Placental metabolic flexibility is affected by maternal obesity

s / Placenta 45 (2016) 63e133 69 Objectives: A critical role for the placenta in fetal programming of adult disease is becoming increasingly evident and the response of the placenta to insults, such as oxidative stress, can have a lifelong impact on the health of the offspring. To accurately investigate such phenomena, we have developed a vascularised in vitro model of the placental barrier using primary human trophoblasts with human umbilical vein endothelial cells (HUVECs) and normal human dermal fibroblasts (NHDFs). Methods: To create a stable confluent trophoblast barrier the ‘cell-accumulation technique’was used, whereby primary human trophoblasts were coated in extra-cellular matrix (ECM) components and seeded onto transwell inserts. We mimicked the connective tissue containing fetal blood vessels, which is found directly beneath the trophoblast layer in vivo, using the same technique to layer NHDFs and HUVECs below the trophoblast barrier to produce a multilayered structure. This 3D microenvironment enables the HUVECs to self-assemble into a vascular network. Results: Using this methodology, we were able to construct functional and intact vascularized in vitro models of the placental barrier, as confirmed using dye transfer assays, electron microscopy, and immunofluorescence staining for cellular markers. Syncytialisation of the trophoblast barriers was observed by immunostaining for the adherens junction marker ECadherin. The development of extensive and uniform vascular networks within the tissue models was clearly visualised by staining for the endothelial cell marker CD31. Conclusions: This vascularised placental barrier model enables investigation of interactions between trophoblasts and endothelial networks in vitro, allowing the response of human trophoblasts to different stimuli and the resultant effect on vasculogenesis to be explored. Additionally, trophoblasts from human placentas associated with pre-eclampsia or other pregnancy disorders can be incorporated into the model and their effects studied. By replicating some of the complexity seen in vivo, this model reproduces the human placental barrier more faithfully than other in vitro models currently available. NI3.06 PLACENTAL METABOLIC FLEXIBILITY IS AFFECTED BY MATERNAL OBESITY Alina Maloyan , James Mele , Suba Muralimanoharan , Leslie Myatt . Oregon Health and Science University, Portland, OR, USA; University of Texas Health Science Center, San Antonio, TX, USA Maternal obesity is a major problem in obstetrics: it impacts pregnancy outcome and programs the offspring for adult metabolic disease. The placenta is involved in obesity-related pregnancy complications and associated adverse outcomes via its roles at the maternal fetal interface. We have shown mitochondrial dysfunction and reduced mitochondrial biogenesis in the placenta with maternal obesity. Metabolic flexibility is the ability to switch between different energy sources. Estrogen-related receptor alpha (ERRa) and peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1a) are the key regulators of the transcriptional machinery governing mitochondrial biogenesis and function. We hypothesized that placental expression of ERRa and PGC1a are affected by maternal obesity. Placentas were collected at term from lean (BMI 30, OB) following Csection (no labor, n1⁄410/group/fetal sex), and used for isolation of total protein or for syncitiotrophoblast (ST) culture. Protein expression of PGC-1a was decreased by 75% in both male and female placentas of OB vs. LN women (p<0.001). In contrast, ERRa expression was decreased only in female placentas of OB vs. LN women (50% reduction, p<0.05). Since ERRa stimulates glycolysis, extracellular acidification rate (ECAR), a measure of glycolytic consumption, was measured. ECAR decreased with maternal obesity, but only reached statistical significance in female placentas (r1⁄4 0.67, p< 0.05). Interestingly, female STs from LN women had the highest glycolytic and respiratory reserves, while male and female STs from OB displayed the lowest glycolytic and respiratory reserves, indicating that these cells operate at their maximal capacity. Protein expression of glycolytic enzymes Hexokinase 2 and Lactate Dehydrogenase A were decreased by 50% in female but not male placentas of OB vs. LN women (p<0.05). This study identifies metabolic inflexibility and an inability to switch to glycolysis in the placenta of obese women with evidence of sexually dimorphic differences between male and female placentas. O1.01 GENOME SCALE ANALYSIS OF MRNA AND MIRNA REGULATION IN PRETERM BIRTH Nathan Price. Institute of Systems Biology, Seattle, USA Preterm birth is a common pregnancy complication related to changes in placental physiology and function throughout pregnancy. There is a pressing clinical need for biomarker development for early identification of women at risk of preterm birth and especially better understanding of mechanisms to drive better therapy strategies. microRNAs are highly expressed in the placenta and circulate throughout maternal serum, making them excellent biomarker candidates to study the pathogenesis of preterm birth over the course of pregnancy. The objective of this analysis is to identify early transcriptional changes and microRNA expression in women destined to undergo spontaneous preterm labor, and to characterize miRNA regulation of transcriptional expression in relation to prematurity. microRNA and mRNA was isolated fromwhole blood of mothers who were 15 weeks pregnant, a subset of which went on to develop preterm labor(26 controls, 15 preterm labor). Sequencing data was processed and differentially expressed genes and microRNAs were identified using negative binomial linear models within DESeq2. We identified 1821 transcripts upregulated, and 2815 transcripts downregulated in preterm birth samples compared to controls. Differentially expressed miRNAs will be mapped to their mRNA targets using mirsystem, a comprehensive target gene prediction database. We will identify intersections between miRNAs and their target genes that are mutually perturbed in relation to perturbed birth, and generate a series of linear (and other) regression models to define the relationship betweenmiRNA andmRNA expression of these genes in preterm and term samples. With this, we can study the relationships between miRNA and mRNA expression in maternal serum and determine how they are perturbed in preterm and term samples. We will examine how expression of these candidate miRNAs in maternal serum relates to placental gene expression using an independent dataset of non pathological mother-infant dyads. This approach will inform us of how miRNA expression in maternal serum is reflective of expression within the placenta. Overall, this analysis will provide a more comprehensive understanding of transcriptional changes that occur in women destined to develop preterm birth, providing key insights relating to the underlying biology and ideal targets for biomarker development. O1.02 MASS SPECTROMETRY-BASED METABOLOMICS: STATE-OF-THE-ART AND THE FUTURE Thomas O. Metz. Pacific Northwest National Laboratory, Portland, USA Metabolomics, and the related field of lipidomics, are the latest and least mature of the omics disciplines. While both rely on some of the same basic analysis technologies and approaches as those for proteomics, there remain distinct challenges in achieving comprehensive coverage of the metabolome, including strategies and informatics tools that provide confident identifications of metabolites in a high throughput manner. This presentation will provide a brief overview of the analytical history of metabolomics, the current state-of-the-art in mass spectrometry-based metabolite measurements and associated data analysis, and conclude with a vision for the future and a tangible paradigm shift towards confident identification of metabolites without the use of authentic reference materials. Metabolomics measurement technologies at Pacific Northwest National Laboratory will also be highlighted. O1.03 CHROMATIN REMODELING DURING FORMATION OF THE SYNCYTIOTROPHOBLAST Andrew Adey, Sarah Vitak, Kevin Kolahi, Kent Thornburg. Department of Molecular and Medical Genetics, Oregon Health and Science University,