摘要
目的 探讨顺铂耐药肺腺癌细胞系A549DDP的耐药、凋亡相关基因的表达及其与亲代细胞A549 的差异,观察差异表达基因的反义寡核苷酸的逆转作用。方法 将人工合成的反义、正义硫代脱氧寡核苷酸(Soligodleoxynucleotide ,SODN) 经脂质体包裹后转染A549DDP细胞,应用RTPCR 检测耐药及凋亡相关基因mRNA 水平,免疫细胞化学及流式细胞抗体(FCMIg) 检测相应蛋白及核增殖抗原Ki67 表达,MTT检测其耐药性改变,TUNEL、DNA 电泳检测细胞凋亡。结果 A549DDP细胞多药耐药基因( MDR1) 、拓朴异构酶Ⅱ(TOPOⅡ) 、cmyc mRNA 及蛋白表达水平接近A549 ,二者cerbB2 均阴性。A549DDPbcl2 阳性,且MRP mRNA 及蛋白水平较A549 高。bcl2 、MRP 反义SODN 可相应地抑制bcl2 、MRP、cmyc 表达,降低A549DDP耐药性,抑制其生长与增殖,促进细胞凋亡。结论 肺腺癌A549 细胞获得顺铂抗药性,与bcl2 和MRP基因高表达有密切关系。
Objective To study the expression of drug resistance and apoptosis related genes of A 549 DDP cells as compared to the parental cell line A 549 , and its reversion by antisense s oligodeoxynucleotide (S ODN) of the differentially expressed genes.Methods Sense and antisense S ODN were transferred into A 549 DDP cells by lipofectin. Expression of genes related to drug resistance and apoptosis was examined by RT PCR, immunocytochemitry and flow cytometry. Apoptosis was identified by DNA electrophoresis and TUNEL, and cell growth by MTT uptake.Results The expression of bcl 2 was positive and that of MRP at mRNA and protein levels was increased in A 549 DDP cells compared to A 549 cells. MDR1, c myc and TOPOⅡ were similarly expressed in the two cell lines. Both cell lines were negative for c erbB 2 expression. In A 549 DDP cells, the expression of bcl 2 and MRP was significantly inhibited by respective antisense S ODN. Antisense S ODN could also significantly inhibit proliferation of A 549 DDP cells, and promote cell apoptosis by reducing its resistance to cisplatin.Conclusion Bcl 2 and MRP genes are responsible for the induced resistance of A 549 DDP cells to cisplatin.
出处
《中华肿瘤杂志》
CAS
CSCD
北大核心
1999年第6期422-426,共5页
Chinese Journal of Oncology
基金
北京自然科学基金资助
关键词
肺肿瘤
细胞凋亡
药物耐受性
基因表达
腺癌
Lung neoplasms A 549 and A 549 DDP cell Apoptosis Antisense oligonucleotide Drug resistance Gene expression