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. 2019 Jul 11;10:3055. doi: 10.1038/s41467-019-11044-9

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© The Author(s) 2019

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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Fig. 5 — LIF suppresses the Hippo-signaling pathway in human pancreatic cancer cells. a Representative HOPFlash luciferase assay showing YAP/TAZ-TEAD transcriptional activity in human pancreatic cancer cells receiving IL-6 or LIF at 100 ng/mL (N = 3, **P < 0.01, ***P < 0.001, ****P < 0.0001). b (Left panel) Representative western blots showing inducible knockdown of LIF in Panc1.0 by using two different hairpin sequences. (Right panel) Representative HOPFlash luciferase assay showing YAP/TAZ-TEAD transcriptional activity in Panc1.0 cells, which express inducible shRNA targeting human LIF (N = 3, **P < 0.01). Error bars from (a, b) represent the standard deviation and P-value was generated by t-test. c Western blots showing subcellular (nuclear) localization of total YAP in Panc1.0 cell line which express inducible shRNA against human LIF. d Representative western blots showing protein expression of phosphorylated LAST1, phosphorylated MOB1, and phosphorylated YAP at S127 in Panc6.03 which express inducible shRNA targeting human LIF. e Representative western blots showing protein expression of phosphorylated LAST1, phosphorylated MOB1, phosphorylated YAP at S127, KRAS, and phosphorylated ERK in human pancreatic cancer cells treated with LIF-neutralizing antibody at 2 μg/mL. f Western blots suggesting expression of phosphorylated YAP at S127 and phospho-STAT3 at Y705 in PDX tumors receiving gemcitabine or gemcitabine along with LIF antibody. The tumor samples were harvested from in vivo assay shown in Fig. 3g. g Sphere-forming efficiency in multiple human pancreatic cancer cell lines where YAP is knocked down by using siRNA in the presence or absence of human LIF in culture medium (N = 6, *P < 0.05, ***P < 0.001, ****P < 0.0001). Error bar represents the standard deviation. P-value was generated by t-test. Western blots in the upper panel showing YAP expression in different cell lines. h (Left panel) IHC suggesting the expression of LIF or phospho-YAP at S127 in human pancreatic tissues, including normal tissues and malignant tumors (Biomax PA484, PAB241). (Right panel) The correlation curve suggested that the expressions of LIF and phospho-YAP at S127 are negatively correlated in human pancreatic tissues. The imagines were taken, and the intensity of was quantified by the KEYENCE BZ-X800 microscope. The Pearson’s correlation coefficient was used to analyze the relationship between the staining index of LIF and phospho-YAP (S127)