摘要
目的:建立一株对紫杉醇(PTX)耐药的食管鳞癌细胞株TE1/PTX,并初步观察其表型特征。方法:取对数生长期TE1细胞,用100倍半数抑制浓度(IC50)的PTX(3 mg/L)诱导2 h,待细胞出现克隆现象后传代2~3次,再次冲击诱导,共冲击3次。之后延长药物作用时间,每次诱导4 h,重复7次。总计冲击10次,共需时8个月。用不同浓度的PTX、ADM、5-FU分别干预TE1和TE1/PTX 24/48、48、72 h,CCK-8法检测细胞活性,计算IC50及耐药指数(RI)。倒置光学显微镜下观察TE1和TE1/PTX细胞的形态,细胞计数法和平板克隆形成实验检测细胞增殖能力,PI染色法检测细胞周期,Transwell小室法检测细胞的侵袭和迁移能力,Western blot法检测耐药相关蛋白(BCRP、MRP1、P-gp)和上皮间质转化相关蛋白(E-cadherin、N-cadherin、Slug、Vimentin)的表达。结果:TE1/PTX对PTX(24 h)、PTX(48 h)、ADM(48 h)和5-FU(72 h)的RI分别为13.01、14.62、1.80、3.50。与亲本细胞TE1比较,TE1/PTX变长且无规则,MRP1、P-gp蛋白表达增加;增殖能力下降,细胞被阻滞在G2/M期,侵袭和迁移能力提高,N-cadherin、Slug蛋白表达升高(P<0.05)。结论:成功建立一株PTX耐药的食管鳞癌细胞株TE1/PTX,其耐药机制可能与耐药相关蛋白和上皮间质转化相关蛋白表达改变有关。
Aim:To establish a drug-resistant esophageal squamous cell carcinoma cell line TE1/PTX and observe its phenotype.Methods:Logarithmic phase TE1 cells were induced with 100 times IC50 PTX(3 mg/L)for 2 hours,then cultured with fresh medium,and subcultured 2-3 times after the clone appeared.Shock was repeated for 3 times.And then,the time of drug induction was prolonged to 4 hours for 7 times.A total of 10 shocks were carried out over a period of 8 months.Different concentrations of PTX,ADM and 5-FU were used to intervene TE1 and TE1/PTX for 24/48 hours,48 hours,and 72 hours,respectively.The cell activity was detected by CCK-8 method,IC50 and drug resistance index(RI)were calculated.The morphology of TE1 and TE1/PTX cells was observed under an inverted optical microscope,the proliferation was examined by cytometry and plate clonogenic assay,cell cycle was examined by PI staining,and invasion and migration abilities were examined by Transwell chamber assay,the expressions of drug resistance associated proteins(BCRP,MRP1,P-gp)and epithelial-mesenchymal transition(EMT)related proteins(E-cadherin,N-cadherin,Slug,Vimentin)were detected by Western blot.Results:RI of TE1/PTX to PTX(24 hours),PTX(48 hours),ADM(48 hours)and 5-FU(72 hours)were 13.01,14.62,1.80 and 3.50,respectively.Compared with the parent cell TE1,TE1/PTX became longer and irregular;MRP1 and P-gp protein expressions increased,the proliferation ability decreased,the cells were arrested at G2/M stage,invasion and migration abilities increased,the expressions of N-cadherin and Slug increased(P<0.05).Conclusion:A PTX resistant esophageal squamous carcinoma cell line TE1/PTX has been successfully established.The mechanism of its drug resistance may be related to the changes of resistance-related proteins and EMT-related proteins expressions.
作者
高娜
李岳衡
杨珍珍
高铮钒
李静
党婧晗
许培荣
范天黎
GAO Na;LI Yueheng;YANG Zhenzhen;GAO Zhengfan;LI Jing;DANG Jinghan;XU Peirong;FAN Tianli(Department of Pharmacology,School of Basic Medical Sciences,Zhengzhou University,Zhengzhou 450001;Translational Medicine Research Center,Zhengzhou People′s Hospital,Zhengzhou 450003;Experimental Center,School of Pharmacy,Zhengzhou University,Zhengzhou 450001)
出处
《郑州大学学报（医学版）》
CAS
北大核心
2020年第6期741-745,共5页
Journal of Zhengzhou University(Medical Sciences)
基金
河南省高等学校重点科研项目(20A310021)。
关键词
食管鳞癌
耐药
紫杉醇
esophageal squamous cell carcinoma
resistance
paclitaxel
