The Study On C-type Natriuretic Peptide Influencing Bovine Oocyte Matured In Vitro And The Expression Of Imprinted Genes And The Apoptosis In Bovine Early Embryos

In vitro maturation (IVM) and in vitro culture (IVC) of bovine oocyte are the key technologies of the reproductive biology which used in livestock production widely. In order to improve the quality of the in vitro maturation of bovine oocyte and the early development embryo, the quality of bovine oocyte pretreated with C-type natriuretic peptide (CNP) which is an oocyte meiosis inhibitor was studed in this paper, and the effects of different oxygen concentrations on embryos apoptosis and the differential expression of the maternal and paternal imprinted genes in early bovine embryo developed in vitro were also studied.The results were as follows:1. To investigated the maturation of bovine oocyte pretreated by CNP for6h, then matured for28h, compared to conventional method of bovine oocytes maturated for24h in vitro. we found that the distribution of mitochondria, the mitochondrial copy number level and the quality of blastocysts derived from matured oocyte of treatment group were superior to the conventional group, indicating indicated that pretreatment by CNP can effectively improve the quality of bovine oocytes matured in vitro.2. The ROS level and GSH concentration of oocytes obtained from conventional maturation culture or CNP pretreated treatment were detected. The results showed that the ROS level of bovine oocytes increased after pretreated for6h and then matured for28h, but the fertilization ability of oocyte and embryo development could be protected due to the scavenge effect of GSH, indicated that the bovine oocytes maturated by two-step process with CNP pretreatment did not cause oocyte aging.3. The effect of different oxygen concentrations on apoptosis and the apoptotic gene expression levels of bovine early embryonic cells were studied. The result demonstrated that hypoxia environment coud be more beneficial for the development of bovine IVF embryos, moreover, oxygen concentration of the bovine embryos culture environment could influence the expression of apoptosis-related genes, such as Mcl-1、Bad、Bak、Bik and Caspase-3, and the products of these genes might synergistically affect the apoptosis of the embryonic cells.4. To detect the expression levels of three paternal imprinted genes Gnas, Grb10, Xist and three maternal imprinted genes Mage12, Sgce and Snrpn in bovine parthenogenetic (PA) and in vitro fertilization (IVF) embryos by Real-time PCR technique. The results showed that there were some differences in the expression patterns between paternal and maternal imprinting gene during the development of IVF and PA embryos, which might act an important role for the normal development of bovine early embryos.In conclusion, two-step process using CNP pretreatment can effectively improve the quality of bovine oocytes matured in vitr. Hypoxic environment can reduce the apoptosis during the development of bovine embryos in vitro, and more beneficial to the development of bovine IVF embryos. The normal development of bovine early embryo is the results of the interaction of the culture conditions, apoptotic genes and imprinted genes.