epidermal-growth-factor and 4-4-dimethylcholesta-8-14-24-trienol

Follicular fluid meiosis-activating sterol (FF-MAS) is regarded as an important compound relevant to meiotic resumption in mammalian oocytes. The objective of this study was to investigate the influence of FF-MAS on germinal vesicle breakdown (GVBD) and first polar body (PBI) extrusion with regard to culture conditions, state of the oocyte and mouse strain. Denuded oocytes (DO) and cumulus-enclosed oocytes (CEO) were retrieved from PMSG-primed Quackenbush or C57BL/6J x DBA/2 (C57) mice and cultured for 20 h in alpha-MEM medium under the following conditions: (i) 250 micromol/l dibutyryl cAMP (dbcAMP) +/- EGF, 1 ng/ml or FF-MAS, 20 micromol/l; (ii) 4 mmol/l hypoxanthine (HX) +/- EGF or FF-MAS; (iii) HX + EGF + FF-MAS; and (iv) HX + FF-MAS 5 h priming and subsequent culture with HX + EGF. Oocyte GVBD and PBI emission were recorded and stained with Hoechst 33342. Very limited meiotic inhibition was observed in Quackenbush mice in comparison with C57 mice. FF-MAS promoted maturation in C57 DO and CEO and Quackenbush DO. In Quackenbush DO and CEO and C57 DO a significant increase in atypical PBI extrusion occurred, but not in C57 CEO as well as in EGF-treated Quackenbush CEO primed or co-cultured with FF-MAS. These results support a meiosis resumption function for FF-MAS and suggest that in its presence, the quality of the MII oocytes retrieved appears to be influenced by the strain of the mice, the state of the oocyte and the presence or absence of growth factors in the culture medium.

Topics: Animals; Cells, Cultured; Cholestenes; Coculture Techniques; Cyclic CMP; Epidermal Growth Factor; Female; Hypoxanthine; Meiosis; Mice; Mice, Inbred C57BL; Mice, Inbred DBA; Oocytes; Species Specificity

In this study, we compared the relative ability of FSH (100 mIU/ml), epidermal growth factor (EGF) (10 ng/ml), and follicular-fluid meiosis-activating sterol (FF-MAS, 10 micromol/l) to induce meiotic resumption and polar body I (PBI) extrusion in mouse oocytes.. Cumulus-enclosed oocytes (CEO) were co-incubated with meiosis-arresting agents, including 4 mmol/l hypoxanthine (Hx), 0.3 mmol/l dibutyryl cAMP (dbcAMP), and 8.5 micromol/l cilostamide, a selective inhibitor of the oocyte-specific phosphodiesterase 3 (PDE 3).. In Hx-treated oocytes, FSH, EGF and FF-MAS induced meiosis resumption at very high rates, but only FSH and EGF also promoted PBI extrusion with high frequency. In experiments conducted in the presence of dbcAMP, FF-MAS was unable to promote an increase in germinal vesicle breakdown (GVBD) rate, whereas FSH and EGF generated a response similar to the Hx groups. Neither FSH, EGF nor FF-MAS caused any change in the meiotic status of CEO when meiotic arrest at the germinal vesicle (GV) stage was maintained by cilostamide. In the presence of Hx, naked oocytes (NkO) co-cultured with their cumulus cells were able to respond to the GVBD-inducing effect of FSH and EGF by resuming meiosis at high rate.. Collectively, these results indicate that: (i) a signal triggered in cumulus cells by either FSH or EGF, but not necessarily coincident with FF-MAS, may contribute to meiotic maturation, supporting GVBD and extrusion of PBI; (ii) the transmission of this signal can occur in a paracrine fashion, at least with reference to the breakdown of the GV. It also appears that concomitant regulation of intra-oocyte cAMP degradation is a prerequisite for meiosis resumption.

Topics: Animals; Bucladesine; Cell Differentiation; Cells, Cultured; Cholestenes; Coculture Techniques; Epidermal Growth Factor; Female; Follicle Stimulating Hormone; Hypoxanthine; Meiosis; Mice; Oocytes; Ovarian Follicle; Quinolones