Epidermal growth factor combined with recombinant human chorionic gonadotrophin improves meiotic progression in mouse follicle-enclosed oocyte culture

Abstract

Using a mouse early preantral follicle culture system, mature full grown oocytes, arrested in prophase I of meiosis, were produced after 12 days using a recombinant gonadotrophin-supplemented medium. This culture medium does not mimic the normal extracellular environment of the oocyte and might therefore modify meiotic regulation and more particularly progression to metaphase II (MII). The aim of this study was to optimize the treatment using recombinant stimulatory ligands which were known to induce germinal vesicle breakdown (GVBD) and completion of meiosis I, metaphase II (MII), namely recombinant follicle stimulating hormone (r-FSH), chorionic gonadotrophin (r-HCG) and epidermal growth factor (EGF). Full-grown intrafollicular oocytes could not resume meiosis when the 'ovulatory' stimulus was r-FSH, used at a 100 times higher dose than during culture. r-FSH did not increase progesterone production. When 1.5 IU/ml r-HCG was used as meiotic trigger, germinal vesicle breakdown was obtained in 95% of the oocytes 64% of which extruded a first polar body. r-HCG induced a dramatic increase in progesterone production. When EGF was administered as sole stimulus on day 12 to the attached follicle-enclosed oocytes, only doses > or =5 ng/ml could cause GVBD, although less effectively than r-HCG (45 versus 95%; P < 0.0001). Oocytes undergoing GVBD by the EGF pulse reached metaphase II at a rate of 54% (not significant versus r-HCG). EGF did not stimulate progesterone production. Addition of increasing doses of EGF (0.5; 5; 10; 50 ng/ml) to r-HCG did not increase the GVBD-rate, but EGF doses >5 ng/ml improved MI to MII transition (P=0.027), thereby improving the final yield of MII oocytes by 12.5%. These data show that up to a dose of 50 ng/ml, EGF on its own could only override the somatic inhibitory stimuli in less than half of the cultured follicles. However, in addition to HCG, EGF (25 ng/ml) had a stimulatory effect on completing the first meiotic division. It was concluded that, under the present culture conditions, EGF in combination with HCG provided optimal nuclear maturation.

Epiregulin can effectively mature isolated cumulus–oocyte complexes, but fails as a substitute for the hCG/epidermal growth factor stimulus on cultured follicles

“…When used in combination with hCG, EREG did repetitively induce a lower PB rate than hCG/EGF (only 53 vs 89% respectively; P!0.05), and 37% of the oocytes remained arrested in GVBD stage. This would suggest that in combination with hCG, EREG is less effective than EGF (Smitz et al 1998). Moreover, it has also been shown that EGF influences the phosphorylation, ubiquitination and internalisation of connexin 43 (GJA1), the predominant functional unit of ovarian gap junctions connecting oocytes and cumulus cells (Lau et al 1992, Leithe & Rivedal 2004, Sela-Abramovich et al 2005.…”

“…Almahbobi et al (1995) reported that EGF or TGFA block antral follicle growth in vitro and reduce oestradiol (E 2 ) production, compared with the control. Smitz et al (1998) reported that when used as an ovulatory stimulus in cultured mouse follicles, EGF could induce meiotic reinitiation; however, the ensuing production of progesterone by luteinising follicles was minimal, suggesting that progesterone production and oocyte meiotic maturation could be independent events. Nevertheless, Jamnongjit et al (2005) showed that EGF induced progesterone production in COCs, and additionally, they showed that progesterone as well as testosterone was able to induce oocyte meiotic resumption.…”

A prospective, randomized and blinded comparison between 10,000 IU urinary and 250 μg recombinant human chorionic gonadotropin for oocyte maturation in in vitro fertilization cycles

“…The removal of urinary contaminants increases both the purity and the local tolerability of hCG. hCG can induce oocyte maturation and ovulation through its ability to trigger germinal vesicle breakdown and extrusion of the first polar body and induce an increase in progesterone production (2).…”

Detection of gene expression and enzyme activity of Cytochrom P450 arom (aromatase) in preantral and early antral bovine follicles depending on culture conditions in vitro

“…An effective possibility of these oocytes maturing up to the fertilization capability would develop a great potential for many bio-technical methods. For small mammals (mice), methods have been described where live offsprings have been produced (EPPIG and SCHROEDER, 1989;SPEARS et al, 1994) using oocytes from preantral follicles. For large mammals, such as cattle, these methods cannot be easily adopted (WU et al, 2001).…”

“…Its expression, therefore, is directly proportional to the ability of the follicle for estradiol synthesis. The measurement of aromatase activity in individual follicles is possible by means of a modified estradiol production assay (FEHRENBACH et al, 1998;PÖHLAND et al, 1998;YUAN and GIUDICE, 1999;SMITZ et al, 1998). It is possible to carry out quantitative examinations with this assay, but the exact site of synthesis cannot be pinpointed in the follicle (oocyte, different somatic cell layers).…”