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. 2020 Nov 12;10:19644. doi: 10.1038/s41598-020-76883-9

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© The Author(s) 2020

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 licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence 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 licence, visit http://creativecommons.org/licenses/by/4.0/.

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IL-8 secretion by LPS induces CXCR2 gene expression and RGS12 inhibits CXCR2-induced signaling activated by LPS in NCI-H292 cells. (a) IL-8 secretion was measured by ELISA. *, p < 0.05 compared with control cells. (b) Cells were treated with consensus PDZ or mutant PDZ peptide for 24 h and then incubated with LPS for four hours. The ELISA for IL-8 was carried out with cell lysates. *p < 0.05 compared with the control; **p < 0.05 compared with LPS only; ***p < 0.05 compared with LPS and consensus PDZ peptide treatment. (c) Cells were incubated with LPS in a time-dependent manner, the lysates were harvested and then analyzed by qRT-PCR and Western blot. *p < 0.05 compared with the control. (d) Cells were treated with consensus PDZ or mutant PDZ peptide for 24 h and then incubated with LPS for four hours. Expression of CXCR2 and RGS12 was measured by qRT-PCR. *p < 0.05 compared with the control; **p < 0.05 compared with LPS only; ***p < 0.05 compared with LPS and consensus PDZ peptide treatment. (e, upper panel) The cells were treated with consensus PDZ or mutant PDZ peptide. After 24 h, the cells were re-trypsinized and seeded at 7000 cells/well into a 96-well plate. The cells were incubated with LPS for four hours, and a cAMP assay was performed according to the manufacturer’s instructions (cAMP-Glo assay; Promega; Madison, WI). *p < 0.05 compared with the control; **p < 0.05 compared with LPS only; ***p < 0.05 compared with LPS and consensus PDZ peptide treatment. (e, lower panel) The cells were transfected with an RGS12 overexpression construct or siRNA-RGS12, the cells were incubated with LPS for 4 h, and a cAMP assay was performed. *p < 0.05 compared with the control; **p < 0.05 compared with LPS only; ***p < 0.05 compared with LPS and RGS12 treatment. All data shown are representative of three independent experiments.