Meiosis-activating sterol promotes resumption of meiosis in mouse oocytes cultured in vitro in contrast to related oxysterols.
The sterol 4,4-dimethyl-5alpha-cholesta-8,14,24-trien-3beta-ol (FF-MAS [follicular-fluid meiosis-activating sterol]) from human follicular fluid has recently been identified as a compound that induces the resumption of meiosis. FF-MAS and various oxysterols have been reported to transactivate the orphan receptor LXRalpha. The objective was to determine the biological activity of synthetic FF-MAS on the resumption of meiosis and final maturation of mouse oocytes in vitro. In order to evaluate whether LXRalpha might mediate FF-MAS action on the oocyte, we compared the capability of various compounds to activate LXRalpha-dependent transcription and to induce resumption of meiosis in the oocyte assay. Ovaries were isolated from immature mice primed with FSH 48 h before collection. Naked oocytes (NkO) and cumulus enclosed oocytes (CEO) were isolated from follicles. The oocytes were cultured in two groups, NkO and CEO, respectively, in media containing either 3 mM hypoxanthine, 5 microM IBMX, or 0.100 mM dbcAMP to maintain the oocytes in the germinal vesicle stage. The resumption of meiosis was assessed by the frequency of germinal vesicle breakdown (GVBD) after 24 h of in vitro culture. FF-MAS overcame the meiotic inhibition by hypoxanthine in both the NkO group and CEO group in a dose-dependent manner within the concentration range 0.07-7 microM. FF-MAS displayed similar potency in all inhibitory agents used. Also, FF-MAS significantly increased the formation of polar bodies in both the CEO and NkO group. The oxysterols 22(R)-hydroxycholesterol (a potent ligand for the LXRalpha receptor), 16-hydroxycholesterol, 25-hydroxycholesterol, and 27-hydroxycholesterol, as well as cholesterol, were tested without any significant effect on maturation compared to that of controls. Oxysterols and FF-MAS were observed to activate LXRalpha. In conclusion, the results reported here clearly demonstrate that synthetic FF-MAS exclusively is capable of mediating resumption of meiosis in vitro in both NkO and CEO irrespective of the inhibitory substance used. In contrast, the oxysterols and cholesterol had no significant biological activity on this oocyte function, and consequently we found no correlation between LXRalpha activation and meiosis stimulation.
Grøndahl
C
C
Health Care Discovery, Novo Nordisk A/S, Copenhagen, Denmark. chgr@novo.dk
Ottesen
J L
JL
Lessl
M
M
Faarup
P
P
Murray
A
A
Grønvald
F C
FC
Hegele-Hartung
C
C
Ahnfelt-Rønne
I
I
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000818:Animals; D003994:Bucladesine / Q000494:pharmacology; D002478:Cells, Cultured; D004268:DNA-Binding Proteins; D005260:Female; D019271:Hypoxanthine / Q000494:pharmacology; D008024:Ligands; D000071518:Liver X Receptors; D008540:Meiosis / Q000187:drug effects*; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D009865:Oocytes / Q000187:drug effects*; D057093:Orphan Nuclear Receptors; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D010957:Plasmids / Q000235:genetics / Q000502:physiology; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics; D013261:Sterols / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000502:physiology
D004268:DNA-Binding Proteins; D008024:Ligands; D000071518:Liver X Receptors; C000605213:NR1H3 protein, human; C000605214:Nr1h3 protein, mouse; D057093:Orphan Nuclear Receptors; D010726:Phosphodiesterase Inhibitors; D018160:Receptors, Cytoplasmic and Nuclear; D013261:Sterols; D019271:Hypoxanthine; D003994:Bucladesine; D015056:1-Methyl-3-isobutylxanthine