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. 2019 Nov 20;9:17213. doi: 10.1038/s41598-019-53610-7

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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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In vitro differentiation of NP23-NHD13 DN thymocytes. (a) In vitro co-culture of DN cells from WT and NP23-NHD13 mice on OP9 and OP9-DL1 feeder layers with the indicated cytokines; DN cells were also Mac1⁻(1 × 10⁵ cells were plated). (b) Differentiation of DN cells on OP9 and OP9-DL1 at indicated timepoints. (c) graph represents Mac1+/Gr1 + percentage of WT and NP23-NHD13 DN cells on OP9. (d,e) Long term co-culture of DN cells from NP23-NHD13 mice on OP9. Mac1⁺/Gr1⁺ and Mac1⁻Gr1⁻CD44^hetCD25^hetThy-1⁺cKit^bright (designated Kit + -OP9) cells (1 × 10⁴ cells/mouse) were transplanted at 18 or 34 days of culture, respectively. (f) Survival of Mac1⁺/Gr1⁺ and Kit + -OP9 recipients. Data are analyzed by Log-rank (Mantel-Cox) test), P = 0.07. Engraftment of donor myeloid cells (CD45.2^+/Mac1⁺) is shown in both (g) Kit + -OP9 and (h) Mac1⁺/Gr1⁺ recipients (dotted circle indicates engraftment data obtained from BM).