CXCR4介导巨噬细胞自噬在下肢缺血后炎症的作用及机制

Inflammation response is involved in tissue injury and repair after hindlimb ischemia. Post-ischemic inflammation is mostly rely on the infiltration and activation of macrophage. Recent studies exhibited that macrophage can suppress inflammatory response through cell autophagy. However, little is known about the molecular mechanism of autophagy in such processes. CXCR4 is a key chemokine receptor for leukocyte recruitment and inflammation regulation. our preliminary data also showed that knockout of cxcr4 in macrophage can promote inflammatory response both in vivo and in vitro through impaired autophagy. Furthermore, macrophage CXCR4 deletion lead to impaired blood flow perfusion in ischemic limb. Therefore, based on our previous works, our project will evaluate the effect of CXCR4 on macrophage autophagy after acute hindlimb ischemia and testify the control of post ischemic inflammatory response required by autophagy pathway by various molecular biology methods and CXCR4 conditional knock-out mice.The launching of this project is to clarify a new mechanism for the macrophage autophagy in regulation of post-ischemic inflammation and to explore how autophagy was involved in modulating inflammatory signaling pathway, finally providing new strategies and potential intervention targets for the treatment of peripheral artery disease.

炎症参与下肢缺血后损伤及修复。缺血后的炎症反应主要是由巨噬细胞的浸润和激活引起的。最近研究表明自噬在调节巨噬细胞促炎能力中发挥重要作用，然而其具体作用机制仍不清楚。CXCR4对于炎症细胞募集、功能调控极为重要。我们的前期工作显示CXCR4敲除使巨噬细胞自噬受损，从而增强其促炎能力，使小鼠缺血下肢炎症因子增多，影响下肢血流恢复。本项目拟采用CXCR4条件性敲除小鼠并结合多种分子生物学手段，通过体内外实验探讨CXCR4调控巨噬细胞自噬的作用，并深入研究自噬调控炎症反应的信号通路，最终为治疗外周血管病提供新的策略和潜在的干预靶点。

背景：血管新生对于缺血引起的组织损伤和改善血流恢复至关重要。缺血后血管新生的调控和潜在的分子机制尚未完全揭示。趋化因子受体CXCR4在白细胞募集和炎症调节中起关键作用。但是，关于CXCR4如何调节血管再生的了解较少。.目的：探索CXCR4阳性的巨噬细胞在缺血后血管新生中的作用及其潜在的分子机制。.方法和结果：通过股动脉结扎后野生型（WT）小鼠后肢，发现CXCR4阳性的炎症细胞在缺血下肢中募集。在骨髓特异性CXCR4基因敲除（MKO）小鼠的缺血肢体中观察到更好的血液再灌注和更高的毛细血管密度。此外，从MKO小鼠到WT小鼠的骨髓移植改善了WT小鼠的血液再灌注，反之亦然。此外，巨噬细胞耗竭后，WT和MKO小鼠的血流恢复均受损。在体外，通过小干扰RNA（siCXCR4）沉默CXCR4的巨噬细胞相对于阴性对照（siNC）的条件培养基促进了人脐带静脉内皮细胞（HUVEC）的管腔形成。并且从机制上讲，Lps激活了AKT-mTOR信号传导并下调了巨噬细胞的自噬,促进了促炎因子的分泌，siCXCR4抑制了这一作用。通过抑制siNC巨噬细胞中的AKT或激活siCXCR4的巨噬细胞中的AKT，我们证明了AKT介导的自噬是CXCR4调节HUVEC中血管生成所必需的。.结论：总之，我们的数据表明CXCR4 +巨噬细胞通过激活AKT和抑制自噬而成为血管生成的负调节剂，表明CXCR4构成了缺血性血管疾病的新型分子靶标。