广东医学
廣東醫學
엄동의학
Guangdong Medical Journal
2015年
18期
2777-2782
,共6页
PEG10%肺癌%β-catenin%上皮间质转化%RNA干扰
PEG10%肺癌%β-catenin%上皮間質轉化%RNA榦擾
PEG10%폐암%β-catenin%상피간질전화%RNA간우
PEG10%lung cancer%β-catenin%epithelial mesenchymal transition%RNAi
目的 通过应用RNAi技术沉默肺癌细胞系A549 中的PEG10 基因,探讨PEG10 在肺癌细胞系A549细胞增殖及侵袭迁移过程中的作用. 方法 RPMI-1640培养肺癌A549细胞. 瞬时转染及建立稳定转染shRNA PEG10细胞系;MTT及软琼脂克隆形成实验检测PEG10基因沉默对A549肺癌细胞增殖的影响;细胞划痕试验和Transwell小室方法观察PEG10基因沉默对A549肺癌细胞迁移能力的影响;逆转录RT-PCR、实时定量qRT-PCR及Western blot检测PEG10、β-catenin和其下游分子cmyc、MMPs、twist、E-cadherin、Vimentin表达水平的变化. 结果 PEG10干扰效率达65%;转染后肺癌细胞划痕愈合速度变慢;穿过Transwell小室的细胞数减少;克隆形成数目也降低,同时检测到β-catenin及c-myc、MMPs、twist、Vimentin表达水平降低,E-cadherin表达水平升高. 结论 靶向封闭PEG10基因可下调Wnt/β-catenin信号通路的关键因子β-catenin的表达水平,降低肺癌A549细胞增殖、侵袭及迁移能力,部分逆转肺癌上皮间质转化.
目的 通過應用RNAi技術沉默肺癌細胞繫A549 中的PEG10 基因,探討PEG10 在肺癌細胞繫A549細胞增殖及侵襲遷移過程中的作用. 方法 RPMI-1640培養肺癌A549細胞. 瞬時轉染及建立穩定轉染shRNA PEG10細胞繫;MTT及軟瓊脂剋隆形成實驗檢測PEG10基因沉默對A549肺癌細胞增殖的影響;細胞劃痕試驗和Transwell小室方法觀察PEG10基因沉默對A549肺癌細胞遷移能力的影響;逆轉錄RT-PCR、實時定量qRT-PCR及Western blot檢測PEG10、β-catenin和其下遊分子cmyc、MMPs、twist、E-cadherin、Vimentin錶達水平的變化. 結果 PEG10榦擾效率達65%;轉染後肺癌細胞劃痕愈閤速度變慢;穿過Transwell小室的細胞數減少;剋隆形成數目也降低,同時檢測到β-catenin及c-myc、MMPs、twist、Vimentin錶達水平降低,E-cadherin錶達水平升高. 結論 靶嚮封閉PEG10基因可下調Wnt/β-catenin信號通路的關鍵因子β-catenin的錶達水平,降低肺癌A549細胞增殖、侵襲及遷移能力,部分逆轉肺癌上皮間質轉化.
목적 통과응용RNAi기술침묵폐암세포계A549 중적PEG10 기인,탐토PEG10 재폐암세포계A549세포증식급침습천이과정중적작용. 방법 RPMI-1640배양폐암A549세포. 순시전염급건립은정전염shRNA PEG10세포계;MTT급연경지극륭형성실험검측PEG10기인침묵대A549폐암세포증식적영향;세포화흔시험화Transwell소실방법관찰PEG10기인침묵대A549폐암세포천이능력적영향;역전록RT-PCR、실시정량qRT-PCR급Western blot검측PEG10、β-catenin화기하유분자cmyc、MMPs、twist、E-cadherin、Vimentin표체수평적변화. 결과 PEG10간우효솔체65%;전염후폐암세포화흔유합속도변만;천과Transwell소실적세포수감소;극륭형성수목야강저,동시검측도β-catenin급c-myc、MMPs、twist、Vimentin표체수평강저,E-cadherin표체수평승고. 결론 파향봉폐PEG10기인가하조Wnt/β-catenin신호통로적관건인자β-catenin적표체수평,강저폐암A549세포증식、침습급천이능력,부분역전폐암상피간질전화.
Objective To investigate the role of PEG10 in the proliferation, invasion and migration in A549 lung cancer cells, by implementing RNAi silencing of the PEG10 genein A549 cells.Methods A549 cells were cultured in RPMI-1640.Cell lines expressing PEG10-shRNAs were established through transient and stable transfection.Cell pro-liferation was assessed by MTT assay and soft agar colony formation assay for the effect of PEG10 silencing on proliferation of A549 cells.Scratch test and transwell assay were used to detect the cell migration and invasion.Expression of PEG10,β-catenin, cmyc, MMPs, twist, E-cadherin and Vimentin at both mRNA and protein levels were evaluated by reverse transcription PCR, real time PCR and Western blot.Results RNAi efficiency of PEG10 was 65%.Healing rate was slo-wer in the scratch test and quantity of viable cells in the transwell assay was lower after transfection, as well as colony forming units.Expression ofβ-catenin, c-myc, MMPs, twist and Vimentin were reduced while expression of E-cad-herin was elevated.Conclusion Blockage of PEG10 in A549 cells downregulates the expression ofβ-catenin, which is the critical factor of the Wnt/β-catenin signal pathway, inhibits proliferation, invasion and migration of the human lung cancer A549 cells, and partially reverses epithelial mesenchymal transition effect in lung cancer.
