Hordenine Effects on Dermal Papilla Cells

DPCs were seeded in a 24-well plate at a density of 3 × 10⁴ and cultured for 24 h. Then, Hordenine at the concentration of 0, 25 and 50 µmol/L was added to treat DPCs for 24 h. Then, cells were washed with PBS for three times, fixed in 4% paraformaldehyde for 15 min, permeabilized with 0.1% Triton X-100, and blocked with 1% FBS for 0.5 h. The cells were then incubated overnight at 4 °C with the specific primary antibodies, followed by incubation for 2 h with fluorescent secondary antibody and staining with DAPI for 5 min. Photographing were conducted under an Olympus microscope. All the primary antibodies used in immunofluorescence staining were as follows: 1:200 anti-Ki67 antibody (Abcam, 16667, Cambridge, UK), 1:200 anti-β-catenin antibody (Proteintech, 51067-2-AP, Chicago, IL, USA), 1:100 anti-ALP antibody (Abcam, 65834, Cambridge, UK), 1:100 anti-Versican antibody (Abcam, 19345, Cambridge, UK).