A study of annexin A2 and implantation

Abstract

Implantation is a critical step in reproduction. It is complicated and well-coordinated consisting of apposition, attachment and invasion of embryo into the endometrium. The mechanism of implantation is unclear. Our previous proteomic study showed an increase of annexin A2 in the endometrium during the implantation window of mice, consistent with the increased annexin A2 expression in the receptive human endometrium. The hypothesis of this project was that annexin A2 mediatedthe embryo-endometrium attachment.

The first objective was to study the spatio-temporal expression of endometrial annexin A2 immunoreactivities in humans and mice. The cyclical change in annexin A2 expression in the mouse and human reproductive cycle suggested the involvement of a steroid regulatory mechanism. Interestingly, annexinA2 was transiently expressed on the membrane between the mouse uterine luminal epithelium and the implanting embryos from Day 4 (pre-implantation) to Day 5 (post-implantation) of pregnancy. No such signal change was observed at the inter-implantation sites, showing that the implanting embryos partially regulated annexin A2 expression. These observations and the high expression of the molecule in the luminal epithelium of human endometrium in the mid-and late luteal phase were consistent with a role of annexin A2 in implantation.

The second objective was to verify the action of steroids on annexin A2 expression. It was found that a combination of 6675 pmol/L of estrogen and 429.8nmol/L of progesterone increased the total and apical surface expression of annexin A2. In mice, estrogen but not progesterone, increased annexin A2 expression in the uterine luminal epithelium of ovariectomized mice.

The third objective was to study the function of annexin A2 in embryo-endometrium attachment using an Ishikawa (endometrial epithelial cells)-JEG-3 trophoblast spheroids (embryo surrogate) coculture model. Knockdown of the expression of annexin A2 in either or both cell lines significantly decreased the attachment rate of the spheroids onto the endometrial cells. The suppressive action on the two cell lines was additive. The attachment was also suppressed in the presence of anti-annexin A2 antibody during coculture. Annexin A2 was also involved in mouse implantation as demonstrated by a significant decrease in implantation sites after injection of anti-annexin A2 antibody into the mouse uterine horn.

The final objective was to study the action of annexin A2 as an adhesive molecule in embryo attachment. It was found that loss of P11, the binding partner of annexin A2, reduced the attachment rate of the JEG-3 spheroids probably by decreasing the translocation of annexin A2 to the surface of the endometrial cells. Recombinant P11 and annexin A2 protein failed to bind significantly to the Ishikawa cells and the JEG-3 cells.

In summary, this study demonstrates the involvement of annexin A2 as an adherent molecule in the embryo-endometrium interaction.