Zhang Tianrui, Zheng Yingying, Kuang Tianya, Yang Lianyu, Jiang Hailong, Wang Heming, Zhao Yicheng, Han Rui, Che Dongsheng
Key Laboratory of Animal Production, Product Quality and Security of Ministry of Education, Jilin Provincial Key Laboratory of Animal Nutrition and Feed Science, College of Animal Science and Technology, Jilin Agricultural University, Changchun, China.
Department of Gastroenterology and Hepatology, Zhongshan Hospital, Fudan University, Shanghai, China.
Front Vet Sci. 2022 Jun 23;9:921406. doi: 10.3389/fvets.2022.921406. eCollection 2022.
Arginine has a positive effect on pre-implantation development in pigs. However, the exact mechanism by which arginine promotes embryonic development is undefined. Here, single-cell RNA sequencing technology was applied to porcine pre-implantation embryos from the zygote to morula stage, it was found that that the expression of arginine metabolism-related genes clearly changed from the 2-cell stage to the 4-cell stage, when zygotic genome activation (ZGA) occurs in porcine embryos. Further analysis showed that arginine metabolism-related genes are significantly correlated with key ZGA genes. To determine the function of arginine in porcine embryos during ZGA, the fertilization embryos were cultured in PZM-3 medium (0.12 mM arginine, Control group), a modified PZM-3 medium (0 mM arginine, Block group) and a modified PZM-3 medium supplemented with arginine (0.12 mM arginine, Block + Arg group). The results showed that the 4-cell arrest rate was significantly increased in the Block group compared to the Control group ( < 0.05). The 4-cell arrest rate in the Block + Arg group was significantly decreased than that in the Block group ( < 0.05). Meanwhile, the expression of ZGA marker genes and SIRT1 protein in 4-cell embryos was significantly decreased in the Block group compared to the Control group, and their expression was significantly increased in the Block + Arg group. In addition, we observed that the glutathione (GSH), ATP levels, and lipid droplet contents were significantly increased, and the reactive oxygen species (ROS) level was decreased in the Block + Arg group compared to the Block group. Compared with Control group, spermine content in culture medium and the mRNA expression of ornithine decarboxylase1 (ODC1) of embryos in the Block group were significantly decreased ( < 0.05), and those in the Block + Arg group were significantly increased compared with the Block group ( < 0.05). Moreover, when difluoromethylornithine (an inhibitor of ODC1) was added to the modified PZM-3 medium supplemented with arginine, the effect of arginine on ZGA was inhibited. In summary, our findings demonstrated that arginine may regulate ZGA under nutrition restriction in porcine embryos by promoting polyamine synthesis.
精氨酸对猪的植入前胚胎发育具有积极作用。然而，精氨酸促进胚胎发育的确切机制尚不清楚。在此，将单细胞RNA测序技术应用于从合子到桑葚胚阶段的猪植入前胚胎，发现精氨酸代谢相关基因的表达在猪胚胎发生合子基因组激活（ZGA）的2细胞阶段到4细胞阶段明显改变。进一步分析表明，精氨酸代谢相关基因与关键ZGA基因显著相关。为了确定ZGA期间精氨酸在猪胚胎中的功能，将受精胚胎在PZM - 3培养基（0.12 mM精氨酸，对照组）、改良的PZM - 3培养基（0 mM精氨酸，阻断组）和添加精氨酸的改良PZM - 3培养基（0.12 mM精氨酸，阻断 + 精氨酸组）中培养。结果显示，与对照组相比，阻断组的4细胞停滞率显著增加（<0.05）。阻断 + 精氨酸组的4细胞停滞率比阻断组显著降低（<0.05）。同时，与对照组相比，阻断组4细胞胚胎中ZGA标记基因和SIRT1蛋白的表达显著降低，而在阻断 + 精氨酸组中它们的表达显著增加。此外，我们观察到，与阻断组相比，阻断 + 精氨酸组的谷胱甘肽（GSH）、ATP水平和脂滴含量显著增加，活性氧（ROS）水平降低。与对照组相比，阻断组培养基中的精胺含量和胚胎鸟氨酸脱羧酶1（ODC1）的mRNA表达显著降低（<0.05），而阻断 + 精氨酸组与阻断组相比显著增加（<0.05）。此外，当将二氟甲基鸟氨酸（ODC1的抑制剂）添加到添加精氨酸的改良PZM - 3培养基中时，精氨酸对ZGA的作用受到抑制。总之，我们的研究结果表明，精氨酸可能通过促进多胺合成在营养限制条件下调节猪胚胎中的ZGA。
