The role of trophoblastic items in oocyte culture

Aim: In this study we aimed to investigate the role of trophoblastic items on oocyte maturation and oocyte arrest. Material methods: Trophoblast stem cells isolated from 14,5 days mouse embriyo plasentas. Prophase I, metaphase I and metaphase II oocytes collected from mice and cultured in M2 culturing media, trofoblast stem cells culturing media (CM) and with trofoblast stem cells for 48 hours. Then the oocytes were seperated into two main groups for whole mount immunohistochemistry, that we anaylsed the immuno reactivity of anti-APC, anti- MAD1, anti-MAD2, anti-RAS, anti-MPF and anti-cyclin B-1, and also for DNA methylation analyses. Results: Trophoblast stem cells were cultured and characterised with cytokeratin 7. After culturing prophase I, metaphase I and metaphase II oocytes in M2, CM, TR medium APC immunoreavtivity was highest in TR prophase I. MAD1 immunoreactivity was only detected in TR metaphase II whereas the other groups was positive at the earlier stages. Although MAD2 immunoreactivity was higher in CM group RAS immunoreactivity was minimum in TR group and the other groups showed the immunoreactivity as they were expected. The immunoreactivity of cyclin B1 was weak in all groups however MPF immunoreactivity detected in all oocytes of CM and metaphase II of M2. While the low level of DNA methylation was detected at CM metaphase II oocytes, the highest ratio was observed in M2 metaphase II oocytes. Discussion: The protocol of trophoblast stem cells derivation was obtained for the first time in our department and characterised. Because of increase of RAS, MPF and MAD2 immunoreactivity and decrease of APC and Cyclin B1 after culturing of oocytes in CM, this culture conditionmight support the in vitro oocyte maturation.