Hsieh Hsi-Lung, Tung Wei-Hsuan, Wu Cheng-Ying, Wang Hui-Hsin, Lin Chih-Chung, Wang Tze-Shyuan, Yang Chuen-Mao
Department of Nursing, Division of Basic Medical Sciences, Chang Gung Institute of Technology, Chang Gung University, Kwei-San, Tao-Yuan, Taiwan.
Arterioscler Thromb Vasc Biol. 2009 Oct;29(10):1594-601. doi: 10.1161/ATVBAHA.109.185801. Epub 2009 Jul 23.
Thrombin upregulates expression of several proteins in vascular smooth muscle cells (VSMCs) which may contribute to atherosclerosis. Here, we investigated the mechanisms underlying thrombin-induced EGF receptor (EGFR) expression and its effect on VSMCs.
Normal rat VSMCs were used. First, Western blotting and RT-PCR analyses showed that thrombin induces the expression of EGFR at transcription and translation levels in VSMCs. Second, pharmacological inhibitors, dominant negative mutants, and short hairpin RNA interference (shRNA) technology enabled us to demonstrate that thrombin-induced EGFR expression is mediated through PKC(delta)/c-Src-dependent transactivation of EGFR linking to PI3K/Akt and ERK1/2. We further investigated whether the transcription factors AP-1 and NF-kappaB are involved in this response by a promoter assay. Finally, data obtained by using EGFR shRNA technology and XTT assay demonstrated that thrombin-enhanced VSMC proliferation was mediated through upregulation of EGFR.
Our results demonstrate that thrombin-enhanced VSMC proliferation was mediated through upregulation of EGFR via a PKC(delta)/c-Src-dependent transactivation of EGFR, PI3K-Akt, and ERK, and AP-1/NF-kappaB pathway.
凝血酶可上调血管平滑肌细胞（VSMC）中多种蛋白质的表达，这可能与动脉粥样硬化的发生有关。在此，我们研究了凝血酶诱导表皮生长因子受体（EGFR）表达的机制及其对VSMC的影响。
使用正常大鼠VSMC。首先，蛋白质免疫印迹法和逆转录-聚合酶链反应（RT-PCR）分析表明，凝血酶在转录和翻译水平诱导VSMC中EGFR的表达。其次，药理学抑制剂、显性负性突变体和短发夹RNA干扰（shRNA）技术使我们能够证明，凝血酶诱导的EGFR表达是通过PKC（δ）/c-Src依赖的EGFR反式激活介导的，该反式激活与PI3K/Akt和ERK1/2相关。我们通过启动子分析进一步研究转录因子AP-1和核因子κB（NF-κB）是否参与此反应。最后，使用EGFR shRNA技术和XTT分析获得的数据表明，凝血酶增强的VSMC增殖是通过EGFR的上调介导的。
我们的结果表明，凝血酶增强的VSMC增殖是通过PKC（δ）/c-Src依赖的EGFR、PI3K-Akt和ERK反式激活以及AP-1/NF-κB途径上调EGFR介导的。
