【作者】 岳丽玲； 李成冲； 周丽； 刘吉成；

【Author】 YUE Li-ling,LI Cheng-chong, ZHOU-li, LIU Ji-Cheng (.Institute of Medicine, Qiqihar Medical College, Qiqihar 161006, Heilongjiang Province, China)

【机构】 齐齐哈尔医学院医药科学研究所；

【摘要】 目的:探讨低分子量姬松茸多糖(LMPAB)对TNF-α诱导HUVECs表达E-selectin和肿瘤细胞体外粘附的影响及其相关机制。方法:本研究以TNF-α诱导HUVECs建立内皮细胞分泌E-selectin模型,流式细胞术及Real-timePCR检测LMPAB对TNF-α诱导的HUVECs细胞E-selectin蛋白和mRNA表达的影响;虎红染料染色法检测HT-29与HUVECs细胞黏附能力的改变;Westernblotting检测HUVECs细胞NF-κBp65蛋白表达;免疫细胞化学染色观察HUVECs细胞NF-κB核转位。结果:10μg·L-1TNF-α刺激HUVECs2h后E-selectin表达量明显增加。与TNF-α刺激组比较,LMPAB预处理HUVECs后E-selectinmRNA和蛋白表达水平明显下降(P<0.05),并具有剂量依赖效应;LMPAB可降低TNF–α诱导的HT-29与HUVECs的粘附,低、中、高剂量组抑制率分别为22.51%,31.03%和33.51%,与TNF-α刺激组比较差异显著(P<0.05);HUVEC经LMPAB处理组,核内NF-κBp65含量较TNF-α刺激组明显减少(P<0.05),并且细胞核内着色强度明显降低,表明LMPAB在一定程度上抑制了NF-κB的活性,阻止NF-κBp65进入核内。结论:LMPAB通过阻断NF-κB活化抑制TNF-α诱导的内皮细胞粘附分子E-selectin表达,对E-selectin介导的肿瘤转移具有一定的防护作用。更多还原

【Abstract】 Objective:Investigate the effect of low molecular weight polysaccharide isolated from agaricus blazei murill on the E-selectin expression in HUVECs cell induced by TNF-α and the cell adhesion of tumour cell to HUVECs. Methods: Flow cytometry and Real-time PCR are applied to assay status of E-selectin protein and mRNA expression in HUVECs cell induced by TNF-α. Rose-Bengal method assay ability of human colon carcinoma HT-29 cell line adhesion to HUVECs.The expression of NF-κB p65 protein in HUVECs cell were detected by Western blotting and the nucleus translocation of NF-κB in HUVECs cell by Immunocytochemistry. Result:The expression of E-selectin has significantly increased by the stimulated with TNF-α 10μg·L-1 for 2 h.Compared with the group stimulated of TNF-a,the expression of E-selectin mRNA and protein in HUVECs cell has apparente decreased by pre-treated with LMPAB. LMPAB can depress the cell adhesion of HT-29 to HUVECs which induced by TNF-α.The Inhibitory rate of low, middle and high dose group was 22.51%,31.03% and 33.51%, respectively. It has obviously disparity compared with TNF-α group(P<0.05).The protein of NF-κB p65 has obviously decreased compare with TNF-α group in HUVEC pre-treated with LMPAB (P<0.05).And Staining intensity has obviously decreased in the nucleus of HUVEC.It indicate that the activity of NF-κB has inhibited in a certain extent and blocked translocation of NF-kB to the nucleus by the action of LMPAB. Conclusion:LMPAB inhibited expression of E-selectin induced by TNF-a in endothelial cells by blocked the JNF-kB activity and has protective effect for tumor metastasis by mediated of E-selectin.更多还原