Functional inhibition of native volume-sensitive outwardly rectifying anion channels in muscle cells and Xenopus oocytes by anti-ClC-3 antibody
Functional inhibition of native volume-sensitive outwardly rectifying anion channels in muscle cells and Xenopus oocytes by anti-ClC-3 antibody
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DOI：
10.1111/j.1469-7793.2001.0437i.x
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发表时间：
2001-03-01
影响因子：
5.5
通讯作者：
Hume, JR
中科院分区：
文献类型：
--
作者：
Duan, DY;Zhong, JM;Hume, JR
1. 用市售的抗ClC-3多克隆抗体（Ab）在NIH/3T3细胞内透析约30分钟，完全抑制豚鼠表达的ClC-3电流（IgpClC-3），而用抗原预吸收的抗clc -5 Ab在细胞内透析不能影响IgpClC-3.2。采用抗ClC-3抗体作为选择性探针，研究了内源性ClC-3表达与豚鼠心肌细胞、犬肺动脉平滑肌细胞和非洲爪蟾卵母细胞中天然体积敏感外向纠偏阴离子通道（VSOACs）的关系。细胞内透析或注射抗clc -3抗体可消除心脏细胞和pasmc中天然VSOAC的功能，并显著降低卵母细胞中的VSOAC。相比之下，抗原预吸收anti- clc - 3ab .3未改变天然VSOAC功能。这表明内源性的ClC-3是负责心脏、平滑肌细胞和爪蟾卵母细胞中天然VSOACs的主要分子实体。Anti-ClC-3 Ab应该是一个有用的实验工具，可以直接测试内源性ClC-3表达与天然VSOAC功能之间的关系，并有助于解决现有的关于多种不同细胞中天然VSOAC的调节和生理作用的争议。
1. Intracellular dialysis of NIH/3T3 cells with a commercially available anti-ClC-3 polyclonal antibody (Ab) for similar to 30 min completely inhibited expressed guinea-pig ClC-3 currents (IgpClC-3), while intracellular dialysis with antigen-preabsorbed anti-ClC-5 Ab failed to affect IgpClC-3.2. Anti-ClC-3 Ab was used as a selective probe to examine the relationship between endogenous ClC-3 expression and native volume-sensitive outwardly rectifying anion channels (VSOACs) in guinea-pig cardiac cells, canine pulmonary arterial smooth muscle cells (PASMCs) and Xenopus laevis oocytes. Intracellular dialysis or injection of anti-ClC-3 Ab abolished native VSOAC: function in cardiac cells and PASMCs and significantly reduced VSOACs in oocytes. In contrast, native VSOAC function was unaltered by antigen-preabsorbed anti-ClC-3 Ab.3. It is suggested that endogenous ClC-3 represents a major molecular entity responsible for native VSOACs in cardiac and smooth muscle cells and Xenopus oocytes. Anti-ClC-3 Ab should be a useful experimental tool to directly test the relationship between endogenous ClC-3 expression and native VSOAC function, and help resolve existing controversies related to the regulation and physiological role of native VSOACs in a wide variety of different cells.