摘要
目的 :探讨let-7i对人卵巢癌SKOV3细胞增殖、迁移、侵袭和顺铂(cisplatin,DDP)耐药性的影响及可能的作用机制。方法:应用实时荧光定量PCR检测人永生化卵巢细胞IOSE、卵巢癌SKOV3细胞和卵巢癌DDP耐药细胞SKOV3/DDP中let-7i的表达。将let-7i过表达质粒(let-7i)、抑制let-7i表达质粒(anti-let-7i)和阴性对照(negative control,NC)质粒(let-7i-NC)分别转染SKOV3细胞后,应用Ed U(5-ethynyl-2’-deoxyuridine)实验、划痕愈合实验和Transwell侵袭实验分别检测细胞的增殖、迁移和侵袭能力。用不同浓度的DDP处理转染了let-7i质粒的SKOV3/DDP细胞后,MTT法检查SKOV3/DDP细胞对DDP敏感性的变化。应用Target Scan、mi Randa和Pic Tar靶基因预测软件以及DAVI(D Database for Annotation,Visualization and Integrated Discovery)数据库对let-7i进行生物信息学分析,实时荧光定量PCR和蛋白质印迹法检测let-7i和anti-let-7i转染组SKOV3细胞中let-7i潜在靶基因Toll样受体4(Toll-like receptor 4,TLR4)m RNA和蛋白的表达水平。结果 :SKOV3细胞中let-7i的表达水平低于IOSE细胞(P<0.01),而SKOV3/DDP细胞中let-7i的表达水平低于SKOV3细胞(P<0.01)。Let-7i转染组SKOV3细胞的增殖、迁移和侵袭能力下降(P<0.05、P<0.01和P<0.05),而anti-let-7i转染组SKOV3细胞增殖、迁移和侵袭能力的改变不显著(P值均>0.05)。转染let-7i质粒后,SKOV3/DDP细胞对DDP的敏感性增强(P<0.05)。生物信息学分析表明,let-7i的靶基因主要富集于丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)信号通路、转化生长因子-β(transforming growth factor-beta,TGF-β)信号通路和黏着斑通路等。Let-7i转染组SKOV3细胞中TLR4 m RNA和蛋白的表达水平下调(P值均<0.05)。结论 :卵巢癌SKOV3和SKOV3/DDP细胞中let-7i低表达。Let-7i过表达可抑制SKOV3细胞的增殖、迁移和侵袭能力,增强SKOV3/DDP细胞对DDP的敏感性。TLR4可能是let-7i的靶基因之一。
Objective： To investigate the effects of let-7i on the proliferation, migration, invasion and cisplatin（DDP）-resistance of ovarian cancer SKOV3 cells, and explore its possible mechanism.Methods： The expression levels of let-7i in immortalized normal human ovarian epithelial IOSE cells, human ovarian cancer SKOV3 cells and DDP-resistant SKOV3/DDP cells were detected by real-time fluorescent quantitative PCR. After let-7i-overexpression plasmid（let-7i）, let-7i-inhibitor plasmid（anti-let-7i） and negative control（NC） plasmid（let-7i-NC） were transfected into SKOV3 cells, respectively, the abilities of proliferation, migration and invasion of SKOV3 cells were measured by 5-ethynyl-2＇-deoxyuridine（Ed U） assay, wound healing assay and Transwell invasion assay, respectively. The DDP sensitivity of SKOV3/DDP cells transfected with let-7i plasmid after treatment with different concentrations of DDP was detected by MTT assay. A combined bioinformatics approach using Target Scan, mi Randa and Pic Tar softwares and Database for Annotation, Visualization and Integrated Discovery（DAVID） was conducted to obtain the bioinformatics data of let-7i. The expression levels of Tolllike receptor 4（TLR4） m RNA and protein in SKOV3 cells transfected with let-7i plasmid and anti-let-7i plasmid were detected using real-time fluorescent quantitative PCR and Western blotting, respectively.Results： The expression level of let-7i in SKOV3 cells was lower than that in IOSE cells（P 0.01）, but was higher than that in SKOV3/DDP cells（P 0.01）. The abilities of proliferation, invasion and migration of SKOV3 cells transfected with let-7i plasmid were decreased（P 0.05, P 0.01 and P 0.05, respectively）, while which of SKOV3 cells transfected with antilet-7i plasmid were not obviously changed（all P 0.05）. DDP sensitivity of SKOV3/DDP cells was enhanced after transfection with let-7i plasmid（P 0.05）. The result of bioinformatics data analysis indicated that the predicted target genes of let-7i wer
出处
《肿瘤》
CAS
CSCD
北大核心
2016年第2期149-157,共9页
Tumor
关键词
卵巢肿瘤
细胞增殖
肿瘤浸润
抗药性
肿瘤
Let-7i
Ovarian neoplasms
Cell proliferation
Neoplasm invasiveness
Drug resistance
neoplasms
Let-7i
