2-GFP(+) cells (∼11%) than 6-week-old teratomas (∼3%). These in vivo-derived NSCs expressed common NSC markers (Nestin and Sox2) and differentiated into terminal neuronal and glial lineages. These results suggest that pure NSC populations exhibiting properties similar to those of brain-derived NSCs can be established through teratoma formation. PMID:27439546

  19. Axonal control of the adult neural stem cell niche.

    PubMed

    Tong, Cheuk Ka; Chen, Jiadong; Cebrián-Silla, Arantxa; Mirzadeh, Zaman; Obernier, Kirsten; Guinto, Cristina D; Tecott, Laurence H; García-Verdugo, Jose Manuel; Kriegstein, Arnold; Alvarez-Buylla, Arturo

    2014-04-01

    The ventricular-subventricular zone (V-SVZ) is an extensive germinal niche containing neural stem cells (NSCs) in the walls of the lateral ventricles of the adult brain. How the adult brain's neural activity influences the behavior of adult NSCs remains largely unknown. We show that serotonergic (5HT) axons originating from a small group of neurons in the raphe form an extensive plexus on most of the ventricular walls. Electron microscopy revealed intimate contacts between 5HT axons and NSCs (B1) or ependymal cells (E1) and these cells were labeled by a transsynaptic viral tracer injected into the raphe. B1 cells express the 5HT receptors 2C and 5A. Electrophysiology showed that activation of these receptors in B1 cells induced small inward currents. Intraventricular infusion of 5HT2C agonist or antagonist increased or decreased V-SVZ proliferation, respectively. These results indicate that supraependymal 5HT axons directly interact with NSCs to regulate neurogenesis via 5HT2C. PMID:24561083

  20. Axonal Control of the Adult Neural Stem Cell Niche

    PubMed Central

    Tong, Cheuk Ka; Chen, Jiadong; Cebrián-Silla, Arantxa; Mirzadeh, Zaman; Obernier, Kirsten; Guinto, Cristina D.; Tecott, Laurence H.; García-Verdugo, Jose Manuel; Kriegstein, Arnold; Alvarez-Buylla, Arturo

    2014-01-01

    SUMMARY The ventricular-subventricular zone (V-SVZ) is an extensive germinal niche containing neural stem cells (NSC) in the walls of the lateral ventricles of the adult brain. How the adult brain’s neural activity influences the behavior of adult NSCs remains largely unknown. We show that serotonergic (5HT) axons originating from a small group of neurons in the raphe form an extensive plexus on most of the ventricular walls. Electron microscopy revealed intimate contacts between 5HT axons and NSCs (B1) or ependymal cells (E1) and these cells were labeled by a transsynaptic viral tracer injected into the raphe. B1 cells express the 5HT receptors 2C and 5A. Electrophysiology showed that activation of these receptors in B1 cells induced small inward currents. Intraventricular infusion of 5HT2C agonist or antagonist increased or decreased V-SVZ proliferation, respectively. These results indicate that supraependymal 5HT axons directly interact with NSCs to regulate neurogenesis via 5HT2C. PMID:24561083

  1. 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

  2. Neural stem cells in the adult human brain

    PubMed Central

    Gonzalez-Perez, Oscar

    2012-01-01

    For decades, it was believed that the adult brain was a quiescent organ unable to produce new neurons. At the beginning of the1960's, this dogma was challenged by a small group of neuroscientists. To date, it is well-known that new neurons are generated in the adult brain throughout life. Adult neurogenesis is primary confined to the subventricular zone (SVZ) of the forebrain and the subgranular zone of the dentate gyrus within the hippocampus. In both the human and the rodent brain, the primary progenitor of adult SVZ is a subpopulation of astrocytes that have stem-cell-like features. The human SVZ possesses a peculiar cell composition and displays important organizational differences when compared to the SVZ of other mammals. Some evidence suggests that the human SVZ may be not only an endogenous source of neural precursor cells for brain repair, but also a source of brain tumors. In this review, we described the cytoarchitecture and cellular composition of the SVZ in the adult human brain. We also discussed some clinical implications of SVZ, such as: stem-cell-based therapies against neurodegenerative diseases and its potential as a source of malignant cells. Understanding the biology of human SVZ and its neural progenitors is one of the crucial steps to develop novel therapies against neurological diseases in humans. PMID:23181200

  3. 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

  4. N-cadherin-mediated cell adhesion restricts cell proliferation in the dorsal neural tube.

    PubMed

    Chalasani, Kavita; Brewster, Rachel M

    2011-05-01

    Neural progenitors are organized as a pseudostratified epithelium held together by adherens junctions (AJs), multiprotein complexes composed of cadherins and α- and β-catenin. Catenins are known to control neural progenitor division; however, it is not known whether they function in this capacity as cadherin binding partners, as there is little evidence that cadherins themselves regulate neural proliferation. We show here that zebrafish N-cadherin (N-cad) restricts cell proliferation in the dorsal region of the neural tube by regulating cell-cycle length. We further reveal that N-cad couples cell-cycle exit and differentiation, as a fraction of neurons are mitotic in N-cad mutants. Enhanced proliferation in N-cad mutants is mediated by ligand-independent activation of Hedgehog (Hh) signaling, possibly caused by defective ciliogenesis. Furthermore, depletion of Hh signaling results in the loss of junctional markers. We therefore propose that N-cad restricts the response of dorsal neural progenitors to Hh and that Hh signaling limits the range of its own activity by promoting AJ assembly. Taken together, these observations emphasize a key role for N-cad-mediated adhesion in controlling neural progenitor proliferation. In addition, these findings are the first to demonstrate a requirement for cadherins in synchronizing cell-cycle exit and differentiation and a reciprocal interaction between AJs and Hh signaling.

  5. Neural tube closure in Xenopus laevis involves medial migration, directed protrusive activity, cell intercalation and convergent extension.

    PubMed

    Davidson, L A; Keller, R E

    1999-10-01

    We have characterized the cell movements and prospective cell identities as neural folds fuse during neural tube formation in Xenopus laevis. A newly developed whole-mount, two-color fluorescent RNA in situ hybridization method, visualized with confocal microscopy, shows that the dorsal neural tube gene xpax3 and the neural-crest-specific gene xslug are expressed far lateral to the medial site of neural fold fusion and that expression moves medially after fusion. To determine whether cell movements or dynamic changes in gene expression are responsible, we used low-light videomicroscopy followed by fluorescent in situ and confocal microscopy. These methods revealed that populations of prospective neural crest and dorsal neural tube cells near the lateral margin of the neural plate at the start of neurulation move to the dorsal midline using distinctive forms of motility. Before fold fusion, superficial neural cells apically contract, roll the neural plate into a trough and appear to pull the superficial epidermal cell sheet medially. After neural fold fusion, lateral deep neural cells move medially by radially intercalating between other neural cells using two types of motility. The neural crest cells migrate as individual cells toward the dorsal midline using medially directed monopolar protrusions. These movements combine the two lateral populations of neural crest into a single medial population that form the roof of the neural tube. The remaining cells of the dorsal neural tube extend protrusions both medially and laterally bringing about radial intercalation of deep and superficial cells to form a single-cell-layered, pseudostratified neural tube. While ours is the first description of medially directed cell migration during neural fold fusion and re-establishment of the neural tube, these complex cell behaviors may be involved during cavitation of the zebrafish neural keel and secondary neurulation in the posterior axis of chicken and mouse.

  6. Nanosized zinc oxide particles induce neural stem cell apoptosis

    NASA Astrophysics Data System (ADS)

    Deng, Xiaoyong; Luan, Qixia; Chen, Wenting; Wang, Yanli; Wu, Minghong; Zhang, Haijiao; Jiao, Zheng

    2009-03-01

    Given the intensive application of nanoscale zinc oxide (ZnO) materials in our life, growing concerns have arisen about its unintentional health and environmental impacts. In this study, the neurotoxicity of different sized ZnO nanoparticles in mouse neural stem cells (NSCs) was investigated. A cell viability assay indicated that ZnO nanoparticles manifested dose-dependent, but no size-dependent toxic effects on NSCs. Apoptotic cells were observed and analyzed by confocal microscopy, transmission electron microscopy examination, and flow cytometry. All the results support the viewpoint that the ZnO nanoparticle toxicity comes from the dissolved Zn2+ in the culture medium or inside cells. Our results highlight the need for caution during the use and disposal of ZnO manufactured nanomaterials to prevent the unintended environmental and health impacts.

  7. 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

  8. The ubiquitous neural cell adhesion molecule (N-CAM).

    PubMed

    Weledji, Elroy P; Assob, Jules C

    2014-09-01

    Adhesive interactions are important for cell trafficking, differentiation, function and tissue differentiation. Neural cell adhesion molecule (NCAM) is involved in a diverse range of contact-mediated interactions among neurons, astrocytes, oligodendrocytes, and myotubes. It is widely but transiently expressed in many tissues early in embryogenesis. Four main isoforms exist but there are many other variants resulting from alternative splicing and post-translational modifications. This review discusses the actions and association of N-CAM and variants, PSA CAM. L1CAM and receptor tyrosine kinase. Their interactions with the interstitial cells of Cajal - the pacemaker cells of the gut in the manifestation of gut motility disorders, expression in carcinomas and mesenchymal tumours are discussed. PMID:25568792

  9. NFL-lipid nanocapsules for brain neural stem cell targeting in vitro and in vivo.

    PubMed

    Carradori, Dario; Saulnier, Patrick; Préat, Véronique; des Rieux, Anne; Eyer, Joel

    2016-09-28

    The replacement of injured neurons by the selective stimulation of neural stem cells in situ represents a potential therapeutic strategy for the treatment of neurodegenerative diseases. The peptide NFL-TBS.40-63 showed specific interactions towards neural stem cells of the subventricular zone. The aim of our work was to produce a NFL-based drug delivery system able to target neural stem cells through the selective affinity between the peptide and these cells. NFL-TBS.40-63 (NFL) was adsorbed on lipid nanocapsules (LNC) whom targeting efficiency was evaluated on neural stem cells from the subventricular zone (brain) and from the central canal (spinal cord). NFL-LNC were incubated with primary neural stem cells in vitro or injected in vivo in adult rat brain (right lateral ventricle) or spinal cord (T10). NFL-LNC interactions with neural stem cells were different depending on the origin of the cells. NFL-LNC showed a preferential uptake by neural stem cells from the brain, while they did not interact with neural stem cells from the spinal cord. The results obtained in vivo correlate with the results observed in vitro, demonstrating that NFL-LNC represent a promising therapeutic strategy to selectively deliver bioactive molecules to brain neural stem cells.

  10. NFL-lipid nanocapsules for brain neural stem cell targeting in vitro and in vivo.

    PubMed

    Carradori, Dario; Saulnier, Patrick; Préat, Véronique; des Rieux, Anne; Eyer, Joel

    2016-09-28

    The replacement of injured neurons by the selective stimulation of neural stem cells in situ represents a potential therapeutic strategy for the treatment of neurodegenerative diseases. The peptide NFL-TBS.40-63 showed specific interactions towards neural stem cells of the subventricular zone. The aim of our work was to produce a NFL-based drug delivery system able to target neural stem cells through the selective affinity between the peptide and these cells. NFL-TBS.40-63 (NFL) was adsorbed on lipid nanocapsules (LNC) whom targeting efficiency was evaluated on neural stem cells from the subventricular zone (brain) and from the central canal (spinal cord). NFL-LNC were incubated with primary neural stem cells in vitro or injected in vivo in adult rat brain (right lateral ventricle) or spinal cord (T10). NFL-LNC interactions with neural stem cells were different depending on the origin of the cells. NFL-LNC showed a preferential uptake by neural stem cells from the brain, while they did not interact with neural stem cells from the spinal cord. The results obtained in vivo correlate with the results observed in vitro, demonstrating that NFL-LNC represent a promising therapeutic strategy to selectively deliver bioactive molecules to brain neural stem cells. PMID:27503706

  11. miR-381 Regulates Neural Stem Cell Proliferation and Differentiation via Regulating Hes1 Expression

    PubMed Central

    Liu, Baoquan; Yang, Chunxiao; Nie, Xuedan; Wang, Xiaokun; Zheng, Jiaolin; Wang, Yue; Zhu, Yulan

    2015-01-01

    Neural stem cells are self-renewing, multipotent and undifferentiated precursors that retain the capacity for differentiation into both glial (astrocytes and oligodendrocytes) and neuronal lineages. Neural stem cells offer cell-based therapies for neurological disorders such as Alzheimer's disease, Parkinson's disease, Huntington's disease and spinal cord injuries. However, their cellular behavior is poorly understood. MicroRNAs (miRNAs) are a class of small noncoding RNAs involved in cell development, proliferation and differentiation through regulating gene expression at post-transcriptional level. The role of miR–381 in the development of neural stem cells remains unknown. In this study, we showed that overexpression of miR–381 promoted neural stem cells proliferation. It induced the neural stem cells differentiation to neurons and inhibited their differentiation to astrocytes. Furthermore, we identified HES1 as a direct target of miR–381 in neural stem cells. Moreover, re-expression of HES1 impaired miR-381-induced promotion of neural stem cells proliferation and induce neural stem cells differentiation to neurons. In conclusion, miR–381 played important role in neural stem cells proliferation and differentiation. PMID:26431046

  12. Neural differentiation from pluripotent stem cells: The role of natural and synthetic extracellular matrix

    PubMed Central

    Li, Yan; Liu, Meimei; Yan, Yuanwei; Yang, Shang-Tian

    2014-01-01

    Neural cells differentiated from pluripotent stem cells (PSCs), including both embryonic stem cells and induced pluripotent stem cells, provide a powerful tool for drug screening, disease modeling and regenerative medicine. High-purity oligodendrocyte progenitor cells (OPCs) and neural progenitor cells (NPCs) have been derived from PSCs recently due to the advancements in understanding the developmental signaling pathways. Extracellular matrices (ECM) have been shown to play important roles in regulating the survival, proliferation, and differentiation of neural cells. To improve the function and maturation of the derived neural cells from PSCs, understanding the effects of ECM over the course of neural differentiation of PSCs is critical. During neural differentiation of PSCs, the cells are sensitive to the properties of natural or synthetic ECMs, including biochemical composition, biomechanical properties, and structural/topographical features. This review summarizes recent advances in neural differentiation of human PSCs into OPCs and NPCs, focusing on the role of ECM in modulating the composition and function of the differentiated cells. Especially, the importance of using three-dimensional ECM scaffolds to simulate the in vivo microenvironment for neural differentiation of PSCs is highlighted. Future perspectives including the immediate applications of PSC-derived neural cells in drug screening and disease modeling are also discussed. PMID:24567784

  13. Two developmentally distinct populations of neural crest cells contribute to the zebrafish heart.

    PubMed

    Cavanaugh, Ann M; Huang, Jie; Chen, Jau-Nian

    2015-08-15

    Cardiac neural crest cells are essential for outflow tract remodeling in animals with divided systemic and pulmonary circulatory systems, but their contributions to cardiac development in animals with a single-loop circulatory system are less clear. Here we genetically labeled neural crest cells and examined their contribution to the developing zebrafish heart. We identified two populations of neural crest cells that contribute to distinct compartments of zebrafish cardiovascular system at different developmental stages. A stream of neural crest cells migrating through pharyngeal arches 1 and 2 integrates into the myocardium of the primitive heart tube between 24 and 30 h post fertilization and gives rise to cardiomyocytes. A second wave of neural crest cells migrating along aortic arch 6 envelops the endothelium of the ventral aorta and invades the bulbus arteriosus after three days of development. Interestingly, while inhibition of FGF signaling has no effect on the integration of neural crest cells to the primitive heart tube, it prevents these cells from contributing to the outflow tract, demonstrating disparate responses of neural crest cells to FGF signaling. Furthermore, neural crest ablation in zebrafish leads to multiple cardiac defects, including reduced heart rate, defective myocardial maturation and a failure to recruit progenitor cells from the second heart field. These findings add to our understanding of the contribution of neural crest cells to the developing heart and provide insights into the requirement for these cells in cardiac maturation.

  14. Neural differentiation from pluripotent stem cells: The role of natural and synthetic extracellular matrix.

    PubMed

    Li, Yan; Liu, Meimei; Yan, Yuanwei; Yang, Shang-Tian

    2014-01-26

    Neural cells differentiated from pluripotent stem cells (PSCs), including both embryonic stem cells and induced pluripotent stem cells, provide a powerful tool for drug screening, disease modeling and regenerative medicine. High-purity oligodendrocyte progenitor cells (OPCs) and neural progenitor cells (NPCs) have been derived from PSCs recently due to the advancements in understanding the developmental signaling pathways. Extracellular matrices (ECM) have been shown to play important roles in regulating the survival, proliferation, and differentiation of neural cells. To improve the function and maturation of the derived neural cells from PSCs, understanding the effects of ECM over the course of neural differentiation of PSCs is critical. During neural differentiation of PSCs, the cells are sensitive to the properties of natural or synthetic ECMs, including biochemical composition, biomechanical properties, and structural/topographical features. This review summarizes recent advances in neural differentiation of human PSCs into OPCs and NPCs, focusing on the role of ECM in modulating the composition and function of the differentiated cells. Especially, the importance of using three-dimensional ECM scaffolds to simulate the in vivo microenvironment for neural differentiation of PSCs is highlighted. Future perspectives including the immediate applications of PSC-derived neural cells in drug screening and disease modeling are also discussed. PMID:24567784

  15. Development of a microfluidic platform with integrated power splitting waveguides for optogenetic neural cell stimulation.

    PubMed

    Feng, Hongtao; Shu, Weiliang; Chen, Xi; Zhang, Yuanyuan; Lu, Yi; Wang, Liping; Chen, Yan

    2015-10-01

    We present a microfluidic platform with integrated power splitting waveguides for optogenetic neural cell stimulation. A liquid-core/PDMS-cladding waveguide with a power splitter design was integrated with a neural cell culture chamber to provide a simple way of precise localized optical stimulation. The parallel on-chip excitation of individual neural cells using a single optical fiber input is demonstrated for optogenetic neural cell studies, and the excitation of each individual waveguide can be independently controlled by pneumatic valves. Light delivery and loss mechanisms through the waveguides were studied and characterized. The waveguide power splitter platform is capable of providing sufficient irradiance to evoke spikes in ChR2-expressing neural cells. The system enables high-resolution stimulation of neural cells in a controllable manner. The microfluidic platform described here represents a novel methodology for studying optogenetics in a compact integrated system with high spatial resolutions. PMID:26371060

  16. Ultrastructural changes in tracheal epithelial cells exposed to oxygen

    NASA Technical Reports Server (NTRS)

    Philpott, D. E.; Harrison, G. A.; Turnbill, C.; Black, S.

    1977-01-01

    White albino rats were sacrificed after 24, 36, 48, 72, and 96 h of exposure to 100% O2 at 1 atm. Tissue was prepared for the scanning electron microscope (SEM) by Critical Point Drying and for the transmission electron microscope (TEM) by plastic embedding. Scanning microscopy showed a loss of microvilli after 48 h of exposure. Cilia appeared relatively normal with SEM, but TEM revealed changes in the outer membrane. In TEM, nonciliated cells appeared swollen and often encroached on the ciliated cells. A heavy mucous blanket remained even after processing. All the changes observed that are induced by oxygen exposure contribute to mucostasis, reducing and/or halting mucociliary clearance.

  17. Proliferation of human mammary cancer cells exposed to 27-hydroxycholesterol

    PubMed Central

    CRUZ, PAMELA; TORRES, CRISTIAN; RAMÍREZ, MARÍA EUGENIA; EPUÑÁN, MARÍA JOSÉ; VALLADARES, LUIS EMILIO; SIERRALTA, WALTER DANIEL

    2010-01-01

    The aim of the present study was to identify the possible mechanisms by which certain estradiol receptor (ER)-positive mammary tumor cells remain resistant to treatment with anti-estrogens or inhibitors of local estradiol (E2) production. To this end, we compared the proliferative effects on mammary cancer cells of the novel selective ER modulator 27-hydroxycholesterol (27OHC) to those of E2, and evaluated their inhibition by ICI 182,780 (ICI). Analysis of the effects on the cell cycle of 27OHC and E2 in the absence or presence of ICI was conducted. In ER-positive mammary tumor cells, we detected the blocking of 27OHC proliferation-stimulatory activity by simvastatin, as well as the inhibition of E2-stimulated proliferation by an α-fetoprotein-derived cyclic nonapeptide. The effects reported herein may be extrapolated to infiltrating mammary cancer, where the activity of local macrophages may stimulate tumor growth. We suggest that increased breast cancer growth in obese patients may be related to increased 27OHC circulatory levels. PMID:22993572

  18. Transplantation of erythropoietin gene-modified neural stem cells improves the repair of injured spinal cord

    PubMed Central

    Wu, Min-fei; Zhang, Shu-quan; Gu, Rui; Liu, Jia-bei; Li, Ye; Zhu, Qing-san

    2015-01-01

    The protective effects of erythropoietin on spinal cord injury have not been well described. Here, the eukaryotic expression plasmid pcDNA3.1 human erythropoietin was transfected into rat neural stem cells cultured in vitro. A rat model of spinal cord injury was established using a free falling object. In the human erythropoietin-neural stem cells group, transfected neural stem cells were injected into the rat subarachnoid cavity, while the neural stem cells group was injected with non-transfected neural stem cells. Dulbecco's modified Eagle's medium/F12 medium was injected into the rats in the spinal cord injury group as a control. At 1–4 weeks post injury, the motor function in the rat lower limbs was best in the human erythropoietin-neural stem cells group, followed by the neural stem cells group, and lastly the spinal cord injury group. At 72 hours, compared with the spinal cord injury group, the apoptotic index and Caspase-3 gene and protein expressions were apparently decreased, and the bcl-2 gene and protein expressions were noticeably increased, in the tissues surrounding the injured region in the human erythropoietin-neural stem cells group. At 4 weeks, the cavities were clearly smaller and the motor and somatosensory evoked potential latencies were remarkably shorter in the human erythropoietin-neural stem cells group and neural stem cells group than those in the spinal cord injury group. These differences were particularly obvious in the human erythropoietin-neural stem cells group. More CM-Dil-positive cells and horseradish peroxidase-positive nerve fibers and larger amplitude motor and somatosensory evoked potentials were found in the human erythropoietin-neural stem cells group and neural stem cells group than in the spinal cord injury group. Again, these differences were particularly obvious in the human erythropoietin-neural stem cells group. These data indicate that transplantation of erythropoietin gene-modified neural stem cells into the

  19. Transplantation of erythropoietin gene-modified neural stem cells improves the repair of injured spinal cord.

    PubMed

    Wu, Min-Fei; Zhang, Shu-Quan; Gu, Rui; Liu, Jia-Bei; Li, Ye; Zhu, Qing-San

    2015-09-01

    The protective effects of erythropoietin on spinal cord injury have not been well described. Here, the eukaryotic expression plasmid pcDNA3.1 human erythropoietin was transfected into rat neural stem cells cultured in vitro. A rat model of spinal cord injury was established using a free falling object. In the human erythropoietin-neural stem cells group, transfected neural stem cells were injected into the rat subarachnoid cavity, while the neural stem cells group was injected with non-transfected neural stem cells. Dulbecco's modified Eagle's medium/F12 medium was injected into the rats in the spinal cord injury group as a control. At 1-4 weeks post injury, the motor function in the rat lower limbs was best in the human erythropoietin-neural stem cells group, followed by the neural stem cells group, and lastly the spinal cord injury group. At 72 hours, compared with the spinal cord injury group, the apoptotic index and Caspase-3 gene and protein expressions were apparently decreased, and the bcl-2 gene and protein expressions were noticeably increased, in the tissues surrounding the injured region in the human erythropoietin-neural stem cells group. At 4 weeks, the cavities were clearly smaller and the motor and somatosensory evoked potential latencies were remarkably shorter in the human erythropoietin-neural stem cells group and neural stem cells group than those in the spinal cord injury group. These differences were particularly obvious in the human erythropoietin-neural stem cells group. More CM-Dil-positive cells and horseradish peroxidase-positive nerve fibers and larger amplitude motor and somatosensory evoked potentials were found in the human erythropoietin-neural stem cells group and neural stem cells group than in the spinal cord injury group. Again, these differences were particularly obvious in the human erythropoietin-neural stem cells group. These data indicate that transplantation of erythropoietin gene-modified neural stem cells into the

  20. Leader Cells Define Directionality of Trunk, but Not Cranial, Neural Crest Cell Migration.

    PubMed

    Richardson, Jo; Gauert, Anton; Briones Montecinos, Luis; Fanlo, Lucía; Alhashem, Zainalabdeen Mohmammed; Assar, Rodrigo; Marti, Elisa; Kabla, Alexandre; Härtel, Steffen; Linker, Claudia

    2016-05-31

    Collective cell migration is fundamental for life and a hallmark of cancer. Neural crest (NC) cells migrate collectively, but the mechanisms governing this process remain controversial. Previous analyses in Xenopus indicate that cranial NC (CNC) cells are a homogeneous population relying on cell-cell interactions for directional migration, while chick embryo analyses suggest a heterogeneous population with leader cells instructing directionality. Our data in chick and zebrafish embryos show that CNC cells do not require leader cells for migration and all cells present similar migratory capacities. In contrast, laser ablation of trunk NC (TNC) cells shows that leader cells direct movement and cell-cell contacts are required for migration. Moreover, leader and follower identities are acquired before the initiation of migration and remain fixed thereafter. Thus, two distinct mechanisms establish the directionality of CNC cells and TNC cells. This implies the existence of multiple molecular mechanisms for collective cell migration.

  1. Correlation between dielectric property by dielectrophoretic levitation and growth activity of cells exposed to electric field.

    PubMed

    Hakoda, Masaru; Hirota, Yusuke

    2013-09-01

    The purpose of this study is to develop a system analyzing cell activity by the dielectrophoresis method. Our previous studies revealed a correlation between the growth activity and dielectric property (Re[K(ω)]) of mouse hybridoma 3-2H3 cells using dielectrophoretic levitation. Furthermore, it was clarified that the differentiation activity of many stem cells could be evaluated by the Re[K(ω)] without differentiation induction. In this paper, 3-2H3 cells exposed to an alternating current (AC) electric field or a direct current (DC) electric field were cultivated, and the influence of damage by the electric field on the growth activity of the cells was examined. To evaluate the activity of the cells by measuring the Re[K(ω)], the correlation between the growth activity and the Re[K(ω)] of the cells exposed to the electric field was examined. The relations between the cell viability, growth activity, and Re[K(ω)] in the cells exposed to the AC electric field were obtained. The growth activity of the cells exposed to the AC electric field could be evaluated by the Re[K(ω)]. Furthermore, it was found that the adverse effects of the electric field on the cell viability and the growth activity were smaller in the AC electric field than the DC electric field.

  2. 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

  3. [The Evolutionary Origin of Placodes and Neural Crest Cells

    NASA Technical Reports Server (NTRS)

    Bronner-Fraser, Marianne

    2003-01-01

    The long-term goal of this NASA-supported research is to understand the evolutionary origin of placodes and neural crest cells, with particular reference to evolution of the inner ear, and their evolutionary and developmental relationships. The cephalochordcate amphioxus, the closest living invertebrate relative of the vertebrates is used as a stand-in for the ancestral vertebrate. The research, which has supported one graduate student, Jr-Kai Yu, has resulted in ten publications by the Holland laboratory in peer-reviewed journals.

  4. History of Neural Stem Cell Research and Its Clinical Application.

    PubMed

    Takagi, Yasushi

    2016-01-01

    "Once development was ended…in the adult centers, the nerve paths are something fixed and immutable. Everything may die, nothing may be regenerated," wrote Santiago Ramón y Cajal, a Spanish neuroanatomist and Nobel Prize winner and the father of modern neuroscience. This statement was the central dogma in neuroscience for a long time. However, in the 1960s, neural stem cells (NSCs) were discovered. Since then, our knowledge about NSCs has continued to grow. This review focuses on our current knowledge about NSCs and their surrounding microenvironment. In addition, the clinical application of NSCs for the treatment of various central nervous system diseases is also summarized. PMID:26888043

  5. 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

  6. Involvement of seven in absentia homolog-1 in ethanol-induced apoptosis in neural crest cells

    PubMed Central

    Sun, Haijing; Chen, Xiaopan; Yuan, Fuqiang; Liu, Jie; Zhao, Yingming; Chen, Shao-yu

    2014-01-01

    Ethanol-induced apoptosis in selected cell populations is a major component of pathogenesis underlying ethanol-induced teratogenesis. However, there is a fundamental gap in understanding how ethanol leads to apoptosis in embryos. In this study, we investigate the role of seven in absentia homolog-1 (Siah1) protein, an E3 ubiquitin ligase, in ethanol-induced apoptosis. Using an in vitro model of neural crest cell (NCC), JoMa1.3 cells, we found that exposure to 100 mM ethanol resulted in a significant increase in Siah1 mRNA expression in NCCs, an ethanol-sensitive cell population implicated in Fetal Alcohol Spectrum Disorders (FASD). Treatment with 100 mM ethanol for 24 hours also significantly increased the protein expression of Siah1 in JoMa1.3 cells. The nuclear translocation and accumulation of Siah1 was evidenced in the cells exposed to ethanol. In addition, we have found that the inhibition of Siah1 function with siRNA prevents ethanol-induced increase in Siah1 protein expression and nuclear translocation in NCCs. Down-regulation of Siah1 by siRNA also greatly diminished ethanol-induced cell death and caspase-3 activation, indicating that inhibition of Siah1 can attenuate ethanol-induced apoptosis. These results strongly suggest that Siah1 plays an important role in ethanol-induced apoptosis in NCCs. PMID:25193017

  7. How Tissue Mechanical Properties Affect Enteric Neural Crest Cell Migration

    NASA Astrophysics Data System (ADS)

    Chevalier, N. R.; Gazguez, E.; Bidault, L.; Guilbert, T.; Vias, C.; Vian, E.; Watanabe, Y.; Muller, L.; Germain, S.; Bondurand, N.; Dufour, S.; Fleury, V.

    2016-02-01

    Neural crest cells (NCCs) are a population of multipotent cells that migrate extensively during vertebrate development. Alterations to neural crest ontogenesis cause several diseases, including cancers and congenital defects, such as Hirschprung disease, which results from incomplete colonization of the colon by enteric NCCs (ENCCs). We investigated the influence of the stiffness and structure of the environment on ENCC migration in vitro and during colonization of the gastrointestinal tract in chicken and mouse embryos. We showed using tensile stretching and atomic force microscopy (AFM) that the mesenchyme of the gut was initially soft but gradually stiffened during the period of ENCC colonization. Second-harmonic generation (SHG) microscopy revealed that this stiffening was associated with a gradual organization and enrichment of collagen fibers in the developing gut. Ex-vivo 2D cell migration assays showed that ENCCs migrated on substrates with very low levels of stiffness. In 3D collagen gels, the speed of the ENCC migratory front decreased with increasing gel stiffness, whereas no correlation was found between porosity and ENCC migration behavior. Metalloprotease inhibition experiments showed that ENCCs actively degraded collagen in order to progress. These results shed light on the role of the mechanical properties of tissues in ENCC migration during development.

  8. Dscam-Mediated Cell Recognition Regulates Neural Circuit Formation

    PubMed Central

    Hattori, Daisuke; Millard, S. Sean; Wojtowicz, Woj M.; Zipursky, S. Lawrence

    2009-01-01

    The Dscam family of immunoglobulin cell surface proteins mediates recognition events between neurons that play an essential role in the establishment of neural circuits. The Drosophila Dscam1 locus encodes tens of thousands of cell surface proteins via alternative splicing. These isoforms exhibit exquisite isoform-specific binding in vitro that mediates homophilic repulsion in vivo. These properties provide the molecular basis for self-avoidance, an essential developmental mechanism that allows axonal and dendritic processes to uniformly cover their synaptic fields. In a mechanistically similar fashion, homophilic repulsion mediated by Drosophila Dscam2 prevents processes from the same class of cells from occupying overlapping synaptic fields through a process called tiling. Genetic studies in the mouse visual system support the view that vertebrate DSCAM also promotes both self-avoidance and tiling. By contrast, DSCAM and DSCAM-L promote layer-specific targeting in the chick visual system, presumably through promoting homophilic adhesion. The fly and mouse studies underscore the importance of homophilic repulsion in regulating neural circuit assembly, whereas the chick studies suggest that DSCA Mproteins may mediate a variety of different recognition events during wiring in a context-dependent fashion. PMID:18837673

  9. How Tissue Mechanical Properties Affect Enteric Neural Crest Cell Migration

    PubMed Central

    Chevalier, N.R.; Gazguez, E.; Bidault, L.; Guilbert, T.; Vias, C.; Vian, E.; Watanabe, Y.; Muller, L.; Germain, S.; Bondurand, N.; Dufour, S.; Fleury, V.

    2016-01-01

    Neural crest cells (NCCs) are a population of multipotent cells that migrate extensively during vertebrate development. Alterations to neural crest ontogenesis cause several diseases, including cancers and congenital defects, such as Hirschprung disease, which results from incomplete colonization of the colon by enteric NCCs (ENCCs). We investigated the influence of the stiffness and structure of the environment on ENCC migration in vitro and during colonization of the gastrointestinal tract in chicken and mouse embryos. We showed using tensile stretching and atomic force microscopy (AFM) that the mesenchyme of the gut was initially soft but gradually stiffened during the period of ENCC colonization. Second-harmonic generation (SHG) microscopy revealed that this stiffening was associated with a gradual organization and enrichment of collagen fibers in the developing gut. Ex-vivo 2D cell migration assays showed that ENCCs migrated on substrates with very low levels of stiffness. In 3D collagen gels, the speed of the ENCC migratory front decreased with increasing gel stiffness, whereas no correlation was found between porosity and ENCC migration behavior. Metalloprotease inhibition experiments showed that ENCCs actively degraded collagen in order to progress. These results shed light on the role of the mechanical properties of tissues in ENCC migration during development. PMID:26887292

  10. Functional 3D Neural Mini-Tissues from Printed Gel-Based Bioink and Human Neural Stem Cells.

    PubMed

    Gu, Qi; Tomaskovic-Crook, Eva; Lozano, Rodrigo; Chen, Yu; Kapsa, Robert M; Zhou, Qi; Wallace, Gordon G; Crook, Jeremy M

    2016-06-01

    Direct-write printing of stem cells within biomaterials presents an opportunity to engineer tissue for in vitro modeling and regenerative medicine. Here, a first example of constructing neural tissue by printing human neural stem cells that are differentiated in situ to functional neurons and supporting neuroglia is reported. The supporting biomaterial incorporates a novel clinically relevant polysaccharide-based bioink comprising alginate, carboxymethyl-chitosan, and agarose. The printed bioink rapidly gels by stable cross-linking to form a porous 3D scaffold encapsulating stem cells for in situ expansion and differentiation. Differentiated neurons form synaptic contacts, establish networks, are spontaneously active, show a bicuculline-induced increased calcium response, and are predominantly gamma-aminobutyric acid expressing. The 3D tissues will facilitate investigation of human neural development, function, and disease, and may be adaptable for engineering other 3D tissues from different stem cell types. PMID:27028356

  11. Functional 3D Neural Mini-Tissues from Printed Gel-Based Bioink and Human Neural Stem Cells.

    PubMed

    Gu, Qi; Tomaskovic-Crook, Eva; Lozano, Rodrigo; Chen, Yu; Kapsa, Robert M; Zhou, Qi; Wallace, Gordon G; Crook, Jeremy M

    2016-06-01

    Direct-write printing of stem cells within biomaterials presents an opportunity to engineer tissue for in vitro modeling and regenerative medicine. Here, a first example of constructing neural tissue by printing human neural stem cells that are differentiated in situ to functional neurons and supporting neuroglia is reported. The supporting biomaterial incorporates a novel clinically relevant polysaccharide-based bioink comprising alginate, carboxymethyl-chitosan, and agarose. The printed bioink rapidly gels by stable cross-linking to form a porous 3D scaffold encapsulating stem cells for in situ expansion and differentiation. Differentiated neurons form synaptic contacts, establish networks, are spontaneously active, show a bicuculline-induced increased calcium response, and are predominantly gamma-aminobutyric acid expressing. The 3D tissues will facilitate investigation of human neural development, function, and disease, and may be adaptable for engineering other 3D tissues from different stem cell types.

  12. Role of endogenous neural stem cells in spinal cord injury and repair.

    PubMed

    Stenudd, Moa; Sabelström, Hanna; Frisén, Jonas

    2015-02-01

    Spinal cord injury is followed by glial scar formation, which has positive and negative effects on recovery from the lesion. More than half of the astrocytes in the glial scar are generated by ependymal cells, the neural stem cells in the spinal cord. We recently demonstrated that the neural stem cell-derived scar component has several beneficial functions, including restricting tissue damage and neural loss after spinal cord injury. This finding identifies endogenous neural stem cells as a potential therapeutic target for treatment of spinal cord injury.

  13. Biology of the adult enteric neural stem cell.

    PubMed

    Estrada-Mondaca, Sandino; Carreón-Rodríguez, Alfonso; Belkind-Gerson, Jaime

    2007-01-01

    An increasing body of evidence has accumulated in recent years supporting the existence of neural stem cells in the adult gut. There are at least three groups that have obtained them using different methodologies and have described them in vitro. There is a growing amount of knowledge on their biology, but many questions are yet unanswered. Among these questions is whether these cells are part of a permanent undifferentiated pool or are recruited in a regular basis; in addition, the factors and genes involved in their survival, proliferation, migration, and differentiation are largely unknown. Finally, with between 10 and 20% of adults suffering from diseases involving the enteric nervous system, most notably irritable bowel syndrome and gastroesophageal reflux, what is the possible role of enteric nervous stem cells in health and disease?

  14. Alcohol-Induced Molecular Dysregulation in Human Embryonic Stem Cell-Derived Neural Precursor Cells

    PubMed Central

    Kim, Yi Young; Roubal, Ivan; Lee, Youn Soo; Kim, Jin Seok; Hoang, Michael; Mathiyakom, Nathan; Kim, Yong

    2016-01-01

    Adverse effect of alcohol on neural function has been well documented. Especially, the teratogenic effect of alcohol on neurodevelopment during embryogenesis has been demonstrated in various models, which could be a pathologic basis for fetal alcohol spectrum disorders (FASDs). While the developmental defects from alcohol abuse during gestation have been described, the specific mechanisms by which alcohol mediates these injuries have yet to be determined. Recent studies have shown that alcohol has significant effect on molecular and cellular regulatory mechanisms in embryonic stem cell (ESC) differentiation including genes involved in neural development. To test our hypothesis that alcohol induces molecular alterations during neural differentiation we have derived neural precursor cells from pluripotent human ESCs in the presence or absence of ethanol treatment. Genome-wide transcriptomic profiling identified molecular alterations induced by ethanol exposure during neural differentiation of hESCs into neural rosettes and neural precursor cell populations. The Database for Annotation, Visualization and Integrated Discovery (DAVID) functional analysis on significantly altered genes showed potential ethanol’s effect on JAK-STAT signaling pathway, neuroactive ligand-receptor interaction, Toll-like receptor (TLR) signaling pathway, cytokine-cytokine receptor interaction and regulation of autophagy. We have further quantitatively verified ethanol-induced alterations of selected candidate genes. Among verified genes we further examined the expression of P2RX3, which is associated with nociception, a peripheral pain response. We found ethanol significantly reduced the level of P2RX3 in undifferentiated hESCs, but induced the level of P2RX3 mRNA and protein in hESC-derived NPCs. Our result suggests ethanol-induced dysregulation of P2RX3 along with alterations in molecules involved in neural activity such as neuroactive ligand-receptor interaction may be a molecular event

  15. OXIDATIVE STRESS INDUCES CELL DEATH IN CD-1 MOUSE CRANIAL NEURAL CREST CELLS IN VITRO

    EPA Science Inventory

    OXIDATIVE STRESS INDUCES CELL DEATH IN CD-1 MOUSE CRANIAL NEURAL CREST CELLS IN VITRO. J.B. Smith, K.K. Sulik, E.S. Hunter III. University of North Carolina at Chapel Hill, Chapel Hill, NC 27599.
    The induction of craniofacial defects by ethanol exposure is mediated in part by...

  16. The Hippo pathway member YAP enhances human neural crest cell fate and migration

    PubMed Central

    Hindley, Christopher J.; Condurat, Alexandra Larisa; Menon, Vishal; Thomas, Ria; Azmitia, Luis M.; Davis, Jason A.; Pruszak, Jan

    2016-01-01

    The Hippo/YAP pathway serves as a major integrator of cell surface-mediated signals and regulates key processes during development and tumorigenesis. The neural crest is an embryonic tissue known to respond to multiple environmental cues in order to acquire appropriate cell fate and migration properties. Using multiple in vitro models of human neural development (pluripotent stem cell-derived neural stem cells; LUHMES, NTERA2 and SH-SY5Y cell lines), we investigated the role of Hippo/YAP signaling in neural differentiation and neural crest development. We report that the activity of YAP promotes an early neural crest phenotype and migration, and provide the first evidence for an interaction between Hippo/YAP and retinoic acid signaling in this system. PMID:26980066

  17. Premigratory and migratory neural crest cells are multipotent in vivo.

    PubMed

    Baggiolini, Arianna; Varum, Sandra; Mateos, José María; Bettosini, Damiano; John, Nessy; Bonalli, Mario; Ziegler, Urs; Dimou, Leda; Clevers, Hans; Furrer, Reinhard; Sommer, Lukas

    2015-03-01

    The neural crest (NC) is an embryonic stem/progenitor cell population that generates a diverse array of cell lineages, including peripheral neurons, myelinating Schwann cells, and melanocytes, among others. However, there is a long-standing controversy as to whether this broad developmental perspective reflects in vivo multipotency of individual NC cells or whether the NC is comprised of a heterogeneous mixture of lineage-restricted progenitors. Here, we resolve this controversy by performing in vivo fate mapping of single trunk NC cells both at premigratory and migratory stages using the R26R-Confetti mouse model. By combining quantitative clonal analyses with definitive markers of differentiation, we demonstrate that the vast majority of individual NC cells are multipotent, with only few clones contributing to single derivatives. Intriguingly, multipotency is maintained in migratory NC cells. Thus, our findings provide definitive evidence for the in vivo multipotency of both premigratory and migrating NC cells in the mouse. PMID:25748934

  18. A requirement for trypsin-sensitive cell-surface components for cell-cell interactions of embryonic neural retina cells

    PubMed Central

    McClay, DR; Godding, LR; Fransen, ME

    1977-01-01

    A quantitative assay was used to measure the rate of collection of a population of embryonic neural retina cells to the surface of cell aggregates. The rate of collection of freshly trysinized cells was limited in the initial stages by the rate of replacement of trypsin-sensitive cell- surface components. When cells were preincubated, or "recovered," and then added to cell aggregates, collection occurred at a linear rate and was independent of protein and glycoprotein synthesis. The adhesion of recovered cells was temperature and energy dependent, and was reversibly inhibited by cytochalasin B. Colchicine had little effect on collection of recovered cells. Antiserum directed against recovered cell membranes was shown to bind to recovered cells by indirect immunofluorescence. The antiserum also was shown to inhibit collection of recovered cells to aggregates, suggesting that at least some of the antigens identified might be involved in the adhesion process. The inhibitory effect of the antiserum was dose dependent . Freshly trypsinized cells absorbed neither the immunofluorescence activity nor the adhesion-inhibiting activity. Recovered cells absorbed away both activities. In specificity studies, dorsal neural retina cells adhered to aggregates of ventral optic tectum in preference to aggregates of dorsal optic tectum. The adhesive specificity of the dorsal retina cells was less sensitive to trypsin than the adhesive specificity of ventral retina cells which adhered preferentially to dorsal tectal aggregates only after a period of recovery. PMID:562349

  19. Neural stem cells induce bone-marrow-derived mesenchymal stem cells to generate neural stem-like cells via juxtacrine and paracrine interactions

    SciTech Connect

    Alexanian, Arshak R. . E-mail: aalexan@mcw.edu

    2005-11-01

    Several recent reports suggest that there is far more plasticity that previously believed in the developmental potential of bone-marrow-derived cells (BMCs) that can be induced by extracellular developmental signals of other lineages whose nature is still largely unknown. In this study, we demonstrate that bone-marrow-derived mesenchymal stem cells (MSCs) co-cultured with mouse proliferating or fixed (by paraformaldehyde or methanol) neural stem cells (NSCs) generate neural stem cell-like cells with a higher expression of Sox-2 and nestin when grown in NS-A medium supplemented with N2, NSC conditioned medium (NSCcm) and bFGF. These neurally induced MSCs eventually differentiate into {beta}-III-tubulin and GFAP expressing cells with neuronal and glial morphology when grown an additional week in Neurobasal/B27 without bFGF. We conclude that juxtacrine interaction between NSCs and MSCs combined with soluble factors released from NSCs are important for generation of neural-like cells from bone-marrow-derived adherent MSCs.

  20. Oligodendrogenesis from neural stem cells: perspectives for remyelinating strategies.

    PubMed

    Grade, Sofia; Bernardino, Liliana; Malva, João O

    2013-11-01

    Mobilization of remyelinating cells spontaneously occurs in the adult brain. These cellular resources are specially active after demyelinating episodes in early phases of multiple sclerosis (MS). Indeed, oligodendrocyte precursor cells (OPCs) actively proliferate, migrate to and repopulate the lesioned areas. Ultimately, efficient remyelination is accomplished when new oligodendrocytes reinvest nude neuronal axons, restoring the normal properties of impulse conduction. As the disease progresses this fundamental process fails. Multiple causes seem to contribute to such transient decline, including the failure of OPCs to differentiate and enwrap the vulnerable neuronal axons. Regenerative medicine for MS has been mainly centered on the recruitment of endogenous self-repair mechanisms, or on transplantation approaches. The latter commonly involves grafting of neural precursor cells (NPCs) or neural stem cells (NSCs), with myelinogenic potential, in the injured areas. Both strategies require further understanding of the biology of oligodendrocyte differentiation and remyelination. Indeed, the success of transplantation largely depends on the pre-commitment of transplanted NPCs or NSCs into oligodendroglial cell type, while the endogenous differentiation of OPCs needs to be boosted in chronic stages of the disease. Thus, much effort has been focused on finding molecular targets that drive oligodendrocytes commitment and development. The present review explores several aspects of remyelination that must be considered in the design of a cell-based therapy for MS, and explores more deeply the challenge of fostering oligodendrogenesis. In this regard, we discuss herein a tool developed in our research group useful to search novel oligodendrogenic factors and to study oligodendrocyte differentiation in a time- and cost-saving manner.

  1. Autosomal mutants of proton-exposed kidney cells display frequent loss of heterozygosity on nonselected chromosomes.

    PubMed

    Grygoryev, Dmytro; Dan, Cristian; Gauny, Stacey; Eckelmann, Bradley; Ohlrich, Anna P; Connolly, Marissa; Lasarev, Michael; Grossi, Gianfranco; Kronenberg, Amy; Turker, Mitchell S

    2014-05-01

    High-energy protons found in the space environment can induce mutations and cancer, which are inextricably linked. We hypothesized that some mutants isolated from proton-exposed kidneys arose through a genome-wide incident that causes loss of heterozygosity (LOH)-generating mutations on multiple chromosomes (termed here genomic LOH). To test this hypothesis, we examined 11 pairs of nonselected chromosomes for LOH events in mutant cells isolated from the kidneys of mice exposed to 4 or 5 Gy of 1 GeV protons. The mutant kidney cells were selected for loss of expression of the chromosome 8-encoded Aprt gene. Genomic LOH events were also assessed in Aprt mutants isolated from isogenic cultured kidney epithelial cells exposed to 5 Gy of protons in vitro. Control groups were spontaneous Aprt mutants and clones isolated without selection from the proton-exposed kidneys or cultures. The in vivo results showed significant increases in genomic LOH events in the Aprt mutants from proton-exposed kidneys when compared with spontaneous Aprt mutants and when compared with nonmutant (i.e., nonselected) clones from the proton-exposed kidneys. A bias for LOH events affecting chromosome 14 was observed in the proton-induced Aprt mutants, though LOH for this chromosome did not confer increased radiation resistance. Genomic LOH events were observed in Aprt mutants isolated from proton-exposed cultured kidney cells; however the incidence was fivefold lower than in Aprt mutants isolated from exposed intact kidneys, suggesting a more permissive environment in the intact organ and/or the evolution of kidney clones prior to their isolation from the tissue. We conclude that proton exposure creates a subset of viable cells with LOH events on multiple chromosomes, that these cells form and persist in vivo, and that they can be isolated from an intact tissue by selection for a mutation on a single chromosome.

  2. Comparative Mitochondrial Proteomic Analysis of Raji Cells Exposed to Adriamycin

    PubMed Central

    Jiang, Yu-Jie; Sun, Qing; Fang, Xiao-Sheng; Wang, Xin

    2009-01-01

    The antitumor mechanisms of adriamycin (ADR) have been thought to contribute to induction of apoptosis and inefficiency of DNA repair, processes that are to a large extent mediated by mitochondria. This study aimed to investigate characteristics of ADR, including its antineoplastic activity, drug resistance, and unexpected toxicity in non-Hodgkin lymphoma (NHL) Raji cells at the mitochondrial proteomic level. The alterations of the mitochondrial proteome of Raji cells treated with ADR were analyzed by two-dimensional differential in-gel electrophoresis (2D-DIGE) coupled with linear ion trap quadrupole–electrospray ionization tandem mass spectrometry (LTQ-ESI-MS/MS).The altered patterns of three identified proteins were validated by Western blot and analyzed by pathway studio software. The results showed that 34 proteins were downregulated and 3 proteins upregulated in the study group compared with the control group. The differentially expressed proteins distributed their functions in reduction-oxidation reactions, DNA repair, cell cycle regulation, transporters and channels, and oxidative phosphorylation. Furthermore, heat shock protein 70 (HSP70), ATP-binding cassette transporter isoform B6 (ABCB6), and prohibitin (PHB) identified in this study may be closely related to chemoresistance and could serve as potential chemotherapeutic targets for NHL. Collectively, these results suggest that specific mitochondrial proteins are uniquely susceptible to alterations in abundance following exposure to ADR and carry implications for the investigation of therapeutic and prognostic markers. Further studies focusing on these identified proteins will be used to predict treatment response and reverse apoptosis resistance,and to explore drug-combination strategies associated with ADR for NHL therapy. PMID:19209238

  3. Neogenin and RGMa control neural tube closure and neuroepithelial morphology by regulating cell polarity.

    PubMed

    Kee, Nigel; Wilson, Nicole; De Vries, Melissa; Bradford, DanaKai; Key, Brian; Cooper, Helen M

    2008-11-26

    In humans, neural tube closure defects occur in 1:1000 pregnancies. The design of new strategies for the prevention of such common defects would benefit from an improved understanding of the molecular events underlying neurulation. Neural fold elevation is a key morphological process that acts during neurulation to drive neural tube closure. However, to date, the molecular pathways underpinning neural fold elevation have not been elucidated. Here, we use morpholino knock-down technology to demonstrate that Repulsive Guidance Molecule (RGMa)-Neogenin interactions are essential for effective neural fold elevation during Xenopus neurulation and that loss of these molecules results in disrupted neural tube closure. We demonstrate that Neogenin and RGMa are required for establishing the morphology of deep layer cells in the neural plate throughout neurulation. We also show that loss of Neogenin severely disrupts the microtubule network within the deep layer cells suggesting that Neogenin-dependent microtubule organization within the deep cells is essential for radial intercalation with the overlying superficial cell layer, thereby driving neural fold elevation. In addition, we show that sustained Neogenin activity is also necessary for the establishment of the apicobasally polarized pseudostratified neuroepithelium of the neural tube. Therefore, our study identifies a novel signaling pathway essential for radial intercalation and epithelialization during neural fold elevation and neural tube morphogenesis.

  4. Differential gene expression in pulmonary artery endothelial cells exposed to sickle cell plasma.

    PubMed

    Klings, Elizabeth S; Safaya, Surinder; Adewoye, Adeboye H; Odhiambo, Adam; Frampton, Garrett; Lenburg, Marc; Gerry, Norman; Sebastiani, Paola; Steinberg, Martin H; Farber, Harrison W

    2005-05-11

    Clinical variability in sickle cell disease (SCD) suggests a role for extra-erythrocytic factors in the pathogenesis of vasoocclusion. We hypothesized that endothelial cell (EC) dysfunction, one possible modifier of disease variability, results from induction of phenotypic changes by circulating factors. Accordingly, we analyzed gene expression in cultured human pulmonary artery ECs (HPAEC) exposed to plasma from 1) sickle acute chest syndrome (ACS) patients, 2) SCD patients at steady state, 3) normal volunteers, and 4) serum-free media, using whole genome microarrays (U133A-B GeneChip, Affymetrix). Data were analyzed by Bayesian analysis of differential gene expression (BADGE). Differential expression was defined by the probability of >1.5 fold change in signal intensity greater than 0.999 and a predicted score of 70-100, measured by cross-validation. Compared with normal plasma, plasma from SCD patients (steady state) resulted in differential expression of 50 genes in HPAEC. Of these genes, molecules involved in cholesterol biosynthesis and lipid transport, the cellular stress response, and extracellular matrix proteins were most prominent. Another 58 genes were differentially expressed in HPAEC exposed to plasma from ACS patients. The pattern of altered gene expression suggests that plasma from SCD patients induces an EC phenotype which is anti-apoptotic and favors cholesterol biosynthesis. An altered EC phenotype elicited by SCD plasma may contribute to the pathogenesis of sickle vasoocclusion.

  5. Applications of Mesenchymal Stem Cells and Neural Crest Cells in Craniofacial Skeletal Research

    PubMed Central

    Ouchi, Takehito; Shibata, Shinsuke; Fujimura, Takumi; Kawana, Hiromasa; Okano, Hideyuki; Nakagawa, Taneaki

    2016-01-01

    Craniofacial skeletal tissues are composed of tooth and bone, together with nerves and blood vessels. This composite material is mainly derived from neural crest cells (NCCs). The neural crest is transient embryonic tissue present during neural tube formation whose cells have high potential for migration and differentiation. Thus, NCCs are promising candidates for craniofacial tissue regeneration; however, the clinical application of NCCs is hindered by their limited accessibility. In contrast, mesenchymal stem cells (MSCs) are easily accessible in adults, have similar potential for self-renewal, and can differentiate into skeletal tissues, including bones and cartilage. Therefore, MSCs may represent good sources of stem cells for clinical use. MSCs are classically identified under adherent culture conditions, leading to contamination with other cell lineages. Previous studies have identified mouse- and human-specific MSC subsets using cell surface markers. Additionally, some studies have shown that a subset of MSCs is closely related to neural crest derivatives and endothelial cells. These MSCs may be promising candidates for regeneration of craniofacial tissues from the perspective of developmental fate. Here, we review the fundamental biology of MSCs in craniofacial research. PMID:27006661

  6. Neural stem cells and cell replacement therapy: making the right cells.

    PubMed

    Bithell, Angela; Williams, Brenda P

    2005-01-01

    The past few years have seen major advances in the field of NSC (neural stem cell) research with increasing emphasis towards its application in cell-replacement therapy for neurological disorders. However, the clinical application of NSCs will remain largely unfeasible until a comprehensive understanding of the cellular and molecular mechanisms of NSC fate specification is achieved. With this understanding will come an increased possibility to exploit the potential of stem cells in order to manufacture transplantable NSCs able to provide a safe and effective therapy for previously untreatable neurological disorders. Since the pathology of each of these disorders is determined by the loss or damage of a specific neural cell population, it may be necessary to generate a range of NSCs able to replace specific neurons or glia rather than generating a generic NSC population. Currently, a diverse range of strategies is being investigated with this goal in mind. In this review, we focus on the relationship between NSC specification and differentiation and discuss how this information may be used to direct NSCs towards a particular fate.

  7. Effects of Triclosan on Neural Stem Cell Viability and Survival.

    PubMed

    Park, Bo Kyung; Gonzales, Edson Luck T; Yang, Sung Min; Bang, Minji; Choi, Chang Soon; Shin, Chan Young

    2016-01-01

    Triclosan is an antimicrobial or sanitizing agent used in personal care and household products such as toothpaste, soaps, mouthwashes and kitchen utensils. There are increasing evidence of the potentially harmful effects of triclosan in many systemic and cellular processes of the body. In this study, we investigated the effects of triclosan in the survivability of cultured rat neural stem cells (NSCs). Cortical cells from embryonic day 14 rat embryos were isolated and cultured in vitro. After stabilizing the culture, triclosan was introduced to the cells with concentrations ranging from 1 μM to 50 μM and in varied time periods. Thereafter, cell viability parameters were measured using MTT assay and PI staining. TCS decreased the cell viability of treated NSC in a concentration-dependent manner along with increased expressions of apoptotic markers, cleaved caspase-3 and Bax, while reduced expression of Bcl2. To explore the mechanisms underlying the effects of TCS in NSC, we measured the activation of MAPKs and intracellular ROS. TCS at 50 μM induced the activations of both p38 and JNK, which may adversely affect cell survival. In contrast, the activities of ERK, Akt and PI3K, which are positively correlated with cell survival, were inhibited. Moreover, TCS at this concentration augmented the ROS generation in treated NSC and depleted the glutathione activity. Taken together, these results suggest that TCS can induce neurodegenerative effects in developing rat brains through mechanisms involving ROS activation and apoptosis initiation. PMID:26759708

  8. Effects of Triclosan on Neural Stem Cell Viability and Survival

    PubMed Central

    Park, Bo Kyung; Gonzales, Edson Luck T.; Yang, Sung Min; Bang, Minji; Choi, Chang Soon; Shin, Chan Young

    2016-01-01

    Triclosan is an antimicrobial or sanitizing agent used in personal care and household products such as toothpaste, soaps, mouthwashes and kitchen utensils. There are increasing evidence of the potentially harmful effects of triclosan in many systemic and cellular processes of the body. In this study, we investigated the effects of triclosan in the survivability of cultured rat neural stem cells (NSCs). Cortical cells from embryonic day 14 rat embryos were isolated and cultured in vitro. After stabilizing the culture, triclosan was introduced to the cells with concentrations ranging from 1 μM to 50 μM and in varied time periods. Thereafter, cell viability parameters were measured using MTT assay and PI staining. TCS decreased the cell viability of treated NSC in a concentration-dependent manner along with increased expressions of apoptotic markers, cleaved caspase-3 and Bax, while reduced expression of Bcl2. To explore the mechanisms underlying the effects of TCS in NSC, we measured the activation of MAPKs and intracellular ROS. TCS at 50 μM induced the activations of both p38 and JNK, which may adversely affect cell survival. In contrast, the activities of ERK, Akt and PI3K, which are positively correlated with cell survival, were inhibited. Moreover, TCS at this concentration augmented the ROS generation in treated NSC and depleted the glutathione activity. Taken together, these results suggest that TCS can induce neurodegenerative effects in developing rat brains through mechanisms involving ROS activation and apoptosis initiation. PMID:26759708

  9. DNA Fragmentation in mammalian cells exposed to various light ions

    NASA Astrophysics Data System (ADS)

    Belli, M.; Cherubini, R.; Dalla Vecchia, M.; Dini, V.; Esposito, G.; Moschini, G.; Sapora, O.; Signoretti, C.; Simone, G.; Sorrentino, E.; Tabocchini, M. A.

    Elucidation of how effects of densely ionizing radiation at cellular level are linked to DNA damage is fundamental for a better understanding of the mechanisms leading to genomic damage (especially chromosome aberrations) and developing biophysical models to predict space radiation effects. We have investigated the DNA fragmentation patterns induced in Chinese hamster V79 cells by 31 keV/μm protons, 123 keV/μm helium-4 ions and γ-rays in the size range 0.023-5.7 Mbp, using calibrated Pulsed Field Gel Electrophoresis (PFGE). The frequency distributions of fragments induced by the charged particles were shifted towards smaller sizes with respct to that induced by comparable doses of γ-rays. The DSB yields, evaluated from the fragments induced in the size range studied, were higher for protons and helium ions than for γ-rays by a factor of about 1.9 and 1.2, respectively. However, these ratios do not adequately reflect the RBE observed on the same cells for inactivation and mutation induced by these beams. This is a further indication for the lack of correlation between the effects exerted at cellular level and the initial yield of DSB. The dependence on radiation quality of the fragmentation pattern suggests that it may have a role in damage reparability. We have analyzed these patterns with a "random breakage" model generalized in order to consider the initial non-random distribution of the DNA molecules. Our results suggest that a random breakage mechanism can describe with a reasonable approximation the DNA fragmentation induced by γ-rays, while the approximation is not so good for light ions, likely due to the interplay between ion tracks and chromatin organization at the loop level.

  10. YAP/TAZ enhance mammalian embryonic neural stem cell characteristics in a Tead-dependent manner

    SciTech Connect

    Han, Dasol; Byun, Sung-Hyun; Park, Soojeong; Kim, Juwan; Kim, Inhee; Ha, Soobong; Kwon, Mookwang; Yoon, Keejung

    2015-02-27

    Mammalian brain development is regulated by multiple signaling pathways controlling cell proliferation, migration and differentiation. Here we show that YAP/TAZ enhance embryonic neural stem cell characteristics in a cell autonomous fashion using diverse experimental approaches. Introduction of retroviral vectors expressing YAP or TAZ into the mouse embryonic brain induced cell localization in the ventricular zone (VZ), which is the embryonic neural stem cell niche. This change in cell distribution in the cortical layer is due to the increased stemness of infected cells; YAP-expressing cells were colabeled with Sox2, a neural stem cell marker, and YAP/TAZ increased the frequency and size of neurospheres, indicating enhanced self-renewal- and proliferative ability of neural stem cells. These effects appear to be TEA domain family transcription factor (Tead)–dependent; a Tead binding-defective YAP mutant lost the ability to promote neural stem cell characteristics. Consistently, in utero gene transfer of a constitutively active form of Tead2 (Tead2-VP16) recapitulated all the features of YAP/TAZ overexpression, and dominant negative Tead2-EnR resulted in marked cell exit from the VZ toward outer cortical layers. Taken together, these results indicate that the Tead-dependent YAP/TAZ signaling pathway plays important roles in neural stem cell maintenance by enhancing stemness of neural stem cells during mammalian brain development. - Highlights: • Roles of YAP and Tead in vivo during mammalian brain development are clarified. • Expression of YAP promotes embryonic neural stem cell characteristics in vivo in a cell autonomous fashion. • Enhancement of neural stem cell characteristics by YAP depends on Tead. • Transcriptionally active form of Tead alone can recapitulate the effects of YAP. • Transcriptionally repressive form of Tead severely reduces stem cell characteristics.

  11. Low-Dose Methylmercury-Induced Apoptosis and Mitochondrial DNA Mutation in Human Embryonic Neural Progenitor Cells.

    PubMed

    Wang, Xinjin; Yan, Mengling; Zhao, Lina; Wu, Qing; Wu, Chunhua; Chang, Xiuli; Zhou, Zhijun

    2016-01-01

    Methylmercury (MeHg) is a long-lasting organic pollutant primarily found in the aquatic environment. The developing brain is particularly sensitive to MeHg due to reduced proliferation of neural stem cell. Although several mechanisms of MeHg-induced apoptosis have been defined in culture models, it remains unclear whether mitochondrial DNA (mtDNA) mutation is involved in the toxic effect of MeHg, especially in the neural progenitor cells. In the present study, the ReNcell CX cell, a human neural progenitor cells (hNPCs) line, was exposed to nanomolar concentrations of MeHg (≤50 nM). We found that MeHg altered mitochondrial metabolic function and induced apoptosis. In addition, we observed that MeHg induced ROS production in a dose-dependent manner in hNPCs cells, which was associated with significantly increased expressions of ND1, Cytb, and ATP6. To elucidate the mechanism underlying MeHg toxicity on mitochondrial function, we examined the ATP content and mitochondrial membrane potential in MeHg-treated hNPCs. Our study showed that MeHg exposure led to decreased ATP content and reduced mitochondrial membrane potential, which failed to match the expansion in mtDNA copy number, suggesting impaired mtDNA. Collectively, these results demonstrated that MeHg induced toxicity in hNPCs through altering mitochondrial function and inducing oxidative damage to mtDNA. PMID:27525052

  12. Isolation and characterization of neural stem cells from the neonatal rat cochlear nucleus.

    PubMed

    Rak, Kristen; Wasielewski, Natalia V; Radeloff, Andreas; Völkers, Johannes; Scherzed, Agmal; Jablonka, Sibylle; Hagen, Rudolf; Mlynski, Robert

    2011-03-01

    Neural stem cells have been identified in multiple parts of the postnatal mammalian brain, as well as in the inner ear. No investigation of potential neural stem cells in the cochlear nucleus has yet been performed. The aim of this study was to investigate potential neural stem cells from the cochlear nucleus by neurosphere assay and in histological sections to prove their capacity for self-renewal and for differentiation into progenitor cells and cells of the neuronal lineage. For this purpose, cells of the cochlear nucleus of postnatal day 6 rats were isolated and cultured for generation of primary neurospheres. Spheres were dissociated and cells analyzed for capacity for mitosis and differentiation. Cell division was detected by cell-counting assay and BrdU incorporation. Differentiated neural progenitor cells showed distinct labeling for Nestin and for Atoh1. Positive staining of ß-III Tubulin, glial fibrillary acid protein (GFAP) and myelin basic protein (MBP) showed differentiation into neurons, astrocytes and oligodendrocytes. Furthermore, Nestin- and BrdU-labeled cells could also be detected in histological sections. In conclusion, the isolated cells from the cochlear nucleus presented all the features of neural stem cells: cell division, presence of progenitor cells and differentiation into different cells of the neuronal lineage. The existence of neural stem cells may add to the understanding of developmental features in the cochlear nucleus. PMID:21258945

  13. Activation of endogenous neural stem cells for multiple sclerosis therapy.

    PubMed

    Michailidou, Iliana; de Vries, Helga E; Hol, Elly M; van Strien, Miriam E

    2014-01-01

    Multiple sclerosis (MS) is a chronic inflammatory disorder of the central nervous system, leading to severe neurological deficits. Current MS treatment regimens, consist of immunomodulatory agents aiming to reduce the rate of relapses. However, these agents are usually insufficient to treat chronic neurological disability. A promising perspective for future therapy of MS is the regeneration of lesions with replacement of the damaged oligodendrocytes or neurons. Therapies targeting to the enhancement of endogenous remyelination, aim to promote the activation of either the parenchymal oligodendrocyte progenitor cells or the subventricular zone-derived neural stem cells (NSCs). Less studied but highly potent, is the strategy of neuronal regeneration with endogenous NSCs that although being linked to numerous limitations, is anticipated to ameliorate cognitive disability in MS. Focusing on the forebrain, this review highlights the role of NSCs in the regeneration of MS lesions.

  14. Activation of endogenous neural stem cells for multiple sclerosis therapy

    PubMed Central

    Michailidou, Iliana; de Vries, Helga E.; Hol, Elly M.; van Strien, Miriam E.

    2015-01-01

    Multiple sclerosis (MS) is a chronic inflammatory disorder of the central nervous system, leading to severe neurological deficits. Current MS treatment regimens, consist of immunomodulatory agents aiming to reduce the rate of relapses. However, these agents are usually insufficient to treat chronic neurological disability. A promising perspective for future therapy of MS is the regeneration of lesions with replacement of the damaged oligodendrocytes or neurons. Therapies targeting to the enhancement of endogenous remyelination, aim to promote the activation of either the parenchymal oligodendrocyte progenitor cells or the subventricular zone-derived neural stem cells (NSCs). Less studied but highly potent, is the strategy of neuronal regeneration with endogenous NSCs that although being linked to numerous limitations, is anticipated to ameliorate cognitive disability in MS. Focusing on the forebrain, this review highlights the role of NSCs in the regeneration of MS lesions. PMID:25653584

  15. Neural stem/progenitor cells in Alzheimer's disease.

    PubMed

    Tincer, Gizem; Mashkaryan, Violeta; Bhattarai, Prabesh; Kizil, Caghan

    2016-03-01

    Alzheimer's disease (AD) is the most prevalent neurodegenerative disease and a worldwide health challenge. Different therapeutic approaches are being developed to reverse or slow the loss of affected neurons. Another plausible therapeutic way that may complement the studies is to increase the survival of existing neurons by mobilizing the existing neural stem/progenitor cells (NSPCs) - i.e. "induce their plasticity" - to regenerate lost neurons despite the existing pathology and unfavorable environment. However, there is controversy about how NSPCs are affected by the unfavorable toxic environment during AD. In this review, we will discuss the use of stem cells in neurodegenerative diseases and in particular how NSPCs affect the AD pathology and how neurodegeneration affects NSPCs. In the end of this review, we will discuss how zebrafish as a useful model organism with extensive regenerative ability in the brain might help to address the molecular programs needed for NSPCs to respond to neurodegeneration by enhanced neurogenesis. PMID:27505014

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

    PubMed

    Waters, M J; Blackmore, D G

    2011-12-01

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

  17. Aging of the subventricular zone neural stem cell niche.

    PubMed

    Conover, Joanne C; Shook, Brett A

    2011-02-01

    The persistence of an active subventricular zone neural stem cell niche in the adult mammalian forebrain supports its continued role in the production of new neurons and in generating cells to function in repair through adulthood. Unfortunately, with increasing age the niche begins to deteriorate, compromising these functions. The reasons for this decline are not clear. Studies are beginning to define the molecular and physiologic changes in the microenvironment of the aging subventricular zone niche. New revelations from aging studies will allow for a more thorough understanding of which reparative functions are lost in the aged brain, the progression of niche demise and the possibility for therauptic intervention to improve aging brain function. PMID:22396866

  18. DETAIL OF ZINC CLEANER CELL INTERIOR (EXPOSED AT F/45 FOR ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    DETAIL OF ZINC CLEANER CELL INTERIOR (EXPOSED AT F/45 FOR DEPTH OF FIELD PURPOSES). NOTE GALIGHER STYLE BAFFLES AND TENDENCY OF ZINC TO BUILD UP ON CELL COMPONENTS. - Shenandoah-Dives Mill, 135 County Road 2, Silverton, San Juan County, CO

  19. Adipose tissue-derived stem cells in neural regenerative medicine.

    PubMed

    Yeh, Da-Chuan; Chan, Tzu-Min; Harn, Horng-Jyh; Chiou, Tzyy-Wen; Chen, Hsin-Shui; Lin, Zung-Sheng; Lin, Shinn-Zong

    2015-01-01

    Adipose tissue-derived stem cells (ADSCs) have two essential characteristics with regard to regenerative medicine: the convenient and efficient generation of large numbers of multipotent cells and in vitro proliferation without a loss of stemness. The implementation of clinical trials has prompted widespread concern regarding safety issues and has shifted research toward the therapeutic efficacy of stem cells in dealing with neural degeneration in cases such as stroke, amyotrophic lateral sclerosis, Parkinson's disease, Alzheimer's disease, Huntington's disease, cavernous nerve injury, and traumatic brain injury. Most existing studies have reported that cell therapies may be able to replenish lost cells and promote neuronal regeneration, protect neuronal survival, and play a role in overcoming permanent paralysis and loss of sensation and the recovery of neurological function. The mechanisms involved in determining therapeutic capacity remain largely unknown; however, this concept can still be classified in a methodical manner by citing current evidence. Possible mechanisms include the following: 1) the promotion of angiogenesis, 2) the induction of neuronal differentiation and neurogenesis, 3) reductions in reactive gliosis, 4) the inhibition of apoptosis, 5) the expression of neurotrophic factors, 6) immunomodulatory function, and 7) facilitating neuronal integration. In this study, several human clinical trials using ADSCs for neuronal disorders were investigated. It is suggested that ADSCs are one of the choices among various stem cells for translating into clinical application in the near future.

  20. Chronic ethanol consumption transiently reduces adult neural progenitor cell proliferation.

    PubMed

    Rice, Ann C; Bullock, M Ross; Shelton, Keith L

    2004-06-11

    Adult neural stem/progenitor cells proliferate throughout the life of the animal in the subependymal zone and the subgranular zone of the dentate gyrus (DG). Treatments such as enriched environment, dietary restriction, running and anti-depressants increase proliferation, however, stress and opiates have been shown to decrease proliferation. While models of binge ethanol drinking decreases proliferation, few studies have characterized the effect chronic ethanol usage has on progenitor cell proliferation. In this study, we have examined changes in the progenitor cell proliferation rate following chronic ethanol consumption. Animals were given a nutritionally balanced liquid diet containing 6.5% v/v ethanol or an isocalorically balanced liquid diet. Bromodeoxyuridine (BrdU) was administered (150 mg/kg x 3) and the animals sacrificed 2 h after the last injection on days 3, 10 or 30 of the ethanol diet. Coronal brain blocks were paraffin embedded and 6 microm sections sliced and immunohistochemically stained for BrdU. Quantitation of the number of BrdU-labeled cells in the subgranular zone of the DG revealed a significant decrease only at the 3-day time-point, with recovery by the 10- and 30-day time-points. Thus, the progenitor cell proliferation rate is transiently decreased by chronic ethanol usage. This data suggests that chronic alcohol use results in a compensatory response that restores the progenitor cell proliferation rate.

  1. Adipose tissue-derived stem cells in neural regenerative medicine.

    PubMed

    Yeh, Da-Chuan; Chan, Tzu-Min; Harn, Horng-Jyh; Chiou, Tzyy-Wen; Chen, Hsin-Shui; Lin, Zung-Sheng; Lin, Shinn-Zong

    2015-01-01

    Adipose tissue-derived stem cells (ADSCs) have two essential characteristics with regard to regenerative medicine: the convenient and efficient generation of large numbers of multipotent cells and in vitro proliferation without a loss of stemness. The implementation of clinical trials has prompted widespread concern regarding safety issues and has shifted research toward the therapeutic efficacy of stem cells in dealing with neural degeneration in cases such as stroke, amyotrophic lateral sclerosis, Parkinson's disease, Alzheimer's disease, Huntington's disease, cavernous nerve injury, and traumatic brain injury. Most existing studies have reported that cell therapies may be able to replenish lost cells and promote neuronal regeneration, protect neuronal survival, and play a role in overcoming permanent paralysis and loss of sensation and the recovery of neurological function. The mechanisms involved in determining therapeutic capacity remain largely unknown; however, this concept can still be classified in a methodical manner by citing current evidence. Possible mechanisms include the following: 1) the promotion of angiogenesis, 2) the induction of neuronal differentiation and neurogenesis, 3) reductions in reactive gliosis, 4) the inhibition of apoptosis, 5) the expression of neurotrophic factors, 6) immunomodulatory function, and 7) facilitating neuronal integration. In this study, several human clinical trials using ADSCs for neuronal disorders were investigated. It is suggested that ADSCs are one of the choices among various stem cells for translating into clinical application in the near future. PMID:25647067

  2. Gene transfer into neural cells in vitro using adenoviral vectors.

    PubMed

    Southgate, T D; Kingston, P A; Castro, M G

    2001-05-01

    Adenoviruses (Ads) have become a very attractive and versatile vector system for delivering genes into brain cells in vitro and in vivo. One of the main attractions of Ads is that they can mediate gene transfer into post-mitotic cells, i.e. neurons. Ads are easy to grow and manipulate, stable, and their biology is very well understood. This unit is designed to help newcomers into the field, to design, prepare and grow replication-defective recombinant adenovirus vectors with the aim of transferring genes into neurons and glial cells in primary culture. It provides step-by-step methods describing the preparation of brain cell cultures, their infection using recombinant adenovirus vectors and also the assessment of transgene expression using a variety of techniques including fluorescence immunocytochemistry and fluorescence activated cell-sorting (FACS) analysis. The methods described will be useful to scientists wishing to enter the adenovirus field to construct adenovirus vectors to be used for gene transfer into neural cells.

  3. Biomaterials coated by dental pulp cells as substrate for neural stem cell differentiation.

    PubMed

    Soria, Jose Miguel; Sancho-Tello, María; Esparza, M Angeles Garcia; Mirabet, Vicente; Bagan, Jose Vicente; Monleón, Manuel; Carda, Carmen

    2011-04-01

    This study is focused on the development of an in vitro hybrid system, consisting in a polymeric biomaterial covered by a dental pulp cellular stroma that acts as a scaffold offering a neurotrophic support for the subsequent survival and differentiation of neural stem cells. In the first place, the behavior of dental pulp stroma on the polymeric biomaterial based on ethyl acrylate and hydroxy ethyl acrylate copolymer was studied. For this purpose, cells from normal human third molars were grown onto 0.5-mm-diameter biomaterial discs. After cell culture, quantification of neurotrophic factors generated by the stromal cells was performed by means of an ELISA assay. In the second place, survival and differentiation of adult murine neural stem cells on the polymeric biomaterials covered by dental pulp stromal cells was studied. The results show the capacity of dental pulp cells to uniformly coat the majority of the material's surface and to secrete neurotrophic factors that become crucial for a subsequent differentiation of neural stem cells. The use of stromal cells cultured on scaffolding biomaterials provides neurotrophic pumps that may suggest new criteria for the design of cell therapy experiments in animal models to assist the repair of lesions in Central Nervous System.

  4. Susceptibility of human embryonic stem cell-derived neural cells to Japanese encephalitis virus infection.

    PubMed

    Shen, Shih-Cheng; Shen, Ching-I; Lin, Ho; Chen, Chun-Jung; Chang, Chia-Yu; Chen, Sheng-Mei; Lee, Hsiu-Chin; Lai, Ping-Shan; Su, Hong-Lin

    2014-01-01

    Pluripotent human embryonic stem cells (hESCs) can be efficiently directed to become immature neuroepithelial precursor cells (NPCs) and functional mature neural cells, including neurotransmitter-secreting neurons and glial cells. Investigating the susceptibility of these hESCs-derived neural cells to neurotrophic viruses, such as Japanese encephalitis virus (JEV), provides insight into the viral cell tropism in the infected human brain. We demonstrate that hESC-derived NPCs are highly vulnerable to JEV infection at a low multiplicity of infection (MOI). In addition, glial fibrillary acid protein (GFAP)-expressing glial cells are also susceptible to JEV infection. In contrast, only a few mature neurons were infected at MOI 10 or higher on the third day post-infection. In addition, functional neurotransmitter-secreting neurons are also resistant to JEV infection at high MOI. Moreover, we discover that vimentin intermediate filament, reported as a putative neurovirulent JEV receptor, is highly expressed in NPCs and glial cells, but not mature neurons. These results indicate that the expression of vimentin in neural cells correlates to the cell tropism of JEV. Finally, we further demonstrate that membranous vimentin is necessary for the susceptibility of hESC-derived NPCs to JEV infection.

  5. Primary brain tumors, neural stem cell, and brain tumor cancer cells: where is the link?

    PubMed Central

    Germano, Isabelle; Swiss, Victoria; Casaccia, Patrizia

    2010-01-01

    The discovery of brain tumor-derived cells (BTSC) with the properties of stem cells has led to the formulation of the hypothesis that neural stem cells could be the cell of origin of primary brain tumors (PBT). In this review we present the most common molecular changes in PBT, define the criteria of identification of BTSC and discuss the similarities between the characteristics of these cells and those of the endogenous population of neural stem cells (NPCs) residing in germinal areas of the adult brain. Finally, we propose possible mechanisms of cancer initiation and progression and suggest a model of tumor initiation that includes intrinsic changes of resident NSC and potential changes in the microenvironment defining the niche where the NSC reside. PMID:20045420

  6. Slits Affect the Timely Migration of Neural Crest Cells via Robo Receptor

    PubMed Central

    Giovannone, Dion; Reyes, Michelle; Reyes, Rachel; Correa, Lisa; Martinez, Darwin; Ra, Hannah; Gomez, Gustavo; Kaiser, Josh; Ma, Le; Stein, Mary-Pat; de Bellard, Maria Elena

    2013-01-01

    SUMMARY Background Neural crest cells emerge by delamination from the dorsal neural tube and give rise to various components of the peripheral nervous system in vertebrate embryos. These cells change from non-motile into highly motile cells migrating to distant areas before further differentiation. Mechanisms controlling delamination and subsequent migration of neural crest cells are not fully understood. Slit2, a chemorepellant for axonal guidance that repels and stimulates motility of trunk neural crest cells away from the gut has recently been suggested to be a tumor suppressor molecule. The goal of this study was to further investigate the role of Slit2 in trunk neural crest cell migration by constitutive expression in neural crest cells. Results We found that Slit gain-of-function significantly impaired neural crest cell migration while Slit loss-of-function favored migration. In addition, we observed that the distribution of key cytoskeletal markers was disrupted in both gain and loss of function instances. Conclusions These findings suggest that Slit molecules might be involved in the processes that allow neural crest cells to begin migration and transitioning to a mesenchymal type. PMID:22689303

  7. Planar cell polarity-mediated induction of neural stem cell expansion during axolotl spinal cord regeneration

    PubMed Central

    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. DOI: http://dx.doi.org/10.7554/eLife.10230.001 PMID:26568310

  8. Human endometrial stem cells as a new source for programming to neural cells.

    PubMed

    Mobarakeh, Zahra Taherian; Ai, Jafar; Yazdani, Farzad; Sorkhabadi, Seyed Mahdi Rezayat; Ghanbari, Zinat; Javidan, Abbas Noroozi; Mortazavi-Tabatabaei, Seyed Abdolreza; Massumi, Mohammad; Barough, Somayeh Ebrahimi

    2012-04-26

    Human EnSC (endometrial-derived stem cell) is an abundant and easily available source for cell replacement therapy. Many investigations have shown the potency of the cells to differentiate into several mesoderm-derived cell lineages, including osteocytes and adipocytes. Here, the potency of EnSC in neural differentiation has been investigated. Flow cytometric analysis showed that they were positive for CD90, CD105, OCT4, CD44 and negative for CD31, CD34, CD133. The characterized cells were induced into neural differentiation by bFGF (basic fibroblast growth factor), PDGF (platelet-derived growth factor) and EGF (epidermal growth factor) signalling molecules, respectively in a sequential protocol, and differentiated cells were analysed for expression of neuronal markers by RT-PCR (reverse transcription-PCR) and immunocytochemistry, including Nestin, GABA (γ-aminobutyric acid), MAP2 (microtubule-associated protein 2), β3-tub (class III β-tubulin) and NF-L (neurofilament-light) at the level of their mRNAs. The expression of MAP2, β3-tub and NF-L proteins in EnSC was confirmed 28 days PT (post-treatment) by immunocytochemistry. In conclusion, EnSC can respond to signalling molecules that are usually used as standards in neural differentiation and can programme neuronal cells, making these cells worth considering as a unique source for cell therapy in neurodegenerative disease.

  9. Daucosterol promotes the proliferation of neural stem cells.

    PubMed

    Jiang, Li-hua; Yang, Nian-yun; Yuan, Xiao-lin; Zou, Yi-jie; Zhao, Feng-ming; Chen, Jian-ping; Wang, Ming-yan; Lu, Da-xiang

    2014-03-01

    Neural stem cells (NSCs) are self-regenerating cells, but their regenerative capacity is limited. The present study was conducted to investigate the effect of daucosterol (a sterolin) on the promotion of NSC proliferation and determine the corresponding molecular mechanism. Results of cell counting kit-8 (CCK-8) assay showed that daucosterol significantly increased the quantity of viable cells and the effectiveness of daucosterol was similar to that of basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF). Flow cytometry detection of CFSE-labeled (CFSE, carboxyfluorescein diacetate succinimidyl ester) NSCs showed that Div Index (or the average number of cell divisions) and % Divided (or the percentage of cells that divided at least once) of the cells were increased, indicating that daucosterol increased the percentage of NSCs re-entering the cell cycle. mRNA microarray analysis showed that 333 genes that are mostly involved in the mitotic cell cycle were up-regulated. By contrast, 627 genes that are mostly involved in differentiation were down-regulated. In particular, insulin-like growth factor I (IGF1) was considered as an important regulatory gene that functionally promoted NSC proliferation, and the increased expression of IGF1 protein was validated by ELISA. In addition, the phosphorylation of AKT was increased, indicating that the proliferation-enhancing activity of daucosterol may be involved in IGF1-AKT pathway. Our study provided information about daucosterol as an efficient and inexpensive growth factor alternative that could be used in clinical medicine and research applications. PMID:24333794

  10. Low oxygen alters mitochondrial function and response to oxidative stress in human neural progenitor cells.

    PubMed

    Lages, Yury M; Nascimento, Juliana M; Lemos, Gabriela A; Galina, Antonio; Castilho, Leda R; Rehen, Stevens K

    2015-01-01

    Oxygen concentration should be carefully regulated in all living tissues, beginning at the early embryonic stages. Unbalances in oxygen regulation can lead to cell death and disease. However, to date, few studies have investigated the consequences of variations in oxygen levels for fetal-like cells. Therefore, in the present work, human neural progenitor cells (NPCs) derived from pluripotent stem cells grown in 3% oxygen (v/v) were compared with NPCs cultured in 21% (v/v) oxygen. Low oxygen concentrations altered the mitochondrial content and oxidative functions of the cells, which led to improved ATP production, while reducing generation of reactive oxygen species (ROS). NPCs cultured in both conditions showed no differences in proliferation and glucose metabolism. Furthermore, antioxidant enzymatic activity was not altered in NPCs cultured in 3% oxygen under normal conditions, however, when exposed to external agents known to induce oxidative stress, greater susceptibility to DNA damage was observed. Our findings indicate that the management of oxygen levels should be considered for in vitro models of neuronal development and drug screening.

  11. Endogenous Neural Stem/Progenitor Cells Stabilize the Cortical Microenvironment after Traumatic Brain Injury

    PubMed Central

    Dixon, Kirsty J.; Theus, Michelle H.; Nelersa, Claudiu M.; Mier, Jose; Travieso, Lissette G.; Yu, Tzong-Shiue; Kernie, Steven G.

    2015-01-01

    Abstract Although a myriad of pathological responses contribute to traumatic brain injury (TBI), cerebral dysfunction has been closely linked to cell death mechanisms. A number of therapeutic strategies have been studied in an attempt to minimize or ameliorate tissue damage; however, few studies have evaluated the inherent protective capacity of the brain. Endogenous neural stem/progenitor cells (NSPCs) reside in distinct brain regions and have been shown to respond to tissue damage by migrating to regions of injury. Until now, it remained unknown whether these cells have the capacity to promote endogenous repair. We ablated NSPCs in the subventricular zone to examine their contribution to the injury microenvironment after controlled cortical impact (CCI) injury. Studies were performed in transgenic mice expressing the herpes simplex virus thymidine kinase gene under the control of the nestinδ promoter exposed to CCI injury. Two weeks after CCI injury, mice deficient in NSPCs had reduced neuronal survival in the perilesional cortex and fewer Iba-1-positive and glial fibrillary acidic protein-positive glial cells but increased glial hypertrophy at the injury site. These findings suggest that the presence of NSPCs play a supportive role in the cortex to promote neuronal survival and glial cell expansion after TBI injury, which corresponds with improvements in motor function. We conclude that enhancing this endogenous response may have acute protective roles after TBI. PMID:25290253

  12. Stem Cell Bioprinting: Functional 3D Neural Mini-Tissues from Printed Gel-Based Bioink and Human Neural Stem Cells (Adv. Healthcare Mater. 12/2016).

    PubMed

    Gu, Qi; Tomaskovic-Crook, Eva; Lozano, Rodrigo; Chen, Yu; Kapsa, Robert M; Zhou, Qi; Wallace, Gordon G; Crook, Jeremy M

    2016-06-01

    On page 1429 G. G. Wallace, J. M. Crook, and co-workers report the first example of fabricating neural tissue by 3D bioprinting human neural stem cells. A novel polysaccharide based bioink preserves stem cell viability and function within the printed construct, enabling self-renewal and differentiation to neurons and supporting neuroglia. Neurons are predominantly GABAergic, establish networks, are spontaneously active, and show a bicuculline induced increased calcium response. PMID:27333401

  13. Neural crest cell migratory pathways in the trunk of the chick embryo.

    PubMed

    Loring, J F; Erickson, C A

    1987-05-01

    Neural crest cells migrate during embryogenesis to give rise to segmented structures of the vertebrate peripheral nervous system: namely, the dorsal root ganglia and the sympathetic chain. However, neural crest cell arise from the dorsal neural tube where they are apparently unsegmented. It is generally agreed that the somites impose segmentation on migrating crest cells, but there is a disagreement about two basic questions: exactly pathways do neural crest cells use to move through or around somites, and do neural crest cells actively migrate or are they passively dispersed by the movement of somite cells? The answers to both questions are critically important to any further understanding of the mechanisms underlying the precise distribution of the neural crest cells that develop into ganglia. We have done an exhaustive study of the locations of neural crest cells in chick embryos during early stages of their movement, using antibodies to neural crest cells (HNK-1), to neural filament-associated protein in growing nerve processes (E/C8), and to the extracellular matrix molecule laminin. Our results show that Some neural crest cells invade the extracellular space between adjacent somites, but the apparent majority move into the somites themselves along the border between the dermatome/myotome (DM) and the sclerotome. Neural crest cells remain closely associated with the anterior half of the DM of developing somites as they travel, suggesting that the basal lamina of the DM may be used as a migratory substratum. Supporting this idea is our observation that the development of the DM basal lamina coincides in time and location with the onset of crest migration through the somite. The leading front of neural crest cells advance through the somite while the length of the DM pathway remains constant, suggesting active locomotion, at least in this early phase of development. Neural crest cells leave the DM at a later stage of development to associate with the dorsal

  14. Review: the role of neural crest cells in the endocrine system.

    PubMed

    Adams, Meghan Sara; Bronner-Fraser, Marianne

    2009-01-01

    The neural crest is a pluripotent population of cells that arises at the junction of the neural tube and the dorsal ectoderm. These highly migratory cells form diverse derivatives including neurons and glia of the sensory, sympathetic, and enteric nervous systems, melanocytes, and the bones, cartilage, and connective tissues of the face. The neural crest has long been associated with the endocrine system, although not always correctly. According to current understanding, neural crest cells give rise to the chromaffin cells of the adrenal medulla, chief cells of the extra-adrenal paraganglia, and thyroid C cells. The endocrine tumors that correspond to these cell types are pheochromocytomas, extra-adrenal paragangliomas, and medullary thyroid carcinomas. Although controversies concerning embryological origin appear to have mostly been resolved, questions persist concerning the pathobiology of each tumor type and its basis in neural crest embryology. Here we present a brief history of the work on neural crest development, both in general and in application to the endocrine system. In particular, we present findings related to the plasticity and pluripotency of neural crest cells as well as a discussion of several different neural crest tumors in the endocrine system.

  15. 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

  16. Neural stem cell apoptosis after low-methylmercury exposures in postnatal hippocampus produce persistent cell loss and adolescent memory deficits.

    PubMed

    Sokolowski, Katie; Obiorah, Maryann; Robinson, Kelsey; McCandlish, Elizabeth; Buckley, Brian; DiCicco-Bloom, Emanuel

    2013-12-01

    The developing brain is particularly sensitive to exposures to environmental contaminants. In contrast to the adult, the developing brain contains large numbers of dividing neuronal precursors, suggesting that they may be vulnerable targets. The postnatal day 7 (P7) rat hippocampus has populations of both mature neurons in the CA1-3 region as well as neural stem cells (NSC) in the dentate gyrus (DG) hilus, which actively produce new neurons that migrate to the granule cell layer (GCL). Using this well-characterized NSC population, we examined the impact of low levels of methylmercury (MeHg) on proliferation, neurogenesis, and subsequent adolescent learning and memory behavior. Assessing a range of exposures, we found that a single subcutaneous injection of 0.6 µg/g MeHg in P7 rats induced caspase activation in proliferating NSC of the hilus and GCL. This acute NSC death had lasting impact on the DG at P21, reducing cell numbers in the hilus by 22% and the GCL by 27%, as well as reductions in neural precursor proliferation by 25%. In contrast, non-proliferative CA1-3 pyramidal neuron cell number was unchanged. Furthermore, animals exposed to P7 MeHg exhibited an adolescent spatial memory deficit as assessed by Morris water maze. These results suggest that environmentally relevant levels of MeHg exposure may decrease NSC populations and, despite ongoing neurogenesis, the brain may not restore the hippocampal cell deficits, which may contribute to hippocampal-dependent memory deficits during adolescence.

  17. Mimicking Neural Stem Cell Niche by Biocompatible Substrates

    PubMed Central

    Regalado-Santiago, Citlalli; Juárez-Aguilar, Enrique; Olivares-Hernández, Juan David; Tamariz, Elisa

    2016-01-01

    Neural stem cells (NSCs) participate in the maintenance, repair, and regeneration of the central nervous system. During development, the primary NSCs are distributed along the ventricular zone of the neural tube, while, in adults, NSCs are mainly restricted to the subependymal layer of the subventricular zone of the lateral ventricles and the subgranular zone of the dentate gyrus in the hippocampus. The circumscribed areas where the NSCs are located contain the secreted proteins and extracellular matrix components that conform their niche. The interplay among the niche elements and NSCs determines the balance between stemness and differentiation, quiescence, and proliferation. The understanding of niche characteristics and how they regulate NSCs activity is critical to building in vitro models that include the relevant components of the in vivo niche and to developing neuroregenerative approaches that consider the extracellular environment of NSCs. This review aims to examine both the current knowledge on neurogenic niche and how it is being used to develop biocompatible substrates for the in vitro and in vivo mimicking of extracellular NSCs conditions. PMID:26880934

  18. A novel role for Lh3 dependent ECM modifications during neural crest cell migration in zebrafish.

    PubMed

    Banerjee, Santanu; Isaacman-Beck, Jesse; Schneider, Valerie A; Granato, Michael

    2013-01-01

    During vertebrate development, trunk neural crest cells delaminate along the entire length of the dorsal neural tube and initially migrate as a non-segmented sheet. As they enter the somites, neural crest cells rearrange into spatially restricted segmental streams. Extracellular matrix components are likely to play critical roles in this transition from a sheet-like to a stream-like mode of migration, yet the extracellular matrix components and their modifying enzymes critical for this transition are largely unknown. Here, we identified the glycosyltransferase Lh3, known to modify extracellular matrix components, and its presumptive substrate Collagen18A1, to provide extrinsic signals critical for neural crest cells to transition from a sheet-like migration behavior to migrating as a segmental stream. Using live cell imaging we show that in lh3 null mutants, neural crest cells fail to transition from a sheet to a stream, and that they consequently enter the somites as multiple streams, or stall shortly after entering the somites. Moreover, we demonstrate that transgenic expression of lh3 in a small subset of somitic cells adjacent to where neural crest cells switch from sheet to stream migration restores segmental neural crest cell migration. Finally, we show that knockdown of the presumptive Lh3 substrate Collagen18A1 recapitulates the neural crest cell migration defects observed in lh3 mutants, consistent with the notion that Lh3 exerts its effect on neural crest cell migration by regulating post-translational modifications of Collagen18A1. Together these data suggest that Lh3-Collagen18A1 dependent ECM modifications regulate the transition of trunk neural crest cells from a non-segmental sheet like migration mode to a segmental stream migration mode.

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

    PubMed

    Christie, Kimberly J; Turnley, Ann M

    2012-01-01

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

  20. Brief Azacytidine Step Allows The Conversion of Suspension Human Fibroblasts into Neural Progenitor-Like Cells

    PubMed Central

    Mirakhori, Fahimeh; Zeynali, Bahman; Kiani, Sahar; Baharvand, Hossein

    2015-01-01

    In recent years transdifferentiation technology has enabled direct conversion of human fibroblasts to become a valuable, abundant and accessible cell source for patient-specific induced cell generation in biomedical research. The majority of transdifferentiation approaches rely upon viral gene delivery which due to random integration with the host genome can cause genome instability and tumorigenesis upon transplantation. Here, we provide a simple way to induce neural progenitor-like cells from human fibroblasts without genetic manipulation by changing physicochemical culture properties from monolayer culture into a suspension in the presence of a chemical DNA methyltransferase inhibitor agent, Azacytidine. We have demonstrated the expression of neural progenitor-like markers, morphology and the ability to spontaneously differentiate into neural-like cells. This approach is simple, inexpensive, lacks genetic manipulation and could be a foundation for future chemical neural transdifferentiation and a safe induction of neural progenitor cells from human fibroblasts for clinical applications. PMID:25870845

  1. Zhichan decoction induces differentiation of dopaminergic neurons in Parkinson's disease rats after neural stem cell transplantation

    PubMed Central

    Shi, Huifen; Song, Jie; Yang, Xuming

    2014-01-01

    The goal of this study was to increase the dopamine content and reduce dopaminergic metabolites in the brain of Parkinson's disease rats. Using high-performance liquid chromatography, we found that dopamine and dopaminergic metabolite (dihydroxyphenylacetic acid and homovanillic acid) content in the midbrain of Parkinson's disease rats was increased after neural stem cell transplantation + Zhichan decoction, compared with neural stem cell transplantation alone. Our genetic algorithm results show that dihydroxyphenylacetic acid and homovanillic acid levels achieve global optimization. Neural stem cell transplantation + Zhichan decoction increased dihydroxyphenylacetic acid levels up to 10-fold, while transplantation alone resulted in a 3-fold increment. Homovanillic acid levels showed no apparent change. Our experimental findings show that after neural stem cell transplantation in Parkinson's disease rats, Zhichan decoction can promote differentiation of neural stem cells into dopaminergic neurons. PMID:25206914

  2. Elements of a neural stem cell niche derived from embryonic stem cells.

    PubMed

    Pierret, Chris; Spears, Kathleen; Morrison, Jason A; Maruniak, Joel A; Katz, Martin L; Kirk, Mark D

    2007-12-01

    Recent studies show that adult neural tissues can harbor stem cells within unique niches. In the mammalian central nervous system, neural stem cell (NSC) niches have been identified in the dentate gyrus and the subventricular zone (SVZ). Stem cells in the well-characterized SVZ exist in a microenvironment established by surrounding cells and tissue components, including transit-amplifying cells, neuroblasts, ependymal cells, blood vessels, and a basal lamina. Within this microenvironment, stem cell properties, including proliferation and differentiation, are maintained. Current NSC culture techniques often include the addition of molecular components found within the in vivo niche, such as mitogenic growth factors. Some protocols use bio-scaffolds to mimic the physical growth environment of living tissue. We describe a novel NSC culture system, derived from embryonic stem (ES) cells, that displays elements of an NSC niche in the absence of exogenously applied mitogens or complex physical scaffolding. Mouse ES cells were neuralized with retinoic acid and plated on an entactin-collagen-laminin-coated glass surface at high density (250,000 cells/cm(2)). Six to eight days after plating, complex multicellular structures consisting of heterogeneous cell types developed spontaneously. NSC and progenitor cell proliferation and differentiation continued within these structures. The identity of cellular and molecular components within the cultures was documented using RT-PCR, immunocytochemistry, and neurosphere-forming assays. We show that ES cells can be induced to form structures that exhibit key properties of a developing NSC niche. We believe this system can serve as a useful model for studies of neurogenesis and stem cell maintenance in the NSC niche as well as for applications in stem cell transplantation.

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

    PubMed Central

    Han, Youngmin

    2016-01-01

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

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

    PubMed Central

    Han, Youngmin

    2016-01-01

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

  5. Postnatal Neural Stem Cells in Treating Traumatic Brain Injury.

    PubMed

    Gazalah, Hussein; Mantash, Sarah; Ramadan, Naify; Al Lafi, Sawsan; El Sitt, Sally; Darwish, Hala; Azari, Hassan; Fawaz, Lama; Ghanem, Noël; Zibara, Kazem; Boustany, Rose-Mary; Kobeissy, Firas; Soueid, Jihane

    2016-01-01

    Traumatic brain injury (TBI) is one of the leading causes of death and disabilities worldwide. It affects approximately 1.5 million people each year and is associated with severe post-TBI symptoms such as sensory and motor deficits. Several neuro-therapeutic approaches ranging from cell therapy interventions such as the use of neural stem cells (NSCs) to drug-based therapies have been proposed for TBI management. Successful cell-based therapies are tightly dependent on reproducible preclinical animal models to ensure safety and optimal therapeutic benefits. In this chapter, we describe the isolation of NSCs from neonatal mouse brain using the neurosphere assay in culture. Subsequently, dissociated neurosphere-derived cells are used for transplantation into the ipsilateral cortex of a controlled cortical impact (CCI) TBI model in C57BL/6 mice. Following intra-cardiac perfusion and brain removal, the success of NSC transplantation is then evaluated using immunofluorescence in order to assess neurogenesis along with gliosis in the ipsilateral coronal brain sections. Behavioral tests including rotarod and pole climbing are conducted to evaluate the motor activity post-treatment intervention. PMID:27604746

  6. A fast neural-network algorithm for VLSI cell placement.

    PubMed

    Aykanat, Cevdet; Bultan, Tevfik; Haritaoğlu, Ismail

    1998-12-01

    Cell placement is an important phase of current VLSI circuit design styles such as standard cell, gate array, and Field Programmable Gate Array (FPGA). Although nondeterministic algorithms such as Simulated Annealing (SA) were successful in solving this problem, they are known to be slow. In this paper, a neural network algorithm is proposed that produces solutions as good as SA in substantially less time. This algorithm is based on Mean Field Annealing (MFA) technique, which was successfully applied to various combinatorial optimization problems. A MFA formulation for the cell placement problem is derived which can easily be applied to all VLSI design styles. To demonstrate that the proposed algorithm is applicable in practice, a detailed formulation for the FPGA design style is derived, and the layouts of several benchmark circuits are generated. The performance of the proposed cell placement algorithm is evaluated in comparison with commercial automated circuit design software Xilinx Automatic Place and Route (APR) which uses SA technique. Performance evaluation is conducted using ACM/SIGDA Design Automation benchmark circuits. Experimental results indicate that the proposed MFA algorithm produces comparable results with APR. However, MFA is almost 20 times faster than APR on the average.

  7. Chemokine-Mediated Migration of Mesencephalic Neural Crest Cells

    PubMed Central

    Rezzoug, Francine; Seelan, Ratnam S.; Bhattacherjee, Vasker; Greene, Robert M.; Pisano, M. Michele

    2011-01-01

    Clefts of the lip and/or palate are among the most prevalent birth defects affecting approximately 7000 newborns in the United States annually. Disruption of the developmentally programmed migration of neural crest cells (NCCs) into the orofacial region is thought to be one of the major causes of orofacial clefting. Signaling of the chemokine SDF-1 (Stromal Derived Factor-1) through its specific receptor, CXCR4, is required for the migration of many stem cell and progenitor cell populations from their respective sites of emergence to the regions where they differentiate into complex cell types, tissues and organs. In the present study, “transwell” assays of chick embryo mesencephalic (cranial) NCC migration and ex ovo whole embryo “bead implantation” assays were utilized to determine whether SDF-1/CXCR4 signaling mediates mesencephalic NCC migration. Results from this study demonstrate that attenuation of SDF-1 signaling, through the use of specific CXCR4 antagonists (AMD3100 and TN14003), disrupts the migration of mesencephalic NCCs into the orofacial region, suggesting a novel role for SDF-1/CXCR4 signaling in the directed migration of mesencephalic NCCs in the early stage embryo. PMID:22015108

  8. Effects of engrafted neural stem cells in Alzheimer's disease rats.

    PubMed

    Xuan, A G; Luo, M; Ji, W D; Long, D H

    2009-01-30

    Cell therapy is thought to have a central role in restorative therapy, which aims to restore the function of the damaged nervous system. Neural stem cells (NSCs) can differentiate into neurons, astrocytes and oligodendrocytes. The purpose of this study was to evaluate the therapeutic effects of transplanting NSCs into rats which have the animal model of Alzheimer's disease (AD). NSCs from the hippocampus and NSCs-derived glial cells labeled with 5'-Bromo-2'-deoxyuridine (BrdU) were transplanted into two groups of transected rat basal forebrain. Nestin staining, glial fibrillary acidic protein (GFAP) staining and double-labeling immunofluorescence were used to detect the engrafted cells in the basal forebrain. Immunohistochemical detection of p75(NGFR) showed that the number of cholinergic neurons of the NSCs-transplanted group was significant higher than that of the glia-transplanted group in medial septum (MS) and vertical diagonal branch (VDB) (P<0.05). Learning and memory abilities were also measured by Y-maze test. The results indicate that transplanted NSCs can differentiate into cholinergic neurons, which may play an important role in the therapeutic effects of transplanted NSCs.

  9. Chandelier Cells in Functional and Dysfunctional Neural Circuits

    PubMed Central

    Wang, Yiqing; Zhang, Peng; Wyskiel, Daniel R.

    2016-01-01

    Chandelier cells (ChCs; also called axo-axonic cells) are a specialized GABAergic interneuron subtype that selectively innervates pyramidal neurons at the axon initial segment (AIS), the site of action potential generation. ChC connectivity allows for powerful yet precise modulation of large populations of pyramidal cells, suggesting ChCs have a critical role in brain functions. Dysfunctions in ChC connectivity are associated with brain disorders such as epilepsy and schizophrenia; however, whether this is causative, contributory or compensatory is not known. A likely stumbling block toward mechanistic discoveries and uncovering potential therapeutic targets is the apparent lack of rudimentary understanding of ChCs. For example, whether cortical ChCs are inhibitory or excitatory remains unresolved, and thus whether altered ChC activity results in altered inhibition or excitation is not clear. Recent studies have shed some light onto this excitation-inhibition controversy. In addition, new findings have identified preferential cell-type connectivities established by cortical ChCs, greatly expanding our understanding of the role of ChCs in the cortical microcircuit. Here we aim to bring more attention to ChC connectivity to better understand its role in neural circuits, address whether ChCs are inhibitory or excitatory in light of recent findings and discuss ChC dysfunctions in brain disorders. PMID:27199673

  10. Space Exploration: A Risk for Neural Stem Cells

    NASA Technical Reports Server (NTRS)

    Encinas, Juan M.; Vazquez, Marcelo E.; Switzer, Robert C.; Chamberland, Dennis W.; Nick, Harry; Levine, Howard G.; Scarpa, Philip J.; Enikolopov, Grigori; Steindler, Dennis A.

    2006-01-01

    During spaceflights beyond low Earth orbit, astronauts are exposed to potentially carcinogenic and tissue damaging galactic cosmic rays, solar proton events, and secondary radiation that includes neutrons and recoil nuclei produced by nuclear reactions in spacecraft walls or in tissue (1). Such radiation risk may present a significant health risk for human exploration of the moon and Mars. Emerging evidence that generation of new neurons in the adult brain may be essential for learning, memory, and mood (2) and that radiation is deleterious to neurogenesis (3-5) underscores a previously unappreciated possible risk to the cognitive functions and emotional stability of astronauts exposed to radiation in space. Here we use a novel reporter mouse line to identify at-risk populations of stem and progenitor cells in the brain and find, unexpectedly, that quiescent stem-like cells (rather than their rapidly dividing progeny) in the hippocampus constitute the most vulnerable cell population. This finding raises concerns about the possible risks facing astronauts on long duration space missions.

  11. Gene array analysis of neural crest cells identifies transcription factors necessary for direct conversion of embryonic fibroblasts into neural crest cells

    PubMed Central

    Motohashi, Tsutomu; Watanabe, Natsuki; Nishioka, Masahiro; Nakatake, Yuhki; Yulan, Piao; Mochizuki, Hiromi; Kawamura, Yoshifumi; Ko, Minoru S. H.; Goshima, Naoki; Kunisada, Takahiro

    2016-01-01

    ABSTRACT Neural crest cells (NC cells) are multipotent cells that emerge from the edge of the neural folds and migrate throughout the developing embryo. Although the gene regulatory network for generation of NC cells has been elucidated in detail, it has not been revealed which of the factors in the network are pivotal to directing NC identity. In this study we analyzed the gene expression profile of a pure NC subpopulation isolated from Sox10-IRES-Venus mice and investigated whether these genes played a key role in the direct conversion of Sox10-IRES-Venus mouse embryonic fibroblasts (MEFs) into NC cells. The comparative molecular profiles of NC cells and neural tube cells in 9.5-day embryos revealed genes including transcription factors selectively expressed in developing trunk NC cells. Among 25 NC cell-specific transcription factor genes tested, SOX10 and SOX9 were capable of converting MEFs into SOX10-positive (SOX10+) cells. The SOX10+ cells were then shown to differentiate into neurons, glial cells, smooth muscle cells, adipocytes and osteoblasts. These SOX10+ cells also showed limited self-renewal ability, suggesting that SOX10 and SOX9 directly converted MEFs into NC cells. Conversely, the remaining transcription factors, including well-known NC cell specifiers, were unable to convert MEFs into SOX10+ NC cells. These results suggest that SOX10 and SOX9 are the key factors necessary for the direct conversion of MEFs into NC cells. PMID:26873953

  12. History of Neural Stem Cell Research and Its Clinical Application

    PubMed Central

    TAKAGI, Yasushi

    2016-01-01

    “Once development was ended…in the adult centers, the nerve paths are something fixed and immutable. Everything may die, nothing may be regenerated,” wrote Santiago Ramón y Cajal, a Spanish neuroanatomist and Nobel Prize winner and the father of modern neuroscience. This statement was the central dogma in neuroscience for a long time. However, in the 1960s, neural stem cells (NSCs) were discovered. Since then, our knowledge about NSCs has continued to grow. This review focuses on our current knowledge about NSCs and their surrounding microenvironment. In addition, the clinical application of NSCs for the treatment of various central nervous system diseases is also summarized. PMID:26888043

  13. Direct Adaptive Aircraft Control Using Dynamic Cell Structure Neural Networks

    NASA Technical Reports Server (NTRS)

    Jorgensen, Charles C.

    1997-01-01

    A Dynamic Cell Structure (DCS) Neural Network was developed which learns topology representing networks (TRNS) of F-15 aircraft aerodynamic stability and control derivatives. The network is integrated into a direct adaptive tracking controller. The combination produces a robust adaptive architecture capable of handling multiple accident and off- nominal flight scenarios. This paper describes the DCS network and modifications to the parameter estimation procedure. The work represents one step towards an integrated real-time reconfiguration control architecture for rapid prototyping of new aircraft designs. Performance was evaluated using three off-line benchmarks and on-line nonlinear Virtual Reality simulation. Flight control was evaluated under scenarios including differential stabilator lock, soft sensor failure, control and stability derivative variations, and air turbulence.

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

    PubMed

    Waldau, B; Shetty, A K

    2008-08-01

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

  15. Hoxb1b controls oriented cell division, cell shape and microtubule dynamics in neural tube morphogenesis

    PubMed Central

    Žigman, Mihaela; Laumann-Lipp, Nico; Titus, Tom; Postlethwait, John; Moens, Cecilia B.

    2014-01-01

    Hox genes are classically ascribed to function in patterning the anterior-posterior axis of bilaterian animals; however, their role in directing molecular mechanisms underlying morphogenesis at the cellular level remains largely unstudied. We unveil a non-classical role for the zebrafish hoxb1b gene, which shares ancestral functions with mammalian Hoxa1, in controlling progenitor cell shape and oriented cell division during zebrafish anterior hindbrain neural tube morphogenesis. This is likely distinct from its role in cell fate acquisition and segment boundary formation. We show that, without affecting major components of apico-basal or planar cell polarity, Hoxb1b regulates mitotic spindle rotation during the oriented neural keel symmetric mitoses that are required for normal neural tube lumen formation in the zebrafish. This function correlates with a non-cell-autonomous requirement for Hoxb1b in regulating microtubule plus-end dynamics in progenitor cells in interphase. We propose that Hox genes can influence global tissue morphogenesis by control of microtubule dynamics in individual cells in vivo. PMID:24449840

  16. 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.

  17. Hoxb1b controls oriented cell division, cell shape and microtubule dynamics in neural tube morphogenesis.

    PubMed

    Zigman, Mihaela; Laumann-Lipp, Nico; Titus, Tom; Postlethwait, John; Moens, Cecilia B

    2014-02-01

    Hox genes are classically ascribed to function in patterning the anterior-posterior axis of bilaterian animals; however, their role in directing molecular mechanisms underlying morphogenesis at the cellular level remains largely unstudied. We unveil a non-classical role for the zebrafish hoxb1b gene, which shares ancestral functions with mammalian Hoxa1, in controlling progenitor cell shape and oriented cell division during zebrafish anterior hindbrain neural tube morphogenesis. This is likely distinct from its role in cell fate acquisition and segment boundary formation. We show that, without affecting major components of apico-basal or planar cell polarity, Hoxb1b regulates mitotic spindle rotation during the oriented neural keel symmetric mitoses that are required for normal neural tube lumen formation in the zebrafish. This function correlates with a non-cell-autonomous requirement for Hoxb1b in regulating microtubule plus-end dynamics in progenitor cells in interphase. We propose that Hox genes can influence global tissue morphogenesis by control of microtubule dynamics in individual cells in vivo.

  18. A novel role for MuSK and non-canonical Wnt signaling during segmental neural crest cell migration.

    PubMed

    Banerjee, Santanu; Gordon, Laura; Donn, Thomas M; Berti, Caterina; Moens, Cecilia B; Burden, Steven J; Granato, Michael

    2011-08-01

    Trunk neural crest cells delaminate from the dorsal neural tube as an uninterrupted sheet; however, they convert into segmentally organized streams before migrating through the somitic territory. These neural crest cell streams join the segmental trajectories of pathfinding spinal motor axons, suggesting that interactions between these two cell types might be important for neural crest cell migration. Here, we show that in the zebrafish embryo migration of both neural crest cells and motor axons is temporally synchronized and spatially restricted to the center of the somite, but that motor axons are dispensable for segmental neural crest cell migration. Instead, we find that muscle-specific receptor kinase (MuSK) and its putative ligand Wnt11r are crucial for restricting neural crest cell migration to the center of each somite. Moreover, we find that blocking planar cell polarity (PCP) signaling in somitic muscle cells also results in non-segmental neural crest cell migration. Using an F-actin biosensor we show that in the absence of MuSK neural crest cells fail to retract non-productive leading edges, resulting in non-segmental migration. Finally, we show that MuSK knockout mice display similar neural crest cell migration defects, suggesting a novel, evolutionarily conserved role for MuSK in neural crest migration. We propose that a Wnt11r-MuSK dependent, PCP-like pathway restricts neural crest cells to their segmental path.

  19. Effects of silver nanoparticles on human and rat embryonic neural stem cells

    PubMed Central

    Liu, Fang; Mahmood, Meena; Xu, Yang; Watanabe, Fumiya; Biris, Alexandru S.; Hansen, Deborah K.; Inselman, Amy; Casciano, Daniel; Patterson, Tucker A.; Paule, Merle G.; Slikker, William; Wang, Cheng

    2015-01-01

    Silver nano-particles (Ag-NPs) are becoming increasingly prevalent in consumer products as antibacterial agents. The increased use of Ag NP-enhanced products will almost certainly increase environmental silver levels, resulting in increased exposures and the potential for increased adverse reactions including neurotoxic effects. In the present study, embryonic neural stem cells (NSCs) from human and rat fetuses (gestational day-16) were used to determine whether Ag-NPs are capable of causing developmental neurotoxicity. The NSCs were cultured in serum free medium supplemented with appropriate growth factors. On the eighth day in vitro (DIV 8), the cells were exposed to Ag-NPs at concentrations of 1, 5, 10, and 20 μg/ml for 24 h. The cultured cells then were characterized by NSC markers including nestin and SOX2 and a variety of assays were utilized to determine the effects of Ag-NPs on NSC proliferation and viability and the underlying mechanisms associated with these effects. The results indicate that mitochondrial viability (MTT metabolism) was substantially attenuated and LDH release was increased significantly in a dose-dependent manner. Ag-NPs-induced neurotoxicity was further confirmed by up-regulated Bax protein expression, an increased number of TUNEL-positively stained cells, and elevated reactive oxygen species (ROS). NSC proliferation was also significantly decreased by Ag-NPs. Co-administration of acetyl-L-carnitine, an antioxidant agent, effectively blocked the adverse effects associated with Ag-NP exposure. PMID:25904840

  20. Effects of silver nanoparticles on human and rat embryonic neural stem cells.

    PubMed

    Liu, Fang; Mahmood, Meena; Xu, Yang; Watanabe, Fumiya; Biris, Alexandru S; Hansen, Deborah K; Inselman, Amy; Casciano, Daniel; Patterson, Tucker A; Paule, Merle G; Slikker, William; Wang, Cheng

    2015-01-01

    Silver nano-particles (Ag-NPs) are becoming increasingly prevalent in consumer products as antibacterial agents. The increased use of Ag NP-enhanced products will almost certainly increase environmental silver levels, resulting in increased exposures and the potential for increased adverse reactions including neurotoxic effects. In the present study, embryonic neural stem cells (NSCs) from human and rat fetuses (gestational day-16) were used to determine whether Ag-NPs are capable of causing developmental neurotoxicity. The NSCs were cultured in serum free medium supplemented with appropriate growth factors. On the eighth day in vitro (DIV 8), the cells were exposed to Ag-NPs at concentrations of 1, 5, 10, and 20 μg/ml for 24 h. The cultured cells then were characterized by NSC markers including nestin and SOX2 and a variety of assays were utilized to determine the effects of Ag-NPs on NSC proliferation and viability and the underlying mechanisms associated with these effects. The results indicate that mitochondrial viability (MTT metabolism) was substantially attenuated and LDH release was increased significantly in a dose-dependent manner. Ag-NPs-induced neurotoxicity was further confirmed by up-regulated Bax protein expression, an increased number of TUNEL-positively stained cells, and elevated reactive oxygen species (ROS). NSC proliferation was also significantly decreased by Ag-NPs. Co-administration of acetyl-L-carnitine, an antioxidant agent, effectively blocked the adverse effects associated with Ag-NP exposure. PMID:25904840

  1. Transplantation of neural progenitor cells in chronic spinal cord injury.

    PubMed

    Jin, Y; Bouyer, J; Shumsky, J S; Haas, C; Fischer, I

    2016-04-21

    Previous studies demonstrated that neural progenitor cells (NPCs) transplanted into a subacute contusion injury improve motor, sensory, and bladder function. In this study we tested whether transplanted NPCs can also improve functional recovery after chronic spinal cord injury (SCI) alone or in combination with the reduction of glial scar and neurotrophic support. Adult rats received a T10 moderate contusion. Thirteen weeks after the injury they were divided into four groups and received either: 1. Medium (control), 2. NPC transplants, 3. NPC+lentivirus vector expressing chondroitinase, or 4. NPC+lentivirus vectors expressing chondroitinase and neurotrophic factors. During the 8 weeks post-transplantation the animals were tested for functional recovery and eventually analyzed by anatomical and immunohistochemical assays. The behavioral tests for motor and sensory function were performed before and after injury, and weekly after transplantation, with some animals also tested for bladder function at the end of the experiment. Transplant survival in the chronic injury model was variable and showed NPCs at the injury site in 60% of the animals in all transplantation groups. The NPC transplants comprised less than 40% of the injury site, without significant anatomical or histological differences among the groups. All groups also showed similar patterns of functional deficits and recovery in the 12 weeks after injury and in the 8 weeks after transplantation using the Basso, Beattie, and Bresnahan rating score, the grid test, and the Von Frey test for mechanical allodynia. A notable exception was group 4 (NPC together with chondroitinase and neurotrophins), which showed a significant improvement in bladder function. This study underscores the therapeutic challenges facing transplantation strategies in a chronic SCI in which even the inclusion of treatments designed to reduce scarring and increase neurotrophic support produce only modest functional improvements. Further

  2. 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

  3. Neural differentiation of adipose-derived stem cells isolated from GFP transgenic mice

    SciTech Connect

    Fujimura, Juri; E-mail: juri-f@nms.ac.jp; Ogawa, Rei; Mizuno, Hiroshi; Fukunaga, Yoshitaka; Suzuki, Hidenori

    2005-07-22

    Taking advantage of homogeneously marked cells from green fluorescent protein (GFP) transgenic mice, we have recently reported that adipose-derived stromal cells (ASCs) could differentiate into mesenchymal lineages in vitro. In this study, we performed neural induction using ASCs from GFP transgenic mice and were able to induce these ASCs into neuronal and glial cell lineages. Most of the neurally induced cells showed bipolar or multipolar appearance morphologically and expressed neuronal markers. Electron microscopy revealed their neuronal morphology. Some cells also showed glial phenotypes, as shown immunocytochemically. The present study clearly shows that ASCs derived from GFP transgenic mice differentiate into neural lineages in vitro, suggesting that these cells might provide an ideal source for further neural stem cell research with possible therapeutic application for neurological disorders.

  4. Alternative Routes to Induced Pluripotent Stem Cells Revealed by Reprogramming of the Neural Lineage.

    PubMed

    Jackson, Steven A; Olufs, Zachariah P G; Tran, Khoa A; Zaidan, Nur Zafirah; Sridharan, Rupa

    2016-03-01

    During the reprogramming of mouse embryonic fibroblasts (MEFs) to induced pluripotent stem cells, the activation of pluripotency genes such as NANOG occurs after the mesenchymal to epithelial transition. Here we report that both adult stem cells (neural stem cells) and differentiated cells (astrocytes) of the neural lineage can activate NANOG in the absence of cadherin expression during reprogramming. Gene expression analysis revealed that only the NANOG+E-cadherin+ populations expressed stabilization markers, had upregulated several cell cycle genes; and were transgene independent. Inhibition of DOT1L activity enhanced both the numbers of NANOG+ and NANOG+E-cadherin+ colonies in neural stem cells. Expressing SOX2 in MEFs prior to reprogramming did not alter the ratio of NANOG colonies that express E-cadherin. Taken together these results provide a unique pathway for reprogramming taken by cells of the neural lineage. PMID:26905202

  5. X-box-binding protein 1-modified neural stem cells for treatment of Parkinson's disease.

    PubMed

    Si, Lihui; Xu, Tianmin; Wang, Fengzhang; Liu, Qun; Cui, Manhua

    2012-04-01

    X-box-binding protein 1-transfected neural stem cells were transplanted into the right lateral ventricles of rats with rotenone-induced Parkinson's disease. The survival capacities and differentiation rates of cells expressing the dopaminergic marker tyrosine hydroxylase were higher in X-box-binding protein 1-transfected neural stem cells compared to non-transfected cells. Moreover, dopamine and 3,4-dihydroxyphenylacetic acid levels in the substantia nigra were significantly increased, α-synuclein expression was decreased, and neurological behaviors were significantly ameliorated in rats following transplantation of X-box-binding protein 1-transfected neural stem cells. These results indicate that transplantation of X-box-binding protein 1-transfected neural stem cells can promote stem cell survival and differentiation into dopaminergic neurons, increase dopamine and 3,4-dihydroxyphenylacetic acid levels, reduce α-synuclein aggregation in the substantia nigra, and improve the symptoms of Parkinson's disease in rats.

  6. Extracellular vesicles released from cells exposed to reactive oxygen species increase annexin A2 expression and survival of target cells exposed to the same conditions.

    PubMed

    Grindheim, Ann Kari; Vedeler, Anni

    2016-01-01

    Annexin A2 (AnxA2) is present in multiple cellular compartments and interacts with numerous ligands including calcium, proteins, cholesterol, negatively charged phospholipids and RNA. These interactions are tightly regulated by its post-translational modifications. The levels of AnxA2 and its Tyr23 phosphorylated form (pTyr23AnxA2) are increased in many cancers and the protein is involved in malignant cell transformation, metastasis and angiogenesis. Our previous studies of rat pheochromocytoma (PC12) cells showed that reactive oxygen species (ROS) induce rapid, simultaneous and transient dephosphorylation of nuclear AnxA2, most likely associating with PML bodies, while AnxA2 associated with F-actin at the cell cortex undergoes Tyr23 phosphorylation. The pTyr23AnxA2 in the periphery of the cells is incorporated into intraluminal vesicles of multivesicular endosomes and subsequently released to the extracellular space. We show here that extracellular vesicles (EVs) from cells exposed to ROS prime untreated PC12 cells to better tolerate subsequent oxidative stress, thus enhancing their survival. There is an increase in the levels of pTyr23AnxA2 and AnxA2 in the primed cells, suggesting that AnxA2 is involved in their survival. This increase is due to an upregulation of AnxA2 expression both at the transcriptional and translational levels after relatively short term (2 h) exposure to primed EVs. PMID:27574537

  7. Resident neural stem cells restrict tissue damage and neuronal loss after spinal cord injury in mice.

    PubMed

    Sabelström, Hanna; Stenudd, Moa; Réu, Pedro; Dias, David O; Elfineh, Marta; Zdunek, Sofia; Damberg, Peter; Göritz, Christian; Frisén, Jonas

    2013-11-01

    Central nervous system injuries are accompanied by scar formation. It has been difficult to delineate the precise role of the scar, as it is made by several different cell types, which may limit the damage but also inhibit axonal regrowth. We show that scarring by neural stem cell-derived astrocytes is required to restrict secondary enlargement of the lesion and further axonal loss after spinal cord injury. Moreover, neural stem cell progeny exerts a neurotrophic effect required for survival of neurons adjacent to the lesion. One distinct component of the glial scar, deriving from resident neural stem cells, is required for maintaining the integrity of the injured spinal cord.

  8. Hypoxic preconditioning involves system Xc- regulation in mouse neural stem cells.

    PubMed

    Sims, Brian; Clarke, Melinda; Francillion, Ludwig; Kindred, Elijah; Hopkins, Elana Shuford; Sontheimer, Harald

    2012-03-01

    In animals, hypoxic preconditioning has been used as a form of neuroprotection. The exact mechanism involved in neuroprotective hypoxic preconditioning has not been described, yet could be valuable for possible neuroprotective strategies. The overexpression of the cystine-glutamate exchanger, system Xc-, has been demonstrated as being neuroprotective (Shih, Erb et al. 2006). Here, using immunohistochemistry, we demonstrate that C57BL/6 mice exposed to hypoxia showed an increase in system Xc- expression, with the highest level of intensity in the hippocampus. Western Blot analysis also showed an almost 2-fold increase in system Xc- protein in hypoxia-exposed versus control mice. The mRNA for the regulatory subunit of system Xc-, xCT, and the xCT/actin ratio were also increased under hypoxic conditions. Experiments using hypoxia-inducible factor (HIF-1α) siRNA showed a statistically significant decrease in HIF-1α and system Xc- expression. Under hypoxic conditions, system Xc- activity, as determined by cystine uptake, increased 2-fold. Importantly, hypoxic preconditioning was attenuated in neural stem cells by pharmacological inhibition of system Xc- activity with S4-carboxyphenylglycine. These data provide the first evidence of hypoxic regulation of the cystine glutamate exchanger system Xc-.

  9. Acrylamide affects proliferation and differentiation of the neural progenitor cell line C17.2 and the neuroblastoma cell line SH-SY5Y.

    PubMed

    Attoff, K; Kertika, D; Lundqvist, J; Oredsson, S; Forsby, A

    2016-09-01

    Acrylamide is a well-known neurotoxic compound and people get exposed to the compound by food consumption and environmental pollutants. Since acrylamide crosses the placenta barrier, the fetus is also being exposed resulting in a risk for developmental neurotoxicity. In this study, the neural progenitor cell line C17.2 and the neuroblastoma cell line SH-SY5Y were used to study proliferation and differentiation as alerting indicators for developmental neurotoxicity. For both cell lines, acrylamide reduced the number of viable cells by reducing proliferation and inducing cell death in undifferentiated cells. Acrylamide concentrations starting at 10fM attenuated the differentiation process in SH-SY5Y cells by sustaining cell proliferation and neurite outgrowth was reduced at concentrations from 10pM. Acrylamide significantly reduced the number of neurons starting at 1μM and altered the ratio between the different phenotypes in differentiating C17.2 cell cultures. Ten micromolar of acrylamide also reduced the expression of the neuronal and astrocyte biomarkers. Although the neurotoxic concentrations in the femtomolar range seem to be specific for the SH-SY5Y cell line, the fact that micromolar concentrations of acrylamide seem to attenuate the differentiation process in both cell lines raises the interest to further investigations on the possible developmental neurotoxicity of acrylamide. PMID:27241584

  10. Acrylamide affects proliferation and differentiation of the neural progenitor cell line C17.2 and the neuroblastoma cell line SH-SY5Y.

    PubMed

    Attoff, K; Kertika, D; Lundqvist, J; Oredsson, S; Forsby, A

    2016-09-01

    Acrylamide is a well-known neurotoxic compound and people get exposed to the compound by food consumption and environmental pollutants. Since acrylamide crosses the placenta barrier, the fetus is also being exposed resulting in a risk for developmental neurotoxicity. In this study, the neural progenitor cell line C17.2 and the neuroblastoma cell line SH-SY5Y were used to study proliferation and differentiation as alerting indicators for developmental neurotoxicity. For both cell lines, acrylamide reduced the number of viable cells by reducing proliferation and inducing cell death in undifferentiated cells. Acrylamide concentrations starting at 10fM attenuated the differentiation process in SH-SY5Y cells by sustaining cell proliferation and neurite outgrowth was reduced at concentrations from 10pM. Acrylamide significantly reduced the number of neurons starting at 1μM and altered the ratio between the different phenotypes in differentiating C17.2 cell cultures. Ten micromolar of acrylamide also reduced the expression of the neuronal and astrocyte biomarkers. Although the neurotoxic concentrations in the femtomolar range seem to be specific for the SH-SY5Y cell line, the fact that micromolar concentrations of acrylamide seem to attenuate the differentiation process in both cell lines raises the interest to further investigations on the possible developmental neurotoxicity of acrylamide.

  11. Molecular Evolution of Drosophila Germline Stem Cell and Neural Stem Cell Regulating Genes

    PubMed Central

    Choi, Jae Young; Aquadro, Charles F.

    2015-01-01

    Here, we study the molecular evolution of a near complete set of genes that had functional evidence in the regulation of the Drosophila germline and neural stem cell. Some of these genes have previously been shown to be rapidly evolving by positive selection raising the possibility that stem cell genes as a group have elevated signatures of positive selection. Using recent Drosophila comparative genome sequences and population genomic sequences of Drosophila melanogaster, we have investigated both long- and short-term evolution occurring across these two different stem cell systems, and compared them with a carefully chosen random set of genes to represent the background rate of evolution. Our results showed an excess of genes with evidence of a recent selective sweep in both germline and neural stem cells in D. melanogaster. However compared with their control genes, both stem cell systems had no significant excess of genes with long-term recurrent positive selection in D. melanogaster, or across orthologous sequences from the melanogaster group. The evidence of long-term positive selection was limited to a subset of genes with specific functions in both the germline and neural stem cell system. PMID:26507797

  12. Alternating Current Electric Fields of Varying Frequencies: Effects on Proliferation and Differentiation of Porcine Neural Progenitor Cells

    PubMed Central

    Lim, Ji-Hey; McCullen, Seth D.; Piedrahita, Jorge A.

    2013-01-01

    Abstract Application of sinusoidal electric fields (EFs) has been observed to affect cellular processes, including alignment, proliferation, and differentiation. In the present study, we applied low-frequency alternating current (AC) EFs to porcine neural progenitor cells (pNPCs) and investigated the effects on cell patterning, proliferation, and differentiation. pNPCs were grown directly on interdigitated electrodes (IDEs) localizing the EFs to a region accessible visually for fluorescence-based assays. Cultures of pNPCs were exposed to EFs (1 V/cm) of 1 Hz, 10 Hz, and 50 Hz for 3, 7, and 14 days and compared to control cultures. Immunocytochemistry was performed to evaluate the expression of neural markers. pNPCs grew uniformly with no evidence of alignment to the EFs and no change in cell numbers when compared with controls. Nestin expression was shown in all groups at 3 and 7 days, but not at 14 days. NG2 expression was low in all groups. Co-expression of glial fibrillary acidic protein (GFAP) and TUJ1 was significantly higher in the cultures exposed to 10- and 50-Hz EFs than the controls. In summary, sinusoidal AC EFs via IDEs did not alter the alignment and proliferation of pNPCs, but higher frequency stimulation appeared to delay differentiation into mature astrocytes. PMID:23961767

  13. Alternating current electric fields of varying frequencies: effects on proliferation and differentiation of porcine neural progenitor cells.

    PubMed

    Lim, Ji-Hey; McCullen, Seth D; Piedrahita, Jorge A; Loboa, Elizabeth G; Olby, Natasha J

    2013-10-01

    Application of sinusoidal electric fields (EFs) has been observed to affect cellular processes, including alignment, proliferation, and differentiation. In the present study, we applied low-frequency alternating current (AC) EFs to porcine neural progenitor cells (pNPCs) and investigated the effects on cell patterning, proliferation, and differentiation. pNPCs were grown directly on interdigitated electrodes (IDEs) localizing the EFs to a region accessible visually for fluorescence-based assays. Cultures of pNPCs were exposed to EFs (1 V/cm) of 1 Hz, 10 Hz, and 50 Hz for 3, 7, and 14 days and compared to control cultures. Immunocytochemistry was performed to evaluate the expression of neural markers. pNPCs grew uniformly with no evidence of alignment to the EFs and no change in cell numbers when compared with controls. Nestin expression was shown in all groups at 3 and 7 days, but not at 14 days. NG2 expression was low in all groups. Co-expression of glial fibrillary acidic protein (GFAP) and TUJ1 was significantly higher in the cultures exposed to 10- and 50-Hz EFs than the controls. In summary, sinusoidal AC EFs via IDEs did not alter the alignment and proliferation of pNPCs, but higher frequency stimulation appeared to delay differentiation into mature astrocytes.

  14. 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

  15. Glioma migration: clues from the biology of neural progenitor cells and embryonic CNS cell migration.

    PubMed

    Dirks, P B

    2001-06-01

    Neural stem cells have recently come to the forefront in neurobiology because of the possibilities for CNS repair by transplantation. Further understanding of the biology of these cells is critical for making their use in CNS repair possible. It is likely that these discoveries will also have spin-offs for neuro-oncology as primary brain tumors may arise from a CNS progenitor cell. An understanding of the normal migratory ability of these cells is also likely to have a very important impact on the knowledge of brain tumor invasion.

  16. Activation of NK cells in subjects exposed to mild hyper- or hypothermic load.

    PubMed

    Lackovic, V; Borecký, L; Vigas, M; Rovenský, J

    1988-06-01

    The effect of mild hyper- and hypothermic stress on release of selected hormones (somatotropin, noradrenaline, etc.), interferon (IFN), and activity of NK cells in the blood was examined in groups of young males during a 30 min exposure to 39 degrees C and 4 degrees C. A quick release of somatotropin was registered in 44% of examinees in the hyperthermic group, while the persons exposed to 4 degrees C reacted with a release of noradrenaline only. Concurrently, an elevation of NK cell activity was observed both in the subgroup releasing somatotropin after hyperthermic stress and in the group exposed to cold. Since these forms of mild stress did not lead to an appearance of IFN in the serum, the possibility of an NK cell activating effect of somatotropin and/or the adrenal hormones was tested. While the adrenal hormones stimulated the NK cell activity in vitro, no support for a similar role for somatotropin was found. PMID:2457640

  17. Determining adaptive and adverse oxidative stress responses in human bronical epithelial cells exposed to zinc

    EPA Science Inventory

    Determining adaptive and adverse oxidative stress responses in human bronchial epithelial cells exposed to zincJenna M. Currier1,2, Wan-Yun Cheng1, Rory Conolly1, Brian N. Chorley1Zinc is a ubiquitous contaminant of ambient air that presents an oxidant challenge to the human lung...

  18. SIGNALING MECHANISMS IN HUMAN AIRWAY EPITHELIAL CELLS EXPOSED TO CARBON ULTRAFINE PARTICLES

    EPA Science Inventory

    SIGNALING MECHANISMS IN HUMAN AIRWAY EPITHELIAL CELLS EXPOSED TO CARBON ULTRAFINE PARTICLES
    Y.M. Kim, A.G. Lenz, R. Silbajoris, I. Jaspers and J.M. Samet. Department of Environmental Sciences and Engineering and Center for Environmental Medicine, University of North Carolina, ...

  19. Non-Viral Generation of Neural Precursor-like Cells from Adult Human Fibroblasts

    PubMed Central

    Maucksch, C; Firmin, E; Butler-Munro, C; Montgomery, JM; Dottori, M; Connor, B

    2012-01-01

    Recent studies have reported direct reprogramming of human fibroblasts to mature neurons by the introduction of defined neural genes. This technology has potential use in the areas of neurological disease modeling and drug development. However, use of induced neurons for large-scale drug screening and cell-based replacement strategies is limited due to their inability to expand once reprogrammed. We propose it would be more desirable to induce expandable neural precursor cells directly from human fibroblasts. To date several pluripotent and neural transcription factors have been shown to be capable of converting mouse fibroblasts to neural stem/precursor-like cells when delivered by viral vectors. Here we extend these findings and demonstrate that transient ectopic insertion of the transcription factors SOX2 and PAX6 to adult human fibroblasts through use of non-viral plasmid transfection or protein transduction allows the generation of induced neural precursor (iNP) colonies expressing a range of neural stem and pro-neural genes. Upon differentiation, iNP cells give rise to neurons exhibiting typical neuronal morphologies and expressing multiple neuronal markers including tyrosine hydroxylase and GAD65/67. Importantly, iNP-derived neurons demonstrate electrophysiological properties of functionally mature neurons with the capacity to generate action potentials. In addition, iNP cells are capable of differentiating into glial fibrillary acidic protein (GFAP)-expressing astrocytes. This study represents a novel virusfree approach for direct reprogramming of human fibroblasts to a neural precursor fate. PMID:24693194

  20. Neural stem cell sparing by linac based intensity modulated stereotactic radiotherapy in intracranial tumors

    PubMed Central

    2013-01-01

    Background Neurocognitive decline observed after radiotherapy (RT) for brain tumors in long time survivors is attributed to radiation exposure of the hippocampus and the subventricular zone (SVZ). The potential of sparing capabilities for both structures by optimized intensity modulated stereotactic radiotherapy (IMSRT) is investigated. Methods Brain tumors were irradiated by stereotactic 3D conformal RT or IMSRT using m3 collimator optimized for PTV and for sparing of the conventional OARs (lens, retina, optic nerve, chiasm, cochlea, brain stem and the medulla oblongata). Retrospectively both hippocampi and SVZ were added to the list of OAR and their dose volume histograms were compared to those from two newly generated IMSRT plans using 7 or 14 beamlets (IMSRT-7, IMSRT-14) dedicated for optimized additional sparing of these structures. Conventional OAR constraints were kept constant. Impact of plan complexity and planning target volume (PTV) topography on sparing of both hippocampi and SVZ, conformity index (CI), the homogeneity index (HI) and quality of coverage (QoC) were analyzed. Limits of agreement were used to compare sparing of stem cell niches with either IMSRT-7 or IMSRT-14. The influence of treatment technique related to the topography ratio between PTV and OARs, realized in group A-D, was assessed by a mixed model. Results In 47 patients CI (p ≤ 0.003) and HI (p < 0.001) improved by IMSRT-7, IMSRT-14, QoC remained stable (p ≥ 0.50) indicating no compromise in radiotherapy. 90% of normal brain was exposed to a significantly higher dose using IMSRT. IMSRT-7 plans resulted in significantly lower biologically effective doses at all four neural stem cell structures, while contralateral neural stem cells are better spared compared to ipsilateral. A further increase of the number of beamlets (IMSRT-14) did not improve sparing significantly, so IMSRT-7 and IMSRT-14 can be used interchangeable. Patients with tumors contacting neither the

  1. Experimental investigation on neural cell survival after dielectrophoretic trapping.

    PubMed

    Heida, T; Rutten, W L C; Marani, E

    2002-12-01

    Negative dielectrophoretic forces can effectively be used to trap cortical rat neurons. The creation of dielectrophoretic forces requires electric fields of high non-uniformity. High electric field strengths, however, can cause excessive membrane potentials by which cells may unrecoverably be changed or it may lead to cell death. In a previous study it was found that cells trapped at 3 Vtt/14 MHz did not change morphologically as compared to cells that were not exposed to the electric field. This study investigates the viability of fetal cortical rat neurons after being trapped by negative dielectrophoretic forces at frequencies up to 1 MHz. A planar quadrupole micro-electrode structure was used for the creation of a non-uniform electric field. The sinusoidal input signal was varied in amplitude (3 and 5 Vtt) and frequency (10 kHz-1 MHz). The results presented in this paper show that the viability of dielectrophoretically trapped postnatal cortical rat cells was greatly frequency dependent. To preserve viability frequencies above 100 kHz (at 3 Vtt) or 1 MHz (5 Vtt) must be used.

  2. Characterization of neural stemness status through the neurogenesis process for bone marrow mesenchymal stem cells.

    PubMed

    Mohammad, Maeda H; Al-Shammari, Ahmed M; Al-Juboory, Ahmad Adnan; Yaseen, Nahi Y

    2016-01-01

    The in vitro isolation, identification, differentiation, and neurogenesis characterization of the sources of mesenchymal stem cells (MSCs) were investigated to produce two types of cells in culture: neural cells and neural stem cells (NSCs). These types of stem cells were used as successful sources for the further treatment of central nervous system defects and injuries. The mouse bone marrow MSCs were used as the source of the stem cells in this study. β-Mercaptoethanol (BME) was used as the main inducer of the neurogenesis pathway to induce neural cells and to identify NSCs. Three types of neural markers were used: nestin as the immaturation stage marker, neurofilament light chain as the early neural marker, and microtubule-associated protein 2 as the maturation marker through different time intervals in the neurogenesis process starting from the MSCs, (as undifferentiated cells), NSCs, production stages, and toward neuron cells (as differentiated cells). The results of different exposure times to BME of the neural markers analysis done by immunocytochemistry and real time-polymerase chain reaction helped us to identify the exact timing for the neural stemness state. The results showed that the best exposure time that may be used for the production of NSCs was 6 hours. The best maintenance media for NSCs were also identified. Furthermore, we optimized exposure to BME with different times and concentrations, which could be an interesting way to modulate specific neuronal differentiation and obtain autologous neuronal phenotypes. This study was able to characterize NSCs in culture under differentiation for neurogenesis in the pathway of the neural differentiation process by studying the expressed neural genes and the ability to maintain these NSCs in culture for further differentiation in thousands of functional neurons for the treatment of brain and spinal cord injuries and defects. PMID:27143939

  3. Effects of Near-Infrared Laser on Neural Cell Activity

    NASA Astrophysics Data System (ADS)

    Mochizuki-Oda, Noriko; Kataoka, Yosky; Yamada, Hisao; Awazu, Kunio

    2004-08-01

    Near-infrared laser has been used to relieve patients from various kinds of pain caused by postherpetic neuralgesia, myofascial dysfunction, surgical and traumatic wound, cancer, and rheumatoid arthritis. Clinically, He-Ne (λ=632.8 nm, 780 nm) and Ga-Al-As (805 ± 25 nm) lasers are used to irradiate trigger points or nerve ganglion. However the precise mechanisms of such biological actions of the laser have not yet been resolved. Since laser therapy is often effective to suppress the pain caused by hyperactive excitation of sensory neurons, interactions with laser light and neural cells are suggested. As neural excitation requires large amount of energy liberated from adenosine triphosphate (ATP), we examined the effect of 830-nm laser irradiation on the energy metabolism of the rat central nervous system and isolated mitochondria from brain. The diode laser was applied for 15 min with irradiance of 4.8 W/cm2 on a 2 mm-diameter spot at the brain surface. Tissue ATP content of the irradiated area in the cerebral cortex was 19 % higher than that of the non-treated area (opposite side of the cortex), whereas the ADP content showed no significant difference. Irradiation at another wavelength (652 nm) had no effect on either ATP or ADP contents. The temperature of the brain tissue was increased 4.5 - 5.0 °C during the irradiation of both 830-nm and 652-nm laser light. Direct irradiation of the mitochondrial suspension did not show any wavelength-dependent acceleration of respiration rate nor ATP synthesis. These results suggest that the increase in tissue ATP content did not result from the thermal effect, but from specific effect of the laser operated at 830 nm. Electrophysiological studies showed the hyperpolarization of membrane potential of isolated neurons and decrease in membrane resistance with irradiation of the laser, suggesting an activation of potassium channels. Intracellular ATP is reported to regulate some kinds of potassium channels. Possible mechanisms

  4. {sub p}53-Dependent Adaptive Responses in Human Cells Exposed to Space Radiations

    SciTech Connect

    Takahashi, Akihisa; Su Xiaoming; Suzuki, Hiromi; Omori, Katsunori; Seki, Masaya; Hashizume, Toko; Shimazu, Toru; Ishioka, Noriaki; Iwasaki, Toshiyasu; Ohnishi, Takeo

    2010-11-15

    Purpose: It has been reported that priming irradiation or conditioning irradiation with a low dose of X-rays in the range of 0.02-0.1 Gy induces a p53-dependent adaptive response in mammalian cells. The aim of the present study was to clarify the effect of space radiations on the adaptive response. Methods and Materials: Two human lymphoblastoid cell lines were used; one cell line bears a wild-type p53 (wtp53) gene, and another cell line bears a mutated p53 (mp53) gene. The cells were frozen during transportation on the space shuttle and while in orbit in the International Space Station freezer for 133 days between November 15, 2008 and March 29, 2009. After the frozen samples were returned to Earth, the cells were cultured for 6 h and then exposed to a challenging X-ray-irradiation (2 Gy). Cellular sensitivity, apoptosis, and chromosome aberrations were scored using dye-exclusion assays, Hoechst33342 staining assays, and chromosomal banding techniques, respectively. Results: In cells exposed to space radiations, adaptive responses such as the induction of radioresistance and the depression of radiation-induced apoptosis and chromosome aberrations were observed in wtp53 cells but not in mp53 cells. Conclusion: These results have confirmed the hypothesis that p53-dependent adaptive responses are apparently induced by space radiations within a specific range of low doses. The cells exhibited this effect owing to space radiations exposure, even though the doses in space were very low.

  5. The novel steroidal alkaloids dendrogenin A and B promote proliferation of adult neural stem cells

    SciTech Connect

    Khalifa, Shaden A.M.; Medina, Philippe de; Erlandsson, Anna; El-Seedi, Hesham R.; Silvente-Poirot, Sandrine; Poirot, Marc

    2014-04-11

    Highlights: • Dendrogenin A and B are new aminoalkyl oxysterols. • Dendrogenins stimulated neural stem cells proliferation. • Dendrogenins induce neuronal outgrowth from neurospheres. • Dendrogenins provide new therapeutic options for neurodegenerative disorders. - Abstract: Dendrogenin A (DDA) and dendrogenin B (DDB) are new aminoalkyl oxysterols which display re-differentiation of tumor cells of neuronal origin at nanomolar concentrations. We analyzed the influence of dendrogenins on adult mice neural stem cell proliferation, sphere formation and differentiation. DDA and DDB were found to have potent proliferative effects in neural stem cells. Additionally, they induce neuronal outgrowth from neurospheres during in vitro cultivation. Taken together, our results demonstrate a novel role for dendrogenins A and B in neural stem cell proliferation and differentiation which further increases their likely importance to compensate for neuronal cell loss in the brain.

  6. Isolation and propagation of primary human and rodent embryonic neural progenitor cells and cortical neurons

    PubMed Central

    Darbinyan, Armine; Kaminski, Rafal; White, Martyn K; Darbinian, Nune; Khalili, Kamel

    2014-01-01

    Summary The research on human neural progenitor cells holds great potential for the understanding the molecular programs that control differentiation of cells of glial and neuronal lineages and pathogenetic mechanisms of neurological diseases. Stem cell technologies provide also opportunities for pharmaceutical industry to develop new approaches for regenerative medicine. Here we describe the protocol for isolation and maintenance of neural progenitor cells and cortical neurons using human fetal brain tissue. This protocol can be successfully adapted for preparation of rodent neural and oligodendrocyte progenitor cells. While several methods for isolation of neural and ologodendrocyte progenitors from rodent brain tissue have been described, including techniques which use gene transfer and magnetisc resonsnce beads, few methods are focused on derivation of human oligodendrocyte progenitor cells. Development of human culture provides the most physiologically relevent system for investigation of mechanisms which regulate function of oligodendrocyte, specifically of human origin. PMID:23975820

  7. Prolonged Treatment with Propofol Transiently Impairs Proliferation but Not Survival of Rat Neural Progenitor Cells In Vitro

    PubMed Central

    Friese, Matthew B.; Cotran, Emily; Moller, Ludde; Boyd, Justin D.; Crosby, Gregory

    2016-01-01

    Neurocognitive dysfunction is common in survivors of intensive care. Prolonged sedation has been implicated but the mechanisms are unclear. Neurogenesis continues into adulthood and is implicated in learning. The neural progenitor cells (NPC) that drive neurogenesis have receptors for the major classes of sedatives used clinically, suggesting that interruption of neurogenesis may partly contribute to cognitive decline in ICU survivors. Using an in vitro system, we tested the hypothesis that prolonged exposure to propofol concentration- and duration-dependently kills or markedly decreases the proliferation of NPCs. NPCs isolated from embryonic day 14 Sprague-Dawley rat pups were exposed to 0, 2.5, or 5.0 μg/mL of propofol, concentrations consistent with deep clinical anesthesia, for either 4 or 24 hours. Cells were assayed for cell death and proliferation either immediately following propofol exposure or 24 hours later. NPC death and apoptosis were measured by propidium iodine staining and cleaved caspase-3 immunocytochemistry, respectively, while proliferation was measured by EdU incorporation. Staurosporine (1μM for 6h) was used as a positive control for cell death. Cells were analyzed with unbiased high-throughput immunocytochemistry. There was no cell death at either concentration of propofol or duration of exposure. Neither concentration of propofol impaired NPC proliferation when exposure lasted 4 h, but when exposure lasted 24 h, propofol had an anti-proliferative effect at both concentrations (P < 0.0001, propofol vs. control). However, this effect was transient; proliferation returned to baseline 24 h after discontinuation of propofol (P = 0.37, propofol vs. control). The transient but reversible suppression of NPC proliferation, absence of cytotoxicity, and negligible effect on the neural stem cell pool pool suggest that propofol, even in concentrations used for clinical anesthesia, has limited impact on neural progenitor cell biology. PMID:27379684

  8. Prolonged Treatment with Propofol Transiently Impairs Proliferation but Not Survival of Rat Neural Progenitor Cells In Vitro.

    PubMed

    Palanisamy, Arvind; Friese, Matthew B; Cotran, Emily; Moller, Ludde; Boyd, Justin D; Crosby, Gregory; Culley, Deborah J

    2016-01-01

    Neurocognitive dysfunction is common in survivors of intensive care. Prolonged sedation has been implicated but the mechanisms are unclear. Neurogenesis continues into adulthood and is implicated in learning. The neural progenitor cells (NPC) that drive neurogenesis have receptors for the major classes of sedatives used clinically, suggesting that interruption of neurogenesis may partly contribute to cognitive decline in ICU survivors. Using an in vitro system, we tested the hypothesis that prolonged exposure to propofol concentration- and duration-dependently kills or markedly decreases the proliferation of NPCs. NPCs isolated from embryonic day 14 Sprague-Dawley rat pups were exposed to 0, 2.5, or 5.0 μg/mL of propofol, concentrations consistent with deep clinical anesthesia, for either 4 or 24 hours. Cells were assayed for cell death and proliferation either immediately following propofol exposure or 24 hours later. NPC death and apoptosis were measured by propidium iodine staining and cleaved caspase-3 immunocytochemistry, respectively, while proliferation was measured by EdU incorporation. Staurosporine (1μM for 6h) was used as a positive control for cell death. Cells were analyzed with unbiased high-throughput immunocytochemistry. There was no cell death at either concentration of propofol or duration of exposure. Neither concentration of propofol impaired NPC proliferation when exposure lasted 4 h, but when exposure lasted 24 h, propofol had an anti-proliferative effect at both concentrations (P < 0.0001, propofol vs. control). However, this effect was transient; proliferation returned to baseline 24 h after discontinuation of propofol (P = 0.37, propofol vs. control). The transient but reversible suppression of NPC proliferation, absence of cytotoxicity, and negligible effect on the neural stem cell pool pool suggest that propofol, even in concentrations used for clinical anesthesia, has limited impact on neural progenitor cell biology. PMID:27379684

  9. Pyrroloquinoline quinone against glutamate-induced neurotoxicity in cultured neural stem and progenitor cells.

    PubMed

    Guan, Shui; Xu, Jianqiang; Guo, Yifu; Ge, Dan; Liu, Tianqing; Ma, Xuehu; Cui, Zhanfeng

    2015-05-01

    Pyrroloquinoline quinone (PQQ), as a well-known redox enzyme cofactor, has been proven to play important roles in the regulation of cellular growth and development in mammals. Numerous physiological and medicinal functions of PQQ have so far been reported although its effect on neural stem and progenitor cells (NS/PCs) and the potential mechanism were even rarely investigated. In this study, the neuroprotective effects of PQQ were observed by pretreatment of NS/PCs with PQQ before glutamate injury, and the possible mechanisms were examined. PQQ stimulated cell proliferation and markedly attenuated glutamate-induced cell damage in a dose-dependent manner. By observing the nuclear morphological changes and flow cytometric analysis, PQQ pretreatment showed its significant effect on protecting NS/PCs against glutamate-induced apoptosis/necrosis. PQQ neuroprotection was associated with the decrease of intracellular reactive oxygen species (ROS) production, the increase of glutathione (GSH) levels, and the decrease of caspase-3 activity. In addition, pretreatment with PQQ also significantly enhanced the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) in the NS/PCs exposed to glutamate. These results suggest that PQQ can protect NS/PCs against glutamate toxicity associated with ROS-mediated mitochondrial pathway, indicating a useful chemical for the clinical application of NS/PCs.

  10. Intracellular transport and cell surface delivery of the neural cell adhesion molecule (NCAM).

    PubMed

    Leshchyns'ka, Iryna; Sytnyk, Vladimir

    2015-01-01

    The neural cell adhesion molecule (NCAM) regulates differentiation and functioning of neurons by accumulating at the cell surface where it mediates the interactions of neurons with the extracellular environment. NCAM also induces a number of intracellular signaling cascades, which coordinate interactions at the cell surface with intracellular processes including changes in gene expression, transport and cytoskeleton remodeling. Since NCAM functions at the cell surface, its transport and delivery to the cell surface play a critical role. Here, we review recent advances in our understanding of the molecular mechanisms of the intracellular transport and cell surface delivery of NCAM. We also discuss the data suggesting a possibility of cross talk between activation of NCAM at the cell surface and the intracellular transport and cell surface delivery of NCAM.

  11. RKIP Regulates Neural Cell Apoptosis Induced by Exposure to Microwave Radiation Partly Through the MEK/ERK/CREB Pathway.

    PubMed

    Zuo, Hongyan; Lin, Tao; Wang, Dewen; Peng, Ruiyun; Wang, Shuiming; Gao, Yabing; Xu, Xinping; Zhao, Li; Wang, Shaoxia; Su, Zhentao

    2015-01-01

    In the present study, we investigated whether Raf-1 kinase inhibitory protein (RKIP) is important for neural cell apoptosis induced by microwave exposure and explored the role of MEK/ERK/CREB pathway regulated by RKIP in the apoptosis. Differentiated PC12 cells were exposed to continuous microwave radiation at 2.856 GHz for 5 min with average power density of 30 mW/cm(2). RKIP sense and anti-sense recombinant plasmids were constructed and transfected into PC12 cells, respectively. Terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) staining and caspase-3 activity assay were used to detect cell apoptosis. The results showed that RKIP was downregulated after microwave exposure while the MEK/ERK/CREB signaling pathway was activated excessively. Moreover, the ratio of Bcl-2/Bax decreased, activity of caspase-3 increased, and thus apoptotic DNA fragmentation increased. RKIP overexpression significantly inhibited the phosphorylation of MEK, ERK, and CREB, while RKIP downregulation had the reverse effect. Furthermore, U0126 was found to antagonize the changes caused by RKIP downregulation after exposure to radiation. In conclusion, RKIP plays an important role in the neural cell apoptosis induced by microwave radiation, and the regulation of cell apoptosis by RKIP is partly through the MEK/ERK/CREB pathway. This suggests that RKIP may act as a key regulator of neuronal damage caused by microwave radiation.

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

    PubMed

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

    2016-02-01

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

  13. Signal transduction of the physical environment in the neural differentiation of stem cells

    PubMed Central

    Thompson, Ryan; Chan, Christina

    2016-01-01

    Neural differentiation is largely dependent on extracellular signals within the cell microenvironment. These extracellular signals are mainly in the form of soluble factors that activate intracellular signaling cascades that drive changes in the cell nucleus. However, it is becoming increasingly apparent that the physical microenvironment provides signals that can also influence lineage commitment and very low modulus surfaces has been repeatedly demonstrated to promote neurogenesis. The molecular mechanisms governing mechano-induced neural differentiation are still largely uncharacterized; however, a growing body of evidence indicates that physical stimuli can regulate known signaling cascades and transcription factors involved in neural differentiation. Understanding how the physical environment affects neural differentiation at the molecular level will enable research and design of materials that will eventually enhance neural stem cell (NSC) differentiation, homogeneity and specificity. PMID:27785459

  14. Twist1 Controls a Cell-Specification Switch Governing Cell Fate Decisions within the Cardiac Neural Crest

    PubMed Central

    Vincentz, Joshua W.; Firulli, Beth A.; Lin, Andrea; Spicer, Douglas B.; Howard, Marthe J.; Firulli, Anthony B.

    2013-01-01

    Neural crest cells are multipotent progenitor cells that can generate both ectodermal cell types, such as neurons, and mesodermal cell types, such as smooth muscle. The mechanisms controlling this cell fate choice are not known. The basic Helix-loop-Helix (bHLH) transcription factor Twist1 is expressed throughout the migratory and post-migratory cardiac neural crest. Twist1 ablation or mutation of the Twist-box causes differentiation of ectopic neuronal cells, which molecularly resemble sympathetic ganglia, in the cardiac outflow tract. Twist1 interacts with the pro-neural factor Sox10 via its Twist-box domain and binds to the Phox2b promoter to repress transcriptional activity. Mesodermal cardiac neural crest trans-differentiation into ectodermal sympathetic ganglia-like neurons is dependent upon Phox2b function. Ectopic Twist1 expression in neural crest precursors disrupts sympathetic neurogenesis. These data demonstrate that Twist1 functions in post-migratory neural crest cells to repress pro-neural factors and thereby regulate cell fate determination between ectodermal and mesodermal lineages. PMID:23555309

  15. 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

  16. Single-Cell Phenotype Classification Using Deep Convolutional Neural Networks.

    PubMed

    Dürr, Oliver; Sick, Beate

    2016-10-01

    Deep learning methods are currently outperforming traditional state-of-the-art computer vision algorithms in diverse applications and recently even surpassed human performance in object recognition. Here we demonstrate the potential of deep learning methods to high-content screening-based phenotype classification. We trained a deep learning classifier in the form of convolutional neural networks with approximately 40,000 publicly available single-cell images from samples treated with compounds from four classes known to lead to different phenotypes. The input data consisted of multichannel images. The construction of appropriate feature definitions was part of the training and carried out by the convolutional network, without the need for expert knowledge or handcrafted features. We compare our results against the recent state-of-the-art pipeline in which predefined features are extracted from each cell using specialized software and then fed into various machine learning algorithms (support vector machine, Fisher linear discriminant, random forest) for classification. The performance of all classification approaches is evaluated on an untouched test image set with known phenotype classes. Compared to the best reference machine learning algorithm, the misclassification rate is reduced from 8.9% to 6.6%.

  17. Neural stem/progenitor cells in Alzheimer’s disease

    PubMed Central

    Tincer, Gizem; Mashkaryan, Violeta; Bhattarai, Prabesh; Kizil, Caghan

    2016-01-01

    Alzheimer’s disease (AD) is the most prevalent neurodegenerative disease and a worldwide health challenge. Different therapeutic approaches are being developed to reverse or slow the loss of affected neurons. Another plausible therapeutic way that may complement the studies is to increase the survival of existing neurons by mobilizing the existing neural stem/progenitor cells (NSPCs) — i.e. “induce their plasticity” — to regenerate lost neurons despite the existing pathology and unfavorable environment. However, there is controversy about how NSPCs are affected by the unfavorable toxic environment during AD. In this review, we will discuss the use of stem cells in neurodegenerative diseases and in particular how NSPCs affect the AD pathology and how neurodegeneration affects NSPCs. In the end of this review, we will discuss how zebrafish as a useful model organism with extensive regenerative ability in the brain might help to address the molecular programs needed for NSPCs to respond to neurodegeneration by enhanced neurogenesis. PMID:27505014

  18. 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

  19. Systems biology approach to imaging of neural stem cells.

    PubMed

    Ma, Li Hua; Li, Yao; Djurić, Petar M; Maletić-Savatić, Mirjana

    2011-01-01

    Over the past decade, the advances in human brain magnetic resonance imaging (MRI) have significantly improved our ability to gain insightful information about the structure and function of the brain. One of the MRI imaging modalities that still awaits more comprehensive data mining is magnetic resonance spectroscopy (MRS). MRS provides information on the functional status of the brain tissue and can detect metabolic abnormalities that precede structural changes. The chemical specificity of proton MRS ((1)H-MRS) allows detection of several biomarkers that are specific for neurons (N-acetyl aspartate, NAA) and astrocytes (myoinositol (mI) and choline (Cho)), the two most abundant cell types present in the brain tissue. However, apart from a dozen metabolites, current methodologies utilized for MRS analysis do not allow further biomarker discoveries. Herein, we introduce a bioinformatics approach to MRS data processing and discuss possible discoveries that such approach may provide. Specifically, we describe the methodology for neural stem/progenitor cell (NPC) detection in vitro and in vivo, utilizing metabolomic profiling and singular value decomposition analyses. PMID:21279615

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

    PubMed Central

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

    2015-01-01

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

  1. Use of Microfluidic Technology to Monitor the Differentiation and Migration of Human ESC-Derived Neural Cells.

    PubMed

    Bae, Jiwoo; Lee, Nayeon; Choi, Wankyu; Lee, Suji; Ko, Jung Jae; Han, Baek Soo; Lee, Sang Chul; Jeon, Noo Li; Song, Jihwan

    2016-01-01

    Microfluidics forms the basis of unique experimental approaches that visualize the development of neural structure using micro-scale devices and aids the guidance of neurite growth in an axonal isolation compartment. We utilized microfluidics technology to monitor the differentiation and migration of neural cells derived from human embryonic stems cells (hESC). We cocultured hESC with PA6 stromal cells and isolated neural rosette-like structures, which subsequently formed neurospheres in a suspension culture. We found that Tuj1-positive neural cells but not nestin-positive neural precursor cells (NPC) were able to enter the microfluidics grooves (microchannels), suggesting a neural cell-migratory capacity that was dependent on neuronal differentiation. We also showed that bundles of axons formed and extended into the microchannels.Taken together, these results demonstrated that microfluidics technology can provide useful tools to study neurite outgrowth and axon guidance of neural cells, which are derived from human embryonic stem cells. PMID:27062598

  2. Polysialic Acid Directs Tumor Cell Growth by Controlling Heterophilic Neural Cell Adhesion Molecule Interactions

    PubMed Central

    Seidenfaden, Ralph; Krauter, Andrea; Schertzinger, Frank; Gerardy-Schahn, Rita; Hildebrandt, Herbert

    2003-01-01

    Polysialic acid (PSA), a carbohydrate polymer attached to the neural cell adhesion molecule (NCAM), promotes neural plasticity and tumor malignancy, but its mode of action is controversial. Here we establish that PSA controls tumor cell growth and differentiation by interfering with NCAM signaling at cell-cell contacts. Interactions between cells with different PSA and NCAM expression profiles were initiated by enzymatic removal of PSA and by ectopic expression of NCAM or PSA-NCAM. Removal of PSA from the cell surface led to reduced proliferation and activated extracellular signal-regulated kinase (ERK), inducing enhanced survival and neuronal differentiation of neuroblastoma cells. Blocking with an NCAM-specific peptide prevented these effects. Combinatorial transinteraction studies with cells and membranes with different PSA and NCAM phenotypes revealed that heterophilic NCAM binding mimics the cellular responses to PSA removal. In conclusion, our data demonstrate that PSA masks heterophilic NCAM signals, having a direct impact on tumor cell growth. This provides a mechanism for how PSA may promote the genesis and progression of highly aggressive PSA-NCAM-positive tumors. PMID:12897159

  3. T-cell activation in pulmonary lymph nodes of mice exposed to ozone

    SciTech Connect

    Dziedzic, D.; White, H.J.

    1985-12-01

    Groups of Cd-1 female mice were exposed to ozone at 0.3, 0.5, and 0.7 ppm, 20 hr per day, 7 days per week for 1-28 days. The effect of ozone exposure on lymphoid cells was determined by studying mediastinal lymph nodes at various times of exposure. It was found that lymphocyte numbers underwent a dose-dependent, four-phased change:cellular depletion (Days 1-2), followed by rapid hyperplasia (Days 3-4), incremental cell number reduction (Days 5-7), and a subsequent subacute phase of elevated lymphocyte numbers (Days 8-28). Using tritiated thymidine it was determined that cells underwent a rapid burst of division by Day 3 of exposure and that mitosis subsequently declined to near baseline values by 2 weeks of exposure. Autoradiographic analysis of histologic sections revealed that the paracortical T-cell areas of the nodes were particularly involved. In addition to the increase in thymidine uptake, several morphologic changes were evident in affected cells. By comparison, the B cells from ozone-exposed animals were virtually unaffected with respect to cell division or morphological alterations. Prior treatment of ozone-exposed animals with a monoclonal antibody that is cytotoxic for T cells eliminated the hyperplastic response. Immunologic aspects of T-cell reactivity were studied. T-cell responsiveness to mitogenic stimulation with concanavalin A showed little alteration during the first days of exposure; however, by Day 14 an increase in reactivity was observed. This change indicated that functional lymphocyte stimulation occurred during ozone exposure.

  4. Analysis of T-cell proliferation and cytokine secretion in the individuals exposed to arsenic.

    PubMed

    Biswas, R; Ghosh, P; Banerjee, N; Das, J K; Sau, T; Banerjee, A; Roy, S; Ganguly, S; Chatterjee, M; Mukherjee, A; Giri, A K

    2008-05-01

    Over six million people in nine districts of West Bengal, India are exposed to very high levels of arsenic primarily through their drinking water. More than 300,000 people showed arsenic-induced skin lesions in these districts. This is regarded as the greatest arsenic calamity in the world. Chronic arsenicosis causes varied dermatological signs ranging from pigmentation changes, hyperkeratosis to non-melanocytic cancer of skin, and also malignancies in different internal organs. Higher incidences of opportunistic infections are found in the arsenic-exposed individuals, indicating that their immune systems may be impaired somehow. We have thus investigated the effect of arsenic on T-cell proliferation and cytokine secretion in 20 individuals with arsenic-induced skin lesions and compared the results with 18 arsenic-unexposed individuals. A marked dose-dependent suppression of Concanavalin A (Con A) induced T-cell proliferation was observed in the arsenic-exposed individuals compared with the unexposed (P < 0.001) individuals. This correlated with a significant decrease in the levels of secreted cytokines by the T cells (TNF-alpha, IFN-gamma, IL2, IL10, IL5, and IL4) in the exposed individuals (P < 0.001). Thus it can be inferred that arsenic exposure can cause immunosuppression in humans.

  5. In vitro metabolism study of normal and tumor cells when exposed to red LED light

    NASA Astrophysics Data System (ADS)

    Stolbovskaya, Olga V.; Khairullin, Radik M.; Saenko, Yuri V.; Krasnikova, Ekaterina S.; Krasnikov, Aleksandr V.; Fomin, Aleksandr A.; Skaptsov, Aleksandr A.

    2016-04-01

    This work presents the results of studying the mitochondrial membrane potential, intracellular ROS, peculiarities of the cell cycle of cancer cells HCT-116 and the normal line of CHO cells when exposed to the red LED light with a wavelength range of 0.620-0.680 μm. A dose-dependent increase in mitochondrial membrane potential and intracellular ROS concentration in cancer cells HCT-116 was established. In normal CHO cell line a dose-dependent reduction of mitochondrial membrane potential and dose-dependent increase in intracellular ROS occur. It has been shown that the sensitivity of the studied cell lines to the red light depends on the stage of the cell cycle.

  6. PSA-NCAM-negative neural crest cells emerging during neural induction of pluripotent stem cells cause mesodermal tumors and unwanted grafts.

    PubMed

    Lee, Dongjin R; Yoo, Jeong-Eun; Lee, Jae Souk; Park, Sanghyun; Lee, Junwon; Park, Chul-Yong; Ji, Eunhyun; Kim, Han-Soo; Hwang, Dong-Youn; Kim, Dae-Sung; Kim, Dong-Wook

    2015-05-12

    Tumorigenic potential of human pluripotent stem cells (hPSCs) is an important issue in clinical applications. Despite many efforts, PSC-derived neural precursor cells (NPCs) have repeatedly induced tumors in animal models even though pluripotent cells were not detected. We found that polysialic acid-neural cell adhesion molecule (PSA-NCAM)(-) cells among the early NPCs caused tumors, whereas PSA-NCAM(+) cells were nontumorigenic. Molecular profiling, global gene analysis, and multilineage differentiation of PSA-NCAM(-) cells confirm that they are multipotent neural crest stem cells (NCSCs) that could differentiate into both ectodermal and mesodermal lineages. Transplantation of PSA-NCAM(-) cells in a gradient manner mixed with PSA-NCAM(+) cells proportionally increased mesodermal tumor formation and unwanted grafts such as PERIPHERIN(+) cells or pigmented cells in the rat brain. Therefore, we suggest that NCSCs are a critical target for tumor prevention in hPSC-derived NPCs, and removal of PSA-NCAM(-) cells eliminates the tumorigenic potential originating from NCSCs after transplantation. PMID:25937368

  7. Transcriptional profiling of adult neural stem-like cells from the human brain.

    PubMed

    Sandberg, Cecilie Jonsgar; Vik-Mo, Einar O; Behnan, Jinan; Helseth, Eirik; Langmoen, Iver A

    2014-01-01

    There is a great potential for the development of new cell replacement strategies based on adult human neural stem-like cells. However, little is known about the hierarchy of cells and the unique molecular properties of stem- and progenitor cells of the nervous system. Stem cells from the adult human brain can be propagated and expanded in vitro as free floating neurospheres that are capable of self-renewal and differentiation into all three cell types of the central nervous system. Here we report the first global gene expression study of adult human neural stem-like cells originating from five human subventricular zone biopsies (mean age 42, range 33-60). Compared to adult human brain tissue, we identified 1,189 genes that were significantly up- and down-regulated in adult human neural stem-like cells (1% false discovery rate). We found that adult human neural stem-like cells express stem cell markers and have reduced levels of markers that are typical of the mature cells in the nervous system. We report that the genes being highly expressed in adult human neural stem-like cells are associated with developmental processes and the extracellular region of the cell. The calcium signaling pathway and neuroactive ligand-receptor interactions are enriched among the most differentially regulated genes between adult human neural stem-like cells and adult human brain tissue. We confirmed the expression of 10 of the most up-regulated genes in adult human neural stem-like cells in an additional sample set that included adult human neural stem-like cells (n = 6), foetal human neural stem cells (n = 1) and human brain tissues (n = 12). The NGFR, SLITRK6 and KCNS3 receptors were further investigated by immunofluorescence and shown to be heterogeneously expressed in spheres. These receptors could potentially serve as new markers for the identification and characterisation of neural stem- and progenitor cells or as targets for manipulation of cellular fate.

  8. Transcriptional Profiling of Adult Neural Stem-Like Cells from the Human Brain

    PubMed Central

    Sandberg, Cecilie Jonsgar; Vik-Mo, Einar O.; Behnan, Jinan; Helseth, Eirik; Langmoen, Iver A.

    2014-01-01

    There is a great potential for the development of new cell replacement strategies based on adult human neural stem-like cells. However, little is known about the hierarchy of cells and the unique molecular properties of stem- and progenitor cells of the nervous system. Stem cells from the adult human brain can be propagated and expanded in vitro as free floating neurospheres that are capable of self-renewal and differentiation into all three cell types of the central nervous system. Here we report the first global gene expression study of adult human neural stem-like cells originating from five human subventricular zone biopsies (mean age 42, range 33–60). Compared to adult human brain tissue, we identified 1,189 genes that were significantly up- and down-regulated in adult human neural stem-like cells (1% false discovery rate). We found that adult human neural stem-like cells express stem cell markers and have reduced levels of markers that are typical of the mature cells in the nervous system. We report that the genes being highly expressed in adult human neural stem-like cells are associated with developmental processes and the extracellular region of the cell. The calcium signaling pathway and neuroactive ligand-receptor interactions are enriched among the most differentially regulated genes between adult human neural stem-like cells and adult human brain tissue. We confirmed the expression of 10 of the most up-regulated genes in adult human neural stem-like cells in an additional sample set that included adult human neural stem-like cells (n = 6), foetal human neural stem cells (n = 1) and human brain tissues (n = 12). The NGFR, SLITRK6 and KCNS3 receptors were further investigated by immunofluorescence and shown to be heterogeneously expressed in spheres. These receptors could potentially serve as new markers for the identification and characterisation of neural stem- and progenitor cells or as targets for manipulation of cellular fate. PMID

  9. Hydrogel formulation determines cell fate of fetal and adult neural progenitor cells.

    PubMed

    Aurand, Emily R; Wagner, Jennifer L; Shandas, Robin; Bjugstad, Kimberly B

    2014-01-01

    Hydrogels provide a unique tool for neural tissue engineering. These materials can be customized for certain functions, i.e. to provide cell/drug delivery or act as a physical scaffold. Unfortunately, hydrogel complexities can negatively impact their biocompatibility, resulting in unintended consequences. These adverse effects may be combated with a better understanding of hydrogel chemical, physical, and mechanical properties, and how these properties affect encapsulated neural cells. We defined the polymerization and degradation rates and compressive moduli of 25 hydrogels formulated from different concentrations of hyaluronic acid (HA) and poly(ethylene glycol) (PEG). Changes in compressive modulus were driven primarily by the HA concentration. The in vitro biocompatibility of fetal-derived (fNPC) and adult-derived (aNPC) neural progenitor cells was dependent on hydrogel formulation. Acute survival of fNPC benefited from hydrogel encapsulation. NPC differentiation was divergent: fNPC differentiated into mostly glial cells, compared with neuronal differentiation of aNPC. Differentiation was influenced in part by the hydrogel mechanical properties. This study indicates that there can be a wide range of HA and PEG hydrogels compatible with NPC. Additionally, this is the first study comparing hydrogel encapsulation of NPC derived from different aged sources, with data suggesting that fNPC and aNPC respond dissimilarly within the same hydrogel formulation.

  10. 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

  11. Using magnetic nanoparticles for gene transfer to neural stem cells: stem cell propagation method influences outcomes.

    PubMed

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

    2015-04-24

    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.

  12. Mature natural killer cells reset their responsiveness when exposed to an altered MHC environment

    PubMed Central

    Joncker, Nathalie T.; Shifrin, Nataliya; Delebecque, Frédéric

    2010-01-01

    Some mature natural killer (NK) cells cannot be inhibited by major histocompatibility complex (MHC) I molecules, either because they lack corresponding inhibitory receptors or because the host lacks the corresponding MHC I ligands for the receptors. Such NK cells nevertheless remain self-tolerant and exhibit a generalized hyporesponsiveness to stimulation through activating receptors. To address whether NK cell responsiveness is set only during the NK cell differentiation process, we transferred mature NK cells from wild-type (WT) to MHC I–deficient hosts or vice versa. Remarkably, mature responsive NK cells from WT mice became hyporesponsive after transfer to MHC I–deficient mice, whereas mature hyporesponsive NK cells from MHC I–deficient mice became responsive after transfer to WT mice. Altered responsiveness was evident among mature NK cells that had not divided in the recipient animals, indicating that the cells were mature before transfer and that alterations in activity did not require cell division. Furthermore, the percentages of NK cells expressing KLRG1, CD11b, CD27, and Ly49 receptors specific for H-2b were not markedly altered after transfer. Thus, the functional activity of mature NK cells can be reset when the cells are exposed to a changed MHC environment. These findings have important implications for how NK cell functions may be curtailed or enhanced in the context of disease. PMID:20819928

  13. Establishment of long term cultures of neural stem cells from adult sea bass, Dicentrarchus labrax.

    PubMed

    Servili, Arianna; Bufalino, Mary Rose; Nishikawa, Ryuhei; Sanchez de Melo, Ivan; Muñoz-Cueto, Jose A; Lee, Lucy E J

    2009-02-01

    Long term cell cultures could be obtained from brains of adult sea bass (Dicentrarchus labrax) up to 5 days post mortem. On three different occasions, sea bass brain tissues were dissected, dispersed and cultured in Leibovitz's L-15 media supplemented with 10% fetal bovine serum. The resulting cellular preparations could be passaged within 2 or 3 weeks of growth. The neural cells derived from the first trial (SBB-W1) have now been passaged over 24 times within two years. These cells have been cryopreserved and thawed successfully. SBB-W1 cells are slow growing with doubling times requiring at least 7 days at 22 degrees C. These long term cell cultures could be grown in suspension as neurospheres that were immunopositive for nestin, a marker for neural stem cells, or grown as adherent monolayers displaying both glial and neural morphologies. Immunostaining with anti-glial fibrillary acidic protein (a glial marker) and anti-neurofilament (a neuronal marker), yielded positive staining in most cells, suggesting their possible identity as neural stem cells. Furthermore, Sox 2, a marker for neural stem cells, could be detected from these cell extracts as well as proliferating cell nuclear antigen, a marker for proliferating cells. SBB-W1 could be transfected using pEGFP-N1 indicating their viability and suitability as convenient models for neurophysiological or neurotoxicological studies.

  14. Accelerating bioelectric functional development of neural stem cells by graphene coupling: Implications for neural interfacing with conductive materials.

    PubMed

    Guo, Rongrong; Zhang, Shasha; Xiao, Miao; Qian, Fuping; He, Zuhong; Li, Dan; Zhang, Xiaoli; Li, Huawei; Yang, Xiaowei; Wang, Ming; Chai, Renjie; Tang, Mingliang

    2016-11-01

    In order to govern cell-specific behaviors in tissue engineering for neural repair and regeneration, a better understanding of material-cell interactions, especially the bioelectric functions, is extremely important. Graphene has been reported to be a potential candidate for use as a scaffold and neural interfacing material. However, the bioelectric evolvement of cell membranes on these conductive graphene substrates remains largely uninvestigated. In this study, we used a neural stem cell (NSC) model to explore the possible changes in membrane bioelectric properties - including resting membrane potentials and action potentials - and cell behaviors on graphene films under both proliferation and differentiation conditions. We used a combination of single-cell electrophysiological recordings and traditional cell biology techniques. Graphene did not affect the basic membrane electrical parameters (capacitance and input resistance), but resting membrane potentials of cells on graphene substrates were more strongly negative under both proliferation and differentiation conditions. Also, NSCs and their progeny on graphene substrates exhibited increased firing of action potentials during development compared to controls. However, graphene only slightly affected the electric characterizations of mature NSC progeny. The modulation of passive and active bioelectric properties on the graphene substrate was accompanied by enhanced NSC differentiation. Furthermore, spine density, synapse proteins expressions and synaptic activity were all increased in graphene group. Modeling of the electric field on conductive graphene substrates suggests that the electric field produced by the electronegative cell membrane is much higher on graphene substrates than that on control, and this might explain the observed changes of bioelectric development by graphene coupling. Our results indicate that graphene is able to accelerate NSC maturation during development, especially with regard to

  15. Accelerating bioelectric functional development of neural stem cells by graphene coupling: Implications for neural interfacing with conductive materials.

    PubMed

    Guo, Rongrong; Zhang, Shasha; Xiao, Miao; Qian, Fuping; He, Zuhong; Li, Dan; Zhang, Xiaoli; Li, Huawei; Yang, Xiaowei; Wang, Ming; Chai, Renjie; Tang, Mingliang

    2016-11-01

    In order to govern cell-specific behaviors in tissue engineering for neural repair and regeneration, a better understanding of material-cell interactions, especially the bioelectric functions, is extremely important. Graphene has been reported to be a potential candidate for use as a scaffold and neural interfacing material. However, the bioelectric evolvement of cell membranes on these conductive graphene substrates remains largely uninvestigated. In this study, we used a neural stem cell (NSC) model to explore the possible changes in membrane bioelectric properties - including resting membrane potentials and action potentials - and cell behaviors on graphene films under both proliferation and differentiation conditions. We used a combination of single-cell electrophysiological recordings and traditional cell biology techniques. Graphene did not affect the basic membrane electrical parameters (capacitance and input resistance), but resting membrane potentials of cells on graphene substrates were more strongly negative under both proliferation and differentiation conditions. Also, NSCs and their progeny on graphene substrates exhibited increased firing of action potentials during development compared to controls. However, graphene only slightly affected the electric characterizations of mature NSC progeny. The modulation of passive and active bioelectric properties on the graphene substrate was accompanied by enhanced NSC differentiation. Furthermore, spine density, synapse proteins expressions and synaptic activity were all increased in graphene group. Modeling of the electric field on conductive graphene substrates suggests that the electric field produced by the electronegative cell membrane is much higher on graphene substrates than that on control, and this might explain the observed changes of bioelectric development by graphene coupling. Our results indicate that graphene is able to accelerate NSC maturation during development, especially with regard to

  16. 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

  17. The endocannabinoid system promotes astroglial differentiation by acting on neural progenitor cells.

    PubMed

    Aguado, Tania; Palazuelos, Javier; Monory, Krisztina; Stella, Nephi; Cravatt, Benjamin; Lutz, Beat; Marsicano, Giovanni; Kokaia, Zaal; Guzmán, Manuel; Galve-Roperh, Ismael

    2006-02-01

    Endocannabinoids exert an important neuromodulatory role via presynaptic cannabinoid CB1 receptors and may also participate in the control of neural cell death and survival. The function of the endocannabinoid system has been extensively studied in differentiated neurons, but its potential role in neural progenitor cells remains to be elucidated. Here we show that the CB1 receptor and the endocannabinoid-inactivating enzyme fatty acid amide hydrolase are expressed, both in vitro and in vivo, in postnatal radial glia (RC2+ cells) and in adult nestin type I (nestin(+)GFAP+) neural progenitor cells. Cell culture experiments show that CB1 receptor activation increases progenitor proliferation and differentiation into astroglial cells in vitro. In vivo analysis evidences that, in postnatal CB1(-/-) mouse brain, progenitor proliferation and astrogliogenesis are impaired. Likewise, in adult CB1-deficient mice, neural progenitor proliferation is decreased but is increased in fatty acid amide hydrolase-deficient mice. In addition, endocannabinoid signaling controls neural progenitor differentiation in the adult brain by promoting astroglial differentiation of newly born cells. These results show a novel physiological role of endocannabinoids, which constitute a new family of signaling cues involved in the regulation of neural progenitor cell function.

  18. Ca2+ ion transport through patch-clamped cells exposed to magnetic fields.

    PubMed

    Höjevik, P; Sandblom, J; Galt, S; Hamnerius, Y

    1995-01-01

    The total current of Ca2+ ions through patch-clamped cell membranes was measured while exposing clonal insulin-producing beta-cells (RINm5F) to a combination of DC and AC magnetic fields at so-called cyclotron resonance conditions. Previous experimental evidence supports the theory that a resonant interaction between magnetic fields and organisms can exist. This experiment was designed to test one possible site of interaction: channels in the cell membrane. The transport of Ca2+ ions through the protein channels of the plasma membrane did not show any resonant behavior in the frequency range studied. PMID:7748201

  19. Zebrafish neural tube morphogenesis requires Scribble-dependent oriented cell divisions.

    PubMed

    Žigman, Mihaela; Trinh, Le A; Fraser, Scott E; Moens, Cecilia B

    2011-01-11

    How control of subcellular events in single cells determines morphogenesis on the scale of the tissue is largely unresolved. The stereotyped cross-midline mitoses of progenitors in the zebrafish neural keel provide a unique experimental paradigm for defining the role and control of single-cell orientation for tissue-level morphogenesis in vivo. We show here that the coordinated orientation of individual progenitor cell division in the neural keel is the cellular determinant required for morphogenesis into a neural tube epithelium with a single straight lumen. We find that Scribble is required for oriented cell division and that its function in this process is independent of canonical apicobasal and planar polarity pathways. We identify a role for Scribble in controlling clustering of α-catenin foci in dividing progenitors. Loss of either Scrib or N-cadherin results in abnormally oriented mitoses, reduced cross-midline cell divisions, and similar neural tube defects. We propose that Scribble-dependent nascent cell-cell adhesion clusters between neuroepithelial progenitors contribute to define orientation of their cell division. Finally, our data demonstrate that while oriented mitoses of individual cells determine neural tube architecture, the tissue can in turn feed back on its constituent cells to define their polarization and cell division orientation to ensure robust tissue morphogenesis.

  20. Arsenic-induced mitochondrial instability leading to programmed cell death in the exposed individuals.

    PubMed

    Banerjee, Nilanjana; Banerjee, Mayukh; Ganguly, Sudipto; Bandyopadhyay, Santu; Das, Jayanta K; Bandyopadhay, Apurba; Chatterjee, Mitali; Giri, Ashok K

    2008-04-18

    In West Bengal, India, more than 6 million people in nine districts are exposed to arsenic through drinking water. It is regarded as the greatest arsenic calamity in the world. Arsenic is a well-documented human carcinogen, which does not induce cancer in any other animal model. Interestingly, at lower concentrations, arsenic is known to induce apoptosis in various cancer cell lines in vitro. We have studied apoptosis in human peripheral blood mononuclear cells (PBMC) of 30 arsenic exposed skin lesion individuals by annexin V-FITC staining and compared with 28 unexposed individuals. The percentage of apoptotic cells in individuals with skin lesions was significantly higher (p<0.001) in comparison to unexposed individuals. In the exposed individuals with skin lesions, there were elevated levels of intracellular reactive oxygen species (ROS), mitochondrial membrane permeability and increased cytochrome c release, leading to increased downstream caspase activity. Arsenic-induced DNA damage was confirmed by DNA ladder formation and confocal microscopy. We also observed that chronic arsenic exposure reduced Bcl-2/Bax ratio and also resulted in cell cycle arrest of PBMC in G0/G1 phase. All these observations indicate that mitochondria-mediated pathway may be responsible for arsenic-induced apoptosis.

  1. α1-Antitrypsin reduces rhinovirus infection in primary human airway epithelial cells exposed to cigarette smoke

    PubMed Central

    Berman, Reena; Jiang, Di; Wu, Qun; Chu, Hong Wei

    2016-01-01

    Human rhinovirus (HRV) infections target airway epithelium and are the leading cause of acute exacerbations of COPD. Cigarette smoke (CS) increases the severity of viral infections, but there is no effective therapy for HRV infection. We determined whether α1-antitrypsin (A1AT) reduces HRV-16 infection in CS-exposed primary human airway epithelial cells. Brushed bronchial epithelial cells from normal subjects and patients diagnosed with COPD were cultured at air–liquid interface to induce mucociliary differentiation. These cells were treated with A1AT or bovine serum albumin for 2 hours and then exposed to air or whole cigarette smoke (WCS) with or without HRV-16 (5×104 50% Tissue Culture Infective Dose [TCID50]/transwell) infection for 24 hours. WCS exposure significantly increased viral load by an average of fivefold and decreased the expression of antiviral genes interferon-λ1, OAS1, and MX1. When A1AT was added to WCS-exposed cells, viral load significantly decreased by an average of 29-fold. HRV-16 infection significantly increased HRV-16 receptor intercellular adhesion molecule-1 messenger RNA expression in air-exposed cells, which was decreased by A1AT. A1AT-mediated reduction of viral load was not accompanied by increased epithelial antiviral gene expression or by inhibiting the activity of 3C protease involved in viral replication or maturation. Our findings demonstrate that A1AT treatment prevents a WCS-induced increase in viral load and for the first time suggest a therapeutic effect of A1AT on HRV infection. PMID:27354786

  2. Pitx2 expression promotes p21 expression and cell cycle exit in neural stem cells.

    PubMed

    Heldring, Nina; Joseph, Bertrand; Hermanson, Ola; Kioussi, Chrissa

    2012-11-01

    Cortical development is a complex process that involves many events including proliferation, cell cycle exit and differentiation that need to be appropriately synchronized. Neural stem cells (NSCs) isolated from embryonic cortex are characterized by their ability of self-renewal under continued maintenance of multipotency. Cell cycle progression and arrest during development is regulated by numerous factors, including cyclins, cyclin dependent kinases and their inhibitors. In this study, we exogenously expressed the homeodomain transcription factor Pitx2, usually expressed in postmitotic progenitors and neurons of the embryonic cortex, in NSCs with low expression of endogenous Pitx2. We found that Pitx2 expression induced a rapid decrease in proliferation associated with an accumulation of NSCs in G1 phase. A search for potential cell cycle inhibitors responsible for such cell cycle exit of NSCs revealed that Pitx2 expression caused a rapid and dramatic (≉20-fold) increase in expression of the cell cycle inhibitor p21 (WAF1/Cip1). In addition, Pitx2 bound directly to the p21 promoter as assessed by chromatin immunoprecipitation (ChIP) in NSCs. Surprisingly, Pitx2 expression was not associated with an increase in differentiation markers, but instead the expression of nestin, associated with undifferentiated NSCs, was maintained. Our results suggest that Pitx2 promotes p21 expression and induces cell cycle exit in neural progenitors.

  3. Sequential Differentiation of Embryonic Stem Cells into Neural Epithelial-Like Stem Cells and Oligodendrocyte Progenitor Cells

    PubMed Central

    Bian, Jing; Zheng, Jiao; Li, Shen; Luo, Lan; Ding, Fei

    2016-01-01

    Background Recent advances in stem cell technology afford an unlimited source of neural progenitors and glial cells for cell based therapy in central nervous system (CNS) disorders. However, current differentiation strategies still need to be improved due to time-consuming processes, poorly defined culture conditions, and low yield of target cell populations. Methodology/Principle Findings This study aimed to provide a precise sequential differentiation to capture two transient stages: neural epithelia-like stem cells (NESCs) and oligodendrocytes progenitor cells (OPCs) derived from mouse embryonic stem cells (ESCs). CHIR99021, a glycogen synthase kinase 3 (GSK-3) inhibitor, in combination with dual SMAD inhibitors, could induce ESCs to rapidly differentiate into neural rosette-like colonies, which facilitated robust generation of NESCs that had a high self-renewal capability and stable neuronal and glial differentiation potentials. Furthermore, SHH combined with FGF-2 and PDGF-AA could induce NESCs to differentiate into highly expandable OPCs. These OPCs not only robustly differentiated into oligodendrocytes, but also displayed an increased migratory activity in vitro. Conclusions/Significance We developed a precise and reliable strategy for sequential differentiation to capture NESCs and OPCs derived from ESCs, thus providing unlimited cell source for cell transplantation and drug screening towards CNS repair. PMID:27192219

  4. Endometrial stem cell transplantation in MPTP- exposed primates: an alternative cell source for treatment of Parkinson's disease.

    PubMed

    Wolff, Erin F; Mutlu, Levent; Massasa, Efi E; Elsworth, John D; Eugene Redmond, D; Taylor, Hugh S

    2015-01-01

    Parkinson's disease (PD) is a neurodegenerative disease caused by the loss of dopaminergic neurons in the substantia nigra. Cell-replacement therapies have emerged as a promising strategy to slow down or replace neuronal loss. Compared to other stem cell types, endometrium-derived stem cells (EDSCs) are an attractive source of stem cells for cellular therapies because of their ease of collection and vast differentiation potential. Here we demonstrate that endometrium-derived stem cells may be transplanted into an MPTP exposed monkey model of PD. After injection into the striatum, endometrium-derived stem cells engrafted, exhibited neuron-like morphology, expressed tyrosine hydroxylase (TH) and increased the numbers of TH positive cells on the transplanted side and dopamine metabolite concentrations in vivo. Our results suggest that endometrium-derived stem cells may provide a therapeutic benefit in the primate model of PD and may be used in stem cell based therapies.

  5. A galvanotaxis assay for analysis of neural precursor cell migration kinetics in an externally applied direct current electric field.

    PubMed

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

    2012-10-13

    The discovery of neural stem and progenitor cells (collectively termed neural precursor cells) (NPCs) in the adult mammalian brain has led to a body of research aimed at utilizing the multipotent and proliferative properties of these cells for the development of neuroregenerative strategies. A critical step for the success of such strategies is the mobilization of NPCs toward a lesion site following exogenous transplantation or to enhance the response of the endogenous precursors that are found in the periventricular region of the CNS. Accordingly, it is essential to understand the mechanisms that promote, guide, and enhance NPC migration. Our work focuses on the utilization of direct current electric fields (dcEFs) to promote and direct NPC migration - a phenomenon known as galvanotaxis. Endogenous physiological electric fields function as critical cues for cell migration during normal development and wound repair. Pharmacological disruption of the trans-neural tube potential in axolotl embryos causes severe developmental malformations(1). In the context of wound healing, the rate of repair of wounded cornea is directly correlated with the magnitude of the epithelial wound potential that arises after injury, as shown by pharmacological enhancement or disruption of this dcEF(2-3). We have demonstrated that adult subependymal NPCs undergo rapid and directed cathodal migration in vitro when exposed to an externally applied dcEF. In this protocol we describe our lab's techniques for creating a simple and effective galvanotaxis assay for high-resolution, long-term observation of directed cell body translocation (migration) on a single-cell level. This assay would be suitable for investigating the mechanisms that regulate dcEF transduction into cellular motility through the use of transgenic or knockout mice, short interfering RNA, or specific receptor agonists/antagonists.

  6. Effect of photobiomodulation on endothelial cell exposed to Bothrops jararaca venom.

    PubMed

    Franco, Ana Tereza Barufi; Silva, Luciana Miato Gonçalves; Costa, Marcília Silva; Zamuner, Silvia Fernanda; Vieira, Rodolfo Paula; de Fatima Pereira Teixeira, Catarina; Zamuner, Stella Regina

    2016-07-01

    Bleeding is a common feature in envenoming caused by Bothrops snake venom due to extensive damage to capillaries and venules, producing alterations in capillary endothelial cell morphology. It has been demonstrated, in vivo, that photobiomodulation (PBM) decreases hemorrhage after venom inoculation; however, the mechanism is unknown. Thus, the objective was to investigate the effects of PBM on a murine endothelial cell line (tEnd) exposed to Bothrops jararaca venom (BjV). Cells were exposed to BjV and irradiated once with either 660- or 780-nm wavelength laser light at energy densities of 4 and 5 J/cm(2), respectively, and irradiation time of 10 s. Cell integrity was analyzed by crystal violet and cell viability/mitochondrial metabolism by MTT assay. The release of lactic dehydrogenase (LDH) was quantified as a measure of cell damage. In addition, cytokine IL1-β levels were measured in the supernatant. PBM at 660 and 780 nm wavelength was able to increase cellular viability and decrease the release of LDH and the loss of cellular integrity. In addition, the concentration of pro-inflammatory cytokine IL1-β was reduced after PBM by both wavelengths. The data reported herein indicates that irradiation with red or near-infrared laser resulted in protection on endothelial cells after exposure to Bothrops venom and could be, at least in part, a reasonable explanation by the beneficial effects of PBM inhibiting the local effects induced by Bothrops venoms, in vivo. PMID:27147074

  7. Maintenance of neural stem cell regional identity in culture.

    PubMed

    Delgado, Ryan N; Lu, Changqing; Lim, Daniel A

    2016-01-01

    Neural stem cells (NSCs) are distributed throughout the ventricular-subventricular zone (V-SVZ) in the adult mouse brain. NSCs located in spatially distinct regions of the V-SVZ generate different types of olfactory bulb (OB) neurons, and the regional expression of specific transcription factors correlates with these differences in NSC developmental potential. In a recent article, we show that Nkx2.1-expressing embryonic precursors give rise to NKX2.1+ NSCs located in the ventral V-SVZ of adult mice. Here we characterize a V-SVZ monolayer culture system that retains regional gene expression and neurogenic potential of NSCs from the dorsal and ventral V-SVZ. In particular, we find that Nkx2.1-lineage V-SVZ NSCs maintain Nkx2.1 expression through serial passage and can generate new neurons in vitro. Thus, V-SVZ NSCs retain key aspects of their in vivo regional identity in culture, providing new experimental opportunities for understanding how such developmental patterns are established and maintained during development. PMID:27606338

  8. 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

  9. IDH1R132H in Neural Stem Cells: Differentiation Impaired by Increased Apoptosis

    PubMed Central

    Rosiak, Kamila; Smolarz, Maciej; Stec, Wojciech J.; Peciak, Joanna; Grzela, Dawid; Winiecka-Klimek, Marta; Stoczynska-Fidelus, Ewelina; Krynska, Barbara; Piaskowski, Sylwester; Rieske, Piotr

    2016-01-01

    Background The high frequency of mutations in the isocitrate dehydrogenase 1 (IDH1) gene in diffuse gliomas indicates its importance in the process of gliomagenesis. These mutations result in loss of the normal function and acquisition of the neomorphic activity converting α-ketoglutarate to 2-hydroxyglutarate. This potential oncometabolite may induce the epigenetic changes, resulting in the deregulated expression of numerous genes, including those related to the differentiation process or cell survivability. Methods Neural stem cells were derived from human induced pluripotent stem cells following embryoid body formation. Neural stem cells transduced with mutant IDH1R132H, empty vector, non-transduced and overexpressing IDH1WT controls were differentiated into astrocytes and neurons in culture. The neuronal and astrocytic differentiation was determined by morphology and expression of lineage specific markers (MAP2, Synapsin I and GFAP) as determined by real-time PCR and immunocytochemical staining. Apoptosis was evaluated by real-time observation of Caspase-3 activation and measurement of PARP cleavage by Western Blot. Results Compared with control groups, cells expressing IDH1R132H retained an undifferentiated state and lacked morphological changes following stimulated differentiation. The significant inhibitory effect of IDH1R132H on neuronal and astrocytic differentiation was confirmed by immunocytochemical staining for markers of neural stem cells. Additionally, real-time PCR indicated suppressed expression of lineage markers. High percentage of apoptotic cells was detected within IDH1R132H-positive neural stem cells population and their derivatives, if compared to normal neural stem cells and their derivatives. The analysis of PARP and Caspase-3 activity confirmed apoptosis sensitivity in mutant protein-expressing neural cells. Conclusions Our study demonstrates that expression of IDH1R132H increases apoptosis susceptibility of neural stem cells and their

  10. BOSS on EXPOSE-R2-Comparative Investigations on Biofilm and Planktonic cells of Deinococcus geothermalis as Mission Preparation Tests

    NASA Astrophysics Data System (ADS)

    Panitz, C.; Rettberg, P.; Frösler, J.; Flemming, H.-C.; Rabbow, E.; Reitz, G.

    2013-09-01

    Biofilms are of interest for Astrobiological investigations since they are one of the oldest clear signs of life on Earth. In the experiment BOSS the hypothesis will be tested if the biofilm form of life with microorganisms embedded and aggregated in their EPS matrix is suited to support long-term survival of microorganisms under the harsh environmental conditions as they exist in space and on Mars and is superior to the same bacteria in the form of planktonic cultures. An additional protective role may be provided by particles associated in biofilms which may shield the organisms against radiation. The experiment will be flown on EXPOSE-R2 attached outside of the ISS on the Russian module. BOSS has participated the Experiment verification tests and will attend the upcoming Science verification test carried out in the Planetary and Space Simulation Facilities at DLR. The launch is scheduled for April 2014.

  11. DNA DAMAGE IN BUCCAL EPITHELIAL CELLS FROM INDIVIDUALS CHRONICALLY EXPOSED TO ARSENIC VIA DRINKING WATER IN INNER MONGOLIA, CHINA

    EPA Science Inventory

    The purpose of this pilot study was to assess DNA damage in buccal cells from individuals chronically exposed to arsenic via drinking water in Ba Men, Inner Mongolia. Buccal cells were collected from 19 Ba Men residents exposed to arsenic at 527.5 ? 23.7 g/L (mean ? SEM) and ...

  12. Phospholipidomic Profile Variation on THP-1 Cells Exposed to Skin or Respiratory Sensitizers and Respiratory Irritant.

    PubMed

    Martins, João D; Maciel, Elisabete A; Silva, Ana; Ferreira, Isabel; Ricardo, Fernando; Domingues, Pedro; Neves, Bruno M; Domingues, Maria Rosário M; Cruz, Maria Teresa

    2016-12-01

    Occupational exposure to low molecular weight reactive chemicals often leads to development of allergic reactions such as allergic contact dermatitis and respiratory allergies. Further insights into the interaction of these chemicals with physiopathological relevant cellular models might provide the foundations for novel non-animal approaches to safety assessment. In this work we used the human THP-1 cell line to determine phospholipidome changes induced by the skin sensitizer 1-fluoro-2,4-dinitrobenzene (DNFB), the respiratory allergen hexamethylene diisocyanate (HDI), and the irritant methyl salicylate (MESA). We detected that these chemicals differently induce lipid peroxidation and modulate THP-1 IL-1β, IL-12B, IL-8, CD86, and HMOX1 transcription. Decreased phosphatidylethanolamine content was detected in cells exposed to MESA, while profound alterations in the relative abundance of cardiolipin species were observed in cells exposed to DNFB. All chemicals tested induced a decrease in the relative abundance of plasmanyl phosphatidylcholine species PC (O-16:0e/18:1) and phosphatidylinositol species PI (34:1), while increasing PI (38:4). An increased abundance of oleic acid was observed in the phospholipids of cells exposed to DNFB while a decreased abundance of palmitic acid was detected in cells treated with MESA or DNFB. We conclude that both specific and common alterations at phospholipidome levels are triggered by the different chemicals, while not allowing a complete distinction between them using a Canonical Analysis of Principal Coordinates (CAP). The common effects observed at phospholipids level with all the chemicals tested might be related to unspecific cell cytotoxic mechanisms that nevertheless may contribute to the elicitation of specific immune responses. J. Cell. Physiol. 231: 2639-2651, 2016. © 2016 Wiley Periodicals, Inc. PMID:26946329

  13. Transcriptional modulation of a human monocytic cell line exposed to PM(10) from an urban area.

    PubMed

    Bastonini, Emanuela; Verdone, Loredana; Morrone, Stefania; Santoni, Angela; Settimo, Gaetano; Marsili, Giovanni; La Fortezza, Marco; Di Mauro, Ernesto; Caserta, Micaela

    2011-08-01

    Insight into the mechanisms by which ambient air particulate matter mediates adverse health effects is needed to provide biological plausibility to epidemiological studies demonstrating an association between PM(10) exposure and increased morbidity and mortality. In vitro studies of the effects of air pollution on human cells help to establish conditions for the analysis of cause-effect relationships. One of the major challenges is to test native atmosphere in its complexity, rather than the various components individually. We have developed an in vitro system in which human monocyte-macrophage U937 cells are directly exposed to filters containing different amounts of PM(10) collected in the city of Rome. Transcriptional profiling obtained after short exposure (1h) of cells to a filter containing 1666μg PM(10) (77.6μg/cm(2)) using a macroarray panel of 1176 genes reveals a significant change in the mRNA level (>2 fold) for 87 genes relative to cells exposed to a control filter. Overall, 9 out of 87 modulated genes were annotated as "lung cancer". qRT-PCR confirmed the induction of relevant genes involved in DNA repair and apoptosis, specifically: ERCC1, TDG, DAD1 and MCL1. In cells exposed for 10min, 1h and 3h to different amounts of PM(10), transcription of TNFα and TRAP1, which code for a key pro-inflammatory cytokine and a mitochondrial protein involved in cell protection from oxidative stress, respectively, was shown to be modulated in a time-dependent, but not a dose-dependent manner. Taken together, these data indicate that it is possible to analyze the effects of untreated particulate matter on human cells by the direct-exposure approach we have developed, possibly providing new clues to traffic-related health hazard.

  14. Identification of differentially expressed non-coding RNAs in embryonic stem cell neural differentiation

    PubMed Central

    Skreka, Konstantinia; Schafferer, Simon; Nat, Irina-Roxanna; Zywicki, Marek; Salti, Ahmad; Apostolova, Galina; Griehl, Matthias; Rederstorff, Mathieu; Dechant, Georg; Hüttenhofer, Alexander

    2012-01-01

    Protein-coding genes, guiding differentiation of ES cells into neural cells, have extensively been studied in the past. However, for the class of ncRNAs only the involvement of some specific microRNAs (miRNAs) has been described. Thus, to characterize the entire small non-coding RNA (ncRNA) transcriptome, involved in the differentiation of mouse ES cells into neural cells, we have generated three specialized ribonucleo-protein particle (RNP)-derived cDNA libraries, i.e. from pluripotent ES cells, neural progenitors and differentiated neural cells, respectively. By high-throughput sequencing and transcriptional profiling we identified several novel miRNAs to be involved in ES cell differentiation, as well as seven small nucleolar RNAs. In addition, expression of 7SL, 7SK and vault-2 RNAs was significantly up-regulated during ES cell differentiation. About half of ncRNA sequences from the three cDNA libraries mapped to intergenic or intragenic regions, designated as interRNAs and intraRNAs, respectively. Thereby, novel ncRNA candidates exhibited a predominant size of 18–30 nt, thus resembling miRNA species, but, with few exceptions, lacking canonical miRNA features. Additionally, these novel intraRNAs and interRNAs were not only found to be differentially expressed in stem-cell derivatives, but also in primary cultures of hippocampal neurons and astrocytes, strengthening their potential function in neural ES cell differentiation. PMID:22492625

  15. Identification of differentially expressed non-coding RNAs in embryonic stem cell neural differentiation.

    PubMed

    Skreka, Konstantinia; Schafferer, Simon; Nat, Irina-Roxanna; Zywicki, Marek; Salti, Ahmad; Apostolova, Galina; Griehl, Matthias; Rederstorff, Mathieu; Dechant, Georg; Hüttenhofer, Alexander

    2012-07-01

    Protein-coding genes, guiding differentiation of ES cells into neural cells, have extensively been studied in the past. However, for the class of ncRNAs only the involvement of some specific microRNAs (miRNAs) has been described. Thus, to characterize the entire small non-coding RNA (ncRNA) transcriptome, involved in the differentiation of mouse ES cells into neural cells, we have generated three specialized ribonucleo-protein particle (RNP)-derived cDNA libraries, i.e. from pluripotent ES cells, neural progenitors and differentiated neural cells, respectively. By high-throughput sequencing and transcriptional profiling we identified several novel miRNAs to be involved in ES cell differentiation, as well as seven small nucleolar RNAs. In addition, expression of 7SL, 7SK and vault-2 RNAs was significantly up-regulated during ES cell differentiation. About half of ncRNA sequences from the three cDNA libraries mapped to intergenic or intragenic regions, designated as interRNAs and intraRNAs, respectively. Thereby, novel ncRNA candidates exhibited a predominant size of 18-30 nt, thus resembling miRNA species, but, with few exceptions, lacking canonical miRNA features. Additionally, these novel intraRNAs and interRNAs were not only found to be differentially expressed in stem-cell derivatives, but also in primary cultures of hippocampal neurons and astrocytes, strengthening their potential function in neural ES cell differentiation. PMID:22492625

  16. Clear cell renal carcinoma in a pregnant DES-exposed patient.

    PubMed

    Mansi, M L

    1989-07-01

    Several decades ago, diethylstilbestrol (DES) was prescribed to support the pregnancy of women who were diabetic, who had had consecutive abortions, or who were threatening to abort. The use of this estrogen substitute to support human gestation had ceased by the 1960s. In 1971, the first report was published in which DES exposure was linked with clear cell carcinoma of the vagina and cervix. Since then, many other documentations have been published on upper genital tract anomalies, poor reproductive performance, and the high incidence of fetal wastage in DES-exposed women. The author describes a case of clear cell carcinoma of the kidney in an 18-year-old pregnant woman with a prior history of vaginal adenosis who had been exposed to DES in utero.

  17. Mutagenicity and genotoxicity in gill erythrocyte cells of Poecilia reticulata exposed to a glyphosate formulation.

    PubMed

    De Souza Filho, José; Sousa, Caio César Neves; Da Silva, Cláudio Carlos; De Sabóia-Morais, Simone Maria Teixeira; Grisolia, Cesar Koppe

    2013-11-01

    Poecilia reticulata were exposed to herbicide Roundup Transorb(®) for micronucleus test, nuclear abnormalities and comet assay. The exposure-concentrations were based on CL50-96 h following 0, 1.41, 2.83, 4.24 and 5.65 μL L(-1) for 24 h. Micronucleus and comets were significantly increased in the gill erythrocyte cells after herbicide exposure compared with the non-exposed group. Results showed a gradual increase in the number of damaged cells, indicating a concentration-dependent effect and that this herbicide was mutagenic and genotoxic to P. reticulata and this effect could be attributed to a combination of compounds contained in the formulation with the active ingredient glyphosate. PMID:24042842

  18. Structural and function changes in organelles of liver cells in rats exposed to magnetic fields

    SciTech Connect

    Gorczynska, E. ); Wegrzynowicz, R. )

    1991-08-01

    Exposure of rats to magnetic fields of 10{sup {minus}3} and 10{sup {minus}2} T for 1 hr daily generated structural changes in hepatocytes mitochondria, endoplasmic reticulum, and ribosomes. Simultaneously there was an increase in the activities of the mitochondrial respiratory enzymes: NADH dehydrogenase, succinic dehydrogenase, and cytochrome oxidase. The extent of the changes in liver cell properties following exposure depend on the duration of exposure to and the strength of the applied magnetic fields. Ultrastructural studies did not reveal any changes in external membranes of hepatocytes or in the membranes of cell nuclei. An increase in the amount of glycogen in hepatocytes of rats exposed to both 10{sup {minus}3} and 10{sup {minus}2} T was noted. The high level of cortisol in serum of exposed rats suggests that magnetic field may be a stress generating factor.

  19. Enteric neural crest cells regulate vertebrate stomach patterning and differentiation.

    PubMed

    Faure, Sandrine; McKey, Jennifer; Sagnol, Sébastien; de Santa Barbara, Pascal

    2015-01-15

    In vertebrates, the digestive tract develops from a uniform structure where reciprocal epithelial-mesenchymal interactions pattern this complex organ into regions with specific morphologies and functions. Concomitant with these early patterning events, the primitive GI tract is colonized by the vagal enteric neural crest cells (vENCCs), a population of cells that will give rise to the enteric nervous system (ENS), the intrinsic innervation of the GI tract. The influence of vENCCs on early patterning and differentiation of the GI tract has never been evaluated. In this study, we report that a crucial number of vENCCs is required for proper chick stomach development, patterning and differentiation. We show that reducing the number of vENCCs by performing vENCC ablations induces sustained activation of the BMP and Notch pathways in the stomach mesenchyme and impairs smooth muscle development. A reduction in vENCCs also leads to the transdifferentiation of the stomach into a stomach-intestinal mixed phenotype. In addition, sustained Notch signaling activity in the stomach mesenchyme phenocopies the defects observed in vENCC-ablated stomachs, indicating that inhibition of the Notch signaling pathway is essential for stomach patterning and differentiation. Finally, we report that a crucial number of vENCCs is also required for maintenance of stomach identity and differentiation through inhibition of the Notch signaling pathway. Altogether, our data reveal that, through the regulation of mesenchyme identity, vENCCs act as a new mediator in the mesenchymal-epithelial interactions that control stomach development.

  20. Ionic mechanisms subserving mechanosensory transduction and neural integration in statocyst hair cells of Hermissenda

    NASA Technical Reports Server (NTRS)

    Farley, Joseph

    1988-01-01

    The neural processing of gravitational-produced sensory stimulation of statocyst hair cells in the nudibranch mollusk Hermissenda was studied. The goal in these studies was to understand how: gravireceptor neurons sense or transduce gravitational forces, gravitational stimulation is integrated so as to produce a graded receptor potential, and ultimately the generation of an action potential, and various neural adaptation phenomena which hair cells exhibit arise. The approach to these problems was primarily electrophysical.

  1. Inhibition of GSK-3β enhances neural differentiation in unrestricted somatic stem cells.

    PubMed

    Dastjerdi, Fatemeh Vahid; Zeynali, Bahman; Tafreshi, Azita Parvaneh; Shahraz, Anahita; Chavoshi, Mahin Sadat; Najafabadi, Irandokht Khaki; Vardanjani, Marzieh Mowlavi; Atashi, Amir; Soleimani, Masoud

    2012-11-01

    GSK-3β is a key molecule in several signalling pathways, including the Wnt/β-catenin signalling pathway. There is increasing evidence suggesting Wnt/β-catenin signalling is involved in the neural differentiation of embryonic, somatic and neural stem cells. However, a large body of evidence indicates that this pathway maintains stem cells in a proliferative state. To address this controversy, we have investigated whether the Wnt/β-catenin pathway is present and involved in the neural differentiation of newly introduced USSCs (unrestricted somatic stem cells). Our results indicate that the components of Wnt/β-catenin signalling are present in undifferentiated USSCs. We also show that the treatment of neurally induced USSCs with BIO (6-bromoindirubin-3'-oxime), a specific GSK-3β inhibitor and Wnt activator, for 5 and 10 days results in increased expression of a general neuronal marker (β-tubulin III). Moreover, the expression of pGSK-3β and stabilized β-catenin increased by BIO in neurally induced USSCs, indicates that the Wnt pathway is activated and functional in these cells. Thus, inhibition of GSK-3β in USSCs enhances their neural differentiation, which suggests a positive role of the Wnt/β-catenin signalling pathway towards neural fate.

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

    PubMed

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

    2013-02-01

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

  3. Characterization of insulin-producing cells derived from PDX-1-transfected neural stem cells.

    PubMed

    Wang, Hailan; Jiang, Zesheng; Li, Aihui; Gao, Yi

    2012-12-01

    Islet cell transplantation is a promising treatment strategy for type-1 diabetes. However, functional islet cells are hard to obtain for transplantation and are in short supply. Directing the differentiation of stem cells into insulin‑producing cells, which serve as islet cells, would overcome this shortage. Bone marrow contains hematopoietic stem cells and mesenchymal stem cells. The present study used bone marrow cells isolated from rats and neural stem cells (NSCs) that were derived from bone marrow cells in culture. Strong nestin staining was detected in NSCs, but not in bone marrow stromal cells (BMSCs). In vitro transfection of the pancreatic duodenal homeobox-1 (PDX-1) gene into NSCs generated insulin‑producing cells. Reverse transcription polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) analysis confirmed that PDX-1-transfected NSCs expressed insulin mRNA and released insulin protein. However, insulin release from PDX-1-transfected NSCs did not respond to the challenge of glucose and glucagon-like peptide-1. These results support the use of bone marrow-derived NSCs as a renewable source of insulin-producing cells for autologous transplantation to treat type-1 diabetes.

  4. Natural killer cells in highly exposed hepatitis C-seronegative injecting drug users.

    PubMed

    Mina, M M; Cameron, B; Luciani, F; Vollmer-Conna, U; Lloyd, A R

    2016-06-01

    Injecting drug use remains the major risk factor for hepatitis C (HCV) transmission. A minority of long-term injecting drug users remain seronegative and aviraemic, despite prolonged exposure to HCV - termed highly exposed seronegative subjects. Natural killer (NK) cells have been implicated in this apparent protection. A longitudinal nested, three group case-control series of subjects was selected from a prospective cohort of seronegative injecting drug users who became incident cases (n = 11), remained seronegative (n = 11) or reported transient high-risk behaviour and remained uninfected (n = 11). The groups were matched by age, sex and initial risk behaviour characteristics. Stored peripheral blood mononuclear cells were assayed in multicolour flow cytometry to enumerate natural killer cell subpopulations and to assess functional activity using Toll-like receptor ligands before measurement of activation, cytokine production and natural cytotoxicity receptor expression. Principal components were derived to describe the detailed phenotypic characteristics of the major NK subpopulations (based on CD56 and CD16 co-expression), before logistic regression analysis to identify associations with exposed, seronegative individuals. The CD56(dim) CD16(+) (P = 0.05, OR 6.92) and CD56(dim) CD16(-) (P = 0.05, OR 6.07) principal components differed between exposed, seronegative individuals and pre-infection samples of the other two groups. These included CD56(dim) CD16(+) and CD56(dim) CD16(-) subsets with CD56(dim) CD16(+) IFN-γ and TNF-α on unstimulated cells, and CD56(dim) CD16(-) CD69(+) , CD107a(+) , IFN-γ and TNF-α following TLR stimulation. The cytotoxic CD56(dim) NK subset thus distinguished highly exposed, seronegative subjects, suggesting NK cytotoxicity may contribute to protection from HCV acquisition. Further investigation of the determinants of this association and prospective assessment of protection against HCV infection are warranted.

  5. Analysis of cell identity, morphology, apoptosis and mitotic activity in a primary neural cell culture system in Drosophila

    PubMed Central

    2012-01-01

    In Drosophila, most neurogenetic research is carried out in vivo. Mammalian research demonstrates that primary cell culture techniques provide a powerful model to address cell autonomous and non-autonomous processes outside their endogenous environment. We developed a cell culture system in Drosophila using wildtype and genetically manipulated primary neural tissue for long-term observations. We assessed the molecular identity of distinct neural cell types by immunolabeling and genetically expressed fluorescent cell markers. We monitored mitotic activity of cell cultures derived from wildtype and tumorous larval brains. Our system provides a powerful approach to unveil developmental processes in the nervous system and to complement studies in vivo. PMID:22554060

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

    PubMed

    Ivanov, Vladimir N; Hei, Tom K

    2015-07-01

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

  7. Mechanisms of mutagenesis in human cells exposed to 55 MeV protons

    NASA Technical Reports Server (NTRS)

    Gauny, S.; Wiese, C.; Kronenberg, A.

    2001-01-01

    Protons represent the major type of charged particle radiation in spaceflight environments. The purpose of this study was to assess mutations arising in human lymphoid cells exposed to protons. Mutations were quantitated at the thymidine kinase (TK1) locus in cell lines derived from the same donor: TK6 cells (wt TP53) and WTK1 cells (mutant TP53). WTK1 cells were much more susceptible to mutagenesis following proton exposure than TK6 cells. Intragenic deletions were observed among early-arising TK1 mutants in TK6 cells, but not in WTK1 cells where all of the mutants arose by LOH. Deletion was the predominant mode of LOH in TK6 cells, while allelic recombination was the major mode of LOH in WTK1 cells. Deletions were of variable lengths, from <1 cM to 64 cM, while mutations that arose by allelic recombination often extended to the telomere. In summary, proton exposures elicited many types of mutations at an autosomal locus in human cells. Most involved large scale loss of genetic information, either through deletion or by recombination.

  8. Neural differentiation of pluripotent mouse embryonal carcinoma cells by retinoic acid: inhibitory effect of serum.

    PubMed

    Pacherník, J; Bryja, V; Esner, M; Kubala, L; Dvorák, P; Hampl, A

    2005-01-01

    In both embryonal carcinoma (EC) and embryonic stem (ES) cells, the differentiation pathway entered after treatment with retinoic acid (RA) varies as it is based upon different conditions of culture. This study employs mouse EC cells P19 to investigate the effects of serum on RA-induced neural differentiation occurring in a simplified monolayer culture. Cell morphology and expression of lineage-specific molecular markers document that, while non-neural cell types arise after treatment with RA under serum-containing conditions, in chemically defined serum-free media RA induces massive neural differentiation in concentrations of 10(-9) M and higher. Moreover, not only neural (Mash-1) and neuroectodermal (Pax-6), but also endodermal (GATA-4, alpha-fetoprotein) genes are expressed at early stages of differentiation driven by RA under serum-free conditions. Furthermore, as determined by the luciferase reporter assay, the presence or absence of the serum does not affect the activity of the retinoic acid response element (RARE). Thus, mouse EC cells are able to produce neural cells upon exposure to RA even without culture in three-dimensional embryoid bodies (EBs). However, in contrast to standard EBs-involving protocol(s), neural differentiation in monolayer only takes place when complex signaling from serum factors is avoided. This simple and efficient strategy is proposed to serve as a basis for neurodifferentiation studies in vitro. PMID:15717849

  9. Bone Morphogenetic Protein 4 Signalling in Neural Stem and Progenitor Cells during Development and after Injury

    PubMed Central

    Cole, Alistair E.; Murray, Simon S.; Xiao, Junhua

    2016-01-01

    Substantial progress has been made in identifying the extracellular signalling pathways that regulate neural stem and precursor cell biology in the central nervous system (CNS). The bone morphogenetic proteins (BMPs), in particular BMP4, are key players regulating neuronal and glial cell development from neural precursor cells in the embryonic, postnatal, and injured CNS. Here we review recent studies on BMP4 signalling in the generation of neurons, astrocytes, and oligodendroglial cells in the CNS. We also discuss putative mechanisms that BMP4 may utilise to influence glial cell development following CNS injury and highlight some questions for further research. PMID:27293450

  10. GSK-3β: a signaling pathway node modulating neural stem cell and endothelial cell interactions.

    PubMed

    Li, Qi; Michaud, Michael; Canosa, Sandra; Kuo, Andrew; Madri, Joseph A

    2011-05-01

    The neurogenic areas of the brain are highly organized structures in which there is dynamic reciprocal modulation of neural stem cells (NSC) and microvascular endothelial cells (BEC) resulting in control of neural stem cell and vascular proliferation, survival and differentiation throughout the life of the individual. Select molecules such as GSK-3β, functioning as signaling nodes, and their downstream signaling components including HIF-1α, HIF-2α and β-catenin participate in regulating and orchestrating the diverse responses involved in this complex process. In this report we demonstrate GSK-3β's role as a signaling node in two mouse strains (C57BL/6, which have been found to respond to and recover from a hypoxic insult from P3 to P11 poorly and CD-1, which have been found to respond to and recover from a hypoxic insult from P3 to P11 well both in vivo and in vitro) which mimic the wide range of responsiveness to hypoxic insult observed in the very low birth weight premature infant population. Differences in levels of neural stem cell and microvascular endothelial cell GSK-3β activation, β-catenin serine phosphorylation, HIF-1α and 2α, BDNF, SDF-1 and VEGF, β-III-tubulin and cleaved notch-1 expression in C57BL/6 and CD-1 subventricular zone tissues, and cultured NSC and BEC were noted. Specifically, CD1 pups, SVZ tissues and isolated NSC and BEC exhibit less GSK-3β and β-catenin serine phoslphorylation and greater HIF-1α and 2α, BDNF, SDF-1 and VEGF, β-III-tubulin and cleaved notch-1 expression compared to C57BL/6. Correlating with these changes were differences of several neural stem cell and microvascular endothelial cell behaviors including proliferation, apoptosis, migration and differentiation with CD1 NSC exhibiting greater proliferation and migration and decreased apoptosis and differentiation and CD1 BEC exhibiting greater angiogenesis. Further, upon treatment with nanomolar concentrations of a GSK-3β inhibitor (SB412682), C57 NSC and BEC

  11. Neural stem/progenitor cell properties of glial cells in the adult mouse auditory nerve

    PubMed Central

    Lang, Hainan; Xing, Yazhi; Brown, LaShardai N.; Samuvel, Devadoss J.; Panganiban, Clarisse H.; Havens, Luke T.; Balasubramanian, Sundaravadivel; Wegner, Michael; Krug, Edward L.; Barth, Jeremy L.

    2015-01-01

    The auditory nerve is the primary conveyor of hearing information from sensory hair cells to the brain. It has been believed that loss of the auditory nerve is irreversible in the adult mammalian ear, resulting in sensorineural hearing loss. We examined the regenerative potential of the auditory nerve in a mouse model of auditory neuropathy. Following neuronal degeneration, quiescent glial cells converted to an activated state showing a decrease in nuclear chromatin condensation, altered histone deacetylase expression and up-regulation of numerous genes associated with neurogenesis or development. Neurosphere formation assays showed that adult auditory nerves contain neural stem/progenitor cells (NSPs) that were within a Sox2-positive glial population. Production of neurospheres from auditory nerve cells was stimulated by acute neuronal injury and hypoxic conditioning. These results demonstrate that a subset of glial cells in the adult auditory nerve exhibit several characteristics of NSPs and are therefore potential targets for promoting auditory nerve regeneration. PMID:26307538

  12. SNPs in the neural cell adhesion molecule 1 gene (NCAM1) may be associated with human neural tube defects

    PubMed Central

    Deak, Kristen L.; Boyles, Abee L.; Etchevers, Heather C.; Melvin, Elizabeth C.; Siegel, Deborah G.; Graham, Felicia L.; Slifer, Susan H.; Enterline, David S.; George, Timothy M.; Vekemans, Michel; McClay, David; Bassuk, Alexander G.; Kessler, John A.; Linney, Elwood; Gilbert, John R.

    2011-01-01

    Neural tube defects (NTDs) are common birth defects, occurring in approximately 1/1,000 births; both genetic and environmental factors are implicated. To date, no major genetic risk factors have been identified. Throughout development, cell adhesion molecules are strongly implicated in cell–cell interactions, and may play a role in the formation and closure of the neural tube. To evaluate the role of neural cell adhesion molecule 1 (NCAM1) in risk of human NTDs, we screened for novel single-nucleotide polymorphisms (SNPs) within the gene. Eleven SNPs across NCAM1 were genotyped using TaqMan. We utilized a family-based approach to evaluate evidence for association and/or linkage disequilibrium. We evaluated American Caucasian simplex lumbosacral myelomeningocele families (n=132 families) using the family based association test (FBAT) and the pedigree disequilibrium test (PDT). Association analysis revealed a significant association between risk for NTDs and intronic SNP rs2298526 using both the FBAT test (P=0.0018) and the PDT (P=0.0025). Using the HBAT version of the FBAT to look for haplotype association, all pairwise comparisons with SNP rs2298526 were also significant. A replication study set, consisting of 72 additional families showed no significant association; however, the overall trend for overtransmission of the less common allele of SNP rs2298526 remained significant in the combined sample set. In addition, we analyzed the expression pattern of the NCAM1 protein in human embryos, and while NCAM1 is not expressed within the neural tube at the time of closure, it is expressed in the surrounding and later in differentiated neurons of the CNS. These results suggest variations in NCAM1 may influence risk for human NTDs. PMID:15883837

  13. Stroke Increases Neural Stem Cells and Angiogenesis in the Neurogenic Niche of the Adult Mouse

    PubMed Central

    Zhang, Rui Lan; Chopp, Michael; Roberts, Cynthia; Liu, Xianshuang; Wei, Min; Nejad-Davarani, Siamak P.; Wang, Xinli; Zhang, Zheng Gang

    2014-01-01

    The unique cellular and vascular architecture of the adult ventricular-subventricular zone (V/SVZ) neurogenic niche plays an important role in regulating neural stem cell function. However, the in vivo identification of neural stem cells and their relationship to blood vessels within this niche in response to stroke remain largely unknown. Using whole-mount preparation of the lateral ventricle wall, we examined the architecture of neural stem cells and blood vessels in the V/SVZ of adult mouse over the course of 3 months after onset of focal cerebral ischemia. Stroke substantially increased the number of glial fibrillary acidic protein (GFAP) positive neural stem cells that are in contact with the cerebrospinal fluid (CSF) via their apical processes at the center of pinwheel structures formed by ependymal cells residing in the lateral ventricle. Long basal processes of these cells extended to blood vessels beneath the ependymal layer. Moreover, stroke increased V/SVZ endothelial cell proliferation from 2% in non-ischemic mice to 12 and 15% at 7 and 14 days after stroke, respectively. Vascular volume in the V/SVZ was augmented from 3% of the total volume prior to stroke to 6% at 90 days after stroke. Stroke-increased angiogenesis was closely associated with neuroblasts that expanded to nearly encompass the entire lateral ventricular wall in the V/SVZ. These data indicate that stroke induces long-term alterations of the neural stem cell and vascular architecture of the adult V/SVZ neurogenic niche. These post-stroke structural changes may provide insight into neural stem cell mediation of stroke-induced neurogenesis through the interaction of neural stem cells with proteins in the CSF and their sub-ependymal neurovascular interaction. PMID:25437857

  14. A robust method to derive functional neural crest cells from human pluripotent stem cells.

    PubMed

    Kreitzer, Faith R; Salomonis, Nathan; Sheehan, Alice; Huang, Miller; Park, Jason S; Spindler, Matthew J; Lizarraga, Paweena; Weiss, William A; So, Po-Lin; Conklin, Bruce R

    2013-01-01

    Neural crest (NC) cells contribute to the development of many complex tissues of all three germ layers during embryogenesis, and its abnormal development accounts for several congenital birth defects. Generating NC cells-including specific subpopulations such as cranial, cardiac, and trunk NC cells-from human pluripotent stem cells will provide a valuable model system to study human development and disease. Here, we describe a rapid and robust NC differentiation method called "LSB-short" that is based on dual SMAD pathway inhibition. This protocol yields high percentages of NC cell populations from multiple human induced pluripotent stem and human embryonic stem cell lines in 8 days. The resulting cells can be propagated easily, retain NC marker expression over multiple passages, and can spontaneously differentiate into several NC-derived cell lineages, including smooth muscle cells, peripheral neurons, and Schwann cells. NC cells generated by this method represent cranial, cardiac and trunk NC subpopulations based on global gene expression analyses, are similar to in vivo analogues, and express a common set of NC alternative isoforms. Functionally, they are also able to migrate appropriately in response to chemoattractants such as SDF-1, FGF8b, and Wnt3a. By yielding NC cells that likely represent all NC subpopulations in a shorter time frame than other published methods, our LSB-short method provides an ideal model system for further studies of human NC development and disease.

  15. A robust method to derive functional neural crest cells from human pluripotent stem cells.

    PubMed

    Kreitzer, Faith R; Salomonis, Nathan; Sheehan, Alice; Huang, Miller; Park, Jason S; Spindler, Matthew J; Lizarraga, Paweena; Weiss, William A; So, Po-Lin; Conklin, Bruce R

    2013-01-01

    Neural crest (NC) cells contribute to the development of many complex tissues of all three germ layers during embryogenesis, and its abnormal development accounts for several congenital birth defects. Generating NC cells-including specific subpopulations such as cranial, cardiac, and trunk NC cells-from human pluripotent stem cells will provide a valuable model system to study human development and disease. Here, we describe a rapid and robust NC differentiation method called "LSB-short" that is based on dual SMAD pathway inhibition. This protocol yields high percentages of NC cell populations from multiple human induced pluripotent stem and human embryonic stem cell lines in 8 days. The resulting cells can be propagated easily, retain NC marker expression over multiple passages, and can spontaneously differentiate into several NC-derived cell lineages, including smooth muscle cells, peripheral neurons, and Schwann cells. NC cells generated by this method represent cranial, cardiac and trunk NC subpopulations based on global gene expression analyses, are similar to in vivo analogues, and express a common set of NC alternative isoforms. Functionally, they are also able to migrate appropriately in response to chemoattractants such as SDF-1, FGF8b, and Wnt3a. By yielding NC cells that likely represent all NC subpopulations in a shorter time frame than other published methods, our LSB-short method provides an ideal model system for further studies of human NC development and disease. PMID:23862100

  16. Short Report: Olfactory Ensheathing Cells Promote Differentiation of Neural Stem Cells and Robust Neurite Extension

    PubMed Central

    Sethi, Rosh; Sethi, Roshan; Redmond, Andy

    2014-01-01

    Aims The goal of this study was to gain insight into the signaling between olfactory ensheathing cells (OECs) and neural stem cells (NSCs). We sought to understand the impact of OECs on NSC differentiation and neurite extension and to begin to elucidate the factors involved in these interactions to provide new targets for therapeutic interventions. Materials and Methods We utilized lines of OECs that have been extremely well characterized in vitro and in vivo along with well studied NSCs in gels to determine the impact of the coculture in three dimensions. To further elucidate the signaling, we used conditioned media from the OECs as well as fractioned components on NSCs to determine the molecular weight range of the soluble factors that was most responsible for the NSC behavior. Results We found that the coculture of NSCs and OECs led to robust NSC differentiation and extremely long neural processes not usually seen with NSCs in three dimensional gels in vitro. Through culture of NSCs with fractioned OEC media, we determined that molecules larger than 30 kDa have the greatest impact on the NSC behavior. Conclusions Overall, our findings suggest that cocultures of NSCs and OECs may be a novel combination therapy for neural injuries including spinal cord injury (SCI). Furthermore, we have identified a class of molecules which plays a substantial role in the behavior that provides new targets for investigating pharmacological therapies. PMID:24996386

  17. Inducible expression of beta defensins by human respiratory epithelial cells exposed to Aspergillus fumigatus organisms

    PubMed Central

    2009-01-01

    Background Aspergillus fumigatus, a saprophytic mould, is responsible for life-threatening, invasive pulmonary diseases in immunocompromised hosts. The role of the airway epithelium involves a complex interaction with the inhaled pathogen. Antimicrobial peptides with direct antifungal and chemotactic activities may boost antifungal immune response. Results The inducible expression of defensins by human bronchial epithelial 16HBE cells and A549 pneumocyte cells exposed to A. fumigatus was investigated. Using RT-PCR and real time PCR, we showed an activation of hBD2 and hBD9 defensin genes: the expression was higher in cells exposed to swollen conidia (SC), compared to resting conidia (RC) or hyphal fragments (HF). The kinetics of defensin expression was different for each one, evoking a putative distinct function for each investigated defensin. The decrease of defensin expression in the presence of heat-inactivated serum indicated a possible link between defensins and the proteins of the host complement system. The presence of defensin peptide hBD2 was revealed using immunofluorescence that showed a punctual cytoplasmic and perinuclear staining. Quantification of the cells stained with anti hBD2 antibody demonstrated that SC induced a greater number of cells that synthesized hBD2, compared to RC or HF. Labelling of the cells with anti-hBD-2 antibody showed a positive immunofluorescence signal around RC or SC in contrast to HF. This suggests co-localisation of hBD2 and digested conidia. The HBD2 level was highest in the supernatants of cells exposed to SC, as was determined by sandwich ELISA. Experiments using neutralising anti-interleukine-1β antibody reflect the autocrine mechanism of defensin expression induced by SC. Investigation of defensin expression at transcriptional and post-transcriptional levels demonstrated the requirement of transcription as well as new protein synthesis during A. fumigatus defensin induction. Finally, induced defensin expression in

  18. Spirulina promotes stem cell genesis and protects against LPS induced declines in neural stem cell proliferation.

    PubMed

    Bachstetter, Adam D; Jernberg, Jennifer; Schlunk, Andrea; Vila, Jennifer L; Hudson, Charles; Cole, Michael J; Shytle, R Douglas; Tan, Jun; Sanberg, Paul R; Sanberg, Cyndy D; Borlongan, Cesario; Kaneko, Yuji; Tajiri, Naoki; Gemma, Carmelina; Bickford, Paula C

    2010-01-01

    Adult stem cells are present in many tissues including, skin, muscle, adipose, bone marrow, and in the brain. Neuroinflammation has been shown to be a potent negative regulator of stem cell and progenitor cell proliferation in the neurogenic regions of the brain. Recently we demonstrated that decreasing a key neuroinflammatory cytokine IL-1beta in the hippocampus of aged rats reversed the age-related cognitive decline and increased neurogenesis in the age rats. We also have found that nutraceuticals have the potential to reduce neuroinflammation, and decrease oxidative stress. The objectives of this study were to determine if spirulina could protect the proliferative potential of hippocampal neural progenitor cells from an acute systemic inflammatory insult of lipopolysaccharide (LPS). To this end, young rats were fed for 30 days a control diet or a diet supplemented with 0.1% spirulina. On day 28 the rats were given a single i.p. injection of LPS (1 mg/kg). The following day the rats were injected with BrdU (50 mg/kg b.i.d. i.p.) and were sacrificed 24 hours after the first injection of BrdU. Quantification of the BrdU positive cells in the subgranular zone of the dentate gyrus demonstrated a decrease in proliferation of the stem/progenitor cells in the hippocampus as a result of the LPS insult. Furthermore, the diet supplemented with spirulina was able to negate the LPS induced decrease in stem/progenitor cell proliferation. In a second set of studies we examined the effects of spirulina either alone or in combination with a proprietary formulation (NT-020) of blueberry, green tea, vitamin D3 and carnosine on the function of bone marrow and CD34+ cells in vitro. Spirulina had small effects on its own and more than additive effects in combination with NT-020 to promote mitochondrial respiration and/or proliferation of these cells in culture. When examined on neural stem cells in culture spirulina increased proliferation at baseline and protected against the

  19. Spirulina Promotes Stem Cell Genesis and Protects against LPS Induced Declines in Neural Stem Cell Proliferation

    PubMed Central

    Bachstetter, Adam D.; Jernberg, Jennifer; Schlunk, Andrea; Vila, Jennifer L.; Hudson, Charles; Cole, Michael J.; Shytle, R. Douglas; Tan, Jun; Sanberg, Paul R.; Sanberg, Cyndy D.; Borlongan, Cesario; Kaneko, Yuji; Tajiri, Naoki; Gemma, Carmelina; Bickford, Paula C.

    2010-01-01

    Adult stem cells are present in many tissues including, skin, muscle, adipose, bone marrow, and in the brain. Neuroinflammation has been shown to be a potent negative regulator of stem cell and progenitor cell proliferation in the neurogenic regions of the brain. Recently we demonstrated that decreasing a key neuroinflammatory cytokine IL-1β in the hippocampus of aged rats reversed the age-related cognitive decline and increased neurogenesis in the age rats. We also have found that nutraceuticals have the potential to reduce neuroinflammation, and decrease oxidative stress. The objectives of this study were to determine if spirulina could protect the proliferative potential of hippocampal neural progenitor cells from an acute systemic inflammatory insult of lipopolysaccharide (LPS). To this end, young rats were fed for 30 days a control diet or a diet supplemented with 0.1% spirulina. On day 28 the rats were given a single i.p. injection of LPS (1 mg/kg). The following day the rats were injected with BrdU (50 mg/kg b.i.d. i.p.) and were sacrificed 24 hours after the first injection of BrdU. Quantification of the BrdU positive cells in the subgranular zone of the dentate gyrus demonstrated a decrease in proliferation of the stem/progenitor cells in the hippocampus as a result of the LPS insult. Furthermore, the diet supplemented with spirulina was able to negate the LPS induced decrease in stem/progenitor cell proliferation. In a second set of studies we examined the effects of spirulina either alone or in combination with a proprietary formulation (NT-020) of blueberry, green tea, vitamin D3 and carnosine on the function of bone marrow and CD34+ cells in vitro. Spirulina had small effects on its own and more than additive effects in combination with NT-020 to promote mitochondrial respiration and/or proliferation of these cells in culture. When examined on neural stem cells in culture spirulina increased proliferation at baseline and protected against the negative

  20. Bi-parental care contributes to sexually dimorphic neural cell genesis in the adult mammalian brain.

    PubMed

    Mak, Gloria K; Antle, Michael C; Dyck, Richard H; Weiss, Samuel

    2013-01-01

    Early life events can modulate brain development to produce persistent physiological and behavioural phenotypes that are transmissible across generations. However, whether neural precursor cells are altered by early life events, to produce persistent and transmissible behavioural changes, is unknown. Here, we show that bi-parental care, in early life, increases neural cell genesis in the adult rodent brain in a sexually dimorphic manner. Bi-parentally raised male mice display enhanced adult dentate gyrus neurogenesis, which improves hippocampal neurogenesis-dependent learning and memory. Female mice display enhanced adult white matter oligodendrocyte production, which increases proficiency in bilateral motor coordination and preference for social investigation. Surprisingly, single parent-raised male and female offspring, whose fathers and mothers received bi-parental care, respectively, display a similar enhancement in adult neural cell genesis and phenotypic behaviour. Therefore, neural plasticity and behavioural effects due to bi-parental care persist throughout life and are transmitted to the next generation.

  1. A cell junction pathology of neural stem cells leads to abnormal neurogenesis and hydrocephalus.

    PubMed

    Rodríguez, Esteban M; Guerra, María M; Vío, Karin; González, César; Ortloff, Alexander; Bátiz, Luis F; Rodríguez, Sara; Jara, María C; Muñoz, Rosa I; Ortega, Eduardo; Jaque, Jaime; Guerra, Francisco; Sival, Deborah A; den Dunnen, Wilfred F A; Jiménez, Antonio J; Domínguez-Pinos, María D; Pérez-Fígares, José M; McAllister, James P; Johanson, Conrad

    2012-01-01

    Most cells of the developing mammalian brain derive from the ventricular (VZ) and the subventricular (SVZ) zones. The VZ is formed by the multipotent radial glia/neural stem cells (NSCs) while the SVZ harbors the rapidly proliferative neural precursor cells (NPCs). Evidence from human and animal models indicates that the common history of hydrocephalus and brain maldevelopment starts early in embryonic life with disruption of the VZ and SVZ. We propose that a "cell junction pathology" involving adherent and gap junctions is a final common outcome of a wide range of gene mutations resulting in proteins abnormally expressed by the VZ cells undergoing disruption. Disruption of the VZ during fetal development implies the loss of NSCs whereas VZ disruption during the perinatal period implies the loss of ependyma. The process of disruption occurs in specific regions of the ventricular system and at specific stages of brain development. This explains why only certain brain structures have an abnormal development, which in turn results in a specific neurological impairment of the newborn. Disruption of the VZ of the Sylvian aqueduct (SA) leads to aqueductal stenosis and hydrocephalus, while disruption of the VZ of telencephalon impairs neurogenesis. We are currently investigating whether grafting of NSCs/neurospheres from normal rats into the CSF of hydrocephalic mutants helps to diminish/repair the outcomes of VZ disruption.

  2. Interplay between autophagy and programmed cell death in mammalian neural stem cells

    PubMed Central

    Chung, Kyung Min; Yu, Seong-Woon

    2013-01-01

    Mammalian neural stem cells (NSCs) are of particular interest because of their role in brain development and function. Recent findings suggest the intimate involvement of programmed cell death (PCD) in the turnover of NSCs. However, the underlying mechanisms of PCD are largely unknown. Although apoptosis is the best-defined form of PCD, accumulating evidence has revealed a wide spectrum of PCD encompassing apoptosis, autophagic cell death (ACD) and necrosis. This mini-review aims to illustrate a unique regulation of PCD in NSCs. The results of our recent studies on autophagic death of adult hippocampal neural stem (HCN) cells are also discussed. HCN cell death following insulin withdrawal clearly provides a reliable model that can be used to analyze the molecular mechanisms of ACD in the larger context of PCD. More research efforts are needed to increase our understanding of the molecular basis of NSC turnover under degenerating conditions, such as aging, stress and neurological diseases. Efforts aimed at protecting and harnessing endogenous NSCs will offer novel opportunities for the development of new therapeutic strategies for neuropathologies. [BMB Reports 2013; 46(8): 383-390] PMID:23977985

  3. Effects of selenocystine on lead-exposed Chinese hamster ovary (CHO) and PC-12 cells

    SciTech Connect

    Aykin-Burns, Nukhet; Ercal, Nuran . E-mail: nercal@umr.edu

    2006-07-15

    Lead is a pervasive environmental toxin that affects multiple organ systems, including the nervous, renal, reproductive, and hematological systems. Even though it is probably the most studied toxic metal, some of the symptoms of lead toxicity still cannot be explained by known molecular mechanisms. Therefore, lead-induced oxidative stress has recently started to gain attention. This in vitro study confirms the existence of oxidative stress due to lead exposure. Administration of lead acetate (PbA) to cultures of Chinese hamster ovary cells (CHO) had a concentration-dependent inhibitory effect on colony formation and cell proliferation. This inhibition was eliminated by 5 {mu}M selenocystine (SeCys). In order to evaluate the nature of SeCys's effect, we measured glutathione (GSH), its oxidized form glutathione disulfide (GSSG), malondialdehyde (MDA), catalase, and GSH peroxidase (GPx) activities in lead-exposed CHO cells both in the presence and absence of SeCys. Increases in MDA, catalase, and GPx activities were observed in cultures that received only PbA, but supplementation with SeCys returned these measures to pretreatment levels. The ratio of GSH to GSSG increased in lead-exposed cells incubated in SeCys-enhanced media but declined in cultures treated with PbA only. In order to determine whether SeCys also reverses lead-induced neurotoxicity, a neuronal cell line, PC-12 cells, was used. Lead's inhibition on neurite formation was significantly eliminated by SeCys in PC-12 cells. Our results suggest that SeCys can confer protection against lead-induced toxicity in CHO cells and neurotoxicity in PC-12 cells.

  4. Expression of AS3MT alters transcriptional profiles in human urothelial cells exposed to arsenite.

    PubMed

    Hester, Sd; Drobná, Z; Andrews, Dmk; Liu, J; Waalkes, Mp; Thomas, Dj; Styblo, M

    2009-01-01

    Inorganic arsenic (iAs) is an environmental toxicant and human carcinogen. The enzymatic methylation of iAs that is catalyzed by arsenic (+3 oxidation state)-methyltransferase (AS3MT) generates reactive methylated intermediates that contribute to the toxic and carcinogenic effects of iAs. We have shown that clonal human urothelial cells (UROtsa/F35) that express rat AS3MT and methylate iAs are more susceptible to acute toxicity of arsenite (iAs(III)) than parental UROtsa cells that do not express AS3MT and do not methylate iAs. The current work examines transcriptional changes associated with AS3MT expression and identifies specific categories of genes expressed in UROtsa and UROtsa/F35 cells in response to a 24-h exposure to 1 or 50 microM iAs(III). Here, the expression of 21,073 genes was assessed using Agilent Human 1A(V2) arrays. Venn analysis showed marked concentration-dependent differences between gene expression patterns in UROtsa and UROTsa/F35 cells exposed to iAs(III). Among 134 genes altered by exposure to subtoxic 1 microM iAs(III), only 14 were shared by both cell lines. Exposure to cytotoxic 50 microM iAs(III) uniquely altered 1389 genes in UROtsa/F35 and 649 genes in UROtsa cells; 5033 altered genes were associated with the chemical alone. In UROtsa, but not UROtsa/F35 cells exposure to 1 microM iAs(III) altered expression of genes associated with cell adhesion. In contrast, expression of genes involved in cell cycle regulation was significantly altered in UROtsa/F35 cells at this exposure level. At 50 microM iAs(III), pathways regulating cell cycle, cell death, transcription, and metabolism were affected in both cell lines. However, only Urotsa/F35 cells showed numerous G-protein and kinase pathway alterations as well as alterations in pathways involved in cell growth and differentiation. These data link the AS3MT-catalyzed methylation of iAs to specific genomic responses in human cells exposed to iAs(III). Further analysis of these responses will

  5. Identification of gene-based responses in human blood cells exposed to alpha particle radiation

    PubMed Central

    2014-01-01

    Background The threat of a terrorist-precipitated nuclear event places humans at danger for radiological exposures. Isotopes which emit alpha (α)-particle radiation pose the highest risk. Currently, gene expression signatures are being developed for radiation biodosimetry and triage with respect to ionizing photon radiation. This study was designed to determine if similar gene expression profiles are obtained after exposures involving α-particles. Methods Peripheral blood mononuclear cells (PBMCs) were used to identify sensitive and robust gene-based biomarkers of α-particle radiation exposure. Cells were isolated from healthy individuals and were irradiated at doses ranging from 0-1.5 Gy. Microarray technology was employed to identify transcripts that were differentially expressed relative to unirradiated cells 24 hours post-exposure. Statistical analysis identified modulated genes at each of the individual doses. Results Twenty-nine genes were common to all doses with expression levels ranging from 2-10 fold relative to control treatment group. This subset of genes was further assessed in independent complete white blood cell (WBC) populations exposed to either α-particles or X-rays using quantitative real-time PCR. This 29 gene panel was responsive in the α-particle exposed WBCs and was shown to exhibit differential fold-changes compared to X-irradiated cells, though no α-particle specific transcripts were identified. Conclusion Current gene panels for photon radiation may also be applicable for use in α-particle radiation biodosimetry. PMID:25017500

  6. Non-coding RNAs in pluripotency and neural differentiation of human pluripotent stem cells.

    PubMed

    Lukovic, Dunja; Moreno-Manzano, Victoria; Klabusay, Martin; Stojkovic, Miodrag; Bhattacharya, Shomi S; Erceg, Slaven

    2014-01-01

    Several studies have demonstrated the important role of non-coding RNAs as regulators of posttranscriptional processes, including stem cells self-renewal and neural differentiation. Human embryonic stem cells (hESCs) and induced pluripotent stem cells (ihPSCs) show enormous potential in regenerative medicine due to their capacity to differentiate to virtually any type of cells of human body. Deciphering the role of non-coding RNAs in pluripotency, self-renewal and neural differentiation will reveal new molecular mechanisms involved in induction and maintenances of pluripotent state as well as triggering these cells toward clinically relevant cells for transplantation. In this brief review we will summarize recently published studies which reveal the role of non-coding RNAs in pluripotency and neural differentiation of hESCs and ihPSC.

  7. Regulation of neural stem cell in the human SVZ by trophic and morphogenic factors

    PubMed Central

    Alvarez-Palazuelos, Lucia E.; Robles-Cervantes, Martha S.; Castillo-Velazquez, Gabriel; Rivas-Souza, Mario; Guzman-Muniz, Jorge; Moy-Lopez, Norma; Gonzalez-Castaneda, Rocio E.; Luquin, Sonia; Gonzalez-Perez, Oscar

    2011-01-01

    The subventricular zone (SVZ), lining the lateral ventricular system, is the largest germinal region in mammals. In there, neural stem cells express markers related to astoglial lineage that give rise to new neurons and oligodendrocytes in vivo. In the adult human brain, in vitro evidence has also shown that astrocytic cells isolated from the SVZ can generate new neurons and oligodendrocytes. These proliferative cells are strongly controlled by a number of signals and molecules that modulate, activate or repress the cell division, renewal, proliferation and fate of neural stem cells. In this review, we summarize the cellular composition of the adult human SVZ (hSVZ) and discuss the increasing evidence showing that some trophic modulators strongly control the function of neural stem cells in the SVZ. PMID:22053150

  8. 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

  9. Cell surface differences of Naegleria fowleri and Naegleria lovaniensis exposed with surface markers.

    PubMed

    González-Robles, Arturo; Castañón, Guadalupe; Cristóbal-Ramos, Ana Ruth; Hernández-Ramírez, Verónica Ivonne; Omaña-Molina, Maritza; Martínez-Palomo, Adolfo

    2007-12-01

    Differences in the distribution of diverse cell surface coat markers were found between Naegleria fowleri and Naegleria lovaniensis. The presence of carbohydrate-containing components in the cell coat of the two species was detected by selective staining with ruthenium red and alcian blue. Using both markers, N. fowleri presented a thicker deposit than N. lovaniensis. The existence of exposed mannose or glucose residues was revealed by discriminatory agglutination with the plant lectin Concanavalin A. These sugar residues were also visualized at the cell surface of these parasites either by transmission electron microscopy or by fluorescein-tagged Concanavalin A. Using this lectin cap formation was induced only in N. fowleri. The anionic sites on the cell surface detected by means of cationized ferritin were more apparent in N. fowleri. Biotinylation assays confirmed that even though the two amoebae species have some analogous plasma membrane proteins, there is a clear difference in their composition.

  10. Wnt-YAP interactions in the neural fate of human pluripotent stem cells and the implications for neural organoid formation.

    PubMed

    Bejoy, Julie; Song, Liqing; Li, Yan

    2016-01-01

    Human pluripotent stem cells (hPSCs) have shown the ability to self-organize into different types of neural organoids (e.g., whole brain organoids, cortical spheroids, midbrain organoids etc.) recently. The extrinsic and intrinsic signaling elicited by Wnt pathway, Hippo/Yes-associated protein (YAP) pathway, and extracellular microenvironment plays a critical role in brain tissue morphogenesis. This article highlights recent advances in neural tissue patterning from hPSCs, in particular the role of Wnt pathway and YAP activity in this process. Understanding the Wnt-YAP interactions should provide us the guidance to predict and modulate brain-like tissue structure through the regulation of extracellular microenvironment of hPSCs. PMID:26901039

  11. Zinc deficiency impairs the renewal of hippocampal neural stem cells in adult rats: involvement of FoxO3a activation and downstream p27(kip1) expression.

    PubMed

    Han, Jingling; Zhao, Jianya; Jiang, Junkang; Ma, Xia; Liu, Xinhang; Wang, Cheng; Jiang, Shengyang; Wan, Chunhua

    2015-09-01

    Zinc plays an important role in the development and maintenance of central neural system. Zinc deficiency has been known to alter normal brain function, whose molecular mechanism remains largely elusive. In the present study, we established a zinc deficiency-exposed rat model, and, using western blot and immunohistochemical analyses, found that the expression of FoxO3a and p27(kip1) was remarkably up-regulated in the rat brain hippocampus. Immunofluorescence assay showed that FOXO3a and p27(kip1) were significantly co-localized with nestin, the marker of neural stem cells (NSCs). Furthermore, we identified that the proportion of proliferating NSCs was markedly decreased in zinc-deficient rat hippocampaus. Using C17.2 neural stem cells, it was revealed that exposure to zinc chelator N,N,N',N'-tetrakis-(2-pyridylmethy) ethylenediamine induced the expression of FoxO3a and p27(kip1) , which coincided with reduced NSC proliferation. Furthermore, depletion of FoxO3a inhibited p27(kip1) expression and restored the growth of NSCs. On the basis of these data, we concluded that FoxO3a/p27(kip1) signaling might play a significant role in zinc deficiency-induced growth impairment of NSCs and consequent neurological disorders. We describe here that zinc deficiency induces the proliferative impairment of hippocampal neural stem cells partially through the activation of FOXO3a-p27 axis in rats. Neural progenitor cells exhibited significantly up-regulated expression of FOXO3a and p27 after zinc deficiency in vivo and in vitro. Depletion of FOXO3a ameliorates zinc deficiency-induced expression of p27 and growth impairment of neural stem cells. We provide novel insight into the mechanisms underlying zinc deficiency-induced neurological deficits.

  12. Effects of high-LET radiation on neural cells in culture: apoptosis induction, cell toxicity and gene expression

    NASA Astrophysics Data System (ADS)

    Vazquez, M.; Otto, S.; Estevez, L.; Rios, D.; Pena, L.; Anderson, C.

    Despite the fact that some in vivo studies suggest that chronic low-dose exposure to HZE particles might produce effects similar to aging and neurodegeneration, the basic mechanisms of HZE particle neurotoxicity remain to be elucidated. The goal of these experiments is to establish neural cellular models to evaluate the capacity of low- and high-LET radiation, to induce cell damage and apoptosis. In the present study we measured apoptosis, cell toxicity and gene expression induced by low fluences-doses of heavy ions, protons and photons using neuronal precursor cells (NT2, STRATAGENE) and post-mitotic neurons as models for adult neural cell system. Using heavy ions accelerated at AGS (BNL) and HIMAC (Chiba, Japan), and protons (Loma Linda) we study the neurotoxic effects of a variety of heavy particles (1 and 0.6 GeV/n Fe, 580 MeV/n Si, 290 MeV/n C, 550 MeV/n Ar; LET ranging from 13 to148 keV/μm), and 255 MeV/n protons. Apoptosis Induction: We measured the induction of apoptosis by flow cytometry using a FACSCalibur to detect the expression of Annexin V, as an early marker in the apoptotic pathway, in NT-2 cells. The ApoAlert Annexin V assay is based on the observation that soon after initiating apoptosis, most cell types translocate phosphatidylserine (PS) from the inner face of the plasma membrane to the cell surface. Once on the cell surface, PS can be easily detected by staining with a FITC conjugate of Annexin V, a protein that has a strong natural affinity for PS. Externalization of PS occurs earlier than the nuclear changes associated with apoptosis, so the ApoAlert Assay detects apoptotic cells significantly earlier than do DNA-based assays. Exposing NT-2 cells to Fe ions and protons induced a strong dose- and time-dependent induction of apoptosis with the peak of apoptosis appearing at 72 hours post-irradiation. It was determined that Fe ion exposure were more effective to induce apoptosis in comparison to protons and gamma rays, suggesting an high RBE

  13. Phenotypic Modifications in Staphylococcus aureus Cells Exposed to High Concentrations of Vancomycin and Teicoplanin

    PubMed Central

    Gonçalves, Fábio D. A.; de Carvalho, Carla C. C. R.

    2016-01-01

    Bacterial cells are known to change the fatty acid (FA) composition of the phospholipids as a phenotypic response to environmental conditions and to the presence of toxic compounds such as antibiotics. In the present study, Staphylococcus aureus cells collected during the exponential growth phase were challenged with 50 and 100 mg/L of vancomycin and teicoplanin, which are concentrations high enough to kill the large majority of the cell population. Colony-forming unit counts showed biphasic killing kinetics, typical for persister cell enrichment, in both antibiotics and concentrations tested. However, fluorescence microscopy showed the existence of viable but non-culturable (VBNC) cells in a larger number than that of possible persister cells. The analysis of the FA composition of the cells showed that, following antibiotic exposure up to 6 h, the survivor cells have an increased percentage of saturated FAs, a significant reduced percentage of branched FAs and an increased iso/anteiso branched FA ratio when compared to cells exhibiting a regular phenotype. This should result in lower membrane fluidity. However, cells exposed for 8–24 h presented an increased branched/saturated and lower iso/anteiso branched FA ratios, and thus increased membrane fluidity. Furthermore, the phenotypic changes were transmitted to daughter cells grown in drug-free media. The fact that VBNC cells presented nearly the same FA composition as those obtained after cell growth in drug-free media, which could only be the result of growth of persister cells, suggest that VBNC and persister phenotypes share the same type of response to antibiotics at the lipid level. PMID:26834731

  14. Phenotypic Modifications in Staphylococcus aureus Cells Exposed to High Concentrations of Vancomycin and Teicoplanin.

    PubMed

    Gonçalves, Fábio D A; de Carvalho, Carla C C R

    2016-01-01

    Bacterial cells are known to change the fatty acid (FA) composition of the phospholipids as a phenotypic response to environmental conditions and to the presence of toxic compounds such as antibiotics. In the present study, Staphylococcus aureus cells collected during the exponential growth phase were challenged with 50 and 100 mg/L of vancomycin and teicoplanin, which are concentrations high enough to kill the large majority of the cell population. Colony-forming unit counts showed biphasic killing kinetics, typical for persister cell enrichment, in both antibiotics and concentrations tested. However, fluorescence microscopy showed the existence of viable but non-culturable (VBNC) cells in a larger number than that of possible persister cells. The analysis of the FA composition of the cells showed that, following antibiotic exposure up to 6 h, the survivor cells have an increased percentage of saturated FAs, a significant reduced percentage of branched FAs and an increased iso/anteiso branched FA ratio when compared to cells exhibiting a regular phenotype. This should result in lower membrane fluidity. However, cells exposed for 8-24 h presented an increased branched/saturated and lower iso/anteiso branched FA ratios, and thus increased membrane fluidity. Furthermore, the phenotypic changes were transmitted to daughter cells grown in drug-free media. The fact that VBNC cells presented nearly the same FA composition as those obtained after cell growth in drug-free media, which could only be the result of growth of persister cells, suggest that VBNC and persister phenotypes share the same type of response to antibiotics at the lipid level. PMID:26834731

  15. Adaptive responses and apoptosis in endothelial cells exposed to carbon monoxide

    PubMed Central

    Thom, Stephen R.; Fisher, Donald; Xu, Y. Anne; Notarfrancesco, Kathy; Ischiropoulos, Harry

    2000-01-01

    Prior studies have shown that exposure to carbon monoxide (CO) will elevate the steady-state concentration of nitric oxide (⋅NO) in several cell types and body organs and that some toxic effects of CO are directed toward endothelial cells. Studies reported in this paper were conducted with bovine pulmonary artery endothelial cells exposed to 10 to 100 ppm CO to achieve concentrations between 11 and 110 nM in air-saturated buffer. Exposure to 11 nM CO increased synthesis of manganous superoxide dismutase and conferred resistance against the lethal effects of 110 nM CO. At concentrations of 88 nM CO or more, exposures for 1 h or longer caused cell death that became apparent 18 h after the exposure ceased. Caspase-1 was activated in response to CO, and cell death was inhibited by a caspase-1 inhibitor. Alteration of proteolytic pathways by CO was indicated by the presence of ubiquitin-containing intracellular inclusion bodies. Morphological changes and caspase activation indicated that cell death was an apoptotic process. Cells exposed to 110 nM CO had higher concentrations of manganous superoxide dismutase and heme oxygenase-1 but no changes in glutathione peroxidase, glucose-6-phosphate dehydrogenase, thiols, or catalase. Elevated levels of antioxidant enzymes and apoptosis were inhibited by the nitric oxide synthase inhibitor, S-isopropylisothiourea, and the peroxynitrite scavenger, selenomethionine. These results show that biochemical effects of CO occur at environmentally relevant concentrations, that apoptotic cell death follows exposure to relatively high concentrations of CO, and that these actions of CO are mediated by nitric oxide. PMID:10655526

  16. Assessment of DNA integrity (COMET assay) in sperm cells of boron-exposed workers.

    PubMed

    Duydu, Yalçin; Başaran, Nurşen; Ustündağ, Aylin; Aydin, Sevtap; Undeğer, Ulkü; Ataman, Osman Yavuz; Aydos, Kaan; Düker, Yalçin; Ickstadt, Katja; Waltrup, Britta Schulze; Golka, Klaus; Bolt, Hermann M

    2012-01-01

    An extension of a male reproductive study conducted in a boric acid/borate production zone at Bandırma, Turkey, is presented. The relation between DNA-strand breaks (COMET assay, neutral and alkaline version) in sperm cells and previously described sperm quality parameters was investigated in boron-exposed males. A correlation between blood boron levels and mean DNA-strand breaks in sperm was weak, and DNA-strand breaks in sperm were statistically not different between control and exposed groups. Therefore, increasing boron exposures had no additional contribution in addition to already pre-existing DNA-strand breaks in the sperm cells. Weak but statistically significant correlations between DNA-strand breaks and motility/morphology parameters of sperm samples were observed in the neutral version of the COMET assay, while correlations between the same variables were statistically not significant in the alkaline version. A likely reason for these negative results, even in highly exposed humans, is that experimental exposures that had led to reproductive toxicity in animals were significantly higher than any boron exposures, which may be reached under realistic human conditions.

  17. Isolation and Culture of Neural Crest Stem Cells from Human Hair Follicles.

    PubMed

    Yang, Ruifeng; Xu, Xiaowei

    2016-01-01

    The hair follicle undergoes lifelong cycling and growth. Previous studies have been focused on epithelial stem cells in the hair follicles. Neural crest stem cells (NCSCs) are pluripotent cells that can persist in adult tissues. We have previously demonstrated that human NCSCs can be isolated from hair follicles. Here, we present a protocol to isolate NCSCs from human hair follicles based on their specific surface-marker expression of CD271/HNK1 or CD271/CD49D (alpha4 integrin). NCSCs can be expanded in the culture as neural spheres or attached cells.

  18. The impact of neural stem cell biology on CNS carcinogenesis and tumor types.

    PubMed

    Kurian, K M

    2011-01-01

    The incidence of gliomas is on the increase, according to epidemiological data. This increase is a conundrum because the brain is in a privileged protected site behind the blood-brain barrier, and therefore partially buffered from environmental factors. In addition the brain also has a very low proliferative potential compared with other parts of the body. Recent advances in neural stem cell biology have impacted on our understanding of CNS carcinogenesis and tumor types. This article considers the cancer stem cell theory with regard to CNS cancers, whether CNS tumors arise from human neural stem cells and whether glioma stem cells can be reprogrammed.

  19. Experimental Advances Towards Neural Regeneration from Induced Stem Cells to Direct In Vivo Reprogramming.

    PubMed

    Dametti, Sara; Faravelli, Irene; Ruggieri, Margherita; Ramirez, Agnese; Nizzardo, Monica; Corti, Stefania

    2016-05-01

    Neuronal loss is a common substrate of many neurological diseases that still lack effective treatments and highly burden lives of affected individuals. The discovery of self-renewing stem cells within the central nervous system (CNS) has opened the doors to the possibility of using the plasticity of CNS as a potential strategy for the development of regenerative therapies after injuries. The role of neural progenitor cells appears to be crucial, but insufficient in reparative processes after damage. In addition, the mechanisms that regulate these events are still largely unknown. Stem cell-based therapeutic approaches have primarily focused on the use of either induced pluripotent stem cells or induced neural stem cells as sources for cell transplantation. More recently, in vivo direct reprogramming of endogenous CNS cells into multipotent neural stem/progenitor cells has been proposed as an alternative strategy that could overcome the limits connected with both the invasiveness of exogenous cell transplantation and the technical issues of in vitro reprogramming (i.e., the time requested and the limited available amount of directly induced neuronal cells). In this review, we aim to highlight the recent studies on in vivo direct reprogramming, focusing on astrocytes conversion to neurons or to neural stem/precursors cells, in the perspective of future therapeutic purposes for neurological disorders.

  20. Morphological changes in human neural cells following tick-borne encephalitis virus infection.

    PubMed

    Růzek, Daniel; Vancová, Marie; Tesarová, Martina; Ahantarig, Arunee; Kopecký, Jan; Grubhoffer, Libor

    2009-07-01

    Tick-borne encephalitis (TBE) is one of the leading and most dangerous human viral neuroinfections in Europe and north-eastern Asia. The clinical manifestations include asymptomatic infections, fevers and debilitating encephalitis that might progress into chronic disease or fatal infection. To understand TBE pathology further in host nervous systems, three human neural cell lines, neuroblastoma, medulloblastoma and glioblastoma, were infected with TBE virus (TBEV). The susceptibility and virus-mediated cytopathic effect, including ultrastructural and apoptotic changes of the cells, were examined. All the neural cell lines tested were susceptible to TBEV infection. Interestingly, the neural cells produced about 100- to 10,000-fold higher virus titres than the conventional cell lines of extraneural origin, indicating the highly susceptible nature of neural cells to TBEV infection. The infection of medulloblastoma and glioblastoma cells was associated with a number of major morphological changes, including proliferation of membranes of the rough endoplasmic reticulum and extensive rearrangement of cytoskeletal structures. The TBEV-infected cells exhibited either necrotic or apoptotic morphological features. We observed ultrastructural apoptotic signs (condensation, margination and fragmentation of chromatin) and other alterations, such as vacuolation of the cytoplasm, dilatation of the endoplasmic reticulum cisternae and shrinkage of cells, accompanied by a high density of the cytoplasm. On the other hand, infected neuroblastoma cells did not exhibit proliferation of membranous structures. The virions were present in both the endoplasmic reticulum and the cytoplasm. Cells were dying preferentially by necrotic mechanisms rather than apoptosis. The neuropathological significance of these observations is discussed. PMID:19264624

  1. Efficient differentiation of neural stem cells induced by the rat bone marrow stromal cells

    PubMed Central

    Gu, Ping; Qiu, Fu-Cheng; Han, Rui; Zhang, Zhong-Xia; Dong, Ci; Zhang, Li-Na; Wang, Yan-Yong; Ma, Qing-Ying; Yan, Bao-Yong

    2015-01-01

    Neural stem cells (NSCs) are valuable self-renewing cells that can maintain the capacity to differentiate into specific brain cell types. NSCs may repair and even replace the brain tissue, and ultimatley promoting the central nervous system regeneration. Therefore, it is important, for scientists and pjysicians, to study the method for efficient culture and differentiation of NSCs. Our previous study demonstrated that Bone Marrow Stromal Cells (BMSCs) can directly regulate the differentiation of NSCs into neurons, and soluble molecules excreted by BMSCs played a key role in this process. Hereby, we further identified the BMSCs-induced neurons could form the synapses, convey dopamine and express voltage-depend and receptor-depend calcium channels. Moreover, the extracellular signal-regulated protein kinase ERK1/2 pathway was founded to be involved in the process of neuron differentiation and proliferation by the in vitro experiments. Finally, by using protein array, we, for the first time, found that the cytokine-induced neutrophil chemoattractant-3 (CINC-3, a small molecule cytokine) can promote the leukocytes invasion into the inflammation site, and have the ability to induce mesencephal NSCs into neurons. Consequently, these positive findings suggested that our BMSCs-induced culture system could provide a useful tool to investigate the molecular mechanisms of neural differentiation of NSCs, which may be benifical for neurodegenerative diseases in the near future. PMID:26221209

  2. Treatment of multiple sclerosis by transplantation of neural stem cells derived from induced pluripotent stem cells.

    PubMed

    Zhang, Chao; Cao, Jiani; Li, Xiaoyan; Xu, Haoyu; Wang, Weixu; Wang, Libin; Zhao, Xiaoyang; Li, Wei; Jiao, Jianwei; Hu, Baoyang; Zhou, Qi; Zhao, Tongbiao

    2016-09-01

    Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS), with focal T lymphocytic infiltration and damage of myelin and axons. The underlying mechanism of pathogenesis remains unclear and there are currently no effective treatments. The development of neural stem cell (NSC) transplantation provides a promising strategy to treat neurodegenerative disease. However, the limited availability of NSCs prevents their application in neural disease therapy. In this study, we generated NSCs from induced pluripotent stem cells (iPSCs) and transplanted these cells into mice with experimental autoimmune encephalomyelitis (EAE), a model of MS. The results showed that transplantation of iPSC-derived NSCs dramatically reduced T cell infiltration and ameliorated white matter damage in the treated EAE mice. Correspondingly, the disease symptom score was greatly decreased, and motor ability was dramatically rescued in the iPSC-NSC-treated EAE mice, indicating the effectiveness of using iPSC-NSCs to treat MS. Our study provides pre-clinical evidence to support the feasibility of treating MS by transplantation of iPSC-derived NSCs. PMID:27233903

  3. Chromosomal Aberrations in Normal and AT Cells Exposed to High Dose of Low Dose Rate Irradiation

    NASA Technical Reports Server (NTRS)

    Kawata, T.; Shigematsu, N.; Kawaguchi, O.; Liu, C.; Furusawa, Y.; Hirayama, R.; George, K.; Cucinotta, F.

    2011-01-01

    Ataxia telangiectasia (A-T) is a human autosomally recessive syndrome characterized by cerebellar ataxia, telangiectases, immune dysfunction, and genomic instability, and high rate of cancer incidence. A-T cell lines are abnormally sensitive to agents that induce DNA double strand breaks, including ionizing radiation. The diverse clinical features in individuals affected by A-T and the complex cellular phenotypes are all linked to the functional inactivation of a single gene (AT mutated). It is well known that cells deficient in ATM show increased yields of both simple and complex chromosomal aberrations after high-dose-rate irradiation, but, less is known on how cells respond to low-dose-rate irradiation. It has been shown that AT cells contain a large number of unrejoined breaks after both low-dose-rate irradiation and high-dose-rate irradiation, however sensitivity for chromosomal aberrations at low-dose-rate are less often studied. To study how AT cells respond to low-dose-rate irradiation, we exposed confluent normal and AT fibroblast cells to up to 3 Gy of gamma-irradiation at a dose rate of 0.5 Gy/day and analyzed chromosomal aberrations in G0 using fusion PCC (Premature Chromosomal Condensation) technique. Giemsa staining showed that 1 Gy induces around 0.36 unrejoined fragments per cell in normal cells and around 1.35 fragments in AT cells, whereas 3Gy induces around 0.65 fragments in normal cells and around 3.3 fragments in AT cells. This result indicates that AT cells can rejoin breaks less effectively in G0 phase of the cell cycle? compared to normal cells. We also analyzed chromosomal exchanges in normal and AT cells after exposure to 3 Gy of low-dose-rate rays using a combination of G0 PCC and FISH techniques. Misrejoining was detected in the AT cells only? When cells irradiated with 3 Gy were subcultured and G2 chromosomal aberrations were analyzed using calyculin-A induced PCC technique, the yield of unrejoined breaks decreased in both normal and AT

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

    PubMed

    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.

  5. Nifurtimox Is Effective Against Neural Tumor Cells and Is Synergistic with Buthionine Sulfoximine

    PubMed Central

    Du, Michael; Zhang, Linna; Scorsone, Kathleen A.; Woodfield, Sarah E.; Zage, Peter E.

    2016-01-01

    Children with aggressive neural tumors have poor survival rates and novel therapies are needed. Previous studies have identified nifurtimox and buthionine sulfoximine (BSO) as effective agents in children with neuroblastoma and medulloblastoma. We hypothesized that nifurtimox would be effective against other neural tumor cells and would be synergistic with BSO. We determined neural tumor cell viability before and after treatment with nifurtimox using MTT assays. Assays for DNA ladder formation and poly-ADP ribose polymerase (PARP) cleavage were performed to measure the induction of apoptosis after nifurtimox treatment. Inhibition of intracellular signaling was measured by Western blot analysis of treated and untreated cells. Tumor cells were then treated with combinations of nifurtimox and BSO and evaluated for viability using MTT assays. All neural tumor cell lines were sensitive to nifurtimox, and IC50 values ranged from approximately 20 to 210 μM. Nifurtimox treatment inhibited ERK phosphorylation and induced apoptosis in tumor cells. Furthermore, the combination of nifurtimox and BSO demonstrated significant synergistic efficacy in all tested cell lines. Additional preclinical and clinical studies of the combination of nifurtimox and BSO in patients with neural tumors are warranted. PMID:27282514

  6. Changes of neural markers expression during late neurogenic differentiation of human adipose-derived stem cells

    PubMed Central

    Razavi, Shahnaz; Khosravizadeh, Zahra; Bahramian, Hamid; Kazemi, Mohammad

    2015-01-01

    Background: Different studies have been done to obtain sufficient number of neural cells for treatment of neurodegenerative diseases, spinal cord, and traumatic brain injury because neural stem cells are limited in central nerves system. Recently, several studies have shown that adipose-derived stem cells (ADSCs) are the appropriate source of multipotent stem cells. Furthermore, these cells are found in large quantities. The aim of this study was an assessment of proliferation and potential of neurogenic differentiation of ADSCs with passing time. Materials and Methods: Neurosphere formation was used for neural induction in isolated human ADSCs (hADSCs). The rate of proliferation was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and potential of neural differentiation of induced hADSCs was evaluated by immunocytochemical and real-time reverse transcription polymerase chain reaction analysis after 10 and 14 days post-induction. Results: The rate of proliferation of induced hADSCs increased after 14 days while the expression of nestin, glial fibrillary acidic protein, and microtubule-associated protein 2 was decreased with passing time during neurogenic differentiation. Conclusion: These findings showed that the proliferation of induced cells increased with passing time, but in early neurogenic differentiation of hADSCs, neural expression was higher than late of differentiation. Thus, using of induced cells in early differentiation may be suggested for in vivo application. PMID:26605238

  7. miR-430 regulates oriented cell division during neural tube development in zebrafish.

    PubMed

    Takacs, Carter M; Giraldez, Antonio J

    2016-01-15

    MicroRNAs have emerged as critical regulators of gene expression. Originally shown to regulate developmental timing, microRNAs have since been implicated in a wide range of cellular functions including cell identity, migration and signaling. miRNA-430, the earliest expressed microRNA during zebrafish embryogenesis, is required to undergo morphogenesis and has previously been shown to regulate maternal mRNA clearance, Nodal signaling, and germ cell migration. The functions of miR-430 in brain morphogenesis, however, remain unclear. Herein we find that miR-430 instructs oriented cell divisions in the neural rod required for neural midline formation. Loss of miR-430 function results in mitotic spindle misorientation in the neural rod, failed neuroepithelial integration after cell division, and ectopic cell accumulation in the dorsal neural tube. We propose that miR-430, independently of canonical apicobasal and planar cell polarity (PCP) pathways, coordinates the stereotypical cell divisions that instruct neural tube morphogenesis.

  8. Neural differentiation of mouse embryonic stem cells studied by FTIR spectroscopy

    NASA Astrophysics Data System (ADS)

    Tanthanuch, Waraporn; Thumanu, Kanjana; Lorthongpanich, Chanchao; Parnpai, Rangsun; Heraud, Philip

    2010-04-01

    Embryonic Stem-derived Neural Cells (ESNCs) hold potential as a source of neurons for a cell-based therapy for the treatment of brain tumors, and other neurological diseases and disorders in the future. The sorting of neural cell types is envisaged to be one of the most important processed for clinical application of these cells in cell-based therapies of the central nervous system (CNS). In this study, laboratory-based FTIR and Synchrotron-FTIR (SR-FTIR) microspectroscopy were used to identify FTIR marker for distinguishing different neural cell types derived from the differentiation of mouse embryonic stem cells (mESCs). Principal Component Analysis (PCA) and Unsupervised Hierarchical Cluster Analysis (UHCA) were shown to be able to distinguish the developmental stage of mESCs into three cell types: embryoid bodies (EBs), neural progenitor cells (NPCs), and ESNCs. Moreover, PCA provided the mean for identifying potential FTIR "marker bands" that underwent dramatic changes during stem cell differentiation along neural lineages. These appeared to be associated with changes in lipids (bands from CH 2 and CH 3 stretching vibrations at ˜2959, 2923 and 2852 cm -1) and proteins (changes in the amide I band at ˜1659 and 1637 cm -1). The results suggested that lipid content of cells increased significantly over the time of differentiation, suggesting increased expression of glycerophospholipids. Changes in the amide I profile, suggested concomitant increases in α-helix rich proteins as mESCs differentiated towards ESNCs, with a corresponding decrease in β-sheet rich proteins, corresponding with changes in cytoskeleton protein which may have been taking place involved with the establishment of neural structure and function.

  9. Dynamic methylation and expression of Oct4 in early neural stem cells.

    PubMed

    Lee, Shih-Han; Jeyapalan, Jennie N; Appleby, Vanessa; Mohamed Noor, Dzul Azri; Sottile, Virginie; Scotting, Paul J

    2010-09-01

    Neural stem cells are a multipotent population of tissue-specific stem cells with a broad but limited differentiation potential. However, recent studies have shown that over-expression of the pluripotency gene, Oct4, alone is sufficient to initiate a process by which these can form 'induced pluripotent stem cells' (iPS cells) with the same broad potential as embryonic stem cells. This led us to examine the expression of Oct4 in endogenous neural stem cells, as data regarding its expression in neural stem cells in vivo are contradictory and incomplete. In this study we have therefore analysed the expression of Oct4 and other genes associated with pluripotency throughout development of the mouse CNS and in neural stem cells grown in vitro. We find that Oct4 is still expressed in the CNS by E8.5, but that this expression declines rapidly until it is undetectable by E15.5. This decline is coincident with the gradual methylation of the Oct4 promoter and proximal enhancer. Immunostaining suggests that the Oct4 protein is predominantly cytoplasmic in location. We also found that neural stem cells from all ages expressed the pluripotency associated genes, Sox2, c-Myc, Klf4 and Nanog. These data provide an explanation for the varying behaviour of cells from the early neuroepithelium at different stages of development. The expression of these genes also provides an indication of why Oct4 alone is sufficient to induce iPS formation in neural stem cells at later stages. PMID:20646110

  10. Optimizing protocols for imaging neural cells and tissues using functionalized quantum dots

    NASA Astrophysics Data System (ADS)

    Pathak, Smita; Silva, Gabriel A.

    2008-02-01

    Chemically functionalized semiconductor quantum dot protocols were optimized for the specific labeling and imaging of neural cells, both neurons and macroglial cells. Beta-tubulin III was used to image primary cortical neurons and PC12 cells while glial fibrillary acidic protein (GFAP) was used to image primary spinal cord and cortical astrocytes and the rMC-1 retinal glial Muller cell line. Both proteins are the main components of intermediate filaments and are specific to the two classes of neural cells. We also specifically labeled and imaged at high resolutions using anti-GFAP conjugated quantum dots glial scars in situ in intact neural sensory retina in a rodent model of macular degeneration.

  11. Low Density Lipoprotein Receptor Related Proteins as Regulators of Neural Stem and Progenitor Cell Function

    PubMed Central

    Landowski, Lila M.; Young, Kaylene M.

    2016-01-01

    The central nervous system (CNS) is a highly organised structure. Many signalling systems work in concert to ensure that neural stem cells are appropriately directed to generate progenitor cells, which in turn mature into functional cell types including projection neurons, interneurons, astrocytes, and oligodendrocytes. Herein we explore the role of the low density lipoprotein (LDL) receptor family, in particular family members LRP1 and LRP2, in regulating the behaviour of neural stem and progenitor cells during development and adulthood. The ability of LRP1 and LRP2 to bind a diverse and extensive range of ligands, regulate ligand endocytosis, recruit nonreceptor tyrosine kinases for direct signal transduction and signal in conjunction with other receptors, enables them to modulate many crucial neural cell functions. PMID:26949399

  12. Vision-based force measurement using neural networks for biological cell microinjection.

    PubMed

    Karimirad, Fatemeh; Chauhan, Sunita; Shirinzadeh, Bijan

    2014-03-21

    This paper presents a vision-based force measurement method using an artificial neural network model. The proposed model is used for measuring the applied load to a spherical biological cell during micromanipulation process. The devised vision-based method is most useful when force measurement capability is required, but it is very challenging or even infeasible to use a force sensor. Artificial neural networks in conjunction with image processing techniques have been used to estimate the applied load to a cell. A bio-micromanipulation system capable of force measurement has also been established in order to collect the training data required for the proposed neural network model. The geometric characterization of zebrafish embryos membranes has been performed during the penetration of the micropipette prior to piercing. The geometric features are extracted from images using image processing techniques. These features have been used to describe the shape and quantify the deformation of the cell at different indentation depths. The neural network is trained by taking the visual data as the input and the measured corresponding force as the output. Once the neural network is trained with sufficient number of data, it can be used as a precise sensor in bio-micromanipulation setups. However, the proposed neural network model is applicable for indentation of any other spherical elastic object. The results demonstrate the capability of the proposed method. The outcomes of this study could be useful for measuring force in biological cell micromanipulation processes such as injection of the mouse oocyte/embryo. PMID:24411067

  13. Vision-based force measurement using neural networks for biological cell microinjection.

    PubMed

    Karimirad, Fatemeh; Chauhan, Sunita; Shirinzadeh, Bijan

    2014-03-21

    This paper presents a vision-based force measurement method using an artificial neural network model. The proposed model is used for measuring the applied load to a spherical biological cell during micromanipulation process. The devised vision-based method is most useful when force measurement capability is required, but it is very challenging or even infeasible to use a force sensor. Artificial neural networks in conjunction with image processing techniques have been used to estimate the applied load to a cell. A bio-micromanipulation system capable of force measurement has also been established in order to collect the training data required for the proposed neural network model. The geometric characterization of zebrafish embryos membranes has been performed during the penetration of the micropipette prior to piercing. The geometric features are extracted from images using image processing techniques. These features have been used to describe the shape and quantify the deformation of the cell at different indentation depths. The neural network is trained by taking the visual data as the input and the measured corresponding force as the output. Once the neural network is trained with sufficient number of data, it can be used as a precise sensor in bio-micromanipulation setups. However, the proposed neural network model is applicable for indentation of any other spherical elastic object. The results demonstrate the capability of the proposed method. The outcomes of this study could be useful for measuring force in biological cell micromanipulation processes such as injection of the mouse oocyte/embryo.

  14. Competitive binding of pentraxins and IgM to newly exposed epitopes on late apoptotic cells.

    PubMed

    Ciurana, Caroline L F; Hack, C Erik

    2006-01-01

    A random distribution of phospholipids among the inner and outer leaflet of the cell membrane occurs during apoptosis and is known as membrane flip-flop. Flip-flopped cells have binding sites for various plasma proteins, such as IgM and the pentraxins C-reactive protein (CRP) and serum amyloid P component (SAP). In this study, we investigated whether pentraxins and IgM antibodies recognize the same binding sites on apoptotic cells, and whether phospholipids constitute these binding sites. Except for SAP which also bound to early apoptotic cells, pentraxins and IgM preferentially bound to late apoptotic cells. Competition experiments with different phosphatemonoesters revealed that CRP and SAP as well as part of the IgM bound to the phospholipids head groups, SAP mainly to phosphorylethanolamine, CRP to phosphorylcholine and phosphorylethanolamine and to a lesser extent to phosphorylserine, and IgM to phosphorylcholine and phosphorylserine. These results were confirmed in experiments in which proteins were adsorbed from plasma with artificial phospholipids particles. IgM and the pentraxins variably competed for the same binding sites on late apoptotic cells, SAP having the highest and CRP the lowest apparent affinity. We conclude that CRP, SAP, and part of the IgM bind to the phospholipid head groups exposed on apoptotic cells. This shared specificity as well as their shared capability to activate complement, suggest that IgM and the pentraxins CRP and SAP exert similar functions in the removal of apoptotic cells.

  15. Single Walled Carbon Nanotubes Exhibit Dual-Phase Regulation to Exposed Arabidopsis Mesophyll Cells

    PubMed Central

    2011-01-01

    Herein we are the first to report that single-walled carbon nanotubes (SWCNTs) exhibit dual-phase regulation to Arabidopsis mesophyll cells exposed to different concentration of SWCNTs. The mesophyll protoplasts were prepared by enzyme digestion, and incubated with 15, 25, 50, 100 μg/ml SWCNTs for 48 h, and then were observed by optical microscopy and transmission electron microscopy, the reactive oxygen species (ROS) generation was measured. Partial protoplasts were stained with propidium iodide and 4'-6- diamidino-2-phenylindole, partial protoplasts were incubated with fluorescein isothiocyanate-labeled SWCNTs, and observed by fluorescence microscopy. Results showed that SWCNTs could traverse both the plant cell wall and cell membrane, with less than or equal to 50 μg/ml in the culture medium, SWCNTs stimulated plant cells to grow out trichome clusters on their surface, with more than 50 μg/ml SWCNTs in the culture medium, SWCNTs exhibited obvious toxic effects to the protoplasts such as increasing generation of ROS, inducing changes of protoplast morphology, changing green leaves into yellow, and inducing protoplast cells' necrosis and apoptosis. In conclusion, single walled carbon nanotubes can get through Arabidopsis mesophyll cell wall and membrane, and exhibit dose-dependent dual-phase regulation to Arabidopsis mesophyll protoplasts such as low dose stimulating cell growth, and high dose inducing cells' ROS generation, necrosis or apoptosis. PMID:27502666

  16. Cytoskeleton Modifications and Autophagy Induction in TCam-2 Seminoma Cells Exposed to Simulated Microgravity

    PubMed Central

    Ferranti, Francesca; Caruso, Maria; Cammarota, Marcella; Fabrizi, Cinzia; Fumagalli, Lorenzo; Schiraldi, Chiara; Catizone, Angela

    2014-01-01

    The study of how mechanical forces may influence cell behavior via cytoskeleton remodeling is a relevant challenge of nowadays that may allow us to define the relationship between mechanics and biochemistry and to address the larger problem of biological complexity. An increasing amount of literature data reported that microgravity condition alters cell architecture as a consequence of cytoskeleton structure modifications. Herein, we are reporting the morphological, cytoskeletal, and behavioral modifications due to the exposition of a seminoma cell line (TCam-2) to simulated microgravity. Even if no differences in cell proliferation and apoptosis were observed after 24 hours of exposure to simulated microgravity, scanning electron microscopy (SEM) analysis revealed that the change of gravity vector significantly affects TCam-2 cell surface morphological appearance. Consistent with this observation, we found that microtubule orientation is altered by microgravity. Moreover, the confocal analysis of actin microfilaments revealed an increase in the cell width induced by the low gravitational force. Microtubules and microfilaments have been related to autophagy modulation and, interestingly, we found a significant autophagic induction in TCam-2 cells exposed to simulated microgravity. This observation is of relevant interest because it shows, for the first time, TCam-2 cell autophagy as a biological response induced by a mechanical stimulus instead of a biochemical one. PMID:25140323

  17. Insights on cryoprotectant toxicity from gene expression profiling of endothelial cells exposed to ethylene glycol.

    PubMed

    Cordeiro, Rui Martins; Stirling, Soren; Fahy, Gregory M; de Magalhães, João Pedro

    2015-12-01

    Cryopreservation consists of preserving living cells or tissues generally at -80 °C or below and has many current applications in cell and tissue banking, and future potential for organ banking. Cryoprotective agents such as ethylene glycol (EG) are required for successful cryopreservation of most living systems, but have toxic side effects whose mechanisms remain largely unknown. In this work, we investigated the mechanisms of toxicity of ethylene glycol in human umbilical vein endothelial cells (HUVECs) as a model of the vascular endothelium in perfused organs. Exposing cells to 60% v/v EG for 2 h at 4 °C resulted in only a slight decrease in subsequent cell growth, suggesting only modest toxicity of EG for this cell type. Gene expression analysis with whole genome microarrays revealed signatures indicative of a generalized stress response at 24 h after EG exposure and a trend toward partial recovery at 72 h. The observed changes involved signalling pathways, glycoproteins, and genes involved in extracellular and transmembrane functions, the latter suggesting potential effects of ethylene glycol on membranes. These results continue to develop a new paradigm for understanding cryoprotectant toxicity and reveal molecular signatures helpful for future experiments in more completely elucidating the toxic effects of ethylene glycol in vascular endothelial cells and other cell types. PMID:26471925

  18. Single Walled Carbon Nanotubes Exhibit Dual-Phase Regulation to Exposed Arabidopsis Mesophyll Cells

    NASA Astrophysics Data System (ADS)

    Yuan, Hengguang; Hu, Shanglian; Huang, Peng; Song, Hua; Wang, Kan; Ruan, Jing; He, Rong; Cui, Daxiang

    2011-12-01

    Herein we are the first to report that single-walled carbon nanotubes (SWCNTs) exhibit dual-phase regulation to Arabidopsis mesophyll cells exposed to different concentration of SWCNTs. The mesophyll protoplasts were prepared by enzyme digestion, and incubated with 15, 25, 50, 100 μg/ml SWCNTs for 48 h, and then were observed by optical microscopy and transmission electron microscopy, the reactive oxygen species (ROS) generation was measured. Partial protoplasts were stained with propidium iodide and 4'-6- diamidino-2-phenylindole, partial protoplasts were incubated with fluorescein isothiocyanate-labeled SWCNTs, and observed by fluorescence microscopy. Results showed that SWCNTs could traverse both the plant cell wall and cell membrane, with less than or equal to 50 μg/ml in the culture medium, SWCNTs stimulated plant cells to grow out trichome clusters on their surface, with more than 50 μg/ml SWCNTs in the culture medium, SWCNTs exhibited obvious toxic effects to the protoplasts such as increasing generation of ROS, inducing changes of protoplast morphology, changing green leaves into yellow, and inducing protoplast cells' necrosis and apoptosis. In conclusion, single walled carbon nanotubes can get through Arabidopsis mesophyll cell wall and membrane, and exhibit dose-dependent dual-phase regulation to Arabidopsis mesophyll protoplasts such as low dose stimulating cell growth, and high dose inducing cells' ROS generation, necrosis or apoptosis.

  19. Cytoskeleton modifications and autophagy induction in TCam-2 seminoma cells exposed to simulated microgravity.

    PubMed

    Ferranti, Francesca; Caruso, Maria; Cammarota, Marcella; Masiello, Maria Grazia; Corano Scheri, Katia; Fabrizi, Cinzia; Fumagalli, Lorenzo; Schiraldi, Chiara; Cucina, Alessandra; Catizone, Angela; Ricci, Giulia

    2014-01-01

    The study of how mechanical forces may influence cell behavior via cytoskeleton remodeling is a relevant challenge of nowadays that may allow us to define the relationship between mechanics and biochemistry and to address the larger problem of biological complexity. An increasing amount of literature data reported that microgravity condition alters cell architecture as a consequence of cytoskeleton structure modifications. Herein, we are reporting the morphological, cytoskeletal, and behavioral modifications due to the exposition of a seminoma cell line (TCam-2) to simulated microgravity. Even if no differences in cell proliferation and apoptosis were observed after 24 hours of exposure to simulated microgravity, scanning electron microscopy (SEM) analysis revealed that the change of gravity vector significantly affects TCam-2 cell surface morphological appearance. Consistent with this observation, we found that microtubule orientation is altered by microgravity. Moreover, the confocal analysis of actin microfilaments revealed an increase in the cell width induced by the low gravitational force. Microtubules and microfilaments have been related to autophagy modulation and, interestingly, we found a significant autophagic induction in TCam-2 cells exposed to simulated microgravity. This observation is of relevant interest because it shows, for the first time, TCam-2 cell autophagy as a biological response induced by a mechanical stimulus instead of a biochemical one.

  20. Cranial neural crest cell migration in the Australian lungfish, Neoceratodus forsteri.

    PubMed

    Falck, P; Joss, J; Olsson, L

    2000-01-01

    A crucial role for the cranial neural crest in head development has been established for both actinopterygian fishes and tetrapods. It has been claimed, however, that the neural crest is unimportant for head development in the Australian lungfish (Neoceratodus forsteri), a member of the group (Dipnoi) which is commonly considered to be the living sister group of the tetrapods. In the present study, we used scanning electron microscopy to study cranial neural crest development in the Australian lungfish. Our results, contrary to those of Kemp, show that cranial neural crest cells do emerge and migrate in the Australian lungfish in the same way as in other vertebrates, forming mandibular, hyoid, and branchial streams. The major difference is in the timing of the onset of cranial neural crest migration. It is delayed in the Australian lungfish in comparison with their living sister group the Lissamphibia. Furthermore, the delay in timing between the emergence of the hyoid and branchial crest streams is very long, indicating a steeper anterior-posterior gradient than in amphibians. We are now extending our work on lungfish head development to include experimental studies (ablation of selected streams of neural crest cells) and fate mapping (using fluoresent tracer dyes such as Dil) to document the normal fate as well as the role in head patterning of the cranial neural crest in the Australian lungfish.

  1. Exposure to 1800 MHz radiofrequency radiation impairs neurite outgrowth of embryonic neural stem cells.

    PubMed

    Chen, Chunhai; Ma, Qinlong; Liu, Chuan; Deng, Ping; Zhu, Gang; Zhang, Lei; He, Mindi; Lu, Yonghui; Duan, Weixia; Pei, Liping; Li, Min; Yu, Zhengping; Zhou, Zhou

    2014-05-29

    A radiofrequency electromagnetic field (RF-EMF) of 1800 MHz is widely used in mobile communications. However, the effects of RF-EMFs on cell biology are unclear. Embryonic neural stem cells (eNSCs) play a critical role in brain development. Thus, detecting the effects of RF-EMF on eNSCs is important for exploring the effects of RF-EMF on brain development. Here, we exposed eNSCs to 1800 MHz RF-EMF at specific absorption rate (SAR) values of 1, 2, and 4 W/kg for 1, 2, and 3 days. We found that 1800 MHz RF-EMF exposure did not influence eNSC apoptosis, proliferation, cell cycle or the mRNA expressions of related genes. RF-EMF exposure also did not alter the ratio of eNSC differentiated neurons and astrocytes. However, neurite outgrowth of eNSC differentiated neurons was inhibited after 4 W/kg RF-EMF exposure for 3 days. Additionally, the mRNA and protein expression of the proneural genes Ngn1 and NeuroD, which are crucial for neurite outgrowth, were decreased after RF-EMF exposure. The expression of their inhibitor Hes1 was upregulated by RF-EMF exposure. These results together suggested that 1800 MHz RF-EMF exposure impairs neurite outgrowth of eNSCs. More attention should be given to the potential adverse effects of RF-EMF exposure on brain development.

  2. Human Embryonic Stem Cells: A Model for the Study of Neural Development and Neurological Diseases

    PubMed Central

    Prajumwongs, Piya; Weeranantanapan, Oratai; Jaroonwitchawan, Thiranut; Noisa, Parinya

    2016-01-01

    Although the mechanism of neurogenesis has been well documented in other organisms, there might be fundamental differences between human and those species referring to species-specific context. Based on principles learned from other systems, it is found that the signaling pathways required for neural induction and specification of human embryonic stem cells (hESCs) recapitulated those in the early embryo development in vivo at certain degree. This underscores the usefulness of hESCs in understanding early human neural development and reinforces the need to integrate the principles of developmental biology and hESC biology for an efficient neural differentiation. PMID:27239201

  3. Neuralization of mouse embryonic stem cells in alginate hydrogels under retinoic acid and SAG treatment.

    PubMed

    Delivopoulos, Evangelos; Shakesheff, Kevin M; Peto, Heather

    2015-08-01

    This paper examines the differentiation of a mouse embryonic stem cell line (CGR8) into neurons, under retinoic acid (RA) and smoothened agonist (SAG) treatment. When stem cells underwent through an embryoid body (EB) formation stage, dissociation and seeding on glass coverslips, immunofluorescent labelling for neuronal markers (Nestin, b-Tubulin III, MAP2) revealed the presence of both immature neural progenitors and mature neurons. Undifferentiated CGR8 were also encapsulated in tubular, alginate-gelatin hydrogels and incubated in differentiation media containing retinoic acid (RA) and smoothened agonist (SAG). Cryo-sections of the hydrogel tubes were positive for Nestin, Pax6 and b-Tubulin III, verifying the presence of neurons and neural progenitors. Provided neural induction can be more precisely directed in the tubular hydrogels, these scaffolds will become a powerful model of neural tube development in embryos and will highlight potential strategies for spinal cord regeneration. PMID:26737053

  4. Live-cell imaging study of mitochondrial morphology in mammalian cells exposed to X-rays.

    PubMed

    Noguchi, M; Kanari, Y; Yokoya, A; Narita, A; Fujii, K

    2015-09-01

    Morphological changes in mitochondria induced by X-irradiation in normal murine mammary gland cells were studied with a live-cell microscopic imaging technique. Mitochondria were visualised by staining with a specific fluorescent probe in the cells, which express fluorescent ubiquitination-based cell-cycle indicator 2 (Fucci2) probes to visualise cell cycle. In unirradiated cells, the number of cells with fragmented mitochondria was about 20 % of the total cells through observation period (96 h). In irradiated cells, the population with fragmented mitochondria significantly increased depending on the absorbed dose. Particularly, for 8 Gy irradiation, the accumulation of fragmentation persists even in the cells whose cell cycle came to a stand (80 % in G1 (G0-like) phase). The fraction reached to a maximum at 96 h after irradiation. The kinetics of the fraction with fragmented mitochondria was similar to that for cells in S/G2/M phase (20 %) through the observation period (120 h). The evidences show that, in irradiated cells, some signals are continually released from a nucleus or cytoplasm even in the G0-like cells to operate some sort of protein machineries involved in mitochondrial fission. It is inferred that this delayed mitochondrial fragmentation is strongly related to their dysfunction, and hence might modulate radiobiological effects such as mutation or cell death.

  5. The experiment on the protein exposed to acoustic wave in model of hair cells.

    PubMed

    Takeda, H

    1976-01-01

    Why do animals have intense troubles mainly in the basal turns of their cochleae by exposure to sound and the ototoxity? I thought that the protein in cells had an important relation to this subject. So I performed this experiment. I made model of hair cells of silicon and poured three kinds of solutions of protein into them. And I checked the ionization of the protein and its denaturation by giving strong energy of oscillation to them like the same idea of photoelectric effect. As the result, as for the protein liquid, I noticed the increase of volts just after it was exposed to sound and about 3 hours later, I observed the figure decreased gradually. From this phenomenon, I presumed that the protein in sensory cells became a factor of the stimulation by receiving strong energy conversely. Low-percent protein liquid had less effect caused by sound. PMID:7097

  6. QSAR model for predicting cell viability of human embryonic kidney cells exposed to SiO₂ nanoparticles.

    PubMed

    Manganelli, Serena; Leone, Caterina; Toropov, Andrey A; Toropova, Alla P; Benfenati, Emilio

    2016-02-01

    A predictive model for the viability (%) of cultured human embryonic kidney cells (HEK293) exposed to 20 and 50 nm silica nanoparticles was built using 'optimal descriptors' as mathematical functions of size, concentration and exposure time. The calculation was carried out with CORAL software (http://www.insilico.eu/coral/) on five random splits of combined systems (particle size-particle concentration-cell exposure time) into training, calibration, and validation sets. The R(2) values of the best models were above 0.68. The average statistical quality of the model for the viability (%) of HEK293 exposed to different concentrations of silica nanoparticles measured by MTT assay is satisfactory. PMID:26439516

  7. Chromosome Aberrations in Normal and Ataxia-Telangiectasia Cells Exposed to Heavy Ions

    NASA Technical Reports Server (NTRS)

    Kawata, T.; Ito, H.; Liu, C.; Shigematsu, N.; George, K.; Cucinotta, F. A.

    2007-01-01

    Although cells derived from Ataxia Telangiectasia (AT) patients are known to exhibit abnormal responses to ionizing radiations, its underlying mechanism still remains unclear. Previously, the authors reported that at the same gamma-irradiation dose AT cells show higher frequencies of misrepair and deletions compared to normal human fibroblast cells. In this study, we investigated the effects of heavy ions beams on chromosomal aberrations in normal and AT cells. Normal and AT fibroblast cells arrested at G0/G1 phase were irradiated with 2 Gy of X-rays, 490 MeV/u Silicon (LET 55 keV/m), 500 MeV/u Iron (LET 185 keV/m) and 200 MeV/u Iron (LET 440 keV/m) particles, and then cells were allowed to repair for 24 hours at 37 degrees before subculture. Calyculin-A induced PCC method was employed to collect G2/M chromosomes and whole DNA probes 1 and 3 were used to analyze chromosomal aberrations such as color-junctions, deletions, simple exchanges (incomplete and reciprocal exanges) and complex-type exchanges. The percentages of aberrant cells were higher when normal and AT cells were exposed to heavy ions compared to X-rays, and had a tendency to increase with increasing LET up to 185 keV/m and then decreased at 440 keV/m. When the frequency of color-junctions per cell was compared after X-ray exposure, AT cells had around three times higher frequency of color-junctions (mis-rejoining) than normal cells. However, at 185 keV/m there was no difference in the frequency of color-junctions between two cell lines. It was also found that the frequency of simple exchanges per cell was almost constant in AT cells regardless LET levels, but it was LET dependent for normal cells. Interestingly, the frequency of simple exchanges was higher for AT cells when it was compared at 185 keV/m but AT cells had more complex-type exchanges at the same LET levels. Heavy ions are more efficient in inducing chromosome aberrations in normal and AT cells compared to X-rays, and the aberration types

  8. Insulin withdrawal-induced cell death in adult hippocampal neural stem cells as a model of autophagic cell death.

    PubMed

    Baek, Seung-Hoon; Kim, Eun-Kyoung; Goudreau, John L; Lookingland, Keith J; Kim, Seong Who; Yu, Seong-Woon

    2009-02-01

    The term "autophagic cell death" was coined to describe a form of cell death associated with the massive formation of autophagic vacuoles without signs of apoptosis. However, questions about the actual role of autophagy and its molecular basis in cell death remain to be elucidated. We recently reported that adult hippocampal neural stem (HCN) cells undergo autophagic cell death following insulin withdrawal. Insulin-deprived HCN cells exhibit morphological and biochemical markers of autophagy, including accumulation of Beclin 1 and the type II form of microtubule-associated protein 1 light chain 3 (LC3) without evidence of apoptosis. Suppression of autophagy by knockdown of Atg7 reduces cell death, whereas promotion of autophagy with rapamycin augments cell death in insulin-deficient HCN cells. These data reveal a causative role of autophagy in insulin withdrawal-induced HCN cell death. HCN cells have intact apoptotic capability despite the lack of apoptosis following insulin withdrawal. Our study demonstrates that autophagy is the default cell death mechanism in insulin-deficient HCN cells, and provides a genuine model of autophagic cell death in apoptosis-intact cells. Novel insight into molecular mechanisms of this underappreciated form of programmed cell death should facilitate the development of therapeutic methods to cope with human diseases caused by dysregulated cell death.

  9. Fate of D3 mouse embryonic stem cells exposed to X-rays or carbon ions.

    PubMed

    Luft, S; Pignalosa, D; Nasonova, E; Arrizabalaga, O; Helm, A; Durante, M; Ritter, S

    2014-01-15

    The risk of radiation exposure during embryonic development is still a major problem in radiotoxicology. In this study we investigated the response of the murine embryonic stem cell (mESC) line D3 to two radiation qualities: sparsely ionizing X-rays and densely ionizing carbon ions. We analyzed clonogenic cell survival, proliferation, induction of chromosome aberrations as well as the capability of cells to differentiate to beating cardiomyocytes up to 3 days after exposure. Our results show that, for all endpoints investigated, carbon ions are more effective than X-rays at the same radiation dose. Additionally, in long term studies (≥8 days post-irradiation) chromosomal damage and the pluripotency state were investigated. These studies reveal that pluripotency markers are present in the progeny of cells surviving the exposure to both radiation types. However, only in the progeny of X-ray exposed cells the aberration frequency was comparable to that of the control population, while the progeny of carbon ion irradiated cells harbored significantly more aberrations than the control, generally translocations. We conclude that cells surviving the radiation exposure maintain pluripotency but may carry stable chromosomal rearrangements after densely ionizing radiation.

  10. Antioxidative capacity and enzyme activity in Haematococcus pluvialis cells exposed to superoxide free radicals

    NASA Astrophysics Data System (ADS)

    Liu, Jianguo; Zhang, Xiaoli; Sun, Yanhong; Lin, Wei

    2010-01-01

    The antioxidative capacity of astaxanthin and enzyme activity of reactive oxygen eliminating enzymes such as superoxide dismutase (SOD), peroxidase (POD), catalase (CAT) and ascorbate peroxidase (APX) were studied in three cell types of Haematococcus pluvialis exposed to high concentrations of a superoxide anion radical (O{2/-}). The results show that defensive enzymes and astaxanthin-related mechanisms were both active in H. pluvialis during exposure to reactive oxygen species (ROS) such as O{2/-}. Astaxanthin reacted with ROS much faster than did the protective enzymes, and had the strongest antioxidative capacity to protect against lipid peroxidation. The defensive mechanisms varied significantly between the three cell types and were related to the level of astaxanthin that had accumulated in those cells. Astaxanthin-enriched red cells had the strongest antioxidative capacity, followed by brown cells, and astaxanthin-deficient green cells. Although there was no significant increase in expression of protective enzymes, the malondialdehyde (MDA) content in red cells was sustained at a low level because of the antioxidative effect of astaxanthin, which quenched O{2/-} before the protective enzymes could act. In green cells, astaxanthin is very low or absent; therefore, scavenging of ROS is inevitably reliant on antioxidative enzymes. Accordingly, in green cells, these enzymes play the leading role in scavenging ROS, and the expression of these enzymes is rapidly increased to reduce excessive ROS. However, because ROS were constantly increased in this study, the enhance enzyme activity in the green cells was not able to repair the ROS damage, leading to elevated MDA content. Of the four defensive enzymes measured in astaxanthin-deficient green cells, SOD eliminates O{2/-}, POD eliminates H2O2, which is a by-product of SOD activity, and APX and CAT are then initiated to scavenge excessive ROS.

  11. Radiation-Induced Bystander Effects in A549 Cells Exposed to 6 MV X-rays.

    PubMed

    Yang, Shuning; Xu, Jing; Shao, Weixian; Geng, Chong; Li, Jia; Guo, Feng; Miao, Hui; Shen, Wenbin; Ye, Tao; Liu, Yazhou; Xu, Haiting; Zhang, Xuguang

    2015-07-01

    The aim of the study is to explore the bystander effects in A549 cells that have been exposed to 6MV X-ray. Control group, irradiated group, irradiated conditioned medium (ICM)-received group, and fresh medium group were designed in this study. A549 cells in the logarithmic growth phase were irradiated with 6MV X-ray at 0, 0.5, 1, 1.5, and 2. In ICM-received group, post-irradiation A549 cells were cultured for 3 h and were transferred into non-irradiated A549 cells for further cultivation. Clone forming test was applied to detect the survival fraction of cells. Annexin V-FITC/PI double-staining assay was used to detect the apoptosis of A549 cells 24, 48, 72, and 96 h after 2-Gy 6MV X-ray irradiation, and the curves of apoptosis were drawn. The changes in the cell cycles 4, 48, 72, and 96 h after 2-Gy 6MV X-ray irradiation were detected using PI staining flow cytometry. With the increase of irradiation dose, the survival fraction of A549 cells after the application of 0.5 Gy irradiation was decreasing continuously. In comparison to the control group, the apoptosis rate of the ICM-received group was increased in a time-dependent pattern, with the highest apoptosis rate observed at 72 h (p < 0.05). Cell count in G2/M stages was obviously increased compared with that of the control group (p < 0.05), with the highest count observed at 72 h, after which G2/M stage arrest was diminished. ICM can cause apparent A549 cell damage, indicating that 6MV X-ray irradiation can induce bystander effect on A549 cells, which reaches a peak at 72 h.

  12. Endothelial cells promote neural stem cell proliferation and differentiation associated with VEGF activated Notch and Pten signaling.

    PubMed

    Sun, Jinqiao; Zhou, Wenhao; Ma, Duan; Yang, Yi

    2010-09-01

    To investigate whether and how endothelial cells affect neurogenesis, we established a system to co-culture endothelial cells and brain slices of neonatal rat and observed how subventricular zone cells differentiate in the presence of endothelial cells. In the presence of endothelial cells, neural stem cells increased in number, as did differentiated neurons and glia. The augmentation of neurogenesis was reversed by diminishing vascular endothelial growth factor (VEGF) expression in endothelial cells with RNA interference (RNAi). Microarray analysis indicated that expression levels of 112 genes were significantly altered by co-culture and that expression of 81 of the 112 genes recovered to normal levels following RNAi of VEGF in endothelial cells. Pathway mapping showed an enrichment of genes in the Notch and Pten pathways. These data indicate that endothelial cells promote neural stem cell proliferation and differentiation associated with VEGF, possibly by activating the Notch and Pten pathways.

  13. Transplantation of placenta-derived mesenchymal stem cell-induced neural stem cells to treat spinal cord injury.

    PubMed

    Li, Zhi; Zhao, Wei; Liu, Wei; Zhou, Ye; Jia, Jingqiao; Yang, Lifeng

    2014-12-15

    Because of their strong proliferative capacity and multi-potency, placenta-derived mesenchymal stem cells have gained interest as a cell source in the field of nerve damage repair. In the present study, human placenta-derived mesenchymal stem cells were induced to differentiate into neural stem cells, which were then transplanted into the spinal cord after local spinal cord injury in rats. The motor functional recovery and pathological changes in the injured spinal cord were observed for 3 successive weeks. The results showed that human placenta-derived mesenchymal stem cells can differentiate into neuron-like cells and that induced neural stem cells contribute to the restoration of injured spinal cord without causing transplant rejection. Thus, these cells promote the recovery of motor and sensory functions in a rat model of spinal cord injury. Therefore, human placenta-derived mesenchymal stem cells may be useful as seed cells during the repair of spinal cord injury.

  14. Adult retinal pigment epithelium cells express neural progenitor properties and the neuronal precursor protein doublecortin.

    PubMed

    Engelhardt, Maren; Bogdahn, Ulrich; Aigner, Ludwig

    2005-04-01

    The adult mammalian retina is devoid of any detectable neurogenesis. However, different cell types have been suggested to potentially act as neural progenitors in the adult mammalian retina in vitro, such as ciliary body (CB), Muller glia, and retinal pigment epithelium (RPE) cells. In rodents and humans, strong evidence for neural stem or progenitor properties exists only for CB-derived cells, but not for other retinal cell types. Here, we provide a comparative analysis of adult rat CB- and RPE-derived cells suggesting that the two cell types share certain neural progenitor properties in vitro. CB and RPE cells expressed neural progenitor markers such as Nestin, Flk-1, Hes1, and Musashi. They proliferated under adherent and neurosphere conditions and showed limited self-renewal. Moreover, they differentiated into neuronal and glial cells based on the expression of differentiation markers such as the young neuronal marker beta-III tubulin and the glial and progenitor markers GFAP and NG2. Expression of beta-III tubulin was found in cells with neuronal and non-neuronal morphology. A subpopulation of RPE- and CB-derived progenitor cells expressed the neurogenesis-specific protein doublecortin (DCX). Interestingly, DCX expression defined a beta-III tubulin-positive CB and RPE fraction with a distinct neuronal morphology. In summary, the data suggest that RPE cells share with CB cells the potential to de-differentiate into a cell type with neural progenitor-like identity. In addition, DCX expression might define the neuronal-differentiating RPE- and CB-derived progenitor population. PMID:15804431

  15. 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.

  16. Detection of 8-oxodG in Dreissena polymorpha gill cells exposed to model contaminants.

    PubMed

    Michel, Cécile; Vincent-Hubert, Françoise

    2012-01-24

    Genotoxic end-points are routinely measured in various sentinel organisms in aquatic environments in order to monitor the impact of water pollution on organisms. As a first step towards the evaluation of oxidative DNA damage (8-oxodG) in organisms exposed to chemical water pollution, we have optimized the association between the comet assay and the hOGG1 enzyme for use on zebra mussel (Dreissena polymorpha) gill cells by in vitro exposure to H₂O₂. Firstly, we observed that in vitro exposure of D. polymorpha gill cells to benzo[a]pyrene (B[a]P, 98.4nM) induced an increase of the Olive Tail Moment (OTM) in both the comet-hOGG1 and comet-Fpg assays, indicating that B[a]P causes oxidative DNA damage. By contrast, methylmethane sulfonate (MMS, 33μM) only induced an increase of the Fpg-sensitive sites, indicating that MMS caused alkylating DNA damage and confirming that hOGG1 does not detect alkylating damage. Thus, the hOGG1 enzyme seems to be more specific towards oxidative DNA damage, such as 8-oxodG than Fpg. Secondly, as was observed in vitro, the in vivo exposure of D. polymorpha to B[a]P (24.6 and 98.4nM) increased oxidative DNA damage in gill cells, whereas only Fpg-sensitive sites were detected in mussels exposed to MMS (240μM). These results show that the comet-hOGG1 assay detects oxidative DNA lesions induced in vitro by H₂O₂ and in vivo with BaP. The comet-hOGG1 assay will be used to detect oxidative DNA lesions (8-oxodG) in mussels exposed in situ. PMID:22009068

  17. Apical accumulation of MARCKS in neural plate cells during neurulation in the chick embryo

    PubMed Central

    Zolessi, Flavio R; Arruti, Cristina

    2001-01-01

    Background The neural tube is formed by morphogenetic movements largely dependent on cytoskeletal dynamics. Actin and many of its associated proteins have been proposed as important mediators of neurulation. For instance, mice deficient in MARCKS, an actin cross-linking membrane-associated protein that is regulated by PKC and other kinases, present severe developmental defects, including failure of cranial neural tube closure. Results To determine the distribution of MARCKS, and its possible relationships with actin during neurulation, chick embryos were transversely sectioned and double labeled with an anti-MARCKS polyclonal antibody and phalloidin. In the neural plate, MARCKS was found ubiquitously distributed at the periphery of the cells, being conspicuously accumulated in the apical cell region, in close proximity to the apical actin meshwork. This asymmetric distribution was particularly noticeable during the bending process. After the closure of the neural tube, the apically accumulated MARCKS disappeared, and this cell region became analogous to the other peripheral cell zones in its MARCKS content. Actin did not display analogous variations, remaining highly concentrated at the cell subapical territory. The transient apical accumulation of MARCKS was found throughout the neural tube axis. The analysis of another epithelial bending movement, during the formation of the lens vesicle, revealed an identical phenomenon. Conclusions MARCKS is transiently accumulated at the apical region of neural plate and lens placode cells during processes of bending. This asymmetric subcellular distribution of MARCKS starts before the onset of neural plate bending. These results suggest possible upstream regulatory actions of MARCKS on some functions of the actin subapical meshwork. PMID:11329360

  18. Convergent genesis of an adult neural crest-like dermal stem cell from distinct developmental origins.

    PubMed

    Jinno, Hiroyuki; Morozova, Olena; Jones, Karen L; Biernaskie, Jeffrey A; Paris, Maryline; Hosokawa, Ryoichi; Rudnicki, Michael A; Chai, Yang; Rossi, Fabio; Marra, Marco A; Miller, Freda D

    2010-11-01

    Skin-derived precursors (SKPs) are multipotent dermal stem cells that reside within a hair follicle niche and that share properties with embryonic neural crest precursors. Here, we have asked whether SKPs and their endogenous dermal precursors originate from the neural crest or whether, like the dermis itself, they originate from multiple developmental origins. To do this, we used two different mouse Cre lines that allow us to perform lineage tracing: Wnt1-cre, which targets cells deriving from the neural crest, and Myf5-cre, which targets cells of a somite origin. By crossing these Cre lines to reporter mice, we show that the endogenous follicle-associated dermal precursors in the face derive from the neural crest, and those in the dorsal trunk derive from the somites, as do the SKPs they generate. Despite these different developmental origins, SKPs from these two locations are functionally similar, even with regard to their ability to differentiate into Schwann cells, a cell type only thought to be generated from the neural crest. Analysis of global gene expression using microarrays confirmed that facial and dorsal SKPs exhibit a very high degree of similarity, and that they are also very similar to SKPs derived from ventral dermis, which has a lateral plate origin. However, these developmentally distinct SKPs also retain differential expression of a small number of genes that reflect their developmental origins. Thus, an adult neural crest-like dermal precursor can be generated from a non-neural crest origin, a finding with broad implications for the many neuroendocrine cells in the body.

  19. Existence of Neural Crest-Derived Progenitor Cells in Normal and Fuchs Endothelial Dystrophy Corneal Endothelium.

    PubMed

    Katikireddy, Kishore Reddy; Schmedt, Thore; Price, Marianne O; Price, Francis W; Jurkunas, Ula V

    2016-10-01

    Human corneal endothelial cells are derived from neural crest and because of postmitotic arrest lack competence to repair cell loss from trauma, aging, and degenerative disorders such as Fuchs endothelial corneal dystrophy (FECD). Herein, we identified a rapidly proliferating subpopulation of cells from the corneal endothelium of adult normal and FECD donors that exhibited features of neural crest-derived progenitor (NCDP) cells by showing absence of senescence with passaging, propensity to form spheres, and increased colony forming efficacy compared with the primary cells. The collective expression of stem cell-related genes SOX2, OCT4, LGR5, TP63 (p63), as well as neural crest marker genes PSIP1 (p75(NTR)), PAX3, SOX9, AP2B1 (AP-2β), and NES, generated a phenotypic footprint of endothelial NCDPs. NCDPs displayed multipotency by differentiating into microtubule-associated protein 2, β-III tubulin, and glial fibrillary acidic protein positive neurons and into p75(NTR)-positive human corneal endothelial cells that exhibited transendothelial resistance of functional endothelium. In conclusion, we found that mitotically incompetent ocular tissue cells contain adult NCDPs that exhibit a profile of transcription factors regulating multipotency and neural crest progenitor characteristics. Identification of normal NCDPs in FECD-affected endothelium holds promise for potential autologous cell therapies. PMID:27639969

  20. RA induces the neural-like cells generated from epigenetic modified NIH/3T3 cells.

    PubMed

    Zhang, Xi-Mei; Li, Qiu-Ming; Su, Dong-Ju; Wang, Ning; Shan, Zhi-Yan; Jin, Lian-Hong; Lei, Lei

    2010-03-01

    Recently, differentiated somatic cells had been reprogrammed to pluripotential state in vitro, and various tissue cells had been elicited from those cells. Epigenetic modifications allow differentiated cells to perpetuate the molecular memory needed for the cells to retain their identity. DNA methylation and histone deacetylation are important patterns involved in epigenetic modification, which take critical roles in regulating DNA expression. In this study, we dedifferentiated NIH/3T3 fibroblasts by 5-aza-2-deoxycytidine (5-aza-dC) and Trichstatin A (TSA) combination, and detected gene expression pattern, DNA methylation level, and differentiation potential of reprogrammed cells. As the results, embryonic marker Sox2, klf4, c-Myc and Oct4 were expressed in reprogrammed NIH/3T3 fibroblasts. Total DNA methylation level was significant decreased after the treatment. Moreover, exposure of the reprogrammed cells to all trans-retinoic acid (RA) medium elicited the generation of neuronal class IIIbeta-tubulin-positive, neuron-specific enolase (NSE)-positive, nestin-positive, and neurofilament light chain (NF-L)-positive neural-like cells. PMID:19263240

  1. Mesenchymal stem cells instruct oligodendrogenic fate decision on adult neural stem cells.

    PubMed

    Rivera, Francisco J; Couillard-Despres, Sebastien; Pedre, Xiomara; Ploetz, Sonja; Caioni, Massimiliano; Lois, Carlos; Bogdahn, Ulrich; Aigner, Ludwig

    2006-10-01

    Adult stem cells reside in different tissues and organs of the adult organism. Among these cells are MSCs that are located in the adult bone marrow and NSCs that exist in the adult central nervous system (CNS). In transplantation experiments, MSCs demonstrated neuroprotective and neuroregenerative effects that were associated with functional improvements. The underlying mechanisms are largely unidentified. Here, we reveal that the interactions between adult MSCs and NSCs, mediated by soluble factors, induce oligodendrogenic fate decision in NSCs at the expense of astrogenesis. This was demonstrated (a) by an increase in the percentage of cells expressing the oligodendrocyte markers GalC and myelin basic protein, (b) by a reduction in the percentage of glial fibrillary acidic protein (GFAP)-expressing cells, and (c) by the expression pattern of cell fate determinants specific for oligodendrogenic differentiation. Thus, it involved enhanced expression of the oligodendrogenic transcription factors Olig1, Olig2, and Nkx2.2 and diminished expression of Id2, an inhibitor of oligodendrogenic differentiation. Results of (a) 5-bromo-2'-deoxyuridine pulse-labeling of cells, (b) cell fate analysis, and (c) cell death/survival analysis suggested an inductive mechanism and excluded a selection process. A candidate factor screen excluded a number of growth factors, cytokines, and neurotrophins that have previously been shown to influence neurogenesis and neural differentiation from the oligodendrogenic activity derived from the MSCs. This work might have major implications for the development of future transplantation strategies for the treatment of degenerative diseases in the CNS. PMID:16763198

  2. Diesel Exhaust Particle-Exposed Human Bronchial Epithelial Cells Induce Dendritic Cell Maturation and Polarization via Thymic Stromal Lymphopoietin

    PubMed Central

    Bleck, Bertram; Tse, Doris B.; Curotto de Lafaille, Maria A.; Zhang, Feijie

    2009-01-01

    Human exposure to air pollutants, including ambient particulate matter, has been proposed as a mechanism for the rise in allergic disorders. Diesel exhaust particles, a major component of ambient particulate matter, induce sensitization to neoallergens, but the mechanisms by which sensitization occur remain unclear. We show that diesel exhaust particles upregulate thymic stromal lymphopoietin in human bronchial epithelial cells in an oxidant-dependent manner. Thymic stromal lymphopoietin induced by diesel exhaust particles was associated with maturation of myeloid dendritic cells, which was blocked by anti-thymic stromal lymphopoietin antibodies or silencing epithelial cell-derived thymic stromal lymphopoietin. Dendritic cells exposed to diesel exhaust particle-treated human bronchial epithelial cells induced Th2 polarization in a thymic stromal lymphopoietin-dependent manner. These findings provide new insight into the mechanisms by which diesel exhaust particles modify human lung mucosal immunity. PMID:18049884

  3. Reversal of Cellular Phenotypes in Neural Cells Derived from Huntington’s Disease Monkey-Induced Pluripotent Stem Cells

    PubMed Central

    Carter, Richard L.; Chen, Yiju; Kunkanjanawan, Tanut; Xu, Yan; Moran, Sean P.; Putkhao, Kittiphong; Yang, Jinjing; Huang, Anderson H.C.; Parnpai, Rangsun; Chan, Anthony W.S.

    2014-01-01

    Summary Huntington’s disease (HD) is a dominant neurodegenerative disorder caused by the expansion of glutamine residues in the N-terminal region of the huntingtin (HTT) protein. The disease results in progressive neuronal loss, leading to motor, cognitive, and psychiatric impairment. Here, we report the establishment of neural progenitor cell (NPC) lines derived from induced pluripotent stem cells (iPSCs) of transgenic HD monkeys. Upon differentiation to neurons, HD neural cells develop cellular features of HD, including the formation of nuclear inclusions and oligomeric mutant HTT (mHTT) aggregates, as well as increased apoptosis. These phenotypes are rescued by genetic suppression of HTT and pharmacological treatment, demonstrating the ability of our HD cell model to respond to therapeutic treatment. The development and reversal of HD-associated phenotypes in neural cells from HD monkeys provides a unique nonhuman primate (NHP) model for exploring HD pathogenesis and evaluating therapeutics that could be assessed further in HD monkeys. PMID:25358787

  4. Comparison of different protocols for neural differentiation of human induced pluripotent stem cells.

    PubMed

    Salimi, Ali; Nadri, Samad; Ghollasi, Marzieh; Khajeh, Khosro; Soleimani, Masoud

    2014-03-01

    Although embryonic stem cells (ESCs) have enormous potentials due to their pluripotency, their therapeutic use is limited by ethical, biological and safety issues. Compared to ESCs, induced pluripotent stem cells (iPSCs) can be obtained from mouse or human fibroblasts by reprogramming. Numerous studies have established many protocols for differentiation of human iPSCs (hiPSCs) into neural lineages. However, the low differentiation efficiency of such protocols motivates researchers to design new protocols for high yield differentiation. Herein, we compared neural differentiation potential of three induction media for conversion of hiPSCs into neural lineages. In this study, hiPSCs-derived embryoid bodies were plated on laminin coated dishes and were treated with three induction media including (1) bFGF, EGF (2) RA and (3) forskolin, IBMX. Immunofluorescence staining and quantitative real-time PCR (qPCR) analysis were used to detect the expression of neural genes and proteins. qPCR analysis showed that the expression of neural genes in differentiated hiPSCs in forskolin, IBMX supplemented media was significantly higher than undifferentiated cells and those in induction media containing bFGF, EGF or RA. In conclusion, our results indicated a successful establishment protocol with high efficiency for differentiation of hiPSCs into neural lineages.

  5. Self-organization of neural tissue architectures from pluripotent stem cells.

    PubMed

    Karus, Michael; Blaess, Sandra; Brüstle, Oliver

    2014-08-15

    Despite being a subject of intensive research, the mechanisms underlying the formation of neural tissue architectures during development of the central nervous system remain largely enigmatic. So far, studies into neural pattern formation have been restricted mainly to animal experiments. With the advent of pluripotent stem cells it has become possible to explore early steps of nervous system development in vitro. These studies have unraveled a remarkable propensity of primitive neural cells to self-organize into primitive patterns such as neural tube-like rosettes in vitro. Data from more advanced 3D culture systems indicate that this intrinsic propensity for self-organization can even extend to the formation of complex architectures such as a multilayered cortical neuroepithelium or an entire optic cup. These novel experimental paradigms not only demonstrate the enormous self-organization capacity of neural stem cells, they also provide exciting prospects for studying the earliest steps of human neural tissue development and the pathogenesis of brain malformations in reductionist in vitro paradigms. PMID:24737617

  6. Proteomic signature of Arabidopsis cell cultures exposed to magnetically induced hyper- and microgravity environments.

    PubMed

    Herranz, Raul; Manzano, Ana I; van Loon, Jack J W A; Christianen, Peter C M; Medina, F Javier

    2013-03-01

    Earth-based microgravity simulation techniques are required due to space research constraints. Using diamagnetic levitation, we exposed Arabidopsis thaliana in vitro callus cultures to environments with different levels of effective gravity and magnetic field strengths (B) simultaneously. The environments included simulated 0 g* at B=10.1 T, an internal 1 g* control (B=16.5 T), and hypergravity (2 g* at B=10.1 T). Furthermore, samples were also exposed to altered gravity environments that were created with mechanical devices, such as the Random Positioning Machine (simulated μg) and the Large Diameter Centrifuge (2 g). We have determined the proteomic signature of cell cultures exposed to these altered-gravity environments by means of the difference gel electrophoresis (DiGE) technique, and we have compared the results with microarray-based transcriptomes from the same samples. The magnetic field itself produced a low number of proteomic alterations, but the combination of gravitational alteration and magnetic field exposure produced synergistic effects on the proteome of plants (the number of significant changes is 3-7 times greater). Tandem mass spectrometry identification of 19 overlapping spots in the different conditions corroborates a major role of abiotic stress and secondary metabolism proteins in the molecular adaptation of plants to unusual environments, including microgravity.

  7. Proteomic signature of Arabidopsis cell cultures exposed to magnetically induced hyper- and microgravity environments.

    PubMed

    Herranz, Raul; Manzano, Ana I; van Loon, Jack J W A; Christianen, Peter C M; Medina, F Javier

    2013-03-01

    Earth-based microgravity simulation techniques are required due to space research constraints. Using diamagnetic levitation, we exposed Arabidopsis thaliana in vitro callus cultures to environments with different levels of effective gravity and magnetic field strengths (B) simultaneously. The environments included simulated 0 g* at B=10.1 T, an internal 1 g* control (B=16.5 T), and hypergravity (2 g* at B=10.1 T). Furthermore, samples were also exposed to altered gravity environments that were created with mechanical devices, such as the Random Positioning Machine (simulated μg) and the Large Diameter Centrifuge (2 g). We have determined the proteomic signature of cell cultures exposed to these altered-gravity environments by means of the difference gel electrophoresis (DiGE) technique, and we have compared the results with microarray-based transcriptomes from the same samples. The magnetic field itself produced a low number of proteomic alterations, but the combination of gravitational alteration and magnetic field exposure produced synergistic effects on the proteome of plants (the number of significant changes is 3-7 times greater). Tandem mass spectrometry identification of 19 overlapping spots in the different conditions corroborates a major role of abiotic stress and secondary metabolism proteins in the molecular adaptation of plants to unusual environments, including microgravity. PMID:23510084

  8. Localizing Protein in 3D Neural Stem Cell Culture: a Hybrid Visualization Methodology

    PubMed Central

    Fai, Stephen; Bennett, Steffany A.L.

    2010-01-01

    The importance of 3-dimensional (3D) topography in influencing neural stem and progenitor cell (NPC) phenotype is widely acknowledged yet challenging to study. When dissociated from embryonic or post-natal brain, single NPCs will proliferate in suspension to form neurospheres. Daughter cells within these cultures spontaneously adopt distinct developmental lineages (neurons, oligodendrocytes, and astrocytes) over the course of expansion despite being exposed to the same extracellular milieu. This progression recapitulates many of the stages observed over the course of neurogenesis and gliogenesis in post-natal brain and is often used to study basic NPC biology within a controlled environment. Assessing the full impact of 3D topography and cellular positioning within these cultures on NPC fate is, however, difficult. To localize target proteins and identify NPC lineages by immunocytochemistry, free-floating neurospheres must be plated on a substrate or serially sectioned. This processing is required to ensure equivalent cell permeabilization and antibody access throughout the sphere. As a result, 2D epifluorescent images of cryosections or confocal reconstructions of 3D Z-stacks can only provide spatial information about cell position within discrete physical or digital 3D slices and do not visualize cellular position in the intact sphere. Here, to reiterate the topography of the neurosphere culture and permit spatial analysis of protein expression throughout the entire culture, we present a protocol for isolation, expansion, and serial sectioning of post-natal hippocampal neurospheres suitable for epifluorescent or confocal immunodetection of target proteins. Connexin29 (Cx29) is analyzed as an example. Next, using a hybrid of graphic editing and 3D modelling softwares rigorously applied to maintain biological detail, we describe how to re-assemble the 3D structural positioning of these images and digitally map labelled cells within the complete neurosphere. This

  9. 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.

  10. Axonal growth and connectivity from neural stem cell grafts in models of spinal cord injury.

    PubMed

    Lu, Paul; Kadoya, Ken; Tuszynski, Mark H

    2014-08-01

    Spinal cord injury (SCI) damages both gray matter and white matter, but white matter damage is responsible for the vast majority of the subsequent functional loss. Neural stem cells (NSCs) have been investigated as a means of improving outcomes after SCI, either through neuroprotective properties that limit secondary damage or by direct cell replacement. This review will focus on cell replacement strategies, and the ability of multipotent NSCs to form new functional synaptic relays across sites of even severe SCI. The ability of these early stage neurons to extend axons from the lesion site in large numbers and over long distances constitutes an important mechanism underlying their potential to promote neural repair.

  11. Transplanted neural stem/precursor cells instruct phagocytes and reduce secondary tissue damage in the injured spinal cord.

    PubMed

    Cusimano, Melania; Biziato, Daniela; Brambilla, Elena; Donegà, Matteo; Alfaro-Cervello, Clara; Snider, Silvia; Salani, Giuliana; Pucci, Ferdinando; Comi, Giancarlo; Garcia-Verdugo, Jose Manuel; De Palma, Michele; Martino, Gianvito; Pluchino, Stefano

    2012-02-01

    Transplanted neural stem/precursor cells possess peculiar therapeutic plasticity and can simultaneously instruct several therapeutic mechanisms in addition to cell replacement. Here, we interrogated the therapeutic plasticity of neural stem/precursor cells after their focal implantation in the severely contused spinal cord. We injected syngeneic neural stem/precursor cells at the proximal and distal ends of the contused mouse spinal cord and analysed locomotor functions and relevant secondary pathological events in the mice, cell fate of transplanted neural stem/precursor cells, and gene expression and inflammatory cell infiltration at the injured site. We used two different doses of neural stem/precursor cells and two treatment schedules, either subacute (7 days) or early chronic (21 days) neural stem/precursor cell transplantation after the induction of experimental thoracic severe spinal cord injury. Only the subacute transplant of neural stem/precursor cells enhanced the recovery of locomotor functions of mice with spinal cord injury. Transplanted neural stem/precursor cells survived undifferentiated at the level of the peri-lesion environment and established contacts with endogenous phagocytes via cellular-junctional coupling. This was associated with significant modulation of the expression levels of important inflammatory cell transcripts in vivo. Transplanted neural stem/precursor cells skewed the inflammatory cell infiltrate at the injured site by reducing the proportion of 'classically-activated' (M1-like) macrophages, while promoting the healing of the injured cord. We here identify a precise window of opportunity for the treatment of complex spinal cord injuries with therapeutically plastic somatic stem cells, and suggest that neural stem/precursor cells have the ability to re-programme the local inflammatory cell microenvironment from a 'hostile' to an 'instructive' role, thus facilitating the healing or regeneration past the lesion.

  12. 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

  13. Novel insights into mammalian embryonic neural stem cell division: focus on microtubules.

    PubMed

    Mora-Bermúdez, Felipe; Huttner, Wieland B

    2015-12-01

    During stem cell divisions, mitotic microtubules do more than just segregate the chromosomes. They also determine whether a cell divides virtually symmetrically or asymmetrically by establishing spindle orientation and the plane of cell division. This can be decisive for the fate of the stem cell progeny. Spindle defects have been linked to neurodevelopmental disorders, yet the role of spindle orientation for mammalian neurogenesis has remained controversial. Here we explore recent advances in understanding how the microtubule cytoskeleton influences mammalian neural stem cell division. Our focus is primarily on the role of spindle microtubules in the development of the cerebral cortex. We also highlight unique characteristics in the architecture and dynamics of cortical stem cells that are tightly linked to their mode of division. These features contribute to setting these cells apart as mitotic "rule breakers," control how asymmetric a division is, and, we argue, are sufficient to determine the fate of the neural stem cell progeny in mammals.

  14. Increasing magnetite contents of polymeric magnetic particles dramatically improves labeling of neural stem cell transplant populations.

    PubMed

    Adams, Christopher F; Rai, Ahmad; Sneddon, Gregor; Yiu, Humphrey H P; Polyak, Boris; Chari, Divya M

    2015-01-01

    Safe and efficient delivery of therapeutic cells to sites of injury/disease in the central nervous system is a key goal for the translation of clinical cell transplantation therapies. Recently, 'magnetic cell localization strategies' have emerged as a promising and safe approach for targeted delivery of magnetic particle (MP) labeled stem cells to pathology sites. For neuroregenerative applications, this approach is limited by the lack of available neurocompatible MPs, and low cell labeling achieved in neural stem/precursor populations. We demonstrate that high magnetite content, self-sedimenting polymeric MPs [unfunctionalized poly(lactic acid) coated, without a transfecting component] achieve efficient labeling (≥90%) of primary neural stem cells (NSCs)-a 'hard-to-label' transplant population of major clinical relevance. Our protocols showed high safety with respect to key stem cell regenerative parameters. Critically, labeled cells were effectively localized in an in vitro flow system by magnetic force highlighting the translational potential of the methods used.

  15. ATM Deficiency Results in Accumulation of DNA-Topoisomerase I Covalent Intermediates in Neural Cells

    PubMed Central

    Sharma, Abhishek; El-Khamisy, Sherif F.

    2013-01-01

    Accumulation of peptide-linked DNA breaks contributes to neurodegeration in humans. This is typified by defects in tyrosyl DNA phosphodiesterase 1 (TDP1) and human hereditary ataxia. TDP1 primarily operates at single-strand breaks (SSBs) created by oxidative stress or by collision of transcription machinery with topoisomerase I intermediates (Top1-CCs). Cellular and cell-free studies have shown that Top1 at stalled Top1-CCs is first degraded to a small peptide resulting in Top1-SSBs, which are the primary substrates for TDP1. Here we established an assay to directly compare Top1-SSBs and Top1-CCs. We subsequently employed this assay to reveal an increased steady state level of Top1-CCs in neural cells lacking Atm; the protein mutated in ataxia telangiectasia. Our data suggest that the accumulation of endogenous Top1-CCs in Atm-/- neural cells is primarily due to elevated levels of reactive oxygen species. Biochemical purification of Top1-CCs from neural cell extract and the use of Top1 poisons further confirmed a role for Atm during the formation/resolution of Top1-CCs. Finally, we report that global transcription is reduced in Atm-/- neural cells and fails to recover to normal levels following Top1-mediated DNA damage. Together, these data identify a distinct role for ATM during the formation/resolution of neural Top1-CCs and suggest that their accumulation contributes to the neuropathology of ataxia telangiectasia. PMID:23626666

  16. Cell surface glycan engineering of neural stem cells augments neurotropism and improves recovery in a murine model of multiple sclerosis.

    PubMed

    Merzaban, Jasmeen S; Imitola, Jaime; Starossom, Sarah C; Zhu, Bing; Wang, Yue; Lee, Jack; Ali, Amal J; Olah, Marta; Abuelela, Ayman F; Khoury, Samia J; Sackstein, Robert

    2015-12-01

    Neural stem cell (NSC)-based therapies offer potential for neural repair in central nervous system (CNS) inflammatory and degenerative disorders. Typically, these conditions present with multifocal CNS lesions making it impractical to inject NSCs locally, thus mandating optimization of vascular delivery of the cells to involved sites. Here, we analyzed NSCs for expression of molecular effectors of cell migration and found that these cells are natively devoid of E-selectin ligands. Using glycosyltransferase-programmed stereosubstitution (GPS), we glycan engineered the cell surface of NSCs ("GPS-NSCs") with resultant enforced expression of the potent E-selectin ligand HCELL (hematopoietic cell E-/L-selectin ligand) and of an E-selectin-binding glycoform of neural cell adhesion molecule ("NCAM-E"). Following intravenous (i.v.) injection, short-term homing studies demonstrated that, compared with buffer-treated (control) NSCs, GPS-NSCs showed greater neurotropism. Administration of GPS-NSC significantly attenuated the clinical course of experimental autoimmune encephalomyelitis (EAE), with markedly decreased inflammation and improved oligodendroglial and axonal integrity, but without evidence of long-term stem cell engraftment. Notably, this effect of NSC is not a universal property of adult stem cells, as administration of GPS-engineered mouse hematopoietic stem/progenitor cells did not improve EAE clinical course. These findings highlight the utility of cell surface glycan engineering to boost stem cell delivery in neuroinflammatory conditions and indicate that, despite the use of a neural tissue-specific progenitor cell population, neural repair in EAE results from endogenous repair and not from direct, NSC-derived cell replacement.

  17. 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

  18. Evaluation of cell types for assessment of cytogenetic damage in arsenic exposed population

    PubMed Central

    Ghosh, Pritha; Basu, Arindam; Singh, Keshav K; Giri, Ashok K

    2008-01-01

    Background Cytogenetic biomarkers are essential for assessing environmental exposure, and reflect adverse human health effects such as cellular damage. Arsenic is a potential clastogen and aneugen. In general, the majority of the studies on clastogenic effects of arsenic are based on frequency of micronuclei (MN) study in peripheral lymphocytes, urothelial and oral epithelial cells. To find out the most suitable cell type, here, we compared cytogenetic damage through MN assay in (a) various populations exposed to arsenic through drinking water retrieved from literature review, as also (b) arsenic-induced Bowen's patients from our own survey. Results For literature review, we have searched the Pubmed database for English language journal articles using the following keywords: "arsenic", "micronuclei", "drinking water", and "human" in various combinations. We have selected 13 studies consistent with our inclusion criteria that measured micronuclei in either one or more of the above-mentioned three cell types, in human samples. Compared to urothelial and buccal mucosa cells, the median effect sizes measured by the difference between people with exposed and unexposed, lymphocyte based MN counts were found to be stronger. This general pattern pooled from 10 studies was consistent with our own set of three earlier studies. MN counts were also found to be stronger for lymphocytes even in arsenic-induced Bowen's patients (cases) compared to control individuals having arsenic-induced non-cancerous skin lesions. Conclusion Overall, it can be concluded that MN in lymphocytes may be superior to other epithelial cells for studying arsenic-induced cytogenetic damage. PMID:18505595

  19. 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.

  20. Neural differentiation of novel multipotent progenitor cells from cryopreserved human umbilical cord blood

    SciTech Connect

    Lee, Myoung Woo; Moon, Young Joon; Yang, Mal Sook; Kim, Sun Kyung; Jang, In Keun; Eom, Young-woo; Park, Joon Seong; Kim, Hugh C.; Song, Kye Yong; Park, Soon Cheol; Lim, Hwan Sub; Kim, Young Jin . E-mail: jin@lifecord.co.kr

    2007-06-29

    Umbilical cord blood (UCB) is a rich source of hematopoietic stem cells, with practical and ethical advantages. To date, the presence of other stem cells in UCB remains to be established. We investigated whether other stem cells are present in cryopreserved UCB. Seeded mononuclear cells formed adherent colonized cells in optimized culture conditions. Over a 4- to 6-week culture period, colonized cells gradually developed into adherent mono-layer cells, which exhibited homogeneous fibroblast-like morphology and immunophenotypes, and were highly proliferative. Isolated cells were designated 'multipotent progenitor cells (MPCs)'. Under appropriate conditions for 2 weeks, MPCs differentiated into neural tissue-specific cell types, including neuron, astrocyte, and oligodendrocyte. Differentiated cells presented their respective markers, specifically, NF-L and NSE for neurons, GFAP for astrocytes, and myelin/oligodendrocyte for oligodendrocytes. In this study, we successfully isolated MPCs from cryopreserved UCB, which differentiated into the neural tissue-specific cell types. These findings suggest that cryopreserved human UCB is a useful alternative source of neural progenitor cells, such as MPCs, for experimental and therapeutic applications.

  1. Neural differentiation of transplanted neural stem cells in a rat model of striatal lacunar infarction: light and electron microscopic observations

    PubMed Central

    Muñetón-Gómez, Vilma C.; Doncel-Pérez, Ernesto; Fernandez, Ana P.; Serrano, Julia; Pozo-Rodrigálvarez, Andrea; Vellosillo-Huerta, Lara; Taylor, Julian S.; Cardona-Gómez, Gloria P.; Nieto-Sampedro, Manuel; Martínez-Murillo, Ricardo

    2012-01-01

    The increased risk and prevalence of lacunar stroke and Parkinson's disease (PD) makes the search for better experimental models an important requirement for translational research. In this study we assess ischemic damage of the nigrostriatal pathway in a model of lacunar stroke evoked by damaging the perforating arteries in the territory of the substantia nigra (SN) of the rat after stereotaxic administration of endothelin-1 (ET-1), a potent vasoconstrictor peptide. We hypothesized that transplantation of neural stem cells (NSCs) with the capacity of differentiating into diverse cell types such as neurons and glia, but with limited proliferation potential, would constitute an alternative and/or adjuvant therapy for lacunar stroke. These cells showed neuritogenic activity in vitro and a high potential for neural differentiation. Light and electron microscopy immunocytochemistry was used to characterize GFP-positive neurons derived from the transplants. 48 h after ET-1 injection, we characterized an area of selective degeneration of dopaminergic neurons within the nigrostriatal pathway characterized with tissue necrosis and glial scar formation, with subsequent behavioral signs of Parkinsonism. Light microscopy showed that grafted cells within the striatal infarction zone differentiated with a high yield into mature glial cells (GFAP-positive) and neuron types present in the normal striatum. Electron microscopy revealed that NSCs-derived neurons integrated into the host circuitry establishing synaptic contacts, mostly of the asymmetric type. Astrocytes were closely associated with normal small-sized blood vessels in the area of infarct, suggesting a possible role in the regulation of the blood brain barrier and angiogenesis. Our results encourage the use of NSCs as a cell-replacement therapy for the treatment of human vascular Parkinsonism. PMID:22876219

  2. Cell wall chitosaccharides are essential components and exposed patterns of the phytopathogenic oomycete Aphanomyces euteiches.

    PubMed

    Badreddine, Ilham; Lafitte, Claude; Heux, Laurent; Skandalis, Nicholas; Spanou, Zacharoula; Martinez, Yves; Esquerré-Tugayé, Marie-Thérèse; Bulone, Vincent; Dumas, Bernard; Bottin, Arnaud

    2008-11-01

    Chitin is an essential component of fungal cell walls, where it forms a crystalline scaffold, and chitooligosaccharides derived from it are signaling molecules recognized by the hosts of pathogenic fungi. Oomycetes are cellulosic fungus-like microorganisms which most often lack chitin in their cell walls. Here we present the first study of the cell wall of the oomycete Aphanomyces euteiches, a major parasite of legume plants. Biochemical analyses demonstrated the presence of ca. 10% N-acetyl-D-glucosamine (GlcNAc) in the cell wall. Further characterization of the GlcNAc-containing material revealed that it corresponds to noncrystalline chitosaccharides associated with glucans, rather than to chitin per se. Two putative chitin synthase (CHS) genes were identified by data mining of an A. euteiches expressed sequence tag collection and Southern blot analysis, and full-length cDNA sequences of both genes were obtained. Phylogeny analysis indicated that oomycete CHS diversification occurred before the divergence of the major oomycete lineages. Remarkably, lectin labeling showed that the Aphanomyces euteiches chitosaccharides are exposed at the cell wall surface, and study of the effect of the CHS inhibitor nikkomycin Z demonstrated that they are involved in cell wall function. These data open new perspectives for the development of antioomycete drugs and further studies of the molecular mechanisms involved in the recognition of pathogenic oomycetes by the host plants. PMID:18806214

  3. HALOACETIC ACIDS AND KINASE INHIBITORS PERTURB MOUSE NEURAL CREST CELLS IN VITRO

    EPA Science Inventory

    HUNTER, E.S.1, J. SMITH2, J. ANDREWS1. 1 Reproductive Toxicology Division, NHEERL, US EPA, Research Triangle Park and 2 Department of Cell and Developmental Biology, UNC-CH, Chapel Hill, North Carolina. Haloacetic acids and kinase inhibitors perturb mouse neural crest cells in vi...

  4. Neural stem cell transplantation as a therapeutic approach for treating lysosomal storage diseases.

    PubMed

    Shihabuddin, Lamya S; Cheng, Seng H

    2011-10-01

    Treating the central nervous system manifestations of subjects with neuropathic lysosomal storage diseases remains a major technical challenge. This is because of the low efficiency by which lysosomal enzymes in systemic circulation are able to traverse the blood brain barrier into the central nervous system. Intracranial transplantation of neural stems cells genetically modified to overexpress the respective deficient enzymes represents a potential approach to addressing this group of diseases. The unique properties of neural stem cells and progenitor cells, such as their ability to migrate to distal sites, differentiate into various cell types and integrate within the host brain without disrupting normal function, making them particularly attractive therapeutic agents. In addition, neural stem cells are amenable to ex vivo propagation and modification by gene transfer vectors. In this regard, transplanted cells can serve not only as a source of lysosomal enzymes but also as a means to potentially repair the injured brain by replenishing the organ with healthy cells and effecting the release of neuroprotective factors. This review discusses some of the well-characterized neural stem cell types and their possible use in treating neuropathic lysosomal storage diseases such as the Niemann Pick A disease.

  5. Extremely low-frequency electromagnetic fields enhance the proliferation and differentiation of neural progenitor cells cultured from ischemic brains.

    PubMed

    Cheng, Yannan; Dai, Yiqin; Zhu, Ximin; Xu, Haochen; Cai, Ping; Xia, Ruohong; Mao, Lizhen; Zhao, Bing-Qiao; Fan, Wenying

    2015-10-21

    In the mammalian brain, neurogenesis persists throughout the embryonic period and adulthood in the subventricular zone of the lateral ventricle and the granular zone (dentate gyrus) of the hippocampus. Newborn neural progenitor cells (NPCs) in the two regions play a critical role in structural and functional plasticity and neural regeneration after brain injury. Previous studies have reported that extremely low-frequency electromagnetic fields (ELF-EMF) could promote osteogenesis, angiogenesis, and cardiac stem cells' differentiation, which indicates that ELF-EMF might be an effective tool for regenerative therapy. The present studies were carried out to examine the effects of ELF-EMF on hippocampal NPCs cultured from embryonic and adult ischemic brains. We found that exposure to ELF-EMF (50 Hz, 0.4 mT) significantly enhanced the proliferation capability both in embryonic NPCs and in ischemic NPCs. Neuronal differentiation was also enhanced after 7 days of cumulative ELF-EMF exposure, whereas glial differentiation was not influenced markedly. The expression of phosphorylated Akt increased during the proliferation process when ischemic NPCs were exposed to ELF-EMF. However, blockage of the Akt pathway abolished the ELF-EMF-induced proliferation of ischemic NPCs. These data show that ELF-EMF promotes neurogenesis of ischemic NPCs and suggest that this effect may occur through the Akt pathway.Video abstract, Supplemental Digital Content 1, http://links.lww.com/WNR/A347.

  6. Dimethyloxalylglycine may be enhance the capacity of neural-like cells in treatment of Alzheimer disease.

    PubMed

    Ghasemi Moravej, Fahimeh; Vahabian, Mehrangiz; Soleimani Asl, Sara

    2016-06-01

    Although using differentiated stem cells is the best proposed option for the treatment of Alzheimer disease (AD), an efficient differentiation and cell therapy require enhanced cell survival and homing and decreased apoptosis. It seems that hypoxia preconditioning via Dimethyloxalylglycine (DMOG) may increase the capacity of MSC to induce neural like stem cells (NSCs). Furthermore, it can likely improve the viability of NSCs when transplanted into the brain of AD rats.

  7. Disrupted Endothelial Cell Layer and Exposed Extracellular Matrix Proteins Promote Capture of Late Outgrowth Endothelial Progenitor Cells.

    PubMed

    Zhao, Jing; Mitrofan, Claudia-Gabriela; Appleby, Sarah L; Morrell, Nicholas W; Lever, Andrew M L

    2016-01-01

    Late outgrowth endothelial progenitor cells (LO-EPC) possess a high proliferative potential, differentiate into vascular endothelial cells (EC), and form networks, suggesting they play a role in vascular repair. However, due to their scarcity in the circulation there is a requirement for ex vivo expansion before they could provide a practical cell therapy and it is currently unclear if they would home and engraft to an injury site. Using an in vitro flow system we studied LO-EPC under simulated injury conditions including EC activation, ischaemia, disrupted EC integrity, and exposed basement membrane. Perfused LO-EPC adhered to discontinuous EC paracellularly at junctional regions between adjacent cells under shear stress 0.7 dyn/cm(2). The interaction was not adhesion molecule-dependent and not enhanced by EC activation. LO-EPC expressed high levels of the VE-Cadherin which may explain these findings. Ischaemia reperfusion injury decreased the interaction with LO-EPC due to cell retraction. LO-EPC interacted with exposed extracellular matrix (ECM) proteins, fibronectin and vitronectin. The interaction was mediated by integrins α5β3, αvβ1, and αvβ3. This study has demonstrated that an injured local environment presents sufficient adhesive signals to capture flow perfused LO-EPC in vitro and that LO-EPC have properties consistent with their potential role in vascular repair.

  8. Disrupted Endothelial Cell Layer and Exposed Extracellular Matrix Proteins Promote Capture of Late Outgrowth Endothelial Progenitor Cells.

    PubMed

    Zhao, Jing; Mitrofan, Claudia-Gabriela; Appleby, Sarah L; Morrell, Nicholas W; Lever, Andrew M L

    2016-01-01

    Late outgrowth endothelial progenitor cells (LO-EPC) possess a high proliferative potential, differentiate into vascular endothelial cells (EC), and form networks, suggesting they play a role in vascular repair. However, due to their scarcity in the circulation there is a requirement for ex vivo expansion before they could provide a practical