钙离子信号蛋白TRPC1及Orai1参与了HUVEC经SOC和ROC介导的钙内流和NO生成

发布时间：2018-03-30 22:11

本文选题：瞬时受体电位通道　切入点：钙释放激活钙通道调节分子　出处：《中国动脉硬化杂志》2017年12期

【摘要】：目的研究瞬时受体电位通道1(TRPC1)及钙释放激活钙通道调节分子1(Orai1)在人脐静脉内皮细胞(HUVEC)经钙池操纵性钙通道(SOC)和受体操纵性钙通道(ROC)介导的Ca2+内流和一氧化氮(NO)生成中的作用。方法取2~3代HUVEC进行随机分组,将构建的TRPC1、Orai1干扰质粒(sh TRPC1、sh Orai1)分别转染入HUVEC,采用real-time PCR和Western blot检测TRPC1、Orai1 mRNA和蛋白的表达及转染抑制效率。将各组细胞分别与钙敏感受体(CaR)激动剂精胺、ROC模拟剂(TPA)+CaR负性变构调节剂Calhex231、蛋白激酶C(PKC)抑制剂Ro31-8220与经典型PKCs和PKCμ抑制剂Go6967孵育后,荧光探针Fura-2/AM及DAF-FM负载方法同步检测[Ca2+]i和NO生成的变化。结果与对照组比较,sh TRPC1组和sh Orai1组mRNA表达(抑制率分别为84.50%、76.10%)和蛋白的表达(抑制率分别为83.98%、71.73%)明显降低(P0.05)。在4种不同药物作用下,sh TRPC1组和sh Orai1组[Ca2+]i△ratio值和NO净荧光强度值均明显降低(P0.05)。结论 TRPC1、Orai1参与了人脐静脉内皮细胞SOC和ROC介导的钙内流和NO生成。
[Abstract]:Objective to study the Ca2 influx and nitric oxide (no) generation mediated by transient receptor potential channel 1 / TRPC 1) and calcium release activated calcium channel regulator 1 / Orai1) via calcium pool operable calcium channel (SOC1) and receptor operable calcium channel (roc) in human umbilical vein endothelial cells (HUVECs). Methods 2 or 3 generations of HUVEC were randomly divided into two groups. Real-time PCR and Western blot were used to detect the expression of TRPC1Orai1 mRNA and protein and their transfection inhibition efficiency. The cells in each group were respectively negative to the Ca ~ (2 +) -sensitive receptor (Ca ~ (1)) agonist, spermatine (TPA) CaR, and the transfected cells were transfected into HUVECs. The expression and transfection inhibition efficiency of TRPC1- Orai1 mRNA and protein were detected by real-time PCR and Western blot, respectively. Ro31-8220, a protein kinase inhibitor, was incubated with classical PKCs and PKC 渭 inhibitor Go6967. Results compared with the control group, mRNA expression (inhibition rate was 84.50 and 76.10%) and protein expression (inhibition rate was 83.98% 71.73%) of mRNA and protein (83.98% 71.73%) were significantly decreased in sh TRPC1 group and sh Orai1 group compared with control group. I ratio and no net fluorescence intensity were significantly decreased in TRPC1 group and sh#en1# group. Conclusion TRPC 1 Orai1 is involved in calcium influx and no production mediated by SOC and ROC in human umbilical vein endothelial cells.
【作者单位】：新疆地方病与民族高发病教育部重点实验室石河子大学医学院病理生理教研室;
【基金】：国家自然科学基金资助项目(31160239;81160018)
【分类号】：R54

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