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. 2021 May 14;9:24. doi: 10.1038/s41413-021-00136-2

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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 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. 2 — NRP2 ablation increases osteoclast differentiation. a Protein analysis of NRP2 expression in a time course (0-6 days) in osteoclasts differentiated under different conditions in comparison to osteoclastic precursors under the conditions of RANKL (100 ng·mL⁻¹) and M-CSF (20 ng·mL⁻¹), LNCaP C4-2B CM and PC3 CM. Right: Comparison of NRP2 protein levels in RANKL and M-CSF, LNCaP C4-2B CM and PC3 CM at day 6. b Graph showing the expression of NRP2 at mRNA level in all the three conditions in the time course of osteoclast differentiation. Knockout of NRP2 in osteoclastic precursors isolated from NRP2 Fl/Fl; CSF1R-Cre transgenic mice by addition of 4-HydroxyTamoxifen in vitro and differentiated into osteoclasts in the presence of RANKL and M-CSF, LNCaP C4-2B CM and PC3 CM. c–e TRAP staining showing osteoclast differentiation at day 4–6 after depletion of NRP2 in RANKL and M-CSF, LNCaP C4-2B CM and PC3 CM, respectively. f–h Quantification of the TRAP positive multinucleated cells per well and comparison between NRP2WT and NRP2KO osteoclasts represented as a graph in RANKL and M-CSF, LNCaP C4-2B CM and PC3 CM, respectively. All values reported as mean± SEM from three independent experiments. Statistically significant P value denoted as * (0.05), ** (0.01), *** (0.001)