Effect of cell-cell contact on HK protection.
(A) HK cells were plated in the lower chamber of transwell plates containing a porous membrane. CLL B cells (2 × 106 cells per milliliter) were plated in the upper chamber (no coculture). As a control, HK cells and CLL B cells were also cocultured together on the same side of filter (“coculture”). Simultaneously, B-CLL cells (2 × 106 cells per milliliter) were cultured with conditioned medium recovered from confluent HK cultures (“HK-soluble factors”). After 24 hours of culture, the CLL B cells were recovered, and the percentage of apoptosis was determined by annexin V–FITC/PI double staining, followed by flow cytometric analysis (n = 4). (B) Conditioned medium (cm) from cultures of HK or OVCAR3 cells was applied to cultures of CLL B cells at 50% (vol/vol), and the percentage of apoptotic cells was determined 1 day later as described above. As controls, CLL B cells were cultured in parallel without (“no coculture”) or with (“HK coculture”) HK cells before measurement of apoptosis percentages.
