Wang Qiwei, Ren Jun, Morgan Stephanie, Liu Zhenjie, Dou Changlin, Liu Bo
Division of Vascular Surgery, Department of Surgery, University of Wisconsin-Madison, Wisconsin, United States of America.
Luye Pharma Group, Yantai, China.
PLoS One. 2014 Mar 14;9(3):e92053. doi: 10.1371/journal.pone.0092053. eCollection 2014.
In abdominal aortic aneurysm (AAA), macrophages are detected in the proximity of aortic smooth muscle cells (SMCs). We have previously demonstrated in a murine model of AAA that apoptotic SMCs attract monocytes and other leukocytes by producing MCP-1. Here we tested whether infiltrating macrophages also directly contribute to SMC apoptosis.
Using a SMC/RAW264.7 macrophage co-culture system, we demonstrated that MCP-1-primed RAWs caused a significantly higher level of apoptosis in SMCs as compared to control macrophages. Next, we detected an enhanced Fas ligand (FasL) mRNA level and membrane FasL protein expression in MCP-1-primed RAWs. Neutralizing FasL blocked SMC apoptosis in the co-culture. In situ proximity ligation assay showed that SMCs exposed to primed macrophages contained higher levels of receptor interacting protein-1 (RIP1)/Caspase 8 containing cell death complexes. Silencing RIP1 conferred apoptosis resistance to SMCs. In the mouse elastase injury model of aneurysm, aneurysm induction increased the level of RIP1/Caspase 8 containing complexes in medial SMCs. Moreover, TUNEL-positive SMCs in aneurysmal tissues were frequently surrounded by CD68(+)/FasL(+) macrophages. Conversely, elastase-treated arteries from MCP-1 knockout mice display a reduction of both macrophage infiltration and FasL expression, which was accompanied by diminished apoptosis of SMCs.
Our data suggest that MCP-1-primed macrophages are more cytotoxic. MCP-1 appears to modulate macrophage cytotoxicity by increasing the level of membrane bound FasL. Thus, we showed that MCP-1-primed macrophages kill SMCs through a FasL/Fas-Caspase8-RIP1 mediated mechanism.
在腹主动脉瘤（AAA）中，可在主动脉平滑肌细胞（SMC）附近检测到巨噬细胞。我们先前在AAA小鼠模型中证明，凋亡的SMC通过产生MCP-1吸引单核细胞和其他白细胞。在此，我们测试了浸润的巨噬细胞是否也直接促成SMC凋亡。
使用SMC/RAW264.7巨噬细胞共培养系统，我们证明与对照巨噬细胞相比，经MCP-1预处理的RAW细胞导致SMC中凋亡水平显著更高。接下来，我们检测到经MCP-1预处理的RAW细胞中Fas配体（FasL）mRNA水平和膜FasL蛋白表达增强。中和FasL可阻断共培养中的SMC凋亡。原位邻近连接分析表明，暴露于预处理巨噬细胞的SMC含有更高水平的包含受体相互作用蛋白-1（RIP1）/半胱天冬酶8的细胞死亡复合物。沉默RIP1赋予SMC抗凋亡能力。在小鼠弹性蛋白酶损伤动脉瘤模型中，动脉瘤诱导增加了中膜SMC中包含RIP1/半胱天冬酶8复合物的水平。此外，动脉瘤组织中TUNEL阳性的SMC经常被CD68(+)/FasL(+)巨噬细胞包围。相反，来自MCP-1基因敲除小鼠的弹性蛋白酶处理的动脉显示巨噬细胞浸润和FasL表达均减少，同时SMC凋亡减少。
我们的数据表明，经MCP-1预处理的巨噬细胞具有更高的细胞毒性。MCP-1似乎通过增加膜结合FasL的水平来调节巨噬细胞的细胞毒性。因此，我们表明经MCP-1预处理的巨噬细胞通过FasL/Fas-半胱天冬酶8-RIP1介导的机制杀死SMC。
