Dulos John, Kaptein Allard, Kavelaars Annemieke, Heijnen Cobi, Boots Annemieke
Department of Pharmacology, N.V. Organon, Oss, The Netherlands.
Arthritis Res Ther. 2005;7(6):R1271-80. doi: 10.1186/ar1819. Epub 2005 Sep 15.
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.
糖皮质激素已成功用于类风湿性关节炎的治疗。数据表明，脱氢表雄酮的一种免疫刺激代谢产物7α-羟基脱氢表雄酮（7α-OH-DHEA）可阻断糖皮质激素诱导的免疫抑制。7α-OH-DHEA的形成由细胞色素P450酶7b（Cyp7b）的活性催化。最近，我们报道肿瘤坏死因子（TNF）-α、IL-1α、IL-1β和IL-17可增强Cyp7b mRNA表达，并导致类风湿性关节炎患者的成纤维样滑膜细胞（FLS）形成7α-OH-DHEA的量随之增加。本研究的目的是阐明FLS中TNF-α介导的Cyp7b活性诱导涉及哪种信号转导途径。我们通过逆转录PCR测量Cyp7b mRNA表达以及测量7α-OH-DHEA的形成，研究了不同信号转导途径抑制剂对FLS中Cyp7b活性的影响。我们应用了SN50，一种转录因子（即活化蛋白-1 [AP-1]和核因子-κB [NF-κB]）核转位的抑制剂；PSI，一种蛋白酶体抑制剂，可防止IkappaB降解，从而阻止NF-κB释放；SP600125，一种c-Jun N端激酶（JNK）抑制剂；以及丝裂原活化蛋白激酶抑制剂PD98059（细胞外信号调节激酶）和SB203580（p38）。Cyp7b在类风湿性关节炎FLS中组成性表达，并且可响应TNF-α而被激活。SN50和PSI可阻止TNF-α诱导的Cyp7b活性增加，而丝裂原活化蛋白激酶抑制剂PD98059和SB203580则无作用。此外，SN50、PSI和SP600125可抑制Cyp7b mRNA表达和活性，这表明NF-κB和AP-1可诱导Cyp7b转录。这些发现表明NF-κB和AP-1参与了TNF-α增强的脱氢表雄酮代谢产物7α-OH-DHEA的形成。我们的结果与Cyp7b启动子中存在AP-1和NF-κB结合位点一致。
