Cardiac Macrophages as Disease Drivers in Chronic Ischemic Heart Failure

衰竭心脏中巨噬细胞的扩张导致组织损伤，并被认为是导致进展的原因之一。 心力衰竭(HF)。衰竭心脏中巨噬细胞扩张的主要因素是自我更新的细胞。 并且不依赖于血液单核细胞池而增殖(因此是C-C趋化因子受体2[CCR2]-)。 然而，这种局部来源的巨噬细胞在慢性左心室(LV)发病机制中的作用 人们对重塑知之甚少。正常心脏有巨噬细胞表达CD206(Mrc1)，以及 基因标记YM1、Fizz1和Arg1，主要是CCR2-。这些CD206巨噬细胞可以被激活 Th2细胞因子，如白介素4/白介素13。此外，分泌的FIZZ1对肌成纤维细胞有贡献 在其他疾病模型中的激活和纤维化。我们的实验数据表明，表达IL-1的CD206巨噬细胞 4受体(IL-4ra)在心力衰竭时显著升高，提示巨噬细胞IL-4/IL-4ra信号转导 促进左室重构和纤维化，部分是通过FIZZ1。因此，我们假设心脏CD206 表达IL-4ra和Fizz1的巨噬细胞是慢性左心室不良重构的先天免疫介质 HF和治疗性免疫调节的关键靶点。三个目标将检验这一假设。在目标1中，在小鼠身上 冠状动脉结扎模型，使用流式细胞术、细胞分类、单细胞RNA测序(ScRNAseq)以及 免疫组织化学，我们将全面定义心脏CD206巨噬细胞的病理变化 在HF中，包括IL-4ra水平和下游信号转导，FIZZ1表达，以及体内细胞丰度， 增殖、周转和吞噬能力。使用scRNAseq，我们将定义新的功能亚群 在小鼠和人类衰竭心脏中CD206巨噬细胞的转录水平。在目标2中，我们将 利用CD45.2诱导的髓系细胞特异性研究CD206 IL-4ra巨噬细胞在心衰发病中的作用 IL-4Ra基因敲除小鼠，慢性心力衰竭时髓系IL-4Ra的缺失及对LV的远期影响 重塑、炎症和纤维化。建立衰竭心脏CD206 IL-4ra巨噬细胞的充分性 诱导组织损伤，我们将过继转移分选的具有完整或缺失IL-2的心脏CD206巨噬细胞 4Ra从心衰小鼠心肌内注入幼稚CD45.1小鼠，6周后评价左室重构。至 评估FIZZ1的作用，我们将类似地将M[IL-4]极化的骨髓巨噬细胞从野生型 和Fizz1-/-老鼠。在目标3中，我们将测试潜在的可翻译疗法来拮抗CD206 IL-4ra 巨噬细胞和减轻慢性左室重构，包括针对IL-1的系统反义寡核苷酸。 4Ra和Fizz1，抗IL-4中和抗体，小分子cFMS激酶抑制剂GW2580。我们会 测量这些疗法对左室重构、心脏巨噬细胞、纤维化和趋化因子的影响。 细胞因子的表达。这些研究将进一步加深我们对心脏先天性免疫基础的理解。 缺血性心力衰竭的炎症，提供了对巨噬细胞IL-4依赖的信号转导和Fizz1在 研究左室重构的发病机制，为心衰的细胞类型特异性免疫调节寻找新的途径。

Macrophage expansion in the failing heart induces tissue injury and is thought to contribute to the progression of heart failure (HF). The main contributors to macrophage expansion in the failing heart are cells that self-renew and proliferate independent of the blood monocyte pool (and hence are C-C chemokine receptor 2 [CCR2]–). However, the role of such locally-sourced macrophages in the pathogenesis of chronic left ventricular (LV) remodeling is poorly understood. The normal heart harbors macrophages expressing CD206 (Mrc1), along with the gene markers Ym1, Fizz1, and Arg1, that are primarily CCR2–. These CD206+ macrophages can be activated by Th2 cytokines such as interleukin(IL)-4/IL-13. Moreover, secreted FIZZ1 contributes to myofibroblast activation and fibrosis in other disease models. Our pilot data indicate that CD206+ macrophages expressing IL- 4 receptora (IL-4Ra) robustly increase in the failing heart, and suggest that macrophage IL-4/IL-4Ra signaling promotes LV remodeling and fibrosis, in part through FIZZ1. Hence, we hypothesize that cardiac CD206+ macrophages expressing IL-4Ra and Fizz1 are innate immune mediators of adverse LV remodeling in chronic HF, and key targets for therapeutic immunomodulation. Three Aims will test this hypothesis. In Aim 1, in a murine coronary ligation model, using flow cytometry, cell sorting, single cell RNA sequencing (scRNAseq), and immunohistochemistry, we will comprehensively define pathological alterations in cardiac CD206+ macrophages in HF, including IL-4Ra levels and downstream signaling, FIZZ1 expression, and in vivo cell abundance, proliferation, turnover and phagocytic capacity. Using scRNAseq, we will define novel functional subpopulations of CD206+ macrophages at the transcriptional level in both murine and human failing hearts. In Aim 2, we will establish the pathogenetic role of CD206+IL-4Ra+ macrophages in HF using CD45.2 inducible myeloid-specific IL-4Ra knockout mice, deleting myeloid IL-4Ra during chronic HF, and assessing the late effects on LV remodeling, inflammation, and fibrosis. To establish sufficiency of failing heart CD206+IL-4Ra+ macrophages to induce tissue injury, we will adoptively transfer sorted cardiac CD206+ macrophages with intact or deleted IL- 4Ra from HF mice into naïve CD45.1 mice via intramyocardial injection and assess LV remodeling 6 w later. To assess the role of FIZZ1, we will similarly transfer M[IL-4] polarized bone marrow macrophages from wild-type and Fizz1-/- mice. In Aim 3, we will test potentially translatable therapies to antagonize CD206+IL-4Ra+ macrophages and alleviate chronic LV remodeling, including systemic anti-sense oligonucleotides against IL- 4Ra and Fizz1, anti-IL-4 neutralizing antibody, and GW2580, a small molecule cFMS kinase inhibitor. We will measure the effects of these therapies on LV remodeling, cardiac macrophages, fibrosis, and chemokine/ cytokine expression. These studies will further our understanding of the innate immune basis for cardiac inflammation in ischemic HF, provide novel insights into macrophage IL-4-dependent signaling and Fizz1 in the pathogenesis of LV remodeling, and identify new approaches for cell type-specific immunomodulation in HF.