Effect of Dicer-mediated let-7 expression in inflammatory macrophages on atherosclerosis

动脉粥样硬化是心肌梗死和中风的主要原因，因此是全球死亡的重要原因。在细胞水平上，不同类型的免疫细胞，主要是巨噬细胞，聚集在动脉壁上，吸收脂蛋白，发育成泡沫细胞。然而，巨噬细胞也可以分化为亲炎性和抗炎性表型，这在先天免疫中起着关键作用。RNase Dicer产生调节巨噬细胞极化的microrna。我们发现Dicer通过产生miR-10a和let-7调节抗炎巨噬细胞和泡沫细胞的线粒体功能，从而减少动脉粥样硬化。此外，我们的初步结果表明，Dicer调节炎症性巨噬细胞中的基因表达，从而促进死亡细胞的清除（efferocytosis），减少细胞死亡（apoptosis）。本课题旨在研究Dicer在动脉粥样硬化炎性巨噬细胞中的作用。我们假设Dicer在炎性巨噬细胞中产生let-7 mirna，通过线粒体蛋白和efferocytosis受体调节efferocytosis和凋亡，从而减少动脉粥样硬化。这一假设将通过使用转基因动脉粥样硬化小鼠模型的三个工作包来解决。在第一部分和第二部分，我们将使用一种新的efferocytosis实验来研究炎性巨噬细胞中Dicer和let-7对efferocytosis的影响。此外，我们将使用共聚焦显微镜进行4D活斑块成像，以评估巨噬细胞中Dicer和let-7对efferocytosis的影响。此外，我们将研究Dicer和let-7如何影响炎性巨噬细胞的线粒体动力学。在提案的第二部分，我们还将研究let-7如何通过确定Dicer调节基因中功能相关的结合位点来介导炎性巨噬细胞中Dicer的功能。我们将使用靶点阻滞剂特异性地抑制let-7与单个潜在靶点之间的相互作用。因此，我们将在第三部分分析这些相互作用对efferocytosis、凋亡、线粒体动力学和动脉粥样硬化的影响。该项目的结果旨在获得炎性巨噬细胞在动脉粥样硬化中的作用的新见解。失去像let-7这样的mirna可能会损害它们的功能，从而促进斑块的形成。

Atherosclerosis is a main cause of myocardial infarction and stroke and thus contributes crucially to worldwide mortality. At a cellular level, different types of immune cells, mainly macrophages, accumulate in the arterial wall, where they take up lipoproteins and develop into foam cells. However, macrophages can also polarize into a pro- and anti-inflammatory phenotype, which play key roles in innate immunity. The RNase Dicer produces microRNAs that regulate macrophage polarization. We found that Dicer regulates mitochondrial function in anti-inflammatory macrophages and foam cells by producing miR-10a and let-7, thus reducing atherosclerosis. Moreover, our preliminary results show that Dicer regulates gene expression in inflammatory macrophages, which promotes the removal of dead cells (efferocytosis) and reduces cell death (apoptosis). This proposal aims to study the role of Dicer in inflammatory macrophages in atherosclerosis. We hypothesize that Dicer's production of let-7 miRNAs in inflammatory macrophages reduces atherosclerosis by regulating efferocytosis and apoptosis through mitochondrial proteins and receptors for efferocytosis. This hypothesis will be addressed by three working packages using genetically modified mouse models of atherosclerosis. In the first and second parts, we will investigate the effect of Dicer and let-7 in inflammatory macrophages on efferocytosis using a novel efferocytosis assay. Moreover, we will perform 4D live plaque imaging using confocal microscopy to evaluate the effect of Dicer and let-7 in macrophages on efferocytosis. In addition, we will study how Dicer and let-7 affect mitochondrial dynamics in inflammatory macrophages. In the second part of the proposal, we will also study how let-7 mediates the function of Dicer in inflammatory macrophages by determining functionally relevant binding sites in the Dicer-regulated genes. We will use target site blockers to specifically inhibit the interaction between let-7 and individual potential targets. Thus, we will analyze in the third part the effect of these interactions on efferocytosis, apoptosis, mitochondrial dynamics, and atherosclerosis. The results of this project aim to gain novel insights into the role of inflammatory macrophages in atherosclerosis. Losing miRNAs like let-7 may impair their function and thus promote plaque formation.