【摘要】：本研究旨在探討表沒(méi)食子兒茶素沒(méi)食子酸酯(epigallocatechin gallate,EGCG)對(duì)脂多糖(lipopolysaccharide,LPS)誘導(dǎo)人視網(wǎng)膜內(nèi)皮細(xì)胞(human retinal endothelial cells,HRECs)炎癥反應(yīng)通路中調(diào)節(jié)活化正常T細(xì)胞表達(dá)與分泌趨化因子(regulated upon activation normal T cell expressed and secreted,RANTES)表達(dá)的影響及可能機(jī)制。將HRECs作為研究對(duì)象,分別用實(shí)時(shí)細(xì)胞計(jì)數(shù)法和非同位素細(xì)胞增殖法檢測(cè)LPS(50～250 ng/mL)和EGCG(0～200μmol/L)對(duì)HRECs的毒性作用,確定合適的實(shí)驗(yàn)藥物濃度。再將細(xì)胞隨機(jī)分為正常對(duì)照組、LPS組和LPS+不同濃度EGCG(100、50、25、12.5、6.25μmol/L)共7組,用不同濃度EGCG預(yù)處理2 h,再加入LPS刺激24 h后,酶聯(lián)免疫吸附法測(cè)定各組培養(yǎng)上清液中RANTES的表達(dá)水平,Western免疫印跡法檢測(cè)蛋白激酶Akt及其磷酸化水平。結(jié)果顯示,LPS可顯著刺激誘導(dǎo)HRECs產(chǎn)生RANTES,EGCG抑制LPS誘導(dǎo)的RANTES表達(dá),作用呈劑量依賴(lài)性。免疫印跡結(jié)果也顯示,LPS對(duì)HRECs炎癥過(guò)程中的Akt通路起重要作用,EGCG可顯著抑制LPS誘導(dǎo)HRECs中Akt信號(hào)分子的蛋白磷酸化水平。以上結(jié)果提示,EGCG能有效抑制LPS誘導(dǎo)HRECs中RANTES的表達(dá),其機(jī)制可能與抑制Akt信號(hào)通路有關(guān)。
[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.
【作者單位】： 杭州職業(yè)技術(shù)學(xué)院;浙江大學(xué)醫(yī)學(xué)院附屬第一醫(yī)院眼科;浙江大學(xué)醫(yī)學(xué)院附屬第一醫(yī)院感染病診治國(guó)家重點(diǎn)實(shí)驗(yàn)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)
【分類(lèi)號(hào)】：R587.2;R774.1

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