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. 2021 May 8;12(5):462. doi: 10.1038/s41419-021-03739-z

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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. 5 — A qPCR analysis of H322 cells with or without FTO shRNA expression or combined expression of FTO shRNA and MYC shRNA was performed. Data represent the means ± SD of triplicate samples. ***P < 0.001. B Immunoblotting analysis of H322 cells with or without FTO shRNA expression or combined expression of FTO shRNA and MYC shRNA was performed with the indicated antibodies. C, D Glucose consumption (C) and lactate production (D) of H322 cells with or without expression of FTO shRNA or combined expression of FTO shRNA and MYC shRNA were determined. Data represent the means ± SD of triplicate samples. **P < 0.01, ***P < 0.001, ****P < 0.0001. E H322 cells with or without FTO shRNA or MYC shRNA were cultured for the indicated periods of time and were harvested for cell counting. The data represent the means ± SD of triplicate samples. ****P < 0.0001. F, G The anchorage-independent growth of H322 cells with or without expression of FTO shRNA or combined expression of FTO shRNA and MYC shRNA was measured by soft agar assays (F). Scale bar: 100 μm. The data represent the means ± SD of triplicate samples. The migration and invasion of the indicated cells were measured (G). Scale bar: 100 μm. The number of migrated and invaded cells were counted from 3 different fields under a microscope. The data represent the means ± SD. **P < 0.01, ***P < 0.001, and ****P < 0.0001. H, I H358 cells with or without expression of FTO shRNA or combined expression of FTO shRNA and MYC shRNA were subcutaneously implanted into the flank regions of nude mice (n = 6). Photographs of tumors, tumor growth curves and tumor weights are shown or calculated (H). Data represent the means ± SD of 6 mice in each group. IHC staining of tumor tissues was performed with indicated antibodies. Representative images are shown (I). Scale bar: 100 μm. **P < 0.01, ***P < 0.001, and ****P < 0.0001. J H358 cells with or without expression of FTO shRNA or combined expression of FTO shRNA and MYC shRNA were injected into the tail veins of NOD/SCID mice (n = 5). Mouse lungs were stained by H&E. Representative images of metastatic tumors in the lungs are shown. The black arrows indicate metastatic tumors. Scale bar: 100 μm. The number of tumors in the lung was counted. Data represent the means ± SD of 5 mice in each group. ****P < 0.0001.