ИСТИНА

Elucidation of mechanisms of mammalian oocyte meiotic maturation is highly important not only for basic research, but also for the development of application-oriented methods, such as ART (assisted reproductive technologies). Nowadays many women who suffer from oncological or autoimmune diseases require fertility- preservation procedures. The most actively developing approaches of female fertility preservation are oocyte cryopreservation, ovarian tissue cryopreservation and ovarian tissue or ovarian follicle in vitro culture [1,2]. Moreover, progress in this field of science is necessary for the development of effective oocyte IVM (in vitro maturation) protocols and for the design of novel contraceptive drugs which will act only on maturing oocytes allowing to reduce negative impact on women’s health. Meiotic arrest occurs at prophase I and can continue for a long time. Prophase I maintenance is caused by action of multiple proteins and signaling molecules. The key molecules which plays an important role in meiotic resumption are the proteins involved in the MPF-complex regulatory pathway and Mos-MAPK molecular cascade. Normal gene expression during oocyte maturation and early embryonic development requires well-timed activation of specific maternal mRNA translation. These mRNAs have been accumulated in the oocyte during first meiotic arrest at prophase I [3-5]. The key proteins which are involved into translational regulation at the early stages of development in mice are ePAB, Dazl, Elavl2. During prophase I meiotic arrest mammalian oocytes grow and accumulate various molecules which will be necessary for early embryo development. In the case of mice size fully-grown oocyte is about 80 µm in diameter [6]. Oocytes with lower sizes resume meiosis in less number of cases [6]. In the current work we have evaluated the expression levels of the key genes involved in meiotic resumption in murine oocytes. Two groups of oocytes were used for the analysis: immature oocytes (with an average diameter about 70 µm) and the fully-grown ones (with an average diameter about 80 µm). Oocytes were extracted from ovarian follicles mechanically at the GV (germinal vesicle) stage. Then, cumulus cells also were removed mechanically by gentle pipetting. The samples for molecular genetic analysis were compound of 10 oocytes in each sample. After that oocytes were subjected for quantitative real-time PCR analysis using the SuperScript® III CellDirect cDNA Synthesis (Invitrogen) kit according to the manufacturer recommendations. It has been shown that the expression levels of important for meiotic resumption genes such as Mos, Cyclin B, CDK1, ePAB, Cyclin H, Wee2 in fully-grown oocytes is statistically (p<0,05) higher in comparison to non-fully-grown oocytes. Elucidation of key mechanisms which affect acquisition of meiotic competence by oocyte plays a pivotal role for the ART. Hence, it can be expected the appearance of a long list of the novel research works in this area.