PI3Kγ is highly expressed in LSCs and promotes AML development. (A) mRNA levels of Pik3cg in normal mouse BM cells (Normal-BM), Lin−Sca-1+c-Kit+CD34+CD135− short-term HSCs (ST-HSCs), Lin−Sca-1+c-Kit+CD34−CD135− long-term HSCs (LT-HSCs), MLL-AF9+ leukemia cells (AML-BM), and Lin−CD127−Sca-1−c-Kit+CD34+CD16/32+ L-GMPs were measured by quantitative RT-PCR (n = 3). (B-C) Representative flow cytometric analysis of leukemia cells (GFP+) in the PB 4 weeks after primary transplantation (B) and quantification data (C, n = 5). (D-E) Lineage analysis with myeloid cell markers (Mac-1/Gr-1) was conducted in WT and Pik3cg-KO PB leukemia cells 4 weeks after primary transplantation (D) and quantification data (E, n = 5). (F-G) Representative images of the size of spleens and livers of recipients 4 weeks after primary transplantation (F) and quantification data (G, n = 5). (H) The overall survival of the recipient mice transplanted with WT or Pik3cg-KO MLL-AF9+ leukemia cells upon primary transplantation (n = 8). (I) Percentages of leukemia cells in the PB 4 weeks after secondary transplantation (n = 5). (J) The percentages of Mac-1+Gr-1− and Mac-1+Gr-1+ cells in the PB leukemia cells 4 weeks after secondary transplantation (n = 5). (K-L) Representative images of the size of spleens and livers of recipients 4 weeks after secondary transplantation (K) and quantification data (L, n = 5). (M) The overall survival of the recipient mice upon secondary transplantation (n = 10). (N-O) The frequencies of leukemia cells in the PB 4 weeks after transplantation (N, n = 5) and overall survival (O, n = 5) were compared among the recipients transplanted with WT, Pik3cg-overexpressing WT, Pik3cg-KO, and Pik3cg-overexpressing KO AML cells. Data are represented as mean ± standard error of the mean (SEM). Student 2-tailed unpaired t test for panels C,I, 1-way analysis of variance (ANOVA) with Tukey multiple comparison test for panels A,N, 2-way ANOVA with Sidak multiple comparison test for panels E,G,J,L, and log-rank test for panels H,M,O were used for the comparison of statistical significance. ∗P < .05; ∗∗P < .01; ∗∗∗P < .001.
Figure 1.

PI3Kγ is highly expressed in LSCs and promotes AML development. (A) mRNA levels of Pik3cg in normal mouse BM cells (Normal-BM), Lin−Sca-1+c-Kit+CD34+CD135− short-term HSCs (ST-HSCs), Lin−Sca-1+c-Kit+CD34−CD135− long-term HSCs (LT-HSCs), MLL-AF9+ leukemia cells (AML-BM), and Lin−CD127−Sca-1−c-Kit+CD34+CD16/32+ L-GMPs were measured by quantitative RT-PCR (n = 3). (B-C) Representative flow cytometric analysis of leukemia cells (GFP+) in the PB 4 weeks after primary transplantation (B) and quantification data (C, n = 5). (D-E) Lineage analysis with myeloid cell markers (Mac-1/Gr-1) was conducted in WT and Pik3cg-KO PB leukemia cells 4 weeks after primary transplantation (D) and quantification data (E, n = 5). (F-G) Representative images of the size of spleens and livers of recipients 4 weeks after primary transplantation (F) and quantification data (G, n = 5). (H) The overall survival of the recipient mice transplanted with WT or Pik3cg-KO MLL-AF9+ leukemia cells upon primary transplantation (n = 8). (I) Percentages of leukemia cells in the PB 4 weeks after secondary transplantation (n = 5). (J) The percentages of Mac-1+Gr-1− and Mac-1+Gr-1+ cells in the PB leukemia cells 4 weeks after secondary transplantation (n = 5). (K-L) Representative images of the size of spleens and livers of recipients 4 weeks after secondary transplantation (K) and quantification data (L, n = 5). (M) The overall survival of the recipient mice upon secondary transplantation (n = 10). (N-O) The frequencies of leukemia cells in the PB 4 weeks after transplantation (N, n = 5) and overall survival (O, n = 5) were compared among the recipients transplanted with WT, Pik3cg-overexpressing WT, Pik3cg-KO, and Pik3cg-overexpressing KO AML cells. Data are represented as mean ± standard error of the mean (SEM). Student 2-tailed unpaired t test for panels C,I, 1-way analysis of variance (ANOVA) with Tukey multiple comparison test for panels A,N, 2-way ANOVA with Sidak multiple comparison test for panels E,G,J,L, and log-rank test for panels H,M,O were used for the comparison of statistical significance. ∗P < .05; ∗∗P < .01; ∗∗∗P < .001.

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