Endocrine Abstracts

Introduction: The production of an oocyte competent of fertilisation and normal embryonic development is controlled by a series of highly organised signals and intracellular events which are known to be affected by physiological glucocorticoids. The actions of these adrenal steroids are modulated by 11β-hydroxysteroid dehydrogenase (11β-HSD) enzymes and we have recently reported that 11β-HSD activity in porcine oocytes increases 20-fold during meiotic maturation. While expression of 11β-HSD1 protein and mRNA have been reported in human and rat oocytes, available evidence suggest that 11β-HSD2 is not present in oocytes. The aim of this study was to characterise the expression of the two cloned 11β-HSD isoenzymes in mouse oocytes during maturation.

Methods: Groups of germinal vesicle (GV) and metaphase II (MII) stage oocytes were snap frozen in liquid nitrogen. Total RNA was extracted using an RNeasy Kit (Qiagen), and cDNA synthesised with a Verso cDNA kit (ABgene) using oligo dT primers. QPCR was performed in an ABI 7000 cycler, using Quantifast (Qiagen) with intron spanning primers designed for the mouse HSD11B1 and HSD11B2 genes. GV and MII stage oocytes were fixed, permeabilised and stained using primary antibodies raised against 11β-HSD1 or 11β-HSD2. Protein expression was visualised using fluorescent secondary antibodies and a Olympus AX81 microscope.

Results and discussion: While HSD11B1 mRNA was expressed at similar levels throughout maturation, levels of HSD11B2 mRNA were increased markedly in MII versus GV stage oocytes. When examined for protein expression, immunofluorescence was observed with both 11β-HSD1 and 11β-HSD2 antibodies in GV and MII eggs. These findings demonstrate the presence of 11β-HSD1 mRNA and protein in GV and MII stage oocytes. Furthermore, the expression of 11β-HSD2, which has not been previously reported in mammalian oocytes, was increased in the MII oocytes consistent with our data showing an increase in 11β-HSD activity during meiotic maturation.