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Spation-temporal Expression of A Novel Neuron-derived Neurotrophic Factor (ndnf) in Mouse Brains During Development

Author: KuangXiuLi
Tutor: SunGuoTao
School: Henan University
Course: Neurobiology
Keywords: Neuron-derived neurotrophic factor brain development secreted protein
CLC: Q42
Type: Master's thesis
Year: 2010
Downloads: 42
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Abstract

Background:The fate of neurons in the developing and adult nervous system is controlled, in part, by members of the neurotrophin family. This consists of four proteins: nerve growth factor (NGF), brain derived neurotrophic factor (BDNF), neurotrophin 3 (NT-3) and neurotrophin 4/5 (NT-4/5). Neurotrophic factors secreted by cells in the target field protect the neurons from apoptosis. Only those neurons that obtain sufficient neurotrophins survive. NGF can regulate the proliferation and differentiation of neuron precursor cells. BDNF can promote the survival of neurons and rescue them from damage, On the other hand, it can induce long-lasting changes in synaptic plasticity, neurotransmitter and neuropeptide production and excitability. Glial cell-line-derived neurotrophic factor (GDNF) is a protein known to enhance the survival of dopaminergic and motor neurons. NT-3 can promote growth and differentiation of different types of neurons both in CNS and PNS including neural nest and oligo dendroglial precursor cells. Lots of research showed that neurotrophins have a very important role in survive, differentiation and synaptic plasticity. They are candidates for treatment of the degenerative motor neural disorders, amyotrophic lateral sclerosis(ALS), spinal muscular atrophy and post-polio syndrome, and also the potential medicine for epileptic attack, degenerative diseases such as AD and Parkinson disease.Objective:Our main objective is to study a new gene, NDNF, which expressed only in the nerve system. We will determin the spatial expression characteristic of NDNF and the relation between NDNF expression and development. And the effect of NDNF on migration and differentiation of neurons will be studied. We hope that it can help us better to know the development process and mechanism of the nerve system, and also hope that it can provide clinical treatment clues for the central nervous system degenerative diseases.Method:The molecular biology technologies are used to study functions of NDNF, and the morphology methods are used to determine the spatiotemporal changes of NDNF expression during development.Results:1. Gene cloning and vector constructionThe sequencing results of cloned NDNF gene are concordant with that in the database Genebank. After eukaryotic expression plasmid was successful constructed, it was transfected into mammalian cells. We found that the intracellular localization of the new protein was consistent with secreted proteins, which are predominantly distributed around the nuclein, representing the endoplasmic reticulum and Golgi apparatus. Additionly glycosylated NDNF is found in the brain tissue. Western blot ting confirmed that NDNF can be secreted into culture medium.2. Neurontrophic functionPrimary cultured hippocampal pyramidal neurons were treated with conditioned medium containing recombinatnt NDNF. Compared with the control group, the axons of neurons in the treatment group were longer and the cell body size was larger. Transwell migration assay further certified that NDNF was not only provided direct trophic support to neurons but also promoted neuronal migration, with BDNF as positive control.3. Distribution of NDNF in tissue and developmental characteristicsTotal proteins were extracted from mouse brain, spinal cord, liver, heart and kidney,and then western blotting was used. The results confirmed that NDNF only exists in brain and spinal cord. Immunofluorescence assay further confirmed this distribution and additionally found that the localization of NDNF in the nervous system is consistent with neuron, not astrocyte. The higher expression level was found in Cajal-Retzius cells of cerebral cortex and Purkinje cells of cerebellum. From embryonic stage to adult, the expression level of NDNF is sustained and is coordinated with the number of neurons.Conclusion:NDNF is a new neurotrophic factor, which is conserved through evolution. NDNF belongs to secreted protein. In vitro assay show that NDNF can promote neuronal migration, differentiation and maturation. NDNF is widly expressed in the nervous system from embryo to adult, and its expression level changes along with the number of neurons.

Spation-temporal Expression of A Novel Neuron-derived Neurotrophic Factor (ndnf) in Mouse Brains During Development

Abstract

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