Effect of a pore-forming protein derived from Flammulina velutipes on the Caco-2 intestinal epithelial cell monolayer
Effect of a pore-forming protein derived from Flammulina velutipes on the Caco-2 intestinal epithelial cell monolayer
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DOI：
10.1271/bbb.68.2230
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发表时间：
2004-11-01
影响因子：
1.6
通讯作者：
Shimizu, M
中科院分区：
文献类型：
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作者：
Narai, A;Watanabe, H;Shimizu, M
我们之前发现了一种从金针菇中提取的经上皮电阻（TEER）降低蛋白，该蛋白与被称为溶血性孔形成蛋白的金针菇毒素（FTX）相同。该蛋白诱导肠上皮Caco-2细胞单层TEER迅速下降，细胞旁通透性平行增加，但无任何细胞毒性。免疫印迹分析显示，ftx诱导的TEER下降伴随着Caco-2细胞表面高分子量复合物的形成。细胞内Ca2+成像显示暴露于FTX引起快速Ca2+内流。电镜观察到，FTX诱导微绒毛肿胀和细胞表面膨胀。荧光phalloidin染色显示ftx处理的细胞中丝状肌动蛋白有明显变化。这些结果表明，TEER还原可以灵敏地检测到FTX在肠上皮中形成的小膜孔，这种小膜孔引起的形态学改变和细胞旁屏障功能的破坏。
We have previously found a transepithelial electrical resistance (TEER)-decreasing protein derived from Flammulina velutipes, which was revealed to be identical to flammutoxin (FTX) that is known as a hemolytic pore-forming protein. This protein induced a rapid decrease in TEER and parallel increase in paracellular permeability in the intestinal epithelial Caco-2 cell monolayer without any cytotoxicity. An immunoblotting analysis revealed that the FTX-induced decrease in TEER was accompanied by the formation of a highmolecular-weight complex on the surface of Caco-2 cells. Intracellular Ca2+ imaging showed that exposure to FTX caused a rapid Ca2+ influx. It was observed by electron microscopy that FTX induced swelling of microvilli and expansion of the cellular surface. Staining with fluorescent phalloidin showed a marked change to filamentous actin in the FTX-treated cells.These results suggest that TEER reduction could sensitively detect small membrane pore formation by FTX in the intestinal epithelium which causes a morphological alteration and disruption of the paracellular barrier function.