High levels of mitochondrial heteroplasmy modify the development of ovine - bovine interspecies nuclear transferred embryos

Abstract

To investigate the effect of mitochondrial heteroplasmy on embryo development, cloned embryos produced using bovine oocytes as the recipient cytoplasm and ovine granulosa cells as the donor nuclei were complemented with 2 pL mitochondrial suspension isolated from ovine (BOOMT embryos) or bovine (BOBMT embryos) granulosa cells; cloned embryos without mitochondrial injection served as the control group (BO embryos). Reverse transcription–quantitative polymerase chain reaction (RT-qPCR) and sodium bisulfite genomic sequencing were used to analyse mRNA and methylation levels of pluripotency genes (OCT4, SOX2) and mitochondrial genes (TFAM, POLRMT) in the early developmental stages of cloned embryos. The number of mitochondrial DNA copies in 2 pL ovine-derived and bovine-derived mitochondrial suspensions was 960 ± 110 and 1000 ± 120, respectively. The blastocyst formation rates were similar in BOBMT and BO embryos (P > 0.05), but significantly higher than in BOOMT embryos (P < 0.01). Expression of OCT4 and SOX2, as detected by RT-qPCR, decreased significantly in BOOMT embryos (P < 0.05), whereas the expression of TFAM and POLRMT increased significantly, compared with expression in BOOMT and BO embryos (P < 0.05). In addition, methylation levels of OCT4 and SOX2 were significantly greater (P < 0.05), whereas those of TFAM and POLRMT were significantly lower (P < 0.01), in BOOMT embryos compared with BOBMT and BO embryos. Together, the results of the present study suggest that the degree of mitochondrial heteroplasmy may affect embryonic development.