表没食子儿茶素没食子酸酯抑制脂多糖诱导人视网膜内皮细胞中调节活化正常T细胞表达与分泌趋化因子的表达

发布时间：2018-09-09 19:22

【摘要】：本研究旨在探讨表没食子儿茶素没食子酸酯(epigallocatechin gallate,EGCG)对脂多糖(lipopolysaccharide,LPS)诱导人视网膜内皮细胞(human retinal endothelial cells,HRECs)炎症反应通路中调节活化正常T细胞表达与分泌趋化因子(regulated upon activation normal T cell expressed and secreted,RANTES)表达的影响及可能机制。将HRECs作为研究对象,分别用实时细胞计数法和非同位素细胞增殖法检测LPS(50～250 ng/mL)和EGCG(0～200μmol/L)对HRECs的毒性作用,确定合适的实验药物浓度。再将细胞随机分为正常对照组、LPS组和LPS+不同浓度EGCG(100、50、25、12.5、6.25μmol/L)共7组,用不同浓度EGCG预处理2 h,再加入LPS刺激24 h后,酶联免疫吸附法测定各组培养上清液中RANTES的表达水平,Western免疫印迹法检测蛋白激酶Akt及其磷酸化水平。结果显示,LPS可显著刺激诱导HRECs产生RANTES,EGCG抑制LPS诱导的RANTES表达,作用呈剂量依赖性。免疫印迹结果也显示,LPS对HRECs炎症过程中的Akt通路起重要作用,EGCG可显著抑制LPS诱导HRECs中Akt信号分子的蛋白磷酸化水平。以上结果提示,EGCG能有效抑制LPS诱导HRECs中RANTES的表达,其机制可能与抑制Akt信号通路有关。
[Abstract]:The aim of this study was to investigate the effect of epigallocatechin gallate (epigallocatechin gallate,EGCG) on the expression of activated normal T cells and the secretion of chemokine (regulated upon activation normal T in the inflammatory response pathway induced by lipopolysaccharide (lipopolysaccharide,LPS) in human retinal endothelial cells (human retinal endothelial cells,HRECs). Cell expressed and secreted,RANTES) expression and its possible mechanism. The toxicity of LPS (50 ~ 250 ng/mL) and EGCG (0 ~ 200 渭 mol/L) to HRECs was determined by real-time cell count method and non-isotope cell proliferation assay. The cells were randomly divided into 7 groups: the control group and the control group. The cells were pretreated with different concentrations of EGCG for 2 h and then stimulated with LPS for 24 h. Enzyme linked immunosorbent assay (Elisa) was used to detect the expression of RANTES in culture supernatant. Western blot was used to detect protein kinase Akt and its phosphorylation. The results showed that lipopolysaccharide could induce HRECs to produce RANTES,EGCG and inhibit RANTES expression induced by LPS in a dose-dependent manner. Western blotting also showed that lipopolysaccharide plays an important role in the Akt pathway during HRECs inflammation. EGCG can significantly inhibit the protein phosphorylation of Akt signaling molecules in HRECs induced by LPS. These results suggest that EGCG can effectively inhibit the expression of RANTES in HRECs induced by LPS, and its mechanism may be related to the inhibition of Akt signaling pathway.
【作者单位】：杭州职业技术学院;浙江大学医学院附属第一医院眼科;浙江大学医学院附属第一医院感染病诊治国家重点实验室;
【基金】：supported by the Public Technology Application Research Grant of Zhejiang Province,China(No.2011C33029) the Natural Science Foundation of Zhejiang Province,China(No.LY13B020002)
【分类号】：R587.2;R774.1

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