MicroRNA-429 functions as a regulator of epithelial-mesenchymal transition by targeting Pcdh8 during murine embryo implantation

Study question: What is the role of miR-429 in murine embryo implantation?

Summary answer: miR-429 functions as a suppressor of epithelial-mesenchymal transition (EMT) during the process of embryo implantation by reverse regulation of Pcdh8.

What is known already: MicroRNAs (miRNAs) may serve as promising regulators of embryo implantation. miR-429 was recently found to be down-regulated during embryo implantation period in a microarray analysis.

Study design, size, duration: The expression profile of miR-429 was clarified in a series of models, and the target gene was confirmed. The in vivo and in vitro effect of miR-429 on embryo implantation was examined.

Participants/materials, setting, methods: Pregnancy was produced by natural mating between female C57BL6/J mice and male mice, and a series of models, including pseudopregnancy, delayed implantation and artificial decidualization, were established. The expression profile of miR-429 during the embryo implantation period was clarified in these models. Candidate target genes of miR-429 were predicted by bioinformatic analysis and tested by luciferase activity assay. The in vivo effects of miR-429 on embryo implantation were also examined. The in vitro effects of miR-429 on EMT were studied by examining migratory and invasive capacities by transwell assay and expression profiles of cadherin family members by western blotting and qRT-PCR.

Main results and the role of chance: The expression profile of miR-429 in animal models suggested its down-regulation should be dependent on the presence and status of blastocysts and on endometrial decidualization. The luciferase activity assay showed that Pcdh8, a member of cadherin gene family, was the target gene of miR-429, and miR-429 suppressed the expression of Pcdh8 mRNA and protein. Gain-of-function of miR-429 in vivo resulted in a significant reduction of the number of implantation sites, but had little effect on fertilization. Up-regulation of miR-429 in vitro led to suppression of mesenchymal marker genes Vim, Cdh2, Zeb1 and Zeb2, and activation of epithelial marker gene Cdh1, resulting in suppression of the migratory and invasive capacities of cells. miR-429 also partially abrogated TGF-beta-induced EMT. The dysregulated expression profiles of EMT markers during embryo implantation period could be partially reversed by gain-of-function of miR-429 in vivo.

Limitations, reasons for caution: The association of miR-429 with other members of the miR-200 family in embryo implantation remains to be determined. The relationship between miR-429 and the cadherin family needs more intensive description and the detailed mechanism of miR-429 in regulating the cadherin family needs to be elucidated.

Wider implications of the findings: Our findings indicate that miR-429 plays a major role in embryo implantation as a suppressor of EMT by targeting Pcdh8. This information could contribute to a better understanding of the mechanisms involved in the miRNA-mediated regulation of embryo implantation, and subsequently improve treatments for infertility. The findings are consistent with that from previous research of the other members in miR-200 family in embryo implantation and in the EMT.

Study funding/competing interests: This study was supported by the Natural Science Foundation of China (Grant number: 81170592), and Special Fund from National Excellent Doctoral Dissertation (Grant number: 201079). There was no conflict of interest.