Li Li-Min, Zheng Bing, Zhang Ruo-Nan, Jin Li-Shuang, Zheng Cui-Ying, Wang Chang, Zhou Pei-Pei, Guo Zong-Wei, Ma Dong, Wen Jin-Kun
Department of Biochemistry and Molecular Biology, Key Laboratory of Neural and Vascular Biology, China Administration of Education, Hebei Medical University, Shijiazhuang, China.
Cell Biochem Funct. 2015 Jun;33(4):226-34. doi: 10.1002/cbf.3108. Epub 2015 Apr 23.
Tongxinluo (TXL) is a compound prescription formulated according to the meridian theory of traditional Chinese medicine. It may play an important role in cardiovascular protection by improving endothelial cell function. The aim of present study was to investigate whether endothelial protection with TXL is related to its regulation of tight junction protein expression. Human cardiac microvascular endothelial cells (HCMECs) were cultured and treated with 10(-7) mol l(-1) angiotensin II (Ang II) and the different doses of TXL; the expression of tight junction proteins occludin, claudin, VE-cadherin and beta-catenin was determined by Western blotting and real-time PCR. Gain-of-function and loss-of-function of Krüppel-like factor 5 (KLF5) were carried out in HCMEC transfected with either KLF5 adenovirus pAd-KLF5 or siRNA specific for KLF5. Angiotensinogen transgenic mice were treated with TXL by oral administration of TXL of 0.75 g kg(-1) day(-1) , and immunohistochemical staining was performed with antioccludin, anticlaudin, anti-VE-cadherin, antibeta-catenin and anti-KLF5 antibodies. Ang II treatment significantly reduced the expression of tight junction proteins occludin, claudin, VE-cadherin and beta-catenin in cultured HCMECs. TXL pretreatment could abrogate the down-regulation of these tight junction proteins induced by Ang II. Ang II treatment also decreased KLF5 expression at the mRNA and protein levels; TXL pretreatment markedly reversed the inhibitory effect of Ang II on KLF5 expression. Gain-of-function and loss-of-function of KLF5 showed that KLF5 mediated the expression of tight junction proteins in HCMECs. TXL-enhanced expression of the tight junction proteins was mediated by KLF5. In angiotensinogen transgenic mice, TXL also increased the tight junction protein levels by inducing KLF5 expression. Chinese medicine TXL increases tight junction protein levels by inducing KLF5 expression in microvascular endothelial cells.
通心络（TXL）是根据中医经络理论配制的复方制剂。它可能通过改善内皮细胞功能在心血管保护中发挥重要作用。本研究的目的是探讨通心络的内皮保护作用是否与其对紧密连接蛋白表达的调节有关。培养人心脏微血管内皮细胞（HCMECs），并用10⁻⁷ mol·L⁻¹血管紧张素II（Ang II）和不同剂量的通心络进行处理；通过蛋白质免疫印迹法和实时定量PCR测定紧密连接蛋白occludin、claudin、血管内皮钙黏蛋白（VE-cadherin）和β-连环蛋白的表达。在转染Krüppel样因子5（KLF5）腺病毒pAd-KLF5或KLF5特异性小干扰RNA（siRNA）的HCMECs中进行KLF5的功能获得和功能缺失实验。给血管紧张素原转基因小鼠口服0.75 g·kg⁻¹·d⁻¹的通心络进行处理，并用抗occludin、抗claudin、抗VE-cadherin、抗β-连环蛋白和抗KLF5抗体进行免疫组织化学染色。Ang II处理显著降低了培养的HCMECs中紧密连接蛋白occludin、claudin、VE-cadherin和β-连环蛋白的表达。通心络预处理可消除Ang II诱导的这些紧密连接蛋白的下调。Ang II处理还降低了KLF5在mRNA和蛋白水平的表达；通心络预处理显著逆转了Ang II对KLF5表达的抑制作用。KLF5的功能获得和功能缺失实验表明，KLF5介导了HCMECs中紧密连接蛋白的表达。通心络增强紧密连接蛋白的表达是由KLF5介导的。在血管紧张素原转基因小鼠中，通心络还通过诱导KLF5表达增加了紧密连接蛋白水平。中药通心络通过诱导微血管内皮细胞中KLF5表达增加紧密连接蛋白水平。
