Stabilization of F-actin prevents cAMP-elicited Cl- secretion in T84 cells.
Stabilization of F-actin prevents cAMP-elicited Cl- secretion in T84 cells.
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F-肌动蛋白的稳定可防止 T84 细胞中 cAMP 引发的 Cl 分泌。
DOI：
10.1172/jci115215
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发表时间：
1991
期刊：
影响因子：
--
通讯作者：
Madara,JL
中科院分区：
文献类型：
--
作者：
Shapiro,M;Matthews,J;Hecht,G;Delp,C;Madara,JL
T84细胞是人类肠道上皮细胞系，是电生氯离子分泌的模型。我们发现，cAMP诱导的T84细胞的Cl-分泌伴随着F-肌动蛋白在细胞的基底外侧部分的显著重新分布。为了防止这种F-肌动蛋白的重新分布，从而评估其对氯离子分泌的重要性，我们定义了这种模型上皮可以负载硝基苯并恶二唑(NBD)-鬼臼毒素的简单条件。这种试剂以高亲和力结合F-肌动蛋白，从而通过防止解聚来稳定F-肌动蛋白细胞骨架，解聚是肌动蛋白微丝动态重新排序所必需的事件。通过乳酸脱氢酶释放、蛋白质合成、跨上皮阻力和负载细胞在顶端添加Na+离子载体时向吸收方向泵送Na+的能力来评估NBD-Pallicidin负载没有细胞毒性。然而，cAMP诱导的F-肌动蛋白重分布和cAMP诱导的Cl-分泌反应在NBD-Pallicidin预加载的T84细胞中均显著受损。相反，NBD-Pallicidin预负荷不能减弱氨甲酰胆碱引起的Cl-分泌反应(钙离子介导)，也不伴随F-肌动蛋白的重新分布。这些发现表明，我们所描述的cAMP引起的细胞骨架重分布是cAMP引起的T84细胞氯离子分泌的一个组成部分。
T84 cells, a human intestinal epithelial cell line, serve as a model of electrogenic Cl- secretion. We find that cAMP-elicited Cl- secretion in T84 cells is accompanied by a marked redistribution of F-actin in the basolateral portion of the cell. To prevent this F-actin redistribution and thereby assess its importance to Cl- secretion, we have defined simple conditions under which this model epithelium can be loaded with nitrobenzoxadiazole (NBD)-phallicidin. This reagent binds F-actin with high affinity thus stabilizing the F-actin cytoskeleton by preventing depolymerization, an event necessary for dynamic reordering of actin microfilaments. NBD-phallicidin loading is not cytotoxic as assessed by lactic dehydrogenase release, protein synthesis, transepithelial resistance, and the ability of the loaded cells to pump Na+ in an absorptive direction in response to the apical addition of a Na+ ionophore. However, cAMP-elicited redistribution of F-actin and the cAMP-elicited Cl- secretory response are both markedly impaired in NBD-phallicidin preloaded T84 cells. In contrast, the carbachol-elicited Cl- secretory response (Ca++ mediated) is not attenuated by NBD-phallicidin preloading nor is it accompanied by redistribution of F-actin. These findings suggest that the cAMP-elicited cytoskeletal redistribution we describe is an integral part of cAMP-elicited Cl- secretion in T84 cells.Images