Tumor necrosis factor alpha induces p53 up-regulated modulator of apoptosis expression in colorectal cancer cell lines.
OBJECTIVE
Inflammatory bowel disease (IBD)-associated colorectal carcinogenesis involves dysregulation of multiple cellular pathways, including p53 signaling and cytokine action. The purpose of the current study was to evaluate the effects of tumor necrosis factor alpha (TNF-alpha) on p53 and p53 up-regulated modulator of apoptosis (PUMA), a downstream effector of p53 in the apoptotic pathway in colorectal cancer cells.


METHODS
The cell lines HT29 (which express mutant p53) and HCT116 (which express wild-type p53) were treated with TNF-alpha (0, 50, 100, or 500 ng/mL) for 1, 12, 24, or 48 hours. Protein expression and subcellular localization of p53 and PUMA were determined by immunoblot and immunofluorescence. Changes in p53 and PUMA mRNA expression were determined by quantitative real time polymerase chain reaction.


RESULTS
Nuclear p53 expression was increased in TNF-alpha-treated HT29 cells; in contrast, expression was decreased or minimally changed in TNF-alpha-treated HCT116 cells, as determined by immunoblot and immunofluorescence. At 24 hours, p53 mRNA transcript levels were minimally increased in HT29 cells, whereas PUMA increased 34-fold.


CONCLUSIONS
TNF-alpha increased nuclear p53 expression in HT29 cells, which express p53 mutation, but not in HCT116 cells, which are wild type for p53. In addition, TNF-alpha markedly up-regulated PUMA mRNA levels in HT29 cells. Our findings suggest that TNF-alpha may be a factor in carcinogenesis in IBD in cells carrying a p53 mutation.
Pastor
Danielle M
DM
Department of Surgery, Penn State Milton S. Hershey Medical Center and College of Medicine, Hershey, Pennsylvania, USA. dpastor@psu.edu
Irby
Rosalyn B
RB
Poritz
Lisa S
LS
D000704:Analysis of Variance; D017209:Apoptosis / Q000187:drug effects*; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism*; D045744:Cell Line, Tumor; D015179:Colorectal Neoplasms / Q000188:drug therapy* / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006801:Humans; D015151:Immunoblotting; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism / Q000494:pharmacology*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D015854:Up-Regulation
D051017:Apoptosis Regulatory Proteins; D014409:Tumor Necrosis Factor-alpha; D016159:Tumor Suppressor Protein p53