Yan Zhan-Peng, Xu Ting-Ting, An Zhen-Tao, Hu Ying, Chen Wan-Zhen, Zhu Fang-Shi
Affiliated Hospital of Integrated Traditional Chinese and Western Medicine, Nanjing University of Chinese Medicine, Nanjing 210028, China.
Jiangsu Province Institute of Traditional Chinese Medicine, Nanjing 210028, China.
Sheng Li Xue Bao. 2018 Jun 25;70(3):262-268.
The aim of this study was to investigate the mechanisms of mono-functional alkylating agent MNNG to damage human gastric epithelial GES-1 cells and roles of Wnt/β-catenin signaling pathway in the process. The GES-1 cells were treated with MNNG (2 × 10 mol/L) for 24 h. The morphological changes of the GES-1 cells were observed under inverted microscope 2 d after treatment. The cell viability was measured by MTT assay. The apoptosis and cell cycle distribution of the GES-1 cells were analyzed by flow cytometry. The mRNA expressions of β-catenin, GSK-3β, c-Met and MMP7 in the GES-1 cells were detected by qPCR. The protein expressions of β-catenin, GSK-3β, p-GSK-3β and c-Met were determined by Western blot. The results showed that MNNG induced the injury of GES-1 cells and changed the normal cell morphology to irregular long spindle shape. MNNG induced the apoptosis of GES-1 cells and blocked the cell cycle progression obviously. MNNG up-regulated the mRNA expressions of β-catenin, GSK-3β, c-Met and MMP7, and increased the protein expressions of β-catenin, GSK-3β and p-GSK-3β. These results suggest that the damage of GES-1 cells induced by MNNG may be related to the activation of Wnt/β-catenin signaling pathway, which will provide the basis for the study of cell model of gastric mucosal cell injury.
本研究旨在探讨单功能烷化剂MNNG损伤人胃上皮GES-1细胞的机制以及Wnt/β-连环蛋白信号通路在此过程中的作用。将GES-1细胞用MNNG（2×10⁻⁵mol/L）处理24小时。处理后2天，在倒置显微镜下观察GES-1细胞的形态变化。通过MTT法检测细胞活力。通过流式细胞术分析GES-1细胞的凋亡和细胞周期分布。通过qPCR检测GES-1细胞中β-连环蛋白、糖原合成酶激酶-3β（GSK-3β）、c-Met和基质金属蛋白酶7（MMP7）的mRNA表达。通过蛋白质免疫印迹法测定β-连环蛋白、GSK-3β、磷酸化GSK-3β（p-GSK-3β）和c-Met的蛋白表达。结果显示，MNNG诱导GES-1细胞损伤，使正常细胞形态变为不规则长梭形。MNNG诱导GES-1细胞凋亡并明显阻断细胞周期进程。MNNG上调β-连环蛋白、GSK-3β、c-Met和MMP7的mRNA表达，并增加β-连环蛋白、GSK-3β和p-GSK-3β的蛋白表达。这些结果表明，MNNG诱导的GES-1细胞损伤可能与Wnt/β-连环蛋白信号通路的激活有关，这将为胃黏膜细胞损伤细胞模型的研究提供依据。
