摘要
目的探讨以透明质酸-聚乙烯亚胺/透明质酸-聚乙二醇(HA-PEI/HA-PEG)作为基因载体传递NAF1-siRNA对人肝细胞LM3进行增殖能力抑制的效果。方法将纳米复合物HA-PEI/HA-PEG与NAF1-siRNA复合,转染LM3细胞建立HNS组,等量NAF1-siRNA转染建立NSI组,另设未转染对照对照组,等量HA-PEI/HA-PEG干预细胞建立HAH组。采用荧光实时定量PCR检测细胞NAF1 mRNA的表达水平,采用Western blotting法检测NAF1、cyclin D1蛋白表达。采用MTT实验和平板克隆形成实验检测细胞增殖抑制效果。4组比较采用单因素方差分析,两两比较采用t检验。结果以对照组NAF1 mRNA的相对表达量为100﹪进行检测。HNS组NAF1 mRNA的相对表达量为(2.12±0.17)﹪,与对照组相比,差异具有统计学意义(t=14.17,P=0.04)。HNS组LM3细胞的NAF1蛋白和Cyclin D1蛋白的相对表达量分别为0.07±0.01、0.06±0.00,而对应蛋白在对照组LM3细胞中的相对表达量分别为0.37±0.08、0.16±0.03,差异具有统计学意义(t=37.72,20.96,P=0.01,0.03)。HNS组细胞测得的细胞增殖抑制率为(73.20±0.43)﹪,与对照组的(0.49±0.02)﹪相比差异具有统计学意义(t=11.92,P=0.01)。HNS组LM3细胞的平板克隆形成率(8.35±0.33)﹪明显低于对照组的(23.61±0.10)﹪(t=17.75,P=0.00,0.02)。结论纳米复合物HA-PEI/HA-PEG可高效递送NAF1-si RNA,对肝癌LM3细胞进行转染,抑制NAF1表达,进而通过降低Cyclin D1表达水平抑制肝癌细胞LM3的增殖。
Objective To investigate the effect of HA-poly （ethyleneimine）/HA-poly （ethylene glycol） （HA-PEI / HA-PEG） as a gene transfer vector effect NAFI-siRNA in human hepatocytes were LM3 proliferation inhibition. Methods The nano-composite HA-PEI / HA-PEG and NAFI-siRNA complexes, cells transfected LM3 establish HNS group, the same amount of NAF1- siRNA transfected establish NSI group, with another Ctrl untransfected control group and the same amount of HA-PEI/HA-PEG transfeeted establish HAIl group. Expression using quantitative real- time PCR to detect cell NAF1 mRNA level, using Western blotting assay NAF1, cyclin D1 protein expression. Using MTT assay and colony formation assay to detect inhibition of cell proliferation effect. 4 groups were compared using ANOVA, pairwise comparisons using t test. Results In the relative expression of Ctrl group NAF1 mRNA were detected as 100 %. Relatively HNS group NAF1 mRNA expression was （2.12±0.17） %, compared with the Ctrl group, a statistically significant difference （t = 14.17, P = 0.04）. NAF 1 protein and the relative expression of CyclinD1 protein HNS group LM3 cells were 0.07±0.01, 0.06±0.00, and the relative expression of the corresponding protein in the Ctrl group LM3 cells were 0.37±0.08, 0.16±0.03, the difference was statistics significance （t = 37.72, 20.96, P = 0.01, 0.03）. HNS cells measured by cell proliferation inhibition rate （73.20±0.43） %, with the Ctrl group （0.49±0.02） % compared to a statistically significant difference （t = 11.92, P = 0.01 ）. HNS Group LM3 cell colony formation rate （8.35± 0.33） %was significantly lower than that （23.61±0.10） % Ctrl group （t= 17.75, P= 0.00, 0.02）. Conclusion Nanocomposite HA-PEI / HA-PEG can efficiently deliver NAFI-siRNA, hepatoma cells were transfected LM3, inhibition NAF1 expression, thereby reducing the expression of Cyclin D1 levels through inhibition LM3 proliferation of liver cancer cells.
出处
《中华细胞与干细胞杂志（电子版）》
2016年第3期141-145,共5页
Chinese Journal of Cell and Stem Cell（Electronic Edition）
基金
国家自然科学基金(81301331
81302550)
教育部高等学校博士点专项科研基金新教师类资助课题(20110171120091)
广东省科技计划项目(2011B080701063)
关键词
纳米技术
基因
RNA
小分子干扰
癌
肝细胞
Nanotechnology
Genes
RNA, Small Interfering
Carcinoma, Hepatocellular
