Elevated maternal levels of the neutral amino acid homocysteine (Hcy) during pregnancy are associated with various complications of pregnancy and adverse neonatal outcomes, suggesting placental transport of Hcy may influence fetal development. The primary aim of this study was to characterise Hcy transport across the microvillous plasma membrane (MVM) of the syncytiotrophoblast, the transporting epithelium of human placenta.

This thesis describes the development of three-dimensional power Doppler angiography (3D-PDA) and finally magnetic resonance imaging (MRI) perfusion techniques for the assessment of the human placental blood flow. Assessment of changes in placental blood flow may inform the clinical management of pregnancies complicated by placental dysfunction and allow modification of care with the ultimate aim of improving fetal outcome and maternal health. Ideally this should involve a non-invasive technique. I hypothesised that 3D-PDA would prove to be a reliable tool to quantify placental blood flow and demonstrate reduction in conditions associated with aberrant vascularity such as pre- eclampsia, fetal growth restriction and diabetes. I began by determining the validity of 3D-PDA and whether the 3D vascular indices would change in a predictable manner, by examining their relationship to blood flow in both the myometrium in labour and placental tissue in a phantom model. This work demonstrated that there was a relationship between the 3D vascular indices and flow rates, erythrocyte concentration and depth. The relationship was, however, complex. I then assessed within- and between-observer reliability for both data measurement and acquisition in the placenta at various gestational ages using different measurement techniques depending upon whether the entire placenta could be acquired or not. These experiments revealed variable degrees of reliability and demonstrated that the 3D-PDA whole placental technique before 21 completed weeks' gestation was most reliable. Preliminary experiments testing the reliability of an MRI perfusion technique as a tool to assess placental vascularity in the third trimester are also described in the penultimate chapter. The most reliable 3D-PDA techniques where then used to examine placental vascularity between clinical groups. Between 12 and 21 weeks whole placental FI.fmbv was Significantly higher in the diabetic compared to the normal group [54.48 (SE 0.68); 50.60 (SE 0.54) p=O.OOl] and there was a significant difference across gestation (p=0.003). Between 12 and 32 weeks the stereology within a volume technique demonstrated significant differences in FrVolBV between normal and diabetic subjects (p=0.036) and across gestation (p<O.OOl) and the significant difference occurred between 16 and 32 weeks. Mean FrVolBV was also Significantly lower in subjects with pre-eclampsia 0.107 (SE 0.012) compared to those without 0.139 (0.006); p=0.021.

There is substantial data indicating that components of the cAMP signalling pathway are differentially expressed in the human myometrium during pregnancy. The effects of cAMP in most tissues and cell types are mainly modulated via protein kinase A, a heterotetrameric protein complex consisting of two regulatory (R) and two catalytic (C) subunits. In this thesis Western-blotting/immuno-precipitation, RT-PCR and functional PKA phosphorylation assays. have been used to determine the PKA holoenzymes that are expressed in the human myometrium throughout pregnancy and labour. As early as the second trimester of pregnancy a significant increase in expression of the regulatory RIIct protein subunit of PKA in the myometrium was seen. This increase in protein expression is also mirrored at the mRNA level indicating transcriptional control throughout pregnancy, whereas during parturition both transcript and protein are significantly decreased. This increase in RIIa protein also resulted in increased particulate PKA activity in the myometrium during gestation, which was subsequently decreased during labour. The RIIa subunit is associated with A kinase anchoring proteins thus directing the cAMP quiescence signal to specific sub-cellular loci within myometrial smooth muscle cells including the contractile machinery at the cytoskeleton, this effect is then removed during parturition.

The human placenta is characterised by a unique circulatory arrangement, with numerous villous trees containing fetal vessels immersed in maternal blood. Placental tissue therefore manifests a multiscale structure balancing microscopic delivery of nutrients and macroscopic flow. The aims of this study are to examine the interaction between these scales and to understand the influence of placental organisation on the effectiveness of nutrient uptake, which can be compromised in pathologies like pre-eclampsia and diabetes. We first systematically analyse solute transport by a unidirectional flow past an array of microscopic sinks, taking up a dissolved nutrient or gas, for both regular and random sink distributions. We classify distinct asymptotic transport regimes, each characterised by the dominance of advective, diffusive or uptake effects at the macroscale, and analyse a set of simplified model problems to assess the accuracy of homogenization approximations as a function of governing parameters (Peclet and Damkohler numbers) and the statistical properties of the sink distribution. The difference between the leading-order homogenization approximation and the exact solute distribution is determined by large spatial gradients at the scale of individual villi (depending on transport parameter values) and substantial fluctuations that can be correlated over lcngthscales comparable to the whole domain. In addition, we consider the nonlinear advective effects of solute-carriers, such as red blood cells carrying oxygen. Homogenization of the solute-carrier-facilitated transport introduces an effective Peclet number that depends on the slowly varying leading-order concentration, so that an asymptotic transport regime can be changed within the domain. At large Peclet and Damkohler numbers (typical for oxygen transport in the human placenta), nonlinear advection due to solute-carriers leads to a more uniform solute distribution than for a linear carrier-free transport, suggesting a "homogenizing" effect of red blood cells on placental oxygen transport. We then use image analysis and homogenization concepts to extract the effective transport properties (diffusivity and hydraulic resistance) from the microscopic images of histological sections of the normal human placenta. The resulting two-dimensional tensor quantities allow us to assess the anisotropy of placental tissue for solute transport. We also show how the pattern of villous centres of mass can be characterised using an integral correlation measure, and identify the minimum spatial scale over which the distribution of villous branches appears statistically homogeneous. Finally, we propose a mathematical model for maternal blood flow in a placental functional unit (a placentone), describing flow of maternal blood via Darcy's law and steady advective transport of a dissolved nutrient. An analytical method of images and computational integration along streamlines are employed to find flow and solute concentration distributions, which are illustrated for a range of governing system parameters. Predictions of the model agree with experimental radioangiographic studies of tracer dynamics in the intervillous space. The model supports the hypothesis that basal veins are located on the periphery of the placentone in order to optimise delivery of nutrients. We also explain the importance of dilatation of maternal spiral arteries and suggest the existence of an optimal volume fraction of villous tissue, which can both be involved in the placental dysfunction. Theoretical studies of this thesis thus constitute a step towards modelling-based diagnostics and treatment of placental disorders.

- / -

- / -