Fatty acid induced lipolysis influences embryo development, gene expression and lipid droplet formation in the porcine cumulus cells†.

The pig oocyte maturation protocol differs from other mammalian species due to dependence on follicular fluid (FF) supplementation. One of the most abundant components of the porcine follicular fluid are fatty acids (FAs). Although evidence from other mammalian models revealed a negative impact of saturated fatty acids (SFA) on developmental competence of oocytes, pig has not yet been widely analyzed. Therefore, we aimed to investigate whether supplementation of IVM medium with 150 µM of stearic acid (SA) and oleic acid (OA) affects lipid content and expression of genes related to fatty acid metabolism in porcine cumulus-oocyte complexes and parthenogenetic embryo development. We found significant influence of fatty acids on lipid metabolism in cumulus cells without affecting the oocyte proper. The expression of ACACA, SCD, PLIN2, FADS1, and FADS2 genes was upregulated (P < 0.01) in cumulus cells, while their expression in oocytes did not change. The increase in gene expression was more pronounced in the case of OA (e.g., up to 30-fold increase in PLIN2 transcript level compared to the control). The number of lipid droplets and occupied area increased significantly in the cumulus cells and did not change in oocytes after SA treatment. Oleic acid improved the blastocyst rate (48 vs 32% in control), whereas stearic acid did not affect this parameter (27%). Additionally, we have discovered a phenotypic diversity of LD in cumulus cells in response to FA supplementation, suggesting extensive lipolysis in response to SA. Stearic acid excess in maturation media led to the formation of multiple micro lipid droplets in cumulus cells.

The consequences of porcine IVM medium supplementation with follicular fluid become reflected in embryo quality, yield and gene expression patterns.

Oocyte and embryo developmental competence are shaped by multiple extrinsic and intrinsic factors. One of the most extensive research areas in the last decade is the regulation of lipid metabolism in oocytes and embryos of different species. We hypothesized that differences in developmental competence of oocytes and embryos between prepubertal and cyclic gilts may arise due to distinct fatty acid profiles in follicular fluid. We found that supplementation of oocyte maturation media with follicular fluid from prepubertal pigs affected quality and development of embryos from prepubertal pigs while embryos of cyclic pigs were not affected. PLIN2, SCD and ACACA transcripts involved in lipid metabolism were upregulated in embryos originating from oocytes of prepubertal pigs matured with autologous follicular fluid. The surface occupied by lipid droplets tend to increase in oocytes matured with follicular fluid from prepubertal pigs regardless oocyte origin. The change into follicular fluid of cyclic pigs increased the efficiency of embryo culture and improved quality, while gene expression was similar to embryos obtained from cyclic gilts. We assume that the follicular fluids of prepubertal and cyclic pigs influenced the quality of oocytes and embryos obtained from prepubertal pigs which are more susceptible to suboptimal in vitro culture conditions.

Mitochondria and mitochondrial DNA in porcine oocytes and cumulus cells--A search for developmental competence marker.

The development of mammalian oocytes is dependent on bidirectional signaling with the surrounding cumulus cells. Among the numerous factors that contribute to oocyte developmental competence, the mitochondria and the mitochondrial DNA play pivotal roles. Although these highly abundant organelles have been well-studied in oocytes, their roles, abundance and metabolism remain elusive in cumulus cells. Therefore, the aim of our study was to analyze the correlation between the mtDNA copy number in cumulus cells and oocytes, as well as the mitochondrial distribution patterns in oocytes, using two groups of animals that differ in terms of the developmental competence of their oocytes. We determined a positive correlation between the mtDNA copy number in the cumulus cells and mtDNA copy number in oocytes of prepubertal pigs and negative correlation in cyclic gilts. These opposing correlations may reflect the differences in the developmental competence of the prepubertal and cyclic oocytes. We also hypothesize that observed differences may reflect different metabolism and energy requirements of the cumulus-oocyte complexes from prepubertal and cyclic gilts. The mitochondrial distribution patterns in the prepubertal and cyclic gilts were not different.