沉默三联基序蛋白25基因对A549/DDP细胞株顺铂耐药性及凋亡的影响  

Effects of tripartite motif containing 25 gene silencing on cisplatin resistance and apoptosis in A549/DDP cells

作　　者：粟林[1] 秦霞[1] 敬怀志 屈灿[1] 邱峰[1] 

机构地区：[1]重庆医科大学第一附属医院药学部,重庆400016

出　　处：《重庆医科大学学报》2014年第11期1604-1607,共4页Journal of Chongqing Medical University

基　　金：重庆市卫生局医学科研重点资助项目(编号:2011-1-036)

摘　　要：目的:三联基序蛋白25(tripartite motif containing 25,TRIM25)为泛素化E3连接酶,通过抑制TRIM25表达,研究其对肺癌顺铂(cis-dichlorodiamine platinum,DDP)耐药细胞株A549/DDP耐药性及凋亡的影响。方法:针对TRIM25基因设计4条不同序列sh RNA,分别构建重组质粒载体pt1、pt2、pt3、pt4。利用脂质体将质粒转染至A549/DDP细胞,采用real-time PCR、Western blot检测抑制效果,MTT实验和流式细胞仪(flow cytometry,FCM)实验分别检测在DDP作用下细胞耐药性改变和凋亡情况。结果:干扰质粒pt1在m RNA和蛋白质水平上均能特异性地抑制TRIM25的表达;MTT法结果显示,抑制TRIM25表达后,干扰组pt1细胞IC50值低于p NC组(P=0.001)及对照组(P=0.000);FCM结果显示,干扰组pt1的细胞凋亡率明显低于p NC组(P=0.005)、对照组(P=0.004)。结论:抑制TRIM25表达后,能有效地降低A549/DDP细胞株的DDP耐药性并促进其凋亡。Objective ：To research the effects of silencing tripartite motif containing 25（TRIM25） gene on cisplatin resistance and apoptosis in cisplatin resistant lung cancer cell lines A549/DDP. Methods：Four types of short hairpin RNA were designed and prepared. The recombinant plasmids （pt1,pt2,pt3,pt4） were constructed respectively,which were transfected into A549/ DDP cells using lipofectamine. Real-time PCR and Western blot were used to detect the inhibitory effects. The resistance of cisplatin to A549/DDP cells was detected by MTT assay. Cell apoptosis was tested by flow eytometry（FCM）. Results：The recombinant plasmid ptl can inhibit mRNA and protein expression of TRIM25 specifically. MTT data showed that the cisplatin IC50 was decreased significantly in ptl group than in pNC group（P=0.000） and control group（P=-0.000） after the TRIM25 gene inhibition. According to the results of FCM, apoptotie cells was significantly decreased in ptl group than in pNC group （P=0.005） and control group （P=0.004）. Conclusion：TRIM25 gene silencing in A549/DDP ceils can effectively reduce eisplatin resistance and promote cells apoptosis.

关 键 词：SIRNA TRIM25 A549/DDP细胞 肿瘤耐药 

分 类 号：R73[医药卫生—肿瘤]