Transforming growth factor beta regulates cyclooxygenase-2 in glomerular mesangial cells.
This study examines the hypothesis that transforming growth factor beta (TGFbeta) regulates cyclooxygenase-2 (COX-2) and induces prostaglandin E synthase (mPGES-1) in rat mesangial cells. COX-2 expression was determined by Northern blot analysis after treatment with either TGFbeta1 or the selective COX-2 inhibitor, NS398. mPGES-1 expression was determined by real-time polymerase chain reaction. The effect of TGFbeta1 on COX-2 gene transcription was assessed using a luciferase reporter assay, and mRNA stability was also determined. To determine whether TGFbeta1 activates elements of the COX-2 promoter, we performed gel shift analyses to examine activation of activator protein-1 (AP-1) and nuclear factor kappaB (NF-kappaB). Prostaglandin E(2) (PGE(2)) and thromboxane B2 (TxB2) production was assayed by enzyme immunoassay. Finally, the pathophysiological relevance of COX-2 inhibition on the downstream effects of TGFbeta was assessed by examining collagen type I mRNA and net collagen production. COX-2 mRNA and mPGES-1 were induced after treatment with TGFbeta1 for 4 h, and this rise was accompanied by a three-fold increase in PGE(2) production that could be antagonized by selective inhibition of COX-2 with NS398. TGFbeta1 increased transcription by approximately 50% and activated both AP-1 and NF-kappaB. These effects were antagonized by co-treatment with NS398. Treatment with TGFbeta1 also doubled the half-life of COX-2 mRNA. Neither collagen type I mRNA nor net collagen production were altered by co-treatment with NS398. In conclusion, these results indicate that TGFbeta stimulates COX-2 and mPGES-1, with additional effects on transcription and stability of COX-2 mRNA.
Department of Pathology and Anatomy, Eastern Virginia Medical School, Norfolk, Virginia, USA. phardin1@hfhs.org
P
P
Harding
L
L
Balasubramanian
J
J
Swegan
A
A
Stevens
W F
WF
Glass
D000818:Animals; D024042:Collagen Type I / Q000235:genetics; D051546:Cyclooxygenase 2 / Q000187:drug effects / Q000235:genetics*; D052246:Cyclooxygenase 2 Inhibitors / Q000494:pharmacology; D015232:Dinoprostone / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007678:Kidney Glomerulus / Q000187:drug effects* / Q000201:enzymology; D050527:Mesangial Cells / Q000187:drug effects* / Q000201:enzymology; D016328:NF-kappa B / Q000378:metabolism; D009578:Nitrobenzenes / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000187:drug effects; D000071876:Prostaglandin-E Synthases; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics; D020871:RNA Stability; D012333:RNA, Messenger / Q000032:analysis / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013449:Sulfonamides / Q000494:pharmacology; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
D024042:Collagen Type I; D052246:Cyclooxygenase 2 Inhibitors; D016328:NF-kappa B; D009578:Nitrobenzenes; D012333:RNA, Messenger; D013449:Sulfonamides; D018808:Transcription Factor AP-1; D016212:Transforming Growth Factor beta; C080955:N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases; D000071876:Prostaglandin-E Synthases; C497818:Ptges protein, rat; D015232:Dinoprostone