巨噬细胞Ly6Clow亚群在诱导心肌梗死后炎症反应向瘢痕修复转化过程中的作用和机制

Cardiac healing following acute myocardial infarction(MI) needs a transition from inflammatory response to scar formation. Because of the catastrophic consequences of excessive or prolonged inflammation on tissue architecture and organ function, repair of injured tissues requires timely activation of endogenous inhibitory pathways that restrain and suppress pro-inflammatory signals. Monocytes/macrophages are cirtical modulating cells of the inflammatory response. Two subpopulations of macrophages, defined as Ly6Chigh and Ly6Clow cells, have been found participating in this modulation with distinct roles. Ly6Chigh cells exhibit pro-inflammatory properties, while the Ly6Clow subset shows significant activities on inflammatory suppression, angiogenic enhancement and collagen synthesis, suggesting that Ly6Clow macrophages may exert proresolving and protective effects on injured tissues. However,the mechnism is unknown. The present study will investigate the modulating function of Ly6Clow macrophages following myocardial infarction, by examining gene expression levels of pro-and anti-inflammatory factors in Ly6Clow cells isolated from infarcted areas and the association between Ly6Clow cell infiltration and tissue collagen expression in infarcts. Specific clearance of Ly6Clow by clodronate liposomes will be performed to illustrate the effect of Ly6Clow macrophages on cardiac remodeling after myocardial infarction. Toll-Like Receptor (TLR) and Interleukin (IL)-1 signaling have been proved to be critically involved in the post-infarction negative inflammatory response and in the pathogenesis of cardiac remodeling. Our prelimilary data showed that Interleukin-1 receptor-associated kinase (IRAK)-M and type II IL1 receptor (IL1RII),known as endogenous inhibitory moleculars of TLR/IL pathways,were upregulated in Ly6Clow cells after MI.The depletion of IRAK-M gene resulted in a reduced anti-inflammatory function of Ly6Clow cells. Therefore, we hypothesized that the inflammatory suppression of Ly6Clow macrophages may mediated by TLR/IL inhibitory signals. In the present study, IRAK-M gene knockout mice will be used to analyze the possible molecular pathways and mechnisms of Ly6Clow macrphages acting as both a negative regulator of inflammation and a positive mediator of scar formation. Our study will demonstrate for the first time the protective effect of Ly6Clow macrophages on infarcted hearts and may identify a specific cellular effector and mechanisms accellerating cardic repair. Understanding of inhibitory and proresolving pathways of Ly6Clow macrophages in the infarcted hearts is important for designing novel therapeutic strategies.

急性心肌梗死后心脏修复经历了从炎症反应到瘢痕形成的转化过程。过度的炎症反应导致瘢痕形成受阻，造成诸如心脏破裂或心脏重塑等灾难性后果。单核/巨噬细胞是心梗后重要的炎症调控细胞，研究显示巨噬细胞分为Ly6C高表达和低表达两个亚群。前者具有刺激炎症作用；后者抑制炎症反应、促进新生血管和胶原形成，提示Ly6Clow亚群具有促心脏修复作用，机制尚不清楚。本课题组前期研究发现IL-1通路的内源性炎症抑制因子如IRAK-M在Ly6Clow细胞上高表达,其基因缺陷造成该亚群细胞抑炎作用减弱，提出Ly6Clow亚群可能通过上述炎症抑制通路促进瘢痕修复的新假说。本研究拟从动物-细胞-分子水平，深入研究Ly6Clow细胞亚群在心肌梗死后炎症调控中的作用；分析内源性炎症抑制因子在Ly6Clow细胞中的表达和功能，阐明该群细胞的心脏保护作用，找到心梗后促心脏愈合的特异性靶细胞和相应机制，为改善心脏重塑提供新方向。

急性心肌梗死后心脏修复经历了从炎症反应到瘢痕形成的转化过程。过度的炎症反应将导致瘢痕形成受阻，造成诸如心脏破裂或心脏重塑等灾难性后果。单核/巨噬细胞是心肌梗死后重要的炎症调控细胞，研究显示巨噬细胞分为Ly6C 高表达和低表达两个亚群。前者具有促进炎症反应的作用；后者则抑制炎症、促进新生血管和胶原形成，提示Ly6Clow 细胞具有促心脏修复的作用，但机制尚不清楚。本课题组前期研究发现IL-1 通路的内源性炎症抑制因子如IRAK-M在心肌梗死后具有抑制炎症，减少心脏重塑的作用。心肌梗死后7天IRAK-M高表达，而心肌梗死后7天的巨噬细胞以Ly6Clow 亚群细胞为主，因而提出Ly6Clow 亚群可能通过激活IRAK-M促进瘢痕修复这一新假说。本研究成功建立了野生型和IRAK-M基因敲除小鼠的缺血再灌注模型，研究Ly6Clow 细胞亚群在心肌梗死后炎症调控中的作用：1)首先在野生型小鼠中明确了心肌梗死后7天IRAK-M的表达主要定位于巨噬细胞内；2)发现高表达IRAK-M的巨噬细胞数量在心肌梗死后7天较3天时增加9倍；3)为了进一步明确梗死心脏中巨噬细胞的亚类及其炎症特性，我们发现缺血再灌注后3天巨噬细胞以Ly6Chi为主，90%的Ly6Chi细胞IL1β为阳性，具有促炎作用。缺血再灌注后7天，Ly6Chi阳性细胞数量下降了90%, 而Ly6Clow细胞则增加了4倍；4) 为了进一步明确IRAK-M基因对Ly6Clow细胞的炎症/抗炎特性的影响，我们将IRAK-M基因敲除小鼠和野生型小鼠进行对比，发现Ly6Clow细胞表达具有抑炎特性的IL1－II型受体（IL1RII）水平为6.0%-9.07%，而Ly6Chi细胞表面则不表达IL1RII；5) 缺血再灌注后72小时，IRAK-M基因敲除小鼠心脏中Ly6Clow细胞数量较野生型小鼠显著降低，且这些Ly6Clow细胞更多的表现为IL1β阳性。提示IRAK-M的缺失使得具有炎症抑制作用的Ly6Clow细胞不仅数量上减少，而且炎性特征也向致炎方向转化。6)我们的研究首次发现，心肌梗死后小鼠心肌成纤维细胞也表达IRAK-M，提示IRAK-M也可能参与心肌梗死后成纤维细胞的心肌修复作用。本研究深入探讨了IRAK-M在巨噬细胞亚群中的表达和功能，阐述了该群细胞的心脏保护作用，提出心肌梗死后促心脏愈合的特异性靶细胞和可能机制。