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. 2017 Jul 14;7:5454. doi: 10.1038/s41598-017-05707-0

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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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Netrin-1 activation of the NF-κB pathway was dependent on its canonical receptor UNC5A. (a) Western blotting analysis of canonical receptors of netrin-1 in the adult mouse brain. (b) Western blotting analysis of canonical receptors of netrin-1 in U87MG, U251 and SHG44 cells. (c) Western blotting analyses of canonical receptors of netrin-1 in glioma tumor tissues (T) and normal brain tissues (N) from clinical samples. (d and e) UNC5A mRNA and protein expression levels were determined by real-time PCR and western blotting in U251 cells infected with lentivirus expressing shCtrl or shUNC5A. (f) Western blotting analysis of p65^ser536 and c-Myc in U251 cells infected with lentivirus expressing shCtrl or shUNC5A with or without netrin-1 stimulation. (g) Quantification of p65^ser536 phosphorylation using Gel-Pro Analyzer 4 software. Phosphorylation of NF-κB p65^ser536 was normalized to total p65, and c-Myc was normalized to GAPDH. (h) CCK-8 assays revealed that the addition of netrin-1 promoted cell proliferation. On the contrary, silencing of UNC5A blocked netrin-1-induced cell proliferation. The data are shown as the mean ± SEM.