Fig. 3

Treatment with a combination of VD and TNFα effectively induces monocyte differentiation in HL60 cells. a NSE staining of HL60 cells treated with the vehicle (control group; Con), 50 nM VD (VD group; VD), and 50 nM VD with 5 ng/mL TNFα (VD-TNF group; VD-TNF). Note that NSE-reactivities in the VD-TNF group become negative by adding 40 mM fluoride (NaF) to the reaction medium, indicating that NSE-positive cells are possibly monocytes. b Frequencies of NSE-positive cells in the control, VD, and VD-TNF groups as determined from three independent experiments. Cells stained brown to dark brown or almost black were defined as NSE-positive cells. At random, three field images per smear sample, 500 cells per image, and thus 1500 cells per sample were counted in each group. NSE-positive cells were 46.4 ± 1.5% and 69.5 ± 2.7% in the VD and VD-TNF group, respectively, and are significantly increased compared to those in the control group (1.5 ± 0.4%). The frequency of NSE-positive cells in the VD-TNF group is 1.50-fold greater and statistically significant compared to that in the VD group. *P < 0.01