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

项目摘要/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。在我们对血管生成的理解中仍然存在严重的差距 影响脂肪组织功能障碍和整体代谢。MicroRNAs(MiRs)与调控有关 对病理生理刺激的血管生成反应。然而，MIR在调节 饮食诱导的胰岛素抵抗的血管生成反应还知之甚少。 使用miRNA-Seq方法，我们发现miR-409-3p在 饮食诱导肥胖(DIO)小鼠和人糖尿病棕色脂肪组织(BAT)内皮细胞的观察 将血浆样本与非糖尿病患者进行比较。MiR-409-3p过表达显著抑制EC 抑制miR-409-3p的作用相反。初步研究表明， MIR-409-3P靶向锌指E盒结合同源框1(ZEB1)的3‘UTRs和丝裂原激活 蛋白激酶3(MAP4K3)。SiRNA抑制内皮细胞ZEB1或MAP4K3的表达 表型复制miR-409-3p过表达并显著抑制EC增殖和 迁移。3T3-L1细胞或人皮肤脂肪类物质与内皮细胞条件培养液共培养 过度表达miR-409-3p降低了BAT标志物(UCP1、CIDA)的表达，而条件性 缺失miR-409-3p的内皮细胞培养上清液可显著增加BAT基因的表达。 从机制上讲，内皮细胞在miR-409-3p抑制下分泌的胎盘生长因子(PLGF) 通过增加UCP1的表达，促进3T3-L1向棕色样脂肪细胞分化。 系统地给DIO小鼠注射LNA-抗miR-409-3p抑制剂显著促进血管生成 CD31染色检测BAT和皮下白色脂肪组织中UCP-1的表达，而 改善血糖、胰岛素耐量和能量消耗。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，可能会改善代谢 DIO的功能障碍。因此，在Aim1中，我们将研究miR-409-3p 调节EC的生长和血管生成。在AIM2中，我们将描述miR-409-3p在 ECS调节脂肪组织中的褐变。最后，在Aim3中，我们将探索miR-409-3P的作用 脂肪组织血管系统的中和与小鼠DIO和胰岛素抵抗的发展。 这些研究的成功完成将有助于深入了解miR-409-3p在 饮食诱导的肥胖和脂肪组织功能障碍的血管生成受损，这一效应可以加以利用 对肥胖引起的胰岛素抵抗进行治疗干预。 好了！

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. !