长春碱增加班马鱼肿瘤耐药基因abcb4的表达  

Vinblastine enhances the expression of tumor resistance gene abcb4 in zebrafish

作　　者：胡荣英 孙琮杰 金璐[2,3] 何思佳 周艳华[2] 舒莉萍[1,2] 何志旭[2,3] 

机构地区：[1]贵州医科大学免疫学教研室 [2]贵州医科大学组织工程与干细胞中心 [3]贵州医科大学儿科学教研室,贵州贵阳550004 [4]成都市妇女儿童中心医院,四川成都610091

出　　处：《基础医学与临床》2017年第6期758-762,共5页Basic and Clinical Medicine

基　　金：国家自然科学基金(31460312);贵州省科技基础条件平台[黔科平台(2009)4005];贵州省重大应用基础专项[黔科合J重大字(2015)2003]

摘　　要：目的通过研究长春碱(VBL)对斑马鱼早期发育过程中abcb4肿瘤耐药基因表达的影响,为进一步建立抗肿瘤药物多药耐药筛选的斑马鱼模型奠定重要的实验基础。方法将长春碱稀释成不同浓度梯度的药液,在受精后0.5~1.5 h的斑马鱼胚胎上完成药物暴露实验,计算出长春碱IC_(50)的数值。设置长春碱为实验组,胚胎培养液为空白对照组,分别作用于受精后0.5~1.5 h的斑马鱼胚胎,观察受精后24~120 h的斑马鱼胚胎发育情况,记录死亡、孵化及畸形的胚胎数目;收集不同组别的斑马鱼胚胎,通过实时荧光定量PCR和胚胎整体原位杂交的方法,检测斑马鱼胚胎中abcb4基因的表达量。结果计算出长春碱在斑马鱼胚胎中的IC_(50)为3.08μmol/L;与对照组比较,实验组的斑马鱼胚胎abcb4基因的mRNA水平明显升高(P<0.05);通过胚胎整体原位杂交的方法,在受精后120 h斑马鱼胚胎的小肠部位发现有abcb4基因阳性杂交信号,且实验组斑马鱼胚胎在脑和心脏部位发现abcb4基因阳性杂交信号。结论长春碱能明显增加斑马鱼早期胚胎中abcb4肿瘤耐药基因的表达水平,提示使用斑马鱼可以复制肿瘤耐药模型。Objective To explore the effects of vinblastine on abcb4 tumor resistance gene expression in early development zebrafish and lay an important foundation for multi-drug resistant antineoplastie screening in zebrafish model. Methods Zebrafish embryos of 0.5 - 1.5 hours post-fertilization were exposured to different concentrations of vinblastine, and then calculated the IC50 of vinblastine. Zebrafish embryos of 0.5 - 1.5 hours post-fertilization were treated with different concentrations of vinblasfine and embryo culture medium respectively, and then observed zebrafish embryo development in 24 - 120 hours post-fertilization and recorded the number of death, hatch and mal- formation. Evaluating the impact of vinblastine on abcb4 gene expression in zebrafish with quantitative real-time PCR and whole-mount in situ hybridization by collecting zebrafish embryos exposed to different concentrations of vinblastine. Results Vinblastine IC50 in zebrafish embryos was 3.08 μmol/L. The mRNA level of abcb4 gene in vinblastine treated embryos was significantly increased compared to blank control group. Moreover, abcb4 gene positive hybridization signals were found in the small intestine of zebrafish embryos after 120 hours post-fertilization and also found in the brain and heart of zebrafish embryos by the method of whole-mount in situ hybridization. Conclusions Vinblastine can significantly increase the expression level of abcb4 tumor resistancegene in early development zebrafish embryos, which indicates that zebrafish can be used as a tumor resistant drug screening model.

关 键 词：斑马鱼 长春碱 abcb4 多药耐药 

分 类 号：R965[医药卫生—药理学]