Skip to main content
. 2020 Jun 23;11(25):2387–2403. doi: 10.18632/oncotarget.27630

Figure 2. BI-D1870 treatment induces mitotic arrest and apoptosis in HL60 cells.

Figure 2

(A) Cell cycle profiles of control and BI-D1870-treated HL60 cells by flow cytometry. HL60 cells were treated with BI-D1870 (5 μM). Cells were stained with 4,6-diamidino-2-phenylindole (DAPI) and antibodies. Bivariate distribution of DNA content versus the level of phosphorylated Histone H3 (p-H3) was detected by multi-parameter flow cytometry. Percentages of cell populations at each cell cycle phase determined by DNA content (DAPI) and levels of p-H3, Cyclin A and Cyclin B were graphed. (B) Protein levels of G2/M phase markers (p-Rb (S780), p-CDC2 (Y15), Cyclin B and Cyclin A), apoptosis marker (Cleaved Caspase 3) and phosphorylated Ribosomal protein S6 (p-RPS6 (S235/236)) in BI-D1870-treated HL60 cells. Cell extracts were prepared at the indicated times after BI-D1870 treatment. β-Actin was used as an internal control. (C) Accumulation of metaphase cells following the treatment of BI-D1870. After treatment with BI-D1870 (5 μM) for the indicated times, HL60 cells were fixed and stained with DAPI and antibodies. Mitotic phases were further characterized in p-H3-positive populations by measuring the levels of Cyclin A and Cyclin B. Data represent the percentages of cell populations residing at each mitotic phase analyzed by the levels of Cyclin A and Cyclin B in the mitotic population. (D) Induction of mitotic arrest in KG1 cells by BI-D1870 treatment. KG1 cells were treated with BI-D1870 (5 μM) for indicated times. Cells were fixed and stained with DAPI and antibodies against p-H3, Cyclin A, and Cyclin B. Cell cycle profiles of DMSO or compound-treated KG1 cells were shown as the bivariate distribution of DNA content versus the level of phosphorylated Histone H3 (top). Each mitotic phase distribution was identified as the cellular expression of Cyclin A and Cyclin B in mitotic cells (bottom). The percentage cell population at each cell cycle stage is shown. Flow cytometric profiles represent one out of three experiments with similar results. Data are graphed as mean ± SEM (n = 3). * p < 0.05; ** p < 0.01; *** p < 0.001.