Figure 4.
Figure 4. Overexpression of MLF1 partly overcomes Trib1-induced AML. (A) Myeloid leukemia–free survival curves of mice transplanted with MLF1/Trib1-cotransduced BM cells compared with Trib1 alone. Results are derived from 3 independent transfer experiments. The P value between Trib1 and MLF1/Trib1 mice was calculated with the log-rank test. P < .01. (B) May-Grünwald Giemsa–stained peripheral blood (PB) smears and cytospins of BM cells from leukemic Trib1 (left panel), leukemic MLF1/Trib1 (middle panel), and nonleukemic MLF1/Trib1 (right panel) mice. Original magnification ×400. (C) A fluorescence-activated cell sorter (FACS) analysis of BM and spleen (SP) cells for immature (Mac-1+Gr-1lo) and differentiated (Mac-1+Gr-1hi) granulocytes. The population of GFP-positive cells in BM and SP is shown in the upper panels. (D-E) A qRT-PCR analysis of Trib1 and MLF1 expression in BM cells from Trib1 (n = 6), leukemic MLF1/Trib1 (n = 5), and nonleukemic MLF1/Trib1 (n = 5) mice. ΔCt values for each sample were standardized by the ΔCt values of β-actin followed by normalization with the percentage of GFP-positive cells. The level of expression in normal BM was set to 1.0 (D). The Trib1/MLF1 ratio of each sample in panel D. P values between leukemic MLF1/Trib1 and nonleukemic MLF1/Trib1 mice were calculated with Student t test. P < .005. (E). Data are presented as scatter diagrams with mean values (horizontal bar). (F-G) Microarray data analysis of the TRIB1 and MLF1 expression in human leukemia. Gene expression data of normal BMs (n = 9) from healthy donors and AML (n = 285) and ALL (n = 284) patient samples were obtained from the GEO data sets (F). Patients with AML were classified into positive groups with C/EBPα mutation (n = 17), EVI1 mutation (n = 20), RAS mutation (n = 34), FLT3 mutation (n = 106), and t(15;17) chromosomal translocation (n = 18). Negative vs positive groups with each gene alteration were analyzed (G). Box plots showed stratification of the TRIB1/MLF1 ratio from each sample. Significance was calculated by the Mann-Whitney U test (*P < .05; **P < .01; ***P < .001; n.s., not significant).

Overexpression of MLF1 partly overcomes Trib1-induced AML. (A) Myeloid leukemia–free survival curves of mice transplanted with MLF1/Trib1-cotransduced BM cells compared with Trib1 alone. Results are derived from 3 independent transfer experiments. The P value between Trib1 and MLF1/Trib1 mice was calculated with the log-rank test. P < .01. (B) May-Grünwald Giemsa–stained peripheral blood (PB) smears and cytospins of BM cells from leukemic Trib1 (left panel), leukemic MLF1/Trib1 (middle panel), and nonleukemic MLF1/Trib1 (right panel) mice. Original magnification ×400. (C) A fluorescence-activated cell sorter (FACS) analysis of BM and spleen (SP) cells for immature (Mac-1+Gr-1lo) and differentiated (Mac-1+Gr-1hi) granulocytes. The population of GFP-positive cells in BM and SP is shown in the upper panels. (D-E) A qRT-PCR analysis of Trib1 and MLF1 expression in BM cells from Trib1 (n = 6), leukemic MLF1/Trib1 (n = 5), and nonleukemic MLF1/Trib1 (n = 5) mice. ΔCt values for each sample were standardized by the ΔCt values of β-actin followed by normalization with the percentage of GFP-positive cells. The level of expression in normal BM was set to 1.0 (D). The Trib1/MLF1 ratio of each sample in panel D. P values between leukemic MLF1/Trib1 and nonleukemic MLF1/Trib1 mice were calculated with Student t test. P < .005. (E). Data are presented as scatter diagrams with mean values (horizontal bar). (F-G) Microarray data analysis of the TRIB1 and MLF1 expression in human leukemia. Gene expression data of normal BMs (n = 9) from healthy donors and AML (n = 285) and ALL (n = 284) patient samples were obtained from the GEO data sets (F). Patients with AML were classified into positive groups with C/EBPα mutation (n = 17), EVI1 mutation (n = 20), RAS mutation (n = 34), FLT3 mutation (n = 106), and t(15;17) chromosomal translocation (n = 18). Negative vs positive groups with each gene alteration were analyzed (G). Box plots showed stratification of the TRIB1/MLF1 ratio from each sample. Significance was calculated by the Mann-Whitney U test (*P < .05; **P < .01; ***P < .001; n.s., not significant).

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