Effect Of Cotreatment With RepSox And LBH589 On The In Vitro Developmental Competence Of Porcine Somatic Cell Nuclear Transfer Embryos

Somatic cell nuclear transfer(SCNT)leads to embryo development retardation due to low nuclear pluripotency and abnormal epigenetic reprogramming.This leads to the abnormal development of somatic clone embryos and the low efficiency of somatic cell nuclear transfer in mammals.At present,there are few studies on simultaneous enhancement of pluripotency and epigenetic reprogramming in embryonic development.RepSox is small molecular compounds found in induced pluripotent stem cells that promote pluripotency in cells.The aim of this experiment is to combine the RepSox and histone deacetylation inhibitor to treat porcine SCNT embryos,improve the pluripotency of embryo development and the epigenetic modification,so as to improve the in vitro development competence and cloning efficiency of pig cells to clone embryos.The best deacetylation inhibitor and the best concentration of RepSox were selected by analyzing the development competence of early embryo in vitro.Analysis totipotency of embryo development by detecting pluripotency-relate genes.The levels of acetylation and methylation were detected by immunofluorescence staining in porcine early embryos to estimated epigenetic reprogramming capability,and embryos quality were determined by TUNEL staining.The following analysis results were obtained in this experiment:1.The development capacity of SCNT embryos treated with 50 nmol/L LBH589 for 24 hr higher than 5 ?mol/L M344 treated for 6 hr,0.2?mol/L MGCD0103 treated for 6 hr,2 mmol/L VPA treated for 24 hr and control groups(P<0.05).2.The rates of 2-cell,4-cell,blastocyst and total cell number had no significant difference between 12.5,25 ?mol/L RepSox treatment groups and control group.12.5,25 ?mol/L groups had a significantly development capacity than 50,100 ?mol/L groups(P<0.05).3.It was significantly higher in the group of 12.5 ?mol/L RepSox co-treatment with 50 nmol/L LBH589 than LBH589 and control about the development capacity,the expression level of pluripotency-relate genes NANOG,POU5F1 and SOX2,the level of epigenetic reprogramming in porcine SCNT embryos.Conclusion:Combination of RepSox and LBH589 can improve the development capacity and quality,the level of pluripotency gene expression and epigenetic reprogramming in porcine SCNT embryo in vitro development.