Macrophage Circadian Clock Disruption and Inflammation in Heart Failure

单核细胞和巨噬细胞基本上来源于骨髓和脾脏-促炎 小鼠中的Ly 6Chi单核细胞（人中的CD 14 hi细胞）和C-C趋化因子受体2（CCR 2）+单核细胞衍生的 巨噬细胞（MDM）-在慢性心脏病组织损伤和左心室（LV）重塑中起重要作用 失败（HF），表明这些细胞可能代表免疫调节的治疗靶点。重要的是， 单核细胞和MDM在多个功能参数中表现出昼夜变化，这在很大程度上是由于 细胞自主分子钟，由CLOCK/BMAL 1转录复合物调节，受试者 与Rev-erba有关的自动调节回路值得注意的是，无论是单核细胞和 MDM在HF中被破坏，其与炎症和疾病进展的机制联系完全未知。 我们的初步研究表明，单核细胞和MDM时钟中断是HF的特征， 单核细胞/巨噬细胞Bmal 1（以及随后的Rev-erba/B）缺失与免疫调节的上调有关。 激活剂E4 bp 4和LV重构加重。基于这些数据，我们假设 单核细胞/MDM时钟在HF中功能失调，导致病理性炎症和心脏重塑， REV-ERBa连接和E4 BP 4依赖的方式，并且时钟校正是 免疫调节三个目标将检验这一假设。在目标1中，我们将定义单核细胞中的改变， 使用小鼠冠状动脉结扎模型测定HF中的昼夜节律钟，并测试单核细胞/巨噬细胞是否时钟 骨髓特异性Bmal 1敲除（MBK）小鼠中的破坏加剧了炎症，先天免疫扩张， 和左心室重构。我们还将测量CD 14 hi单核细胞中的炎症和时钟基因， 人类HF在目的2中，我们将描述单核细胞定位的E4 bp 4的作用， 炎症转录因子和Rev-erba的直接靶点，在HF中通过评估炎症和LV 在骨髓特异性E4 bp 4敲除（MEK）小鼠中冠状动脉结扎后的重构。我们还将评估 单核细胞E4 BP 4抑制挽救了单核细胞/巨噬细胞观察到的加重的LV重构 通过诱导骨髓特异性Bmal 1/E4 bp 4双敲除小鼠中的HF来破坏生物钟。在目标3中，我们 通过测试，从机制上确定靶向生物钟作为HF治疗方法的潜力 用SR9009（一种合成的REV-ERBa/B激动剂）治疗是否有利地调节单核细胞和MDM 并改善或逆转已建立HF的野生型小鼠的LV重塑，然后在MEK HF小鼠中， 确定SR9009的作用是否需要单核细胞/巨噬细胞E4 bp 4。我们还将评估前 人HF CD 14 hi单核细胞对激动剂的体内活化反应。这些研究将进一步促进我们的 了解巨噬细胞生物钟的变化如何调节炎症和疾病 HF的进展，并使用遗传和药理学方法测试免疫调节的新方法 纠正由时钟中断引起的病理变化的策略。

Monocytes and macrophages fundamentally originating from the bone marrow and spleen - pro-inflammatory Ly6Chi monocytes in mice (CD14hi cells in humans) and C-C chemokine receptor 2 (CCR2)+ monocyte-derived macrophages (MDMs) - contribute importantly to tissue injury and left ventricular (LV) remodeling in chronic heart failure (HF), suggesting that these cells may represent therapeutic targets for immunomodulation. Importantly, monocytes and MDMs exhibit circadian variation in multiple functional parameters, in large part due to a robust cell-autonomous molecular clock, which is regulated by the CLOCK/BMAL1 transcriptional complex and subject to an autoregulatory loop involving Rev-erba. Notably, whether the intrinsic circadian clock in monocytes and MDMs is disrupted in HF and its mechanistic link to inflammation and disease progression is entirely unknown. Our pilot studies suggest that monocyte and MDM clock disruption is characteristic of HF, and that monocyte/macrophage Bmal1 (and subsequent Rev-erba/b) loss is associated with upregulation of the immune activator E4bp4, and aggravation of LV remodeling. Based on these data, we hypothesize that the monocyte/MDM clock is dysfunctional in HF, leading to pathological inflammation and cardiac remodeling in a REV-ERBa-linked and E4BP4-dependent manner, and that clock correction is a key molecular target for immunomodulation. Three Aims will test this hypothesis. In Aim 1, we will define alterations in the monocyte circadian clock in HF using a murine coronary ligation model, and test whether monocyte/macrophage clock disruption in myeloid-specific Bmal1 knockout (MBK) mice exacerbates inflammation, innate immune expansion, and LV remodeling during HF. We will also measure inflammatory and clock genes in CD14hi monocytes from humans with HF. In Aim 2, we will delineate the role of monocyte-localized E4bp4, a clock-controlled inflammatory transcription factor and direct target of Rev-erba, in HF by assessing inflammation and LV remodeling after coronary ligation in myeloid-specific E4bp4 knockout (MEK) mice. We will also evaluate whether monocyte E4BP4 suppression rescues the aggravated LV remodeling observed with monocyte/macrophage clock disruption by inducing HF in myeloid-specific Bmal1/E4bp4 double knockout mice. In Aim 3, we will mechanistically establish the potential of targeting the circadian clock as a therapeutic approach in HF, by testing whether treatment with SR9009, a synthetic REV-ERBa/b agonist, favorably modulates monocytes and MDMs and ameliorates or reverses LV remodeling in wild-type mice with established HF, and then in MEK HF mice to determine whether the effects of SR9009 require monocyte/macrophage E4bp4. We will also evaluate the ex vivo activation responses of human HF CD14hi monocytes to the agonist. These studies will further our understanding of how changes in the macrophage circadian clock modulate both inflammation and disease progression in HF, and test novel approaches to immunomodulation using genetic and pharmacological strategies to correct the pathological changes induced by clock disruption.