Research Paper Volume 16, Issue 18 pp 12651—12666

Figure 2. Immortalized NHEK with terminated expression of SV40T showed phenotypes of cell senescence. (A) Immunofluorescence detection of SA-β-GAL in NHEKs and reversibly immortalized keratinocytes with PBS or Dox treatment. Scale bar = 20 μm. (B) Statistical analysis of (B) (****P < 0.0001, ns indicates not significant). (C) Flow cytometry analysis of SA-β-GAL in NHEKs and reversibly immortalized keratinocytes with PBS or Dox treatment. (D) mRNA levels of p16 and p21 were tested by qPCR with corresponding treatment (*P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, ns indicates not significant). (E) Protein levels of p16 and p21 were tested by Western blot in NHEKs and reversibly immortalized keratinocytes with corresponding treatment. (F) Immunofluorescent staining of γ-H2A.X, F-actin and DAPI of NHEKs and reversibly immortalized keratinocytes with PBS or Dox treatment. Scale bar = 20 μm.