Constitutive expression of prostaglandin endoperoxide G/H synthetase (PGHS)-2 but not PGHS-1 in hum an tracheal epithelial cells in vitro.
Primary cultures of human tracheal epithelial (HTE) cells cultured in vitro, in defined serum-free media, express prostaglandin endoperoxide G/H synthase (PGHS) activity and produce prostaglandin E2 (PGE2). In contrast to every other cell type studied to date, HTE cells appear to constitutively express PGHS-2, the 'inducible' form of the enzyme, while expressing little or no PGHS-1, the 'housekeeping' isoenzyme in vitro. Prostaglandin synthesis in HTE cells was reduced by a selective PGHS-2 inhibitor, N-[2-cyclohexyloyl-4-nitrophenyl] methane-sulfonamide (NS398), with an IC50 of approximately 1 microM. Immunoblotting and immunoprecipitation of enzymatic activity with isozyme-specific antisera revealed only the PGHS-2 isoform. Full length human cDNA probes detected only PGHS-2 message in Northern blots. Neither PGHS-2 activity nor mRNA levels were dependent on, nor stimulated by peptide growth factors present in the defined serum-free growth medium, or by serum. Prolonged maintenance in the absence of retinoic acid, however, lead to a decline in PGHS activity. Phorbol-myristate acetate (PMA) induced PGHS-2 activity and mRNA and neither PMA-induced, nor constitutive PGHS-2 expression was suppressed by corticosteroids. Actinomycin D-treatment for six hours reduced the PGHS-2 activity and mRNA to only 50% that of untreated cells, suggesting that PGHS-2 mRNA is extremely stable in these cells. HTE cells, at least in vitro, appear unique among prostaglandin-producing cells in that they express PGHS-2, constitutively, independent of regulation by growth factors, serum, or corticosteroids and fail to express PGHS-1 under any culture condition studied.
Walenga
R W
RW
Department of Pediatrics, Case Western Reserve University, Cleveland, OH 44106, USA. rxw13@po.cwru.edu
Kester
M
M
Coroneos
E
E
Butcher
S
S
Dwivedi
R
R
Statt
C
C
D000305:Adrenal Cortex Hormones / Q000494:pharmacology; D015152:Blotting, Northern; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free; D003513:Cycloheximide / Q000494:pharmacology; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells; D004848:Epithelium / Q000187:drug effects / Q000378:metabolism; D006133:Growth Substances / Q000494:pharmacology; D006801:Humans; D015151:Immunoblotting; D007213:Indomethacin / Q000494:pharmacology; D007527:Isoenzymes / Q000096:biosynthesis* / Q000187:drug effects* / Q000502:physiology; D008565:Membrane Proteins; D009578:Nitrobenzenes / Q000494:pharmacology; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis* / Q000187:drug effects* / Q000502:physiology; D012333:RNA, Messenger / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014132:Trachea / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*
D000305:Adrenal Cortex Hormones; D016895:Culture Media, Serum-Free; D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors; D006133:Growth Substances; D007527:Isoenzymes; D008565:Membrane Proteins; D009578:Nitrobenzenes; D012333:RNA, Messenger; D013449:Sulfonamides; C080955:N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide; D003513:Cycloheximide; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; C496511:PTGS1 protein, human; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; D013755:Tetradecanoylphorbol Acetate; D007213:Indomethacin