caspase-8靶向小发夹RNA抑制肝细胞凋亡的研究  

The inhibitive effects of shRNA for caspase-8 on Hepa 1-6 cells apoptosis

作　　者：林森[1] 刘霞[2] 孔德晓[1] 曲云东[1] 张利宁[3] 张源朝[4] 

机构地区：[1]山东大学第二医院消化科,山东济南250033 [2]山东大学齐鲁儿童医院儿研所,山东济南250022 [3]山东大学医学免疫研究所,山东济南250012 [4]山东省立医院风湿免疫科,山东济南250021

出　　处：《中国病原生物学杂志》2008年第10期728-730,724,F0004,共5页Journal of Pathogen Biology

摘　　要：目的研究针对caspase-8 mRNA的小发夹RNA(shRNA)真核表达载体对肝Hepa1-6细胞凋亡的干扰作用。方法TNF-α诱导Hepa1-6细胞凋亡,脂质体法将shRNA表达载体转染入Hepa1-6细胞中,24h后应用荧光实时定量PCR和Western blot法分别在mRNA和蛋白水平测定其caspase-8表达差异,流式细胞仪观察细胞凋亡的变化。结果TNF-α作用后Hepa1-6细胞caspase-8 mRNA由(0.050±0.006)升高至(0.286±0.063)(P<0.01),而shRNA表达载体Pgenesil-casp8使其下降至(0.098±0.037)(P<0.01)。转染Pgenesil-casp8重组质粒后caspase-8蛋白表达与mRNA荧光实时定量PCR结果吻合。TNF-α诱导Hepa1-6细胞凋亡率由(4.70±0.89)%升高至(17.40±2.21)%(P<0.01),Pgenesil-casp8作用后下降至(1.20±0.32)%(P<0.05)。结论针对caspase-8 mRNA的shRNA真核表达载体可有效抑制肝细胞Hepa1-6凋亡。Objective To explore the interfering effects of small hairpin RNA （shRNA） eukaryotic expression vector for caspase-8 mRNA on apoptosis of Hepa 1-6 cells. Methods The apoptosis model of Hepa 1-6 cells was constructed using TNF-α, and the eukaryotic expression vector Pgenesil-casp8 expressing shRNA for caspase-8 mRNA were transfect- ed to Hepa 1-6 cell by lipofectamine 2000. After 24 hours, the expression of caspase-8 mRNA and protein were detected by Real-time Quantitative PCR and Western blot. And the apoptosis of Hepa 1-6 cells was observed by flow cytometry. Results The expression of caspase-8 mRNA in Hepa 1-6 cells was （0. 050±0. 006）, which was increased to （0. 286±0. 063） （P〈0.01） after treated with TNF-m After applying for Pgenesil-casp8, expression of caspase-8 mRNA were decreased to （0. 098±0. 037） （P〈0.01）. And the results of Western blot were identical to the change of caspase-8 mRNA. The apoptosis in Hepa 1-6 cells induced by TNF-α increased to （17.40±2.21）% from （1. 20±0.32）%, while decreased to （4. 70±0.89）% （P〈0.05） after treated with Pgenesil casp8. Conclusion The expression vectors of shRNA for easpase-8 mRNA inhibit the apoptosis of Hepa 1-6 cells induced by TNF-α efficiently.

关 键 词：CASPASE-8 RNA干扰 凋亡 

分 类 号：R392[医药卫生—免疫学]