摘要
目的:探讨环巴胺阻断Hedgehog通路对食管癌EC109细胞上皮间质转化(epithelial-mesenchymal transition,EMT)的作用及其可能机制。方法:实验分为实验组和对照组,实验组应用Hedgehog通路特异性阻断剂环巴胺作用食管癌EC109细胞48 h。通过real-time PCR检测Gli1的表达变化,观察细胞形态变化,通过血管拟态实验检测血管形成能力变化,Transwell小室侵袭和迁移实验、黏附实验分别检测细胞侵袭、迁移和黏附能力变化,real-time PCR及Western印迹方法检测EMT相关标志物E钙粘蛋白(E-cadherin)、β连环蛋白(β-catenin)、波形蛋白(Vimentin)及EMT调控因子Snail、Twist1等的表达变化。结果:环巴胺阻断EC109细胞Hedgehog信号通路Gli1的m RMA表达明显减少为(41.819±20.150)%,形态发生明显变化,血管拟态个数较对照组[(2.780±0.424)vs.(5.080±0.634)]明显减少(t=-16.919,P=0.000),侵袭实验较对照组[(24.800±2.588)vs.(55.400±4.879)]和迁移实验较对照组[(23.200±1.924)vs.(65.400±4.775)]均明显减少(t=-12.390,P=0.000;t=-18.331,P=0.000),同种细胞间黏附增强(F=9.327,P=0.009),上皮表型标志物E-cadherin表达较对照组[(0.388±0.565)vs.(0.228±0.582)]明显上调(t=3.421,P=0.027),而间质表型Vimentin、β-catenin的表达水平较对照组[(0.588±0.109)vs.(0.507±0.051);(0.998±0.128)vs.(0.756±0.038)]明显下调(t=4.221,P=0.013;t=6.781,P=0.002);与对照组相比,实验组细胞中转录因子Snail的表达较对照组[(0.401±0.021)vs.(0.756±0.038)]下调(t=6.774,P=0.002),Twist1的m RNA表达量相对于对照组也明显下调为(74.987±9.031)%。结论:环巴胺阻断Hh信号通路能明显逆转EC109细胞EMT过程,其机制可能与下调转录因子Snail及Twist1表达有关。
Objective：To explore the effect of cyclopamine on epithelial-mesenchymal transition in human esophageal cancer EC109 cells and the possible mechanism. Methods：EC109 cells were cultured in vitro. After adding cyclopamine for 48 h,the expression of Gli1 was tested by real-time PCR,and morphological change of cells was observed by inverted microscope. The transwell chamber assay was used to examine the invasive and metastatic ability of EC109 cells treated with cyclopamine for 48 h. The ability of vascularization was observed by the method of vascilogenic mimicry and the ability of adhension was detected by adhension experiment.Real-time PCR and Western blot were used to detect the expressions of such EMT markers as E-cadherin,β-catenin and vimentin and such transcription factors as Snail,Twist1. Results：The inhibition of hedgehog signaling significantly reduced the m RNA expression of Gli1（41.819 ±20.150）%,and the invasion [（24.800 ±2.588） vs.（55.400 ±4.879）],migration capacities [（23.200 ±1.924） vs.（65.400±4.775）] of EC109 cells in vitro were decreased（t=-12.390,P=0.000;t=-18.331,P=0.000）. Homogeneous cells intercellular adhesion was increased（F=9.327,P=0.009）,while heterogeneous cells intercellular adhesion was decreased（F=20.459,P=0.000）.Angiogenesis capacity[（2.780±0.424）vs.（5.080±0.634）] was decreased（t=-16.919,P=0.000）. The expression of E-cadherin increased[（0.388±0.565）vs.（0.228±0.582）] significantly（t=3.421,P =0.027）,while Vimentin and β-catenin in the cyclopamine treatment group [（0.588 ±0.109） vs.（0.507 ±0.051）;（0.998 ±0.128）vs.（0.756±0.038）] were significantly down-regulated（t =4.221,P =0.013;t =6.781,P =0.002）. Compared to those in control group,the expressions of Snail were significantly reduced in the cyclopamine treatment group [（0.756 ±0.038） vs.（0.401 ±0.021）]（t=-6.455,P=0.023）. Also,the m RNA expressions of Twist1 were significantly reduced（74.987±9.031）%. Conclusion：The inhibitio
出处
《重庆医科大学学报》
CAS
CSCD
北大核心
2015年第8期1118-1122,共5页
Journal of Chongqing Medical University
关键词
食管癌
环巴胺
HEDGEHOG
上皮间质化
细胞黏附
esophageal cancer
cyclopamine
hedgehog
epithelial-mesenchymal transition
intercellular adhesion
