| Purification and transfection of cochlear Schwann cells. | |
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MedLine Citation:
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PMID: 20837108 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Schwann cells line nerve fibers in the peripheral nervous system (PNS) and synthesize myelin. In addition, they support neuronal survival, neurite growth and regeneration. In dissociated cultures of postnatal mouse spiral ganglia, regenerating neurites spontaneously associate with Schwann cells. However, the mechanisms and consequences of interactions between cochlear Schwann cells and spiral ganglion neurites have not been examined. Further, the similarities and differences between cochlear Schwann cells and other PNS Schwann cells have not been studied. Experiments to examine these questions will rely on the ability to purify and characterize cochlear Schwann cells. Here we present methods for purifying Schwann cells from postnatal mouse cochleas and for transfecting them with expression plasmids. Dissociated spiral ganglia were plated on poly-D-lysine/laminin in medium containing neurotrophins, leukemia inhibitory factor (LIF), N2 supplement and serum and maintained for 5 days. Cells were harvested with trypsin/EDTA and subjected to an immuno-magnetic purification procedure. After 24 h in vitro, cultures were >85% Schwann cells. Nucleofection of purified Schwann cells with pMax-green fluorescent protein (pMax-GFP) plasmid, or with pEGFP-C-vimentin plasmid returned >45% transfection efficiency. These methods will allow the in-depth characterization of cochlear Schwann cells and an evaluation of their biochemical, functional, and genetic mechanisms that may promote neurite growth from the spiral ganglion. |
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Authors:
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D S Whitlon; D Tieu; M Grover |
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Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't Date: 2010-09-15 |
Journal Detail:
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Title: Neuroscience Volume: 171 ISSN: 1873-7544 ISO Abbreviation: Neuroscience Publication Date: 2010 Nov |
Date Detail:
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Created Date: 2010-10-18 Completed Date: 2011-02-02 Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 7605074 Medline TA: Neuroscience Country: United States |
Other Details:
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Languages: eng Pagination: 23-30 Citation Subset: IM |
Copyright Information:
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Copyright © 2010 IBRO. Published by Elsevier Ltd. All rights reserved. |
Affiliation:
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Department of Otolaryngology-Head and Neck Surgery, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA. whitlon@northwestern.edu |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Animals Animals, Newborn Cell Separation / methods Cells, Cultured Green Fluorescent Proteins / genetics Mice Neurites / metabolism Receptors, Nerve Growth Factor / metabolism SOXE Transcription Factors / metabolism Schwann Cells / cytology*, physiology* Spiral Ganglion / cytology*, growth & development* Transfection / methods* Tubulin / metabolism Vimentin / metabolism |
| Chemical | |
Reg. No./Substance:
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0/Receptors, Nerve Growth Factor; 0/SOXE Transcription Factors; 0/Sox10 protein, mouse; 0/TNFRSF16 protein, mouse; 0/Tubulin; 0/Vimentin; 0/beta3 tubulin, mouse; 147336-22-9/Green Fluorescent Proteins |
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