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. 2020 Oct 7;10:16686. doi: 10.1038/s41598-020-73840-4

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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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The mechanism of action of both NESF and NLP in the somatotrophs involves the modulation of the cAMP/PKA/CREB signalling pathway. The effects of NESF or NLP pre-incubation on the CREB phosphorylation ratio by forskolin (FK) (a), CPT (b), or GRL (c) were evaluated. Representative immunoreactive bands of phosphorylated CREB (P-CREB), total CREB (T-CREB) and β-actin, and quantification of band intensity of P-CREB normalized to T-CREB. Four independent experiments with triplicates (n = 3 wells/treatment/experiment) were performed for each study. Data from all four experiments were pooled to conduct statistical analyses and are shown as mean ± SEM (n = 12 wells) and presented as a fold change over Control. Different letters indicate significant differences (p < 0.05) between the different treatments detected by one-way ANOVA test followed by Tukey’s multiple comparison test. When two groups share one letter, they are not statistically different.