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. 2021 Apr 27;28:32. doi: 10.1186/s12929-021-00726-6

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© The Author(s) 2021

Open AccessThis 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/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

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Fig. 4 — NKX6-1 promotes cancer stemness properties in LMS cells. Cells were seeded in ultralow attachment plates to assess sphere-forming ability. a Representative spheroid images from different NKX6-1 transfectants. NKX6-1-overexpressing MES-SA cells formed spheroids with round morphology (upper), and NKX6-1 knockdown MES-SA/DX5 cells formed clusters of loosely associated cells (lower). The scale bar represents 100 μm. b The number of spheres was quantified using ImageJ software. The total number of spheres increased upon NKX6-1 overexpression in MES-SA cells and decreased upon NKX6-1 knockdown in MES-SA/DX5 cells. The expression of cancer stemness-related genes correlated with sphere formation ability. c qRT-PCR was performed to evaluate stemness markers (including NESTIN, NANOG, KLF4, KLF8, MYC, OCT, SOX2, CD44, and CD49F) in MES-SA cells transfected with either NKX6-1 or vector control. d Expression of KLF8, MYC, and CD49F in MES-SA/DX5 cells transfected with either shNKX6-1 or scrambled control was determined by qRT-PCR. *p < 0.05