The COX-2 selective inhibitor-independent COX-2 effect on colon carcinoma cells is associated with the Delta1/Notch1 pathway.
BACKGROUND
Cyclooxygenase-2 (COX-2) is a key factor in the development of colorectal cancer, and non-steroidal anti-inflammatory drugs (NSAIDs) have anti-colorectal cancer activity. However, the potential molecular mechanism of the COX-2 selective inhibitor effect on proliferation and apoptosis of colon cancer cells is unclear. In this study, we have demonstrated for the first time that the Delta1/Notch1 signal transduction pathway mediates the COX-2 selective inhibitor effect on colorectal cancer cells, and we reveal the mechanism of the Notch1 pathway in terms of regulating the proliferation and apoptosis of colorectal cancer cells.


RESULTS
Colon cancer cell lines HT-29 and SW480 were treated with NS-398 (a COX-2 selective inhibitor) and DAPT (a gamma-secretase inhibitor). The colormetric MTT cell proliferation assay and flow cytometry were used to measure cell proliferation and apoptosis. Reverse transcriptase (RT)-PCR and ELISA analyses were used to detect the levels of COX-2 mRNA expression and prostaglandin E2 (PGE2) concentration from the two cell lines, respectively. The expression of the Notch1, Delta1, ICN, Hes1 and NF-kappaB2 proteins was measured by Western blot. Immunohistochemistry results showed that Notch1 was expressed mainly in the cytoplasm and ICN mainly in the nucleus. COX-2 mRNA was highly expressed in HT-29 cells but not in SW480 cells. Both COX-2 mRNA expression and PGE2 concentration decreased in HT-29 cells treated with NS-398; however, PGE2 levels did not change in SW480 cells treated with NS-398. NS-398 and DAPT inhibited cell proliferation and induced apoptosis in a dose time-dependent manner accompanied by significantly decreased Notch1 activity (P < 0.01), and resulted in a significant down-regulation of Hes1 and NF-kappaB2 (P < 0.01).


CONCLUSIONS
Our results show that the selective COX-2 inhibitor may inhibit the proliferation and induce apoptosis in colon cancer cells through the COX-2-dependent pathway (HT-29) by decreasing the COX-2 mRNA/PGE2 levels and the activity of the COX-2-independent pathway (SW480). The Notch1 signal pathway mediates the effects of the COX-2 inhibitor on the proliferation and apoptosis of colon cancer cells. This may be a new target of the selective COX-2 inhibitor effect on colon cancer.
Zhang
Hui
H
Surgical Oncology Laboratory and Department of Gastroenterological Surgery, Peking University People's Hospital, Beijing, PR China.
Ye
Yingjiang
Y
Bai
Zhigang
Z
Wang
Shan
S
D053829:Amyloid Precursor Protein Secretases / Q000037:antagonists & inhibitors / Q000378:metabolism; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D051792:Basic Helix-Loop-Helix Proteins / Q000378:metabolism; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects; D003110:Colonic Neoplasms / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D003124:Colorimetry; D051546:Cyclooxygenase 2 / Q000235:genetics / Q000378:metabolism*; D052246:Cyclooxygenase 2 Inhibitors / Q000494:pharmacology*; D015232:Dinoprostone / Q000378:metabolism; D004151:Dipeptides / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D019073:HT29 Cells; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins / Q000378:metabolism*; D052003:NF-kappa B p52 Subunit / Q000378:metabolism; D009578:Nitrobenzenes / Q000494:pharmacology*; D051881:Receptor, Notch1 / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects*; D013449:Sulfonamides / Q000494:pharmacology*; D013997:Time Factors; D000072056:Transcription Factor HES-1
D053829:Amyloid Precursor Protein Secretases; D000970:Antineoplastic Agents; D051792:Basic Helix-Loop-Helix Proteins; D051546:Cyclooxygenase 2; D052246:Cyclooxygenase 2 Inhibitors; D015232:Dinoprostone; D004151:Dipeptides; D018398:Homeodomain Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D052003:NF-kappa B p52 Subunit; D009578:Nitrobenzenes; D051881:Receptor, Notch1; D013449:Sulfonamides; D000072056:Transcription Factor HES-1; C419410:N-(N-(3,5-difluorophenacetyl)alanyl)phenylglycine tert-butyl ester; C497761:NOTCH1 protein, human; C064045:delta protein; C080955:N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide; C078493:HES1 protein, human; C496540:PTGS2 protein, human