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. 2021 Apr 30;12:2482. doi: 10.1038/s41467-021-22750-8

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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. 2 — a Kaplan–Meier survival curve of Cux1^+/−;Flt3^ITD (n = 14, magenta line), Flt3^ITD (n = 9, orange line), Cux1^+/fl;Vav-iCre⁺ (Cux1^+/−) (n = 11, red line) and control (Vav-iCre^+/− or Cre-negative littermates, Con, n = 11, blue line) mice. Increased mortality in Cux1^+/−;Flt3^ITD mice (median survival 28 weeks; control Con versus Cux1^+/−;Flt3^ITD, P < 0.0001; Cux1^+/− versus Cux1^+/−;Flt3^ITD, P < 0.0001; Flt3^ITD versus Cux1^+/−;Flt3^ITD, P = 0.0002; log-rank test). Open square symbols indicate those mice diagnosed with MDS/MPN and black circles indicate AML cases. b–e Red blood cell count (b), red blood cell mean cell volume (c) neutrophil count (d) and monocyte count (e) in control Con (n = 13), Cux1^+/− (n = 10), Flt3^ITD (n = 9) and Cux1^+/−;Flt3^ITD (n = 15) mice at 10 weeks of age. f Representative May-Grünwald-Giemsa (MGG)-stained blood smears from 10-week-old Flt3^ITD (left) and Cux1^+/−;Flt3^ITD (right) mice. Insets show an abnormal monocyte and blast cell (left); dysplastic neutrophil (right). Scale bar, 20 µm. Five Flt3^ITD and 10 Cux1^+/−;Flt3^ITD mice were assessed. g Plot of necropsy spleen weights normalized to body weight (left) from control Con (n = 7), Cux1^+/− (n = 8), Flt3^ITD (n = 6) and Cux1^+/−;Flt3^ITD (n = 6) mice. Photograph of representative spleen from 10-week-old Cux1^+/−;Flt3^ITD mouse (right). h–k Representative MGG-stained bone marrow cytospin (h) and hematoxylin and eosin (H&E)-stained bone marrow (i), spleen (j) and liver (k) sections from 17-week-old Flt3^ITD (left) and Cux1^+/−;Flt3^ITD (right) mice. Bone marrow and tissues are infiltrated with excess myelomonocytic cells in the Cux1^+/−;Flt3^ITD mouse. Scale bars; black = 20 µm, white = 100 µm. Five Flt3^ITD and 10 Cux1^+/−;Flt3^ITD mice were assessed. l Flow cytometric plot showing distribution of Gr1⁺ and Mac1⁺ cells in bone marrow (left) and spleen (right) from a 15-week-old Cux1^+/−;Flt3^ITD mouse diagnosed with CMML. Percentage of Gr1⁺Mac1⁺ cells is shown. m Immunohistochemistry of liver section from a 15-week-old Cux1^+/−;Flt3^ITD mouse diagnosed with MDS/MPN showing infiltration with myeloperoxidase (MPO)-expressing cells. Scale bar, 100 µm. Three mice were assessed. n Representative MGG-stained blood smear showing blast cells from a 30-week-old Cux1^+/−;Flt3^ITD mouse diagnosed with AML. Scale bar, 20 µm. Five mice were assessed. o Flow cytometric plot showing percentage of c-Kit⁺ AML cells in bone marrow from a 30-week-old Cux1^+/−;Flt3^ITD mouse diagnosed with AML. All plots show mean + s.e.m. Each circle represents one mouse. One-way ANOVA with Tukey’s test for multiple comparisons.