Hamdollah Zadeh M A, Glass Catherine A, Magnussen Anette, Hancox Jules C, Bates David O
Bristol Heart Institute, Department of Physiology and Pharmacology, School of Veterinary Sciences, University of Bristol, United Kingdom.
Microcirculation. 2008 Oct;15(7):605-14. doi: 10.1080/10739680802220323.
Vascular endothelial growth factor (VEGF)-induced vascular permeability has been shown to be dependent on calcium influx, possibly through a transient receptor potential cation channel (TRPC)-mediated cation channel with properties of the TRPC3/6/7 subfamily. To investigate further the involvement of this subfamily, we determined the effects of dominant negative TRPC6 overexpression on VEGF-mediated changes of human microvascular endothelial cell (HMVEC) calcium, proliferation, migration, and sprouting.
Cytoplasmic calcium concentration was estimated by fura-2 fluorescence spectrophotometry, migration by Boyden chamber assay, sprouting by immunofluorescence imaging of stimulated endothelial cells, and proliferation by flow cytometry.
Overexpression of a dominant negative TRPC6 construct in HMVECs inhibited the VEGF-mediated increases in cytosolic calcium, migration, sprouting, and proliferation. In contrast, overexpression of a wild-type TRPC6 construct increased the proliferation and migration of HMVECs.
TRPC6 is an obligatory component of cation channels required for the VEGF-mediated increase in cytosolic calcium and subsequent downstream signaling that leads to processes associated with angiogenesis.
血管内皮生长因子（VEGF）诱导的血管通透性已被证明依赖于钙内流，可能是通过一个具有瞬时受体电位阳离子通道（TRPC）特性的阳离子通道介导，该通道属于TRPC3/6/7亚家族。为了进一步研究该亚家族的作用，我们测定了显性负性TRPC6过表达对VEGF介导的人微血管内皮细胞（HMVEC）钙变化、增殖、迁移和芽生的影响。
通过fura-2荧光分光光度法估算细胞质钙浓度，通过Boyden小室试验检测迁移，通过对受刺激内皮细胞进行免疫荧光成像检测芽生，通过流式细胞术检测增殖。
在HMVEC中过表达显性负性TRPC6构建体可抑制VEGF介导的胞质钙增加、迁移、芽生和增殖。相反，过表达野生型TRPC6构建体可增加HMVEC的增殖和迁移。
TRPC6是VEGF介导的胞质钙增加及随后导致血管生成相关过程的下游信号传导所需阳离子通道的必需组成部分。
