Bradykinin stimulates protein kinase D-mediated colonic myofibroblast migration via cyclooxygenase-2 and heat shock protein 27.
Bradykinin stimulates protein kinase D-mediated colonic myofibroblast migration via cyclooxygenase-2 and heat shock protein 27.
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DOI：
10.1016/j.jss.2016.10.014
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发表时间：
2017-03
期刊：
影响因子：
--
通讯作者：
Yoo J
中科院分区：
文献类型：
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作者：
Chu E;Saini S;Liu T;Yoo J
炎症性肠病（IBD）的特征是间歇性肠道损伤和修复。肌成纤维细胞是调节修复过程的胃肠道（GI）基质细胞，并且是包括缓激肽（BK）在内的炎症介质的已知靶点。然而，炎症调节肌纤维母细胞诱导的伤口愈合的机制仍不完全清楚。在这里，我们证明，第一次，BK刺激肌成纤维细胞迁移通过蛋白激酶D（PKD）介导的激活的考克斯-2和HSP 27途径。CCD-18 Co是从第8-14代使用的人结肠肌成纤维细胞系。体外划痕试验评估了在存在或不存在几种抑制剂（CID 755673（10 µM）和NS 398（10 µM））的情况下，BK（100 nM）对肌成纤维细胞迁移的影响。Hsp 27 siRNA评价Hsp 27对结肠肌成纤维细胞迁移的作用。pPKD、pHsp 27和考克斯-2的抗体通过Western印迹评估表达水平。BK刺激肌成纤维细胞迁移超过24小时。BK还导致PKD在Ser-916处的快速和持续磷酸化，HSP 27在Ser-82处的快速磷酸化，并在4小时内增加考克斯-2表达。BK介导的考克斯-2表达和Hsp 27磷酸化均被PKD抑制剂CID 755673抑制。类似地，BK诱导的肌成纤维细胞迁移被CID 755673（p<0.05）、直接考克斯-2抑制剂NS 398（p<0.05）和Hsp 27 siRNA（p<0.05）显著抑制。BK通过PKD介导的考克斯-2和Hsp 27的活化刺激肌成纤维细胞迁移。PKD、考克斯-2和HSP 27似乎都调节肌成纤维细胞迁移，肌成纤维细胞迁移是一种可能在炎症环境中的粘膜愈合中发挥重要作用的基质群体。
Inflammatory bowel disease (IBD) is characterized by episodic intestinal injury and repair. Myofibroblasts are gastrointestinal (GI) tract stromal cells that regulate the reparative process, and are known targets of inflammatory mediators including bradykinin (BK). However, the mechanisms through which inflammation regulates myofibroblast-induced wound healing remain incompletely understood. Here, we demonstrate, for the first time, that BK stimulates myofibroblast migration through protein kinase D (PKD)-mediated activation of the COX-2 and Hsp27 pathways. CCD-18Co is a human colonic myofibroblast cell line used from passages 8–14. An in vitro scratch assay assessed the effect of BK (100nM) on myofibroblast migration over 24h in the presence or absence of several inhibitors (CID755673 (10 µM) and NS398 (10 µM)). Hsp27 siRNA evaluated the effect of Hsp27 on colonic myofibroblast migration. Antibodies to pPKD, pHsp27, and COX-2 evaluated expression levels by Western blot. BK stimulated myofibroblast migration over 24h. BK also led to rapid and sustained phosphorylation of PKD at Ser-916, rapid phosphorylation of Hsp27 at Ser-82, and increased COX-2 expression over 4h. BK-mediated COX-2 expression and Hsp27 phosphorylation were both inhibited by the PKD inhibitor CID755673. Similarly, BK-induced myofibroblast migration was significantly inhibited by CID755673 (p<0.05), by the direct COX-2 inhibitor NS398 (p<0.05), and by Hsp27 siRNA (p<0.05). BK stimulates myofibroblast migration through PKD-mediated activation of COX-2 and Hsp27. PKD, COX-2 and Hsp27 all appear to regulate myofibroblast cell migration, a stromal population that may play an important role in mucosal healing in the setting of inflammation.