PU.1 induces apoptosis in primary cHL cells from patients. (A) Morphology of purified, primary cHL cells transduced with control lentivirus (Case 1, 4 left panels) or PU.1 expressing lentivirus (Case 1, 4 right panels). A number of primary classic Hodgkin cells expressing PU.1 contained various-sized vacuoles Images were acquired using a BX60 microscope and DP70 digital camera with DP controller software (Olympus; ×1000 magnification). (B) Primary cHL cells transduced with PU.1 lentivirus tended to undergo apoptosis compared with cells transduced with empty vector. Apoptosis was assessed in Venus-positive cells by flow cytometry after staining with PI and allophycocyanin-conjugated annexin V. Stable transduction of primary cHL cells from case 1 with PU.1 lentivirus led to a decrease in the percentage of live PI⁻/annexin V⁻ cells and an increased percentage of PI⁻/annexin V⁺ preapoptotic cells, compared with control vector. Stable transduction of primary cHL cells from case 2 with PU.1 lentivirus also led to a decrease in the percentage of live PI⁻/annexin V⁻ cells and an increased percentage of PI⁺ apoptotic cells. Stable transduction of primary cHL cells from case 3 with PU.1 lentivirus also led to a decrease in the percentage of live PI⁻/annexin V⁺ cells and an increased percentage of PI⁺ apoptotic cells.