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. 2017 Jul 10;7:4928. doi: 10.1038/s41598-017-05401-1

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© The Author(s) 2017

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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LepR⁺Runx2-GFP^low cells differentiate into osteoblasts through multilayered cell formation in response to PTH-induced anabolic effects. (A–F) Z-stack confocal images of thick bone sections of 6 week-old LepR-Cre/Tomato/Runx2-GFP mice (A and B), tamoxifen-administered iOsx/Tomato/Runx2-GFP mice (C and D), and tamoxifen-administered iOsx/Tomato/Col1(2.3)-GFP mice (E and F) with vehicle (A,C and E) and hPTH(1–34) (B,D and F) intermittent treatment. Arrows: LepR-Cre-derived Tomato⁺(LepR/Tomato⁺)Runx2-GFP⁺ cells (A and B), iOsx/Tomato⁺Runx2-GFP⁺ cells (D) and iOsx/Tomato⁺Col1(2.3)-GFP⁺ cells (F). Arrowheads: iOsx/Tomato⁻Runx2-GFP⁺ cells (C and D) and iOsx/Tomato⁺Col1(2.3)-GFP⁻ cells (F). *: Bone-lining mature osteoblasts. Nuclei were visualized with To-PRO-3 (blue).