摘要
目的研究BLCAP基因RNA过编辑对肝癌SMMC7721、Focus细胞的生长增殖能力的影响,并探讨其在肝癌发生发展的作用。方法采用半定量PCR的方法分析BLCAP基因在11株肝癌细胞以及2株正常肝脏细胞中的表达情况,选出2株BLCAP基因相对低表达的肝癌细胞株。以基因转染技术,将空质粒(pc DNA3.1,对照组)、BLCAP_WT(野生型)和BLCAP_RDD(过编辑型)质粒(实验组)分别转染肝癌细胞株,通过Western blot法检验转染效率。利用CCK-8生长曲线试验和平板克隆形成实验检测肝癌细胞株的增殖能力。结果半定量PCR结果显示BLCAP基因在肝癌SMMC7721、Focus细胞中的表达量相对较低。Western blot的结果显示,实验组细胞BLCAP基因的蛋白表达水平较对照组细胞明显增高(P<0.05),表明转染较成功。CCK-8生长曲线试验和平板克隆形成实验的结果显示,肝癌SMMC7721和Focus细胞在转染BLCAP_RDD质粒后,其增殖能力较空质粒对照组及BLCAP_WT组细胞明显增强(P<0.05)。结论 BLCAP基因过编辑可促进肝癌SMMC7721和Focus细胞的增殖,提示其在肝癌细胞中的过编辑与肝癌的发生发展密切相关。
Objective To study the influence of BLCAP RNA over-editing on SMMC7721, Focus cell growth and proliferation, and to presume the mechanism of its action in hepatocarcinogenesis. Methods Semi-quantitative PCR method was used to analyze the expression of BLCAP in 11 hepatoma cells and 2 normal liver cells, then selected two hepatoma cells whose expression of BLCAP were in relatively low level. Transfected hepatoma cells with empty vector （pcDNA3.I, control group）, BLCAP_WT （wild type） and BLCAP- RDD （over-editing type, experimental group） respectively, detect transfection efficiency with Western Blot, and cell proliferation level with CCK-8 growth curve and colony-forming unit assay. Results Semi-quantitative PCR showed that the expression of BLCAP were with a lower level in SMMC7721 and Focus cells. Western Blot showed that the protein expression level of BLCAP in experimental group was significantly higher （P 〈0.05） than control group, and transfected successfully. CCK-8 growth curve and colony-forming unit assay showed that SMMC7721 and Focus cells which transfected with BLCAP_RDD were with a significantly higher proliferation （P 〈0.05） than transfected with pcDNA3.1 and BLCAP-WT. Conclusion BLCAP over-editing could promote SMMC7721 and Focus cells proliferation level, and prompt that over-editing in hepatoma cells is closely related to hepatocarcinogenesis.
出处
《中国热带医学》
CAS
2016年第1期1-5,共5页
China Tropical Medicine
基金
国家自然科学基金(No.81372227)
深圳市科技计划项目(No.JCYJ20130329171031740)
