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. 2017 Aug 14;7:8004. doi: 10.1038/s41598-017-08336-9

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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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Effect of endotoxins in the presence P. aeruginosa phage PNM. (A) Gene expression analysis of 12 immunity-related genes by means of RT-qPCR after 20 h of stimulation of PBMCs. PBMCs were stimulated with P. aeruginosa phage PNM, either a phage lysate (10⁵ pfu/PBMC; 0.1 EU/PBMC) or a highly purified phage preparation (10⁵ pfu/PBMC; 10⁻⁵ EU/PBMC) in combination with 0.1 EU/PBMC. The pro-inflammatory markers IL1A, IL1B, CXCL1 and CXCL5, and the anti-inflammatory markers SOCS3, IL10, IL1RN and IL6 are upregulated The addition of 0.1 EU/PBMC to the highly purified P. aeruginosa phage preparation does not revert the observed immune response to that of the phage lysate for IL10, IL1A, IL6, TNFA and CXCL1. (B) Principal components analysis of P. aeruginosa phage PNM with or without the addition of endotoxins. The immune response induced by the highly purified phage PNM () differs from the one induced by the phage PNM lysate (), as these two groups are visibly separated. When endotoxins are added to a final concentration of 0.1 EU/PBMC to the highly purified phage (), the immune response is similar to the highly purified phage () and not towards the phage lysate (), indicating that the observed difference is not due to the presence of LPS but due to bacterial proteins present in the phage lysate.

Inline graphic — Effect of endotoxins in the presence P. aeruginosa phage PNM. (A) Gene expression analysis of 12 immunity-related genes by means of RT-qPCR after 20 h of stimulation of PBMCs. PBMCs were stimulated with P. aeruginosa phage PNM, either a phage lysate (10⁵ pfu/PBMC; 0.1 EU/PBMC) or a highly purified phage preparation (10⁵ pfu/PBMC; 10⁻⁵ EU/PBMC) in combination with 0.1 EU/PBMC. The pro-inflammatory markers IL1A, IL1B, CXCL1 and CXCL5, and the anti-inflammatory markers SOCS3, IL10, IL1RN and IL6 are upregulated The addition of 0.1 EU/PBMC to the highly purified P. aeruginosa phage preparation does not revert the observed immune response to that of the phage lysate for IL10, IL1A, IL6, TNFA and CXCL1. (B) Principal components analysis of P. aeruginosa phage PNM with or without the addition of endotoxins. The immune response induced by the highly purified phage PNM () differs from the one induced by the phage PNM lysate (), as these two groups are visibly separated. When endotoxins are added to a final concentration of 0.1 EU/PBMC to the highly purified phage (), the immune response is similar to the highly purified phage () and not towards the phage lysate (), indicating that the observed difference is not due to the presence of LPS but due to bacterial proteins present in the phage lysate.