Figure 2
Figure 2. Serial transplantation of Ezh2Δ/Δ hematopoietic cells caused heterogeneous hematologic malignancies. (A) Summary of secondary transplantation. A total of 2.5 × 106 BM cells from MDS and MDS/MPN mice ∼10 months after the deletion of Ezh2 were transplanted into secondary recipients irradiated at a lethal (8.5 Gy) or sublethal (6.5 Gy) dose. The engraftment status is indicated. (B) A Kaplan-Meier survival curve of WT (n = 4) and Ezh2Δ/Δ (n = 12) mice in secondary transplantation. Four recipient mice, which developed CD45.1+ T-ALL of a host cell origin, were excluded. *P < .05. Statistical significance was measured by log-rank test. (C) A pie graph showing the types of hematologic malignancies observed in Ezh2Δ/Δ mice (n = 12) in the secondary transplantation. Four recipient mice, which developed CD45.1+ T-ALL of a host cell origin, were excluded. (D) PB cell counts of moribund mice and those surviving at 370 days after secondary transplantation. WBC, Hb, and Plt counts in the PB of Ezh2Δ/Δ MDS, LPD, and T-ALL/lymphoma mice are plotted as dots and the mean values are indicated as bars. (E) Flow cytometric profiles of donor-derived CD45.2+ T-ALL/lymphoma cells in the PB and thymus obtained from representative mice with T-ALL/lymphoma (11F8_004, supplemental Table 2) at 52 weeks posttransplantation. Unstained cells served as a control for Tcrβ staining. (F) Gene expression profiling of Ezh2Δ/Δ T-ALL cells. RNA sequence was performed using Ezh2Δ/Δ BM CD8+ T-ALL cells obtained from a Ezh2Δ/Δ mouse with T-ALL and WT BM CD8+ T cells. GSEA was performed using the RNA-sequence data and GSEA plots are shown. The NES, NOM P value, and FDR q value are indicated. (G) Scatter diagrams showing RNA-sequence data. The signal levels of RefSeq genes (RPKM + 1 in log2) in Ezh2Δ/Δ BM CD8+ T-ALL cells and WT BM CD8+ T cells are plotted. The light gray lines represent the boundaries for twofold increase and twofold decrease, respectively. The number of genes upregulated and downregulated more than twofold in Ezh2Δ/Δ T-ALL cells compared with WT cells are indicated. The representative direct target genes of Notch1 are shown as red dots. FDR, false discovery rate; NES, normalized enrichment score; NOM, nominal.

Serial transplantation of Ezh2Δ/Δ hematopoietic cells caused heterogeneous hematologic malignancies. (A) Summary of secondary transplantation. A total of 2.5 × 106 BM cells from MDS and MDS/MPN mice ∼10 months after the deletion of Ezh2 were transplanted into secondary recipients irradiated at a lethal (8.5 Gy) or sublethal (6.5 Gy) dose. The engraftment status is indicated. (B) A Kaplan-Meier survival curve of WT (n = 4) and Ezh2Δ/Δ (n = 12) mice in secondary transplantation. Four recipient mice, which developed CD45.1+ T-ALL of a host cell origin, were excluded. *P < .05. Statistical significance was measured by log-rank test. (C) A pie graph showing the types of hematologic malignancies observed in Ezh2Δ/Δ mice (n = 12) in the secondary transplantation. Four recipient mice, which developed CD45.1+ T-ALL of a host cell origin, were excluded. (D) PB cell counts of moribund mice and those surviving at 370 days after secondary transplantation. WBC, Hb, and Plt counts in the PB of Ezh2Δ/Δ MDS, LPD, and T-ALL/lymphoma mice are plotted as dots and the mean values are indicated as bars. (E) Flow cytometric profiles of donor-derived CD45.2+ T-ALL/lymphoma cells in the PB and thymus obtained from representative mice with T-ALL/lymphoma (11F8_004, supplemental Table 2) at 52 weeks posttransplantation. Unstained cells served as a control for Tcrβ staining. (F) Gene expression profiling of Ezh2Δ/Δ T-ALL cells. RNA sequence was performed using Ezh2Δ/Δ BM CD8+ T-ALL cells obtained from a Ezh2Δ/Δ mouse with T-ALL and WT BM CD8+ T cells. GSEA was performed using the RNA-sequence data and GSEA plots are shown. The NES, NOM P value, and FDR q value are indicated. (G) Scatter diagrams showing RNA-sequence data. The signal levels of RefSeq genes (RPKM + 1 in log2) in Ezh2Δ/Δ BM CD8+ T-ALL cells and WT BM CD8+ T cells are plotted. The light gray lines represent the boundaries for twofold increase and twofold decrease, respectively. The number of genes upregulated and downregulated more than twofold in Ezh2Δ/Δ T-ALL cells compared with WT cells are indicated. The representative direct target genes of Notch1 are shown as red dots. FDR, false discovery rate; NES, normalized enrichment score; NOM, nominal.

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