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. 2018 Sep 13;8:13741. doi: 10.1038/s41598-018-32205-8

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© The Author(s) 2018

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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Suppression of cell growth by PIP/56-1 or PIP/56-2 treatment in EVI1^high AML cell lines. (a) Cell growth rates of two EVI1^high AML cell lines (UCSD/AML1 and Kasumi-3) and two EVI1^low AML cell lines (HEL and F36P) were examined after 1 μM PIP/AP2 (AP2), PIP/56-1 or PIP/56-2 treatment and parental cells (P) with no treatment or with DMSO only (C) as controls. Data represent the mean ± S.D. of triplicate determinations. *P < 0.05, ** P < 0.01 between parental control and treatments (Student’s t-test). Data are representative of three independent experiments. (b) Two EVI1^high AML cell lines, UCSD/AML1 and Kasumi-3, treated with 1 μM PIP/56-1 for 24 hours were transfected with GPR56 expression vector or empty vector, and cell proliferation rates were determined by Trypan blue staining. Data represent the means ± S.D. of triplicate determinations. *P < 0.05, **P < 0.01 (Student’s t-test). (c) Cell cycle profiles were determined by BrdU labeling from two EVI1^high AML cell lines (UCSD/AML1 and Kasumi-3) treated with 1 μM PIP/56-1, PIP/56-2, PIP/AP2 or DMSO solvent (C) for three days and parental cells (P). Data represent the mean ± S.D. of three independent experiments. *P < 0.05 between parental control and treatments (Student’s t-test). (d) Percentage of apoptotic cells determined in two AML cell lines (UCSD/AML1 and Kasumi-3) under the same conditions as in (a,b) by Annexin V staining. Data are expressed as the percentage Annexin V-positive cells 24 hours after treatment and represent the mean ± S.D. of three independent experiments. *P < 0.05 between parental control (P) and treatments (Student’s t-test).