Effect of natsudaidain isolated from Citrus plants on TNF-alpha and cyclooxygenase-2 expression in RBL-2H3 cells.
OBJECTIVE
Flavonoids inhibit the activity of chemical mediators released from mast cells. Our aim was to investigate the effects of natsudaidain, a polymethoxyflavone isolated from Citrus plants, on mast cells.


METHODS
We investigated the inhibitory effects of natsudaidain, which is a polymethoxyflavone isolated from Citrus plants, on histamine release, tumour necrosis factor-alpha production and cyclooxygenase-2 expression in Ca ionophore-stimulated rat basophilic leukemia cells (A23187-stimulated RBL-2H3 cells) by spectrofluorometric, ELISA and immunoblotting methods.


RESULTS
The percent of histamine release from A23187-stimulated RBL-2H3 cells pretreated with natsudaidain at 5, 25 and 50 microM was not changed as compared with non-treated A23187-stimulated cells. At 100 and 200 microM, natsudaidain pretreatment resulted in slightly reduced histamine release (% histamine release, 89.8+/-3.5% and 71.5+/-5.6% at 100 and 200 microM). Thus, natsudaidain hardly affects histamine release from RBL-2H3 cells, except at high concentrations. On the other hand, natsudaidain dose-dependently inhibited tumour necrosis factor-alpha protein and mRNA levels in A23187-stimulated RBL-2H3 cells; a concentration of 6.8 microM was required for a 50% reduction. In addition, all concentrations of this compound that we tested also inhibited cyclooxygenase-2 protein expression. The mRNA levels of cyclooxygenase-2 in A23187-stimulated RBL-2H3 cells treated with natsudaidain were also markedly decreased. The phosphorylated-p38 MAPK protein levels in A23187-stimulated RBL-2H3 cells treated with natsudaidain were lower than in the non-treated cells.


CONCLUSIONS
These findings suggest that natsudaidain inhibits tumour necrosis factor-alpha and cyclooxygenase-2 production by suppressing p38 MAPK phosphorylation but not p65 NFkappaB phosphorylation, and that natsudaidain might alleviate inflammatory diseases.
Matsui
Takuya
T
Faculty of Pharmacy, Meijo University, Tempaku-ku, Nagoya, Aichi, Japan. tmatsui@aichi-med-u.ac.jp
Ito
Chihiro
C
Itoigawa
Masataka
M
Okada
Tadashi
T
Furukawa
Hiroshi
H
D000704:Analysis of Variance; D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D045744:Cell Line, Tumor; D002851:Chromatography, High Pressure Liquid; D002957:Citrus / Q000737:chemistry*; D051546:Cyclooxygenase 2 / Q000235:genetics / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D047309:Flavones / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D005419:Flavonoids / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D006636:Histamine Release / Q000187:drug effects; D015151:Immunoblotting; D007476:Ionophores / Q000494:pharmacology; D015394:Molecular Structure; D010766:Phosphorylation / Q000187:drug effects; D018515:Plant Leaves / Q000737:chemistry; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism*; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism
D047309:Flavones; D005419:Flavonoids; D007476:Ionophores; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; C092904:natsudaidain; D000001:Calcimycin; C008661:nobiletin; D051546:Cyclooxygenase 2; D048051:p38 Mitogen-Activated Protein Kinases