In vitro growth, differentiation and biological characteristics of neural stem cells

OBJECTIVE： To summarize the biological characteristics of neural stem cells, and the separation, purification. differentiation and source of neural stem cells. DATA SOURCES ： An online search of Pubmed database was undertaken to identify English articles about the growth of neural stem cells in vitro published from January 2000 to October 2006 by using the keywords of ＂neural stem cells, bone marrow mesenchymal stem cells （BMSCs）, umbilical cord blood stem cells, embryonic stem cells （ESC）, separation methods, neural growth factor＂. And relevant articles published in IEEE/IEE Electronic Library （IEL） database, Springer Link database and Kluwer Online Journals were also searched, Chinese relevant articles published between January 2000 to October 2006 were searched with the same keywords in Chinese in Chinese journal full-text database. STUDY SELECTION ： The articles were primarily screened, and then the full-texts were searched. Inclusive criteria： （1） Articles relevant to the biological characteristics and classification of neural stem cells; （2） Articles about the source, separation and differentiation of the ESCs, BMSCs and umbilical cord blood stem cells. The repetitive studies and reviews were excluded. DATA EXTRACTION ： Thirty articles were selected from 203 relevant articles according to the inclusive criteria Articles were excluded because of repetition and reviews. DATA SYNTHESES ： Neural stem cells have the ability of self-renewing and high differentiation, and they are obtained from ESCs, nerve tissue, nerve system, BMSCs and umbilical cord blood stem cells. ESCs can be separated by means of mechanical dissociation is better than that of the trypsin digestion, BMSCs by density gradient centrifuge separation, hemolysis, whole-blood culture, etc., and umbilical cord blood stem ceils by Ficoil density gradient centrifugation, hydroxyethyl starch （HES） centrifugation sedimentation, etc. Neural growth factor （NGF） and other factors play an important role in the growth of NSCs, such as transforming growth factor （TGF） is an important player in repairing organs, NGF accelerates the process of growth, insulin-like growth factor serves importantly in the differentiation of stem cells into neuron-like cells. CONCLUSION ： As unipotent stem cells, NSCs have the abilities of self-renewal and potential of high differentiation. The method of mechanical dissociation is better than trypsin digestion in e separating ESCs. However, density gradient centrifuge separation is better than other methods in the separation of the BMSCs. NGF and other factors play an important role in the growth of NSCs.

Denervated hippocampus provides a favorable microenvironment for neuronal differentiation of endogenous neural stem cells

Fimbria-fornix transection induces both exogenous and endogenous neural stem cells to differentiate into neurons in the hippocampus.This indicates that the denervated hippocampus provides an environment for neuronal differentiation of neural stem cells.However,the pathways and mechanisms in this process are still unclear.Seven days after fimbria fornix transection,our reverse transcription polymerase chain reaction,western blot assay,and enzyme linked immunosorbent assay results show a significant increase in ciliary neurotrophic factor m RNA and protein expression in the denervated hippocampus.Moreover,neural stem cells derived from hippocampi of fetal（embryonic day 17） Sprague-Dawley rats were treated with ciliary neurotrophic factor for 7 days,with an increased number of microtubule associated protein-2-positive cells and decreased number of glial fibrillary acidic protein-positive cells detected.Our results show that ciliary neurotrophic factor expression is up-regulated in the denervated hippocampus,which may promote neuronal differentiation of neural stem cells in the denervated hippocampus.

P2X receptors in maintenance and differentiation of neural progenitor cells

Purinergic receptors are among the first cell surface receptors expressed during embryonic development （Burnstock and Ulrich, 2011）. These are characterized based on their pharmacological properties of being activated by adenosine or purine/pyrimidine nucleotides as P1 and P2 receptors. P2 receptors are further classified by their structure as P2Y metabotropic and P2X ionotropic receptors.

Correlation between receptor-interacting protein 140 expression and directed differentiation of human embryonic stem cells into neural stem cells

Overexpression of receptor-interacting protein 140（RIP140） promotes neuronal differentiation of N2 a cells via extracellular regulated kinase 1/2（ERK1/2） signaling.However,involvement of RIP140 in human neural differentiation remains unclear.We found both RIP140 and ERK1/2 expression increased during neural differentiation of H1 human embryonic stem cells.Moreover,RIP140 negatively correlated with stem cell markers Oct4 and Sox2 during early stages of neural differentiation,and positively correlated with the neural stem cell marker Nestin during later stages.Thus,ERK1/2 signaling may provide the molecular mechanism by which RIP140 takes part in neural differentiation to eventually affect the number of neurons produced.

Editor's Choice——Umbilical cord blood mesenchymal stem cell differentiation and transplantation in neural regeneration

Previous in vivo experiments have shown that human umbilical cord blood mesenchymal stem cells can promote the proliferation and differentiation of damaged celts, and help to repair damaged sites, Recent studies have reported that umbilical cord blood-derived mesenchymal stem cells can differentiate into neurons and glial cells. Recent studies have reported that the repair mechanisms underlying cord blood stern cells involve the replacement of damaged cells and mediation of the local micro-environment.

Biochemical properties of norepinephrine as a kind of neurotransmitter secreted by bone marrow-derived neural stem cells induced and differentiated in vitro

BACKGROUND： It has been proved by many experimental studies from the aspects of morphology and immunocytochemistry in recent years that bone marrow stromal cells （BMSCs） can in vitro induce and differentiate into the cells possessing the properties of nerve cells. But the functions of BMSCs-derived neural stem cells（NSCs） and the differentiated neuron-like cells are still unclear. OBJECTIVE： To observe whether bone marrow-derived NSCs can secrete norepinephrine （NE） under the condition of in vitro culture, induce and differentiation, and analyze the biochemical properties of BMSCs-derived NSCs. DESIGN： A non-randomized and controlled experimental observation SETTING ： Institute of Neuromedicine of Chinese PLA, Zhujiang Hospital, Southern Medical University MATERIALS： This experiment was carried out in the Institute of Neuromedicine of Chinese PLA, Zhujiang Hospital, Southern Medical University. The bone marrow used in the experiment was collected from 1.5- month-old healthy New Zealand white rabbits. METHODS： This experiment was carried out in the Institute of Neuromedicine of Chinese PLA, Zhujiang Hospital, Southern Medical University. The bone marrow used in the experiment was collected from 1.5 month-old healthy New Zealand white rabbits. BMSCs of rabbits were isolated and performed in vitro culture, induce and differentiation with culture medium of NSCs and differentiation-inducing factor, then identified with immunocytochemical method. Experimental grouping： ①Negative control group： L-02 hepatic cell and RPMI1640 culture medium were used. ② Background culture group： Only culture medium of NSCs as culture solution was added into BMSCs to perform culture, and 0.1 volume fraction of imported fetal bovine serum was supplemented 72 hours later. ③Differentiation inducing factor group： After culture for 72 hours, retinoic acid and glial cell line-derived neurotrophic factors were added in the culture medium of BMSCs and NSCs as corresponding inducing factors. The level of NE in each group was detected on the day of culture and 5, 7, 14 and 20 days after culture with high performance liquid chromatography （HPLC）. The procedure was conducted 3 times in each group.Standard working curve was made according to the corresponding relationship of NE concentration and peak area. The concentration of NE every 1×10^7 cells was calculated according to standard curve and cell counting. MAIN OUTCOME MEASURES ： The level of NE of cultured cells was detected with HPLC; immunocytochemistrical identification of Nestin and neuron specific nuclear protein was performed. RESULTS： ① On the 14^th day after cell culture, BMSCs turned into magnus and round cells which presented Nestin-positive antigen, then changed into neuron-like cells with long processus and presented neuron specific nuclear protein -positive antigen at the 20^th day following culture. ② The ratio of NE concentration and peak area has good linear relationship, and regression equation was Y=1.168 36＋0.000 272 8X,r=-0.998 4. Coefficient variation （CV） was 〈 5% and the recovery rate was 92.39%（ Y referred to concentration and X was peak area）.③NE was well detached within 10 minutes under the condition of this experiment. ④ NE was detected in NSCs and their culture mediums, which were cultured for 7, 14 and 20 days respectively, but no NE in BMSCs, NSCs-free culture medium and L-02 hepatic cell which were as negative control under the HPLC examination. Analysis of variance showed that the level of NE gradually increased following the elongation of culture time （P 〈 0.01 ）. No significant difference in the level of NE existed at the same time between differentiation inducing factor group and basic culture group（P 〉 0.05）. CONCLUSION ： BMSCs of rabbits can proliferate in vitro and express Nestin antigen; They can differentiate into neuron-like cells, express specific neucleoprotein of mature neurons, synthesize and secrete NE as a kind of neurotransmitter.

Improvement of neurological function in rats with spinal cord injury after the transplantation of neural stem cells directly differentiated from bone marrow mesenchymal stem cells

Objective To study the effect and mechanism of neurological function recovery in rats with spinal cord injury ( SCI) rats after transplantation of neural stem cells which are directly differentiated from bone marrow mesenchymal stem cells ( BMSC ) ,and to investigate the suitable engraftment time. Methods BMSC at 3rd passage were differentiated into neural stem cells ( NSC) , and immunofluorescence staining was used to

Transformation of human amniotic epithelial cells into neuron-like cells in the microenvironment of traumatic brain injury in vivo and in vitro

Survival and differentiation of transplanted cells is closely related to the local microenvironment.The present study cultured human amniotic epithelial cells（HAECs） in a simulated microenvironment in vitro comprising RPMI 1640 culture medium and the solution extracted from injured brain tissues.Some HAECs were round,triangular in form or irregularly shaped,with extended neuron-like processes;some of the processes were interconnected,representing neuron-like morphology and some HAECs were microtubule-associated protein 2-positive.HAECs survived for at least 4 weeks following transplantation into the center and edges of the trauma focus with traumatic brain injury,and were microtubule-associated protein 2-positive.Moreover,the motor function of rat hind limbs was significantly improved.

Effects of halobenzoquinone and haloacetic acid water disinfection byproducts on human neural stem cells

Human neural stem cells（h NSCs） are a useful tool to assess the developmental effects of various environmental contaminants; however, the application of h NSCs to evaluate water disinfection byproducts（DBPs） is scarce. Comprehensive toxicological results are essential to the prioritization of DBPs for further testing and regulation. Therefore, this study examines the effects of DBPs on the proliferation and differentiation of h NSCs. Prior to DBP treatment, characteristic protein markers of h NSCs from passages 3 to 6 were carefully examined and it was determined that h NSCs passaged 3 or 4 times maintained stem cell characteristics and can be used for DBP analysis. Two regulated DBPs, monobromoacetic acid（BAA） and monochloroacetic acid（CAA）, and two emerging DBPs, 2,6-dibromo-1,4-benzoquinone（2,6-DBBQ） and 2,6-dichloro-1,4-benzoquinone（2,6-DCBQ）, were chosen for h NSC treatment. Both 2,6-DBBQ and 2,6-DCBQ induced cell cycle arrest at S-phase at concentrations up to 1 μmol/L. Comparatively, BAA and CAA at 0.5 μmol/L affected neural differentiation. These results suggest DBP-dependent effects on h NSC proliferation and differentiation. The DBP-induced cell cycle arrest and inhibition of normal h NSC differentiation demonstrate the need to assess the developmental neurotoxicity of DBPs.

Biological effect of velvet antler polypeptides on neural stem cells from embryonic rat brain

Background Velvet antler polypeptides (VAPs), which are derived from the antler velvets, have been reported to maintain survival and promote growth and differentiation of neural cells and, especially the development of neural tissues This study was designed to explore the influence of VAPs on neural stem cells in vitro derived from embryonic rat brain Methods Neural stem cells derived from E12 14 rat brain were isolated, cultured, and expanded for 7 days until neural stem cell aggregations and neurospheres were generated The neurospheres were cultured under the condition of different concentration of VAPs followed by immunocytochemistry to detect the differentiation of neural stem cells Results VAPs could remarkablely promote differentiation of neural stem cells and most neural stem cells were induced to differentiate towards the direction of neurons under certain concentration of VAPs Conclusion Neural stem cells can be successfully induced into neurons by VAPs in vitro , which could provide a basis for regeneration of the nervous system