Effect of the coculture of porcine luteal cells during in vitro maturation on the DNA methylation pattern of porcine blastocysts

Abstract

In pigs, the in vitro embryo development is still an inefficient biotechnology. The coculture with somatic cells is an alternative to improve suboptimal in vitro culture conditions. Aberrant DNA methylation patterns of genes required for development are common in in vitro produced embryos and may indicate carryover effects of suboptimal culture conditions on epigenetic signatures. In the present study, we investigated the potential effect of a coculture system of porcine luteal cells during in vitro maturation (IVM) on blastocyst development and DNA methylation of embryos using the EmbryoGENE DNA Methylation Array (EDMA). The cumulus-oocyte complexes were matured in vitro in TCM199 supplemented with human menopausal gonadotrophin (control) or in coculture with porcine luteal cells from passage 1 (PLC-1). The coculture with PLC-1 significantly increased the blastocyst rate (blastocyst/cleavage) (control: 14.4%; PLC-1 21.2%; Fisher´s test p<0.05). DNA methylome array was used to identify differentially methylated regions between the control and the coculture. Among the over 180000 probes on the DNA microarray, the signal intensities of 57075 probes were found to be higher than the background signal in all four arrays of a given condition, and 49936 were commonly detected in the two sample groups. Differential methylated regions (DMRs) analysis indicated that the coculture with PLC-1 during IVM has a very mild global impact on the DNA methylation profile; a similar number of hypermethylated probes were significant in each condition (245 in control vs 268 in treatment). In the coculture group, more regions classified as hypermethylated were present in distal promoters, promoters, exonic and intronic than in the control. Also, a similar number of hypermethylated regions were present in repetitive elements, but low-complexity repetitive elements were more hypermethylated in the coculture. More hypermethylated regions in the control were located in CpG shore, CpG shelf, and open sea. On the other hand, the coculture exhibited more hypermethylated regions in CpGs islands. Therefore, this finding indicates that the culture condition during IVM induces changes in the DNA methylation landscape of the resulting blastocysts and affects embryo development.