Figure 7

Pan-SHIP inhibitor 1PIE causes cell cycle arrest and induces apoptosis in MM cells. (A) MM cell lines were cultured in the presence or absence of 10 µmol/L 1PIE for 48 h and 10 µmol/L BrdU for 14 h. Cells were subsequently labeled with APC-anti-BrdU and 7AAD, and cell cycle status was analyzed by flow cytometry A representative example of two independent experiments is shown. ▩, sub G0/G1; ▦ G0/G1; ▤, G2/M; ▥, S. (B) MM cells were treated with increasing concentrations of 1PIE for 48 h and Annexin V-positive cells were determined by FACS analysis. Results represent the mean increase in apoptotic cells in two independent experiments. ▤, U266; ▩, RPMI8226; ▦, OPM2. (C) MM cells were treated for 48 h with increasing concentrations of 1PIE. After cell lysis, proteins were separated by SDS-PAGE, and blots were probed with antibodies against cleaved caspase 3, cleaved caspase 9, cleaved LC3B-II and cleaved PARP. Equal loading was confirmed with antibodies against actin. conc, concentration.