Thyroid hormones are critical to growth and development of the human fetus. In particular, the fetal central nervous system is extremely sensitive to the actions of the active ligand, tri-iodothyronine (T3). The placenta is the organ during pregnancy that allows transport between the mother and her baby by close interaction of the maternal and fetal circulations. Endogenous fetal thyroid hormone production does not occur until the beginning of the second trimester. However, there appears to be transplacental transport of thyroid hormones to the fetus earlier in gestation and organs, such as the central nervous system, appear to be exquisitely sensitive to their actions. The content of this Thesis describes my work, published in peer reviewed papers over the last fifteen years. It outlines the molecular mechanisms controlling the delivery and actions of thyroid hormones to the fetus.

Overall, signaling events in human sperm, together with the identity of many of the molecules stimulating them, remain very poorly characterised. A two-pronged approach involving characterisation of sperm receptors in a model-cell system, followed by single cell [Ca2+]i imaging of large numbers of capacitated human spermatozoa, was employed to start to unravel some of these possibilities. Data were compared to responsiveness of the best-characterised physiological ligand of sperm – the steroid hormone progesterone. Proteomic data strongly indicates the presence of OR2AE1, an olfactory receptor, in mature human sperm. OR2AE1 was cloned into the mammalian expression vector pcDNA3 and expressed in HEK293 cells. Functional expression of OR2AE1 (5.6±1.1% of cells showing correct localisation of FLAG-tagged recombinant receptor at the plasma membrane level) was demonstrated. Despite this relatively low level of functional expression, this system still allowed identification of a potent volatile agonist for OR2AE1 – omega () – via an elimination-panel system used alongside [Ca2+]i imaging. Around 3% of cells in the heterologous system generated a response to the agonist, indicating correct functional expression. When sperm were exposed to 50 μM , [Ca2+]i imaging revealed a transient response which occurred in 30-40% of capacitated cells. 10 μM NNC 55-0396, an antagonist of the CatSper channels, inhibited around 80% of these responses. Similar inhibition levels were demonstrated by inhibitors of the cAMP pathway (100 μM SQ22536 and 25 μM cAMPS-Rp), but no substantial effect was observed by inhibition of PLC by 5 μM U73122. Chemotactic assays indicated that 50 μM  had an optimal chemoattractive effect, with 500 μM  significantly stimulating acrosome reaction. An unrelated family of chemicals, the boronic acids, were also examined for their ability to affect sperm motility, migration and [Ca2+]i dynamics. A significant effect on the number of sperm which migrated from semen when exposed to these chemicals was observed, with pathway inhibition distinctly indicating a bifurcating signal propagating mainly via IP3 but also involving the cAMP pathway. Manipulation of the signalling events described, with inherent modulation of sperm motility may underlie the ability to develop future novel therapy, including contraceptive and diagnostic treatment regimes for the human male.

To achieve fertilisation, human sperm have to navigate and interact with the female reproductive tract (FRT) on molecular and mechanical levels. The current knowledge of some aspects of both types of interactions are limited and they were examined in this research. Proteomic analysis of crude and depleted human follicular fluid (hFF) by three proteomic approaches identified 479 hFF-proteins of which 22% were novel. A table of hFF-proteins, compiled from twenty-four hFF proteomic studies, resulted in 1586 hFF proteins; a resource for folliculogenesis and discovery of hFF biomarkers. A comparative proteomic study of media-capacitated human sperm versus capacitated sperm in the presence of hFF revealed certain hFF proteins were acquired by sperm during capacitation. Comparative metabolomics revealed some elevated metabolites in the sperm capacitation-media following 6-hour incubation compared to 1-hour swim-up, which may have relevance to sperm energy metabolism and potentially to sperm signalling mechanisms in the FRT causing remodelling for fertilisation and preparation for implantation. Sperm micro-particle image-velocimetry revealed an average fluid velocity around the motile sperm of ≤ 25- 45 μm/s and ≤ 20-35 μm/s in low and high viscosity media respectively. The averaged fluid vorticity manifested a trail of spatially confined mixing of the fluid surroundings motile sperm. To the best of our knowledge, this work is the first to conduct a comparative proteomic analysis of human sperm versus sperm-hFF interaction and to perform metabolomics analysis of human sperm capacitation media at two time points. The novel perspective of mechanical aspects of sperm motility by studying fluid velocity and vorticity around motile sperm adds a new approach to the study of sperm motility and chemotaxis. In conclusion, sperm-FRT interactions involve complex molecular and physical interactions and regulatory events. Further research of these interactions may enhance our understanding of potential applications for improved assisted reproductive techniques’ outcome and possible diagnostic approaches of infertility.

Uncommon disorders are difficult to study because routine information sources are unreliable, and comprehensive studies would require a large collaboration to identify relatively few cases. This PhD developed a national network and research platform for the study of serious and uncommon disorders in early pregnancy: The UK Early Pregnancy Surveillance Service (UKEPSS). A national collaboration can enable high quality research to improve our knowledge and help us provide better care for women with rare early pregnancy conditions. Using the UKEPSS platform, with an early pregnancy network of 86 UK hospitals and Early Pregnancy Units (EPUs), nation-wide prospective cohort and case control studies of caesarean scar pregnancy were performed. The second section of this thesis comprises of systematic reviews in priority questions in miscarriage management, exploring the effectiveness of progesterone to reduce miscarriage in women presenting with early pregnancy bleeding; the effect of ethnicity on miscarriage; and the effect of hydrosalpinx and its management on miscarriage risk.

This thesis included a systematic literature review, yielding studies on three employment categories: hairdressers, cosmetologists and laboratory workers. The original research examined pregnancy outcomes for Finnish singleton births between 1990 and 2010. The parameters were increased male gender, low birth-weight, high birth-weight, pre-term delivery, post-term delivery, small for gestational age, large for gestational age, stillbirth and early neonatal death. Finnish Birth Registry data included 507,659 prima gravida women who delivered singletons with at least 22 weeks’ gestation 1990- 2010. There were 12,854 hairdressers, 1841 cosmetologists and 3587 laboratory workers. Control populations: 40,405 teachers, 1968 musicians and 447,004 women-general population. When hairdressers were compared to the general public, there were three marginally statistically significant results for new-borns: SGA, (OR 1.01, 95% CI 1.00 - 1.02), LGA, (OR 1.02, 95% CI 1.00 - 1.03) and post-term delivery (OR 1.06, 95% CI 1.02 -1.11). Marginal increased statistically significant results for pooled effect size (ES) from the meta-analysis were found for LBW among hairdressers ES 1.083 (95% CI, 1.017-1.153) and SGA infants among hairdressers ES 1.077 (95% CI, 1.006-1.153). Hairdressers may be at a marginally increased risk of low birth weight and small size for gestational age. This may be due to occupational exposure.

This thesis sheds light on chronic pelvic pain in following areas: 1. Summarising prevalence of chronic pelvic pain 2. Exploring the risk factors in chronic pelvic pain, 3. Exploring beliefs on laparoscopic uterosacral nerve ablation and 4. Developing a protocol for a Multicentre randomised controlled trial of laparoscopic uterosacral nerve ablation.

COTS provided the necessary information for planning a definitive trial investigating the clinical effectiveness of monofilament non-braided suture materials in reducing pregnancy loss rate following cervical cerclage compared to the traditional multifilament braided sutures. COTS study was a stepwise process, which initially involved retrospective data analysis and later was supported by the evidence from national survey and systematic review. Evidence from retrospective analysis and systematic review confirmed that the research question about the suitability of multifilament/braided sutures in cervical cerclage; and that they may be associated with poor obstetric outcome. The Systematic review confirmed that at the time of writing there were no RCTs addressing this issue. Our national survey proved that this scientific question is of significant interest to the Obstetrical community and that the practice with cerclage varies across the country. Based on the findings of COTS trial, funding was sought from the NIHR HTA programme, and we were successful in being awarded £1.2 million (co-applicant) to conduct a multi-centre randomised controlled trial (RCT): The C-STICH trial Cerclage Suture Type for an Insufficient Cervix and its effect on Health outcomes Trial.

Research presented in this thesis was driven by the need to identify risk factors that predict local recurrence (LVR) in patients with vulval cancer (VSCC), and the need for more effective treatments for women with vulvar intraepithelial neoplasia (VIN). To identify the risk factors that predispose women to LVR, a multivariate analysis was performed on a well-characterized cohort of women treated for VSCC. This analysis revealed that the only independent predictor of LVR was the presence of Lichen Sclerosis (LS). These women were five times more likely to recur than those without LS. VIN is a recognised precursor lesion of HPV-positive VSCC. Topical application of Epigallocatechin-3-gallate (EGCG), a green tea polyphenol, has been shown to be an effective treatment for genital warts; a condition caused by HPV. Although the mechanism(s) by which EGCG influences the growth of HPV-associated proliferative disorders are unknown, I demonstrate that EGCG inhibits cell proliferation and promotes apoptosis, an effect that was accompanied by down-regulation of the E6 and E7 proteins and the induction of p53, p21 and pRb. Biochemical analysis revealed that EGCG did not stimulate E6 degradation by enhancing poly-ubiquitination and proteasome-mediated degradation, suggesting that EGCG-mediated E6 proteolysis occurred through other mechanisms.

Background: Vulnerability is universally present but experienced biopsychosocially on an individual level. Stigma develops when populations are labelled vulnerable. Individual vulnerability can be lessened by resources accessed to assist in developing resilience. A deeper analysis of vulnerability and resilience is required to inform policy, ethics, law and social life. Design: Qualitative, quantitative and mixed method approaches were used. Sample: Five papers represented the perspectives of 102 women, 21 clinicians and 13 student midwives. A further paper presented a concept analysis which included the perspectives of 10,067 women and 325 clinicians (total sample size women n=10,169; clinicians n=346; student midwives n=13). Methods: Gadamer’s ontological perspective of time, place and culture and was seen through Engel’s biopsychosocial lens. Epistemologically, truth originated from multiple realities. Methodologically, women’s experiences were captured via mixed methods. 7 Analysis: Thematic analysis and descriptive statistics were synthesised via framework analysis. Findings: A coherent theme of vulnerability in maternity care was apparent. Women’s concerns were trivialised. The professional’s style of communication determined the women’s experience of maternity care. Clinician control of care provision undermined women’s ability to choose. Women developed resilience in adverse circumstances via: accessing other supportive members of society, identifying their need for information, talking to others and developing accommodative coping strategies. Conclusion: The new conceptual model, in this thesis, should be evaluated via mixed methods. A biopsychosocial approach should underpin informed choice. Clinicians need raised awareness about how interaction can lower women’s self-esteem and build resilience in others. Higher education needs to challenge preconceived biases in safe environments via reflective processes. Research should explore women’s influential circle in decision making during maternity care. Women should be involved in the design of research to inform how best to capture their complex lived experience. Funders of research and ethics committees should request information about how implementation of evidence may be influenced by the current clinical environment. Impact should be measured post implementation. Social policy should be informed by a deeper, conceptual analysis of vulnerability and resilience.

Introduction: Pelvic organ prolapse is present in up to 50% of women. It is commoner in older women, often above 50 years of age and associated with hormonal e.g. oestrogen decline, and stiffer vaginal tissue. Pregnancy, on the contrary is a physiologic condition with higher oestrogen level and exhibits reversible structural and mechanical changes in pelvic tissues. Advanced glycation products, the ageing markers, stiffen connective tissues. There has been no previous systematic study on the relationship and action mechanisms of oestrogen, glycation level and mechanical property of vaginal tissues. Aim: The aim of this project was to study the ultrastructural and mechanical properties of vaginal tissues, and understand the relationship and possible mechanisms of accumulation of glycation (pentosidine), vaginal wall mechanics and oestrogen receptor (ER-a) expression in the vaginal tissues in both pregnancy and prolapse. It was hypothesised that prolapse is a disease of accelerated ageing and that mechanical and ultrastructural changes in prolapsed tissues and oestrogen decline are related to the elevation of glycation content in the tissues. Methods: Following ethical approval, vaginal tissues from 49 women with prolapse and 16 controls were obtained and proformas containing information on known and suspected prolapse associations were completed for each participant. Female Sprague Dawley rats‘ vaginal tissues were used for the pregnancy study. Nano-scale, micro-scale and tissue level mechanical characterizations of the tissues were performed using ball indentation technique, scanning electron microscopy, peakforce nanomechanical property mapping atomic force microscopy, and optical coherence elastography, which was applied for the first time to the study of vaginal tissues. The glycation contents of vaginal tissues in pregnancy and prolapse were quantified by high performance liquid chromatography and values obtained were analysed in comparison to medical comorbidities. Tissues were qualified by histological and immunological staining for structure (haematoxylin &eosin, trichrome and picrosirus red stainings), glyoxalase I, ER-a, elastin and neural stain. A sulphated glycosaminoglycan (sGAG)-collagen model was used to study the role of sGAG in collagen fibrillogenesis. Results: Rat vaginal tissues in pregnancy contained significantly lower amounts of pentosidine, higher oestrogen receptor-a and glyoxalase I (antioxidant enzyme) expression with larger creep, lower elastic modulus, larger fibril diameter and higher sGAG content than their non-pregnant counterparts. Observed morphological changes of the collagen fibrils in pregnancy were attributed to sGAG, which was noted to influence collagen fibril aggregation and bundling. Skin pentosidine content was reflective of vaginal tissue pentosidine in the same subjects. Pentosidine was significantly higher in prolapsed tissues and increased with age; with more age-dependent increase observed in the prolapse population and also significantly different between the 6th and 7th decades. Glyoxalase I and ER-a were poorly expressed in the prolapsed tissues in comparison to controls. Prolapsed tissues had notably disorganized ultrastructure and higher collagen fibril modulus. At all levels of tissue organization, prolapsed tissues were stiffer than controls, with increased stiffening at the more superficial layers of the tissue. Hypertension and smoking were associated with higher glycation and prolapse. In both pregnancy and prolapse, higher expression of glyoxalase I and ER-a were associated with lower glycation content of the vaginal tissues and lower modulus. These observations have led to the suggestions that oestrogen plays an important role in increase or reduction of glycation through an oestrogen-gluthathione-glyoxalase (antioxdant) pathway, which directly affects vaginal tissues‘ mechanics. Future implications: These findings have implications to the current understanding of how prolapse may occur and can inspire future translational research on improved treatment of women with prolapse. Oestrogen may significantly influence the temporary and permanent mechanics of pelvic tissues such as the vaginal wall through its modulation of glycation accumulation within the tissues. Oestrogen thus shows promise of a potential future medical treatment for early stages of prolapse. The knowledge of new prolapse comorbidities can aid the early detection and possible prevention of prolapse through a high index of suspicion.