MiR-1 is a Critical Regulator of VEGF-Induced Angiogenesis

MiR-1 是 VEGF 诱导的血管生成的关键调节因子

• 批准号: 8714026
• 负责人: Seyedtaghi Takyar
• 金额: $ 24.4万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-05 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8714026
• 关键词: Angiogenic Factor; Biological Process; Blood Circulation; Blood Vessels; Cell Culture Techniques; Cell Differentiation process; Cell Proliferation; Cells; Chronic; Data; Development; Disease; Down-Regulation; ENG gene; Embryonic Development; Endothelial Cells; Functional RNA; Future; Gene Expression; Genetic Translation; Growth; Growth Factor; Healed; Hemorrhage; In Vitro; Invaded; Lung; Maintenance; Malignant Neoplasms; Measures; Messenger RNA; MicroRNAs; Mus; Neoplasm Metastasis; Neoplasms; PECAM1 gene; PTPRC gene; Pathologic Processes; Pathway interactions; Permeability; Phosphorylation; Play; Precipitation; Process; Property; RNA-Induced Silencing Complex; Recruitment Activity; Reverse Transcriptase Polymerase Chain Reaction; Role; Route; Signal Pathway; Signal Transduction; Small RNA; Sorting - Cell Movement; Staining method; Stains; Supplementation; Technology; Testing; Tissues; Trachea; Transgenic Mice; Transgenic Model; Transgenic Organisms; Translations; Tube; VEGF165; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor B; Vascular Endothelial Growth Factor C; Vascular Endothelial Growth Factors; Wound Healing; Wounds and Injuries; angiogenesis; carcinogenesis; healing; in vivo; injury and repair; migration; neovascularization; novel; public health relevance; research study; response; stem; tumor; vascular bed; wound

DESCRIPTION (provided by applicant): Project Summary VEGF (Vascular Endothelial Growth Factor) is one of the main angiogenic factors involved in the development and maintenance of blood vessels and is shown to play a central role in various pathological processes such as wound healing, carcinogenesis and metastasis. MicroRNAs are a recently recognized group of non-coding small RNA molecules that regulate gene expression by inhibiting the translation of mRNAs or facilitating their degradation. We characterized the lung microRNA profile of the CC10-rtTA-VEGF transgenic mice in microarray experiments and confirmed our findings by stem-loop RT-PCR. The level of miR-1 was quantified in (CD45-, CD31+) and (CD45-, CD105+) cells separated from total lung by FACS sorting. We measured the level of miR- 1 in primary mouse lung endothelial cells in vitro after stimulation with VEGF. The effect of miR-1 supplementation on VEGF-induced proliferation and cord formation was studied in cell culture. The in vivo effect of miR-1 on angiogenesis after intranasal delivery was characterized by staining of the trachea from VEGF transgenic mice with CD31. We found that the levels of miR-1 were consistently diminished in the total lung RNAs from CC10-rtTa- VEGF transgenic mice, endothelial cells of these mice separated by FACS sorting, and mouse primary lung endothelial cells stimulated by VEGF in vitro, each compared to appropriate controls. MiR-1 supplementation downregulated VEGF-induced endothelial cell proliferation and cord formation in vitro and intranasal delivery of miR-1 inhibited angiogenesis in the bronchial circulation in vivo. Hypothesis: MiR-1 is a critical regulator of VEGF-induced endothelial cell responses To test the validity of this hypothesis we propose to: Aim #1: Characterize the spectrum of VEGF-induced endothelial cell responses regulated by miR-1 in vitro. Aim #2: Characterize the effect of miR-1 over-expression on VEGF responses in vivo (by A. direct delivery and B. transgenic modeling). Future Directions: Define the mechanism of miR-1 effects (by A. characterizing the effect of miR-1 on signaling and, B: identifying mRNAs recruited to RNA Induced Silencing complex (RISC).

描述（由申请人提供）：项目概述VEGF（血管内皮生长因子）是参与血管发育和维持的主要血管生成因子之一，并显示在各种病理过程中发挥核心作用，如伤口愈合、致癌和转移。MicroRNA是近年来发现的一类非编码小RNA分子，通过抑制mRNA的翻译或促进其降解来调节基因表达。 我们在微阵列实验中表征了CC 10-rtTA-VEGF转基因小鼠的肺microRNA谱，并通过茎环RT-PCR证实了我们的发现。在通过FACS分选从总肺分离的（CD 45-，CD 31+）和（CD 45-，CD 105+）细胞中定量miR-1的水平。我们在体外检测了VEGF刺激后原代小鼠肺内皮细胞中miR- 1的水平。在细胞培养中研究了miR-1补充对VEGF诱导的增殖和索形成的影响。通过用CD 31对VEGF转基因小鼠的气管进行染色来表征鼻内递送后miR-1对血管生成的体内作用。 我们发现，与适当的对照相比，来自CC 10-rtTa-VEGF转基因小鼠、通过FACS分选分离的这些小鼠的内皮细胞和体外由VEGF刺激的小鼠原代肺内皮细胞的总肺RNA中的miR-1水平一致地降低。miR-1补充下调VEGF诱导的内皮细胞增殖和体外索形成，并且miR-1的鼻内递送抑制体内支气管循环中的血管生成。假设：miR-1是VEGF诱导的内皮细胞反应的关键调节剂为了测试这一假设的有效性，我们提出：目的#1：表征由miR-1在体外调节的VEGF诱导的内皮细胞反应的谱。目的#2：表征miR-1过表达对体内VEGF应答的影响（通过A.直接交付和B.转基因建模）。未来方向：定义miR-1效应的机制（A.表征miR-1对信号传导的作用，以及，B：鉴定募集到RNA诱导沉默复合物（RISC）的mRNA。