沉默XIST通过上调miR-488-3p抑制宫颈鳞状细胞癌细胞的顺铂耐药性  

Silencing XIST inhibits cisplatin resistance in cervical squamous cell carcinoma cells by up-regulating miR-488-3p

作　　者：吴莉敏 吴莉娜[2] 付丽丽[1] 李宝娣 刘婷婷 寇卫华[3] 李荣[4] WU Limin;WU Lina;FU Lili;LI Baodi;LIU Tingting;KOU Weihua;LI Rong(Department ofPediatric Surgery, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710004, China;Operating Theatery,the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710004, China;Department of Oncology,the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710004, China)

机构地区：[1]西安交通大学第二附属医院妇产科,陕西西安710004 [2]西安交通大学第二附属医院手术室,陕西西安710004 [3]西安交通大学第二附属医院.肿瘤科,陕西西安710004 [4]西安交通大学第二附属医院.小儿外科,陕西西安710004

出　　处：《现代医学》2019年第4期415-421,共7页Modern Medical Journal

摘　　要：目的:探索长链非编码RNA XIST对宫颈鳞状细胞癌细胞顺铂耐药性的作用及机制。方法:建立宫颈鳞状细胞癌细胞顺铂耐药系CASKI/cDDP;构建沉默XIST的慢病毒载体(si-XIST)及miR-488-3p沉默载体miR-inhibitor;将si-XIST单独稳定转入CASKI/cDDP细胞或将si-XIST与miR-inhibitor共转入CASKI/cDDP细胞,qRT-PCR检测XIST、miR-488-3p水平变化;MTT法检测细胞存活率变化,蛋白质印迹法检测多药耐药基因1(MDR1)表达变化,RNA拉下实验确定miR-488-3p与XIST的结合关系。结果:与CASKI相比,CASKI/cDDP中XIST表达升高(P <0. 05);si-XIST转染CASKI/cDDP细胞48 h后,XIST的水平显著下调(P <0. 05)、细胞的存活率下降(P <0. 05)、MDR1表达水平下降(P <0. 05)。CASKI/cDDP细胞XIST沉默可增加miR-488-3p水平(P <0. 05);生物信息学预测宫颈癌关键标志物microRNA-488-3p与XIST结合;RNA拉下实验确认XIST直接结合miR-488-3p。CASKI/c DDP细胞共转染si-XIST与miR-488-3p-inhibitor;miR-488-3p-inhibitor部分逆转si-XIST的功能,提高CASKI/c DDP在顺铂处理下的细胞活性(P <0. 05)。结论:沉默XIST通过上调miR-488-3p抑制宫颈鳞状细胞癌细胞的顺铂耐药性。Objective: To investigate the effect and mechanism of long non-coding( lnc) RNA XIST on the cisplatin( c DDP) resistance in cervical carcinoma cells. Methods: CASKI/cDDP,the cDDP resistant cell line of cervix squamous cell carcinoma was constructed. Additionally,the lentivirus vector for silencing XIST( si-XIST)and miR-488-3 p silencing vector( miR-inhibitor) were constructed. Then,si-XIST was stably transfected into CASKI/cDDP cell alone or si-XIST and miR-inhibitor were co-transfected into CASKI/cDDP cell. The changes of XIST and miR-488-3 p levels were tested by qRT-PCR. Cell proliferation activity was detected by MTT. The expression of multidrug resistance gene 1( MDR1) was detected by western blot. The relationship between miR-488-3 p and XIST was studied by RNA pull down assay. Results: The expression of XIST in CASKI/cDDP was higher than that in CASKI( P < 0. 05). The expression of XIST and MDR1 and the cell proliferation activity in siXIST-transfected CASKI/cDDP cells were downregulated( P < 0. 05). MiR-488-3 p,a key marker of cervical cancer,was predicted to bind with XIST by bioinformatics prediction. RNA pull down assay confirmed that XIST binded directly to miR-488-3 p. Co-transfection of si-XIST and miR-inhibitor into CASKI/cDDP cells partially reversed the function of si-XIST,and increased the cell viability of CASKI/cDDP treated with cisplatin( P <0. 05). Conclusion: Silencing XIST inhibits the cisplatin resistance in cervical squamous cell carcinoma by upregulating miR-488-3 p.

关 键 词：宫颈鳞状细胞癌细胞 顺铂耐药性 长链非编码RNA XIST 微小RNA-488-3p RNA拉下实验 

分 类 号：R737.33[医药卫生—肿瘤]