Reproduction Abstracts

Introduction: The poly(ADP-ribosyl)ation (PARylation) has involvement with pro-survival autophagy. However, there has been no report about PARylation and autophagy during pre-implantation development. Here, we investigated the mechanism between PARylation and pro-survival autophagy in pre-implantation development.

Materials and methods: IVF embryos were cultured in the medium containing 3-aminobenzamide (3ABA), parp inhibitor. Autophagy and PARylation were evaluated by immunocytochemistry and quantitative real time-PCR in pig embryos.

Results and discussions: The expression level of Parp-1 mRNA was increased in 6 hours of post-insemination (hpi) and 12 hpi embryos and decreased from 20 hpi to blastocyst stage. The distribution of PAR was diffused in cytoplasm at metaphase II oocytes. After fertilization, PAR localized in nucleus was transiently induced at 6 and 12 hpi, however, the PAR was restored in cytoplasm again from 20 hpi to blastocyst. To investigate the effect of PARP on embryo development, the treatment of 3ABA from one cell stage to blastocyst. There was no significant difference of development to the morula stage (20.77±4.19 vs 24.55±5.19%), whereas the blastocyst development was significantly reduced (3.63±2.65 vs 12.26±3.37%). The blastocyst formation was significantly reduced by inhibition of PARP in morula stage (19.64±4.61 vs 41.39±5.26%), especially expanded blastocysts (4.69±2.99 vs 28.1±6.06%). The expression level of autophagy related genes (Atg5, Beclin1 and Lc3) and LC3 protein synthesis were reduced in 3ABA treated blastocysts. In addition, total cell number of 3ABA treated blastocysts was significantly lower than control group and apoptosis was increased. Collectively, these data suggest that PARylation may be involved in pro-survival autophagy and embryo quality in pig IVF embryos.

Keywords: Autophagy; Poly(ADP-ribosyl)ation; pig embryos; blastocyst.