Iwasaka C, Tanaka K, Abe M, Sato Y
Department of Vascular Biology, Tohoku University, Sendai, Japan.
J Cell Physiol. 1996 Dec;169(3):522-31. doi: 10.1002/(SICI)1097-4652(199612)169:3<522::AID-JCP12>3.0.CO;2-7.
The coordinate induction of protease activities and cell migration is a principal feature of endothelial cells (ECs) invading the interstitial space in the initial step of angiogenesis. However, the molecular mechanisms of these events are not fully characterized. Ets-1 is a member of the ets gene family of transcription factors, which binds to the Ets binding motif in the cis-acting elements and regulates the expression of certain genes. Four typical angiogenic growth factors, aFGF, bFGF, VEGF, and EGF, induced the expression of ets-1 mRNA in either human umbilical vein endothelial cells (HUVECs), ECV-304 cells (immortalized HUVECs), or human omental microvascular endothelial cells (HOMECs). The expression of ets-1 reached its maximum at 2 hr after factor addition and then decreased to the basal level by 12 hr. For characterization of the role of Ets-1 in angiogenesis, ets-1 antisense and sense oligodeoxynucleotides (ODNs) were constructed. The ets-1 antisense ODN but not sense ODN efficiently blocked the synthesis of Ets-1 protein by human ECs in response to angiogenic growth factors. Moreover, the ets-1 antisense ODN but not sense ODN almost completely abolished the binding of endothelial cell extract to DNA containing the Ets binding motif. The expression of urokinase-type plasminogen activator and matrix metalloproteinase-1 and the migration of ECs in response to growth factors were significantly inhibited by ets-1 antisense ODN but not by sense ODN. Tube formation by HOMECs in type 1 collagen gel stimulated with EGF was abrogated by ets-1 antisense ODN. Finally, the expression of Ets-1 protein in ECs during angiogenesis in vivo was confirmed by an immunohistochemical analysis using a murine angiogenesis model. These results indicate that the induction of ets-1 mRNA is a mutual phenomenon in ECs stimulated with angiogenic growth factors. Ets-1 appears to play an important role in angiogenesis, regulating the expression of proteases and the migration of ECs.
蛋白酶活性与细胞迁移的协同诱导是血管生成初始阶段内皮细胞（ECs）侵入间质空间的一个主要特征。然而，这些事件的分子机制尚未完全明确。Ets-1是ets转录因子基因家族的成员，它与顺式作用元件中的Ets结合基序结合并调节某些基因的表达。四种典型的血管生成生长因子，即酸性成纤维细胞生长因子（aFGF）、碱性成纤维细胞生长因子（bFGF）、血管内皮生长因子（VEGF）和表皮生长因子（EGF），均可在人脐静脉内皮细胞（HUVECs）、ECV-304细胞（永生化HUVECs）或人网膜微血管内皮细胞（HOMECs）中诱导ets-1 mRNA的表达。ets-1的表达在添加因子后2小时达到最大值，然后在12小时时降至基础水平。为了阐明Ets-1在血管生成中的作用，构建了ets-1反义寡脱氧核苷酸（ODNs）和正义寡脱氧核苷酸。ets-1反义ODN而非正义ODN能够有效阻断人内皮细胞在血管生成生长因子作用下合成Ets-1蛋白。此外，ets-1反义ODN而非正义ODN几乎完全消除了内皮细胞提取物与含有Ets结合基序的DNA的结合。ets-1反义ODN可显著抑制尿激酶型纤溶酶原激活剂和基质金属蛋白酶-1的表达以及内皮细胞对生长因子的迁移反应，而正义ODN则无此作用。ets-1反义ODN可消除EGF刺激下HOMECs在I型胶原凝胶中形成管样结构的能力。最后，通过使用小鼠血管生成模型的免疫组织化学分析证实了体内血管生成过程中内皮细胞中Ets-1蛋白的表达。这些结果表明，ets-1 mRNA的诱导是血管生成生长因子刺激内皮细胞时的一种共同现象。Ets-1似乎在血管生成中起重要作用，调节蛋白酶的表达和内皮细胞的迁移。
