...
  • 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Journal of Molecular and Cellular Cardiology >ADAM17 silencing by adenovirus encoding miRNA-embedded siRNA revealed essential signal transduction by angiotensin II in vascular smooth muscle cells
【24h】

ADAM17 silencing by adenovirus encoding miRNA-embedded siRNA revealed essential signal transduction by angiotensin II in vascular smooth muscle cells

机译：编码miRNA嵌入siRNA的腺病毒对ADAM17的沉默揭示了血管紧张素II在血管平滑肌细胞中的重要信号转导

获取原文
获取原文并翻译 | 示例
           
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

Small interfering RNA (siRNA) mediated gene silencing has been utilized as a powerful molecular tool to study the functional significance of a specific protein. However, due to transient gene silencing and insufficient transfection efficiency, this approach can be problematic in primary cell culture such as vascular smooth muscle cells. To overcome this weakness, we utilized an adenoviral-encoded microRNA (miRNA)-embedded siRNA "mi/siRNA"-based RNA interference. Here, we report the results of silencing a disintegrin and metalloprotease 17 (ADAM17) in cultured rat vascular smooth muscle cells and its functional mechanism in angiotensin II signal transduction. 3 distinct mi/siRNA sequences targeting rat ADAM17 were inserted into pAd/CMV/V5-DEST and adenoviral solutions were obtained. Nearly 90% silencing of ADAM17 was achieved when vascular smooth muscle cells were infected with 100 multiplicity of infection of each ADAM17 mi/siRNA encoding adenovirus for 3. days. mi/siRNA-ADAM17 but not mi/siRNA-control inhibited angiotensin II-induced epidermal growth factor receptor trans-activation and subsequent extracellular signal-regulated kinase activation and hypertrophic response in the cells. mi/siRNA-ADAM17 also inhibited angiotensin II-induced heparin-binding epidermal growth factor-like factor shedding. This inhibition was rescued with co-infection of adenovirus encoding mouse ADAM17 but not by its cytosolic domain deletion mutant or cytosolic Y702F mutant. As expected, angiotensin II induced tyrosine phosphorylation of ADAM17 in the cells. In conclusion, ADAM17 activation via its tyrosine phosphorylation contributes to heparin-binding epidermal growth factor-like factor shedding and subsequent growth promoting signals induced by angiotensin II in vascular smooth muscle cells. An artificial mi/siRNA-based adenoviral approach appears to be a reliable gene-silencing strategy for signal transduction research in primary cultured vascular cells.
机译：小干扰RNA（siRNA）介导的基因沉默已被用作研究特定蛋白质功能重要性的强大分子工具。然而，由于瞬时基因沉默和不足的转染效率，这种方法在原代细胞培养如血管平滑肌细胞中可能是有问题的。为了克服这一弱点，我们利用了腺病毒编码的microRNA（miRNA）嵌入的基于siRNA“ mi / siRNA”的RNA干扰。在这里，我们报告了在培养的大鼠血管平滑肌细胞中沉默整联蛋白和金属蛋白酶17（ADAM17）的结果及其在血管紧张素II信号转导中的功能机制。将靶向大鼠ADAM17的3个不同的mi / siRNA序列插入pAd / CMV / V5-DEST中，并获得腺病毒溶液。当血管平滑肌细胞被编码腺病毒的每种ADAM17 mi / siRNA感染100多重感染3天后，ADAM17的沉默率几乎达到90％。 mi / siRNA-ADAM17而非mi / siRNA对照抑制了血管紧张素II诱导的表皮生长因子受体反式激活，并随后抑制了细胞中细胞外信号调节的激酶激活和肥大反应。 mi / siRNA-ADAM17还抑制血管紧张素II诱导的肝素结合表皮生长因子样因子脱落。通过共感染编码小鼠ADAM17的腺病毒来挽救这种抑制作用，但不能通过其胞质结构域缺失突变体或胞质Y702F突变体来挽救这种抑制作用。如预期的那样，血管紧张素II诱导细胞中ADAM17的酪氨酸磷酸化。总之，ADAM17通过其酪氨酸磷酸化的激活有助于肝素结合表皮生长因子样因子的脱落，并随后由血管紧张素II诱导血管平滑肌细胞中的生长促进信号。基于mi / siRNA的人工腺病毒方法似乎是用于原代培养血管细胞信号转导研究的可靠基因沉默策略。

著录项

相似文献

  • 外文文献
  • 中文文献
  • 专利
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号