Background: Uterine fibroids are neoplasms of the myometrium and are the most common benign tumour afflicting women of reproductive age. We have reported previously that both NR2F2 and CTNNB1 expression are altered in fibroids compared to myometrium. NR2F2 and CTNNB1 have established roles in tumorigenesis and pathophysiologies of the reproductive tract. Furthermore, over-expression of CTNNB1 leads to uterine mesenchymal tumors in mice; and CTNNB1, retinoic acid (RA) and Sonic Hedgehog (Shh) can regulate NR2F2 expression.
Objective: To investigate NR2F2 and CTNNB1 expression in vivo and regulation in vitro in human myometrium (M) and fibroids (F).
Methods: Samples were collected from pre-menopausal, cycling women, free from hormone therapy prior to surgery. NR2F2 and CTNNB1 mRNA expression were analysed by qRT-PCR and protein expression and localisation by Western blot and immunohistochemistry (IHC). For regulation studies, M and F primary cultures were stimulated by Shh, P (100nM) and/or RA (1µM) for 24 h followed by gene expression analysed.
Results: NR2F2 mRNA was reduced in F compared to M, while protein levels were unchanged. Both CTNNB1 mRNA and protein were higher in F compared to M. Semi-quantitative IHC demonstrated significantly stronger expression in M vasculature compared to the rest of the tissue for both NR2F2 and CTNNB1, while in F only NR2F2 was more strongly expressed in blood vessels. In vitro studies demonstrated that while the Shh pathway was activated (as shown by increased Gli1 and PTCH1 mRNA), it did not affect NR2F2 or CTNNB1 expression in M or F cells. While RA combined with P significantly decreased NR2F2 but not CTNNB1 expression in M but not F cells.
Conclusion: In conclusion, we demonstrate aberrant expression and regulation of NR2F2 and CTNNB1 in uterine fibroids compared to normal myometrium, consistent with the hypothesis that these factors may play a causal role uterine fibroid development.