三维微环境Wnt/β-catenin通路在胰腺癌化疗耐药中的作用机理

In this study, we investigated the relationship between chemoresistance in pancreatic adenocarcinoma cells of MCTS and Wnt/β-catenin signaling pathway in the three-dimensional culture. Wnt signaling pathway has been reported to be involved in embryonic development and cancer development in recent years. Our previous study found that the cells based on single-layer in two-dimensional model were shown to exhibit significantly higher chemosensitivity in vitro than the cells with in a three-dimensional growing model.Pancreatic cancer cells, in three-dimensional culture, were used for assessing the chemoresistance and Wnt/β-catenin signaling pathway activity.We studied the mechanism of Wnt/β-catenin signaling pathway for pancreatic cancer chemoresistance in the three-dimensional cancer microenvironment with real-time quantitativeRT-PCR and Western blotting. Reporter assays were used for Wnt/β-catenin signaling pathway activity, in which TOPflash or FOPflash.We chose rWnt3a and rDkk-1 for alternating the Wnt signaling pathway activity. rWnt3a was used as agonist and rDkk-1 as antagonist.we investigated chemoresistance in pancreatic cancer at pancreatic cancer cellular level, animal studies of pancreatic adenocarcinoma and pancreatic cancer patients clinical study.The purpose of our study tried to provide the relationship between pancreatic cancer resistance to chemotherapy and the signaling pathway.

项目以三维微环境中调控胚胎发育和肿瘤发生的共同途径Wnt/β-catenin通路为切入点，结合我们前期研究:更接近于体内肿瘤状态的三维微环境中胰腺癌细胞化疗敏感性与平面模型有显著差别，但其化疗耐药机制仍未明确。项目拟在前期基础上，从三维细胞模型，Wnt通路在不同化疗药敏特性球体细胞中活性及Wnt通路对化疗耐药调控机理三方面入手，首先利用三维培养多细胞球体作为理想的化疗模型，接着用荧光素酶报告系统检测不同药敏特性球体Wnt通路活性, 同时采用RT-PCR及Western blot在体内和体外实验体系深入分析不同药敏特性球体Wnt通路β-catenin及其下游靶基因与蛋白表达差异。接着分别激活和阻断Wnt通路，调节其活化状态，观察其对化疗敏感性的变化和对下游靶基因及蛋白表达的影响，同时利用动物模型结合患者标本以期在细胞模型，动物实验，患者研究三个水平探索信号传导网络对胰腺癌化疗耐药的调控规律。