Metalloproteinases and transforming growth factor-α mediate substance P-induced mitogen-activated protein kinase activation and proliferation in human colonocytes
Metalloproteinases and transforming growth factor-α mediate substance P-induced mitogen-activated protein kinase activation and proliferation in human colonocytes
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DOI：
10.1074/jbc.m408523200
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发表时间：
2004-10-29
影响因子：
4.8
通讯作者：
Pothoulakis, C
中科院分区：
文献类型：
--
作者：
Koon, HW;Zhao, DZ;Pothoulakis, C
P物质（SP）通过与G蛋白偶联神经激肽-1受体（NK-1 R）结合并从结肠上皮细胞释放促炎细胞因子参与急性肠道炎症。SP还通过表皮生长因子（EGF）受体（EGFR）的反式激活和丝裂原活化蛋白激酶（MAPK）的激活刺激细胞增殖，这是慢性结肠炎期间组织愈合的关键事件。在这里，我们研究了SP诱导EGFR和MAPK激活的机制。我们使用稳定转染人NK-1 R的非转化人NCM 460集落细胞（NCM 460-NK-1 R细胞）以及表达高水平内源性NK-1 R的未转染U373 MG细胞。将两种细胞系暴露于SP（10（-7）M）刺激EGFR活化（1分钟），随后是细胞外信号调节蛋白激酶（ERK 1/2）活化（2-5分钟）。SP诱导的ERK 1/2激活通过用金属蛋白酶抑制剂Batimastat/GM 6001、EGFR磷酸化抑制剂AG 1478和肿瘤坏死因子-α-转化酶（TACE）抑制剂TAPI-1预处理来阻断。用针对潜在EGFR配体的抗体预处理表明，转化生长因子-α（TGF α），而不是其他EGFR配体EGF，肝素结合EGF或双调蛋白，介导SP诱导的EGFR反式激活。SP刺激TGF α释放到细胞外间隙，在2分钟内可测量，这种释放被金属蛋白酶抑制剂和TACE抑制剂TAPI-1抑制。SP还诱导MAPK介导的细胞增殖，其被TACE、基质金属蛋白酶（MMP）、EGFR和MEK 1抑制剂抑制。因此，在人类结肠细胞中，NK-1 R诱导的EGFR和MAPK活化和细胞增殖涉及基质金属蛋白酶（最有可能是TACE）和TGF α的释放。这些信号传导机制可能参与了NK-1 R在慢性结肠炎中的保护作用。
Substance P (SP) participates in acute intestinal inflammation via binding to the G-protein-coupled neurokinin-1 receptor (NK-1R) and release of proinflammatory cytokines from colonic epithelial cells. SP also stimulates cell proliferation, a critical event in tissue healing during chronic colitis, via transactivation of the epidermal growth factor (EGF) receptor (EGFR) and activation of mitogen-activated protein kinase (MAPK). Here we examined the mechanism by which SP induces EGFR and MAPK activation. We used non-transformed human NCM460 colonocytes stably transfected with the human NK-1R (NCM460-NK-1R cells) as well as untransfected U373 MG cells expressing high levels of endogenous NK-1R. Exposure of both cell lines to SP (10(-7) M) stimulated EGFR activation (1 min) followed by extracellular signal-regulated protein kinase (ERK1/2) activation (2-5 min). SP-induced ERK1/2 activation was blocked by pretreatment with the metalloproteinase inhibitor Batimastat/GM6001, the EGFR phosphorylation inhibitor AG1478, and the tumor necrosis factor-alpha-converting enzyme (TACE) inhibitor TAPI-1. Pretreatment with antibodies against potential EGFR ligands suggested that transforming growth factor-alpha (TGFalpha), but not the other EGFR ligands EGF, heparin-binding EGF, or amphiregulin, mediates SP-induced EGFR transactivation. SP stimulated TGFalpha release into the extracellular space that was measurable within 2 min, and this release was inhibited by metalloproteinase inhibitors and the TACE inhibitor TAPI-1. SP also induced MAPK-mediated cell proliferation that was inhibited by TACE, matrix metalloproteinase (MMP), EGFR, and MEK1 inhibitors. Thus, in human colonocytes, NK-1R-induced EGFR and MAPK activation and cell proliferation involve matrix metalloproteinases (most likely TACE) and the release of TGFalpha. These signaling mechanisms may be involved in the protective effects of NK-1R in chronic colitis.