Indomethacin-induced apoptosis in esophageal adenocarcinoma cells involves upregulation of Bax and translocation of mitochondrial cytochrome C independent of COX-2 expression.
The prolonged use of nonsteroidal anti-inflammatory drugs (NSAIDs) has been shown to exert a chemopreventive effect in esophageal and other gastrointestinal tumors. The precise mechanism by which this occurs, however, is unknown. While the inhibition of COX-2 as a potential explanation for this chemopreventive effect has gained a great deal of support, there also exists evidence supporting the presence of cyclooxygenase-independent pathways through which NSAIDs may exert their effects. In this study, immunohistochemical analysis of 29 Barrett's epithelial samples and 60 esophageal adenocarcinomas demonstrated abundant expression of the COX-2 protein in Barrett's epithelium, but marked heterogeneity of expression in esophageal adenocarcinomas. The three esophageal adenocarcinoma cell lines, Flo-1, Bic-1, and Seg-1, also demonstrated varying expression patterns for COX-1 and COX-2. Indomethacin induced apoptosis in all three cell lines, however, in both a time- and dose-dependent manner. In Flo-1 cells, which expressed almost undetectable levels of COX-1 and COX-2, and in Seg-1, which expressed significant levels of COX-1 and COX-2, indomethacin caused upregulation of the pro-apoptotic protein Bax. The upregulation of Bax was accompanied by the translocation of mitochondrial cytochrome c to the cytoplasm, and activation of caspase 9. Pre-treatment of both cell lines with the specific caspase 9 inhibitor, z-LEHD-FMK, as well as the broad-spectrum caspase inhibitor, z-VAD-FMK, blocked the effect of indomethacin-induced apoptosis. These data demonstrate that induction of apoptosis by indomethacin in esophageal adenocarcinoma cells is associated with the upregulation of Bax expression and mitochondrial cytochrome c translocation, and does not correlate with the expression of COX-2. This may have important implications for identifying new therapeutic targets in this deadly disease.
Aggarwal
S
S
Department of Surgery, University of Michigan Medical School, Ann Arbor 48109, USA.
Taneja
N
N
Lin
L
L
Orringer
M B
MB
Rehemtulla
A
A
Beer
D G
DG
D000230:Adenocarcinoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D001471:Barrett Esophagus / Q000235:genetics / Q000473:pathology; D051546:Cyclooxygenase 2; D003574:Cytochrome c Group / Q000378:metabolism*; D004938:Esophageal Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D004947:Esophagus; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006801:Humans; D007213:Indomethacin / Q000494:pharmacology*; D007527:Isoenzymes / Q000235:genetics; D007700:Kinetics; D008565:Membrane Proteins; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D009092:Mucous Membrane / Q000378:metabolism / Q000473:pathology; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics*; D021381:Protein Transport / Q000187:drug effects; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D019253:Proto-Oncogene Proteins c-bcl-2; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051028:bcl-2-Associated X Protein
C494826:BAX protein, human; D003574:Cytochrome c Group; D007527:Isoenzymes; D008565:Membrane Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; D007213:Indomethacin