南蛇藤提取物联合miR-451抑制胃癌细胞侵袭和迁移的机制研究

Effects of the extract of Celastrus orbiculatus extracts combined with miR-451 on inhibiting the invasion and migration of human gastric cancer cells

为探讨南蛇藤提取物联合miR-451抑制人胃癌AGS细胞侵袭和迁移的作用机制,用慢病毒转染技术构建稳定过表达miR-451的人胃癌AGS细胞,记作AGS/miR-451^(+)细胞。MTT试验检测不同浓度南蛇藤提取物(20、40、80、160或320μg·mL^(-1))对AGS/miR-451^(+)细胞增殖的影响。伤口愈合试验和Transwell试验观察南蛇藤提取物对细胞迁移的影响。qRT-PCR试验检测南蛇藤提取物处理后细胞内mTOR mRNA表达量的变化。Western blot试验分析南蛇藤提取物对上皮间质转化(epithelial-mesenchymal transformation, EMT)和PI3K/AKT/mTOR信号转导通路相关蛋白质表达的影响。结果表明:南蛇藤提取物处理24 h后明显抑制AGS/miR-451^(+)细胞的侵袭和迁移能力。细胞中E-Cadherin表达量增加,Vimentin和N-cadherin表达量降低。mTOR通路相关蛋白质,如mTOR、PI3K、AKT、4EBP1的总蛋白质表达水平和磷酸化蛋白质表达水平均显著降低。综上,南蛇藤提取物协同miR-451可抑制AGS/miR-451^(+)细胞的增殖、侵袭和迁移,其分子机制与PI3K/AKT/mTOR信号转导通路相关。

To investigate the mechanisms and the effects of Celastrus orbiculatus extracts(COE) combined with miR-451 on inhibiting the invasion and migration in human gastric cancer AGS cells. The human gastric cancer AGS cells overexpressing miR-451 were constructed using lentiviral transfection technique. After the AGS/miR-451^(+)cells were treated with different concentrations(20, 40, 80, 160 or 320 μg·mL^(-1)) of the extract of Celastrus orbiculatus for 24 hours, the effects on the proliferation were determined by MTT. The Wound healing assay and Transwell assay were used to study the effects on the migration. The expression of mammalian target of rapamycin(mTOR) was quantified by qRT-PCR after the AGS/miR-451^(+)cells were treated with 200 μg·mL^(-1)of the extract of Celastrus orbiculatus for 24 hours.The expressions of EMT-related proteins and the PI3K/AKT/mTOR signal transduction pathway were detected by Western blot.The results showed that the extract of Celastrus orbiculatus significantly inhibited the invasion and migration of AGS/miR-451^(+)cells.The expression of E-cadherin was significantly increased after the treatment with the extract of Celastrus orbiculatus for 24 hours.Meanwhile,the proteins expressions of Vimentin,N-cadherin,mTOR,p-mTOR,4EBP1,p-4EBP1,PI3K,p-PI3K,AKT,and p-AKT were significantly decreased.This study suggests that the extract of Celastrus orbiculatus synergy miR-451 inhibits the invasion and migration in gastric cancer cells.The molecular mechanisms may be related to the PI3K/AKT/mTOR signal pathway.