Salilew-Wondim Dessie, Hölker Micheal, Rings Franca, Phatsara Chirawath, Mohammadi-Sangcheshmeh Abdollah, Tholen Ernst, Schellander Karl, Tesfaye Dawit
Institute of Animal Science, Animal Breeding and Husbandry Group, University of Bonn, Endenicher Allee 15, Bonn, Germany.
Reprod Fertil Dev. 2010;22(3):564-79. doi: 10.1071/RD09112.
Baculoviral inhibitors of apoptosis repeat-containing 6 (BIRC6) is believed to inhibit apoptosis by targeting key cell-death proteins. To understand its involvement during bovine preimplantation embryo development, two consecutive experiments were conducted by targeted knockdown of its mRNA and protein using RNA interference. In Experiment 1, the effect of BIRC6 knockdown during the early stages of preimplantation embryo development was assessed by injecting zygotes with long double-stranded RNA (ldsRNA) and short hairpin RNA (shRNA) against BIRC6 mRNA followed by in vitro culturing until 96 h post insemination (hpi). The results showed that in RNA-injected zygote groups, reduced levels of BIRC6 mRNA and protein were accompanied by an increase (P < 0.05) in the proportion of 2- and 4-cell and uncleaved embryos and a corresponding decrease (P < 0.05) in the number of 8-cell embryos. In Experiment 2, the effect of BIRC6 knockdown on blastocyst formation, blastocyst total cell number and the extent of apoptosis was investigated. Consequently, zygotes injected with ldsRNA and shRNA resulted in lower (P < 0.05) blastocyst formation and total blastocyst cell number. Moreover, the apoptotic cell ratio, CASPASE 3 and 7 activity, BAX to BCL-2 ratio and levels of SMAC and CASPASE 9 were higher in blastocysts derived from the ldsRNA and shRNA groups, suggesting increased apoptosis in those blastocysts. The results of this study reveal the importance of BIRC6 expression for embryo survival during bovine preimplantation embryo development. However, whether BIRC6 is essential for implantation and fetal development during bovine pregnancy needs further research.
杆状病毒凋亡抑制重复序列蛋白6（BIRC6）被认为通过靶向关键细胞死亡蛋白来抑制细胞凋亡。为了解其在牛植入前胚胎发育过程中的作用，通过RNA干扰靶向敲低其mRNA和蛋白进行了两个连续的实验。在实验1中，通过向受精卵注射针对BIRC6 mRNA的长双链RNA（ldsRNA）和短发夹RNA（shRNA），然后进行体外培养至授精后96小时（hpi），评估BIRC6敲低在植入前胚胎发育早期阶段的影响。结果表明，在注射RNA的受精卵组中，BIRC6 mRNA和蛋白水平降低，同时2细胞和4细胞以及未分裂胚胎的比例增加（P < 0.05），而8细胞胚胎的数量相应减少（P < 0.05）。在实验2中，研究了BIRC6敲低对囊胚形成、囊胚总细胞数和凋亡程度的影响。因此，注射ldsRNA和shRNA的受精卵导致囊胚形成率和囊胚总细胞数降低（P < 0.05）。此外，来自ldsRNA和shRNA组的囊胚中凋亡细胞比例、CASPASE 3和7活性、BAX与BCL-2比例以及SMAC和CASPASE 9水平较高，表明这些囊胚中的细胞凋亡增加。本研究结果揭示了BIRC6表达在牛植入前胚胎发育过程中对胚胎存活的重要性。然而，BIRC6在牛妊娠期间对植入和胎儿发育是否至关重要，还需要进一步研究。
