Complete reversal of epithelial to mesenchymal transition requires inhibition of both ZEB expression and the Rho pathway.
Complete reversal of epithelial to mesenchymal transition requires inhibition of both ZEB expression and the Rho pathway.
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DOI：
10.1186/1471-2121-10-94
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发表时间：
2009-12-21
期刊：
影响因子：
--
通讯作者：
Mertz JE
中科院分区：
文献类型：
--
作者：
Das S;Becker BN;Hoffmann FM;Mertz JE
转化生长因子-β（TGF-β）诱导的上皮细胞向间充质细胞转化（EMT）是器官发生、癌症和器官纤维化中的重要细胞事件。逆转EMT的过程尚未建立。我们的目的是确定维持TGF-β诱导的间充质状态的信号通路和转录因子。评价了与EMT有关的5种激酶的抑制剂，TGF-β I型受体激酶（TβRI）、p38丝裂原活化蛋白激酶（p38 MAPK）、MAP激酶激酶/细胞外信号调节激酶激活剂激酶（MEK 1）、c-Jun NH末端激酶（JNK）和Rho激酶（ROCK），以逆转肾小管上皮细胞诱导的间充质状态。单药不能完全逆转EMT，如通过细胞形态学和基因表达所确定的。然而，暴露于TβRI抑制剂SB 431542，结合ROCK抑制剂Y27632，消除了可检测的肌动蛋白应力纤维和间充质基因表达，同时恢复上皮E-钙粘蛋白和肾特异性钙粘蛋白（Ksp-钙粘蛋白）表达。第二种组合，TβRI抑制剂SB 431542与p38 MAPK抑制剂SB 203580一起，在逆转EMT方面部分有效。此外，JNK抑制剂SP 600125可抑制TβRI抑制剂SB 431542逆转EMT的作用。为了探索EMT逆转的分子基础，我们还靶向了转录抑制因子ZEB 1和ZEB 2/SIP 1。用shRNA降低小鼠乳腺细胞中ZEB 1和ZEB 2的表达足以上调上皮蛋白如E-钙粘蛋白的表达并重建上皮特征。然而，皮质F-肌动蛋白的完全恢复需要与ROCK抑制剂Y27632结合ZEB 1/2敲低孵育。我们证明EMT的逆转需要通过TβRI和ROCK持续和独立的信号传导重建上皮转录和结构成分。这些发现表明，靶向多种激酶的组合小分子疗法可能是逆转疾病状况所必需的。
Epithelial to Mesenchymal Transition (EMT) induced by Transforming Growth Factor-β (TGF-β) is an important cellular event in organogenesis, cancer, and organ fibrosis. The process to reverse EMT is not well established. Our purpose is to define signaling pathways and transcription factors that maintain the TGF-β-induced mesenchymal state. Inhibitors of five kinases implicated in EMT, TGF-β Type I receptor kinase (TβRI), p38 mitogen-activated protein kinase (p38 MAPK), MAP kinase kinase/extracellular signal-regulated kinase activator kinase (MEK1), c-Jun NH-terminal kinase (JNK), and Rho kinase (ROCK), were evaluated for reversal of the mesenchymal state induced in renal tubular epithelial cells. Single agents did not fully reverse EMT as determined by cellular morphology and gene expression. However, exposure to the TβRI inhibitor SB431542, combined with the ROCK inhibitor Y27632, eliminated detectable actin stress fibers and mesenchymal gene expression while restoring epithelial E-cadherin and Kidney-specific cadherin (Ksp-cadherin) expression. A second combination, the TβRI inhibitor SB431542 together with the p38 MAPK inhibitor SB203580, was partially effective in reversing EMT. Furthermore, JNK inhibitor SP600125 inhibits the effectiveness of the TβRI inhibitor SB431542 to reverse EMT. To explore the molecular basis underlying EMT reversal, we also targeted the transcriptional repressors ZEB1 and ZEB2/SIP1. Decreasing ZEB1 and ZEB2 expression in mouse mammary gland cells with shRNAs was sufficient to up-regulate expression of epithelial proteins such as E-cadherin and to re-establish epithelial features. However, complete restoration of cortical F-actin required incubation with the ROCK inhibitor Y27632 in combination with ZEB1/2 knockdown. We demonstrate that reversal of EMT requires re-establishing both epithelial transcription and structural components by sustained and independent signaling through TβRI and ROCK. These findings indicate that combination small molecule therapy targeting multiple kinases may be necessary to reverse disease conditions.
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