Increased levels of testosterone associated with polycystic ovary syndrome (PCOS) negatively affects HOXA10 and integrin beta-3 expression in endometrial cells and embryo attachment using a trophoblast cell spheroid model

PCOS is a reproductive endocrine disorder that affects up to 10% of women of a reproductive age, with 70-80% of patients defined as infertile. There are many different symptoms that could impact fertility in women with PCOS such as anovulation, hyperandrogenism and insulin resistance. A trophoblast (BeWo cell) spheroid attachment assay was used as an in vitro model of embryo implantation to examine the effect of testosterone on the receptivity of endometrial (Ishikawa cell) monolayers. Attachment of BeWo spheroids to Ishikawa cell monolayers was significantly reduced following pre-treatment of Ishikawa cells with 10nM testosterone for 24h (33% compared to 80% with untreated Ishikawa cells; P < 0.0001, n = 3). Flutamide (an androgen receptor antagonist, 1μM) added simultaneously with testosterone reversed the effect of treatment with testosterone alone, returning attachment rates to control levels (75%; p>0.05 compared to untreated cells, n = 3). The transcription factor HOXA10 and the cell surface adhesion receptor αVβ3 have previously been shown to play key roles in implantation. Testosterone treatment significantly decreased the expression of HOXA10 (P < 0.001, n = 4) and integrin β3 (P < 0.002, n = 4) in Ishikawa cells detected by qPCR. Immunocytochemistry showed a significant reduction in αVβ3 protein expression in testosterone-treated ishikawa cells (P < 0.05, n = 2, >300 cells analysed per condition). Co-treatment of flutamide and testosterone rescued the levels of HOXA10 and β3 gene expression (p>0.05 compared to untreated cells, n = 3) and αVβ3 protein expression (p>0.05 compared to untreated cells; n = 2, >150 cells analysed). These data suggest that the hyperandrogenism observed in women with PCOS results in reduced endometrial receptivity through the decreased expression of HOXA10 gene and αVβ3 protein and this may in part be responsible for infertility in these individuals.