Park Sae Young, Kim Eun Young, Jeon Kilsoo, Cui Xiang-Shun, Lee Won Don, Kim Nam-Hyung, Park Se Pill, Lim Jin Ho
Maria Infertility Hospital Medical Institute/Maria Biotech, Seoul, and Chungbuk National University, Chungbuk, South Korea.
Mol Reprod Dev. 2007 May;74(5):582-90. doi: 10.1002/mrd.20641.
Survivin, an inhibitor of apoptotic protein containing a single baculoviral inhibit apoptotic protein repeat domain, is a bifunctional protein that suppresses apoptosis and regulates cell division. Thus, we used double stranded RNA (dsRNA) interference to manipulate survivin expression in bovine embryos and analyze its role in blocking apoptosis and facilitating development of pre-implantation embryos. In vitro fertilized embryos (1-cell) were injected with survivin dsRNA, and expression of survivin mRNA was evaluated by real-time quantitative RT-PCR. To analyze survivin protein expression, we performed immunocytochemistry using a rabbit anti-bovine suvivin antibody. Expression levels of survivin mRNA and protein were decreased in the dsRNA group compared to the sham group. Rates of in vitro blastocyst development were lower in the survivin dsRNA-injected group than in the sham-injected group. Also, the total cell number seen in blastocysts was decreased in the dsRNA group. TUNEL assays of DNA fragmentation indicated an increased apoptotic index in the dsRNA group compared to the sham group. These results indicate that survivin is important for optimal development of bovine blastocysts and confirm that survivin expression suppresses apoptosis of pre-implantation embryos.
存活素是一种含有单个杆状病毒凋亡抑制蛋白重复结构域的凋亡抑制蛋白，是一种双功能蛋白，可抑制细胞凋亡并调节细胞分裂。因此，我们使用双链RNA（dsRNA）干扰来调控牛胚胎中存活素的表达，并分析其在阻止细胞凋亡和促进植入前胚胎发育中的作用。将体外受精胚胎（1细胞期）注射存活素dsRNA，通过实时定量RT-PCR评估存活素mRNA的表达。为了分析存活素蛋白的表达，我们使用兔抗牛存活素抗体进行免疫细胞化学检测。与假注射组相比，dsRNA组中存活素mRNA和蛋白的表达水平均降低。注射存活素dsRNA组的体外囊胚发育率低于假注射组。此外，dsRNA组囊胚中的总细胞数减少。DNA片段化的TUNEL检测表明，与假注射组相比，dsRNA组的凋亡指数增加。这些结果表明，存活素对牛囊胚的最佳发育很重要，并证实存活素的表达可抑制植入前胚胎的细胞凋亡。
