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The placenta is a fast-evolving organ with large morphological and histological differences across eutherians, but the genetic changes driving placental evolution have not been fully elucidated. Transposable elements, through their capacity to quickly generate genetic variation and affect host gene regulation, may have helped to define species-specific trophoblast gene expression programs. Here we assess the contribution of transposable elements to human trophoblast gene expression as enhancers or promoters. Using epigenomic data from primary human trophoblast and trophoblast stem-cell lines, we identified multiple endogenous retrovirus families with regulatory potential that lie close to genes with preferential expression in trophoblast. These largely primate-specific elements are associated with inter-species gene expression differences and are bound by transcription factors with key roles in placental development. Using genetic editing, we demonstrate that several elements act as transcriptional enhancers of important placental genes, such as CSF1R and PSG5. We also identify an LTR10A element that regulates ENG expression, affecting secretion of soluble endoglin, with potential implications for preeclampsia. Our data show that transposons have made important contributions to human trophoblast gene regulation, and suggest that their activity may affect pregnancy outcomes.

Endometrial glands are essential for fertility, consisting of ciliated and secretory cells that facilitate a suitable uterine environment for embryo implantation. This study sought to determine whether an endometrial gland-specific transcriptome and splicing profile are altered in women with recurrent pregnancy loss. Our data provide a comprehensive catalogue of cilia and progestagen-associated endometrial protein (PAEP) gene isoforms and relative exon usage in endometrial glands. We report a previously unannotated endometrial gland cilia transcript GALNT11 and its susceptibility to exon skipping. Key endometrial receptivity gene transcripts are also reported to change in endometrial glands of women with recurrent pregnancy loss. The endometrial gland cilia and PAEP targets identified in this study could be used to identify a perturbed endometrium, isolate causes of recurrent pregnancy loss and develop targeted therapies in personalised medicine. Lay summary Successful embryo implantation is a trade-off between the lining of the womb which receives an implanting embryo, termed the endometrium, and a good quality embryo. For days 21–24 of the menstrual cycle, the endometrium undergoes changes into a receptive state in which it can receive an implanting embryo. Inappropriate endometrial receptivity is thought to underlie recurrent pregnancy loss. Improving pregnancy success in women with recurrent pregnancy loss requires an increased understanding of the endometrium at the molecular level. Genes contain the instructions for the cell and which genes are turned on or off determine how well it can do its role. We sought to determine a gene expression pattern of human endometrial glands in women with recurrent pregnancy loss (n = 5) vs a control group (n = 5). We identify target genes altered in women with recurrent pregnancy loss. Endometrial gland markers could be used to identify inappropriate endometrial receptivity.

Endocytosis is an essential mechanism for cellular uptake in many human tissues. A range of endocytic mechanisms occur including clathrin-dependent and -independent mechanisms. However, the role of endocytosis in the placenta and the spatial localisation of individual mechanisms is not well understood. The two principal cell layers that comprise the placental barrier to maternal-fetal transfer are the syncytiotrophoblast and fetal capillary endothelium. Endocytic uptake into the syncytiotrophoblast has been demonstrated for physiological maternal molecules such as transferrin-bound iron and low density lipoprotein (LDL) and may play an important role in the uptake of several other micronutrients, serum proteins, and therapeutics at both major placental cell barriers. These mechanisms may also mediate placental uptake of some viruses and nanoparticles. This review introduces the mechanisms of cargo-specific endocytosis and what is known about their localisation in the placenta, focussing predominantly on the syncytiotrophoblast. A fuller understanding of placental endocytosis is necessary to explain both fetal nutrition and the properties of the placental barrier. Characterising placental endocytic mechanisms and their regulation may allow us to identify their role in pregnancy pathologies and provide new avenues for therapeutic intervention.

Context Single nucleotide polymorphisms (SNPs) in vitamin D metabolism pathway genes are associated with circulating 25-hydroxyvitamin D (25(OH)D) in adults. Less is known about the relationships between mother and offspring SNPs and umbilical cord blood 25(OH)D. Objective (1) To undertake a meta-analysis of the relationships of maternal and offspring SNPs in the vitamin D metabolism pathway and cord blood 25(OH)D in pregnant women including novel data; and (2) to examine these relationships in women who received antenatal cholecalciferol supplementation in a clinical trial. Methods Novel data analysis from an observational mother–offspring cohort study (Southampton Women’s Survey) and the MAVIDOS double-blind, randomized, placebo-controlled trial of 1000 IU/day cholecalciferol supplementation in pregnancy, and an electronic literature search of published studies in PubMed up to 31 July 2021. Studies reporting associations between rs12785878 (DHCR7), rs10741657 (CYP2R1), rs6013897 (CYP24A1), or rs2282679 (GC) and cord blood 25(OH)D. One published study was included in addition to the novel data analysis. Associations between both maternal and offspring SNPs at rs2282679 (GC) and rs12785878 (DHCR7), and cord blood 25(OH)D were identified. When maternal genotype was adjusted for offspring genotype, and vice versa, there was persisting evidence for associations with maternal rs12785878 (β [95% CI] 1.6 nmol/L [0.3, 2.8] per common allele), and offspring rs2282679 (β 3.1 nmol/L ]2.0, 4.4] per common allele). Maternal and offspring SNPs at rs1074657 and rs613897 were not associated with cord blood 25(OH)D. Result Associations between both maternal and offspring SNPs at rs2282679 (GC) and rs12785878 (DHCR7), and cord blood 25(OH)D were identified. When maternal genotype was adjusted for offspring genotype, and vice versa, there was persisting evidence for associations with maternal rs12785878 (β [95% CI] 1.6 nmol/L [0.3, 2.8] per common allele), and offspring rs2282679 (β 3.1 nmol/L ]2.0, 4.4] per common allele). Maternal and offspring SNPs at rs1074657 and rs613897 were not associated with cord blood 25(OH)D. Conclusion Common genetic variation in the vitamin D metabolism pathway is associated with umbilical cord blood 25(OH)D.

Introduction Pericytes are a common feature in the placental microvasculature but their roles are not well understood. Pericytes may provide physical or endocrine support for endothelium and in some tissues mediate vasoconstriction. Methods This study uses serial block-face scanning electron microscopy (SBFSEM) to generate three-dimensional (3D) reconstructions of placental pericytes of the terminal villi and transmission electron microscopy (TEM) to study pericyte endothelial cell interactions. The proportion of endothelial cell junctions covered by pericytes was determined. Results The detailed 3D models of placental pericytes show pericyte structure at a new level of detail. Placental pericytes have many fingers extending from the cell body which can span multiple capillary branches. The proportion of endothelial cell-cell junctions covered by pericytes was significantly higher than pericyte coverage of capillary endothelium as a whole (endothelium: 14%, junctions: 43%, p 3 μm away from trophoblast (27% vs 62% respectively, p, Highlights • Three-dimensional imaging highlights the structure of placental pericytes. • Placental pericytes preferentially cover endothelial junctions. • The proportion of covered junctions decreased in regions adjacent to trophoblasts. • The localisation of placental pericytes suggests endothelial coverage is non-random. • Junction coverage may alter capillary permeability in key regions of exchange.

Funder: Gerald Kerkut Charitable Trust; FundRef: http://dx.doi.org/10.13039/100012166, Funder: Rank Prize, Funder: NIHR Clinical Lectureship, Pregnancy 25-hydroxyvitamin D [25(OH)D] concentrations are associated with maternal and fetal health outcomes. Using physiological human placental perfusion and villous explants, we investigate the role of the placenta in regulating the relationships between maternal 25(OH)D and fetal physiology. We demonstrate active placental uptake of 25(OH)D3 by endocytosis, placental metabolism of 25(OH)D3 into 24,25-dihydroxyvitamin D3 and active 1,25-dihydroxyvitamin D [1,25(OH)2D3], with subsequent release of these metabolites into both the maternal and fetal circulations. Active placental transport of 25(OH)D3 and synthesis of 1,25(OH)2D3 demonstrate that fetal supply is dependent on placental function rather than simply the availability of maternal 25(OH)D3. We demonstrate that 25(OH)D3 exposure induces rapid effects on the placental transcriptome and proteome. These map to multiple pathways central to placental function and thereby fetal development, independent of vitamin D transfer. Our data suggest that the underlying epigenetic landscape helps dictate the transcriptional response to vitamin D treatment. This is the first quantitative study demonstrating vitamin D transfer and metabolism by the human placenta, with widespread effects on the placenta itself. These data demonstrate a complex interplay between vitamin D and the placenta and will inform future interventions using vitamin D to support fetal development and maternal adaptations to pregnancy., CS was funded by a Gerald Kerkut Charitable Trust studentship and BA by Rank Prize and University of Southampton Vice Chancellor���s Studentships plus the MRC. KMG was supported by the UK Medical Research Council (MC_UU_12011/4), the National Institute for Health Research (NIHR Senior Investigator [NF-SI-0515-10042], NIHR Southampton 1000DaysPlus Global Nutrition Research Group [17/63/154], and NIHR Southampton Biomedical Research Centre [IS-BRC-1215-20004]), British Heart Foundation (RG/15/17/3174) and the US National Institute on Aging of the National Institutes of Health (Award No. U24AG047867). KSJ was supported by the National Institute for Health Research (NIHR) Cambridge Biomedical Research Centre (ISBRC-1215-20014). The NIHR Cambridge Biomedical Research Centre is a partnership between Cambridge University Hospitals NHS Foundation Trust and the University of Cambridge, funded by the NIHR. The views expressed are those of the authors and not necessarily those of the NHS, the NIHR, or the Department of Health and Social Care. Experimental work performed by KSJ and FH at MRC. EWL was supported by Dr Ann Prentice (UK Medical Research Council U105960371). The SWS has been supported by grants from Medical Research Council (MRC) (4050502589 [MRC LEU]), Dunhill Medical Trust, British Heart Foundation, Food Standards Agency, National Institute for Health Research (NIHR) Southampton Biomedical Research Centre, University of Southampton and University Hospital Southampton NHS Foundation Trust, NIHR Oxford Biomedical Research Centre, University of Oxford, and the European Union���s Seventh Framework Programme (FP7/2007-2013), project EarlyNutrition, under grant agreement 289346 and the European Union���s Horizon 2020 research and innovation program (LIFECYCLE, grant agreement no. 733206). EC has been supported by the Wellcome Trust (201268/Z/16/Z) and an NIHR Clinical Lectureship. Work leading to these results was supported by the BBSRC (HDHL-Biomarkers, BB/P028179/1), as part of the ALPHABET project, supported by an award made through the ERA-Net on Biomarkers for Nutrition and Health (ERA HDHL), Horizon 2020 grant agreement number 696295. The proteomic analyses (SDG and AM) were financially supported by the National Institutes of Health (R21AI122389) and the Beckman Institute at the California Institute of Technology. This project has received funding from the European Union���s Horizon 2020 research and innovation program under the Marie Sk��odowska-Curie grant agreement InvADeRS no. 841172 to JMF. The electron microscopy image in Figure 2 was produced with help of the Biomedical imaging unit, Faculty of Medicine, University of Southampton.

The isolated perfused placental cotyledon technique has led to numerous advances in placental biology. Combining placental perfusion with mathematical modelling provides an additional level of insight into placental function. Mathematical modelling of perfusion data provides a quantitative framework to test the understanding of the underlying biology and to explore how different processes work together within the placenta as part of an integrated system. The perfusion technique provides a high degree of control over the experimental conditions as well as regular measurements of functional parameters such as pressure, solute concentrations and pH over time. This level of control is ideal for modelling as it allows placental function to be studied across a wide range of different conditions which permits robust testing of mathematical models. By placing quantitative values on different processes (e.g. transport, metabolism, blood flow), their relative contribution to the system can be estimated and those most likely to become rate-limiting identified. Using a combined placental perfusion and modelling approach, placental metabolism was shown to be a more important determinant of amino acid and fatty acid transfer. In contrast, metabolism was a less important determinant of placental cortisol transfer than initially thought. Identifying the rate-limiting factors in the system allows future work to be focused on the factors that are most likely to underlie placental dysfunction. A combined experimental and modelling approach using placental perfusions promotes an integrated view of placental physiology that can more effectively identify the processes leading to placental pathologies.

The placental microvasculature is a conduit for fetal blood allowing solute exchange between the mother and the fetus. Serial block‐face scanning electron microscopy (SBF SEM) allows ultrastructure to be viewed in three dimensions and provides a new perspective on placental anatomy. This study used SBF SEM to study endothelial cells within the human placental microvasculature from uncomplicated pregnancies. Term human placental villi were aldehyde‐fixed and processed for imaging by SBF SEM. Manual segmentation was carried out on a terminal villous capillary and an intermediate villous arteriole and venule. Twenty‐seven SBF SEM stacks from terminal villi were analysed using stereological approaches to determine the volumes of microvascular components and the proportions of pericyte coverage. SBF SEM analysis of capillary endothelial cells revealed the presence of interendothelial protrusions (IEPs) originating from the donor cell at the endothelial junction and forming deep thin projections up to 7 μm into the adjacent endothelial cells. IEP density was estimated to be in the order of 35 million cm–3 placental tissue. Pericytes cover 15% of the fetal capillary surface area in terminal villi. In comparison, the cytotrophoblast covered 24% of the syncytiotrophoblast basal membrane. A trans‐endothelial channel was observed in a region of the vasculo‐syncytial capillary. Pericyte coverage was extensive in both arteriole and venule. Three‐dimensional imaging of the placental microvasculature identified novel ultrastructural features and provided an insight into factors that may influence capillary permeability and placental function. We hypothesise that the IEPs may allow mechanosensing between adjacent endothelial cells to assist in the maintenance of vessel integrity. The numbers of endothelial junctions, the presence of trans‐endothelial channels and the extent of pericyte coverage all provide an insight into the factors determining capillary permeability., Serial block‐face scanning electron microscopy to create a 3D reconstruction of endothelial cells. Interendothelial protrusions were observed originating from a donor endothelial cell and tunnelling inside a recipient endothelial cell. These structures constitute a novel intercellular connection.

INTRODUCTION: Extracellular vesicles are now believed to be important mediators of placental-maternal communication. However, little is known about the formation of extracellular vesicles by human placenta. This study uses nanoscale three-dimensional imaging to investigate how and where placental extracellular vesicles form.METHODS: Term and first trimester human placental villi were imaged by serial block face scanning electron microscopy. These images were analysed to quantify vesicle surface density. Segmentation was performed to reconstruct three-dimensional images of extracellular vesicles. Live imaging light microscopy of first trimester villous explants was performed.RESULTS: Vesicles were observed on the tips of placental microvilli in term and first trimester placenta. In term placenta these microvillous tip vesicles had a median size of 0.55 μm and their surface area density exceeded 22000 per mm2. Microvillous tip vesicle membranes had a lower electron density than the microvillous plasma membrane. Thirty seven percent of vesicles had a complex membrane structure including double membranes, internal vesicles and vesicle chains. Budding of smaller secondary vesicles from microvillous tip vesicle membranes was observed. Live imaging of a first trimester villus explant observed formation of vesicles which were larger but visually similar to the secondary vesicles observed by electron microscopy.DISCUSSION: These observations suggest that extracellular vesicles are forming on the tips of placental microvilli prior to release into maternal blood. However, it cannot be discounted that there are maternal extracellular vesicles that have bound to microvilli. In either case, the high surface area density of microvillous tip vesicles is consistent with an important role in placental-maternal signalling.

The placental syncytiotrophoblast, a syncytium without cell-cell junctions, is the primary barrier between the mother and the fetus. Despite no apparent anatomical pathway for paracellular diffusion of solutes across the syncytiotrophoblast size-dependent paracellular diffusion is observed. Here we report data demonstrating that the syncytiotrophoblast is punctuated by trans-syncytial nanopores (TSNs). These membrane-bound TSNs directly connect the maternal and fetal facing sides of the syncytiotrophoblast, providing a pathway for paracellular diffusion between the mother and fetus. Mathematical modelling of TSN permeability based on their 3D geometry suggests that 10-37 million TSNs per cm3 of placental tissue could explain experimentally observed placental paracellular diffusion. TSNs may mediate physiological hydrostatic and osmotic pressure homeostasis between the maternal and fetal circulations but also expose the fetus to pharmaceuticals, environmental pollutants and nanoparticles.

INTRODUCTION: The placental syncytiotrophoblast is the primary barrier between the mother and the fetus. To cross the placenta, nutrients and wastes must be transported across the apical microvillous and basal plasma membranes. While the syncytiotrophoblast basal plasma membrane is typically represented as relatively smooth, it has been shown to have invaginations that may increase its surface area. This study aimed to quantify how folding of the syncytiotrophoblast basal membrane contributes to its surface area and to visualise three-dimensional structures of the basal membrane and cytotrophoblast cell structures.METHODS: Transmission electron microscope images of human term placenta were analysed using stereological approaches to quantify how folding of the syncytiotrophoblast basal plasma membrane affected surface area. Serial block-face scanning electron microscopy was used to visualise the three-dimensional structure of the syncytiotrophoblast basal membrane and cytotrophoblast cells.RESULTS: Syncytiotrophoblast basal membrane covered 69.1% of the basal lamina, with cytotrophoblast cells covering the remaining 30.9%. In basal lamina adjacent to syncytiotrophoblast, 34% was adjacent to smooth basal membrane and 66% to folded basal membrane. Syncytiotrophoblast basal membrane folds increased the surface area adjacent to basal lamina by 305%. Including regions overlying the cytotrophoblast cells, basal membrane folds increased syncytiotrophoblast basal membrane surface area by 4.4-fold relative to the basal lamina in terminal villi. Terminal and intermediate villi were similar in terms of trophoblast coverage of the basal lamina and basal membrane folding. The three-dimensional structures of the syncytiotrophoblast basal plasma membrane and cytotrophoblast cells were generated from serial block-face scanning electron microscopy image stacks.DISCUSSION: These findings indicate that the surface area of the syncytiotrophoblast basal plasma membrane is far larger than had been appreciated. We suggest that these folds increase the surface area available for transport to and from the fetus. Changes in the extent of basal membrane folding could affect nutrient transfer capacity and underlie pathological fetal growth, including fetal growth restriction and macrosomia.

The principal role of the placenta is to protect the fetus from harmful substances while actively ensuring appropriate transfer of the nutrients essential for optimal fetal growth and development. Failure of the placenta to protect or nourish the fetus can have significant consequences for the offspring, both before birth and throughout its postnatal life. Poor fetal growth is associated with increased risk of many common noncommunicable disorders in adult life, including coronary heart disease, diabetes, and some forms of cancer. These long-term consequences have been referred to as “developmental programming.” The placenta is the interface between the mother and the fetus, influencing both the intrauterine environment and fetal growth, and placental function is therefore likely to be a major determinant of programming effects. Understanding the role of the placenta in developmental programming may provide new avenues for prevention and treatment of at-risk individuals.

Endometrial glands are essential for fertility, consisting of ciliated and secretory cells that facilitate a suitable uterine environment for embryo implantation. This study sought to determine whether an endometrial gland specific transcriptome and splicing profile are altered in women with recurrent pregnancy loss. Our data provide a comprehensive catalogue of cilia and PAEP gene isoforms and relative exon usage in endometrial glands. We report a previously unannotated endometrial gland cilia transcript GALNT11 and its susceptibility to exon skipping. Key endometrial receptivity gene transcripts are also reported to change in endometrial glands of women with recurrent pregnancy loss. The endometrial gland cilia and PAEP targets identified in this study could be used to identify a perturbed endometrium, isolate causes of recurrent pregnancy loss and develop targeted therapies in personalised medicine.

Study question Do endometrial gland factors influence recurrent pregnancy loss? Summary answer The endometrial gland transcriptome during the window of implantation is altered in women with recurrent pregnancy loss compared to controls. What is known already Secretions from endometrial glands contribute to the uterine environment that supports the attachment and implantation of the embryo in early pregnancy. Studies have attempted to identify an endometrial gene expression pattern associated with recurrent pregnancy loss however, the cellular heterogeneity within the endometrium may obscure important differences in specific cell populations. Study design, size, duration An observational study comparing controls and women with recurrent pregnancy loss. Participants/materials, setting, methods Endometrial samples were collected during the implantation period of the menstrual cycle from five matched participant egg donor controls and women with recurrent pregnancy loss. Endometrial glands were isolated from fresh endometrial biopsies and RNA sequencing was performed. A differential gene expression analysis and a gene ontology enrichment analysis was performed between egg donor controls and women with recurrent pregnancy loss. Main results and the role of chance This study reports a glandular epithelium specific gene expression profile and demonstrates differential gene expression of endometrial glands from women with recurrent pregnancy loss compared to controls. 18 genes were upregulated and 1 gene was downregulated in the endometrial glands from women with recurrent pregnancy loss compared to controls (5% false discovery rate). Biological processes which contain genes that were differentially expressed in women with recurrent pregnancy loss compared to controls include epithelial cell migration and regulation of secretion by the cell. Limitations, reasons for caution This is an observational study with a relatively small sample size. Wider implications of the findings: This study identified differences in gene expression in women with recurrent pregnancy loss that are specifically associated with endometrial glands rather than endometrium as a whole. These differences could be used to identify a perturbed endometrium, isolate causes of recurrent pregnancy loss and develop targeted therapies. Trial registration number Not applicable

Introduction Placental oxidative stress features in pregnancy pathologies but in clinical trials antioxidant supplementation has not improved outcomes. N-acetylcysteine (NAC) stimulates glutathione production and is proposed as a therapeutic agent in pregnancy. However, key elements of N-acetylcysteine biology, including its cellular uptake mechanism, remains unclear. This study explores how the cystine/glutamate transporter xCT may mediate N-acetylcysteine uptake and how N-acetylcysteine alters placental redox status. Methods The involvement of xCT in NAC uptake by the human placenta was studied in perfused placenta and Xenopus oocytes. The effect of short-term N-acetylcysteine exposure on the placental villous proteome was determined using LC-MS. The effect of N-acetylcysteine on Maxi-chloride channel activity was investigated in perfused placenta, villous fragments and cell culture. Results Maternoplacental N-acetylcysteine administration stimulated intracellular glutamate efflux suggesting a role of the exchange transporter xCT, which was localised to the microvillous membrane of the placental syncytiotrophoblast. Placental exposure to a bolus of N-acetylcysteine inhibited subsequent activation of the redox sensitive Maxi-chloride channel independently of glutathione synthesis. Stable isotope quantitative proteomics of placental villi treated with N-acetylcysteine demonstrated changes in pathways associated with oxidative stress, apoptosis and the acute phase response. Discussion This study suggests that xCT mediates N-acetylcysteine uptake into the placenta and that N-acetylcysteine treatment of placental tissue alters the placental proteome while regulating the redox sensitive Maxi-chloride channel. Interestingly N-acetylcysteine had antioxidant effects independent of the glutathione pathway. Effective placental antioxidant therapy in pregnancy may require maintaining the balance between normalising redox status without inhibiting physiological redox signalling.

Placental function is essential for fetal development and establishing the foundations for lifelong health. The placental villous stroma is a connective tissue layer that supports the fetal capillaries and villous trophoblast. All the nutrients that cross the placenta must also cross the stroma, and yet little is known about this region. This study uses high‐resolution three‐dimensional imaging to explore the structural complexity of this region within the placental villi. Serial block‐face scanning electron microscopy and confocal microscopy were used to image the placental villous stroma in three‐dimensions. Transmission electron microscopy (TEM) was used to generate high resolution two‐dimensional images. Stereological approaches were used to quantify volumes of stromal constituents. Three‐dimensional imaging identified stromal extracellular vesicles, which constituted 3.9% of the villous stromal volume. These stromal extracellular vesicles were ovoid in shape, had a median length of 2750 nm (range 350–7730 nm) and TEM imaging confirmed that they were bounded by a lipid bilayer. Fifty‐nine per cent of extracellular vesicles were in contact with a fibroblast‐like stellate cell and these vesicles were significantly larger than those where no contact was observed. These stellate cells formed local networks with adherent junctions observed at contact points. This study demonstrates that the villous stroma contains extracellular macrovesicles which are considerably larger than any previously described in tissue or plasma. The size and abundance of these macrovesicles in the villous stroma highlight the diversity of extracellular vesicle biology and their roles within connective tissues., Extracellular vesicles are membrane‐bound structures released from cells into the plasma or stroma. Extracellular vesicles are thought to contain chemical messengers and have a signalling function. The two main classes of extracellular vesicle are exosomes and microvesicles, although because of their overlapping size distributions these cannot always be readily distinguished. Stromal extracellular vesicles are typically

Intrahepatic cholestasis of pregnancy (ICP) causes increased transfer of maternal bile acids to the fetus and an increased incidence of sudden fetal death. Treatment includes ursodeoxycholic acid (UDCA), but it is not clear if UDCA protects the fetus. This study explores the placental transport of the bile acid taurocholate (TC) by the organic anion-transporting polypeptide, (OATP)4A1, its effects on the placental proteome and vascular function, and how these are modified by UDCA. Various methodological approaches including placental villous fragments and Xenopus laevis oocytes were used to investigate UDCA transport. Placental perfusions and myography investigated the effect of TC on vasculature. The effects of acute TC exposure on placental tissue were investigated using quantitative proteomics. UDCA inhibited OATP4A1 activity in placental villous fragments and oocytes. TC induced vasoconstriction in placental and rat vasculature, which was attenuated by UDCA. Quantitative proteomic analysis of villous fragments showed direct effects of TC on multiple placental pathways, including oxidative stress and autophagy. The effects of TC on the placental proteome and vasculature demonstrate how bile acids may cause fetal distress in ICP. UDCA inhibition of OATP4A1 suggests it will protect the mother and fetus against the vascular effects of TC by inhibiting its cellular uptake. UDCA may protect the fetus in ICP by inhibiting OATP4A1-mediated bile acid transfer and TC-induced placental vasoconstriction. Understanding the physiologic mechanisms of UDCA may allow better therapeutic interventions to be designed specifically for the fetus in the future.-Lofthouse, E. M., Torrens, C., Manousopoulou, A., Nahar, M., Cleal, J. K., O'Kelly, I. M., Sengers, B. G., Garbis, S. D., Lewis, R. M. Ursodeoxycholic acid inhibits uptake and vasoconstrictor effects of taurocholate in human placenta.

ObjectivesTo investigate associations between placental volume (PV) at 11 weeks' gestation and offspring bone outcomes at birth, 6 years and 8 years.Methods3D ultrasound scanning was used to assess 11 week PV in a subset (n = 236) of the Southampton Women's Survey (a prospective mother-offspring cohort). Maternal anthropometric measures and lifestyle information were obtained pre-pregnancy and at 11 weeks' gestation. Offspring dual-energy x-ray absorptiometry scanning was performed within 2 weeks postnatally and at 6 and 8 years. Linear regression was used to assess associations between PV and bone outcomes, adjusting for offspring age at DXA and sex, and maternal age, height, smoking status, walking speed and triceps skinfold thickness. β are SD change in bone outcome per SD change in PV.ResultsIn adjusted models, 11 week PV was positively associated with bone area (BA) at all time points, with evidence of persisting associations with increasing childhood age (birth: n = 80, β = 0.23 [95%CI = 0.03, 0.42], 6 years: n = 110, β = 0.17 [-0.01, 0.36], 8 years: n = 85, β = 0.13 [-0.09, 0.36]). Similar associations between 11 week PV and bone mineral content (BMC) were observed. Associations with size-corrected bone mineral content were weaker at birth but strengthened in later childhood (birth: n = 78, β = 0.07 [-0.21, 0.35], 6 years: n = 107, β = 0.13 [-0.08, 0.34], 8 years: n = 71, β = 0.19 [-0.05, 0.43]).Conclusions11 week PV is associated with DXA bone measures at birth, with evidence of persisting associations into later childhood. Further work is required to elucidate the contributions of placental morphology and function to gestational influences on skeletal development.

Transfer and metabolism of vitamin D across the human placenta is required for fetal development. However, these fundamental mechanisms are not well understood and model systems are required to help understand them. The BeWo choriocarcinoma cell line is derived from extravillous trophoblast but is used as a model for villous syncytiotrophoblast and the placental barrier. Questions have been raised about the suitability of the BeWo cell line as a model for villous trophoblast. This study compares the expression of amino acid transporters and vitamin D related genes in human term placenta with the BeWo and human embryonic kidney (HEK)293 cell lines. HEK293 cells, as transporting epithelium may be more similar to placenta. Gene expression in term placenta was much more similar to HEK293 than BeWo. This study provides further evidence that the BeWo cell line is not an appropriate model for villous trophoblast and a model that more closely represents the human placenta is now required to investigate the effects of vitamin D on the placenta ex-vivo.

Placental amino acid transfer is a complex process that is essential for fetal development. Impaired amino acid transfer causes fetal growth restriction, which may have lifelong health consequences. Transepithelial transfer of amino acids across the placental syncytiotrophoblast requires accumulative, exchange and facilitated transporters on the apical and basal membranes to work in concert. However, transporters alone do not determine amino acid transfer and factors that affect substrate availability, such as blood flow and metabolism, may also become rate-limiting for transfer. In order to determine the rate-limiting processes, it is necessary to take a systems approach which recognises the interdependence of these processes. New technologies have the potential to deliver targeted interventions to the placenta and help poorly growing fetuses. While many factors are necessary for amino acid transfer, novel therapies need to target the rate-limiting factors if they are going to be effective. This review will outline the factors which determine amino acid transfer and describe how they become interdependent. It will also highlight the role of computational modelling as a tool to understand this process.

An obesogenic diet adversely affects the endogenous mammalian circadian clock, altering daily activity and metabolism, and resulting in obesity. We investigated whether an obese pregnancy can alter the molecular clock in the offspring hypothalamus, resulting in changes to their activity and feeding rhythms. Female mice were fed a control (C, 7% kcal fat) or high fat diet (HF, 45% kcal fat) before mating and throughout pregnancy. Male offspring were fed the C or HF diet postweaning, resulting in four offspring groups: C/C, C/HF, HF/C, and HF/HF. Daily activity and food intake were monitored, and at 15 weeks of age were killed at six time-points over 24 h. The clock genes Clock, Bmal1, Per2, and Cry2 in the suprachiasmatic nucleus (SCN) and appetite genes Npy and Pomc in the arcuate nucleus (ARC) were measured. Daily activity and feeding cycles in the HF/C, C/HF, and HF/HF offspring were altered, with increased feeding bouts and activity during the day and increased food intake but reduced activity at night. Gene expression patterns and levels of Clock, Bmal1, Per2, and Cry2 in the SCN and Npy and Pomc in the ARC were altered in HF diet-exposed offspring. The altered expression of hypothalamic molecular clock components and appetite genes, together with changes in activity and feeding rhythms, could be contributing to offspring obesity.

The placenta mediates the transfer of maternal nutrients into the fetal circulation while removing fetal waste products, drugs and environmental toxins that may otherwise be detrimental to fetal development. This study investigated the role of drug transporters and protein binding in the transfer of the antidiabetic drug glibenclamide across the human placental syncytiotrophoblast using placental perfusion experiments and computational modeling. In the absence of albumin, placental glibenclamide uptake from the fetal circulation was not affected by competitive inhibition with bromosulphothalein (BSP), indicating that OATP2B1 does not mediate placental glibenclamide uptake from the fetus. In the presence of maternal and fetal albumin, BSP increased placental glibenclamide uptake from the fetal circulation by displacing glibenclamide from BSA, increasing the free fraction of glibenclamide driving diffusive transport. The P-gp and BCRP inhibitor GF120918 did not affect placental glibenclamide uptake from the maternal circulation and as such this study did not find any evidence for the apical efflux transporters in placental glibenclamide transfer. Computational modeling confirmed that albumin binding and not transporter activity, is the dominant factor in the transfer of glibenclamide across the human placenta. The effect of BSP binding to albumin on promoting the diffusive transfer of glibenclamide highlights the importance of drug-protein binding interactions and their interpretation using computational modeling.

Organic anion transporters (OATs) and organic anion transporting polypeptides (OATPs) are transport proteins that mediate exchange of metabolites, hormones and waste products. Directional transport by these transporters can occur when exchange is coupled to the gradients of other substrates. This study investigates whether the activity of OATP4A1 and OATP2A1 on the maternal facing microvillus membrane of the placental syncytiotrophoblast is coupled to the glutamate gradient. OAT and OATP transporter proteins were over expressed in Xenopus oocytes to study their transport characteristics. Further transport studies were performed in term human placental villous fragments. Xenopus oocytes expressing OATP4A1 mediated glutamate uptake. No glutamate transport was observed in oocytes expressing OAT1, OAT3, OAT7 or OATP2A1. In oocytes expressing OATP4A1, uptake of estrone sulphate, thyroid hormones T3 and T4 and the bile acid taurocholate stimulated glutamate efflux. In term placental villous fragments addition of estrone sulphate and taurocholate trans-stimulated glutamate efflux. Coupling of OATP4A1 to the glutamate gradient may drive placental uptake of estrone-sulphate and thyroid hormone while also facilitating uptake of potentially harmful bile acids. In contrast, if OATP2A1 is not coupled to a similar gradient, it may function more effectively as an efflux transporter, potentially mediating efflux of prostaglandins to the mother. This study provides further evidence for glutamate as an important counter-ion driving transport into the placenta., Highlights • OATP4A1 and OATP2A1 are present on the maternal facing surface of the placenta. • OATP4A1, but not OATP2A1, activity was coupled to glutamate efflux. • Placental estrone-sulphate and thyroid hormone uptake is coupled to glutamate efflux. • Glutamate acts as a counter-ion for OATP4A1, driving transport into the placenta.

Objective MicroRNAs (miRNAs) may serve as potential biomarkers in a variety of pathologies. The aim of this study was to determine whether miRNAs could serve as blood-based markers of isolated coronary artery calcification (CAC) defined as CAC in the absence of an underlying metabolic abnormality. Methods 24 age-matched and sex-matched patients who had been referred for elective CT coronary calcium score and angiography as part of investigation for cardiac chest pain were recruited. Peripheral venesection was performed and an Agatston calcium score was derived from the CT coronary angiogram using default software. RNA was extracted using the LeukoLOCK Total RNA Isolation System for Toray’s microarray analysis and quantitative reverse transcription PCR (qRT-PCR). Results The patients were well matched for age, sex and conventional risk factors for coronary artery disease. Microarray analysis identified lower expression of miRNA-138-2-3p, miRNA-1181, miRNA-6816-3p and miRNA-8059 in patients with coronary artery calcium score (CACS)=0 vs CACS>100. qRT-PCR confirmed significant downregulation of miRNA-8059 in patients with CACS>100 (CACS=0 vs CACS>100; P=0.03). Conclusion miRNA-8059 may serve as a peripheral blood-based biomarker for the presence of CAC, as well as provide a platform for studying the pathophysiological basis of isolated CAC. Trial registration number NCT01992848; Results.

The organic anion transporter OAT4 (SLC22A11) and organic anion transporting polypeptide OATP2B1 (SLCO2B1) are expressed in the basal membrane of the placental syncytiotrophoblast. These transporters mediate exchange whereby uptake of one organic anion is coupled to efflux of a counter-ion. In placenta, these exchangers mediate placental uptake of substrates for oestrogen synthesis as well as clearing waste products and xenobiotics from the fetal circulation. However, the identity of the counter-ion driving this transport in the placenta, and in other tissues, is unclear. While glutamate is not a known OAT4 or OATP2B1 substrate, we propose that its high intracellular concentration has the potential to drive accumulation of substrates from the fetal circulation. In the isolated perfused placenta, glutamate exchange was observed between the placenta and the fetal circulation. This exchange could not be explained by known glutamate exchangers. However, glutamate efflux was trans-stimulated by an OAT4 and OATP2B1 substrate (bromosulphothalein). Exchange of glutamate for bromosulphothalein was only observed when glutamate reuptake was inhibited (by addition of aspartate). To determine if OAT4 and/or OATP2B1 mediate glutamate exchange, uptake and efflux of glutamate were investigated in Xenopus laevis oocytes. Our data demonstrate that in Xenopus oocytes expressing either OAT4 or OATP2B1 efflux of intracellular [(14)C]glutamate could be stimulated by conditions including extracellular glutamate (OAT4), estrone-sulphate and bromosulphothalein (both OAT4 and OATP2B1) or pravastatin (OATP2B1). Cycling of glutamate across the placenta involving efflux via OAT4 and OATP2B1 and subsequent reuptake will drive placental uptake of organic anions from the fetal circulation.

Introduction Placental transfer of amino acids via amino acid transporters is essential for fetal growth. Little is known about the epigenetic regulation of amino acid transporters in placenta. This study investigates the DNA methylation status of amino acid transporters and their expression across gestation in human placenta. Methods BeWo cells were treated with 5-aza-2′-deoxycytidine to inhibit methylation and assess the effects on amino acid transporter gene expression. The DNA methylation levels of amino acid transporter genes in human placenta were determined across gestation using DNA methylation array data. Placental amino acid transporter gene expression across gestation was also analysed using data from publically available Gene Expression Omnibus data sets. The expression levels of these transporters at term were established using RNA sequencing data. Results Inhibition of DNA methylation in BeWo cells demonstrated that expression of specific amino acid transporters can be inversely associated with DNA methylation. Amino acid transporters expressed in term placenta generally showed low levels of promoter DNA methylation. Transporters with little or no expression in term placenta tended to be more highly methylated at gene promoter regions. The transporter genes SLC1A2, SLC1A3, SLC1A4, SLC7A5, SLC7A11 and SLC7A10 had significant changes in enhancer DNA methylation across gestation, as well as gene expression changes across gestation. Conclusion This study implicates DNA methylation in the regulation of amino acid transporter gene expression. However, in human placenta, DNA methylation of these genes remains low across gestation and does not always play an obvious role in regulating gene expression, despite clear evidence for differential expression as gestation proceeds.

Placental transport of vitamin D and other nutrients (e.g. amino acids, fats and glucose) to the fetus is sensitive to maternal and fetal nutritional cues. We studied the effect of maternal calorific restriction on fetal vitamin D status and the placental expression of genes for nutrient transport [aromatic T-type amino acid transporter-1 (TAT-1); triglyceride hydrolase/lipoprotein uptake facilitator lipoprotein lipase (LPL)] and vitamin D homeostasis [CYP27B1; vitamin D receptor (VDR)], and their association with markers of fetal cardiovascular function and skeletal muscle growth. Pregnant sheep received 100% total metabolizable energy (ME) requirements (control), 40% total ME requirements peri-implantation [PI40, 1–31 days of gestation (dGA)] or 50% total ME requirements in late gestation (L, 104–127 dGA). Fetal, but not maternal, plasma 25-hydroxy-vitamin D (25OHD) concentration was lower in PI40 and L maternal undernutrition groups (PPPPP

The endogenous timing system within the suprachiasmatic nuclei (SCN) of the hypothalamus drives the cyclic expression of the clock molecules across the 24 h day–night cycle controlling downstream molecular pathways and physiological processes. The developing fetal clock system is sensitive to the environment and physiology of the pregnant mother and as such disruption of this system could lead to altered physiology in the offspring. Characterizing the gene profiles of the endogenous molecular clock system by quantitative reverse transcription polymerase chain reaction is dependent on normalization by appropriate housekeeping genes (HKGs). However, many HKGs commonly used as internal controls, although stably expressed under control conditions, can vary significantly in their expression under certain experimental conditions. Here we analyzed the expression of 10 classic HKG across the 24 h light–dark cycle in the SCN of mouse offspring exposed to normal chow or a high fat diet during early development and in postnatal life. We found that the HKGs glyceraldehyde-3-phosphate dehydrogenase, beta actin and adenosine triphosphate synthase subunit to be the most stably expressed genes in the SCN regardless of diet or time within the 24 h light–dark cycle, and are therefore suitable to be used as internal controls. However SCN samples collected during the light and dark periods did show differences in expression and as such the timing of collection should be considered when carrying out gene expression studies.

Introduction: placental glutamine synthesis has been demonstrated in animals and is thought to increase the availability of this metabolically important amino acid to the fetus. Glutamine is of fundamental importance for cellular replication, cellular function and inter-organ nitrogen transfer. The objective of this study was to investigate the role of glutamate/glutamine metabolism by the isolated perfused human placenta in the provision of glutamine to the fetus.Methods: glutamate metabolism was investigated in the isolated dually perfused human placental cotyledon. U–13C-glutamate was used to investigate the movement of carbon and 15N-leucine to study movement of amino-nitrogen. Labelled amino acids were perfused via maternal or fetal arteries at defined flow rates. The enrichment and concentration of amino acids in the maternal and fetal veins were measured following 5 h of perfusion.Results: glutamate taken up from the maternal and fetal circulations was primarily converted into glutamine the majority of which was released into the maternal circulation. The glutamine transporter SNAT5 was localised to the maternal-facing membrane of the syncytiotrophoblast. Enrichment of 13C or 15N glutamine in placental tissue was lower than in either the maternal or fetal circulation, suggesting metabolic compartmentalisation within the syncytiotrophoblast.Discussion: placental glutamine synthesis may help ensure the placenta's ability to supply this amino acid to the fetus does not become limiting to fetal growth. Glutamine synthesis may also influence placental transport of other amino acids, metabolism, nitrogen flux and cellular regulation.Conclusions: placental glutamine synthesis may therefore be a central mechanism in ensuring that the human fetus receives adequate nutrition and is able to maintain growth

Introduction: transfer of glucose across the human placenta is directly proportional to maternal glucose concentrations even when these are well above the physiological range. This study investigates the relationship between maternal and fetal glucose concentrations and transfer across the placenta.Methods: transfer of d-glucose, 3H-3-o-methyl-d-glucose (3H-3MG) and 14C-l-glucose across the isolated perfused human placental cotyledon was determined for maternal and fetal arterial d-glucose concentrations between 0 and 20 mmol/l.Results: clearance of 3H-3MG or 14C-l-glucose was not affected by maternal or fetal d-glucose concentrations in either circulation.Discussion: based on the arterial glucose concentrations and the reported KM for GLUT1, the transfer of d-glucose and 3H-3MG would be expected to show signs of saturation as d-glucose concentrations increased but this did not occur. One explanation for this is that incomplete mixing of maternal blood and the rate of diffusion across unstirred layers may lower the effective concentration of glucose at the microvillous membrane and subsequently at the basal membrane. Uncertainties about the affinity of GLUT1 for glucose, both outside and inside the cell, may also contribute to the difference between the predicted and observed kinetics.Conclusion: these factors may therefore help explain why the observed and predicted kinetics differ and they emphasise the importance of understanding the function of transport proteins in their physiological context. The development of a computational model of glucose transfer may improve our understanding of how the determinants of placental glucose transfer interact and function as a system

During pregnancy, nutrient partitioning between the mother and fetus must balance promoting fetal survival and maintaining nutritional status of the mother for her health and future fertility. The nutritional status of the pregnant woman, reflected in her body composition, may affect placental function with consequences for fetal development. We investigated the relationship between maternal body composition and placental system A amino acid transporter activity in 103 term placentas from Southampton Women's Survey pregnancies. Placental system A activity was measured as Na(+)-dependent uptake of 10 mumol/L (14)C-methylaminoisobutyric acid (a system A specific amino acid analogue) in placental villous fragments. Maternal body composition was measured at enrollment pre-pregnancy; in 45 infants neonatal body composition was measured using dual-energy x-ray absorptiometry. Term placental system A activity was lower in women with smaller pre-pregnancy upper arm muscle area (r = 0.27, P = 0.007), but was not related to maternal fat mass. System A activity was lower in mothers who reported undertaking strenuous exercise (24.6 vs 29.7 pmol/mg/15 min in sedentary women, P = 0.03), but was not associated with other maternal lifestyle factors. Lower placental system A activity in women who reported strenuous exercise and had a lower arm muscle area may reflect an adaptation in placental function which protects maternal resources in those with lower nutrient reserves. This alteration may affect fetal development, altering fetal body composition, with long-term consequences.

The placenta is the interface between the mother and the fetus. Its specialized structure and function provide a physical barrier, support fetal growth, and enable the fetus to adapt to environmental changes in utero. The placenta can respond to environmental stimuli by altering its function. These changes are mediated in the short term by humoral and hormonal stimuli, while in the longer term they may be mediated by epigenetic changes. These placental adaptations may buffer the fetus from harmful in utero exposures such as low or high nutrients, reduced uterine blood flow, high cortisol, and toxins. The placenta mediates the transfer of nutrients and removes toxins and waste products from the fetal circulation. Appropriate placental function is vital for optimal fetal development, and failure of the placenta to protect or nourish has significant consequences for the baby. This has long-term implications, as a reduction or acceleration in fetal growth influences the susceptibility to chronic diseases in adulthood.

Fetal growth depends on placental transfer of amino acids from maternal to fetal blood. The mechanisms of net amino acid efflux across the basal membrane (BM) of the placental syncytiotrophoblast to the fetus, although vital for amino acid transport, are poorly understood. We examined the hypothesis that facilitated diffusion by the amino acid transporters TAT1, LAT3 and LAT4 plays an important role in this process, with possible effects on fetal growth. Amino acid transfer was measured in isolated perfused human placental cotyledons (n = 5 per experiment) using techniques which distinguish between different transport processes. Placental TAT1, LAT3 and LAT4 proteins were measured, and mRNA expression levels (measured using real-time quantitative-PCR) were related to fetal and neonatal anthropometry and dual-energy X-ray absorptiometry measurements of neonatal lean mass in 102 Southampton Women's Survey (SWS) infants. Under conditions preventing transport by amino acid exchangers, all amino acids appearing in the fetal circulation were substrates of TAT1, LAT3 or LAT4. Western blots demonstrated the presence of TAT1, LAT3 and LAT4 in placental BM preparations. Placental TAT1 and LAT3 mRNA expression were positively associated with measures of fetal growth in SWS infants (P < 0.05). We provide evidence that the efflux transporters TAT1, LAT3 and LAT4 are present in the human placental BM, and may play an important role in the net efflux of amino acids to the fetus. Unlike other transporters they can increase fetal amino acid concentrations. Consistent with a role in placental amino acid transfer capacity and fetal growth TAT1 and LAT3 mRNA expression showed positive associations with infant size at birth.

Objective: to investigate whether a noninvasive fetal electrocardiography (fECG) system can identify cardiovascular responses to fetal hypoxaemia and validate the results using standard invasive fECG monitoring techniques. Design: prospective cohort study. Setting: biological research facilities at The University of Southampton. Population or Sample: late gestation ovine fetuses; n = 5. Methods: five fetal lambs underwent implantation of vascular catheters, umbilical cord occluder and invasive ECG chest electrodes under general anaesthesia (3% halothane/O2) at 119 days of gestation (term ?147 days of gestation). After 5 days of recovery blood pressure, blood gases, glucose and pH were monitored. At 124 and 125 days of gestation following a 10-minute baseline period a 90-second cord occlusion was applied. Noninvasive fetal ECG was recorded from maternal transabdominal electrodes using advanced signal-processing techniques, concurrently with invasive fECG recordings. Main outcome measures: comparison of T:QRS ratios of the ECG waveform from noninvasive and invasive fECG monitoring systems. Results: our fECG monitoring system is able to demonstrate changes in waveforms during periods of hypoxaemia similar to those obtained invasively, which could indicate fetal distress. Conclusions: these findings may indicate a future use for noninvasive electrocardiography during human fetal monitoring both before and during labour in term and preterm pregnancies.

Environmental perturbations during early mammalian development can affect aspects of offspring growth and cardiovascular health. We have demonstrated previously that maternal gestational dietary protein restriction in mice significantly elevated adult offspring systolic blood pressure. Therefore, the present study investigates the key mechanisms of blood pressure regulation in these mice. Following mating, female MF-1 mice were assigned to either a normal-protein diet (NPD; 18 % casein) or an isocaloric low-protein diet throughout gestation (LPD; 9 % casein), or fed the LPD exclusively during the pre-implantation period (3·5 d) before returning to the NPD for the remainder of gestation (Emb-LPD). All offspring received standard chow. At 22 weeks, isolated mesenteric arteries from LPD and Emb-LPD males displayed significantly attenuated vasodilatation to isoprenaline (P = 0·04 andP = 0·025, respectively), when compared with NPD arteries. At 28 weeks, stereological analysis of glomerular number in female left kidneys revealed no significant difference between the groups. Real-time RT-PCR analysis of type 1a angiotensin II receptor, Na+/K+ATPase transporter subunits and glucocorticoid receptor expression in male and female left kidneys revealed no significant differences between the groups. LPD females displayed elevated serum angiotensin-converting enzyme (ACE) activity (P = 0·044), whilst Emb-LPD males had elevated lung ACE activity (P = 0·001), when compared with NPD offspring. These data demonstrate that elevated offspring systolic blood pressure following maternal gestational protein undernutrition is associated with impaired arterial vasodilatation in male offspring, elevated serum and lung ACE activity in female and male offspring, respectively, but kidney glomerular number in females and kidney gene expression in male and female offspring appear unaffected.

An appropriate foetal cardiovascular (CV) response to reduced substrate supply (e.g. oxygen or other nutrients) is vital for growth and development, and may impact on CV control. The prevailing nutritional environment and associated CV changes may influence subsequent CV responses to challenges during late gestation, for example, umbilical cord occlusion (UCO). We investigated the effect of low-circulating glucose on foetal CV control mechanisms and response to UCO. Under general anaesthesia, late gestation foetal sheep (n = 7, 119 days gestational age (dGA), term ∼147 days) were implanted with vascular catheters, a bladder catheter, electrocardiogram electrodes and an umbilical cord occluder. Mean arterial pressure (MAP), heart rate (HR) and kidney function were monitored during maternal saline (MSAL, 125dGA) and insulin (MINS, 126dGA) infusion, and foetal CV responses were assessed during incremental doses of angiotensin II, a 90-s total UCO, and administration of phenylephrine to assess baroreflex function. During MINS infusion, the decrease in maternal and foetal blood glucose was associated with a small but significant decrease in foetal HR and reduced foetal baroreflex sensitivity (P < 0.05). The increase in foetal MAP during a 90-s UCO was greater during hypoglycaemia (P < 0.05). The MAP response to angiotensin II was not affected by hypoglycaemia. Decreased foetal HR and baroreflex sensitivity and increased CV responsiveness to UCO during hypoglycaemia indicates altered CV homoestatic mechanisms. The combination of altered nutrition and a CV challenge, such as UCO, during late gestation may have a cumulative effect on foetal CV function.

The nutritional environment during development and even prior to conception may contribute to cardiovascular risk. In mature adult female sheep, we investigated the effect of preconceptional and periconceptional maternal nutritional restriction on the vascular reactivity of arteries from four vascular beds supplying the heart, thorax, kidney and hindlimb. Welsh Mountain ewes received 100% of nutrient requirements throughout gestation (control group, C, n = 18), or 50% of nutrient requirements for 30 days prior to conception (preconceptional group, PRE, n = 20) or for 15 days either side of conception (periconceptional group, PERI, n = 31) and 100% thereafter. In 3.5-year-old female offspring, the left anterior descending coronary (LAD), left internal thoracic (LITA), right renal and second and third order femoral arteries were dissected and their reactivity was assessed by organ bath or wire myography. Vasoconstrictor responses were greater in both LAD and LITA from PERI offspring compared with C (P < 0.01), while vasoconstriction was unaffected by maternal diet in arteries from the renal and femoral circulations (P = n.s.). Endothelium-dependent and -independent vasodilatation was attenuated in third order femoral arteries of PRE and PERI groups compared with C (P < 0.05). Endothelium-independent vasodilatation was attenuated in both the LAD and renal arteries in the PERI group compared with C (P < 0.05). These data show that moderate maternal undernutrition either prior to or around conception affects vascular function of adult offspring. The effect depends on the timing of the insult, but also on the vascular bed studied and vessel hierarchy in the vascular tree.

Undernourished late gestation fetuses display asymmetric growth restriction, suggestive of a redistribution of nutritional resources. The modification of fetal organ blood supply in response to acute hypoxia is well characterized, but it is not known whether similar responses occur in response to acute reductions in nutrition, or if such late gestation responses can be influenced by early gestation nutrition. In pregnant sheep, total nutrient requirements were restricted during the peri-implantation period (PI40, 40%; PI50, 50% of total, days 1–31) or in late gestation (L, 50% total, days 104–postmortem). Control animals were fed 100% nutrient requirements. Fetal organ blood flows were measured at baseline, and during acute fetal hypoglycaemia induced by maternal insulin infusion at 125 dGA. Baseline heart rate was increased in PI40 fetuses. During hypoglycaemia, an initial rise in fetal heart rate was followed by a slower fall. Fetal femoral artery blood flow decreased, and adrenal blood flow and femoral vascular resistance increased in all fetuses during hypoglycaemia. These changes were accompanied by increased fetal plasma adrenaline and cortisol, and reduced plasma insulin levels. The maximum femoral artery blood flow response to hypoglycaemia occurred earlier in PI50 and PI40 compared with control fetuses. The late gestation fetal cardiovascular response to acute hypoglycaemia was consistent with a redistribution of combined ventricular output away from the periphery and towards central organs. One element of the peripheral vascular response was modified by peri-implantation nutrient restriction, indicating that nutritional challenges early in gestation can have an enduring impact on cardiovascular control.