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Optimized Culture System to Induce Neurite Outgrowth From Retinal Ganglion Cells in Three-Dimensional Retinal Aggregates Differentiated From Mouse and Human Embryonic Stem Cells

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Title: Optimized Culture System to Induce Neurite Outgrowth From Retinal Ganglion Cells in Three-Dimensional Retinal Aggregates Differentiated From Mouse and Human Embryonic Stem Cells
Other Titles: マウス及びヒト胚性幹細胞より分化した三次元網膜様組織内の網膜神経節細胞からの神経突起伸長を誘導する培養系
Authors: 前川, 有紀
Authors (alternative): Maekawa, Yuki
Issue Date: 30-Sep-2016
Publisher: Taylor & Francis
Citation: Nagasaki University (長崎大学), 博士(医学) (2016-09-30)
Abstract: Purpose: To establish a practical research tool for studying the pathogenesis of retinal ganglion cell (RGC) diseases, we optimized culture procedures to induce neurite outgrowth from three-dimensional self-organizing optic vesicles (3D-retinas) differentiated in vitro from mouse and human embryonic stem cells (ESCs). Materials and methods: The developing 3D-retinas isolated at various time points were placed on Matrigel-coated plates and cultured in media on the basis of the 3D-retinal culture or the retinal organotypic culture protocol. The number, length, and morphology of the neurites in each culture condition were compared. Results: First, we confirmed that Venus-positive cells were double-labeled with a RGC marker, Brn3a, in the 3D-retina differentiated from Fstl4::Venus mouse ESCs, indicating specific RGC-subtype differentiation. Second, Venus-positive neurites grown from these RGC subsets were positive for beta-III tubulin and SMI312 by immunohistochemistry. Enhanced neurite outgrowth was observed in the B27-supplemented Neurobasal-A medium on Matrigel-coated plates from the optic vesicles isolated after 14 days of differentiation from mouse ESCs. For the differentiated RGCs from human ESCs, we obtained neurite extension of >4 mm by modifying Matrigel coating and the culture medium from the mouse RGC culture. Conclusion: We successfully optimized the culture conditions to enhance lengthy and high-frequency neurite outgrowth in mouse and human models. The procedure would be useful for not only developmental studies of RGCs, including maintenance and projection, but also clinical, pathological, and pharmacological studies of human RGC diseases.
Description: 長崎大学学位論文 学位記番号:博(医歯薬)甲第885号 学位授与年月日:平成28年9月30日 / Author: Yuki Maekawa, Akishi Onishi, Keizo Matsushita, Naoshi Koide, Michiko Mandai, Kiyoshi Suzuma, Takashi Kitaoka, Atsushi Kuwahara, Chikafumi Ozone, Tokushige Nakano, Mototsugu Eiraku & Masayo Takahashi / Citation: Current Eye Research, 41(4), pp.558-568; 2016
Keywords: Culture procedure / neuritogenesis / pluripotent stem cells / retinal ganglion cell / three-dimensional optic vesicle
URI: http://hdl.handle.net/10069/37762
ISSN: 02713683
DOI: 10.3109/02713683.2015.1038359
Relational Links: http://hdl.handle.net/10069/36853
Rights: © 2016 Taylor & Francis, LLC. / This is an Accepted Manuscript of an article published by Taylor & Francis in Current Eye Research on 16 April 2015, available online: http://www.tandfonline.com/10.3109/02713683.2015.1038359.
Type: Thesis or Dissertation
Text Version: ETD
Appears in Collections:dissertation

Citable URI : http://hdl.handle.net/10069/37762

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