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. 2017 Jun 22;7:4092. doi: 10.1038/s41598-017-04460-8

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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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HUVEC co-culture and HUVEC-conditioned medium significantly enhanced rat DRG (rDRG) axonal growth. To confirm that HUVEC mediated axonal growth was applicable to mammalian systems, rDRG explants were cultured (A) on PDL wells (control), (B) in HUVEC-conditioned medium (conditioned), and (C) in direct co-culture with HUVECs (co-culture). Axons grew towards clusters of HUVECs when cultured in direct contact with HUVECs (arrows), and axons from DRGs grown on PDL well or with HUVEC-conditioned medium displayed radially symmetrical growth. Scale = 200 µm. Axonal outgrowth from (D) direct co-culture between low densities of HUVECs and rDRGs and from HUVEC-conditioned medium is significantly higher than control axons cultured on PDL wells (controls) and cultures supplemented with neutralizing antibody. Additionally, direct co-culture of HUVECs and rDRGs supported significantly longer axonal outgrowth than outgrowth from rDRGs cultured in HUVEC-conditioned medium. Data are presented as mean ± standard error. *(p < 0.05), ***(p < 0.001) and brackets indicate significance from indicated treatments by one-way ANOVA with Holm-Sidak post hoc analysis and ^††(p < 0.01) indicates significance between indicated treatments as determined two-tailed Student’s t-test (DRG sample size for each group shown in bars from 2–3 independent replicates).