Laboratory of Cellular Pharmacology, School of Pharmacy, Aichi-Gakuin University, 1-100 Kusumoto, Chikusa, Nagoya 464-8650, Japan.
Life Sci. 2013 Mar 12;92(4-5):317-24. doi: 10.1016/j.lfs.2013.01.002. Epub 2013 Jan 16.
The vanilloid type 4 transient receptor potential channel (TRPV4) is a potential environmental sensor to multiple stimuli in many types of cells. In this study, we show that TRPV4 activated by 4α-phorbol 12,13-didecanoate (4αPDD) and hypo-osmotic stimulation (HOS) is a regulator of intracellular calcium (Ca(2+)) in human brain capillary endothelial cells (HBCEs), and its activation can partially regulate cell proliferation of HBCEs.
The expression of TRPV4 in HBCEs was analyzed at the mRNA and protein levels. The function of TRPV4 in HBCEs was evaluated using a TRPV4 agonist, 4αPDD, and HOS while measuring Ca(2+) and membrane currents.
Analysis of the mRNA transcripts of the TRPV subfamily revealed that TRPV2 and TRPV4 were expressed in HBCEs. Immunoreactivity to the TRPV4 protein was also detected in HBCEs, which were positively stained by von Willebrand factor and CD31. When 4αPDD was applied, Ca(2+) in HBCEs was elevated in a concentration-dependent manner. In addition, exposure of HBCEs to HOS at 228mOsm induced an elevation of Ca(2+). Application of 4αPDD also activated non-selective cation currents (NSCCs). Pretreatment of HBCEs with short interference RNA targeting TRPV4 (siRNA) significantly reduced the 4αPDD-induced elevation of Ca(2+). When HBCEs were treated for 24h with concentrations of 4αPDD between 0.3 and 3 μM, the cell proliferation was potentiated in a concentration-dependent manner. The potentiation was partially inhibited in HBCEs treated with siRNA.
These data suggest that endogenous TRPV4, which functions as a regulator of Ca(2+) in HBCEs, partially controls cell proliferation.
香草素型 4 瞬时受体电位通道（TRPV4）是多种细胞对多种刺激的潜在环境传感器。在这项研究中，我们表明，4α-佛波醇 12,13-二癸酸酯（4αPDD）和低渗刺激（HOS）激活的 TRPV4 是人类脑毛细血管内皮细胞（HBCEs）细胞内钙离子（Ca(2+)）的调节剂，其激活可以部分调节 HBCEs 的细胞增殖。
在 mRNA 和蛋白质水平上分析 HBCEs 中 TRPV4 的表达。使用 TRPV4 激动剂 4αPDD 和 HOS 评估 TRPV4 在 HBCEs 中的功能，同时测量Ca(2+)和膜电流。
TRPV 亚家族的 mRNA 转录本分析显示 TRPV2 和 TRPV4 在 HBCEs 中表达。TRPV4 蛋白的免疫反应性也在 HBCEs 中检测到，HBCEs 对 von Willebrand 因子和 CD31 呈阳性染色。当应用 4αPDD 时，HBCEs 中的Ca(2+)呈浓度依赖性升高。此外，将 HBCEs 暴露于 228mOsm 的低渗溶液中会引起Ca(2+)的升高。应用 4αPDD 还激活非选择性阳离子电流（NSCCs）。用靶向 TRPV4 的短发夹 RNA（siRNA）预处理 HBCEs 可显著降低 4αPDD 诱导的Ca(2+)升高。当 HBCEs 用 0.3 至 3μM 之间浓度的 4αPDD 处理 24 小时时，细胞增殖呈浓度依赖性增强。在用 siRNA 处理的 HBCEs 中，这种增强作用部分受到抑制。
这些数据表明，作为 HBCEs 中Ca(2+)调节剂的内源性 TRPV4 部分控制细胞增殖。
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