We have made considerable progress towards understanding the cellular and molecular mechanisms that regulate the proliferation, differentiation and survival of neural progenitor cells in the developing and adult central nervous system. We found that SDFalpha, activates CXCR4 in glial progenitor cells resulting in increased migration and differentation of those cells. Our recent research has revealed a new molecular signaling system that regulates the fate of neural stem cells in the cerebral cortex. We used antibody-blocking and genetic experiments to reveal an requirement for laminin/integrin interactions in apical process adhesion and neural stem cell regulation. Transient abrogation of integrin binding and signalling using blocking antibodies to specifically target the ventricular region in utero results in abnormal cerebral cortex development. Using a multidisciplinary approach to analyse stem cell behaviour by expression of fluorescent transgenes and multiphoton time-lapse imaging revealed that the transient embryonic disruption of laminin/integrin signalling resulted in substantial layering defects in the postnatal neocortex. In other studies we found that mice lacking the monoamine metabolic enzymes MAO A and MAO B (MAO AB-deficient mice) exhibit diminished proliferation of neural stem cells (NSC) in the developing telencephalon beginning in late gestation embryonic day (E) 17.5, a deficit that persists in neonatal and adult mice. The results suggest that a MAO-dependent long-lasting alteration in the proliferation capacity of NSC occurs late in embryonic development and is mediated by serotonin. Glioblastoma brain tumors harbor a small population of cancer stem cells that are resistant to conventional chemotherapeutic and radiation treatments, and are believed responsible for tumor recurrence and mortality. The identification of the epigenetic molecular mechanisms that control self-renewal of glioblastoma stem cells will foster development of targeted therapeutic approaches. The transcriptional repressor REST, best known for its role in controlling cell fate decisions in neural progenitor cells, may also be crucial for cancer stem cell self-renewal. We discovered that reduced TRF2 binding to REST targets REST for proteasomal degradation and thereby inhibits cancer stem cell proliferation. Neurological side effects of treatments that target REST and TRF2 may be less severe than conventional brain tumor treatments because postmitotic neurons do not express REST and have relatively stable telomeres. Apart from protecting telomeres, nuclear TRF2 interacts with the master neuronal gene-silencer repressor element 1-silencing transcription factor (REST), and disruption of this interaction induces neuronal differentiation. We discovered the existence of a developmental switch from the expression of TRF2 in proliferating neural progenitor cells to expression of a unique short nontelomeric isoform of TRF2 (TRF2-S) as neurons establish a fully differentiated state. Unlike nuclear TRF2, which enhances REST-mediated gene repression, TRF2-S is located in the cytoplasm where it sequesters REST, thereby maintaining the expression of neuronal genes, including those encoding glutamate receptors, cell adhesion, and neurofilament proteins. In neurons, TRF2-S-mediated antagonism of REST nuclear activity is greatly attenuated by either overexpression of TRF2 or administration of the excitatory amino acid kainic acid. Overexpression of TRF2-S rescues kainic acid-induced REST nuclear accumulation and its gene-silencing effects. Thus, TRF2-S acts as part of a unique developmentally regulated molecular switch that plays critical roles in the maintenance and plasticity of neurons. Recently,we found that TLR3 protein is present in brain cells in early embryonic stages of development, and in cultured neural stem/progenitor cells (NPC). NPC from TLR3-deficient embryos formed greater numbers of neurospheres compared with neurospheres from wild-type embryos. Numbers of proliferating cells, as assessed by phospho histone H3 and proliferating cell nuclear antigen labeling, were also increased in the developing cortex of TLR3-deficient mice compared with wild-type mice in vivo. Treatment of cultured embryonic cortical neurospheres with a TLR3 ligand (polyIC) significantly reduced proliferating (BrdU-labeled) cells and neurosphere formation in wild type but not TLR3(-/-)-derived NPCs. Our findings reveal a novel role for TLR3 in the negative regulation of NPC proliferation in the developing brain. In a recent screen of a panel of botanical pesticides, we identified plumbagin as having neuroprotective activity. Recently, we determined if plumbagin could modify the developmental fate of rat E14.5 embryonic neural progenitor cells (NPC). Plumbagin exhibited no cytotoxicity when applied to cultured NPC at low concentrations. At intermediate concentrations plumbagin significantly enhanced the proliferation of NPC as indicated by a 17% increase in the percentage of cells incorporating bromo-deoxyuridine. Plumbagin at a low concentration stimulated the production of astrocytes as indicated by increased GFAP expression. Plumbagin selectively induced the proliferation and differentiation of glial progenitor cells without affecting the proliferation or differentiation of neuron-restricted progenitors. Plumbagin rapidly activated the transcription factor signal transducer and activator of transcription 3 (Stat3) in NPC, and a Stat3 inhibitor peptide prevented both plumbagin-induced astrocyte formation and proliferation. These findings demonstrate the ability of a low molecular weight naturally occurring phytochemical to control the fate of glial progenitor cells by a mechanism involving the Stat3 signaling pathway. Human embryonic stem cell (hESC)-derived dopaminergic (DA) neurons hold potential for treating Parkinson's disease (PD) through cell replacement therapy. Generation of DA neurons from hESCs has been achieved by coculture with the stromal cell line PA6, a source of stromal cell-derived inducing activity (SDIA). However, the factors produced by stromal cells that result in SDIA are largely undefined. We previously reported that medium conditioned by PA6 cells can generate functional DA neurons from NTera2 human embryonal carcinoma stem cells. Here we show that PA6-conditioned medium can induce DA neuronal differentiation in both NTera2 cells and the hESC I6 cell line. To identify the factor(s) responsible for SDIA, we used large-scale microarray analysis of gene expression combined with mass spectrometric analysis of PA6-conditioned medium (CM). The candidate factors, hepatocyte growth factor (HGF), stromal cell-derived factor-1 (SDF1), secreted frizzled-related protein 1 (sFRP1), and vascular endothelial growth factor D (VEGFD) were identified, and their concentrations in PA6 CM were established by immunoaffinity capillary electrophoresis. Upon addition of SDF1, sFRP1, and VEGFD to the culture medium, we observed an increase in the number of cells expressing tyrosine hydroxylase (a marker for DA neurons) and III-tubulin (a marker for immature neurons) in both the NTera2 and I6 cell lines. These results indicate that SDF1, sFRP1, and VEGFD are major components of SDIA and suggest the potential use of these defined factors to elicit DA differentiation of pluripotent human stem cells for therapeutic intervention in PD.

Agency
National Institute of Health (NIH)
Institute
National Institute on Aging (NIA)
Type
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
1ZIAAG000324-10
Application #
8552367
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
10
Fiscal Year
2012
Total Cost
$146,327
Indirect Cost
Name
National Institute on Aging
Department
Type
DUNS #
City
State
Country
Zip Code
Yang, Jenq-Lin; Lin, Yu-Ting; Chuang, Pei-Chin et al. (2014) BDNF and exercise enhance neuronal DNA repair by stimulating CREB-mediated production of apurinic/apyrimidinic endonuclease 1. Neuromolecular Med 16:161-74
Chigurupati, Srinivasulu; Mughal, Mohamed R; Okun, Eitan et al. (2013) Effects of cerium oxide nanoparticles on the growth of keratinocytes, fibroblasts and vascular endothelial cells in cutaneous wound healing. Biomaterials 34:2194-201
Barak, Boaz; Okun, Eitan; Ben-Simon, Yoav et al. (2013) Neuron-specific expression of tomosyn1 in the mouse hippocampal dentate gyrus impairs spatial learning and memory. Neuromolecular Med 15:351-63
Schwartz, Catherine M; Tavakoli, Tahereh; Jamias, Charmaine et al. (2012) Stromal factors SDF1*, sFRP1, and VEGFD induce dopaminergic neuron differentiation of human pluripotent stem cells. J Neurosci Res 90:1367-81
Stranahan, Alexis M; Mattson, Mark P (2012) Recruiting adaptive cellular stress responses for successful brain ageing. Nat Rev Neurosci 13:209-16
Hou, Yan; Ouyang, Xin; Wan, Ruiqian et al. (2012) Mitochondrial superoxide production negatively regulates neural progenitor proliferation and cerebral cortical development. Stem Cells 30:2535-47
Petralia, Ronald S; Wang, Ya-Xian; Mattson, Mark P et al. (2012) Subcellular distribution of patched and smoothened in the cerebellar neurons. Cerebellum 11:972-81
Mitchell, Nicholas; Petralia, Ronald S; Currier, Duane G et al. (2012) Sonic hedgehog regulates presynaptic terminal size, ultrastructure and function in hippocampal neurons. J Cell Sci 125:4207-13
Dees, Clara; Zerr, Pawel; Tomcik, Michal et al. (2011) Inhibition of Notch signaling prevents experimental fibrosis and induces regression of established fibrosis. Arthritis Rheum 63:1396-404
Okun, Eitan; Griffioen, Kathleen J; Mattson, Mark P (2011) Toll-like receptor signaling in neural plasticity and disease. Trends Neurosci 34:269-81

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