TLR-4 may mediate signaling pathways of Astragalus polysaccharide RAP induced cytokine expression of RAW264.7 cells
TLR-4 may mediate signaling pathways of Astragalus polysaccharide RAP induced cytokine expression of RAW264.7 cells
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TLR-4可能介导黄芪多糖RAP诱导RAW264.7细胞细胞因子表达的信号通路
DOI：
10.1016/j.jep.2015.12.060
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发表时间：
2016-02-17
影响因子：
5.4
通讯作者：
Han, Quan-Bin
中科院分区：
文献类型：
--
作者：
Wei, Wei;Xiao, Hai-Tao;Han, Quan-Bin
民族药理学相关性：黄芪多糖（黄芪（菲施）Bge.;黄芪）能够诱导巨噬细胞产生细胞因子，被认为是这种常用草药的免疫增强作用的主要活性成分。研究目的：研究黄芪多糖RAP的免疫调节活性的分子机制，RAP是一种平均分子量为1334 kDa的超支化杂聚糖。ELISA法检测细胞因子的产生，MIT法检测细胞活力。Western blot法测定丝裂原活化蛋白激酶（MAPK）的蛋白质含量。结果：RAP本身对小鼠乳腺癌4 T1细胞无细胞毒作用，但能显著增强RAW264.7细胞培养上清对4 T1细胞的细胞毒作用。RAP能促进RAW264.7细胞NO和细胞因子的产生，并显著上调TNF-α、IL-6、iNOS基因表达。这些生物活性均被TLR 4（Toll样受体4）抑制剂阻断，提示TLR 4是RAP的受体，并介导RAP的免疫调节活性。进一步的分析表明，RAP快速激活TLR 4相关的MAPK，包括磷酸化ERK、磷酸化JNK和磷酸化p38，并诱导NF-κ B B易位以及I κ B-α降解。结论：RAP可能通过TLR 4介导的MAPK和NF-κ B B的活化而诱导RAW 264. 7细胞产生细胞因子。(C)2016爱思唯尔爱尔兰有限公司版权所有。
Ethnopharmacological relevance: Polysaccharides of Radix Astragali (Astragalus membranaceus (Fisch) Bge.; Huangqi) are able to induce cytokine production of macrophages and are considered the main active ingredient for the immune-enhancing effect of this commonly used medicinal herb.Aim of study: To investigate the molecular mechanism of immunomodulating activities of a reported Astragalus polysaccharide, RAP, which is a hyperbranched heteroglycan with average molecular weight of 1334 kDa.Materials and methods: The cytokine production of RAW264.7 cells were analyzed by using ELISA assays while cell viability was assessed by MIT method. Western blot analysis was used for determining protein contents of mitogen-activated protein kinases (MAPKs). In addition, the level of IL-6, iNOS, and TNF-alpha mRNA was determined by RT-PCRResults: It has been found that RAP itself did not have any cytotoxic effect on mouse mammary carcinoma 4T1 cells, but it significantly enhanced cytotoxicity of the supernatant of RAW264.7cells on 4T1 cells. Furthermore, RAP enhanced the production of NO and cytokines in RAW264.7 cells, and significantly up-regulated gene expressions of TNF-alpha IL-6, iNOS. All these bioactivities were blocked by the inhibitor of TLR4 (Toll-like receptor 4), suggesting that TLR4 is a receptor of RAP and mediates its immunomodulating activity. Further analyses demonstrated that RAP rapidly activated TLR4-related MAPKs, including phosphorylated ERK, phosphorylated JNK, and phosphorylated p38, and induced translocation of NF-kappa B as well as degradation of I kappa B-alpha. These results are helpful to better understand the immunomodulating effects of Radix Astragali.Conclusions: RAP may induce cytokine production of RAW264.7 cells through TLR4-mediated activation of MAPKs and NF-kappa B. (C) 2016 Elsevier Ireland Ltd. All rights reserved.