TGF-β1诱导胃癌细胞上皮间质转化及其wnt信号通路的研究

发布时间：2018-10-13 17:14

【摘要】：[目的]胃癌是最常见的消化道恶性肿瘤,最近几年发病率逐年升高。超过半数患者初次就诊时疾病已进展至中晚期,即使在通过手术根治+辅助性放化疗,治疗效果仍不理想。胃癌转移是肿瘤最重要的恶性特征,也是导致肿瘤患者死亡的最主要原因。肿瘤的转移是一个多步骤的过程,在肿瘤转移初期过程中肿瘤细胞发生上皮间质转化(Epithelial-Mesenchymal Transition,EMT)。目前,研究发现,转化生长因子与Wnt/β-catenin信号通路在肿瘤EMT进程中发挥着重要作用。我们的目的是探讨胃癌转移过程中发生EMT后的表型改变以及Wnt信号通路在其中的作用机制,为胃癌转移提供新的理论依据,为胃癌转移的靶向治疗提供实验依据。[方法]将人胃癌细胞株MKN28的细胞接种于培养基中,待细胞长至对数期80%汇合度时,进行传代。随后加入不同浓度TGF-β 1(5/10/20/50ng/ml),做72h诱导处理后,通过CCK8、细胞周期试验、侵袭实验及在倒置显微镜下实时观察细胞表型变化并与亲代MKN28细胞株作对比。并且通过Western Blot方法检测MET标志物β-catenin,以及Wnt/β-catenin信号通路相关基因Wnt3a、β-catenin、CyclinD1的表达与原肿瘤细胞做对比。[结果]实验发现不同浓度TGFβ 1在体外能够诱导胃癌细胞发生上皮间质转化。使得胃癌细胞出现细胞形态改变、细胞之间黏附力减弱、细胞活力增加,增殖、侵袭能力增强。与此同时还可激活胃癌细胞中的Wnt3a信号通路,上调Wnt3a、β-catenin、CyclinD1的表达;但实验结果与TGFβ1的浓度未出现相关性,在该试验中存在最佳诱导浓度为1Ong/ml。[结论]1.胃癌细胞在不同浓度TGFβ1的诱导下可发生EMT转化,其细胞形态改变、细胞之间黏附力减弱、细胞活力增加,增殖、侵袭能力增强。2.TGFβ 1可以有效激活Wnt3a信号通路促进胃癌细胞进一步发生EMT。3.TGFβ 1在诱导过程中胃癌细胞表达产物的变化与其浓度未出现相关性。
[Abstract]:Objective: gastric cancer is the most common malignant tumor of digestive tract. More than half of the patients had advanced to advanced stage at the time of first visit, and the effect of treatment was still not satisfactory even after radical operation with adjuvant radiotherapy and chemotherapy. Metastasis of gastric cancer is the most important malignant feature and the main cause of death in cancer patients. Tumor metastasis is a multistep process in which tumor cells undergo epithelial interstitial transformation (Epithelial-Mesenchymal Transition,EMT) in the early stages of tumor metastasis. At present, it has been found that transforming growth factor and Wnt/ 尾-catenin signaling pathway play an important role in tumor EMT process. Our aim is to investigate the phenotypic changes after EMT and the mechanism of Wnt signaling pathway in gastric cancer metastasis, to provide a new theoretical basis for gastric cancer metastasis, and to provide experimental evidence for the targeted treatment of gastric cancer metastasis. [methods] the cells of human gastric cancer cell line MKN28 were inoculated into culture medium and subcultured when the cells reached the logarithmic phase of 80% confluence. Then different concentrations of TGF- 尾 1 (5/10/20/50ng/ml) were added. After 72 hours of induction, the phenotypic changes of the cells were observed by CCK8, cell cycle test, invasion test and real-time observation under inverted microscope. The results were compared with those of the parent MKN28 cell line. The expression of MET marker 尾-catenin, and Wnt/ 尾-catenin signaling pathway related gene Wnt3a, 尾-catenin,CyclinD1 was detected by Western Blot method. [results] it was found that different concentrations of TGF 尾 1 could induce epithelial mesenchymal transformation of gastric cancer cells in vitro. The cell morphology of gastric cancer cells was changed, the adhesion between cells was weakened, the cell viability was increased, proliferation and invasion were enhanced. At the same time, the Wnt3a signaling pathway was activated and the expression of Wnt3a, 尾-catenin,CyclinD1 was up-regulated in gastric cancer cells, but there was no correlation between the results and the concentration of TGF 尾 _ 1. The optimal induction concentration was 1 Ong / ml. [conclusion] 1. Under the induction of different concentrations of TGF 尾 1, gastric cancer cells can undergo EMT transformation. The cell morphology changes, the adhesion between cells decreases, the cell viability increases and the cell proliferates. 2.TGF 尾 1 could effectively activate the Wnt3a signaling pathway to promote the further development of EMT.3.TGF 尾 1 in gastric cancer cells. There was no correlation between the expression of EMT.3.TGF 尾 1 and the concentration of EMT.3.TGF 尾 1 in gastric cancer cells.
【学位授予单位】：昆明医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.2

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