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. 2017 Jun 23;7:4153. doi: 10.1038/s41598-017-03592-1

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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HUCPVC Conditioned Media (CM) induces axonal growth in CNS neurons. (A,B) Effect of CM in axonal outgrowth on cortical and hippocampal neurons. At DIV3 neurons were stimulated for 14 hours with CM. Axonal outgrowth was assessed by immunocytochemistry using an antibody against Tau, an axonal specific marker. Images were taken from random neurons using an AxioObserver Z1 fluorescent microscope with a PlanApochromat 20× objective. (C–F) Quantification of axonal length and axonal longest segment. Results show that axonal network increase after 14 h of CM stimulation (C,D), and in addition neurons stimulated with CM have the longest axonal segments (E,F), demonstrating that global application of CM to both hippocampal and cortical neurons induce an increase in axonal outgrowth. Axonal length and longest axonal segments analysis was performed with Image J 1.45e software. Bars and plots represent the mean ± SEM of approximately 45 neurons randomly selected of 3 independent experiments. (C) ****Represents p < 0.0001; (D) *Represents p = 0.0025; (E) ***Represents p = 0.0010; (F) *Represents p = 0.0083 by Mann Whitney unpaired t-test when compared to Ctr. The scale bar is 25 µm.