Hepatocyte growth factor stimulates the migration of gastric epithelial cells by altering the subcellular localization of the tight junction protein ZO-1
Hepatocyte growth factor stimulates the migration of gastric epithelial cells by altering the subcellular localization of the tight junction protein ZO-1
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DOI：
10.1007/s00535-012-0615-y
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发表时间：
2013-02-01
影响因子：
6.3
通讯作者：
Tsubouchi, Hirohito
中科院分区：
文献类型：
--
作者：
Nasu, Yuichiro;Ido, Akio;Tsubouchi, Hirohito
肝细胞生长因子（HGF）是上皮细胞恢复所必需的，这是一个在胃肠道粘膜损伤后上皮细胞迅速迁移以覆盖脱落上皮的过程。本研究旨在通过分析HGF介导的胃上皮结构重建中紧密连接蛋白的表达和亚细胞动力学，探讨HGF介导的胃上皮结构重建的分子机制，以人胃上皮细胞MKN 74为研究对象，观察HGF对细胞迁移和增殖的影响以及紧密连接蛋白的表达和亚细胞动力学;同时，我们研究了HGF对大分子的细胞旁通透性的影响（使用异硫氰酸荧光素[FITC]-葡聚糖）HGF以剂量依赖的方式显著刺激MKN 74细胞的迁移，但不刺激其增殖。HGF不影响紧密连接蛋白的表达，包括claudin-1、claudin-3、claudin-4和claudin-7; occludin;和zonula occludens（ZO）-1。然而，荧光免疫染色显示，在细胞膜中，ZO-1的水平，但不是闭合蛋白或claudin-4的水平，在HGF治疗后1小时短暂下降。通过蛋白质印迹进一步证实了该结果：HGF减少了细胞膜组分中ZO-1蛋白的量，同时增加了细胞质中ZO-1的量。HGF可降低ZO-1与occludin之间的相互作用，诱导occludin的酪氨酸磷酸化，而ZO-1的磷酸化状态不受HGF的影响。尽管减少ZO-1/occludin相互作用，HGF并没有影响细胞旁的大分子通透性。HGF改变ZO-1的亚细胞定位，可能通过酪氨酸磷酸化的occludin，这可能会导致上皮细胞恢复过程中的分散。
Hepatocyte growth factor (HGF) is essential for epithelial restitution, a process in which epithelial cells rapidly migrate to cover desquamated epithelium after mucosal injury in the gastrointestinal tract. In this study, we aimed to elucidate the molecular mechanisms of the HGF-mediated reconstitution of gastric epithelial structures by analyzing the expression and subcellular dynamics of tight junction proteins.We treated human gastric epithelial MKN74 cells with HGF, and examined the effects of HGF on cell migration and proliferation, and the expression and subcellular dynamics of tight junction proteins; as well, we investigated the effect of HGF on paracellular permeability to macromolecules (using fluorescein isothiocyanate [FITC]-dextran).HGF significantly stimulated the migration of MKN74 cells, but not their proliferation, in a dose-dependent manner. HGF did not affect the expression of tight junction proteins, including claudin-1, -3, -4 and -7; occludin; and zonula occludens (ZO)-1. However, fluorescence immunostaining revealed that, in the cell membrane, the levels of ZO-1, but not those of occludin or claudin-4, were transiently decreased 1 h after HGF treatment. The results were further confirmed by western blotting: HGF reduced the amount of ZO-1 protein in the cell membrane fraction concomitantly with an increase in cytoplasmic ZO-1. Furthermore, HGF reduced the interaction between ZO-1 and occludin, and induced the tyrosine phosphorylation of occludin, whereas the phosphorylation status of ZO-1 was not affected by exposure to HGF. Despite a decrease in the ZO-1/occludin interaction, HGF did not affect paracellular permeability to macromolecules.HGF alters the subcellular localization of ZO-1, probably through the tyrosine phosphorylation of occludin, which may induce cell dispersion during epithelial restitution.