Endocrine Abstracts

Successful embryo implantation requires changes in endometrial gene expression of biomarkers which contribute to uterine receptivity to facilitate blastocyst attachment. Osteopontin (OPN), expressed at the maternal–foetal interface may facilitate implantation in processes, such as cell adhesion, migration and angiogenesis which require OPN signalling via CD44 and integrin receptors. As coordinate expression of OPN and receptors is shown to be cycle-dependent and up-regulated in human, mice and ovine endometrium at implantation, it is logical to conclude regulation of OPN may require ovarian steroids and embryonic signals. Aims of this study were to analyse steroid regulation and cyclical expression of OPN and receptors in ovine uterus using both in vitro and in vivo endometrial models.

Uteri were obtained from sheep sacrificed at follicular, luteal cycle phases, early gestation (d.17 and 35) and ovariectomised animals intramuscularly treated with E₂ (3 μg/ml)±P₄ (12.5 mg/ml). OPN mRNA expression was determined by qRT-PCR and protein levels of OPN and receptors were detected by SDS–PAGE-western immunoblotting and immunohistochemistry. Populations of luminal epithelia and stromal cells were isolated from luteal phase endometrium, and grown in the presence or absence of E₂±P₄, to analyse steroid hormone effects on OPN expression in isolated cell types.

Analysis of OPN and receptors in cyclical and early pregnant ewes confirmed up-regulation of OPN in luteal phase endometrium and early implantation stages. Western-blot analysis revealed the presence of 70-, 32- and 15-kDa OPN isoforms in endometrium which may result from progesterone regulation of posttranslational modifications of the native 70-kDa form. In addition, progesterone (10 ng/ml) significantly increased OPN expression in stromal cells, the effect of which was blocked by progesterone receptor antagonist, but P₄-regulated OPN expression was not shown in epithelial cells. These results demonstrate the coordinate expression of OPN and receptors regulated by progesterone coincide with the timing of implantation.