Studies On The Effect Of Small Chemical Compounds And HiPS Medium On Pig Somatic Cell Nuclear Transplantation

So far, the cloned pigs via somatic cell nuclear transfer (SCNT) have extensive applications in animal husbandry and human clinics. However, the efficiency of SCNT has been low, limiting the applications of cloned pigs. The cloning efficiencies may be attributed to many factors, such as incomplete epigenetic modifications, inappropriate culture condition, and chromosomal abnormalities, etc. In the present study, we first investigated the effect of Lomeguatrib on developmental competence of porcine NT embryos. Moreover, pig SCNT early blastocysts were cultured in human-induced pluripotent stem cell medium and analyzed this interesting result. The results of the present study are as follows:1. In this chapter, pig SCNT embryos were treated with different concentrations of Lomeguatrib for 24 h, and pig SCNT embryos were treated with optimal concentration for different time. The result showed that treatment with 100 nM Lomeguatrib significantly improved the proportion of porcine SCNT embryos. Compared with the control groups or treatment with 50 nM or 200 nM Lomeguatrib, the blastocyst rate was higher (18.3% vs.9.8%,12.3% and 5.2%, respectively, P< 0.05). Also, pig SCNT embryos treated with 100 nM Lomeguatrib for 24 h has a higher proportion of embryos reaching the blastocyst stage in comparison to the control groups or embryos treated for 12 h or 48 h (8.0% vs.13.7%,4.7% and 11.1%, respectively). Pig SCNT embryos treated with 100 nM for 24 h were transferred into two surrogate mothers, resulting in one pregnancy. These results suggested that Lomeguatrib can give positive impacts on the in vitro developmental competence of pig somatic cell nuclear transfer embryos.2. Here, we produced and evaluated blastocysts from two different in vitro culture (IVC) systems, namely one-step and two-step culture systems. In the one-step culture system, porcine NT blastocysts were cultured in North Carolina State University 37 medium for 7 days, with a change of Day 5. In the two-step culture system, on the other hand, porcine NT embryos were cultured in NCSU-37 medium for 5 days, followed by an additional 2 days in hiPS medium (7 days in total). Subsequently, we examined the morphological quality of porcine blastocysts from these two IVC systems. Differences in the ratios of extruding embryonic diameter, perimeter and area from the zona to the embryonic diameter, and perimeter and area inside the zona were observed between one-step and two-step culture systems (3.01±0.20 versus 1.59±0.19, P<0.01; 3.30±0.06 versus 1.38±0.18, P<0.01; and 10.23±0.33 versus 1.12±0.37, P<0.01, respectively). Thus, we could obviously discover that blastocysts obtained from the two-step IVC system were larger than those obtained from the one-step. Furthermore, the number of cells in blastocysts obtained from the two-step system was 3-fold higher than those obtained from the one-step system. These results indicate that hiPS medium can significantly improve the morphology and quality of porcine blastocysts using somatic cell nuclear transfer.