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. 2017 Jun 15;7:3594. doi: 10.1038/s41598-017-03932-1

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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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DENV-infected BHK21 activates ISGs in neighboring cells via gap junctions. (A) BHK21 cells, seeded with increasing proximal distance, were infected with DENV2-PDK53 at MOI 1 for 24 hrs before DENV infection was measured by DENV RNA detection. Results are normalized to HPRT expression and represent mean ± SD of at least two independent experiments. (B) BHK21 cells were seeded at either 1.2 × 10⁵ or 2 × 10⁴ cells per well and either uninfected or infected with DENV2-PDK53 at MOI 1 for 24 hrs. CXCL10 expression was then measured via RT-qPCR and normalized to HPRT expression. (C,D) BHK21 cells were treated with increasing concentration of CBX before infection with DENV2-PDK53 at MOI 1 for 24 hrs. Viral load was assessed by detection of DENV viral RNA with RT-qPCR (C) and plaque assay (D). RT-qPCR results are normalized to HPRT expression and represent mean ± SD of at least seven independent experiments. Plaque assay results shown represent triplicate samples. (E) Spreading of DENV2-PDK53 at 72 hrs post infection in BHK21 cells treated with 300 μM CBX was assessed using immunofluorescent staining of DENV E-protein (red) and nuclei (DAPI, blue). All data are represented as mean ± SD, and *depicts P < 0.05 and ***depicts P < 0.001.

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