应用Bhas42细胞实验检测佛波酯对22个肿瘤促长基因表达的影响  

Assessment of 22 tumor promotion marker genes for predicting tumor promotion potential of chemicals in Bhas42 cells

作　　者：宋征 韩峻松[2] 汪溪洁 马璟 

机构地区：[1]中国医药工业研究总院国家上海新药安全评价研究中心,上海201203 [2]生物芯片上海国家工程研究中心,上海201203

出　　处：《中国药理学与毒理学杂志》2014年第2期274-278,共5页Chinese Journal of Pharmacology and Toxicology

基　　金："十二五"科技重大专项(2012ZX09505001-003)~~

摘　　要：目的探索目标基因与Bhas42细胞转化试验结合成为检测化合物致癌促长活性新的筛选方法的可能性。方法取促癌化合物佛波酯(TPA)作为阳性物,采用Bhas42转化细胞作为实验体系。接种当日为第0天(d 0),d 4将培养基换成含有TPA 0.05 mg·L-1或0.5%DMSO的DF5F培养基,在加入相应受试化合物的24,48及72 h后取样。样品经过预处理后对抽提的RNA进行质量评价,采用RT-PCR方法检测Ccnb1,Rif1,Mcm3,Chek1,Jun,Fosl1,Hells,Vegfa,Stmn1,Prl2c3,Scarb1,Phex,Orm1,Orm2,Nup54,Slc2a1,Il1rl1,Rad51ap1,Tfrc,Ab1,Car13和Pik3r5在不同时间点的表达。结果与阴性对照组相比,TPA组加样后24 h有3个基因出现上调,分别为Vegfa,Il1rl1和Pik3r5;加样后48 h有11个基因出现上调,分别为Chek1,Hells,Mcm3,Rad51ap1,Vegfa,Il1rl1,Prl2c3,Slc2a1,Tfrc,Stmn1和Rif1;加样后72 h有10个基因出现上调,分别是Chek1,Hells,Rad51ap1,Vegfa,Il1rl1,Prl2c3,Slc2a1,Tfrc,Stmn1和Ccnb1。结论 TPA处理后所选取的22个促长阶段相关基因的表达在不同时间点呈现出不同的敏感性,说明22个肿瘤促长基因与Bhas42细胞体系结合,有望成为一种更快捷高速的致癌性筛选方法。OBJECTIVE To evaluate whether the tumor promoting activity syste m based on the ex-pression of 22 tumor pro motion marker genes in Bhas 42 cell transformation assay can be developed to be a new screening technique for predicting tumor promoting potential of che mical.METHODS Chose the 12-O-tetradecanoylphorbol-13-acetate （TPA）as positive che mical and use the Bhas 42 cell lines as test syste m.Define the day of seeding cells as d 0.On d 4,medium in each well was changed with the medium DF5F containing 0.05 mg·L -1 TPA or 0.5 %DMSO,after treat for 24,48 and 72 hrs,collect the cell samples.Extract the RNA fro m the cell samples and detect the expression of Ccnb1 ,Rif1 , Mc m3,Chek1 ,Jun,Fosl1 ,Hells,Vegfa,St mn1 ,Prl2c3,Scarb1 ,Phex,Orm1 ,Orm2,Nup54, Slc2a1 ,Il1 rl1 ,Rad51 ap1 ,Tfrc,Ab1 ,Car13 and Pik3r5 at different ti mepoints by RT-PCR.RESULTS Co mpared to the negative group,the expression of 3 genes was increased after treat with TPA for 24 hr which are Vegfa,Il1 rl1 and Pik3r5;the expression of 12 genes was increased after treat with TPA for 48 hr which are Chek1 ,Hells,Mc m3,Rad51 ap1 ,Vegfa,Il1 rl1 ,Prl2c3,Slc2a1 ,Tfrc,Ab1 ,St mn1 and Rif1 ;the expression of 10 genes was increased after treat with TPA for 72 hr which are Chek1 ,Hells, Rad51 ap1 ,Vegfa,Il1 rl1 ,Prl2c3,Slc2a1 ,Tfrc,St mn1 and Ccnb1 .CONCLUSION The 22 tumor pro-motion marker genes showed different sensitivity to the positive control che mical at different ti mepoints after treat with TPA.The result was consistent with the pro motion activity of TPA.The 22 tumor pro motion marker genes combined with the Bhas 42 cell transformation assay can be developed to be a new screening technique for predicting tumor promoting potential of che mical.

关 键 词：Bhas42细胞 致癌 基因标志物 

分 类 号：R99[医药卫生—毒理学]