Signal Transduction Pathways Mediating Neurotensin-stimulated Interleukin-8 Expression in Human Colonocytes*
Signal Transduction Pathways Mediating Neurotensin-stimulated Interleukin-8 Expression in Human Colonocytes*
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DOI：
10.1074/jbc.m104942200
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发表时间：
2001-11
期刊：
影响因子：
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通讯作者：
Dezheng Zhao;A. C. Keates;Sabina Kuhnt-Moore;M. Moyer;C. Kelly;C. Pothoulakis
中科院分区：
文献类型：
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作者：
Dezheng Zhao;A. C. Keates;Sabina Kuhnt-Moore;M. Moyer;C. Kelly;C. Pothoulakis
神经降压素(NT)是一种胃肠道对多种刺激做出反应而释放的神经肽，参与了结肠炎的病理生理过程。然而，介导这一促炎反应的分子机制(S)仍不清楚。我们发现，未转化的人类结肠细胞NCM460表达一种功能性的高亲和力NT受体，该受体介导NT诱导的ERK激活。通过使用稳定转染NTR1NCM460细胞，我们发现NTR1I激活导致了IL-8的分泌，这种分泌是通过依赖于NF-κB和ERK的途径来实现的。此外，NT刺激的NF-κB激活依赖于细胞内钙释放。NT刺激的Erk活性需要RAS的激活，因为显性负的RAS突变体RAS-17N的过表达几乎完全抑制了ERK的激活。此外，如RAS-GTP下拉实验所示，NT直接刺激RAS-GTP的形成。通过使用在IL-8启动子中含有靶向替换的报告基因构建，我们证明了在NT诱导的IL-8基因表达中，IL-8启动子区域的NF-κB和AP-1以及少量的C/EBP位点是IL-8基因表达所必需的。综上所述，我们的结果表明，NT刺激钙依赖的NF-κB和RAS依赖的ERK通路，介导未转化的人类结肠细胞释放IL-8。我们推测，这些与NT相关的促炎通路在结肠炎的病理生理学中起重要作用。
Neurotensin (NT), a neuropeptide released in the gastrointestinal tract in response to several stimuli, is involved in the pathophysiology of colonic inflammation. However, the molecular mechanism(s) mediating this proinflammatory response remains unclear. We found that NCM460, non-transformed human colonocytes, express a functional high affinity NT receptor that mediates NT-induced Erk activation. By using NCM460 cells stably transfected with NTR1, we show that NTR1 activation leads to interleukin (IL)-8 secretion that is mediated via both NF-κB- and Erk-dependent pathways. In addition, NT-stimulated NF-κB activation is dependent on intracellular calcium release. NT-stimulated Erk activity requires Ras activation because overexpression of the dominant negative Ras mutant Ras-17N almost completely inhibits the Erk activation. Furthermore, NT directly stimulates Ras-GTP formation as shown by a Ras-GTP pull-down assay. By using reporter gene constructs containing targeted substitutions in the IL-8 promoter, we show that the NF-κB, AP-1, and to a lesser degree the C/EBP sites in the IL-8 promoter region are required for IL-8 gene expression induced by NT. In summary, our results demonstrate that NT stimulates calcium-dependent NF-κB and Ras-dependent Erk pathways that mediate the release of IL-8 from non-transformed human colonocytes. We speculate that these NT-related proinflammatory pathways are important in the pathophysiology of colonic inflammation.