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. 2021 Oct 4;11:19614. doi: 10.1038/s41598-021-99090-6

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© The Author(s) 2021

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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Gpnmb mRNA and protein levels of differently polarized Gpnmb^-/- and wildtype macrophages. Cells were treated with IL-4 and 20 ng/mL IL-13, or TGFβ, or LPS and IFNγ. (A–C) Transcript levels were measured by qRT-PCR after 4 h (A), 24 h (B) or 48 h (C). (D–F) Gpnmb protein expression and release of differently polarized macrophages after 48 h. (D) Exemplary Western blot of stimulated cell extracts. Undiluted lysates were loaded onto a SDS gel. Cropped image, the respective full-length blots are presented in Fig. S9. (E) Quantification of the two characteristic Gpnmb bands (indicated by arrows in D) of wildtype macrophage extracts, normalized to Gapdh signal as loading control. (F) Quantification of released soluble Gpnmb in cell culture supernatant by ELISA. n = 3, meaning BMDMs from three individual mice per group, mean ± SEM. Statistical differences were determined by a Two-way ANOVA with Bonferroni post-hoc tests; * p < 0.05; *** p < 0.001.