■脓毒症是由宿主对感染的反应失调引起的危及生命的炎症。细胞外冷诱导RNA结合蛋白(eCIRP)是最近发现的一种与损伤相关的分子模式,可引起败血症中的炎症和器官损伤。Kupffer细胞可以被激活并极化为炎性M1表型,通过产生促炎介质促进组织损伤。我们假设eCIRP促进脓毒症中Kupffer细胞M1极化。
■我们用重组小鼠(rm)CIRP刺激了从野生型(WT)和TLR4-/-小鼠分离的Kupffer细胞(即,eCIRP)并通过ELISA评估上清液IL-6和TNFα水平。M1和M2标记的iNOS和CD206的mRNA表达,分别,通过qPCR评估。我们通过盲肠结扎和穿孔(CLP)在WT和CIRP-/-小鼠中诱导脓毒症,并通过流式细胞术评估iNOS和CD206在Kupffer细胞中的表达。
■eCIRP剂量和时间依赖性增加了WTKupffer细胞中IL-6和TNFα的释放。在TLR4-/-Kupffer细胞中,eCIRP刺激后它们的增加被阻止。eCIRP显著增加iNOS基因表达,而它没有改变WTKupffer细胞中CD206的表达。在TLR4-/-Kupffer细胞中,然而,eCIRP刺激后,与WTKupffer细胞相比,iNOS表达显着降低。WT小鼠CLP后20h,Kupffer细胞中iNOS的表达显着增加。相比之下,CLP后,与WT小鼠相比,CIRP-/-小鼠中的枯否细胞iNOS表达明显降低。Kupffer细胞中的CD206表达在所有组中没有差异。Kupffer细胞M1/M2比值在WT脓毒症小鼠中显著增加,而与CLP后的WT小鼠相比,CIRP-/-小鼠的显着降低。
■我们的数据清楚地表明,在脓毒症中,eCIRP通过TLR4途径诱导Kupffer细胞M1极化,导致炎性细胞因子的过量产生。eCIRP可能是一个有希望的治疗靶标,通过预防脓毒症中的Kupffer细胞M1极化来减轻炎症。
UNASSIGNED: Sepsis is a life-threatening inflammatory condition caused by dysregulated host responses to infection. Extracellular cold-inducible RNA-binding protein (eCIRP) is a recently discovered damage-associated molecular pattern that causes inflammation and organ injury in sepsis. Kupffer cells can be activated and polarized to the inflammatory M1 phenotype, contributing to tissue damage by producing proinflammatory mediators. We hypothesized that eCIRP promotes Kupffer cell M1 polarization in sepsis.
UNASSIGNED: We stimulated Kupffer cells isolated from wild-type (WT) and TLR4-/- mice with recombinant mouse (rm) CIRP (i.e., eCIRP) and assessed supernatant IL-6 and TNFα levels by ELISA. The mRNA expression of iNOS and CD206 for M1 and M2 markers, respectively, was assessed by qPCR. We induced sepsis in WT and CIRP-/- mice by cecal ligation and puncture (CLP) and assessed iNOS and CD206 expression in Kupffer cells by flow cytometry.
UNASSIGNED: eCIRP dose- and time-dependently increased IL-6 and TNFα release from WT Kupffer cells. In TLR4-/- Kupffer cells, their increase after eCIRP stimulation was prevented. eCIRP significantly increased iNOS gene expression, while it did not alter CD206 expression in WT Kupffer cells. In TLR4-/- Kupffer cells, however, iNOS expression was significantly decreased compared with WT Kupffer cells after eCIRP stimulation. iNOS expression in Kupffer cells was significantly increased at 20 h after CLP in WT mice. In contrast, Kupffer cell iNOS expression in CIRP-/- mice was significantly decreased compared with WT mice after CLP. CD206 expression in Kupffer cells was not different across all groups. Kupffer cell M1/M2 ratio was significantly increased in WT septic mice, while it was significantly decreased in CIRP-/- mice compared to WT mice after CLP.
UNASSIGNED: Our data have clearly shown that eCIRP induces Kupffer cell M1 polarization via TLR4 pathway in sepsis, resulting in overproduction of inflammatory cytokines. eCIRP could be a promising therapeutic target to attenuate inflammation by preventing Kupffer cell M1 polarization in sepsis.