Tumour necrosis factor-alpha stimulates dehydroepiandrosterone metabolism in human fibroblast-like synoviocytes: a role for nuclear factor-kappaB and activator protein-1 in the regulation of expression of cytochrome p450 enzyme 7b.
Glucocorticoids have successfully been used in the treatment of rheumatoid arthritis. Data suggest that 7alpha-hydroxy-dehydroepiandrosterone (7alpha-OH-DHEA), an immunostimulating metabolite of dehydroepiandrosterone, can block glucocorticoid-induced immune suppression. Formation of 7alpha-OH-DHEA is catalyzed by activity of cytochrome p450 enzyme 7b (Cyp7b). Recently, we reported that tumour necrosis factor (TNF)-alpha, IL-1alpha, IL-1beta and IL-17 enhance Cyp7b mRNA expression and induce a concomitant increase in the formation of 7alpha-OH-DHEA by fibroblast-like synoviocytes (FLS) from rheumatoid arthritis patients. The aim of this study was to elucidate which signal transduction pathway is involved in the TNF-alpha-mediated induction of Cyp7b activity in FLS. We studied the effects of inhibitors of different signal transduction pathways on Cyp7b activity in FLS by measuring Cyp7b mRNA expression using reverse transcription PCR and by measuring the formation of 7alpha-OH-DHEA. We applied SN50, an inhibitor of nuclear translocation of transcription factors (i.e. activator protein-1 [AP-1] and nuclear factor-kappaB [NF-kappaB]); PSI, a proteasome inhibitor that prevents IkappaB degradation and thereby NF-kappaB release; SP600125, a c-Jun N-terminal kinase (JNK) inhibitor; and the mitogen-activated protein kinase inhibitors PD98059 (extracellular signal-regulated kinase) and SB203580 (p38). Cyp7b is constitutively expressed in RA FLS and can be activated in response to TNF-alpha. SN50 and PSI prevented the TNF-alpha-induced increase in Cyp7b activity, whereas the mitogen-activated protein kinase inhibitors PD98059 and SB203580 had no effect. In addition, inhibition of Cyp7b mRNA expression and activity was observed with SN50, PSI and SP600125, suggesting that NF-kappaB and AP-1 induce Cyp7b transcription. These findings suggest that NF-kappaB and AP-1 are involved in the TNF-alpha-enhanced formation of the dehydroepiandrosterone metabolite 7alpha-OH-DHEA. Our results are in accordance with presence of AP-1 and NF-kappaB binding sites in the Cyp7b promoter.
Dulos
John
J
Department of Pharmacology, N.V. Organon, Oss, The Netherlands. john.dulos@organon.com
Kaptein
Allard
A
Kavelaars
Annemieke
A
Heijnen
Cobi
C
Boots
Annemieke
A
D001172:Arthritis, Rheumatoid / Q000378:metabolism*; D001483:Base Sequence; D002460:Cell Line; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D000072474:Cytochrome P450 Family 7; D003687:Dehydroepiandrosterone / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006801:Humans; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000378:metabolism*; D009842:Oligopeptides / Q000494:pharmacology; D010455:Peptides / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013250:Steroid Hydroxylases / Q000235:genetics / Q000378:metabolism*; D013583:Synovial Membrane / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D016328:NF-kappa B; D009842:Oligopeptides; D010455:Peptides; D012333:RNA, Messenger; C118258:SN50 peptide; D018808:Transcription Factor AP-1; D014409:Tumor Necrosis Factor-alpha; C090535:benzyloxycarbonyl-isoleucyl-glutamyl(O-tert-butyl)-alanyl-leucinal; D003687:Dehydroepiandrosterone; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; D000072474:Cytochrome P450 Family 7; C499571:CYP7B1 protein, human