TSA处理供体细胞对组蛋白乙酰化和核重编程效果的影响  

Effect of Trichostatin A on Reprogramming of Bovine Fetal Fibroblast Cells

作　　者：张东[1] 杨鹭[1] 王勇胜[1] 刘根胜[1] 刘利杰[1] 万敏[1] 张涌[1] 

机构地区：[1]西北农林科技大学生物工程研究所,杨凌712100

出　　处：《畜牧兽医学报》2009年第7期1007-1012,共6页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家"863"计划资助项目(2004AA213072)

摘　　要：克隆胚胎基因组的不完全重编程是克隆动物成功率低的主要原因。试验中以第5代牛胎儿成纤维细胞作为供体核,以牛卵母细胞作为受体胞质进行体细胞核移植,用75 nmol.L-1曲古抑菌素A(Trichostatin A,TSA)分别处理供体细胞6、12和24 h,通过核移植检测克隆胚胎发育率,并应用激光共聚焦显微镜技术和流式细胞术检测处理细胞和克隆囊胚组蛋白H3K18乙酰化水平和细胞周期。结果显示:随着TSA处理时间的延长,供体细胞组蛋白H3K18乙酰化水平不断提高;以75 nmol.L-1TSA处理供体细胞12 h的克隆胚的囊胚发育率显著高于未处理组(23.5%vs 15.7%,P<0.05);供体细胞经TSA处理的克隆囊胚组蛋白H3K18乙酰化水平与未处理组相比差异不显著(P>0.5);处理组和对照组细胞G0/G1期和S期比例间存在显著差异(P<0.05)。结论:TSA对核供体细胞的处理存在时间效应,75 nmol.L-1TSA处理12 h的牛胎儿成纤维细胞更易被卵母细胞重编程,显著提高了克隆胚的体外发育能力,初步证实TSA是通过提高供体细胞组蛋白乙酰化水平来促进供体细胞重编程的。It has been widely suggested that poor ability of oocyte cytoplasmic factors to globally erase the epigenetic modifications of a somatic cell is the major reason of the inefficiency of nuclear transfer. In this study, bovine fetal fibroblasts passaged 5 times were seeded into DMEM plus 10% FBS containing 75 nmol·L^-1 of TSA for 6, 12, 24 h , respectively. These levels were selected to produce time dependent effects. Cells were subsequently subjected to essay for histone acetylation of H3K18 levels by confocal microscope. The effect of Trichostatin A treatment donor cells on in vitro development of bovine nuclear transfer （NT） embryos which were constructed with bovine fetal fibroblast cells and cell cycle was investigated. These results showed that TSA increased the levels of histone acetylation of H3K18 after bovine fetal fibroblast cells treated with 75 nmol·L^-1 TSA for 12 or 24 h（P〈0.05） ;75 nmol·L^-1 TSA treatment of donor cells for 12 h promoted blastocyst development compared to control （23.5% vs15.7%, P〈0.05） ; Marked difference existed in the percentage of cells at G0/G1 and S stages between treated and control groups（P〈0.05）. It was concluded that TSA enhances the reprogramming in terms of high histone acetylation of doner cells.

关 键 词：核移植 曲古抑菌素A 重编程 激光共聚焦显微术 

分 类 号：S823[农业科学—畜牧学] S814.8[农业科学—畜牧兽医]