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本文旨在探讨红景天苷(salidroside,Sal)对脂多糖(lipopolysaccharide,LPS)诱导小鼠巨噬细胞系J774.1炎性活化的影响及其可能机制。J774.1细胞分为PBS对照组、LPS(0.5μg/m L)刺激组和不同剂量(5、25、125μg/m L)Sal预处理+LPS组。CCK-8比色法检测细胞活性,ELISA测定培养上清中TNF-α、MCP-1和MIP-2含量,硝酸还原酶法测定上清中NO含量,RT-PCR检测细胞i NOS m RNA表达,Western blot检测胞浆i NOS蛋白和胞浆与胞核NF-κB/p65蛋白表达,Trans AMTM NF-κB/p65活性检测试剂盒测定NF-κB/p65 DNA结合活性。结果显示,0.5μg/m L LPS以及不同剂量(5、25、125μg/m L)Sal处理细胞12 h对J774.1细胞活力无影响;与LPS刺激组比较,LPS刺激前Sal预处理J774.1细胞,培养上清中TNF-α、MCP-1、MIP-2和NO含量呈剂量依赖性降低(P<0.05),细胞i NOS m RNA和蛋白表达水平下调(P<0.05),胞核NF-κB/p65蛋白表达降低(P<0.05)而胞浆NF-κB/p65蛋白相应增加(P<0.05),且NF-κB/p65 DNA结合活性呈剂量依赖性降低(P<0.05)。以上结果提示,Sal预处理能够降低LPS诱导的巨噬细胞炎性活化,其机制可能通过干扰LPS/TLR4/NF-κB信号通路,从而降低炎性介质及细胞因子的过度表达和分泌。
This article aims to investigate the effect of salidroside (Sal) on inflammatory activation of mouse macrophage cell line J774.1 induced by lipopolysaccharide (LPS) and its possible mechanism. J774.1 cells were divided into PBS control group, LPS (0.5μg / m L) stimulation group and Sal pretreatment + LPS group (5,25,125μg / mL). The activity of TNF-α, MCP-1 and MIP-2 in culture supernatant were determined by CCK-8 colorimetric assay. The content of NO in supernatant was determined by nitrate reductase method. The expression of iNOS mRNA was detected by RT- The expression of NF-κB / p65 protein in cytoplasm and iNOS were detected by Western blot. The NF-κB / p65 DNA binding activity was detected by Trans AMTM NF-κB / p65 activity assay kit. The results showed that LPS stimulated J774 cells with LPS at a concentration of 0.5μg / mL and Saline at different doses (5, 25, 125μg / mL) Sal had no effect on the viability of J774.1 cells. 1, and the content of TNF-α, MCP-1, MIP-2 and NO in culture supernatant decreased in a dose-dependent manner (P <0.05) The NF-κB / p65 protein expression was decreased (P <0.05) and the cytoplasmic NF-κB / p65 protein was increased correspondingly (P <0.05), and the NF-κB / p65 DNA binding activity was decreased in a dose-dependent manner (P <0.05). These results suggest that Sal pretreatment can reduce LPS-induced inflammatory activation of macrophages, and its mechanism may be through interference with LPS / TLR4 / NF-κB signaling pathway, thereby reducing the inflammatory mediators and cytokines over-expression and secretion.