Role of dicer and drosha for endothelial MicroRNA expression and angiogenesis
Role of dicer and drosha for endothelial MicroRNA expression and angiogenesis
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DOI：
10.1161/circresaha.107.153916
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发表时间：
2007-07-06
影响因子：
20.1
通讯作者：
Dimmeler, Stefanie
中科院分区：
文献类型：
--
作者：
Kuehbacher, Angelika;Urbich, Carmen;Dimmeler, Stefanie
microRNA（miRNA）是小的非编码RNA，通过与细胞转录物结合，导致靶mRNA的翻译抑制或降解来调节基因表达。 DICER和DROSHA是miRNA成熟所需的miRNA加工酶。在这里，我们研究了DICER和DROSHA在血管生成中的作用。将内皮细胞用siRNA针对DICER和DROSHA转染，以抑制miRNA生物发生。 DICER和DROSHA的遗传沉默显着降低了内皮细胞的毛细血管发芽和导管形成活性。在转染的细胞中，内皮细胞的迁移显着降低，而drosha siRNA无效。 DICER的沉默，但不能使Drosha的沉默减少体内血管生成。接下来，我们试图鉴定内皮细胞中表达的miRNA。使用实时PCR对168个人miRNA的筛查分析表明，在内皮细胞中高度表达了let-7家族，mir-21，mir-126，mir-221和mir-222的成员。 DICER和DROSHA siRNA降低了LEF-7F和miR-27b表达。针对Let-7f和miR-27b的抑制剂还降低了发芽形成，表明Let-7F和miR-27b通过靶向抗血管生成基因来促进血管生成。在对预测靶标的硅分析中 - 鉴定内源性血管生成抑制剂血小板蛋白-1。的确，DICER和DROSHA siRNA显着增加了血小板传播1的表达。瞬时减少miRNA-调节miRNA-调节酶DICER损害血管生成的酶diCAIR-DICER障碍了血管生成的作用。体外和体内，而Drosha siRNA诱导了较小的抗血管生成效应体外，在体内无效。 Let-7家族和miR-27b似乎是调节血管生成的吸引力靶标。
MicroRNAs ( miRNAs) are small noncoding RNAs that regulate gene expression by binding to the cellular transcript leading to translational repression or degradation of the target mRNA. Dicer and Drosha are the miRNA processing enzymes that are required for the maturation of miRNAs. Here, we investigated the role of Dicer and Drosha for angiogenesis. Endothelial cells were transfected with siRNA against Dicer and Drosha to inhibit miRNA biogenesis. Genetic silencing of Dicer and Drosha significantly reduced capillary sprouting of endothelial cells and tube forming activity. Migration of endothelial cells was significantly decreased in Dicer siRNA - transfected cells, whereas Drosha siRNA had no effect. Silencing of Dicer but not of Drosha reduced angiogenesis in vivo. Next, we attempted to identify miRNAs expressed in endothelial cells. A screening analysis of 168 human miRNAs using real- time PCR revealed that members of the let- 7 family, mir- 21, mir- 126, mir- 221, and mir- 222 are highly expressed in endothelial cells. Dicer and Drosha siRNA reduced lef- 7f and mir- 27b expression. Inhibitors against let- 7f and mir- 27b also reduced sprout formation indicating that let- 7f and mir- 27b promote angiogenesis by targeting antiangiogenic genes. In silico analysis of predicted targets for let- 7 cluster identified the endogenous angiogenesis inhibitor thrombospondin- 1. Indeed, Dicer and Drosha siRNA significantly increased the expression of thrombospondin- 1. Taken together, transient reduction of the miRNA- regulating enzyme Dicer impairs angiogenesis in vitro and in vivo, whereas Drosha siRNA induced a minor antiangiogenic effect in vitro and was not effective in vivo. The let- 7 family and mir- 27b appear to be attractive targets for modulating angiogenesis.