白藜芦醇和褪黑素对绵羊胎儿成纤维细胞转染及卵母细胞体外成熟的影响

doi:10.16431/j.cnki.1671-7236.2021.12.018

Abstract

Abstract: This experiment was aimed to use resveratrol (Re) and melatonin (MT) to improve the efficiency of electroporation transfection of sheep fetal fibroblasts, and optimize the in vitro maturation system of oocytes, in order to improve the production efficiency of transgenic animals.Sheep fetal fibroblasts were divided into control group, high, medium and low concentrations of resveratrol groups (R^-5, R^-6 and R^-7), resveratrol and melatonin combined groups (R^-5M^-5and R^-6M^-7), 6 groups in total.The optimization scheme of electrotransfer system was determined by electrotransfection efficiency and cell apoptosis.The mechanism was studied by detecting cell cycle, mitochondrial membrane potential and reactive oxygen species (ROS).Sheep oocytes were divided into control group and combined addition group of resveratrol and melatonin (R^-5M^-5, R^-6M^-7), the effect of ovine oocytes on in vitro maturation was detected by cumulus cell expansion, first polar body excretion and cleavage rate of in vitro fertilized embryos.The results showed that the P3 generation of isolated sheep fetal fibroblasts proliferated vigorously, and the cell growth curve showed a typical S shape.When compared with control group, the electrotransfection efficiency of sheep fetal fibroblasts in all experimental groups was significantly increased (P<0.05), R^-5M^-5 group had the highest electrotransfection efficiency, R^-6M^-7 group significantly decreased the cell apoptosis rate (P<0.05), R^-5, R^-6, R^-5M^-5 and R^-6M^-7 groups significantly promoted the cell proliferation (P<0.05), R^-6, R^-7, R^-5M^-5 and R^-6M^-7 groups increased the proportion of G1 phase cells, while S phase cells decreased relatively, but the difference was not significant (P>0.05).ROS level was significantly decreased in R^-6, R^-7, R^-5M^-5 and R^-6M^-7 groups (P<0.05), and mitochondrial membrane potential was significantly increased in R^-6, R^-5M^-5 and R^-6M^-7 groups (P<0.05).R^-6M^-7 group significantly promoted the expansion of cumulus cells, and significantly increased the rate of first polar body extrusion and the development ability of in vitro fertilization embryo (P<0.05).In conclusion, resveratrol and melatonin could optimize the electroporation transfection system, improve the in vitro maturation efficiency of oocytes, and provide references for the application of resveratrol and melatonin in the production of transgenic animals.

Key words: resveratrol; melatonin; sheep; fetal fibroblast; oocytes

CLC Number:

S828

YAO Yujun, LI Guangdong, WU Hao, MA Wenkui, YANG Hai, GUAN Shengyu, LYU Dongying, FU Yao, ZHU Tianqi, JI Pengyun, TAN Xinxing, ZHAO Wanmin, LIAN Zhengxing, ZHANG Lu, LIU Guoshi. Effects of Resveratrol and Melatonin on Transfection of Sheep Fetal Fibroblasts and in vitro Maturation of Oocytes[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(12): 4497-4507.

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Effects of Resveratrol and Melatonin on Transfection of Sheep Fetal Fibroblasts and in vitro Maturation of Oocytes

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