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Title: Characterization of human endometrial glandular epithelium in vitro and in vivo
Author: Barros, Flavio
ISNI:       0000 0004 6348 9384
Awarding Body: University of Warwick
Current Institution: University of Warwick
Date of Award: 2017
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Endometrial glands provide histiotrophic support for the developing conceptus prior to the onset of the haemochorial placenta. Several lines of evidence suggest that glands also play an important role in endometrial receptivity, decidualization of the stromal compartment, and in maternal immune tolerance during pregnancy. However, glandular epithelial cells isolated during the luteal phase become acutely arrested in vitro, precluding in-depth analysis of this cellular compartment of the human endometrium. In this thesis, I tested various approaches to overcome the senescence-associated cell cycle block of primary human endometrial epithelial cells (HEECs). I demonstrate that conditional reprogramming of HEECs, using conditioned medium of irradiated fibroblast and a Rho kinase inhibitor (Y-27632), partially reverses the senescent phenotype and enables expansion of primary HEEC cultures. However, the responsiveness of reprogrammed HEECs to embryo-derived signals and hormonal cues remained highly variable. To overcome this hurdle, I used a novel 3D culture system that enabled formation of glandular structures from clonal HEECs seeded in Matrigel and cultured in modified adult stem cell medium. Treatment of glandular organoids with cyclic AMP and steroid hormones induced the expression of PAEP, a glandular differentiation marker. Organoid-forming efficiency experiments revealed that missed miscarriage, characterized by early-onset foetal growth retardation, is associated with glandular progenitor cell deficiency. To validate this observation, I used laser capture microdissection coupled to RNA-sequencing to compare mid-luteal glandular gene expression between missed miscarriage cases and control subjects. Gene ontology analysis of differentially expressed genes revealed that miscarriage may be caused by bioenergetics defects in the glands, exemplified by altered expression of mitochondrial-related genes. Taken together, the ability to grow and differentiate endometrial glands from isolated clonal HEECs provides a powerful new tool to study the mechanisms underpinning reproductive failure.
Supervisor: Not available Sponsor: Conselho Nacional de Pesquisas (Brazil)
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: QP Physiology