Correlation of MAP kinases with COX-2 induction differs between MKN45 and HT29 cells.
BACKGROUND
Mitogen-activated protein (MAP) kinases, including extracellular signal-regulated kinases (ERK),c-Jun NH2-terminal kinases (JNK) and p38 MAP kinase (p38 MAPK) are important intermediates of the signal-transduction pathway from the cell surface to the nucleus. Expression of cyclooxygenase (COX)-2, associated with proliferation, apoptosis or both of gastrointestinal cancer cells, is mediated through MAP kinase families. However, the correlation between respective MAP kinase signals and COX-2 in the proliferation of gastric and colon cancer cells has not been well elucidated.


OBJECTIVE
We examined the effect of selective inhibitors of MAP kinases and COX-2 on serum-induced proliferation of gastric (MKN45) and colon (HT29) cancer cells.


METHODS
After 24-h serum starvation, cancer cells were stimulated with 2% serum and COX-2 inhibitors (NS398 10 micromol/L, or etodolac 100 micromol/L) or 1 h after preincubation with inhibitors for ERK (PD98059 20 micromol/L) or p38 MAPK (SB203580 10 micromol/L). Phosphorylated MAP kinases and COX-2 protein were evaluated by Western blotting, and the proliferation of cancer cells was estimated by 3H-thymidine incorporation. Transcription factors nuclear factor-kappaB and CREB were assayed by an electorophoretic mobility shift assay.


RESULTS
Serum increased the proliferation of MKN45 and HT29 cells by 280% and 200%, respectively, compared with the control levels (100%). In both cancer cells, phosphorylated MAP kinases were increased within 30 min after stimulation. PD98059 and SB203580 inhibited the serum-induced proliferation of MKN45 by 21% and 51% and of HT29 by 81% and 69%, respectively. NS398 and etodolac inhibited the proliferation of HT29 by 21% and 41%, respectively, but not that of MKN45. PD98059 and SB203580 also suppressed serum-induced expression of COX-2 protein in HT29 cells. In addition to the activation of MAP kinases and COX-2, activities of nuclear factor-kappaB and CREB were also increased during HT29 cell proliferation.


CONCLUSIONS
These results suggest that the correlation of MAP kinases with COX-2 induction for cell proliferation differs between MKN45 and HT29 cells.
Tominaga
K
K
Department of Gastroenterology, Osaka City University, Graduate School of Medicine, Osaka, Japan. tomy@med.osaka-cu.ac.jp
Higuchi
K
K
Sasaki
E
E
Suto
R
R
Watanabe
T
T
Fujiwara
Y
Y
Oshitani
N
N
Matsumoto
T
T
Kim
S
S
Iwao
H
H
Arakawa
T
T
D015153:Blotting, Western; D002455:Cell Division; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000201:enzymology*; D051546:Cyclooxygenase 2; D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D019073:HT29 Cells; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008565:Membrane Proteins; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism*; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism*; D011725:Pyridines / Q000494:pharmacology; D044967:Serum / Q000502:physiology; D013274:Stomach Neoplasms / Q000201:enzymology*
D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors; D004791:Enzyme Inhibitors; D005419:Flavonoids; D007093:Imidazoles; D007527:Isoenzymes; D008565:Membrane Proteins; D011725:Pyridines; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; D020929:Mitogen-Activated Protein Kinase Kinases; C093642:SB 203580; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one