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Selective inhibition of cyclooxygenase-2 enhances mitomycin-C-induced apoptosis
C T Hsueh1, C F Chiu, D P Kelsen
1Department of Internal Medicine, China Medical College Hospital, Taichung, Taiwan. d7198@hpd.cmch.org.tw
Purpose:
Cyclooxygenase-2 (COX-2) is involved in antiapoptosis signaling, and its induction may require activation of protein kinase C (PKC). Safingol (SAF), a PKC inhibitor, has been shown to enhance apoptosis induced by mitomycin-C (MMC) in human gastric cancer MKN-74 cells. The aim of this study was to identify the role of COX-2 in MMC-induced apoptosis in MKN-74 cells.
Methods:
Protein expression of COX-2 and Bcl-2 and activation of PKCalpha were examined by Western blot analysis. Apoptosis induction was examined by staining with bisbenzimide trihydrochloride (Hoechst-33258) of condensed chromatin, which characterizes the cells undergoing apoptosis. COX-2 mRNA levels were examined by Northern blot analysis.
Results:
After exposure for 1-2 h to 1 microg/ml MMC, upregulation of COX-2 and Bcl-2 protein expression was noted. The activation of PKCalpha occurred within 1 h of MMC exposure, and temporally preceded the induction of COX-2. Similar results were observed in cells exposed to the PKC activator, 3-phorbol 12-myristate 13-acetate. Cotreatment with SAF and MMC abolished the induction of COX-2 by MMC. Furthermore, NS-398, a selective COX-2 inhibitor, significantly enhanced MMC-induced apoptosis by fivefold from 4 +/- 2% (MMC alone) to 20 +/- 2% (MMC plus NS-398). There was no discernible change in COX-2 mRNA levels after a 2-h exposure to MMC but a twofold increase after a 24-h exposure.
Conclusions:
MMC upregulates COX-2 expression, which appears to be an antiapoptotic signal downstream of PKC. Selective inhibition of COX-2 can therefore provide a novel way to enhance MMC-induced apoptosis independent of inhibiting PKC.
Insights
Mitomycin-C upregulates cyclooxygenase-2 (COX-2) as an antiapoptotic signal in gastric cancer cells. Inhibiting COX-2 enhances apoptosis, offering a new therapeutic strategy.
Area of Science:
- Molecular Biology
- Cancer Research
- Cell Signaling
Background:
- Cyclooxygenase-2 (COX-2) plays a role in anti-apoptosis signaling pathways.
- Protein kinase C (PKC) activation may be necessary for COX-2 induction.
- Safingol (SAF), a PKC inhibitor, enhances apoptosis induced by mitomycin-C (MMC) in gastric cancer cells.
Purpose of the Study:
- To investigate the role of COX-2 in mitomycin-C (MMC)-induced apoptosis in human gastric cancer MKN-74 cells.
- To elucidate the relationship between PKC activation, COX-2 expression, and apoptosis.
Main Methods:
- Western blot analysis was used to assess protein expression of COX-2, Bcl-2, and PKCalpha activation.
- Apoptosis was quantified by Hoechst-33258 staining of condensed chromatin.
- Northern blot analysis was performed to examine COX-2 mRNA levels.
Main Results:
- MMC exposure led to increased COX-2 and Bcl-2 protein levels and PKCalpha activation.
- PKCalpha activation preceded COX-2 induction, and inhibition of PKC with SAF blocked COX-2 upregulation.
- Selective COX-2 inhibition with NS-398 significantly enhanced MMC-induced apoptosis by fivefold.
Conclusions:
- Mitomycin-C upregulates COX-2, which functions as an antiapoptotic signal downstream of PKC.
- Selective inhibition of COX-2 enhances MMC-induced apoptosis independently of PKC inhibition.
- Targeting COX-2 presents a novel strategy to augment apoptosis in gastric cancer treatment.