MiR-409-3p regulates angiogenesis, brown fat adiposity, and insulin resistance

MiR-409-3p 调节血管生成、棕色脂肪肥胖和胰岛素抵抗

• 批准号: 10052898
• 负责人: Basak Icli
• 金额: $ 60.33万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-01 至 2020-11-01
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10052898
• 关键词: 3' Untranslated Regions; 3T3-L1 Cells; Ablation; Adipocytes; Adipose tissue; Age; Angiogenesis Inhibition; Angiogenic Factor; Animal Model; Binding; Biological Assay; Blood Vessels; Brown Fat; Cell Proliferation; Coculture Techniques; Code; Development; Diet; Endothelial Cells; Energy Metabolism; Excision; Fatty acid glycerol esters; Functional disorder; Gene Expression; Genes; Growth; Growth Factor; High Fat Diet; Homeobox; Hormones; Human; Impairment; Insulin; Insulin Resistance; Knock-out; Knockout Mice; Length; MAP Kinase Kinase Kinase; Measures; Mediating; Metabolic; Metabolic Diseases; Metabolic dysfunction; Metabolism; MicroRNAs; Molecular; Mus; Non-Insulin-Dependent Diabetes Mellitus; Nucleotides; Nutrient; Obese Mice; Obesity; Organ; Organoids; PECAM1 gene; PGF gene; Patients; Phenocopy; Phosphotransferases; Plasma; Play; Proteins; Regulation; Reporter; Role; Sampling; Signal Pathway; Signal Transduction; Skin; Small Interfering RNA; Stains; Stimulus; Temperature; Testing; Therapeutic Intervention; Thermogenesis; Tissues; Untranslated RNA; Vascular Endothelial Growth Factors; Vascularization; Waste Products; Zinc Fingers; angiogenesis; base; cadherin 5; cell growth; cell motility; conditional knockout; diabetic; disorder risk; endothelial dysfunction; glucose tolerance; improved; in vivo; inhibitor/antagonist; insight; insulin sensitivity; insulin tolerance; knock-down; miRNA expression profiling; mouse model; non-diabetic; obesity development; overexpression; programs; promoter; response; subcutaneous; transcriptome sequencing; uncoupling protein 1

PROJECT SUMMARY/ABSTRACT White and brown adipose tissues are highly vascularized organs, capable of plasticity based on metabolic demands and energy expenditure. Critical gaps remain in our understanding of how angiogenesis impacts adipose tissue dysfunction and overall metabolism. MicroRNAs (miRs) are implicated in the regulation of the angiogenic response to pathophysiological stimuli. However, the role of miRs in regulating the angiogenic response in diet-induced insulin resistance is poorly understood. Using a miRNA-Seq approach, we identified that miR-409-3p expression was significantly increased in endothelial cells (ECs) of brown adipose tissue (BAT) of diet-induced obese (DIO) mice and in human diabetic plasma samples compared to non-diabetic patients. Overexpression of miR-409-3p markedly inhibited EC growth and migration, whereas miR-409-3p inhibition had the opposite effects. Preliminary studies indicate that miR-409-3p targets the 3'UTRs of Zinc Finger E-box binding Homeobox 1 (ZEB1) and Mitogen-activated protein kinase kinase kinase kinase 3 (MAP4K3). SiRNA knockdown of ZEB1 or MAP4K3 expression in ECs phenocopied the effects of miR-409-3p overexpression and significantly decreased EC proliferation and migration. 3T3-L1 cells or human skin fat organoids co-cultured with conditioned media from ECs overexpressing miR-409-3p had decreased expression of BAT markers (UCP1, Cidea), whereas conditioned media from ECs deficient in miR-409-3p markedly increased expression of BAT gene expression. Mechanistically, Placental Growth Factor (PLGF), secreted from ECs in response to miR-409-3p inhibition promoted 3T3-L1 differentiation towards brown-like adipocytes as measured by increased UCP1 expression. Systemic delivery of LNA-anti-miR-409-3p inhibitor to DIO mice significantly increased angiogenesis as measured by CD31 staining, UCP-1 expression in BAT and subcutaneous white adipose tissue (sWAT), while improving glucose and insulin tolerance and energy expenditure. Therefore, we hypothesize that miR-409-3p serves as a critical regulator of EC growth and angiogenesis in adipose tissue and may improve metabolic dysfunction in DIO. Thus, in Aim1 we will investigate the molecular mechanisms by which miR-409-3p regulates EC growth and angiogenesis. In Aim2, we will delineate the mechanisms by which miR-409-3p in ECs regulates browning in adipose tissues. Finally, in Aim3, we will explore the effect of miR-409-3p neutralization in the vasculature of adipose tissues and development of DIO and insulin resistance in mice. Successful completion of these studies will shed insights on the regulatory role of miR-409-3p between impaired angiogenesis in diet-induced obesity and adipose tissue dysfunction, an effect that could be exploited for therapeutic intervention in obesity-induced insulin resistance. !

项目概要/摘要 白色和棕色脂肪组织是高度血管化的器官，能够基于 代谢需求和能量消耗。我们对血管生成如何发生的理解仍存在关键差距 影响脂肪组织功能障碍和整体代谢。 MicroRNA (miR) 参与调控 血管生成对病理生理刺激的反应。然而，miRs 在调节 饮食引起的胰岛素抵抗中的血管生成反应尚不清楚。 使用 miRNA-Seq 方法，我们发现 miR-409-3p 表达在 饮食诱导肥胖 (DIO) 小鼠和人类糖尿病患者棕色脂肪组织 (BAT) 的内皮细胞 (EC) 血浆样本与非糖尿病患者的比较。 miR-409-3p 过表达显着抑制 EC 生长和迁移，而 miR-409-3p 抑制则具有相反的效果。初步研究表明 miR-409-3p 靶向锌指 E-box 结合同源框 1 (ZEB1) 和丝裂原激活的 3'UTR 蛋白激酶激酶激酶 3 (MAP4K3)。 EC 中 ZEB1 或 MAP4K3 表达的 SiRNA 敲低 表型复制了 miR-409-3p 过表达的影响并显着降低了 EC 增殖和 迁移。 3T3-L1 细胞或人皮肤脂肪类器官与来自 EC 的条件培养基共培养 过表达 miR-409-3p 会降低 BAT 标记物（UCP1、Cidea）的表达，而条件化 来自缺乏 miR-409-3p 的 EC 的培养基显着增加了 BAT 基因的表达。 从机制上讲，胎盘生长因子（PLGF）是 EC 响应 miR-409-3p 抑制而分泌的 通过增加 UCP1 表达来测量，促进 3T3-L1 向棕色脂肪细胞分化。 向 DIO 小鼠全身递送 LNA-抗 miR-409-3p 抑制剂可显着增加血管生成 通过 CD31 染色测量 BAT 和皮下白色脂肪组织 (sWAT) 中的 UCP-1 表达，同时 改善葡萄糖和胰岛素耐受性以及能量消耗。因此，我们假设 miR-409-3p 作为脂肪组织中 EC 生长和血管生成的关键调节因子，并可能改善代谢 DIO 功能障碍。因此，在 Aim1 中，我们将研究 miR-409-3p 的分子机制 调节 EC 生长和血管生成。在 Aim2 中，我们将描述 miR-409-3p 在 ECs 调节脂肪组织的褐变。最后，在Aim3中，我们将探讨miR-409-3p的作用 小鼠脂肪组织脉管系统的中和以及 DIO 和胰岛素抵抗的发展。 这些研究的成功完成将深入了解 miR-409-3p 之间的调节作用 饮食引起的肥胖和脂肪组织功能障碍中血管生成受损，这是可以利用的效应 用于肥胖引起的胰岛素抵抗的治疗干预。 ！