NK Cell-Induced Tumor Cell Differentiation

Human NK cells were purified from healthy donors' PBMCs as described above. NK cells were left untreated or treated with a combination of IL-2 (1000 units/mL) and anti-CD16mAb (3 μg/mL), (a condition to induce split anergy in NK cells) for 18- 24 hours before the supernatants were removed and used in differentiation experiments. The amounts of IFN-γ produced by activated split-anergized NK cells were assessed with IFN-γ ELISA (Biolegend, San Diego, CA, USA). Differentiation of OSCSCs and MP2 cells was conducted with an average total of 1500-2000 pg and 4000-7000 pg of IFN-γ from IFN-γ containing supernatants, respectively, over 4-5 days as described previously 18 (link). Differentiation of A549 and A375 cells was conducted with an average total of 6000 pg and 15000 pg of IFN-γ containing supernatants, respectively over the course of 5 days. Differentiation of X02GB required 35000 pg of IFN-γ containing supernatants as described previously 57 . Initially, 1X10⁶ tumor cells were cultured and treated with split-anergized NK supernatants for differentiation. Afterward, target cells were rinsed with 1X PBS, detached and used for experiments.