Effects Of Ghrelin On The Development Of Pigs Preimplantation Embryos And The Expression Of GHSR In Preimplantation Embryos Of Pigs And Mice

Ghrelin has been proved to play an important role in mammalian reproduction andembryonic development. However, there were few documents about ghrelin effect onmammalian preimplantation embryos, and the results in different studies werecontroversial. Here we investigated the effect of Ghrelin on the development of pigspreimplantational parthengenesis embryos and the expression pattern of ghsrinpreimplantation embryos of pigs and mice. There were two pars in this study:1. The effect of Ghrelin on the early porcine PA embryos:Porcine parthenogenesis embryos were cultured in basic culture medium PZM3supplemented with different concentration of Ghrelin (0ng/ml (contral),0.1ng/ml,1ng/ml,10ng/ml and100ng/ml). Then, the cleaved rates, the blastocyst rates and thedevelopmental competences were observed. Result showed that the4-cell ratee,8-cell rate, morula rate and blastocyst rate of porcine PA embryos cultured in PZM3supplemented with10ng/ml ghrelin were significantly higher than contral group (P <0.05); the4-cell ratee and8-cell rate of PA embryos cultured with100ng/ml ghrelinwere significantly higher than0.1ng/ml group (P <0.05); no significant differencesbetween any two groups in the cleaved rate (P>0.05).2. The expression of ghsrin different stages of pre-implantation embryos.The transcription of ghsrmRNA and the expression of protein in differentdevelopmental stages of embryos, which included porcine PA embryos, mouse invivo fertilized embryos, IVF embryos and PA embryos before implantation, werechecked using semi-quantitative pcr and Immunofluorescence technique. Resultsshowed that:(1). The mRNA transcription of ghsrwas observed at different early stage of porcinePA embryo (2-Cell,4-Cell,8-Cell, morula and blastocyst), which down-regulated alittle at4-Cell stage, while it was highest at8-Cell stage. Afterwords, thetranscription level of ghsrwould decrease sequentially during following development,and the down-regulation would be significant at blastocyst; The analysis ofimmunostaining has detected the protein of ghsrat early stage of PA embryo duringdevelopment, while nothing obviously difference was found between each stage.(2). Prior to implantation, the mRNA expression of ghsr could be detected at eachstage of mouse PA embryo, which level was significantly higher than naturallyfertilized embryo and IVF embryo at the same stage of development. The level ofmRNA would decrease continuously from2-Cell stage to8-Cell stage, then it would upgrade until blastocyst. For the protein of ghsr, result of immunostaining hasrevealed their expression in each early stage of mouse PA embryo duringdevelopment.(3).As to mouse IVF embryo, the mRNA of ghsr and protein of GHSR were observedat its each early stage of development, and the transcription level of ghsr was highestat morula, and the expression level of ghsrwas highest at Blastcoyst.(4) For the naturally fertilized embryo of mouse,the mRNA transcription of ghsr andprotein of GHSR were observed at its each early stage of development, and thetranscriptional level of ghsr was lowst at morula.