Involvement of p53 Mutation and Mismatch Repair Proteins Dysregulation in NNK-Induced Malignant Transformation of Human Bronchial Epithelial Cells
Figure 4
Occurrence of p53-R273H mutation in NHBE-NNK8 cells. (a) TP53 mutation detected by directing sequencing in NHBE and NHBE-NNK8 cells. Mutation at codon 273 in exon 8 was shown in electropherograms. Base changed from CGT to CAT (black arrow). (b) NHBE and NHBE-NNK8 cells were treated with 1 μM doxorubicin (Dox) or DMSO (d) vehicle control for the indicated times. p53 and p21 expression levels were assessed using Western blot analysis. GAPDH was used as loading control. (c) Quantitative data of (b) from three independent experiments (means ± SD). , , in comparison to NHBE cells treated with DMSO. (d) Quantitative RT-PCR analysis of p53 target genes, CDKN1A, P53R2, and PUMA, after treating NHBE and NHBE-NNK8 cells with 1 μM doxorubicin (Dox) for 8 hours. (e) Luciferase activity analysis of CDKN1A promoter after 1 μM doxorubicin (Dox) treatment. Luciferase activity is normalized for the transfection efficiency using cotransfection of pRL-SV40. Data are presented as means ± SD from three independent experiments. , , in comparison to NHBE cells treated with DMSO.
