Epigallocatechin-3-gallate-induced stress signals in HT-29 human colon adenocarcinoma cells.
Epigallocatechin-3-gallate (EGCG), a major component in green tea polyphenols, has been proven to suppress colonic tumorigenesis in animal models and epidemiological studies. As EGCG is retained in the gastrointestinal tract after oral administration, this pharmacokinetics property gives it the potential to function as a chemopreventive agent against colon cancer. In this study, human colorectal carcinoma HT-29 cells were treated with EGCG to examine the anti-proliferative and pro-apoptotic effects of EGCG, as well as the molecular mechanism underlying these effects. Cell viability assay, nuclear staining, DNA fragmentation, caspase assay, cytochrome c release, DiOC6(3) staining, mitogen-activated protein kinases (MAPK) phosphorylation and trypan blue exclusion assays, were utilized to dissect the signaling pathways induced by EGCG. After 36 h treatment, EGCG inhibited HT-29 cell growth with an IC50 of approximately 100 microM. HT-29 cells treated with doses higher than 100 microM showed apparent nuclear condensation and fragmentation, which was confirmed by DNA laddering. Caspase-3 and -9 activation was detected after 12 h treatment, accompanied by mitochondrial transmembrane potential transition and cytochrome c release. Activation of MAPKs was detected as early signaling event elicited by EGCG. Inhibition of c-Jun N-terminal kinase (JNK) pathway showed the involvement of JNK in EGCG-induced cytochrome c release and cell death. EGCG-induced JNK activation was blocked by the antioxidants glutathione and N-acetyl-l-cysteine, suggesting that the cell death signaling was potentially triggered by oxidative stress. In summary, our results from this study suggest that in HT-29 human colon cancer cells (i) EGCG treatment causes damage to mitochondria, and (ii) JNK mediates EGCG-induced apoptotic cell death.
Chen
Chi
C
Department of Pharmaceutics, Ernest-Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, NJ 08854, USA.
Shen
Guoxiang
G
Hebbar
Vidya
V
Hu
Rong
R
Owuor
Edward D
ED
Kong
A-N Tony
AN
D000230:Adenocarcinoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D016588:Anticarcinogenic Agents / Q000494:pharmacology*; D000975:Antioxidants / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D002374:Catalase / Q000494:pharmacology; D002392:Catechin / Q000031:analogs & derivatives* / Q000494:pharmacology*; D003110:Colonic Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D003574:Cytochrome c Group / Q000378:metabolism; D004273:DNA, Neoplasm / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016166:Free Radical Scavengers / Q000494:pharmacology; D005978:Glutathione / Q000494:pharmacology; D019073:HT29 Cells; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016877:Oxidants / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects*; D010766:Phosphorylation; D014568:Urokinase-Type Plasminogen Activator / Q000037:antagonists & inhibitors; D048051:p38 Mitogen-Activated Protein Kinases
D016588:Anticarcinogenic Agents; D000975:Antioxidants; D003574:Cytochrome c Group; D004273:DNA, Neoplasm; D004791:Enzyme Inhibitors; D016166:Free Radical Scavengers; D016877:Oxidants; D002392:Catechin; C045651:epigallocatechin gallate; D002374:Catalase; D048031:JNK Mitogen-Activated Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D014568:Urokinase-Type Plasminogen Activator; C505407:CASP3 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases; D005978:Glutathione