Alique M, Moreno V, Kitamura M, Xu Q, Lucio-Cazana F J
Department of Physiology, Faculty of Medicine, University of Alcala, Alcala de Henares, Madrid, Spain.
Br J Pharmacol. 2006 Sep;149(2):215-25. doi: 10.1038/sj.bjp.0706842. Epub 2006 Aug 7.
Preliminary results in human mesangial cells (MC) suggested that all-trans retinoic acid (ATRA) increased the expression of COX-2 and the production of prostaglandin E2 (PGE2), a PG with anti-inflammatory effects in MC. The aim of this work is to confirm that ATRA increases the expression of COX-2 in MC and to examine the mechanisms involved.
Cultured MC were treated with ATRA. COX expression and kinase activity were analyzed by Western blot. Transcriptional mechanisms were analyzed by Northern blot, RT-PCR and promoter assays.
COX-2 and COX-1 expression and PGE2 production were increased by ATRA. COX-2 played a role in PGE2 production as production was only partially inhibited by COX-1 inhibitor SC-560. COX-2 up-regulation by ATRA was due to transcriptional mechanisms as pre-incubation with actinomycin D abolished it and ATRA increased the expression of COX-2 mRNA and the activity of a human COX-2 promoter construct, whereas post-transcriptional mechanisms were not found. Retinoic acid receptors (RAR) were not involved in the up-regulation of COX-2 by ATRA since it was not inhibited by RAR-pan-antagonists and the RAR-pan-agonist TTNPB did not up-regulate COX-2. Instead ATRA might act through a sustained activation of extracellular signal-regulated kinase 1/2 (ERK1/2) since up-regulation of COX-2 was prevented by inhibition of the activation of ERK1/2 with PD098059. Also ERK1/2, as well as downstream signalling proteins from ERK1/2, remained phosphorylated when COX-2 increased 24 h later.
These results highlight the relevance of RAR-independent mechanisms to the biological effects of ATRA.
在人系膜细胞（MC）中的初步结果表明，全反式维甲酸（ATRA）可增加环氧化酶-2（COX-2）的表达以及前列腺素E2（PGE2）的生成，PGE2是一种在MC中具有抗炎作用的前列腺素。本研究的目的是证实ATRA可增加MC中COX-2的表达，并探讨其相关机制。
用ATRA处理培养的MC。通过蛋白质印迹法分析COX的表达和激酶活性。通过Northern印迹法、逆转录-聚合酶链反应（RT-PCR）和启动子分析来分析转录机制。
ATRA可增加COX-2和COX-1的表达以及PGE2的生成。COX-2在PGE2生成中发挥作用，因为COX-1抑制剂SC-560仅部分抑制了PGE2的生成。ATRA对COX-2的上调是由于转录机制，因为与放线菌素D预孵育可消除这种上调，并且ATRA可增加COX-2 mRNA的表达以及人COX-2启动子构建体的活性，而未发现转录后机制。维甲酸受体（RAR）不参与ATRA对COX-2的上调，因为它不受RAR泛拮抗剂的抑制，并且RAR泛激动剂TTNPB也不能上调COX-2。相反，ATRA可能通过细胞外信号调节激酶1/2（ERK1/2）的持续激活起作用，因为用PD098059抑制ERK1/2的激活可阻止COX-2的上调。此外，当24小时后COX-2增加时，ERK1/2以及ERK1/2的下游信号蛋白仍保持磷酸化状态。
这些结果突出了不依赖RAR的机制对ATRA生物学效应的相关性。
