Coculture with stroma sensitizes hematopoietic cells to ligand-induced apoptosis. We had shown previously that KG1a cells responded to TNF-α–induced apoptosis when cocultured with HS5 or HS27a stroma cells.²⁴  This is further illustrated in panel A. KG1a cells, cultured alone or in coculture with HS5, and treated with TNF-α (100 ng/mL) or camptothecin (10 μM), were labeled with annexin V, and apoptosis was determined by flow cytometry. Shown are representative tracings. (B) Cell surface expression of TNFR1 and TNFR2 as determined by flow cytometry was not altered by coculture (mean plus or minus SEM of 3 experiments). (C) Early apoptosis in KG1a cells occurred in a TNF-α dose-dependent fashion in cocultures, but not in KG1a cells cultured without stroma. Early apoptosis was also determined in CD34⁺ hematopoietic precursor cells collected from patients with MDS (D) or healthy volunteers (E) and cultured alone or in coculture with HS5 or HS27a cells. TNF-α was used at 100 ng/mL. After 16 to 20 hours, cells in cultures were labeled with anti-CD34 antibody (PE conjugated), and apoptosis was assessed with annexin V and PI as described in “Methods.” The experiment was repeated 4 times with independently isolated primary cells from different patients and healthy volunteers. Consistently, TNF-α induced apoptosis in MDS-derived CD34⁺ marrow cells cocultured with stroma, but not in CD34⁺ cells from healthy donors.