Figure 1.

VD₃ effects on proliferation of cultured hDPCs. (A): Proliferation of hDPCs cultured for 1–7 days (d1, d2, d3, d5, and d7) in Dulbecco's modified Eagle's medium (DMEM)/10% fetal bovine serum (FBS) supplemented with various concentrations of VD₃ (0, 1, 10, or 100 nM) (n = 3). *, Significant differences from VD₃-free (0 nM) medium (p < .05). (B): DNA synthesis (5-bromo-2′-deoxyuridine incorporation) compared from hDPCs cultured in DMEM/10% FBS in the presence of various concentration of VD₃ (0, 1, 10, or 100 nM) or in DMEM/SFM (n = 4). **, Significant differences from the control (VD₃-free [0 nM] serum-containing medium) (p < .01). (C): Representative images of immunocytochemical staining of cultured hDPCs with Hoechst 33342, Annexin V, and PI. Arrow and arrowhead show representative images of an apoptotic cell and a necrotic cell, respectively. Scale bars = 50 μm. (D): Percentage of Hoechst+/Annexin+/PI− (apoptotic) and Hoechst+/Annexin+/PI+ (necrotic) cells cultured in media with each VD₃ concentration. SFM showed a significantly higher apoptotic or necrotic cell number compared with VD₃-free medium. However, there were no significant differences among hDPCs cultured with different VD₃ doses (n = 4). *, Significant difference from VD₃-free medium (p < .05). Abbreviations: d, day; hDPC, human dermal papilla cell; PI, propidium iodide; SFM, serum-free medium; VD₃, 1α,25-dihydroxyvitamin D₃.