Tumor necrosis factor-alpha induces apoptosis associated with poly(ADP-ribose) polymerase cleavage in HT-29 colon cancer cells.
BACKGROUND
Tumor necrosis factor-alpha (TNF-alpha) is known for its selective cytotoxic activity on tumour cells. We analysed the response of HT-29 human colon carcinoma cells to this cytokine.


METHODS
After TNF-alpha treatment, cell proliferation, cell cycle, reactive oxygen species (ROS) production (flow cytometry), the amount of apoptotic cells (flow cytometry, fluorescence microscopy), cleavage of poly (ADP-ribose) polymerase (PARP) and caspase-3 activity (Western blotting) were detected.


RESULTS
TNF-alpha induced a decrease of cell growth and viability, an accumulation of cells in the S-phase of the cell cycle, an increase of subdiploid cell population and nuclear chromatin condensation and fragmentation, but not sooner than 96-120 hours. However, earlier events characteristic of apoptosis occurred, such as caspase-3 activation, PARP cleavage to 89 kDa fragment and changes in ROS production.


CONCLUSIONS
We demonstrated that, in addition to being an early marker of apoptosis, activation of caspase-3 and degradation of PARP may play a causative role in HT-29 cell death induced by TNF-alpha.
Vaculová
Alena
A
Laboratory of Cytokinetics, Institute of Biophysics, Academy of Sciences, Brno, Czech Republic.
Hofmanova
Jirina
J
Soucek
Karel
K
Kovariková
Martina
M
Kozubík
Alois
A
D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D019073:HT29 Cells / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D006801:Humans; D007700:Kinetics; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases