College of Veterinary Medicine, Chungnam National University, 34134, Daejeon, Republic of Korea; Faculty of Veterinary Medicine, Hawassa University, 05, Hawassa, Ethiopia.
College of Veterinary Medicine, Chungnam National University, 34134, Daejeon, Republic of Korea.
Theriogenology. 2022 Apr 15;183:90-97. doi: 10.1016/j.theriogenology.2022.02.019. Epub 2022 Feb 22.
The in vitro embryo's competence is lower than in vivo counterparts and miRNAs found to affect the developmental competence, affecting pluripotency, stress level and apoptosis in embryos. We aimed to investigate the effect of miRNA-155 on parthenogenically activated early development porcine embryos at the preimplantation blastocyst stage. We designed miRNA-155 mimics and inhibitors and microinjected them into post-activated oocytes. The embryonic cleavage rate, blastocyst rate, and embryo development quality in terms of stress and apoptosis levels were investigated by confocal microscopy. Furthermore, we selected target genes, analyzed gene interaction and prediction networks, and compared the gene expression level in treatments and controls. miRNA-155 inhibition improved in vitro developmental competence by increasing cell numbers and reducing stress and apoptosis levels. The cleavage rate in the miRNA-155 inhibitor group was significantly higher (P < 0.05) than that in the miRNA-155 mimic group, but not in the control, whereas the blastocyst rate of the miRNA-155 inhibitor group was statistically significantly higher (P < 0.05) than in both control and miRNA-155 mimic groups. The relative gene expression level analysis showed downregulation of mRNAs related to stress and apoptosis, BAX, and the stress-induced autophagy gene ATF4, and TNF-ɑ in the miRNA-155 inhibitor group. Moreover, miRNA-155 inhibition showed upregulation of the relative expression of OCT4, ZEB2, BCL2, and IL-1 mRNA compared to control and mimic groups. However, the miRNA-155 mimic-injected group showed lower cleavage rates and blastocyst development rates than the other two groups. In conclusion, miRNA-155 inhibition in porcine in vitro embryos improved their preimplantation developmental competence and in vitro embryo production.
体外胚胎的能力低于体内对应物，并且已经发现 miRNA 会影响胚胎的发育能力，影响多能性、应激水平和凋亡。我们旨在研究 miRNA-155 对孤雌激活的早期发育猪胚胎在植入前囊胚阶段的影响。我们设计了 miRNA-155 模拟物和抑制剂，并将它们微注射到激活后的卵母细胞中。通过共聚焦显微镜研究胚胎分裂率、囊胚率以及胚胎发育质量（应激和凋亡水平）。此外，我们选择了靶基因，分析了基因相互作用和预测网络，并比较了处理组和对照组的基因表达水平。miRNA-155 抑制通过增加细胞数量和降低应激和凋亡水平来提高体外发育能力。miRNA-155 抑制剂组的分裂率显著高于 miRNA-155 模拟物组（P<0.05），但对照组则不然，而 miRNA-155 抑制剂组的囊胚率显著高于对照组和 miRNA-155 模拟物组（P<0.05）。相对基因表达水平分析显示，应激和凋亡、BAX 以及应激诱导的自噬基因 ATF4 和 TNF-ɑ 的相关 mRNAs 在 miRNA-155 抑制剂组下调。此外，与对照组和模拟物组相比，miRNA-155 抑制剂组中 OCT4、ZEB2、BCL2 和 IL-1 mRNA 的相对表达上调。然而，miRNA-155 模拟物注射组的分裂率和囊胚发育率均低于其他两组。总之，抑制猪体外胚胎中的 miRNA-155 可提高其植入前发育能力和体外胚胎生产。
