Toll样受体在LPS诱导的胆管上皮-间叶样表型转化过程中的作用实验研究-普外科专业论文.docxVIP

遵义医学 遵义医学院硕士学位论文 刘公伟 万方数据 万方数据 Toll 样受体在 LPS 诱导的胆管上皮-间叶样表型转化过程中作 用实验研究 中文摘要 目的：检测胆管上皮细胞与脂多糖（LPS）共培养情况下，Toll样受体（TLR4 ，Toll-like receptor 4）、上皮标志物细胞角蛋白（CK7，Cytokeratin 7）以及间叶标志物波形蛋 白（Vimentin）的表达变化，从而探讨TLR4在LPS诱导胆管上皮细胞上皮-间叶样表 型转化中的可能作用。 方法：用最适浓度LPS（2μg/ml）诱导胆管上皮细胞24小时、48小时、72小时，用实 时荧光定量PCR检测目的基因TLR4、CK7、以及Vimentin在mMRA水平的表达变化情 况，用SPSS17.0统计软件进行统计分析。 结果：LPS诱导胆管上皮细胞，不同时间点检测TLR4、CK7以及Vimentin，实验结 果显示：TLR4在LPS刺激24小时后表达明显上升，48小时达高峰，72小时表达降低， 但扔维持较高水平。72小时内上皮标志物CK7目的基因在mRNA上表达明显下降（P ﹤0.05），且随诱导时间的增加而逐渐降低。间叶标志Vimentin目的基因在mRNA上 表达明显上升（P﹤0.05），且随诱导时间的增加而逐渐增加。同时，TLR4表达量的 增加伴随着CK7表达的下降和Vimentin表达的升高。 结论：LPS刺激下，胆管上皮细胞可发生上皮-间质转变（Epithelial-mesenchymal transitions，EMT），且TLR4在LPS诱导的胆管上皮细胞EMT过程中可能发挥了 重要作用。 关键词：Toll 样受体 4 细胞角蛋白 7 波形蛋白 肝胆管上皮细胞 脂多糖 上 皮-间质转变 2 Study on the roles of Toll-like receptor in LPS induced epithelial-mesenchymal transition of bile duct epithelial cells Abstract Objective: To investigate the expression alterations of Toll-like receptor 4(TLR4) and epithelial marker cytokeratin 7 (CK7) and mesenchymal marker Vimentin (VIM) in human bile duct epithelial cell with stimulation with LPS, and to further explore the roles of TLR4 in LPS-induced EMT of biliary epithelial cells. Methods : The in vitro cultured human bile duct epithelial cells were exposed to LPS (2μg /ml) for 24h, 48h and 72h，respectively. With polymerase chain reaction (PCR), the mRNA levels of CK7,Vimentin and TLR4 were investigated. Statistical analysis was performed with SPSS17.0 statistical software. Results: In bile duct epithelial cells with LPS stimulation, the mRNA expression levels of TLR4 were induced 24h after administration, reached up a peak 48h after administration, and started to decline after 72h with a maintained high level，The mRNA expression levels of Epithelial marker CK 7 gradually descended with the extension of incubation time (P0.01) ， and Vimentin mRNA expression significantly raised (P0.01)with LPS stimulation. Conclusion: With exposure to LPS, the expression of TLR4 was induce