人血管内皮细胞特异性表达TMEM 16B

Human vascular endothelial cells express TMEM 16B specifically

目的探讨人血管内皮细胞上TMEM16A、TMEM16B蛋白的表达情况,并初步探讨后者的功能。方法采用实时PCR方法检测人脐带静脉内皮细胞(human umbilical vein endothelial cells,HUVECs)中TMEM16A、TMEM16B的基因表达情况。采用Western-Blot及免疫荧光染色方法进一步证实HUVECs中TMEM16B的表达情况。Fura-2am标记HUVECs后采用Flex-station检测细胞内钙变化情况。结果 HUVECs细胞表达TMEM16B,而未表达TMEM16A。Western-Blot及免疫荧光方法从蛋白水平证明HUVECs表达TMEM16B。在使用TMEM16B的si RNA干预HUVECs后,无论是有钙及无钙溶液,在50 n M血管内皮细胞生长因子(vascular endothelial growth factor,VEGF)刺激下,钙浓度峰值均增大,而且峰值出现时间提前。结论人血管内皮细胞特异性表达TMEM16B,生理情况下TMEM16B对VEGF刺激引起的Ca2+的释放过程可能起着负向调节作用。

Objective To investigate the expression of TMEM16 A and TMEM16 B protein in human vascular endothelial cells and try to clarify the function of TMEM16 B.Methods Real time PCR was used to check the expression of TMEM16 A and TMEM16 B in human umbilical vein endothelial cells（HUVECs）.Western-Blot and immunofluorescence methods were used to confirm the expression of TMEM16 B in HUVECs.Flex-station was performed to detect the change of intracellular calcium in HUVECs marked with fura-tam.Results HUVECs expressed TMEM16 B but not TMEM16 A.Western-Blot and immunofluorescence methods both proved that HUVECs expressed TMEM16 B.After being intervened by siRNA of TMEM16 B,with the stimulation of 50 nM VEGF,HUVECs in solution with normal Ca^（2＋） showed not only an increased peak value but also an advanced latency of peak value.Above effect remained when HUVECs were in solution without Ca^（2＋）.Conclusion HUVECs expressed TMEM16 B specifically,and it could play an negative feedback function of intracellular calcium releasing evoked by VEGF in physiological condition.