Integrin β3 in rat blastocysts and epithelial cells is essential for implantation in vitro: studies with Ishikawa cells and small interfering RNA transfection.

BACKGROUND

Integrins are involved in the process of embryo-endometrium interaction during implantation. We investigated the localization of integrin β3 in the rat blastocyst and Ishikawa cells using an in vitro co-culture model of implantation.

METHODS

Zona pellucida-free rat blastocysts were co-cultured with the Ishikawa cells (endometrial adenocarcinoma cell line) to observe the attachment between the embryo and endometrium. Immunofluorescence staining was used to investigate the localization of integrin β3 in rat embryos at different stages of development (each n= 3 embryos) and at the embryo/endometrium interface, observed by confocal microscopy. The Ishikawa cells were transfected with integrin β3 small interfering RNA (siRNA) for 48 h and then co-cultured with Day 5 rat blastocysts to observe the effect on attachment.

RESULTS

Integrin β3 staining in the rat embryos increased at the blastocyst stage being highly concentrated in the cytoplasm of trophoblast cells (n= 9 embryos). Integrin β3 was localized on the apical surface of the Ishikawa cells (n= 3 experiments). However, integrin β3 relocated to the apical membrane of trophoblast cells in response to attachment to Ishikawa cells (n= 6 embryos). Moreover, when Ishikawa cells were transfected with integrin β3 siRNA, blastocyst attachment was significantly reduced compared with those transfected with control siRNA (16.7 versus 92.3%, respectively, P< 0.05).

CONCLUSIONS

Integrin β3, localized apically in the blastocyst and the Ishikawa cells, is important during initial attachment of the blastocyst to endometrial cells. This study provides further evidence of the importance of integrins during implantation and may aid in elucidating the molecular mechanism of implantation failure and infertility in women.