CAJ | 학술논문

目的体外实验研究Ras基因家族同源物A(ras homolog gene family,member A,RhoA)小干扰RNA(small interfering RNA,siRNA)在舌癌转移中的作用.方法登录Genebank确定人RhoA基因序列,利用RNA干扰技术针对RhoA的基因序列设计4条短链RNA,构建干扰表达载体,利用LipofectamineTM2000介导法将RhoA siRNA转染至舌癌细胞系Tca8113.转染细胞后48 h检测瞬时表达效率,经杀稻瘟菌素筛选及克隆化培养获得稳定抗性克隆转染株后,蛋白质印迹法检测RhoAsiRNA转染后的抑制效应,细胞计数绘制细胞生长曲线以观察RhoA siRNA转染前后细胞株的生长速度;划痕实验和Millicell小室实验检测转染细胞株的迁移力与侵袭力.结果舌癌细胞转染RhoA siRNA后:RhoA蛋白表达下降;细胞倍增时间从38.0 h延长至45.7 h;细胞克隆形成率由35.2%降低至15.8%;细胞迁移能力减弱;细胞侵袭能力由100%降至58.9%,显著减弱.结论 RhoA siRNA能有效抑制舌癌Tca8113细胞中RhoA的表达,从而降低细胞增殖水平以及细胞侵袭力和迁移力,表明RhoA siRNA具有抑制舌癌细胞生物学特性的能力,提示RhoA在舌癌转移中可能发挥重要作用.
목적 체외실험연구Ras기인가족동원물A(ras homolog gene family,member A,RhoA)소간우RNA(small interfering RNA,siRNA)재설암전이중적작용.방법 등록Genebank학정인RhoA기인서렬,이용RNA간우기술침대RhoA적기인서렬설계4조단련RNA,구건간우표체재체,이용LipofectamineTM2000개도법장RhoA siRNA전염지설암세포계Tca8113.전염세포후48 h검측순시표체효솔,경살도온균소사선급극륭화배양획득은정항성극륭전염주후,단백질인적법검측RhoAsiRNA전염후적억제효응,세포계수회제세포생장곡선이관찰RhoA siRNA전염전후세포주적생장속도;화흔실험화Millicell소실실험검측전염세포주적천이력여침습력.결과 설암세포전염RhoA siRNA후:RhoA단백표체하강;세포배증시간종38.0 h연장지45.7 h;세포극륭형성솔유35.2%강저지15.8%;세포천이능력감약;세포침습능력유100%강지58.9%,현저감약.결론 RhoA siRNA능유효억제설암Tca8113세포중RhoA적표체,종이강저세포증식수평이급세포침습력화천이력,표명RhoA siRNA구유억제설암세포생물학특성적능력,제시RhoA재설암전이중가능발휘중요작용.
Objective To investigate the effect of RhoA on the metastasis of tongue squamous cell carcinoma Tca8113 cells in vitro. Methods A group of RhoA specific small interfering RNAs (siRNA) was constructed and confined by sequencing analysis. The siRNA of RhoA gene was transfected into human tongue squamous cell carcinoma Tca8113 cells line by Lipofectamine? 2000. The protein transient expression of RhoA in the transfectants was examined 48 h after transfection. The cell line with stable expression of siRNA of RhoA was obtained by blasticidin screening and colony culture. Cell growth rate was determined with a cell counting. Millicell chambermodel and wound healing assay were used to examine the abilities of migration and invasion, respectively in vitro. Results RhoA was overexpressed in tongue squamous cell carcinoma Tca8113 cell line. Silencing of endogenous RhoA gene expression in Tca8113 cells resulted in inhibition of the proliferation, adhesion, chemotaxis and invasion of Tca8113 cells in vitro. Conclusions RhoA siRNA inhibits the proliferation, adhesion, chemotaxis and invasion of oral squamous cell carcinoma Tca8113 cell lines. siRNA of RhoA is a potential factor controlling the proliferation and metastasis of Tca8113 cells. RhoA may play an important role in metastasis of oral squamous cell carcinoma.