摘要
目的建立人肝癌多药耐药细胞系并研究其耐药特性及机制。方法应用人肝癌细胞株HepG2,采用阿霉素(ADM)浓度梯度递增诱导法,建立多药耐药细胞系HepG2/ADM。相差显微镜观察细胞,用MTT法检测该耐药细胞株对多种化疗药物的多药耐药性,采用流式细胞术检测该耐药细胞表面多药耐药基因(mdr)的表达产物P糖蛋白(P-gp)、多药耐药相关蛋白(MRP)及谷胱甘肽硫转移系统(GSH/GST)的表达情况,应用RT-PCR检测MDR、MRP基因表达水平。结果HepG2/ADM对多种抗癌药物产生耐药,对阿霉素的耐药性提高了89.38倍。流式细胞仪分析发现(45.5±5.1)%的HepG2/ADM细胞表面P-gp表达阳性,而对照细胞HepG2表达仅为(11.3±1.2)%,二者差异有显著性(P<0.01);HepG2/ADM和HepG2细胞的MRP及GSH/GST表达无明显变化(P>0.05)。RT-PCR检测发现HepG2/ADM中MDR mRNA高表达。结论HepG2/ADM具有明确的多药耐药性,其多药耐药相关基因MDR mRNA表达升高。
Objective To establish a multidrug resistance cell line of human hepatic carcinoma and study the mechanism of multidrug resistance. Methods Human hepatoceUular carcinoma cell line HepG2 was induced to multidrug resistance cell line HepG2/ ADM by intermittent administration of high dose ADM. The multidrug resistance was detected by using MTT assay. The expressions of mdr-1 gene-coded P-glycoportein （P-gp）, multidrug resistance-associated protein （MRP）, and GSH / GST were examined by flow cytometric assay. The expressions of mdr-1 gene-coded P-glycoportein （P-gp）, multidrug resistance-associated protein （MRP）, and GSH / GST were examined by flow cytometric assay. The expression of MDR and MRP gene were also detected using RT-PCR in both HepG2/ADM and HepG2 cell lines.Results HepG2/ADM was resistant to many anti-tumor agents. The IC50 of ADM was 89.38 times higher than that of HepG2. It＇s expression of P-gp was increased significantly to （ 45.5 ± 5.1） %, while that of HepG2 were only （ 11.3 ± 1.2 ） %. There was no difference of the MRP and GSH/GST expressions between HepG2/ADM and HepG2 cells. The level of MDR mRNA expression was relatively high in HepG2/ADM. Conclusion HepG2/ADM was a model with multidrug resistance and the level of MDR mRNA expression were higher in HepG2/ADM than that in HepG2.
出处
《中国实验诊断学》
2006年第5期460-463,共4页
Chinese Journal of Laboratory Diagnosis
基金
辽宁省科技厅自然科学基金资助项目(2001101033)
