Abstract
The choice of medium and supplements can affect meiotic regulation and may have an impact on the
regulation of mammalian oocyte growth and embryonic cell function. The aim of the present study
was to assess the effects of oxygen concentration and endogenous lanosterol on the in vitro maturation
(IVM) media without serum and based on recombinant human chorionic gonadotrophin in prepubertal
ewe oocytes. Firstly, the effect of varying oxygen concentrations (5% and 20%) during IVM in TCM-199
supplemented (4 mg/ml bovine serum albumin (BSA), 100 M cysteamine, 0.3 mM sodium pyruvate,
0.1 UI/ml recombinant follicle-stimulating hormone (r-FSH; Gonal-F R 75 UI, Serono, Italy), 0.1 UI/ml
recombinant leuteinizing hormone (r-LH; Lhadi R 75 UI, Serono, Italy) and 1 g/ml estradiol-17) on
subsequent nuclear maturation of oocytes examined under ultraviolet light following staining with
bisbenzimide (Hoechst 33342) was investigated. Secondly, two concentrations of lanosterol (0, 10 and
50 M) were added to the IVM medium. Nuclear maturation of oocytes was examined as previously.
Lipid content in oocytes, an important indicator of cytoplasmic maturity, was also measured using Nile red fluorescent stain. The results showed that low oxygen concentration affected the nuclear maturation. Similarly, a significantly higher rate of meiosis resumption was observed with 10 M (72.3%) of lanosterol compared with the control (51.8%) or 50 M of lanosterol (59.4%). A significantly higher content of lipids was also observed with 10 and 50 M of lanosterol (7.3 ± 0.2 × 106 and 7.4 ±0.2 × 106 arbitrary units of fluorescence) compared with the control (6.7 ± 0.2 × 106 arbitrary units of fluorescence). The results indicate that 10 M lanosterol during IVM in medium without serum and based on recombinant human chorionic gonadotrophin has a positive effect on maturation of prepubertal ewe oocytes.
