Figure 2.

ZA inhibits CSC properties in OSCC cells. (A) The effect of ZA on the migration ability of OSCC cell lines (SCC4 and UM17B) was determined using Transwell chambers. Migration ability was described as the number of migrated cells per field, with data presented as mean ± standard deviation for 3 randomly selected fields. *P<0.01 vs. the untreated control (CTL) by t-test. Representative images of the migration assay are shown on the right. (B) The effect of ZA on the radiosensitivity of SCC4 cells was determined by clonogenic survival assay. The cells were irradiated (IR) with 5 Gy and cultured in the growth medium containing 0 (−) or 1 µM ZA (+) for 10 days, and the surviving colonies were stained and counted. (C) The effect of ZA on the chemosensitivity of SCC4 cells was determined by MTT assay. The cells were incubated in culture medium containing 40 µM etoposide (Eto) or 40 µM of methotrexate (Met), with or without ZA, for 4 days. Relative cell survival was expressed and normalized to untreated control. (D) The effect of ZA on CSC signaling pathways (Notch and Wnt) was determined by western blot analysis using NICD and β-catenin antibody. GAPDH antibody was used as loading control. Cells were treated with the indicated concentrations of ZA for 2 days and harvested for the assay. ZA, zoledronic acid; CSC, cancer stem-like cells; OSCC, oral/oropharyngeal squamous cell carcinoma; NICD, Notch intracellular domain.