RNA靶向干扰DcR3促进人肝癌HepG2细胞凋亡及机制研究  

Apoptosis of hepatoma cells line HepG2 induced by DcR3 siRNA and its mechanism

作　　者：黄秋林[1] 曹超[1] 雷俊悦[1] 刘璇[1] 

机构地区：[1]南华大学附属第一医院,湖南衡阳421001

出　　处：《中国现代医学杂志》2012年第13期36-40,共5页China Journal of Modern Medicine

基　　金：湖南省科技厅计划立项项目(No:2010SK3034);湖南省自然科学基金(No:11JJ3114)

摘　　要：目的研究靶向干扰DcR3基因对人肝癌HepG2细胞凋亡的影响并探讨其可能的分子机制。方法设计并合成DcR3基因的特异性DcR3 siRNA(实验组)和非特异性DcR3 siRNA(对照组),在脂质体介导下分别转染HepG2细胞株。Western blot检测两组DcR3 siRNA转染HepG2细胞24和48 h后DcR3蛋白表达水平;MTT法检测转染48 h细胞生长增殖;流式细胞术、DNA琼脂糖凝胶电泳检测转染24 h细胞凋亡;免疫组化法检测转染24 h细胞caspase3蛋白的表达。结果实验组细胞24和48 h DcR3蛋白相对表达水平分别为(0.532±0.051)和(0.417±0.044);对照组分别为(1.978±0.246)和(2.018±0.275);两组比较,相同时间点相对表达水平差异有显著性(P<0.05);实验组24和48 h DcR3蛋白相对表达水平比较差异有显著性(P<0.05),对照组差异无显著性(P>0.05)。两组细胞转染48 h抑制率分别为(59.8±5.4)%和(0.8±0.1)%,两者比较差异有显著性(P<0.05);两组细胞转染24 h凋亡率分别为(19.7±3.2)%和(6.9±0.8)%,两者比较差异有显著性(P<0.05);转染24 h后两组caspase3蛋白阳性率分别为72.8%和38.9%,平均光密度值分别为(0.41±0.06)和(0.21±0.03),两者比较差异有显著性(P<0.05)。结论特异性DcR3 siRNA有抑制人肝癌HepG2细胞DcR3蛋白表达、细胞增殖和诱导凋亡的作用,其机制可能与上调caspase 3的表达有关。【Objective】 To investigate the effect of apoptosis of hepatoma cells line HepG2 induced by DcR3 siRNA and its molecular mechanisms.【Methods】 Specific DcR3 siRNA（experimental group） and nonspecific DcR3 siRNA（control group） were synthesized and transfected to HepG2 cells.DcR3 siRNA transfected HepG2 cells in two groups 24 h,48 h after the DcR3 protein levels were analyzed by Western blot.The growth inhibition was examined by MTT assay.The apoptosis was explored by flow cytometry and DNA ladder.Immunohisto-chemical technique was used to determine the expression level of caspase3 protein.【Results】 Experimental group cells 24h,48h DcR3 protein expression levels were（0.532±0.051）,（0.417±0.044）;the control group were（1.978±0.246）,（2.018±0.275）;comparison between the two,relative expression levels at the same point were significantly different（P 〈0.05）,24 h,48 h relative expression level of DcR3 protein was significantly different in experimental group（P 〈0.05）,no significant difference in control group（P 〉0.05）.Two groups of cells transfected after 48h inhibition rates were（59.8±5.4）%,（0.8±0.1）%,both had significant difference（P 〈0.05）,apoptosis rates were transfected after 24 h were（19.7±3.2）%,（6.9±0.8）%,both significant differences compared（P 〈0.05）;positive rates of caspase 3 protein were 72.8%,38.9%,the average optical density were（0.41±0.06）,（0.21±0.03）,both had significant differences（P 〈0.05）.【Conclusion】 Specific DcR3 siRNA could significantly inhibit HepG2 cells DcR3 protein expression,proliferation and induce apoptosis,the mechanism may be related to increased caspase 3 expression.

关 键 词：DCR3 RNA干扰 HEPG2细胞 凋亡 

分 类 号：R394.3[医药卫生—医学遗传学]