Novel insights into endometrial architecture and function in reproductive health and disease

Abstract

Successful embryo implantation is a trade-off between a receptive endometrium and a good quality embryo. Endometrial glands are important for successful pregnancy, creating a favourable uterine environment for the implanting embryo. The 3D structure and gene expression profile of endometrial glands during endometrial receptivity are yet to be determined. The European Society of Human Reproduction and Embryology guidelines suggest recurrent pregnancy loss occurs in approximately 2% of women. While the pathogenesis of recurrent pregnancy loss remains unclear, it has been associated with reduced endometrial gland secretions and altered endometrial receptivity.
In this thesis, multi-modal 3D imaging and RNA sequencing are applied to the endometrium from control women and women with recurrent pregnancy loss, to identify 3D structural alterations and a gene expression profile of endometrial glands during the implantation window. This thesis takes a multi-scale approach to investigating endometrial glands. Beginning with the relationship between endometrial glands and endometrial stromal cells in whole endometrium, investigating endometrial glands only, and finally glandular ciliated cell components.
This thesis used 3D imaging to further our understanding of the spatial relationships between stromal cells and endometrial glands and their relationship to recurrent pregnancy loss. Quantitative 3D distance maps demonstrated for the first time that endometrial cells from women who experience recurrent pregnancy loss have altered 3D spatial relationships compared to controls. This 3D spatial information may help
understand the underlying biology and the changes that are observed in disease states. Endometrial function may underlie a range of fertility disorders and these 3D imaging approaches could ultimately lead to more informative diagnostic testing and new treatments.
3D imaging provided insight into glandular structure and was combined with high-speed video to quantify endometrial gland cilia movements. This study made a number of observations that would not have been obvious from 2D imaging. One of these was that microvesicles appear to be forming on the tips of glandular microvilli. Secretion of glandular microvesicles into the uterine lumen may help mediate a favourable uterine environment during embryo implantation. The microvillus surface of the glandular lumen was interspersed with ciliated cells. The role of glandular cilia is not clear but it is hypothesised that they may facilitate the movement of microvesicles to the uterine lumen. The cilia beat pattern observed in endometrial glands from controls and women with recurrent pregnancy loss however, was uncoordinated. High-speed video could be developed in the future and provide a fast diagnostic of glandular cilia function.
This thesis used 3D imaging and RNA sequencing to further our understanding of the gene expression profile of endometrial glands and glandular beta-tubulin from women with recurrent pregnancy loss. Differentially expressed genes were identified in endometrial glands from women with recurrent pregnancy loss compared to controls and beta-tubulin enrichment in glandular ciliated cells was established. Combined 3D imaging and gene expression analysis also demonstrated that beta-tubulin isoforms were altered on a gene transcript level and a protein level in endometrial glands from women with recurrent pregnancy loss. Combining 3D imaging and gene expression data allows gene expression changes to be located in endometrial tissue. Teasing out beta-tubulin changes in the glandular ciliated epithelium could lead to specific endometrial gland biomarkers of a recurrent pregnancy loss phenotype.
This multi-scale and multi-modal approach has established a four dimensional view of endometrial glands during the window of implantation. Alterations in the endometrial gland architecture and their spatial relationship to the endometrial stroma could be associated with recurrent pregnancy loss. This new approach can be applied to other patient groups who suffer from unsuccessful pregnancy, such as infertility and recurrent implantation failure, and in future, contribute to the development of diagnostics.

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Identifiers

ORCID for Ying Cheong: orcid.org/0000-0001-7687-4597

ORCID for Rohan Lewis: orcid.org/0000-0003-4044-9104

ORCID for Jane Cleal: orcid.org/0000-0001-7978-4327

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