Role of ulipristal acetate in regulating endometrial gene expression and spheroids attachment

Abstract

The novel emergency contraceptive Ulipristal acetate (UPA) belongs to the progesterone receptor modulator family. A single oral dosage of 30mg UPA within 120 hours of unprotected intercourse could delay ovulation and differentiation of endometrium. Yet, whether UPA affect embryo implantation remains largely unknown. This study aims to investigate whether UPA affect endometrial gene expressions and embryo attachment onto endometrial epithelial cells.

The PR-expressing human endometrial carcinoma cell line Ishikawa was used and treated with 10nM estrogen, 1μM progesterone or 4μM UPA for 24 hours. Changes in transcriptome profiles were analyzed by Affymetrix Human Gene 1.0 ST array GeneChip. Gene clustering showed the gene expression pattern after UPA treatment was similar to control (0.1% ethanol); while estrogen treated group was different from all the other groups. Totally, 8 genes were significantly increased and 1 was decreased (≥2-fold, p<0.05) after UPA treatment. All except one of the 8 up-regulated genes were also up-regulated by estrogen; while only one of them increased after progesterone treatment. Most genes that were altered by UPA were involved in angiogenesis and vascular remodeling.

The effect of UPA on human embryo-endometrium attachment was carried out using an in vitro multi-cellular spheroids-endometrial epithelial cell co-culture model. Human choriocarcinoma cell line JAR and Ishikawa were used. UPA (0.04-4μM) treatment for up to 48 hours did not affect the proliferation of JAR or Ishikawa cells. Similarly, the attachment of JAR spheroids onto Ishikawa cells after 1 hour co-culture was not affected by UPA treatment. The molecules of Wnt/β-catenin signaling pathway, a pathway that is actively involved in embryo implantation, such as the β-catenin and GSK-3β, and endometrial receptive marker E-cadherin were not changed after UPA treatment. In Ishikawa cells, the expression of PR-A was induced after UPA (0.04-4μM) treatment; while PR-B increased when 0.04 or 4μM UPA used. However, the PR-A/PR-B ratio remained unchanged after all concentration of UPA treatment.

The effect of UPA on spheroids attachment was further investigated with cultured human primary endometrial epithelial cells. Endometrial glandular epithelial cells were digested and isolated from endometrial biopsy taken from IVF patients on day 7 after luteinizing hormone surge (LH+7). A co-culture assay was optimized with JAR spheroids and endometrial epithelial cells that were growing on Matrigel. The attachment rate of JAR spheroids is approximately 60% after 3 hours incubation. However, after 24 hours of exposure to 4μM UPA, the attachment remained comparable to that of the control group. In conclusion, UPA could alter the expression of genes in Ishikawa cells mainly related to angiogenesis. It is likely that UPA may affect stromal decidualization and blastocyst invasion after attachment. However, UPA did not affect the expression of Wnt-signaling molecules and attachment of JAR spheroids onto either Ishikawa or human primary endometrial epithelial cell.