Mechanisms of chloride secretion induced by thermostable direct haemolysin of Vibrio parahaemolyticus in human colonic tissue and a human intestinal epithelial cell line
Mechanisms of chloride secretion induced by thermostable direct haemolysin of Vibrio parahaemolyticus in human colonic tissue and a human intestinal epithelial cell line
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DOI：
10.1099/0022-1317-49-9-801
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发表时间：
2000-09-01
影响因子：
3
通讯作者：
Honda, T
中科院分区：
文献类型：
--
作者：
Takahashi, A;Sato, Y;Honda, T
由副溶血性弧菌产生的耐热直接溶血素（TDH）被认为在由该菌引起的严重腹泻中起重要作用。本研究考察了TDH对人肠细胞的肠毒性。在Ussing腔中，将TDH添加到人结肠组织的粘膜侧会导致短路电流（Isc）增加，这一过程被4,4 '-二异硫氰酸二苯乙烯-2,2'-二磺酸（DIDS）（一种Ca2+激活的氯离子（Cl-）通道抑制剂）所抑制。在人结肠上皮（ccao -2）细胞中，在细胞单层的顶端侧添加TDH后，检测到高Ise和细胞内Ca2+浓度（[Ca2+](in)）， Ise随着添加DIDS而降低，但与格列本脲、5-硝基-2-（3-苯基丙基氨基）苯甲酸或氯化钆不一致，当培养基中的Cl-被葡萄糖酸盐取代或Ca2+耗尽时，未观察到Ise随TDH的增加。在细胞外Ca2+消耗后，TDH没有升高[Ca2+](in), R7 （TDH的一种突变形式）降低了TDH对Isc和[Ca2+]（in）的影响，正如蛋白激酶C （PKC）抑制剂一样。因此，TDH增加人结肠上皮细胞的Cl-分泌，显然是通过细胞结合和Ca2+内流的机制，随后与PKC磷酸化相关的[Ca2+]（in）升高。
Thermostable direct haemolysin (TDH) produced by Vibrio parahaemolyticus is thought to play an important role in the severe diarrhoea caused by this organism. This study investigated the enterotoxicity of TDH for human intestinal cells. Addition of TDH to the mucosal side of human colonic tissue in Ussing chambers caused increased short circuit currents (Isc), a process that was inhibited by 4,4 '-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS), an inhibitor of Ca2+-activated chloride (Cl-) channels. With human colonic epithelial (Caco-2) cells, high Ise and intracellular Ca2+ concentrations ([Ca2+](in)) were detected after the addition of TDH to the apical side of the cell monolayer, The Ise decreased with the addition of DIDS, but not with glybenclamide, 5-nitro-2-(3-phenylpropylamino) benzoic acid, or gadolinium chloride, No Ise increase with TDH was observed when the Cl- in the medium was replaced by gluconate or when Ca2+ was depleted, Similarly, TDH did not raise [Ca2+](in) after depletion of extracellular Ca2+, R7, a mutant form of TDH, reduced the effects of TDH on Isc and [Ca2+](in) as did protein kinase C (PKC) inhibitors. Thus, TDH increases Cl- secretion in human colonic epithelial cells, apparently through mechanisms involving cell binding and Ca2+ influx, followed by elevation of [Ca2+](in) associated with PKC phosphorylation.