Dependence of regulatory volume decrease on transient receptor potential vanilloid 4 (TRPV4) expression in human corneal epithelial cells
Dependence of regulatory volume decrease on transient receptor potential vanilloid 4 (TRPV4) expression in human corneal epithelial cells
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DOI：
10.1016/j.ceca.2008.01.008
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发表时间：
2008-10-01
期刊：
影响因子：
4
通讯作者：
Reinach, Peter S.
中科院分区：
文献类型：
--
作者：
Pan, Zan;Yang, Hua;Reinach, Peter S.
TRPV4 是一种具有中等钙渗透性的非选择性阳离子通道，可通过低渗状态激活。这种压力会导致人角膜上皮细胞（HCEC）的调节体积减少（RVD）行为。我们假设 TRPV4 通道介导 HCEC 中的 RVD。免疫组织化学显示 TRPV4 在角膜组织中集中和表面集中定位。免疫细胞化学和荧光激活细胞分选仪 (FACS) 分析鉴定了 TRPV4 膜表面和胞浆表达。 RT-PCR 和蛋白质印迹分析分别鉴定了 HCEC 中 TRPV4 基因和蛋白的表达。此外，4 α-PDD 或 50% 低渗介质诱导瞬时细胞内 Ca2+ ([Ca2+](i)) 增加多达三倍。 TRPV4 siRNA HCEC 转染后，其蛋白表达水平下降了 64%，从而消除了这些 [Ca2+](i) 瞬变。同样，暴露于钌红或不含 Ca2+ 的林格氏溶液也消除了这种反应。在这些转染的细胞中，RVD 下降了 51%，而在未转染的细胞中，钌红和不含 Ca2+ 的溶液分别抑制 RVD 54% 和 64%。相比之下，TRPV1 拮抗剂辣椒西平未能抑制 [Ca2+](i) 瞬变和 RVD。 TRPV4 激活会导致 RVD，因为 TRPV4 表达和活性的下降与该反应的抑制相关。总之，HCEC中存在TRPV4功能性表达。 (C) 2008 Elsevier Ltd. 保留所有权利。
TRPV4 is a non-selective cation channel with moderate calcium permeability, which is activated by exposure to hypotonicity. Such a stress induces regulatory volume decrease (RVD) behavior in human corneal epithelia[ cells (HCEC). We hypothesize that TRPV4 channel mediates RVD in HCEC. Immunohistochemistry revealed centrally and superficially concentrated TRPV4 localization in the corneal tissue. Immunocytochemical and fluorescence activated cell sorter (FACS) analyses identified TRPV4 membrane surface and cytosolic expression. RT-PCR and Western blot analyses identified TRPV4 gene and protein expression in HCEC, respectively. In addition, 4 alpha-PDD or a 50% hypotonic medium induced up to threefold transient intracellular Ca2+ ([Ca2+](i)) increases. Following TRPV4 siRNA HCEC transfection, its protein expression level declined by 64%, which abrogated these [Ca2+](i) transients. Similarly, exposure to either ruthenium red or Ca2+-free Ringer's solution also eliminated this response. In these transfected cells, RVD declined by 51% whereas in the non-transfected counterpart, ruthenium red and Ca2+-free solution inhibited RVD by 54 and 64%, respectively. In contrast, capsazepine, a TRPV1 antagonist, failed to suppress [Ca2+](i) transients and RVD. TRPV4 activation contributes to RVD since declines in TRPV4 expression and activity are associated with suppression of this response. In conclusion, there is TRPV4 functional expression in HCEC. (C) 2008 Elsevier Ltd. All rights reserved.