【摘要】 目的构建人肌腱蛋白C(TNC)真核表达质粒,利用稳定过表达TNC的胰腺癌细胞系以及探索TNC在胰腺癌转移中的作用及分子机制。方法采用独特的引物设计方法构建TNC表达质粒,经转染和G418筛选获得稳定过表达TNC的胰腺癌PANC1细胞系。通过RT-qPCR和Western blot对细胞系中TNC的表达进行鉴定。采用Transwell小室法和Western blot检测TNC对细胞迁移、侵袭能力以及上皮间质转化(EMT)相关分子表达的影响。结果成功构建TNC质粒,稳定过表达TNC的PANC1细胞迁移、侵袭能力显著增强(P<0. 05),上皮标志物E-cadherin表达减少而间质标志物N-cadherin表达增加(P<0. 05)。结论 TNC真核表达质粒以及TNC稳定表达胰腺癌细胞系的建立,可用于TNC对细胞EMT、迁移和侵袭等方面的机制研究,也是研究TNC生物学功能和胰腺癌发生发展机制的基础。更多还原

【Abstract】 Objective To construct human TNC expression plasmid for the investigation of the role of TNC in pancreatic cancer metastasis by using pancreatic cancer cell strain with stable over-expression of TNC. Methods TNC expression plasmid was constructed by unique primer design method,and then pancreatic cancer cell strain PANC1 with stable TNC expression were obtained by transfection and G418 screening. RT-qPCR and Western blot examined the expression of TNC. Transwell and Western blot were used to detecte the effects of TNC on cell migration,invasion and EMT-related molecules. Results TNC plasmid was successfully constructed. TNC promoted cell migration and invasion( P<0. 05),decreased epithelial marker E-cadherin expression and upregulated mesenchymal marker N-cadherin expression( P<0. 05). Conclusions The TNC expression plasmid and TNC stable expression pancreatic cancer cell strain may be used to explore the biological function of TNC and the pathogenesis of pancreatic cancer.更多还原