Delineating cell types and toxicity in human ovaries from birth to sexual maturity

Abstract

The ovary contains the female germ cells known as oocytes, which are essential for female fertility. The immature oocytes reside within primordial follicles in the ovarian cortex and are referred to as the ovarian reserve. Disruption of ovaries by various external factors, such as environmental pollutants, medical conditions, and gonadotoxic treatments can impair fertility. While the pathways behind disruption have been studied in adult ovarian tissue, much remains to be elucidated, especially regarding paediatric ovarian tissue. The molecular and transcriptomic landscape of ovarian tissue from children remains poorly characterised, making it even more challenging to estimate its sensitivity to ovarian disruption. Ovarian tissue cryopreservation is an established method for preserving fertility in adults requiring gonadotoxic treatments, resulting in nearly 300 live births. Paediatric ovarian tissue is cryopreserved using the protocols as in adults, despite previous studies showing differences in the competency of residing follicles. Additionally, patients requiring fertility preservation often have received gonadotoxic treatments or have medical conditions associated with infertility prior to fertility preservation, urging further study of their effect on the ovarian reserve. This thesis aims to provide a better understanding of i) the basic biology of the ovary from birth to reproductive age, and ii) the effect of chemical exposures and medical intervention on the ovary through histologic and transcriptomic assessment of the ovarian cortex in patients from child- to adulthood.

Paper I and paper II histologically quantified cortical follicles in biopsied ovarian tissue to elucidate the associations between different exposures and the ovarian reserve. In paper I the association between serum levels of 31 environmental pollutants and both the ovarian reserve and odds of infertility were investigated in a cohort of pregnant women. Some pollutants correlated negatively with odds of infertility and healthy follicle population, but not with atretic follicles. In paper II, a reference standard for ovarian reserve size was established for women up to 25 years old, allowing assessment of the normality of a patient’s ovarian reserve size using Z-scores. This standard was tested in patient cohorts with different medical conditions and exposures to gonadotoxic treatments, identifying patients at risk of reduced ovarian reserve. Particularly, the youngest patients were found to be at risk of a decreased ovarian reserve due to their treatments. These papers illustrated the reduction of the ovarian reserve either through gonadotoxic chemicals or underlying genetic conditions. Paper II prompts further studies of ovarian reserve quality and size especially in the youngest patients in need of fertility preservation.

In Paper III and Paper IV transcriptomic analysis of cortical follicles and cells from child and adult ovaries were carried to identify differences possible agedependent changes. In paper III, sequencing of whole isolated follicles from both child and adult ovarian cortex revealed two transcriptomically distinct, yet morphologically similar follicle types. Type 1 follicles exhibited expected gene expression profiles for oocytes and granulosa cell, while Type 2 showed lower oocyte gene expression and higher predicted signalling activity. Although, gene expression patterns underlying follicle development were similar in children and adults, differences were discovered in extracellular matrix and theca cell biology. Additionally, previous chemotherapy exposure associated with increased interferon signalling in child follicles. In Paper IV, single cell sequencing of cells from child and adult ovarian cortex identified 10 distinct cell types of which four had not been previously described in human ovarian cortex: theca cells, Schwann Cells, lymphatic endothelial cells and epithelial cells. Furthermore, this study revealed changes in stromal populations over age, possibly suggesting differences in its function during different stages towards sexual maturity. These findings confirm the dissimilarities between child and adult ovarian cortex and underscore the need for further research into the effect of age and different exposures on ovarian tissue and the reserve.