Notch 信号调控巨噬细胞极化参与增生性瘢痕形成的作用及机制

Hypertrophic scar form most often at the sites of trama and burns. The disease incidence rate is high in our country and there has been still no effective prevention and treatment methods. We found that the inhibition of angiogenesis at the early phase of wound healing can inhibit the formation of hypertrophic scar, but the initiative factors of continuous vascularization and scar hyperplasia was not clear. We also found that there were a large number of macrophages infiltrating in hypertrophic scar. The high expression of Notch signal Moleculars were also detected and the inhibition of Notch signaling can reduce scar formation. Because the Notch signal is the key signal regulating macrophage polarization, and macrophages can produce various key factors of vascularization and fibrosis, we propose that the Notch signal might initiate the angiogenesis and fibrosis through regulating macrophage polarization. This project aims to collect human wound / scar specimens in different periods of wound healing, and analysis of relationships between the macrophage polarization and scar hyperplasia and the level of Notch signal expression. We also aims to regulate Notch signal in coculture model of macrophages and endothelial cell / fibroblast and to reveal the molecular mechanism use macrophage specific Notch signal knockout mice wound model. This study is expected to further clarify the pathogenesis of hypertrophic scar and to provide a new target for its prevention and treatment.

增生性瘢痕常发生于烧创伤后，我国发病率高，尚无有效的防治方法。我们发现在瘢痕形成早期抑制血管生成能够抑制瘢痕增生，但瘢痕持续血管化及增生的始动因素还不清楚。我们前期还发现增生性瘢痕中大量巨噬细胞浸润，同时存在Notch信号高表达，抑制Notch信号可以减轻瘢痕增生。由于Notch信号是调节巨噬细胞极化的关键信号，而巨噬细胞可产生多种促血管化及纤维化的关键因子，因此我们提出Notch信号可能通过调控巨噬细胞极化启动瘢痕形成中的血管新生和纤维化。本项目拟收集不同时期的人创面/瘢痕标本，进一步分析Notch信号表达与巨噬细胞极化和瘢痕增生的关系；在巨噬细胞与内皮细胞/成纤维细胞共培养模型中调节Notch信号，及利用巨噬细胞特异性敲除Notch信号的小鼠创面模型，研究Notch信号调控巨噬细胞对瘢痕血管化及纤维化的作用，揭示其分子机制。本研究有望进一步阐明增生性瘢痕的发生机制，为其防治提供新靶点。

增生性瘢痕是整形外科、烧伤科、皮肤科常见病，其发病率高，烧伤后增生性瘢痕发病率高达70%。目前的治疗措施效果均不理想，究其根本原因是对增生性瘢痕的发病机制不够了解。巨噬细胞可产生多种促血管化及纤维化的关键因子，可能是引起瘢痕组织血管化和纤维增生的始动因素。然而不同亚群的巨噬细胞功能不同，Notch信号可以改变巨噬细胞的极化状态。为验证Notch信号通路是否通过调控巨噬细胞极化，从而启动瘢痕形成中的血管新生和纤维化，我们收集了临床不同时期创面和增生性瘢痕组织128例，组装成组织芯片，对其Notch信号和M1/M2型巨噬细胞进行了染色和PCR鉴定，发现创面早期以M1型巨噬细胞为主，晚期及瘢痕组织以M2型巨噬细胞为主，Notch信号活化程度与M1/M2型巨噬细胞比例呈显著正相关。进一步采用体外实验分离培养人及小鼠巨噬细胞和成纤维细胞，共培养模型验证了Notch信号调控巨噬细胞极化抑制成纤维细胞I型胶原的产生。此外，我们还发现Notch信号本身也可以抑制成纤维细胞I型胶原、a-SMA、TGFβ-1等促纤维化因子的表达。同时，我们构建了小鼠增生性瘢痕模型。我们还对M2型巨噬细胞亚型进行了进一步的分析和研究，发现转录调节因子c-MAF上调IL-10的表达，IL-10诱导M0型巨噬细胞向M2c型巨噬细胞极化，并且维持M2c型巨噬细胞的持续存在和TGF-β1的持续分泌，促进成纤维细胞向肌成纤维细胞转化，从而造成增生性瘢痕的形成。本课题实施过程中所组装的临床标本组织芯片对于进一步研究创面愈合和增生性瘢痕发病机制提供了优质可靠的材料。我们发现的Notch信号通路、c-MAF和IL-10为增生性瘢痕的防治提供了潜在的治疗靶点，具有重要的临床转化意义和应用前景。