Fig. 3. Chronic TNFα exposure increases the CSC phenotype in HPV-immortalized oral keratinocytes.

a Levels of key stemness transcription factors, KLF4, Lin28, Nanog and Oct4, were measured by qPCR and normalized to the expression of GAPDH. *P < 0.01 compared to HOK-16B cells by two-tailed Student’s t test. b CD44 and CD24 stem cell surface markers were measured by flow cytometry analysis. Dots that fell in the upper left quadrant represent the CD44^high/CD24^low CSC population. c Migration ability was determined by transwell migration assay. Representative images of the transwell migration assay are shown on the right. *P < 0.01. The assay was performed in the absence of TNFα. d Chemosensitivity assay. Five hundred cells were seeded in 96-well plates and treated with 40 µmol·L⁻¹ cisplatin. At each incubation period, cell viability was measured using the MTT assay. *P < 0.05. e Radiosensitivity assay. Two hundred cells were seeded in 6-well plates and irradiated with different doses. After 10 days, surviving colonies were stained and counted. The assays were performed in the absence of TNFα.