LPS诱导巨噬细胞释放包含CXCL2外泌体趋化中性粒细胞激活氧化应激促进脓毒症相关性肝衰竭的机制研究

Exosomes are important mediators of cell-to-cell communication. Early recruitment of neutrophils is an important phenomenon in sepsis-associated liver failure. How the exosomes mediate the recruitment of neutrophils to the liver is rarely reported. Our previous work found that exosomes released by sepsis patients and LPS-stimulated macrophages contained high levels of CXCL2, and exosomes released by LPS-stimulated macrophages recruited neutrophils to liver and caused oxidative stress, but the underly mechanism is unknown. Therefore, we hypothesized that LPS stimulated the release of macrophage exosomes that contained CXCL2, which recruited neutrophil through CXCR2 and activated neutrophil PIP2/DAG/PKC/NADPH to promote the development of sepsis-associated liver failure. This study was to investigate the mechanism of LPS-induced macrophage exosomal CXCL2 chemotaxis of neutrophils by CXCR2 and increase neutrophil oxidative stress through CXCR2 activation in vivo and in vitro. This study provides new thougts and targets toward macrophage exosomes in the development of sepsis-associated liver failure.

外泌体是细胞间通讯的重要媒介。中性粒细胞的早期募集是脓毒症相关性肝衰竭的重要现象。外泌体如何影响中性粒细胞在肝脏募集鲜见报道。我们前期工作发现，脓毒症患者血浆外泌体和LPS刺激巨噬细胞释放的外泌体均表达高水平CXCL2，LPS刺激的巨噬细胞释放的外泌体可趋化中性粒细胞至肝脏并引起氧化应激水平增加，然而具体机制不明。因此，我们设想：LPS刺激巨噬细胞释放含CXCL2外泌体通过CXCR2趋化并激活中性粒细胞PIP2/DAG/PKC/NADPH促进脓毒症相关性肝衰竭的发展。本研究拟从体内、外层面探讨LPS诱导巨噬细胞CXCL2通过CXCR2趋化中性粒细胞及中性粒细胞CXCR2的激活引起氧化应激水平增加的分子机制。本研究为拓展巨噬细胞外泌体在脓毒症相关性肝衰竭的发生发展提供新思路和作用靶点。

胞外囊泡是细胞间通讯的重要媒介。中性粒细胞的早期募集是脓毒症相关性肝衰竭的重要现象。胞外囊泡如何影响中性粒细胞在肝脏募集鲜见报道。我们从体内、外探讨探讨LPS刺激巨噬细胞释放含CXCL2胞外囊泡趋化并激活中性粒细胞氧化应激促进脓毒症相关性肝衰竭的发展机制。体内研究发现：Raw264.7释放的EVs参与了中性粒细胞向肝脏的招募,LPS-Raw264.7 -EVs上的CXCL2在体内募集中性粒细胞，并通过CXCR2/PKC/NOX4途径激活中性粒细胞。体外研究发现：我们首先完成了Raw264.7细胞释放的EVs的表征鉴定，并在体外验证了LPS-巨噬细胞-EVs对中性粒细胞的趋化作用。此外，我们通过蛋白质芯片检测了LPS-Raw-264.7-EVs的细胞因子谱。使用Western blot，我们证实了LPS-巨噬细胞-EVs比对照组-巨噬细胞-EVs表达更多的CXCL2。最后，LPS-Raw264.7 -EVs上的CXCL2在体内外募集中性粒细胞，并通过CXCR2/PKC/NOX4途径激活中性粒细胞。临床数据方面，脓毒症患者血清中的EVs趋化中性粒细胞并在体外激活中性粒细胞CXCR2/PKC/NOX4途径。本项目拟研究LPS刺激巨噬细胞释放含CXCL2胞外囊泡趋化并激活中性粒细胞氧化应激促进脓毒症相关性肝衰竭的发展机制。本研究为拓展巨噬细胞胞外囊泡在脓毒症相关性肝衰竭的发生发展提供作用靶点。