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. 2021 Feb 8;12(2):159. doi: 10.1038/s41419-021-03454-9

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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. 4 — A, B Flow cytometry analysis (A) and its quantification (B) showing the production of mitochondrial ROS in tetraarsenic hexoxide-treated TNBC cells. Cells were treated with 2.5 and 5 μM tetraarsenic hexoxide for 24 h and then were stained with MitoSox Red. **P < 0.01, ***P < 0.001 versus control cells; C Fluorescent microscopy images showing MitoSox Red staining in 5 μM tetraarsenic hexoxide-treated TNBC cells for 24 h. Original magnification, ×50. Scale bar. D Representative immunoblot analysis showing the phosphorylation of STAT3 in tetraarsenic hexoxide–treated TNBC cells for the indicated times. E, F Representative immunoblot analysis (E) and densitometric quantitation (F) showing the phosphorylation of STAT3 in mitochondria and cytoplasmic fractions. 4T1 cells were treated with 5 μM tetraarsenic hexoxide for 24 h and then mitochondria and cytoplasm were isolated. ***P < 0.0005 versus control cells. G, H Flow cytometry analysis (G) and fluorescent microscopy images (H) showing the production of mitochondrial ROS in Stat3-knockdown 4T1 cells. 4T1 cells were transiently transfected with Stat3 siRNA and then treated with 5 μM tetraarsenic hexoxide for 24 h, followed by staining with MitoSox Red. Original magnification, ×50. Scale bar. i Representative immunoblot analysis showing cleaved caspase-3, PARP, and GSDME from tetraarsenic hexoxide-treated 4T1 cells transiently transfected with Stat3 siRNA. J LDH release by tetraarsenic hexoxide in Stat3-knockdown 4T1 cells. The data represent the mean ± S.D. of three independent experiments. ***P < 0.001 versus tetraarsenic hexoxide-untreated cells; ^#P < 0.01 versus tetraarsenic hexoxide-treated control cells. All P values were calculated by unpaired two-tailed Student’s t-tests. The data represent the mean ± S.D. of three independent experiments. FL full length, N N-terminus.